Pulsed-field Gel Electrophoresis Analysis Research Articles

Wound colonization with methicillin-resistant Staphylococcus aureus and hypotheses about acquisition routes in rural healthcare settings in Sub-Saharan Africa: Perspective from a Center devoted to the treatment of cutaneous neglected tropical diseases.

We identified a high prevalence (46.4%) of wound colonization with methicillin-resistant Staphylococcus aureus (MRSA) in patients hospitalized in a Center devoted to the treatment of cutaneous tropical diseases in Benin. The proportion of MRSA among S. aureus isolates was 54.3%. Thirty percent of these MRSA were identified in outpatients. Concurrently, 51% of MRSA were identified in patients for whom the time spent in the Center was between 20 days and 228 days. The analysis of pulsed-field gel electrophoresis demonstrated an important diversity of strains but also identified eight small clusters containing between two and four isolates suggesting cross-transmission. Based on these data, hypotheses about acquisition routes were suggested and measures for limiting the burden of antimicrobial resistance were envisaged.

Colonization by Extended-Spectrum β-Lactamase-Producing Enterobacterales and Bacteremia in Hematopoietic Stem Cell Transplant Recipients.

Assessing the risk of multidrug-resistant colonization and infections is pivotal for optimizing empirical therapy in hematopoietic stem cell transplants (HSCTs). Limited data exist on extended-spectrum β-lactamase-producing Enterobacterales (ESBL-E) colonization in this population. This study aimed to assess whether ESBL-E colonization constitutes a risk factor for ESBL-E bloodstream infection (BSI) and to evaluate ESBL-E colonization in HSCT recipients. A retrospective analysis of ESBL-E colonization and BSI in HSCT patients was conducted from August 2019 to June 2022. Weekly swabs were collected and cultured on chromogenic selective media, with PCR identifying the β-lactamase genes. Pulsed-field gel electrophoresis (PFGE) and whole-genome sequencing (WGS) assessed the colonizing strains' similarities. Of 222 evaluated HSCT patients, 59.45% were colonized by ESBL-E, with 48.4% at admission. The predominant β-lactamase genes were blaTEM (52%) and blaSHV (20%). PFGE analysis did not reveal predominant clusters in 26 E. coli and 15 K. pneumoniae strains. WGS identified ST16 and ST11 as the predominant sequence types among K. pneumoniae. Thirty-three patients developed thirty-five Enterobacterales-BSIs, with nine being third-generation cephalosporin-resistant. No association was found between ESBL-E colonization and ESBL-BSI (p = 0.087). Although the patients presented a high colonization rate of ESBL-E upon admission, no association between colonization and infection were found. Thus, it seems that ESBL screening is not a useful strategy to assess risk factors and guide therapy for ESBL-BSI in HSCT-patients.

Epidemiological Characteristics of Shiga Toxin-Producing Escherichia coli Responsible for Infections in the Polish Pediatric Population.

Shiga toxin-producing Escherichia coli (STEC) are zoonotic pathogens causing hemorrhagic colitis and hemolytic uremic syndrome (HUS) in children and the elderly. Stool samples were collected from 180 children hospitalized in five pediatric centers in Poland in 2018-2022. Direct stx1/stx2 gene detection by PCR in feces and E. coli isolates was performed. Antibiotic susceptibility was tested according to EUCAST v.12. Randomly selected isolates were serotyped with O157 antiserum and genotyped by pulsed-field gel electrophoresis (PFGE). A total of 44 E. coli isolates were confirmed as STEC by PCR. Among them, 84.4% were positive for stx2, and equally 6,8% for only stx1 and both stx1 and stx2 genes. The stx1 gene was also found in one Citrobacter freundii isolate. E. coli serotype O157 was present in 97.6% of the isolates. STEC infections most often occurred between June-October with a peak in July and August (51%). The highest, 77.8% of STEC isolates were found in the 1-5 years old group. No extended-spectrum β-lactamases (ESBL) were found. Resistance only to amoxicillin/clavulanic acid (24.4%), piperacillin/tazobactam (3%), cefotaxime (6%), gentamicin (6%), ciprofloxacin (3%), azithromycin (3%), trimethoprim/sulfamethoxazole (24,2%) was detected. PFGE analysis showed 18 PFGE types with no clonal distribution. Eight isolates with A, B, and C PFGE types showed genetic relatedness in the type with no detection of transmission way of distribution. STEC strains pose a serious threat to human health, therefore demographic and epidemiological characteristics are crucial for their surveillance.

Usefulness and Limitations of PFGE Diagnosis and Nucleotide Sequencing Method in the Analysis of Food Poisoning Pathogens Found in Cooking Employees.

In the case of a food poisoning outbreak, it is essential to understand the relationship between cooking workers and food poisoning. Many biological diagnostic methods have recently been developed to detect food poisoning pathogens. Among these diagnostic tools, this study presents PCR-based pulsed-field gel electrophoresis and nucleotide sequencing diagnostic analysis results for diagnosing food poisoning outbreaks associated with cooking employees in Chungcheongnam-do, Republic of Korea. Pulsed-field gel electrophoresis was useful in identifying the food poisoning outbreaks caused by Staphylococcus aureus and Enteropathogenic Escherichia coli. In the case of Norovirus, nucleotide sequencing was used to identify the relationship between cooking workers and the food poisoning outbreak. However, it is difficult to determine whether cooking employees directly caused the food poisoning outbreaks based on these molecular biological diagnostic results alone. A system is needed to integrate epidemiological and diagnostic information to identify a direct correlation between the food poisoning outbreak and cooking employees.

High Prevalence of Carbapenem-Resistant Enterobacterales Producing OXA-48 among Carbapenem-Resistant Isolates in a Regional Hospital in Central Taiwan.

In response to the increasing number of carbapenem-resistant Enterobacterales (CRE), we investigated carbapenemase-producing Klebsiella pneumoniae and non-K. pneumoniae epidemiology and genetics. We collected 76 clinical Enterobacterales and 4 stool surveillance Escherichia coli isolates resistant to ertapenem or imipenem. Using polymerase chain reaction (PCR) and DNA sequencing, we assessed carbapenemases, extended-spectrum β-lactamases, and AmpC β-lactamases. Molecular typing via pulsed-field gel electrophoresis (PFGE) and conjugation experiments were conducted to examine resistance gene transfer. Among the 80 isolates, 96.2% harbored at least one carbapenemase gene, with blaOXA-48 in 87.5%. KPC-2 and IMP-8 carbapenemases were found in 15.0 and 22.5% of the isolates, respectively, with 27.5% having 2 or more carbapenemase genes. The PFGE analysis revealed the presence of diverse genotypes. PCR-based plasmid replicon typing identified IncA/C as the most prevalent type among K. pneumoniae isolates (26/29), and IncF and IncFIB among E. coli isolates (22/28). Conjugal transfer was successful for plasmids encoding OXA-48, CTX-M-3, CTX-M-14, CMY-2, and other β-lactamases, except the KPC-2 gene. In conclusion, our study highlights high carbapenemase prevalence in CRE, primarily OXA-48. Multiple carbapenemases within strains were common, and PFGE showed diverse patterns in these carbapenem-resistant isolates.

High prevalence of carbapenem resistance and clonal expansion of blaNDM gene in Klebsiella pneumoniae isolates in an Iranian referral pediatric hospital

BackgroundThe increasing global concern regarding antibiotic resistance necessitates in-depth studies to comprehend the phenotypic and genotypic characteristics of resistant bacterial strains. This study aimed to investigate the prevalence, antibiotic resistance profiles, and molecular characteristics of carbapenem-resistant Klebsiella pneumoniae (CRKP) isolates in an Iranian referral pediatrics hospital. Methods: In this study, we examined CRKP isolates collected from hospitalized pediatric patients across various wards. The isolates underwent antimicrobial susceptibility testing, the polymerase chain reaction (PCR) analysis for carbapenemase genes (blaNDM, blaVIM and blaIMP), and genetic relatedness assessment using pulsed-field gel electrophoresis (PFGE).ResultsAmong 166 K. pneumoniae isolates, 54 (32.5%) exhibited resistance to carbapenems. Notably, all these resistant isolates were resistant to imipenem, with 35 (65%) displaying resistance to both imipenem and meropenem. Of the 54 CRKP isolates, 24 (44%) were metallo-β-lactamases (MBL)-producing. The prevalence of the blaNDM gene among CKCP and MBL-producing isolates was 20% (n = 11) and 44% (n = 24), respectively. The blaVIM and blaIMP genes were not detected in any of the isolates. Twenty-six CRKP isolates (48%) were recovered from ICUs. PFGE analysis of CRKP isolates revealed 20 clusters, with cluster S being the most prevalent, comprising 24% of the total (n = 13).ConclusionOur study reveals a concerning prevalence of carbapenem resistance in K. pneumoniae isolates. Specifically, the detection of the blaNDM gene in 20% of CRKP isolates, with a significant proportion (82%) observed in isolated CRKP from the ICUs and emergency departments, underscores the potential clonal expansion of these resistant strains within these critical hospital wards.

Salmonellosis outbreaks linked to eggs at 2 gimbap restaurants in the Republic of Korea.

Salmonellosis outbreaks occurred at 2 restaurants 2 days apart, and an epidemiological investigation was conducted to determine whether the outbreaks were connected. Case studies were conducted for both outbreaks. Stool samples were collected from individuals, and food samples were collected from the restaurants. Pulsed-field gel electrophoresis (PFGE) and whole-genome sequencing analyses were performed on outbreak-related Salmonella enterica serovar Enteritidis (Salmonella Enteritidis) isolates. Traceback investigations were also conducted for the ingredients from gimbap restaurants A and B. In total, 106 people from gimbap restaurant A and 5 from gimbap restaurant B met the case definition. Salmonella Enteritidis was detected in samples from 2 food handlers, 22 patients, and 1 food (iceberg lettuce) at gimbap restaurant A and from 1 patient at gimbap restaurant B. According to PFGE, all isolates were identified as SEGX01.089. The molecular typing of all isolates showed the same pattern, and the genetic distance was close according to phylogenetic analysis. Eggs were the only food ingredient that was supplied to both gimbap restaurants. The outbreaks were caused by Salmonella Enteritidis, and the source of infections was suspected to be contaminated eggs. To prevent foodborne outbreaks of Salmonella, restaurants should heat eggs sufficiently, and egg farms need to establish management systems that prevent Salmonella infections.

Whole genome sequence-based molecular characterization of blood isolates of carbapenem-resistant Enterobacter cloacae complex from ICU patients in Kolkata, India, during 2017-2022: emergence of phylogenetically heterogeneous Enterobacter hormaechei subsp. xiangfangensis.

Blood-borne infections caused by the carbapenem-resistant Enterobacter cloacae complex (CR-ECC) are major public threats with respect to the challenges encountered during treatment. This study describes the whole genome sequencing-based molecular characteristics of blood isolates (n = 70) of CR-ECC from patients admitted to the intensive care unit of tertiary care hospitals in Kolkata, India, during 2017-2022 with respect to species identification, antimicrobial resistance (AMR) profiling, mechanism of drug resistance, and molecular subtypes. Vitek2 MALDI and species-specific PCR identified Enterobacter hormaechei subsp. xiangfangensis (47.14%) as the emerging CR-ECC subspecies in Kolkata. The predominating carbapenemase and extended-spectrum β-lactamase genes found were blaNDM-1 (51.42%) and blaCTX-M-15 (27%), respectively. Besides, blaNDM-4, blaNDM-5, blaNDM-7, blaCMH-3, blaSFO-1, blaOXA-181, blaOXA-232, blaKPC-3, and blaDHA-7 genes were also detected, which were not previously reported from India. A multitude of Class 1 integrons (including In180, In4874, In4887, and In4888, which were novel) and plasmid replicon types (IncFIB, IncFII, IncX3, IncHI1-HI2, IncC, and IncR) involved in AMR dissemination were identified. Reverse transcription-PCR and western blot revealed that carbapenem resistance in non-carbapenemase-producing CR-ECC isolates was contributed by elevated levels of ampC, overexpression of acrAB, and loss of ompF. A total of 30 distinct sequence types (STs) were ascertained by multi-locus sequence typing; of which, ST2011, ST2018, ST2055, ST2721, and ST2722 were novel STs. Pulsed-field gel electrophoresis analysis showed heterogeneity (69 pulsotypes with a similarity coefficient of 48.40%) among the circulating isolates, suggesting multiple reservoirs of infections in humans. Phylogenetically and genetically diverse CR-ECC with multiple AMR mechanisms mandates close monitoring of nosocomial infections caused by these isolates to forestall the transmission and dissemination of AMR.IMPORTANCEThe emergence and extensive dissemination of the carbapenem-resistant Enterobacter cloacae complex (CR-ECC) have positioned it as a critical nosocomial global pathogen. The dearth of a comprehensive molecular study pertaining to CR-ECC necessitated this study, which is the first of its kind from India. Characterization of blood isolates of CR-ECC over the last 6 years revealed Enterobacter hormaechei subsp. xiangfangensis as the most prevalent subsp., exhibiting resistance to almost all antibiotics currently in use and harboring diverse transmissible carbapenemase genes. Besides the predominating blaNDM-1 and blaCTX-M-15, we document diverse carbapenemase and AmpC genes, such as blaNDM-4, blaNDM-7, blaOXA-181, blaOXA-232, blaKPC-3, blaCMH-3, blaSFO-1, and blaDHA-7, in CR-ECC, which were not previously reported from India. Furthermore, novel integrons and sequence types were identified. Our findings emphasize the need for strengthened vigilance for molecular epidemiological surveillance of CR-ECC due to the presence of epidemic clones with a phylogenetically diverse and wide array of antimicrobial resistance genes in vulnerable populations.

Characterization of nontyphoidal Salmonella strains from a tertiary hospital in China: serotype diversity, multidrug resistance, and genetic insights.

Nontyphoidal Salmonella is a significant public health concern due to its ability to cause foodborne illnesses worldwide. This study aims to characterize the nontyphoidal Salmonella strains isolated from patients in China. A total of 19 nontyphoidal Salmonella strains were characterized through serovar identification, antimicrobial susceptibility testing (AST), biofilm formation assessment. Genetic relatedness was determined using pulsed-field gel electrophoresis (PFGE). WGS was employed to decipher the resistance mechanism and to contextualize the S. serovar Mbandaka strains among previously sequenced isolates in China. The biofilm associated mrkA gene was examined by PCR. The predominant serovar identified was S. Enteritidis, followed by S. Mbandaka, S. Thompson, S. Livingston, S. Alachua, and S. Infantis. PFGE analysis indicated a notable genetic similarity among the S. Mbandaka isolates. Phylogenetic analysis suggested that these strains were likely derived from a single source that had persisted in China for over five years. One multidrug resistance (MDR) S. Enteritidis isolate carried a highly transferable IncB/O/K/Z plasmid with bla CTX-M-15. One S. Thompson strain, harboring the mrkABCDF operon in an IncX1 plasmid, isolated from cutaneous lesions, demonstrated robust biofilm formation. However, no mrkABCDF loci were detected in other strains. Our study emphasizes the importance of persisted surveillance and prompt response to Salmonella infections to protect public health. The dissemination of bla CTX-M-15-harboring IncB/O/K/Z plasmid and the spread of virulent mrkABCDF operon among Salmonella in China and other global regions warrant close monitoring.

Foodborne Outbreak by Salmonella enterica Serovar Weltevreden in West Bengal, India.

Foodborne gastroenteritis outbreaks owing to Salmonella enterica serovar Weltevreden (Salmonella Weltevreden) represent a significant global public health problem. In the past two decades, Salmonella Weltevreden has emerged as a dominant foodborne pathogen, especially in South-East Asian countries. This report describes a community foodborne outbreak of gastroenteritis caused by Salmonella Weltevreden in August 2022 following consumption of panipuri from a street vendor in the Polba block in Hooghly district, West Bengal, India. This food item was consumed by 185 people, of whom 129 had acute watery diarrhea with other clinical symptoms and 65 of them were admitted to different District hospitals for treatment. Stool specimens collected from hospitalized cases were positive for S. enterica, and further serotyped as Salmonella Weltevreden. All the Salmonella Weltevreden strains possessed the Salmonella pathogenicity islands associated genes (invA/E, orgA, ttrc, ssaQ, mgtC, misL, spi4D), the enterotoxin (stn), and hyperinvasive locus gene (hilA). Except erythromycin, all the strains were susceptible for commonly used antimicrobials in the treatment of diarrhea. The XbaI-based pulsed-field gel electrophoresis analysis indicated that all the isolates responsible for the recent outbreak were similar, but diverged from other Salmonella Weltevreden that were previously reported in West Bengal. This report indicates that foodborne infection is a major public health concern in India and demands to strengthen capacity-building measures at the local health care levels for linking causative agents of outbreaks.

A Food Poisoning Caused by Salmonella Enterica (S. Enteritidis) ST11 Carrying Multi-Antimicrobial Resistance Genes in 2019, China.

This study was to identify and analyze the pathogen responsible for food poisoning in a tourist group traveling from Macao to Zhuhai. Samples were obtained from 27 patients of 96 cases, as well as samples of contaminated food in Macau. The collected samples were subjected to serological identification, drug sensitivity analysis, drug resistance gene identification, virulence factor analysis, and tracing. Twenty-six isolates and the salad isolate were S. enteritidis ST11. Isolates from patients were exhibited significant resistance to Penicillin AMP (Ampicillin) and quinolones NAL (Nalidixic acid). Among these isolates, 21 strains were resistant to two or more antibiotics, indicating the multi-drug resistance (MDR). Genomic characteristics and phylogenetic analysis were performed on 9 of the isolates using whole genome sequencing (WGS). The analysis revealed that the resistance to AMP and NAL was primarily caused by a gryA mutation D87Y (9/9, 100%), and the presence of beta-lactam resistance genes blaOXA-1 (1/9, 11.11%), blaTEM-141 (1/9, 11.11%), and blaTEM-1B (8/9, 88.89%). It was also found a strains isolated from patients had two resistance genes to quinolones or beta-lactam drugs (1/8, 12.5%), respectively. The strains were found to possess 165 virulence genes, one adherence class virulence factor, one invasion class virulence factor and various pathogenicity islands, including SPI-1, SPI-2, SPI-3, SPI-4, SPI-5, SPI-9, SPI-10, SPI-13, SPI-14, SPI-15, SGI 1, CS54_island, and C63PI-1. Additionally, the virulence plasmids were detected, including IncFIB(s)-IncFII(s)-IncX1 (55.56%), IncFIB(s)-IncFII(s) (33.33%), and IncFIB(s)-IncFII(s)-IncHI2-IncHI2A (11.11%). PFGE (Pulsed Field Gel Electrophoresis) and phylogenetic tree analysis revealed a high degree of similarity between Salmonella isolates from patients and food samples from Macao. This study identified Salmonella enterica ST11 as the cause of the food poisoning outbreak. The findings highlight the importance of phenotypic characterization and next-generation sequencing (NGS) tools in epidemiological studies and emphasize the potential risk of a new emerging multi-antibiotic ST11 clone for S. enteritidis.

ST11 KPC-2-Producing Klebsiella pneumoniae Isolated from Patient with Acute Myelocytic Leukemia.

The emergence of the ST11-CRKP (ST11-CRKP) strain is expected to become a serious public health problem in China. As one of the most serious complications in patients with acute myeloid lymphoma, infections can cause systemic infection and life-threatening sepsis, seriously affecting the morbidity, mortality, and quality of life of patients. Thus, ST11-CRKP infections in patients with acute myeloid lymphoma are worthy of our attention. To investigate the occurrence and genetic characteristics of the ST11-CRKP from a patient with acute myeloid lymphoma. Species identification was determined by MALDI-TOF MS. Antimicrobial susceptibility testing (AST) was conducted by VITEK 2 system with AST-N335 panel. Whole-genome sequencing was performed on the Illumina NovaSeq 6000 platform. Phylogenetic analyses were performed using Snippy based on the core-genome SNPs. S1 nuclease pulsed-field gel electrophoresis (S1-PFGE), Southern blot and Whole-genome analysis indicated blaKPC-2 genes were located on plasmids with a conserved genetic environment. Moreover, the eight ST11-CRKP strains carry a variety of antimicrobial resistance genes (ARGs) and virulence factors. The ability of biofilm formation of eight strains was verified by a crystal violet assay. Core genome single-nucleotide polymorphism (cgSNP) analysis suggesting a possible bacterial translocation event. We performed a comprehensive analysis of ST11-CRKP strains from a patient with acute myelocytic leukemia. Our study emphasized the need for continuous surveillance of ST11-CRKP in the clinic especially in the immunocompromised population.

Pulsed-field gel electrophoresis typing and molecular characterization of <em>Listeria monocytogenes</em> isolates in raw milk samples from Polonnaruwa District, Sri Lanka

Listeria monocytogenes is an important food borne human pathogen associated with severe diseases in humans and animals. Human infection is associated with high mortality rates. Our previous studies done in 2012 on milk and milk products revealed 4b as the dominant serotype followed by 1/2a, 1/2b and 1/2c. Listeria monocytogenes is an important food borne human pathogen associated with severe diseases in humans and animals. Human infection is associated with high mortality rates. Our previous studies done in 2012 on milk and milk products revealed 4b as the dominant serotype followed by 1/2a, 1/2b and 1/2c. Therefore, this study was conducted to determine the sources of contamination and to identify the strain differences (pulsotypes) by pulsed-field gel electrophoresis (PFGE) typing and serotype profiles of L. monocytogenes isolates from farm collected raw milk samples from randomly identified locations in Grama Niladhari divisions of the Polonnaruwa district, Sri Lanka. Eighty isolates obtained from raw milk samples were confirmed as L. monocytogenes by Polymerase Chain Reaction, belonging to serotypes 4b, 1/2a, 1/2b and 1/2c. The strains were identified by PFGE. PFGE analysis digested with AscI and ApaI enzymes revealed different banding patterns. All the pulsotypes were found to be serotype 4b. The sub typing indicated the diversity of the Listeria species. The presence of serotypes 1/2a, 1⁄2b, 1/2c, and 4b in raw milk is a public health concern, as these serotypes are frequently associated with foodborne outbreaks and sporadic cases of human listeriosis. In our study, PFGE analysis allowed discrimination among isolates of the same serogroup. Further PFGE analysis showed heterogeneity among isolates recovered from both same sampling areas and different areas.

High Prevalence of Carbapenem-Resistant Enterobacterales Colonization Among Intensive Care Unit Patients in a Tertiary Hospital, China.

Intestinal colonization with carbapenem-resistant Enterobacterales (CRE) has been shown as a significant risk factor for subsequent CRE infections, especially in intensive care units (ICUs). The aim of this study was to determine the prevalence of intestinal CRE colonization among ICU patients in a Chinese tertiary hospital. Fecal sample screenings for CRE were performed on ICU patients weekly. Antibiotic-susceptibility profile of CRE strains was determined using the Vitek-2 analysis system and broth microdilution method. The carbapenemases of all isolates were determined by phenotypes and genotypes. Clonal relatedness was analyzed by pulsed-field gel electrophoresis (PFGE). Whole-genome sequencing was used to identify the multilocus sequence type (ST), plasmid replicons, and insertion sequences (ISs) of isolates. The overall colonization rate of CRE was 40.4% (82/203). A total of 84 CRE strains were detected, mostly with Klebsiella pneumoniae (92.9%). Antibiotic susceptibility testing profile revealed that 84 CRE strains were resistant to most antibiotics except for tigecycline and colistin. The carbapenemase-encoding genes including blaKPC-2, blaNDM-1, and blaIMP-4 were detected, and blaKPC-2 was the predominant genotype (90.8%). A total of 9 STs were identified among 84 CRE strains, and ST11 was the most common type (83.3%). A variety of mobile genetic elements, including plasmids and ISs, were detected via online tool prediction. PFGE analysis of the 78 K. pneumoniae strains showed 8 different pulsotypes, and pulsotype A was highly prevalent. This study found that the prevalence of CRE colonization was alarmingly high in the ICU, and that effective infection control measures are urgently needed to prevent the dissemination of CRE.

Genotypic and phenotypic analysis of Elizabethkingia meningoseptica in bullfrog Rana catesbeiana isolated in Taiwan.

Elizabethkingia meningoseptica is a hazardous bacterium for agriculture production and human health. The present study identified E. meningoseptica from the bullfrog, human and reference strain BCRC 10677 by API 20NE, 50S ribosome protein L27 sequencing and pulse field gel electrophoresis to differentiate isolates of E. meningoseptica from aquatic animals and humans. All isolates from bullfrogs and humans were identified as E. meningoseptica by DNA sequencing with 98.8%-100% sequence identity. E. meningoseptica displayed significant genetic diversity when analysed using pulsed-field gel electrophoresis (PFGE). There were six distinct pulsotypes, including one pulsotype found in bullfrog isolates and five pulsotypes found in human isolates. However, E. meningoseptica from bullfrog exhibited one genotype only by PFGE. Overall, molecular epidemiological analysis of PFGE results indicated that the frog E. meningoseptica outbreaks in Taiwan were produced by genetically identical clones. The bullfrog isolates were not genetically related to other E. meningoseptica from human and reference isolates. This research provided the first comparisons of biochemical characteristics and genetic differences of E. meningoseptica from human and bullfrog isolates.

Pulsed-Field Gel Electrophoresis Analysis of Bovine Associated Staphylococcus aureus: A Review.

For decades now, DNA fingerprinting by means of pulsed-field gel electrophoresis (PFGE) continues to be the most widely used to separate large DNA molecules and distinguish between different strains in alternating pulses. This is done by isolating intact chromosomal DNA and using restriction enzymes with specific restriction sites to generate less than 30 restriction fragments from 50 Kb to 10 Mbp. These results make clone-specific band profiles easy to compare. Specialized equipment is required for the optimization of DNA separation and resolution, among which a contour-clamped homogeneous electric field (CHEF) apparatus is the most commonly used. As a result, the PFGE analysis of a bacterial genome provides useful information in terms of epidemiological investigations of different bacterial pathogens. For Staphylococcus aureus subtyping, despite its limitations and the emergence of alternative methods, PFGE analysis has proven to be an adequate choice and the gold standard for determining genetic relatedness, especially in outbreak detection and short-term surveillance in the veterinary field.

Dissemination of ArmA- and OXA-23-co-producing Acinetobacter baumannii Global Clone 2 in Switzerland, 2020-2021.

Following the observation of an increased number of isolation of OXA-23- and ArmA-producing Acinetobacter baumannii at the national level, our aim was to evaluate whether some given clone(s) might actually be spreading and/or emerging in Switzerland. To evaluate this possibility, our study investigated and characterized all A. baumannii isolates harboring both the blaOXA-23 and armA genes that had been collected at the Swiss National Reference Center for Emerging Antibiotic Resistance (NARA) from 2020 to 2021. Most isolates were obtained from infections rather than colonization with the majority being obtained from respiratory specimens. Pulsed-field gel electrophoresis (PFGE) analysis of 56 isolates identified nine profiles. Then, whole-genome sequencing that was performed on a subset of 11 isolates including at least one representative isolate of each PFGE profile identified three STs; one each of ST25 and ST1902, and nine ST2 (a member of Global Clone 2 (GC-2). The blaOXA-23 gene was always found embedded within Tn2006 structures, as commonly described with GC-2 (ST2) isolates. Susceptibility testing showed that most of those isolates, despite being highly resistant to all carbapenems and all aminoglycosides, remained susceptible to colistin (94.6%), sulbactam-durlobactam (87.5%), and cefiderocol (83.9% or 91.1% according to EUCAST or CLSI breakpoints, respectively). Overall, this study identified that the A. baumannii co-producing OXA-23 and ArmA are increasing in incidence in Switzerland, largely due to the dissemination of the high-risk GC-2. This highlights the importance of themonitoring of such MDR A. baumannii strains, in order to contribute to reduce their potential further spread.

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.

Tracking spatio-temporal distribution and transmission of antibiotic resistance in aquatic environments by using ESBL-producing Escherichia coli as an indicator

Wastewater treatment plants (WWTPs) play an important role in the production, and transmission of antibiotic resistant bacteria (ARB) and antibiotic resistance genes (ARGs) as a convergence for human, animal, and environmental wastewater. The aim of this study was to investigate the spatio-temporal variation and influencing factors of ARB in different functional areas of the urban WWTP and the connecting rivers for 1-year monitoring using extended-spectrum β-lactamase-producing Escherichia coli (ESBL-Ec) as an indicator bacteria, and to study the transmission patterns of ARB in the aquatic environment. The results showed that ESBL-Ec isolates were identified from the WWTP (n = 219), including influent (n = 53), anaerobiotic tank (n = 40), aerobiotic tank (n = 36), activated sludge tank (n = 31), sludge thickner tank (n = 30), effluent (n = 16), and mudcake storage area (n = 13). The dehydration process can significantly remove the ESBL-Ec isolates; however, ESBL-Ec was still detected in samples collected from the effluent of the WWTP (37.0%). The detection rate of ESBL-Ec was significantly different across seasons (P < 0.05), and ambient temperature was negatively correlated with the detection rate of ESBL-Ec (P < 0.05). Furthermore, a high prevalence of ESBL-Ec isolates (29/187, 15.5%) was detected in samples collected from the river system. These findings emphasize that the high majority of ESBL-Ec in aquatic environments is alarming because it poses a significant threat to public health. Clonal transmission of ESBL-Ec isolates between the WWTP and rivers based on the spatio-temporal scale was observed by pulsed-field gel electrophoresis analysis, ST38 and ST69 ESBL-Ec clone were selected as prioritized isolates for antibiotic resistance monitoring in the aquatic environment. Further phylogenetic analysis showed human-associated (feces, blood) E. coli was the main source contributing to the presence of antibiotic resistance in aquatic environments. Longitudinal and targeted monitoring of ESBL-Ec in WWTPs and the development of effective wastewater disinfection strategies before effluent discharge from WWTPs are urgently required, to prevent and control the spread of antibiotic resistance in the environment.

Clonal diversity of Staphylococcus aureus isolates in clinical specimens from selected health facilities in Ethiopia

Staphylococcus aureus is among the top three causative agents of nosocomial infection in Ethiopia. The majority of studies in Ethiopia have focused on the epidemiology of S. aureus in hospital settings, with limited molecular genotyping results. Molecular characterization of S. aureus is essential for identification of strains, and contributes to the control and prevention of S. aureus infection. The aim of the current study was to determine the molecular epidemiology of methicillin-susceptible S. aureus (MSSA) and methicillin-resistant S. aureus (MRSA) isolates recovered from clinical specimens in Ethiopia. A total of 161 MSSA and 9 MRSA isolates were characterized using pulsed-field gel electrophoresis (PFGE) and staphylococcal protein A (spa) typing. Based on the PFGE analysis, MSSA isolates were grouped into eight pulso-types groups (from A to I), while MRSA isolates clustered into three (A, B and C) pulso-types with more than 80% similarity. The spa typing analysis showed diversity of S. aureus with 56 distinct spa types. Spa type t355 was most prevalent (56/170, 32.9%), while eleven new spa types were detected including t20038, t20039, and t20042. The identified spa types were clustered into 15 spa-clonal complexes (spa-CCs) using BURP analysis; novel/unknown spa types were further subjected to MLST analysis. The majority of isolates belonged to spa-CC 152 (62/170, 36.4%), followed by spa-CC 121 (19/170, 11.2%), and spa-CC 005 (18 /170, 10.6%). Of the nine MRSA isolates, 2 (22.2%) were spa-CC 239 with staphylococcal cassette chromosome (SCC)mec III. These findings highlight the diversity of S. aureus strains in Ethiopia, as well as the presence of potentially epidemic strains circulating in the country necessitating further characterization of S. aureus for antimicrobial resistance detection and infection prevention purposes.