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This study aimed to investigate the potential mechanisms associated with the persistence of chloramphenicol (CHP) resistance in Escherichia coli and Salmonella enterica isolated from pigs, pork, and humans in Thailand. The CHP-resistant E. coli (n = 106) and Salmonella (n = 57) isolates were tested for their CHP susceptibility in the presence and absence of phenylalanine arginine β-naphthylamide (PAβN). The potential co-selection of CHP resistance was investigated through conjugation experiments. Whole genome sequencing (WGS) was performed to analyze the E. coli (E329, E333, and E290) and Salmonella (SA448, SA461, and SA515) isolates with high CHP MIC (32-256 μg/mL) and predominant plasmid replicon types. The presence of PAβN significantly reduced the CHP MICs (≥4-fold) in most E. coli (67.9%) and Salmonella (64.9%). Ampicillin, tetracycline, and streptomycin co-selected for CHP-resistant Salmonella and E. coli-transconjugants carrying cmlA. IncF plasmids were mostly detected in cmlA carrying Salmonella (IncFIIAs) and E. coli (IncFIB and IncF) transconjugants. The WGS analysis revealed that class1 integrons with cmlA1 gene cassette flanked by IS26 and TnAs1 were located on IncX1 plasmid, IncFIA(HI1)/HI1B plasmids and IncFII/FIB plasmids. IncFIA(HI1)/HI1B/Q1in SA448 contained catA flanked by IS1B and TnAs3. In conclusion, cross resistance through proton motive force-dependent mechanisms and co-selection by other antimicrobial agents involved the persistence of CHP-resistance in E. coli in this collection. Dissemination of CHP-resistance genes was potentially facilitated by mobilization via mobile genetic elements., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Puangseree et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

The impact of antimicrobial resistance (AMR) on global public health has been widely documented. AMR in the environment poses a serious threat to both human and animal health but is frequently overlooked. This study aimed to characterize the association between phenotype and genotype of AMR, virulence genes and Extended-Spectrum β-Lactamase (ESBL) production from estuarine environment. The Salmonella (n = 126) and E. coli (n = 409) were isolated from oysters and estuarine water in Thailand. The isolates of Salmonella (96.9%) and E. coli (91.4%) showed resistance to at least one antimicrobial agent. Multidrug resistance (MDR) was 40.1% of Salmonella and 23.0% of E. coli. Resistance to sulfamethoxazole was most common in Salmonella (95.2%) and E. coli (77.8%). The common resistance genes found in Salmonella were sul3 (14.3%), followed by blaTEM (11.9%), and cmlA (11.9%), while most E. coli were blaTEM (31.5%) and tetA (25.4%). The ESBL production was detected in Salmonella (1.6%, n = 2) of which one isolate was positive to blaTEM-1. Eight E. coli isolates (2.0%) were ESBL producers, of which three isolates carried blaCTX-M-55 and one isolate was blaTEM-1. Predominant virulence genes identified in Salmonella were invA (77.0%), stn (77.0%), and fimA (69.0%), while those in E. coli isolates were stx1 (17.8%), lt (11.7%), and stx2 (1.2%). Logistic regression models showed the statistical association between resistance phenotype, virulence genes and ESBL production (p < 0.05). The findings highlighted that estuarine environment were potential hotspots of resistance. One Health should be implemented to prevent AMR bacteria spreading., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2023 Jeamsripong et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

This study aimed to analyze three ESBL-producing E. coli co-harboring mcr and ESBL genes from a healthy fattening pig (E. 431) and two sick pigs (ECP.81 and ECP.82) in Thailand using Whole Genome Sequencing (WGS) using either Illumina MiSeq or HiSeq PE150 platforms to determine their genome and transmissible plasmids. E. 431 carrying mcr-2.1 and mcr-3.1 belonged to serotype O142:H31 with ST29 sequence type. ECP.81 and ECP.82 from sick pigs harboring mcr-1.1 and mcr-3.1 were serotype O9:H9 with ST10. Two mcr-1.1 gene cassettes from ECP.81 and ECP.82 were located on IncI2 plasmid with 98% identity to plasmid pHNSHP45. The mcr-2.1-carrying contig in E. 431 showed 100% identity to plasmid pKP37-BE with the upstream flanking sequence of IS1595. All three mcr-3.1-carrying contigs contained the ΔTnAs2-mcr-3.1-dgkA core segment and had high nucleotide similarity (85-100%) to mcr-3.1-carrying plasmid, pWJ1. The mobile elements i.e. IS4321, ΔTnAs2, ISKpn40 and IS3 were identified in the flanking regions of mcr-3. Several genes conferring resistance to aminoglycosides (aac(3)-IIa, aadA1, aadA2b, aph(3'')-Ib, aph(3')-IIa and aph(6)-Id), macrolides (mdf(A)), phenicols (cmlA1), sulphonamide (sul3) and tetracycline (tet(A) and tet(M)) were located on plasmids, of which their presence was well corresponded to the host's resistance phenotype. Amino acid substitutions S83L and D87G in GyrA and S80I and E62K in ParC were observed. The blaCTX-M-14 and blaCTX-M-55 genes were identified among these isolates additionally harbored blaTEM-1B. Co-transfer of mcr-1.1/blaTEM-1B and mcr-3.1/blaCTX-M-55 was observed in ECP.81 and ECP.82 but not located on the same plasmid. The results highlighted that application of advanced innovation technology of WGS in AMR monitoring and surveillance provide comprehensive information of AMR genotype that could yield invaluable benefits to development of control and prevention strategic actions plan for AMR., Competing Interests: The authors have declared that no competing interests exist.

The emergence and dissemination of extended-spectrum β-lactamase (ESBL)-producing Escherichia coli is a global health issue. Food-producing animals, including pigs, are significant reservoirs of antimicrobial resistance (AMR), which can be transmitted to humans. Thus, the rapid detection of ESBLs is required for efficient epidemiological control and treatment. In this study, multiplex recombinase polymerase amplification (RPA) combined with a single-stranded tag hybridization chromatographic printed-array strip (STH-PAS), as a lateral flow strip assay (LFA), was established for the rapid and simultaneous detection of multiple bla genes in a single reaction. Visible blue lines, indicating the presence of the blaCTX-M, blaSHV, and blaOXA genes, were observed within 10 min by the naked eye. The limit of detection of all three genes was 2.5 ng/25 μL, and no cross-reactivity with seven commensal aerobic bacteria was observed. A total of 93.9% (92/98) and 96% (48/50) of the E. coli isolates from pork meat and fecal samples, respectively, expressed an ESBL-producing phenotype. Nucleotide sequencing of the PCR amplicons showed that blaCTX-M was the most prevalent type (91.3-95.83%), of which the main form was blaCTX-M-55. The sensitivity and specificity of the RPA-LFA were 99.2% and 100%, respectively, and were in almost perfect agreement (κ = 0.949-1.000) with the results from PCR sequencing. Thus, the RPA-LFA is a promising tool for rapid and equipment-free ESBL detection and may facilitate clinical diagnosis in human and veterinary medicine, as well as AMR monitoring and surveillance., Competing Interests: The authors have declared that no competing interests exist.