Study On The Distribution, Aflatoxin Production And Genetic Diversity Of Aspergillus Flavus In Soils Of Peanut Fields In Four Agroecological Zones Of China

Aflatoxin contamination is an important limiting factor for the growth of China’s peanutindustry. Aflatoxins are the secondary metabolites formed mainly from the Aspergillus flavus andA. parasiticus. Soil is the main source of inoculum for A. flavus and since peanut developunderground, pods are in direct contact with the soil fungal populations. The study was firstly,systematically researched on the distribution and production of aflatoxin of A. flavus in soils ofpeanut fields in four agroecological zones （Southeast coast、Yangtze River Basin、Yellow RiverBasin and Northeast area） of China and revealed the distribution and aflatoxin production of A.flavus. And the genetic diversity of A. flavus from the four agroecological zones was studied inorder to grasp genetic variation and it revealed the relationship of genetic variation and geographicorigin. The study could provide a theoretical basis and technical support for the prevention ofaflatoxin contamination in peanut.The results are listed as follows:1. Screened the suitable medium for the isolation of A. flavus. Seven agar media used to isolatemildew were compared for utility in isolating fungi in the A. flavus from soils of peanut fields.The seven agar media were Dicloran18%glycerol medium、High salt Czapek medium、Patatodextrose agar medium、Modified rose Bengal agar medium、YES medium、Bengal medium、Dicloran Bengal agar medium. The results showed that there was a big difference in seven mediain isolating A. flavus. DG18was the most useful for studying the population biology of this groupbecause it permitted both identification of the greatest number of A. flavus group strains andgrowth of the fewest competing fungi. DG-18supported about7.3times more A. flavus coloniesthan the PDA while reducing the number of mucorales colonies by81%than the DRBC. So DG18should be the most useful for studies on the population biology of the A. flavus group.2. Isolated324strains of A. flavus and20strains of Aspergillus parasiticus.344strains ofaflatoxin producing Aspergillus species were isolated from600soil samples of20peanut fields infour agroecological zones.323（94.2%） were A. flavus and20（5.8%） were A. parasiticus. Inpeanut soil population of aflatoxin producing Aspergillus species, A. flavus was found to be thedominant species.3. Mastered the distribution of A. flavus in soils of peanut fields in four agroecologicalzones of China. The distribution of A. flavus isolates significantly varied among the districts andagroecological zones of China. The largest number and the most widely distribution of A. flavuswas Yangtze River Basin (the densities of A. flavus in soils was1039.3cfu·g^-1 and the positiverate of A. flavus in soils was80.7%). The second was Southeast coast, and the third was YellowRiver Basin. The last was Northeast area (The densities of A. flavus in soils was only2.4cfu·g^-1and the positive rate was6.6%). The positive rate of A. flavus in soils and the temperature had thenotable positive correlation. There was the notable positive correlation between the densities of A.flavus in soils and the temperature. And the densities of A. flavus and the longitude had negative correlation. The A. flavus NS-strain was the most commonly isolated member of A. flavus（41.59%） across the four examined agroecological zones, and the second was A. flavus S-strain.The last was L-strain.4. Mastered the aflatoxin production of A. flavus in soils of peanut fields in fouragroecological zones of China. The aflatoxin producing ability was quantified for343isolatedstrains by liquid fermentation with HPLC.69.19%of the tested isolates were toxigenic A. flavusand the percent of atoxigenic A. flavus was30.81%. The aflatoxin producing potential of A. flavusisolates significantly varied among the districts and agroecological zones of China. A. flavusisolate of the Southeast coast zones had the largest aflatoxin producing potential (averagedaflatoxin production was5836ng·mL^-1), the second was Yellow River Basin, the third wasNortheast area and the last was Yangtze River Basin. Incidence of toxigenic A. flavusisolates(>1000ng·mL^-1) was largest in the Southeast area （54.4%）, the second was Yellow RiverBasin,（31.8%）, the last was Yangtze River Basin （10.2%）.The distribution of the toxigenic andatoxigenic A. flavus isolates varied among the districts and agroecological zones of China.Incidence of atoxigenic A. flavus isolates was largest in the Yangtze River Basin （40.2%）, thesecond was Northeast area （36.4%）. The last was Southeast coast zones （15.7%）.5. Estimated the risk of aflatoxin contamination of peanut. Yangtze River Basin had thelargest aflatoxin producing potential in1g soils (1627544ng·mL^-1in1g soils), the second wasNortheast area (1117594ng·mL^-1), the third was Yellow River Basin (18602ng·mL^-1). The lastwas Northeast area (4478ng·mL^-1). So highest risk of aflatoxin contamination of peanut was inYangtze River Basin, the lowest was Northeast area.6. Established ISSR-PCR method for genetic diversity of A. flavus. The optimal reactionsystem of ISSR-PCR amplification was optimized. Of the100primers, the primers UBC834,UBC809, UBC817, UBC895, and UBC899were selected for for genetic diversity of A. flavus.The range of genetic similarity coefficients was from0.59to0.90. The study showed that theISSR technology is an effective molecular approach for studying diversity of A. flavus.7. Mastered the genetic diversity of A. flavus in soils of peanut fields in four agroecologicalzones of China. The genetic diversity of343A. flavus isolates was researched with ISSR. Thecluster analysis showed that the ISSR groups were closely related with geography origin. Theisolates were classified into four large groups. All the strains of Northeast area were in group Ⅰ,and all the strains Yellow River Basin were in group IV.88.9%strains of Yangtze River Basinwere in group Ⅱ. All the strains of Yellow River Basin excepted for8strains were in group Ⅲ.The ISSR groups were not notable correlation with sclerotia character and aflatoxin production.