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. 2017 Sep 22;9(10):296.
doi: 10.3390/toxins9100296.

Monitoring the Temporal Expression of Genes Involved in Ochratoxin A Production of Aspergillus carbonarius under the Influence of Temperature and Water Activity

Iliada K Lappa  1 , Dimosthenis Kizis  2   3 , Efstathios Z Panagou  4
Affiliations

Monitoring the Temporal Expression of Genes Involved in Ochratoxin A Production of Aspergillus carbonarius under the Influence of Temperature and Water Activity

Iliada K Lappa et al. Toxins (Basel). .

Abstract

The objective of this study was to investigate the effect of environmental factors, namely temperature and water activity, on genes involved in the regulation of ochratoxin A (OTA) production over time. For this purpose, the previously characterized toxigenic Aspergilluscarbonarius Ac29 isolate from Greek vineyards and the A. carbonarius ITEM 5010 reference strain were subjected to combined temperature and water activity (aw) treatments to study OTA production and relative gene expression. The fungal isolates were grown on a synthetic grape juice liquid medium (SGM) under different temperature (20 °C, 25 °C and 30 °C) and aw (0.94 and 0.98) regimes. The expression of the AcOTApks, AcOTAnrps, and laeA OTA related genes was investigated using real time PCR. Gene expression was monitored at the same time points, along with fungal biomass and OTA accumulation at three, six and nine days of incubation. In gene expression analysis, stimulation of the biosynthetic genes was observed a few days before any toxin could be detected. This fact may underline a possible early indicator of potential toxin contamination of grapes. However, the transcript levels varied with respect to the different combinations of ecophysiological conditions and time, highlighting a complex regulation of OTA related gene expression of A. carbonarius in the specific medium.

Keywords: Aspergillus carbonarius; OTA; SGM; biosynthetic genes; temperature; water activity.

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Conflict of interest statement

The authors declare no conflicts of interest.

Figures

Figure 1
Figure 1
Two-dimensional contour plots showing the effect of temperature and incubation time on biomass production (g) for: Aspergillus carbonarius Ac29 (A,B); and A. carbonarius 5010 (C,D).
Figure 2
Figure 2
Two-dimensional contour plots showing the effect of temperature and incubation time on OTA production (ng/mL) for: Aspergillus carbonarius Ac29 (A,B); and A. carbonarius 5010 (C,D).
Figure 3
Figure 3
Effect of temperature and aw on transcript levels of: AcOTAnrps (A); AcOTApks (B); and laeA (C) genes of Aspergillus carbonarius Ac29 through time. Error bars indicate standard error of three replicates. In the case of absence of fungal growth at Day 3, no values are displayed.
Figure 4
Figure 4
Effect of temperature and aw on transcript levels of: AcOTAnrps (A); AcOTApks (B); and laeA (C) of reference strain Aspergillus carbonarius ITEM 5010 through time. Error bars indicate standard error of three replicates. In the case of absence of fungal growth at Day 3, no values are displayed.
Figure 5
Figure 5
Plot of: variables (A); and cases (B) of the principal components analysis illustrating the correlation between aw, temperature, fungal biomass, OTA production and temporal expression of Aspergillus carbonarius OTA related genes by Ac29 and reference strain ITEM 5010, grown on SGM. (Cases are coded as follows: W and R correspond to wild and reference fungal strain, respectively; the following two numbers correspond to incubation temperature; the next two digits indicate aw level; and the last digit incubation time).
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