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This study aimed to characterize the alteration of the fecal microbiome and antimicrobial resistance (AMR) determinants in 24 piglets at day 3 pre-weaning (D. − 3), weaning day (D.0), days 3 (D.3) and 8 post-weaning (D.8), using whole-genome shotgun sequencing. Distinct clusters of microbiomes and AMR determinants were observed at D.8 when Prevotella (20.9%) was the major genus, whereas at D. − 3–D.3, Alistipes (6.9–12.7%) and Bacteroides (5.2–8.5%) were the major genera. Lactobacillus and Escherichia were notably observed at D. − 3 (1.2%) and D. − 3–D.3 (0.2–0.4%), respectively. For AMR, a distinct cluster of AMR determinants was observed at D.8, mainly conferring resistance to macrolide–lincosamide–streptogramin (mefA), β-lactam (cfxA6 and aci1) and phenicol (rlmN). In contrast, at D. − 3–D.3, a high abundance of determinants with aminoglycoside (AMG) (sat, aac(6')-aph(2''), aadA and acrF), β-lactam (fus-1, cepA and mrdA), multidrug resistance (MDR) (gadW, mdtE, emrA, evgS, tolC and mdtB), phenicol (catB4 and cmlA4), and sulfonamide patterns (sul3) was observed. Canonical correlation analysis (CCA) plot associated Escherichia coli with aac(6')-aph(2''), emrA, mdtB, catB4 and cmlA4 at D. − 3, D.0 and/or D.3 whereas at D.8 associations between Prevotella and mefA, cfxA6 and aci1 were identified. The weaning age and diet factor played an important role in the microbial community composition.

This study aimed to investigate antimicrobial resistance (AMR) characteristics of Escherichia coli isolates from pig origin (including pigs, pig carcass, and pork) and humans in Thailand and Lao Pe...

Whole-genome sequencing (WGS) was conducted to characterize

This study aimed to determine the epidemiology and association of antimicrobial resistance (AMR) among

Introduction. Antimicrobial resistance (AMR) is a One Health issue concerning humans, animals and the environment and a unified One Health approach is required to contain this problematic issue. Dogs and cats are popular pet animals and are known to carry many bacterial pathogens that are of public health importance, including Salmonella . However, data on AMR in companion animals is limited. Gap statement. Scant AMR data from bacteria originating from companion animals limits an accurate assessment of the impacts of pet-animal-related AMR on public health. Purpose. This study aimed to phenotypically and genetically investigate AMR in Salmonella isolated from pet dogs and cats in Thailand. Methodology. Salmonella enterica were isolated from pet dogs (n=159) and cats (n=19) in Thailand between 2016 and 2019. All isolates were serotyped. Phenotypic and genotypic antimicrobial resistance was examined. PCR-based replicon typing, replicon sequence typing and plasmid multilocus sequence typing were conducted to characterize plasmids. Results. Seventy-seven serovars were identified, with serovars Weltevreden (9.6%) and Stockholm (9.0%) the most common. Most of the isolates (34.3%) were multidrug-resistant. The serovar Stockholm was an ESBL-producer and carried the β-lactamase genes bla TEM-1 and bla CTX-M-55. The plasmid-mediated quinolone resistance (PMQR) gene, qnrS, was also detected (10.1%). Class 1 integrons carrying the dfrA12-aadA2 cassette array were most frequent (45.9%). Five plasmid replicon types as IncA/C (0.6%), N (1.1%), IncFIIA (28.7%), IncHI1 (2.2%), and IncI1 (3.4%) were identified. Based on the pMLST typing scheme (n=9), plasmids were assigned into five different STs including IncA/C-ST6 (n=1), IncH1-ST16 (n=4), IncI1-ST3 (n=1), IncI1-ST60 (n=1) and IncI1-ST136 (n=1). The ST 16 of IncHI1 plasmid was a novel plasmid ST. Subtyping F-type plasmids using the RST scheme (n=9) revealed four different combinations of replicons including S1:A-:B- (n=4), S1:A-:B22 (n=2), S3:A-:B- (n=1) and S-:A-:B47 (n=1). Conclusions. Our findings highlight the role of clinically healthy household dogs and cats as carriers of AMR Salmonella strains with different R plasmid. The implementation of AMR phenotypes instigation and genotypic monitoring and surveillance programmes in companion animals are imperative as integral components of the One Health framework.

Produce-associated food-borne outbreaks have been increasingly implicated as the significant proportion of the annual incidence of food-borne illness worldwide. The objectives of this study were to determine the concentrations of indicator bacteria and the presence of Salmonella spp., Shigella spp., Escherichia coli (E. coli) O157:H7, and Listeria monocytogenes (L. monocytogenes), and to characterize predictors associated with Salmonella contamination of retail produce from fresh markets and supermarkets in Bangkok, Thailand. A total of 503 samples were collected during May 2018 and February 2019, comprised of sweet basil, spring onion, coriander, cabbage, lettuce, cucumber, and tomato, with conventional items from fresh open-air markets (n=167), conventional items from supermarkets (n=168), and organic items from supermarkets (n=168). The overall prevalence in these 503 items for fecal coliforms and E. coli was 84.3% and 71.4%, with mean concentrations (±standard deviation) of fecal coliforms and E. coli being (3.0×105±1.3×106) most probable number (MPN)/g and (1.8×105±1.1×105) MPN/g, respectively. The concentrations of fecal coliforms and E. coli were higher in produce sampled from fresh open-air markets than produce from supermarkets; similarly, these bacterial indicators were higher from produce grown under conventional methods than certified organic produce. The prevalence of Salmonella and Shigella was 4.8% and 0.4%, respectively, but no positives were found for E. coli O157:H7 and L. monocytogenes. The predominant Salmonella serovar was Stanley (30.8%). Based on logistic regression, the odds of Salmonella contamination were significantly (P

Background Colistin and carbapenem-resistant bacteria have emerged and become a serious public health concern, but their epidemiological data is still limited. Objectives This study examined colistin and carbapenem resistance in Escherichia coli and Salmonella from pigs, pig carcasses, and pork in Thailand, Lao PDR, and Cambodia border provinces. Methods The phenotypic and genotypic resistance to colistin and meropenem was determined in E. coli and Salmonella obtained from pigs, pig carcasses, and pork (n = 1,619). A conjugative experiment was performed in all isolates carrying the mcr gene (s) (n = 68). The plasmid replicon type was determined in the isolates carrying a conjugative plasmid with mcr by PCR-based replicon typing (n = 7). The genetic relatedness of mcr-positive Salmonella (n = 11) was investigated by multi-locus sequence typing. Results Colistin resistance was more common in E. coli (8%) than Salmonella (1%). The highest resistance rate was found in E. coli (17.8%) and Salmonella (1.7%) from Cambodia. Colistin-resistance genes, mcr-1, mcr-3, and mcr-5, were identified, of which mcr-1 and mcr-3 were predominant in E. coli (5.8%) and Salmonella (1.7%), respectively. The mcr-5 gene was observed in E. coli from pork in Cambodia. Two colistin-susceptible pig isolates from Thailand carried both mcr-1 and mcr-3. Seven E. coli and Salmonella isolates contained mcr-1 or mcr-3 associated with the IncF and IncI plasmids. The mcr-positive Salmonella from Thailand and Cambodia were categorized into two clusters with 94%–97% similarity. None of these clusters was meropenem resistant. Conclusions Colistin-resistant E. coli and Salmonella were distributed in pigs, pig carcasses, and pork in the border areas. Undivided-One Health collaboration is needed to address the issue.

This study aimed to assess the cecal microbiome, antimicrobial resistance (AMR), and heavy-metal resistance genes (MRGs) in fattening pigs raised under antibiotic-free (ABF) conditions compared with ordinary industrial pigs (control, C) using whole-genome shotgun sequencing. ABF pigs showed enrichment of Prevotella (33%) and Lactobacillus (13%), whereas Escherichia coli (40%), Fusobacterium, and Bacteroides (each at 4%) were notably observed in the C group. Distinct clusters of cecal microbiota of ABF and C pigs were revealed; however, microbiota of some C pigs (C1) appeared in the same cluster as ABF and were totally separated from the remaining C pigs (C2). For AMR genes, the most abundant genes, tet(Q) (35.7%) and mef(A) (12.7%), were markedly observed in the ABF group, whereas tet(Q) (26.2%) and tet(W) (10.4%) were prominently shown in the C group. tet(Q) was positively correlated with Prevotella in ABF and C1 samples. In the C2 group, the prominent tet(W) was positively correlated with Fusobacterium and Bacteroides. The pigs studied here have never received tetracycline, but pregnant sows received chlortetracycline once every 7 days before parturition. Chromosomal Cu and Zn resistance genes were also shown in both groups regardless of the received Cu and Zn feed additives. A higher abundance of multimetal resistance genes was observed in the C group (44%) than with the ABF group (41%). In conclusion, the microbiome clusters in some C pigs were similar to that in ABF pigs. High-abundance tetracycline resistance genes interrelated to major bacteria were observed in both ABF and C pigs. MRGs were also observed. IMPORTANCE AMR is an increasing problem in farm animals, and raising farm animals without antibiotics is one method that could solve this problem. Our study showed that only some tetracycline and macrolide resistance genes, tet(Q), tet(W), and mef(A), were markedly abundant in ABF and C groups. The tet(Q) and tet(W) genes interrelated to different predominant bacteria in each group, showing the potential role of major bacteria as reservoirs of AMR genes. In addition, chromosomal Cu and Zn resistance genes were also observed in both pig groups, independent of the use of Cu and Zn additives on both farms. The association of MRGs and AMR genotypes and phenotypes, together with the method to resensitize bacteria to antibiotics, should be studied further to unveil the cause of genes conferring high-level resistance and solve these problems.

Background Colistin and carbapenem-resistant bacteria have emerged and become a serious public health concern, but their epidemiological data is still limited. Objectives This study examined colistin and carbapenem resistance in Escherichia coli and Salmonella from pigs, pig carcasses, and pork in Thailand, Lao PDR, and Cambodia border provinces. Methods The phenotypic and genotypic resistance to colistin and meropenem was determined in E. coli and Salmonella obtained from pigs, pig carcasses, and pork (n = 1,619). A conjugative experiment was performed in all isolates carrying the mcr gene (s) (n = 68). The plasmid replicon type was determined in the isolates carrying a conjugative plasmid with mcr by PCR-based replicon typing (n = 7). The genetic relatedness of mcr-positive Salmonella (n = 11) was investigated by multi-locus sequence typing. Results Colistin resistance was more common in E. coli (8%) than Salmonella (1%). The highest resistance rate was found in E. coli (17.8%) and Salmonella (1.7%) from Cambodia. Colistin-resistance genes, mcr-1, mcr-3, and mcr-5, were identified, of which mcr-1 and mcr-3 were predominant in E. coli (5.8%) and Salmonella (1.7%), respectively. The mcr-5 gene was observed in E. coli from pork in Cambodia. Two colistin-susceptible pig isolates from Thailand carried both mcr-1 and mcr-3. Seven E. coli and Salmonella isolates contained mcr-1 or mcr-3 associated with the IncF and IncI plasmids. The mcr-positive Salmonella from Thailand and Cambodia were categorized into two clusters with 94%-97% similarity. None of these clusters was meropenem resistant. Conclusions Colistin-resistant E. coli and Salmonella were distributed in pigs, pig carcasses, and pork in the border areas. Undivided-One Health collaboration is needed to address the issue.

The Thai Journal of Veterinary Medicine, 51, 2, 303

Extended-spectrum β-lactamase (ESBL)-producing Escherichia coli is a serious-global public health issue. A total of 292 E. coli isolates obtained from fecal samples of pigs in Central (n = 103) and Northeastern (n = 189) provinces of Thailand were included in this study. Eighty-six E. coli isolates were phenotypically confirmed to be β-lactamase producers (29.5%) and screened for the presence of β-lactamase genes. The genes in CTX-M family was most frequently found (90.7%). The blaCTX-M-15 gene (59.3%) was predominantly identified CTX-M genotype, followed by blaCTX-M-14 (31.4%) and blaCTX-M-4 (25.6%). The blaTEM-1 gene was prevalent (75.6%). The blaCTX-M-4 and blaCTX-M-14 genes were located on conjugative plasmid. The results highlight healthy pigs as reservoirs of ESBL-producing E. coli carrying ESBL genes that could be horizontally transferred.

A total of 594 Vibrio parahaemolyticus isolates from cultivated oysters (n = 361) and estuarine water (n = 233) were examined for antimicrobial resistance (AMR) phenotype and genotype and virulence genes. Four hundred forty isolates (74.1%) exhibited resistance to at least one antimicrobial agent and 13.5% of the isolates were multidrug-resistant strains. Most of the V. parahaemolyticus isolates were resistant to erythromycin (54.2%), followed by sulfamethoxazole (34.7%) and trimethoprim (27.9%). The most common resistance genes were qnr (77.8%), strB (27.4%) and tet(A) (22.1%), whereas blaTEM (0.8%) was rarely found. Four isolates (0.7%) from oysters (n = 2) and estuarine water (n = 2) were positive to tdh, whereas no trh-positive isolates were observed. Significantly positive associations among AMR genes were observed. The SXT elements and class 1, 2 and 3 integrons were absent in all isolates. The results indicated that V. parahaemolyticus isolates from oysters and estuarine water were potential reservoirs of resistance determinants in the environment. This increasing threat of resistant bacteria in the environment potentially affects human health. A ‘One Health’ approach involved in multidisciplinary collaborations must be implemented to effectively manage antimicrobial resistance.

A total of Escherichia coli isolated from pigs (n = 643), pork (n = 111) and pig carcasses (n = 110) in Thailand during 2008–2018 were included. Susceptibilities to antibiotics, disinfectants (i.e. triclosan (TCS), chlorhexidine digluconate (CHX), benzalkonium chloride (BKC), glutaraldehyde (GLU), formaldehyde (FOR)) and heavy metals (i.e. zinc chloride and copper sulfate) were examined. The E. coli isolates susceptible to all antibiotics (n = 24) were included for in vitro exposure experiment. The effect of phenylalanine arginine β-naphthylamide (PAβN) on the susceptibilities was determined. The results showed that the majority of E. coli (89%) from pigs, pig carcass and pork were multidrug resistant strains. The MICs of all disinfectants and heavy metals, except triclosan were clustered under 1 or 2 concentrations, indicating that there was no development or development at limited degree of resistance to these substances. TCS MICs for E. coli isolated in 2011–2014 were significantly higher than that of the strains in 2008–2010. The weak correlation between biocide MICs (BKC, GLU, TCS, CHX and CuSO4) and antibiotic resistance was observed (p

This study aimed to examine the prevalence and antimicrobial resistance (AMR) of Salmonella isolates from broiler chickens, pigs and their associated meat products in the Thailand–Cambodia border provinces. A total of 941 samples were collected from pigs and broiler chickens at slaughter houses and from carcasses at local fresh markets in Sa Kaeo, Thailand (n = 554) and Banteay Meanchey, Cambodia (n = 387) in 2014 and 2015. From these samples, 345 Salmonella isolates were collected from Sa Keao (n = 145; 23%) and Banteay Meanchey (n = 200; 47%) and assayed for antimicrobial susceptibility, class 1 integrons and extended-spectrum β-lactamase (ESBL) genes. Serovars Typhimurium (29%) and Rissen (29%) were the most common serotypes found in Thai and Cambodian isolates, respectively. Multidrug resistance was detected in 34% and 52% of isolates from Sa Keao and Banteay Meanchey, respectively. The majority of the Thai isolates were resistant to ampicillin (72.4%), whereas most Cambodian isolates were resistant to sulfamethoxazole (71%). Eleven isolates from Sa Keao and 44 from Banteay Meanchey carried class 1 integrons comprising resistance gene cassettes. The most common gene cassette array was dfrA12-aadA2 (61.1%). Six isolates were ESBL producers. The β-lactamase genes found included blaTEM-1, blaCTX-M-55 and blaCMY-2. Some of these class 1 integrons and ESBL genes were located on conjugative plasmid. In conclusion, multidrug-resistant Salmonella are common in pigs, chickens and their products in the Thailand–Cambodia border provinces. Our findings indicate that class 1 integrons play a role in spread of AMR in the strains in this study.

Nine hundred and forty-one samples were collected in Sa Keao, Thailand (n = 554) and Banteay Meanchey, Cambodia (n = 387) from July 2014 to January 2015. A total of 667 Escherichia coli isolates (381 isolates from Sa Keao and 286 isolates from Banteay Meanchey) were obtained and examined for antimicrobial susceptibility, class 1 integrons, ESBL genes and horizontal transfer of resistance determinants. Prevalence of E. coli in pig and broiler carcass samples from slaughterhouses and fresh markets was 36–85% in Sa Keao and 11–69% in Banteay Meanchey. The majority of these isolates were multidrug resistant (75.3%). Class 1 integrons were common in both Thai (47%) and Cambodian (62%) isolates, of which four resistance gene cassette arrays including aadA1, dfrA1-aadA1, dfrA12-aadA2 and aadA2-linF were identified. Class 1 integrons in two broiler isolates from Sa Keao (dfrA12-aadA2) and one broiler isolate from Banteay Meanchey (dfrA1-aadA1) were horizontally transferable. Sixteen isolates were confirmed to be ESBL-producing strains with ESBL gene blaCTX-M-15, broad spectrum β-lactamase gene blaTEM-1 and the AmpC gene blaCMY-2 being detected. The blaTEM-1 gene was most prevalent and located on a conjugative plasmid.

It is generally believed that the Pseudomonas aeruginosa biofilm matrix itself acts as a molecular sieve or sink that contributes to significant levels of drug resistance, but it is becoming more apparent that multidrug efflux pumps induced during biofilm growth significantly enhance resistance levels. We present here a novel transcriptional regulator, PA3898, which controls biofilm formation and multidrug efflux pumps in P. aeruginosa. A mutant of this regulator significantly reduced the ability of P. aeruginosa to produce biofilm in vitro and affected its in vivo fitness and pathogenesis in Drosophila melanogaster and BALB/c mouse lung infection models. Transcriptome analysis revealed that PA3898 modulates essential virulence genes/pathways, including multidrug efflux pumps and phenazine biosynthesis. Chromatin immunoprecipitation sequencing (ChIP-seq) identified its DNA binding sequences and confirmed that PA3898 directly interacts with promoter regions of four genes/operons, two of which are mexAB-oprM and phz2. Coimmunoprecipitation revealed a regulatory partner of PA3898 as PA2100, and both are required for binding to DNA in electrophoretic mobility shift assays. PA3898 and PA2100 were given the names MdrR1 and MdrR2, respectively, as novel repressors of the mexAB-oprM multidrug efflux operon and activators for another multidrug efflux pump, EmrAB. The interaction between MdrR1 and MdrR2 at the promoter regions of their regulons was further characterized via localized surface plasmon resonance and DNA footprinting. These regulators directly repress the mexAB-oprM operon, independent of its well-established MexR regulator. Mutants of mdrR1 and mdrR2 caused increased resistance to multiple antibiotics in P. aeruginosa, validating the significance of these newly discovered regulators.

Background and Aim: The genetic relationship among serotypes of Salmonella enterica from food animals, food of animal origin, and human is of interest as the data could provide an important clue for the source of human infection. This study aimed to determine the genetic relatedness of S. enterica from pig production and human in Thailand–Laos border provinces. Materials and Methods: A total of 195 S. enterica serotypes isolated from pig and pork (n=178) and human (n=17) including four serotypes (Typhimurium, Rissen, Derby, and Stanley) were randomly selected to examine their genetic relatedness using highly conserved sequence of three genes (fim A, man B, and mdh). Results: The results showed that 195 Salmonella isolates of four different serotypes were grouped into five different clusters, and members of the same Salmonella serotypes were found in the same cluster. Salmonella isolated from pig production and human in Thailand–Laos border provinces represented overlapping population and revealed a high degree of similarity, indicating close genetic relationship among the isolates. Conclusion: The results support that the determination of Salmonella serotyping combined with analysis of phylogenetic tree can be used track the clonal evolution and genetic diversity of Salmonella serotypes in different host species.

This study characterized microbiological and chemical contamination of oyster meat and estuarine water in Phang Nga, Thailand. Pooled oyster meats (n = 144), estuarine waters (n = 96) and environmental parameters were collected from March, 2016 to February, 2017, and assessed for levels of total coliforms (TC), fecal coliforms (FC), Escherichia coli (EC), and Vibrio parahaemolyticus (VP), presence of Salmonella and Shigella and levels of heavy metals (Mn, Pb and Cd). The prevalence of TC, FC and EC were in 99.3%, 94.4% and 93.1% of oyster meat and 94.8%, 79.2%, and 78.1% of water, respectively. The average VP levels was 8.5 &times, 107 most probable number (MPN)/g oyster. Prevalence of Shigella and Salmonella in the pooled oysters were 7.6% and 30.6%, respectively. The dominant Salmonella serovars were Paratyphi B followed by Seremban, and Kentucky. In contrast, the prevalence of Shigella were 27.1%, but Salmonella was not detected in estuarine water. Factors statistically associated with EC accumulation in oyster were level of FC, 7-day average precipitation, temperature, relative humidity, and presence of Salmonella in the sample. The optimal cutoff value of EC to predict Salmonella in oyster was 420 MPN/g. Results indicate this area has relatively safe levels of heavy metals, whereas bacterial contamination was very high for oysters.

This study aimed to determine prevalence, antimicrobial resistance, virulence genes, and Class 1 integrons of Enterococcus faecium (E. faecium) and Enterococcus faecalis (E. faecalis) from pigs, pork and humans in Thailand-Laos border provinces.Six hundred and forty-eight rectal and carcass swab samples from border provinces of Thailand (n=359) and Lao PDR (n=289) were collected and examined from September 2013 to October 2014.The overall prevalence of Enterococcus species was 483 of 648 (75%), comprising E. faecium (359 of 483, 74.3%) and E. faecalis (124 of 483, 25.7%). The occurrence of E. faecium in pigs, pig carcasses, retail pork, and humans in Thailand was 80.6%, 73.8%, 77.6%, and 67%, respectively. The prevalence of E. faecium was higher in Laos (65.7%) than Thailand (47.1%) (P0.001). Conversely, E. faecalis was more common in Thailand (24.2%) than Laos (12.8%) (P0.001). The E. faecium and E. faecalis isolates were resistant to all antimicrobials except vancomycin. High resistance was first observed to tetracycline, erythromycin and streptomycin, followed by gentamicin, ampicillin and chloramphenicol. Both E. faecium (7%) and E. faecalis (0.8%) carried empty Class 1 integrons: E. faecium carried gel (6.4%) and esp (0.8%), while E. faecalis carried agg (41.9%), cylA (36.3%), gel (60.5%), and esp (42.7%).This study revealed a variable distribution of antimicrobial resistance and virulence genes among E. faecium and E. faecalis from pigs, pig products and humans in Thai-Laos border provinces. These pathogens may serve as potential reservoirs for the maintenance and widespread dissemination of antimicrobial resistance and virulence determinants from animals to humans via the food chain.

The aim of this study was to detect the expression of 4 clinically-important efflux pumps in the Resistance-Nodulation-Cell Division (RND) family including MexAB-OprM, MexXY, MexCD-OprJ and MexEF-OprN in Pseudomonas aeruginosa using a combination of resistance-phenotypic markers and multiplex RT-PCR (mRT-PCR). The antibiotic substrates specific for each Mex systems were used as phenotypic markers including carbenicillin, MexAB-OprM, erythromycin, MexCD-OprJ, norfloxacin and imipenem, MexEF-OprN and gentamicin, MexXY-OprM. The methods were validated with reference strains with known genotypes of the Mex systems and the potential applicability in clinical practice was tested with clinical isolates. The results for the reference strains support that the combination of resistance phenotype and mRT-PCR is a potential-attractive method for diagnosis of efflux-mediated resistance in P. aeruginosa. Further development to make it more practical for clinical use and study in a larger number of clinical isolates is required.

One hundred Acinetobacter baumannii clinical isolates were examined for inhibitory effect of reserpine and carbonyl cyanide m-chlorophenylhydrazone (CCCP) on the antimicrobial susceptibility and expression of 4 resistant–nodulation–cell division (RND)-type multidrug efflux systems, including AdeABC, AdeDE, AdeIJK, and AdeFGH, using RT-PCR. Ten A. baumannii isolates expressing AdeABC, AdeIJK, or AdeFGH were randomly selected for determination of transcription level and regulatory mutations. While all the isolates were resistant to multiple drugs, the reserpine and CCCP experiment showed that the multidrug resistance phenotype in most A. baumannii isolates was associated with efflux pumps. Most isolates expressed at least one of the RND-type efflux pumps tested (97%). AdeIJK expression was most common (97%), but none of the isolates produced AdeDE. Fifty-two percent of the A. baumannii isolates simultaneously produced up to 3 RND-type efflux systems (i.e., AdeABC, AdeFGH, and AdeIJK). No good correlation between the expression of RND-type efflux pumps and the type of antimicrobial resistance was observed. Overexpression of AdeABC, AdeIJK, and AdeFGH was not always related to the presence of mutations in their corresponding regulatory genes. This study highlights (i) the universal presence of the RND-type efflux pumps with variable levels of expression level among the A. baumannii in this collection and (ii) the complexity of their regulation of expression.

Salmonella enterica isolates (n = 122), including 32 serotypes from 113 dogs and 9 cats, were obtained from household dogs (n = 250) and cats (n = 50) during 2012–2015. The isolates were characterized by serotyping, antimicrobial resistance phenotyping and genotyping, and virulence gene screening. Serovars Weltevreden (15.6%) and Typhimurium (13.9%) were the most common. The majority (43%) of the isolates were multidrug resistant. The dog isolates (12.3%) harbored class 1 integrons, of which the dfrA12-aadA2 cassette was most frequent (66.7%). The only class integron in serovar Albany was located on a conjugative plasmid. Two ESBL-producing isolates (i.e., a serovar Krefeld and a serovar Enteritridis) carried blaTEM and blaCTX-M, and the blaTEM gene in both was horizontally transferred. Of the plasmid-mediated quinolone resistance genes tested, only qnrS (4.9%) was detected. Most Salmonella isolates harbored invA (100%), prgH (91.8%), and sipB (91%). Positive associations between resistance and virulence genes were observed for blaPSE-1/orgA, cmlA/spaN, tolC, and sul1/tolC (p < 0.05). The results suggest that companion dogs and cats are potential sources of S. enterica strains that carry resistance and virulence genes and that antimicrobial use in companion animals may select for the examined Salmonella virulence factors.

A total of 344 commensal Escherichia coli isolates from clinically healthy pigs were examined for antimicrobial resistance phenotypes, class 1 integrons, resistance genes, virulence gene profile, and phylogenetic groups. The majority of E. coli isolates were resistant to tetracycline (96.2%) and ampicillin (91.6%). Up to 98% were multidrug resistant. Seventy-three percent of the isolates carried class 1 integrons. Inserted-gene cassette arrays in variable regions included incomplete sat, aadA22, aadA1, dfrA12-aadA2, and sat-psp-aadA2, of which the aadA2 gene cassette was most prevalent (42.9%). Horizontal transfer was detected in eight E. coli isolates carrying class 1 integrons with dfrA12-aadA2 gene cassette array. Sixteen resistance genes were identified among the E. coli isolates with corresponding resistance phenotype. Ten virulence genes (including elt, estA, estB, astA, faeG, fasA, fedA, eaeA, paa, and sepA) were detected, of which fasA was most commonly found (98.3%). Most of the E. coli isolates belonged to phylogenetic group B1. Significantly positive associations were observed between some virulence genes and some resistance phenotypes and genotypes (p0.05). The results support a finding that commensal E. coli have a role as reservoirs for antimicrobial resistance-encoding genes and virulence determinants.

Global gene transcription was assessed by microarray experiments following treatment of a triclosan-susceptible Δ(mexAB-oprM) Pseudomonas aeruginosa strain with subinhibitory concentrations of triclosan. Expression patterns of selected genes were verified by quantitative real-time PCR analysis. The results showed that triclosan exposure had a profound effect on gene expression, affecting 44% of the genes present on the Affymetrix GeneChip(®), with 28% of genes being significantly upregulated and 16% being significantly downregulated in triclosan-treated cells. Genes encoding membrane proteins, transporters of small molecules, aspects of amino acid metabolism, and transcriptional regulators were significantly over-represented among the more strongly upregulated or downregulated genes in triclosan-treated cells. Quorum sensing-regulated genes were among the most strongly downregulated genes, presumably because of decreased acyl-acyl carrier protein pools and the resulting reduced acyl-homoserine lactone molecule synthesis. Surprisingly, iron homeostasis was completed perturbed in triclosan-exposed cells, with iron acquisition systems being strongly downregulated and iron storage systems significantly upregulated, thus mimicking conditions of excess iron. The profound perturbations of cellular metabolism via specific and global mechanisms may explain why triclosan is such a potent antimicrobial in susceptible bacteria.

The aim of this study was to determine antimicrobial resistance of Aeromonas hydrophila isolated from farmed Nile Tilapia. A total of 50 A. hydrophila isolates from clinical cases were screened for the presence of class 1, 2 and 3 integrons and all the strains resistant to enrofloxacin and/or ciprofloxacin (n=19) examined for mutation in the quinolone resistance-determining regions (QRDRs) of gyrA and parC. The intI1 gene was detected in 23 A. hydrophila strains (46%) but no intl2 and intl3 were detected. Among these, 14 isolates (60.8%) carried gene cassettes inserted in variable regions i.e., partial aadA2, aadA2, dfrA1-orfC and dfrA12-aadA2, of which the most common gene cassette array was dfrA12-aadA2 (26.09%). Conjugal transfer of class 1 integrons with resistance gene array was detected. All the A. hydrophila strains resistant to enrofloxacin and/or ciprofloxacin possessed mutations in the QRDRs of gyrA and parC. Only a Ser-83-Ile substitution was identified in GyrA and only a Ser-80-Ile amino change was found in ParC. The data confirms that A. hydrophila from farm-raised Nile Telapia serve as a reservoir for antimicrobial resistance determinants.

A total of 83 Vibrio isolates from farmed marine shrimps (Penaeus monodon) were tested for the presence of class 1, 2 and 3 integrons, SXT constin and tetracycline resistance-encoding genes. Mutations in the quinolone resistance-determining regions (QRDRs) of the gyrA and parC genes were determined in fluoroquinolone-resistant Vibrio strains (n=17). Five isolates were found to carry class 1 integrons, of which only one contained the partial rumA gene in the variable region. All the Vibrio strains were devoid of class 2 and 3 integrons. Seven isolates harbored SXT constin. None of the Vibrio isolates were positive to the tet(K), tet(L), tet(M), tet(O) and tet(S) genes. Ten fluoroquinolone-resistant Vibrio strains carried a point mutation G-248-T in the gyrA QRDR, leading to a Ser-83-Ile substitution in GyrA, but none of these strains had mutations in the QRDR of the parC gene.

A total of 184 resistant Salmonella enterica isolated from poultry and swine were investigated for the distribution of and associations between various antimicrobial resistance genes. All the isolates were screened for the presence of class 2 and 3 integrase genes and 18 resistance genes corresponding to their resistance phenotypes. None of the isolates carried class 2 and 3 integrons. The investigated resistance genes were responsible for resistance in 78% of the isolates. All the strains harboring more than one resistance gene were resistant to three or more antimicrobials. The bla(TEM), cmlA, tetA, dfrA12, sul3, aadA1 genes were detected in the majority of strains resistant to ampicillin (87%), chloramphenicol (63%), tetracycline (60%), trimethoprim (42%), sulphonamides (42%) and streptomycin/spectinomycin (61%), respectively. Two ciprofloxacin-resistant isolates had a single point mutation leading to Ser-83-Phe in GyrA or Thr-57-Ser in ParC. Statistical analysis revealed good correlation between the presence of antimicrobial resistance genes and corresponding resistance phenotype (p

Two hundred eleven Salmonella enterica strains representing 35 serotypes isolated from healthy poultry (n=103) and swine (n=108) were used in this study. The occurrence and characteristics of class 1 integrons were investigated. Salmonella genomic islands (SGIs) and the horizontal transfer of integrons were assessed. One hundred eighty-six isolates (88%) were resistant to at least one antimicrobial and 140 isolates (66%) were multidrug resistant. The intI1 gene was present in 54 isolates (25.6%), of which 33 (15.6%) carried gene cassettes with sizes ranging from 0.7 to 2.3 kb. Sequence analysis revealed 11 distinct integron profiles in which resistance genes bla(PSE-1), dfrA1, dfrA12, aadA2, aadA4a, and silB were present. The gene cassette array dfrA12-aadA2 was the most prevalent among the isolates whereas most integrons were located on conjugative plasmids. SGI1 variants (SGI1-A and -F) were present in nine isolates belonging to serovars Albany, Emek, Kedougou, and Kingston.

Salmonella enterica isolates from poultry (n=125) and swine (n=132) in Thailand were investigated for antibiotic resistance, susceptibility to disinfectants (benzalkonium chloride (BKC), chlorhexidine digluconate (CHX), zinc chloride and copper sulfate) and cyclohexane tolerance. Forty-two percent were of multiple resistance to antibiotics. The minimum inhibitory concentrations (MICs) of all antibiotics against isolates from swine were higher than that against the isolates from poultry. There were generally few variations in MICs to all disinfectants, indicating that the isolates had either no or only a limited degree of developed resistance to the disinfectants tested. Only 5 isolates (1.9%) were tolerant to cyclohexane. The proton-dependent efflux systems did not play a major role in the reduced susceptibility to BKC and CHX, since susceptibility was not restored when an efflux inhibitor, carbonyl cyanide m-chlorophenylhydrazone (CCCP) was present. Successive exposure to subinhibitory concentrations of BKC and CHX generated mutants resistant to BKC and CHX. A spontaneous BKC-resistant derivative expressed cross-resistance to antibiotics, chloramphenicol and erythromycin. The mechanism responsible for cross-resistance between BKC and antibiotics was not driven by the proton motif force (PMF).

A total of 221 Salmonella enterica from raw pork (n=64), raw chicken (n=80), and humans (n=77) were characterized for antimicrobial resistance phenotypes and genotypes and virulence plasmid-associated genes. Most Salmonella isolates (95.9%) were multidrug resistant and exhibited high resistance to sulfamethoxazole (96.4%), streptomycin (93.2%), spectinomycin (76.5%), tetracycline (73.3%), ampicillin (70.1%), and trimethoprim (60.2%). Forty-one percent of all isolates were intI1-positive, of which 60% carried class 1 integrons with variable region ranging in size from 0.2 to 2.0 kb. Six integron profiles (IP-I to IP-VI) were defined. The dfrA12-aadA2 cassette was most prevalent (66.7%). Class 1 integrons with the dfrA12-aadA2 cassette in five pork isolates could be horizontally transferred. Three pork isolates carried Salmonella genomic island 1 (SGI1), of which a serovar Anatum harbored SGI1 gene cluster located between thdF and int2. Two single-point mutations (i.e., G-259-T and C-248-T) in gyrA leading to Asp-87-Tyr and Ser-83-Phe substitutions in GyrA, respectively, were detected. Of all plasmid-mediated quinolone resistance genes tested, only qnrS (4.1%) and qnrB (1.8%) were found. The virulence plasmid-associated genes including spvC, pefA, and rck were identified in 8.1%, 1.8%, and 1.4% of all Salmonella isolates, respectively.