THE 4th TROPICAL WEED SCIENCE CONFERENCE 2013

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 49

from strong allelopathic plants, might successfully overcome this problem. The objectives of this
research were: (i) to determine the allelopathic effects of aqueous extracts and dried powder and pellet
of A. odorata on the germination and initial seedling growth of Echinochloa crus-galli and Phaseolus
lathyroides under laboratory and pot culture conditions and (ii) to evaluate the herbicidal potential of A.
odorata granule applied to three soil types on the emergence and growth of large crabgrass (Digitaria
adscendens) and horse purslane (Trianthema portulacastrum) in an experimental greenhouse and (iii)
to evaluate the herbicidal potential of A. odorata granule to control weeds and effects on growth and
yield of maize under natural field conditions.

Material and method
Plant material

Plant materials of Chinese rice flower plants were collected before flowering and were
separated into the leaves and branches. They were dried in hot-air oven at 45 °C for 72 h.
and cut into 1 cm pieces, powdered in a blender and sieved through 40 mesh (420 µm) sieve.

Preparation of A. odorata in 3 forms and seed bioassay under laboratory conditions
In this experiment there were two factors; (i) two test weed species (E. crus-galli and P.

lathyroides) (ii) A. odorata in 3 forms (aqueous extract, dried leaf powder and pellet). The aqueous
extracts were prepared from powdered leaves and branches of A. odorata which extracted with distilled
water for 72 h at 10 °C, followed by filtration to remove any debris. Dried leaf powder was prepared
from dried leaves which were powdered in a blender. The pellets were prepared by mixing 50% A.
odorata dried leaf powder, 25% cassava glue and 25% CaCO3 powder were dried in hot-air oven at
45°C for 3 days. Weed seeds were transferred to Petri-dishes containing filter paper moistened with 5
mL of distilled water and 4 application doses of each different form. Petri dishes were kept in a growth
chamber. The number of germinating seeds was counted and seedling growth was measured as the root
and shoot lengths at seven days after treatment. Aqueous extract form was applied at 6.25, 12.5, 25 and
50 g dried leaf L–1. The dried powder of A. odorata was added at 31.25, 62.5, 125 and 250 mg/Petri dish
and pellets were applied at 62.5, 125, 250, and 500 mg per Petri dish which equivalent to 6.25, 12.5, 25
and 50 g dried leaf L–1, respectively.

Soil application bioassay
The influence of A. odorata pellets was explored in pot studies done in experimental

house, on the seedlings emergence and growth of E. crus-galli and P. lathyroides. Number
of emerged weeds plants was counted at 14 days after sowing, while plant height and their
biomass were determined at 28 days.

Effect of three soil types on the efficacy of the organic herbicide from A. odorata granules
The effect of the A. odorata granules on the emergence and seedling growth of D. adscendens

and T. portulacastrum was explored under pot conditions in an experimental greenhouse. Clay soil,
sandy loam soil and sand were used as the three soil types. The number of plants of each bioassay
species that emerged was counted at 14 days after treatment and the plant height was determined at 28
days after initiating the treatments. The biomass was determined separately at 28 days after initiating
the treatments.

Field experiment: Effect on growth of weeds and maize
A. odorata granule, a control with the herbicide atrazine and another control without

organic herbicides or atrazine were applied under maize field conditions. After planting 3 weeks
was assessed the emergence of weeds and 12 weeks after planting dry weight was assessed
in two 1 m2 quadrates placed randomly in each plot at maize harvest. Weeds were counted by
species, clipped at ground level within each quadrate. The weed is dried by oven drying to
constant weight at 72°C, for comparison their dry weight. Maize silage yields can be obtained by

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 50

harvesting at the maize silage (12 weeks after planting); In each plot was harvested at stage 20
plants were harvested from the center rows and compared of crop yield for all plant treatments.

Statistical analysis
Inallexperiments,Dataweresubjectedtoone-wayanalysisofvariance(ANOVA)andcomparison
is statistically significant between treatments mean with Tukey’s Studentized Range test at p < 0.05.

Results

The A. odorata in 3 forms at all test concentrations markedly reduced the seed germination
(%) of both test species compared to control (Table 1).The leaf and branch aqueous extracts at all
concentrations inhibited seed germination and seedling growth, and the degree of inhibition increased
with the incremental extracts concentration but the leaf extract was slightly more inhibitory than the
branch extract. The aqueous extracts of leaf and branches of A. odorata inhibited the germination and
seedling growth of E. crus-galli and P. lathyroides. The degree of toxicity of different A. oderata forms
can be classified in order of decreasing inhibition as pellet > dried leaf powder > aqueous extract. In
pot culture, pellet reduced the seedlings emergence, plant height and dry biomass of both test weeds:
E. crus-galli and P. lathyroides (Table 2). The reduction in emergence of E. crus-galli at the lowest
dose (0.5 t ha-1) was 67% and was only 2% in P. lathyroides. At the highest dose (2.0 t ha-1 ), the
emergence of E. crus-galli was reduced by 92% and that of wild pea only by 30%.With increasing doses
of application, the weed emergence as well as the plant height and dry biomass were decreased further.
The pellets were more inhibitory to E. crus-galli than to P. lathyroides.

The effects of applying A. odorata granules to the three soil types on emergence, plant height,
and dry weight of D. adscendens and T. portulacastrum are shown in Fig. 1 and 2, respectively. The
emergence, plant high and dry mass of D. adscendens and T. portulacastrum were affected by soil type
and varied with the amount of A. odorata granules applied. The degree of toxicity of different soil types
can be classified in order of decreasing inhibition as sand > sandy loam >clay. The emergence, plant
height, and dry weight of T. portulacastrum were less affected than those of D. adscendens. These
results indicate that A. odorata granules has strong herbicidal potential for controlling D. adscendens.
Under maize field conditions, the number of D. adscendens plants was affected significantly at all A.
odorata granule doses and with the atrazine application, except for the lowest A. odorata granule dose
(0.25 t ai ha-1) that was applied (Fig.3). The decrease in the number and biomass of the emerged plants
of D. adscendens plants in the 1 t ai ha-1 A. odorata granule-applied plots was similar to that observed
in the plots that received atrazine. Two additional broad-leaved weed species, T. portulacastrum and
Amaranthus gracilis, were found in the experimental plots that receive A. odorata granule, however,
at the highest dose of application (1 t ai ha-1), the emergence and dry weight of the two weed species
were lower than those in the control plot. The A. odorata granules did not provide the same level of
broad-leaved weed (T. portulacastrum and A. gracilis) control as the synthetic herbicide (atrazine), but
the broad-leaved weeds that remained after applying the A. odorata granules seemed to be unable to
compete with the maize. The yield of the maize plants growing in the plots that received the 0.25 t ai
ha-1 A. odorata granule treatment was lower than those in the 0.5 and 1 t ai ha-1 treatment plots (Fig.4).
The increase in the maize yield in the plots receiving the A. odorata granule treatment of 1 t ai ha-1 was
5.5% greater than that in the plots receiving the herbicidal treatment.

Discussion

Leaf and branch aqueous extracts of A. odorata were assayed for their effects on seed germination
and early seedling growth of E. crus-galli and P. lathyroides L. The leaf and branch aqueous extracts at
all concentrations inhibited seed germination and seedling growth, and the degree of inhibition increased
with the incremental extracts concentration but the leaf extract was slightly more inhibitory than the
branch extract. The results of this study suggest that branch and leaf of A. odorata contain water-soluble

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 51

allelochemicals. Further work is needed to specify and verify the allelochemicals produced by this plant
at different forms under laboratory conditions. The degree of toxicity of different A. odorata forms
can be classified in order of decreasing inhibition as pellet > dried leaf powder > aqueous extract. The
inhibitory effects of A. odorata powder were stronger than the effects of the aqueous extracts because
in aqueous extract, the allelochemicals were extracted partially from the leaf powder in water, during
the soaking at 10 °C for 72 h. However, the allelochemicals in leaf powder might have released
continuously during the experimental period. Interestingly, the inhibitory effects of pellets prepared
from the dried leaves were stronger than equal amount of leaf powder. This phenomenon might be
attributed to the moisture of cassava glue acting as a compatibile medium during the pellets preparation.
The A. odorata pellets under pot culture conditions also suppress the growth and yield of E. crus-galli
which may be exploited to create a successful biorational herbicide.

The potential use of A. odorata granules as a weed suppressant was determined under experimental
greenhouse and field condition. Under experimental greenhouse, the emergence and seedling growth of
D. adscendens and T. portulacastrum was inhibited but varied with soil type; the inhibitory effect of the
A. odorata granules depended on soil type; the inhibitory effect was extremely low in clay, as compared
to sand, indicating that the allelochemical activity of A. odorata granules was greatly influenced by
the soil physicochemical properties such as organic mater content. This result agreed with Kobayashi
(2004) who reported that the sensitivity of weed species to phytotoxins depends on the physiological
and biochemical characteristics of each species and the activity of phytotoxins become weak in the
soil (Laosinwattana et al., 2010). Under natural field conditions, A. odorata granules could be used to
suppress D. adscendens weed emergence and growth in maize field and had no adverse effect on maize
growth and silage yield. The lack of a reduction in maize silage yield by A. odorata granule agrees with
the results reported by Dhima et al., (2010) who found that the use of aromatic plants incorporated as
green manure for a maize crop did not significantly affect maize development.

Table 1 Effect of application of A. odorata in 3 forms on germination and seedling growth of
Echinochloa crus-galli and Phaseolus lathyroides in Petri dish.

------------------------------------------- inhibition (%) over control ------------------------------------------

Conc. Aqueous extract Dried leaf powder Pellet
(g dried

leaf /L) Seed Root Shoot Seed Root Shoot Seed Root Shoot

germination length length germination length length germination length length

Echinochloa crus-galli

6.25 2.5 -1.68 -3.3 25 24.04 0 30 16.83 17.09

12.5 21.75 5.05 62.91 50 69.95 62.91 70 74.76 83.49

25 70 34.14 95.53 78 82.21 97.48 88 91.83 98.06

50 100 100 100 100 100 100 100 100 100

Phaseolus lathyroides

6.25 0 -0.81 2.91 0 16.26 -12.62 3 6.02 2.91

12.5 11 4.55 62.14 20 36.42 39.32 25 53.01 62.14

25 47.5 37.72 88.83 54.5 65.37 64.08 69.5 85.85 95.15

50 74.5 80 91.26 100 100 100 100 100 100

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 52

Table 2 Emergence (14 DAT) and growth (28 DAT) of Echinochloa crus-galli and Phaseolus
lathyroides in response to the use of A. odorata pellet formulation as soil surface mulch in the pot
experiment.

---------------------------------- inhibition (%) over control ---------------------------------

Pellet dose Echinochloa crus-galli Phaseolus lathyroides
(t/ha)

Emergence Plant height Biomass Emergence Plant height Biomass

0.5 67 34.62 82.43 2 4.86 10.34

1 92 62.81 96.74 15 2.43 18.1

2 100 100 100 30 27.32 41.81

120

100

% of control 80

60

40

20

0 0.25 0.5 1 0 0.25 0.5 1 0 0.25 0.5 1
0 clay soil Sandy loam soil Sand soil

Dose (ton (ai)/ha) Dry weight

Emergence Plant height

Fig. 1 Emergence, plant height, and dry weight of Digitaria adscendens in clay, sandy loam, and sand
amended with different amounts of A. odorata granules in test pots.

120

100

80

% of control 60

40

20

0 0.25 0.5 1 0 0.25 0.5 1 0 0.25 0.5 1
0 clay soil Sandy loam soil Sand soil

Dose (ton (ai)/ha) Dry weight

Emergence Plant height

Fig. 2 Emergence, plant height, and dry weight of Trianthema portulacastrum in clay, sandy loam, and
sand amended with different amounts of A. odorata granules in test pots.

% of control 53
TWSC 2013
The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand

100

80

60

40

20

0 Dry mass De n s i ty Dry mass De n s i ty Dry mass
De n s i ty

D. adsxe nde ns T. portulacastrum A. gracillis

non-weed control 0.25 ton/ha A.odorata granule
0.5 ton/ha A. odorata granule 1 kg/ha Atrazine
2 kg/ha Atrazine

Fig. 3 Effect of A. odorata granules applied as a pre-emergent herbicide on weed density and dry
biomass under field conditions.

2.5 kg/ha Atrazine

1 ton/ha A.odorata 20 40 60 80 100 120
granule % of control

0.5 ton/ha A. odorata
granule

0.25 ton/ha A.odorata
granule

0

Dry Mass Grain yield

Fig. 4 Effect of A. odorata granules applied as a pre-emergent herbicide on dry biomass grain yield of
maize under field conditions.

Conclusion

Leaf and branch from A. odorata contained certain allelochemicals inhibitory to E. crus-galli
and P. lathyroides germination and growth. The degree of toxicity of different A. odorata forms can be
classified in order of decreasing inhibition as pellet > dried leaf powder > aqueous extract. J. odorata
pellet forms under pot culture condition also suppress the growth of E. crus-galli. Under natural field
conditions, an organic herbicide produced from A. odorata in the form of a powder product could be
used to suppress D. adscendens weed emergence and growth in maize field and had no adverse effect
on maize growth and silage yield.

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 54

Acknowledgements

This research was supported by grants from The Research Foundation of King Mongkut’s
Institute of Technology Ladkrabang and Thailand Toray Science Foundation, Bangkok, Thailand.

References

Batish, D.R, Kaur, M., Singh, H.P. and Kohli, R.K. (2006)a. Phytotoxicity of a Medicinal Plant,
Anisomeles indica, Against Phalaris minor and Its Potential Use as Natural Herbicide in Wheat
Fields, Crop Prot. 26: 948-952.

Dhima, K.V., Vasilakoglou, I.B., Gatsis, Th.D., Panou-Philotheou, E. and Eleftherohorinos, I.G. (2010).
Effects of Aromatic Plants Incorporated as Green Manure on Weed and Maize Development.
Field Crops Res. 110: 235-241.

Duke, S.O. (1986). Naturally Occurring Chemical Compounds as Herbicides. Rev. Weed Sci. 2: 15-
44.

Einhellig, F.A. (1995(. Allelopathy: Current Status and Future Goals. In: Inderjit, Dakshini, K.M.M.,
Einhellig, F.A. (Eds.). Allelopathy: Organisms, Processes and Applications. American Chemical
Society, Washington, DC, pp. 1-24.

Fujii, Y. (2001). Screening and Future Exploitation of Allelopathic Plants as Alternative Herbicides
with Special Reference to Hairy Vetch. Crop Prot. 4: 257-275.

Inderjit, Streibig, J.C. and Olofsdotter, M. (2002). Joint of Action of Phenolic Acid Mixtures and Its
Significance in Allelopathy Research. Physiol. Plant. 144: 422-428.

Khanh, T.D., Hong, N.H., Nhan, D.Q., Kim, S.L., Chun, I.M. and Xuan, T.D. (2006). Herbicidal Activity
of Stylosanthes guianensis and Its Phytotoxic Components. J. Agron. Crop Sci. 192: 427-433.

Kohli, R.K., Batish, D.R. and Singh, H.P. (1998). Allelopathy and Iits Implications in Agroecosystems.
J. Crop Prod. 1, 169–202.

Kobayashi, K. (2004). Factors Affecting Phytotoxin Activity of Allelochemicals in Soil. Weed Biol.
Manag. 4: 1-7.

Laosinwattana, C., Boonleom, C., Teerarak, M., Thitavasanta, S. and Charoenying, P. (2010). Potential
Allelopathic Effects of Suregada multiflorum and The Influence of Soil Type on Its Residue’s
Efficacy. Weed Biol. Manag. 10: 153-159.

Lin, D., Sugitomo, Y., Dong, Y., Terao H. and Matsuo, M. (2006). Natural Herbicidal Potential of
Saururaceae (Houttuynia cordata Thunb) Dried Powders on Paddy Weeds in Transplanted
Rice. Crop Prot. 25: 1126-1129.

Singh, H.P., Batish, D.R., Kaur, S. and Kohli, R. (2003)a. Phytotoxic Interference of Ageratum
conyzoides with Wheat (Triticum aestivum). J. Agron. Crop Sci. 189: 341-346.

Xuan, T.D., Tawata, S.T., Khanh, D. and Chung, I.M. (2005). Biological Control of Weeds and Plant
Pathogens in Paddy Rice by Exploiting Plant Allelopathy. Crop Prot. 24: 197-206.

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 55

Diversity of Hyphomycetes Fungi from Diseased Weeds

Duangporn Suwanagul1, Jitra Kokaew2 and Anawat Suwanagul2
1Faculty of Biotechnology, Rangsit University Pathum Thani 12000, Thailand
2Thailand Institute of Scientific and Technological Research, Pathum Thani 12120, Thailand
E-mail addresses: [email protected], 76 [email protected], [email protected]

Abstract

Diseased weeds showing leaf spot and leaf blight were collected from vegetable plots in
Central of Thailand. Seven weed hosts are included Cyperus rotundus (Cyperaceae), Cyperus
brevifolius (Cyperaceae), Brachiaria mutica (Poaceae), Eleusine indica (Poaceae), Dactyloctenium
aegyptium (Poaceae), Pennisetum polystachyon (Gramineae) and Oryzae sativa f. spontanea
(Poaceae). Tissue transplanting and moist chamber methods were used to isolate microfungi.
Identification was base on growth rate, colony color and other microscope features as observed on
artificial media. Microscopic characteristics were examined under stereo and light microscopes.
A total of 20 fungi isolates, comprising of 9 species, were found. These are included Alternaria
alternate, Bipolaris bicolor, Curvularia intermedia, Curvularia pallescens, Drechslera holmii,
Exserohilum rostratum, Fusarium oxysporum, Myrothecium verrucaria and Nigrospora oryzae.
Five different preservation methods, including PDA slant, liquid paraffin, filter paper, grain culture
and 15% glycerol method, were employed to maintain pure culture of all fungal for future use.

Keywords: Weeds, Fungi, Htphomycetes, Biopesticides

Introduction

The Hyphomycetes fungi is importance to human, animals and plants (Sivanesan, 1987; De
Hoog et al., 2000). Manoch et al. (2004) reported Alternaria alternata, Stemphylium solani, Curvularia
lunata, Thielaviopsis basicola and T. thielavioides caused disease in plant. De Hoog et al. (2000) and
Moss (2003) reported human and animals diseases caused by Hyphomycetes fungi such as Arthrinium
phaeospermum, Exerohilum rostratum, Memnoniella echinata, Pithomyces chartarum and Stachybotrys
chartarum. In the other hand, the Hyphomycetes is beneficial fungi because they can produced many
secondary metabolites such as nigrosporin A & B from Nigrospora oryzae can inhibited Bacillus
subtilis be equal to streptomycin in vitro (Tanaka et al., 1997). Oidiodrendron griseum produced
10-methoxydihydrofuscin, fuscinarin and fuscin that is efficiency to inhibited HIV-1 (Yoganathan et
al., 2003 ). Sin et al., 2002 reported Gilmaniella bambusae, Periconia minmibissia and Tetraploa
aristata produced enzyme cellulase and xylanase. Nigrospora oryzae produced secondary metabolite
to inhibit spore germination of fungal pathogens such as Fusarium avenaceum, F. culmorum, F.
equiseti, F. gramminearum, F. oxysporum, F. lateritium and Botrytis cinerea (Szewczuk et al., 1991).

The benefit and importance of Hyphomycetes fungi have been reported by many
mycologist. Simmons (2004) reported new species of Dematiaceous Hyphomycetes from many
plants. Varma et al. (2006) reported Curvularia lunata caused leaf spot disease on grass this fungi
produced 4-epiradicinol inhibit bacterial growth. Nikonov et al. (2007) reported Alternaria
alternata and Curvularia geniculata produced enzyme laccase to degrade lignin and humus.
The investigations on Hyphomycetous fungi causing weed diseases have been reported
by many researchers (Auld and Mc Rae, 1999; Babu et al., 2003). The exploitation of fungal plant
pathogens as biological weed control agents has gained considerable importance due to the secondary
metabolite they produced (Bonilla et al., 1999; Boyette et al., 2002; Domsch et al., 1993). The
purposes of this study were to isolate and identify Hyphomycetes fungal species from diseased weeds.

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 56

Materials and Methods

1. Moist chamber method
Each excrement sample was placed in a moist chamber consisting of a glass bowl or plastic box lined

with damp cotton or tissue-paper and placed by the window. They were incubated for 2-7 days or longer at
28oC and observation was made under stereomicroscope. Transferred needle was used to transfer spores or
fruitingstructuresonaslideandmountwithadropofdistilledwater,thencoveredwithcoverslipandexamined
under light microscope with Normaski Interference Contrast. Photomicrographs were taken were used.

2. Tissue transplanting method
Small pieces of diseased weeds were surface sterilized in 10 % clorox for 3-5 min, rinsed with

sterile distilled water, transferred to sterilized tissue paper and placed on potato dextrose agar (PDA) in
a petridish. They were incubated at 28°C for a few days. Hyphal tips were transferred on to PDA slants.

3. Identification of Fungi
Macroscopic features were studied including colony growth pattern, color, texture on

different agar media. Fungal growth rate was measured on PDA, CMA. CZA and MEA. Microscopic
characters were observed on a slide preparation using sterile distilled water and lectophenol as
mounting media and examined under a light microscope (Olympus BH-2 with Normaski Interference
Contrast). Photomicrographs of fungal structure were taken under stereo, light microscopes.

4. Preservation of fungi
Five different preservation methods were used to maintain all fungal pure cultures.

4.1 PDA slant method (Monoch et al., 2005; Smith and Onions, 1994)
Pure cultures of fungi were maintained on PDA slants at 28o C. Sub-culturing was carried out
every 6 months.

4.2 Liquid paraffin method (Monoch et al., 2005)
Pure cultures were maintained on PDA agar slant in a vial (1 dram). Liquid paraffin was placed
in a vial and autoclaved twice. Covering the pure culture on agar slant with sterile liquid paraffin about
2/3 of a vial and stored at 28o C. in order to prevent dehydration and slows down metabolic activity and
growth through reduced oxygen tension.

4.3 Filter paper method (Jeamjitt, 2007)
Fifteen pieces (0.5 x 0.5 cm2) of sterile filter paper Whatman no. 1
were placed on PDA in sterile Petri-dishes. The mycelia were transferred on PDA and incubated for
7-14 days depend on the species. The filter papers with mycelium were transferred to new sterile Petri
dishes by using sterile forcep and placed in an electric dessicator for 7-10 days. Dried filter papers
covered with mycelium and fruiting bodies were kept in the alumnium foil in a sterile plastic bags,
labeled and placed in a box and strorage at - 20o C.

4.4 Grain culture
Grain of barley were autoclaved at 121°C for 15 minutes and placed on PDA in sterile Petri-
dishes. Mycelial puge of endophytic fungi were transferred to PDA for 7-14 days. The plant tissue
with mycelium were transferred to plastic tube by using sterile forcep, labeled and placed in a box and
strorage at -20o C.

4.5 15% glycerol
15 % glyceral in vial were autoclaved at 121°C for 15 minutes. Mycelial puge of endophytic
fungi were transferred to 15% glycerol and kept in a box and strorage at 4o C.
All fungal pure cultures were maintained at Microbiological Resource Centre , Thailand Institute
of Scientific and Technological Research (TISTR), Phathum Thani, Thailand for future use.

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 57

Result and Discussion

NinespeciesofHyphomycetesfungiwerefoundon7diseasedweedsasfollows(Table1,2;Figure1).

Table 1 Weeds species and Locations of Collection.

No. Weeds species Family Location

1 Cyperus rotundus Cyperaceae Bangkok
2 Cyperus brevifolius Cyperaceae Chanthaburi
3 Brachiaria mutica Chanthaburi
4 Eleusine indica Poaceae Pathum Thani
5 Dactyloctenium aegyptium Poaceae Pathum Thani
6 Pennisetum polystachyon Poaceae Pathum Thani
7 Oryzae sativa f. spontanea Poaceae
Poaceae Bangkok

Figure 1 Disease symptoms of weeds A; Cyperus rotundus, B; Dactyloctenium aegyptium and C;
Eleusine indica

Altenaria alternate (Fr.) Keissler
Colonies on PDA attaining 9.0 cm in diam. in 7 days at 28°C, cottony, olivaceous black.
Conidiophore geniculate, smooth 2.5-4.5 x 50 µm. Conidia formed in branched chain, 10-15 x 21-55
µm. producing beak, 2.5-5 µm long. A. alternate has been studied and evaluated the potential use of an
indigenous isolate of as a mycoherbicide to control Lantana camara (Saxena and Pandy, 2002)

Bipolaris bicolor Paul & Parbery
Colonies on PDA attaining 9.0 cm in diam. in 7 days at 28°C, hairy, dark brown. Conidiophore
geniculate, smooth 2.5-5 x 54 µm. Conidia strain, cylindrical 18-20 x 55-120 µm., with 7-9 pseudoseptate,
hilum flat dark. B. bicolor has been reported the application of to control Johnson grass (Sorghum
helepense) Bonilla et al. (1999).

Curvularia intermedia Boedijn
Colonies on PDA attaining 8.5 cm in diam. in 7 days at 28°Cbrown, cottony. Conidiophore
straight, smooth 4.7-9.0 x 25-754 µm. Conidia strain or curve, 3 distoseptate, 13-20 x 25-35 µm. C.
intermedia isolated from diseased crabgrass (Digitaria sp.) has been reported the potential use as
microherbicide to control large crabgrass (D. sanguinalis) (Tilley and Walker, 2002).

Curvularia pallescens Boedijn
Colonies on PDA attaining 8.0 cm in diam. in 7 days at 28°C grey. Conidiophore geniculate,

5.5-6.0-9.0 x 85-180 µm. Conidia strain or curve, 3 distoseptate, slightly curve, ellipsoidal, pale to
brown, smooth, 7.2-11.5 x 17-33 µm.
C. pallescens isolated from disease leaves of Cyperus rotundus, Dactylotenium aegyptium,
Echinochloa colona, Eleusine indica, Digitaria ciliaris, Brachairia reptans, Rottboellia cochinchinensis
and Pennisetum polystachyon has showen the symptom very similar to our study. This fungus has been
reported to produce Endo-1,4D-glucanase (Kokaew 2005).

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 58

Drechslera holmii (Luttrell) Subran. & Jain
Colonies on PDA attaining 9.0 cm in diam. in 7 days at 28°C velvety. Conidiophore geniculate,

dark brown, 5.5-10.0 x 185-250 µm. Conidia obclavate and rotrate, with 6-11 pseudosepta, end cell
pall and cut off by dark thick septa, intermediate cell, golden brown, smooth, 20-31 x 60-125µm, hilum
protuberant, dark. Meanwhile, a study of Ellis (1971) reported that D. holmii causing reddish-brown
spot and strips on leaves of Dactyloctenium sp. in USA.

Exserohilum rostratum (Drechsler) Leonard & Suggs
Colonies on PDA attaining 9.0 cm in diam. in 7 days at 28°C velvety. Conidiophore geniculate,

brown, 5.5-7.2 x 125-200 µm. Conidia slightly cuve, obclavate, rotrate, with 4-12 pseudosepta, smooth,
14-22 x 40-180 µm, hilum distinctly protuberant, bipolar germination. This fungus was successfully
applied as foliar spay to control seven weedy grasses in citrus orchard (Chandramohan and Charudattan
2001).

Fusarium oxysporum Schlecht
Colonies on PDA attaining 9.0 cm in diam. in 7 days at 28°C cottony. Conidia of two types,

microconidia 1- celled, hyaline, 2.5-3.0 x 5.5-12.0 lightly cuve, obclavate, rotrate, with 4-12 µm and
macroconidia 3 to many septate, fusiform to falcate, hyaline, 3.2-5.5 x 21.0-60.0 µm. F. oxysporum
has been reporeted as mycoherbicide to control Striga hermonthica in Wastern Africa (Clotala et al.
1995).

Table 2 Micro fungi isolated from weed diseases at different locations.

Fungi Class Host plant

Alternaria alternate Hyphomycetes 1,2

Bipolaris bicolor Hyphomycetes 1,3

Curvularia intermedia Hyphomycetes 1,3,4

Curvularia pallescens Hyphomycetes 1,2,4,6

Drechslera holmii Hyphomycetes 5

Exserohilum rostratum Hyphomycetes 1,2,5,6

Fusarium oxysporum Hyphomycetes 2,3,6

Myrothecium verrucaria Hyphomycetes 2,3,4

Nigrospora oryzae Hyphomycetes 1,2,3

* 1= Cyperus rotundus, 2 = Cyperus brevifolius, 3= Brachiaria mutica, 4= Eleusine indica,
5=Dactyloctenium aegyptium, 6= Pennisetum polystachyon

Myrothecium vurrucaria (Alb. & Schw.) Ditm. Ex fr.

Colonies on PDA attaining 9.0 cm in diam. in 7 days at 28°C Sporodochia, green to black
surrounded by a zone of white. Conidia in dark green slimy masses, cylindrical, navicular to ellipsoidal,
hyaline, smooth, 2.0-4.5 x 6-10 µm.

Boyett et al. (2002) reported the application of M. verrucaria to control Kudzu (Pueraria
lobata). Secondary metabolites produced by this fungus such as Verrucaria A,B,C,D,E,F,G,J; Roridins
A,B,D,E,H; Muconomycin; Coprogen B; Gliotoxin and Antifungal

Nigrospora oryzae Hudson
Colonies on PDA attaining 9.0 cm in diam. in 7 days at 28°C at first white, turned to grey. Conidia

broadly ellipsoidal, black, 10-15 µm in diam. A report of Kokaew (2005) that isolated C. pallescens from
disease leaves of Cyperus rotundus, Dactylotenium aegyptium, Echinochloa colona, Eleusine indica,
Digitaria ciliaris, Brachairia reptans, Rottboellia cochinchinensis and Pennisetum polystachyon
which was very similar to the present study. This fungus produced Verrucaria A,B,C,D,E,F,G,J;
Muconomycin; Coprogen B; Gliotoxin

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 59

Conclusion

Atotalof20isolatesofHyphomycetesfungifoundondiseasedweedcollectedfromvegetableplotsin
Bangkok, Pathum Thani and Chanthaburi provinces were common leaf spot and leaf blight diseases. There
were Alternaria alternate, Bipolaris bicolor, Curvularia intermedoia, Curvularia pallescens, Drechslera
holmii, Exserohilum rostratum, Fusarium oxysporum, Myrothecium verrucaria, Nigrospora oryzae.

Acknowledgment

We would like to thank the authorities of Faculty of Biotechnology, Rangsit University and
Thailand Institute of Scientific and Technological Research for extending facilities on this research.

Reference

Auld, B.A., Mc Rae, C., 1999. Emerging technologies in plant protection-bioherbicides. In: Proceedings
of New Zealand Plant Protection Society (NZPPS), New Zealand, pp. 1–4.

Babu, R.M. , A. Sajeena and K. Seetharaman. 2003. Bioassay of the potentiality of Alternaria
alternata (Fr.) Keissler as a bioherbicide to control waterhyacinth and other aquatic weeds. Crop
Protection. 22(8): 1005 – 1013

Bonilla, T., M.O. Lopez, J. Mena, K. Rodriguez, E. Perez and Tomas. 1999. Mycobiota of Sorghum
halepense (L) Pres. And evaluation of Revista de Protection Vegetal. 14 (1); 65-68.

Boyette, C.D., H.L. Walker and H.K. Abbas. 2002. Biological control of Kudzu (Pueraria lobata) with
and isolate of Myrothecium verrucaria. Biologicalcontrol Science. Technology. 12: 75-82.

Chandramohan, S. and R. Charudattan. 2001. Control of seven grasses with a mixture of three fungal
pathogens with restricted host ranges. Biological Control. 22: 246-255.

Clotola, M., A.K. Watson and S.G. Hallett. 1995. Discovery of an isolate of Fusarium oxysporum with
potential to control Striga hermonthica in Africa. Weed Research. 35: 303-309.

Daniel, J.T., Templeton, G.E., Smith Jr., R.J., Fox, W.T., 1973. Biological control of northern joint vetch
in rice with an endemic fungal disease. Weed Sci. 21, 303–307.

De Hoog G.S., Guarro J., Gene J., Figueras M.J. (2000) Atlas of Clinical Fungi. 2 ed. Centraalbureau
Voor Schimmelcultures, Utrecht. 1126 p.

Domsch, K. H., W. Gams and T.H. Anderson. 1993. Compendium of Soil Fungi. Vol. 1, 2 ed. Academic
Press, London. 859, 405 p.

Ellis, M.B. 1971. Dematiaceous Hyphomycetes. Commonwealth Mycological Instritute, Kew, Surrey.
608 p.

Kokaew, J. 2005. Diversity of fungi on major weed diseases in vegetable graden plots and their
potential uses as biological weed control. M.S. thesis. Kasetsart University. 150 p.

Kokaew, J., L. Manoch, N. Visarathanon and D. Suwanagul. 2006. Diseases of Weed in Vegetable
Garden Plots and Their Potential Use for Their Biological Weed Control. In Abstracts Book of
the 8th International Mycological congress. Cairns Convention Centre Queensland, Australia.
21-25 August 2006.

Manoch, L., O. Jeamjitt, T. Dethoup and P. Athipunyakom. 2004. Spoilage fungi on fruit, vegetable
and food commodity. In Proceedings of the 1st KMITL International Conference on Integration
of Science & Technology for Sustainable Development. Bangkok. Thailand. 25-26 August 2004.
Vol.2: 438-441.Moss, M.O. 2003. Mycotoxin review-3. Houses and pasture. Mycologist 17(2):
79-83.

Nikonov, I.N., Yu.S. Osledkin and V.I. Safronova. 2007. The search of producers of laccase among
filamentous fungi from the all-Russia research institute for agricultural microbiology culture
collection. p. 198-199. In Abstracts of The XV Congress of European Mycologists. St. Petersburg,
Russia16-21 September. 2007.

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 60

Saxena, S. and A.K. Pandy. 2002. Evolution of indigenous isolate of Alternaria alternate (LC≠508)
for use as a mycoherbicide for Lantana camara L. Crop Protection. 21(1): 71-73.

Simmons, E.G. 2004. Novel dematiaceous hyphomycetes. Studies in Mycology 50: 109-118.
Sin, M.K.W., K.D. Hyde and S.B. Pointing. 2002. Comparative enzyme production by fungi from

diverse lignocellulosic substrates. Journal of Microbiology. 40(3): 241-244.
Sivanesan, A. 1987. Graminicolous species of Bipolaris, Curvularia, Drechslera, Exserohilum and

Their Teleomorphs. CAB. International Mycological Institute.
Wallingford. Szewczuk, V., W. Kita, B. Jarosz, W. Truszkowska, Prof. Dr. A. Siewinski 1991. Growth

inhibition of some phytopathogenic fungi by organic extracts from Nigrospora oryzae (Berkeley
and Broome) Petch. Journal of Basic Microbiology 31(1): 69-73.
Tanaka M., Fukushima T., Tsujino Y., Fujimori T. (1997) Nigrosporins A and B, new phytotoxic and
antibacterial metabolites produced by a fungus Nigrospora oryzae. Bioscience biotechnology and
biochemistry 61(11): 1848-1852.
Tilley, A.M. and H.L. Walker. 2002. Evaluation of Curvularia intermedia (Cochiobolus intermedius)
as a potential microbial herbicide for large crabgrass (Digitaria sanguinalis). Biological and
Chamical Diversity. Pure. Appl. Chem. 70 (11). www.iupac.org.
Varma G.B., M.O. Fatope, R.G. Marwah, M.E. Deadman and F.K. Al-Rawahi. 2006. Production of
phenylacetic acid derivatives and 4-epiradicinol in culture by Curvularia lunata. Phytochemistry
67(17):1925-1930.
Yoganathan,K., C. Rossant, S. Ng,Y. Huang, M.S. Butler andA.D. Buss. 2003. 10- Methoxydihydrofuscin,
fuscinarin, and fuscin, novel antagonists of the human CCR5 receptor from Oidiodendron griseum.
Journal of Natural Product 66(8): 1116-1117.

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 61

Impacts of meadowfoam seed meal amendment on weeds
and soil microbial activity

Suphannika Intanon1, Andrew Hulting1, David Myrold1, and Carol Mallory-Smith1
1 Department of Crop and Soil Science, Oregon State University.109 Crop Science Building, Corvallis,

Oregon 97331, USA.
[email protected]

Abstract

Meadowfoam (LimnanthusalbaHartw. exBenth) seed meal, a by-product of meadowfoam oil
extraction, has glucosinolate degradation compounds that are similar to those from Brassicaceae. The
compounds are reported to be herbicidal. Two field studies were conducted to evaluate the application
of meadowfoam seed meal for weed control in lettuce and the effect of meadowfoam seed meal on soil
microbial activity. Meadowfoam seed meal was applied either as 2.86 kg m-2 on day 0 or as 1.43 kg m-2
on day 0 followed by 1.43 kg m-2 on day 7. To account for the fertilizer effect of the seed meal, urea
was used as a nitrogen source and applied eitheras 16.8 g m-2 on day 0 or as 8.4 g m-2 on day 0 followed
by 8.4 g m-2 on day 7. Meadowfoam seed meal treatment suppressed weed emergence and growth.
A split application of meadowfoam seed meal provided the best control of spiny sowthistle. Lettuce
aboveground biomass was similar between urea and meadowfoam seed meal amended treatments. Soil
microbial activity was greater in meadowfoam seed meal treatmentscompared to either urea or non-
amended plots. Both fertilizer and bioherbicide effects were found with the use of meadowfoam seed
meal.

Keywords: meadowfoam seed meal, LimnanthusalbaHartw. exBenth,glucosinolate, soil amendment

Introduction

Meadowfoam (LimnanthusalbaHartw. ex Benth.) is known as an industrial oil seed crop. It is
a winter annual crop in the Limnanthaceae family, which is native to southern Oregon and northern
California. It is used as a winter rotation crop in grass seed production systems in the Pacific Northwest.
The oil extracted from meadowfoam seed possesses a unique oxidative stability that makes it useful in
a wide range of cosmetic and personal care formulations. About 70% of the extracted seed remains after
oil extraction. At present, this by-product, known as meadowfoam seed meal (MSM), has little value.
Interestingly, MSM has potentially herbicidal glucosinolate degradation compounds that are
similar to those from Brassicaceae species. Previous research suggests that the effectiveness of MSM as
a soil amendment depends on the concentration applied. Low levels of MSM may be a growth stimulant
for vegetable crops (Vaughn et al. 2008). At greater concentrations, MSM has been found to inhibit seed
germination and possess herbicidal properties (Lindermanet al. 2007; Machado, 2007; Stevens et al.
2009).
A greenhouse study confirmed the herbicidal effect of MSM on seeding emergence and growth
compared to the control (Intanonet al. 2011). A field study suggested that the meadowfoam seed meal
has both fertilizer and bioherbicide effects (Intanonet al. 2012). However, bioherbicide effects were
much less than those observed in the greenhouse at the same concentration.The fertilizer effect was
observed when MSM was applied at 1.22 kg m-2 and at 2.04 kg m-2, whereas there was a bioherbicide
effect observed at 2.86 kg m-2. Although concentration seems to be a key factor in the application
of MSM for weed control,with respect to developing MSM as a bioherbicide, the application of an
effective concentration and the method of the meal used under field condition need to be investigated.
In addition to the investigation of the bioherbicide effect, theimpact of MSM amendment on
microbial activity was included in this study. Soil microorganisms, mainly bacteria and fungi, play
important roles in nutrients availabilityfor plants (Wardle and Ghani, 1995). The variability in a microbial

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 62

community can be used to indicate the change in soil quality (Breure, 2005).However, there are no data
on soil microbial activity changes related to MSM application so the effects of soil amendment with
MSM on microbial activity need to be identified.
The goal of this study was to evaluate the application of MSM for weed control in transplanted
lettuce. The specific objectives were:1) to determine the crop yield and additional nutrients from MSM
application, 2)to investigate the effect of MSM application on weed emergence and growth, 3) to
determinethe effect of meadowfoam seed meal on soil microbial activity via basal respiration, and 4)
to evaluate the relationship between methods of MSM application, plant growth, and soil microbial
activity.


Material and methods
Soil
A field study was conductedon a Lewis-Brown Horticulture Research Farm, Oregon State
University, Oregon, USA. The soil was a mollisol classified as a Malabonsilty clay loam.

Field study
In the summer of 2012, two field experiments were conducted from July 9 to September 12 for
the first experiment and from August 1 to October 5 for the second experiment using a randomized
complete block design with four replications. There were five treatments which consisted of two
amendment materials (urea and activated MSM) with two application methods (either full or split rate
application) and one control treatment (non-amended treatment). Meadowfoam seed meal was passed
through a 1 mm-sieve before use. Activated MSM consisted of 1% ground meadowfoam seed and 99%
seed meal by weight. Activated MSM was applied either as 2.86 kg m-2 on day 0 or as 1.43 kg m-2 on
day 0 followed by 1.43 kg m-2 on day 7. To account for the fertilizer effect of the seed meal, urea was
used as a inorganic nitrogen source and applied eitheras 16.8 g m-2 on day 0 or as 8.4 g m-2 on day 0
followed by 8.4 g m-2 on day 7.The treatment plot size was 1.6 m2 with 1.07 m border between plots and
1.07 m border around the entire site. All plots were cleared of weeds before starting the experiments.
A hundred seeds of Sonchusasper (spiny sowthistle) and of Echinochloafrumentacea (Japanese millet)
were sown in an evenly spaced rowin each plot right after meal incorporation. Plots were irrigated using
sprinkler irrigation system at an average application rate of 254 mm hr-1.Nine lettuce seedlings (18 days
old) were transplanted in the middle row of the plot at7 days after incorporation (DAI). Lettuce was
transplanted at 15 cm in-row-spacing. On 35 DAI, seven lettuce plantswere harvestedfor aboveground
biomass. Seedlings of S. asper and E.frumentaceawere counted and then harvested for aboveground
biomass. Weed sample area was 1.14 m2 in each plot. Other emerged weeds wereseparated by species,
counted,and harvested for aboveground biomass on35 DAI and re-harvested on 65 DAI. Lettuce and
weeds were dried at 60 C for 72 hr, and weighed.Dry lettuce tissue was used to measuretotal nitrogen
(N)and sulfur(S)by CNS-Analyzer (Leco CNS-2000).

Soil measurement
Soil samples were taken in each plot to 5-cm depth within an areaof 0.46 m2to studysoil microbial
activity. The samples were conducted on 0, 7, 14, and 28 DAI. Three soil cores were sampled in each plot
and composited. The moist soil was passed through a 3-mm sieve and incubatedin the dark for 48 hr at
25 C before monitoring the total CO2basal respiration by using Isotopic CO2Analyser(PicarroG2101).

Data analysis
The aboveground biomass, emergence, total NS, and basal respiration data for each treatment
were analyzed using ANOVA. Means were separated by a least significant difference (LSD) test using
PROC GLM in SAS v. 9.2.

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 63

Results

Effect of soil amended treatments on lettuce growth and plant nutrient availability
Total lettuce biomass in Experiment 1 was greater than Experiment 2 (Table 1). The total plant
sulfur in MSM-amended treatments was 24% and 38% greater in MSM amended treatments compared
to non-amended and urea-amended treatments in Experiment 1 and 2, respectively. In Experiment 1,
total plant nitrogen in amended plots with urea and MSM was 49% greater than non-amended plots.
In Experiment 2, total plant nitrogen in urea-amended treatmentswas not different from non-amended
treatment andthe MSM amended treatments had 34% greater plant-available nitrogen compared to non-
amended and urea-amended treatments.

Table 1. Drybiomass of seven lettuce plants and chemical analysis per gram of lettuce shoot biomass
grown in the middle row of sample plots for 18 days in the greenhouse and 35 days in the field with or
without amended materials.Data are represented as means with SE within parentheses. Different letters
within a column indicate significant differences at the 0.05 levelwithin an experiment.

Expt Trta Lettuce biomassb (g) Total sulfur (g) Total nitrogen (g)
1 NC 33.6 (2.74) nsc 1.0 (0.11) b
CU 47.2 (4.44) 0.068 (0.0043) b+ 2.1 (0.18) a
CR 41.8 (3.06) 2.0 (0.13) a
MS 37.2 (3.89) 0.092 (0.0101) ab 1.9 (0.15) a
MR 36.3 (5.16) 1.8 (0.27) a
0.093 (0.0073) ab

0.112 (0.0092) a

0.110 (0.0164) a

2 NC 25.5 (1.92) a 0.061 (0.0046) bc 1.0 (0.09) b

CU 24.6 (2.55) a 0.058 (0.0074) c 1.3 (0.03) ab

CR 25.9 (0.77) a 0.056 (0.0043) c 1.1 (0.05) b

MS 36.9 (5.21) b 0.099 (0.0159) ab 1.7 (0.28) a

MR 32.4 (2.40) ab 0.091 (0.0072) a 1.7 (0.12) a

atreatments: NC, non-amended; CU, full rate of urea amendment; CR, split rate of urea amendment;
MS, full rate ofmeadowfoam seed meal amendment and MR, split rate of meadowfoam amendment
baverage dry biomass for seven lettuce plants (n=4)
cnot significant
+significant difference at 0.01 level

Effect of soil amended treatments on weed emergence and growth
The effect of treatments on the total emergence and dry biomass of natural occurring weeds, S.
asper, and E.frumentaceadiffered across treatment (Table 2 and 3; Fig. 1). At 35 DAI, total dry biomass
of naturally occurring weeds in MSM-amended treatments was reduced more than 85% for Experiment
1 and more than 74% for Experiment 2 (Table 2). There was no difference in total dry biomass of
naturally occurring weeds harvested at 65 DAI. However, the biomass from Experiment 1 (on average
0.91 g m-2) was much less than those from Experiment 2 (on average 26.72 g m-2). In both experiments,
dry weight per plant of E.frumentacea was not different across treatments (on average of 0.82 g plant-1
for Experiment 1 and 0.66 g plant-1 for Experiment 2). In Experiment 1, a full rate application of MSM
treatment prevented S. asper germination, whereas in Experiment 2, MSM treatments of both full and
split rate applications preventedS. asper germination.
In general, the emergence of naturally occurring weedson 35 and 65 DAI for Experiment 1 was less
than Experiment 2 and the emergence of naturally occurring weeds, S. asper , and E. frumentaceawas
reduced in both MSM treatments (Table 3). On 35 DAI, MSM treatments provided 89% and 85%
suppression of weed emergence when compared to urea-amended and non-amended treatments for

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 64

Table 2. Aboveground biomass of naturally occurring weeds, Sonchusasper and Echinochloa
frumentacea harvested on different days after meal or urea incorporation (DAI). Data are represented as
means with SE within parentheses. Different letters within a column indicate significant differences at
the 0.05 levelwithin an experiment.

Expt Trta DW naturally DW naturally S. asper E. frumentacea
occurring weeds occurring weeds (g plant-1) (g plant-1)
1
NC (g m-2) (g m-2) 35 DAI ab 35 DAI ns
CU 35 DAI 65 DAI 0.13 (0.007) ab 0.48 (0.052)
CR 0.44 (0.079) nsb 0.14 (0.012) a 0.98 (0.066)
MS 41.0 (10.79) b 0.15 (0.017) b 0.97 (0.077)
MR 2.20 (0.933) 0.08 (0.046) c 0.60 (0.209)
83.1 (9.45) a 1.06 (0.715)
0.64 (0.238) 0
55.9 (13.21) b
0.17 (0.087)
6.3 (0.21) c
1.08 (0.986)
4.8 (0.32) c

2 35 DAI 65 DAI 35 DAI 35 DAI
0.54 (0.195)
NC 66.5 (13.74) a 29.6 (1.33) ns 0.07 (0.011) a 0.36 (0.023) ns
0.54 (0.087)
CU 60.6 (6.66) ab 30.7 (6.28) 0.07 (0.010) a 0.91 (0.174)
0.97 (0.253)
CR 40.1 (2.96) b 27.7 (3.80) 0.07 (0.018) a

MS 9.8 (2.02) c 20.0 (7.88) 0 b

MR 10.3 (0.87) c 25.6 (3.93) 0 b

a treatments: NC=non-amended, CU=full rate of urea amendment, CR=split rate of urea amendment,
MS=full rate of meadowfoam seed meal amendment, MR=split rate of meadowfoam amendment

b not significant

Table 3.Total emergence of naturally occurring weeds, Sonchusasper and Echinochloa
frumentaceaharvested at different days after meal or urea incorporation (DAI). Data are represented as
means with SE within parentheses. Different letters within a column indicate significant differences at
the 0.05 levelwithin an experiment.

Expt Trta Total emergence Total emergence S. asper emergence E. frumentaceaemergence

1 naturally occurring naturally occurring (% of sown seeds)b (% of sown seeds)b
NC weeds (seedlings m-2) weeds (seedlings m-2)
CU
CR 35 DAI a 65 DAI ab+ 35 DAI a 35 DAI a
MS 150.9 (29.6) a 4.8 (0.6) a 25.8 (2.84) ab 11.8 (2.14) a
MR 173.9 (22.1) a 8.8 (2.3) ab 24.0 (2.12) b 12.3 (1.11) a
182.7 (39.3) b 7.2 (1.1) b 19.5 (3.01) c 9.5 (1.55) b
23.2 (5.0) b 2.0 (0.9) b 1.3 (0.95) c 3.0 (1.08) b
13.2 (1.4) 2.9 (2.3) 1.3 (0.48)
0

2 35 DAI 65 DAI a 35 DAI a 35 DAI ab
NC 308.8 (23.5) a 229.2 (32.4) a 29.0 (3.29) a 10.3 (2.36) a
CU 276.3 (38.5) a 212.1 (13.4) b 26.8 (3.64) a 10.8 (1.89) ab
CR 166.4 (6.6) b 160.5 (6.3) c 20.8 (3.61) b 8.5 (2.63) bc
MS 38.6 (7.2) c 49.8 (10.4) c b 3.8 (1.89) c
MR 37.9 (3.1) c 58.1 (5.2) 0 1.8 (0.48)
0

a treatments: NC, non-amended; CU, full rate of urea amendment; CR, split rate of urea amendment;
MS, full rate of meadowfoam seed meal amendment and MR, split rate of meadowfoam amendment

baverage percentage of emerged seeds of the total100-sown seeds
+significant difference at 0.01 level

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 65

A)

NC CU CR MS MR
B)

NC CU CR MS MR

Fig. 1. Lettuce growth and weed community across soil amended with different materials on 35 DAI in
A) Experiment 1and B) Experiment 2. NC, non-amended; CU, full rate of urea amendment; CR, split
rate of urea amendment; MS, full rate of meadowfoam seed meal amendment and MR, split rate of
meadowfoam amendment.

Effect of soil amended treatments on total CO2 basal respiration
In general, there were greater total CO2basal respiration in MSM-amended treatments compared
to non-amended and urea-amended treatments (Table 4). Within 28 days, the change of the basal
respiration slightly decreased over time in non-amended and urea-amended treatments but steeply
decreased in MSM-amended treatments. No difference of total CO2basal respiration was detected across
non-amended and urea amended treatments for both experiments. In Experiment 1, the total CO2of
MSM-amended treatments were 98%, 96.4%, 95.6%, and 92.6% greater than those of non-amended
and urea amended treatments on initial, 7, 14, and 28 days, respectively. In Experiment 2, total CO2of
MSM-amended treatments were 97.8%, 97.3%, 96.5%, and 92% greater than those of non-amended
and urea amended treatments on initial, 7, 14, and 28 days, respectively.

Effect of the methods of MSM application on plant growth and soil microbial activity
Based on the results of lettuce growth, weed emergence, and soil microbial activity, the effects
of either a full or a split rate of MSM application treatments on plant growth were not obviously
differentiated. The split MSM-amended treatment provided a better control of S. asper in early summer
than a full rate application (Table 2 and 3).The differences between a full or split rate of MSM application
can be observed in basal respiration across the experiments. In Experiment 1, the soil microbial activity
from a full MSM application was greater than a split MSM application on initial and 7 DAI but less on
14 and 28 DAI. While in Experiment 2, the soil microbial activity from a full MSM application was less
than a split MSM application for all measurement days (Table 4).

Table 4. Total CO2of basal respiration after incubation in the dark for 48 hr at 25 C. Soil sample were
collected at initial, 7, 14, and 28 days after meal or urea incorporation (DAI). Data are represented as
means with SE within parentheses. Different letters within a column indicate significant differences at
the 0.05 levelwithin an experiment.

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 66

Expt Trta Basal respiration (μgCO2-C g-1 soil hr-1)

1 0 DAI 7 DAI 14 DAI 28 DAI
NC
CU 3.7 (0.42) b 2.8 (0.30) c 2.5 (0.13) b 1.4 (0.32) b
CR
MS 3.8 (1.05) b 2.6 (0.30) c 2.6 (0.16) b 2.0 (0.07) b
MR
7.2 (2.02) b 3.6 (0.82) c 2.1 (0.29) b 1.3 (0.19) b

376.2 (92.37) a 96.9 (6.40) a 44.0 (6.44) a 17.0 (1.51) a

118.0 (52.27) b 68.8 (17.43) b 63.1 (18.42) a 25.7 (6.73) a

2 0 DAI 7 DAI 14 DAI 28 DAI

NC 4.6 (0.31) b 2.5 (0.20) c 1.9 (0.26) c 1.7 (0.04) bc

CU 6.3 (1.35) b 2.7 (0.33) c 1.8 (0.30) c 0.8 (0.29) c

CR 5.2 (1.39) b 4.0 (1.02) c 1.8 (0.27) c 1.3 (0.06) c

MS 238.3 (57.97) a 82.4 (12.96) b 37.7 (11.30) b 9.8 (1.39) b

MR 255.2 (44.54) a 142.6 (10.22) a 66.3 (6.90) a 22.2 (5.64) a

a treatments: NC, non-amended; CU, full rate of urea amendment; CR, split rate of urea amendment;
MS, full rate of meadowfoam seed meal amendment and MR, split rate of meadowfoam amendment

Experiment 1 and 2, respectively. On 65 DAI, MSM treatments inhibited 65% and 73% of total weed
emergence for Experiment 1 and 2, respectively. However, there was no difference of total weed
emergence between a full rate and a split rate of the MSM application treatments. When considering
new emerged weedswithin the same MSM-amended treatment, the amount of new weed emergence
on 65 DAI was greater than those on 35 DAI. S. asper emergence was inhibited more than 94% by
MSM-amended treatments, whereas E. frumentaceaemergence was suppressed 80.8% for Experiment
1 and 71.7% for Experiment 2. S. asperwas found in situin Lewis-Brown Horticulture Research Farm,
whereasE.frumentaceawas not. The dry biomass and emergence data of S. asper in Table 2 and 3 did
not include the in situ S. asper in the statistical analyses.

Discussion

The growth of transplanted lettuce plants responded positively to MSM-amended treatments
(Table 1; Fig. 1). It is possible that MSMprovided the essential nutrients to support lettuce growth,
especially plant-available nitrogen and sulfur. MSM treatment appeared to be an organic source of
nitrogen and sulfur. The nitrogen and sulfur were reported as structural elements of MSM based on the
chemical analysis of an allelochemical compound (Stevens et al. 2009). The organic nitrogen supplied
from MSM was available for lettuce onamount similar and/or greatercompared to an inorganic fertilizer,
urea. Similar results were found in conifer seedlings which grew faster in amendment of potting medium
with MSM compared to non-amended medium (Lindermanet al. 2007). The conifer seedlings in MSM
treatment had high total nitrogen and lacked nutrient deficiency symptoms, especially phosphorus.
Lettuce growth in early summer (Experiment 1) was greater than late summer (Experiment 2; Table
1), probably because in early summer, the weather was still cool enough to support lettuce growth and
development. In the split MSM treatment of Experiment 2, 44% of lettuce seedlings were replaced after
a week of transplanting. The crop injury would imply to the less benefit of a split MSM application
compared to a full rate application. The greater activity of MSM amendment in a split MSM application
of Experiment 2 compared to those in Experiment 1 was also confirmed in the basal respiration,
especially on the initial and 7 DAI (Table 4).Therefore, crop safety date should be longer than a week
after MSM application when other environment factors influence crop growth and development.
There was less weed pressure in early summer compared to late summer. In Experiment 2, there
was more late summer grass and E.arvense infestation (Fig. 1B). The emergence and growth of weeds
were inhibitedin MSM-amended treatments (Table 2 and 3). The inhibition was better observed in weed

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 67

emergence compared to weed biomass data. It is possible because both fertilizer and bioherbicide effects
were observed with the use of MSM as a soil amendment. The bioherbicide property was detected
mainly at the first harvest (35 DAI). At the second harvest (65 DAI), 30 days after the first harvest
with regular irrigation, a fertilizer effect was found, especially in Experiment 2 (Table 2). Brassicaceae
seed meal as a soil amendment material for weed control also had similar results of co-occurrence of
bioherbicide and fertilizer effects (Jonhson-Maynard et al. 2005). Emergence ofnaturally occurring
weedsfrom MSM-amended treatments in Experiment 2 wasgreater than those from the first harvest
(Table 3). The non-consistent results across the experiments may be due to greater weed pressure in
late summer. According to the planted species, MSM had high level of suppression on emergence
and growth ofS. asperbut not on the emergence and growth of E.frumentacea (Table 2 and 3; Fig.
1). Emerged E.frumentaceaseedlings from MSM-amended treatments were fewer but larger compared
to non-amended and urea-amended treatments because they may take advantage of the addition of
plant-available nutrientsfrom MSM. Although, MSM application did not have obvious selectivity for
a specific weed species, it can generally suppress annual dicotsand monocots butwas less effective on
some perennial plants such as E. arvense (data not shown).
In this study, the effect of MSM on soil microbial activity focused only on total CO2 production
from basal respiration. Basal respiration reflects the availability of carbon for microbial growth and
maintenance and is a measure of the basic turnover rates in soil (Insamet al. 1991). Greater basal
respiration ratein MSM-amended treatments compared to non-amended and urea-amended treatments
was due to organic inputs from MSM (Table 4). The carbon inputs from MSM increased the gross
metabolic activity of mixed microbial population. However, basal respiration aloneis just one measure
of soil microbialactivity. Further investigations on the effects of MSM on soil microbial activity will
involve multi-parameter approaches including microbial enzyme activity and specific carbon substrates
for soil microbial community.
The effect of MSM did not last longer more than a week. The repeat application after one
week was assumed to increase the bioherbicide effect. Jonhson-Maynardet al. (2005) suggested that
the reapplication of seed meal may provide adequate control weeds throughout the growing season
but this can also increase late-season weed biomass due to the increase in plant-available nitrogen.
However, based on the results of this study, there was no clear evidence of the differences between a
full and a split rate of MSMapplication treatments on weed suppression and soil microbial activity.

Conclusions

MSM treatments applied as soil incorporation at 2.86 kg m-2 can inhibit weed emergence and
growth. The split rate application of MSM provided a significant benefit for weed control similar to
the full rate application. Application time of MSM showed the different results on plant and microbial
activity. The early summer application provided better weed control than the late summer application
which waspossibly due to less weed pressure in the early summer. However, the other environmental
conditions (e.g. temperature and soil moisture) may possibly be involved. The effects of MSM treatment
on soil microbial activity and soil quality still need to be determined using multi-parameter approaches
and longer time span. In summary, a single MSM application as a pre-emergence soil amendment
benefits crop yield, weed suppression, and soil carbon inputs.

Acknowledgements

I would like to thank Dr. John Hart for providing helpful suggestionsand assistance on plant
nutrient analysis. I would also like to thank Megan McGinnisfor her advice and support on basal
CO2respiration.

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 68

References

Breure, A. M. 2005. Ecological soil monitoring and quality assessment, in:P. Doelman and H. J.
P. Eijsackers (Eds.). Vital soil, function value and properties. Elsevier, Amsterdam. pp. 281-305.

Insam, H., C. C. Mitchell, and J. F. Dormaar. 1991. Relationship of soil microbial biomass and activity
withfertilization practice and crop yield of three ultisols. Soil Biol.Biochem. 23: 459-464.

Intanon, S., A. Hulting, F. Stevens, J. Kling, R. Reed, and C. Mallory-Smith. 2011. The use of
meadowfoam seed meal as a soil amendment to suppress seedling emergence. Proceedings of
Western Society of Weed Science, 63rd annual meeting. Spokane, WA, March 7-10, 2011.p 29.

Intanon, S., A. Hulting, J. Kling, and C. Mallory-Smith. 2012. Evaluation of Meadowfoam Seed
Meal as a Potential Bioherbicide. Proceedings of the Weed Science Society of America, 52nd
annual meeting. Waikoloa, HI, February 6-9, 2012.

http://wssaabstracts.com/public/9/proceedings.html. Accessed: November 2, 2012.
Johnson-Maynard, J. M.,Morra, L. Dandurand, C. William, and M. J. Butters. 2005. Brassicaceae seed

meal application for weed reduction and improved nitrogen management in organic farming
systems. Project report, University of Idaho, Moscow, ID.
Linderman, R. G., E. A. Davis, and C. J. Masters. 2007. Response of conifer seedlings to potting
medium amendment with meadowfoam seed meal, in:J. Janick and A. Whipkey (Eds.). Issues in
New Crops and New Use. Alexandria, VA: ASHS Press.pp. 138-142.
Machado, S. 2007. Allelopathic potential of various plant species on downy brome: implications for
weed. Agron. J. 99:127–132.
Stevens, J. F., R. L., Reed, S. Alber, L. Pritchett, and S. Machado. 2009. Herbicidal activity of
glucosinolate degradation products in fermented meadowfoam (Limnanthesalba) seed meal. J.
Agric. Food Chem. 57: 1821-1826.
Vaughn, S. F., M. A. Berhow, and B. Tisserat. 2008. Stimulation of plant growth by (3-methoxyphenyl)
acetonitrile applied as a foliar spray in vivo or as a medium amendment in vitro. HortSci. 43:
372-375.
Wardle, D. A. and A. Ghani. 1995. A critique of the microbial metabolic quotient (qCO2) as a bioindicator
of disturbance and ecosystem development. Soil Biol. Biochem. 27: 1601-1610.

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 69

Imidazolinone tolerance variety for weedy rice control in
direct-seeded rice: The Malaysian Experience

Azmi, M1., Yim, K. M2 and George, T. V2.
1 Malaysian Agricultural Research and Development Institute (MARDI), P. O. Box 12301, General Post

Office,50774 Kuala Lumpur, Malaysia. E-mail: [email protected]
2BASF (Malaysia) Sdn Bhd, 2 Jalan U8/87, Bukit Jelutong, 40706 Shah Alam, Selangor, Malaysia.

E-mail:[email protected], [email protected]

Abstract

Weedy rice (Oryza sativa complex) poses the greatest threat to direct-seeded rice (DSR) because
of its taxonomic and physiological similarities to cultivated rice. Farmers in Malaysia have considered
weedy rice as of significant and major importance since there are no selective herbicides available for
weedy rice control prior to the advent of Herbicide Tolerant Rice (HTR). The development of local
imidazolinone tolerant rice (IMI-TR) was through a collaborative project between MARDI and BASF
which started in 2003 at MARDI Experimental Station in Seberang Perai, Penang. IMI-TR Line No.
1770 from United States was crossed with popular local rice cultivar of high yielding variety, MR
220. The goal of the project is to offer farmers a new innovative and effective solution to the weedy
rice problem. After undergoing many trials and breeding selection, by 2008 two potential varieties,
MR 220CL1 and MR 220CL2, were identified. The introduction of these cultivars is justified by the
need to offer a new efficient and innovative alternative approach and method to manage weedy rice in
DSR. The combination of IMI-TR variety with imidazolinone herbicides is known as the Clearfield®
Production System (CPS) and this was launched in Malaysia on 8th July 2010. This was also the
first launch of Clearfield® Production System for rice in the Asia Pacific region. This new technology
is able to effectively control weedy rice that no other herbicides can control in DSR. The use of
the CPS has benefited the rice industry in Malaysia by providing an effective management of weedy
rice alongside with other noxious weeds in rice cultivation. Clearfield® Production System for rice
offers many advantages and benefits namely it helps to simplify the process of weed management and
control the weedy rice with a single OnDutyTM (imazapic/imazapyr) herbicide application. Most of
all, it is an innovative and effective agronomic solution that delivers significant value and benefits to
rice farmers through effective weed control using farmers normal cultural practices, increased yield
potential and improved crop harvest quality. Studies have shown Clearfield® plots yielded 2.5 t/ha more
than conventional plots with correspondingly net income being higher by more than USD900/ha. The
CPS is non-GMO and signifies the beginning of a paradigm shift in modern agriculture for effective
weedy rice and weed control. In 2012 season, about 30% of rice granaries in Malaysia had adopted
this technology. The demand for this technology is expected to be increased from season to season in
the future as shown from farmers acceptance and adoption since the second season of 2010 till now.
In short, the CPS is tested and proven to be technically feasible, economically viable and culturally
acceptable in the Malaysian experience.

Introduction

A major factor that contributes to a higher production cost for rice is weed control. Currently,
rice farmers throughout the world face a unique weed problem. A weedy relative of cultivated rice
known as weedy rice has invaded and severely infested direct-seeded rice (DSR) fields. Weedy rice has
long been a major threat in the DSR culture in Asia, especially in Malaysia. Weedy rice matures earlier
than cultivated rice, shatters and lodges easily. And if they lodge, they also cause the neighbouring
cultivated rice plants to lodge along side. Under moderate weedy rice infestation (15-20 panicles/m2),

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 70

yield loss is approximately 12 to 15%; under high infestation (21 to 30 panicles/m2) yield loss is 15 to
22%; while under heavy infestation (more than 50 panicles/m2) lodging of weedy rice plants may occur
and can cause total yield loss under tropical climatic conditions (Azmi and Karim 2008).

Since there are no selective weedy rice herbicides in the pipeline, an alternative is to explore
herbicide tolerant rice technology. Rice tolerant to imidazolinone herbicides was developed from a
single plant that survived a chemically induced mutation trial in 1993 at Louisiana State University
Agricultural Center (Sanders et al., 1998). The method used for obtaining herbicide-tolerant rice
was chemical mutagenesis with ethyl methane sulfonate. The acetohydroxy acid synthase (AHAS)
enzyme catalyses the first step in the biosynthesis of the branched chain amino acids leucine, isoleucine
and valine in plants. The mutation in AHAS enzyme results in alteration to the binding site for the
imidazolinone class herbicide, therefore increasing the plant’s tolerance to those herbicides. The other
properties of the AHAS enzyme are unaffected. This rice line is considered non transgenic because it
was developed through seed mutagenesis and not through gene transfer. Thus it is a non-Genetically
Modified Organism (GMO) variety. Genetic engineering involves adding a gene from another organism.
The resulting plant is termed GMO, and any subsequent progenies developed from this plant that
possess the introduced traits are GMOs as well. Clearfield rice is not a GMO since it did not contain
any inserted gene from another organism, and maintains pure DNA from rice in its genome structure.
As such, it has been accepted in the world rice market. Clearfield rice (tolerance to imadozolinones)
was registered in 2001 and fully released by BASF in 2002 in USA.

The imidazolinone herbicides were developed in the late 1970s and 1980s for use in a range of crops
including oil palms, rubber, sugarcane, forestry, soy bean, cereals, sunflower, and lentils. Imidazolinone
herbicides include imazapyr, imazapic, imazethapyr, imazamox, imazamethabenz, and imazaquin.
They have a broad spectrum of weed control, controlling both monocotyledonous and dicotyledonous
weeds. These chemicals can be absorbed through both roots and leaves; hence weeds can be controlled
through both the soil and foliar application (Shaner, 1991). They are translocated to the growing points
of plants where they inhibit the enzyme AHAS thereby disrupting protein synthesis and cell growth.

Breeding and Selection
The research to develop imidazolinone-tolerant rice (IMI-TR) lines started in the main season

2003/04 at the MARDI Seberang Perai Experimental Station. Clearfield line No.1770 (from Louisiana
State University, USA) was crossed with local popular variety, MR 220. The cross was made to
incorporate the PW-16 gene (a dominant gene tolerant to imidazolinone herbicides) from the donor
line 1770 to these recurrent parents. The crossed seeds were planted in the off-season 2004. Screening
for herbicide tolerance was not done on the F1 plants. The F1 plants were backcrossed to the recurrent
parents. The seeds harvested were later planted in the main season 2004/05. Following the BASF
Protocol R-50, imazamox was applied at the rate of 70g a.i./ha at 10 days after sowing the B1F1 plants.
The B1F1 plants were screened for imidazolinone tolerance together with the donor and recurrent parents
as checks. The homozygous susceptible plants (about 50% of the population) and the recurrent parents
died at about 15 days after application. The heterozygous tolerant B1F1 plants showed very slight crop
injury while the homozygous donor parent showed no crop injury. The surviving B1F1 seedlings were
later transplanted in a trough at about 30 days after application. At flowering stage, the B1F1 plants
were backcrossed again to the recurrent parent, MR 220. The backcrosses to the recurrent parent was
repeated in the off season 2005. The treatment with imazamox was carried out as in the main season
2004/05. No backcrosses were done in the main season 2005/06. The B3F1 plants were allowed to
grow for seed multiplication. Screening with imazamox proceeded as before.

Individual plant selection on the tolerant plants started on the B3F2 bulk population in the off
season 2006. At this stage, treatment with imazamox at 200 g a.i./ha was applied at 8 days after sowing.
This was done to eliminate both the homozygous susceptible and heterozygous tolerant plants; leaving
behind the homozygous tolerant plants to grow. At maturation stage, 18 plants were selected. The
line selection was carried out until the off season 2007, where 2 lines were selected in the main season
2006/07 and five lines were selected in the off season 2007.

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 71

By the off-season 2007, two potential imidazolinone tolerant lines were selected for purification
and evaluation for yield, agronomic performance, resistance against major pests and diseases as well
as physical and chemical properties of the grain. The lines were further evaluated in main season
2007/08, from which two lines, MR 220CL1 and MR 220CL2, were identified as potential lines. They
were promoted in the off season 2008 for further evaluation under large plot testing in Felcra Padi
Estate, Seberang Perak and Sungai Limau Dalam, MADA District IV, Kedah. The introduction of these
cultivars is justified by the need to offer a new efficient and innovative alternative method to manage
weedy rice in DSR culture. The combination of IMI-TR with imidazolinone herbicides were known
as Clearfield Production System (CPS). Clearfield rice production system was launched in Malaysia in
October 2010 specifically for control of weedy rice in DSR.

Comparisons between Clearfield Production System and major crop establishment methods
The CPS is basically the same as wet seeding technique (Azmi et al. 2011). The field is

thoroughly puddle and leveled before sowing. Sowing can be carried out using motor-blower or line
seeders such as drum seeder or knapsack row seeder. Under water seeding culture, water is retained in
the field for seeding. On the other hand, transplanting requires saturated soil without standing water for
good crop establishment. Other comparisons are in Table 1.

Table 1. Comparisons between Clearfield Production System, mechanical transplanting, water seeding
and wet seeding technique

Clearfield Mechanical Water seeding Wet seeding
Production System transplanting

Optimal field Saturated Saturated 5-10 cm water Saturated
conditions depth
Crop Low incidence of Low incidence of High incidence of High infestation of
establishment golden apple snail weedy rice and golden apple snail weedy rice and low
attack incidence of golden
Cost of golden apple snail apple snail
establishment Low Low
Cost of weed Low High
control
Optimal season Moderate - High Low Low Moderate - High
Equipment
Off season Main season Main season Off season
Weeds associated Transplanter Motor-blower
with planting Motor-blower, Motor-blower, drum
method drum seeders, Broadleaves but seeders, knapsack
knapsack row other weeds i.e. row seeder
Weed control seeder grasses and sedges
depend on time of Broadleaves are A wider range of
A wider range of flooding the dominant weeds i.e. grasses,
weeds i.e. grasses, 2,4-D, Sulfonyl weeds broadleaves and
broadleaves and ureas, pre- sedges
sedges. emergence
herbicide and 2,4-D, Sulfonyl Graminicide, 2,4-D
Total weed roguing of weedy ureas and roguing and sulfonyl urea
control with rice. of weedy rice. product followed by
imidazolinone roguing of weedy
herbicides with rice.
minimum roguing
of weedy rice.

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 72

Large scale evaluation
A large scale evaluation of CPS cultivation (47.62 ha) was carried out in the off season 2010 in

fields seriously affected by weedy rice infestation (>30% infestation level) in the previous season (main
season 2009) in FELCRA Seberang Perak rice estate (Azmi et al. 2012). Weedy rice populations were
effectively controlled in the fields (from >30% to <1% level) resulting in an average net yield increment
of 0.76 ton/ha from 4.93 ton/ha (main season 2009) to 5.69 ton/ha (off season 2010) giving a better B/C
ratio from 2.55 to 3.42. Further analysis of its economic viability was done with sensitivity analysis
which showed that with a corresponding decrease of 10% in revenue, the B/C ratio is 3.08 while a
corresponding increase of 10% in costs, the B/C ratio is 3.11. Thus this showed the CPS technology is
not sensitive or affected by a 10% decrease in revenue or a 10% increase in costs (Table 2).

Table 2. Financial Analysis of Clearfield Production System in Felcra Seberang Perak Rice Estate, off

season 2009

Main Off

Block Area Season* Season Yield Increment Increase In Income
(47.62 ha) (ha) 2009/10 2010**
  (ton/ha) (RM**/ha)

Yield (ton/ha)

T 5A 10.68 4.11 5.43 1.32 659.16

T 5B 10.88 3.84 4.61 0.77 384.51

T 6A 14.76 5.61 6.51 0.90 449.43

T 6B 11.30 6.14 6.20 0.06 29.96

Av/ha   4.93 5.69 0.76 380.76

B/C ratio   2.55 3.42

B/C ratio*   2.29 3.08 * Sensivity analysis - return 10%

B/C ratio**   2.32 3.11 ** Sensitivity analysis- cost increase
10%

* Felcra management, ** Clearfield Production System; ***1 USD = RM3.00

Comparison between Clearfield Production System versus Conventional System
A preliminary economic sample survey was carried out on farmers planting non-CPS for season

1/2012 versus CPS for season 2/2012 on their same plots over two seasons. Each respondent or farmer
is asked in questionnaire form to compare of before CPS adoption (conventional system) and after CPS
adoption on the same rice plot of cultivation. The result showed that the costs of weeds management
of conventional system per ha was RM 857 which was about 10% higher than CPS of RM 782 mainly
due to the more usage and cost of herbicides applied namely for grasses, broadleaf and sedges control
with generally two rounds of application. On the other hand, for CPS only one single application of
OnDutyTM without any other herbicides was sufficient to control most of the weed species. The notable
and important point here is that the conventional system cannot control weedy rice infestation causing
it to increase its infestation rate over each subsequent season of planting rice but with CPS, the weedy
rice is effectively contained and controlled. Further analysis showed that for half of the respondents, the
costs and frequency of pests and diseases control is lower for CPS as compared to conventional system
i.e. three rounds of application averaging RM 140 as compared to six rounds of application of RM 520.
This invariably showed that CPS is more tolerant to pests and diseases in the fields resulting in lower
costs and number of chemical applications while boosting farmers’ confidence in the product. Next
when roguing costs is factored in, the CPS clearly showed a mark and significant reduction in weedy

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 73

rice costs control. In conventional plots, all the farmers’ results indicated that roguing need to be carried
out two rounds totalling RM 500 per ha plot while for CPS plots, minimum roguing is required. The
bottom line is of course the yield comparison with conventional plots averaging 6.9 ton/ha and when
CPS was adopted, the yield increase was 7.9 ton/ha which is about 15% higher. This directly gives a net
yield increase of one ton/ha amounting to an additional RM 1,250 to the farmers’ income per ha. The
survey was conducted in two sample granary areas with but nevertheless many farmers in more severe
weedy rice infested areas do get yield increases of more than two to five ton per ha.

Conclusion

The use of the CPS has benefited the rice industry in Malaysia by providing an effective management
of weedy rice alongside with other noxious weeds in rice cultivation. Clearfield® Production System
for rice offers many advantages and benefits namely it helps to substantially reduce the cost of weed
management and control the weedy rice with a single OnDutyTM (imazapic/imazapyr) herbicide
application. Most of all, it is an innovative and effective agronomic solution that delivers significant
value and benefits to rice farmers through effective weed control, reduced cost of production, increased
yield potential and improved crop harvest quality. Studies have shown Clearfield® plots yielded 2.5 ton/
ha more than conventional plots with correspondingly net income being higher by more than USD900/
ha. In 2012 season, about 30% of rice granaries in Malaysia adopted this technology. The demand for
this technology is expected to be increased from season to season. In short, the CPS is tested and proven
to be technically feasible, economically viable and culturally acceptable in the Malaysian experience.

References

Azmi, M., Azlan, S., Chew, S. E., George, T. V., Lim, F. W., Hadzim, K. dan Yim, K. M. (2012).
Clearfield Production System for Weed Rice Control in Direct-Seeded Rice (in Malay), MARDI
Report N0. 214 (2012): 15 pp.

Azmi, M, Azlan, A. George, T.V., Chew, S. E. and Yim, K. (2011). Control of weedy rice in direct-
seeded rice using the Clearfield Production System. Proceedings of Asian-Pacific Weed Science
Society Conference, 26-29 Sept. 2011, Cairns, Queensland, Australia. Pp. 50-54.

Azmi, M. and Rezaul, K. (2008). Weedy Rice - Biology, Ecology and Management. MARDI Publication.
p. 56.

Sanders, D. E., Strahan, R. E. Linscombe, S. D. And Croughan, T. P. (1998). Control of red rice (Oryza
sativa) in imidazolinone tolerant rice. Proc. South. Weed Sci. Soc. 51, 36-37.

Shaner, D. L. (1991). Imidazolinone herbicides. Pesticide Outlook. 2 (4): 21-24.

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 74

Phylogenetic relationships of Echinochloa species
based on phenotypic and SSRs markers

Eun-Jeong Lee, Min-Jung Yook, Do-Soon Kim*
Department of Plant Science, Seoul National University, Seoul 151-742, Korea

*Corresponding author: [email protected]

Abstract

Echinochloa species are one of the most important weeds, causing many troubles in rice
cultivation, and difficult to distinguish among species due to their morphological similarity. Therefore,
this study was conducted to find SSR markers and morphological traits that can be used for identification
and classification of Echinochloa species. The relationships among 77 different Echinochloa accessions
including 57 genotypes from Korea and 5 reference species were studied by applying 23 SSR markers
derived from allied species and assessing the 14 morphological traits. E. oryzicola accessions were clearly
clustered as a distinctive group from E. crus-galli and other Echinochloa species in both SSRs analysis
and morphological trait analysis. All E.crus-galli varieties were clustered in same position together, and
we could not find the differences among these varieties. Among 23 SSR markers, we also found 5 SSR
makers that could exactly discriminate E. oryzicola from E. crus-galli and other Echinochloa species.
Although no clear consensus between the results from SSR marker and morphological trait analyses
was founded in this study, our results indicate that both SSR markers and morphological traits can be
useful tools to distinguish among Echinochloa species.

Keywords: Echinochloa, Morphological analysis, SSRs analysis

Introduction

The genus Echinochloa (Poaceae) has about 20-50 annual and perennial species that can be
found throughout the world and some of them are listed most troublesome weeds in the world (Holm et
al, 1977). As rice (Oryza sativa L.) is the main food crop, E. crus-galli (L.) Beauv. (2n=6x=54) and E.
oryzicola Vasing. (2n=4x=36), which spread widely in both dry or water flooded soil, are the greatest yield
limiting and economical loss making weeds in Korea (Chung, 2001). From a taxonomical perspective,
there are many limitations in classification of Echinochloa species because these two species are hard
to distinguish as they show morphologic variation and some morphological trait frequently overlap
among and within species (Yabuno, 1966). Meanwhile, different morphological trait can reflect different
competition ability to crop and response to herbicide. So, it is critical to determination of the level of
variation within species for effective control strategies. In this paper, SSR analysis and morphological
analysis were conducted to assess the polymorphism of intraspecific variation and provide phylogenic
relation of interspecies. In addition, we also expect to make and find genetic and morphological markers
that related each other and can distinguish interspecies clearly from this study.

Materials and Methods

77 Echinochloa accessions were collected in different site (mostly in Korea region) or supplied
by Herbiseed (www.herbiseed.com). Seeds were germinated in round Petri dishes and seedlings were
transplanted in pots located in the glasshouse. Genomic DNA was extracted from young fresh leaves
of individual plants according to serial steps of Sneller et al (1997). DNA were amplified with 23 SSR
markers previous designed for its relative species such as pearl millet, maize, sugar cane, sorghum,
foxtail millet, Echinochloa and Miscanthus. Data cluster was conducted for SSRs analysis with

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 75

NTSYS-pc 2.1 software (Rohlf, 2000) using Jaccard’s coefficients in order to make an unweighted
pair-group (UPGMA) dendrograms. Fourteen morphological characteristics were examined for each
population. The characters used on leaf, stem, spikelet, awn and panicle. Moreover, 10 agronomic
traits also examined in this study such as growth traits and growth date. Data cluster was conducted for
morphological trait analysis with NTSYS-pc 2.1 software (Rohlf, 2000) using correlation coefficients
in order to make an unweighted pair-group (UPGMA) dendrograms.

Results and Discussion

SSRs analysis
Among 79 scorable alleles, 68 had a polymorphism with 86% of polymorphism percentage. The

polymorphism information content (PIC) was obtained for each marker. It varied from a minimum of 0
for CORN05 to a maximum of 0.81 for PSMP2235. Especially, SSR markers designed from sugarcane,
pearl millet and Echinochloa species had relatively high PIC value than others. Therefore, these markers
might be good candidates for getting polymorphism information.

The cluster analysis using UPGMA methods revealed mainly divided 4 groups (I, II, III, and
IV) as marked in Figure 1. First of all, reference species were clearly separated from Echinochloa
accessions by developing group IV. Group I contained most of E. oryzicola significantly distinguished
from E. crus-galli Group (II and III). Group II mainly composed E. crus-galli accessions. Other E. crus-
galli and Echinochloa species created Group III together. In this study, all E. crus-galli varieties were
clustered in same position, Group III, and we could not find the differences among these varieties. In
addition, unclassified Echinochloa accessions (from Taiwan, Vietnam and Sri Lanka) might be estimated
to be classified in E. crus-galli.

As compared electrophoresis gel image and phylogenic tree from SSRs analysis, we found 5
SSR markers that could discriminate E. oryzicola against E. crus-galli and other Echinochloa species. It
was considered that these 5 SSR markers mainly contributed to making such dendrogram of Figure 1.

Morphological trait analysis
In morphological traits analysis, only 77 Echinochloa accessions were evaluated besides

reference species. The cluster analysis using UPGMA methods showed mainly divided 3 groups (I, II
and III) with similar pattern to SSR analysis. In Group I, most of E. oryzicola and 1 E. crus-galli var.
crus-galli were included in it. Group II composed 25 of E. crus-galli var. crus-galli, two of E. crus-galli
var. practicola, one of E. crus-galli var. formosensis, 4 of unclassified Echinochloa accessions and 3
of E.oryzicola. One remained E. crus-galli var. crus-galli and other Echinochloa species were together
in Group III. E. crus-galli varieties and unclassified Echinochloa has the same tendency with those of
SSRs analysis.

The most influential morphological traits were only in seed parts; spikelet size, awn length,
hairs on empty glume and first empty glume area. When analyzing Echinochloa accessions with these
4 seeds traits again, we got the results which were significantly parted as 2 groups (E. oryzicola and
E. crus-galli). Therefore, we can suggest that everyone easily distinguish E.oryzicola and E.crus-galli
with using only these 4 traits.

Through SSRs analysis and morphological trait analysis, we could get the 2 phylogenic trees
that were closely alike each other in overall clustering. E. oryzicola accessions were clearly clustered
as a distinctive group from E. crus-galli and other Echinochloa species with the lowest phylogenetic
relationship among Echinochloa species in both SSRs analysis and morphological trait analysis. However,
no clear consensus between the results from these two analyses was founded in this study. This might
be because SSR markers were non genetic related markers whereas morphological traits were generally
related with gene. Nevertheless, our results indicate that both SSR markers and morphological traits
can be useful tools to distinguish among Echinochloa species. In addition, these two analyses will be
necessary with complementary view point for enhancing the reliance of each marker. Some individual
SSR markers, especially derived from sugarcane, pearl millet and Echinochloa, could discriminate E.
oryzicola from E. crus-galli. For uncertain or misclassified Echinochloa accessions, further study using
more SSR markers and morphological traits will be needed for more clear classification.

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 76

Fig. 1. Phylogenic tree for Echinochloa accessions and relative species based on SSR marker analysis.

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 77

References

Holm, L.G., Plucknett, D.L., Pancho, J.V., and Herberger, J.R., (1977). The World’s Worst Weeds:
Distribution and Biology, East-West Center Press, USA.

Chung, I.M., Ahn, J.K., and Yun, S.J. (2001). Assessment of Allelopathic Potential on Barnyard grass
(Echinochloa crus-galli) of Rice Cultivars. Crop Prot. 20: 921-928.

Yabuno, T. (1966). Biosystematic Study of The Genus Echinochloa. Jpn. J. Bot. 19: 277-323.
Sneller, C.H., Miles, J.W. and Hoyt, J.M., (1997).Agronomic Performance of Soybean Plant Introductions

and Their Genetic Similarity to Elite Lines. Crop Science. 37: 1595.
Rohlf, F.J. (2000). NTSYS-pc: Numerical Taxonomy and Multivariate Analysis System, 2.1 ed. Applied

Biostatistics, New York.

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 78

Eco-efficient weed management approaches for rice in tropical Asia

A.N. Rao*. and A. Nagamani**
* Former Agronomist (Weed Scientist),
International Rice Research Institute (IRRI) and
President, Society for Participatory Development and Research, Plot: 1294A; Road: 63A; Jubilee Hills;
Hyderabad – 500033, Andhra Pradesh, India.

E mail: [email protected]
** Professor, Department of Botany, University College of Science, Osmania University, Saifabad,

Hyderabad – 500004.
E mail: [email protected]

Abstract

Weeds continue to be the major impediments of rice production in Asia, in spite of research
efforts made so far. The yield losses due to weeds range from 12 to 80 percent depending on the
location, associated environment, weed community, rice establishment method, weed management
practices used by farmers. Weeds may cause complete rice crop failure, if appropriate weed management
measures are not taken up. None of the available weed management options, alone, could suppress the
weeds effectively in rice. Weeds are dynamic and in recent years weeds such as weedy rices are posing
severe threat to the efforts to enhance rice productivity to meet the demands of increasing population
in Asia. Innovative approaches need to be evolved for effectively combating the weed menace in rice
in Asia. Eco-efficient weed management in rice is concerned with the efficient and sustainable use
of resources in rice production to shift the crop weed balance in favor of rice. It encompasses both
ecological and economic dimensions of sustainable rice production. Key components of eco-efficient
weed management include: regular weed monitoring to identify shifts in weed populations and identify
problematic weeds from time to time, weed seed depleting soil and other management practices,
weed competitive rice based cropping systems and rice establishment methods, weed smothering rice
varieties, water and nutrient management options that are detrimental to weeds in rice, exploring the
feasibility of weeds usage, use of eco-friendly and economic herbicides in combination with cultural
practices, and adoption of appropriate weed dissemination prevention strategies. Available knowledge
on each of the components of eco-efficient weed management in rice in Asia is synthesized and future
research needs are enlisted.

Introduction

Rice is the major staple crop of tropical Asia. Tropical Asia, which extends over 80 degrees of
longitude (from 70°E to 150°E) and 40 degrees of latitude (from 30°N to 10°S). The 16 countries that
make up the region (Table 1). The region is physically diverse and ecologically rich in natural and crop-
weed related biodiversity. The vast variation in environmental conditions in tropical Asia make wide
variety of weeds (Moody, 1989; Caton et al., 2004) competing with rice and causing severe yield losses
ranging from 12 to 80 percent depending on the location, associated environment, weed community,
rice establishment method, weed management practices used by farmers (Rao et al., 2007). Weeds may
cause complete rice crop failure, if appropriate weed management measures are not taken up.

Weeds continue to be the major impediments of rice production in Asia, in spite of research
efforts made so far. Global climate changes are occurring and will result in further increases in
atmospheric greenhouse gases and temperature (> 0.2°C per decade), soil degradation and competing
claims for land and water (IPCC, 2007). Agricultural productivity in Asia is likely to suffer severe
losses because of high temperature, severe drought, flood conditions, and soil degradation; food
security of many developing countries in the region would be under tremendous threat. Climate change
is projected to compound the pressures on natural resources and the environment associated with

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 79

rapid urbanisation, industrialisation and economic development (IPPC, 2007). The rapidly changing
environmental conditions will affect the future rice production and weed management problem. Weed
species’ distribution and their competitiveness within a weed population and within a rice crop will
change with changes in atmospheric CO2 levels, rainfall, temperature and other growing conditions
(Mahajan et al., 2012*). This may necessitate adaptations in crop management practices, which in turn
will affect weed growth or proliferation of certain weed species.

The present total population in tropical Asia is about 1.6 billion, and the population is projected
to increase to 2.4 billion by 2025. It is estimated that demand for food and non-food commodities is
likely to increase by 75–100% globally between 2010 and 2050 (Keating et al., 2010; Tilman et al.,
2011). The increase in demand in tropical Asia is expected to be at least as much. The demand can be
met by bridging the yield gap through alleviation of impediments such as weed menace. Depending on
the magnitude of environmental changes within the Asian tropical region, environmental conditions
also have a large impact on the effectiveness of cultural, physical, mechanical and chemical weed
management practices.

In addition to impact of climate change, the continuing losses due to weeds in rice based agro-
ecosystems in Asian tropics (Rao et al., 2007), the evolution of herbicide resistance (Valverde and
Itho, 2001), shifts in the weed flora in response to weed management (Ramanjaneyulu et al., 2006), the
chemical contamination of water sources (Srivastava et al., 2010) and soil erosion through excessive
cultivation (Garrity, 1993) all attest to the need to develop systems of weed management, that are
sustainable and eco-efficient. Thus alternative weed management strategies are needed for enhancing
rice productivity to meet the increasing demand of population in tropical Asia through effective and
economical management of weeds in rice. The eco-efficient weed management is one of such innovative
approach.

What is eco-efficiency and Eco-efficient weed management ?
Eco-efficiency is concerned with the efficient and sustainable use of resources in farm

production and land management (Balasubramanian et al., 2012). Eco-efficiency was debated earlier
(British Crop Protection Council (BCPC) Forum (2004; Atkinson and Wilkins 2004), without a single
accepted definition of eco-efficiency. However there was an indication that eco-efficiency is related to
both ‘ecology’ and ‘economy’ and is concerned with the efficient and sustainable use of resources in
farm production and land management. Even though there are no absolute standards which a system
needs to satisfy in order to be classed as being eco-efficient, eco-efficiency will be increased when
a given level of production is achieved using less resources, with less losses to the environment and
without sacrifice to the productive potential of the land or economic performance. The efficient use of
plant nutrients, pesticides and energy and the minimization of greenhouse gas emissions are all key
concerns that affect eco-efficiency.

The BCPC Forum (2004) concluded that eco-efficient farming should satisfy the identified five
key attributes. Eco-efficient weed management is the approach of managing weeds which satisfy the
five key attributes identified by BCPC forum (2004), Viz. (i) it makes the maximum use of renewable
inputs by utilising resources efficiently and economically, (ii) it is locally non-polluting and does
not transfer pollution to elsewhere, (iii) it provides a predictable output, (iv) it conserves functional
biodiversity in relation to strengthening ecological processes, reducing greenhouse gas emission and
pollution generally and limiting soil erosion, and (v) it is capable of responding rapidly to changes in
the social, economic and physical environment. It is also crucial that eco-efficient weed management
must satisfy economic criteria in relation to farm profitability.

The eco-efficient weed management must thus form an integrated component of the rice
farming system, in order to realise the ultimate goal of eco-efficient rice production. Approaches to
enhance eco-efficiency in weed management in rice must include: (i) adoption of the method of rice
establishment and crop management practices that are eco-efficient, (ii) Increased ability to predict
interactions between rice and weed populations through better knowledge of weeds and their ecology
(ii) identifying and integrating weed management practices that make substantial reduction in external
inputs, manage weeds with higher resource use efficiency, with minimal adverse impact on environment
and result in higher rice productivity in an economic manner.

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 80

In this review, the possible eco-efficient weed management practices that may profitably used
are reviewed.

A. Adoption of the eco-efficient method of rice establishment and crop management practices.
In tropical Asia, transplanting seedlings into puddled soil (land prepared by wet tillage) is the

major method of rice crop establishment with 74% of rice is established by transplanting (Table 1). Due
to bigger size of rice seedling at the time of transplanting, puddle transplanted (CT-TPR) rice competes
better with emerging weeds and hence preferred by farmers as major method of rice establishment (Rao
et al., 2007). Reducing water percolation losses, easy seedling establishment, and creating anaerobic
conditions to enhance nutrient availability are the other benefits derived from puddling soil prior to rice
transplanting (Sanchez, 1973). However, puddling and transplanting require large amounts of water and
labor, which are becoming scarce and expensive. Higher labor wages of scarce labor are making rice
production less profitable in addition to the drudgery involved in transplanting by women. Thus, rice
cultivation in Asia is severely constrained by multiple stresses, especially water and labor shortages and
adverse effects of puddling on soil physical properties (Sharma et al., 2003) and a succeeding non-rice
crop (Hobbs and Gupta, 2003). Flooded rice culture with puddling and transplanting is considered one
of the major source of CH4 emission because of prolong flooding resulting in an anaerobic soil condition.
It accounts for 10-20% (50-100 Tg yr-1) of total CH4 emission globally on annual basis (Houghton et al.,
1996; Reiner and Milkha, 2000). Direct-seeding method of rice (DSR) establishment (both wet and dry)
have shown potential to reduce CH4 emissions compared to puddled transplanted rice (Wassmann et al.,
2004). Wassmann et al. (2004) suggested that CH4 mitigation effects can be further enhanced if Wet or
Dry-direct-seeded rice (DSR) is combined with mid-season drainage. Compared to CT-TPR, emissions
of CH4 decreased and of N2O increased in Dry-DSR. Therefore, DSR especially in dry conditions with
zero-tillage may be an effective mitigation option in Asia. All these factors demand a major shift from
CT-TPR to DSR in irrigated areas. Depending on water and labor scarcity, farmers are changing either
their rice establishment methods only [from transplanting to direct seeding in puddled soil (Wet-DSR)]
or both tillage and rice establishment methods [puddled transplanting to dry direct-seeding (Dry-DSR)]
in combination with resource-conserving technologies (RCTs) following the principles of conservation
agriculture (CA) which have been shown to increase the productivity and eco-efficiency of agriculture
at the farm level (Hobbs and Gupta, 2003; Sharma et al., 2005; Gupta and Seth, 2007; Ladha et al.,
2009). However, weeds menace is much severe in these alternative methods of rice establishment and
effective eco-efficient weed management strategies are the prerequisite for attainment of optimal rice
productivity in an eco-efficient manner.

B. Increased ability to predict interactions between rice and weed populations through better
knowledge of weeds and their ecology

Both rice and weeds need same resources for growth and establishment resulting in crop-
weed competition which necessitated the management of weeds to realise higher rice productivity.
Understanding the interactions between rice and weeds is essential to create microenvironment
favourable to rice and detrimental to weeds. Such an understanding would thus help in enhancing eco-
efficiency of weed management strategies. A few examples to illustrate the role of weed ecology in
managing weeds eco-efficiently are cited below.

Light plays a key role in the germination and growth of weeds and the germination response
of different weed species to light and darkness varies. A few rice weeds (eg. Cyperus iria, Eclipta
prostrate) require light for germination and do not germinate in darkness, some weeds do not require
light for germination (eg. Melochia corchorifolia, Mimosa invisa) and some weeds do not need light
but light stimulates their germination (Echinochloa colona, Digitaria ciliaris, Amaranthus viridis)
(Chauhan and Johnson, 2010). Under eco-efficient zero-tillage systems, the weeds species that require
light for germination or light simulates germination have the potentiality to become severe as they are
retained on soil surface where light is readily available. Such species can be managed by having stale
seed bed technique as component of eco-efficient weed management.

Light in the crop canopy is known to influence the growth of weed species. Possibility exists
to manage weeds that are susceptible to shading by selecting a rice variety that covers the ground fast

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 81

and allows less light penetration below its canopy. More research efforts are needed to identify the
weed species susceptible shade and adopted to shade in rice agro-ecosystems to enhance light resource
efficiency.

Adoption of eco-efficient CA practices such as zero-till systems are likely to leave a large
proportion of the weed seed bank on or near the soil surface after rice sowing. Higher loss in seed
viability and greater seed mortality occurs for weed seeds present on or close to the soil surface than the
buried weed seeds (Mohler, 1993). Weed seeds are most vulnerable to surface-dwelling seed predators
when on the soil surface and a tropical Asia study, showed 87% post dispersal weed seed predation was
from the soil surface (Chauhan et al. 2010).

Much more research needs to be conducted on weed seed bank dynamics, weed shifts with time,
identifying and recognising the biological and ecological factors leading to the long term persistence
of species in weed communities, and overall understanding of the ecology of weeds to manage weeds
eco-efficient manner in the age of changing climate.

C. Components of integrated eco-efficient weed management in rice in tropical Asia
Eco-efficiency in the simplest of terms is about achieving more with less-more agricultural

outputs, in terms of quantity and quality, for less input of land, water, nutrients, energy, labor, or capital
(Keating, et al., 2010). The eco-efficient conservation agriculture (CA) is characterised by minimum
soil disturbance, retention of residue for soil cover and rotation of major crops. Those components
could be used for managing weeds in rice. Eco-efficient weed management could not be achieved by
any of the single weed management practice and integration of available weed management practices is
needed to attain that objective. Some of the components that may be integrated include:

i).Avoiding weed seed contamination in rice: In recent years, weedy rice is becoming major problem
in Asian rice ecosystems.. Sowing seed contaminated by weedy rice is likely the primary cause of
invasiveness of weedy rice in rice fields (Rao et al., 2007). Rice seed soaking in herbicide solution for
controlling rice seed contaminants (Rao and Moody, 1996) or the use of certified seed have proved to
be an essential component in weed management (Rao et al., 2007).

ii) Sanitation: Sanitation is one means of minimizing the likelihood of weed introductions and dispersal
of existing weeds throughout a farm, especially herbicide-resistant weeds. All farm machinery should be
washed well to remove weed seeds and propagules of perennials in attached-soils from the neighbouring
weed infested fields before being moved into clean paddy fields.

iii) Soil solarisation: Soil solarization is a preventive method that exploits solar heating to kill weed
seeds and therefore reduce weed emergence. In this method, heating of the soil’s surface is done by
using transparent low-density polyethylene (LDPE film) sheets placed on the soil’s surface to trap solar
radiation (Khan et al., 2003). The use of transparent and black LDPE sheets reduces weed growth and
increases rice yield (Khan et al., 2003).

iv) Water management: Variation in soil moisture conditions can affect weed seed emergence and
viability differently (Mercado, 1979). Ismail et al. (1995) indicated that emergence of Echinochloa
crus-galli, E. colona, Cyperus iria, Ludwigia hyssopifolia and Rhyncospora corymbosa was lower in
soils flooded up to 4 cm water depth compared to seeds sown at all sowing depths in saturated soil.
However, some aquatic weeds germinate under water and this behavior has been used as a guide in their
control (Mercado, 1979). In fields known to be heavily infested with ‘aquatic’ weeds, germination of
the seeds can be stimulated by flooding the field and, after seedlings emerged, control measures can be
applied to these weed (Mercado, 1979). Even though water management got the potentiality to use as
a component of weed management strategy, water availability is becoming scarce in many of the rice
growing Asian countries.

v) Crop rotation: Differentiation of crops grown over time on the same field is a well-known primary
means of preventive weed control. Different crops obviously bring about different cultural practices,

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 82

which act as a factor in disrupting the growing cycle of weeds and, as such, preventing selection of the
flora towards increased abundance of problem species (Karlen, 1994). In contrast, continuous cropping
selects the weed flora by favouring those species that are more similar to the crop and tolerant to
the direct weed control methods used (e.g. herbicides) via repeated application of the same cultural
practices year after year.

vi) Mulching: Major component of CA is retention of residue cover on the soil surface. The spreading
of mulch on the soil surface reduces evaporation, saves water, protects from wind and water erosion,
and suppresses weed growth (Singh et al., 2007). Residue as mulch can suppress emergence and growth
of many weed species depending on the amount and type of crop residue. The rice straw mulch (4t/
ha) was effective in weed management under wet-seeded rice, but not in dry-seeded rice in Sri Lanka
(Devasinghe, et al., 2011). Mulching + dry land weeder at 20 DAS proved more effective in dry-seeded
rice grown without herbicide use (Hussain and Gogoi, 1996). In Nepal, straw mulch + bispyribac-
sodium reduced the population of Alternanthera philoxeroides, Ammannia sp., Commelina diffusa,
Cyperus difformis, Cyperus iria, and D. junceum 8 weeks after seeding dry-seeded rice (Ranjit and
Suwanketnikom, 2005).

vii) Brown manuring: Brown manuring is a no-till‘ version of green manuring, using a herbicide to
desiccate the intercrop (and weeds) at flowering instead of using cultivation. The plant residues are left
standing. In ‘Brown Manuring‘ practice both rice and Sesbania crops are seeded together and allowed
to grow for 30 days. Subsequently Sesbania intercrop is knocked down with 2, 4-D at 500 g ha-1 (Singh
et al., 2007). This technology reduces weed population by nearly half without any adverse effect on rice
yield. Sesbania surface mulch decomposes very fast to supply N and other recycled nutrients.

viii) Stale seedbed technique: Weed density could be considerably reduced in the eco-efficient CA
adopted systems by using the stale seedbed technique. Weeds population during the crop growing
season could be reduced by inducing weed seeds to germinate by giving light irrigations before land
preparation followed by non-selective herbicides or tillage operations for killing the emerged weed
seedlings. Some of the weed species species sensitive to the stale seedbed technique are Cyperus iria,
Digitaria ciliaris, Echinochloa colona, Eclipta prostrata, Leptochloa chinensis, Ludwigia hyssopifolia
and Portulaca oleracea (Chauhan and Mahajan, 2011). Practising stale seedbed for 14 days gave the
highest benefit:cost ratio (Sindhu et al., 2011). In rice-wheat cropping system, the energy utilization for
weed management (which accounted for 1.47 to 3.40 per cent of total input energy) was found slightly
higher in traditional seedbed (925 to 1788 MJ/ha) than stale seedbed (768 to 1364 MJ/ha) (Chaudhary
et al., 2006).

ix) Enhancing crop competitiveness: The detrimental effects of weeds on the crop may be reduced by
making the crop more competitive. Improving rice competitiveness against weeds would provide a low-
cost and safe tool for the eco-efficient weed management strategy. Any cultural practice that facilitates
rapid rice growth and results in rice canopy covering soil surface, and shade out weeds, increase crop
competitiveness. Practices that contribute rice competitiveness include: early sowing, selection of
varieties with early growth (Zhao et al. 2006; Namuco et al. 2009). optimal seed rates (Zhao et al.
2007); close spacing (Chauhan and Johnson 2010a); adequate plant population and fertilization (Rao et
al., 2007) intercropping (Musthafa and Potty, 2001, Duary et al., 2005), and deferred sowing of black
gram in rice (30 cm) after one weeding (Midya et al., 2005).

x) Bioherbicides: Micro-organisms also are used as tools for weed management and have a range of
properties that make them desirable for ecological weed management in direct-seeded rice. COLLEGO,
a powder formulation of Colletotrichum gloeosporioides (Penz.) Sacc. f. sp. aeschynomene, was
registered in 1982 for the control of northern jointvetch (Aeschynomene virginica (L.) B.S.P.) in rice. The
successful mycoherbicide, Rhynchosporium alismatis had synergistic controlling effect on Damasonium
minus (R.Br.) Buch. when bensulfuron-methyl was applied before fungal inoculation (Jahromi et al.,
2001). In Asia, the bioherbicide research is yet to reach the practical usage stage.

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 83

xi)Weeds use: In several countries of Asia, weeds are normally used in villages, as food, medicine
and other purposes. In the long run, with proper planning and support, the growing and marketing of
medicinal herbs which are currently categorized as weeds could become a very good source of income
to farmers, while providing an additional, easily available source of natural medicines to the rural
population (Ediriweera, 2007). However, practical feasibility and economics of such usage on large
scale needs to be determined before it becomes a practical proposition.

xii) Mechanical weeding: Weeding by mechanical devices reduces the cost of labor and also saves time
(Subudhi, 2004). The labor involved was least with the Phulbani weeder (57 person-days ha-1), saving
nearly 57% labor compared with hand weeding (127 person-days ha-1). It also had better weed control
efficiency (Subudhi, 2004). Dry-seeded rice row seeding with interrow weeding using hoes and without
any herbicide achieved higher grain yield (Satoshi et al., 2009). Mnguu (2010) opined that rice farmers
can use rotary weeding instead of herbicide in controlling weeds and achieve the same grain yield of
wet-seeded rice.

xiii) Manual method: Where ever labor is available, manual method could be used as a component
of eco-efficient weed management. However, in tropical Asia the labour availability is decreasing and
the cost is increasing. Even in regions where labour is available, availability of labour in time is a
constraint which is resulting in damage due to weeds even before the manual weeding is done late on
their availability.

xiv) Herbicides: In the eco-efficient CA systems, herbicide use is an important component of weed
management and choosing an appropriate herbicide and timing is critical. A variety of herbicides have
been screened and found effective for burn down, pre‐emergence and post‐emergence weed control in
dry‐seeded systems (Kumar and Ladha, 2011). Virtually all rice farmers who practice direct-seeding
adopt chemical herbicides because they reduce weed control time in dry‐seeded crops by 500 hours
per hectare in comparison to hand‐weeded transplanted rice (Mazid et al., 2006, Ho, 1996). Herbicide
resistant rice cultivars may become an important component of eco-efficient weed management in
future, especially for managing recently emerging problematic weeds such as weedy rice. However,
educating farmers in Asia is needed to effectively use that technological component.

Integration of above components for managing weeds must ultimately lead to enhancement of
the eco-efficiency of agro-ecosystems of the tropical Asia.

Can we quantify the eco-efficiency of the components of eco-efficient weed management:
Most of the components of the eco-efficient weed management are enlisted here are based on

their qualitative characteristics pertaining to their eco-efficiency as reported in the literature. However,
a method to quantify the eco-efficiency is needed to select and integrate the components of eco-efficient
weed management. Eco-efficiency was used as a tool to compare herbicide resistant and conventional
cropping systems (Cobourn and Kniss, 2012). The formulae used by Cobourn and Kniss, (2012) for
calculating eco-efficiency may be used for quantifying the eco-efficiency of different components of eco-
efficient weed management. To compare the eco-efficiency of different herbicides, the Environmental
Impact co-efficient (EIQ) suggested by Kovach et al., (2010) may be used. Thus it is possible to quantify
the eco-efficiency of the components of eco-efficient weed management.

Future research efforts to develop weed management technologies must consider eco-efficiency
as the criteria to attain optimal rice yields in an eco-efficient manner in tropical Asia.

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 84

Acknowledgement
Thanks are due to Ms. Jaya Adusumilli for her help.

Table 1. Area, production, productivity of rice and the percentage of area under direct seeding and
transplanting in tropical Asia (2010) (Compiled based data from IRRI and Rao et al., 2007)

Rank Area Area Productivity Production Direct-seeded Transplanted
(000 MT) Rice area* (%) rice area (%)
( 000 ha) (t/ha)
120620 28 72
1 India* 36950 3.26 66411.5 18 82
2 Indonesia* 13244.2 5.01 49355 19 81
3 Bangladesh* 11800 4.18 39988.9 39-47 53-61
4 Viet Nam* 7513.70 5.32 33204.5 9 91
5 Myanmar* 8051.70 4.12 31597.20 34 66
6 Thailand* 10990.1 2.88 15771.7 42 58
7 Philippines* 4354.16 3.62 8245.32 10 90
8 Cambodia* 2776.51 2.97 4300.62 77 23
9 Sri Lanka* 1060.36 4.06 4023.82 N**
10 Nepal* 1481.29 2.72 3006 33 67
11 Laos 3.46 2548 71 29
12 Malaysia 870 3.78 N-n N
13 Bhutan 673.75 2.83 61.70 N N
14 Brunei 1.27 1.4
21.80
Darussalam 1.1
Tropical Asia
99788.67 3.53 379135.66 26 74
Asia 136550 4.45 607320 20.2 79.8
World 153650 4.37 672016 22.7 77.3

*From Rao et al., 2007
N = Information missing, N-n = negligible DSR – mainly transplanting)
** Manual transplanting is the dominant crop establishment method in lowland ecologies, while
direct seeding of seeds to dry soils is dominant in upland ecologies (FAO, 2002).

References

Atkinson, D. and Wilkins, R. J. 2004 The future opportunities for enhancing eco-efficiency in UK
agriculture. (Paper prepared for BCPC Forum: Enhancing the eco-efficiency of agriculture.) See
http://www.bcpc.org/reports.

Balasubramanian, V., Adhya, T.K. and Ladha, J.K. 2012. Enhancing eco-efficiency in the intensive
cereal-based systems of the Indo-Gangetic Plains. Chapter 7, in Issues in Tropical Agriculture
- Eco-Efficiency: From Vision to Reality, CIAT, Km 17, Recta Cali-Palmira., Apartado Aéreo
6713., Cali, Colombia, pp.1-7.

BCPC Forum. 2004. Eco-efficiency in the future pattern of British Agriculture. Setting the agenda for
crop production. BCPC, Hampshire, U.K.

Cobourn, C. and Kniss, A. 2012. Eco-efficiency as a tool to compare herbicide-resistant and conventional
cropping system. Proc. 65th meeting of Western Society of Weed Science, Reno, Nevada, UDA.
65, 35 http://www.wsweedscience.org/Proceedings%20Archive/2012.pdf

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 85

Caton, B.P., Mortimer, M., and Hill, J.E. 2004. A practical field guide to weedsof rice in Asia. Los
Baños (Philippines): International Rice Research Institute. 116 p.

Chaudhary, V., Sharma, S., Pandey, D. and Gangwar, B. 2006. Energy assessment of different weed
management practices for rice-wheat cropping system in India. International Commission
of Agricultural Engineering (CIGR, Commission Internationale du Genie Rural) E-Journal
Manuscript EE 05 008. Vol. VIII.

Chauhan, B. S. and Johnson, D. E. 2010. The role of seed ecology in improving weed management
strategies in the tropics. Advances in Agronomy. 105, 221-262.

Chauhan, B. S. and D. E. Johnson. 2010 a. Implications of narrow crop row spacing and delayed
Echinochloa colona and Echinochloa crus-galli emergence for weed growth and crop yield loss
in aerobic rice. Field Crops Res. 117, 177–182.

Chauhan, B.S. and Mahajan, G. 2011. Role of integrated weed management strategies in sustaining
conservation agriculture systems. Current Science. 103, 135-136.

Chauhan, B. S., Migo, T., Westerman, P. R. and Johnson, D. E. 2010. Post-dispersal predation of weed
seeds in rice fields. Weed Res. 50, 553–560.

Devasinghe, D.A.U.D., Premarathnel, K.P. and Sangakkara, U.R. 2011. Weed management by rice
straw mulching in direct seeded lowland rice (Oryza sativa L.). Tropical Agricultural Research.
22, 263 - 272 .

Duary, B., Hossain, A. and Mondal, D. C. 2005. Integrated weed management in direct seeded dry
sown rice in lateritic belt of West Bengal. Indian Journal of Weed Science; 37, 101-102.

Ediriweera, E.R.H.S.S.. 2007. A Review on Medicinal uses of Weeds in Sri Lanka. Tropical Agricultural
Research & Extension 10, 11-16.

FAO. 2002. ‘‘Rice Information,’’ Vol. 3. FAO, Rome (www.FAO.org)
Garrity, D. P. 1993. Sustainable land-use systems for sloping uplands in Southeast Asia. In Technologies

for sustainable agriculture in the tropics, (eds) Ragland J and Lal R. American Society of
Agronomy Special Publ., 56, 41 - 66.
Gupta, R.K. and Seth,A. 2007.Areview of resource conserving technologies for sustainable management
of the rice–wheat cropping system of the Indo-Gangetic Plains (GCP). Crop Protection 26, 436–447.
Ho, Nai‐Kin. 1996. Current status of rice herbicide use in the tropics. JIRCAS International Symposium
Series. No. 4, 77‐86.
Hobbs, P.R. and Gupta, R.K. 2003. Resource-conserving technologies for wheat in the rice–wheat
system. In: Ladha et al., eds. Improving the productivity and sustainability of rice–wheat systems:
Issues and impacts. ASA Spec. Pub. 65ASA, CSSA, and SSSA, Madison, WI, USA. P. 149–171.
Houghton, J. T., Meira Filho, L. G., Callander, B. A., Harris, N., Katterberg, A. and Maskell, K. 1996.
IPCC report on climate change: The science of climate change. “WG1 Contribution to the IPCC
Second Assessment Report on Methane Emission from Rice Cultivation”, Cambridge University
Press, Cambridge, UK.
Hussain, A. and Gogoi, A. K. (1996). Non-chemical weed management in upland direct-seeded rice.
Indian J. Weed Sci. 28, 89-90.
IPCC. 2007. Climate Change 2007: Synthesis Report. Contribution of Working Groups I, II and III to
the Fourth Assessment Report of the Intergovernmental Panel on Climate Change [Core Writing
Team, Pachauri RK and Reisinger A eds]. IPCC, Geneva, Switzerland. BCPC Forum 2004
Enhancing the eco-efficiency of agriculture. Alton, UK: British Crop Protection Council.
Ismail, B.S., Faezah, Z.N., Ho, N.K. 1995 Weed populations and their buried seeds in rice fields of the
Muda area, Kedah, Malaysia. Pertanika. 18, 21-28
Karlen, D.L., Varvel, G.E., Bullock, D.G. and Cruse, R.M. 1994. Crop rotations for the 21th century.
Advances in Agronomy 53, 1-45.
Keating, B. A. Carberry, P. S. Bindraban, P. S. Asseng, S. Meinke. and H. Dixon, J. 2010. Eco-efficient
agriculture: concepts, challenges, and opportunities. Crop Science; 50: Supplement 1, 109-119.
Khan A. R., Srivastava R. C., Ghorai A.K. and Singh, S.R. 2003. Efficient soil solarization for weed
control in the rain-fed upland rice ecosystem. Int. Agrophys. 17, 99–103.
Kovach J., Petzoldt, C., Degni, J. and Tette, J. 2010. A method to measure the environmental impact of
pesticides. http://nysipm.cornell.edu/publications/eiq/.

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 86

Kumar, V. and Ladha, J.K. 2011. Direct seeding of rice: recent developments and future research needs.
Advances in Agronomy. 111, 297-413.

Ladha, J.K., Kumar, V., Alam, M.M., Sharma, S., Gathala, M.K., Chandna, P., Sahrawat, Y.S.,
Balasubramanian, V. 2009. Integrating crop and resource management technologies for enhanced
productivity, profitability, and sustainability of the rice–wheat system in South Asia. In: Ladha
JK; Yadvinder-Singh; Erenstein O; Hardy B, eds. Integrated crop and resource management in
the rice–wheat system of South Asia, International Rice Research Institute (IRRI), Los Baños,
Philippines. p 69–108.

Mahajan, G., Singh, S. and Chauhan, B.S. 2012. Impact of climate change on weeds in the rice–wheat
cropping system. Current Science. 102, 1244-1255.

Mazid, M.A., Riches, C.R., Mortimer, A.M., Wade, L.J. and Johnson, D.E. 2006. Improving rice‐based
cropping systems in north‐west Bangladesh. Fifteenth AustraliConference. 331‐334

Mercado, B.L. 1979. Introduction to weed science. South East Asia Regional Center for Graduate Study
and Research in Agriculture, Laguna, Philippines. P. 229

Midya, A., Bhattacharjee, K., Ghose, S. S., Banik, P. 2005. Deferred seeding of blackgram (Phaseolus
mungo L.) in rice (Oryza sativa L.) field on yield advantages and smothering of weeds. Journal of
Agronomy and Crop Science; 191, 195-201.

Mohler, C.L. 1993. A model of the effects of tillage on emergence of weed seedlings. Ecological
Applications 3, 53–73.

Moody, K. 1989. Weeds reported in rice in South and Southeast Asia. Los Baños (Philippines):
International Rice Research Institute. 442 p.

Mnguu, Y. O. 2010. Effect of crop management practices on Nitrogen (N) availability and N use
efficiency in direct wet-seeded Rice. Journal of Animal & Plant Sciences. 6, 589- 596.

Musthafa, K. and Potty, N. N. 2001. Effect of in situ green manuring on weeds in rice. Journal of
Tropical Agriculture 39, 172-174.

Namuco, O.S., Carins, J.E., Johnson, D.E. 2009. Investigating early vigour in upland rice (Oryza sativa L.):
part I. seedling growth and grain yield in competition with weeds. Field Crops Res 113: 197-206.

Ramanjaneyulu, A.V., Sharma, R. and Giri, G. 2006. Weed shift in rice based cropping systems - a
review. Agric. Rev., 27, 73 – 78.

Ranjit, J. D. and Suwanketnikom, R. 2005. Response of weeds and yield of dry direct seeded rice to
tillage and weed management. Kasetsart Journal, Natural Sciences; 39, 165-173.

Rao, A. N. and Moody, K. 1996. Seed soaking in herbicide solution for controlling Ischaemum rugosum,
a rice seed contaminant. In: Proc. of the 15th Asian Pacific Weed Science Society Conf. 24th–28th
July 1995, Tsukuba, Japan.

Rao, A. N., Mortimer, A. M.., Johnson, D. E., Sivaprasad, B. and Ladha, J. K. 2007. Weed management
in direct-seeded rice. Advances in Agronomy. 93, 155-257.

Reiner, W., and Milkha, S. A. 2000. The role of rice plants in regulating mechanisms of methane
emissions. Biol. Fertil. Soils 31, 20–29

Sanchez, P.A. 1973. Puddling tropical rice soils. 2. Effect of water losses. Soil Sci. 115, 303–308.
Satoshi, H., Anuchart, K., Boonrat, B., Akihiko, K. and Junko, Y. 2009. Spatial variability in the growth

of direct-seeded rainfed lowland rice (Oryza sativa L.) in northeast Thailand. Field crops research.
111, 251-261.
Sharma, P. K., Ladha, J. K., and Bhushan, L. 2003. Soil physical effects of puddling in rice-wheat
cropping systems. In “Improving the Productivity and Sustainability of Rice-Wheat Systems:
Issues and Impacts” (J. K. Ladha, J. E. Hill, J. M. Duxbury, R. K. Gupta, and R. J. Buresh, Eds.),
pp. 97–113. ASA, CSSA, SSSA, Madison, WI.
Sindhu, P. V., Thomas, C. G. and Abraham, C. T. 2011. Stale seedbed and green manuring as an effective
weed management strategy in dry-seeded rice (Oryza sativa). Indian Journal of Agronomy. 56,
109-115.
Singh, S., Ladha, J. K., Gupta, R. K., Bhushan, L., Rao, A. N., Shiva Prasad, B. and Singh, P. P. 2007.
Evaluation of mulching, intercropping with Sesbania and herbicide use for weed management in
dry-seeded rice (Oryza sativa). Crop Protection. 26, 518-524.

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 87

Srivastava, S.; Goyal, P. & Mohan Srivastava, M. 2010. Pesticides: Past, present, and future. In:
Handbook of Pesticides: Methods of Pesticide Residues Analysis, Nollet, L.M.L. & Rathore, H.S.
(Eds.), pp. 47-65, Taylor & Francis, Boca Raton, Florida

Subudhi, C. R. 2004. Evaluation of weeding devices for upland rice in the Eastern Ghats of Orissa,
India. IRRN. 29: 79-80.

Tilman, D., Baizer, C., Hill, J., Befort, B.L. 2011. Global food demand and the sustainable intensification
of agriculture. Proceedings of the National Academy of Sciences (PNAS). 108, 20260–20264.

Valverde, B E and Itho, K. 2001. World rice and herbicide resistance. In: Powels, S.B., Shaner, D.L.
(eds.): Herbicide Resistance and World Grains, CRC Press, Florida, 196-249

Wassmann, R., Neue, H. U., Ladha, J. K. and Aulakh, M. S. 2004. Mitigating greenhouse gas emissions
from rice–wheat cropping system in Asia. Environ. Dev. Sustain. 6, 65–90

Jahromi, F., Cother, E. and Ash, G. 2001. Weed Control in Rice Crops - Suitability of Rhynchosporium
alismatis as a Mycoherbicide for Integrated Management of Damasonium minus in Rice Fields.
RIRDC Publication No 01/39. RIRDC Project No UCS 7A. (RIRDC, Canberra, Australia).

Zhao, D.L., Altin, G.N., Bastiaans, L. and Spiertz, J.H.J. 2006. Cultivar- weeds Competitiveness in
aerobic rice: Heritability, correlated traits, and the potential for indirect selection in weed- Free
environments.CropSci46, 372-380.

Zhao, D.L., Bastiaans, L., Atlin, G.N. and Spiertz, J.H.J. 2007. Interaction of genotype × management
on vegetative growth and weed suppression of aerobic rice. Field Crops Res.100, 327-340.

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 88

Morphological and physiological responses of Miscanthus spp.
to varying temperature and light intensity

Jastin Edrian Revilleza1, Soo-Hyun Lim1, Ji-Hoon Chung1, Do-Soon Kim1
1Department of Plant Science, Seoul National University, Seoul 151-742, Korea

Corresponding author: E-mail: [email protected]

Abstract

C4 plants are known to have a higher yield compared to C3 plants. Generally, C4 species are best
suited to tropical and subtropical climates. Miscanthus, commonly known as a rhizomatous grass species
and an important bioenergy weed crop, is a highly exceptional C4 species due to its high productivity
in temperate climates. However, recent reports were focused only in the comparison between Maize
and Miscanthus response to such unfavorable conditions. Therefore, this study was conducted to
investigate photosynthetic responses of different Miscanthus genotypes to low temperature conditions
and sub-optimal light intensity. Various Miscanthus genotypes collected from different locations were
grown under three different temperature levels, 25/20oC, 20/15oC and 15/10oC during a 35-day period
and at five different light intensity regimes namely 100%, 75%, 55%, 25% and 5% during a 70-day
period. Necessary morphological assessments include plant height, number of leaves and leaf area and
essential physiological assessments include photosynthetic rate, chlorophyll content and chlorophyll
fluorescence. Results show that different Miscanthus genotypes showed the same trend within the
different environmental conditions. However, biomass yield were not affected by these conditions,
suggesting that it is independent of the following physiological response. This report demonstrates an
overview of Miscanthus tolerance to low temperature and sub-optimal light conditions.

Keywords: Bioenergy crop, biomass, Miscanthus, photosynthesis

Introduction

Miscanthus x giganteus is exceptional among C4 species for its high productivity in temperate
climates (Dohleman and Long, 2009) and has been cultivated for biomass production in Europe and
USA. Miscanthus genusis composed of 14 major species, of which M. sinensis, M. sacchariflorus and
M. floridulus are native to the Eastern Asian region including Korea, China and Japan. Miscanthus
x giganteus is the only Miscanthus species being commercially cultivated for biomass production
due to its high biomass yield (Chung and Kim, 2012). However, the study of C4 photosynthesis on
different environmental conditions are limited and is focused mainly on the difference between Maize
and Miscanthus spp. (Naidu et al., 2003; Wu and Wedding, 1987) and focused mainly on Miscanthus
spp. grown in Europe and USA with little emphasis on the eastern parts of Asia (Beale et al, 1996).
Therefore, the objectives of this study were to investigate photosynthetic responses of different
Miscanthus genotypes to low temperature conditions and sub-optimal light intensity and to compare
the biomass yield among the different genotypes with relation to its photosynthesis ability.

Materials and Methods

Various genotypes belonging to Miscanthus x giganteus (reference genotype), M. lutarioriparius,
M. sinensis and M. sacchariflorus were selected from 300 genotypes collected from various locations in
Korea and neighboring East Asian countries such as China, Japan and Russia (Table 1).

To give the different low temperature treatments, the Miscanthus spp. plants were grown for
at least 25 cm tall and were transferred to a growing chamber (Hanbaek, Korea) with varying day and

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 89

night temperatures of 25/20°C, 20/15°C, and 15/10°C,. Assessments were made on a 7-day interval
for 35 days. To give the different light intensity treatments, the Miscanthus spp. plants were grown
for 60 days and were transferred to different light intensity conditions: 100%, 75%, 45%, 25%, and
5%. Assessments were made on a 7-day interval for 70 days. To assess the photosynthesis rate various
Miscanthus spp. genotypes on field conditions, three-year stands of 53 genotypes of Miscanthus spp.
planted on the germplasm field in the experimental farm station in Suwon, South Korea. Three leaves
from different tillers of the same genotype were used in measuring the different parameters during the
summer months of 2012; June, August and September.
For the morphological assessments, plant height, leaf number, tiller number and leaf area were
measured. For the physiological assessments, LI-6400XT (LI-COR, USA) was used to measure the
photosynthesis rate, SPAD (Konica Minolta, Japan) was used to measure leaf chlorophyll content and
Handy PEA (Hansatech Instruments, UK) was used to measure the leaf fluorescence. The biomass yield
was measured at the end of the observation period.

Results and Discussion

Plant growth and increase in tiller number was halted at lower temperature and lower light
intensity. However, leaf number was not significantly different among the different temperature range
and light intensity. Photosynthesis rate, chlorophyll content (SPAD) and leaf fluorescence (Pi) was
significantly affected at low temperature and low light intensity at 28 and 56 days after transferring
(DAT), respectively. The various Miscanthus spp. genotypes have different rates of responses on these
various temperature and light intensity regimes. There was also no clear correlation of photosynthesis to
chlorophyll content and leaf fluorescence. Moreover, there was no clear correlation of photosynthesis rate
to biomass yield at different temperature ranges and suboptimal light intensities (Figure 1). Therefore,
there is diversity with response of different Miscanthus spp. to various environmental conditions such
as temperature and light intensity.

AB

Figure 1. Relationship between biomass yields and photosynthetic abilities of M. x giganteus (●), M.
lutarioriparius (■), M. sinensis (▲) and M. sacchariflorus (▼) for varying temperature (A) and varying
light intensity (C).

Photosynthetic abilities of 56 Miscanthus genotypes grown in the field condition showed very
diverse depending on species, genotypes and months when the measurement was made. Some genotypes
have a higher biomass yield even with low photosynthesis rate, while some have a lower biomass yield
even with high photosynthesis rate (Table 1). Nonetheless we selected 4 genotypes, two M. sinensis
genotypes, one M. lutarioriparius genotype and M. x giganteus (reference) having high biomass yield
potential with high photosynthetic ability. Therefore, these three genotypes except M. x giganteus will
be further studied for developing a new Miscanthus variety for biomass production.

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 90

Table 1. Miscanthus spp genotypes with high photosynthesis rate (and high biomass yield.

Miscanthus spp. Genotype Carbon dioxide assimilation (µCO2 m-2s-1) Biomass yield (g)
June August September
63.05
M x giganteus M-57 12.71 10.55 9.75 66.21
M lutarioriparius M-271 9.38
M sacchariflorus M-07 9.18 8.59 7.83 17.91
M sacchariflorus M-46 20.77
M sacchariflorus M-90 8.58 11.23 8.60 10.28
M sacchariflorus M-93 9.14
M sacchariflorus M-157 8.80 7.70 11.79 19.75
M sacchariflorus M-180 26.86
M sacchariflorus M-243 12.42 10.23 8.12 18.27
M sacchariflorus M-289 54.03
M-37 8.76 11.01 11.52 28.64
M sinensis M-104 42.28
M sinensis M-257 8.34 10.12 8.26
M sinensis
7.75 7.60 7.59

11.97 9.25 8.02

9.93 11.20 11.71

8.74 7.38 12.66

8.19 9.10 8.63

8.02 10.90 8.32

References

Beale, C.V., Blint, D.A. and Long. S.P. (1996). Leaf Photosynthesis in the C4-grass Miscanthus x
giganteus, Growing in the Cool Temperate Climate of Southern England. J. Exp. Bot. 47: 267-273.

Chung, J.H. and Kim, D.S. (2012). Miscanthus as a Potential Bioenergy Crop in East Asia. J. Crop Sci.
Biotech.. 15(2): 65-77.

Dohleman, F.G. and Long, S.P. (2009). More Productive Than Maize in the Midwest: How Does
Miscanthus Do it?. Plant Physiol. 150: 2104-2115.

Naidu, S.L., Moose, S.P., Al-Shoaibi, A.K., Raines, C.A. and Long, S.P. (2003). Cold Tolerance of
C4 Photosynthesis in Miscanthus x giganteus: Adaptation in Amounts and Sequence of C4
Photosynthetic Enzymes. Plant Physiol. 132: 1688-1697.

Wedding R.T. and Wu, M.-X. (1987). “Temperature Effects on Phosphoenolpyruvate Carboxylase from
a CAM and a C4 Plant.” Plant Physiol. 85: 497-501.

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 91

Flucetosulfuron performance improved by adjuvant

Jin Won Kim1, Seong-Hyu Shin2, Jong-Nam Lee3, Se-Eun Lim3, Soo-Hyun Lim1, Do-Soon Kim1,*
1Department of Plant Science, Seoul National University, Seoul, Korea
2National Research Institute of Crop Science, RDA, Suwon, Korea
3Division of Specialty Chemicals, LG Life Science Ltd., Seoul, Korea
* Corresponding author: E-mail: [email protected]

Abstract

This study was conducted to investigate the effects of adjuvant on herbicidal performance of
flucetosulfuron (FTS, Fluxo®, LG Life Science Ltd.) against Echinochloa crus-galli and Aeschynomene
indica. Adjuvant which reduced contact angles of herbicide droplet generally increased herbicidal
performance of FTS. HLB (hydrophilicity and lipophilicity balance) of adjuvant tested are also related
with herbicidal performance of FTS. The GR50 values of FTS dramatically decreased with increasing
adjuvant concentration in the same manner as the herbicide dose-response. Leaf excision test revealed
that FTS uptake and translocation was also enhanced by adjuvant, depending on adjuvant concentration.
As a result, we selected several adjuvant showing much more improved performance in FTS activity
against E. crus-galli and A. indica and produced new FTS formulation for foliar application. In this
presentation, we will also introduce new FTS formulations improved by adding a new adjuvant, which
may help to manage paddy weeds in tropical rice cultivation.

Keywords: adjuvant, Aeschynomene indica, contact angle, flucetosulfuron, HLB

Introduction

There are significant differences in foliar uptake of herbicide formulations among plant species
(Chamel et al., 1992; Santier and Chamel, 1996; Gouret et al., 1993; Price and Anderson, 1985; Baker
et al., 1992; Knoche and Bukovac, 1993). In general, increasing amounts of surfactants will increase
agrochemical uptake, although in some cases, such as with glyphosate, if this markedly increases the
contact area of the droplet with the leaf surface, it may reduce the concentration per unit area and reduce
the uptake of the herbicide (Liu and Zabkiewicz, 1998).

Contact angle was more closely correlated with diuron efficacy than surface tension, and the
coefficient of determination between the contact angle and the fresh weight or control percent of
barnyardgrass was 0.41. Kinetic, an organosilicones, applied before or in combination with diuron
significantly enhanced diuron efficacy. However, Kinetic sprayed after diuron did not affect the efficacy.
Kinetic influenced the retention and absorption of diuron rather than its deposition and translocation
(Singh et al., 2002).

Surfactants having a high hydrophilic-lipophilic balance (HLB) are absorbed into the cuticle and
enhance the water-holding capacity (hydration state) of the cuticle. With increased cuticle hydration, the
permeance of hydrophilic herbicides into the cuticle is increased, which increases the herbicide diffusion
rate at a constant concentration gradient. Surfactants having a low HLB are absorbed into the cuticle
and increase the fluidity of waxes, as measured by a small reduction in melting point. This increased
fluidity increases the permeance of lipophilic herbicides in the cuticle, which, in turn, increases their
diffusion rate at a set concentration gradient (Hess and Foy, 2000). Therefore, this study was conducted
to investigate the effects of adjuvant on the herbicidal performance of flucetosulfuron and to select a
suitable adjuvant for tank mix or pre-mix with flucetosulfuron.

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 92

Materials and Methods

Whole plant response to adjuvants
Indian joint vetch (Aeschynomene indica L.) was seeded on the pot containing sandy loam soil

and was grown up to 40 cm tall. Forty seven adjuvants were prepared at two concentrations, 0.05 and
0.1% (v/v), and then flucetosulfuron was mixed with the prepared adjuvants. The mixtures were applied
to A. indica at 15 g flucetosulfuron a.i. ha-1 and 300 L ha-1 (30 psi) of spray volume with a laboratory track
sprayer. Herbicidal efficacy was evaluated with visual-rating from 0 to 10 at 14 days after application.
All treatments were performed three times and arranged by randomized block design.

Contact angle measurement
A 10 μL of 0.05 and 0.1% adjuvant solutions was dropped on a slide glass and its contact angle

was measured three or four times by Drop Shape Analysis System (DSA 10-MK2, Krüss, Germany) at
23oC. Contact angle of distilled water was 30.5o.

Concentration effect of adjuvants on flucetosulfuron performance
OA14 (HLB 13.6) was prepared at 0.025, 0.05, and 0.1%, and then flucetosulfuron was mixed

with the prepared OA14 solutions. The mixture was applied to A. indica at 3.75, 7.5, 15, and 30 g
flucetosulfuron a.i. ha-1 under same conditions as whole plant assay. All treatments were performed
three times and arranged by randomized block design. Herbicidal efficacy was evaluated with visual-
rating from 0 to 10 at 14 days after application.

Results

Relationship between contact angle and FTS activity
Contact angles of solutions containing tested adjuvants at 0.1% were correlated with visual

efficacy of tested adjuvants in controlling A. indica. Contact angle showed significantly negative
correlation with visual efficacy with -0.79 (Figure 1). However, several adjuvants e.g. Silwet L-77,
SN10, SN20, TN20, and LN20 were away from such a relationship. Silwet L-77 is an organosilicone
adjuvant and showed relatively poor efficacy of flucetosulfuron in spite of less than 1 degree of contact
angle. SN10, SN20, TN20, and LN20 belong to plyoxyethylene alkyl amine series, which showed
excellent efficacy of flucetosulfuron in spite of large contact angle, ranged from 22.4 through 28.0
degrees. Nevertheless, it is clear that decrease in contact angle by adding adjuvant significantly improves
herbicidal performance of flucetosulfuron. Therefore, contact angle can be also a decisive consideration
for choosing adjuvant for flucetosulfuron.

Concentration effect of adjuvants on flucetosulfuron performance
Applications of 15 and 30 g a.i. ha-1 of flucetosulfuron with 0.1% adjuvant had six and four

times herbicidal efficacy as good as those without an adjuvant, respectively, regardless of adjuvant type.
Regardless of the applied flucetosulfuron dose, there was concentration effect of OA14 on efficacy of
flucetosulfuron, showing the more the adjuvant concentration the better the flucetosulfuron efficacy
(Figure 2) is. Particularly, without OA14, the herbicidal efficacy of flucetosulfuron was very poor,
unable to estimate GR50 value (Figure 2). The GR50 of flucetosulfuron was 29.9, 13.4, 5.39 g a.i. ha-1
at 0.025, 0.05, and 0.1% of OA14, respectively, demonstrating flucetosulfuron performance improved
with increasing adjuvant concentration (Figure 2).

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 93

12Visual efficacy

r = -0.79

10

8

6

4

2

0
0 5 10 15 20 25

Contact angle (o)

Figure 1. Correlation between contact angle of adjuvant solutions and visual efficacy against
Aeschynomene indica of flucetosulfuron sprayed with adjuvants at 0.1% (v/v).

10 No Adjuvant
0.025%
GR50 (g a.i. ha-1) 0.05%
No adjuvant: Not calcu- 0.1%

8 lable

0.025%: 29.9
0.05%: 13.4

6 0.1%: 5.39

4

Visual efficacy 2

0
0 5 10 15 20 25 30

Flucetosulfuron (g a.i. ha-1)

Figure 2. Visual efficacy against A. indica of flucetosulfuron sprayed with OA14 at different
concentrations. The continuous lines are fitted values by using the log-logistic model. GR50 is the dose
of flucetosulfuron to cause the 50%-growth retardation of A. indica.

References

Baker, E.A., Hayes, A.L. and Butler, R.C. (1992). Physiochemcial Properties of Agrochemicals: Their
Effects on Foliar Penetration. Pesticide Science. 34: 167-182.

Chamel, A., Gambonnet, B. and Coret, J. (1992). Effects of Two Ethoxylated Nonylphenols on Sorption
and Penetration of [14C]Isoproturon through Isolated Plant Cuticles. Plant Physiology and
Biochemistry. 30: 713-721.

Gaskin, R.E., Steele, K.D. and Forster, W.A. (2005). Characterising Plant Surfaces for Spray Adhesion
and Retention. New Zealand Plant Protection, 58 : pp. 179-183.

Gouret, E., Rohr, R. and Chamel, A., (1993). Ultrastructure and Chemical Composition of Some Isolated
Plant Cuticles in Relation to their Permeability to the Herbicide, Diuron. New Phytologists. 124:
423-431.

Griffin, W.C. (1949). Classification of Surface-Active Agents by HLB. Journal of the Society of
Cosmetic Chemists. 1: 311.

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 94

Hess, F.D. and Foy, C.L. (2000). Interaction of Surfactants with Plant Cuticles. Weed Technology. 14:
807-813.

Knoche, M. and Bukovac, M. (1993). Interaction of Surfactants and Leaf Surface in Glyphosate
Absorption. Weed Science. 41: 87-93.

Liu, Z.Q. and Zabkiewicz, J.A. (1998). Organosilicone Surfactant Mediated Cuticular Uptake of
Glyphosate into Grasses. In: Proceedings of 5th International Symposium on Adjuvants for
Agrochemicals, Memphis, USA. pp. 119-124

Price, C.E. and Anderson, N.H. (1985). Uptake of Chemicals from Foliar Deposits: Effects of Plant
Species and Molecular Structure. Pesticide Science. 16: 369-377.

Santier, S. and Chamel, A. (1996). Penetration of Triolein and Methyl Oleate through Isolated Plant
Cuticles and their Effect on Penetration of [14C]Quizalofop-Ethyl and [14C]Fenoxaprop-Ethyl.
Weed Research. 36: 174-176.

Sanyal, D., Reddy, K.N. and Bhowmik, P.C. (2007). Surfactants Enhance Primisulfuron Activity in
Common Lambsquarters (Chenopodium album L.). Proceedings of 21st Asian Pacific Weed
Science Society (APWSS) Conference. pp. 432-436.

Seefeldt, S.S., Jensen, J.E. and Fuerst, E.P. (1995). Log-Logistic Analysis of Herbicide Dose-Response
Relationships. Weed Technology. 9: 218-227.

Singh, M., Tan, S. and Sharma, S.D. 2002. Adjuvants Enhance Weed Control Efficacy of Foliar-Applied
Diuron. Weed Technology. 16: 74-78.

Sun, J. (1996). Characterization of Organosilicone Surfactants and their Effects on Sulfonylurea
Herbicide Activity. Ph.D. Thesis, Virginia Polytechnic Institute and State University, pp. 133.

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 95

Baseline sensitivity of Echinochloa crus-galli to alternative herbicides
selected for managing herbicide resistant Echinochloa species

Ji-Soo Lim1, Soo-Hyun Lim1, Do-Soon Kim1
1 Department of Plant Science, CALS, Seoul National University, Seoul 151-742, Korea

Corresponding author: Email:[email protected]

Abstract

Acetolactate (ALS) and acetyl CoA carboxylase (ACCase) inhibitor resistant Echinochloa
species now become problematic in Korean rice cultivation. Alternative herbicides with different modes
of action can be used to control these herbicide resistant Echinochloa species. However, continuous
uses of these alternative herbicides will eventually make the Echinochloa species become resistant
to these herbicides as well. Therefore, this study was conducted to evaluate baseline sensitivity of
Echinochloa crus-galli to alternative herbicides selected for managing herbicide resistant Echinochloa
species. 63 accessions of Echinochloa crus-galli collected across Korea were tested by whole plant
assay. Six early post-emergence herbicides such as mefenacet, pretilachlor, fentrazamide, cafenstrole,
oxadiargyl, and oxaziclomefone at a range of their doses were directly treated to the flooded soil when
Echinochloa reached the 2 leaf stage. The sensitivity indices estimated by comparing the maximum
GR80 value and the minimum GR80 value was lowest for cafenstrole, 2.05, and greatest for mefenacet,
46.8. These sensitivity indices thus suggest that continuous use of these herbicides may be eventually
resulted in herbicide resistance in Echinochloa species.

Keywords: Baseline sensitivity, Echinochloa crus-galli, herbicide resistance

Introduction

Herbicide resistance is a serious worry to worldwide farming system (Holt and Lebaron, 1990;
Warwick, 1991). For management of herbicide resistance problem, EPPO suggested changing weed
management system, and suggested to investigate baseline sensitivity of about dose response(EPPO,1999).
Nowadays, in Korea, resistance herbicide weeds are also reported, about ALS inhibitor resistance
herbicide in paddy fields (Hwang, Lee, et al.,2001) and cross-resistance is also detected (Kuk, Kim,
et al.,2004). So, Korea farmer need alternative herbicides to manage herbicide resistant weeds. The
previous study suggested that some alternative herbicides could control Echinochloa spp. including
ACCase inhibitor herbicide resistant species. However, these herbicides also have resistance risk if they
are continuously used for Echinochloa control. Our understanding of baseline sensitivity of E. crus-
galli to these herbicides may help us pre-estimate potential risk of herbicide resistance development to
these herbicides but no study has been done in Korea. Therefore, this study was conducted to quantify
baseline sensitivity of Korean E. crus-galli accessions to the alternative herbicides and to assess potential
resistance risk of Echinochloa crus-galli in Korean paddy fields.

Materials and Methods

Collecting sufficient number of accessions representing a region or a country is an essential
prerequisite for the baseline sensitivity study. More than three hundred Echinochloa spp. were collected
across Korea in 2009 and 2011. Among them, 61 E. crus-galli accessions were selected based on
their regional distribution for this study. ACCase inhibitor resistant and susceptible accessions (Im, et
al.,2009), and one accession collected in Japan in 2010 were also included as a reference.

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 96

Pre-germinated seeds of each accession were transplanted in a well of open multiple-well acryl
plate, specially designed for this study, placed on paddy soil in a 0.185 m2 rectangle tray. Seedlings were
then thinned to 4 plants per well. The trays were placed in the 30/20 oC glasshouse at the Experimental
Farm Station of Seoul National University, Suwon, Korea. All experiments were consisted with three
replications of a completely randomized block design.

Herbicides were chosen based on our previous study to select alternative herbicides to control
herbicide resistant Echinochloa spp. in Korea (Bae et al. 2009, not published). Six herbicides, mefenacet,
oxaziclomefone, cafenstrole, oxadiargy, pretilachlor and fentrazamide were selected as they well
controlled Echinochloa spp. even resistant accessions. Echinochloa crus-galli accessions were treated
with mefenacet at 65.63 ~ 525 g ha-1, oxaziclomefone at 7.5 ~ 60 g ha-1, cafenstrole at 30 ~ 240 g ha-1,
oxadiargyl at 8.5 ~ 68 g ha-1, pretilachlor at 69.4 ~ 555 g ha-1, and fentrazamide at 11.9 ~ 95 g ha-1 at 7
days after sowing after flooding the tray with water at 4 to 5 cm water depth.

Visual assessments were made at 10 days and 20 days after treatment (DAT). The visual efficacy
data were fitted to the standard dose-response model (Streibig,1980) by using Genstat 5 (Genstat
Committee 1997) to estimate GR50 and GR80 values.

Results and Discussion

A total of the 63 accessions of E. cruss-galli were assessed for their sensitivity to mefenacet,
pretilachlor, fentrazamide, cafenstrole, oxadiargyl, and oxaziclomefone. GR80 values of mefenacet,
pretilachlor, fentrazamide, cafenstrole, oxadiargyl, and oxaziclomefone ranged 12.67 - 3544.84 g a.i.
ha-1, 12.20 - 372.98 g a.i. ha-1, 2.68 - 58.16 g a.i. ha-1, 34.35 - 95.21 g a.i. ha-1, 4.68 - 461.50 g a.i. ha-1,
and 0.34 - 25.81 g a.i. ha-1, respectively. Their mean values were 525.06, 122.36, 24.86, 53.31, 90.66,
and 7.25, respectively.

To analyze distribution of sensitivity, GR80 values were fitted to cumulative distribution function
by using Genstat 5 (Genstat Committee 1997). All distributions of sensitivity by each herbicide were
confirmed normal distribution. Skewness on sensitivity of E. crus-galli to mefenacet, pretilachlor,
fentrazamide, cafenstrole, oxadiargyl, and oxaziclomefone were 4.34, 1.11, 0.63, 0.82, 1.72, and 1.46,
with kurtosis of 24.44, 2.50, 1.93, -0.12, 2.32, and 2.54, respectively. All distributions of sensitivity
were right-skewed and sensitivity of mefenacet and oxadiargyl were more skewed than the others.
According to previous study, skewness could be a clue of creeping resistance. Therefore, skewed
distributions of mefenacet and oxadiargyl could suggest some processing of resistance development
(Espeby et al.,2011). On the other hand, the distributions of cafenstrole and fentrazamide indicate no or
little processing in resistance development.

4000 400

) )

-1 -1

3000 300

2000 200

1000 100

Mefenacet (g a.i. ha Pretilachlor ( g a.i. ha

0 0

Accessions

Figure 1. Sensitivity of the 63 accessions of E. cruss-galli collected in Korea from 2009 to 2010.
Scatter graph showed herbicide dose for the 80 % growth reduction (GR80 value) in sensitivity test. Each
spot represents GR80 value of each accession. The GR80 value was calculated by non-linear regression
analysis against log herbicide dose.

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 97

25 40
20
15 30
10
20
5
0 10

0 30 60 90 120 150 180 210 240 270 300 330 360

Pretilachlor (g a.i. ha-1)
% of accessions 0 0 200 400 600 800 1000 1200 ~
% of accessions
Mefenacet (g a.i. ha-1)

Figure 2. Frequency distribution of E. cruss-galli accessions for each herbicide dose for the 80 %
growth reduction (GR80 value) in sensitivity test with mefenact and pretilachlor. The GR80 value was
calculated by non-linear regression analysis against log herbicide dose.

References

EPPO. 1999. Resistance risk. EPPO Bulletin 29: 325-347. doi:10.1111/j.1365-2338. 1999.tb00838.x.
Espeby, L.Å., H. Fogelfors and P. Milberg. 2011. Susceptibility variation to new and established

herbicides: Examples of inter-population sensitivity of grass weeds. Crop Protection 30: 429-435.
doi:http://dx.doi.org/10.1016/j.cropro.2010.12.022.
Holt, J.S. and H.M. Lebaron. 1990. Significance and distribution of herbicide resistance. Weed
Technology 4: 141-149.
Hwang, I.T., K.H. Lee, S.H. Park, B.H. Lee, K.S. Hong, S.S. Han, et al. 2001. Resistance to acetolactate
synthase inhibitors in a biotype of Monochoria vaginalis discovered in Korea. Pesticide
Biochemistry and Physiology 71: 69-76. doi:10.1006/pest.2001.2565.
Im, S.H., M.W. Park, M.J. Yook and D.S. Kim. 2009. Resistance to ACCase inhibitor cyhalofop-butyl
in Echinochloa crus-galli var. crus-galli collected in Seosan, Korea. Korean Journal of Weed
Science 29: 7.
Kuk, Y.I., K.H. Kim, O.D. Kwon, D.J. Lee, N.R. Burgos, S. Jung, et al. 2004. Cross-resistance pattern
and alternative herbicides for Cyperus difformis resistant to sulfonylurea herbicides in Korea. Pest
Management Science 60: 85-94. doi:10.1002/ps.786.
Streibig, J.C. 1980. Models for curve-fitting herbicide dose response data.ActaAgriculturae Scandinavica
30: 59-64. doi:10.1080/00015128009435696.
Warwick, S.I. 1991. Herbicide resistance in weedy plants: Physiology and population biology. Annual
Review of Ecology and Systematics 22: 95-114.

The 4th Tropical Weed Science Conference, 23-25 January 2013, Chiang Mai, Thailand TWSC 2013 98

Weed and Weedy Rice Control by Imidazolinone Herbicides in
ClearfieldTM Paddy in Vietnam

Duong Van Chin1, Tran Cong Thien2, Huynh Hong Bi1, Nguyen Thi Nhiem1 and
Tran Thi Ngoc Son1

1Cuu Long Delta Rice Research Institute, Can Tho, Vietnam.
2BASF Representative Office in Ho Chi Minh City, Vietnam

[email protected] ; [email protected]

Abstract

Two experiments with two types of rice culture , dry- seeded and wet- seeded rice, were carried
out at the Experimental Farm of the Cuulong Delta Rice Research Institute (CLRRI) in Vietnam during
Spring-Summer and Summer- Autumn seasons of 2006. The tested variety was OM5749-5, an indica
rice genotype developed by plant breeders at CLRRI by crossing between promising Vietnamese indica
rice with IMI-tolerance japonica rice from Louisiana State University, USA. The tested herbicides are:
imazapic, imazapyr, imazapic+imazapyr and imazethapyr+imazapyr. Results revealed that common
weeds observed in the experimental field including Echinochloa crus- galli, Leptochloa chinensis,
Cyperus iria, Cyperus difformis, Ludwigia octovalvis and especially weedy rice (Oryza sativa) were
controlled successfully by the herbicides. The density and dry weight of weeds were brought down
significantly as compared to those under untreated check. In dry seeded rice, the average rice grain yield
of six imidazolinone treatments is 1.83 t/ha, 101.1% higher than check (0.91 t/ha). The corresponding
data in wet-seeded rice are 2.15 t/ha and 0.88 t/ha with 143.9% increment. Rice quality is also improved
by the treatments. The number of contaminated weedy rice seeds in rice product, seeds dropped in the
soil surface, the percentage of red grains in milled rice is also reduced under herbicide treatments as
compared to that of check statistically.

Keywords: ClearfieldTM paddy, herbicide tolerant variety, imidazolinone, weedy rice control

Introduction

Weedy rice, commonly considered as ecotypes of Oryza sativa, is a new pest in rice growing
countries in the world including Vietnam. In tropical areas, weedy rice is progenies of crosses between
wild rice and cultivated rice or come from degradation of cultivated rice varieties. The major characteristic
of weedy rice is easy shattering. Other characteristics are observed as taller plants, fewer tillers, and
high percentage of red rice in milled rice (Chin et al. 2000). Weedy rice competes with cultivated
rice for sunlight, water and nutrients resulting in reduction in rice yield. The quality of milled rice is
reduced due to contaminated red rice. Weedy rice infestation in rice fields is dangerous because seeds
in seed bank increase over time with self-regeneration and there is no effective selective herbicide for
controlling weedy rice. Recently, a new option for controlling weedy rice and also common weeds in
rice fields has been initiated by exploring the integration of imidazolinone herbicides and tolerant trait
containing variety (which is called CLEARFIELD TM rice). Imidazolinone herbicides controls weeds
by inhibiting the plant specific enzyme acetohydroxyacid synthase (AHAS), which is involved in the
biosynthesis pathway of the branched-chain amino acids as valine, leucine and isoleucine. This inhibition
causes a disruption of protein synthesis, which interferes DNA synthesis and cell growth (Shanner and
Connor 1991). CLEARFIELDTM rice has been developed by Louisiana State University Agricultural
Center breeders through a combination of mutagenesis and conventional plant breeding, which is
tolerant to imidazolinone herbicides. This is characterized as a non-GMO variety. In Vietnam, CLRRI
plant breeders have successfully developed indica rice genotypes, which are tolerant to imidazolinone
herbicides. The research aims at determining whether the integration of imidazolinone herbicides and
tolerant trait in the genotypes can be used for controlling weedy rice and common weeds in rice fields