Identification and quantification of fumonisin-producing Fusarium species in grain and soil samples from Egypt and the Philippines

Abstract

Fumonisins are considered among the important mycotoxins associated with human esophageal cancer and livestock diseases. These mycotoxins are mainly produced by Fusarium verticillioides in tropical and subtropical regions such as the Philippines and Egypt and humid temperate regions of the world. The classical taxonomy of fumonisin-producing fungi is challenging, and species-specific PCR reactions are commonly used to clearly identify species within these complexes. The aim of this study was to isolate, identify and quantify fumonisin-producing species in maize, wheat and soil samples from Egypt and the Philippines, and to test Eppendorf-Agar as a long term preservation method. We isolated 44 single spore isolates (39 from Egypt and five from the Philippines) from the collected samples (25 isolates from maize, five from wheat and 14 from soil). In addition, we quantified the content of fumonisin-producing fungi DNA from 15 maize samples and six wheat samples from Egypt, and from six maize samples from the Philippines. morphological and microscopic identification indicated that 21 isolates from Egypt and five from the Philippines were F. verticillioides, one isolate was F. proliferatum and two isolates were F nygamai. Molecular identification indicated that all these isolates belonged to F. verticillioides. Most were from maize, four were from soil and only one was from wheat. Other Fusarium species isolated included F. oxysporum and F. solani. No F. graminearum isolates were found. The quantitative PCR (qPCR) results obtained using the Taqfum-2f, Vpgen-3R primer pair and the FUMp probe for quantification of fumonisin-producing Fusarium species showed that fumonisin-producing Fusarium isolates were present in four maize samples from the Philippines and eight maize samples from Egypt. The Fusarium DNA levels from fumonisin-producing isolates were in the range of 13 x 10(-3) to 61 x 10(-1) ng ng(-1) total DNA in positive samples, except in one maize sample from the Philippines with high concentration of >0.5 ng ng(-1) total DNA. This indicates that >50 % of all DNA was Fusarium DNA. No fumonisin-producing Fusarium DNA was detected in the wheat samples and in the remaining maize samples. These results showed that PCR-techniques based on qPCR can be used to identify fumonisin-producing Fusarium species and quantify risks of mycotoxin contaminated grains.