A Simple Procedure to Evaluate Competitiveness of Toxigenic and Atoxigenic Isolates of Aspergillus flavus in Solid and Liquid Media

محورهای موضوعی :

Mohammad Moradi ¹ (Pistachio Research Center, Horticultural Sciences Research Institute, Agricultural Research, Education and Exten-sion Organization (AREEO), Rafsanjan, Iran)
Seyed Reza Fani ² (Plant Protection Research Department, Yazd Agricultural and Natural Resources Research and Education Center, AREEO, Yazd, Iran)
Rosa Dargahi ³ (Pistachio Research Center, Horticultural Sciences Research Institute, Agricultural Research, Education and Exten-sion Organization (AREEO), Rafsanjan, Iran)
Mehdi Mohammadi Moghadam ⁴ (Plant Protection Research Department, Agricultural and Natural Resources Research Center of Semnan Province (Shahrood), AREEO, Shahrood, Iran)
Abdolhamid Sherafati ⁵ (Horticulture Crops Research Department, Khorasan Razavi Agricultural and Natural Resources Research and Edu-cation Center, AREEO, Mashhad, Iran)

تاریخ دریافت : 1396/02/06 تاریخ پذیرش : 1397/08/07 تاریخ انتشار : 1396/02/06

کلید واژه: biocontrol, Competitiveness, Aflatoxin, Thin layer chromatography, Cultural methods, Ammonium hydrox-ide,

چکیده مقاله :

Application of atoxigenic strains of Aspergillus flavus to reduce aflatoxin levels is the most successful strategy applied in some agricultural crops. The role of ammonium hydroxide for preliminary screening of the competitiveness of atoxigenic A. flavus isolates to interfere with aflatoxin production by highly toxigenic isolates were evaluated. Out of 270 A. flavus isolates, 17 were detected as true atoxigenic using cultural methods and confirmed by analytical assays from different pistachio agro-ecological zones during 2013. For assessment competitive ability among atoxigenic isolates of A. flavus with highly toxigenic one, rice flour, coconut agar and coconut broth medium substrates were inoculated with mixtures including combinations of toxigenic and atoxigenic isolates, simultaneously. The rice flour substrate was used to quantify the content of aflatoxin in either co-inoculations or toxigenic isolate alone on thin layer chromatography plates with a scanning densitometer. While the culture media were used to determine the intensity of color change on exposing to ammonium hydroxide vapor. The reduction rates of aflatoxin B1 in co-inoculations were varied and ranged from 2%-82%. Based on the intensity of colony color changes, the competitiveness of the isolates was classified into five groups. Atoxigenic isolates with high competitiveness have shown low color changes in culture media and high aflatoxin reduction in TLC assays with a ratio of higher than 78%. The method will facilitate preliminary screening of efficient atoxigenic isolates for mitigation of aflatoxins in food and feed as a cheap, simple and quick method.

منابع و مأخذ:

1. Dorner J.W., 2004. Biological control of aflatoxin contamination of crops. J Toxicol. 23, 425-450.
2. Moradi M., Hokmabadi H., 2011. Control of Mycotoxin Bioactives in Nuts: Farm to Fork. InTokusoglu, O., Hal III, C. (Eds), Fruit and Cereal Bioactives, Sources, Chemistry, and Applications (pp. 291-315). Turkey: CRC Press.
3. Cotty P.J., Mellon J.E., 2006. Ecology of aflatoxin producing fungi and biocontrol of aflatoxin contamination. Mycol Res. 22, 110-117.
4. Cotty P.J., 1990. Effect of atoxigenic strains of Aspergillus flavus on aflatoxin contamination of developing cotton seed. Plant Dis. 74, 233-235.
5. Bandyopadhyay R., Kiewnick S., Atehnkeng J., Donner M., Cotty P.J., Hell K., 2005. Biological con-trol of aflatoxin contamination in maize in Africa. In Abstr. Tropentag 2005 Conf. Int. Agric. Res. Dev. Swiss Federal Institute of Technology, Zurich, Switzerland (p. 66).
6. Dorner J.W., 2008. Management and prevention of mycotoxins in peanuts. Food Addit Contam. 25(2), 203-208.
7. Abbas H.K., Weaver M.A., Horn B.W., Carbone I., Monacell J.T., Shier W.T., 2011. Selection of Aspergillus flavus isolates for biological control of aflatoxins in corn. Toxin Rev. 30(2-3), 59-70.
8. Doster M.A., Cotty P.J., Michailides T.J., 2014. Evaluation of the atoxigenic Aspergillus flavus strain AF36 in pistachio orchards. Plant Dis. 98(7), 948-956.
9. Yin Y.N., Yan L.Y., Jiang J.H., Ma Z.H., 2008. Bio-logical control of aflatoxin contamination of crops. J Zhejiang Univ Sci B. 9(10), 787-792.
10. Nasir M.S., Jolley M.E., 2002. Development of a fluorescence polarization assay for the determination of aflatoxins in grains. J Agric Food Chem. 50(11), 3116-3121.
11. Fente C.A., Ordaz J.J., VazquezB.I., Franco C.M., Cepeda A., 2001. New additive for culture media for rapid identification of aflatoxin-producing Aspergillus strains. Appl Environ Microb. 67(10), 4858-4862.
12. Ordaz J.J., Fente C.A., Vázquez B.I., Franco C.M., Cepeda A., 2003. Development of a method for direct visual determination of aflatoxin production by colonies of the Aspergillus flavus group. Int J Food Microbiol. 83(2), 219-225.
13. Gupta A., Gopal M., 2002. Aflatoxin production by Aspergillus flavus isolates pathogenic to coconut insect pests. World J Microbiol Biotechnol. 18(4), 329-335.
14. Fani S.R., Moradi M., Probst C., Zamanizadeh H.R., Mirabolfathy M., Haidukowski M., Logrieco A.F., 2014a. A critical evaluation of cultural methods for the identification of atoxigenic Aspergillus flavus isolates for aflatoxin mitigation in pistachio orchards of Iran. Eur J Plant Pathol. 140(4), 631-642.
15. Saito M., Machida S., 1999. A rapid identification method for aflatoxin-producing strains of Aspergillus flavus and A. parasiticus by ammonia vapor. Mycoscience. 40, 205-208.
16. Cotty P.J., 1988. Simple fluorescence method for rapid estimation of aflatoxin levels in a solid culture medium. Appl Microbiol Biot. 54(1), 274-276.
17. Abbas H.K., Shier W.T., Horn B.W., Weaver M.A., 2004. Cultural methods for aflatoxin detection. J Toxicol. Toxin Rev. 23, 295-319.
18. Bhatnagar D., Ehrlich K.C., Cleveland T.E., 2003. Molecular genetic analysis and regulation of aflatoxin biosynthesis. Appl Microbiol Biotechnol. 61, 83-93.
19. Gourama H., Bullerman L.B., 1995. Aspergillus flavus and Aspergillus parasiticus: aflatoxigenic fungi of concern in foods and feeds: A review. J Food Prot. 58(12), 1395-1404.
20. Klich M.A., 2002. Identification of common Aspergillus species. Utrecht: Centraalbureau voor Schimmel cultures, Utrecht, The Netherlands.
21. Fani S.R, Moradi M., Zamanizadeh H.R., Mira-bolfathy M., Probst C., 2014b. Distribution of nontoxigenic strains of Aspergillus flavus throughout pistachio growing areas in Iran. Appl Entomol Phytopathol. 81(2), 179-189.
22. Probst C., Bandyopadhyay R., Price L.E., Cotty P.J., 2011. Identification of atoxigenic Aspergillus flavus isolates to reduce aflatoxin contamination of maize in Kenya. Plant Dis. 95, 212-218.
23. Davis N.D., Iyer S.K., Diener U.L., 1987. Improved method of screening for aflatoxin with a coconut agar medium. Appl Environ Microbiol. 53(7), 1593-1595.
24. Wei D., Jong S., 1986. Production of aflatoxins by strains of the Aspergillus flavus group maintained in ATCC. Mycopathologia. 93, 19-24.
25. Dorner J.W., Cole R.J., 2002. Effect of application of nontoxigenic strains of Aspergillus flavus and A. parasiticus on subsequent aflatoxin contamination of peanuts in storage. J Stored Prod Res. 38(4), 329-339.
26. Stroka J., AnklamE., 2000. Development of a simplified densitometer for the determination of aflatoxins by thin-layer chromatography. J Chromatogr A. 904(2), 263-268.
27. Wiseman H.G., Jacobson W.C., Harmeyer W.C., 1967. Note on removal of pigments from chloroform extracts of aflatoxin cultures with copper carbonate. J Assoc off Agric Chem. 50, 982-983.
28. Kumar S., Shekhar M., Ali K.A., Sharma P., 2007. A rapid technique for detection of toxigenic and non-toxigenic strain of Aspergillus flavus from maize grain. Indian Phytopathol. 1, 31-34.
29. Shier W.T., Lao Y., Steele T.W., Abbas H.K., 2005.Yellow pigments used in rapid identification of aflatoxin-producing Aspergillus strain are anthraquinones associated with the aflatoxin biosynthetic pathway. Bioorg Chem. 33, 426-438.