Gut colonization by extended-spectrum ß-lactamase-producing Escherichia coli in dairy herd in Brazil: successful dissemination of a One Health clone.

The overuse of antimicrobials in livestock has contributed to the emergence and selection of clinically relevant multidrug-resistant bacteria. In Brazil, there is no conclusive information on the occurrence of Escherichia coli producing extended-spectrum ß-lactamase (ESßL) in cattle breeding, which is an important sector of agribusiness in this country. Herein, we investigated the presence of ESßL-positive E. coli strains in dairy cattle from a commercial farm with routine practice of therapeutic cephalosporins. Ninety-five rectal swab samples were collected from healthy dairy calves and cows under treatment with ceftiofur. Samples were screened for the presence of ESßL producers, and positive isolates were identified by MALDI-TOF, with subsequent screening for genes encoding ESßL variants by PCR and sequencing. The presence of ESßL (CTX-M-15)-producing E. coli was confirmed in calves, and lactating and dry cows. Most ESßL strains with genetic homologies ≥ 90% were grouped into two major PFGE clusters, confirming the suscessful expansion of clonally related lineages in animals from different lactating cycles, on the same property. Four representatives CTX-M-15-positive E. coli strains had their genomes sequenced, belonging to the clonal complex (CC) 23 and sequence type (ST) 90. A phylogeographical landscape of ST90 was performed revealing a global One Health linkage. Our results highlight the intestinal microbiota of dairy cattle as a hotspot for the spread of critical priority ESßL-producing E. coli and demonstrate that ST90 is an international clone genomically adapted to human and animal hosts, which deserve additional investigation to determine its zoonotic potential and impact in food chain.

Exploring extended-spectrum beta lactamase resistance in Vibrio parahaemolyticus and Vibrio cholerae within the tropical mangrove ecosystem of southwest India.

Multidrug-resistant pathogenic vibrios are a crisis of concern as they cause multiple illnesses, including gastroenteritis in humans and acute hepatopancreatic necrosis in aquaculture. In the current study, we investigated the prevalence of the beta-lactamase gene CTX-M-group 1 in Vibrio spp. (Vibrio cholerae and Vibrio parahaemolyticus) from the water and sediment of urban tropical mangrove ecosystems of Kerala, southwest India. A total of 120 isolates of Vibrio spp. were tested for antibiotic susceptibility to 14 antibiotics. In water, ampicillin resistance was very high in isolates of V. cholerae (94.1%, n = 17) and V. parahaemolyticus (89.1%, n = 46). 26.9% of V. parahaemolyticus and 14.2% of V. cholerae harbored the CTX-M-group 1 gene in water samples. Compared to V. cholerae, the CTX-M-group 1 gene was exclusively hosted by V. parahaemolyticus (49%) in sediment samples. A significant difference in the prevalence of the CTX-M-group 1 gene was observed among Vibrio spp. in both water and sediment samples (p < 0.05). The results revealed the presence of multidrug-resistant and beta-lactamase harboring Vibrio spp. in mangrove ecosystems, which may have evolved as a consequence of the misuse and abuse of broad-spectrum antibiotics as prophylaxis in human health care and aquaculture.

Extended-spectrum ß-lactamase-producing Plesiomonas shigelloides isolated from the stool of a Japanese traveler returning from Rwanda: A case report.

A 21-year-old previously healthy Japanese woman visited an outpatient clinic because of abdominal pain, watery diarrhea, vomiting, and mild fever that had started on the previous day. She traveled to rural and urban areas of Rwanda and returned to Japan 3 days before. Stool culture yielded the Plesiomonas shigelloides strain TMCH301018, against which minimum inhibitory concentrations of cefotaxime and cefotaxime-clavulanate were 128 and ≤0.12/4 µg/mL, respectively. The strain had the blaCTX-M-27 gene and an IncA/C replicon-type plasmid. Moreover, a transformant produced by introduction of an IncA/C plasmid extracted from TMCH301018 into Escherichia coli DH5α was positive for the blaCTX-M-27 gene and fulfilled the criteria of extended-spectrum ß-lactamase (ESBL) production described by the Clinical and Laboratory Standards Institute, indicating that TMCH301018 produced ESBL of CTX-M-27 and the ESBL-encoding gene was located on an IncA/C plasmid. Pathogenicity of TMCH301018 for the patient's complaints was uncertain because a molecular assay detected other enteropathogens in the stool specimen and the symptoms improved within 2 days with administration of oral ciprofloxacin, to which TMCH301018 was not susceptible. To our knowledge, this is the first report describing the isolation of ESBL-producing P. shigelloides.

Distribution and antimicrobial susceptibility pattern of CTX-M-type extended-spectrum ß-lactamase-producing Escherichia coli isolated in Chubu region, Japan.

The widespread prevalence of extended-spectrum ß-lactamase (ESBL)-producing Escherichia coli limits treatment options and is a worldwide problem. The aim of this study was to investigate the antimicrobial susceptibility and ESBL-type of 204 strains of CTX-M-type ESBLs-producing E. coli isolated from 2011 to 2017 in the Chubu region of Japan. Minimal inhibitory concentrations were determined in accordance with the guidelines of the Clinical and Laboratory Standards Institute. Genes encoding CTX-M group ß-lactamases were detected by PCR amplification. The CTX-M subtypes were determined using sequence analysis. The CTX-M-9 group was the most frequently detected ESBL group, and CTX-M-27 was the most frequently detected ESBL gene. CTX-M-15-producing strains showed significantly lower rates of susceptibility to tazobactam/piperacillin (TAZ/PIPC) than those by CTX-M-14 and -27-producing strains. Additional analysis of secondary ß-lactamases revealed that most of the OXA-1-positive strains were CTX-M-15-producing strains (94.7%). These strains displayed significantly lower susceptibility rates to TAZ/PIPC (47.4%), sulbactam/ampicillin (SBT/ABPC) (0.0%), and amikacin (AMK) (73.7%) than those by OXA-1-negative strains, suggesting that the high non-susceptibility rate of the CTX-M-15-producing strain was due to the co-carriage of OXA-1. The CTX-M-15-producing strains showed reduced susceptibility to TAZ/PIPC, SBT/ABPC, and AMK, presumably due to the co-carriage of OXA-1.

ß-Lactamase and Macrolide Resistance Gene Carriage in Escherichia coli Isolates Among Children Discharged From Inpatient Care in Western Kenya: A Cross-sectional Study.

Background: Antimicrobial resistance (AMR) is a global threat to infectious disease control, particularly among recently hospitalized children. We sought to determine the prevalence and mitigating factors of resistance in enteric Escherichia coli among children discharged from health facilities in western Kenya. Methods: Between June 2016 and November 2019, children aged 1 to 59 months were enrolled at the point of discharge from the hospital. E coli was isolated by microbiological culture from rectal swabs at baseline. ß-Lactamases and macrolide resistance-conferring genes were detected by polymerase chain reaction. A modified Poisson regression model was used to assess the predictors mph(A) and CTX-M-type extended-spectrum ß-lactamase (ESBL). Results: Of the 238 children whose E coli isolates were tested, 91 (38.2%) and 109 (45.8%) had detectable CTX-M-type ESBL and mph(A) genes, respectively. Antibiotic treatment during hospitalization (adjusted prevalence ratio [aPR], 2.47; 95% CI, 1.12-5.43; P = .025), length of hospitalization (aPR, 1.42; 95% CI, 1.00-2.01; P = .052), and the practice of open defecation (aPR, 2.47; 95% CI, 1.40-4.36; P = .002) were independent predictors for CTX-M-type ESBL and mph(A) genes. Pneumococcal vaccination was associated with a 43% lower likelihood of CTX-M-type ESBL (aPR, 0.57; 95% CI, .38-.85; P = .005), while measles vaccination was associated with a 32% lower likelihood of mph(A) genes (aPR, 0.68; 95% CI, .49-.93; P = .017) in E coli isolates. Conclusions: Among children discharged from the hospital, history of vaccination, shorter hospital stay, lack of in-hospital antibiotic exposure, and improved sanitation were associated with a lower likelihood of AMR genes. To mitigate the continued spread of AMR, AMR control programs should consider strategies beyond antimicrobial stewardship, including improvements in sanitation, increased vaccine coverage, and the development of novel vaccines.

Dissemination of Ceftriaxone-Resistant Salmonella Enteritidis Harboring Plasmids Encoding blaCTX-M-55 or blaCTX-M-14 Gene in China.

Salmonella Enteritidis was the primary foodborne pathogen responsible for acute gastroenteritis. The growing ceftriaxone resistance poses a significant threat to public health. Infection with S. Enteritidis has emerged as a major public health concern, particularly in developing countries. However, research on ceftriaxone-resistant S. Enteritidis (CRO-RSE) remains limited, particularly concerning its resistance mechanism, plasmid structure, and transmission characteristics. This study aims to address these gaps comprehensively. We collected 235 S. Enteritidis isolates from Hangzhou First People's Hospital between 2010 and 2020. Among these, 8.51% (20/235) exhibited resistance to ceftriaxone. Whole-genome analysis revealed that 20 CRO-RSE isolates harbored blaCTX-M-55 or blaCTX-M-14 on the plasmid. Moreover, the dissemination of the blaCTX-M-type gene was associated with IS26 and ISEcp1. Plasmid fusion entailing the integration of the p1 plasmid with antibiotic resistance genes and the p2 (pSEV) virulence plasmid was observed in certain CRO-RSE. Additionally, the structural analysis of the plasmids unveiled two types carrying the blaCTX-M-type gene: type A with multiple replicons and type B with IncI1 (Alpha) replicon. Type B plasmids exhibited superior adaptability and stability compared to type A plasmids within Enterobacteriaceae. Interestingly, although the type B (S808-p1) plasmid displayed the potential to spread to Acinetobacter baumannii, it failed to maintain stability in this species.

Loss and gain of ceftazidime-avibactam susceptibility in a non-carbapenemase-producing K1-ST23 hypervirulent Klebsiella pneumoniae.

OBJECTIVES: This study aimed at revealing the underlying mechanisms of the loss and gain of ceftazidime-avibactam susceptibility in a non-carbapenemase-producing hypervirulent Klebsiella pneumoniae (hvKp). METHODS: Here we longitudinally recovered 3 non-carbapenemase-producing K1-ST23 hvKp strains at a one-month interval (KP29105, KP29499 and KP30086) from an elderly male. Antimicrobial susceptibility testing, whole genome sequencing, transcriptomic sequencing, gene cloning, plasmid conjugation, quantitative real-time PCR (qRT-PCR), and SDS-PAGE (sodium dodecyl sulfate-polyacrylamide gel electrophoresis) were conducted. RESULTS: Among the 3 hvKp strains, KP29105 was resistant to the third- and fourth-generation cephalosporins, KP29499 acquired resistance to both ceftazidime-avibactam and carbapenems, while KP30086 restored its susceptibility to ceftazidime-avibactam, imipenem and meropenem but retained low-level resistance to ertapenem. KP29105 and KP29499 carried plasmid-encoded genes blaCTX-M-15 and blaCTX-M-71, respectively, but KP30086 lost both. Cloning of gene blaCTX-M-71 and conjugation experiment of blaCTX-M-71-carrying plasmid showed that the transformant and transconjugant were susceptible to ceftazidime-avibactam but had a more than 8-fold increase in MICs. Supplementation with an outer membrane permeabilizer could reduce the MIC of ceftazidime-avibactam by 32 folds, indicating that porins play a key role in ceftazidime-avibactam resistance. The OmpK35 of the 3 isolates was not expressed, and the OmpK36 of KP29499 and KP30086 had a novel amino acid substitution (L359R). SDS-PAGE and qRT-PCR showed that the expression of porin OmpK36 of KP29499 and KP30086 was significantly down-regulated compared with KP29105. CONCLUSIONS: In summary, we reported the rare ceftazidime-avibactam resistance in a non-carbapenemase-producing hvKp strain. Resistance plasmid carrying blaCTX-M-71 and mutated OmpK36 had a synergetic effect on the resistance.

Hospital Wastes as Potential Sources for Multi-Drug-Resistant ESBL-Producing Bacteria at a Tertiary Hospital in Ethiopia.

The hospital environment is increasingly becoming an important reservoir for multi-drug-resistant (MDR) Gram-negative bacteria, posing serious challenges to efforts to combat antimicrobial resistance (AMR). This study aimed to investigate the role of hospital waste as a potential source of MDR ESBL-producing bacteria. Samples were collected from multiple sources within a hospital and its vicinity, including surface swabs, houseflies, and sewage samples. The samples were subsequently processed in a microbiology laboratory to identify potential pathogenic bacteria and confirmed using MALDI-TOF MS. Bacteria were isolated from 87% of samples, with the predominant isolates being E. coli (30.5%), Klebsiella spp. (12.4%), Providencia spp. (12.4%), and Proteus spp. (11.9%). According to the double disc synergy test (DDST) analysis, nearly half (49.2%) of the bacteria were identified as ESBL producers. However, despite exhibiting complete resistance to beta-lactam antibiotics, 11.8% of them did not test positive for ESBL production. The characterization of E. coli revealed that 30.6% and 5.6% of them carried blaCTX-M group 1 type-15 and blaNDM genes, respectively. This finding emphasizes the importance of proper hospital sanitation and waste management practices to mitigate the spread of AMR within the healthcare setting and safeguard the health of both patients and the wider community.

Epidemiological characteristics of human- and chicken-derived CTX-M-type extended-spectrum ß-lactamase-producing Escherichia coli from China.

Bacterial resistance to ß-lactams is mainly attributed to CTX-M-type extended-spectrum ß-lactamases (ESBLs). However, the predominant sequence type (ST) of blaCTX-M-carrying Escherichia coli (blaCTX-M-Ec) in chickens, an important food animal, in China and its contribution to human ß-lactam resistance are not investigated. In this study, approximately 1808 chicken-derived strains collected from 10 provinces from 2012 to 2020 were screened for blaCTX-M-Ec, and 222 blaCTX-M-Ec were identified. Antimicrobial susceptibility tests, whole genome sequencing and conjugation experiment were performed. All quality-controlled 136 chicken-derived blaCTX-M-Ec and 1193 human-derived blaCTX-M-Ec genomes were downloaded from NCBI and EnteroBase to comprehensively analyze the prevalence of blaCTX-M-Ec in China. blaCTX-M-55 (153/358, 42.7% in chicken isolates; 312/1193, 26.2% in human isolates) and blaCTX-M-14 (92/358, 25.7% in chicken isolates; 450/1193, 37.7% in human isolates) were dominant in blaCTX-M-Ec. The STs of blaCTX-M-Ec were diverse and scattered, with ST155 (n = 21) and ST152 (n = 120) being the most abundant in chicken- and human-derived isolates, respectively. Few examples indicated that chicken- and human-derived blaCTX-M-Ec have 10 or less core genome single nucleotide polymorphisms (cgSNPs). Genetic environment analysis indicated that ISEcp1, IS26 and IS903B were closely associated with blaCTX-M transfer. The almost identical pc61-55 and pM-64-1161 indicated the possibility of plasmid-mediated transmission of blaCTX-M between humans and chickens. Although the genomes of most blaCTX-M-Ec isolated from chickens and humans were quite different, the prevalence and genetic environment of blaCTX-M variants in both hosts were convergent. CTX-M-mediated resistance is more likely to spread through horizontal gene transmission than bacterial clones.

The draft genome sequence of blaCTX-M-270-carrying Kluyvera ascorbata isolated from human sputum.

Kluyvera ascorbata is recognized as an opportunistic pathogen of the Enterobacteriaceae family but remains less studied. We report the draft genome of a K. ascorbata clinical strain recovered from human sputum, comprising approximately 5.18 million bases and harboring an intrinsic gene encoding the extended-spectrum ß-lactamase CTX-M-270.

Prevalence and characteristics of ESBL-producing Escherichia coli in clinically healthy pigs: implications for antibiotic resistance spread in livestock.

AIM: This study aimed to compare and characterize the resistance profile and the presence of extended-spectrum beta-lactamase (ESBL) related genes in Escherichia coli isolated from healthy finishing pigs fed with or without antibiotics in their diets. METHODS AND RESULTS: A total of 27 ceftiofur-resistant E. coli isolates were obtained from 96 healthy pigs. The antibiotic resistance profile was tested, and all 27 isolates were classified as multidrug-resistant (MDR). A high proportion of isolates were resistant to cephalosporins, ampicillin, ciprofloxacin, and tetracyclines. The ESBL production was observed in 85% of isolates by double-disc synergy test. The MDR-E. coli isolates harbored ESBL genes, such as blaTEM, blaCTX-M-1, blaCTX-M-2, and blaCTX-M-8,25. In addition, other antibiotics resistance genes (ARGs) were also detected, such as sul2, ant(3″)-I, tetA, and mcr-1. The mobilization of the blaCTX-M gene was confirmed for nine E. coli isolates by conjugation assays. The presence of blaCTX-M on mobile genetic elements in these isolates was demonstrated by Southern blot hybridization, and the resistance to cephalosporins was confirmed in the transconjugants. Our results indicate the prevalence of CTX-M-producing E. coli strains harboring mobile genetic elements in the normal microbiota of healthy pigs. CONCLUSIONS: These findings highlight the significance of ESBL genes as a global health concern in livestock and the potential spread of antimicrobial resistance to other members of the gastrointestinal tract microbiota.

Sensitive and Specific Loop-Mediated Isothermal Amplification Assays for Detection of Salmonella, CTX-M-1 Group Genes, mph(A), and ermB in Stool and Blood Samples Based on Orange to Green Visible Dye.

Salmonella is a globally prevalent foodborne bacterium, and ceftriaxone and azithromycin have been regarded as drugs of choice for treating Salmonella infections, particularly in children. With the growing incidence of ceftriaxone and azithromycin resistance in Salmonella, there is an urgent requirement for a rapid and dependable gene testing approach to enhance the efficacy of treating Salmonella infections. Utilizing the orange to green visible dye approach, this study developed loop-mediated isothermal amplification (LAMP) assays for the sensitive and specific detection of Salmonella, ceftriaxone and azithromycin resistance genes (including CTX-M-1 group, mph(A), and ermB genes) in stool and blood samples. The specificity and sensitivity of primers during the LAMP assays for detection of Salmonella, CTX-M-1 group, mph(A), and ermB genes were determined in this study. The detection threshold for Salmonella was found to be 1.5 × 103 colony-forming units (CFU)/mL, while it was 1.5 × 102 CFU/mL for CTX-M-1 group genes (including blaCTX-M-3, blaCTX-M-15, and blaCTX-M-55), 1.5 × 102 CFU/mL for mph(A), and 1.5 × 102 CFU/mL for ermB, showing 10-103-fold, 103-fold, and 105-fold increased sensitivity compared with the polymerase chain reaction assay, respectively. Results indicated that the LAMP primers designed for Salmonella, CTX-M-1 group, mph(A), and ermB genes possess high specificity (100%) and sensitivity (over 94%). This novel approach advocates its application in detecting Salmonella, CTX-M-1 group, mph(A), and ermB genes.

CTX-M-127 with I176F mutations found in bacteria isolates from Bangladeshi circulating banknotes.

Extended-spectrum beta-lactamase (ESBL)-producing organisms are widely recognized as clinically relevant causes of difficult-to-treat infections. CTX-M has formed a rapidly growing family distributed worldwide among a wide range of clinical bacteria, particularly members of Enterobacteriaceae. Circulating banknotes, exchanged daily among people, pose a potential vehicle for transmitting multidrug resistance. We screened for ESBL-carrying bacteria in the present study and reported CTX-M mutations in Bangladesh's banknotes. We sequenced the genes and performed homology modeling using the Swiss model with CTX-M-15 (4HBT) as a template. Then, we performed molecular docking of mecillinam with the template and the generated model using Autodock 4.2 (Release 4.2.6). After docking, we visually inspected the complexes built using Autodock tools for polar contacts and pi-pi interactions in PyMOL 2.5.4. Our partially sequenced blaCTX-M was related to blaCTX-M-10 and blaCTX-M-15. We observed multiple single-nucleotide substitution mutations, i.e., G613T (silent mutation), A626T (I176F), and A503G (N135D). Homology modeling showed high similarity when the model was superimposed over the template. The orientation of Asn (135) in the template and Asp (135) in the model does not show a significant difference. Likewise, Ile (176) in the template and Phe (176) in the model offer the same orientation. Our generated model could bind to Lys237, Ser240, and Asp135 residues with the lowest binding energy on docking. Our predicted binding of the mecillinam to the mutated D-135 residue in the model indicates contributions and supports previous reports proposing CTX-M-15 to CTX-M-127 mutational conversion on the mecillinum resistance phenotype.

Horizontal transfer characterization of ColV plasmids in blaCTX-M-bearing avian Escherichia coli.

Extended-spectrum-ß-lactamases (ESBLs)-producing Escherichia coli conferred resistance to most ß-lactams, except for carbapenems. To date, the transmission mechanism of blaCTX-M, as the most common ESBLs subtype, in E. coli has received sustained attention around the worldwide, but the research on the pathogenicity of blaCTX-M-bearing E. coli is still scarce. The aims of this study were to discern the spread characteristics of ColV (encoding colicin V) plasmids in blaCTX-M-positive E. coli. The multi-drug resistance traits, phylogroups, and ColV plasmid profilings were screened in 76 blaCTX-M-positive E. coli. Thereafter, the genetic profiles of E. coli G12 and GZM7 were determined by whole genome sequencing, conjugation and S1-pulsed-field gel electrophoresis. The median lethal dose was analyzed in E. coli G12 and TG12A, the ColV-plasmid transconjugant of G12. Of all 76 blaCTX-M-bearing E. coli, 67.11% exhibited resistance to at least 2 drugs in addition to ceftiofur, 14.47% carried ColV-positive plasmids, and 53.95% were phylogroup C. Further studies demonstrated that the blaCTX-M-bearing E. coli G12 was assigned to the predominant lineage O78:H4-ST117 of phylogroup G. In addition, its ColV-positive plasmid simultaneously carried multiple resistance genes, and could be independently transferred to confer partial pathogenicity on its host by plasmid mating. E. coli GZM7 was O53:H9-ST23 of phylogroup C, which belonged to another representative lineage of APEC (avian pathogenic E. coli). Its ColV-positive plasmid could complete conjugation with the help of the other coexisting-resistance conjugative plasmid, although it failed to transfer alone. Our findings highlight the flexibly horizontal transfer of ColV plasmids along with multidrug-resistant genes among blaCTX-M-bearing E. coli poses a threat to poultry health and food safety, which contributes to elucidate the concept of "One Health" and deserves particular concern.

Genomic characterisation of the population structure and antibiotic resistance of Salmonella enterica serovar Infantis in Chile, 2009-2022.

Background: Multidrug-resistant (MDR) Salmonella Infantis has disseminated worldwide, mainly linked to the consumption of poultry products. Evidence shows dissemination of this pathogen in Chile; however, studies are primarily limited to phenotypic data or involve few isolates. As human cases of Salmonella Infantis infections have substantially increased in recent years, this study aimed to characterise the genomic epidemiology and antimicrobial-resistance profiles of isolates obtained from different sources, aiming to inform effective surveillance and control measures. Methods: We sequenced 396 Salmonella Infantis genomes and analysed them with all publicly available genomes of this pathogen from Chile (440 genomes in total), representing isolates from environmental, food, animal, and human sources obtained from 2009 to 2022. Based on bioinformatic and phenotypic methods, we assessed the population structure, dissemination among different niches, and antimicrobial resistance (AMR) profiles of Salmonella Infantis in the country. Findings: The genomic and phylogenetic analyses showed that Salmonella Infantis from Chile comprised several clusters of highly related isolates dominated by sequence type 32. The HC20_343 cluster grouped an important proportion of all isolates. This was the only cluster associated with pESI-like megaplasmids, and up to 12 acquired AMR genes/mutations predicted to result in an MDR phenotype. Accordingly, antimicrobial-susceptibility testing revealed a strong concordance between the AMR genetic determinants and their matching phenotypic expression, indicating that a significant proportion of HC20_343 isolates produce extended-spectrum ß-lactamases and have intermediate fluoroquinolone resistance. HC20_343 Salmonella Infantis were spread among environmental, animal, food, and human niches, showing a close relationship between isolates from different years and sources, and a low intra-source genomic diversity. Interpretation: Our findings show a widespread dissemination of MDR Salmonella Infantis from the HC20_343 cluster in Chile. The high proportion of isolates with resistance to first-line antibiotics and the evidence of active transmission between the environment, animals, food, and humans highlight the urgency of improved surveillance and control measures in the country. As HC20_343 isolates predominate in the Americas, our results suggest a high prevalence of ESBL-producing Salmonella Infantis with intermediate fluoroquinolone resistance in the continent. Funding: Partially supported by the Food and Drug Administration (FDA) of the U.S. Department of Health and Human Services as part of an award, FDU001818, with 30% percent funded by FDA/HHS; and by Agencia de Investigación y Desarrollo de Chile (ANID) through FONDECYT de Postdoctorado Folio 3230796 and Folio 3210317, FONDECYT Regular Folio 1231082, and ANID-Millennium Science Initiative Program-ICN2021_044.

Emergence of a clinical Salmonella enterica serovar 1,4,[5], 12: i:-isolate, ST3606, in China with susceptibility decrease to ceftazidime-avibactam carrying a novel blaCTX-M-261 variant and a blaNDM-5.

PURPOSE: The detection rate of Salmonella enterica serovar 1,4,[5], 12: i: - (S. 1,4,[5], 12: i: -) has increased as the most common serotype globally. A S. 1,4,[5], 12: i: - strain named ST3606 (sequence type 34), isolated from a fecal specimen of a child with acute diarrhea hospitalized in a tertiary hospital in China, was firstly reported to be resistant to carbapenem and ceftazidime-avibactam. The aim of this study was to characterize the whole-genome sequence of S. 1,4,[5], 12: i: - isolate, ST3606, and explore its antibiotic resistance genes and their genetic environments. METHODS: The genomic DNA of S. 1,4,[5], 12: i: - ST3606 was extracted and performed with single-molecule real-time sequencing. Resistance genes, plasmid replicon type, mobile elements, and multilocus sequence types (STs) of ST3606 were identified by ResFinder 3.2, PlasmidFinder, OriTfinder database, ISfinder database, and MLST 2.0, respectively. The conjugation experiment was utilized to evaluate the conjugation frequency of pST3606-2. Protein expression and enzyme kinetics experiments of CTX-M were performed to analyze hydrolytic activity of a novel CTX-M-261 enzyme toward several antibiotics. RESULTS: Single-molecule real-time sequencing revealed the coexistence of a 109-kb IncI1-Iα plasmid pST3606-1 and a 70.5-kb IncFII plasmid pST3606-2. The isolate carried resistance genes, including blaNDM-5, sul1, qacE, aadA2, and dfrA12 in pST3606-1, blaTEM-1B, aac(3)-lld, and blaCTX-M-261, a novel blaCTX-M-1 family member, in pST3606-2, and aac(6')-Iaa in chromosome. The blaCTX-M-261 was derived from blaCTX-M-55 by a single-nucleotide mutation 751G>A leading to amino acid substitution of Val for Met at position 251 (Val251Met), which conferred CTX-M increasing resistance to ceftazidime verified by antibiotics susceptibility testing of transconjugants carrying pST3606-2 and steady-state kinetic parameters of CTX-M-261. pST3606-1 is an IncI1-α incompatibility type that shares homology with plasmids of pC-F-164_A-OXA140, pE-T654-NDM-5, p_dm760b_NDM-5, and p_dmcr749c_NDM-5. The conjugation experiment demonstrated that pST3606-2 was successfully transferred to the Escherichia coli recipient C600 with four modules of OriTfinder. CONCLUSION: Plasmid-mediated horizontal transfer plays an important role in blaNDM-5 and blaCTX-M-261 dissemination, which increases the threat to public health due to the resistance to most ß-lactam antibiotics. This is the first report of blaCTX-M-261 and blaNDM-5 in S. 1,4,[5], 12: i: -. The work provides insights into the enzymatic function and demonstrates the ongoing evolution of CTX-M enzymes and confirms urgency to control resistance of S. 1,4,[5], 12: i: -.

CTX-M, TEM, and SHV Genes in Escherichia coli, Klebsiella pneumoniae, and Enterobacter spp Isolated from Hematologic Cancer Patients with Bacteremia in Uganda.

Purpose: We determined the phenotypic resistance to third-generation cephalosporins, phenotypic extended spectrum beta-lactamase (ESBL) prevalence, and genotypic prevalence of ESBL-encoding genes blaCTX-M, blaTEM, and blaSHV in Enterobacteriaceae isolated from hematologic cancer patients with febrile neutropenia and bacteremia at the Uganda Cancer Institute (UCI). Patients and Methods: Blood cultures from hematologic cancer patients with febrile neutropenia were processed in BACTEC 9120. E. coli, K. pneumoniae, and Enterobacter spp. isolates were identified using conventional biochemical methods. Antimicrobial susceptibility tests, phenotypic ESBL characterization, and genotypic characterization of the ESBL-encoding genes blaCTX-M, blaTEM, and blaSHV were determined for pure isolates of E. coli, K. pneumoniae, and Enterobacter spp. Results: Two hundred and two patients were included in the study. Median age of patients was 19 years (IQR: 10-30 years). Majority (N=119, 59%) were male patients. Sixty (30%) of the participants had at least one febrile episode due to Enterobacteriaceae. Eighty-three organisms were isolated with E. coli being predominant (45, 54%). Seventy-nine (95%) Enterobacteriaceae were multidrug resistant. The ESBL phenotype was detected in 54/73 (74%) of Enterobacteriaceae that were resistant to third-generation cephalosporins. A higher proportion of Enterobacteriaceae with ESBL-positive phenotype were resistant to piperacillin-tazobactam (p=0.024), gentamicin (p=0.000), ciprofloxacin (p=0.000), and cotrimoxazole (p=0.000) compared to Enterobacteriaceae, which were sensitive to third-generation cephalosporins. The organisms were more susceptible to carbapenems and chloramphenicol than resistant. ESBL-encoding genes (blaCTX-M, blaTEM, and blaSHV) were detected in 55 (75%) of the 73 Enterobacteriaceae that were resistant to third-generation cephalosporins. BlaCTX-M, was the most common ESBL-encoding gene identified with 50 (91%). Conclusion: ESBL-producing Enterobacteriaceae are a predominant cause of bacteremia in hematologic cancer patients at UCI. The most common ESBL-encoding gene identified in the ESBL-PE was blaCTX-M. Resistance to imipenem and meropenem was low.

In silico analysis of luteolin derivatives as antibacterial agents targeting DNA gyrase and CTX-M-15 extended-spectrum ß-lactamase of Escherichia coli.

Luteolin exhibited antibacterial activity against Escherichia coli and its chemical structure similar to that of ciprofloxacin (CPF) which works by inhibiting DNA gyrase. Filtrate from passion fruit extract containing luteolin and its derivatives could inhibit extended-spectrum ß-lactamase (ESBL)-producing E. coli. Antibacterial compounds that can also inhibit ESBL will be valuable compounds to overcome the problem of resistant bacteria. This study aimed to ensure the potency of luteolin and luteolin derivatives targeting DNA gyrase and ESBL by in silico approach. Docking simulation of ligands L1-L14 was performed using AutoDock Vina, and pharmacokinetics and toxicity (absorption, distribution, metabolism, excretion, and toxicity) profiles were predicted by pKCSM online. The docking result revealed higher binding affinity on DNA gyrase (PDB.1KZN) of 12 luteolin derivatives (energy <-7.6 kcal/mol) compared to CPF and higher affinity (energy <-6.27 kcal/mol) of all compounds than clavulanic acid against ESBL CTX-M-15 (PDB.4HBU). The compounds could be absorbed through the human intestine moderately, which showed low permeability to blood-brain barrier, nontoxic and nonhepatotoxic. The most active luteolin glycoside (L6) is capable to inhibit DNA gyrase and ESBL from E. coli which provided the potential against resistant bacteria and was promoted as lead compounds to be developed further.

Genomic characteristics of two strains of ESBL-producing Klebsiella pneumoniae ST268 isolated from different samples of one patient.

OBJECTIVES: This study reports the whole-genome sequences of two strains of extended-spectrum beta-lactamase (ESBL)-producing and multidrug-resistant (MDR) K. pneumoniae ST268 and explores their acquired antibiotic resistance genes (ARGs) and the mobile genetic elements (MGEs). METHODS: Two strains of K. pneumoniae ST268 were isolated from different samples of one patient. Assessment of antimicrobial susceptibility was performed, and then whole-genome sequencing was conducted. Acquired ARGs, insertion sequences, and transposons harboured by the two strains of K. pneumoniae ST268 were identified, and then the genetic contexts associated with the ARGs were analysed systematically. RESULTS: Two strains of K. pneumoniae ST268 were found to carry the 118.6-kb hybrid IncFIIK:IncQ1:repBR1701 plasmid. All the acquired ARGs carried by the IncF plasmid were found to be situated on the 25.3-kb MDR region bracketed by ISKpn19 and IS26, which was widely present in the plasmids in 14 STs of strains in K. pneumoniae but also in IncF plasmids from Shigella flexneri and Klebsiella quasipneumoniae. Notably, the IncF plasmids harbouring the 25.3-kb MDR region were geographically distributed mainly in China, and the pKP161637-1/pKP160802-1 in our study was the first report on the IncF plasmid carrying the 25.3-kb MDR region bracketed in K. pneumoniae ST268. CONCLUSIONS: Two strains of ESBL-producing K. pneumoniae ST268 with a MDR IncF plasmid were identified in a hospital in China. The ARGs were identified on the 25.3-kb MDR region, bracketed by ISKpn19 and IS26, of the IncF plasmids, which were present not only in the K. pneumoniae but also in the S. flexneri and K. quasipneumoniae.

Epidemiology of Cefotaxime-Hydrolysing ß-Lactamase-Producing Escherichia coli in Children with Diarrhoea Reported Globally between 2012 and 2022.

The global spread of cefotaxime-hydrolysing ß-lactamase (CTX-M)-producing Escherichia coli (E. coli) and its associated impact on paediatric diarrhoeal treatment and management has become a public health concern. This review assessed surveillance studies on CTX-M-producing E. coli associated with diarrhoea in children published between 2012 and 2022 globally. A total of thirty-eight studies were included for data analysis, categorised into continental regions, and tabulated. The majority (68%) of studies were conducted in Asian countries while few studies were conducted in Europe (11%) and Africa (18%), respectively. On the African continent, the majority (11%) of studies were conducted in Northern Africa while no studies were reported in East Africa. On the American continent, 3% of the studies were reported from South America. The studies included were classified into diarrheagenic E. coli (74%; 28/38) and faecal carriage (26%; 10/38). Of all the E. coli pathotypes associated with CTX-M production, EPEC was frequently reported. The prevalence of CTX-M-producing E. coli including the CTX-M-15-producing variants ranged between 1% and 94%. About 37% of the studies generalised the report as blaCTX-M-positive E. coli. The use of sequencing in characterising the CTX-M-producing E. coli was reported in only 32% of all the studies. This review provides information on the epidemiology of CTX-M-15-producing E. coli in paediatric diarrhoea and the extent to which surveillance is being performed. This is relevant in informing clinical practice for the management of diarrhoea as well as the design of future surveillance studies.