Molecular identification and quantification of tetracycline and erythromycin resistance genes in Spanish and Italian retail cheeses. - Related Documents




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279501.0000Molecular 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
279610.9999Distribution of specific tetracycline and erythromycin resistance genes in environmental samples assessed by macroarray detection. A macroarray system was developed to screen environmental samples for the presence of specific tetracycline (Tc(R)) and erythromycin (erm(R)) resistance genes. The macroarray was loaded with polymerase chain reaction (PCR) amplicons of 23 Tc(R) genes and 10 erm(R) genes. Total bacterial genomic DNA was extracted from soil and animal faecal samples collected from different European countries. Macroarray hybridization was performed under stringent conditions and the results were analysed by fluorescence scanning. Pig herds in Norway, reared without antibiotic use, had a significantly lower incidence of antibiotic resistant bacteria than those reared in other European countries, and organic herds contained lower numbers of resistant bacteria than intensively farmed animals. The relative proportions of the different genes were constant across the different countries. Ribosome protection type Tc(R) genes were the most common resistance genes in animal faecal samples, with the tet(W) gene the most abundant, followed by tet(O) and tet(Q). Different resistance genes were present in soil samples, where erm(V) and erm(E) were the most prevalent followed by the efflux type Tc(R) genes. The macroarray proved a powerful tool to screen DNA extracted from environmental samples to identify the most abundant Tc(R) and erm(R) genes within those tested, avoiding the need for culturing and biased PCR amplification steps.200717298370
279720.9998Widespread 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
360330.9998Diversity of tet resistance genes in tetracycline-resistant bacteria isolated from a swine lagoon with low antibiotic impact. Tetracycline resistance has been extensively studied and shown to be widespread. A number of previous studies have clearly demonstrated that a variety of tetracycline resistance genes are present in swine fecal material, treatment lagoons, and the environments surrounding concentrated animal feeding operations (CAFOs). The diversity of tetracycline resistance within a swine lagoon located at a CAFO that used only bacitricin methylene disalicylate as an antibiotic was evaluated by screening 85 tetracycline-resistant isolates for the presence of 18 different genes by performing PCR with primers that target tetracycline efflux genes of Gram-negative bacteria and ribosomal protection proteins. In addition, partial 16S rRNA sequences from each of these isolates were sequenced to determine the identity of these isolates. Of the 85 isolates examined, 17 may represent potential novel species based on BLAST results. Greater than 50% of the isolates (48 out of 85) were found to not contain targeted tet efflux genes. Though minimum inhibitory concentrations ranged widely (16 - >256 mg/L), these values did not give an indication of the tet genes present. Ten new genera were identified that contain at least one tet efflux gene. Five other genera possessed tet efflux genes that were not found in these organisms previously. Interestingly, none of the isolates possessed any of the selected ribosomal protection protein genes. Though tetracycline resistance was found in bacteria isolated from a swine CAFO lagoon, it appears that the limited antibiotic use at this CAFO might have impacted the presence and diversity of tetracycline resistance genes.200718059563
281840.9998Tetracycline resistance associated with commensal bacteria from representative ready-to-consume deli and restaurant foods. Proper knowledge of antibiotic resistance (AR) dissemination is essential for effective mitigation. This study examined the profiles of tetracycline-resistant (Tetr) commensal bacteria from representative ready-to-consume food samples from salad bars at local grocery stores and restaurants. Out of 900 Tetr isolates examined, 158 (17.6%) carried one or more of tetM, tetL, tetS, and tetK genes by conventional PCR, 28 harbored more than one Tetr determinants. The most prevalent genotype was tetM, which was detected in 70.9% of the AR gene carriers, followed by tetL (31.6%), tetS (13.9%), and tetK (2.5%). Identified AR gene carriers included Enterococcus, Lactococcus, Staphylococcus, Brochothrix, Carnobacterium, Stenotrophomonas, Pseudomonas, and Sphingobacterium, by 16S rRNA gene sequence analysis. AR determinants were successfully transmitted, and led to resistance in Streptococcus mutans via natural gene transformation and Enterococcus faecalis via electroporation, suggesting the functionality and mobility of the AR genes from the food commensal bacteria. In addition, the AR traits in many isolates are quite stable, even in the absence of the selective pressure. The identification of new commensal carriers for representative AR genes revealed the involvement of a broad spectrum of bacteria in the horizontal transmission of AR genes. Meanwhile, the spectrum of the antibiotic-resistant bacteria differed from the spectrum of the total bacteria (by denaturing gradient gel electrophoresis) associated with the food items. Our data revealed a common avenue in AR exposure and will assist in proper risk assessment and the development of comprehensive mitigation strategies to effectively combat AR.201021067672
282050.9998Direct detection of antibiotic resistance genes in specimens of chicken and pork meat. Antibiotic resistance (AR) in bacteria, a major threat to human health, has emerged in the last few decades as a consequence of the selective pressure exerted by the widespread use of antibiotics in medicine, agriculture and veterinary practice and as growth promoters in animal husbandry. The frequency of 11 genes [tet(M), tet(O), tet(K), erm(A), erm(B), erm(C), vanA, vanB, aac (6')-Ie aph (2'')-Ia, mecA, blaZ] encoding resistance to some antibiotics widely used in clinical practice was analysed in raw pork and chicken meat and in fermented sausages as well as in faecal samples from the relevant farm animals using a molecular approach based on PCR amplification of bacterial DNA directly extracted from specimens. Some of the 11 AR genes were highly prevalent, the largest number being detected in chicken meat and pig faeces. The genes found most frequently in meat were tet(K) and erm(B); vanB and mecA were the least represented. All 11 determinants were detected in faecal samples except mecA, which was found only in chicken faeces. erm(B) and erm(C) were detected in all faecal samples. The frequency of AR genes was not appreciably different in meat compared to faecal specimens of the relevant animal except for vanB, which was more prevalent in faeces. Our findings suggest that AR genes are highly prevalent in food-associated bacteria and that AR contamination is likely related to breeding rather than processing techniques. Finally, the cultivation-independent molecular method used in this work to determine the prevalence of AR genes in foods proved to be a rapid and reliable alternative to traditional tools.200717005283
279460.9998Influence of soil use on prevalence of tetracycline, streptomycin, and erythromycin resistance and associated resistance genes. This study examined differences in antibiotic-resistant soil bacteria and the presence and quantity of resistance genes in soils with a range of management histories. We analyzed four soils from agricultural systems that were amended with manure from animals treated with erythromycin and exposed to streptomycin and/or oxytetracycline, as well as non-manure-amended compost and forest soil. Low concentrations of certain antibiotic resistance genes were detected using multiplex quantitative real-time PCR (qPCR), with tet(B), aad(A), and str(A) each present in only one soil and tet(M) and tet(W) detected in all soils. The most frequently detected resistance genes were tet(B), tet(D), tet(O), tet(T), and tet(W) for tetracycline resistance, str(A), str(B), and aac for streptomycin resistance, and erm(C), erm(V), erm(X), msr(A), ole(B), and vga for erythromycin resistance. Transposon genes specific for Tn916, Tn1549, TnB1230, Tn4451, and Tn5397 were detected in soil bacterial isolates. The MIC ranges of isolated bacteria for tetracycline, streptomycin, and erythromycin were 8 to >256 μg/ml, 6 to >1,024 μg/ml, and 0.094 to >256 μg/ml, respectively. Based on 16S rRNA gene similarity, isolated bacteria showed high sequence identity to genera typical of soil communities. Bacteria with the highest MICs were detected in manure-amended soils or soils from agricultural systems with a history of antibiotic use. Non-manure-amended soils yielded larger proportions of antibiotic-resistant bacteria, but these had lower MICs, carried fewer antibiotic resistance genes, and did not display multidrug resistance (MDR).201222203596
312770.9998Characterization of Bacteria and Antibiotic Resistance in Commercially Produced Cheeses Sold in China. ABSTRACT: The consumption of cheese in the People's Republic of China is increasing rapidly. Little is known about the microbiota, the presence of antibiotic-resistant bacteria, or the distribution of antibiotic resistance genes (ARGs) in commercially produced cheeses sold in China. This information is important for evaluating quality and safety. This study was conducted using 16S rRNA gene sequencing to assess the metagenomics of 15 types of cheese. Fourteen bacterial genera were detected, and Lactococcus, Lactobacillus, and Streptococcus were dominant based on number of sequence reads. Multidrug-resistant lactic acid bacteria (i.e., resistant to two or more types of antibiotic) were isolated from most of the types of cheese. Of these isolates, 100 and 91.7% were resistant to streptomycin and sulfamethoxazole, respectively, and genes involved in acquired resistance to streptomycin (strB) and sulfonamides (sul2) were detected with high frequency. To analyze the distribution of ARGs in the cheeses overall, 309 ARGs from eight categories and nine transposase genes were profiled. A total of 169 ARGs were detected in the 15 cheeses; their occurrence and abundance varied significantly between cheeses. Our study revealed diverse bacteria and ARGs in cheeses sold in China. The risks associated with multidrug resistance among dominant lactic acid bacteria are of great concern.202234855936
358980.9998Prevalence and diversity of tetracycline resistant lactic acid bacteria and their tet genes along the process line of fermented dry sausages. In order to study the prevalence and diversity of tetracycline resistant lactic acid bacteria (Tc(r) LAB) along the process line of two different fermented dry sausage (FDS) types, samples from the raw meat, the meat batter and the fermented end product were analysed quantitatively and qualitatively by using a culture-dependent approach. Both the diversity of the tet genes and their bacterial hosts in the different stages of FDS production were determined. Quantitative analysis showed that all raw meat components of both FDS types (FDS-01 and FDS-08) contained a subpopulation of Tc(r) LAB, and that for FDS-01 no Tc(r) LAB could be recovered from the samples after fermentation. Qualitative analysis of the Tc(r) LAB subpopulation in FDS-08 included identification and typing of Tc(r) LAB isolates by (GTG)5-PCR fingerprinting, plasmid profiling, protein profiling and a characterization of the resistance by PCR detection of tet genes. Two remarks can be made when the results of this analysis for the different samples are compared. (i) The taxonomic diversity of Tc(r) LAB varies along the process line, with a higher diversity in the raw meat (lactococci, lactobacilli, streptococci, and enterococci), and a decrease after fermentation (only lactobacilli). (ii) Also the genetic diversity of the tet genes varies along the process line. Both tet(M) and tet(S) were found in the raw meat, whereas only tet(M) was found after fermentation. A possible relationship was found between the disappearing of species other than lactobacilli and tet(S), because tet(S) was only found in lacotocci, enterococci, and streptococci. These data suggest that fermented dry sausages are among those food products that can serve as vehicles for Tc(r) LAB and that the raw meat already contains a subpopulation of these bacteria. Whether these results reflect the transfer of resistant bacteria or of bacterial resistance genes from animals to man via the food chain is difficult to ascertain and may require a combination of cultivation-dependent and cultivation-independent approaches.200312866855
282190.9998Antibiotic resistant enterococci and staphylococci isolated from flies collected near confined poultry feeding operations. Use of antibiotics as feed additives in poultry production has been linked to the presence of antibiotic resistant bacteria in farm workers, consumer poultry products and the environs of confined poultry operations. There are concerns that these resistant bacteria may be transferred to communities near these operations; however, environmental pathways of exposure are not well documented. We assessed the prevalence of antibiotic resistant enterococci and staphylococci in stored poultry litter and flies collected near broiler chicken houses. Drug resistant enterococci and staphylococci were isolated from flies caught near confined poultry feeding operations in the summer of 2006. Susceptibility testing was conducted on isolates using antibiotics selected on the basis of their importance to human medicine and use in poultry production. Resistant isolates were then screened for genetic determinants of antibiotic resistance. A total of 142 enterococcal isolates and 144 staphylococcal isolates from both fly and poultry litter samples were identified. Resistance genes erm(B), erm(A), msr(C), msr(A/B) and mobile genetic elements associated with the conjugative transposon Tn916, were found in isolates recovered from both poultry litter and flies. Erm(B) was the most common resistance gene in enterococci, while erm(A) was the most common in staphylococci. We report that flies collected near broiler poultry operations may be involved in the spread of drug resistant bacteria from these operations and may increase the potential for human exposure to drug resistant bacteria.200919157515
2798100.9998The Distribution of Eight Antimicrobial Resistance Genes in Streptococcus oralis, Streptococcus sanguinis, and Streptococcus gordonii Strains Isolated from Dental Plaque as Oral Commensals. It has been proposed that oral commensal bacteria are potential reservoirs of a wide variety of antimicrobial resistance genes (ARGs) and could be the source of pathogenic bacteria; however, there is scarce information regarding this. In this study, three common streptococci of the mitis group (S. oralis, S. sanguinis, and S. gordonii) isolated from dental plaque (DP) were screened to identify if they were frequent reservoirs of specific ARGs (blaTEM, cfxA, tetM, tetW, tetQ, ermA, ermB, and ermC). DP samples were collected from 80 adults; one part of the sample was cultured, and from the other part DNA was obtained for first screening of the three streptococci species and the ARGs of interest. Selected samples were plated and colonies were selected for molecular identification. Thirty identified species were screened for the presence of the ARGs. From those selected, all of the S. sanguinis and S. oralis carried at least three, while only 30% of S. gordonii strains carried three or more. The most prevalent were tetM in 73%, and blaTEM and tetW both in 66.6%. On the other hand, ermA and cfxA were not present. Oral streptococci from the mitis group could be considered frequent reservoirs of specifically tetM, blaTEM, and tetW. In contrast, these three species appear not to be reservoirs of ermA and cfxA.202337999618
5921110.9998Prevalence 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
2800120.9998Occurrence 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
2793130.9997Investigating Antibiotic Resistance Genes in Marketed Ready-to-Eat Small Crickets (Acheta domesticus). The present investigation was aimed at evaluating the occurrence of transferable genes conferring resistance to tetracyclines, macrolide-lincosamide-streptogramin B (MLS(B) ), vancomycin, beta-lactams, and aminoglycosides in 32 samples from eight batches of ready-to-eat crickets (Acheta domesticus) commercialized by four European Union producers (two batches per producer). Bacterial DNA extracted directly from the insects was subjected to optimized polymerase chain reaction (PCR) and nested-PCR assays for the qualitative detection of 12 selected antibiotic resistance (AR) genes. Microbial enumeration demonstrated high counts of spore-forming bacteria and total mesophilic aerobes. Statistical analyses revealed significant differences between different producers and insect batches. Regarding AR genes, a high prevalence of genes conferring resistance to tetracycline [tet(M), tet(O), tet(K), tet(S)] was observed, together with the presence of genes conferring resistance to erythromycin [erm(B), erm(C)], beta-lactams (blaZ and mecA), and aminoglycosides [aac(6')-Ie aph(2")-Ia]. We performed a principal component analysis based on the AR gene frequencies that differentiated samples of batch 1 from those of batch 2. This analysis provided evidence for a difference between the producer from France and all the other producers among the batch 1 samples. PRACTICAL APPLICATION: Overall, an intrabatch variation was seen in the transferable resistances among different producers. This evidence, coupled with the observed differences in the viable counts, suggests a low standardization of the production processes. Hence, a prudent use of antimicrobials during the rearing of insects destined for human consumption is strongly recommended, as well as a need for a full standardization of production technologies.201931600843
2828140.9997The distribution of antibiotic resistance genes in chicken gut microbiota commensals. Antibiotic resistance in bacterial pathogens or several indicator bacteria is commonly studied but the extent of antibiotic resistance in bacterial commensals colonising the intestinal tract is essentially unknown. In this study, we aimed to investigate the presence of horizontally acquired antibiotic resistance genes among chicken gut microbiota members in 259 isolates with known whole genomic sequences. Altogether 124 isolates contained at least one gene coding for antibiotic resistance. Genes coding for the resistance to tetracyclines (detected in 101 isolates), macrolide-lincosamide-streptogramin B antibiotics (28 isolates) and aminoglycosides (25 isolates) were the most common. The most frequent tetracycline resistance genes were tet(W), tet(32), tet(O) and tet(Q). Lachnospiraceae and Ruminococcaceae frequently encoded tet(W). Lachnospiraceae commonly coded also for tet(32) and tet(O). The tet(44) gene was associated with Erysipelotrichaceae and tet(Q) was detected in the genomes of Bacteroidaceae and Porphyromonadaceae. Without any bias we have shown that antibiotic resistance is quite common in gut commensals. However, a comparison of codon usage showed that the above-mentioned families represent the most common current reservoirs but probably not the original host of the detected resistances.202133558560
2830150.9997Diverse tetracycline resistant bacteria and resistance genes from coastal waters of Jiaozhou Bay. Environmental microbiology investigation was carried out in Jiaozhou Bay to determine the source and distribution of tetracycline-resistant bacteria and their resistance mechanisms. At least 25 species or the equivalent molecular phylogenetic taxa in 16 genera of resistant bacteria could be identified based on 16S ribosomal deoxyribonucleic acid sequence analysis. Enterobacteriaceae, Pseudomonadaceae, and Vibrionaceae constituted the majority of the typical resistant isolates. Indigenous estuarine and marine Halomonadaceae, Pseudoalteromonadaceae, Rhodobacteraceae, and Shewanellaceae bacteria also harbored tetracycline resistance. All the six resistance determinants screened, tet(A)-(E) and tet(G), could be detected, and the predominant genes were tet(A), tet(B), and tet(G). Both anthropogenic activity-related and indigenous estuarine or coastal bacteria might contribute to the tet gene reservoir, and resistant bacteria and their molecular determinants may serve as bioindicators of coastal environmental quality. Our work probably is the first identification of tet(E) in Proteus, tet(G) in Acinetobacter, tet(C) and tet(D) in Halomonas, tet(D) and tet(G) in Shewanella, and tet(B), tet(C), tet(E), and tet(G) in Roseobacter.200817668262
5905160.9997Characterization of Antibiotic Resistance Genes from Lactobacillus Isolated from Traditional Dairy Products. Lactobacilli are widely used as starter cultures or probiotics in yoghurt, cheese, beer, wine, pickles, preserved food, and silage. They are generally recognized as safe (GRAS). However, recent studies have shown that some lactic acid bacteria (LAB) strains carry antibiotic resistance genes and are resistant to antibiotics. Some of them may even transfer their intrinsic antibiotic resistance genes to other LAB or pathogens via horizontal gene transfer, thus threatening human health. A total of 33 Lactobacillus strains was isolated from fermented milk collected from different areas of China. We analyzed (1) their levels of antibiotic resistance using a standardized dilution method, (2) their antibiotic resistance gene profiles by polymerase chain reaction (PCR) using gene-specific primers, and (3) the transferability of some of the detected resistance markers by a filter mating assay. All Lactobacillus strains were found to be resistant to vancomycin, but susceptible to gentamicin, linezolid, neomycin, erythromycin, and clindamycin. Their susceptibilities to tetracycline, kanamycin, ciprofloxacin, streptomycin, quinupristin/dalfopristin, trimethoprim, ampicillin, rifampicin, and chloramphenicol was different. Results from our PCR analysis revealed 19 vancomycin, 10 ciprofloxacin, and 1 tetracycline-resistant bacteria that carried the van(X), van(E), gyr(A), and tet(M) genes, respectively. Finally, no transferal of the monitored antibiotic resistance genes was observed in the filter mating assay. Taken together, our study generated the antibiotic resistance profiles of some milk-originated lactobacilli isolates and preliminarily assessed their risk of transferring antibiotic gene to other bacteria. The study may provide important data concerning the safe use of LAB.201728182844
5646170.9997Dispersion and persistence of antimicrobial resistance genes among Staphylococcus spp. and Mammaliicoccus spp. isolated along a swine manure treatment plant. Staphylococcus spp. and Mammaliicoccus spp. colonize the skin and mucosa of humans and other animals and are responsible for several opportunistic infections. Staphylococci antibiotic resistance may be present in the environment due to the spread of treated and untreated manure from the livestock industry due to antibiotic use to disease control or growth promoter. In this work, we analyzed the species distribution and antimicrobial susceptibility of Staphylococcus and Mammaliicoccus species along different sites of a swine manure treatment plant from Southeastern Brazil. Bacterial colonies were obtained on mannitol salt agar, selected after catalase test and Gram staining, and finally identified by mass spectrometry and sequencing of the tuf gene. According to the results, S.cohnii and S. simulans were the most prevalent species. Antibiotic resistance test revealed that several strains were resistant to multiple drugs, with high levels of chloramphenicol resistance (98%), followed by erythromycin (79%), tetracycline (73%), gentamicin (46%), ciprofloxacin (42%), cefoxitin (18%), sulfamethoxazole + trimethoprim (12%), and linezolid (4%). In addition, gene detection by PCR showed that all strains carried at least 2 resistance genes and one of them carried all 11 genes investigated. Using the GTG(5)-PCR approach, a high genetic similarity was observed between some strains that were isolated from different points of the treatment plant. Although some were seemingly identical, differences in their resistance phenotype and genotype suggest horizontal gene transfer. The presence of resistant bacteria and resistance genes along the treatment system highlights the potential risk of contamination by people in direct contact with these animals and the soil since the effluent is used as a biofertilizer in the surrounding environment.202336515883
2829180.9997Prevalence of streptomycin-resistance genes in bacterial populations in European habitats. The prevalence of selected streptomycin (Sm)-resistance genes, i.e. aph (3''), aph (6)-1d, aph (6)-1c, ant (3'') and ant (6), was assessed in a range of pristine as well as polluted European habitats. These habitats included bulk and rhizosphere soils, manure from farm animals, activated sludge from wastewater treatment plants and seawater. The methods employed included assessments of the prevalence of the genes in habitat-extracted DNA by PCR, followed by hybridisation with specific probes, Sm-resistant culturable bacteria and exogenous isolation of plasmids carrying Sm-resistance determinants. The direct DNA-based analysis showed that aph (6)-1d genes were most prevalent in the habitats examined. The presence of the other four Sm-modifying genes was demonstrated in 58% of the tested habitats. A small fraction of the bacterial isolates (8%) did not possess any of the selected Sm-modifying genes. These isolates were primarily obtained from activated sludge and manure. The presence of Sm-modifying genes in the isolates often coincided with the presence of IncP plasmids. Exogenous isolation demonstrated the presence of plasmids of 40-200 kb in size harbouring Sm-resistance genes from all the environments tested. Most plasmids were shown to carry the ant (3'') gene, often in combination with other Sm-resistance genes, such as aph (3'') and aph (6)-1d. The most commonly found Sm-modifying gene on mobile genetic elements was ant (3''). Multiple Sm-resistance genes on the same genetic elements appeared to be the rule rather than the exception. It is concluded that Sm-resistance genes are widespread in the environmental habitats studied and often occur on mobile genetic elements and ant (3'') was most often encountered.200219709288
3593190.9997Genes homologous to glycopeptide resistance vanA are widespread in soil microbial communities. The occurrence of d-Ala : d-Lac ligase genes homologous to glycopeptide resistance vanA was studied in samples of agricultural (n=9) and garden (n=3) soil by culture-independent methods. Cloning and sequencing of nested degenerate PCR products obtained from soil DNA revealed the occurrence of d-Ala : d-Ala ligase genes unrelated to vanA. In order to enhance detection of vanA-homologous genes, a third PCR step was added using primers targeting vanA in soil Paenibacillus. Sequencing of 25 clones obtained by this method allowed recovery of 23 novel sequences having 86-100% identity with vanA in enterococci. Such sequences were recovered from all agricultural samples as well as from two garden samples with no history of organic fertilization. The results indicated that soil is a rich and assorted reservoir of genes closely related to those conferring glycopeptide resistance in clinical bacteria.200616734783