Occurrence of tetracycline resistance genes tet(M) and tet(S) in bacteria from marine aquaculture sites. - Related Documents




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280001.0000Occurrence of tetracycline resistance genes tet(M) and tet(S) in bacteria from marine aquaculture sites. Occurrence of tetracycline resistance genes encoding ribosomal protection proteins was examined in 151 tetracycline-resistant bacterial isolates from fish and seawater at coastal aquaculture sites in Japan and Korea. The tet(M) gene was detected in 34 Japanese and Korean isolates, which included Vibrio sp., Lactococcus garvieae, Photobacterium damsela subsp. piscicida, and unidentified Gram-positive bacteria. The majority of these bacterial isolates displayed high-level resistance with a minimum inhibitory concentrations (MICs) equal to or greater than 250 microg/ml of oxytetracycline and only four isolates had MICs less than 31.3 microg/ml. 16S rDNA RFLP typing of tet(M)-positive Vibrio isolates suggests that these are clonal populations of the same phylotype specific to a particular location. One Vibrio clone (phylotype III), however, is widely disseminated, being detected during different sampling years, at different locations, and in different fish species in both Japan and Korea. The tet(S) gene was detected in L. garvieae from yellowtail in Japan and in Vibrio sp. from seawater in Korea. This is the first report of tet(S) occurrence in Gram-negative facultative anaerobes. These results suggest that tet(M) and tet(S) genes are present in fish intestinal and seawater bacteria at aquaculture sites and could be an important reservoir of tetracycline resistance genes in the marine environment.200415268950
592110.9999Prevalence of tetracycline resistance genes in oral bacteria. Tetracycline is a broad-spectrum antibiotic used in humans, animals, and aquaculture; therefore, many bacteria from different ecosystems are exposed to this antibiotic. In order to determine the genetic basis for resistance to tetracycline in bacteria from the oral cavity, saliva and dental plaque samples were obtained from 20 healthy adults who had not taken antibiotics during the previous 3 months. The samples were screened for the presence of bacteria resistant to tetracycline, and the tetracycline resistance genes in these isolates were identified by multiplex PCR and DNA sequencing. Tetracycline-resistant bacteria constituted an average of 11% of the total cultivable oral microflora. A representative 105 tetracycline-resistant isolates from the 20 samples were investigated; most of the isolates carried tetracycline resistance genes encoding a ribosomal protection protein. The most common tet gene identified was tet(M), which was found in 79% of all the isolates. The second most common gene identified was tet(W), which was found in 21% of all the isolates, followed by tet(O) and tet(Q) (10.5 and 9.5% of the isolates, respectively) and then tet(S) (2.8% of the isolates). Tetracycline resistance genes encoding an efflux protein were detected in 4.8% of all the tetracycline-resistant isolates; 2.8% of the isolates had tet(L) and 1% carried tet(A) and tet(K) each. The results have shown that a variety of tetracycline resistance genes are present in the oral microflora of healthy adults. This is the first report of tet(W) in oral bacteria and the first report to show that tet(O), tet(Q), tet(A), and tet(S) can be found in some oral species.200312604515
592020.9998Study on acquisition of bacterial antibiotic resistance determinants in poultry litter. Antibiotic resistance and the mode of transmission were investigated in bacteria isolated from poultry litter. Total aerobic heterotrophic bacteria were screened and identified for their resistance to different antibiotics such as ampicillin, streptomycin, erythromycin, tetracycline, chloramphenicol, kanamycin, tobramycin, and rifampicin. The distribution of bacteria found in the litter was Staphylococcus (29.1%), which was the predominant group, followed by Streptococcus (25%), Micrococcus (20.8%), Escherichia coli (12.5%), Salmonella (8.3%), and Aeromonas (4.1%). Fifty percent of these isolates were susceptible to ampicillin, 57% to erythromycin, 25% to tetracycline, 4% to chloramphenicol, 40% to kanamycin, 75% to streptomycin, 54% to tobramycin, and 4% to rifampicin. Three randomly selected isolates representing Staphylococcus, Streptococcus, and Micrococcus were examined for plasmids, and plasmid-curing and plasmid-induced transformation studies were conducted. Streptococcus and Micrococcus harbored a plasmid of 4.2 and 5.1 kb, respectively, whereas Staphylococcus did not harbor any plasmids. Plasmids were cured in Streptococcus and Micrococcus at a concentration of 75 and 100 microg/ mL of acridine orange, respectively, and transformation of 4.2- and 5.1-kb plasmids isolated from the Streptococcus and Micrococcus to plasmid-free E. coli DH5alpha strain was possible. In conjugation experiments, the antibiotic resistance profiles of transconjugant cells were found to be the same as the donors with the exception of Staphylococcus. The results of this study suggest that transformation and conjugation could be an important mechanism for horizontal gene transfer between bacteria in poultry litter. An understanding of the mechanism and magnitude of resistance gene transfer may provide a strategy to reduce the potential for dissemination of these genes.200919531707
292130.9998Diversity of tetracycline resistance genes in bacteria from aquaculture sources in Australia. AIMS: To determine the genetic determinants responsible for tetracycline resistance in oxytetracycline resistant bacteria from aquaculture sources in Australia. METHODS AND RESULTS: Twenty of 104 (19%) isolates tested were resistant to oxytetracycline (MIC > or = 16 microg ml(-1)). Using polymerase chain reaction (PCR) amplification, one or more tet genes were detected in 15/20 (75%) isolates tested, but none were found in 5/20 (25%). tetM (50%) was the most common determinant, followed by tetE (45%), tetA (35%) and tetD (15%). Five of 12 oxytetracycline resistant isolates studied were able to transfer their R-plasmid to Escherichia coli recipients of chicken, pig and human origin. tetA, tetD and tetM were found to be transferred while tetE was not transferred. Southern hybridization and PCR were used to confirm transfer of determinants. CONCLUSIONS: Bacterial isolates from aquaculture sources in Australia harbour a variety of tetracycline resistance genes, which can be transferred to other bacteria of different origin. SIGNIFICANCE AND IMPACT OF THE STUDY: Bacteria from aquaculture sources in Australia contribute to the resistance gene pool reservoir. The in vitro transfer of tetracycline R-plasmid from aquatic bacteria to E. coli isolates from various sources is an indication of the potential public health risk associated with these resistance determinants.200717953612
279940.9998Genetic and physiological characterization of oxytetracycline-resistant bacteria from giant prawn farms. Four hundred and thirteen oxytetracycline-resistant bacteria were recovered from six freshwater giant prawn farms with a history of oxytetracycline use. Most oxytetracyclineresistant isolates were Gram-negative bacteria. Six groups of oxytetracycline-resistant bacteria were classified using cluster analysis based on a comparison of levels of oxytetracycline resistance. Complex fingerprint patterns were obtained for 71 isolates studied. In general, the band patterns of isolates from different ponds were very similar, and the data indicated that the isolates were closely related. The exploration for crossresistance found that most of the 71 oxytetracycline-resistant isolates were also resistant to tetracycline and chlortetracycline, but had a relatively low resistance to doxycycline. Many isolates showed higher chlortetracycline resistance than oxytetracycline resistance. Additionally, the oxytetracyclineresistant isolates were examined for the presence of tetracycline resistance (tet) genes. Fifty percent of the isolates carried one of the 14 known tet genes examined. The most common determinants were TetA and TetD. However, TetB, TetC, TetE, TetK, TetL, and TetM were also found with various frequencies.200818309262
291050.9998Phenotypic and genotypic characterization of tetracycline and minocycline resistance in Clostridium perfringens. The aim of this study was to determine the incidence of tetracycline resistance and the prevalence of tetracycline-resistance genes in strains of Clostridium perfringens isolated from different sources between 1994 and 2005. Susceptibility to tetracycline and minocycline in strains from humans (35 isolates), chickens (15 isolates), food (21 isolates), soil (16 isolates) and veterinary sources (6 isolates) was determined, and tetracycline-resistance genes were detected. Resistance was most common in strains isolated from chickens, followed by those from soils, clinical samples and foods. The most highly resistant strains were found among clinical and food isolates. tetA(P) was the most common resistance gene, and along with tetB(P) was found in all resistant strains and some sensitive strains. One tetracycline-resistant food isolate had an intact tet(M) gene. However, PCR fragments of 0.4 or 0.8 kb with high degrees of identity to parts of the tet(M) sequences of other bacteria were found, mainly in clinical isolates, and often in isolates with tetB(P). No correlation between level of sensitivity to tetracycline or minocycline and the presence of tetA(P), tetB(P) or part of tet(M) was found. The presence of part of tet(M) in some strains of C. perfringens containing tetB(P) may have occurred by recent gene transfer.201020661548
291660.9998The identification of a tetracycline resistance gene tet(M), on a Tn916-like transposon, in the Bacillus cereus group. In order to investigate whether resistance genes present in bacteria in manure could transfer to indigenous soil bacteria, resistant isolates belonging to the Bacillus cereus group (Bacillus cereus, Bacillus anthracis and Bacillus thuringiensis) were isolated from farm soil (72 isolates) and manure (12 isolates) samples. These isolates were screened for tetracycline resistance genes (tet(K), tet(L), tet(M), tet(O), tet(S) and tet(T)). Of 88 isolates examined, three (3.4%) isolates carried both tet(M) and tet(L) genes, while four (4.5%) isolates carried the tet(L) gene. Eighty-one (92.1%) isolates did not contain any of the tested genes. All tet(M) positive isolates carried transposon Tn916 and could transfer this mobile DNA element to other Gram-positive bacteria.200212351239
279770.9998Widespread distribution of tetracycline resistance genes in a confined animal feeding facility. We sought to determine the distribution of resistance and the tetracycline resistance genes among bacteria isolated from a swine confined animal feeding facility where tetracycline-containing feed had been in use for over 20 years. Samples collected from feed, hogs, hog houses, waste lagoon, soil, surface water and well water were screened for the presence of (a) resistant Escherichia coli and enterococci and (b) tetracycline-resistant strains of all species. Genomic DNA was extracted from the latter strain collection and fragments from 16S rDNA and ten tetracycline resistance genes (tetA, tetB, tetC, tetE, tetH, tetL, tetM, tetS, tetT and rumB) were polymerase chain reaction-amplified and a partial nucleotide sequence was obtained. In this environment, 77% of E. coli and 68% of enterococci isolated were tetracycline resistant. Tetracycline resistance was found in 26 different bacterial genera and in 60 species. Single resistance gene alleles (as defined by nucleotide sequence) were present in multiple species. There was evidence of gene recombination and multiple different tetracycline resistance genes were present in single bacterial isolates. These data provide further evidence for the widespread distribution of resistance genes in microbial populations in settings in which there is ongoing subtherapeutic antimicrobial use.200717287111
599680.9998Molecular characterization of intrinsic and acquired antibiotic resistance in lactic acid bacteria and bifidobacteria. The minimum inhibitory concentrations (MICs) of 6 different antibiotics (chloramphenicol, clindamycin, erythromycin, streptomycin, tetracycline and vancomycin) were determined for 143 strains of lactic acid bacteria and bifidobacteria using the Etest. Different MICs were found for different species and strains. Based on the distribution of these MIC values, most of the strains were either susceptible or intrinsically resistant to these antibiotics. However, the MIC range of some of these antibiotics showed a bimodal distribution, which suggested that some of the tested strains possess acquired antibiotic resistance. Screening for resistance genes was performed by PCR using specific primers, or using a DNA microarray with around 300 nucleotide probes representing 7 classes of antibiotic resistance genes. The genes identified encoded resistance to tetracycline [tet(M), tet(W), tet(O) and tet(O/W)], erythromycin and clindamycin [erm(B)] and streptomycin [aph(E) and sat(3)]. Internal portions of some of these determinants were sequenced and found to be identical to genes described in other bacteria. All resistance determinants were located on the bacterial chromosome, except for tet(M), which was identified on plasmids in Lactococcus lactis. The contribution of intrinsic multidrug transporters to the antibiotic resistance was investigated by cloning and measuring the expression of Bifidobacterium breve genes in L. lactis.200817957105
289690.9998Resistance gene patterns of tetracycline resistant Escherichia coli of human and porcine origin. Resistance transfer from animals to humans (and vice versa) is a frequently discussed topic in human and veterinary medicine, albeit relevant studies focus mainly on phenotypic antibiotic resistance. In order to get a comparative insight regarding the distribution of selected resistance genes [tet(A/B/C/D/M/K/L/O/S/W/Z), sulI, II, III, str(A/B), aad(A)] in Escherichia coli of different origins, phenotypically tetracycline resistant isolates of porcine and human origin (n=137 and 152) were investigated using PCR. The most common gene was tet(A) in porcine, but tet(B) in human isolates (>55%). Tet(C/M/D) were rare (1-7%); tet(K/L/O/S/W/Z) were not detected. Co-occurrence of tet(A) and tet(B) was more frequent in human strains (11% vs. 2%). 88% of the porcine isolates had one, and 9% had two tet-genes. By contrast, only 69% of the human strains had one tet-gene, whereas 17% were carriers of two tet-determinants. The most common sulfonamide resistance gene was represented by sulII (40% in porcine, 62% in human isolates), followed by sulI. SulIII was present in eight isolates. Streptomycin resistance was mostly mediated by str(A)/str(B) in porcine, and by str(A)/str(B)/aad(A) in human strains (35% each). In one E. coli of human origin, 7 resistance genes were simultaneously detected. Co-occurrence of 5 or 6 resistance genes was more present in human strains, whereas porcine isolates carried more often only 1-4 genes. The huge diversities between gene patterns of bacteria of human and porcine origin indicate that genetic transfers between microorganisms from different sources are less frequent than transfers within populations of the same source.201019939589
2904100.9998The maintenance in the oral cavity of children of tetracycline-resistant bacteria and the genes encoding such resistance. OBJECTIVES: To investigate the maintenance of tetracycline-resistant oral bacteria and the genes encoding tetracycline resistance in these bacteria in children (aged 4--6 years) over a period of 12 months. METHODS: Plaque and saliva samples were taken from 26 children. Tetracycline-resistant bacteria were isolated and identified. The types of resistance genes and their genetic locations were also determined. RESULTS: Fifteen out of 18 children harboured tetracycline-resistant (defined as having a MIC>or=8 mg/L) oral bacteria at all three time points. The median percentage of tetracycline-resistant bacteria at 0, 6 and 12 months was 1.37, 1.37 and 0.85%, respectively; these were not significantly different. The MIC(50) of the group was 64 mg/L at all three time points compared with the MIC(90), which was 64 mg/L at 0 months, and 128 mg/L at 6 and 12 months. The most prevalent resistant species were streptococci (68%), which were isolated at all three time points in 13 children. The most prevalent gene encoding tetracycline resistance was tet(M) and this was found in different species at all three time points. For the first time, tet(32) was found in Streptococcus parasanguinis and Eubacterium saburreum. PCR and Southern-blot analysis (on isolates from three of the children) showed that the tet(M) gene was located on a Tn916-like element and could be detected at all three time points, in four different genera, Streptococcus, Granulicatella, Veillonella and Neisseria. CONCLUSIONS: The results of this study show that tetracycline-resistant bacteria and tet(M) are maintained within the indigenous oral microbiota of children, even though they are unlikely to have been directly exposed to tetracycline.200516027144
5997110.9998Resistance of potential probiotic lactic acid bacteria and bifidobacteria of African and European origin to antimicrobials: determination and transferability of the resistance genes to other bacteria. Probiotic bacteria and starter cultures of Lactobacillus, Weissella and Bifidobacterium of African and European origins were studied and compared for their susceptibility to antimicrobials. The study included, for all isolates, determination of MICs (Minimal Inhibitory Concentration) for 24 antimicrobials, detection of resistance genes by PCR reactions using specific primers and sequencing of positive amplicons. The ability of Lb. reuteri from Africa to transfer the erythromycin resistance gene erm(B) to closely related bacteria was investigated by conjugation. Variations were observed and high levels of intrinsic resistance were found among the tested species. Positive amplicons were obtained for resistance genes encoding aminoglycoside (aph(3')-III, aadA, aadE) and tetracycline (tet(S)) from isolates from Europe and macrolide (erm(B)) from an isolate from Africa. However, only the erm(B) gene found in Lb. reuteri L4: 12002 from Africa contained a homologous sequence to previously published sequences. This gene could be transferred in vitro to enterococci. Higher prevalence of phenotypic resistance for aminoglycoside was found in isolates from Europe.200818063151
2922120.9998Tetracycline-resistance genes in gram-negative isolates from estuarine waters. AIMS: To investigate the diversity and dissemination of tetracycline resistance genes in isolates from estuarine waters. METHODS AND RESULTS: Forty-two out of 164 multi-resistant isolates previously obtained were resistant or less-susceptible to tetracycline, as evaluated by the disc diffusion method. Minimal inhibitory concentration for resistant bacteria ranged from 16 to 256 mg l(-1). Screening of tet genes by polymerase chain reaction showed that 88% of the isolates carried at least one of the genes tested, namely tet(A) (present in 13 isolates), tet(B) (present in 13 isolates), tet(C) (present in 3 isolates), tet(D) (present in 1 isolate), tet(E) (present in 6 isolates) and tet(M) (present in 1 isolate). One isolate carried tet(A) and tet(M). To our knowledge, this study presents the first description of a tet(D) gene in Morganella morganii. Hybridization revealed that tet genes were plasmid-located in 31% of the isolates. Those isolates were included as donors in conjugation experiments and 38% transferred tetracycline resistance. CONCLUSIONS: A considerable diversity of tet genes was detected in the estuary. Frequently, these genes were associated with plasmids and could be transferred to Escherichia coli. SIGNIFICANCE AND IMPACT OF THE STUDY: The results presented provide further evidence of the role played by estuarine reservoirs in antibiotic resistance maintenance and dissemination.200819120920
5935130.9998Antibiotic resistance genes in anaerobic bacteria isolated from primary dental root canal infections. Fourty-one bacterial strains isolated from infected dental root canals and identified by 16S rRNA gene sequence were screened for the presence of 14 genes encoding resistance to beta-lactams, tetracycline and macrolides. Thirteen isolates (32%) were positive for at least one of the target antibiotic resistance genes. These strains carrying at least one antibiotic resistance gene belonged to 11 of the 26 (42%) infected root canals sampled. Two of these positive cases had two strains carrying resistance genes. Six out of 7 Fusobacterium strains harbored at least one of the target resistance genes. One Dialister invisus strain was positive for 3 resistance genes, and 4 other strains carried two of the target genes. Of the 6 antibiotic resistance genes detected in root canal strains, the most prevalent were blaTEM (17% of the strains), tetW (10%), and ermC (10%). Some as-yet-uncharacterized Fusobacterium and Prevotella isolates were positive for blaTEM, cfxA and tetM. Findings demonstrated that an unexpectedly large proportion of dental root canal isolates, including as-yet-uncharacterized strains previously regarded as uncultivated phylotypes, can carry antibiotic resistance genes.201223108290
3555140.9998Antimicrobial resistance and antimicrobial resistance genes in marine bacteria from salmon aquaculture and non-aquaculture sites. Antimicrobial resistance (AR) detected by disc diffusion and antimicrobial resistance genes detected by DNA hybridization and polymerase chain reaction with amplicon sequencing were studied in 124 marine bacterial isolates from a Chilean salmon aquaculture site and 76 from a site without aquaculture 8 km distant. Resistance to one or more antimicrobials was present in 81% of the isolates regardless of site. Resistance to tetracycline was most commonly encoded by tetA and tetG; to trimethoprim, by dfrA1, dfrA5 and dfrA12; to sulfamethizole, by sul1 and sul2; to amoxicillin, by blaTEM ; and to streptomycin, by strA-strB. Integron integrase intl1 was detected in 14 sul1-positive isolates, associated with aad9 gene cassettes in two from the aquaculture site. intl2 Integrase was only detected in three dfrA1-positive isolates from the aquaculture site and was not associated with gene cassettes in any. Of nine isolates tested for conjugation, two from the aquaculture site transferred AR determinants to Escherichia coli. High levels of AR in marine sediments from aquaculture and non-aquaculture sites suggest that dispersion of the large amounts of antimicrobials used in Chilean salmon aquaculture has created selective pressure in areas of the marine environment far removed from the initial site of use of these agents.201424612265
2905150.9998Scarce detection of mobile erm genes associated with tetQ in Bacteroides and Parabacteroides from Costa Rica. The frequency of finding of clindamycin-resistant anaerobic bacteria in clinical samples has doubled from 2008 to 2010 in Costa Rica. To determine whether this increase is due to dissemination of erm genes aided by tetQ elements, we analyzed 100 isolates of Bacteroides or Parabacteroides from a regional hospital, a national hospital, and the community. Antimicrobial susceptibilities were recorded with a broth micro-dilution method and erm genes were detected by PCR and Southern blotting. In addition, plasmid isolation and mating experiments were performed to clarify the location and mobility of the detected erm genes. Resistance to clindamycin was by far more frequent in the regional hospital (72%) than in the national hospital (29%) and the community (26%). Resistance to tetracycline was even more common, with the community (85%) outweighing the hospitals (71-72%). While MIC of clindamycin were higher in the hospitals than in the community (P < 0.05), the opposite was seen for tetracycline (P < 0.0001). Of the sought-after genes, only ermG (n = 2), ermA (n = 1), and ermF (n = 1) were detected in the hospitals and ermF in the community (n = 2). In opposition to the low frequency of finding of erm genes, 71% of the isolates were positive for tetQ. None of the detected genes were encoded on plasmids. Only three isolates from the hospitals transferred their erm genes laterally. By contrast, 13 hospital isolates and two community isolates transferred tetQ. Despite the widespread finding of tetracycline-resistant tetQ-positive bacteria, mobile erm genes were rare in our bacterial collection. We conclude that the detected erm genes are likely not included in typical conjugative transposons of Bacteroides and Parabacteroides.201323528984
5862160.9998Diversity of tetracycline resistance genes in bacteria from Chilean salmon farms. Twenty-five distinct tetracycline-resistant gram-negative bacteria recovered from four Chilean fish farms with no history of recent antibiotic use were examined for the presence of tetracycline resistance (tet) genes. Sixty percent of the isolates carried 1 of the 22 known tet genes examined. The distribution was as follows. The tet(A) gene was found in six isolates. The tet(B) gene was found in two isolates, including the first description in the genus Brevundimonas: Two isolates carried the tet(34) and tet(B) genes, including the first description of the tet(34) gene in Pseudomonas and Serratia and the first description of the tet(B) gene in Pseudomonas: The tet(H) gene was found in two isolates, which includes the first description in the genera Moraxella and Acinetobacter: One isolate carried tet(E), and one isolate carried tet(35), the first description of the gene in the genus Stenotrophomonas: Finally, one isolate carried tet(L), found for the first time in the genus Morganella: By DNA sequence analysis, the two tet(H) genes were indistinguishable from the previously sequenced tet(H) gene from Tn5706 found in Pasteurella multocida. The Acinetobacter radioresistens isolate also harbored the Tn5706-associated 1,063-bp IS element IS1597, while the Moraxella isolate carried a 1,026-bp IS-like element whose 293-amino-acid transposase protein exhibited 69% identity and 84% similarity to the transposase protein of IS1597, suggesting the presence of a novel IS element. The distribution of tet genes from the Chilean freshwater ponds was different than those that have previously been described from other geographical locations, with 40% of the isolates carrying unidentified tetracycline resistance genes.200312604516
2795170.9998Molecular identification and quantification of tetracycline and erythromycin resistance genes in Spanish and Italian retail cheeses. Large antibiotic resistance gene pools in the microbiota of foods may ultimately pose a risk for human health. This study reports the identification and quantification of tetracycline- and erythromycin-resistant populations, resistance genes, and gene diversity in traditional Spanish and Italian cheeses, via culturing, conventional PCR, real-time quantitative PCR (qPCR), and denaturing gradient gel electrophoresis (DGGE). The numbers of resistant bacteria varied widely among the antibiotics and the different cheese varieties; in some cheeses, all the bacterial populations seemed to be resistant. Up to eight antibiotic resistance genes were sought by gene-specific PCR, six with respect to tetracycline, that is, tet(K), tet(L), tet(M), tet(O), tet(S), and tet(W), and two with respect to erythromycin, that is, erm(B) and erm(F). The most common resistance genes in the analysed cheeses were tet(S), tet(W), tet(M), and erm(B). The copy numbers of these genes, as quantified by qPCR, ranged widely between cheeses (from 4.94 to 10.18log10/g). DGGE analysis revealed distinct banding profiles and two polymorphic nucleotide positions for tet(W)-carrying cheeses, though the similarity of the sequences suggests this tet(W) to have a monophyletic origin. Traditional cheeses would therefore appear to act as reservoirs for large numbers of many types of antibiotic resistance determinants.201425302306
5933180.9997Novel macrolide-resistance genes, mef(C) and mph(G), carried by plasmids from Vibrio and Photobacterium isolated from sediment and seawater of a coastal aquaculture site. The aim of this study was to determine whether mef(C) and mph(G), originally found on the transferable multi-drug plasmid pAQU1 from Photobacterium damselae subsp. damselae isolated from seawater of a fish farm, are responsible for conferring macrolide resistance. Since these genes are localized head-to-tail on pAQU1 and only four nucleotides exist between them, the single- and combination-effect of these genes was examined. When mph(G) alone was introduced to Escherichia coli, the minimum inhibitory concentrations (MICs) against erythromycin, clarithromycin and azithromycin increased, whereas introduction of mef(C) alone did not influence macrolide susceptibility. Introduction of both mef(C) and mph(G) dramatically increased the MICs to the same three macrolides, i.e. >512 μg ml(-1) , >512 μg ml(-1) and 128 μg ml(-1) respectively. These results suggest that the macrolide phosphotransferase encoded by mph(G) is essential for macrolide resistance, while the efflux pump encoded by mef(C) is required for high-level macrolide resistance. The tandem-pair arrangements of the mef(C) and mph(G) genes were conserved on plasmids ranging in size from 240 to 350 kb of the 22 erythromycin-resistant strains belonging to Vibrio and Photobacterium obtained from the fish farm. Sixteen of 22 plasmids ranged in size from 300 to 350 kb. This is the first report of novel macrolide resistance genes originating from a marine bacterium. SIGNIFICANCE AND IMPACT OF THE STUDY: In this study, mef(C) and mph(G) were found to be novel macrolide-resistance genes, and this is the first report of macrolide-resistance genes originating from a marine bacterium. These genes may be responsible for previously reported cases of the emergence of erythromycin-resistant bacteria in aquaculture sites by an unknown mechanism. The introduction of the tandem arrangement of the mef(C) and mph(G) genes in Escherichia coli increased the MICs to erythromycin, clarithromycin and azithromycin, suggesting a novel mechanism conferring high-level macrolide resistance via combined expression of the efflux pump and macrolide phosphotransferase.201525765542
2909190.9997Determination of the prevalence of antimicrobial resistance genes in canine Clostridium perfringens isolates. Clostridium perfringens is a well documented cause of a mild self-limiting diarrhea and a potentially fatal acute hemorrhagic diarrheal syndrome in the dog. A recent study documented that 21% of canine C. perfringens isolates had MIC's indicative of resistance to tetracycline, an antimicrobial commonly recommended for treatment of C. perfringens-associated diarrhea. The objective of the present study was to further evaluate the antimicrobial susceptibility profiles of these isolates by determining the prevalence of specific resistance genes, their expression, and ability for transference between bacteria. One hundred and twenty-four canine C. perfringens isolates from 124 dogs were evaluated. Minimum inhibitory concentrations of tetracycline, erythromycin, tylosin, and metronidazole were determined using the CLSI Reference Agar Dilution Method. All isolates were screened for three tetracycline resistance genes: tetA(P), tetB(P) and tetM, and two macrolide resistance genes: ermB and ermQ, via PCR using primer sequences previously described. Ninety-six percent (119/124) of the isolates were positive for the tetA(P) gene, and 41% (51/124) were positive for both the tetA(P) and tetB(P) genes. No isolates were positive for the tetB(P) gene alone. Highly susceptible isolates (MIC< or = 4 microg/ml) were significantly more likely to lack the tetB(P) gene. One isolate (0.8%) was positive for the ermB gene, and one isolate was positive for the ermQ gene. The tetM gene was not found in any of the isolates tested. Two out of 15 tested isolates (13%) demonstrated transfer of tetracycline resistance via bacterial conjugation. Tetracycline should be avoided for the treatment of C. perfringens-associated diarrhea in dogs because of the relatively high prevalence of in vitro resistance, and the potential for conjugative transfer of antimicrobial resistance.200616330169