Repetitive Intergenic Consensus Polymerase Chain Research Articles

High prevalence of multidrug-resistant Enterobacterales carrying extended-spectrum beta-lactamase and AmpC genes isolated from neonatal sepsis in Ahvaz, Iran

ObjectivesIn the recent years, multidrug resistant (MDR) neonatal septicemia-causing Enterobacterales has been dramatically increased due to the extended-spectrum beta-lactamases (ESBLs) and AmpC enzymes. This study aimed to assess the antibiotic resistance pattern, prevalence of ESBLs/AmpC beta-lactamase genes, and Enterobacterial Repetitive Intergenic Consensus Polymerase Chain Reaction (ERIC-PCR) fingerprints in Enterobacterales isolated from neonatal sepsis.ResultsIn total, 59 Enterobacterales isolates including 41 (69.5%) Enterobacter species, 15 (25.4%) Klebsiella pneumoniae and 3 (5.1%) Escherichia coli were isolated respectively. Resistance to ceftazidime and cefotaxime was seen in all of isolates. Furthermore, all of them were multidrug-resistant (resistant to three different antibiotic categories). The phenotypic tests showed that 100% of isolates were ESBL-positive. Moreover, AmpC production was observed in 84.7% (n = 50/59) of isolates. Among 59 ESBL-positive isolates, the highest percentage belonged to blaCTX−M−15 gene (66.1%) followed by blaCTX−M (45.8%), blaCTX−M−14 (30.5%), blaSHV (28.8%), and blaTEM (13.6%). The frequency of blaDHA, blaEBC, blaMOX and blaCIT genes were 24%, 24%, 4%, and 2% respectively. ERIC-PCR analysis revealed that Enterobacterales isolates were genetically diverse. The remarkable prevalence of MDR Enterobacterales isolates carrying ESBL and AmpC beta-lactamase genes emphasizes that efficient surveillance measures are essential to avoid the more expansion of drug resistance amongst isolates.

Stray Dogs (Mongrels) Are Potent Reservoir of Drug-Resistant Pathogens: A Study in Peri-Urban Areas of Kolkata, India.

This study depicts the drug-resistance and phylogenomic characteristics of 365 Escherichia coli (EC) and 76 Klebsiella pneumoniae (KP) isolated from stray dogs (293) in and around Kolkata, India. Initial screening found 59 isolates, including 48 E. coli and 11 KP multidrug resistant, which included 33 extended-spectrum β-lactamase, 41 AmpC β-lactamase and 18 metallo-β-lactamase producers carrying blaNDM-1 (11) and blaNDM-5 (7) genes. Majority of them had the resistant genes such as blaCTX-M (33), blaTEM (18), blaSHV (4), blaOXA (17), blaFOX (2), blaDHA (2), blaCITM (15), blaCMY-2 (13), blaGES (2) and blaVEB (2), qnrS (15), qnrB (3), aac-6'-Ib-cr (14), tetA (26), tetB (14), sul-1 (25), armA (2) and rmtB (6), in addition to adherence genes such as csgA (33), fimA (27), fliC (13), sdiA (33), rcsA (38), and rpoS (39). They also carried plasmid of diverse replicon types of which IncFIA and FIB were the most frequent. Phylogrouping categorized most of the MDR E. coli in phylogroup A (20), B1 (14), and B2 (6). Enterobacteriaceae repetitive intergenic consensus-polymerase chain reaction (ERIC-PCR) showed genetic diversity of multidrug resistant isolates irrespective of their origin, resistance, and virulence types, differentiating the EC in five clades (A-E) and KP in four clades (A-D). As these stray dogs, which had no history or scope of previous antimicrobial therapy, were found to have contracted potential antimicrobial resistance pathogens, the role of environment in spread of such pathogens and further possibility of human infections cannot be ruled out.

Microbiological quality of irrigation water on highly diverse fresh produce smallholder farms: elucidating environmental routes of contamination.

To evaluate the microbiological safety, potential multidrug-resistant bacterial presence and genetic relatedness (DNA fingerprints) of Escherichia coli isolated from the water-soil-plant nexus on highly diverse fresh produce smallholder farms. Irrigation water (n=44), soil (n=85), and fresh produce (n=95) samples from six smallholder farms with different production systems were analysed for hygiene indicator bacterial counts and the presence of shigatoxigenic E. coli and Salmonella spp. using standard microbiological methods. Identities of isolates were confirmed using matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS), and the genetic relatedness of the E. coli isolates determined using enterobacterial repetitive intergenic consensus polymerase chain reaction (ERIC-PCR) analysis. Irrigation water E. coli levels ranged between 0 and 3.45 log MPN/100 ml-1 with five farms having acceptable levels according to the World Health Organization limit (3 log MPN/100 ml-1). Fresh produce samples on four farms (n=65) harboured E. coli at low levels (<1 log CFU/g-1) except for one sample from kale, spring onion, green pepper, onion, and two tomato samples, which exceeded international acceptable limits (100 CFU/g-1). Only one baby carrot fresh produce sample tested positive for Salmonella spp. Of the 224 samples, E. coli isolates were identified in 40% (n=90) of all water, soil, and fresh produce types after enrichment. Additionally, the DNA fingerprints of E. coli isolates from the water-soil-plant nexus of each respective farm clustered together at high similarity values (>90%), with all phenotypically characterized as multidrug-resistant. The clustering of E. coli isolated throughout the water-soil-plant nexus, implicated irrigation water in fresh produce contamination. Highlighting the importance of complying with irrigation water microbiological quality guidelines to limit the spread of potential foodborne pathogens throughout the fresh produce supply chain.

Molecular typing of clinical multidrug-resistant Klebsiella pneumoniae isolates.

Klebsiella pneumoniae is an opportunistic pathogen which is an important cause of hospital-acquired and antibiotic resistance infections. Therefore, this study aimed to determine the frequency of resistance to antibiotics, as well as the molecular typing of the associated isolates, and compare multiple-locus VNTR analysis (MLVA) and Enterobacterial Repetitive Intergenic Consensus-Polymerase Chain Reaction (ERIC-PCR) methods to specify the degree to which distinctions can be separated from each other. One hundred K. pneumoniae isolates were obtained from different sources of infections from patients admitted to hospitals. Antibiotic susceptibility testing was then performed by applying the Kirby-Bauer disk diffusion method. Typing of K. pneumoniae was done by utilizing MLVA and ERIC-PCR methods. Eighty-six multidrug-resistant (MDR) K. pneumoniae isolates were identified, which resistance to ampicillin, trimethoprim/sulfamethoxazole, and ceftriaxone was the most frequent in the considered isolates (100, 93, and 93%, respectively). A total of 50 different antibiotic susceptibility patterns were observed among the MDR K. pneumonia, with the most frequent pattern being resistance to all antibiotics (12.79%) and resistance to all antibiotics except amikacin (10.47%). The isolates were then divided into 37 different MLVA types and seven clonal complexes were obtained from the minimum spanning tree analysis. Finally, the isolates were assigned to 38 different ERIC types. The discriminatory power of MLVA and ERIC methods also showed a value of 0.958, and 0.974. Both PCR-typing methods with phenotypic patterns can be useful for the epidemiological typing of K. pneumoniae isolates with the highest performance in discriminating isolates.

Serotype and genotyping of &lt;i&gt;Salmonella&lt;/i&gt; sp. isolated from street foods in N’Djaména, Chad

Group foodborne illness infections with Salmonella are often caused by the simultaneous consumption by several people of heavily contaminated food. The objective of this work was to determine the phylogenetic relationships between dietary strains of Salmonella sp. isolated from our samples and confirmed clinical strains. A total of 447 samples were collected and analyzed using standard food microbiology methods. Enterobacterial Repetitive Intergenic Consensus-Polymerase chain Reaction (ERIC-PCR) technique allowed confirmation that all our strains belong to the genus Salmonella. From ten (10) strains characterized, eight (8) different genotype profiles were identified. The different DNA fragments of the isolates were generated by ERIC-PCR and then revealed on the agarose gel and had bands ranging from 2 to 6 bands with sizes between 150 and 1500 bp. The two food strains of S. Mbadaka (1 and 2) presented identical profiles. However, it is noted that two (2) strains of S. Anatum had different ERIC-PCR profiles. There is also a very high resemblance between the strain of S. Anatum (strain 5) and the confirmed clinical strains (H5: Salmonella Typhi). The presence of risky germs in the food analyzed poses a public health problem and should call on the competent authorities and prompt the taking of corrective measures.

Salmonella enterica serovars linked with poultry in India: antibiotic resistance profiles and carriage of virulence genes.

Salmonella is an important poultry pathogen with zoonotic potential. Being a foodborne pathogen, Salmonella-contaminated poultry products can act as the major source of infection in humans. In India, limited studies have addressed the diversity of Salmonella strains of poultry origin. This study represented 26 strains belonging to Salmonella serovars Typhimurium, Infantis, Virchow, Kentucky, and Agona. The strains were tested for resistance to 14 different antimicrobial agents using the Kirby-Bauer disk-diffusion assay. The presence of the invA, hilA, agfA, lpfA, sopE, and spvC virulence genes was assessed by polymerase chain reaction (PCR), and the genetic diversity was assessed by Enterobacterial Repetitive Intergenic Consensus Polymerase Chain Reaction (ERIC-PCR). The highest resistance to tetracycline (n = 17; 65.38%) followed by nalidixic acid (n = 16; 61.53%) was detected among the strains. Among the strains (n = 17) phenotypically resistant to tetracycline, 94% (n = 16) were also positive for the tetA gene. Based on the presence of virulence genes, the strains were characterized into three virulence profiles (PI, P2, and P3). Among the investigated virulence genes, invA, hilA, agfA, and lpfA were present inall strains. The sopE gene was mostly associated with serovars Virchow (n = 3; 100%) and Typhimurium (n = 8; 80%), whereas spvC gene was exclusive for two Typhimurium strains that lacked sopE gene. ERIC-PCR profiling indicated clusters correlating their serovar, geographical, and farm origins. These results demonstrate that Salmonella isolates with a wide genetic range, antibiotic resistance, and virulence characteristics can colonize poultry. The presence of such strains is crucial for both food safety and public health.

Pathogenicity assessment of Arcobacter butzleri isolated from Canadian agricultural surface water

BackgroundWater is considered a source for the transmission of Arcobacter species to both humans and animals. This study was conducted to assess the prevalence, distribution, and pathogenicity of A. butzleri strains, which can potentially pose health risks to humans and animals. Cultures were isolated from surface waters of a mixed-use but predominately agricultural watershed in eastern Ontario, Canada. The detection of antimicrobial resistance (AMR) and virulence-associated genes (VAGs), as well as enterobacterial repetitive intergenic consensus-polymerase chain reaction (ERIC-PCR) assays were performed on 913 A. butzleri strains isolated from 11 agricultural sampling sites.ResultsAll strains were resistant to one or more antimicrobial agents, with a high rate of resistance to clindamycin (99%) and chloramphenicol (77%), followed by azithromycin (48%) and nalidixic acid (49%). However, isolates showed a significantly (p < 0.05) high rate of susceptibility to tetracycline (1%), gentamycin (2%), ciprofloxacin (4%), and erythromycin (5%). Of the eight VAGs tested, ciaB, mviN, tlyA, and pldA were detected at high frequency (> 85%) compared to irgA (25%), hecB (19%), hecA (15%), and cj1349 (12%) genes. Co-occurrence analysis showed A. butzleri strains resistant to clindamycin, chloramphenicol, nalidixic acid, and azithromycin were positive for ciaB, tlyA, mviN and pldA VAGs. ERIC-PCR fingerprint analysis revealed high genetic similarity among strains isolated from three sites, and the genotypes were significantly associated with AMR and VAGs results, which highlight their potential environmental ubiquity and potential as pathogenic.ConclusionsThe study results show that agricultural activities likely contribute to the contamination of A. butzleri in surface water. The findings underscore the importance of farm management practices in controlling the potential spread of A. butzleri and its associated health risks to humans and animals through contaminated water.

Clinical and epidemiological characteristics of multi-drug resistant Enterobacterales isolated from King Fahad Hospital of the University, AlKhobar, Saudi Arabia.

Multi-drug resistant (MDR) Enterobacterales remain a major clinical problem. Infections caused by carbapenem-resistant strains are particularly difficult to treat. This study aimed to assess the clinical and epidemiological characteristics of MDR Enterobacterales isolates. A total of 154 non-repetitive clinical isolates, including Escherichia coli (n = 66), Klebsiella pneumoniae (n = 70), and other Enterobacterales (n = 18), were collected from the Diagnostic Microbiology Laboratory at King Fahad Hospital of the University. Most E. coli isolates were collected from urine specimens (n = 50, 75.8%) and resistance against the third and fourth-generation cephalosporins (ceftriaxone, ceftazidime, cefixime, and cefepime) and fluoroquinolones (ciprofloxacin and levofloxacin) was assessed. Clonal relatedness analysis using enterobacterial repetitive intergenic consensus polymerase chain reaction (ERIC-PCR) revealed two clones (E. coli A and B), each comprising two strains. Most K. pneumoniae samples were collected from respiratory specimens (27.1%, 20 samples), and the strains showed overall resistance to most of the antimicrobials tested (54%‒100%). Moreover, clonal-relatedness analysis using ERIC-PCR revealed seven major clones of K. pneumoniae. These findings suggest nosocomial transmission among some identical strains and emphasize the importance of strict compliance with infection prevention and control policies and regulations. Environmental reservoirs could facilitate this indirect transmission, which needs to be investigated.

Isolation and Characterization of Cefotaxime and Ciprofloxacin Co-Resistant Escherichia coli in Retail Chicken Carcasses

Transmission of antimicrobial-resistant bacteria to humans through the food chain is of great importance for public health. In this study, it was aimed to isolate and characterize the cefotaxime and ciprofloxacin-resistant Escherichia coli in retail chicken meat samples sold in Hatay. The isolates were subjected to phylogenetic group typing and antimicrobial susceptibility testing. The genetic relatedness of the isolates was determined using Enterobacterial Repetitive Intergenic Consensus Polymerase Chain Reaction (ERIC-PCR) technique. The isolates were also screened for the presence of both antimicrobial and plasmid-mediated quinolone resistance (PMQR) genes by PCR. Cefotaxime and ciprofloxacin co-resistant E. coli isolates with diverse genetic origins were recovered in 42.3% (22/52) of retail chicken carcasses. The E. coli isolates belonged to the phylogenetic group D2 were dominant (40.9%, 9/22), followed by B1 (27.3%, 6/22), B23 (18.2%, 4/22), and A1 (13.6%, 3/22), respectively. Based on dendrogram analysis, the ERIC-PCR method differentiated the isolates into 10 clusters (I-X). The multidrug resistance (MDR) was observed in 81.8% (18/22) of the isolates. PMQR determinants were not identified in any isolates tested. Molecular analysis revealed one or more β-lactamase-encoding genes in all isolates as a single or in combination: blaCTX-M-blaTEM (n=5), blaCMY-2 (n=5), blaCTX-M (n=5), blaCMY-2-blaSHV (n=3), blaCMY-2-blaTEM (n=3), and blaCTX-M-blaCMY-2 (n=1). This study highlights that retail chicken meat is an important reservoir of cefotaxime and ciprofloxacin co-resistant E. coli isolates. It is necessary to evaluate their contribution to the community and hospital infections.

Ecological incidence, genetic diversity, multidrug resistance of Salmonella enteritidis recovered from broiler and layer chicken farms

Salmonella is a significant foodborne pathogen that has a significant impact on public health, and different strains of multidrug resistance (MDR) have been identified in this genus. This study used a combination of phenotypic and genotypic approaches to identify distinct Salmonella species collected from poultry broiler and layer farms, and antibiotic sensitivity testing was performed on these species. A total of 56 Salmonella isolates were serotyped, and phenotypic antibiotic resistance was determined for each strain. The enterobacterial repetitive intergenic consensus polymerase chain reaction (ERIC-PCR) method was also used to provide a genotypic description, from which a dendrogram was constructed and the most likely phylogenetic relationships were applied. Salmonella isolates were detected in 20 (17%) out of 117 samples collected from small-scale broiler flocks. Salmonella isolates were classified as MDR strains after showing tolerance to four antibiotics, but no resistance to cloxacillin, streptomycin, vancomycin, or netilmicin was observed. From a genotypic perspective, these strains lack dfrD, parC, and blasfo-1 resistant genes, while harboring blactx-M, blaDHA-L, qnrA, qnrB, qnrS, gyrA, ermA, ermB, ermC, ermTR, mefA, msrA, tet A, tet B, tet L, tet M resistance genes. The genotyping results obtained with ERIC-PCR allowed isolates to be classified based on the source of recovery. It was determined that Salmonella strains displayed MDR, and many genes associated with them. Additionally, the ERIC-PCR procedure aided in the generation of clusters with biological significance. Extensive research on Salmonella serotypes is warranted, along with the implementation of long-term surveillance programs to monitor MDR Salmonella serotypes in avian-derived foods.

Sociodemographic distributions and molecular characterization of colonized Enterococcus faecium isolates from locality hospitals in Khartoum, Sudan.

Enterococcus faecium is an opportunistic pathogen of humans with diverse hosts, encompassing animals as well as human beings. In the past twenty years, there has been a rise in the instances of nosocomial infections that are linked to antibiotic-resistant Enterococcus faecium. The acquisition of diverse antimicrobial resistance factors has driven the global development of robust and convergent adaptive mechanisms within the healthcare environment. The presence of microorganisms in hospitalized and non-hospitalized patient populations has been significantly aided by the facilitation of various perturbations within their respective microbiomes. This study aimed to determine the antimicrobial profile, demographic and clinical characteristics, along with the detection of virulence encoding genes, and to find out the clonal genetic relationship among colonized E. faecium strains. A hospital-based cross-sectional study was carried out between October 2018 and March 2020 at four Khartoum locality hospitals in Sudan. The study comprised a total of 108 strains of E. faecium isolated from patients admitted to four locality hospitals in Khartoum. A self-structured questionnaire was used to gather information on sociodemographic traits. Data were analyzed using chi-square test. In all cases, P value ≤ 0.05 with a corresponding 95% confidence interval was considered statistically significant. Moreover, enterobacterial repetitive intergenic consensus-polymerase chain reaction (ERIC-PCR) was utilized to assess the prevalence of clonal relationships, and the gel was analyzed using CLIQS software. In this study, the isolation rate of colonized E. faecium strains was 108/170 (63.5%). The colonization of E. faecium and its association with various sociodemographic and clinical features was examined. 73 (67.6%) of patients had multidrug-resistant (MDR), and 22 (20.4%) had extensively drug-resistant (XDR), 73 (67.6%) of patients engaged in self-medication practices. Eighty patients (74.1%) were non-adherence to prescribed antibiotics, while 70 (64.8%) patients reported recent antibiotic usage within the 3 months. The present study suggests that demographic factors may not be significantly associated with the incidence of E. faecium infection except for patients who had a prior history of antibiotic use (P≤0.005). The analysis of virulence genes showed a high prevalence of asa1 gene (22.2%) among strains. In ERIC-PCR the genetic relatedness of E. faecium showed seven identical clusters (A-G) with 100% genetic similarity. This implies clonal propagation in hospitals and communities. This study found that the incidence of E. faecium isolated from locality hospitals in Khartoum was likely due to the spread of E. faecium clones, thereby highlighting the need for intensifying infection control measures to prevent the spreading of nosocomial infection.

Molecular Typing of Multidrug-Resistant Acinetobacter baumannii Isolates from Clinical Specimens by ERIC-PCR and MLVA.

Acinetobacter baumannii, a Gram-negative and oxidase-negative bacterium, is a major cause of nosocomial infections, leading to high mortality rates in hospitalized patients. The use of 2 prominent molecular typing methods (i.e., enterobacterial repetitive intergenic consensus-polymerase chain reaction [ERIC-PCR] and multiple-locus variable-number tandem repeat [VNTR] analysis [MLVA]) for genotyping A. baumannii isolates has proven to be an effective approach in assessing the clonal relation of these isolates and managing their outbreaks. A total of 100 A. baumannii isolates were collected from immunocompromised patients hospitalized in the intensive care unit (ICU) of a hospital in Zanjan City, Iran. Their antibiotic resistance ability (especially aminoglycoside resistance) was studied by disc diffusion tests. The genetic typing of A. baumannii was studied using ERIC-PCR and MLVA methods. All isolates were resistant to 3 or more antibiotics and regarded as multidrug-resistant (MDR). Additionally, 32% of the isolates were resistant to all antibiotics tested, and 91% were extensively drug-resistant (XDR). The increased rate of aminoglycoside-resistant A. baumannii in ICU patients, with an increased incidence of aminoglycoside-modifying enzymes of aac (6')-Ib, ant (3″)-I, and aph (2″)-Id. ERIC-PCR has likewise shown an increased level of diversity in A. baumannii isolates. According to the ERIC-PCR patterns, isolates were classified as 4 clusters, while according to the MLVA patterns, isolates were classified as 9 distinct clusters. ERIC-PCR and MLVA assays serve as useful genotyping methods to assess the genetic variety or clonal relatedness of A. baumannii isolates.

Primary Carrier Status and Alteration in the Colonization of the Extended-Spectrum β-Lactamase Producing Enterobacteriaceae Among Children in Pediatric Intensive Care Unit During the Hospital Stay

Background: Widespread and inappropriate use of antibiotics has led to an increase in antibiotic-resistant bacteria worldwide. Extended-spectrum β-lactamases (ESBLs) are among the most important resistance mechanisms in members of Enterobacteriaceae, which can pose a threat to patients. Objectives: This study aimed to investigate the carrier status and alteration in the fecal transmission of ESBL-producing Enterobacteriaceae (ESBL-E) on admission and during the hospital stay, as well as its correlation with the usage of antibiotics among children in a pediatric intensive care unit (PICU). Molecular typing between the primary and secondary isolates was done to detect the homotypic clonal strains. Methods: Demographic and medical data of PICU children were collected, and the carrier status of ESBL-E was investigated in pairs of their rectal swab samples at the admission and discharge time. Detection of ESBL phenotype and antimicrobial susceptibility to 12 antibiotics were performed by double-disk synergy and disc diffusion methods, respectively. Polymerase chain reaction for detection of blaTEM, blaSHVblaPER, blaCTX-M, and blaVEB genes was performed using specific primers. The phylogenetic relations were analyzed by the enterobacterial repetitive intergenic consensus-polymerase chain reaction (ERIC-PCR) method. Results: Extended-spectrum β-lactamase-producing Enterobacteriaceae was detected in 48% of the samples at admission and 42% at discharge time. The highest frequency of resistance was observed in cephazolin, amoxicillin-clavulanic acid, and ampicillin. Children with a history of meropenem administration showed a significantly higher frequency of meropenem resistance Enterobacteriaceae in their samples. Moreover, the administration of metronidazole increased the isolation of ESBL-Escherichia species in hospitalized children. The most common gene associated with the ESBL phenotype was blaCTX-M, while blaPER and blaVEB were not detected in the ESBL-E isolates. The phylogenetic analysis did not confirm the occurrence of the cross-transmission of ESBL-E in the patients during hospitalization. Conclusions: Our results showed a high frequency of ESBL-E in the feces of children upon admission to the PICU, which did not change significantly during the hospital stay. Although the prescription of metronidazole showed an association with the enrichment of ESBL-E due to observed diversity in the molecular types and resistance phenotypes of the isolates, the community seems to be the primary source of ESBL-E transmission in children. Further investigations are needed to understand the role of hospital stay in the colonization and enrichment of ESBL-E in the intestinal tract of children and their association with infections during the medical interventions in the PICU.

Endogenous Bacteremia Caused by Intestinal Colonization of Carbapenem-Resistant Enterobacteriaceae (CRE) in Immunocompromised Children.

Carbapenem-resistant Enterobacteriaceae (CRE) infection is life-threatening, especially for immunocompromised children. The source tracking of CRE could prevent bacteremia during hospitalization. In this study, the intestinal colonization of CRE and their translocation to blood were investigated. Stool samples from immunocompromised pediatric patients were collected after admission, and secondary stool and blood samples were collected in case of fever. After CRE phonotypic detection, the OXA-48, NDM-1, VIM, IMP, and KPC genes were detected by PCR. Enterobacterial Repetitive Intergenic Consensus Polymerase Chain Reaction (ERIC-PCR) was used to determine the phylogenic relatedness of the blood and fecal isolates. Bacteremia was recorded in 71.4% of the patients. Enterobacteriaceae spp. were recorded in 100% of the stool samples and 31% of the blood samples. The correlation between the length of stay (LOS), days of fever, chemotherapy regimens, and death rate was significant (p-value ≤ 0.05). OXA-48 was present in all CRE isolates in both the primary and the secondary stool samples and the blood samples. According to the phylogenetic data, 58.33% of the patients with bacteremia had identical blood and stool isolates. The death rate was 24.4% in children with CRE bacteremia. The primary intestinal colonization with CRE in immunocompromised pediatrics and their translocation to blood was established in this study. The implementation of infection control programs and the application of infection prevention strategies for immunocompromised children is necessary.

Evidence-Based Interventions to Reduce the Incidence of Common Multidrug-Resistant Gram-Negative Bacteria in an Adult Intensive Care Unit.

Background Multidrug-resistant Gram-negative bacteria (MDR-GNB) present a significant and escalating hazard to healthcare globally. Context-specific interventions have been implemented for the prevention and control of MDR-GNB in several healthcare facilities. The objective of this study was to implement and evaluate the effectiveness of evidence-based interventions in the incidence and dissemination of MDR-GNB. Methods This was a pre-and post-intervention study conducted in three phases at King Abdulaziz Medical City Jeddah, Saudi Arabia. During Phase-1, the data on each of the four MDR-GNB (Acinetobacter baumannii, Klebsiella pneumoniae, Pseudomonas aeruginosa, and Escherichia coli) were collected prospectively. Genomic fingerprinting was performed on isolates using enterobacterial repetitive intergenic consensus-polymerase chain reaction (ERIC-PCR) to determine clonality and establish a link between different strains within and between the hospital wards/units. In the second phase, targeted interventions were implemented in the adult intensive care unit (ICU) based on previously determined risk factors and includedthe education of healthcare workers on hand hygiene, disinfection of patients' surrounding, daily chlorhexidine baths, and disinfection rooms on discharge with hydrogen peroxide fogging after MDR-GNB patients were discharged. An antibiotic restriction protocol was simultaneously implemented as part of the hospital antibiotic stewardship program. In the third phase, the effectiveness of the interventions was evaluated by comparing the incidence rate and clonality (using ERIC-PCR genetic fingerprints) of MDR-GNB before and after the intervention. Results A significant reduction of MDR-GNB was observed in Phase-2 and Phase-3 compared with Phase-1. The mean incidence rate of MDR-GNB per 1000 patient days in Phase-1 (pre-intervention) was 11.08/1000, followed by 6.07 and 3.54/1000 in Phase-2 and Phase-3, respectively. A statistically significant reduction was observed in the incidence rate of MDR-GNB in the adult ICU (P=0.007), whereas no statistically significant decrease (P=0.419) was observed in areas other than the adult ICU. Two A. baumannii strains appear to be circulating within the ICU environment with reduced frequency in Phase-2 and Phase-3 compared to Phase-1. Conclusion There was a significant reduction in the incidence of MDR-GNB in the adult ICU due to the successful implementation of both infection control and stewardship interventions, albeit challenging to ascertain the relative contribution of each.

Phenotypic and genetic screening of Klebsiella pneumoniae isolates from human UTI patients for beta-lactamases and their genetic diversity analysis by ERIC and REP PCRs.

Klebsiella pneumoniae is one of the major nosocomial pathogens responsible for pneumoniae, septicaemia, liver abscesses, and urinary tract infections. Coordinated efforts by antibiotic stewardship and clinicians are underway to curtail the emergence of antibiotic-resistant strains. The objective of the present study is to characterize K. pneumoniae strains through antibiotic resistance screening for production of beta-lactamases (β-lactamases) such as extended spectrum beta lactamases (ESBLs), AmpC β-lactamases, and carbapenemases by phenotypic and genotypic methods and genetic fingerprinting by enterobacterial repetitive intergenic consensus-polymerase chain reaction (ERIC-PCR) and repetitive element palindromic PCR (REP-PCR). A total of 85 K. pneumoniae strains isolated from 504 human urinary tract infections (UTI) were used in this study. Only 76 isolates showed positive in phenotypic screening test (PST), while combination disc method (CDM) as phenotypic confirmatory test (PCT) confirmed 72 isolates as ESBL producers. One or more β-lactamase genes were detected by PCR in 66 isolates (91.66%, 66/72) with blaTEM gene being the most predominant (75.75%, 50/66). AmpC genes could be detected in 21 isolates (31.8%, 21/66) with FOX gene being the predominant (24.24%, 16/66), whereas NDM-I was detected in a single strain (1.51%, 1/66). Genetic fingerprinting using ERIC-PCR and REP-PCR revealed wide heterogeneity among β-lactamase producing isolates with discriminatory power of 0.9995 and 1, respectively.

Microbiological Characterization and Clinical Facets of Elizabethkingia Bloodstream Infections in a Tertiary Care Hospital of Eastern India.

Elizabethkingia is an emerging non-fermenting Gram-negative bacillus (NFGNB) causing bloodstream infections (BSI) associated with high mortality. It demonstrates a unique antimicrobial profile in showing susceptibility to antimicrobials effective against Gram-positive bacteria. This study was undertaken to determine the overall frequency of Elizabethkingia BSI, associated risk factors, microbiological susceptibility, and clonal relationship of Elizabethkingia isolates using Enterobacterial Repetitive Intergenic ConsensusPolymerase Chain Reaction (ERIC-PCR). Elizabethkingia isolates obtained from the blood culture of admitted patients (August 2020-December 2021) were identified by the VITEK 2 system and subjected to an antimicrobial susceptibility test by standard procedures. Demographics, co-morbidities, risk factors for survival, and outcome were summarized and analyzed by Chi-square test, Kaplan-Meier curve, and Cox regression. Clonal relatedness between Elizabethkingia isolates was analyzed using ERIC‑PCR fingerprinting with the "PAST: Paleontological statistics software package". Of 13,747 blood samples received during the study period, 13.59% were culture positive, and 14.60% were NFGNBs. The frequency of Elizabethkingia spp. among all NFGNBs in BSI was 29.30%, and the overall prevalence in BSI was 4.21%. In patients with Elizabethkingia BSI, Foley's catheter was present in 81.25% of the cases. 100% susceptibility was observed to linezolid, followed by vancomycin (98.75%) and chloramphenicol (89.5%). The 30-day mortality rate in the patients of Elizabethkingia BSI was 26.25%. The Presence of COVID-19, pneumonia, diabetes mellitus (DM), mechanical ventilation (MV), and prior antibiotics were significantly different (p<0.05) between the survival and death groups. ERIC-PCR profile dendrogram of Elizabethkingia isolates showed ten major clusters indicating high genetic diversity. Elizabethkingia was responsible for one-third of NFGNB BSI in a single-center study, with approximately 26% of 30-day all-cause mortality. Most isolates were susceptible to linezolid, vancomycin, and chloramphenicol. COVID-19 was the most significant risk factor associated with mortality. ERIC-PCR of Elizabethkingia isolates exhibited high genetic diversity.

Isolation and Molecular Characterization of Corynebacterium pseudotuberculosis: Association with Proinflammatory Cytokines in Caseous Lymphadenitis Pyogranulomas.

Corynebacterium pseudotuberculosis (C. pseudotuberculosis) is a causative agent of numerous chronic diseases, including caseous lymphadenitis (CLA) in sheep and goats, which has a zoonotic potential in humans in addition to a poor therapeutic response. In this study, out of 120 collected samples, only 12 (10%) were positive for C. pseudotuberculosis by PCR and by intraperitoneal injection of male Guinea pigs and then characterized for antimicrobial susceptibility and its genetic-relatedness by enterobacterial repetitive intergenic consensus polymerase chain reaction (ERIC-PCR), which showed 2-4 bands ranging from 100 to 3000 bp that can be clustered into four clusters (C1-C4). Despite the serotype biovar 1 only infecting sheep and goats, ERIC-PCR reveals intra-subtyping variation. Examination of affected LNs and organs revealed marked enlargement with either thick creamy green pus or multiple abscesses of variable sizes with a central caseated core surrounded by dense fibrous capsule. A histopathological examination revealed a central necrotic core surrounded by a peripheral mantle of mononuclear cells and a fibrous capsule. Positive immune expression of nuclear factor kappa B (NF-κB/p65) and interleukin-1β (IL-1β) and negative expression of tumor necrosis factor (TNF) in CLA is the first report to our knowledge. Conclusion: In CLA pyogranulomas, IL1β is a more crucial proinflammatory cytokine than TNF in the regulation of C. pseudotuberculosis infection, which is accompanied by marked NF-κB immunoexpression. Therefore, the NF-κB/p65 signaling pathway is involved in the activation of IL1β, and additional immunohistochemical studies are required to determine the various roles of NF-κB/p65 in the inflammatory response within CLA pyogranulomas to control this pathogen.

First Report of Potentially Pathogenic Klebsiella pneumoniae from Serotype K2 in Mollusk Tegillarca granosa and Genetic Diversity of Klebsiella pneumoniae in 14 Species of Edible Aquatic Animals.

Klebsiella pneumoniae can cause serious pneumonitis in humans. The bacterium is also the common causative agent of hospital-acquired multidrug-resistant (MDR) infections. Here we for the first time reported the genetic diversity of K. pneumoniae strains in 14 species of edible aquatic animals sampled in the summer of 2018 and 2019 in Shanghai, China. Virulence-related genes were present in the K. pneumoniae strains (n = 94), including the entB (98.9%), mrkD (85.1%), fimH (50.0%), and ybtA (14.9%) strains. Resistance to sulfamethoxazole-trimethoprim was the most prevalent (52.1%), followed by chloramphenicol (31.9%), and tetracycline (27.7%), among the strains, wherein 34.0% had MDR phenotypes. Meanwhile, most strains were tolerant to heavy metals Cu2+ (96.8%), Cr3+ (96.8%), Zn2+ (91.5%), Pb2+ (89.4%), and Hg2+ (81.9%). Remarkably, a higher abundance of the bacterium was found in bottom-dwelling aquatic animals, among which mollusk Tegillarca granosa contained K. pneumoniae 8-2-5-4 isolate from serotype K2 (ST-2026). Genome features of the potentially pathogenic isolate were characterized. The enterobacterial repetitive intergenic consensus polymerase chain reaction&amp;nbsp;(ERIC-PCR)-based genome fingerprinting classified the 94 K. pneumoniae strains into 76 ERIC genotypes with 63 singletons, demonstrating considerable genetic diversity in the strains. The findings of this study fill the gap in the risk assessment of K. pneumoniae in edible aquatic animals.

Epidemiology, Evolution of Antimicrobial Profile and Genomic Fingerprints of Pseudomonas aeruginosa before and during COVID-19: Transition from Resistance to Susceptibility.

The purpose of the study was to describe the epidemiological implication of Pseudomonas aeruginosa between 2017-2022 in a tertiary hospital from Romania, including the molecular fingerprinting of similar phenotypic strains (multidrug-resistant isolates), which would have an important health impact. The study also describes the resistance profile of P. aeruginosa before and during COVID-19, which might bring new information regarding the management of antibiotic treatments. Information regarding wards, specimen types, species, and antibiotic resistance profile of 1994 strains of Pseudomonas spp. Isolated over a period of 6 years in Mures Clinical County Hospital, Romania, was collected from the WHONET database. From 50 multidrug-resistant isolates, molecular fingerprinting was performed by Enterobacterial Repetitive Intergenic Consensus Polymerase Chain Reaction (ERIC-PCR) to prove the potential clonal distribution. A number of 1994 Pseudomonas spp. were isolated between 2017-2022, from which P. aeruginosa was the most frequent species, 97.39% (n = 1942). P. aeruginosa was most frequently isolated in 2017 (n = 538), with the dermatology department as the main source, mainly from pus secretion. A drop in the harvesting rate was noted in 2020 due to COVID-19 restrictions. Regarding the resistance profile, there are a few modifications. The susceptibility of P. aeruginosa to carbapenems, piperacillin-tazobactam, and amikacin suffered alterations before and during COVID-19. The molecular fingerprinting showed three P. aeruginosa clusters, including strains with 80-99% similarity.