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. 2015 Nov 27;7(12):5079-93.
doi: 10.3390/toxins7124864.

Biodegradation of Ochratoxin A by Bacterial Strains Isolated From Vineyard Soils

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Free PMC article

Biodegradation of Ochratoxin A by Bacterial Strains Isolated From Vineyard Soils

Palmira De Bellis et al. Toxins (Basel). .
Free PMC article

Abstract

Ochratoxin A (OTA) is a mycotoxin with a main nephrotoxic activity contaminating several foodstuffs. In the present report, five soil samples collected from OTA-contaminated vineyards were screened to isolate microorganisms able to biodegrade OTA. When cultivated in OTA-supplemented medium, OTA was converted in OTα by 225 bacterial isolates. To reveal clonal relationships between isolates, molecular typing by using an automated rep-PCR system was carried out, thus showing the presence of 27 different strains (rep-PCR profiles). The 16S-rRNA gene sequence analysis of an isolate representative of each rep-PCR profiles indicated that they belonged to five bacterial genera, namely Pseudomonas, Leclercia, Pantoea, Enterobacter, and Acinetobacter. However, further evaluation of OTA-degrading activity by the 27 strains revealed that only Acinetobacter calcoaceticus strain 396.1 and Acinetobacter sp. strain neg1, consistently conserved the above property; their further characterization showed that they were able to convert 82% and 91% OTA into OTα in six days at 24 °C, respectively. The presence of OTα, as the unique OTA-degradation product was confirmed by LC-HRMS. This is the first report on OTA biodegradation by bacterial strains isolated from agricultural soils and carried out under aerobic conditions and moderate temperatures. These microorganisms might be used to detoxify OTA-contaminated feed and could be a new source of gene(s) for the development of a novel enzymatic detoxification system.

Keywords: Acinetobacter; biodegradation; detoxification; ochratoxin; soil bacteria.

Figures

Figure 1
Figure 1
Degradation of OTA (grey) in OTα (white) by microbial populations associated to five soil samples in three liquid substrates.
Figure 2
Figure 2
Ochratoxin A degradation by microbial populations associated with the three soil samples respectively cultured in presence of nystatine or chloramphenicol. The OTA degradation in absence of antibiotic addition is also shown.
Figure 3
Figure 3
Gel-like image obtained using the Agilent Expert software of the 27 different rep-PCR profiles identified after the analysis of the 225 OTA-degrading isolates. The profiles were obtained using primers REP-1R-Dt/REP-2R-Dt. L: DNA 7500 ladder.
Figure 4
Figure 4
Percentage of OTA degradation by the twenty-seven strains identified by the REP-PCR analysis. Cultures were incubated at 24 °C and mycotoxin degradation was evaluated by HPLC-FLD assay after six days. Bars represent standard errors. (A) Pseudomonas taiwanensis; (B) Leclercia adecarboxylata; (C) Acinetobacter sp. neg1; (D) Pantoea agglomerans; (E) Pseudomonas reinekei; (F) Pseudomonas koreensis; (G) Enterobacter aerogenes; (H) Acinetobacter calcoaceticus; and (I) Enterobacter xiangfangensis.
Figure 5
Figure 5
OTA degradation by Acinetobacter sp. neg1 (A) and A. calcoaceticus 396.1; and (B) in MMP medium added with 1 µg/mL of OTA. Cultures were incubated at 22, 24 and 28 °C and mycotoxin degradation was evaluated by HPLC-FLD assay at 13 and six days post inoculation.
Figure 6
Figure 6
LC-HR-MS Total Ion Current (TIC) and extracted ion chromatograms (XIC) filtered on the accurate mass of ochratoxin A and ochratoxin alpha from the supernatant of liquid culture sample added with ochratoxin A and inoculated Acinetobacter sp. neg1. In the upper panels are reported (A) the full mass chromatogram; (B) the extracted ion chromatogram filtered on the accurate mass of ochratoxin alpha; and (C) the extracted ion chromatogram filtered on the accurate mass of ochratoxin A.

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