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180000.9986Unveiling the ecological landscape of bacterial β-lactam resistance in Delhi-national capital region, India: An emerging health concern. Inappropriate antibiotic use not only amplifies the threat of antimicrobial resistance (AMR), moreover exacerbates the spread of resistant bacterial strains and genes in the environment, underscoring the critical need for effective research and interventions. Our aim is to assess the prevalence and resistance characteristics of β-lactam resistant bacteria (BLRB) and β-lactamase resistant bacterial genes (BLRBGs) under various environmental conditions within Delhi NCR, India. Using a culture-dependent method, we isolated 130 BLRB from 75 different environmental samples, including lakes, ponds, the Yamuna River, agricultural soil, aquatic weeds, drains, dumping yards, STPs, and gaushalas. Tests for antibiotic susceptibility were conducted in addition to phenotypic and genotypic identification of BLs and integron genes. The water and sediment samples recorded an average bacterial abundance of 3.6 × 10(6) CFU/mL and an average ampicillin-resistant bacterial count of 2.2 × 10(6) CFU/mL, which can be considered a potent reservoir of BLRB and BLRBGs. The majority of the BLRB discovered are opportunistic pathogens from the Bacillus, Aeromonas, Pseudomonas, Enterobacter, Escherichia, and Klebsiella genera, with Multiple Antibiotic Resistance (MAR) index ≥0.2 against a wide variety of β-lactams and β-lactamase (BLs) inhibitor combinations. The antibiotic resistance pattern was similar in the case of bacteria isolated from STPs. Meanwhile, bacteria isolated from other sources were diverse in their antibiotic resistance profile. Interestingly, we discovered that 10 isolates of various origins produce both Extended Spectrum BLs and Metallo BLs, as well as found harboring bla(TEM), bla(CTX), bla(OXA), bla(SHV), int-1, and int-3 genes. Enterobacter cloacae (S50/A), a common nosocomial pathogen isolated from Yamuna River sediment samples at Nizamuddin point, possesses three BLRBGs (bla(TEM), bla(CTX), and bla(OXA)) and a MAR index of 1.0, which is a major cause for concern. Therefore, identifying the source, origin and dissemination of BLRB and BLRGs in the environment is of the utmost importance for designing effective mitigation approaches to reduce a load of antimicrobial resistance factors in the environmental settings.202438850900
179810.9986Impacts of Domestication and Veterinary Treatment on Mobile Genetic Elements and Resistance Genes in Equine Fecal Bacteria. Antimicrobial resistance in bacteria is a threat to both human and animal health. We aimed to understand the impact of domestication and antimicrobial treatment on the types and numbers of resistant bacteria, antibiotic resistance genes (ARGs), and class 1 integrons (C1I) in the equine gut microbiome. Antibiotic-resistant fecal bacteria were isolated from wild horses, healthy farm horses, and horses undergoing veterinary treatment, and isolates (9,083 colonies) were screened by PCR for C1I; these were found at frequencies of 9.8% (vet horses), 0.31% (farm horses), and 0.05% (wild horses). A collection of 71 unique C1I(+) isolates (17 Actinobacteria and 54 Proteobacteria) was subjected to resistance profiling and genome sequencing. Farm horses yielded mostly C1I(+) Actinobacteria (Rhodococcus, Micrococcus, Microbacterium, Arthrobacter, Glutamicibacter, Kocuria), while vet horses primarily yielded C1I(+) Proteobacteria (Escherichia, Klebsiella, Enterobacter, Pantoea, Acinetobacter, Leclercia, Ochrobactrum); the vet isolates had more extensive resistance and stronger P(C) promoters in the C1Is. All integrons in Actinobacteria were flanked by copies of IS6100, except in Micrococcus, where a novel IS5 family element (ISMcte1) was implicated in mobilization. In the Proteobacteria, C1Is were predominantly associated with IS26 and also IS1, Tn21, Tn1721, Tn512, and a putative formaldehyde-resistance transposon (Tn7489). Several large C1I-containing plasmid contigs were retrieved; two of these (plasmid types Y and F) also had extensive sets of metal resistance genes, including a novel copper-resistance transposon (Tn7519). Both veterinary treatment and domestication increase the frequency of C1Is in equine gut microflora, and each of these anthropogenic factors selects for a distinct group of integron-containing bacteria. IMPORTANCE There is increasing acknowledgment that a "one health" approach is required to tackle the growing problem of antimicrobial resistance. This requires that the issue is examined from not only the perspective of human medicine but also includes consideration of the roles of antimicrobials in veterinary medicine and agriculture and recognizes the importance of other ecological compartments in the dissemination of ARGs and mobile genetic elements such as C1I. We have shown that domestication and veterinary treatment increase the frequency of occurrence of C1Is in the equine gut microflora and that, in healthy farm horses, the C1I are unexpectedly found in Actinobacteria, while in horses receiving antimicrobial veterinary treatments, a taxonomic shift occurs, and the more typical integron-containing Proteobacteria are found. We identified several new mobile genetic elements (plasmids, insertion sequences [IS], and transposons) on genomic contigs from the integron-containing equine bacteria.202336988354
506520.9985Locus of Heat Resistance (LHR) in Meat-Borne Escherichia coli: Screening and Genetic Characterization. Microbial resistance to processing treatments poses a food safety concern, as treatment tolerant pathogens can emerge. Occasional foodborne outbreaks caused by pathogenic Escherichia coli have led to human and economic losses. Therefore, this study screened for the extreme heat resistance (XHR) phenotype as well as one known genetic marker, the locus of heat resistance (LHR), in 4,123 E. coli isolates from diverse meat animals at different processing stages. The prevalences of XHR and LHR among the meat-borne E. coli were found to be 10.3% and 11.4%, respectively, with 19% agreement between the two. Finished meat products showed the highest LHR prevalence (24.3%) compared to other processing stages (0 to 0.6%). None of the LHR(+)E. coli in this study would be considered pathogens based on screening for virulence genes. Four high-quality genomes were generated by whole-genome sequencing of representative LHR(+) isolates. Nine horizontally acquired LHRs were identified and characterized, four plasmid-borne and five chromosomal. Nine newly identified LHRs belong to ClpK1 LHR or ClpK2 LHR variants sharing 61 to 68% nucleotide sequence identity, while one LHR appears to be a hybrid. Our observations suggest positive correlation between the number of LHR regions present in isolates and the extent of heat resistance. The isolate exhibiting the highest degree of heat resistance possessed four LHRs belonging to three different variant groups. Maintenance of as many as four LHRs in a single genome emphasizes the benefits of the LHR in bacterial physiology and stress response.IMPORTANCE Currently, a "multiple-hurdle" approach based on a combination of different antimicrobial interventions, including heat, is being utilized during meat processing to control the burden of spoilage and pathogenic bacteria. Our recent study (M. Guragain, G. E. Smith, D. A. King, and J. M. Bosilevac, J Food Prot 83:1438-1443, 2020, https://doi.org/10.4315/JFP-20-103) suggests that U.S. beef cattle harbor Escherichia coli that possess the locus of heat resistance (LHR). LHR seemingly contributes to the global stress tolerance in bacteria and hence poses a food safety concern. Therefore, it is important to understand the distribution of the LHRs among meat-borne bacteria identified at different stages of different meat processing systems. Complete genome sequencing and comparative analysis of selected heat-resistant bacteria provide a clearer understanding of stress and heat resistance mechanisms. Further, sequencing data may offer a platform to gain further insights into the genetic background that provides optimal bacterial tolerance against heat and other processing treatments.202133483306
552030.9985Emergence of highly virulent and multidrug-resistant Escherichia coli in breeding sheep with pneumonia, Hainan Province, China. BACKGROUND: Sheep are a rarely raised livestock in Hainan Island, China, because of the unfavorable tropical marine climate. Here, this article reports a severe pneumonia in the sheep breeding and domestication facility caused acute mortality during the winter 2021-2022. METHODS: Six sheep were clinically dissected and histopathologically observed. The bacteria were isolated and cultured by traditional methods and identified by 16S rRNA sequencing. The genotypes, serotypes, virulence genes and antimicrobial resistance genes were analyzed by PCR and whole genome sequencing. The pubMLST website was used for phylogenetic analysis of related strains. Kirby-Bauer disk diffusion method was used for antimicrobial susceptibility test. The antimicrobial susceptibility test standard was referred to the Clinical and Laboratory Standards Institute (CLSI). The virulence of bacteria was detected by mouse infection model. RESULTS: Etiology and histopathology examination of the pneumonia reveled pulmonary abscess and alveolar neutrophilia and pulmonary fibrinous exudates. Escherichia coli was the only bacterial species isolated, primarily from the lungs and blood of the six dead or moribund sheep, a total of 29 E. coli strains were isolated. Antimicrobial resistance profiling shows that all the isolates were resistant to six agents (penicillin, ampicillin, cephalothin, neomycin, erythromycin, and vancomycin) belonging to five classes of antibiotics, classifying them as multi drug resistant (MDR). Furthermore, genotyping analysis revealed all strains were common with 11-17 virulence factors indicating high pathogenicity. The lab mice infection model shows that all strains severely affect the health status particularly weight loss, lethargy, pneumonia and shortly lead to death. The molecular epidemiological analysis indicated most strains share the same genotype as previously reported strains in humans and other farmed animals this suggests a high possibility of cross-species transmission (CST) of virulent and MDR isolates. This CST could be from sheep to humans and other farmed animals or from humans and other farmed animals to sheep. CONCLUSION: Therefore, this study indicates that E. coli is an emerging threat that causes sheep pneumonia in Hainan, and the quarantine of contacts is important to control the spread of virulent E. coli and the transmission of acquired resistance genes between humans and farmed animals such as sheep.202439507338
225440.9984Hospitalized Pets as a Source of Carbapenem-Resistance. The massive and irrational use of antibiotics in livestock productions has fostered the occurrence and spread of resistance to "old class antimicrobials." To cope with that phenomenon, some regulations have been already enforced in the member states of the European Union. However, a role of livestock animals in the relatively recent alerts on the rapid worldwide increase of resistance to last-choice antimicrobials as carbapenems is very unlikely. Conversely, these antimicrobials are increasingly administered in veterinary hospitals whose role in spreading bacteria or mobile genetic elements has not adequately been addressed so far. A cross-sectional study was carried out on 105 hospitalized and 100 non-hospitalized pets with the aim of measuring the prevalence of carbapenem-resistant Gram-negative bacteria (GNB) colonizing dogs and cats, either hospitalized or not hospitalized and estimating the relative odds. Stool samples were inoculated on MacConkey agar plates containing 1 mg/L imipenem which were then incubated aerobically at 37°C ± 1 for 48 h. Isolated bacteria were identified first by Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry and were confirmed by 16S rRNA sequencing. The genetic basis of resistance was investigated using PCR methods, gene or whole genome sequencing (WGS). The prevalence of pets harboring carbapenem-resistant bacteria was 11.4 and 1.0% in hospitalized and not-hospitalized animals, respectively, with an odds ratio of 12.8 (p < 0.01). One pet carried two diverse isolates. Overall, 14 gram-negative non-fermenting bacteria, specifically, one Acinetobacter radioresistens, five Acinetobacter baumannii, six Pseudomonas aeruginosa and two Stenotrophomonas maltophilia were isolated. The Acinetobacter species carried acquired carbapenemases genes encoded by bla (NDM-1) and bla (OXA-23). In contrast, Pseudomonas phenotypic resistance was associated with the presence of mutations in the oprD gene. Notably, inherent carbapenem-resistant isolates of S. maltophilia were also resistant to the first-line recommended chemotherapeutic trimethoprim/sulfamethoxazole. This study estimates the risk of colonization by carbapenem-resistant non-fermenting GNB in pets hospitalized in veterinary tertiary care centers and highlights their potential role in spreading resistance genes among the animal and human community. Public health authorities should consider extending surveillance systems and putting the release of critical antibiotics under more strict control in order to manage the infection/colonization of pets in veterinary settings.201830574124
156650.9984Accumulation of Antibiotic Resistance Genes in Carbapenem-Resistant Acinetobacter baumannii Isolates Belonging to Lineage 2, Global Clone 1, from Outbreaks in 2012-2013 at a Tehran Burns Hospital. The worldwide distribution of carbapenem-resistant Acinetobacter baumannii (CRAB) has become a global concern, particularly in countries where antibiotic prescription is not tightly regulated. However, knowledge of the genomic aspects of CRAB from many parts of the world is still limited. Here, 50 carbapenem-resistant A. baumannii isolates recovered at a single hospital in Tehran, Iran, during several outbreaks in 2012 and 2013 were found to be resistant to multiple antibiotics. They were examined using PCR mapping and multilocus sequence typing (MLST). All Iranian strains belonged to sequence type 328 in the Institut Pasteur MLST scheme (ST328(IP)), a single-locus variant of ST81(IP,) and all Iranian strains contained two carbapenem resistance genes, oxa23 and oxa24. The oxa23 gene is in the transposon Tn2006 in AbaR4, which interrupts the chromosomal comM gene. Phylogenetic analysis using whole-genome sequence (WGS) data for 9 isolates showed that they belonged to the same clade, designated the ST81/ST328 clade, within lineage 2 of global clone 1 (GC1). However, there were two groups that included either KL13 or KL18 at the K locus (KL) for capsular polysaccharide synthesis and either a tet39 or an aadB resistance gene, respectively. The genetic context of the resistance genes was determined, and the oxa24 (OXA-72 variant) and tet39 (tetracycline resistance) genes were each in a pdif module in different plasmids. The aadB gene cassette (which encodes gentamicin, kanamycin, and tobramycin resistance) was harbored by pRAY*, and the aphA6 gene (which encodes amikacin resistance) and sul2 gene (which encodes sulfamethoxazole resistance) were each harbored by a different plasmid. The sequences obtained here will underpin future studies of GC1 CRAB strains from the Middle East region.IMPORTANCE Carbapenem-resistant Acinetobacter baumannii strains are among the most critical antibiotic-resistant bacteria causing hospital-acquired infections and treatment failures. The global spread of two clones has been responsible for the bulk of the resistance, in particular, carbapenem resistance. However, there is a substantial gap in our knowledge of which clones and which specific lineages within each clone are circulating in many parts of the world, including Africa and the Middle East region. This is the first genomic analysis of carbapenem-resistant A. baumannii strains from Iran. All the isolates, from a single hospital, belonged to lineage 2 of global clone 1 (GC1) but fell into two groups distinguished by genes in the locus for capsule biosynthesis. The analysis suggests a potential origin of multiply antibiotic-resistant lineage 2 in the Middle East region and highlights the ongoing evolution of carbapenem-resistant GC1 A. baumannii strains. It will enhance future studies on the local and global GC1 population structure.202032269158
159060.9984Molecular characterization of highly prevalent Escherichia coli and Escherichia marmotae resistant to extended-spectrum cephalosporins in European starlings (Sturnus vulgaris) in Tunisia. European starlings are widespread migratory birds that have already been described as carrying bacteria resistant to extended-spectrum cephalosporins (ESC-R). These birds are well known in Tunisia because they spend the wintertime in this country and are hunted for human consumption. The goal of our study was to estimate the proportion of ESC-R in these birds and to characterize the collected isolates using whole-genome sequencing. Results showed that 21.5% (42/200) of the birds carried either an extended-spectrum beta-lactamase (ESBL) or an acquired AmpC gene. Diverse bla (CTX-M) genes were responsible for the ESBL phenotype, bla (CTX-M-14) being the most prevalent, while only bla (CMY-2) and one bla (CMY-62) were found in AmpC-positive isolates. Likewise, different genetic determinants carried these resistance genes, including IncHI2, and IncF plasmids for bla (CTX-M) genes and IncI1 plasmids for bla (CMY-2) genes. Three chromosomally encoded bla (CTX-M-15) genes were also identified. Surprisingly, species identification revealed a large proportion (32.7%) of Escherichia marmotae isolates. This species is phenotypically indistinguishable from Escherichia coli and has obviously the same capacity to acquire ESC-R genes. Our data also strongly suggest that at least the IncHI2/pST3 plasmid can spread equally between E. coli and E. marmotae. Given the potential transmission routes between humans and animals, either by direct contact with dejections or through meat preparation, it is important to closely monitor antimicrobial resistance in European starlings in Tunisia and to set up further studies to identify the sources of contamination of these birds. IMPORTANCE The One Health concept highlighted knowledge gaps in the understanding of the transmission routes of resistant bacteria. A major interest was shown in wild migratory birds since they might spread resistant bacteria over long distances. Our study brings further evidence that wild birds, even though they are not directly submitted to antibiotic treatments, can be heavily contaminated by resistant bacteria. Our results identified numerous combinations of resistance genes, genetic supports, and bacterial clones that can spread vertically or horizontally and maintain a high level of resistance in the bird population. Some of these determinants are widespread in humans or animals (IncHI2/pST3 plasmids and pandemic clones), while some others are less frequent (atypical IncI1 plasmid and minor clones). Consequently, it is essential to be aware of the risks of transmission and to take all necessary measures to prevent the proportions of resistant isolates from increasing uncontrollably.202337772831
189470.9984Phenotypic and Genotypic Characterization of Multidrug-Resistant Enterobacter hormaechei Carrying qnrS Gene Isolated from Chicken Feed in China. Multidrug resistance (MDR) in Enterobacteriaceae including resistance to quinolones is rising worldwide. The plasmid-mediated quinolone resistance (PMQR) gene qnrS is prevalent in Enterobacteriaceae. However, the qnrS gene is rarely found in Enterobacter hormaechei (E. hormaechei). Here, we reported one multidrug resistant E. hormaechei strain M1 carrying the qnrS1 and bla(TEM-1) genes. This study was to analyze the characteristics of MDR E. hormaechei strain M1. The E. hormaechei strain M1 was identified as Enterobacter cloacae complex by biochemical assay and 16S rRNA sequencing. The whole genome was sequenced by the Oxford Nanopore method. Taxonomy of the E. hormaechei was based on multilocus sequence typing (MLST). The qnrS with the other antibiotic resistance genes were coexisted on IncF plasmid (pM1). Besides, the virulence factors associated with pathogenicity were also located on pM1. The qnrS1 gene was located between insertion element IS2A (upstream) and transposition element ISKra4 (downstream). The comparison result of IncF plasmids revealed that they had a common plasmid backbone. Susceptibility experiment revealed that the E. hormaechei M1 showed extensive resistance to the clinical antimicrobials. The conjugation transfer was performed by filter membrane incubation method. The competition and plasmid stability assays suggested the host bacteria carrying qnrS had an energy burden. As far as we know, this is the first report that E. hormaechei carrying qnrS was isolated from chicken feed. The chicken feed and poultry products could serve as a vehicle for these MDR bacteria, which could transfer between animals and humans through the food chain. We need to pay close attention to the epidemiology of E. hormaechei and prevent their further dissemination. IMPORTANCE Enterobacter hormaechei is an opportunistic pathogen. It can cause infections in humans and animals. Plasmid-mediated quinolone resistance (PMQR) gene qnrS can be transferred intergenus, which is leading to increase the quinolone resistance levels in Enterobacteriaceae. Chicken feed could serve as a vehicle for the MDR E. hormaechei. Therefore, antibiotic-resistance genes (ARGs) might be transferred to the intestinal flora after entering the gastrointestinal tract with the feed. Furthermore, antibiotic-resistant bacteria (ARB) were also excreted into environment with feces, posing a huge threat to public health. This requires us to monitor the ARB and antibiotic-resistant plasmids in the feed. Here, we demonstrated the characteristics of one MDR E. hormaechei isolate from chicken feed. The plasmid carrying the qnrS gene is a conjugative plasmid with transferability. The presence of plasmid carrying antibiotic-resistance genes requires the maintenance of antibiotic pressure. In addition, the E. hormaechei M1 belonged to new sequence type (ST). These data show the MDR E. hormaechei M1 is a novel strain that requires our further research.202235467399
156880.9984Genomic Characteristion of Opportunistic Pathogen Kluyvera Reveals a Novel CTX-M Subgroup. A rising incidence of clinical infections has been caused by Kluyvera, a significant opportunistic pathogen. Meanwhile, Kluyvera acts as an important reservoir of bla(CTX-Ms), which are the dominant genes of class A extended-spectrum β-lactamases (ESBLs). In this work, 60 strains of Kluyvera were subjected to phylogenetic relationship reconstruction, antimicrobial susceptibility testing, and antibiotic resistance genes prediction. All mature bla(CTX-Ms) were gathered to perform subgroup reclassification. The findings demonstrate that Kluyvera has a large gene pool with significant genetic flexibility. Notably, 25% of strains showed simultaneous detection of ESBLs and carbapenem resistance genes. The genotypes of fourteen novel bla(CTX-Ms) were identified. A new subgroup classification approach for bla(CTX-Ms) was defined by using 20 amino acid site variants, which could split bla(CTX-Ms) into 10 subgroups. The results of the subgroup division were consistent with the phylogenetic clustering. More significantly, we proposed a novel bla(CTX-M) subgroup, KLUS, that is chromosomally encoded in K. sichuanensis and the new species put forward in this study, showing amino acid differences from the currently known sequences. Cloning and transformation tests demonstrated that the recipient bacteria had a robust phenotype of cefotaxime resistance. Closely related Kluyvera species had bla(CTX-Ms) in the same subgroup. Our research lays the groundwork for a deeper comprehension of Kluyvera and emphasizes how important a bla(CTX-M) reservoir it is. We provide an update on bla(CTX-M) subgroups reclassification from the aspects of phylogenetic relationship, amino acid differences, and the new subgroup KLUS, which needs to be strengthen monitored due to its strong resistance phenotype to cefotaxime.202338137980
183090.9984Shifts in bla genes and Class 1 integron prevalence in beta-lactamase-producing bacteria before and after the COVID-19 pandemic in Mendoza, Argentina. This study analyzes the molecular epidemiology of bla genes and Class 1 integron in broad-spectrum beta-lactamase (BSBL) and extended-spectrum beta-lactamase (ESBL) producing strains of bacteria isolated from clinical samples of hospitalized and ambulatory patients before and after the COVID-19 pandemic. Isolates obtained in two periods were compared: the first corresponding to the years November 2019-March 2020, and the second to the years November 2021-April 2022. We evaluate changes in resistance patterns of antibiotics associated with pressures on the healthcare system and social lockdowns. A total of 156 isolates were analyzed: 78 from the first period (61 hospitalized, 17 ambulatory) and 78 from the second period (47 hospitalized, 31 ambulatory). Escherichia coli and Klebsiella pneumoniae were the predominant bacterial species, representing 85% of the isolates in both periods. The frequency of ambulatory ESBL-producing isolates increased significantly, from 22% (17/78) to 40% (31/78; P < 0.01) in the second period. The prevalence of bla(SHV) increased from 24% (19/78) to 72% (56/78; P < 0.01) in the second period, while the bla(CTX-M-2) group, absent in the first period, was detected in 43% (34/78) of isolates from the second period. Strains from the second period exhibited greater genetic complexity, with an increased prevalence of combinations involving three or more bla genes, including isolates carrying up to five of such genes. Class 1 integron showed a strong correlation with resistance to ciprofloxacin and trimethoprim-sulfamethoxazole. The gene bla(OXA-1), previously associated with resistance to beta-lactamase inhibitors, did not show a clear pattern in the second period.IMPORTANCEAntimicrobial resistance associated with the production of extended-spectrum beta-lactamase (ESBL) represents a critical global health challenge, particularly due to the limited development of new antibiotics. This is the first report from Argentina's central-west region examining the prevalence of beta-lactamase-encoding genes, providing a framework for future research. Our findings reveal a significant increase in bacteria with the ESBL phenotype, particularly among ambulatory populations post-pandemic, suggesting a concerning spread of multidrug-resistant bacteria outside hospital environments. This could compromise empirical antibiotic treatments for ambulatory patients, increasing the risk of severe complications. Our results highlight the urgent need for ongoing surveillance to detect virulent strains before clonal spread or horizontal gene transfer occurs in the community. They also emphasize the importance of strategies to ensure the prudent use of antimicrobials and mitigate the increasing prevalence of resistance genes, which threatens the effectiveness of current therapeutic options.202540662585
1683100.9984Colonization of a hand washing sink in a veterinary hospital by an Enterobacter hormaechei strain carrying multiple resistances to high importance antimicrobials. BACKGROUND: Hospital intensive care units (ICUs) are known reservoirs of multidrug resistant nosocomial bacteria. Targeted environmental monitoring of these organisms in health care facilities can strengthen infection control procedures. A routine surveillance of extended spectrum beta-lactamase (ESBL) producers in a large Australian veterinary teaching hospital detected the opportunistic pathogen Enterobacter hormaechei in a hand washing sink of the ICU. The organism persisted for several weeks, despite two disinfection attempts. Four isolates were characterized in this study. METHODS: Brilliance-ESBL selective plates were inoculated from environmental swabs collected throughout the hospital. Presumptive identification was done by conventional biochemistry. Genomes of multidrug resistant Enterobacter were entirely sequenced with Illumina and Nanopore platforms. Phylogenetic markers, mobile genetic elements and antimicrobial resistance genes were identified in silico. Antibiograms of isolates and transconjugants were established with Sensititre microdilution plates. RESULTS: The isolates possessed a chromosomal Tn7-associated silver/copper resistance locus and a large IncH12 conjugative plasmid encoding resistance against tellurium, arsenic, mercury and nine classes of antimicrobials. Clusters of antimicrobial resistance genes were associated with class 1 integrons and IS26, IS903 and ISCR transposable elements. The blaSHV-12, qnrB2 and mcr-9.1 genes, respectively conferring resistance to cephalosporins, quinolones and colistin, were present in a locus flanked by two IS903 copies. ESBL production and enrofloxacin resistance were confirmed phenotypically. The isolates appeared susceptible to colistin, possibly reflecting the inducible nature of mcr-9.1. CONCLUSIONS: The persistence of this strain in the veterinary hospital represented a risk of further accumulation and dissemination of antimicrobial resistance, prompting a thorough disinfection of the ICU. The organism was not recovered from subsequent environmental swabs, and nosocomial Enterobacter infections were not observed in the hospital during that period. This study shows that targeted routine environmental surveillance programs to track organisms with major resistance phenotypes, coupled with disinfection procedures and follow-up microbiological cultures are useful to control these risks in sensitive areas of large veterinary hospitals.202033087168
1672110.9984Colonization of extended-spectrum beta-lactamase-producing bacteria in healthy pregnant women and its impact on perinatal care: A cross-sectional study. BackgroundColonization of extended-spectrum beta-lactamase (ESBL)-producing organisms is increasing becoming more frequent not only in hospitalized patients but also in healthy individuals. Although these bacteria are thought to be transmitted to newborns on their way through the birth canal, molecular evidence for this is scarce. In this study, we aimed to survey the current prevalence of resistant bacterial colonization in this area by examining the colonization carriage of this organism before and after delivery.MethodsWe examined the colonization rate of ESBL-producing bacteria in healthy pregnant women, the colonization rate in newborns, and the transmission rate from pregnant women who are carriers of the bacteria to their newborns. We also performed resistance gene and similarity analyses for each strain in pairs of mother-child carriers.ResultsOf 494 pregnant women, 33 carried ESBL-producing bacteria, all of whom were identified as Escherichia coli. The colonization carriage rate among pregnant women was 6.7%. Among newborns, the rate rose from 1.0% immediately after birth to 6.9% at the one-month checkup. Furthermore, of the 13 strains detected among mothers and children, 10 pairs had matching resistance genes.ConclusionsSome ESBL-producing bacterial carriers exist even among healthy pregnant women, and about half of them go on to infect their newborns. However, routes of transmission beyond vertical transmission cannot be ruled out. Therefore, it is important to promote infection control in the healthcare environment and in the families of newborns, as well as antimicrobial stewardship among pregnant women.202540152933
1696120.9984Assessment of the presence of Acinetobacter spp. resistant to β-lactams in commercial ready-to-eat salad samples. Acinetobacter baumannii is a well-known nosocomial infection causing agent. However, other Acinetobacter spp. have also been implicated in cases of human infection. Additionally, these bacteria are known for the development of antibiotic resistance thus making the treatment of the infections they cause, challenging. Due to their relevance in clinical setups less attention has been paid to their presence in foods, and its relation with infection/dissemination routes. In the current study commercial Ready-To-Eat (RTE) salads were analyzed seeking for antibiotic resistant Acinetobacter spp. A preliminary screening allowed us to recover Gram-negative bacteria resistant to β - lactams using cefotaxime, third generation cephalosporins, as the selective agent, and this was followed by identification with CHROMagar™ Acinetobacter and 16S rDNA sequencing. Finally, the isolates identified as Acinetobacter spp. were reanalyzed by PCR to determine the presence of nine potential Extended Spectrum β Lactamases (ESBL). Two commercial RTE salad brands were included in the study (2 batches per brand and 8 samples of each batch making a total of 32 independent samples), and compared against an organic lettuce. High concentrations of β - lactam, resistant bacteria were found in all the samples tested (5 log CFU/g). Additionally, 209 isolates were phenotypically characterized on CHROMagar Acinetobacter. Finally, PCR analysis identified the presence of different ESBL genes, being positive for blaACC, blaSHV, blaDHA and blaVEB; out of these, blaACC was the most prevalent. None of the isolates screened were positive for more than one gene. To conclude, it is important to highlight the fact that pathogenic species within the genus Acinetobacter spp., other than A. baumannii, have been identified bearing resistance genes not typically associated to these microorganisms highlight the importance of continuous surveillance.202438049272
1897130.9984Plasmid-based replicon typing: Useful tool in demonstrating the silent pandemic of plasmid-mediated multi-drug resistance in Enterobacterales. BACKGROUND: Multi-drug resistant Enterobacterales increasingly isolated in hospital settings have a significant impact on therapy and overall treatment costs. Conjugative plasmids carrying relevant resistance genes have been described as the most frequent mechanism of acquisition and spread of resistance. PCR-based replicon typing (PBRT) is a method for plasmid identification and incompatibility typing which helps detect the presence of plasmid families in these bacteria. This study was undertaken to provide an insight into the prevalence of resistance plasmids in MDR Enterobacterales in our tertiary care setting. METHODS: A selection of one hundred multi-drug resistant Enterobacterale isolates sourced from clinical samples were subjected to PCR-based replicon typing. RESULTS: A total of 21 plasmid replicon types were detected from 85% of the isolates out of the 28 families described in literature. Majority of the isolates (54%) showed three or more replicons. IncF was the most frequent plasmid family detected with FIA being the most common replicon type (43%) followed by FII (29%) and FIB (28%) replicons. Among the IncX plasmid family, X3 replicon was the commonest (14%). IncF and IncX plasmid families are known to carry a large spectrum of resistance genes. CONCLUSION: The presence of these plasmids engenders emergent steps to be taken for prevention of their transmission in the form of strict infection control measures in the hospital and adoption of novel methods of plasmid curing to eliminate the plasmids from these organisms rendering them susceptible to the currently used antimicrobials.202540463599
1842140.9984Emergence of mcr-9.1 in Extended-Spectrum-β-Lactamase-Producing Clinical Enterobacteriaceae in Pretoria, South Africa: Global Evolutionary Phylogenomics, Resistome, and Mobilome. Extended-spectrum-β-lactamase (ESBL)-producing Enterobacteriaceae are critical-priority pathogens that cause substantial fatalities. With the emergence of mobile mcr genes mediating resistance to colistin in Enterobacteriaceae, clinicians are now left with few therapeutic options. Eleven clinical Enterobacteriaceae strains with resistance to cephems and/or colistin were genomically analyzed to determine their resistomes, mobilomes, and evolutionary relationships to global strains. The global phylogenomics of mcr genes and mcr-9.1-bearing genomes were further analyzed. Ten isolates were ESBL positive. The isolates were multidrug resistant and phylogenetically related to global clones but distant from local strains. Multiple resistance genes, including bla (CTX-M-15) bla (TEM-1), and mcr-9.1, were found in single isolates; ISEc9, IS19, and Tn3 transposons bracketed bla (CTX-M-15) and bla (TEM-1) Common plasmid types included IncF, IncH, and ColRNAI. mcr-9 was of close sequence identity to mcr-3, mcr-5, mcr-7, mcr-8, and mcr-10. Genomes bearing mcr-9.1 clustered into six main phyletic groups (A to F), with those of this study belonging to clade B. Enterobacter species and Salmonella species are the main hosts of mcr-9.1 globally, although diverse promiscuous plasmids disseminate mcr-9.1 across different bacterial species. Emergence of mcr-9.1 in ESBL-producing Enterobacteriaceae in South Africa is worrying, due to the restricted therapeutic options. Intensive One Health molecular surveillance might discover other mcr alleles and inform infection management and antibiotic choices.IMPORTANCE Colistin is currently the last-resort antibiotic for difficult-to-treat bacterial infections. However, colistin resistance genes that can move from bacteria to bacteria have emerged, threatening the safe treatment of many bacterial infections. One of these genes, mcr-9.1, has emerged in South Africa in bacteria that are multidrug resistant, further limiting treatment options for clinicians. In this work, we show that this new gene is disseminating worldwide through Enterobacter and Salmonella species through multiple plasmids. This worrying observation requires urgent action to prevent further escalation of this gene in South Africa and Africa.202032430406
1856150.9984Whole-Genome Sequencing-Based Species Classification, Multilocus Sequence Typing, and Antimicrobial Resistance Mechanism Analysis of the Enterobacter cloacae Complex in Southern China. Members of the Enterobacter cloacae complex (ECC) are important opportunistic nosocomial pathogens that are associated with a great variety of infections. Due to limited data on the genome-based classification of species and investigation of resistance mechanisms, in this work, we collected 172 clinical ECC isolates between 2019 and 2020 from three hospitals in Zhejiang, China and performed a retrospective whole-genome sequencing to analyze their population structure and drug resistance mechanisms. Of the 172 ECC isolates, 160 belonged to 9 classified species, and 12 belonged to unclassified species based on ANI analysis. Most isolates belonged to E. hormaechei (45.14%) followed by E. kobei (13.71%), which contained 126 STs, including 62 novel STs, as determined by multilocus sequence typing (MLST) analysis. Pan-genome analysis of the two ECC species showed that they have an "open" tendency, which indicated that their Pan-genome increased considerably with the addition of new genomes. A total of 80 resistance genes associated with 11 antimicrobial agent categories were identified in the genomes of all the isolates. The most prevailing resistance genes (12/29, 41.38%) were related to β-lactams followed by aminoglycosides. A total of 247 β-lactamase genes were identified, of which the bla(ACT) genes were the most dominant (145/247, 58.70%), followed by the bla(TEM) genes (21/247, 8.50%). The inherent ACT type β-lactamase genes differed among different species. bla(ACT-2) and bla(ACT-3) were only present in E. asburiae, while bla(ACT-9), bla(ACT-12), and bla(ACT-6) exclusively appeared in E. kobei, E. ludwigii, and E. mori. Among the six carbapenemase-encoding genes (bla(NDM-1), bla(NDM-5), bla(IMP-1), bla(IMP-4), bla(IMP-26), and bla(KPC-2)) identified, two (bla(NDM-1) and bla(IMP-1)) were identified in an ST78 E. hormaechei isolate. Comparative genomic analysis of the carbapenemase gene-related sequences was performed, and the corresponding genetic structure of these resistance genes was analyzed. Genome-wide molecular characterization of the ECC population and resistance mechanism would offer valuable insights into the effective management of ECC infection in clinical settings. IMPORTANCE The presence and emergence of multiple species/subspecies of ECC have led to diversity and complications at the taxonomic level, which impedes our further understanding of the epidemiology and clinical significance of species/subspecies of ECC. Accurate identification of ECC species is extremely important. Also, it is of great importance to study the carbapenem-resistant genes in ECC and to further understand the mechanism of horizontal transfer of the resistance genes by analyzing the surrounding environment around the genes. The occurrence of ECC carrying two MBL genes also indicates that the selection pressure of bacteria is further increased, suggesting that we need to pay special attention to the emergence of such bacteria in the clinic.202236350178
1870160.9984Novel Insights into bla(GES) Mobilome Reveal Extensive Genetic Variation in Hospital Effluents. Mobile genetic elements contribute to the emergence and spread of multidrug-resistant bacteria by enabling the horizontal transfer of acquired antibiotic resistance among different bacterial species and genera. This study characterizes the genetic backbone of bla(GES) in Aeromonas spp. and Klebsiella spp. isolated from untreated hospital effluents. Plasmids ranging in size from 9 to 244 kb, sequenced using Illumina and Nanopore platforms, revealed representatives of plasmid incompatibility groups IncP6, IncQ1, IncL/M1, IncFII, and IncFII-FIA. Different GES enzymes (GES-1, GES-7, and GES-16) were located in novel class 1 integrons in Aeromonas spp. and GES-5 in previously reported class 1 integrons in Klebsiella spp. Furthermore, in Klebsiella quasipneumoniae, bla(GES-5) was found in tandem as a coding sequence that disrupted the 3' conserved segment (CS). In Klebsiella grimontii, bla(GES-5) was observed in two different plasmids, and one of them carried multiple IncF replicons. Three Aeromonas caviae isolates presented bla(GES-1), one Aeromonas veronii isolate presented bla(GES-7), and another A. veronii isolate presented bla(GES-16). Multilocus sequence typing (MLST) analysis revealed novel sequence types for Aeromonas and Klebsiella species. The current findings highlight the large genetic diversity of these species, emphasizing their great adaptability to the environment. The results also indicate a public health risk because these antimicrobial-resistant genes have the potential to reach wastewater treatment plants and larger water bodies. Considering that they are major interfaces between humans and the environment, they could spread throughout the community to clinical settings. IMPORTANCE In the "One Health" approach, which encompasses human, animal, and environmental health, emerging issues of antimicrobial resistance are associated with hospital effluents that contain clinically relevant antibiotic-resistant bacteria along with a wide range of antibiotic concentrations, and lack regulatory status for mandatory prior and effective treatment. bla(GES) genes have been reported in aquatic environments despite the low detection of these genes among clinical isolates within the studied hospitals. Carbapenemase enzymes, which are relatively unusual globally, such as GES type inserted into new integrons on plasmids, are worrisome. Notably, K. grimontii, a newly identified species, carried two plasmids with bla(GES-5), and K. quasipneumoniae carried two copies of bla(GES-5) at the same plasmid. These kinds of plasmids are primarily responsible for multidrug resistance among bacteria in both clinical and natural environments, and they harbor resistant genes against antibiotics of key importance in clinical therapy, possibly leading to a public health problem of large proportion.202235880869
5621170.9984Comparative Genomics of DH5α-Inhibiting Escherichia coli Isolates from Feces of Healthy Individuals Reveals Common Co-Occurrence of Bacteriocin Genes with Virulence Factors and Antibiotic Resistance Genes. Background/Objectives: The presence of multi-drug-resistant (MDR) bacteria in healthy individuals poses a significant public health concern, as these strains may contribute to or even facilitate the dissemination of antibiotic resistance genes (ARGs) and virulence factors. In this study, we investigated the genomic features of antimicrobial-producing Escherichia coli strains from the gut microbiota of healthy individuals in Singapore. Methods: Using a large-scale screening approach, we analyzed 3107 E. coli isolates from 109 fecal samples for inhibitory activity against E. coli DH5α and performed whole-genome sequencing on 37 representative isolates. Results: Our findings reveal genetically diverse strains, with isolates belonging to five phylogroups (A, B1, B2, D, and F) and 23 unique sequence types (STs). Bacteriocin gene clusters were widespread (92% of isolates carried one or more bacteriocin gene clusters), with colicins and microcins dominating the profiles. Notably, we identified an hcp-et3-4 gene cluster encoding an effector linked to a Type VI secretion system. Approximately 40% of the sequenced isolates were MDR, with resistance for up to eight antibiotic classes in one strain (strain D96). Plasmids were the primary vehicles for ARG dissemination, but chromosomal resistance determinants were also detected. Additionally, over 55% of isolates were classified as potential extraintestinal pathogenic E. coli (ExPEC), raising concerns about their potential pathogenicity outside the intestinal tract. Conclusions: Our study highlights the co-occurrence of bacteriocin genes, ARGs, and virulence genes in gut-residing E. coli, underscoring their potential role in shaping microbial dynamics and antibiotic resistance. While bacteriocin-producing strains show potential as probiotic alternatives, careful assessment of their safety and genetic stability is necessary for therapeutic applications.202541009839
1821180.9984Emergence and dissemination of bla(KPC-31) and bla(PAC-2) among different species of Enterobacterales in Colombia: a new challenge for the microbiological laboratories. Ceftazidime/avibactam (CZA) is a promising treatment option for infections caused by carbapenem-resistant Enterobacterales (CRE). However, CZA resistance is increasingly reported worldwide, largely due to the emergence of KPC variants and increase of metallo-β-lactamases (MBL). This study describes the mechanisms associated with CZA resistance in circulating Enterobacterales isolates from Colombia, highlighting the challenge this represents for microbiological identification. Between 2021 and 2024, 68 CZA-resistant Enterobacterales isolates were identified by automated methods in seven Colombian cities. Resistance to CZA was subsequently confirmed by broth microdilution and E-test. Carbapenemase production was evaluated using phenotypic tests, such as the mCIM test, Carba NP, lateral flow assay, and qPCR (bla(KPC), bla(NDM), bla(VIM), bla(IMP), and bla(OXA-48)). Whole-genome sequencing was performed on 15 isolates that tested negative for MBL genes. Whole-genome sequencing of these 15 isolates revealed a variety of resistance determinants: six isolates harbored bla(KPC-31), one bla(KPC-33), one bla(KPC-8), five harbored bla(PAC-2), and two co-harbored bla(PAC-2) and bla(KPC-2). Notably, bla(PAC-2) was located on an IncQ plasmid. However, some of these variants were not detected by phenotypic assays, likely due to their low or undetectable carbapenemase activity. CZA resistance in non-MBL producing Enterobacterales in Colombia is primarily mediated by the presence of bla(KPC-31) and emergence of bla(PAC-2). These resistance mechanisms pose significant diagnostic, therapeutic, and epidemiological challenges, as they frequently go undetected by conventional microbiological methods. In this context, enhanced molecular surveillance and improved diagnostic strategies are urgently needed to enable early detection, guide antimicrobial therapy, and support infection control and stewardship efforts.IMPORTANCEAntibiotic resistance is a serious global health threat. Ceftazidime/avibactam (CZA) is a key treatment option for multidrug-resistant (MDR) Enterobacterales often used when other antibiotics fail. However, bacteria are now developing resistance to this drug as well, making infections increasingly difficult to treat. In this study, we examined CZA-resistant bacteria from multiple cities in Colombia and found uncommon resistance genes across several bacterial species. These genes are frequently missed, as they often do not test positive due to the limitations of most routinely used laboratory tests. Importantly, some of these genes can be transferred between bacteria, increasing the likelihood of indiscriminate dissemination in the hospital setting. Therefore, our findings highlight the urgent need for improved diagnostic tools and molecular surveillance. Early detection will help physicians select effective treatments quickly and prevent the wider dissemination of these MDR-resistant bacteria.202541070989
5741190.9984Multidrug-Resistant Klebsiella variicola Isolated in the Urine of Healthy Bovine Heifers, a Potential Risk as an Emerging Human Pathogen. Klebsiella variicola, a member of Klebsiella pneumoniae complex, is found to infect plants, insects, and animals and is considered an emerging pathogen in humans. While antibiotic resistance is often prevalent among K. variicola isolates from humans, this has not been thoroughly investigated in isolates from nonhuman sources. Prior evidence suggests that K. variicola can be transmitted between agricultural products as well as between animals, and the use of antibiotics in agriculture has increased antibiotic resistance in other emerging pathogens. Furthermore, in animals that contain K. variicola as a normal member of the rumen microbiota, the same bacteria can also cause infections, such as clinical mastitis in dairy cows. Here, we describe K. variicola UFMG-H9 and UFMG-H10, both isolated from the urine of healthy Gyr heifers. These two genomes represent the first isolates from the urine of cattle and exhibit greater similarity with strains from the human urinary tract than isolates from bovine fecal or milk samples. Unique to the UFMG-H9 genome is the presence of flagellar genes, the first such observation for K. variicola. Neither of the sampled animals had symptoms associated with K. variicola infection, even though genes associated with virulence and antibiotic resistance were identified in both strains. Both strains were resistant to amoxicillin, erythromycin, and vancomycin, and UFMG-H10 is resistant to fosfomycin. The observed resistances emphasize the concern regarding the emergence of this species as a human pathogen given its circulation in healthy livestock animals. IMPORTANCE Klebsiella variicola is an opportunistic pathogen in humans. It also has been associated with bovine mastitis, which can have significant economic effects. While numerous isolates have been sequenced from human infections, only 12 have been sequenced from cattle (fecal and milk samples) to date. Recently, we discovered the presence of K. variicola in the urine of two healthy heifers, the first identification of K. variicola in the bovine urinary tract and the first confirmed K. variicola isolate encoding for flagella-mediated motility. Here, we present the genome sequences and analysis of these isolates. The bovine urinary genomes are more similar to isolates from the human urinary tract than they are to other isolates from cattle, suggesting niche specialization. The presence of antibiotic resistance genes is concerning, as prior studies have found transmission between animals. These findings are important to understand the circulation of K. variicola in healthy livestock animals.202235416681