Proceedinq« of the 7th Internatumal Worh't11JConference on Stored-product Protection - Yolume 1

Studies on seed-borne fungi in hybrid rice and controlling effects of some fungicides

Lmo Xiaolan, Luo Kuan1

Abstract The Isolation of Seed-borne Fungi Seed-borne fungi in 19 hybnd rice vane ties and 3 One hundred seeds were selected randomly from each conventional nee vane ties from Southern, Central and sample (more than 100 seeds for same samples) and shelled. Northern parts of Provmce were exammed The fungi- mfected place and the mechamsm of thetr mfection to The dilution platmg method was used for isolation, and the tested seeds were also studied. The relation between the enumeration and Identification work was performed after seed mfection rate and seed germmation rate was exammed incubated at 26 t: for 7 days. and analyzed. Several effective fungicides were selected for Effect of seed-borne fungi on seed germination seed treatment. One hundred seeds are selected randomly from each sample (more than 100 seeds for same samples) and shelled. Soaked m water for 24 hours, then put on wet filter paper in stenlized perti dishes. The germinatmg rates of the seeds Introduction were counted after mcubatmg at 26°C for 10 days. The effect of fungicides on seed germination Extensive and deep studies on seed-borne fungi of nee have been carried out all over the world (Imelenm E. D. 1983; One hundred seeds were randomly selected from each Zainum W. and Nik H. W. 1977). In , some studies sample (more than 100 seeds for same samples). After on pathogens as Piricularui Oryzae, Helminthoeporiuni soaked in water for 24 hours, the seeds were left soak in one oryzae, Gibberella fUJ'/,kuroi wollen-weber earned by seeds of the followmg fungicide solutions for another 24 hours: of conventional nce have been made, but httle on that of Copper hydroxide (1 500 ), Mancozeb (1 . 500), 40 % hybrid rice (Zhou Zhenyue 1988; Liao Xiaolan, Luo kuan. Miejunwei (l 500), 45 % Thiabendazole (1 1000), 50 % 1993) This study covers the type and quantity of seed-born Procymidone (1 1000), 50% Metalaxy + DT (1 500), fungi in hybrid rice in some districts of Hunan Province, and 25 % Pukela (1 2000), t 500 Zhongjunling. Or first soaked investigates their effects on seed germination and the in water for 12 hours and then in TCCA (1 400) for another controlling effects of some fungicides. The research alms to 12 hours The treated seeds were washed With water, and provide theoretical and practical baSIS from the pathological put on moistened filter papers in sterilized petn dishes. aspect of seed to mcrease the germmatmg rate of hybrid nee Checked the numbers of germinated seeds and counted the seeds germmatmg rates after mcubated for 10 days at 26t: . The effect of fungicides on seed-borne fungi Materials and Methods Treated as above described, the seeds were shelled, and Experimental Material analyzed for fungi by the direct plating method. Check the number of seeds contammated by fungi after incubation for 7 Twenty two samples of nee seed (rnamly hybnd nce ) days and count the fungal contaminated rates and fungicidal were collected from Hunan Academy of Agncultural SCIence, effects. Hunan Agricultural University, Hejiasan Farm, County, Prefecture and Prefecture, among Pathogenic characteristics of Alternaria padwickii these samples, 3 has been stored for 2 years, one was the Dunng bootmg stage and heading stage, hybrid nee parent of hybrid nee and 4 were conventional rice vanety V64, V6, Shangyou 63 and V20A were inoculated with Alternaria padwickii strams AP-1, AP-1, AP-2, AP-3 and AP-4 by spraymg and ejecting their spore suspension 1 Hunan Agncultural College, Changsha 410128, Hunan, PRof (1.05 X 108 spores per ml ). Carefully harvest paddy kernels Chma and examme their fungal contammated rates at late dough 268 Proceedzngs of the 7th Internatwnal Worhng Conference on Stored-product Protection - Volume 1 stage In seeds stored for less than 1 year, vanety V46 held the highest contammated rate of 35 % and variety V64 the Check infecting place on seeds and infecting lowest of 12% In seeds stored for 2 years, the highest rate mechanism of fungal contammation was 40% m vanety Imyou 307, and After paddy husk removed, nee seeds of vanety V64 from the lowest was 24 % m vanety Jmyou 080. Hejiasan Farm, Shangyou 64 from Hunan Academy of Comparison of fungus contamination between Agncultural Science, were subjected to separation of embryo hybrid and common rice and endosperm albumen by traditional method, and exammed the fungus contammated rates of embryo and Fungus contammation rates in seeds of hybrid and endosperm albumen respectively. RIce seeds of vanety V64, common nce from Hunan Academy of Agricultural Science V6, Shangyou 63 and V20A from Hejiasan Farm were sown were detected The results are showed m table3. Both m cement pools, 25 to 30 growmg plants were Isolated and hybrid nee and common nee carry fungi in seeds A. exammed for their contammated SItuation withm 7 days at 3 puiunckii IS the most common detected fungus, with a leave stage, internode enlongmg stage, booting stage and detected rate of 66% -74% for hybrid nce and 60% -75% headmg stage. for common nee. The results show that hybnd and common rice are similar m types and rate of contammated fungi. Resul ts and Discussion Relation between fungus infection and germination of seed Fungi carrying situation of hybrid paddies in different areas Seeds WIth different fungus infection rates were used to do gerrnmation test Results show that the fungus infection Hybnd nce seeds from 7 areas of Hejiasan Farm and the rate has no sigruficant effect on seed germination. Fungus others were exammed for fungi. The resul t showed tha t the contammation rates for seeds of vanety Shanyou 62, Weiyou hybnd nee seeds from all different areas m Hunan Province 6, Weiyou 64 and V20A from Hejiashan Farm were carried fungi, WIth the highest contammated rate of 97% 95.81 %, 93.23%, 91.29% and 61.61%, and their found on seeds of vanety V49 from Anren County and the germmation rates were 96.00%, 98.00%, 91.67% and lowest rate of 57% on seeds of Shangyou 64 from Changsha. 84.33% respectively. Fungus contammation rates for seeds Alternaria padwickeh was the most dominate one Its of vanety Shanyou 63, Shanyou 64, Weiyou 46 from contammated rates were all above 50%, WIth the highest of were 62.00 %, 57.23 %, 71.00 %, and 92 % (table 1). The contammated rates of the other strams their germmation rates were 78.40%, 73.00%, 31.20% as Illeuabar clayana, H. oruzae , Fuscrium sp., respectively Fungus contaminauon rates for seeds of Curoularui sp , Alternarui sp were 3% - 9%, 4% - vanety Shanyou 64, Shanyouwan 3, Weiyouhuabian 2 from 10%, 1% - 7%, 2% - 10% and 2% - 6% respectively. Hunan Academy of Agncul tural Science were 68.00 % , Table 1 also shows that there were ObVIOUSdifferences m 72.00%, 84.00%, and their germmation rates were fungal contammated rates between parents and F1' s , fungal 76.50%, 85.71%, 76.02% respectively. The correlation contammated rates of F1 ' s of vanety Shangyou 63, V6 and coefficient between fungus contammation and gerrmnation V64 reached more than 90%, whereas that of parent V20A rate IS 0.3523, which IS not srgmficant at 0.05 level in the was 61.61 % T-test. The results show that the germmation rate of seeds Situation of fungus contamination in seed of m next year IS not closely related to fungus infection rate in different hybrid combination hybrid rice. The germmation rates of both hybrid nee and common Fungal contaminated rates m different F1 ' s were also nee seeds WIth different fungus infection rates in the same exammed The results showed (Table 2) that all the seeds area were also compared. The fungus infection rates of were contammated by fungi at a relatrvely low rate, WIth hybnd nce Shanyou 64, Shanyouwan 3 and Welyouhuahan 2 the hIghest of 40 % occurred m vanety Jmyou 307. The were 68 00 %, 72.00 %, 84.00 % respectively, and their contammated rates m seeds stored for 2 years were hIgher germmation rates were 76.70 %, 85.71 %, 76.02 %. The than that stored for 1 year. Alternarw padwwkh was still fungus infection rates of common nce Xlangzhongxmg 2, the most dominate fungus, WIth a contammated rate Xiangzhongxmg 3, 86 - 106 were 79.00 %, 87.00 % , between 32 % and 92.3 %; Curvularza spp were the 82.00%, and theIr germmation rates were 95.60%, second frequent detected fungI; Fusarzurn spp., 90.40 %, 90. 19 % respectively. The germmation rate of Cladosporzum spp. and ASpergLllus spp. were also found common nce seeds IS hIgher than that of hybnd nce seeds.

269 Proceedings of the 7th Internatumal Workwg Conjerence on Stored-product Protectwn - Volume 1

Table 1. Isolatmg results of seed-born fungi in hybrid rice in different areas

Number of Number of Infected A. puiunckai Ratio of C Area Hybrids seeds (A) fungi (B) rate (%) number (C) toB(%)

Hejiashan Farm (Northern) 4 1240 1060 85.48 874 70.48

Changsha County (Central) 3 300 190 63.33 168 88.47

Aren County (Southern) 1 100 97 97 00 92 92.00

Table 2. Isolating result of seed-born fungi m different hybrid rices.

No hybrids Seeds F-l F-2 F-3 F-4 F-5 F-6 F-7 Total Infected Rate of used A fungi rate/ %) A(%)

1 Shangyou 63 100 10 3 0002 16 31 31 32.26 2 Shangyou 77 100 12 1 0 020 0 15 15 80.00 3V46100 15 7 0 12 1 0 03535 42.86 4 V64 100 10 0 0 101012 12 83.33 5 V77 100 18 0 0 2 2 1 0 23 23 78.26 6 Peilianyou 288 100 12 0 0 100 0 13 13 92.30 7 YUCh1231-8 100 29 28 2303 2 67 67 43.28 8 Jinyou 080 100 4 2 0 0221424 24 16.67 9 Jmyou 091 100 7 1 0 0041527 27 25.93 10 Jmyou 307 100 23 4 0 0121040 40 57.50

Note: (1) No 1 to 6 are seeds of hybrid nee stored for 1 year; No 7 IS seed of common nee stored for 1 year; No 8 to 10 are seeds of hybrid nee stored for 2 years (2) F-l, F-2, F-3, F-4, F-5, F-6 and F-7 stands for Alternaria puiuncku , Curoularui sp , Tilleiui barclayana , Fusanu:m spp , Aspergtllus spp , Cuuiosporium spp respectively

Table 3. Fungus mfected rate in seeds of hybrid and common nee.

Number of Fungus Fungus A padwwkii A. padwichi Type Variety seeds used mfected mfected Infected mfected rate seeds rate (%) seeds (%)

Common nee A 100 79 79.00 60 60.00

B 100 87 87.00 75 75.00

C 100 68 68.00 66 66.00

Hybrid nee D 100 68 68.00 66 66.00 E 100 72 72.00 69 69.00

F 100 84 84.00 74 74.00 Note-A, B, C, D, E and F stands for nee vanety Xiangzhongxmg 2, Xiangzhongxing 3, 86 - 106, Shangyou 64, Shangyouwan 3 and Weiyouhuahan 2 respectively

500 Metalaxy + DT (50%), i 500 Zhongjunlmg, 1 2000 Effects of fungicides on seed germination fungus Pukela (25%) for vanety Weihualian 2 (FIgure 1). Among infection the 9 kinds of fungicides, 1 500 Zhongjunlmg showed the After the seeds were treated by fungicides mentioned best fungus-killmg effect, the isolation rate of seedborn above, no adverse effect on seed germmation was observed Alternaria padunckeii IS 0% and the fungus-killing except m the treatment of variety V20 seed WIth 1: 500 of efficiency is 100% after the treatement. The second best copper hydroxide one is Miejunwei (density 40%), ItS fungus-killing rate IS The seed germination rate mcreases after treatred WIth 1 between 25.89 % to 79.81 %. The effects of the other 100 procymidone (50 %) for vanety Shangyouwan 3 and 1 fungicides are ineffective or greatly varied

270 Proceedings of the 7th International Worktng Conference an Siored-proiuci Protectuni - Volume 1

100 ..... - • • 100 80 • • 80 60 ,A g- .... -2 60 Jj.··· ... ···-A·.-_A··· 40 40 ~-iJ-2 20 20

II o 0

r=+=1l l.::!::1J

2 3 4

Fig.l. Effect of fungicide treatment on seed gemunation and A. padwicken mfection rates m seeds

Note: (1) Figure left-up shows the results of Weiyou 6; mark 1, 2, 3, 4 and 5 at the abscissa stand for treatments of 1 500 copper hydroxide, 1 500 Mancozeb, 1 500 Trichloroisocyan nne acid, 1 500 Miejunwei (densrty 40%) and the control (water) respectively; curve 1 and curve

2 show germmation rates ( % ) and Aiternaria padwtcken-carrymg rates (%) (2) Figure nght-up 1S for V20A; mark 1, 2, 3, 4 and 5 at the abscissa stand for treatments of 1 500 copper hydroxide, 1 500 mancozeb, 1 500 Tnchloroisocyan nne acid, 1 500 Miejunwei (density 40%) and the control (water) respectively; curve 1 and curve 2 show the germmation rates (%) and Alternarui paduncken-carrymg rates (%) respectively (3) Figure down-left 1S for Shangyouwan 3; mark 1, 2, 3, 4 and 5 at the abscissa stand for treatments of 1 500 Thiabendazole (45 % ), 1 100 procyrmdone (50 % ), 1 500 Metalaxy + DT (50 % ), 1 500 Zhongjunlmg, 1 2000 Pukela (5 %) and control (water) respectively; curve 1 and curve 2 mean germination rate (%) and Alierruirui padwtCken-carrymg rate (%) (4) Figure down-nght 1S the results of Weihuahan 2; mark 1, 2, 3 and 4 at the abscissa stand for treatments of 1 500 metalaxy (50%), 1 500 Zhongjunlmg , 1 2000 Pukela (5 %) and control (water) respectively; curve 1 and curve 2 show the germmation rates (%) and Alternarui padWtCken-carrymg rates (%)

shows that there are fungi in both embryo and endosperm Pathogenic characteristic of Alternaria padwickii albumen. Fungal infected rates of variety Weiyou 64 are The detected rates of Alternarw podioickeii in WelYou 6 50% - 70% in embryo and 75% - 87.5% m endosperm and Shangyou 63 increased after inoculated WIth 4 strains by albumen Those of vanety Shangyou 64 are 2.5 % - 7.5 % two inocula tmg methods The increase in detected rates was in embryo and 10% -15% m endosperm 1 % to 33% and 2% to 24% respectively (see table 4). In the systematic detection of fungus infection, only one Browning kernels increased after spike ejection moculation, nee plant m each vanety of Weiyou 6 and Weiyou 64 was whereas only strain Ap-3 caused an increase m browning detected With A. padwicketi infection m three leave stage, kernels by solution-spraying inoculation. Table 4 also shows booting stage and heading stage. The result indicates that that there IS no correlation between detected rate of among the A padunckeu. that contaminated the seed Alternana padwickell and kernel brownmg embryo, httle could cause systematic infection m the outgrowing plant and ItS new seeds. Checking of infection place on seeds and mechanism of infection

Test of infected places on hybnd seed by A padwickeit

271 Proceeunas of the 7th Internatwnal Worki'tlf}Conference on Stored-product Protection. - Volume 1

Table 4. Test of pathogenic character of Alternaria txuluncki: (AP for short)

Black Blackenmg Tested AP Rate Hybrid Inocula non Inoculated Number pointed rate by AP seed mfected mcreased vanety method strain of seeds kernels (% ) numbers ratet %) (% ) AP-1 660 23 3.48 100 61.00 8.00 spike- AP-2 533 20 3.75 100 61.00 8.00 ejecting AP-3 537 55 10.24 100 86.00 33.00 AP-4 734 13 1.77 100 60.00 7.00 WeIYou AP-1 1176 4 0.34 100 55.00 2.00 6 AP-2 1155 3 0.26 100 76.00 23.00 Solution AP-3 1030 10 0.97 100 54.00 1.00 spraymg AP-4 709 14 1.97 100 58.00 5.00 CK 584 4 0.68 100 53.00 AP-1 464 15 3.23 100 60.00 24.00 Spike- AP-2 310 7 2.26 100 38.00 2.00 ejecting AP-3 472 5 1.06 100 39.00 3.00 AP-4 811 6 0.74 100 59.00 23.00 Shan you AP-1 560 4 0.71 100 42.00 6.00 63 AP-2 490 3 0.61 100 52.00 16.00 Solution AP-3 638 11 1.72 100 58.00 22.00 spraymg AP-4 873 6 0.69 100 50.00 14.00 CK 482 5 1.03 100 36.00

Reddy O. R. (1984) and Shetty S. A. (1989) reported Conclusions that Tnfonne, Carboxin, Thiophanate-methyl plus thiram, Kasugamycm, Mancozeb and the extracts of the Strychnos Shetty S. A (1987) reported that A. puiunckei was the fruit, garlic, ginger have controllmg effects on A. pathogen of nee spot disease. Tolentmo V. (1989) studied 9 padwickeii. In this expenment, 9 fungicides were used to types of fungi m 2 india varieties stored for 20 years, and treat seed with Zhongjunlmg from Hunan Dongkou Factory, found that both A. padioickeu. and Curoularui sp were which demonstrated the best effect and Mancozeb showed detected in these 2 vane ties , but Fusarium. spp. and different effects III different nee vaneties. Further research Drechslera spp. were detected only in one vanety. IS needed concerning the optimum workmg concentration The results of the present study are smular to those of and mechanism of Zhongjunhng. former researchers in that A. padwickeii IS the dommant seed-born fungus m both common nee and hybrid nee, and References few A padunckeii m seeds mfects the filial generation through seed gerrrunation It IS necessary to do further Chen Lianbi. 1990. Observation of endosperm's albumen of research work on the source of pnmary infection, time and nee seed during germmation WIth electronic-scope. Journal approach of A. padwicke~i infection and etc. of Natural Science of Hunan Normal University, 13, 1, 71 QUite a lot of fungi as A. puiunckeu , Curoularui spp. , -73 Tuletia barclayana, Fusarium spp. , Aspergillus spp , Imelenm E. D. 1983. Rice seed-born fungi and theireffect Cladosporium spp. and some other fungi were detected m on seed germmation. Plant DIsease, 67, 1334 -1336. new and stored seeds. That means seeds m the seed Liao Xiao-lan, Luo kuan. 1993. Study on seed-born fungi of production base of Hunan Province carry many types of fungi hybrid and treatmg seeds WIth chemical. Hybrid Rice, 6, mcluding some pathogens. 28-32 For hybrid seeds, there IS no significant correlation Reddy.O. R. 1984. Evaluation of tnfonne (Saprol) against between the fungal infected rates and their germmation seed-borne pathogens of rice (Oryza sativa). Int. J rates m next year. The gerrrunation rate of common rice IS Trop. Plant Dis. , 2, 133 - 138 higher than that of hybrid nee after 2 years' storage. Shetty, SA, Prakash, H. S. and Shetty, H. S. 1989 Further research IS needed to investigate the effect of fungal Efficacy of certam plant extracts against seed-borne infection on seed germination of hybrid rice infection of Trichoconiella padwickii infection in paddy 272 Proceedings of the 7th International Worktng Conference an Stored-product Protectwn - Volume 1

(oryza sativa). Can. J Bot. 67, 1956 - 1958. fungi of nee in Malaysia. Malay. Appl. Bioi, 6, 67 -74. Shetty, S. A., Sharada, R. and She tty , H. S. 1987. Zheng J inchai. 1989. The course and measure of low rate of Technique to detect grain spot of rice caused by hybnd rice seed, Hybrid RIce, 1, 13 -15. Trichoconwlla padwickii (Ganguly) Jam. Curro Sci., Zhou Zhenyue. 1988. The mechanisms of low germination 56, 32 - 33. rate and seedhng-becoming rate of hybnd rice. Crop Zamum W. and Nik H. W 1977. A survey of seed-borne Research, 2, 4, 23

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