Comprehensive molecular system to study the presence, growth and ochratoxin A biosynthesis of Penicillium verrucosum in wheat

• Published in: Food additives & contaminants. Part A, Chemistry, analysis, control, exposure & risk assessment Vol. 25; no. 8; pp. 989 - 996
• Main Authors: Schmidt-Heydt, M., Richter, W., Michulec, M., Buttinger, G., Geisen, R.
• Format: Journal Article
• Language: English
• Published: England Taylor & Francis Group 01-08-2008
• Subjects: cereals; DNA, Fungal - genetics; Food Handling - methods; Food Microbiology; Gene Expression; Gene Expression Profiling - methods; Humans; molecular biology; mycotoxins; ochratoxin A; Ochratoxins - analysis; Ochratoxins - biosynthesis; Oligonucleotide Array Sequence Analysis; Penicillium - genetics; Penicillium - growth & development; Penicillium - metabolism; polymerase chain reaction (PCR); Polymerase Chain Reaction - methods; Time; Triticum - microbiology
• ISSN: 1944-0049; 1944-0057
• DOI: 10.1080/02652030801961305

Based on the sequence of the ochratoxin A polyketide synthase gene (otapksPV), a polymerase chain reaction (PCR) system for the specific detection of Penicillium verrucosum in wheat has been developed. In a further approach, a real-time PCR system has been applied to determine the growth kinetics of P. verrucosum in wheat at cell numbers above 10
3
colony-forming units (cfu) ml
−1
. The data obtained by real-time PCR correlated well with the data obtained by the plate count technique. For this purpose, the DNA was isolated directly from contaminated wheat without any further enrichment step. In a reverse transcriptase real-time PCR, the expression of the otapksPV gene in wheat was detected 22 days after inoculation and storage at ambient temperature. Reasonable amounts of ochratoxin A, however, could not be detected before day 30. This early activation of ochratoxin A related genes was confirmed by microarray analysis.