Biochemical and Molecular Characterization of Tetracycline-Resistant Aeromonas veronii Isolates from Catfish

Abstract

Eighty-one tetracycline-resistant Aeromonas sp. strains were isolated from farm-raised catfish. Morphological and biochemical characteristics indicated that 23 of the 81 aeromonads were Aeromonas hydrophila, 7 isolates were Aeromonas trota, 6 isolates were Aeromonas caviae, 42 isolates were Aeromonas veronii, and 3 isolates were Aeromonas jandaei. However, the AluI and MboI restriction fragment length polymorphism (RFLP) patterns of the PCR-amplified 1.4-kb 16S rRNA gene from all 81 tetracycline-resistant aeromonads from catfish were identical to the RFLP banding patterns of A. veronii ATCC 35626, indicating that all 81 isolates were strains of A. veronii. A multiplex PCR assay successfully amplified the 5 tetracycline-resistant genes (tetA to E) from the genomic DNA of all 81 isolates. The assay determined that tetE was the dominant gene occurring in 73/81 (90.0%) of the aeromonads. Plasmids (2.0 to 20 kb) were isolated from 33 of the 81 isolates. Dendrogram analysis of the SpeI pulsed-field gel electrophoresis identified 15 distinct macrorestriction patterns among the isolates. Our results indicate the need for use of 16S rRNA in the identification of Aeromonas spp. and the prevalence of catfish as a reservoir of tet genes.The United States produces more than $500 million per annum of pond-raised channel catfish (Ictalurus punctatus), primarily in the southeastern part of the country (4). Infectious diseases reduce catfish production by nearly 10% every year. Motile aeromonad septicemia, caused by Aeromonas hydrophila, is one of the common diseases accounting for the reduced production (5). Antimicrobials agents, such as oxytetracycline and Romet 30 (sulfadimethoxine-ormetoprim), are used to prevent the outbreak of infectious diseases (5, 32). However, widespread use of drugs may result in the selection of tetracycline-and sulfonamide-resistant bacteria in the aquaculture environment and may play a role in the dissemination of antibiotic resistance genes to clinical aeromonad strains (20,22).Aeromonads have been implicated in the cause of numerous human infections, such as gastroenteritis, cellulitis, meningitis, bacteremia, soft-tissue infections, peritonitis, and bronchopulmonary infections (8,15,(18)(19). Direct contact with contaminated water and soil is the most frequent cause of gastrointestinal and wound infections in humans (15). Broad-spectrum antibiotics, such as tetracycline, are prescribed clinically for the treatment of such infections.The molecular epidemiology of tetracycline-resistant aeromonads, especially Aeromonas salmonicida, has been well documented (2,3,29). These studies indicate that tetracycline resistance is plasmid-encoded and that, among the different classes of tetracycline-resistant genes, tetA is predominant. Schmidt et al. (29) reported the isolation and characterization of oxytetracycline-sulfonamide/trimethoprim-resistant aeromonads from Danish rainbow trout farms. Their PCR data indicate tetE as the predominant determinant, followed by tetA and tetD. Earlier, DePaola et al. ...

The involvement of tetA and tetE tetracycline resistance genes in plasmid and chromosomal resistance of Aeromonas in Brazilian strains

“…The association of this gene with the chromosome was also observed (DePaola & Roberts 1995, Roberts 1996, which might explain its limited distribution among environmental samples, being more prevalent in polluted marine sediments (Andersen & Sandaa 1994, Chopra & Roberts 2001. Nawaz et al (2006), while describing the tetE gene, observed the high frequency of this determinant in Aeromonas spp. isolates.…”

“…There are some reports showing that the tetracyclineresistance determinants most predominant in Aeromonas species are tetA and tetE (Schmidt et al 2001b, Nawaz et al 2006). Our work is in agreement with the literature.…”

“…The molecular characterization of the tet genes and plasmids that carry these resistance determinants may help to explain how human and the environment have been affected by the increase and indiscriminate use of tetracycline. The tetracycline resistance determinants most commonly observed in Aeromonas species are tetA and tetE (Schmidt et al 2001b, Nawaz et al 2006. For that reason, those genes were chosen for the amplification reactions performed in this work.…”

Molecular Detection and Characterization of Furunculosis and Other Aeromonas Fish Infections

“…(Figueras et al, 2011b). Biochemical identifications in fact lack precision and tend to generate a lot of inconsistent results in comparison to those obtained by molecular methods (Beaz-Hidalgo et al, 2010;Figueras et al, 2011b;Kozińska et al, 2002, Kozińska, 2007Nawaz et al, 2006).…”

“…However, the importance of the species A. hydrophila is overestimated because biochemical identification systems erroneously identify up to 70-80% of the strains of Aeromonas as belonging to this species, when in fact they are many different species when identified by molecular methods (Beaz-Hidalgo et al, 2010;Figueras, 2005;2011b;Soler et al, 2003a). Prevalent motile species associated to diseased fish following molecular identification include A. veronii, associated with catfish, A. sobria with tilapia and trout, A. hydrophila with trout, A. bestiarum with carp and the recently described A. piscicola recovered from diseased salmonids and turbots (Beaz-Hidalgo et al, 2010;Kozińska, 2007;Li & Cai, 2011;Martino et al, 2011;Nawaz et al, 2006;Soriano-Vargas et al, 2010). Other motile species, such as A.…”