Proteomic characterization and discrimination of Aeromonas species recovered from meat and water samples with a spotlight on the antimicrobial resistance of Aeromonas hydrophila

Abstract

Aeromonas is recognized as a human pathogen following ingestion of contaminated food and water. One major problem in Aeromonas identification is that certain species are phenotypically very similar. The antimicrobial resistance is another significant challenge worldwide. We therefore aimed to use mass spectrometry technology for identification and discrimination of Aeromonas species and to screen the antimicrobial resistance of Aeromonas hydrophila (A. hydrophila). A total of 150 chicken meat and water samples were cultured, and then, the isolates were identified biochemically by the Vitek^® 2 Compact system. Proteomic identification was performed by MALDI-TOF MS and confirmed by a microchannel fluidics electrophoresis assay. Principal component analysis (PCA) and single-peak analysis created by MALDI were also used to discriminate the Aeromonas species. The antimicrobial resistance of the A. hydrophila isolates was determined by Vitek^® 2 AST cards. In total, 43 samples were positive for Aeromonas and comprised 22 A. hydrophila, 12 Aeromonas caviae (A. caviae), and 9 Aeromonas sobria (A. sobria) isolates. Thirty-nine out of 43 (90.69%) Aeromonas isolates were identified by the Vitek^® 2 Compact system, whereas 100% of the Aeromonas isolates were correctly identified by MALDI-TOF MS with a score value ≥2.00. PCA successfully separated A. hydrophila, A. caviae and A. sobria isolates into two groups. Single-peak analysis revealed four discriminating peaks that separated A. hydrophila from A. caviae and A. sobria isolates. The resistance of A. hydrophila to antibiotics was 95.46% for ampicillin, 50% for cefotaxime, 45.45% for norfloxacin and pefloxacin, 36.36% for ceftazidime and ciprofloxacin, 31.81% for ofloxacin and 27.27% for nalidixic acid and tobramycin. In conclusion, chicken meat and water were tainted with Aeromonas spp., with a high occurrence of A. hydrophila. MALDI-TOF MS is a powerful technique for characterizing aeromonads at the genus and species levels. Future studies should investigate the resistance of A. hydrophila to various antimicrobial agents.

Keywords: Aeromonas spp.; antimicrobial resistance; differentiation; microchannel electrophoresis; protein fingerprinting.

Figure 1

Mass spectrum protein profiles of 43 Aeromonas spp.; (a) Distribution of peaks within the line spectra ranging from 2,000 to 11,000 Da; (b) The gel profile of protein spectra in which the varied color of spots was the gathering of spectra with several contents

Figure 2

Mass spectral profiles of 43 Aeromonas spp.; (a) higher strength peaks were scattered within the line spectra ranging from 4,000 to 10,000 Da; (b) The gel profile of protein spectra distributed within the same range

Figure 3

The dimensional image from PCA displays the difference between 43 Aeromonas spp.; (a) the grouping of A. hydrophila (red), A. caviae (green), and A. sobria (blue) in the first three model of PC (PC1, PC2, PC3); (b) the influence of ten principal components to the profiling classification in plot of percentage explained variance of PC. The contributions of PC1, PC2, and PC3 were around 45%, 17%, and 9%, correspondingly

Figure 4

Higher peaks intensity (3,194, 4,030, 5,383, and 7,611 Da) were detected in A. hydrophila (red), whereas they were missed in A. sobria (green) and A. caviae (blue)

Figure 5

Higher peaks intensity (3,667, 4,351, 7,335 and 9,635 Da) were detected in A. sobria (green), whereas they were missed in A. hydrophila (red) and A. caviae (blue). Moreover, a higher peak intensity (7,347 Da) was detected in A. caviae (blue) while it was missed in A. hydrophila (red) and A. sobria (green)