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Biocontrol activity of volatile organic compounds from Streptomyces alboflavus TD-1 against Aspergillus flavus growth and aflatoxin production
Mingguan Yang , Laifeng Lu , Jing Pang , Yiling Hu , Qingbin Guo , Zhenjing Li , Shufen Wu , Huanhuan Liu , Changlu Wang
J. Microbiol. 2019;57(5):396-404.   Published online May 6, 2019
DOI: https://doi.org/10.1007/s12275-019-8517-9
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  • 43 Web of Science
  • 42 Crossref
AbstractAbstract
Aspergillus flavus is a saprophytic fungus that contaminates crops with carcinogenic aflatoxin. In the present work, the antifungal effects of volatile organic compounds (VOCs) from Streptomyces alboflavus TD-1 against A. flavus were investigated. VOCs from 8-day-old wheat bran culture of S. alboflavus TD-1 displayed strong inhibitory effects against mycelial growth, sporulation, and conidial germination of A. flavus. Severely misshapen conidia and hyphae of A. flavus were observed by scanning electron microscopy after exposure to VOCs for 6 and 12 h, respectively. Rhodamine 123 staining of mitochondria indicated that mitochondria may be a legitimate antifungal target of the VOCs from S. alboflavus TD-1. Furthermore, the VOCs effectively inhibited aflatoxin B1 production by downregulating genes involved in aflatoxin biosynthesis. Dimethyl trisulfide and benzenamine may play important roles in the suppression of A. flavus growth and production of aflatoxin. The results indicate that VOCs from S. alboflavus TD-1 have tremendous potential to be developed as a useful bio-pesticide for controlling A. flavus.

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Characterization of the velvet regulators in Aspergillus flavus
Tae-Jin Eom , Heungyun Moon , Jae-Hyuk Yu , Hee-Soo Park
J. Microbiol. 2018;56(12):893-901.   Published online October 25, 2018
DOI: https://doi.org/10.1007/s12275-018-8417-4
  • 46 View
  • 0 Download
  • 32 Crossref
AbstractAbstract
Fungal development and secondary metabolism are closely associated via the activities of the fungal NK-kB-type velvet regulators that are highly conserved in filamentous fungi. Here, we investigated the roles of the velvet genes in the aflatoxigenic fungus Aspergillus flavus. Distinct from other Aspergillus species, the A. flavus genome contains five velvet genes, veA, velB, velC, velD, and vosA. The deletion of velD blocks the production of aflatoxin B1, but does not affect the formation of sclerotia. Expression analyses revealed that vosA and velB mRNAs accumulated at high levels during the late phase of asexual development and in conidia. The absence of vosA or velB decreased the content of conidial trehalose and the tolerance of conidia to the thermal and UV stresses. In addition, double mutant analyses demonstrated that VosA and VelB play an inter-dependent role in trehalose biosynthesis and conidial stress tolerance. Together with the findings of previous studies, the results of the present study suggest that the velvet regulators play the conserved and vital role in sporogenesis, conidial trehalose biogenesis, stress tolerance, and aflatoxin biosynthesis in A. flavus.

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An efficient Agrobacterium-mediated transformation method for aflatoxin generation fungus Aspergillus flavus
Guomin Han , Qian Shao , Cuiping Li , Kai Zhao , Li Jiang , Jun Fan , Haiyang Jiang , Fang Tao
J. Microbiol. 2018;56(5):356-364.   Published online May 2, 2018
DOI: https://doi.org/10.1007/s12275-018-7349-3
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AbstractAbstract
Aspergillus flavus often invade many important corps and produce harmful aflatoxins both in preharvest and during storage stages. The regulation mechanism of aflatoxin biosynthesis in this fungus has not been well explored mainly due to the lack of an efficient transformation method for constructing a genome-wide gene mutant library. This challenge was resolved in this study, where a reliable and efficient Agrobacterium tumefaciens-mediated transformation (ATMT) protocol for A. flavus NRRL 3357 was established. The results showed that removal of multinucleate conidia, to collect a homogenous sample of uninucleate conidia for use as the transformation material, is the key step in this procedure. A. tumefaciens strain AGL-1 harboring the ble gene for zeocin resistance under the control of the gpdA promoter from A. nidulans is suitable for genetic transformation of this fungus. We successfully generated A. flavus transformants with an efficiency of ~ 60 positive transformants per 106 conidia using our protocol. A small-scale insertional mutant library (~ 1,000 mutants) was constructed using this method and the resulting several mutants lacked both production of conidia and aflatoxin biosynthesis capacity. Southern blotting analysis demonstrated that the majority of the transformants contained a single T-DNA insert on the genome. To the best of our knowledge, this is the first report of genetic transformation of A. flavus via ATMT and our protocol provides an effective tool for construction of genome-wide gene mutant libraries for functional analysis of important genes in A. flavus.

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Isolation and characterization of Aspergillus flavus strains in China
Firew Tafesse Mamo , Bo Shang , Jonathan Nimal Selvaraj , Yan Wang , Yang Liu
J. Microbiol. 2018;56(2):119-127.   Published online February 2, 2018
DOI: https://doi.org/10.1007/s12275-018-7144-1
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AbstractAbstract
Important staple foods (peanuts, maize and rice) are susceptible to contamination by aflatoxin (AF)-producing fungi such as Aspergillus flavus. The objective of this study was to explore non-aflatoxin-producing (atoxigenic) A. flavus strains as biocontrol agents for the control of AFs. In the current study, a total of 724 A. flavus strains were isolated from different regions of China. Polyphasic approaches were utilized for species identification. Non-aflatoxin and non-cyclopiazonic acid (CPA)-producing strains were further screened for aflatoxin B1 (AFB1) biosynthesis pathway gene clusters using a PCR assay. Strains lacking an amplicon for the regulatory gene aflR were then analyzed for the presence of the other 28 biosynthetic genes. Only 229 (32%) of the A. flavus strains were found to be atoxigenic. Smaller (S) sclerotial phenotypes were dominant (51%) compared to large (L, 34%) and non-sclerotial (NS, 15%) phenotypes. Among the atoxigenic strains, 24 strains were PCR-negative for the fas-1 and aflJ genes. Sixteen (67%) atoxigenic A. flavus strains were PCRnegative for 10 or more of the biosynthetic genes. Altogether, 18 new PCR product patterns were observed, indicating great diversity in the AFB1 biosynthesis pathway. The current study demonstrates that many atoxigenic A. flavus strains can be isolated from different regions of China. In the future laboratory as well as field based studies are recommended to test these atoxigenic strains as biocontrol agents for aflatoxin contamination.

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Research Support, Non-U.S. Gov'ts
Molecular Characterization of Atoxigenic Aspergillus flavus Isolates Collected in China
Dandan Wei , Lu Zhou , Jonathan Nimal Selvaraj , Chushu Zhang , Fuguo Xing , Yueju Zhao , Yan Wang , Yang Liu
J. Microbiol. 2014;52(7):559-565.   Published online May 30, 2014
DOI: https://doi.org/10.1007/s12275-014-3629-8
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AbstractAbstract
Aspergillus flavus strains were isolated from peanut fields of Liaoning, Shandong, Hubei and Guangdong Provinces in China, and identified through phenotypic and molecular approaches. Of the 323 A. flavus strains isolated, 76 strains did not produce aflatoxins detectable by UPLC. The incidence of atoxigenic A. flavus strains decreased with increase in temperature and increased with increase in latitude in different geographical locations. Amplification of all the aflatoxin genes in the aflatoxin gene cluster in the atoxigenic isolates showed that there were 25 deletion patterns (A-Y), with 22 deletion patterns identified for the first time. Most of the atoxigenic A. flavus isolates with gene deletions (97%) had deletions in at least one of the four genes (aflT, nor-1, aflR, and hypB), indicating that these four genes could be targeted for rapid identification of atoxigenic strains. The atoxigenic isolates with gene deletions, especially the isolates with large deletions, are potential candidates for aflatoxin control.

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The Proportion of Non-Aflatoxigenic Strains of the Aspergillus flavus/oryzae Complex from Meju by Analyses of the Aflatoxin Biosynthetic Genes
Seung-Beom Hong , Mina Lee , Dae-Ho Kim , Soo-Hyun Chung , Hyeon-Dong Shin , Robert A. Samson
J. Microbiol. 2013;51(6):766-772.   Published online December 19, 2013
DOI: https://doi.org/10.1007/s12275-013-3128-3
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AbstractAbstract
Strains of the Aspergillus flavus/oryzae complex are frequently isolated from meju, a fermented soybean product, that is used as the starting material for ganjang (soy sauce) and doenjang (soybean paste) production. In this study, we examined the aflatoxin producing capacity of A. flavus/oryzae strains isolated from meju. 192 strains of A. flavus/oryzae were isolated from more than 100 meju samples collected from diverse regions of Korea from 2008 to 2011, and the norB-cypA, omtA, and aflR genes in the aflatoxin biosynthesis gene cluster were analyzed. We found that 178 strains (92.7%) belonged to non-aflatoxigenic group (Type I of norB-cypA, IB-L-B-, IC-AO, or IA-L-B- of omtA, and AO type of aflR), and 14 strains (7.3%) belonged to aflatoxin-producible group (Type II of norB-cypA, IC-L-B+/B- or IC-L-B+ of omtA, and AF type of aflR). Only 7 strains (3.6%) in the aflatoxin-producible group produced aflatoxins on Czapek yeast-extract medium. The aflatoxin-producing capability of A. flavus/ oryzae strains from other sources in Korea were also investigated, and 92.9% (52/56) strains from air, 93.9% (31/33) strains from rice straw, 91.7% (11/12) strains from soybean, 81.3% (13/16) strains from corn, 82% (41/50) strains from peanut, and 73.2% (41/56) strains from arable soil were included in the non-aflatoxigenic group. The proportion of non-aflatoxigenicity of meju strains was similar to that of strains from soybean, air and rice straw, all of which have an effect on the fermentation of meju. The data suggest that meju does not have a preference for non-aflatoxigenic or aflatoxin-producible strains of A. flavus/oryzae from the environment of meju. The non-aflatoxigenic meju strains are proposed to be named A. oryzae, while the meju strains that can produce aflatoxins should be referred to A. flavus in this study.

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