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Alternanthera Philoxeroodes Biocontrol Potential of Pathogenic Fungi Research

Author: WangXiaoYang
Tutor: DongLiYao
School: Nanjing Agricultural College
Course: Pesticides
Keywords: Nimbya alternantherae toxin filtrate crude toxin herbicidal bioactivity 2-(2-acetyl-3,5-dihydroxyphenyl) acetic acid
CLC: S451
Type: Master's thesis
Year: 2009
Downloads: 10
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Abstract

Twenty fungi strains (named as NCH1, NCH2......NCH20) were isolated from the infected Alternanthera philoxeroides. The fungi were taupe in PSA culture medium when at the beginning of separate. The russet coloring matter was produced, and the colour of the mycelial was from achromaticity to sandy beige. The inverse of colony was snuff color. But the conidiophore was not produced. It was identified as Nimbya alternantherae in laboratory of antiseptic, college of plant protection, Nanjing agricultural university. Isolate NCH1 was selected to test its biological characteristics. NCH1 grows more quickly than other strains on OA, PC A, V8 and ZZJZ culture medium. The colony diameter of NCH1 was 7.90±0.00cm when cultured for 13 days. But the growth speed was the slowest on Czapek culture medium, NCH1 diameter was only 2.66±0.62cm when cultured for 13 days. The strain of NCH1 can not produce spore in 10 kinds of culture medium when cultured for 7 days, 14days and 21 days.The optimum strains and culture conditions for N. alternantherae toxin production was studied. The results showed that NCH4 was the optimum strain of toxin production. The lowest percentage of germination of E. colonum was 23.33%treated by culturing filtrates of NCH4, while the inhibition rate of root and shoot length were the highest (96.51%and 100.00%, respectively). The results showed that the optimum culture conditions were listed as follows:25℃with full dark, pH 7.0 and shake cultivation (150rpm) for 7 days.According to optimum conditions for toxin production obtaind the toxin filtrate, which was plenitude extraction 5 times by equal to volume ethyl acetate. Combination ethyl acetate, the solvents were removed under reduced pressure in a rotary evaporator at 35℃and the toxin was received. According to seed germination bioassay, the herbicidal bioactivity of crude toxin to twelve common weeds and the satety to six crops were studied. The results showed that the toxin could inhibit the seeds germination, root and shoot length, and the inhibition was increased by the increasing of concentration. The percentage germination of E. colonum, Capsella bursa-pastoris and Digitaria ciliaris was restrained in a certain extent in phytotoxin concentration 50μg·mL-1. When the phytotoxin concentration was 400μg·mL-1, the percentage germination of C. bursa-pastoris and Leptochloa chinensis were both zero. The root and shoot length of Eclipta prostrate, E. colonum, D. ciliaris, Beckmannia syzigachne and Alopecurus aequalis were also inhibited at phytotoxin concentration of 50μg·mL-1. The crude toxin had very strong inhibition with root and shoot length of C. bursa-pastoris, L. chinensis, Poa annua and Amaranthus retroflexus in phytotoxin concentration 400μg·mL-1. The inhibition of root and shoot length were 100%, 100%; 100%,100%; 92.11%,74.49%; 90.31%,59.60%; respectively. The crude toxin had different effection on the germination of different crops. It couldn’t inhibit the germination of Arachis hypogaea. On the contrary, it could promote the root length of A. hypogaea. When the phytotoxin concentration was less than 400μg·mL-1, the germination, root and shoot length of Brassia campestris were not inhibited. The crude toxin was not safe to Triticum aestivum.By silica gel column chromatography and Sephadex LH-20, the toxin was separated and purified. The results showed that the eluent of 100:2 (chloromethane/methanol) (v/v) exhibited the strongest herbicidal bioactivity. When the phytotoxin concentration was 400μg·mL-1, the inhibition rate of root and shoot length of C. bursa-pastoris were 71.99% and 62.83%, respectively.100:8 (Chloromethane/methanol) (v/v) also had strong herbicidal bioactivity. When the phytotoxin concentration was 400μg·mL-1, the inhibition rate of root and shoot length of C. bursa-pastoris were 64.86%and 47.20%, respectively. In the toxin the herbicidal bioactivity substance most focus on 100:2 and 100:8. We gained a compound from 100:2. The compound was curvulinic acid according to MS,13C-NMR and 1H-CMR. At a concentration of 600μg·mL-1, curvulinic acid caused 73.54%and 66.67%growth inhibition of root and shoot with EC50 value of 204.67 and 281.08μg·mL-1, respectively. And the percentage of germination of C. bursa-pastoris was 73.33%.

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