Antimicrobial Susceptibility of Coagulase-Positive and Coagulase-Negative Staphylococci in Ready-to-Eat Sashimi

The aim of this study was to investigate the antimicrobial susceptibility pattern of Staphylococcus spp. isolated from sashimi meals collected in Japanese restaurants from northern Portugal. One hundred and thirty-six isolates of Staphylococcus spp. were recovered from sashimi samples. The antimicrobial susceptibility test showed high levels of resistance to β-lactams, macrolides, and lincosamides. All Staphylococcus isolates were sensitive to trimethoprim-sulfamethoxazole, ciprofloxacin, and chloramphenicol. Multidrug-resistant staphylococci (25%) were found in sashimi samples. It is of the utmost need to improve the hygienic practices in handling in order to reduce the occurrence of Staphylococcus species, potentially resistant to antimicrobial agents.     

Methods for identification and differentiation of different Shewanella spp. isolates for diagnostic use

Shewanella spp. are Gram‐negative, rod‐shaped, motile bacteria that are widely distributed in marine and freshwater environments. The bacteria are present in the physiological microflora of fish from temperate waters and are known as fish spoilage species. From clinically healthy fish and from fish with skin ulcerations, Shewanella spp. is regularly isolated, indicating a possible role as fish pathogen. In this study, 74 isolates of Shewanella spp. were analysed. For species identification, biochemical techniques, 16S rRNA sequencing, MALDI‐TOF MS and the Sherlock Microbial Identification System (MIS) based on the composition of fatty acid ethyl esters were compared. The phylogenetic relationship, cytotoxicity in vitro and resistance against antibiotics were tested. The most reliable method for species identification was 16S rRNA sequencing. From diseased fish, clinically healthy fish and the aquatic environment, different Shewanella species were isolated. This indicates that Shewanella spp. is widespread in the aquatic milieu and acts as a secondary pathogen. The virulence of Shewanella spp. is probably not depending on the species but on the isolate itself. Many isolates of Shewanella spp. were showing multiresistances against antibiotic substances, especially in samples derived from retailers and in routine diagnostics, all Shewanella spp. should therefore be tested for resistances against antibiotic agents.     

High prevalence of antimicrobial resistance in Escherichia coli,Salmonella spp. and Staphylococcus aureus isolated from fish samples in India

Antibiotic resistance, driven by inappropriate use of antimicrobial agents for fish farming, is an expanding global health threat. Demand of animal protein for human consumption is entraining at remarkable pace which in turn, upturning the non‐judicial use of life‐saving drugs in these modern animal production practices at an incautious rate. Hence, study was devised to observe the antimicrobial resistance (AMR) pattern of human prominence bacteria, against drugs of critically and highly importance of both human and veterinary medicine, circulating over the fish farming in India. One hundred sixty fish faecal samples were obtained from distinct geographical locales of Rajasthan, India. Overall, 202 isolates comprising of Escherichia coli (81, 51%), Salmonella spp. (72, 45%) and Staphylococcus aureus (49, 31%) were isolated to screen for resistance. Antimicrobial resistance has been analysed using broth microdilution method. In case of E. coli and Salmonella spp., highest resistance (>95%) was observed against streptomycin while for S. aureus, it was observed against trimethoprim, along with high resistance to other antibiotics. Majority of the isolates were found to be resistant to more than three antibiotics implying multi‐drug resistance (>89%) relative to the critical and highly important antibiotics. The high AMR is a reflection of misuse of these agents in the fish farming, which is a concern of serious health issues. Our findings persuade the prudent use of antimicrobial agents in fish farming and other aquaculture settings that may assist in drafting of the new policies for the judicial use of these life‐saving drugs in these sectors.     

Diagnostic methods for identifying different Aeromonas species and examining their pathogenicity factors,their correlation to cytotoxicity and adherence to fish mucus

Aeromonas spp. are ubiquitous in the aquatic environment, acting as facultative or obligate pathogens for fish. Identifying Aeromonas spp. is important for pathogenesis and prognosis in diagnostic cases but can be difficult because of their close relationship. Forty‐four already characterized isolates of Aeromonas spp. were analysed by 16S rRNA gene sequencing, by gyrase B sequencing, by analysing their fatty acid profiles, by biochemical reactions and by MALDI‐TOF MS. To determine their pathogenicity, cytotoxicity, adhesion to mucus and the expression of 12 virulence factors were tested. The susceptibility of the isolates towards 13 different antibiotics was determined. MALDI‐TOF MS was found to be an acceptable identification method for Aeromonas spp. Although the method does not detect all species correctly, it is time‐effective and entails relatively low costs and no other methods achieved better results. A high prevalence of virulence‐related gene fragments was detected in almost all examined Aeromonas spp., especially in A. hydrophila and A. salmonicida, and most isolates exhibited a cytotoxic effect. Single isolates of A. hydrophila and A. salmonicida showed multiple resistance to antibiotics. These results might indicate the potentially pathogenic capacity of Aeromonas spp., suggesting a risk for aquatic animals and even humans, given their ubiquitous nature.     

Microbiological Quality of Ready-to-Eat Pickled Fish Products

The microbiological quality of 18 commercially available in Spain ready-to-eat fish products containing Engraulidae was evaluated through application of the corresponding ISO procedures for total mesophilic aerobic microbial counts, detection and enumeration of enterobacteria, and detection of Staphylococcus spp. All isolates were identified to the species level using two different biochemical methods: the API® test and the Biolog® identification system. The most commonly occurring contaminants found were Enterobacteriaceae—such as Citrobacter freundii and other Citrobacter species, Enterobacter cloacae, Cronobacter sakazakii, Hafnia alvei, Pantoea, Proteus ssp., and Escherichia coli. The presence of such opportunistic pathogens and contaminant microflora was confirmed in 61% of the foods sampled.     

Identification and pathogenicity study of emerging fish pathogens Acinetobacter junii and Acinetobacter pittii recovered from a disease outbreak in Labeo catla (Hamilton, 1822) and Hypophthalmichthys molitrix (Valenciennes, 1844) of freshwater wetland in West Bengal,India

The disease outbreaks in aquaculture system of wetlands are the major cause of fish mortality. Among various bacterial septicaemic diseases, fish mortality caused by Acinetobacter spp. is recently reported in different fish species. Fish disease outbreak was investigated in a wetland of West Bengal, India to identify the aetiological factors involved. The moribund fish were examined and subjected to bacterial isolation. Two bacterial causative agents were identified as Acinetobacter junii and Acinetobacter pittii by biochemical characterization and 16S rRNA gene amplification. Both the isolates were oxidase‐negative, nitrate‐negative, catalase‐positive and indole‐negative. The molecular identification using 16S rRNA gene sequencing and phylogenetic tree analysis further confirmed the two Acinetobacter spp. with 97%–99% similarity. The antibiotic resistance patterns of these two bacteria revealed that both of them were resistant to β‐lactam, cefalexin, cephalothin, amoxyclav, cefuroxime, cefadroxil, clindamycin, vancomycin and penicillin. In addition, A. pittii was also resistant to other antibiotics of cephams group such as ceftazidime and cefotaxime. In the challenge experiment, both A. junii and A. pittii were found to be pathogenic with LD₅₀ of 1.24 × 10⁵ and 1.88 × 10⁷ cfu/fish respectively. Histopathological examination of gill, liver and kidney revealed prominent changes supporting bacterial septicaemia. The investigation reports for the first time on concurrent infection by A. junii and multidrug‐resistant (MDR)‐A. pittii as emerging fish pathogens to cause severe mortality in Labeo catla and Hypophthalmichthys molitrix in a freshwater wetland.     

Physicochemical and Microbiological Changes During Drying of Wolf Herring (Chirocentrus dorab) and Coastal Trevally (Carangoides coeruleopinnatus)

The aim of the present study was to assess the physicochemical and microbiological changes during sun drying of salted wolf herring (Chirocentrus dorab) and coastal trevally (Carangoides coeruleopinnatus). For that purpose, the pH value, moisture, sodium chloride (NaCl) content, total volatile base nitrogen (TVB-N), histamine, cadaverine, putrescine, tryptamine, tyramine, spermine, and spermidine, as well as total aerobic mesophilic count, amine forming bacteria, total coliforms, Staphylococcus spp., and Bacillus spp., were determined. The initial pH value was 6.4 and increased during the salt drying process to 6.9 in both cases. The initial moisture, salt, and TVB-N levels of C. dorab and C. coeruleopinnatus were 64.3 and 60.3%, 2.55 and 2.70%, and 22.8 and 16.2 mg/100 g, respectively. At the end of drying, moisture decreased to 31.3 and 35.6%, respectively; salt increased to 13.71 and 16.04%; and TVB-N increased to 35.9 and 33.13 mg/100 g, respectively. Regarding total aerobic mesophilic count, amine forming bacteria, total coliforms, Staphylococcus spp., and Bacillus spp., a statistically significant (P < 0.05) reduction of the population was observed in both cases. Regarding the biogenic amine forming bacteria, Morganella morganii and Klebsiella pneumoniae were isolated from C. dorab; Bacillus cereus, Staphylococcus xylosus, and Providencia rettgeri were isolated from C. coeruleopinnatus. During sun drying, the amount of histamine, cadaverine, putrescine, and tryptamine was reduced; spermine was detected in C. dorab only during the first day, whereas spermidine was not detected. This reduction may be attributed to the presence of biogenic amine decomposing bacteria. However, further research is necessary in order to verify in situ this capacity and exploit potential applications for fish and fishery products.     

Prevalence and antimicrobial resistance of vibrios of human health significance in inland saline aquaculture areas

This study was conducted to evaluate the prevalence, potential pathogenicity and antimicrobial resistance of Vibrio isolates from 65 soil/water/fish samples collected from inland saline aquaculture areas. Depending on the sample type, presumptive Vibrio counts ranged from 2.50 to 6.16 log₁₀ CFU/ml (or/g). Among the 119 confirmed Vibrio isolates, Vibrio cholerae was found to most dominant (91.6%) and it was detected in all the samples from inland saline areas. Seven other Vibrio spp. including Vibrio parahaemolyticus and Vibrio vulnificus were also detected. Except one O139 serotype, rest of the V. cholerae isolates were found belonging to non‐O1/non‐O139 serogroups. None of the V. cholerae isolate was found positive for ctx gene. Antimicrobial susceptibility testing against 7 commonly used antibiotics revealed highest resistance (50.4%) against ampicillin. Very high intermediate resistance (87.4%) was also observed against erythromycin. Contrary to previous studies, high susceptibility (>70%) to chloramphenicol, nalidixic acid, tetracycline and trimethoprim was observed in Vibrio isolates obtained in present study. Almost 20% of Vibrio isolates were resistant to two or more antibiotic classes with multiple antibiotic resistance (MAR) index value of ≥0.28. Presence of V. cholerae isolates with very high MAR index value of 0.85 also suggested that these multidrug‐resistant environment isolates could serve as reservoir of antibiotic‐resistant genes in aquatic systems. The presence of multiple drug resistance vibrios in emerging inland saline aquaculture systems emphasizes the need for their routine monitoring for developing the risk assessment and mitigation strategies.     

Identification and partial characterization of potential probiotic lactic acid bacteria in freshwater Labeo rohita and Cirrhinus mrigala

Carps are the most diversified freshwater fish belonging to family Cyprinidae. Numerous probiotic and pathogenic lactic acid bacteria (LAB) have been characterized from carps. However, the diversity of these ecologically important bacteria is entirely unknown in freshwater fish of Pakistan. The present study aimed to characterize and identify the lactic acid bacteria from two carps viz. Laboe rohita and Cirrhinus mrigala and determine their antagonistic activity. Seventeen bacterial isolates were purified from the gastrointestinal tract and gills of these fish and characterized morphologically. Initially, seven isolates were screened as LAB using agar supplemented with CaCO₃. Subsequently, only two isolates CILB2 and RIL10 were selected as LAB after high‐performance liquid chromatography analysis for lactic acid production. Isolates CILB2 and RIL10 were genetically identified as Enterococcus faecalis and Weissella sp., respectively after 16S rRNA gene sequence analysis. Both strains exhibited significant antagonistic activity against common fish pathogens Streptococcus agalactiae, Staphylococcus aureus and Escherichia coli. Enterococcus faecalis CILB2 and Weissella sp. RIL10 were also found negative for haemolysis and gelatinase activities and were sensitive to ampicillin, amoxicillin, doxycycline, erythromycin, chloramphenicol and co‐trimoxazole antibiotics. The identified LAB strains may further be investigated for their potential probiotic application in fish feed and food preservation techniques.     

Characterization of spaC‐type Erysipelothrix sp. isolates causing systemic disease in ornamental fish

Since 2012, low‐to‐moderate mortality associated with an Erysipelothrix sp. bacterium has been reported in ornamental fish. Histological findings have included facial cellulitis, necrotizing dermatitis and myositis, and disseminated coelomitis with abundant intralesional Gram‐positive bacterial colonies. Sixteen Erysipelothrix sp. isolates identified phenotypically as E. rhusiopathiae were recovered from diseased cyprinid and characid fish. Similar clinical and histological changes were also observed in zebrafish, Danio rerio, challenged by intracoelomic injection. The Erysipelothrix sp. isolates from ornamental fish were compared phenotypically and genetically to E. rhusiopathiae and E. tonsillarum isolates recovered from aquatic and terrestrial animals from multiple facilities. Results demonstrated that isolates from diseased fish were largely clonal and divergent from E. rhusiopathiae and E. tonsillarum isolates from normal fish skin, marine mammals and terrestrial animals. All ornamental fish isolates were PCR positive for spaC, with marked genetic divergence (<92% similarity at gyrB, <60% similarity by rep‐PCR) between the ornamental fish isolates and other Erysipelothrix spp. isolates. This study supports previous work citing the genetic variability of Erysipelothrix spp. spa types and suggests isolates from diseased ornamental fish may represent a genetically distinct species.     

Diversity of siderophore‐producing bacteria isolated from the intestinal tracts of fish along the Japanese coast

This study examined the diversity of siderophore‐producing bacteria in the intestinal tracts of coastal fish in Japan and screened candidate bacterial strains for probiotic use in aquaculture. Of the 2637 bacteria isolated from the 27 fish specimens (13 species) and six environmental samples collected in this study, 266 isolates exhibited the ability to produce siderophores. Siderophore producers were detected in the intestines of 18 of the fish specimens caught (68%) at densities of 2.3 × 10⁴–2.3 × 10⁸ CFU g^?1, in all three seawater samples at 2.0 × 10²–1.3 × 10³ CFU mL^?1 and in all three sand samples at 2.6 × 10¹–2.8 × 10⁴ CFU g^?1. These findings suggest that siderophore‐producing bacteria are widely distributed in the intestinal tracts of coastal fish and their environments. Analysis of 16S rRNA gene sequences revealed that the siderophore producers belonged to 38 species, of which Vibrio splendidus, Vibrio ichthyoenteri and Vibrio crassostreae accounted for 32.7%, 19.5% and 11.3% of the 266 isolates, respectively, suggesting that these bacteria are indigenous to the intestinal tract of coastal fish. Six bacterial species, Enterovibri norvegicus, Photobacterium leiognathi, Photobacterium phosphoreum, Photobacterium rosenbergii, V. crassostrea and Vibrio scophthalmi were identified as possible candidates for use as probionts in fish aquaculture.     

The effect of dietary inulin on aerobic bacteria associated with hindgut of Arctic charr (Salvelinus alpinus L.)

The primary aim of the present study was to evaluate the population level of adherent (autochthonous) aerobic and facultative anaerobic bacteria in the hindgut of healthy Arctic charr (Salvelinus alpinus L.) fed dextrin or inulin. This was assessed by the dilution plate technique, and visualized using both transmission and scanning electron microscopy. A population level of 4.8 × 10⁵ adherent bacteria per gram wet mass was found in the hindgut of fish fed a casein‐based diet supplemented with 15% dextrin. However, substituting dextrin with 15% inulin reduced the bacterial population level in the hindgut (3.56 × 10⁴). A total of 217 bacterial isolates were identified by key phenotypical and biochemical characteristics. In addition, 22 strains were also identified by partial sequencing of the 16S rRNA gene. The composition of bacteria colonizing the hindgut of Arctic charr fed dextrin was dominated by the genera Staphylococcus, Pseudomonas, Micrococcus, Psychrobacter glacincola and Streptococcus. However, bacteria colonizing the hindgut of fish fed inulin were dominated by Gram‐positive bacteria of the genera Staphylococcus, Streptococcus, Carnobacterium and Bacillus. While Carnobacterium divergens‐like strains were isolated from charr fed dextrin, Carnobacterium maltaromicus‐like strains were isolated from the hindgut of fish fed inulin. Electron microscopical analysis of hindgut regions confirmed traditional culture‐based microbial analysis as fewer bacterial cells were observed between microvilli and associated with the surfaces of enterocytes of fish fed inulin rather than dextrin.     

Phenotypic and genotypic characterization of Streptococcus spp. isolated from tilapia (Oreochromis spp.) cultured in river-based cage and earthen ponds in Northern Thailand

Streptococcus spp. are major pathogenic bacteria associated with massive mortality in tilapia. This study investigated the phenotypic and genotypic characterization of Streptococcus agalactiae (GBS) and Streptococcus iniae (S. iniae) isolated from tilapia in river-based floating cage and earthen pond farms in northern Thailand. Isolates were identified by biochemical and molecular analyses. Capsular typing, enterobacterial repetitive intergenic consensus polymerase chain reaction and multilocus sequence typing were performed to investigate the genetic relatedness. Six and one isolates were confirmed as GBS and S. iniae, respectively. All Streptococcus spp. isolates were obtained from 4 river-based cage farms (4/33), while samples collected from earthen pond farms (N = 28) were negative for streptococcosis. All GBS with serotype Ⅲ and sequence type (ST) 283 was observed. The β-haemolytic GBS isolates were resistant to five antimicrobials, while the S. iniae was susceptible to all antimicrobials. This study indicates both GBS and S. iniae are the major bacterial pathogens responsible for streptococcosis infection in farmed tilapia of northern Thailand with GBS as dominant species. This survey highlights that the river-based cage farms seriously impact on the healthy development of the tilapia industry.     

Molecular Screening and Biocontrol of Aflatoxigenic Fungi in Fish Feed

ABSTRACT

Contamination of fish feed (FF) with aflatoxigenic fungi is an ongoing hazardous risk and is the first route for transferring aflatoxins to consumers from fish products. Prevention is the gold standard to deal with microbial food contamination. Different FF samples (n = 38) were screened for the occurrence of fungal contamination, and identified Aspergillus flavus isolates were assessed for the presence of aflatoxigenic production genes (Nor-1, Ver-1, and Omt-A) and the ability to produce aflatoxins. Biocontrol of aflatoxigenic A. flavus was proposed via exposure to plant (Cinnamon bark, Athl, and Lilac leaves) smoldering fumes (PSF). Numerous fungal species were identified from FF; Aspergillus spp. was the prevalent genus, followed by Penicillium spp. and Fusarium spp. The complete set of aflatoxigenic genes was detected in 54.8% of A. flavus isolates and was correlated with their aflatoxin-producing ability. The PSF exposure was successful for inhibiting A. flavus growth in agar media and in contaminated fish feed. Cinnamon PSF was the most effective and could entirely inhibit fungal growth at a proportion of 20 g/m³ in both treatments. The PSF exposures could be suggested as effectual techniques for complete biocontrol of aflatoxigenic fungi in fish feed to protect fish and consumers from their threatening effects.     

Identification of bacterial agents causing mortality in postlarvae of giant freshwater prawn (Macrobrachium rosenbergii) in south‐west coastal districts of Bangladesh

Following an incidence of freshwater prawn Macrobrachium rosenbergii postlarvae (PL) mortality in hatcheries in summer 2012, samples from dead PL, rearing water and prawn feed from two south‐west coastal districts of Bangladesh were collected to isolate, identify and characterize the agents causing PL mortality. Antibiogram profile of sixteen randomly selected bacteria, isolated from dead PL, that grew on TCBS, to 20 different antibiotics belonging to 12 major groups revealed that the drug resistance pattern varied from moderate (56% to the drugs: ciprofloxacin, cefotaxime, nitrofurantoin, kanamycin) to complete (to penicillin, ceftazidime and oxacillin) level. To identify the isolates, amplified rDNA restriction analysis (ARDRA) classified them in to four groups, and RAPD (randomly amplified polymorphic DNA) typing yielded nine different types of isolates within these four ARDRA groups. The 16S rDNA gene sequences identified that the groups were genotypically diverse belonging to the bacterial species: Enterobacter cloacae, Klebsiella pneumoniae, Exiguobacterium profundum and Enterococcus casseliflavus, respectively, that all demonstrated their killing potential to PLs in a simulated environment. The study therefore identified four different bacterial pathogens, one of which, Exiguobacterium profundum is reported for the first here in Bangladesh, that demand special consideration for disease management strategy.     

Natural outbreak of Streptococcus agalactiae (GBS) infection in wild giant Queensland grouper, Epinephelus lanceolatus (Bloch), and other wild fish in northern Queensland, Australia

Ninety‐three giant Queensland grouper, Epinephelus lanceolatus (Bloch), were found dead in Queensland, Australia, from 2007 to 2011. Most dead fish occurred in northern Queensland, with a peak of mortalities in Cairns in June 2008. In 2009, sick wild fish including giant sea catfish, Arius thalassinus (Rüppell), and javelin grunter, Pomadasys kaakan (Cuvier), also occurred in Cairns. In 2009 and 2010, two disease epizootics involving wild stingrays occurred at Sea World marine aquarium. Necropsy, histopathology, bacteriology and PCR determined that the cause of deaths of 12 giant Queensland grouper, three wild fish, six estuary rays, Dasyatis fluviorum (Ogilby), one mangrove whipray, Himantura granulata (Macleay), and one eastern shovelnose ray, Aptychotrema rostrata (Shaw), was Streptococcus agalactiae septicaemia. Biochemical testing of 34 S. agalactiae isolates from giant Queensland grouper, wild fish and stingrays showed all had identical biochemical profiles. The 16S rRNA gene sequences of isolates confirmed all isolates were S. agalactiae; genotyping of selected S. agalactiae isolates showed the isolates from giant Queensland grouper were serotype Ib, whereas isolates from wild fish and stingrays closely resembled serotype II. This is the first report of S. agalactiae from wild giant Queensland grouper and other wild tropical fish and stingray species in Queensland, Australia.     

Genotypic diversity among Shewanella spp. collected from freshwater fish

Different Shewanella species are isolated both from healthy and from diseased fish. To date, contemporary methods do not provide sufficient insight to determine species and detail differentiation between tested strains. Bacteria isolated from cultured (n = 33), wild (n = 12) and ornamental (n = 6) fish, as well as several reference strains, were tested by 16S rRNA gene sequencing, ERIC‐PCR and pulsed‐field gel electrophoresis (PFGE) assays. Our study indicates that isolates collected from freshwater fish were genetically diverse. Based on 16S rRNA gene sequences, bacteria were clustered into groups S. putrefaciens, S. xiamenensis and S. oneidensis. Some isolates were classified only to genus Shewanella; thus, 16S rRNA gene analyses were not enough to determine the species. ERIC‐PCR revealed 49 different genotype profiles indicating that the method might be useful for differentiation of Shewanella isolates irrespectively to species identification, contrary to PFGE which is not suitable for Shewanella typing.     

Multilocus sequence analysis revealed a high genotypic diversity of Aeromonas hydrophila infecting fish in Uganda

A multilocus sequence analysis (MLSA) was carried out to delineate Aeromonas hydrophila from fish in Uganda. Five housekeeping genes including recA, gyrB, metG, gltA and pps; and the 16S rRNA gene were amplified and sequenced from a total of nine A. hydrophila isolates. The obtained sequences were edited, and consensus sequences generated for each gene locus. The housekeeping gene sequences were concatenated and phylogenetic analysis performed in MEGA version 7.0.2. Pairwise distances ranged from 0.000 to 0.118, highest within the gltA gene locus and lowest within the 16S rRNA gene. The average evolutionary diversity within isolates from the same source ranged between 0.002 and 0.037, and it was 0.033 between the different sources. Similar tree topologies were obtained from the different gene loci with recA, metG and gyrB being more consistent in discriminating isolates according to sources while the 16S rRNA gene had the lowest resolution. The concatenated tree had the highest discriminatory power. This study revealed that A. hydrophila strains infecting fish in Uganda are of diverse genotypes suggesting different sources of infection in a given outbreak. Efforts to minimize spread of the bacteria across sources should be emphasized to control infections of mixed genotypes.     

Incidence of class 1 integron and other antibiotic resistance determinants in Aeromonas spp. from rainbow trout farms in Australia

There is limited information on antibiotic resistance determinants present in bacteria of aquaculture origin in Australia. The presence of integron and other resistance determinants was investigated in 90 Aeromonas isolates derived from nine freshwater trout farms in Victoria (Australia). Polymerase chain reaction was carried out for the detection of integrase genes Int1, Int2 and Int3, gene cassette array, integron‐associated aadA, sul1 and qac1 genes, streptomycin resistance genes strA‐strB, β‐lactamase resistance genes bla_TEM and bla_SHV, and tetracycline resistance genes tetA‐E and tetM. Clonal analysis was performed by pulsed‐field gel electrophoresis (PFGE). Class 1 integrons were detected in 28/90 (31%) and class 2 and class 3 in none of the strains, aadA gene in 19/27 (70%) streptomycin‐resistant strains, sul1 in 13/15 (86.7%) sulphonamide‐resistant strains and qac1 gene in 8/28 (28.6%) integron‐bearing strains. Five strains from two different farms carried gene cassettes of 1000 bp each containing the aadA2 gene and PFGE analysis revealed genetic relatedness. tetC was detected in all and tetA in 9/18 (50%) tetracycline‐resistant strains. The strA‐strB, bla_TEM or bla_SHV genes were not detected in any of the strains. Aeromonas spp. carrying integrons and other resistance genes are present in farm‐raised fish and sediments even though no antibiotics were licensed for use in Australian aquaculture at the time of the study.     

Antibiotic resistance and repetitive‐element PCR fingerprinting in Aeromonas veronii isolates

This study evaluated antibiotic resistance and the related genes in total 47 Aeromonas veronii isolates from pet fish, eel (Anguilla japonica) and koi (Cyprinus carpio) in Korea. In comparison with the antibiotic susceptibilities of isolates from eel and koi, those of pet fish were more resistant to ceftiofur, aminoglycosides, tetracycline and nitrofurantoin. And isolates from pet fish showed high prevalences of class 1 integron, quinolones and tetracycline resistance determinants than those from eel and koi. Repetitive‐element palindromic PCR (rep‐PCR) showed larger diversities among A. veronii isolates. Collectively, pet fish may be a reservoir for multiple clones of A. veronii involved in antibiotic resistance. In this aspect, imported fish in the aquaculture trade should be steadily and continually screened for bacterial antibiotic resistance and related genes.