Pulsed-field Gel Electrophoresis Patterns Research Articles

Pathogenic characteristics of an aggregated diarrhea event caused by Plesiomonas shigelloides from stream.

This study aimed to investigate the cause of a foodborne disease outbreak in Huzhou on August 14, 2023. Multiple enteropathogens were detected using FilmArray, and the pathogen was subsequently isolated and cultured from anal swabs of the cases and stream water. The isolated strains were identified using VITEK MS, and antimicrobial susceptibility test, pulsed field gel electrophoresis (PFGE) molecular typing, and whole genome sequencing (WGS) were performed on the isolates of Plesiomonas shigelloides. Gene annotation and sequence alignment were used to analyze the virulence genes and drug resistance genes of the strains. A phylogenetic tree was constructed based on single nucleotide polymorphism (SNP), and homology analysis was conducted to trace the origin of P. shigelloides. A total of 7 strains of P.shigelloides were isolated, with 3 from stream water and 4 from anal swabs. All 7 strains exhibited the same PFGE pattern and showed resistance to amikacin, trimethoprim-sulfamethoxazole, chloramphenicol, tetracycline, cefazolin, streptomycin, and florfenicol. The isolated strains carried the same resistance genes and virulence factors. In the sequences of the isolated strains from this outbreak, 11 mutation sites were detected. The phylogenetic tree based on SNP sites showed that these strains were homologous. This foodborne disease outbreak caused by P.shigelloides was the first reported in Huzhou. WGS can be used as a complementary method to PFGE for epidemiological investigations of disease outbreaks.

Antibiotic-Resistance Profiles and Genetic Diversity of Shigella Isolates in China: Implications for Control Strategies.

The urgent need for comprehensive and systematic analyses of Shigella as the key pathogen led us to meticulously explore the epidemiology and molecular attributes of Shigella isolates. Accordingly, we procured 24 isolates (10 from Xinjiang and 14 from Wuhan, China) and performed serotype identification and antimicrobial susceptibility testing. Resistance gene detection and homology analysis by polymerase chain reaction and pulsed-field gel electrophoresis (PFGE), respectively, were performed for genetic diversity analysis. All isolates were identified as Shigella flexneri, with 70% (35.4-91.9%) and 30% (8.1-64.6%) of the Xinjiang isolates and 85.7% (56.2-97.5%) and 14.3% (2/14, 2.5-43.9%) of the Wuhan isolates belonging to serotype 2a and serotype 2b, respectively. All isolates displayed resistance to at least two antibiotics and complete resistance to ampicillin. Multidrug resistance (MDR) was recorded in 70.8% (48.8-86.6%) of isolates, with Xinjiang isolates exhibiting relatively higher resistance to ampicillin-sulbactam, piperacillin, ceftriaxone, and aztreonam. Conversely, Wuhan isolates displayed higher MDR and resistance to tetracycline, ciprofloxacin, levofloxacin, and cefepime relative to Xinjiang isolates. Molecular scrutiny of antibiotic-resistance determinants revealed that blaTEM was the main mechanism of ampicillin resistance, blaCTX-M was the main gene for resistance to third- and fourth-generation cephalosporins, and tetB was the predominant gene associated with tetracycline resistance. Four Xinjiang and seven Wuhan isolates shared T1-clone types (>85%), and two Xinjiang and one Wuhan isolates were derived from the T6 clone with a high similarity of 87%. Six PFGE patterns (T1, T2, T5, T6-3, T8, and T10) of S. flexneri were associated with MDR. Thus, there is a critical need for robust surveillance and control strategies in managing Shigella infections, along with the development of targeted interventions and antimicrobial stewardship programs tailored to the distinct characteristics of Shigella isolates in different regions of China.

Flagella Phenotypic Variations of ST34 Type Salmonella Typhimurium and Variants.

Salmonella enterica serovar Typhimurium and its variants are the most common serotypes of human salmonellosis cases. Serotyping Salmonella Typhimurium and its variants has always been challenging. Our previous work found that among 14 Salmonella Typhimurium and variant strains, some different antigenic formulas had 100% pulsed-field gel electrophoresis (PFGE) similarity. The 14 strains were sorted into 3 groups; in each group, the different antigenic formulas had the same PFGE patterns. This phenomenon suggested that different antigenic formula identification might originate from a common ancestor subtyped by PFGE. To assess whether the serotyping method on Salmonella Typhimurium and variant strains reflected the genetic relationship, we improved the discrimination for the phylogenetic relationship among the 14 Salmonella Typhimurium and variant strains using Fourier-transform infrared spectroscopy (FTIR) and whole-genome multilocus sequence typing (wgMLST). We compared the wgMLST assay of 14 Salmonella Typhimurium and variant strains from this study with 50 public ST34 strain data of Salmonella Typhimurium and variant strains. We also compared flagella (H antigen)-related genes based on the whole genome of 14 strains and the other 293 ST34 public database for further understanding of this question. The phylogenetic results (PFGE) showed no regularity between the antigenic formulas and genotypes. The results of the higher discrimination power assays (FTIR and whole-genome multilocus sequence typing) also showed no regularity between the antigenic formulas and genotypes (or phenotypes). The 58 flagella encoding genes of different antigenic formulas were sorted into 13 patterns. However, a similar phenomenon was found: the same flagella encoding gene patterns could express different antigenic formulas. In conclusion, there is no consistency between the antigenic formulas and phylogenetic relationships among ST34 Salmonella Typhimurium and variant strains, even in flagella antigenic formula and flagella encoding genes.

Case reports involving coinfection with Avibacterium paragallinarum and Ornithobacterium rhinotracheale in broiler chickens and Avibacterium endocarditis in broiler breeding hens in Poland

ABSTRACT The study describes three clinical cases of infection with Avibacterium spp.. In case no. 1, respiratory clinical signs and high mortality (0.7–4.2% daily; total 21.2%) in Ross 308 broiler chickens were shown to be caused by coinfection with sequence type 9 of O. rhinotracheale presumptive serotype A and A. paragallinarum presumptive serotype B. The identical (pulsed-field gel electrophoresis) restriction pattern (pulsotype) of seven A. paragallinarum isolates indicated that infectious coryza in broilers was caused by the same clone. In cases 2 and 3, sudden increased deaths in Ross 308 broiler breeders (especially males) with lesions in the endocardium (valvular or mural endocarditis) were shown to be caused by A. endocarditis. Among nine antibiotics tested, florfenicol was the only antibiotic to which all A. paragallinarum and O. rhinotracheale isolates were susceptible. Out of the eight antibiotics tested, 11 A. endocarditis isolates from both clinical cases of infective endocarditis were susceptible to penicillin, amoxicillin, doxycycline and florfenicol. The A. endocarditis isolates tested in both clinical cases had different PFGE patterns (pulsotypes), but identical within a case. The causes of infectious coryza and infective endocarditis in the cases presented have not been determined. In the prevention of infectious diseases in large-scale livestock farming, it is very important to follow the rules of biosecurity.

Pseudomonas aeruginosa from river water: antimicrobial resistance, virulence and molecular typing.

Pseudomonas aeruginosa isolates were recovered from surface river water samples in La Rioja region (Spain) to characterise their antibiotic resistance, molecular typing and virulence mechanisms. Fifty-two P. aeruginosa isolates were isolated from 15 different water samples (45.4%) and belonged to 23 different pulsed-field electrophoresis (PFGE) patterns. All isolates were susceptible to all antibiotics tested, except one carbapenem-resistant P. aeruginosa that showed a premature stop codon in OprD porin. Twenty-two sequence types (STs) (six new ones) were detected among 29 selected P. aeruginosa (one strain with a different PFGE pattern per sample), with ST274 (14%) being the most frequent one. O:6 and O:3 were the predominant serotypes (31%). Seven virulotypes were detected, being 59% exoS-exoY-exoT-exoA-lasA-lasB-lasI-lasR-rhlAB-rhlI-rhlR-aprA-positive P. aeruginosa. It is noteworthy that the exlA gene was identified in three strains (10.3%), and the exoU gene in seven (24.1%), exoS in 18 (62.1%), and both exoS and exoU genes in one strain. High motility ranges were found in these strains. Twenty-seven per cent of strains produced more biofilm biomass, 90% more pyorubin, 83% more pyocyanin and 65.5% more than twice the elastase activity compared with the PAO1 strain. These results highlight the importance of rivers as temporary reservoirs and sources of P. aeruginosa transmission, and show the importance of their epidemiological surveillance in the environment.

Antimicrobial Resistance and Molecular Characterization of Salmonella Rissen Isolated in China During 2008-2019.

This study aimed to provide epidemiological features of Salmonella enterica serovar Rissen, determine antimicrobial susceptibility, virulence gene profiles, and describe the potential association of S. Rissen from different sources in China. During 2008-2019, a total of non-repetitive 228 S. Rissen isolates were collected from human, animals and environment in China. The antimicrobial susceptibility test, screening of antimicrobial and virulence genes by PCR, and pulsed-field gel electrophoresis (PFGE) were performed. Among the 154 isolates from human, the majority of the cases (80.5%) occurred in summer, and S. Rissen was mainly detected in people aged 21-40 (37.7%) and 41-60 (28.6%) years old, and 74 non-human source S. Rissen strains were identified, with pork being the most common source. About 93.4% isolates were resistant to at least one of the 12 tested antimicrobial agents, and high frequencies of resistance were observed for tetracyclines (91.2%), trimethoprim-sulfamethoxazole (74.1%) and ampicillin (67.5%). A total of 171 (75%) isolates were resistant to at least three categories of antimicrobials, and the most common resistance profile was Tetracycline(s)-β-Lactams-Sulfonamides. The resistance rates to chloramphenicol, quinolones and sulfafurazole were significantly higher in strains isolated from human compared to non-human source strains. Among these isolates, the β-Lactams resistance was mainly associated with gene blaTEM (54.7%), sulfonamide resistance with sul2 (45.7%) and sul3 (54.3%), tetracycline resistance with tetA (81.3%). All the isolates harbored virulence genes hilA, sopB, sciN, stn and ssrB, and most of them harbored ssaQ (98.7%), mgtC (98.7%) and invA (98.2%). The majority (91.7%) of S. Rissen isolates showed high similarity (>80%) with each other in PFGE patterns and came from human, animals and environment. The high frequencies of multidrug resistance and probable clonal dissemination in this serovar call for the necessity of systematic surveillance on S. Rissen in China.

Different cellular and molecular responses of Bovine milk phagocytes to persistent and transient strains of Streptococcus uberis causing mastitis.

Streptococcus uberis is frequently isolated from milk collected from dairy cows with mastitis. According to the host's immunity, bacterial virulence, and their interaction, infection with some strains can induce persistent subclinical inflammation, while infection with others induces severe inflammation and transient mastitis. This study compared the inflammatory response of milk-isolated white blood cells (mWBCs) to persistent and transient S. uberis strains. Quarter milk samples were collected aseptically for bacterial culture from all lactating cows once a week over a 10-week period. A transient and noncapsular strain with a 1-week intramammary infection duration was selected from this herd, while a persistent and capsular S. uberis strain with an intramammary infection longer than 2 months from our previous study was selected based on an identical pulse field gel electrophoresis pattern during the IMI episode. Cellular and molecular responses of mWBCs were tested, and the data were analyzed using repeated analysis of variance. The results showed a higher response in migration, reactive oxygen species generation, and bacterial killing when cells were stimulated with transient S. uberis. In contrast, the persistent strain led to increased neutrophil extracellular trap release. This study also highlighted several important molecular aspects of mWBCs. Gene expression analyses by real-time RT-PCR revealed a significant elevation in the expression of Toll-like receptors (TLR-1, TLR-2, TLR-6) and proinflammatory cytokines (tumor necrosis factor-alpha or TNF-α) with the transient strain. Additionally, Streptococcus uberis capsule formation might contribute to the capability of these strains to induce different immune responses. Altogether, these results focus on the immune function of activated mWBCs which demonstrate that a transient strain can elicit a stronger local immune response and, subsequently, lead to rapid recovery from mastitis.

Distribution of Staphylococcus aureus carriage among healthcare workers in a Japanese convalescent and rehabilitation hospital.

Earlobes, nasal cavities, and fingers of 145 healthcare workers in convalescent and rehabilitation hospital (60 nurses and 85 rehabilitation healthcare workers) were sampled. Of the 3 sites sampled, Staphylococcus aureus was detected in one or more sites in 25 nurses and 27 rehabilitation workers. S. aureus was detected in all 3 sites in 2 (8.0%) nurses and 2 (7.4%) rehabilitation workers, and the S. aureus isolates in each case showed related PFGE pattern. S. aureus was detected in both the fingers and nasal cavities of 5 (18.5%) of the rehabilitation healthcare workers; in all 5 cases, the PFGE patterns of the S. aureus isolates from each site belonged to same cluster based on PFGE. Of the 2 cases in which methicillinresistant S. aureus (MRSA) was recovered from earlobes, fingers, and nasal cavities, in both cases, MRSA isolates from the 3 sites were the same clone according to PFGE analysis and SCCmec type IV. As S. aureus was detected in pierced earlobes of nurses, hand hygiene must be practiced after touching pierced earlobes and before patient contact. The same S. aureus clone in the nasal cavity and earlobes indicates that the route of transmission is through the fingers.

Investigation of some changes and clonal relationship in enterococci isolates due to relocation of a hospital.

To investigate the isolation rates, antimicrobial resistance rates, minimum inhibitory concentration values of antimicrobial agents, and clonal relationships of Enterococcus faecalis and Enterococcus faeciumdue to the relocation of a hospital to a newly constructed building. The comparative, prospective study was conducted at adult general intensive care units of the Mus State Hospital, Mus, Turkey, in two phases; before the relocation from January 25 to December 1, 2014, and after the relocation from February 10 to May 24, 2015. Rectal swab samples were collected 72 hours post-hospitalisation. Identification of Enterococcus faecalis and Enterococcus faeciumisolates was determined by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry, and antimicrobial resistance with minimum inhibitory concentration values was detected with Vitek 2 system. The clonal relatedness among the strains was investigated by pulsed-field gel electrophoresis. Data was analysed using SPSS 23. Of the 69 patients, 37(53.62%) were related to pre-relocation phase; 20(54.1%) females and 17(45.9%) males with mean age 62.81±21.71 years. There were 32(46.37%) patients in the post-relocation phase; 13(40.6%) females and 19(59.4%) males with mean age 62.69±21.35 years (p>0.05). Of the 84 enterococci strains isolated, 51(60.7%) were Enterococcus faecium; 28(55%) before relocation and 23(45%) after relocation (p=0.77). The remaining 33(39.3%) isolates were Enterococcus faecalis; 16(48.5%) before relocation and 17(51.5%) after relocation (p=0.73). Multiple strains were located in 7(18.9%) patients before relocation and in 7(21.9%) after relocation. In 1(3.1%) patient after relocation, 2(8.7%) Enterococcus faecium isolates with different resistance and pulsed-field gel electrophoresis patterns were detected. There were no significant differences between the isolation and antibiotic resistance rates before and after relocation (p>0.05), and a clonal relation between the isolates was not detected (p>0.05). Decreased minimum inhibitory concentration values were noted for some antibiotics. Clonal relationship between the isolates and change in the rates of isolation and antimicrobial resistance of Enterococcus faecalis and Enterococcus faecium was not detected due to relocation. Minimum inhibitory concentration values could be used to reveal relocation-related changes in isolates obtained from patients hospitalised in intensive care units.

Presence of Extended-Spectrum Beta-Lactamase-Producing Escherichia coli in Food-Producing and Companion Animals and Wildlife on Small-Holder Farms of Floreana Island, Galápagos Islands.

Background: Antimicrobial resistance (AR) has led to increasing human and animal morbidity and mortality and negative consequences for the environment. AR among Escherichia coli (EC) is on the rise, with serious concerns about extended-spectrum β-lactamase-producing E. coli (ESBL-EC). In the Galápagos Islands, where antimicrobials are available without a prescription, growing demands for food production can drive antimicrobial use. Food producing animals are at the interface of wildlife and environmental health on the smallest human-inhabited Galápagos Island, Floreana. We sought to determine if ESBL-EC were present in Floreana Island farm animal species and nearby wildlife and the relatedness of ESBL-EC isolates identified. Materials and Methods: During July 4-5, 2022, we visited 8 multispecies farms, representing 75% of food-producing animal production on Floreana, and collected 227 fecal samples from farm animals and wildlife. Each sample was plated on MacConkey agar supplemented with cefotaxime (4 μg/mL). Results: ESBL-EC was isolated from 20 (9%) fecal samples collected from pigs (N = 10), chickens (N = 6), wildlife (N = 3), and dog (N = 1). All ESBL-EC isolates were from samples taken at three (38%) of the eight farms. Fifteen (75%) of the ESBL-EC isolates were from a single farm. All ESBL-EC isolates were multidrug resistant. The most prevalent ESBL genes belonged to the blaCTX-M group. Among the typeable isolates from the farm with the largest proportion of ESBL-EC isolates (N = 14), we observed nine unique pulsed-field gel electrophoresis (PFGE) patterns, with identical patterns present across pig and chicken isolates. PFGE patterns in the three farms with ESBL-EC isolates were different. Conclusions: These results lend support for future routine AR monitoring activities at the livestock-wildlife interface in Galápagos to characterize potential interspecies transmission of AR bacteria and AR genes in this unique protected ecosystem, and the related human, animal, and environmental health impacts, and to formulate interventions to reduce AR spread in this setting.

Dissemination and characteristics of carbapenem-resistant Klebsiella pneumoniae in nine district hospitals in southwestern China.

Carbapenem-resistant Klebsiella pneumoniae (CRKP) is epidemically transmitted globally, but few studies focused on the prevalence in district-level hospitals. In this study, we investigated CRKP strains collected from nine district hospitals from September 2019 to September 2020, aiming to determine the resistance mechanisms, virulence profiles, and molecular epidemiological characteristics of CRKP in district hospitals in Southwest China. A total of 51 CRKP strains were collected from 9 district-level hospitals. Matrix-assisted laser desorption/ionization-time of flight mass spectrometer was used for strain identification review, and the micro-broth dilution method was used for antibiotic sensitivity detection. Molecular epidemiological investigation of strains was performed by multilocus sequence typing (MLST) and pulsed-field gel electrophoresis (PFGE) methods. PCR and efflux pump inhibition tests were used to detect CRKP resistance mechanisms. PCR and serum killing tests were used to detect capsular serotype, virulence-related genes, and virulence validation. The CRKP strains in district hospitals presented high levels of MIC50 and MIC90 in carbapenem antibiotics especially ertapenem and meropenem. A total of 90.2% (46/51) CRKP strains were detected as carbapenemase producers, and the proportion of strains co-expressing carbapenemases was 11.8% (6/51). All CRKP strains were grouped into eight MLST types, and ST11 was the most prevalent genotype. A total of 11.8% (6/51) CRKP isolates were positive for the string test, and three strains of hypervirulent and carbapenem-resistant K. pneumoniae (HV-CRKP) were positive in serum killing test. The molecular typing of all the CRKP isolates was grouped into 29 different PFGE patterns, and 40 ST11 isolates belonged to 20 different PFGE clusters. CRKP strains showed high-level antibiotic resistance and virulence phenotype in district hospitals in Southwest China, which suggested that we should immediately pay attention to the rapid dissemination of the CRKP in regional hospitals. Our study will provide new insights into the epidemiology of CRKP in regional hospitals, which will help regional hospitals develop nosocomial infection prevention and control policies tailored to local conditions.

Genomic and phenotypic comparison of two variants of multidrug-resistant Salmonella enterica serovar Heidelberg isolated during the 2015-2017 multi-state outbreak in cattle.

Salmonella enterica subspecies enterica serovar Heidelberg (Salmonella Heidelberg) has caused several multistate foodborne outbreaks in the United States, largely associated with the consumption of poultry. However, a 2015-2017 multidrug-resistant (MDR) Salmonella Heidelberg outbreak was linked to contact with dairy beef calves. Traceback investigations revealed calves infected with outbreak strains of Salmonella Heidelberg exhibited symptoms of disease frequently followed by death from septicemia. To investigate virulence characteristics of Salmonella Heidelberg as a pathogen in bovine, two variants with distinct pulse-field gel electrophoresis (PFGE) patterns that differed in morbidity and mortality during the multistate outbreak were genotypically and phenotypically characterized and compared. Strain SX 245 with PFGE pattern JF6X01.0523 was identified as a dominant and highly pathogenic variant causing high morbidity and mortality in affected calves, whereas strain SX 244 with PFGE pattern JF6X01.0590 was classified as a low pathogenic variant causing less morbidity and mortality. Comparison of whole-genome sequences determined that SX 245 lacked ~200 genes present in SX 244, including genes associated with the IncI1 plasmid and phages; SX 244 lacked eight genes present in SX 245 including a second YdiV Anti-FlhC(2)FlhD(4) factor, a lysin motif domain containing protein, and a pentapeptide repeat protein. RNA-sequencing revealed fimbriae-related, flagella-related, and chemotaxis genes had increased expression in SX 245 compared to SX 244. Furthermore, SX 245 displayed higher invasion of human and bovine epithelial cells than SX 244. These data suggest that the presence and up-regulation of genes involved in type 1 fimbriae production, flagellar regulation and biogenesis, and chemotaxis may play a role in the increased pathogenicity and host range expansion of the Salmonella Heidelberg isolates involved in the bovine-related outbreak.

Molecular Typing and Resistance Profile of Acinetobacter baumannii Isolates during the COVID-19 Pandemic: Findings from the "EPIRADIOCLINF" Project.

Due to the COVID-19 pandemic, there has been a shift in focus towards controlling the spread of SARS-CoV-2, which has resulted in the neglect of traditional programs aimed at preventing healthcare-associated infections and combating antimicrobial resistance. The present work aims to characterize the colonization or infection with Acinetobacter baumannii of COVID-19 patients and to identify any clonality between different isolates. Specifically, data and resistance profiles of A. baumannii isolates were prospectively collected from patients recruited by the EPIRADIOCLINF project. Pulsed-field gel electrophoresis (PFGE) and multi-locus sequence typing (MLST) were used for molecular typing. Overall, we analyzed 64 isolates of A. baumannii from 48 COVID-19 patients. According to our analysis, we have identified the spread of a clonally related isolate, referred to as B. The PFGE pattern B includes four subtypes: B1 (consisting of 37 strains), B2 (11), B3 (5), and B4 (2). Furthermore, in the isolates that were examined using MLST, the most observed sequence type was ST/281. In terms of resistance profiles, 59 out of the total isolates (92.2%) were found to be resistant to gentamicin, carbapenems, ciprofloxacin, and tobramycin. The isolation and identification of A. baumannii from COVID-19 patients, along with the high levels of transmission observed within the hospital setting, highlight the urgent need for the implementation of effective prevention and containment strategies.

Diagnostic tool for surveillance, detection and monitoring of the high-risk clone K. pneumoniae ST15

The global spread of Klebsiella pneumoniae ST15, causing multi-continental outbreaks, contributes to the movement of resistance genes between clones increasing the antimicrobial resistance crisis. The genomic traits providing it with the ability to outcompete other bacteria and cause epidemics remain unclear. To identify the specific genomic traits of K.pneumoniae ST15 to develop a diagnostic test. An outbreak caused by K.pneumoniae occurred in Hospital A Coruña, Spain. Antimicrobial susceptibility analysis and molecular typing (PGFE and MLST) were performed. One isolate of each sequence type was selected for whole-genome sequencing analysis. Comparative analysis of genomes was performed using RAST. BLASTn was used to evaluate the presence of the fhaC and kpiD genes. Two hundred and ninety-four K.pneumoniae from a Spanish nationwide collection were analysed by PCR. Genotyping showed that 87.5% of the isolates tested belonged to a clone with a unique PFGE pattern which corresponded to ST15. Comparative genomic analysis of the different STs enabled us to determine the specific genomic traits of K.pneumoniae ST15. Two adherence-related systems (Kpi and KpFhaB/FhaC) were specific markers of this clone. Multiplex-PCR analysis with kpiD and fhaC oligonucleotides revealed that K.pneumoniae ST15 is specifically detected with a sensitivity of 100% and a specificity of 97.76%. The PCR results showed 100% concordance with the MLST and whole-genome sequencing data. K.pneumoniae ST15 possesses specific genomic traits that could favour its dissemination. They could be used as targets to detect K.pneumoniae ST15 with high sensitivity and specificity.

High Prevalence of GES-5 Variant and Co-Expression of VIM-2 and GES-45 among Clinical Pseudomonas aeruginosa Strains in Tunisia.

Carbapenem-resistant Pseudomonas aeruginosa (CRPA) are a global health concern. The antimicrobial resistance, virulence, and molecular typing of 57 CRPA isolated from 43 patients who attended a specific Tunisian hospital from September 2018 to July 2019 were analyzed. All but one were multidrug-resistant CRPA, and 77% were difficult-to-treat-resistant (DTR) isolates. The blaVIM-2 gene was detected in four strains (6.9%), and among the 36 blaGES-positive CRPA (62%), the blaGES-5 gene was the predominant variant (86%). Three strains co-harbored the blaVIM-2 and blaGES-45 genes, and seven CRPA carried the blaSHV-2a gene (14%). OprD alterations, including truncations by insertion sequences, were observed in 18 strains. Regarding the 46 class 1 integron-positive CRPA (81%), the blaGES-5 gene was located in integron In717, while the blaGES-29 and blaGES-45 genes were found in two new integrons (In2122 and In4879), and the blaVIM-2 gene was found in In1183 and the new integron In2142. Twenty-four PFGE patterns and thirteen sequence types (three new ones) were identified. The predominant serotype O:11 and exoU (81%) were mostly associated with ST235 and the new ST3385 clones. The seven blaSHV-2a-CRPA from different patients belonged to ST3385 and the same PFGE pattern. The blaGES-5- and blaVIM-2 + blaGES-45-positive CRPA recovered mostly from ICU patients belonged to the high-risk clone ST235. Our results highlight the alarming prevalence of blaGES-5- and ST235-CRPA, the co-existence of blaGES-45 and blaVIM-2, and their location within integrons favoring their dissemination.

Dissemination of the high-risk cloneST147 carbapenem-resistant klebsiella pneumoniae from a local tertiary care hospital in the Republic of Korea

BackgroundThe emergence of carbapenem-resistant Enterobacterales (CRE) infections is rapidly increasing and represents a serious public threat. In 2020, a total of 16,883 carbapenemase-producing Enterobacterales strains were collected; among these isolates, 21 strains were repeatedly isolated in a local tertiary care hospital.MethodsAntimicrobial susceptibility testing was performed using the broth microdilution method. All 21 strains of CRKP were analyzed by PFGE after XbaI digestion. The 21 CRKP strains were sequenced on the Illumina Miseq and Oxford Nanopore GridION platforms.ResultsThese 21 CRKP isolates showed an identical antimicrobial resistance profile, including resistance to ampicillin, carbapenems, cephems, chloramphenicol, fluoroquinolone, macrolides and trimethoprim/sulfamethoxazole. Based on whole-genome analysis, these 21 CRKP isolates shared a common genetic structure (ISAba125-IS630-blaNDM−1-bleMBL) and harbored additional resistance determinants (blaOXA−1, blaCTX−M−15, blaSHV−11, blaSHV−67, aac(6’)-Ib-cr, qnrS1, OqxA, OqxB, catB3, mph(A), sul1, and dfrA12) and mutations in the quinolone resistance-determining regions of gyrA (S83I) and parC (S80I). These isolates belonged to the ST147 and KL64 capsular types, which were carried on IncFIB replicon plasmids. The 21 CRKP strains collected from one hospital were divided into five PFGE patterns, and they were closely related with a minimum similarity value of 95.2%. These isolates were found to be highly related based on the presence of between 2 and 27 SNPs.ConclusionsThese findings indicate that NDM-1-producing K. pneumoniae ST147 may have been introduced via a common source, implying nosocomial transmission; furthermore, continuous monitoring is necessary to prevent endemic transmission.

The emergence of Klebsiella pneumoniae sequence type 395 non-susceptible to carbapenems and colistin from Turkey

PurposeOXA-48 producing Klebsiella pneumoniae is an emerging threat and outbreaks due to specific sequence types have been commonly reported. Here, we report an outbreak due to multidrug-resistant ST395 K. pneumoniae ST395. To the best of our knowledge, this is the first outbreak of K. pneumoniae ST395 harbouring blaOXA-48 genes in our country. MethodsThe strains were characterized by antimicrobial susceptibility, extended-spectrum β-lactamase (ESBL) and carbapenemase production, plasmid-mediated colistin, high-level aminoglycoside, and quinolone resistance. Also multidrug efflux pumps and porin coding genes were investigated. Pulsed-field gel electrophoresis (PFGE), multilocus sequence typing (MLST), wzi typing and plasmid analysis were used for the epidemiological relationships. ResultsAll strains were positive for blaOXA-48 with at least one of the β-lactamase genes (blaCTX-M, blaTEM, blaSHV) and harboured IncL plasmids. 16 of 20 (80%) isolates carried qnrA. All isolates were positive for aac(6’)-1b, acrAB-tolC, ompK35, and ompK36 genes but none of them harboured 16s rRNA methyltransferase, mcr-1-5, qepA, oqxAB, and mdtK genes. All strains had the same PFGE pattern, that is, wzi type K2 and found to be ST395 with MLST. ConclusionThe association of ST395 with OXA-48-producers could be an emerging threat for Turkey and continuous monitoring is crucial to prevent the spread of these powerful strains.

Antibiotic resistance and genetic diversity of Salmonella enterica serovar 1,4,[5],12:i:- isolated from animal-derived foods and humans in Shanghai, China

Salmonella enterica serovar 1,4,[5],12:i:- (S. 1,4,[5],12:i:-) is an emerging pathogen that poses a significant risk of foodborne outbreaks, particularly through the contamination of animal-derived foods. This study aimed to investigate the antibiotic susceptibility and genetic diversity of 32 S. 1,4,[5],12:i:- isolates from animal-derived foods and humans in Shanghai, China. The results revealed a high prevalence of resistance among these isolates, particularly against ampicillin (71.9%), tetracycline (65.6%), tobramycin (53.1%), trimethoprim-sulfamethoxazole (40.6%), and ciprofloxacin (40.6%). Furthermore, more than half of the isolates (56.3%) exhibited multidrug resistance. Pulsed-field gel electrophoresis analysis revealed ten distinct PFGE patterns. Several pork and clinical isolates grouped into the same pattern with 100% genetic similarity, indicating significant transmission of S. 1,4,[5],12:i:- through pork. Additionally, nine plasmid incompatibility groups (Inc) were identified, with IncQ1, IncHI2A, and IncHI2 being the most prevalent types. Interestingly, 15 isolates from foods and humans contained both IncHI2A-HI2 plasmids. These plasmids carried numerous resistance genes (e.g., blaCTX-M, blaTEM, blaOXA), suggesting their potential role in the transfer of resistance genes. The findings underscore the high prevalence of MDR S. 1,4,[5],12:i:- isolates in animal-derived foods and humans. Strengthening surveillance of animal-derived food isolates is crucial to prevent and control their spread in the food chain.

Characteristics of Virulent ST5-SCCmec II Methicillin-Resistant Staphylococcus aureus Prevalent in a Surgery Ward.

To investigate the transmission pathway of a MRSA prevalence in a pancreatic surgery ward in a Chinese teaching hospital. Molecular epidemiology investigations were carried out combined PFGE, MLST, SCCmec typing and whole-genome sequencing for 20 successive MRSA isolates (2 isolates from the ward environment). Resistance and virulence genes were detected using specific PCR. Bacterial identification and AST were performed using the Vitek 2 Compact System. Clinical data of enrolled cases were retrieved from electronic case records. From January 2020 to May 2020, successive isolated 20 MRSA strains were clarified to 2 PFGE patterns (A = 19, B = 1) in the ward. Both isolates from environment and patients belonged to sequence type ST5-SCCmec II-spa type t311. MRSA-related resistance genes mecA, blaZ, ermA, ant(4')-Ia and norA were found in each clone. All 20 isolates carried tst, hlg, hla, eta, eap, fnbA and seo virulence genes, other virulence genes such as sea, sec, seb, seg, sei, sem, sen, ebpS and fnbB were also found in partial stains. All patients had fever symptom, 27.8% were accompanied by diarrhea, 88.9% had undergone surgery or invasive procedures within 30 days. Finally, 94.4% of these patients recovered. This study confirmed a prevalence of ST5-MRSA-II-t311 clone in a surgery ward, indicated MRSA is a risk factor for post-surgery nosocomial infection and hand hygiene and environmental surveillance should not be ignored.

Whole-Genome Sequencing of Shiga Toxin-Producing Escherichia coli for Characterization and Outbreak Investigation.

Shiga toxin-producing Escherichia coli (STEC) causes high frequencies of foodborne infections worldwide and has been linked to numerous outbreaks each year. Pulsed-field gel electrophoresis (PFGE) has been the gold standard for surveillance until the recent transition to whole-genome sequencing (WGS). To further understand the genetic diversity and relatedness of outbreak isolates, a retrospective analysis of 510 clinical STEC isolates was conducted. Among the 34 STEC serogroups represented, most (59.6%) belonged to the predominant six non-O157 serogroups. Core genome single nucleotide polymorphism (SNP) analysis differentiated clusters of isolates with similar PFGE patterns and multilocus sequence types (STs). One serogroup O26 outbreak strain and another non-typeable (NT) strain, for instance, were identical by PFGE and clustered together by MLST; however, both were distantly related in the SNP analysis. In contrast, six outbreak-associated serogroup O5 strains clustered with five ST-175 serogroup O5 isolates, which were not part of the same outbreak as determined by PFGE. The use of high-quality SNP analyses enhanced the discrimination of these O5 outbreak strains into a single cluster. In all, this study demonstrates how public health laboratories can more rapidly use WGS and phylogenetics to identify related strains during outbreak investigations while simultaneously uncovering important genetic attributes that can inform treatment practices.