absorption, transport, metabolism and excretion. Nutrition and Feeds 177
Maintaining particular intensity and colour is critical to
achieving a marketable product. confer disease resistance in shrimp farming. Establishment
and maintenance of these changes in the gut microbiome
The use of enzymes in aquafeeds has gained interest in are founded on yielding enhanced growth and increased
an effort to eliminate the adverse effects on production efficiency of digestion, and increased immunocompe-
of certain compounds that exist in plant feedstuffs. The tence/disease resistance. Successful use of probiotics is
primary limitation to use of exogenous enzymes in aqua- dependent on the ability to preserve the viability of the live
feeds is cost and the loss of activity that generally ceases organisms under the conditions of feed manufacture.
around 95 °C. Thus, the enzymes cannot withstand the Nonetheless, some evidence suggests that dead cells,
much higher temperatures used in the preparation of freeze dried cells or extracts can be added to diets to serve
mixtures of feed ingredients prior to compressed or as a vehicle for the control of pathogenesis. Some probiot-
extruded pelletisation. Capitalizing on the potential ben- ics have been produced for introduction into the culture
efit of enzyme additives lies in technology that will suc- environment, but the claims of success lack documenta-
cessfully protect them from high temperatures or surface tion of consistency and reliability.
application subsequent to the manufacture of the feed.
In contrast to probiotics, prebiotics are compounds that,
Phytase, a microbial‐derived enzyme, has been used to when added to an organism’s diet, promote the growth of
improve the availability of phosphorous that composes bacterial species and/or enhance the metabolic activity
part of the phytate compound found in plants. It eliminates of resident intestinal bacteria to improve feed efficiency.
the ability of phytate to bind micro‐elements (see sec- Oligofructose and inulin are two inert/indigestible com-
tion 8.4.6) and cause them to be unavailable for uptake. As pounds that have been shown to alter the intestinal micro-
the phosphate is enzymatically removed from the phytate biome and enhance nutrient transport in the small intestine
molecule, the capacity to bind to micro‐e lements is elimi- of mice fed diets containing them. This observation estab-
nated. The release of phosphorous confers an improved lished an early foundation for the potential benefit of their
availability, as manifested by increased amounts of phos- use as additives to diets for monogastric aquatic animals.
phorous retained in bone and less phosphorous appearing Other potentially effective prebiotics include oligosaccha-
in faeces. This combined result has been repeatedly dem- rides such as mannanoligosaccharides, fructooligosaccha-
onstrated for several species of fish fed diets containing rides and galatooligosaccharides (see section 8.4.3). Even
phytase. The ideal phytase would be heat stable, effective at certain dietary fatty acids and carbohydrates have altered
low pH and low phytate concentrations, and demonstrate a the composition of microbial populations in the gastroin-
sustained activity within the gastric region. There is some testinal tract of fish. These demonstrated effects are good
evidence of the beneficial effect of the addition of carboxy- evidence to expect that dietary prebiotics will continue to
hydrases in diets containing carbohydrates whereby elicit interest and continued testing in aquafeeds whereby
increased digestibility has been determined, but convinc- the altered intestinal microbiome confers a higher resist-
ing evidence as that found from studies with pigs and poul- ance to disease. Such an achievement will serve as a p ositive
try is lacking. A mix of directly fed microbials (DFM) and contribution to the elimination of use of sub‐therapeutic
enzymes, i.e., spores of three Bacillus strains, and xylanase, levels of antibiotics as part of the management of pro-
amylase and protease improved the gut health, perfor- duction systems and thereby hinder the occurrence of
mance (body weight gain associated with improved feed unwanted antibiotic resistant strains of pathogens.
efficiency) and welfare of poultry. Synergistic activities
between the DFM and the enzymes increased the digestion Currently, documentation of the beneficial effects of
of nutrients, thereby decreasing the FCR. With the addi- probiotic and prebiotic additives to feed to improve
growth rate, digestion, immunity and resistance to path-
tional of this microbial and enzyme mix, energy utilization ogenic infections is encouraging. However, before these
from high fibre diets was found to be as good as that from additives become routine additions to aquafeed, exten-
a diet containing a lower amount of fibre. These results sive standarised evaluation of response and cost effec-
tiveness will be required for application to the diverse
may have applicability to aquatic organisms. array of aquaculture species.
Use of probiotics and prebiotics in aquaculture has con-
Recent investigations have demonstrated the effective-
tinued to engender interest and widespread application ness of the addition of short chain fatty acids (SCFAs),
in aquaculture enterprise may eventually be realised. often termed organic acids, as dietary additives that con-
Probiotics are live microbials that are generally added to fer antibiotic activity. Low concentrations of formic acid
have been effective in inhibiting the growth of five differ-
the diet of an organism to modify the microbial composi- ent species of Vibrio grown in liquid media. Enhanced
tion (microbiome) of the gastrointestinal tract. Recognised immune responses to Steptococcus galactiae in red hybrid
probiotics include bacteria of the genus Bacillus and dif- tilapia and to Vibrio harveyi in Penaeus monodon have
been observed in response to specific dietary SCFAs.
ferent lactic acid bacteria such as Lactobacillus sp. and
Streptococcus sp. Dietary probiotics have been found to
178 Aquaculture with intensive systems will lie in the purchase, storage
and provision of a nutritionally complete and balanced
8.7 Nutrition Management feed. Development of feeding practices for formulated
Strategies feeds and, in some cases, capitalizing on natural food
organisms as a complementary resource within the pro-
Applying knowledge of nutritional requirements to the duction unit itself will be essential for ensuring economic
formulation and manufacture of feeds is complemented and environmental sustainability. The potential interest
by an effective provision of these nutrients combined in production systems that address the trophic feeding
with control of anti‐nutrient compounds, the manage- behaviour of two or more species simultaneously has
ment of feedstuffs and the use of additives. In addition, continued to gain interest, evaluation and application.
natural food whose growth can be promoted and sus- These systems are termed Integrated Multi‐Trophic
tained in ponds can also play a noteworthy role in satis- Aquaculture (IMTA) and they serve to meet the impor-
faction of nutrient requirements and reductions in costs tant goal of funnelling nutrient excess that can be
of production. The common goal of these strategies is to characteristic of intensive monoculture systems into
achieve the highest feed efficiency (lowest FCR). production of another species.
The expected increase in the global demand for animal 8.7.1 Use of Natural Food as
protein, particularly demand for farmed seafood, cou- Complementary Nutrient Sources
pled with restricted resources (water, land, energy and Other practices that together use formulated feed,
capital) and global environmental change will undoubt- organic fertilisers and live food resources derived from a
edly impact how aquatic animals will be farmed. The pond production unit itself have evolved to reduce feed
characteristics of different production strategies and costs and ensure good water quality for culture. A note-
their comparative relationships to meet the challenges worthy advance is founded in biofloc technology (BFT)
are presented in Figure 8.6. Under most scenarios, that which has improved farming in traditional freshwater
movement will require a transition to highly intensive and saltwater ponds by conservation of water and land
culture systems (NRC, 2011) that preserve sustainability
with the goal of achieving the highest production per
unit of area (space) and other resources utilised. In most
cases, a large proportion of the variable costs associated
Integrated Multitrophic Aquaculture (IMTA)
Polyculture
Direct Source of Nutrients Indirect Source of Nutrients
Nutritionally complete and balanced Nutritionally incomplete
Nutrient enhancement
• Extruded
feeds • Organic fertilisation
• Pelleted feeds • Extruded/pelleted feeds
• Farm-made • Farm-made feeds
• Uneaten feed
feeds • Faeces
• Bio ocs
Intensive Aquaculture Semi-intensive Aquaculture
Systems Systems
Ponds Ponds
Tanks Raceways
Raceways
RAS
BFT
PAS
Figure 8.6 Sources of nutrients and associated aquaculture production systems; RAS‐recirculating aquaculture systems; BFT‐biofloc
technology; PAS‐partitioned aquaculture systems.
resources in concert with biosecurity and higher feed Nutrition and Feeds 179
efficiency. The technology promotes the production of
bioflocs which are aggregates of algae, protozoans, bac- used to satisfy nutrient requirements throughout the
teria and particulate organic matter that help to maintain duration of the grow‐out period in ponds as long as pop-
good quality water for culture through uptake of nitro- ulations of natural food organisms were sustained and
gen and phosphorous. In addition, animal consumption consumed by prawns as part of their diet. As a result, it
of bioflocs is a good source of nutrients (nitrogen) and has been observed that commercially‐prepared pel-
will usually permit a reduction in the protein content of letised feeds of a crude protein content of no less than
the feed that translates into a corresponding cost reduc- 18%, such as corn gluten pellets, range cubes, or wheat
tion of the feed. BFT systems are a good example of middlings, are sufficient to achieve the same growth as
addressing the special challenges of intensive aquacul- that achieved with nutritionally complete, commercially‐
ture through management techniques designed to manufactured prawn feed that commonly contains
e stablish ecological stability. These systems are impres- approximately 30% crude protein. Levels of macro‐ or
sive and proven, having been in operation at the com- micro‐nutrients within the pellets were insufficient to
mercial level for shrimp and tilapia species since the satisfy nutritional requirements, but were fulfilled
beginning of the 21st century. Variations of this biotech- through the consumption of the secondary productivity
nology exist and warrant the need to understand and in the pond. Comparison of nutrient (fatty acid) compo-
establish common principles whereby a successful tran- sition of the pellet versus the tissue supported this
sition for application to other species can be realised. assumption. Tissue analysis revealed that EFAs were
completely absent in the pelleted feed, but requirements
The biology of some farmed species dictates a different could be satisfied through consumption of natural biota.
approach whereby the best approach lies in semi‐i ntensive This management strategy also allowed use of a feed that
production systems where use of nutritionally complete contained no fishmeal or fish oil.
and balanced feeds is not needed. Two good examples are
crayfish and freshwater prawns. Crayfish, Procambarus The semi‐intensive systems described for crayfish and
clarkii and P. acutus, are often grown in the USA in shal- freshwater prawn culture are highly sustainable, repre-
low ponds where a crop grown in rows (s ection 23.4.5.2), senting a trade‐off of intensive production for a reduc-
often certain strains of rice, has been previously planted tion in the risk of the incidence of economic loss. Based
and then filled with water to a shallow depth. Upon filling on different feeding strategies that would be characteris-
of the pond, crayfish emerge from burrows where they tic of intensive and these semi‐intensive systems, the
had previously retreated when the water was drained. cost of production per unit of protein would probably be
The plant, which may or may not be harvested, does not comparable.
serve as a principal source of nutrition per se, but rather
as an indirect source of nutrition by serving as a substrate 8.7.2 Integrated Multi‐Trophic
(detrital food chain) whereby secondary productivity is Aquaculture (IMTA)
enhanced and is consumed by the crayfish. The crayfish IMTA systems are designed to improve the efficiency of
are not fed any commercially‐produced feed. production by combining a species receiving a manufac-
tured feed with an extractive species that derives nutri-
Highly efficient, i.e., sustainable, freshwater prawn, ents from faeces, excess (uneaten) feed or other nutrient
Macrobrachium rosenbergii, farming is realised within overload originating from the fed species. For example,
semi‐intensive systems to manage as much as possible fish or shrimp would be the fed species and extractive
the density dependent growth rates and the existence of species could be other fish or shrimp, echinoderms (sea
intraspecific aggressive and agonistic behaviour. Under cucumbers, sea urchins) or molluscs (mussels, clams and
the semi‐intensive production system, consumption of oysters). This approach helps to meet important goals of
benthic populations of natural pond organisms (mac- environmental sustainability through efficient nutrient
roinvertebrates) contributes to the satisfaction of nutri- cycling, as well as economic sustainability.
ent requirements. With the use of distiller’s dried grains
and solubles (DDGS) either applied as a fertiliser or pro- For example, intensive cage culture of species of fish in
vided in the form of a pelleted feed, standing populations freshwater ponds or in the ocean offers the opportunity
of macroinvertebrates in prawn production ponds were to increase yield by incorporating a species that is an
significantly higher than those in the ponds where nei- inhabitant of the benthos as part of a polyculture system.
ther feed nor fertiliser was used. Consumption of b enthic Uneaten feed and faecal material fall through the cage
macroinvertebrate populations occurred, and certain and essentially become substrates for the colonization of
taxonomic groups were preferentially consumed. The nutrient rich microorganisms. Under the cages, benthic
results suggest that at semi‐intensive stocking densities, communities proliferate and become a source of food for
less expensive nutritionally incomplete feeds could be a farmed benthic species. Preliminary evaluations of the
potential for culture of the sea cucumber, Apostichopus
180 Aquaculture body weight, estimated mean weight and survival are
made based on whether no feed or excess feed is found in
japonicus, in the water column below fish cages in Japan, the feeding tray after a certain period of feeding. The
have been conducted. Such systems offer an opportunity weight, generally age, of a species of aquatic organism
to achieve both environmental and economic sustaina- has a specific feeding rate (% of body weight per day) that
bility. Technological advances in IMTA may also find will achieve the lowest FCR and that feeding rate may be
application to RAS systems whereby extractive species divided among several feedings. Feeding rates based on
may assist in the improvement of water quality for cul- temperature and individual body weight of the Pacific
ture of the fed species. white shrimp, Litopenaeus vannamei have been devel-
oped and applied in grow‐out systems.
8.8 Feed Management
Feed consumption, growth and accordingly feed con-
As already discussed, feed performance is founded in its version efficiency may be affected by feeding frequency
formulation, manufacture, transport and storage. or time of day. Generally, the feeding of larvae involves
Growth/Ration (G/R) curves addressed in section 8.3 several feedings per day, due to a high energy demand.
attest to the fact that maximum growth and high feed For feeding of juvenile, sub‐adult or adult aquatic organ-
efficiency do not coincide at the same feeding rate. The isms, the effect of feeding once or twice per day, or within
amount of formulated feed consumed per aquatic organ- certain intervals, i.e., every two days has been studied.
ism per day is dependent on an array of factors that are An increase in daily consumption is commonly associ-
both abiotic, i.e., water temperature, level of dissolved ated with an increase in frequency of feeding. If these
oxygen, salinity, photoperiod, barometric pressure and feed management schemes do have a positive effect on
biotic, i.e., weight, intraspecific behaviour, health, age, feed efficiency, this effect must be weighed against the
life history, location in moult cycle (in crustaceans), nat- economic cost. No significant differences in amount of
ural food availability and whether a hybrid is the choice feed fed, weight gain or estimated survival were found
for culture. The time of day that feed is provided may when juvenile channel catfish were fed to satiation once
also affect consumption, growth and feed efficiency. daily, either in the morning or the afternoon or twice
However, a plan to feed at a certain time of the day may daily, morning and afternoon. Feeding to satiation every
not be logistically possible due to the sheer size of com- other day reduced weight gain but improved the FCR
mercial operations, thereby resulting in feeding occuring and this same result was observed with hybrid catfish.
over an extended interval of time during the day. Restricted feeding rates (feeding to a maximum/pond
area/day) also reduced weight gain and net yield while
Because the amount of feed consumed is influenced by FCR correspondingly decreased. When more feed is fed
many factors, many operations offer feed to satiation. under satiation feeding, weight gain and yield is higher
This method of feed management offers the opportunity but FCR also increases. The advantage of optimum
to address the combined effect of all variables and pre- weight gain rather than maximum weight gain is a con-
sumably minimises the loss of uneaten feed which con- cept that many farmers fail to acknowledge despite opti-
tributes to adverse environmental conditions, which mum weight gain being the obvious choice to achieve
include low oxygen concentration and increased phos- economic sustainability. A comprehensive review of feed
phorous levels. For the culture of certain species of fish, management strategies used globally in aquaculture is
this ability can be accomplished through the use of float- available (Shipton and Hasan, 2013).
ing or slow sinking feeds whereby feeding behaviour can
be observed. In order to feed to satiation, the act of feed- The use of demand feeders has gained popularity for
ing and the point of satiation must be observed, and this feeding rainbow trout. The demand feeder is cone
method of feeding can actually increase FCRs because shaped and attached to it is a rod that extends into the
determining satiation can be highly subjective. Higher water column. Inside the cone is a disc that has a diame-
amounts of feed are accordingly provided, weight gain is ter that is slightly smaller than the diameter of the cone
highest but FCRs are also higher. For those aquatic spe- at that location. The disc holds the feed from falling into
cies that are benthic in nature, for example, shrimp, sea the water. However, when the rod is ‘hit’ by a fish the
urchins and abalone, the amount of uneaten feed must be disc moves and permits feed to fall into the water. The
monitored to adjust feeding rates, particularly as bio- location of the disc can be changed to increase the
mass and age increases. For intensive culture of marine amount of food that is released when the rod is triggered
shrimp, feeding trays, positioned at different locations in by the fish. This method of feeding minimises incidence
a pond are used to monitor feed consumption of poor water quality and diminishes the detrimental
(Figure 22.17). A certain amount of feed is placed in a effects on growth of fish that would result from aggres-
feeding tray based on temperature and size of shrimp. sive encounters among fish of different size or some
Adjustments to the feeding rate, based on a percent of other behavioural hierarchy. Methods for distributing
feed to cultured fish and shrimp are described in later Nutrition and Feeds 181
chapters (see Figures 17.14, 17.15, 22.15 and 22.16).
food needs to be consumed per body weight increase
8.9 Emerging Research Areas (protein deposition) relative to terrestrial animals.
Thus, this aspect of aquatic animal production is a
Nutrigenomics, the study of the effects of foods and food defining characteristic of sustainable farming to meet
constituents on gene expression, and metabolomics, the the future global needs of animal protein.
study of the production and identification of chemical ●● Additional noteworthy advances in the nutrition of
biomarkers influenced by genetic and environmental fac- farmed aquatic animals can be more readily achieved by
tors, are areas of research with high potential for applica- embracing a systems‐based holistic approach founded
tion to the development of feeds with both environmentally on a continued focus on the determination of nutrient
and economically sustainable properties. Metabolomics requirements and a complementary understanding of
represents the molecular phenotype and may focus on the metabolism and utilization of nutrients and their
the metabolic products of the whole organism, or tissue, possible interactions.
cell or cell organelle level. The objective is to understand ●● The sheer number of aquatic species being farmed
how alternative feedstuffs or different nutrients can warrants a goal of the establishment of feeds and feed-
impact and regulate gene expression and ultimately influ- ing practices that will have common application based
ence specific products of metabolism. With knowledge of on similar phylogeny, feeding habits, life history and
the full gene sequence maps of aquaculture organisms, natural environment (temperature, salinity, etc.).
control of gene expression through diet and breeding of ●● Selection of new and alternative feedstuffs for use in
organisms can be used to optimise responses to changes aquafeeds, particularly sources of protein and fishmeal
in the environment and available resources. The capacity and fish oil substitutes, must be consistent with meet-
assumes particular significance in how changes in nutri- ing the standards of environmental as well as economic
tion and feed management can effectively address regional sustainability. Choice of ingredients should also focus
changes in environments and their available resources as on alternatives that will achieve equivalent or greater
imposed by the phenomenon of global warming. benefits to animal health (disease prevention) and
increase feed efficiency and thereby contribute to
Some recent research results are based on an approach, global food security.
geometric framework (GF) that contrasts with the tradi- ●● Application of environmentally and economically sus-
tional nutrient requirement evaluation responses to tainable probiotic and prebiotic feed additives whereby
graded dietary levels. GF methodology investigates the intestinal microbiomes are altered to achieve
whether animals, if provided with an array of dietary greater production efficiency and pathogenic resist-
choices, will self‐control the intake of specific nutrients to ance must be a future focus of aquaculture nutrition.
ultimately satisfy requirements. For example, consump- ●● The provision of nutrients to meet requirements in
tion of dietary protein by adult sea urchins, Lytechinus association with sustainable aquaculture should be
variegatus, has been observed to be regulated and not composed of several species‐specific strategies that
affected by either ratios or levels of other dietary macro- include not only direct provision of feeds but also indi-
nutrients, whereas consumption of dietary carbohydrate rect consumption of nutrients through the enhance-
is not regulated. Knowledge of whether such selectivity is ment of primary and secondary productivity of pond
a strategy exhibited by other farmed aquatic animals has production systems as well as the implementation of
the potential to be applied to species and age‐specific successful IMTA.
formulation of aquafeeds. ●● With the development of full gene sequence maps for
species of aquaculture importance, an increase in the
8.10 Summary knowledge of the effects of nutrients on gene expres-
sion (nutrigenomics), is possible. This information
●● A significant amount of knowledge of nutrition, feed offers the ability to establish an understanding of the
production and feed management for farming of ani- interaction of diet with such responses as susceptibil-
mal aquatic species has been generated over the past ity to disease and feed efficiency. In addition, identifi-
50 years and has had an important impact on the cation of biomarkers, metabolic products that reflect
extraordinary increases in the volume and value of changes in physiological response (metabolomics) can
global aquaculture production. lead to a successful program of breeding that would
promote efficient response under specific culture con-
●● Due to reduced needs of aquatic organisms for energy, ditions. This knowledge will be useful in making
aquaculture organisms are more efficient because less adjustments at the molecular level to regional changes
in environments and their available resources arising
from the phenomenon of global warming.
182 Aquaculture Izquierdo, M. S., Fernandez‐Palacios, H. and Tacon, A. G. J.
(2001). Effect of broodstock nutrition on reproductive
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183
9
Hatchery and Larval Foods
Paul C. Southgate
CHAPTER MENU 9.6 Harvesting Natural Plankton, 199
9.1 Introduction, 183 9 .7 Pond Fertilisation as a Food Source for Aquaculture, 199
9 .2 Foods for Hatchery Culture Systems, 183 9.8 Summary, 200
9.3 Feeding Strategy for Larval Culture, 196
9 .4 Compound Hatchery Feeds, 196 References, 201
9 .5 Development of Microdiets for Fish Larvae, 197
9.1 Introduction (Figure 9.1) are the most widely used microalgae in
aquaculture. As their name suggests, the cells of flagel-
Hatcheries are those aquaculture facilities concerned lates possess one or more flagellae, which provide motil-
with the holding and conditioning of broodstock, ity (Figure 9.2), whereas diatoms lack a flagellum and are
spawning induction, larval rearing, and early juvenile or non‐motile. Diatoms contain silica in their cell walls and
nursery culture. The range of foods used in aquaculture may possess siliceous spines. Some species of diatoms
hatcheries must therefore be appropriate for adults, exist as single cells (e.g., Chaetoceros gracilis), while
larvae and juveniles of the target species. Intensive other species have cells that are joined to form
rearing of the larval stages of fish and shellfish currently chains (e.g., Skeletonema costatum). The generalised
relies on the availability of live food organisms that are morphology of microalgae used in aquaculture is shown
usually cultured on‐site at the hatchery and include in Figure 9.1.
microalgae, rotifers, brine shrimp and copepods.
9.2.2 Culture Methods
9.2 Foods for Hatchery Culture The simplest method of microalgal production is to
Systems bloom local species of phytoplankton in ponds or tanks.
This is achieved by filling a pool or pond with local water,
9.2.1 Microalgae which is filtered to remove zooplankton, detritus and
Cultured microalgae have a central role as a food source other unwanted particulates while retaining the smaller
in aquaculture. Microalgae are used directly as a food phytoplankton. With the addition of fertiliser (usually an
source for larval, juvenile and adult bivalves, and for inorganic fertiliser), adequate light and aeration, blooms
early larval stages of some crustaceans and fish. They are of natural phytoplankton will develop. Although
also very important as a food source for rearing zoo- inexpensive, this method can be unreliable because the
plankters, such as rotifers and brine shrimp, which, in bloom is not guaranteed and there is little control
turn, are used to feed crustacean and fish larvae. over the species composition of the bloom. As such, the
nutritional value of microalgae produced in this way is
Golden‐brown flagellates (Prymnesiophyceae and unpredictable.
Chrysophyceae), green flagellates (Prasinophyceae
and Chlorophyceae) and diatoms (Bacillariophyceae) More commonly in aquaculture, monospecific cultures
of microalgae are intensively reared in systems where
Aquaculture: Farming Aquatic Animals and Plants, Third Edition. Edited by John S. Lucas, Paul C. Southgate and Craig S. Tucker.
© 2019 John Wiley & Sons Ltd. Published 2019 by John Wiley & Sons Ltd.
184 Aquaculture shown in Figure 9.3. Stock cultures are maintained under
Flagellum controlled conditions of temperature and light. During
the scaling‐up process, microalgae are usually transferred
e.g., Isochrysis sp. from container to container under axenic conditions
Pavlova sp. using a laminar‐flow or sterile transfer cabinet. Culture
vessels receiving the inoculant contain seawater that has
FLAGELLATE been filtered (usually to 0.20–0.45 µm) and sterilised by
autoclaving (Figure 9.4). Inoculating microalgae cultures
Siliceous under these conditions reduces the possibility of bacterial
appendages contamination. However, it is impracticable to use this
method for culture vessels with volumes greater than ca.
Single-celled species 20 L. In larger bag and tank cultures, efforts are made to
e.g., Chaetoceros sp. reduce the bacterial population of the culture water by
fine filtration (0.20–0.45 µm) and this may be followed
DIATOM Chain-forming species by passage of the water through an ultraviolet (UV)
e.g., Skeletonema sp. steriliser. Flasks, bags or cylinders of microalgae may
finally be transferred to indoor or outdoor ponds or pools
Figure 9.1 Diagram showing general morphology of a flagellate where a large aquaculture operation requires large vol-
and diatom. umes of microalgae.
Figure 9.2 Cells of Pavlova sp. (Prymnesiophyceae) viewed using For growth, microalgae cultures require:
a microscope. This species is a golden‐brown flagellate and their
flagellae are visible in the photo. Source: Photograph by CSIRO. ●● aeration;
Reproduced under the terms of the Creative Commons license, ●● a suitable nutrient medium; and
CC BY‐SA 3.0. ●● light.
efforts have been made to minimise or exclude bacterial At each transfer, a clean vessel containing filtered
contamination. Monospecific axenic (bacteria‐free) (p referably sterile) seawater, to which culture medium
starter cultures of many species of microalgae are availa- has been added, is inoculated with microalgae. The
ble to the aquaculture industry from specialised laborato- newly inoculated vessel is provided with a filtered
ries. These are the basis for microalgae production, which (0.20–0.45 µm) air supply to maintain microalgae cells in
involves scaling‐up the culture volume and the density of suspension and to supply sufficient carbon dioxide (CO2)
algal cells by maintaining favourable conditions for algal for their growth. The air supply may also be supple-
growth. An example of a suitable scale‐up procedure is mented directly with CO2 gas to further stimulate
growth. Microalgae cultures are maintained under a
controlled light and temperature regime, e.g., suitable
conditions for most species of microalgae are provided
by a photoperiod of 12–16 h, providing irradiance of
70–80 mE/m2/s at 20–25 °C. The growth of microalgae
follows a distinct pattern and consists of a number of
different phases (Figure 9.5).
1) A lag phase occurs following inoculation and is char-
acterised by a steady cell density.
2) The exponential or log phase is marked by a rapid
increase in cell density within the culture. This is the
time when microalgae generally have optimal nutri-
tional value.
3) The stationary phase is reached as nutrients in the
culture medium start to become limiting, and increas-
ing cell density results in reduced light intensity
within the culture; the rate of cell division slows and
cell density reaches a plateau.
4) The death phase is reached as, eventually, cells within
the culture begin to die as nutrients in the culture
medium become exhausted and the culture enters the
phase characterised by declining cell density.
Hatchery and Larval Foods 185
or
20-mL or 200-mL 500-mL culture
starter cultures 5-litre culture
20-litre carboy
500-litre bag culture
2000-litre pond
Figure 9.3 Typical scale‐up of microalgae cultures from starter cultures. Source: Reproduced from Brown et al. (1989) CSIRO Marine
Laboratories Report 205 Nutritional Aspects of Microalgae Used in Mariculture: A Literature Review, with permission from CSIRO Publishing.
3
4
2
Cell density
Figure 9.4 Sterile flask culture of microalgae. Such cultures will be 1
scaled‐up to useable volumes in larger flasks, bags and tanks (see
Figure 9.3). Source: Photograph by CSIRO. Reproduced under the Time
terms of the Creative Commons license, CC BY‐SA 3.0. Figure 9.5 General pattern of changes in cell density over time in
microalgae batch cultures. 1, Lag phase; 2, exponential or log
The cell density of a microalgae culture is usually deter- phase; 3, stationary phase; and 4, death phase.
mined by counting cells on a haemocytometer grid using
a microscope, or use of high‐speed electronic particle 3) Continuous culture involves harvesting cultured
counters. microalgae on a continuous basis and the volume
removed from the culture vessel is continually replaced
There are three main methods for culturing microalgae: by new water and fresh nutrient medium.
1) Batch culture involves growing a microalgae culture
The objective of continuous and semi‐continuous cul-
to a point at which it is completely harvested. tures is to maintain maximal growth rate (exponential
2) Semi‐continuous culture involves partial periodic phase) to maximise production and reduce variability in
harvesting of culture vessels that are topped up with
new water and fresh nutrient medium.
186 Aquaculture 9.2.4 Nutritional Value of Microalgae
When considering the suitability of various species of
the biochemical composition (and nutritional value) of microalgae as a food source, the first concern is their
resulting microalgae. In batch cultures, the nutrient physical characteristics. Factors such as:
composition can vary widely according to the growth ●● cell size
phase and age of the culture. ●● thickness of cell wall
●● digestibility
9.2.3 Nutrient Media ●● presence of spiny appendages, and
●● chain formation (e.g., diatoms)
Many nutrient media have been developed for microal- influence the nutritional value of a particular species.
gae culture. They generally contain macronutrients to Clearly, microalgae must have suitable physical character-
provide nitrogen and phosphorus (e.g., sodium nitrate, istics to enable ingestion and, once ingested, must be
sodium glycerophosphate), trace metals and vitamins. A digestible. Cultured invertebrate larvae vary in their feed-
commonly used medium is the f/2 medium of Guillard ing and digestive mechanisms, and this greatly influences
(1972), whose composition is shown in Table 9.1. Nutrient the sizes and kinds of microalgae that can be ingested and
media are made up from distilled water to which nutri- digested. For example, shrimp larvae have a complete set of
ents are added. It is convenient to make up concentrated setous mouthparts adapted to feeding on chain diatoms.
standard stock solutions of media, which are then added However, such diatoms cannot be captured and ingested
to microalgae culture vessels to provide appropriate by the ciliated feeding structures of bivalve mollusc larvae.
nutrient levels. In general, 1–3 mL of stock nutrient solu-
tions is added to each litre of culture water. Given the Assuming suitable physical characteristics appropriate
structural importance of silica in diatoms, a source of for ingestion, and subsequent digestion, the nutritional
silica (usually sodium metasilicate) should be included in value of a given microalga is determined by its biochemi-
nutrient media used to culture diatoms. A stock solution cal or nutrient composition. Biochemical composition
is prepared by dissolving sodium metasilicate (40 g) in varies greatly between species (Table 9.2). It is also influ-
1 L of distilled water, and 0.2–0.4 mL of the resulting enced by the growth phase from which the microalgae
solution is added per litre of culture water. Nutrient cells were harvested, and by abiotic factors such as:
media are generally added to smaller culture containers ●● light (photoperiod, intensity and wavelength);
(e.g., glass flasks) before they are sterilised by autoclave. ●● temperature;
Once cooled, containers are inoculated to begin new
microalgae cultures.
Table 9.1 The composition of f/2 medium for microalgae culture.
Nutrient Concentration/L Table 9.2 The gross nutritional composition of microalgae
commonly used in aquaculture.
NaNO3 75 mg
NaH2PO4 5 mg Composition (%)
*Na2SiO3 15–30 mg
Species Protein Carbohydrate Lipid
Trace metals 4.36 mg
Na2EDTA 3.15 mg Golden‐brown flagellates 44 9 25
FeCl3.6H2O 0.01 mg Isochrysis clone T‐ISO 41 5 21
CuSO4.5H2O 0.022 mg Isochrysis galbana 49 31 12
ZnSO4.7H2O 0.01 mg Pavlova lutheri
CoCl2.6H2O 0.18 mg 33 17 10
MnCl2.4H2O 0.006 mg Diatoms 33 24 10
Na2MoO4.2H2O Chaetoceros calcitrans 37 21 7
0.5 µg Phaeodactylum tricornutum
Vitamins 0.5 µg Skelotenema costatum 57 32 9
Cyanocobalamin 100 µg 39 8 7
Biotin Green flagellates
Thiamine HCl Dunaliella salina
Tetraselmis suecica
* Required for diatom cultures only.
Source: Data complied from Parsons et al. (1961), Utting (1986)
and Whyte (1987).
●● nutrient medium (composition and concentration); Hatchery and Larval Foods 187
●● salinity;
●● nitrogen availability; and compositions of various species of microalgae, which vary
●● CO2 availability. widely between species. Golden‐brown flagellates and
For example, protein levels decrease while lipid and car- diatoms generally contain relatively high levels of essen-
bohydrate levels typically increase during the stationary tial fatty acids (EFAs), whereas others, notably species of
phase of a culture. Similarly, the protein content of green algae, contain low levels of EFAs or none at all.
microalgae is greatly influenced by the nitrogen content
of the culture medium. Culture conditions also influence The carbohydrate content of microalgae is another
levels of micronutrients such as fatty acids and vitamins important factor in determining nutritional value.
in the resulting microalgae. As such, the conditions Assuming that dietary EFA requirements are met,
under which microalgae are grown, and the stage at research has shown that growth and condition of bivalve
which they are harvested, may greatly influence their larvae are correlated with dietary carbohydrate content.
nutritional value. More information on the factors influ- Dietary carbohydrate is used primarily as an energy
encing the nutrient content of cultured microalgae is source and is considered to spare dietary protein and
provided by Brown et al. (2013). lipid, which can then be utilised for tissue growth (sec-
tion 8.2). Diets consisting of more than one species of
The nutritional requirements of cultured aquatic organ- microalgae are generally considered nutritionally supe-
isms are discussed in Chapter 8. Numerous growth trials, rior to a single‐species diet. They are thought to provide a
using different species of microalgae as food, have shown better balance of nutrients by minimising any nutritional
that differences in the food value of microalgae are related deficiencies present in any of the component species.
primarily to their fatty acid and carbohydrate composi-
tions. As detailed in section 8.4.2, marine fish and shell- Choosing an appropriate species of microalgae for use
fish larvae have an essential dietary requirement for n‐3 in an aquaculture hatchery requires careful consideration
highly unsaturated fatty acids (HUFAs). As such, n‐3 of their suitability for culture and use under local condi-
HUFA content is an important factor in determining the tions. This is particularly important when microalgae are
nutritional value of microalgae, and it is generally accepted cultured in outdoor tanks. The three most important fac-
that species containing the essential fatty acids eicosapen- tors to consider for outdoor culture are temperature,
taenoic acid (EPA, 20:5n‐3) and docosahexaenoic salinity and light intensity. For example, in the tropics, the
acid (DHA, 22:6n‐3) will be of high nutritional value ability of microalgae to tolerate fluctuating salinity and
for cultured animals. Table 9.3 shows the n‐3 fatty acid temperature is particularly important in areas where cul-
tures may experience periods of high rainfall and high
temperatures. Microalgae vary in their optimal tempera-
ture and salinity ranges (Table 9.4). Light intensity also
Table 9.4 Salinity and temperature tolerances of microalgae
commonly used in aquaculture.
Table 9.3 The n‐3 fatty acid compositions (% total fatty acids) Species Salinity Temperature
of selected species of microalgae used in aquaculture. tolerance (‰) tolerance (°C)
Fatty acid Golden‐brown flagellates 15–30
Isochrysis sp. (T‐ISO) 15–30
Species 18:3n‐3 20:5n‐3 22:6n‐3 Pavlova salina 7–35 10–25
Pavlova lutheri 21–35
Golden‐brown flagellates 3.6 0.2 8.3 7–35 10–30
Isochrysis clone T‐ISO 1.8 19.7 9.4 Diatoms 15–30
Pavlova lutheri Chaetoceros calcitrans 7–35 10–20
Trace 5.7 0.4 Chaetoceros gracilis 7–35 10–20
Diatoms 0.1 11.1 0.8 Thalassiosira pseudonana
Chaetoceros gracilis 19.3 3.9 Skeletonema costatum 14–35 10–30
Chaetoceros calcitrans 11.1 10–30
Thalassiosira pseudonana 43.5 4.3 Trace Green flagellates 7–35 10–30
21.7 Tetraselmis suecica 7–35 10–30
Green flagellates 3.2 Trace Dunaliella tertiolecta 7–35
Tetraselmis suecica Nannochloris atomus 7–35
Dunaliella tertiolecta Nanochloropsis oculata
Nannochloris atomus
Source: Data from Volkman et al. (1989). Source: Data from Jeffrey et al. (1992).
188 Aquaculture have microalgae culture facilities which could result in
considerable savings in infrastructure and running costs.
affects the growth rates of microalgae and may alter their
biochemical composition and therefore their nutritional 9.2.5.1 Microalgae Concentrates
value. Again, this is of particular importance in areas of Microalgae concentrates are prepared by removing the
high natural light intensity such as the tropics. culture medium from the microalgae culture to produce
a thick paste of concentrated microalgae cells. The
On the basis of their known ranges of tolerance to resulting microalgae cells are intact and remain in sus-
these parameters, microalgae can be categorised accord- pension with minimal agitation of the culture water to
ing to their suitability for culture and use in different which they are added. Microalgae concentrates have very
environments. For example, Jeffrey et al. (1992) divided a high cell densities (billions of cells/mL) and can be stored
range of microalgae species according to their tempera- for months or years in a refrigerator or frozen. A number
ture tolerances into: of species of microalgae are now available in concen-
1) excellent universal species, which show good growth trated form from commercial suppliers. They include
species from genera commonly used in aquaculture such
at 10–30 °C (e.g., Tetraselmis suecica, T. chuii, as Tetraselmis, Isochrysis, Pavlova and the diatoms
Nannochloris atomus) Thalassiosira. Microalgae concentrates composed of a
2) excellent tropical and sub‐tropical species, which mixture of microalgae species are also available commer-
show good growth at 15–30 °C (e.g., Isochrysis clone cially and have particular application as foods for shell-
T‐ISO, Chaetoceros gracilis, Pavlova salina) fish broodstock and zooplankton production. Microalgae
3) good temperate species, which show good growth at concentrates have found increasing application in aqua-
10–20 °C (e.g., Chroomonas salina, Skeletonema cos- culture hatcheries over recent years (Reed and Henry,
tatum, Thalassiosira pseudonana). 2014) and have been used successfully as a total replace-
ment for live cultured microalgae as a food for the larvae
9.2.5 Recent Developments in Microalgae and early juveniles of a number of invertebrate species
Production (Southgate et al., 2016; Duy et al., 2017).
Despite its central nutritional role in aquaculture hatch-
eries, production and use of cultured live microalgae 9.2.5.2 Dried Microalgae
presents a number of potential problems for aquaculture Dried microalgae preparations have been produced from
hatcheries: microalgae grown heterotrophically. This technique
1) On‐site production of microalgae is labour intensive involves growing microalgae in the dark, using sugars
rather than light as an energy source (section 15.3.4).
and is associated with high running costs (up to Growth under these conditions produces microalgae
30–50% of hatchery operating costs). with a considerably different nutritional composition to
2) On‐site production of microalgae requires specialised that of the same species grown using conventional cul-
facilities and dedicated personnel. There are substan- ture methods with light. Although the number of micro-
tial establishment costs, and microalgae culture algae species that can be produced in this manner is
requires hatchery space that could otherwise be limited, some have been produced commercially. A
devoted to production of the target species. number of studies have shown the value of dried micro-
3) Microalgae cultures can crash through failure of the algae as a food source for crustacean and fish larvae, and
culture system or infection with contaminant or path- for the larvae and spat of bivalve molluscs (Knauer and
ogenic organisms. Either case may result in a shortage Southgate, 1999). Although dried microalgae prepara-
of food for the culture organisms. tions offer prolonged shelf life, significant structural
Research to overcome some of these problems has damage to the cell can result for the drying process.
resulted in the development of microalgae products that
have become commercially available to aquaculture 9.2.6 Zooplankton
hatcheries including: As well as cultured microalgae, hatcheries that culture fish
●● microalgae concentrates; and and crustacean larvae also rely on zooplankton as a larval
●● dried microalgae. food source. The two major organisms cultured for this
Both allow microalgae to be stored in concentrated form purpose are rotifers and brine shrimp. However, recent
until required. The major advantage of using concen- years have seen considerable research effort directed
trated or dried microalgae is that the microalgae is towards the development of mass‐culture techniques for
cultured at a central facility and then distributed to copepods and their use as live feeds in aquaculture.
hatcheries. This eliminates the need for hatcheries to
9.2.7 Rotifers Hatchery and Larval Foods 189
Rotifers (Brachionus species) are widely used in aquacul-
ture as a food for the larvae of fish and crustaceans, and Figure 9.6 A female rotifer, Brachionus plicatilis, with a single egg
their use in aquaculture was reviewed by Dhont and (at bottom). The body is enclosed within a shell or lorica and the
Dierckens (2013). Rotifers have a number of characteristics ciliated head is visible at the top of the animal. The flexible foot
that confer suitability as a live food for use in aquaculture extends to the right‐hand side of the photo. Source: Photograph
hatcheries including: by Sofdrakou. Reproduced under the terms of the Creative
1) suitable size range for ingestion by fish larve; Commons license, CC BY‐SA 4.0.
2) suitable for mass cultured at high densities;
3) a high reproductive rate under favourable conditions; males if not fertilised. If fertilised, they become resting
4) broad temperature and salinity tolerances; and eggs that have a dehydration‐resistant outer shell. They
5) slow swimming behaviour that incites prey capture can remain dormant for several years and hatch into
females when conditions become favourable. The pres-
responses. ence of males in rotifer cultures therefore indicates poor
The species typically used in marine aquaculture hatch- culture conditions.
eries are: 9.2.8 Rotifer Culture
●● Brachionus plicatilis (known as large or L‐type), which 9.2.8.1 Rotifer Culture Methods
Brachionus plicatilis, B. ibericus and B. rotundiformis are
is ca. 130–340 µm in length; euryhaline and productive at salinities between 4‰ and
●● B. ibericus (known as small or S‐type) which is ca. 35‰. Optimal water temperature varies between species,
with B. rotundiformis and B. plicatilis most productive at
100–230 µm in length; and high (30–35 °C) and low (15–25 °C) water temperatures
●● B. rotundiformis (known as super small or SS‐type), respectively. Rotifer cultures are generally maintained at
a salinity of 20–35‰, within a temperature range of
which is ca. 90–190 µm in length. 20–30 °C and with gentle aeration. Successful rotifer cul-
However, there is some confusion regarding the taxon- ture requires the maintenance of constant conditions.
omy of rotifers and it has been suggested, for example, Water quality must be maintained by regular cleaning to
that the B. plicatilis complex may be composed of at least prevent the build‐up of detritus and faecal matter.
14 species. Marine rotifers are generally unsuitable as a Conventional rotifer cultures can be very productive and
food for freshwater fish larvae because of their limited may reach densities of 700–1000 rotifers/mL; however,
tolerance to freshwater. However, the freshwater rotifer, ultra‐high‐density culture methods with 10 000–30 000
Brachionus calicyflorus, has shown potential as a larval rotifers/mL have been developed.
food source for freshwater species and its culture and use
in freshwater fish larviculture were discussed by Arimoro Rotifers are usually cultured using either batch, semi‐
(2006). An important consideration in rotifer culture is continuous or continuous methods. Batch culture is the
selection of a strain best suited to the mouth size of the most widely used method and is conducted with either
target species. constant culture volume with increasing rotifer density,
or constant rotifer density with increasing volume. When
Rotifers consist of a lorica or body shell from which the cultured at a constant density, new clean water is added
foot extends ventrally and the head extends dorsally
(Figure 9.6). The head has two bands of cilia used for the
capture of food particles and for locomotion. The life
cycle of the rotifer (egg‐juvenile‐adult) takes 7–10 days.
Cultures may contain both males and females, but males
are rare and considerably smaller than females. Rotifers
reproduce sexually or asexually depending on culture
conditions. Under favourable conditions, reproduction
is asexual and the female produces diploid amictic eggs,
which she carries until they hatch into females:
Female rotifer amictic egg 2n female rotifer
amictic egg 2n female rotifer, etc.
Most reproduction in cultured rotifer populations
occurs by this method. However, in unfavourable culture
conditions, reproduction occurs sexually and females
produce smaller haploid mictic eggs (n) that hatch into
190 Aquaculture culture. The microalga Nannochloropsis oculata and
bakers’ yeast are considered excellent foods for maintain-
to the culture vessel as rotifer density increases and this ing rotifer cultures. Food must be provided to rotifers
helps maintain appropriate water quality and supports several times a day and it is not uncommon for rotifer
improved production. Batch cultures are generally har- cultures to have continuous food input.
vested after two to four days when a small volume of the
culture can be used to start a new batch culture. During The nutritional value of cultured rotifers is largely
harvest rotifers are separated from culture water and determined by their food. For example, rotifers reared on
from uneaten clumps of food and flocs using mesh sieves. bakers’ yeast, which is deficient in essential fatty acids
They can be used immediately after harvest or stored at (EFA), are themselves deficient in these fatty acids. For
a low temperature (4 °C) for up to five days for later use. this reason, rotifers reared on yeasts or other foods with
low levels of EFA are usually fed microalgae (in fresh or
Use of recirculating aquaculture systems (RAS) in roti- concentrated form), or artificial feeds that are high in
fer culture supports higher rotifer densities and higher EFA, prior to their use as a larval food source. This pro-
production rates, and allows rotifer cultures to be main- cess is generally known as enrichment and is outlined in
tained for several weeks. Rotifer density in such systems section 9.2.14.
may reach > 10 000 rotifers/mL and because of this
greater technical input is required including input of 9.2.9 Brine Shrimp
oxygen or ozone to maintain dissolved oxygen levels Brine shrimp (Artemia species) are found worldwide in
above 4 ppm. salt lakes and similar habitat, in environments with salin-
ities ranging from approximately 10 to 340 ‰. Their
The health of rotifer cultures can be assessed by moni- inactive dry cysts can be harvested in large quantities
toring swimming activity and the number of eggs pre- and stored in a dry state for many years. When immersed
sent. Rotifers should be free‐swimming (not attached to in saline water, the cysts rehydrate and become spherical,
the surfaces of culture vessels) and healthy cultures con- and the embryo inside begins to metabolise. The cyst
tain actively swimming females with many carrying more ruptures after ca. 24 h and a free‐swimming nauplius
than one egg (Figure 9.6). Rotifer cultures should be sam- emerges (Figure 9.7). This first larval stage (instar I) is
pled regularly to assess culture health. Determining the generally 400–500 µm long and brown‐orange in colour.
ratio of the number of eggs over the total number of It has a single red eye and three sets of appendages,
females allows an estimation of both the health and which have sensory, locomotory and feeding functions.
imminent productivity of the culture. The egg ratio Instar I larvae do not feed as their digestive tract is not
should be no lower than 10% and the presence of male
rotifers indicates production problems because sexual 2 3
reproduction occurs only when environmental condi- 1
tions become unfavourable.
4
9.2.8.2 Foods for Rotifer Cultures
Rotifers are hardy and are easily mass cultured on a wide 6
variety of foods. Mass culture of rotifers is usually initi-
ated by inoculating a culture of microalgae with rotifers. 5
Under suitable conditions, the rotifers consume the Figure 9.7 Hatching and development of brine shrimp (Artemia
microalgae and their population rapidly increases. species). 1, Dry cyst; 2, hydrated cyst; 3, breaking; 4, hatching;
Consumption of microalgae must be monitored regularly 5, nauplius; 6, larger metanauplius.
and more microalgae added when required; it is impor-
tant to ensure constant food availability. A portion of the
culture water is usually removed from the rearing vessels
on a daily basis and replaced with a similar volume of
microalgae culture. Water can be removed by siphoning
through a 60‐µm sieve, which prevents removal of
rotifers. Bakers’ yeast (Saccharomyces cerevisiae) and
commercially produced modified yeast are also com-
monly used as a food for rotifers, either singly or in com-
bination with microalgae. Rotifers ingest food particles
ranging in size from 0.3 µm up to 21 µm. Various species
of microalgae are used to culture rotifers, including
Nannochloropsis, Tetraselmis, Isochrysis and Pavlova,
and commercially‐available microalgae concentrates
(section 9.3.4.1) are increasingly used as a food in rotifer
Figure 9.8 Adult male and female Artemia sp. The male (upper) is Hatchery and Larval Foods 191
holding the female using his claspers and eggs can be seen in the
ovisac or broodpouch of the female. Source: Photograph by Hans more than 200 boats to harvest cysts from the lake. But
Hillewaert. Reproduced under the terms of the Creative Commons this is a natural ecosystem influenced by climatic condi-
license, CC BY-SA 4.0. tions that result in periodic fluctuations in cyst produc-
tion and inconsistent supply to the aquaculture industry.
yet functional. After ca. 12 h the nauplius moults to the The ’Artemia crisis’ of the 1990s saw greatly reduced
instar II stage, which has a functional gut and begins to harvests of Artemia cysts from GSL because of declin-
ingest small particles such as microalgae. ing salinity (Leger, 1999). Production at GSL has now
recovered to supply more than 90% of global demand,
Brine shrimp undergo ca. 15 moults over 8–14 days to but has fluctuated from ca. 2000–12000 t/yr over the
produce mature adults 10–20 mm in length. The body is last decade. The Artemia crisis of the 1990s highlighted
covered with a thin, chitinous exoskeleton that is shed the need for diversification in supply of Artemia cysts to
periodically, although females must moult prior to ovula- the aquaculture industry and other production sites
tion. Like rotifers, brine shrimp can reproduce sexually or were established by introducing Artemia into areas
asexually (Figure 9.8). Under favourable conditions, where they do not naturally occur. Production of
females produce free‐swimming nauplii (ovoviviparous Artemia cysts for aquaculture production now occurs in
reproduction) and under optimal conditions, brine countries such as Australia, China, Vietnam, Russia and
shrimp can reproduce at a rate of 300 nauplii every 4 days. Kazakhstan. Current global demand for Artemia cysts
However, under unfavourable conditions, such as high is ca. 3000 t/yr with greatest annual consumption of
salinity and low oxygen, the shell glands of the female ca. 1500 t in China.
become active and secrete a thick shell around the devel-
oping gastrula, which enters a dormant state (diapause). The majority of Artemia cysts supplied to the global
These embryos are released by the female as cysts (ovipa- aquaculture industry are made up by Artemia francis-
rous). Females are able to switch between these two cana from GSL. This species has also been introduced
modes of reproduction depending on environmental con- to other areas both deliberately (e.g., Vietnam) or as a
ditions. Production of cysts has obvious advantages for result of gradual dispersal and out‐competing local pop-
aquaculture. Dry cysts can be easily stored and live feed ulations (e.g., India, Sri Lanka, Australia and coastal
(in the form of nauplii) can be produced when required. areas in the Mediterranean and China). Cysts are now
marketed with emphasis on quality criteria relating to
9.2.10 Production and Sources of Brine yield (e.g., hatch rate and cysts size) and nutritional
Shrimp Cysts composition (e.g., fatty acids and vitamin contents).
The two primary and traditional sources of Artemia Development of Artemia resources other than GSL is
cysts for aquaculture are the coastal salt works in San likely to result in greater selectivity for aquaculture
Francisco Bay in California (USA) and the Great Salt hatchery managers.
Lake in Utah (USA) (Figure 9.9). The Great Salt Lake
(GSL) has become the primary source for Artemia cysts 9.2.11 Hatching Brine Shrimp Cysts
used in aquaculture. At its peak the GSL industry was Although cysts can be successfully incubated in full‐
composed of around 40 companies that used a fleet of strength (35‰) seawater, the hatching rate is generally
superior at low salinities and a salinity of 5‰ is optimal.
Cysts are incubated at densities up to 5 g/L culture
medium, which is maintained at 25–30 °C with vigorous
aeration. Dissolved oxygen content must be maintained
above 2 mg/L and, to facilitate good aeration and water
movement, culture vessels are usually V‐shaped or coni-
cally based. Cultures require a pH of 8–9 and constant
illumination at the water surface. Culture conditions
must be constant during incubation. Within 24 h, the
majority of cysts will have hatched.
To harvest hatched nauplii, aeration is stopped, caus-
ing the cyst shells to float to the top of the culture vessel.
Nauplii are positively phototaxic, and this behaviour can
be used to concentrate them prior to harvesting by
siphon. It is important that the number of cyst shells
accompanying the nauplii is limited. Cyst shells have the
potential to introduce disease and bacteria into larval
192 Aquaculture
Figure 9.9 Salt evaporation ponds used for production of Artemia cysts on the western shore of San Francisco Bay, California. Source:
Photograph by Doc Searls. Reproduced under the terms of the Creative Commons license, CC BY‐SA 2.0.
cultures and can cause digestive disorders in fish larvae. Any residual chlorine can be removed from decapsu-
Contamination with cyst shells can be minimised if the lated cysts by washing in 0.1% sodium thiosulphate solu-
cysts are decapsulated prior to incubation. tion for 1 min. The decapsulated cysts are then washed
and placed into a medium for hatching, or they may
9.2.12 Decapsulation of Cysts be stored at 4 °C for a short period before hatching.
The process in which the outer shell or chorion is Decapsulation offers major advantages in limiting
removed from hydrated brine shrimp cysts is decapsula- potential digestive and disease problems caused by cyst
tion. This is achieved by treating hydrated cysts with shells; it disinfects brine shrimp embryos and improves
hypochlorite solution, which dissolves the chorion with- hatch rate. Decapsulated cysts can also be offered
out damaging the embryo inside. Prior to decapsulation, directly as a larval food source, with a major advantage
dried cysts are rehydrated in freshwater for 60–90 min being that, prior to hatching, embryos have their maxi-
at the rate of 1 g of cysts per 30 mL of water. Approximately mum energy content. For further information on hatch-
20–30 mL of liquid bleach (sodium hypochlorite NaOCl) ing and decapulation of Artemia see Dhont and Van
is added per gram of cysts, and the solution stirred con- Stappen (2003).
tinuously. The colour of the solution changes as the cho-
rion is dissolved, and decapsulation is complete within 9.2.13 Culturing Brine Shrimp
24 min when the solution becomes orange in colour. The For the production of large or adult brine shrimp, nauplii
solution is then poured through a sieve to remove the are reared in tanks at an initial stocking density of ca.
chlorine solution. The decapsulated cysts retained on 1000–3000/L. Nauplii are initially fed microalgae at a
the sieve are washed thoroughly with seawater or fresh- d ensity of approximately 5 × 105 to 1 × 106 cells/mL; the
water until no further chlorine smell can be detected. feeding rate is adjusted as the brine shrimp grow and
more food is required. Best growth of brine shrimp Hatchery and Larval Foods 193
c ultures occurs with:
●● good aeration; levels (or a total lack) of certain EFA that are required for
●● good water quality; normal growth and development of marine larvae (sec-
●● a readily available food supply; tion 8.4.2). To overcome these deficiencies, the EFA con-
●● low light conditions; tent of rotifers and brine shrimp has to be manipulated
●● 25–30°C; and using fatty acid enrichment techniques. This process
●● 30–35‰ salinity. involves feeding a nutrient source rich in EFA to the rotifers
Culture tanks must be cleaned regularly to remove or brine shrimp prior to feeding them to the cultured
detritus and faecal matter to maintain water quality. larvae. Various materials can be used for enrichment,
Under suitable conditions, production rates in the order including microalgae, oil suspensions and self‐emulsifying
of 57 kg (wet weight) of brine shrimp per cubic metre are concentrates, microencapsulated and microparticulate
achievable using batch culture techniques. products and yeasts, and a number of enrichment prepara-
tions are available commercially. Enrichment results in an
Brine shrimp nauplii have their greatest energy content increase in the EFA content of the live food organism that
at hatching and, because instar I nauplii do not feed, there increases dietary EFA intake by the target species, support-
is a 24% reduction in organic content and a 27% reduction ing improved survival.
in lipid content between instar I and instar II. Although
the lipid and fatty acid content of brine shrimp varies The degree to which EFA are incorporated into rotifers
according to the geographical origin of the cysts, they are and brine shrimp during enrichment is influenced by a
generally considered to be deficient in essential fatty acids number of factors including:
(EFA). The nutritional value of instar II nauplii, particu- ●● the duration of the enrichment procedure;
larly their fatty acid content, can be significantly improved ●● the density of rotifers or brine shrimp; and
using an appropriate enrichment procedure. ●● the density and EFA content of the enrichment material.
Although enrichment procedures were developed to
9.2.14 Enrichment of Rotifers and Brine improve the EFA composition of live food organisms
Shrimp such as rotifers and Artemia, this process is also used to
Rotifers and brine shrimp are extensively used in aquacul- improve dietary energy intake and consumption of other
ture, primarily because they are amenable to mass culture, important nutrients such as protein, phospholipids, ster-
not because they are an ideal food source (Table 9.5). ols and vitamins. Live food enrichment is also used as a
Rotifers and many strains of brine shrimp have very low means of presenting therapeutic compounds, pigments,
antioxidants and enzymes to the larvae of the target spe-
Table 9.5 Some potential problems associated with use of rotifers cies via their foods.
and brine shrimp.
The organic load of the culture medium and live food
Disadvantage Comments organisms can become very high during the enrichment
Nutritional process and bacterial load can increase significantly.
deficiency Both brine shrimp and rotifers have Thorough rinsing of enriched live food organisms is
Nutritional inadequate fatty acid compositions for therefore necessary after their harvest and before they
inconsistency marine larvae. They must be artificially are introduced to larval culture tanks. The enrichment
Reliability of enriched prior to their use as food, which process is generally kept as short as possible to minimise
supply adds to the expense of food production bacterial load within enrichment vessels.
Contamination Live food organisms may vary in their
nutritional composition according to the 9.2.15 Copepods
source of stock (i.e., source of Artemia Copepods occur in all aquatic systems and are natural
cysts) and the nutritional composition of prey for virtually all fish larvae. There are over 10 000
their food(s) known species, with most planktonic forms ranging
Hatcheries in most parts of the world between 0.5 mm and 2.5 mm in size. Given their impor-
rely on a supply of brine shrimp cysts tance as prey for wild fish larvae, there is clear potential
from North America. Availability may for the use of copepods in aquaculture and research with
fluctuate from year to year and imports fish has shown that when copepods are included in the
may be subject to quarantine problems larval diet, survival, growth rates, pigmentation and gut
Live foods can be vectors for inadvertent development are all improved.
introduction of disease and
contaminants to larval cultures Developments in this field were recently reviewed by
Dhont and Dierckens (2013). Research has focused pri-
marily on the harpacticoid and calanoid copepods:
194 Aquaculture
Figure 9.10 Harpacticoid copepod adult female with eggs Figure 9.11 Calanoid copepod (Parvocalanus sp.) female. Source:
(right) and nauplius (left). Source: Reproduced with permission Reproduced with permission from Dr T. Camus.
from Dr T. Camus.
that resembles the adult. The copepodite goes through a
●● Harpacticoid copepods (Figure 9.10) are distinguished futher six moults before becoming an adult. The small
by a very short pair of 1st antennae with biramous 2nd size of newly hatched nauplii is appropriate for the larvae
antennae and a joint between the 4th and 5th body of fish with small mouth gape (Figure 26.9) but copepods
segments. They are generally epibenthic. can be used as a live food in aquaculture hatcheries at the
nauplius, copepodite and adult stages, depending on the
●● Calanoid copepods (Figure 9.11) are distinguished by mouth size of the target larvae.
1st antennae that are at least half the body length with
biramous 2nd antennae and a joint between the 5th Copepods are readily digested by fish larvae and are
and 6th body segments. They are generally pelagic. superior to rotifers and brine shrimp in their nutritional
value. In particular, they contain high levels of n‐3 HUFA
Most copepods reproduce sexually with eggs being and a wide range of copepod species have been used as
released directly into the water column (calanoids) or food for marine fish larvae (see section 20.3.1.4;
nauplii hatching directly from egg sacs (harpacticoids Table 9.6). Nauplii and copepodites of calanoid copep-
and cyclopiods). Nauplii develop through six stages, sep- ods are valuable dietary components for a wide range of
arated by moults, before reaching the copepodite stage marine fish species including flatfish such as halibut and
Table 9.6 Some copepods used as food for fish larvae.
Type Species Type Species
Calanoid Harpacticoid
Acartia clausi Euterpina acutifrons
Acartia tonsa Nitokra lacustris
Acartia sinjiensis Tigriopus japonicus
Acartia tsuensis Tigriopus californicus
Acartia tranteri Tigriopus brevicornis
Centropages spp. Tisbe furcata
Eurytemora affinis Tisbe holothuriae
Gladioferens imparipes Amphiascoides atopus
Paracalanus sp.
Parvocalanus sp.
Pseudocalanus sp.
Pseudodiaptomus spp.
Temora spp.
flounder, for which they are a preferred first food for Hatchery and Larval Foods 195
larvae supporting enhanced development, pigmentation Table 9.7 Culture parameters used for the calanoid copepod
and improved survival (section 20.13). However, much (Acartia tonsa) and the harpacticoid (Tigriopus sp.).
of our knowledge of the nutritional value of copepods is
based on small‐scale (experimental) culture and, where Culture Calanoid Harpacticoid
copepods are used as a larval food on a larger scale, they Parameter
are often obtained serendipitously from blooms in local
water bodies or from blooms in dedicated ponds follow- Tank: 200 L (1500 × 50 cm) 500 L
ing fertilisation (section 9.7). Water: 1‐µm filtered, 35 ‰ 1‐µm filtered, 35‰
Watyer system: Batch (5 % change/day) Semi flow‐through
9.2.15.1 Copepod Culture Temperature: 16–18°C 24–26 °C
Culture of copepods and their use as live prey in aqua- Stocking density <100/L 20–70/mL
culture was reviewed by Lee et al. (2005). Copepods used Female:Male 1:1 Stocked with
as live foods in aquaculture are commonly produced in ratio gravid females
extensive or semi‐extensive outdoor culture systems Aeration Gentle Gentle
such as ponds or lagoons. The ponds are filled with Food type Microalgae Chaetoceros
crudely filtered (to 40 µm) sea water to which fertilisers (Rhodomonas) gracilis or green
are added to promote a bloom of local microalgae. Once flagellate
the bloom is established, copepods obtained from filter- Food ration 8 × 108 cells/day 5 × 104–2 × 105
ing local sea water are added to the ponds, providing a cells/mL
mixture of species (and sizes) that will proliferate within
the ponds. Copepod densities of 300/L to more than Source: Data from Lavens and Sorgeloos (1996).
1000 nauplii/L are possible in such systems. Fish larvae
are generally stocked into the ponds at relatively low and harvest tanks, and the parameters shown are those in
densities once good copepod populations have become the basis tanks. In this system, 10 L of culture water (con-
established. For more information about pond fertilisa- taining eggs) is siphoned from the bottom of the basis
tion as a means of live food production see section 9.7. tank and replaced by clean filtered sea water. The eggs
contained in the siphonate are retained on a 40 µm mesh
Copepods are generally difficult to mass culture in and transferred to the growth tank where maximal den-
intensive culture systems that are characterised by vari- sity may reach 6000 eggs/L. Nauplii hatch after 24 hours
able and unreliable production. The productivity of such and the microalgae Isochrysis is provided at a density of
systems is influenced by factors such as cannibalism (e.g., 1000 cell/mL, increasing to 1500 cells/mL after 10 days
Acartia spp.) and appropriate system design for epiben- with the addition of a second microalgae, Rhodomonas.
thic species. The challenge remains to develop, cost‐ The time required to reach 50% fertilised female copepods
effective large‐scale copepod production methods that in the growth tank (generation time) is about 20 days
can match those used for traditional live prey organisms when adults are collected using a 180 µm mesh and
(rotifers and Artemia). Approximately 60 copepod spe- transferred to either newly established basis tanks, for
cies have been successfully cultured. Maximum densities the production cycle to begin again, or to the harvest
of cultivated calanoid copepods with potential in aqua- tanks for use as a larval food. Production from basis tanks
culture are ca. 100–2000 adults/L although cyclopoid can be around 95 000 eggs/day or 25 eggs/female/day.
copepods may achieve densities of ca. 5000/L. Most har- Basis tanks are emptied and cleaned 2–3 times a year.
pacticoids can be grown at relatively high densities (ca.
10 000 to 400 000/L) in high volume systems incorporat- Harpacticoid Culture
ing three‐dimensional structures. Production of up to Harpacticoid copepods are generally considered to have
8.2 million nauplii/day was reported for a 266 L system a number of characteristics that are well suited to
holding Nitokra lacustris. aquaculture production including:
●● high fecundity and short generation time;
Calanoid Culture ●● acceptance of a wide variety of foods including rice
Calanoids have limited tolerance to high density culture
with cannibalism resulting from increased encounters bran, yeast and microalgae;
between individuals. This can be minimised by separat- ●● ability to achieve high density in culture (e.g., Tigriopus
ing nauplii from adults during culture. Basic culture
parameters for Acartia tonsa are shown in Table 9.7. fed on rice bran increased in density from 0.5/mL to
These are based on a continuous production system con- 9.5/mL in 12 days); and
sisting of three culture units: basis tanks, growth tanks ●● broad environmental tolerance such as a salinity range
of 15–70 mg/L and temperature range of 17–30 °C.
196 Aquaculture The volume of a microalgae, rotifer or brine shrimp
culture that has to be added to larval rearing tanks to
Basic culture parameters for Tigriopus sp. are shown in obtain the desired density of food organisms is calcu-
Table 9.7. The culture is based on 10–100 gravid females lated as:
that are used to inoculate culture tanks and will support
a population growth rate of around 15% per day to an A B C /D
optimal stocking density of 20–70 copepods/mL. The where A is the required volume (L) of the live food cul-
generation time is around 8–11 days. Early nauplii can ture; D is the density of the live food culture (number/
be collected from the culture tank using a 37 µm mesh mL); B is the required density of microalgae, rotifers or
and copepodites can be collected on a 100 µm mesh. brine shrimp in the larval tank (number/mL); and C is
the volume of the larval tank (L).
It is likely that copepods will assume increasing impor-
tance as a hatchery food, as more reliable mass‐culture The feeding regimen is an important aspect of
techniques are developed, and more species are investi- hatchery management. Overfeeding is wasteful and
gated for their culture potential. expensive. It also compromises water quality, which
can lead to disease and affect larval performance.
9.3 Feeding Strategy for Larval Underfeeding reduces growth rates, thereby increas-
Culture ing hatchery running costs. It is important to monitor
the presence of food in larval tanks to avoid these
9.3.1 Feeding Protocols problems.
A generalised feeding protocol for marine fish larvae
begins with rotifers at first feeding followed by brine 9.3.2 Some Disadvantages of Live Feed
shrimp nauplii and larger brine shrimp as larvae increase Organisms
in size (Figures 9.12 and 26.10). Formulated diets are A number of disadvantages are common to intensive cul-
then introduced and larvae are weaned from live food ture of microalgae, rotifers and brine shrimp. Some of
organisms. Fish hatcheries also culture microalgae as a these have been outlined for microalgae production in
food source for rotifers and brine shrimp and, as such, section 9.2.5, but they also apply to rotifer and brine
they generally culture three different live foods to feed shrimp production. Other potential problems relating
the larvae of a single target species. Excellent coverage specifically to rotifers and brine shrimp are listed in
of the practical use of live feeds and microdiets in Table 9.5. In response to these potential problems, par-
marine fish hatcheries is provided in Chapter 20. Shrimp ticularly the high costs associated with live food culture,
hatcheries generally begin feeding with microalgae there has been considerable research and commercial
(usually a diatom such as Chaetoceros species), which interest in developing compound or formulated hatchery
are usually followed by rotifers and brine shrimp or just feeds, sometime called microdiets, microfeeds or wean-
brine shrimp as the larvae grow. Bivalve hatcheries rely ing diets, as alternatives to live foods. Characteristics and
exclusively on cultured microalgae as a larval food potential of these feeds used with fish larvae was
source. reviewed by Kolkovski (2013).
(90–250 µm) (450 µm) 9.4 Compound Hatchery Feeds
Rotifers Artemia nauplii
9.4.1 Advantages
Weaning The high cost of live food production in aquaculture
(> 500 µm) hatcheries could be reduced by cheaper production of live
Artemia metanauplii foods and earlier weaning to formulated feeds in the case
of crustaceans and fish. Complete or significant replace-
Arti cial Diets ment of live foods would considerably reduce hatchery
running costs and provide ‘off‐the‐shelf’ convenience and
Larval Age nutritional consistency. Perhaps the greatest potential
advantage of appropriate compound larval diets is that,
Figure 9.12 A generalised feeding protocol for marine fish larvae unlike live foods, the size of the food particle and diet
begins with rotifers at first feeding followed by brine shrimp composition can be adjusted to suit the exact nutritional
nauplii and larger brine shrimp as larvae increase in size. Larvae
are then weaned to artificial formulated feeds. Source: Southgate
and Partridge 1998. Reproduced with permission from Elsevier.
Table 9.8 Desired characteristics of compound feeds for aquatic Hatchery and Larval Foods 197
larvae.
ingredients and the culture water, there is potential for
Characteristic Comments nutrient leaching and they are susceptible to direct bac-
terial attack.
Acceptability Must be attractive and readily ingested. Diet
particles must be of suitable size for ingestion 9.4.3 Microencapsulated Diets
Stability and must elicit a feeding response from the Microencapsulated diets (MED) consist of dietary mate-
Digestibility larvae. Diet particles must remain available in rials enclosed within a microcapsule wall or membrane.
Nutrient the water column This greatly reduces nutrient leaching and the suscepti-
composition Diet particles must maintain integrity in bility of the diet to bacterial attack. MED have been used
Storage aqueous suspension and nutrient leaching with some success as a replacement for microalgae for
must be minimal. Some nutrient leaching may bivalve larvae and juveniles (Knauer and Southgate,
be beneficial in enhancing diet attractability 1999). MED and other microdiets have been commer-
Diet particles must be digestible and their cially available for shrimp larvae for a number of years
nutrients easily assimilated and are widely used in shrimp hatcheries (section 22.6.3).
Diets must have an appropriate nutritional It is generally accepted that a combination of microdiet
composition. Material added to the diet as and live feeds, a practice known as co‐feeding, supports
binders or the components of microcapsule superior growth and survival of shrimp larvae than either
walls must have some nutritional value feed alone. However, it is now possible to completely
Diets must be suitable for long‐term (6–12 replace live feeds with microdiets in penaeid shrimp
months) storage with nutrient composition hatcheries. The development and use of formulated diets
and particle integrity remaining stable for crustacean larvae was reviewed by Holme et al.
(2009). Despite the development of successful artificial
requirements of the larvae. However, they must satisfy a diets for shrimp larvae and their routine use in shrimp
number of criteria (Table 9.8). hatcheries, similar success has not been achieved with
fish larvae.
Various materials have been assessed for their poten-
tial to replace live microalgae as a food for bivalves. 9.4.4 Microcoated Diets
These include dried and concentrated microalgae (sec- Microcoated diets (MCD) are primarily composed of
tion 9.2.5), dried and pulverised macroalgae, yeasts and MBD particles that are coated with lipids or lipoproteins
cereal products (Knauer and Southgate, 1999). They also to reduce nutrient leaching from the food particles.
include compound or formulated diets, known generi-
cally as microdiets, that encompass microbound diets, 9.4.5 Microextrusion Marumerisation Diets
microencapsulated diets, microcoated diets and diet Microextrusion marumerisation (MEM) diet particles
particles manufactured using microextrusion marumeri- are prepared by processes such as spray‐drying, fluidised
sation (Knauer and Southgate, 1999; Kolkovski, 2013). bed drying and particle‐assisted rotational agglomera-
Although dried microalgae have also been used in the tion, and a number of commercially‐available hatchery
culture of crustacean and fish larvae, much of the feed products are produced using these methods. These
research to develop alternatives to live feeds has focused processes can be used to manufacture diet particles cov-
on microdiets. ering a size range of 50‐1000 µm and containing a broad
range of nutrients.
9.4.2 Microbound Diets
In microbound diets (MBD), nutrients (both particulate 9.5 Development of Microdiets
and dissolved) are bound within a particle matrix con- for Fish Larvae
sisting of a binding material such as agar, gelatin, alginate
or carrageenan. Dietary ingredients are mixed with the 9.5.1 Limited Success
binder to form a slurry, which is then dried, ground and Numerous studies have been conducted to assess the
sieved to produce food particles of the desired size. MBD nutritional value of microdiets for marine fish larvae. In
allow precise manipulation of dietary contents and, for general, they have resulted in lower survival and poorer
this reason, have been used extensively in research to growth of larvae than those fed live foods and they often
determine nutritional requirements of larvae, particu-
larly crustacean larvae (e.g., Holme et al., 2009). However,
because MBD particles have no barrier between dietary
198 Aquaculture 9.5.3 Weaning Diets
Larvae reared on live feeds during the hatchery phase
lead to a higher incidence of deformity. These results require weaning to formulated feeds towards the end of
indicate that total replacement of live prey with artificial the larval period (Figure 9.12; section 20.3.1.5). The
diets is still not possible for the larvae of most marine weaning process usually involves providing live feeds
fish (Kolkovski, 2013). Despite this, partial replacement together with formulated food particles over a period
of live foods can result in cost savings, and some studies when the live feed component of the diet is gradually
have shown that between 50% and 80% of a live feed reduced and the formulated component is increased.
ration can be replaced with a microdiet without affecting The duration of the weaning process varies, but weaning
larval growth. Weaning fish larvae to a formulated diet at is usually completed within 30 days. A wide variety of
the earliest possible age is another means of reducing weaning diets are available commercially in the form of
feed costs and is a major goal for commercial fish hatch- MBD, MED, MEM, flake diets, crushed pellets (crum-
eries (section 20.3.1.5). It has been estimated that wean- bles) and yeast‐based diets. Development of successful
ing European bass 15 days earlier enables savings in brine formulated diets for fish larvae would eliminate the need
shrimp production of up to 80%. to wean larvae from live to formulated foods becaues lar-
vae could simply be fed larger food particles as they grow.
9.5.2 Constraints to Developing Microdiets
for Marine Fish Larvae 9.5.4 Use of Microdiets in Hatcheries
The relatively poor performance of formulated diets in Formulated microdiet particles are negatively buoyant
studies with marine fish larvae is thought to result and this presents problems maintaining food particles in
primarily from reduced rates of ingestion and poor suspension. This may reduce the availability of food to
digestion. the larvae, and food particles that settle at the bottom of
larval culture tanks may pollute culture water and
Successful formulated diets must be ingested at a enhance bacterial activity. In contrast, live foods generally
similar rate to live foods. This is a particular problem remain motile in larval culture tanks and this maximizes
with carnivorous fish larvae, which require the vis- their availability to larvae and reduces contamination of
ual stimulus of moving prey to initiate a prey capture the culture water from uneaten food. Tank design and
response. Attempts to overcome or reduce this problem aeration systems are important in maximizing microdiet
have included: particle buoyancy and for maintaining particle movement.
●● inclusion of various chemicals (using light refraction) The use of microdiets requires careful consideration of
tank design and aeration, and regular monitoring of feed-
into diet particles to impart a sense of motion; ing rates. Adding small quantities of food a number of
●● incorporation of food dyes into diets to simulate the times per day optimises water quality and maximises the
food available to the larvae.
colour of brine shrimp nauplii; and
●● use of amino acids that naturally emanate from live 9.5.5 Further Development of Formulated
Hatchery Feeds
food organisms to enhance larval feeding response. As described above, use of commercially produced micr-
They act as feed attractants and may be incorporated odiets is commonly practiced in shrimp hatcheries (sec-
into artificial diets to improve attractability. tion 22.6.3). However, the situation is not as good for
Most marine fish larvae are poorly developed at hatch, marine fish and bivalve hatcheries. Although both fish
and in many species the digestive tract does not develop and bivalve larvae have developed to the early juvenile
fully until they are early juveniles. Marine fish larvae stage when fed microdiets under laboratory conditions,
also have low gut enzyme activity compared to adult commercial fish and bivalve hatcheries still rely on live
fish and, again, secretion of some enzymes begins in food production for the majority, if not all of the larval
early juveniles when a functional stomach is present. culture period. This situation is now changing for bivalves
Marine fish larvae generally improve in their ability to where there is increasing hatchery use of commercially‐
digest artificial food particles with age. Live food available microalgae concentrates (section 9.2.5.1); how-
organisms consumed by larvae assist digestion by ever, development of formulated hatchery feeds or
donating their digestive enzymes either by autolysis or microdiets for fish and bivalves has been hindered by our
as zymogens, which activate endogenous digestive lack of knowledge of their nutritional requirements, and
enzymes within the larval gut. Inclusion of digestive
enzymes (particularly proteases) in artificial diets has
been shown to improve nutrient assimilation by up to
30%, resulting in superior larval growth. Similarly,
inclusion of digestive system neuropeptides in micro-
diets may also improve nutrient assimilation and
growth.
by problems relating to the attractability and digestion of Hatchery and Larval Foods 199
formulated food particles and their use in culture sys-
tems. Development of more suitable formulated feeds for 9.7.1 Fertilisers
marine fish larvae will require further research in the fol- Inorganic fertilisers are chemical fertilisers that contain
lowing areas: at least one of the primary nutrients nitrogen (N), phos-
●● improved ingestion and digestion of artificial diets; phorous (P) and potassium (K). Commercially‐available
●● greater understanding of the nutritional requirements agricultural fertilisers such as ammonium sulphate and
superphosphate are widely used in aquaculture. Animal
of larval stages; and manures are probably the commonest organic fertilisers
●● development of more appropriate culture system designs. used in aquaculture, although decomposed plant materi-
The potential cost savings offered by the use of suitable als are also widely used. Use of organic fertilisers in
formulated diets will ensure that research in this field is aquaculture is an ancient practice and is an economical
ongoing. means of increasing production in aquaculture ponds.
There is greater reliance on organic fertilisers in devel-
9.6 Harvesting Natural Plankton oping countries as they are more readily available than
chemical fertilisers. They are also more economical to
Natural sources of zooplankton represent a large, rela- use and more efficient if pond culture is integrated with
tively untapped, potential food source for aquaculture crop or animal production. In developing countries, ter-
hatcheries. A large portion of this plankton is composed restrial and aquatic animals (usually fish) are often
of copepods, which can occur naturally at densities up to reared together in integrated systems (section 2.4).
10 000 per cubic metre. Utilising this potential food Readers are directed to Chapter 4 (section 4.4.2) for
source requires efficient extraction of zooplankton from more information about the chemical fertlisers used in
very large volumes of water. This can be achieved by aquaculture.
pumping water through sieves that collect zooplankton.
Plankton harvesting machines have been developed that 9.7.2 Production in Fertilised Ponds
harvest and grade plankton by size. One of the draw- Fertilisation encourages primary productivity and pro-
backs of harvesting from natural waters is that plankton motes a succession of organisms within the pond.
densities may vary between locations and, as such, the Initially, fertilisation results in blooms of protozoa and
reliability of the food supply is questionable. However, bacteria, which are generally followed by blooms of algae
this may be overcome by harvesting from dedicated and then zooplankton. The natural food organisms pre-
plankton ponds in which high plankton loads can be sent in ponds consist of:
encouraged by fertilisation. The use of harvested natural ●● bacteria and protozoans
zooplankton as a food source for aquaculture is not ●● plants (phytoplankton, periphyton, macrophytes)
widespread; however, the variety of organisms compos- ●● animals (mainly invertebrates: zooplankton, zooben-
ing natural zooplankton would undoubtedly provide a
nutritionally superior diet to the standard rotifers/brine thos, small nekton)
shrimp diets used routinely in aquaculture hatcheries. ●● fish.
Fertilisation increases the biomass of potential food
9.7 Pond Fertilisation as a Food organisms present in a pond. For example, zooplankton
Source for Aquaculture levels of <0.055 g/m3 and 3.3424 g/m3 have been reported
in non‐manured and manured ponds, respectively.
Extensive and semi‐intensive pond culture of herbivo-
rous and omnivorous species is usually based on food Ecological conditions within a pond determine which
production through pond fertilisation (section 2.3). organisms are present, the proportion of each and their
Fertilisation of ponds for semi‐intensive culture of tila- abundance. All the listed organisms in the pond form the
pia, for example, is outlined in detail in section 18.7.3. biocenosis (self‐regulating ecological community) of the
Although this system is more commonly used for grow‐ pond, which therefore contains all potential food sources
out, pond fertilisation has also been used successfully for for cultured organisms. However, a given species will
larval fish culture (section 9.7.3 and Chapter 20). The only feed on a certain portion of the biocenosis and this
fertilisers used for this purpose may be inorganic or is dictated by its feeding habit (e.g., carp species;
organic in nature, or a combination of both. Figure 2.4). The specific portion of the biocenosis con-
sumed by an organism is its trophic basis, and the trophic
basis of a particular species may differ during its life
cycle, e.g., the larvae of many herbivorous fish consume
zooplankton.
200 Aquaculture densities of culture animals are higher than in equiva-
lent extensive systems that rely on natural foods alone.
The finite biomass of natural food in a pond can only In many developing countries, however, the use of sup-
support a finite standing crop of animals under culture. plementary feeds may not be feasible because it raises
When the standing crop is low, the amount of available the cost of production. These countries may also have
food exceeds the requirements of the culture population a limited capability to manufacture or import com-
and so each animal is able to find sufficient food to pound feeds.
support its energy requirement for maintenance and
maximum growth. However, an increase in the cul- 9.7.3 Pond Culture of Fish Larvae
ture population brings about an increase in their Freshwater and marine/estuarine species can be reared
food requirement. At a certain population density, the extensively in earthen ponds with blooms of natural
amount of food required by the culture population for plankton as their food source. The larvae of barramundi
maintenance and growth exceeds that available in the (Lates calcarifer) are a good example of larvae that may
pond. Since the maintenance requirement must be satis- be reared in this manner. This method has proved to be
fied, the amount of food energy that can be utilised for very successful, and achieves ca. 50% survival through
growth is reduced and growth rates decline as a result. larval rearing and growth rates greater than those
This standing crop is termed the critical standing crop achieved using intensive culture systems. Other examples
(CSC). A continued increase in the standing crop further of pond culture of fish are provided in Chapter 20 and
limits the amount of food that can be utilised for growth, further information on establishing copepod culture as a
and the point at which the natural food available in the basis for pond culture of marine fish larvae is provided in
pond is sufficient to support only the maintenance energy section 9.2.15.
requirement of the culture crop is known as the carrying
capacity of the pond. Fertilisation increases the CSC and 9.8 Summary
carrying capacity of a pond, and growth rates of the cul-
ture population can only be increased above these levels ●● Cultured live microalgae has a central role in aquacul-
if supplementary feed is added to the pond (Table 9.9). ture hatcheries where it is used as a food for all stages
of bivalve molluscs, for some crustacean and fish lar-
Supplementary feeds are generally classified into sim- vae, and for rotifers and Artemia that are themselves
ple feeds and compound (or compounded) feeds. Simple used as live foods for fish and crustacean larvae. Recent
feeds may be of animal origin (e.g., trash fish, slaughter- developments include potential for replacing live
house waste and fishmeal), or of plant origin (e.g., forage, microalgae with commercially‐available microalgae
oil meals, rice bran and sorghum). Many simple feeds are concentrates.
relatively cheap agricultural by‐products. The simple
feeds used by a given aquafarm are dictated by what is ●● Rotifers and Artemia are the major live foods used for
locally available. As would be expected, the nutritional crustacean and fish larvae in aquaculture hatcheries.
composition of simple feeds varies greatly. Those of plant Both are deficient in essential fatty acid content and
origin are usually rich in carbohydrate, whereas others, this must be corrected using enrichment procedures
such as oil meals and animal products, are rich in protein. prior to feeding to larvae.
Compound feeds consist of mixtures of ingredients that
are commonly bound together to form doughs or pellets. ●● Copepods have superior nutritional value to rotifers
and Artemia and are well suited as a food for fish lar-
An aquaculture system that utilises supplementary vae with small mouth gapes. Broader use of copepods
feeds is classed as semi‐intensive (section 2.2), and the as a live food in aquaculture hatcheries is hampered by
difficulties in intensive culture systems, that are char-
Table 9.9 The effect of fertilisation and supplemental feeding acterised by variable and unreliable production. It is
on critical standing crop and carrying capacity of freshwater likely that copepods will assume increasing impor-
ponds. tance as a hatchery food as more reliable mass‐culture
techniques are developed and more species are inves-
Treatment Critical standing Carrying capacity tigated for their culture potential.
crop (kg/ha) (kg/ha)
●● Potential problems with production and use of
No feeding, 65 130 live foods in aquaculture hatcheries (microalgae,
no fertilisation 140 480 rotifers, Artemia and copepods) include high cost,
No feeding, 550 2500 technical and infrastructure requirements, nutritional
fertilisation inconsistency and deficiency, and the potential for live
Feeding and
fertilisation
Source: Reproduced from Hepher (1988) with permission
from Cambridge University Press.
food to become vectors for contamiantion and disease Hatchery and Larval Foods 201
introdcution to larval cultures.
●● Potential alternatives to hatchery‐based culture of live lack of knowledge of their nutritional requirements
food organisms include dried and concentrated micro- and poor rates of ingestion and digestion of microdiet
algae, yeast‐based food particles and various formulated particles.
microdiets. Advances have been made with shrimp lar- ●● Live food organisms for larval culture can be harvested
vae that can be culture on microdiets without live foods, from natural water bodies or encouraged by fertilisa-
and with bivalve mollusc larvae that can be cultured tion of dedicated ponds and embayments. Extensive
using microalgae concentrates. However, culture of fish culture of copepods and other live food organisms in
larvae still relies on live foods. Development of ponds used to culture fish larvae is well established,
formulated microdiets for fish larvae is hampered by and can be very successful with careful management of
the plankton population and stocking density of the
target species.
R eferences development of alternative and artificial diets for bivalve
aquaculture. Reviews in Fisheries Science, 7, 241–280.
Arimoro, F. O., 2006. Culture of the freshwater rotifer, Kolkovski, S. 2013. Microdiets as alternatives to live feeds
Brachionus calicyflorus, and its application in fish for fish larvae in aquaculture: improving the efficiency
larviculture technology. African Journal of of feed particle utilisation. In: Allan, G. and Burnell, G.
Biotechnology. 5 (7): 536–541. (Eds). Advances in aquaculture hatchery technology.
Woodhead Publishing, Oxford. 203–245.
Brown, M. R., Jeffrey, S. W. & Garland, C. D. (1989). Lavens, P., Sorgeloos, P. (Editors) 1996. Manual on the
Nutritional Aspects of Microalgae Used in Mariculture: production and use of live food for Aquaculture. FAO
A Literature Review. CSIRO Marine Laboratories report Fisheries technical paper 361. FAO, Rome. 295 pp.
205. CSIRO, Hobart. Leger, P. (1999). The Artemia crisis…. and solutions, poor
yields at the Great Salt Lake. The Advocate, December
Brown, M.R., Blackburn, S.I. 2013. Live microalgae as feeds 1999, 79–82.
in aquaculture hatcheries. In: Allan, G. and Burnell, G. Lee, C.S., OBryen, P.J., Marcus, N.H. 2005. Copepods in
(Eds). Advances in aquaculture hatchery technology. Aquaculture. Blackwell Publishing, Oxford.
Woodhead Publishing, Oxford. 117–156. Parsons, T. R., Stephens, K. & Strickland, J. D. H. (1961).
On the chemical composition of eleven species of
Dhont, J. & Dierckens, K. 2013. Rotifers, Artemia and marine phytoplankters. Journal of the Fisheries
copepods as live feeds for fish larvae in aquaculture. In: Research Board of Canada, 18, 1001–1016.
Allan, G. & Burnell, G . (Eds), Advances in aquaculture Reed, T. & Henry, E. (2014). The case for concentrates.
hatchery technology, Woodhead Publishing, Oxford. Hatchery International, 15(4), 24–25.
pp. 157–202. Southgate, P. C. & Partridge, G. J. (1998). Development of
artificial diets for marine finfish larvae: problems and
Dhont, J. & Van Stappen, G. 2003. Biology, tank production prospects. In: Tropical Mariculture (Ed. by S. DeSilva),
and nutritional value of Artemia. In: Stottrup, J.G. & Academic Press, London. pp. 151–169.
McEvoy, L.A. (Eds), Live Feeds in Marine Aquaculture. Southgate, P.C., Beer, A.C. & Ngaluafe, P. (2016). Hatchery
Blackwell Science, Oxford. pp. 65–121. culture of the winged pearl oyster, Pteria penguin, without
living micro‐algae. Aquaculture, 451, 121–124.
Duy, N.D.Q., Francis, D.S., Southgate, P.C. 2017. The Utting, S. D. (1986). A preliminary study on growth of
nutritional value of live and concentrated micro‐algae Crassostrea gigas larvae and spat in relation to dietary
for early juveniles of sandfish, Holothuria scabra. protein. Aquaculture, 58, 123–138
Aquaculture, 473, 97–104. Whyte, J. N. C. (1987). Biochemical composition and
energy content of six species of Phytoplankton used in
Hepher, B. (1988). Nutrition of Pond Fishes. Cambridge mariculture of bivalves. Aquaculture, 60, 231–241.
University Press, Cambridge. Volkman, J. K., Jeffrey, S. W., Nichols, P. D., Rogers, G. I. &
Garland, C. D. (1989). Fatty acid and lipid composition
Holme, M., Zeng, C. and Southgate, P.C. 2009. A review of 10 species of microalgae used in mariculture.
of recent progress towards development of formulated Journal of Experimental Marine Biology and Ecology,
microbound diet for mud crab, Scylla serrata, larvae 128, 21940.
and their nutritional requirements. Aquaculture 286,
164–175.
Jeffrey, S. W., Leroi, J. M. & Brown, M. R. (1992).
Characteristics of microalgal species needed for
Australian mariculture. In: Proceedings of the
Aquaculture Nutrition Workshop (Ed. by G. L. Allan
& W. Dall), pp. 164–173. NSW Fisheries, Australia.
Knauer, J. & Southgate, P. C. (1999) A review of the
nutritional requirements of bivalves and the
203
10
Disease Principles
Leigh Owens
CHAPTER MENU 10.4 Generalised Disease Management Techniques, 206
10.1 Introduction to Disease, 203 10.5 Major Diseases, 208
1 0.2 General Principles of Infectious Diseases in 10.6 Summary, 216
Aquaculture, 203 References, 216
1 0.3 The Philosophy of Disease Control, 205
10.1 Introduction to Disease example, that the Asian shrimp industry lost at least
USD 20 billion over the last decade as a result of disease.
To put the impact of disease into perspective, it is inter- One of the most serious is white spot syndrome virus
esting to evaluate the future of the human race and its that typically results in an 80–100% loss of stock.
needs. It has been estimated that for mankind to main- Infections in 3907 hectares of shrimp ponds in the
tain its consumption of seafood at current levels, aqua- Mekong Delta, Vietnam in 2015 caused estimated losses
culture needs to produce over 80 million tonnes (t) by of more than USD 8 million.
2030 in order to maintain current per capita consump-
tion (Chapter 27). Thus, aquaculture will need to pro- Diseases include both infectious (see Table 10.1) and
duce an additional 30 million t of seafood in less than a non‐infectious (environmental, nutritional and genetic)
decade and a half. There is probably not enough land or problems. The non‐infectious diseases are solely due to
suitable marine areas for this to occur without massive management practices and are often limited to particular
disruptions to multiple ecosystems. However, about 40% farms. However, infectious diseases have the potential to
of all aquaculture production is lost to disease, as it is threaten whole industries and will therefore form the
broadly defined below. So, by simply removing or limiting basis of this chapter.
disease impacts, mankind could almost meet sea-
food requirements without changing any land utilising 10.2 General Principles of Infectious
practices. Diseases in Aquaculture
Disease can be defined as ‘any process that limits the 10.2.1 Interaction between Host, Pathogen
productivity of a system’ and is one of the most serious and Environment
factors in aquaculture (Kabata, 1985). Estimates of the The Sneizko three rings, Venn diagram of the interac-
economic cost of a disease outbreak are complicated by tions between host (the aquaculture species), pathogen
the complex interplay of numerous factors associated and environment is well known (Figure 10.1). It illus-
with a specific incident, which may range from direct trates the fact that, to occur, most infectious disease is a
production losses to socio‐economic impacts on liveli- three‐way interaction needing all components:
hoods and industries associated with the primary pro- ●● pathogen;
ducer. A recent review of economic costs attributable to ●● host;
a range of key parasite pathogens within the world’s ●● environment.
major marine and brackish water aquaculture industries
was presented by Shinn et al. (2015). It is estimated, for
Aquaculture: Farming Aquatic Animals and Plants, Third Edition. Edited by John S. Lucas, Paul C. Southgate and Craig S. Tucker.
© 2019 John Wiley & Sons Ltd. Published 2019 by John Wiley & Sons Ltd.
204 Aquaculture Epizootic Haematopoietic Necrosis Virus in redfin perch,
Table 10.1 Some major pathogens of aquaculture species. and crayfish plague (Aphanomyces astaci) in signal cray-
fish are two such examples of obligate pathogens produc-
Group Genera, etc. ing disease in the most pristine conditions.
Viruses
Bacilliform viruses, herpesvirus, iridovirus, For specific diseases of cultured species, the three‐ring
Bacteria nodavirus, rhabdovirus, coronavirus, model can be modified by changing the size of the
birnavirus rings to reflect the relative importance of the various
Fungi Rickettsiales, Aeromonas, Enterococcus, components.
Protozoa Flavobacterium, Flexibacter,
Pseudoalteromonas, Pseudomonas, 10.2.2 Density and Disease
Helminths Streptococcus, Vibrio The spread of pathogens is a density‐dependent process
Nematodes Aphanomyces, Branchiomyces, Lagenidium, and is therefore affected by stocking rates. There is a
Annelids Saprolegnia, Sirolipidium relationship such that the greater the density, the smaller
Crustaceans Amoebae: Neoparamoeba the distance between neighbours. This leads to a higher
Flagellates: Hexamita, Ichthyobodo likelihood of pathogens crossing the distance between
Gastropods Ciliates: Ichthyophthirius, Trichodina hosts in a viable state.
Sporozoans: Bonamia, Loma, Marteilia,
Perkinsus Immobile pathogens such as viruses, non‐motile bac-
Dactylogyrus teria, sporozoans and parasite eggs basically follow the
diffusion laws and, therefore, in still water conditions a
Polydora concentration gradient of the pathogens will form
Fish ‘lice’: Isopods: around an infected individual. Other pathogens such as
Fish ‘lice’: Branchiura: Argulus bacteria, fungal zoospores, protozoa and metazoans gen-
Copepods: Lernaea, Ergasilus, Mytilicola erally have active but variable dispersal capabilities. As
Crabs: Pinnotherids distance increases, fewer pathogens will be able to reach
Pyramidellids susceptible hosts to establish or continue a disease epi-
demic (outbreak). As there is natural attrition of patho-
Pathogen gens in the environment, if the pathogen does not reach
a susceptible host in a defined period of time, the chance
Obligate Probiotics of establishing a new infection is almost zero.
pathogens
Higher densities lead to genetic selection of mutant
Host Infectious Microbial pathogens that are virulent:
disease environment
1) In the natural environment, a newly‐mutated patho-
Non-infectious gen does not ‘know’ where the next susceptible host
disease will be encountered. In such a case, pathogens that
slowly release progeny are selected for because this
Physical will give a random‐moving infected host a chance to
environment spread the pathogen in the environment and to the
next host. That is, a pathogen does not ‘want’ to
Figure 10.1 A modified Sneizko three‐ring model depicting the destroy its current host, but rather allow it to function
interaction between host, pathogen and the environment. as close to normal as possible, thus increasing the
chance of an encounter with another susceptible host.
Various modifications of this model have been made to
illustrate specific points. Non‐infectious disease is an 2) Under culture conditions the next host is very close. A
interaction only between the host and environment. The mutant that uses all the available resources of a host to
area of overlap between pathogen and host represents produce pathogenic progeny, no matter what the con-
obligate pathogens: the most threatening group as they do sequences to the host, will be selected for (i.e., a viru-
not need environmental stress to cause clinical disease. lent organism). One of its large number of progeny is
the most likely to infect the next host as opposed to a
conservative, slow‐release adapted pathogen (1 above).
Once a virulent, resource‐utilising pathogen arises by
mutation, it is very quickly selected for in an aquacul-
ture environment. A single mutation in a nucleo-
tide that begins a codon for a single amino acid can
change virulence from non‐virulent to highly virulent.
Disease Principles 205
Disease etc. 1950s 1960s 1970s 1980s 1990s 2000s
Perkinsus
Haplosporidium
Thanatostrea
Gill Iridovirus
Marteilia
Bonamia
(a) Oysters
Vibriosis
Nocardiosis
Ichthyophoniasis
Pseudotuberculosis
Streptococcosis
Lymphocystis
Nodavirosis
(b) Yellowtail
Figure 10.2 Sequential appearance of microbial diseases of cultured animals. The dates are approximate. (a) Atlantic oysters
(Crassostrea virginica and Ostrea edulis), (b) Japanese yellowtail (Seriola quinqueradiatus).
This may contribute to the sequential rise of pathogen immediately after control is completed. Clearly, the erad-
problems that beset so many aquaculture industries ication philosophy is untenable in such areas.
(Figure 10.2).
There are a number of factors to consider when decid-
10.2.3 The Effect of Aquaculture on Life ing on control measures:
Cycles of Pathogens 1) The cost of the control measure. Some pathogens
By stocking facilities with monocultures, aquaculture
excludes both predators and competitors of the species make culture uneconomical in their presence and
being grown. A large number of the prey items of the they must be totally removed from the culture system
cultured species are also excluded. Exclusion of cohabit- (e.g., highly pathogenic organisms like crayfish plague).
ing animals results in removal of intermediate hosts and Others might be self‐limiting and only reduce stand-
definitive hosts from the aquaculture ecosystem. This ing stocks by a small percentage (e.g., peritrichous
effectively breaks the life cycle of many of the multi‐host ciliates on crustaceans). In this latter case, living with
helminths (e.g., digeneans and cestodes), which conse- the pathogen is more cost effective than trying to
quently have less of a role in disease in aquaculture than eradicate it. In most cases, however, there are no
in wild populations. Sea cages are much less effective at accurate cost estimates of losses due to a pathogen.
breaking these life cycles than ponds or recirculation Rather, a general and often very inaccurate ‘feel’ for
systems. the costs associated with a disease agent are the best
estimates available. However, this should not stop the
10.3 The Philosophy of Disease Control aquaculturalist from trying to economically evaluate
the cost of the control method and the downstream
Disease control in aquaculture is usually attempted on benefits. A classic example of the inaccuracy of the
the assumption that an absence of pathogens is the ‘feel’ of an impact of pathogens occurs in the freshwa-
desired state. However, the chance of beginning an aqua- ter crayfish industry of Australia. This industry has
culture venture without any potential pathogens in the three ‘orphan viruses’ (viruses that cause no overt
system is very slim and the question arises as to whether mortalities, so they are not considered important);
it is cost effective to achieve a pathogen‐free state. This Cherax bacilliform virus, Cherax giardiavirus and
‘total elimination of pathogens’ strategy is the classic Cherax reovirus. Typically, crayfish with these viruses
approach to disease control: the pathogenocentric grow to 35 g in 7 months – standard practice. However,
approach. This may not always be the best strategy. If, for once these viruses were removed by hatchery rearing
instance, the chance of reinfection from the local envi- of surface‐sterilised eggs, the average weight of
ronment is very high, the infection starts to re‐establish 6‐month‐old crayfish was 70 g!
2) The likelihood of reinfection. Ideally, there should be
almost no chance of the pathogen being re‐acquired
from the environment or from wild stocks in the vicin-
ity. Alternatively, infection with a pathogen and subse-
quent treatment will often allow the vertebrate immune
206 Aquaculture ●● live alternative hosts;
Table 10.2 Relationship between a diagnostic test and the real ●● frozen carcasses for human consumption;
presence of a pathogen. ●● aquaculture feeds; and
●● bait.
Result of test Pathogen present Pathogen absent
The majority of new introductions of pathogens to unin-
Positive A B fected systems are due to the unrestrained movement of
Negative C D contaminated animals. Sometimes this is unavoidable as
A + C B + D aquaculture does not exist without either broodstock or
live juveniles for stocking. However, the biosecurity of
Sensitivity (%) = A/(A + C) × 100. broodstock, the number one contaminator, has often
Specificity (%) = D/(B + D) × 100. been neglected and should be the first point considered.
Thus, in Europe it is mandatory to have broodstock
system to be primed and thus further infections are lim- tested for a broad range of notifiable diseases (bacterial
ited, e.g., white spot (Ichthyophthirius multifilis) on fish. and viral) before transfer is permitted.
Sometimes, however, the environment is supersaturated
with the pathogen from adjoining farms suffering the If pathogen‐free broodstock are not available, what is
same problem and unless the farm can be isolated, treat- the pathogen status of the broodstock that is being
ment may be almost useless (e.g., vibriosis in Southeast used? For example, in marine finfish aquaculture and
Asian shrimp farms). In sea cage situations, wild stock shrimp aquaculture, viral encephalopathy and retinopa-
will often congregate outside the cages where they can thy and white spot syndrome virus, respectively, are
easily recontaminate caged stock that have been treated. both spread vertically from broodstock to larvae and
Therefore, an understanding of the probability of rein- then distributed through infected postlarvae and juve-
fection is needed to correctly assess the control strategy. niles to farms.
Again, this information is usually lacking.
3) An adequate assay for the pathogen. It must be possible Whilst it is impossible to have strategies that will work
to accurately identify the pathogen to be able to assess for all pathogens, there are a number of procedures that
the effect of the control measures on the pathogen. In can help limit pathogens within culture systems.
the first instance this relies on an accurate diagnosis and
later on a sampling regime that will determine true posi- 10.4.1 Batch Culture
tives (sensitivity) and true negatives (specificity)
(Table 10.2). The sampling regime is constructed by Batch culture works on the ‘all in, all out’ principle.
assuming a certain prevalence which is the maximum Continuous culture eventually suffers from the early
allowable for that pathogen and the confidence level that batches acting as pathogen ‘factories’ which contami-
is acceptable to the farmer. If the test is insensitive, the nate the environment to levels where later batches
sample size or frequency of sampling must be increased cannot be raised (e.g., viral encephalopathy and retin-
to compensate for the low discriminatory ability of the opathy ‐ barramundi nodavirus). Furthermore, young
test. If the test is non‐specific, then animals will be animals are often susceptible to levels of a pathogen
assessed as infected when they are not, and a pathogen that only mildly affect older stages. This is due to a
control or treatment regime may be discounted when it naïve immune response in young animals and, having
had worked at the level required. The numbers needed fewer cells in a target organ, the animal is more com-
for accurate sampling of low prevalences are large and promised when cells lose their function due to a patho-
few commercial ventures willingly part with the neces- gen. Drying out and sterilisation of the culture system
sary numbers without monetary compensation or it and associated items between batches stops the ampli-
being mandatory compliance to legislation. fication in numbers of pathogens for later batches. This
is widely practiced in hatcheries, but it is often not used
10.4 Generalised Disease in grow‐out systems, where individuals from a number
Management Techniques of spawnings and of different ages may be mixed to
stock to an optimum density. However, batch culture
The most important factor for the movement and intro- during grow‐out (single year class practice) and
duction of pathogens to farms and, indeed on any geo- techniques to reduce pathogens between batches, such
graphical scale, is the movement of animals. This includes: as drying out ponds and leaving ponds or sea cage
●● live broodstock in particular; areas fallow, are employed by the marine shrimp and
●● live larval forms for stocking; salmon sea cage industries (see Chapters 22 and 17,
respectively).
10.4.2 Incoming Water Treatment Percentage of shrimp in Disease Principles 207
size class
Treatment of incoming water is essential in recirculating reaching the next host is reduced on an exponential
culture systems and more useful in hatcheries than grow‐ scale. On theoretical grounds, disease epidemics will fall
out situations due to the sheer volume of water involved to extinction unless a threshold number of hosts are pre-
in the latter. Water treatment includes chemical sterilisa- sent in a given area. Simplistically, each infected host
tion (chlorine, iodophores, ozone) and physical sterilisa- must infect at least two other hosts as it succumbs, or the
tion (UV light) (see Chapter 4). All are greatly enhanced epidemic will not propagate. Furthermore, lowering
by the inclusion of good settlement ponds preceding the stocking densities will also decrease the level of sibling‐
treatment. Particulate matter, producing high turbidity, interaction‐induced stress and competition for space
offers a substrate, nutrition and protection for pathogens, and food.
particularly bacteria, which prefer to be attached to sub-
strates. It has been shown that such settlement for nine 10.4.4 Single Spawning Stockings
days causes Vibrio bacteria to die out and be replaced by Differential growth is a good indicator of poor health in
oligotrophic, less pathogenic, species (section 11.3.6). a captive population. Runts are very useful to screen for
diseases as they are either stunted by pathogens, or
The bodies of each generation of bacteria are used as behaviourally and nutritionally stressed by being at the
nutrient for a progressively smaller biomass of bacteria, so bottom of a pecking order (Figure 10.3). Such stressed
that the numbers and biomass spiral downwards. This animals will also express pathogens. If a mixed spawning
phenomenon is called ‘self‐cure’. Settlement ponds reduce population is used to stock a culture system, the differen-
the particulate load of incoming water. Sterilisation is tial growth due to age, genetics or variations in hatching
most effective against obligate pathogens that cannot use conditions will obscure pathogen‐caused, differential
an alternative life cycle to build up their numbers (e.g., growth. Thus, stocking with a single spawning is of par-
viruses, rickettsia, chlamydia, sporozoans). However, aer- ticular benefit for an aquatic pathobiologist. This tech-
osols arising from vigorous aeration are very common in nique is not quite as useful in many finfish species where
hatcheries. They allow facultative pathogens to bypass the size grading is a normal part of culture (e.g., eels, salmon
sterilisation processes, build up to threshold numbers and and trout), but it works well for invertebrates (e.g., fresh-
then cause problems. For example, bacteria in shrimp water crayfish). This technique also highlights the prob-
hatcheries have been shown to travel 8 m as aerosols to lem of a very common practice amongst fish farmers. At
contaminate other tanks. harvest, most fish farmers will put the runts that are too
small to meet market needs into a pond to allow them to
Chlorine is very dangerous to use as a sterilising agent in grow to market size. This overlooks the most likely
organically‐polluted waters due to the formation of chlo- reasons for their failure to reach market size: they are
ramines. These are highly reactive, have a long half‐life compromised by having a disease. Therefore, in reality,
and are not neutralised by chemicals used to remove free the farmer is keeping a reservoir of diseased individuals
chlorine. Many farmers have neutralised free chlorine and on the farm to infect the next stocking.
subsequently watched in horror as their fry proceeded to
die from chloramine toxicity when the water was used. 50 Harvest weight
40
Ozone is a particularly useful sterilising chemical, 30
especially for recirculation where its application can be 20 runts
tied into reduction‐oxidation (redox) measuring devices. 10
When reduction potential is high, anaerobic bacteria
easily produce electrons from electron‐donating sub- 0
stances and electron acceptors are relatively rare. These 10 12 14 16 18 20 22 24 26 28 30 32
electrons are highly reactive and can cause cellular dam- Weight of shrimp (g)
age as a means of stabilising. At high oxidation potential,
electron acceptors are abundant and free electron dam- Figure 10.3 A typical proportion of runts in a population of
age is low. For aquaculturists, oxidative conditions are shrimp carrying a suite of diseases. Note the spread of sizes of
preferable, and ozone produces such conditions. Ozone the infected shrimp (■) across more size classes compared to
is also safe as it and its ozide derivatives have a very short uninfected shrimp (□).
half‐life and the final product, oxygen, is very useful.
10.4.3 Lower Stocking Density
By lowering the stocking density, the average ‘inter‐fish’
distance is increased and the probability of a pathogen
208 Aquaculture 10.4.7 Vaccination
Vaccination basically works on the premise that an immu-
10.4.5 Specific Pathogen‐Free Broodstock nological memory exists and that prior exposure to a
Most pathogens are more virulent to the younger stages pathogen will allow a stronger and quicker immune
of a host. By producing offspring from broodstock free of response. This technology is treated in detail in Chapter 12.
specific pathogens, the offspring have a good chance of
growing to a non‐susceptible size before being infected 10.5 Major Diseases
and thus a crop can be produced even in an area where
disease regularly affects animals. This can also work if all The following rendition gives a broad brush to com-
life stages are equally susceptible, but by late infection of monly‐occurring disease problems experienced in major
the host, the crop can be grown to harvest before the aquaculture industries. The diversity of infectious bio-
disease has a chance to establish. This is the approach logical agents in aquaculture is treated in detail in
taken for Infectious Hypodermal and Haematopoietic Chapter 11.
Necrosis Virus in marine shrimp (Litopenaeus van-
namei) culture. 10.5.1 Molluscs
10.5.1.1 Bivalves
Use of the words ‘specific pathogen‐free’ (SPF) is greatly Worldwide, protozoan parasites are the most significant
misunderstood by farmers and scientists alike who, in cause of losses to bivalve industries. This predominance
their minds, hear the words ‘free of all diseases’. The term of protozoan parasites is reflected in a guide to diseases
SPF should be followed by the pathogens that the host has for the mollusc farmer (Elston, 1990). Of the 11 ‘Notable
been tested for and found to be free of as part of the defi- Oyster Diseases’ described in this guide, seven are caused
nition to clear up misinterpretations. by protozoans:
●● Perkinsus marina;
10.4.6 Stress Reduction ●● Haplosporidium nelson;
Stress is often used as an excuse for problems when no ●● Haplosporidium costalis;
other logical explanation is available. Despite this nebu- ●● Bonamia mackini;
lous use of the concept of stress, it does have a real physi- ●● Bonamia ostrea;
ological basis and consequences (Pickering, 1981). ●● Marteilia refringens; and
Unfavourable conditions lead to an adaptive response ●● Hexamita nelson.
and a new level of homeostasis is achieved. If this is not Although this guide to diseases is based primarily on the
achieved, then exhaustion follows along with the over‐ experiences of the North American and European
production of the stress hormones (including, in fish, cultured bivalve industries, similar diseases affect bivalve
the corticosteroids cortisol, cortisone, corticosterone, industries around the world.
11‐deoxycortisol and adrenocorticodrophic hormones).
High plasma levels of corticosteroids: For example, the haplosporidian parasite Marteilia
●● induce lymphopaenia; sydneyi causes QX disease in Sydney rock oysters
●● reduce phagocytosis; (Saccostrea glomerata) in Australia. A freshwater flush-
●● reduce access of lymphocytes to inflammatory sites; ing of rivers usually brings on an outbreak of the diseases
●● deplete vitamin C reserves and consequent wound (epizootic) with deaths in about six weeks. The digestive
gland is attacked, first in the Leidig tissue and later the
repair; and epithelium. The gland becomes pale yellow and watery,
●● increase protein catabolism (gluconeogenesis) leading and gonadal condition may be greatly reduced. No con-
trol is known, but growers try not to have oysters on
to muscle wasting, low antibody and collagen synthe- their leases over the wet summer months. One interme-
sis, which again restricts wound repair. diate host appears to be a polychaete that scavenges the
Levels of the stress hormones do not correlate well with tissues of dead and dying oysters, although further inter-
levels of stress as some fish pass through the high levels mediate hosts cannot be excluded. Marteilia refringens
of secretion to a new, highly‐stressed state with no secre- causes similar problems in Ostrea edulis in France and
tion of the hormones. The two most practical ways of Spain, with infection occurring first in summer and most
limiting stress are to double the aeration, thus alleviating deaths in winter. At least one intermediate host, the
any oxygen stress that may be occurring, particularly copepod Paracartia grani, has been implicated. As the
during hot summers, and to lower stocking density as
mentioned above. In marine shrimp ponds, it has been
found that keeping the morning dissolved oxygen (DO)
>4.5 ppm is the single biggest factor that is able to limit
viral mortalities and keep survival at 85%.
parasite can be transmitted from oysters to P. grani, but Disease Principles 209
not vice versa, further hosts are likely, and at least five
species of copepod have been reported to be parasitised Iridoviral infections cause disease of the larval velum
by M. refringens. in Pacific oysters in western USA (OVVD), and gill
necrosis and haemocytic diseases in adult Crassostrea
The Bonamia group of intracellular parasites cause angulata and, lately, Pacific oysters in France. There has
problems worldwide by infecting both the cupped and been speculation that the same iridovirus is responsible
flat oysters (section 11.5.2). They are characterised by for all diseases and was imported into France from the
causing mortality in oysters 3 years or older when the USA with Pacific oysters.
oysters enter the female stage of their life cycle. Bonamia
roughleyi causes winter mortality in Sydney rock oysters Bacterial infections have from time to time caused mor-
and requires high salinity (30–35 ‰). Small yellow to tality in Tasmanian oyster hatcheries. Vibrio (13 strains),
brown pustules occur on the gills, palps and mantle. Alteromonas (10 strains), Pseudomonas (8 strains) and
Bonamia mackini occurs in North America and causes Flavobacterium (3 strains) were all involved. Five of the
similar problems in the Pacific oyster (Crassostrea gigas). Vibrio and two of the Alteromonas strains caused mortality
at 107 and 105 bacteria/mL, respectively, with the lower
Bonamia ostreae is a major problem in the cultured doses taking longer. All strains were lethal at 108 bacteria/
flat oysters of Western Europe. It appears that it was mL. Bacterial virulence for oysters has been strongly asso-
originally taken to France with shipments of oysters from ciated with a low molecular weight (500–1000 kDa), heat‐
western USA, similar to the route of the iridovirus, stable toxin. The toxin stops cilia movement (ciliostasis),
Oyster Velum Viral Disease (OVVD). Bonamia exitiosa which in turn reduces feeding and swimming in larvae, and
has also caused local problems in New Zealand bluff oys- inhibits the cleaning function of the gills of older oysters.
ters (Ostrea chiliensis), where it destroyed over 80% of
the industry over a six‐year period. In Australia in 1991, Some diseases affect the oyster’s shell and not the soft
the Victorian flat oyster industry also lost 90% of its pro- tissues. The shell may be much weakened structurally in
duction to Bonamia. Early this century, boats took live extreme cases, thus exposing the oyster to predation. In
bluff oysters from New Zealand to Hobart and then to lesser infections there is malformation of the shell, mak-
Melbourne, because the local fisheries had collapsed. ing the oyster less presentable or unacceptable to the
Speculation suggests that this translocation spread the market, which is as good as killing it from the viewpoint
Bonamia from New Zealand to Australia, but the ques- of the industry.
tion of what first caused the collapse of these oyster
industries is unclear. The occurrence of Bonamia in There is a shell disease caused by a fungus that grows
Western Australia suggests that the parasite is native to as filaments through the shell, weakening the shell and
Australia. causing dark raised ’warts’ on the inner surface of the
shell. It occurs in the European flat oyster, where it has
Bonamia first infects the tissues below the gut and caused heavy mortalities in some regions. Similarly, bor-
later invades the haemocytes that are involved in resorb- ing sponges, Cliona species, riddle the shells of some
ing unspent gonadal material, especially the ovaries. bivalves, including clams and pearl oysters, with very
Bonamia is a density‐dependent disease. Reducing the deleterious effects on marketability. However, Cliona is
stocking density and growing the oysters on hanging apparently not a problem with table oysters.
lines to keep them off the bottom should allow marketa-
ble crops to be grown in areas where Bonamia is endemic. Boring polychaete worms (Polydora species) invade
the shells of oysters, some other benthic bivalves (e.g.,
It is not just protozoans that cause disease in molluscs, scallops) and even abalone. They have a widespread dis-
however, viruses and bacteria are also involved. tribution, including Norway, France and Australia.
Polydora is responsible for the use of intertidal stick cul-
A herpes virus (131 nm diameter) has been associated ture of Sydney rock oysters in northern New South
with larval mortalities in Pacific oysters in northern New Wales. These ’mudworms’ bore through the shell, caus-
Zealand. Feeding ceased between days 3 and 4, and 60 to ing blisters on the inner surface. By culturing oysters in
the intertidal zone, the racks dry out during low tides
100% mortality occurred in days 7–11. The cells infected and newly‐settled polychaete larvae are killed. Polydora
were the fibroblasts and presumptive phagocyte precur- also affects abalone farms and benthic mussels, however,
sors, which displayed enlarged, marginated nuclei with rafted mussels are almost free of infection.
both intranuclear and cytoplasmic inclusion material.
Herpes viruses have been reported from oysters in the 10.5.1.2 Other Molluscs
There is a large family of ectoparasitic gastropods, the
north‐eastern USA, northern Wales and northern France, Pyramidellidae and some parasitise oysters and other
with the Ostreid herpes virus OsHV‐1 being one of the bivalves (Figure 10.4). Pyramidellids are small gastro-
more prevalent and damaging diseases of oysters in pods which suck the blood or other tissue fluids of the
host by using a long, penetrating proboscis. The host has
Europe. Predisposing conditions for all herpesvirus‐related
disease included elevated temperatures and crowding.
210 Aquaculture A related species, Perkinsus marinus, has caused kills of
table oyster in southeastern USA during summer. More
Figure 10.4 Pyramidellid parasite (Turbonilla sp.) on a juvenile significantly a herpes virus causing ganglioneuritis is also
giant clam (Tridacna gigas) during ocean nursery culture. Note the a major problem in abalone fisheries in Taiwan (Haliotis
pyramidellids’s long white proboscis that it is using to pierce and diversicolor supertexta) and in the state of Victoria
suck fluid from the mantle edge of the clam. Source: Reproduced (Haliotis laevigata and H. rubra) on the south coast of
with permission from John Lucas. Australia. The virus can cause in excess of 90% mortality
within two weeks, and is of major concern in wild stocks
less energy for growth in mild infections. In heavy infec- and fisheries as well as aquaculture. The major diseases of
tions, the host may die of tissue and fluid loss. The pyra- abalone are covered in detail in section 25.7.
midellid, Boonea impressa, is an important parasite of
the American oyster. There are several families of decapod crustaceans
which include species that live within the mantle cavities
Epizootics have been observed in cultured pearl indus- of large bivalves, such as large clams, mussels and oys-
tries. There were considerable long‐term mortalities in ters. These are species of the shrimp family Pontoniidae
black‐lip pearl oysters (Pinctada margaritifera) in French and the crab family Pinnotheridae (section 11.11.4).
Polynesia during the late 1980s, but no causative agent These crustaceans often live in pairs within the host.
was identified. In the silver‐lip pearl oyster (P. maxima) They usually occur on the gills, where they feed from the
industry in Australia, a bacterium, Vibrio harveyi, was host’s food grooves. They cause mechanical damage to
isolated from the haemolymph of dying oysters. Cold the host’s gill tissue. There is no evidence that these para-
water temperatures (19°C) apparently predisposed the sites cause mortality, but they have some minor adverse
pearl oysters to infection and crowded transportation effect on the host.
with very little water circulation allowed the bacteria to
establish. More recently rapid die-off of spat due to an 10.5.2 Crustaceans
unknown disease, likely to be of viral etiology has caused 10.5.2.1 Marine Shrimp
serious concern in the Australian pearl oyster industry. Viruses have caused hatchery mortalities and considera-
Of some interest, is the association of V. harveyi ble grow‐out problems in marine shrimp culture. The most
with mortalities of scallop spat in temperate, central devastating virus known to date is whitespot syndrome
New South Wales. virus (WSSV). It started in 1993 in China where it destroyed
almost USD1 billion worth of Fenneropenaeus chinensis.
A protozoan parasite, Perkinsus, has been associated When infected kuruma (Marsupenaeus japonicus)
with mortalities of pearl oysters and abalone (sec- prawns were imported from China and Korea to Japan,
tion 11.5.3). When water temperatures are elevated, the the virus was spread with devastating consequences.
cellular immune system of the abalone cannot encapsu- Further spread of WSSV to Taiwan, then to Thailand and
late and destroy the Perkinsus. Treatment consists of low- the rest of southeast and central Asia ensued. Later the
ering the water temperature by 8°C in a holding tank. This virus was moved to Texas, USA, presumably via frozen
will stop even a progressing disease outbreak in two days. commodity shrimp. Central and South America fol-
lowed, so by 1999 only the Philippines, Australia and
Oceania were known to be free of this virus. Unfortunately,
illegal movement of broodstock from the Indo‐Malaysian
archipelago to the Philippines infected that country. In
late 2016, there was an outbreak in southeast Queensland,
Australia, where the disease was detected in seven prawn
farms as well as wild prawns in the region. On a regional
scale, the spread of this virus was always with live animals
or frozen commodity shrimp. Interestingly, other rod‐
shaped viruses have been described from crabs, yet crabs
do not seem as to be as susceptible to the virus as shrimp.
The original outbreak of this virus coincided with the
upsurge in the practice of feeding raw crustaceans, par-
ticular mud crabs (Scylla species), to broodstock shrimp
as a maturation diet.
Shrimp stocks have become partially resistant to
WSSV so that carrier animals are common. Carriers can
be detected biochemically using nested polymerase Disease Principles 211
chain reactions. The most highly infected animals are
detected at the first stage of the reaction and are dis- Fenneropenaeus merguiensis. Control measures, devel-
carded. Should these infected progeny be stocked, the oped in Tahiti, involve separating floating eggs from the
risk of crop failure is 95%. Progeny of broodstock that are spawning female, surface sterilising them and then using
positive on the second stage of the reaction and those the phototactic response of the nauplii to separate the
that are negative (ideally) are used to stock shrimp ponds. larvae from the egg shells and moribund larvae.
The risk of crop failure from these shrimp is 31%. The
risk of crop failure is enhanced by stresses (osmotic, pH, Infectious hypodermal and haematopoietic necrosis
oxygen) associated with the wet season. Failure rate virus (IHHNV) has all but destroyed the aquaculture
might be around 19% during the dry season, but it may industry based on Litopenaeus stylirostris in the Americas.
leap to 70% with onset of the wet season. These same Interestingly, as L. stylirostris was not susceptible to Taura
crop failures, which are enhanced by the wet season, virus, IHHNV‐resistant strains made a temporary come-
occurred in Australia, where they were called Midcrop back as an alternative crop to L. vannamei before the virus
Mortality Syndrome (MCMS). Stresses from infections mutated and started killing L. stylirostris. Intranuclear
with other pathogens also greatly affect the outcome of inclusions were prominent throughout all tissues except
an infection with WSSV. hepatocytes. However, the inclusions from Australian
shrimp did not cross‐hybridise with an IHHNV gene
Taura syndrome virus has been responsible for wide- probe that was 90% specific for the American IHHNV
spread mortalities in Litopenaeus vannamei in the strain. A Philippine IHHNV strain did cross‐hybridise
Americas after initial outbreaks near the Taura River, with the American gene probe suggesting that the
Ecuador. Taura syndrome was initially believed to be due Australian IHHNV is a different strain. Nor did the gene
to the toxicity of fungicides used to control black sigatoka probe react with lymphoidal parvovirus that may be a vari-
disease on banana crops. The washing of the fungicides ant in expression of IHHNV. Similarly, an American‐
into the waterways allowed it to accumulate in the shrimp produced hepatopancreatic parvovirus (HPV) gene probe
ponds. The fungicides were very similar to crustacean based on virus from Korean Fenneropenaeus chinensis did
moult inhibitors and this is believed to be the mode of not cross‐react well with the Indonesian and Australian
action. A massive lawsuit against the European chemical shrimp strains nor with HPV from Malaysian freshwater
companies that produced the fungicides was launched prawns (Macrobrachium rosenbergii). HPV has not been
for compensation. Subsequently, cell‐free bioassays and found to be a problem in the aquaculture of Australian
electron microscopy unequivocally demonstrated the Penaeus monodon, even though it is common in wild
presence of a picornavirus that produced the characteris- stocks of Australian banana shrimp (Penaeus merguiensis)
tic hypodermal, buck‐shot lesions and mortalities. The and causes 28% losses in farmed P. merguiensis. However,
disease seems to be most aggressive in L. vannamei so recent information about Penaeus monodon in Thailand
that L. stylirostris had become an alternative crop in demonstrated HPV to cause severe stunting and therefore
infected areas. Unfortunately, survivors of the epizootic considerable loss of income to farmers.
are chronic carriers and infected L. vannamei have been
introduced to Taiwan, China and Indonesia. Spawner mortality virus (SMV) (a parvo‐like virus) in
conjunction with gill associated virus (ronivirus) has
Monodon‐type baculoviruses (MBV) can infect all spe- caused major losses in P. monodon broodstock and grow‐
cies of shrimp, although their ability to produce disease out. This Midcrop Mortality Syndrome (MCMS) has
differs between shrimp species. MBV has been impli- become a major grow‐out problem in northern Australia
cated as one of the causative agents of the mass mortali- with most farms losing 50% of their stock. Around A$44
million was estimated to have been lost over the years
ties that swept through Taiwanese shrimp farms in 1995–97. The disease outbreak is slow in its onset and
1987/88 and resulted in the crash of that industry. This shrimp grow to around 12–15 g before dying. Much time,
may not be correct as yellow head ronivirus (initially labour, feed and money are thus invested in a crop which
will be devastated if not harvested. A symptom of MCMS
called a baculovirus then a rhabdovirus in the literature) is the presence of sick, weak and reddened shrimp at the
(YHV), which was undiagnosed at the time, seems to edge of grow‐out ponds. The number of days the stock is
have had a very prominent role. All life stages of the host in ponds is a major risk factor with the probability of epi-
after mysis 1 are susceptible to MBV, but it is a disease of zootics increasing dramatically after 120 days. Partial
hatcheries rather than later stages, which are largely harvests to decrease stocking density allow some ani-
mals to be grown beyond 120 days.
asymptomatic when infected. In an Australian study,
MBV was found in 8 of a total of 13 monitored hatcheries Bacteria are still a major constraint to hatchery pro-
for P. monodon. However, it has been largely managed duction with very few batches of postlarvae being able to
be produced without antibiotics. Strategic use of antibi-
out of existence in P. monodon, but it has been a otics during larval and early post‐larval development
major problem with the establishment for aquaculture of
212 Aquaculture 1986/87 the crayfish industry in Turkey was also devas-
tated by the introduction of this fungus across the natural
(nauplii VI to protozoea I and again at mysis III to post- barrier of the Bosphorus Straits. This led to the introduc-
larvae I) is usual. The Vibrionaceae predominate in isola- tion into Europe of the signal crayfish (Pacifastacus leni-
tions from shrimp hatcheries with 8 of 37 strains usculus) from the USA; a freshwater crayfish that is
producing significant mortality in larval bioassays. highly resistant to the fungus.
Vibrio species, including V. harveyi and V. tubiashi, and
Photobacterium damselae are involved. On a worldwide WSSV has recently been identified in the Louisiana
basis, the non‐sucrose utilising bacteria (those produc- crayfish‐ranching industry infecting red swamp crayfish
ing green colonies on thiosulphate citrate bile salts (Procambarus clarkii). Whilst the route of introduction
sucrose agar, TCBS) are often involved in disease in crus- of WSSV is unknown and since crustaceans in the Gulf
taceans. Application of sucrose can alter the bacterial of Mexico waterways are not known to be infected with
balance favouring the less pathogenic bacteria and hence the virus, commodity shrimp are suspected.
reduce losses.
With the exception of the devastating crayfish ‘plague’
V. harveyi has been involved in massive mortalities in in Europe and WSSV in USA, freshwater crayfish are
Ecuador and major mortalities in two grow‐out farms in generally considered to be relatively disease free. This
Australia. In one study in Australia, over 50% of the bac- reflects the extensive culture methods used for crayfish,
teria at the completion of grow‐out were V. harveyi and whereby only very low stocking densities (5 /m2) are
V. alginolyticus. Strains of V. harveyi in Australia and the used. With the development of more intensive culture
Philippines have been shown to have very strong antibi- methods, disease will become more of a problem. A list
otic resistance with one virulent Australian strain having of pathogens already documented from farmed crayfish
a plasmid coding for four different antibiotic resistances. in Australia is presented in Table 10.3. The most impor-
V. penaecida has been associated with mortalities in tant agents causing mortalities so far are:
ponds in Japan (Marsupenaeus japonicus) and New
Caledonia (Litopenaeus stylirostris); and V. nigripulchri- ●● the rickettsia Coxiella cheraxi;
tudo has been associated with mortalities in ponds in ●● Cherax baculovirus/ bacteraemia; and
New Caledonia. ●● bacterial erosion of the eyeballs associated with
In the early years of shrimp culture, the fungi Aeromonas hydrophila and A. sobria.
Lagenidium callinectes and Sirolipidium were frequently
involved in causing larval mortality, especially in hatch- The rickettsia‐ and the baculovirus have been introduced
eries where there was excessive use of antibiotics. Lobster into at least Ecuador and the baculovirus into the USA
eggs have been shown to have a surface commensal with trans‐shipments of crayfish from Australia.
Pseudoalteromonas species, which protects against Emerging pathogens include microsporidiosis in Western
fungi. Antibiotics inhibit this commensal allowing the
fungi to become established. A Pseudoalteromonas spe- Table 10.3 Pathogens found in farmed Australian freshwater
cies has also been found associated with shrimp eggs and crayfish.
so the mechanism is presumed to be similar. Use of the
agent orange‐based herbicides has effectively controlled Type Pathogen
fungi in the hatchery. Viruses
Bacteria Cherax bacilliform virus, Cherax
While the fungus Fusarium solangi is present in grow‐ giardiavirus, parvovirus, reovirus,
out ponds in Australia, it is only seen when moulting fre- Microsporidia picornavirus
quency is slowed due to cool water temperatures or Fungi Coxiella cheraxi, Aeromonas hydrophila,
another disease problem. It is thus a good indicator of Temnocephalids A. sobria, Citrobacter freundii, Klebsiella
retarded growth. Similarly, the peritrich ciliate protozo- Ciliates pneumoniae, Plesiomonas shigelloides,
ans also indicate suboptimum conditions and, at their Pseudomonas, Shewenella putrifaciens,
highest fouling rates on the shrimp gills, can cause death Streptococcus
by suffocation if any further stress is added. Peritrich cili- Agmasoma (= Thelohania), Vavraia
ates need high organic loads to feed. So, by lowering the parastacida
amount of uneaten food with water changes or by Achlya, Mucor, Psorospermium
restricting supplemental feeding, the peritrich numbers Craspedella spenceri, Diceratocephala,
can be reduced within a week. Didymorchis, Notadactylus,
Temnocephala dendyi, T. minor
10.5.2.2 Freshwater Crayfish Corthunia, Epistylus, Lagenophrys,
The European noble crayfish (Astacus astacus) industry, Tetrahymena, Vorticella, Zoothamnium
which was a major aquaculture industry, has been deci-
mated by a fungus, Aphanomyces astaci. Furthermore, in
Australia and a systemic parvovirus in the redclaw cray- Disease Principles 213
fish, Cherax quadricarinatus.
birds. Therefore, they are very difficult to eradicate.
10.5.3 Finfish Primarily a disease of young freshwater salmonids, IPNV
now increasingly causes significant mortalities in marine
10.5.3.1 Carp farmed post‐smolts, with outbreaks usually occurring
As outlined in Chapter 16, the carp and their relatives are within 8 weeks of seawater transfer. Control in post‐
the largest group of finfish or shellfish produced from smolts by early vaccination has had some effect, but con-
aquaculture. They made up around 30% of global finfish trol in young freshwater fish by this method is impractical.
and shellfish production in 2015, and two‐thirds of all Such is the economic significance of this disease that the
finfish and shellfish from freshwater. The Disease section first genetically selected lines of Atlantic salmon with
in Chapter 16 (section 16.6) refers the reader to a mono- genetic markers for resistance to IPNV are available
graphic treatment of diseases in carps and their relatives commercially in Norway.
by Hoole et al. (2001).
Infectious haematopoietic necrosis virus is one of the
10.5.3.2 Salmonids most studied as it infects a range of salmon species includ-
The finfish group that dominates the aquaculture litera- ing Atlantic salmon, rainbow trout, sockeye salmon, chi-
ture on a worldwide basis is the salmonids because, nook salmon and coho salmon. While this rhabdovirus is
unlike carp, they are cultured in so many countries endemic to the north‐eastern Pacific it has been spread
around the world. Over 20 viral diseases are recognised across the northern Pacific as far south as China and
in salmonids and approximately half are considered to Taiwan, eastern USA, France and Italy. It primarily affects
have moderate to high virulence. fish less than 6 months old and, as it is found in the sexual
secretions of both male and female broodstock, it has
Infectious salmon anaemia virus, ISAV, is currently the ready access to young fish for infection.
most economically damaging disease in Atlantic salmon
aquaculture worldwide and is the first of the diseases Viral haemorrhagic septicaemia (VHS) has a broad
classed on the List One of the European Commission’s host and geographic range and multiple strains of this
fish health control regimen. List One diseases require virus have been described (see section 11.2.6). It only
eradication of all infected stock as part of the con- grows at temperatures below 16°C so its geographic
trol measures. ISAV, a member of the family range is limited and losses can be ameliorated by increas-
Orthomyxoviridae, is a single stranded RNA virus that ing temperature. Recently, VHS has been found in wild
infects blood cells causing anaemia. Symptoms include Southern Californian pilchards and mackerel that are
pale gills, liver and spleen inflammation and necrosis, used extensively as bait and fodder for other aquaculture
and circulatory failure as the system becomes ‘clogged’ industries such as bluefin tuna.
with dead cells. Initially discovered in Norway during
the 1980s, ISAV is now found in Scotland, Canada, the Bacteria are major pathogens of cultured salmonids.
Faroes, and is devastating the Chilean aquaculture Antibiotics have been widely used to control outbreaks,
industry (for further information see section 11.2.5). but they are becoming less and less acceptable in food
animals due to concerns about antibiotic resistance
Pancreas disease (PD) caused by the salmonid alphavi- spreading to pathogenic bacteria of humans. Bacteria in
rus (SAV) was first discovered in Scotland in 1976. the sediments of abandoned aquaculture sites have been
Initially associated with poor performance or ‘runts’, assayed for antibiotic resistance 10 years after the culture
attributed to loss of centroacinar cells in the pancreas operation ceased and they were found to still have major
rather than overt mortality, the last decade has seen a resistance against oxytetracycline. This type of problem
resurgence of a more acute form of the disease. There are has led to the production of effective vaccines. The 30%
at least nine subtypes of SAV that can cause mortality. rise in Atlantic salmon production in Norway is due to
With significant losses in Norway, Scotland and Ireland, control of furunculosis caused by Aeromonas salmoni-
the Chilean government moved to block the import of cida subsp. salmonicida, and winter disease caused by
salmonid eggs in November 2009 to prevent introduction Vibrio salmonicida. In fact, A. salmonicida is probably
of the disease. the most important finfish pathogen in all marine aqua-
culture industries (see section 11.3.1). Whilst many of
The Aquabirnavirus, e.g., infectious pancreatic necro- the original causes of generalised septicaemias in salmo-
sis virus (IPNV), and similar viruses are found world- nids have been successfully controlled through vaccina-
wide (see section 11.2.2). These viruses have a great tion, emerging infections with intracellular bacteria are
propensity for replicating or surviving in hosts other proving more difficult to control. Bacterial Kidney
than fish. These other hosts include bivalves, crayfish, Disease (BKD) caused by Renibacterium salmoninarum
marine shrimp, leeches and the intestines of fish‐eating has been an intracellular pathogen of note throughout
salmonid aquaculture for decades (see section 11.3.8).
In the last 15 years, rickettsia‐like infections caused
214 Aquaculture that was attributed to Neoparamoeba species. The amoe-
bae appeared on marine farms and were quickly eradi-
predominantly by Piscirickettsia salmonis have also cated from the fish gills by exposing them to freshwater,
become prevalent in major Atlantic salmon farming e.g., towing the fish cages from seawater into freshwater
nations (see section 11.3.5). Most recently, significant or by treating the fish in freshwater baths. However, the
mortalities associated with members of the bacteria Neoparamoeba are developing resistance to the freshwa-
genus Francisella have occurred in salmonid aquaculture ter treatment and it is only now effective for about
in Norway, Canada and Chile. Whilst the fish isolates 30 days. A parasitic amoeba, Neoparameoba perurans
(F. noatunensis and F. asiatica) do not appear to be caused problems at the outset of the salmonid sea cage
pathogenic to humans, interest has been high as the industry in Tasmania, Australia. Fish in their first year in
closely‐related F. tularensis almost invariably causes seawater develop a proliferative gill disease especially if
fatal diseases in humans. sea cages are fouled or water temperature is above 15°C.
Amoebic gill disease (AGD) may kill up to 2% of fish per
Septicaemia caused by Lactococcus garvieae is a major day if untreated and represents the major disease prob-
problem in rainbow trout in Europe, Asia, South Africa lem of Tasmanian farmed salmon. Chemical baths are
and southern Australia. These pathogens appear to largely ineffective but freshwater baths of a few hours, or
evade host macrophages as part of their colonisation moving the whole sea cage to freshwater, is reasonably
strategy. L. garvieae is encapsulated (coated with a poly- effective. Once the cycle is broken, the immune system
saccharide) reducing the ability of the lytic complement may prevent further heavy infestations. Unfortunately,
pathway to lyse the cells and impeding phagocytosis. there is strong evidence that the freshwater treatment is
Hence, macrophage engulfment is compromised, and becoming less effective. Instead of a one‐off bath, bath-
host antibody production is slowed. ing frequency has been increased to every few weeks
during the warmer weather, incurring enormous cost to
Of the parasitic infections of farmed salmonids, per- the industry. Research into vaccination against this para-
haps the two most significant are sea lice (see sec- site is underway.
tion 11.11.3) and proliferative kidney disease (PKD) (see
section 11.6). Infestations by calagid copepods of the The salmonid aquaculture industry in Australia merits
genera Lepeophtheirus and Caligus are probably the some attention as these animals are farmed at the extreme
most prevalent pathological and environmental con- upper limit of their temperature tolerance. Most culinary
cerns facing Atlantic salmon farmers in the dominant experts prefer rainbow trout to Atlantic salmon and
producing countries. Treatment is generally via chemi- accordingly the Australian sea cage industry started with
cals. Treatment by bathing in organophosphates was rainbow trout. Enterococcus (previously Streptococcus)
phased out in the early 1990s with pyrethroids becom- biovar I killed 30% of rainbow trout in freshwater and as
ing licensed as replacements. Latterly, treatment has high as 60% in marine waters associated with the accli-
been via feed with emamectin benzoate, although matisation stress. Therefore, the industry swapped en
resistance is emerging and biological solutions are being masse to Atlantic salmon, which is highly resistant to this
sought. The causative agent of PKD was first identified bacterium.
as a myxozoan parasite, Tetracapsuloides bryosalmo-
nae, and is probably the most significant parasitic Yersinia ruckeri serovar III (the Australian strain)
disease of salmonids (particularly trout) in Europe and
North America causing up to 90% mortality in infected causes some problems mainly in young Atlantic salmon
stock. Severe wild fish kills have also been reported. particularly when water temperatures are above 15°C.
Treatment with malachite green was favoured until it its Some fish may develop the pathogonomic ‘red mouth’
use was banned. The fungicide fumagillin has been tri- due to haemorrhagic lesions and in chronic cases, bilat-
alled in both the USA and UK with some positive indi- eral exophthalmia and anorexia may predominate. The
cations. Survivors of infection tend to be resistant to bacteria can be spread in the faeces of carrion feeding
reinfection thus control by vaccination may be possible
in the future. raptors. Vibrio anguillarum serovar C or 01 has been
isolated from Tasmanian rainbow trout and Atlantic
Kent and Margolis (1995) outlined the diseases caused salmon. Clinical signs include hyperaemic prolapsed
in seawater‐reared salmonids by parasitic protozoans.
There are numerous and diverse protozoans that para- rectum, congestion at the bases of the fins and reddened
sitise aquatic vertebrates in freshwater and marine habi- ulcers on the flanks. The muscles may contain enclosed
tats and these cause various levels of debilitation (see lesions and severe peritonitis can occur. Overseas, viru-
section 11.5). For example, the amoeboid protozoan, lence in V. anguillarum has been linked to a mostly plas-
Neoparamoeba species, is an opportunistic pathogen
that infects the gills of fish under some conditions (see mid‐encoded siderophore (iron sequestering) system
section 11.5.2). On a coho salmon farm in Washington with the most virulent strain known having four separate
State, Kent and Margolis (1995) observed 25% mortality siderophores. Both of these conditions can be controlled
by efficient vaccines.
The typical, non‐motile strain of Aeromonas salmoni- Disease Principles 215
cida (subsp. salmonicida) usually forms large boils or
furuncles in salmonids. This obligate pathogen exists in widespread malnutrition, starvation and socioeconomic
Victoria as the atypical, non‐salmonid infecting, motile disruption of subsistence lifestyles. After many compet-
strain in goldfish Carassius auratus. The bacterium ing hypotheses on the aetiological agent including
seems to have been imported to Victoria from Japan in r habdoviruses and Aeromonas hydrophila bacteria, A.
1974. Despite there being a restriction on the movement invadans was identified by a group of researchers in
of goldfish from Victoria to Western Australia and Australia. In the fullness of time, EUS has been traced
Tasmania, there was an isolation of a typical strain of back to early outbreaks in the 1970s in Australia and
A. salmonicida from flounder in Tasmania. Similarly, a Papua New Guinea. The earliest outbreak was in Japan in
silver perch farm in New South Wales was quarantined 1970. One interesting outbreak occurred only in the fish-
due to this bacterium. pond at the Sri Lankan international airport, where pre-
sumably a passenger had thrown some fish scraps from a
Since the eradication of infectious haematopoietic meal into the pond. Lately, EUS has become a problem in
necrosis virus during the initial outbreak, only three the establishment of the jade perch (Scortum barcoo) as
viruses have been found in Australian salmon. The chum an aquaculture species.
salmon Aquareovirus was isolated from ovarian fluid of
Atlantic salmon and it apparently only causes disease in Barramundi (Lates calcarifer) is cultured throughout
fry of chum and chinook salmon. This virus has more tropical Asia and is the only fish seriously cultured in the
significance as a trade barrier for exclusion of competing tropics of Australia. Whilst most farms have intermittent
Australian products by importing countries than as a problems, the most common pathogens are Cytophaga
disease threat. Infectious Pancreatic Necrosis virus was johnsonae causing saddle back, which may lead to dirty
isolated from sea‐caged Atlantic salmon displaying yellow ulcers and white spot from Cryptocaryon irritans
pinhead. in sea cages (see section 11.5.6; Figure 10.5). Problems
are associated with the onset of cooler temperatures,
10.5.3.3 Tropical and Subtropical Finfish usually in May. Streptococcus iniae is emerging as the
Amongst the farmed fish that have shown substantial most important disease in barramundi in estuarine sea-
increases in production in recent years in tropical and cage operations, freshwater ponds and in recirculation
subtropical freshwater are tiliapia and catfish species. systems (see section 11.3.9). The initial episode starts
The diseases of tilapia and their management are out- after displacement of soil in the watershed where the
lined in section 18.9. The diseases of channel catfish, barramundi are grown. Losses approach 30% and are not
Ictalurus punctatus, in the southern USA are outlined in ameliorated by winter water temperatures. Also, Vibrio
section 19.3.4. The diseases of pangasids largely mirror harveyi is emerging as major enteric disease in barra-
those found in ictalurids including enteric scepticaemias mundi hatcheries.
caused by Edwardsiella ictaluri and motile Aeromonads,
particularly of the A. hydrophila group. In Vietnam the Politically, there have been major problems associated
pangasid hatchery industry is becoming more advanced. with nodaviruses which cause viral encephalopathy and
Control of infection is largely by antibiotics, but resistant retinopathy of larval barramundi, and kill young,
in both these diseases is not uncommon. Vaccination crowded larvae, especially in later batches through a
with live attenuated E. ictaluri has proved effective in the hatchery. The growing of barramundi outside their
USA, but introduction of a live agent is generally poorly
accepted by farmers due to concerns over reversion to Figure. 10.5 Barramundi (Lates calcarifer) infected with ‘white
virulence under farm conditions. The A. hydrophila spot’ (Cryptocaryon irritans). Source: Reproduced with permission
group of organisms present a particularly intriguing from Professor Bob Lester.
challenge for vaccination due to their high intraspecific
diversity and range of different species, subspecies and
subgroups that cause disease. With demand of upwards
of 100 million doses of effective vaccine in Vietnam
alone, research into these diseases is intense.
Epizootic Ulcerative Syndrome (EUS) is caused by the
fungus Aphanamyces invadans. EUS came into promi-
nence in the 1980s as a devastating pathogen of snake-
head and other fish of southeastern and central Asia.
Snakehead fish are used for protein supplementation in
the diets of the rural poor. Destruction of these fish led to
216 Aquaculture rity appears to be most promising means of control in
the future. Bacterial diseases of warm water marine fin-
natural range in the Murray‐Darling watershed, south- fish include Photobacterium damselae subsp. piscicida,
eastern Australia, and the potential impact of the noda- vibriosis caused by a number of different Vibrio species,
virus on the endemic freshwater fish have been hotly and streptococcosis caused by Streptococcis iniae and
debated. A similar, if not identical virus, has been found S. agalactiae. As the major proportion of aquaculture
in barramundi in Tahiti and Singapore, and in most fish developments is in warm water, it is here that new
species (about 30 species at present) around the world. diseases are most likely to arise as significant problems
Almost all fish raised in marine hatcheries end up being for the future.
infected with this virus, once the correct tools for inves-
tigation have been used. 10.6 Summary
Other pathogens seen from time to time include lym- ●● The nature of aquaculture and the best anti‐disease
phocystis virus associated with fouled sea cages, vibriosis, technologies are the direct nemeses of each other.
epitheliocystis, and Epieimeria. Experimental infections
with Bohle iridovirus, a systemic Ranavirus have shown ●● Aquaculture is about maximising profit by growing as
barramundi to be acutely susceptible in both fresh and many animals as possible in the smallest possible vol-
marine waters. Total mortalities usually occur within ume of water, whereas anti‐disease methodologies
10 days. include keeping culture densities as low as possible.
Other warm water species of note include the various ●● As profit is the major goal in aquaculture and culture
species of grouper farmed throughout Asia. Nodaviruses densities maximised because of this, disease problems
are the major issue in larvae, fry and juvenile fish with are amplified.
broodstock screening and hatchery biosecurity the
major means of control currently. Although experimen- ●● Because of the above, aquatic pathobiologists will always
tal vaccines appear to hold promise in larger fish be in demand and the challenges faced make a very
(Chapter 12), protection of fry is more problematic. interesting and rewarding career path for students.
A combination of vaccination of larger juveniles and
broodstock, then carefully managed hatchery biosecu-
R eferences Kent, M.L., Margolis, L. 1995. Parasitic protozoa of seawater‐
reared salmonids. Aquaculture Magazine, 21, 64–74.
Elston, R.A. 1990. Mollusc Diseases: Guide for the Shellfish
Farmer. Washington Sea Grant Program, University of Pickering, A.D. (Ed.) 1981. Stress and Fish. Academic
Washington Press, Seattle. Press, London.
Hoole, D., Bucke, D., Burgess, P., Wellby, I. 2001. Diseases Shinn, A.P., Pratoomyot, J., Bron, J.E., Paladini, D., Brooker,
of Carp and Other Cyprinid Fishes. Fishing News Books, E.E., Brooker, A.J. 2015. Economic costs of protistan and
Oxford. metazoan parasites to global mariculture. Parasitology,
142, 196–270.
Kabata, Z. 1985. Parasites and Diseases of Fish Cultured in
the Tropics. Taylor and Francis, London.
217
11
Pathogens and Parasites
Kate S. Hutson and Kenneth D. Cain
CHAPTER MENU 11.9 Acanthocephalans, 240
11.1 Introduction, 217 11.10 Leeches, 240
11.2 Viruses, 218 11.11 Crustaceans, 241
11.3 Bacteria, 222 11.12 Fishborne Zoonotic Agents and Aquaculture, 244
11.4 Fungi, 229 11.13 Aquaponics, 245
11.5 Protozoans, 230 11.14 Summary, 246
11.6 Myxozoans, 234
11.7 Platyhelminths, 235 References, 246
11.8 Nematodes, 239
11.1 Introduction (e.g., vaccination) or reactionary methods (i.e., treat-
ments) are needed to minimise the effects of specific
There is an enormous diversity of pathogens and para- pathogens.
sites that can infect aquatic organisms. Pathogens, as
described here are infectious organisms (viruses, bacte- This chapter provides an introduction to the diversity
ria, and fungi) that cause disease and will harm the host of infectious biological agents commonly encountered
or cell. Parasites may be considered pathogens if they when farming aquatic animals and highlights specific
cause disease in a host species, but a parasite may live and broader impacts of each group. Specific case studies
on or within a host for all or part of its life but not cause are developed for particularly harmful pathogens and
clinical disease. Conditions associated with intensive parasites in aquatic fish and invertebrates. Current
monoculture of aquatic animals mean that only a lim- approaches to reducing infection intensities are outlined
ited diversity of disease causing agents can successfully for each group and potential human diseases associated
propagate, proliferate and harm aquaculture stock. Host with the production of aquatic animals are also consid-
specificity is arguably the most important property of a ered. Plants grown in aquaponics systems are subject to
pathogen or parasite because it can determine whether many of the same pests and diseases that affect food
it has the potential to become established in aquacul- crops which have been well documented in other
ture, although there is potential for the emergence of sources, thus strategies to disease management are high-
new pathogens and parasitic diseases or for free‐living lighted herein. The World Organisation for Animal
organisms to switch to a parasitic life style. Aquaculture Health (OIE) provides a current list of notifiable diseases
systems and biosecurity practices impact pathogen and (diseases that are required by law to be reported to gov-
parasite diversity and virulence by either promoting ernment authorities) for molluscs, crustaceans and fish
exponential growth or possibly eliminating conditions on their website and there are considerable economic
necessary for some species to survive. Disease manage- costs attributable to a range of key parasites and patho-
ment strategies are most effective if host contact with gens in the world’s major marine and brackish water
pathogens can be avoided. However, in many aquacul- aquaculture production industries (e.g., Shinn et al.,
ture systems fish are naturally exposed to endemic path- 2015). There are several text books dedicated specifically
ogens and other strategies aimed at preventing disease to aquatic animal diseases and disorders where further
information can be sourced.
Aquaculture: Farming Aquatic Animals and Plants, Third Edition. Edited by John S. Lucas, Paul C. Southgate and Craig S. Tucker.
© 2019 John Wiley & Sons Ltd. Published 2019 by John Wiley & Sons Ltd.
218 Aquaculture There have been hundreds of different viruses isolated
from infected fish, but only a small percentage cause sig-
11.2 Viruses nificant impact to aquaculture or are considered to be of
regulatory significance and notifiable disease agents as
Viruses are small particles that are infectious and require designated by the OIE. As both freshwater and marine
a host cell to replicate. Virus particles are referred to as aquaculture expands globally, there are greater opportuni-
virions and are made up of either DNA or RNA sur- ties for the transmission and spread of aquatic viruses, and
rounded by a protein coat or capsid that provides protec- viral diseases will continue to cause substantial impact on
tion for this genetic material. In some cases, viruses may aquaculture production. New and emerging viral patho-
be enveloped; meaning the virion has a lipid envelope gens impacting aquaculture are continually being discov-
usually acquired from the host cell membrane as it repli- ered. One example is the recent discovery of tilapia lake
cates and buds from the cell to form a new virus particle. virus (TiLV) found to be the cause of significant mortality
Virus shapes can be complex and range from simple heli- in tilapia farms in Israel and Ecuador. In this chapter, we
cal forms to icosahedral shapes. Most viruses can only be present a range of viral groups and specific viral agents that
observed using electron microscopy. If special stains and affect cultured as well as wild fish. Shrimp and prawn cul-
markers are used, it is possible to visualise virally infected ture have also been heavily impacted by viral diseases (see
cells using light microscopy, but virus particles them- sections 10.5.2.1 and 22.10). For example, white spot syn-
selves are too small for this. To diagnose or detect viruses, drome virus (WSSV) has decimated shrimp production in
they must be detected in tissue culture by infecting cell many regions due to its rapid spread and heavy mortality.
types (cell lines) they are able to replicate in. These cell It is important to note that not all viral groups and families
lines are grown in the laboratory in nutrient media and are represented here and only a fraction of the many viral
when a sample is applied that contains a virus, the diag- pathogens that impact aquaculture is described in this
nostician will observe cell destruction and lysis under chapter. There are many books and reviews available that
the microscope. This is usually referred to as cytopathic provide in depth coverage of these and other important
effect (CPE) of the virus. Viruses can infect many organ- viral pathogens of fish, crustaceans and molluscs.
isms and a range of cell lines have been developed from
plants, insects, mammals, and fish. Some viruses (i.e., 11.2.1 Betanodaviruses
bacteriophages) can even infect bacterial cells and may
have specificity to particular types of bacteria. The genus Betanodavirus is in the family Nodaviridae.
Viruses in this genus are linked to disease outbreaks in
In aquaculture, viruses can spread through specific vec- wild and cultured fish species, while the other genus in this
tors (e.g., blood‐sucking parasites), via the faecal–oral family Alphanodavirus causes disease in insects. Many
route, physical contact and/or entering the body from different species of marine and freshwater fish are suscep-
food or water. Viral infections in animals may elicit an tible to infection with Betanodaviruses and the disease is
immune response that can eliminate the infecting virus. most commonly referred to as Viral Nervous Necrosis
However, some viruses are effective at evading the host’s (VNN), but it is now recognised and has been renamed
immune system and may even utilise part of the host cell viral encephalopathy and retinopathy (VER) by OIE. This
to form a viral envelope as they replicate and bud from a VER virus has been isolated from fish on all continents
cell. This makes it difficult for the host to recognise the except South America; however, there has been one report
virus particle as foreign and mount an immune response on the identification of nodavirus positive samples from
to neutralise it. Vaccines have been developed for a few the brains of two freshwater aquarium species imported to
select viral diseases that impact aquaculture, but their use South Korea from the Amazon. Nodavirus infections
is not currently widespread. Those that are available typi- appear to be highly prevalent in areas where marine fish
cally consist of a non‐virulent or killed virus, or some com- culture is widespread and have been linked to severe dis-
ponent of the virus of interest (e.g., protein or DNA) that is ease outbreaks primarily in larval and juvenile marine as
delivered to the animal by injection, immersion, or orally well as freshwater fish. In Australia, VER was first detected
in the feed. Following a period of time (often temperature in barramundi, and the virus has been isolated from over
dependent) the host’s immune system responds to the 40 species globally (Colorni and Diamant, 2014).
viral ‘antigens’ and confers a level of acquired immunity
that will result in the production of antibodies to the Nervous Necrosis Virus (NNV) species are small
virus. These antibodies then provide specific protection (25–30 nm) single stranded RNA viruses that are non‐
and neutralization of the target viral pathogen if the ani- enveloped and possess an icosahedral capsid (Colorni and
mal comes into contact at a later time. Diamant, 2014). An important aspect of nodavirus infec-
tions is that they are transmitted vertically (from adults
Viral pathogens are a significant challenge for aquacul- to their offspring) and horizontally (among individuals).
ture and can be devastating in hatcheries or when out-
breaks occur at grow out sites. For most viral diseases,
there are limited control or prevention options available.
Pathogens and Parasites 219
It is likely that infected broodstock transmit VER virus to This virus grew on a range of fish cell lines and reacted
progeny during spawning or rearing. Clinical signs vary with commercial IPNV antisera; however, clinical dis-
but the virus is known to affect nervous tissues and can ease associated with this was never observed and it was
cause erratic swimming, whirling, loss of equilibrium, subsequently isolated from a range of other species in
and blindness; possibly due to its affinity for retinal tis- the marine environment. The first report of IPNV
sue, which is a primary site of viral replication (Colorni in Mexico came from a clinical outbreak in farmed
and Diamant, 2014). Because surviving fish often become rainbow trout (Oncorhynchus mykiss) where gross
asymptomatic carriers and the virus impacts larval and and microscopic pathology was consistent with IPN.
juvenile fish, biosecurity is important. Outbreaks have Further characterisation identified this as the Buhl
been reported in European sea bass (Dicentrarchus lab- strain of IPNV.
rax) in the 1990s, and the first report of VER virus in
North America occurred in juvenile California white sea The contagious nature of IPNV has been problematic
bass (Atractoscion nobilis) being cultured for population and fish that survive a disease outbreak often become
recovery efforts. Control measures for VER are limited asymptomatic carriers that are able to horizontally trans-
and no commercial vaccine is available. Because fish that mit IPNV without exhibiting clinical signs of disease.
survive outbreaks can become carriers, the best method The best control strategy for IPN or any viral disease
to reduce impacts from VER is to eliminate its presence affecting finfish is to avoid contact through proper bios-
through strict biosecurity and sanitation measures. ecurity. It is not always possible to avoid contact; there-
Infected fish may need to be culled from the population fore, other management options such as vaccination are
and new fish should be quarantined and tested prior to available in some countries.
mixing with the general population.
11.2.3 Herpesviruses
11.2.2 Birnaviruses
Two significant fish viruses in the order Herpesvirales and
Viruses in the family Birnaviridae are non‐enveloped the family Alloherpesviridae are Channel Catfish Virus
double‐stranded RNA (dsRNA) viruses with an icosahe- (CCV) and Koi Herpesvirus (KHV). These viruses are
dral shape that infects fish and shellfish. The genus highly contagious. CCV and KHV have resulted in sub-
Aquabirnavirus comprises three species, the first being stantial economic impact to the US catfish aquaculture
the type species Infectious Pancreatic Necrosis Virus industry and ornamental koi producers, respectively.
(IPNV), the second being the marine aquabirnavirus
(MABV) that infects yellowtail (Seriola quinqueradiata) CCV is considered endemic in the USA and channel
and is referred to as yellowtail ascites virus (YAV), and catfish virus disease (CCVD) emerged as a problem
the third being the Tellina virus (TV‐1) that was isolated during the early years of commercial catfish farming. In
from the marine mollusc Tellina tenuis. It should be the 1960s high mortalities in channel catfish (Ictalurus
noted that YAV has since been shown to infect a variety punctatus) were reported primarily in fry and finger-
of marine hosts. lings following introduction of fish from the hatchery to
the fry ponds and in 1971, CCV was identified as a
IPNV is one of the most significant finfish viruses in herpes virus. It is now present in nearly all regions in
this family and causes the disease Infectious Pancreatic the US that produce catfish. There are no prevention or
Necrosis (IPN). Although viral diseases were suspected treatment options for CCVD, but there have been
in hatcheries in the early to mid‐1900s, it was not until attempts to develop vaccines to CCVD. It was shown
the establishment of cell lines from specific fish species that a DNA vaccine could provide protection against
that IPNV and other fish viruses were identified. IPNV CCVD; however, the use of this under practical condi-
causes acute disease and is highly contagious, most often tions is limited by the need for injection delivery. In
affecting very young salmonid fry. In some cases, mortal- fish, CCV can persist in a carrier state and transmission
ity in the hatchery is up to 100% (Crane and Hyatt, 2011). of the virus is thought to be both vertical from the
IPNV is characterised as an aquatic birnavirus and since broodstock and horizontal through the water column.
the time of its discovery similar IPN‐like viruses (often In catfish ponds, water temperature and other environ-
non‐virulent) have been found in many salmonid and mental stressors appear to play a role in disease occur-
non‐salmonid hosts in both fresh and sea water. rence and severity.
Interesting case reports have identified both non‐ KHV (also known as cyprinid herpesvirus 3; CyHV‐3)
virulent forms of aquatic birnaviruses and virulent has a more recent history compared to CCV as it was
forms of IPNV that have impacted aquaculture facili- first reported in the UK in 1996. It has since been reported
ties. In Australia, the first report of an aquatic birnavi- and confirmed in nearly all countries that produce koi or
rus was from routine sampling of Atlantic salmon farms. other common carp, except Australia and New Zealand.
Interestingly, Australia is exploring the possibility that
220 Aquaculture of double‐stranded DNA that is methylated in a way simi-
lar to viruses affecting vertebrates.
KHV could be used as a potential biological control agent
against invasive common carp in areas where their popu- In general, ranaviruses and megalocytiviruses are
lations have expanded. KHV is listed by OIE as a notifia- important emerging pathogens for cultured and wild fish
ble disease and this creates regulatory concern for and affect fish in both marine and freshwater environ-
aquaculture producers and koi growers. Once a fish ments. Lymphocystis virus disease (LVD) affects fibro-
becomes infected or survives KHV disease (KHVD) they blastic cells in the skin and connective tissue resulting in
are potential carriers of the virus. superficial lesions on fish. LVD is known to affect more
than 125 wild and cultured fish species in marine and
There is no effective treatment for KHVD, however, an freshwater and is widespread in distribution. Transmission
attenuated live vaccine was approved for the prevention is horizontal and incidence rates for this disease may be
of this disease. It has been shown to generate high anti- as high as 70%. It has been speculated that another iri-
body titres specific for KHV and to protect common dovirus (WSIV) may be transmitted vertically as disease
carp or koi following challenge. One major issue is that outbreaks have been observed in progeny from adult
import and export control regulations may not allow white sturgeon (Acipenser transmontanus) collected
movement of KHV‐vaccinated fish. This relates to the from the wild and spawned for conservation or commer-
live nature of the vaccine and the inability of diagnostic cial aquaculture programs. The potential that WSIV
tests to differentiate between naturally infected or vac- could be transmitted vertically was investigated following
cinated fish. For the koi hobbyist or carp producer it is collection and disinfection of gametes from wild caught
important to maintain strict biosecurity and prevent sturgeon. Vertical transmission did not occur but prog-
exposure of fish to KHV. eny from eggs incubated using a river water source all
became infected. This and further cohabitation experi-
11.2.4 Iridoviruses ments clearly showed that WSIV was horizontally trans-
There are a number of viruses in the family iridoviridae mitted and highly contagious. WSIV does not cause a
that infect fish. A brief overview of the more significant systemic infection and appears to be primarily localised
species is provided here, but a critical review of fish iri- to the epidermis of the skin, barbells, oropharynx, and
doviruses is available in the literature (Whittington et al., gills of sturgeon. Although clinical signs vary, and limited
2010). Fish iridoviruses fall into three genera within this pathology or tissue distribution is observed, haemor-
family,Megalocytivirus,Ranavirus,andLymphocystivirus. rhaging can occur, and mortality may be high in heavily
There are also fish iridoviruses that are yet to be assigned infected fish due to secondary bacterial infection or pos-
to a genus, such as white sturgeon iridovirus (WSIV) and sibly a wasting syndrome resulting in emaciation and
Erythrocytic necrosis virus (Whittington et al., 2010). eventual death (Figure 11.2).
Iridoviruses are icosahedral in shape (Figure 11.1), large
in structure (120–300 nm) and may appear enveloped or Epizootic haematopoietic necrosis virus (EHNV) was
non‐enveloped in some cases. They consist of a genome isolated in 1985 in Australia and represents the first iri-
dovirus known to cause systemic infection and high
mortality in finfish. It appears to be restricted to
Australia, where large mortality events in wild redfin
perch (Perca fluviatilis) and severe outbreaks in farmed
rainbow trout (Oncorhynchus mykiss) have been
reported. Due to its virulence and localization, the dis-
ease epizootic heamatopoietic necrosis (EHN) is notifi-
able to OIE. In general, redfin perch are highly susceptible
and considered to be important carriers of the virus;
however, occurrence of mortality in farmed rainbow
trout highlights the potential risk should this virus spread
outside endemic areas.
Figure 11.1 Electron micrograph of white sturgeon, Acipenser 11.2.5 Orthomyxoviruses
transmontanus, iridovirus (WSIV) particles showing the Viruses in the family Orthomixoviridae consist of six
characteristic icosahedral shape. Source: Reproduced with genera. They include Influenza Virus A, B, and C, which
permission from Dr J. Drennan, Colorado Parks & Wildlife. infect humans, other mammals, and birds; Isavirus,
which infects salmon; Thogotovirus, infecting insects
and mammals, including humans; and Quaranjavirus, a
Pathogens and Parasites 221
(a) (b)
Figure 11.2 (a) WSIV infected cells in the gill lamellae of white sturgeon. Arrows indicate infected cells; and (b) juvenile white sturgeon
with signs of emaciation due to lack of food intake following infection with WSIV. Source: Reproduced with permission from Dr J. Drennan,
Colorado Parks & Wildlife.
new genus primarily infecting arthropods and birds. development and although the efficacy of such vaccines
Infectious salmon anaemia virus (ISAV) is an ortho- is not completely clear, there are vaccines available and
myxovirus in this family and considered the type virus in these have been used in North America.
the Isavirus genus. ISAV is the causative agent of infec-
tious salmon anaemia (ISA), a disease of Atlantic salmon 11.2.6 Rhabdoviruses
(Salmo salar). This disease that initially affected farmed Within the family Rhabdoviridae there are six genera;
Atlantic salmon in Norway, causes a systemic condition however, those that infect fish are primarily from the
resulting in severe anaemia in fish and is often linked to genus Novirhabdovirus. Viruses in this genus are gener-
necrosis and haemorrhaging of internal organs. An out- ally well characterised and infectious haematopoietic
break may result in high cumulative mortality over time, necrosis virus (IHNV) is the type species. Other fish
but the disease may be more chronic in nature with low rhabdoviruses were tentatively placed in the genus
level daily mortality of 0.05–0.1%. ISA is considered a Vesiculovirus, but two new genera have been recently
major disease impacting Atlantic salmon aquaculture proposed as rhabdoviruses that are not congruent
and has been isolated from fish in Eastern Canada, the with Novirhabdovirus (Kurath, 2012). These include
UK, Faroe Islands, and Chile. There are also reports of Sprivivirus of which spring viremia of carp virus (SVCV)
ISAV being discovered in tissues of farmed Atlantic is the type strain, and Perhavirus of which perch rhab-
salmon from the coast of British Columbia, Canada; dovirus is the type species. Rhabdoviruses of fish infect a
however, only sequences from the highly polymorphic broad range of host species and may have broad geo-
region (HPR) of ISAV variants have been identified by graphical distribution, including freshwater and sea
polymerase chain reaction (PCR) and to date ISAV has water environments. Some of the most severe and conta-
not been isolated in tissue culture. The lack of disease gious viruses impacting aquaculture and wild fish fall
outbreaks and limitations in interpreting positive results within this family. New fish rhabdoviruses continue to be
from molecular detection methods brings into question discovered and due to their significance, there is contin-
the presence of this virus in this region. It is clear that ual work in the areas of diagnostics and development of
identification of ISAV in British Columbia salmon farms new and better control/prevention methods.
would be particularly concerning since other salmonids
including Pacific salmon (Oncorhynchus tshawytscha) Rhabdoviruses that infect fish are usually bullet‐
can become infected. However, at this time, it does not shaped enveloped viruses with virions typically measur-
appear that clinical disease develops in species other ing approximately 75 nm wide and 180 nm long, replicate
than Atlantic salmon, but it can be speculated that viru- in the cytoplasm of cells and contain single‐stranded
lent strains of ISAV could emerge in Pacific salmon spe- RNA encoding five to six proteins. Some rhabdoviruses
cies if it were to persist in a carrier state. are of particular concern in wild and cultured fish, cause
diseases that are considered notifiable by OIE, and
Following infection from ISAV, fish have been shown include SVCV, IHNV, and viral haemorrhagic septicae-
to be resistant to re‐infection and develop a protective mia virus (VHSV). Of these, IHNV and VHSV have been
immune response. This has led to research on vaccine
222 Aquaculture severity in different host species. The ability of fish to
develop immunity to IHNV is well documented and
extensively characterised and many sources are available research in the area of vaccine development has been
in the literature that provide comprehensive descriptions substantial. The most effective vaccines against IHN are
and reviews (Kurath, 2012). DNA vaccines that must be injected into fish. The first
commercial DNA vaccine was developed and licensed by
IHNV causes infectious haematopoietic necrosis (IHN) the Canadian Food Inspection Agency (CFIA) in 2005
and was originally described following disease outbreaks and is available as a plasmid DNA‐based vaccine
in Sockeye (Oncorhynchus nerka) and Chinook salmon
(Oncorhynchus tshawytsch) in the Pacific northwest USA (Apex®‐IHN).
in the 1950s. It has since been a significant concern in
freshwater salmonid aquaculture (Figure 11.3) but can Another important fish rhabdovirus is VHSV. This
infect fish in both fresh and sea water environments. virus causes viral haemorrhagic septicaemia (VHS), con-
Outbreaks have occurred in Atlantic salmon farms in sidered the greatest disease problem in the European
Canada, but IHN remains a primary concern for wild trout farming industry. This virus was originally thought
fish or hatchery‐reared Pacific salmon in North America to be restricted to Europe, but it is now clear that it has a
as well as commercial rainbow trout farms in the USA broad host and geographic range and multiple strains of
and elsewhere. Originally considered a viral disease this virus have been described. In 1988 an interesting
restricted to North America, it has since spread to case occurred in Washington State where for the first
European and Asian countries where salmonids are cul- time VHSV was discovered in returning adult Pacific
tured. This disease affects young salmonids and an acute salmon. This discovery and isolation of VHSV resulted
outbreak can result in mortalities up to 100%; however, in the compulsory euthanasia of millions of hatchery fish
most outbreaks are less severe and surviving fish may that where the progeny of infected adults. This occurred
develop immunity to re‐infection. The best way to man- despite an actual disease outbreak and it was later found
age around IHN is to avoid contact but in areas where it that this strain of VHSV was distinct from the European
is endemic this can be difficult. As with most diseases in strain and appeared to be associated with a range of
fish, environment and stress during culture can impact marine species such as herring (Clupea pallasii) and cod
the severity of an outbreak. Five genogroups are known (Gadus macrocephalus). It is virulent to herring but
to exist for IHNV and these have been linked to outbreak unlike the European isolate it is avirulent in rainbow
trout. In 2003, VHSV emerged in wild fish in the
Laurentian Great Lakes in North America. Fish affected
by VHS were dying in large numbers and washing up on
shore in many areas. The number of fish and species
affected was alarming with over 1823 cases and 19 fish
species that tested positive for VHSV. This has led to fur-
ther characterization of VHSV strains and now a series
of four genogroups and additional subgroups exist, with
the Great Lakes strain designated as genotype IVb.
Figure 11.3 Severe exophthalmia in rainbow trout fry infected 11.3 Bacteria
with IHNV. Source: Reproduced with permission from G. Kurath.
Bacteria are categorised into a large domain of prokary-
otic microorganisms. They are larger than viruses, are
typically visualised using light microscopy, and may be
rod‐shaped, spherical, or spiral‐shaped. Many bacteria
are harmless or may even be beneficial to their host.
They can be found in the environment, may be part of
many animals’ natural gut microbiota, or in some cases
may invade an organism and cause disease. Bacterial dis-
ease outbreaks in aquaculture operations result in sig-
nificant economic losses in both public (resource
enhancement and stocking) and private sectors. In some
cases, there are vaccines used to prevent bacterial dis-
ease outbreaks, and antibiotics delivered in feed have
commonly been used to treat bacterial infections in fish
and other animals. Such treatments have been important Pathogens and Parasites 223
factors in disease management but represent risks due to
the ability of bacteria to develop resistance to common OIE as a notifiable disease; however, other regulatory
antibiotic treatments. Infection to humans due to aquatic agencies may restrict fish movement to non‐endemic
pathogens is relatively rare; however, bacteria are the areas (e.g., USA state or federal agencies acting under the
most common aquatic pathogens found to cause U.S. Fish and Wildlife Service Aquatic Animal Health
zoonotic infections by transmission from fish or culture Program (FWS‐AAHP) Title 50 act).
water to humans (see section 11.12).
Control of infection from A. salmonicida is most
In this section, an overview of select bacterial pathogens effective by preventing exposure. In areas where this
known to infect fish and cause significant economic pathogen is found in the environment, spring or ground
impact for aquaculture is provided. It is not possible to water should be used as a water source for aquaculture
cover all new and emerging bacterial pathogens of fish, facilities because they are typically free of most patho-
crustaceans and shellfish here, but it should be noted that gens. If an open water source such as a river, lake, or
at this time there have been 13 genera of bacteria reported marine environment is utilised then preventative meas-
as pathogenic to aquatic animals. These include Gram‐ ures such as vaccination should be incorporated if pos-
negative pathogens such as Aeromonas, Edwardsiella, sible. Due to the long history of furunculosis, some of
Flavobacterium, Francisella, Photobacterium, Piscirickettsia, the earliest attempts at vaccinating fish were with A. sal-
Pseudomonas, Tenacibaculum, Vibrio and Yersinia; and monicida. Currently, there are a range of furunculosis
Gram‐positive bacterial pathogens within the genera vaccines commercially available. The use of these and
Lactococcus, Renibacterium and Streptococcus. other bacterial vaccines has nearly eliminated the need
for antibiotic treatment in Norway’s Atlantic salmon
11.3.1 Aeromonas salmonicida industry. This is not the case everywhere and for some
species and regions, antibiotic treatments are adminis-
Furunculosis, caused by the Gram‐negative bacteria tered when fish show clinical signs of furunculosis. Due
Aeromonas salmonicida was one of the earliest described to the concerns over antibiotics, they should be used as
fish diseases and was named due the formation of large a last resort.
boils (furuncles) just under the skin in clinically sick
fish. First described in the late 1800s in cultured and 11.3.2 Edwardsiella ictaluri
wild fish in Europe, it is widely distributed and infects
many salmonid and non‐salmonid species in marine Enteric septicaemia of catfish (ESC) is caused by the
and freshwater worldwide. Although the taxonomy of Gram‐negative bacteria Edwardsiella ictaluri. This is
Aeromonas is not always agreed upon, there are four considered one of the most severe bacterial diseases in
primary subspecies of A. salmonicida that are consid- the catfish industry in the USA and may account for 30%
ered infectious. The ‘typical’ strain is considered A. sal- or greater losses. Cases of ESC are most often reported
monicida salmonicida primarily affecting salmonids, in the spring around May and June and also in autumn in
while the subspecies masoucida, achromogenes, and September and October due to the bacterium’s prefer-
smithia are ‘atypical’ strains that affect salmonids, and ence for temperatures between 22 and 28 °C. In general,
A. salmonicida nova is an ‘atypical’ strain that is infec- channel catfish (Ictaluri punctatus) are the primary spe-
tious for non‐salmonid fish. cies that is susceptible to ESC, but infection with E. icta-
luri along with minor pathology has been reported in
Furunculosis is widely reported in commercial and other species of catfish and non‐ictalurids. It has been a
‘resource’ hatcheries in the USA that stock fish into primary problem in the USA, but outbreaks, sometimes
public waters and is also known to impact wild fish. This severe, have been reported in Australia, Indonesia, Japan,
disease became problematic in the Atlantic salmon Thailand, and Vietnam.
industry due to outbreaks in smolts when they were
moved from freshwater to seawater. When the disease Edwardsiella ictaluri infections can lead to acute or
affects young fish, mortality can be acute and clinical chronic forms of ESC. In the acute form the infection is
signs may be limited to darkening of fish, lethargy, and systemic with bacteria moving into the blood stream.
anorexia. The formation of haemorrhaged boils or This causes septicaemia and results in ulcerations and
furuncles may appear more often in fish that are chroni- necrosis in multiple organs. Haemorrhaging may appear
cally affected. If the furuncles rupture and release bacte- around the mouth and operculum and along the base of
ria, toxins or necrotic cells, an increase in bacterial load the fins (Figure 11.4). Interestingly, in the chronic form,
can occur and thus an increase in the risk of horizontal E. ictaluri may enter the olfactory organ or nasal passage
transmission to other fish. As with other bacterial diseases and move into the skull and skin of the head to form a
of aquatic organisms, A. salmonicida is not considered by lesion, leading to a condition known as ‘hole in the head’.
Control of ESC in commercial catfish operations
has relied on antibiotic treatment, and such treatments
224 Aquaculture
Figure 11.4 Channel catfish (Ictalurus punctatus) showing Figure 11.5 Light micrograph or Flavobacterium psychrophilum
external clinical signs of haemorrhaging near the mouth and base cells showing long thin rod‐shaped bacteria. Source: Reproduced
of the fins following laboratory challenge with E. ictaluri. Source: with permission from Dr B. LaFrentz.
Reproduced with permission from Dr B. LaFrentz.
following nano‐injection, which resulted in high mortal-
continue to be used to control losses due to this disease. ity in eyed eggs and early hatched fry due to F. psychro-
There is a commercial immersion vaccine consisting of a philum. Such findings make broodstock selection critical
live‐attenuated strain of E. ictaluri available in the USA. and highlight the potential for this bacterium to persist
The potential to prevent major impacts due to ESC within a hatchery environment.
through vaccination is high, but some practical difficul-
ties exist because delivery of the vaccine to fish when Flavobacterium psychrophilum is a Gram‐negative
they are fully immunocompetent is often not possible. bacterium with cell morphology consistent with other
Catfish production in many cases is on a large scale and Flavobacterium species. Although strain differences
ponds are often stocked shortly after the fry hatch, which exist, most bacterial cells are rod shaped (Figure 11.5) and
is usually the only time fish are handled until harvest and range in size from 0.2–0.75 × 2–7 μm. This bacterium has
therefore the only time a vaccine can be administered. gliding motility and a gliding mobility protein (GldN)
may affect the bacterium’s ability to enter and infect cells.
11.3.3 Flavobacterium psychrophilum
Bacterial coldwater disease (BCWD) and rainbow trout Cases of BCWD and/or RTFS are widespread and
fry syndrome (RTFS) are caused by Flavobacterium psy- include most salmonid producing countries. It is wide-
chrophilum. This bacterial pathogen and a number of spread in the USA and European trout industries, and
other bacteria in the Flavobacteriaceae family can cause F. psychrophilum has been reported in trout and other
disease in cultured and wild fish. Flavobacterium psy- species in a number of countries, including Australia
chrophilum is one of the best‐known pathogens in this where it was reported in Atlantic salmon during the
genus (along with F. columnare) due to the significant freshwater stages of production. There have been a
impact BCWD has on salmonid aquaculture worldwide. large number of F. psychrophilum strains isolated and
Nearly all salmonids are susceptible to F. psychrophilum such strains can have varying levels of virulence.
and it has been reported to cause disease in a number of Although mortality may be high from BCWD or RTFS,
non‐salmonid species as well. Young fish are primarily commercial aquaculture also experiences losses due to
affected, but when early hatched fry are infected, mortal- growth and performance impacts as well as increased
ity may be greater than 50%. Although considered ubiq- levels of deformities (resulting in a lower grade product)
uitous in the aquatic environment, it appears that following an outbreak. In the USA, public hatcheries
F. psychrophilum can be transmitted both horizontally that rear steelhead (ocean run rainbow trout) and
and in some cases vertically from parent to progeny salmon for stocking purposes are also heavily impacted
(Cain and Polinski, 2014). It has been isolated from sal- by BCWD, and this disease causes greater overall losses
monid eggs and egg contents, and it has been shown that in these facilities than any other fish disease (Cain and
F. psychrophilum could survive inside fertilised eggs Polinski, 2014).
Clinical signs of BCWD/RTFS will vary depending on
the species and age of fish infected. There are acute forms
of the disease that cause septicaemia and high mortality,
but lingering chronic infections may also occur. Fish
infected with F. psychrophilum exhibit behavioural
changes such as spiral swimming or lack of feeding Pathogens and Parasites 225
response. Internally, this bacterium has an affinity for the
spleen but it can be isolated from many organs and tis- 11.3.4 Flavobacterium columnare
sues depending on the severity of infection. Externally, a Flavobacterium columnare is the causative agent of
range of clinical signs may appear including frayed fins, columnaris and has historically been a major disease in
exophthalmia, dark pigmentation, or haemorrhages on warmwater aquaculture; however, columnaris is emerg-
the skin or bases of fins. In some cases, the caudal pedun- ing as a significant problem in salmonid aquaculture
cle region may be severely eroded (Figure 11.6), which with infections increasing in recent years. A wide range
relates to this disease originally being referred to as of species are affected by columnaris, and in the catfish
‘peduncle disease’. industry, it has been estimated that infection by
F. columnare resulted in mortality and 39% losses in
Currently, control of BCWD/RTFS relies on good fish 2009. Clinical disease occurs most often in young fish
culture practices and antibiotic treatment following con- and transmission of this pathogen is considered horizon-
firmation of an outbreak. However, there has been exten- tal. Good water quality is important as outbreaks can be
sive work aimed at developing an effective vaccine to linked to stress associated with poor environmental con-
prevent and/or limit disease impacts. Due to the small ditions. Temperature plays an important role in infection
size and the production requirements for species such as with occurrence in channel catfish being most prevalent
rainbow trout, the key to a successful vaccine has been at temperatures between 25–32 °C (Lio‐Po and Lim,
creating one that can be mass delivered to very young 2014). In wild adult salmon in the Pacific northwest of
fish. Recently, a live‐attenuated F. psychrophilum strain the USA, columnaris has been an issue during migration
that provides protection following immersion vaccina- from the ocean to spawning grounds since many of the
tion has been developed. This shows great potential for reservoirs they must pass through have elevated water
commercial development, and more recent improve- temperatures in spring and summer months.
ments in the formulation have improved efficacy in the
laboratory and in the field. Other alternative control Similar to other Flavobacteria, F. columnare is Gram‐
strategies may include the use of natural gut bacteria that negative and represented as a long rod‐shaped bacte-
can be incorporated into feed as probiotics. This has rium. One characteristic that gives columnaris its name
been shown to reduce mortality from BCWD following relates to the tendency of these bacteria to form ‘hay
feeding of two bacterial strains (Enterobacter C6‐6 and stacks’ or columns in wet mounts of the gills or other
C6‐8) isolated from the gut of healthy rainbow trout. infected tissues (Figure 11.7). Clinical signs of disease
Further work has demonstrated that reduced mortality is often include frayed fins, skin lesions that may appear
due to the expression of an anti‐microbial peptide by the yellowish in colour or in some cases depigmented
Enterobacter. (Figure 11.8), or gill damage due to colonisation of bacte-
ria (Figure 11.9). It has been suggested that acute mortal-
ity results more often when F. columnare is associated
with the gills (Lio‐Po and Lim, 2014), but infections can
become systemic and result in high mortalities and in
some cases minimal pathology to infected organs. To
Figure 11.6 Laboratory challenged rainbow trout showing severe Figure 11.7 Columns or ‘hay stacks’ formed by Flavobacterium
erosion in the dorsal musculature at the site of Flavobacterium columnare visible in a wet mount of infected gill tissue. Source:
psychrophilum injection. Source: Reproduced with permission Reproduced with permission from Dr A. E. Goodwin.
from Dr B. LaFrentz.
226 Aquaculture 11.3.5 Piscirickettsia salmonis
Salmonid rickettsial septicaemia (piscirickettsiosis)
Figure 11.8 Depigmented lesions following laboratory challenge caused by the intracellular Gram‐negative bacteria
of tilapia Oreochromis sp. with Flavobacterium columnare. Source: Piscirickettsia salmonis is a disease that impacts fish pri-
Reproduced with permission from Dr B. LaFrentz. marily in seawater. It infects salmonids and was first
reported in coho salmon in Chile. This disease has had
Figure 11.9 Gill necrosis following laboratory challenge of the greatest impact on salmonid aquaculture in Chile,
rainbow trout with Flavobacterium columnare. Source: Reproduced but P. salmonis has been found in a number of countries
with permission from Dr B. LaFrentz. including Europe, North America, and Australia, where
a rickettsia‐like organism was isolated in Tasmania. In
prevent or control columnaris, it is critical to maintain Chile, cases resulting in up to 90% mortality of coho
fish in optimal conditions. Should an outbreak occur and salmon were reported in the 1980s with clinical disease
represent primarily a systemic infection, then antibiotic occurring between 6–8 weeks post transfer to seawater
treatment with medicated feed may be beneficial. cages. Transmission of P. salmonis appears to be primar-
External and gill associated infections may require appli- ily horizontal, but it has been suggested that vertical
cation of chemical therapeutants, such as potassium transmission can occur.
permanganate to the water (Lio‐Po and Lim, 2014).
Currently, in the USA there is a commercially available The intracellular nature of P. salmonis makes isolation
vaccine aimed at prevention of columnaris in channel and confirmation of disease more difficult. Typically, pis-
catfish; however, it is not clear how widespread its use is cirickettsiosis is diagnosed following histological exami-
or if this live‐attenuated vaccine is efficacious in other nation and immunohistochemistry of cells infected with
fish species. P. salmonis. However, it can be isolated using fish cell
lines in a similar manner to virus isolations. Cytopathic
effect due to P. salmonis is evident on CHSE‐214 cells
when incubated for up to two weeks at temperatures of
15–18 °C. Other serological and molecular methods such
as PCR are available to confirm infection.
Fish that are affected by P. salmonis may be lethargic,
go off feed, become dark in colour, and have pale gills.
Internally, the posterior kidney and spleen may be
enlarged, and the liver may develop nodules that if rup-
tured will result in crater‐like lesions. In some species,
the nervous system can be affected, and fish will swim in
an irregular manner.
Prevention and control options for piscirickettsiosis
have been limited in the past and the use of antibiotics
appears to have been less successful due to the intracel-
lular nature of P. salmonis. Vaccination has shown prom-
ise and commercial vaccines have recently become
available. Although the efficacy of these vaccines is still
in question, it is clear that prevention of an outbreak
through methods such as vaccination offer the best
opportunity to manage this disease in aquaculture. Good
management practices including low density stocking of
pens and removal of mortalities early in an outbreak are
important. Other considerations include potential
screening of broodstock and rejection of eggs from fish
that are positive for infection.
11.3.6 Vibrio spp.
Vibriosis is caused by a number of bacterial species in the
family vibrionaceae and affects both coldwater and some
warmwater species of fish, crustaceans and molluscs in
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