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. 2014 Feb;51(2):401-7.
doi: 10.1007/s13197-013-1190-9. Epub 2013 Oct 11.

Detection of hemolytic strains of Aeromonas hydrophila and A . sobria along with other Aeromonas spp. from fish and fishery products by multiplex PCR

I A Hussain  1 , G Jeyasekaran  1 , R Jeya Shakila  1 , K T Raj  1 , E Jeevithan  1
Affiliations

Detection of hemolytic strains of Aeromonas hydrophila and A . sobria along with other Aeromonas spp. from fish and fishery products by multiplex PCR

I A Hussain et al. J Food Sci Technol. 2014 Feb.

Abstract

Hemolytic strains of Aeromonas spp. from fish and fishery products were detected by multiplex PCR. The selected primers for the amplification of segments of ahh1, asa1 and 16S rRNA gene yielded products with the size of 130 bp, 249 bp and 356 bp, respectively. This assay was found to be highly sensitive, as it could detect 7 and 9 cells of Aeromonas hydrophila and A. sobria with a detection limit of 1 pg of pure genomic DNA. The assay, when screened for 73 commercial fish and fishery product samples consisting of freshwater, marine fish and shellfish, showed 56 % positive for Aeromonas spp., 16 % for Aeromonas hydrophila and 13 % for A. sobria. This assay provides specific and reliable results and can be a powerful tool for the simultaneous detection of hemolytic strains of A. hydrophila A. sobria and other Aeromonas spp. from fish and fishery products.

Keywords: Aeromonas hydrophila; Fish and fishery products; Hemolytic strains; Multiplex PCR; Species-specific detection.

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Figures

Fig. 1
Fig. 1
Detection of A. hydrophila, A. sobria and other Aeromonas spp by uniplex and multiplex PCR. Lane 1- A. hydrophila; Lane 2- A. sobria; Lane 3- Aeromonas spp.; Lane 4- multiplex for A. hydrophila, A. sobria and other Aeromonas spp.; Lane M-100 bp DNA marker
Fig. 2
Fig. 2
Specificity of MPCR assay for the detection of A. hydrophila, A. sobria and other Aeromonas spp. Lane 1- A. hydrophila; Lane 2- A. sobria; Lane 3- A. caviae; Lane 4- A. liquefaciens; Lane 5- A. hydrophila and A. sobria; Lane 6- Salmonella typhi; Lane 7- S. paratyphi A; Lane 8- Vibrio cholerae; Lane 9- V. parahaemolyticus; Lane 10- V. vulnificus; Lane 11- V. alginolyticus; Lane M-100 bp DNA marker
Fig. 3
Fig. 3
Sensitivity of the MPCR assay for the detection of cells of A. hydrophila, A. sobria and other Aeromonas spp. after 12 h of enrichment. Lane 1- > 250 cells/ml in 10−6 dilution; Lane 2–75 cells/ml of A. hydrophila and 96 cells/ml of A. sobria in 10−7 dilution; Lane 3–7 cells/ml of A. hydrophila and 9 cells/ml of A. sobria in10−8 dilution; Lane 4-Positive control; Lane M- 100 bp DNA marker
Fig. 4
Fig. 4
Sensitivity of the MPCR assay for the detection of genomic DNA of A. hydrophila, A. sobria and other Aeromonas spp. Lane 1–1,000 pg; Lane 2–100 pg; Lane 3–10 pg; Lane 4–1 pg; Lane 5–100 fg; Lane 6–10 fg; Lane M, 100 bp DNA marker
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