Tetracycline Resistance Gene Tet Research Articles

Molecular characterisation of methicillin-resistant Staphylococcus pseudintermedius from dogs and the description of their SCCmec elements

In recent years an increasing number of methicillin-resistant S. pseudintermedius (MRSP) has been observed in both, healthy and clinically infected dogs. The aim of the study was the characterisation of MRSP isolates from clinical routine diagnostics of a German laboratory in order to assess the abundancy of resistance genes and SCCmec elements. 97 isolates from 96 dogs were analysed using microarrays detecting resistance genes and SCCmec-associated markers.All isolates harboured mecA and blaZ. Other abundant resistance markers (in >80% of isolates) included aacA-aphD, aphA3 and sat as well as erm(B). Tetracycline resistance genes (tet(K), tet(M)) and cat also were common (in >20%). The vast majority (n = 59) of isolates carried SCCmec III elements. SCCmec IV and V elements were identified in 21 and 15 isolates, respectively. Irregular or pseudo-SCCmec elements were found in 2 isolates.The high degree of uniformity of hybridisation patterns of tested strains suggest that the majority of MRSP infections was caused by one single strain and comparison to previously published reports and sequences suggest that this was the ST71-SCCmec III strain that also predominates elsewhere in Western Europe.

Multiple-antibiotic resistance of Enterococcus faecalis in an endangered olive ridley sea turtle (Lepidochelys olivacea): A case report

There has recently been an alarming increase in the prevalence of antibiotic resistant bacteria in marine environments. In this study, an Enterococcus sp. was isolated from bacteriologic cultures of the elbow joint of an olive ridley turtle (Lepidochelys olivacea ). The enterococci isolate was identified at the species level as Enterococcus faecalis using PCR and the RAPID ID 32 STREP system. Analysis of the antimicrobial resistance patterns and the antibiotic-resistance genes of the E. faecalis isolated in this study revealed multiple-antibiotic-resistance (MAR). Specifically, the isolate presented resistance to doxycycline, enrofloxacin, erythromycin, neomycin, oxytetracycline, gentamicin, amikacin, ciprofloxacin, spiramycin, chloramphenicol, ceftiofur, and azithromycin. We also identified the tetracycline resistance gene tet(M) in the E. faecalis isolate. Further research into the antimicrobial resistance patterns of Enterococcus spp. should be conducted to promote the conservation of sea turtles.

Effects of Environmental Factors on Tetracycline and Macrolides Resistance Genes in Cattle Manure Composting Systems

The effects of environmental factors and bacterial numbers on typical tetracycline resistance genes (tetQ, tetW, tetM, tetG, tetA, and tetX) and macrolide resistance genes (erm35, erm36, ermB, ermF, ermT, and ermX) during cow manure composting were investigated and quantified via real-time quantitative PCR. The results suggested that the abundance of tetQ, tetW, tetG, and tetX increased by 1.7, 0.3, 84.8, and 4.5 times respectively, whereas tetM decreased by 83.1% during the heating stages of cow manure composting. The abundance of tetG and tetX increased by 23.8 and 11.5 times respectively, whereas tetW, tetQ, and tetM decreased by 90% during the maturity stage. However, the macrolide resistance genes (erm35, erm36, ermB, ermF, ermT, and ermX) increased by 2.1, 430.4, 0.6, 11.5, 2.1, and 49.1 times, respectively. Redundancy analysis showed that ORP, temperature, and VS were crucial factors that could explain the variations of 54.8%, 34%, and 11.3%, respectively, in ARGs. ORP, temperature, and VS also had the greatest positive correlation with ermX, ermB, tetQ, ermF, and tetW, in that order.

Draft genome sequences of KPC-2- and CTX-M-15-producing Klebsiella pneumoniae ST437 isolated from a clinical sample and urban rivers in Sao Paulo, Brazil.

KPC-producing Klebsiella pneumoniae is considered one of the most worrisome multidrug-resistant micro-organisms in nosocomial infections. It has also been reported in wastewater and urban rivers in the city of Sao Paulo, Brazil. Here we report the draft genome sequences of three KPC-2- and CTX-M-15-producing K. pneumoniae sequence type 437 (ST437) isolates obtained from two urban rivers and from a clinical sample of a patient in Sao Paulo. A genomic library was constructed using a Nextera XT Kit. An Illumina platform was used to perform whole-genome sequencing (WGS). WGS of environmental isolates Kp148/PINH-4900 and Kp196/TIET-4200 and clinical isolate Kp314/11 resulted in estimated genome sizes of 5464058, 5437723 and 5319218bp, respectively. Resistome analysis of the environmental and clinical strains revealed the presence of resistance genes to the following antimicrobials in all strains: aminoglycosides [aac(6')-Ib-cr]; β-lactams (blaOXA-1, blaSHV-11, blaCTX-M-15 and blaKPC-2); fluoroquinolones [aac(6')-Ib-cr, oqxA and oqxB]; fosfomycin (fosAKP); macrolides [mph(A)]; phenicols (catB4); sulfonamides (sul1); and trimethoprim (dfrA30). The tetracycline resistance gene tetA was identified in Kp148/PINH-4900 and Kp314/11 only; the aminoglycoside resistance gene aph(3')-Ia was found only in environmental isolates, and aadA2 only in Kp314/11; and the phenicol resistance gene catA1 was identified only in Kp148/PINH-4900. The draft genome sequences of these strains help us to elucidate the dissemination of resistance genes in micro-organisms inside and outside the hospital and are useful for further comparisons between clinical and environmental strains.

Phenotypic and genotypic drug resistance profile of Salmonella serovars isolated from poultry farm and processing units located in and around Mumbai city, India.

Background and Aim:The extensive use of antimicrobials in poultry has led to an increase in bacterial multidrug resistance, and the emergence of multidrug-resistant nontyphoidal Salmonella is a global problem. This study was performed to detect antibiotic-resistant Salmonella serovars in poultry farming and processing environment.Materials and Methods:A total of 956 various samples, comprising 432 farm origin, 324 poultry processing stage wise and environmental, and 154 product processing stages and environmental samples, were collected from poultry farms and processing units located in and around Mumbai city. Of a total of 71 recovered isolates, 42 randomly selected Salmonella isolates were subjected for antibiotic susceptibility testing by disk diffusion method and serotyping. A total of 31 serotypically confirmed isolates were characterized for the presence of tetA, tetB, blaTEM, and CTX-M gene.Results:Higher resistance was recorded against Doxycycline (100%), followed by Oxytetracycline (97.62%), Neomycin (88.10%), Erythromycin (83.33%), Tetracycline (78.57%), and Ceftizoxime (35.71%). Resistance from 0.00 to 26.19 percent was found to antimicrobials, namely Norfloxacin (26.19%), Ampicillin (21.43%), Azithromycin (21.43%), Ciprofloxacin (19.05%), Colistin (4.76%), Streptomycin (16.67%), Cefotaxime (14.19%), Enrofloxacin (14.29%), Amoxyclav (14.29%), Gentamicin (7.14%), Chloramphenicol (4.76%), Amikacin (4.76%), and Ceftazidime (0.0%). Results demonstrate that the Salmonella Virchow dominated and all serotypes were found to carry Tetracycline resistance gene tetA, 5 isolates were found to be positive for blaTEM, whereas none of the isolates were carrying tetB and CTX-M gene.Conclusion:This study revealed that there is a significant rise of Tetracycline resistance with the presence of tetA gene in Salmonella spp. which indicates selective pressure for adopting resistance against tetracycline group of antibiotics.

Characterization of Erythromycin and Tetracycline Resistance in Lactobacillus fermentum Strains.

Lactobacillus fermentum colonizing gastrointestinal and urogenital tracts of humans and animals is widely used in manufacturing of fermented products and as probiotics. These bacteria may function as vehicles of antibiotic resistance genes, which can be transferred to pathogenic bacteria. Therefore, monitoring and control of transmissible antibiotic resistance determinants in these microorganisms is necessary to approve their safety status. The aim of this study was to characterize erythromycin and tetracycline resistance of L. fermentum isolates and to estimate the potential transfer of resistance genes from lactobacilli to the other Gram-positive and Gram-negative bacteria. Among six L. fermentum strains isolated from human feces and commercial dairy products, five strains demonstrated phenotypic resistance to tetracycline. PCR screening for antibiotic resistance determinants revealed plasmid-located tetracycline resistance genes tet(K) and tet(M) in all strains and erythromycin resistance genes erm(B) in the chromosome of L. fermentum 5-1 and erm(C) in the plasmid of L. fermentum 3-4. All tested lactobacilli lacked conjugative transposon Tn916 and were not able to transfer tetracycline resistance genes to Staphylococcus aureus, Staphylococcus epidermidis, Listeria monocytogenes, Acinetobacter baumannii, Citrobacter freundii, and Escherichia coli by filter mating. Staphylococcus haemolyticus did not accept erythromycin resistance genes from corresponding Lactobacillus strains. Thus, in the present study, L. fermentum was not implicated in the spread of erythromycin and tetracycline resistance, but still these strains pose the threat to the environment and human health because they harbored erythromycin and tetracycline resistance genes in their plasmids and therefore should not be used in foods and probiotics.

High removal efficiency of antibiotic resistance genes in swine wastewater via nanofiltration and reverse osmosis processes

Swine wastewater treatment plant has become one of the main sources of antibiotic resistance genes (ARGs). Membrane treatment processes are promising solutions for removal of the emerging contaminants. However, limited studies have investigated the effects of nanofiltration and reverse osmosis treatment in removing ARGs in swine wastewater. In this study, the presence and the fate of common ARGs including sul1, sul2, tetA, tetM and tetW, as well as intI1 and 16S rRNA gene, were investigated in a medium-sized (6500) pig farm wastewater treatment plant (WWTP) equipped with conventional biological treatment and advanced membrane processing system. All of the genes were detected with highly abundance in the raw sewage. The biological treatments of the swine wastewater treatment plant did not reduce the quantity of the ARGs. As expected, nanofiltration and reverse osmosis treatment reduced the absolute gene copy number of ARGs efficiently (4.98–9.52 logs removal compared to raw sewage). Compared to the reverse osmosis effluent, however, the absolute abundance of ARGs in the artificial wetland increased by 1.00–2.06 logs. Meanwhile, the relative abundance of sulfonamide resistant genes were basically unchanged, while tetracycline resistance genes (tetA, tetM and tetW) decreased by 0.88, 3.47, 2.51 log, respectively. The results demonstrated that advanced membrane treatments are capable of removing various kinds of ARGs efficiently, as well as some common nitrogen and phosphorus contaminants. This study suggested a mature alternative method for the removal of ARGs from livestock wastewater.

Distribution and relationship between antimicrobial resistance genes and heavy metals in surface sediments of Taihu Lake, China.

Heavy metals, pharmaceuticals, and other wastes released into the environment can significantly influence environmental antibiotic resistance. We investigated the occurrence of 22 antimicrobial resistance genes (ARGs) and 10 heavy metal concentrations, and the relationship between ARGs and heavy metals in surface sediment from seven sites of Lake Taihu. The results showed significant correlations (p < 0.05) between sediment ARG levels, especially for tetracycline and sulfonamides (e.g., tet(A), tet(D), tet(E), tet(O), sul1, sul2 and int-1) and specific heavy metals (Fe, Mn, Cr, Cu, Zn, among others) in the Lake. In the surface sediments, heavy metals showed an interaction with resistance genes, but the strength of interaction was diminished with increasing depth. For most of the heavy metals, the concentration of elements in the top sediments was higher than that in other depths. Tetracycline resistance genes (tet(A), tet(B), tet(D), tet(E) and tet(O), β-lactam resistance genes (SHV, TEM, CTX, OXA and OXY) and sulfonamide resistance genes (sulA, sul1, sul2, sul3 and int-1) were detected. They showed a trend which inferred a statistically significant increase followed by decreases in the relative abundance of these ARGs (normalized to 16S rRNA genes) with increasing depth. This study revealed that tet(A), tet(O), TEM, OXY, int-1, sul1 and sul3 were widespread in surface sediments with high abundance, indicating that these genes deserve more attention in future work.

Classification and phylogenetic analysis of the genus Mobiluncus based on matrix-assisted laser de-sorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) and whole genome sequen-cing

Objective To identify Mobiluncus species with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) and 16S rRNA gene sequencing and to analyze antibiotic resistance genes and phylogenetic relationships among Mobiluncus species with whole genome sequencing. Methods Twenty-five Mobiluncus strains were isolated after anaerobic culture of 65 vaginal secretion samples of patients with bacterial vaginosis (BV) and identified to species level by MALDI-TOF MS and 16S rRNA gene sequencing. The whole genome DNA of each strain was extracted for next-generation sequencing. SPAdes was used to assembly genomes. Resistance genes were searched in ARGD database. The core genome of all isolates was analyzed by Harvest to construct phylogenetic tree. Results MALDI-TOF MS could only identify Mobiluncus curtisii, while 16S rRNA gene sequencing could identify both Mobiluncus mulieris and Mobiluncus curtisii. Results of the whole genome sequencing showed that tetracycline resistance gene tet(o) and macrolides resistance gene erm(x) were the two predominant acquired resistance genes of Mobiluncus with a positive rate of 84.7% and 61.5% respectively. Intra-species relationships of the two Mobiluncus species were close, but a distant phylogenetic relationship was found between the two species. Conclusion This study shows that MALDI-TOF MS can′t be used to identify Mobiluncus mulieris at present. Mobiluncus strains have potential resistance to tetracycline and macrolides. Intra-species evolution of Mobiluncus is slow, which indicates that there is no growing trend towards new species for the time being. Key words: Mobiluncus; Bacterial vaginosis (BV); MALDI-TOF MS; Whole genome sequencing; Resistance gene

Antimicrobial Resistance Profiles and Tetracycline Resistance Genes of Escherichia coli in Mediterranean Mussel and Sea Snails Collected from Black Sea, Turkey

Aquatic environments are often affected by and exposed to anthropogenic pollutants including antimicrobials used as disease prevention and feed additives. Antimicrobial resistance is a major problem both in animal and in human health worldwide. In this study, Mediterranean mussel (Mytilus galloprovincialis) and Sea snail (Rapana venosa) samples were collected seasonally from the coastline of Black Sea. A total of 54 Escherichia coli were isolated from Mediterranean mussel and Sea snail collected from the coast of Artvin, Rize, Trabzon and Giresun, Turkey. Antimicrobial resistance and the presence of tetracycline (tet) resistance genes (tetA, tetB, tetC, tetD, and tetE) in E. coli isolates were investigated. Antimicrobial susceptibility test determined that 83.3 % of the isolates exhibited resistance to sulfamethoxazole. Resistance to ampicillin and aztreonam was as 66.7 % and 37.0 % among the tested antimicrobials, respectively. The lowest resistant antimicrobial was florfenicol (1.9 %). tetC resistance gene was detected in more than 50% of the isolates. Among the tet resistance genes, tetC was found in the most common gene followed by tetB, tetA, tetE, and tetD. At least one tet gene was detected in 88% of the isolates, and 46% of the isolates had two or more tet genes. The presence of tet resistance genes in E. coli in aquatic environments indicates that these isolates may be a reservoir of tet resistance genes. They may also exhibit an important role in the spread of genes among the pathogenic and non-pathogenic bacteria.

Persistence of bacterial pathogens, antibiotic resistance genes, and enterococci in tidal creek tributaries

Intertidal creeks form the primary hydrologic link between estuaries and land-based activities on barrier islands. Fecal indicators Enterococcus spp. (Entero1), pathogens Shigella spp. (ipaH), Salmonella spp. (invA), E. coli of EHEC/EPEC groups (eaeA), E. coli of EAEC, EIEC, and UPEC groups (set1B), E. coli of STEC group (stx1); and tetracycline resistance genes (tet(B), tet(C), tet(D), tet(E), tet(K), tet(Q), tet(W), and tet(X); TRG) were detected in the headwater of Oakdale Creek (Sapelo Island, GA) receiving runoffs from Hog Hammock village. Excavation of drainage ditches around the village caused a high increase in the incidence of the above determinants. Water samples were collected from the headwater, transferred to diffusion chambers, submersed in the headwater, saltmarsh, and mouth of the creek; and the determinants were monitored for 3 winter months. With some exceptions, their persistence decreased in order headwater > saltmarsh > mouth. Genes associated with Enterococcus spp. were the most persistent at all the sites, following in the headwater with determinants for Salmonella spp. and E. coli of EAEC, EIEC, and UPEC groups. In the mouth, the most persistent gene was eaeA indicating EHEC, EPEC, and STEC. Tet(B) and tet(C) persisted the longest in headwater and saltmarsh. No TRG persisted after 11 days in the mouth. Most determinants revealed correlations with temperature and pH, and inverse correlations with dissolved oxygen. Decay rates of the above determinants varied in the range of −0.02 to −0.81/day, and were up to 40 folds higher in the saltmarsh and mouth than in the headwater. Our data demonstrated that water parameters could to some extent predict a general trend in the fate of virulence and antibiotic resistance determinants in tidal creek tributaries but strongly suggested that their persistence in these tributaries cannot be predicted from that of enterococci, or extrapolated from one biological contaminant to another.

Long-term impact of a tetracycline concentration gradient on the bacterial resistance in anaerobic-aerobic sequential bioreactors

Wastewater treatment systems are considered as hotspots for release of antibiotic resistance genes (ARGs) into the environment. Anaerobic-aerobic sequential (AAS) bioreactors now are intensively used for wastewater treatment worldwide. However, the occurrence of ARGs in wastewater treatment systems exposed to low-level (i.e., sub-inhibitory) antibiotic is poorly known. Here, we studied the distribution patterns of seven tetracycline resistance genes (tet genes) including tet(A), tet(C), tet(G), tet(X), tet(M), tet(O), and tet(W), as well as one mobile element [class 1 integron (intI1)] in AAS bioreactors under exposure to tetracycline from 50 μg/L to 500 μg/L. Additionally, effect on the removal performance of nutrients and tetracycline in both anaerobic and aerobic units was also investigated. A tetracycline concentration gradient selected for bacterial resistance in the anaerobic reactor, with the exception of tet(A) and tet(W), and the tetracycline removal deteriorated by 47%. However, the abundance of tet and intI1 genes reduced in the subsequent aerobic unit, and the removal of tetracycline, soluble COD, and NH4+-N maintained at average efficiencies of 91%, 90%, and 93%, respectively. The level of tet(X) was largely unaffected by AAS treatment. It is notable that intI1 genes probably played a crucial role on the horizontal dissemination of tet genes. The tetracycline levels and intI1 genes appear to be the primary factors influencing the occurrence of tet genes in AAS bioreactors. Nonetheless, AAS treatments still show promise for reducing antibiotics, ARGs and mobile elements without affecting nutrient removal, and need further research for practical applications.

Tetracycline resistance gene tet(M) of a marine bacterial strain is not accumulated in bivalves from seawater in clam tank experiment and mussel monitoring

Antibiotic resistance genes (ARGs) are found in marine as well as terrestrial bacteria. Bivalves are known to accumulate chemical pollutants and pathogenic microbes, however, the fate of ARGs in bivalves after the intake of ARG-possessing bacteria is not known. Here we show that the copy number of oxytetracycline resistance gene tet(M) increased rapidly in the clam digestive tract by filtering water, then remained constant over 96h in a tank experiment even with the addition of tet(M)-possessing bacteria every 24h. >99.9% of the added tet(M) was decomposed, reaching a balanced state. Environmental sampling of mussel digestive tract and seawater supported the hypothesis that tet(M) was decomposed in bivalves as tet(M) was present in seawater from April to October at a concentration of 10−5 to 10−6 copies/16S, whereas tet(M) in mussels was mostly below the detection limit. Two (April) and three (July and October) individual mussels were positive for tet(M) with a concentration equivalent to that of seawater. We therefore conclude that bivalves do not accumulate tet(M) from seawater.

Antimicrobial susceptibility and genetic relatedness of respiratory tract pathogens in weaner pigs over a 12-month period

The collaboration project VASIB aims at reducing the antibiotic consumption in pig production by integrating information from consulting expertise in clinical inspection, hygiene, epidemiology, microbiology and pharmacology. In this VASIB subproject, we investigated the antimicrobial susceptibility and relatedness of porcine respiratory tract pathogens. Bordetella bronchiseptica (n = 47), Pasteurella multocida (n = 18) and Streptococcus suis (n = 58) were obtained from weaner pigs at two farms. Antimicrobial susceptibility testing was performed by broth microdilution according to CLSI standards. Resistance genes were detected via specific PCR assays. Macrorestriction analysis was conducted to determine the relatedness of the isolates and to identify clones. The B. bronchiseptica isolates showed indistinguishable (farm 1) or two closely related XbaI-patterns (farm 2). Different SmaI-PFGE patterns of P. multocida isolates were obtained at three different time points. In contrast, PFGE analysis of S. suis indicated more than one fragment pattern per pig and time point. Isolates exhibiting indistinguishable PFGE patterns were considered to represent the same clone. This study showed that only two closely related B. bronchiseptica clones were present in both farms, which had low MICs to all antimicrobials, except to β-lactams. Different P. multocida clones were present at the three time points. They showed overall low MIC values, with two clones being resistant and one intermediate to tetracycline. S. suis clones were resistant to tetracycline (n = 19) and/or erythromycin/clindamycin (n = 16). They harboured the tetracycline resistance genes tet(O), tet(M) or tet(L) and/or the macrolide/lincosamide/streptogramin B resistance gene erm(B). Five penicillin-resistant S. suis clones were also detected.

Tetracyclines, sulfonamides and quinolones and their corresponding resistance genes in the Three Gorges Reservoir, China

The Three Gorges Project significantly impacted water quality and ecological balance in this area. The special engineered aquatic environment could be an important reservoir for antibiotic resistance genes (ARGs). Fifteen ARGs corresponding to three groups of antibiotics (tetracyclines, sulfonamides and quinolones) were determined in surface water, soil and sediment in this study. Total concentrations of antibiotics ranged from 21.55 to 536.86ng/L, 3.69 to 438.76ng/g, 15.78 to 213.84ng/g in water, soil and sediment, respectively. Polymerase chain reaction (PCR) of ARGs revealed the presence of two sulfonamide resistance genes (sul1, sul2), five tetracycline resistance genes (tetA, tetB, tetM, tetQ, tetG) and class 1 integron gene (intI1) in all samples. And the relative abundance of sulfonamide resistance genes was generally higher than tetracycline resistance genes in three matrices. Significant correlations (p<0.05) were found between the concentrations of intI1 and ARGs (tetA, tetB, tetM, tetQ, tetG, sul1, sul2), indicating intI1 may facilitate the proliferation and propagation of these genes. Redundancy analysis (RDA) showed distribution of ARGs was related to the certain antibiotics residues, which may exert selective pressure on bacteria and thus enrich the abundance of ARGs. The results of this study could provide useful information for both better understanding and management of the contamination caused by ARGs and related antibiotics in engineered aquatic environments.

Characteristics and drug resistance of non-O157 Shiga toxin-producing E. coli in animal feces, from Shandong Province

Objective: To understand the infection status, characteristics and drug resistance of non-O157 Shiga toxin-producing E. coli (STEC) in animal feces in Shandong Province. Methods: From 2015 to 2016, convient sampling method was used to collect 1 022 fresh feces of animals in Weishan county and Laizhou city, and 24 non-O157 STEC were isolated. The serotypes of non-O157 STEC strains were confirmed through serum agglutination test. The susceptibility was explored through the antimicrobial sensitivity experiments. ESBLs activity was confirmed by double-disc diffusion. PCR method was used to detect the resistance genes. PFGE typing was operated to assess the relatedness and variability of the strains. The multi-locus sequence typing (MLST) was adopted to get the allelic profile and ST sequence of strains. Analysis was made on the evolutionary relationship between different ST groups was made through CLC Sequence Viewer and Counting Express. Results: A total of 24 non-O157 STEC were isolated from animal feces. 23 strains were from pig feces, and 1 strain was from cow feces, and the serotypes were more dispersed. All of the 24 strains carried stx2 genes. The highest resistance rate was sulfamethoxazole(22 strains), the mount of cotrimoxazole and nalidixic acid was 18 strains, chloramphenicol was 13 strains, tetracycline was 19, and there was a phenomenon of multiple drug resistance. The drug resistance spectrum was sulfamethoxazole tetracycline-compound novammin-naphthidine-chloramphenicol. All strains were sensitive to cefepime and imipenem. The ESBLs confirmatory test showed that 4 strains of non O157 STEC produced beta lactamase. PCR detected 7 resistance genes, and 4 tetracycline resistance genes (Tet A, Tet B, tetC and tetD) were detected. The beta lactamase resistance genes (blaSHV-1, bla CTX-M, bla TEM) were all negative. 24 strains were divided into 15 PFGE types, and their clustering results were more dispersed and no dominant PFGE type. There were 11 kinds of MLST types, most of them are ST540 and ST5133 types, each of which was 4 strains, and clustered into 1 MLST genomes. Conclusion: The serotypes of non-O157 STEC in animal feces O157 STEC were dispersed, and the resistant rate to common antibiotic was high. MLST typing results presents obvious polymorphism. Surveillance and manage ment of these strains should be strengthened.

Antibiotic-mediated changes in the fecal microbiome of broiler chickens define the incidence of antibiotic resistance genes

BackgroundAntimicrobial agents have been widely used in animal farms to prevent and treat animal diseases and to promote growth. Antimicrobial agents may change the bacterial community and enhance the resistome in animal feces. We used metagenome-wide analysis to investigate the changes in bacterial community, variations in antibiotic resistance genes (ARGs), and their bacterial hosts in the feces of broiler chickens over a full-treatment course of chlortetracycline at low and therapeutic dose levels.ResultsThe effects of chlortetracycline on resistome were dependent on the specific ARG subtypes and not simply the overall community-level ARGs. Therapeutic dose of chlortetracycline promoted the abundance of tetracycline resistance genes (tetA and tetW) and inhibited multidrug resistance genes (mdtA, mdtC, mdtK, ompR, and TolC). The therapeutic dose of chlortetracycline led to loss of Proteobacteria mainly due to the decrease of Escherichia/Shigella (from 72 to 58%). Inhibition of Escherichia by chlortetracycline was the primary reason for the decrease of genes resistant to multiple drugs in the therapeutic dose group. The ARG host Bifidobacterium were enriched due to tetW harbored by Bifidobacterium under chlortetracycline treatment. Escherichia was always the major host for multidrug resistance genes, whereas the primary host was changed from Escherichia to Klebsiella for aminoglycoside resistance genes with the treatment of therapeutic dose of chlortetracycline.ConclusionsWe provided the first metagenomic insights into antibiotic-mediated alteration of ARG-harboring bacterial hosts at community-wide level in chicken feces. These results indicated that the changes in the structure of antibiotic-induced feces microbial communities accompany changes in the abundance of bacterial hosts carrying specific ARGs in the feces microbiota. These findings will help to optimize therapeutic schemes for the effective treatment of antibiotic resistant pathogens in poultry farms.Graphical abstractResistome variations in faecal microbiome of chickens exposed to chlortetracycline

Seasonal dynamics of tetracycline resistance gene transport in the Sumas River agricultural watershed of British Columbia, Canada

Environmental transport of contaminants that can influence the development of antibiotic resistance in bacteria is an important concern in the management of ecological and human health risks. Agricultural regions are locales where practices linked to food crop and livestock production can introduce contaminants that could alter the selective pressures for the development of antibiotic resistance in microbiota. This is important in regions where the use of animal manure or municipal biosolids as waste and/or fertilizer could influence selection for antibiotic resistance in pathogenic bacterial species. To investigate the environmental transport of contaminants that could lead to the development of antibiotic resistance in bacteria, a watershed with one of the highest levels of intensity of agricultural activity in Canada was studied; the Sumas River located 60 km east of Vancouver, British Columbia. This two-year assessment monitored four selected tetracycline resistance genes (tet(O), tet(M), tet(Q), tet(W)) and water quality parameters (temperature, specific conductivity, turbidity, suspended solids, nitrate, phosphate and chloride) at eight locations across the watershed. The tetracycline resistance genes (Tcr) abundances in the Sumas River network ranged between 1.47 × 102 and 3.49 × 104 copies/mL and ranged between 2.3 and 6.9 copies/mL in a control stream (located far from agricultural activities) for the duration of the study. Further, Tcr abundances that were detected in the wet season months ranged between 1.3 × 103 and 2.29 × 104 copies/mL compared with dry season months (ranging between 0.6 and 31.2 copies/mL). Highest transport rates between 1.67 × 1011 and 1.16 × 1012 copies/s were observed in November 2005 during periods of high rainfall. The study showed that elevated concentrations of antibiotic resistance genes in the order of 102–104 copies/mL can move through stream networks in an agricultural watershed but seasonal variations strongly influenced specific transport patterns of these genes.

Prevalence and Molecular Characterization of Antimicrobial Resistance in Escherichia coli Isolated from Raw Milk and Raw Milk Cheese in Egypt

The goal of this study was to examine antimicrobial resistance and characterize the implicated genes in 222 isolates of Escherichia coli from 187 samples of raw milk and the two most popular cheeses in Egypt. E. coli isolates were tested for susceptibility to 12 antimicrobials by a disk diffusion method. Among the 222 E. coli isolates, 66 (29.7%) were resistant to one or more antimicrobials, and half of these resistant isolates showed a multidrug resistance phenotype (resistance to at least three different drug classes). The resistance traits were observed to tetracycline (27.5%), ampicillin (18.9%), streptomycin (18.5%), sulfamethoxazole-trimethoprim (11.3%), cefotaxime (4.5%), kanamycin (4.1%), ceftazidime (3.6%), chloramphenicol (2.3%), nalidixic acid (1.8%), and ciprofloxacin (1.4%). No resistance to fosfomycin and imipenem was observed. Tetracycline resistance genes tetA, tetB, and tetD were detected in 53 isolates, 9 isolates, and 1 isolate, respectively, but tetC was not detected. Aminoglycoside resistance genes strA, strB, aadA, and aphA1 were detected in 41, 41, 11, and 9 isolates, respectively. Sulfonamide resistance genes sul1, sul2, and sul3 were detected in 7, 25, and 3 isolates, respectively. Of 42 ampicillin-resistant isolates, blaTEM, blaCTX-M, and blaSHV were detected in 40, 9, and 3 isolates, respectively, and 10 (23.8%) ampicillin-resistant isolates were found to produce extended-spectrum β-lactamase. Each bla gene of extended-spectrum β-lactamase-producing E. coli was further subtyped to be blaCTX-M-15, blaCTX-M-104, blaTEM-1, and blaSHV-12. The class 1 integron was also detected in 28 resistant isolates, and three different patterns were obtained by PCR-restriction fragment length polymorphism. Sequencing analysis of the variable region revealed that four isolates had dfrA12/orfF/aadA2, two had aadA22, and one had dfrA1/aadA1. These data suggest that antimicrobial-resistant E. coli are widely distributed in the milk production and processing environment in Egypt and may play a role in dissemination of antimicrobial resistance to other pathogenic and commensal bacteria.

Characteristics of ARG-carrying plasmidome in the cultivable microbial community from wastewater treatment system under high oxytetracycline concentration.

Studies on antibiotic production wastewater have shown that even a single antibiotic can select for multidrug resistant bacteria in aquatic environments. It is speculated that plasmids are an important mechanism of multidrug resistance (MDR) under high concentrations of antibiotics. Herein, two metagenomic libraries were constructed with plasmid DNA extracted from cultivable microbial communities in a biological wastewater treatment reactor supplemented with 0 (CONTROL) or 25mg/L of oxytetracycline (OTC-25). The OTC-25 plasmidome reads were assigned to 72 antibiotic resistance genes (ARGs) conferring resistance to 13 types of antibiotics. Dominant ARGs, encoding resistance to tetracycline, aminoglycoside, sulfonamide, and multidrug resistance genes, were enriched in the plasmidome under 25mg/L of oxytetracycline. Furthermore, 17 contiguous multiple-ARG carrying contigs (carrying ≥ 2 ARGs) were discovered in the OTC-25 plasmidome, whereas only nine were found in the CONTROL. Mapping of the OTC-25 plasmidome reads to completely sequenced plasmids revealed that the conjugative IncU resistance plasmid pFBAOT6 of Aeromonas caviae, carrying multidrug resistance transporter (pecM), tetracycline resistance genes (tetA, tetR), and transposase genes, might be a potential prevalent resistant plasmid in the OTC-25 plasmidome. Additionally, two novel resistant plasmids (containing contig C301682 carrying multidrug resistant operon mexCD-oprJ and contig C301632 carrying the tet36 and transposases genes) might also be potential prevalent resistant plasmids in the OTC-25 plasmidome. This study will be helpful to better understand the role of plasmids in the development of MDR in water environments under high antibiotic concentrations.