Applied Mycology Group, Cranfield Health, Cranfield University, Bedford, UK.
J Appl Microbiol. 2010 Dec;109(6):1914-22. doi: 10.1111/j.1365-2672.2010.04820.x.
A relative quantification system (RQ-PCR) was used to monitor the correlations between the activity of the nor-1 (=aflD) gene of Aspergillus flavus using real-time PCR in relation to phenotypic aflatoxin B(1) (AFB(1) ) production and populations of A. flavus in stored peanuts at three water activity levels (a(w) , 0·95, 0·90 and 0·85) for 6 weeks.
Real-time PCR was used to amplify the nor-1 gene (target gene), and benA56 (β-tubulin gene) used as a control gene. Expression of three structural genes, nor-1 (=aflD), ver-1 (=aflM), and omtA (=aflP), and the regulatory gene aflR of the aflatoxin biosynthetic pathway were also assayed. There were significant differences between nor-1 gene expression at the three a(w) levels; higher expression at 0·90 a(w) in weeks 1-3, when compared to 0·95. In contrast, in the driest treatment (0·85 a(w) ) none or very low nor-1 expression occurred. The populations of A. flavus colony-forming units (CFUs g(-1) ) increased over time with the highest at 0·95 a(w) . Highest AFB(1) production was at 0·90 and 0·95 a(w) from weeks 3-6. A(w) had a significant effect on aflR transcription at 0·95 a(w) over the 6-week period, while at 0·90 a(w) , only in the last 2 weeks.
Correlations between different factors showed that log AFB(1) × log CFUs, log AFB(1) × a(w) , and log CFUs × a(w) were statistically significant, while log CFUs × RQ-PCR and RQ-PCR × a(w) were not. The AflR gene may not have an important role in the regulation of nor-1 expression in food matrices (e.g. peanuts).
Determination of correlations between nor-1 expression and aflatoxin production by A. flavus in raw peanuts under different a(w) levels could be helpful to predict potential risk of aflatoxin production during storage of this hygroscopic food product and minimize contamination with the AFB(1) .
采用相对定量实时 PCR（RQ-PCR）系统，监测黄曲霉 nor-1（= aflD）基因的活性与表型黄曲霉毒素 B1（AFB1）的产生以及在 3 种水分活度（a w ，0.95、0.90 和 0.85）下储存花生中黄曲霉种群之间的相关性，为期 6 周。
采用实时 PCR 扩增 nor-1 基因（靶基因），并以 benA56（β-微管蛋白基因）作为对照基因。还测定了黄曲霉毒素生物合成途径的三个结构基因 nor-1（= aflD）、ver-1（= aflM）和 omtA（= aflP）以及调节基因 aflR 的表达。在 3 种 a w 水平下，nor-1 基因的表达存在显著差异；在第 1-3 周，在 0.90 a w 时表达更高，与 0.95 相比。相比之下，在最干燥的处理（0.85 a w ）中，nor-1 的表达几乎不存在或非常低。黄曲霉菌落形成单位（CFU g（-1））的种群随时间增加，在 0.95 a w 时最高。在第 3-6 周，在 0.90 和 0.95 a w 时，AFB1 的产量最高。在 6 周的时间里，a w 对 aflR 转录有显著影响在 0.95 a w ，而在 0.90 a w ，仅在最后 2 周。
不同因素之间的相关性表明，logAFB1×logCFUs、logAFB1×a w 和 logCFUs×a w 具有统计学意义，而 logCFUs×RQ-PCR 和 RQ-PCR×a w 则没有。AflR 基因可能在调节食品基质（如花生）中 nor-1 表达方面没有重要作用。
在不同 a w 水平下测定生花生中黄曲霉 nor-1 表达与黄曲霉毒素产生之间的相关性，有助于预测这种吸湿性食品在储存过程中产生黄曲霉毒素的潜在风险，并最大程度地减少 AFB1 的污染。
