Enterobacterial Repetitive Intergenic Consensus PCR Research Articles

Distribution characteristics of integrons and correlation analysis of antibiotic resistance in urine isolated Enterobacter cloacae.

This study aims to understand the distribution of integrons among Enterobacter cloacae isolated from clinical urine specimens in our hospital, as well as the molecular characteristics of the variable region resistance gene cassette of integron-positive strains and its relationship with drug resistance. We collected a total of 80 strains of Enterobacter cloacae isolated from urine specimens of hospitalized patients in our hospital between August 2019 and July 2023, and conducted drug sensitivity testing on them. Polymerase Chain Reaction (PCR) technology was employed to screen these strains for Class 1, 2, and 3 integrons. Following this, the promoter and variable regions of integron-positive strains were amplified and sequenced. Additionally, Enterobacterial Repetitive Intergenic Consensus PCR (ERIC-PCR) was utilized for homology analysis of integron-positive strains. Among the 80 clinical strains, Class 1 integrons were detected in 31 (38.8%) strains, and the following resistance gene cassettes were identified: aadA2, aadA1, aadB, aac(6'), and catB8. Three types of variable region promoters were observed: PcS (4 strains), PcW (7 strains), and PcH1 (17 strains), with consistently inactive downstream P2 promoters. Additionally, Class 2 integrons were detected in 5 (6.3%) strains, carrying the variable region resistance gene cassette dfrA1-sat2-aadA1. The promoters for Class 2 integrons were uniformly of the Pc2D-Pc2A-Pc2B-Pc2C type. No Class 3 integrons were detected. The strains containing integrons showed significantly higher resistance rates to ciprofloxacin, compound sulfamethoxazole, levofloxacin, gentamicin, amikacin, and tobramycin compared to those without integrons (P<0.05). 35 strains of Enterobacter cloacae carrying integrons are primarily classified into three genotypes: A, B, and C. These genotypes are mainly distributed in the urology department and Intensive Care Unit (ICU). The distribution of variable region gene boxes and promoter types is relatively concentrated in the same genotype. Our study confirmed that Enterobacter cloacae isolated from urine samples predominantly carries Class 1 integrons with an extended array of antibiotic-resistant genes. For future research, it is recommended to explore additional resistance mechanisms and evaluate the effectiveness of new therapeutic strategies. Clinicians should be vigilant about the possibility of clonal dissemination and implement enhanced infection control measures in hospital settings.

Characterization of blaOXA-232 carrying carbapenem-resistant Klebsiella pneumoniae (CRKP) & their expression profiles under selective carbapenem pressure: An in-depth study from India.

Background & objectives OXA-232 is a five amino acid substitution variant of OXA-48 and is reported in carbapenem-resistant Klebsiella pneumoniae (CRKP), which is associated with nosocomial infections among immunocompromised patients in the intensive care unit. This study aimed to characterise blaOXA-232 in CRKP of clinical origin and investigate its transcriptional response against sub-inhibitory levels of carbapenems. Methods CRKP was isolated from blood (pathogens) and stool cultures (colonisers) of neonates and was characterized for blaOXA-232. Co-existing resistance determinants were investigated in blaOXA-232 positive isolates, followed by horizontal gene transferability assay and PCR-based replicon typing (PBRT). Cloning of blaOXA-232 was performed, and expression of blaOXA-232 in the isolates and their clones under sub-inhibitory concentrations of carbapenems was checked via RT-PCR. Mobile genetic elements associated with blaOXA-232 were investigated, followed by DNA fingerprinting through enterobacterial repetitive intergenic consensus (ERIC) PCR. Results blaOXA-232 with co-carriage of extended-spectrum beta-lactamases (ESBLs), sulphonamides and quinolones were identified in seven CRPK isolates recovered from blood samples of neonates. Transformation and cloning of blaOXA-232 was successful. The sub-inhibitory concentration of carbapenems induces elevated expression of this resistant determinant. ISEcp1 was associated with blaOXA-232 in the upstream region within two haplotypes of CRKP isolates of clinical origin. Interpretation & conclusions Selective carbapenem pressure resulted in higher expression of this gene, which could account for treatment failure. With frequent reports of occurrence among clinical isolates, monitoring and further investigation of this novel variant are necessary to understand its transmission dynamics and to thwart its further dissemination.

Prevalence of plasmid-mediated quinolone resistance genes and biofilm formation in different species of quinolone-resistant clinical Shigella isolates: a cross-sectional study

BackgroundThe purpose of this study was to look into the presence of plasmid-mediated quinolone resistance (PMQR) genes and biofilm formation in several species of clinical Shigella isolates that were resistant to quinolones.MethodsThe stool samples of 150 patients (younger than 10 years) with diarrhea were collected in this cross-sectional study (November 2020 to December 2021). After cultivation of samples on Hektoen Enteric agar and xylose lysine deoxycholate agar, standard microbiology tests, VITEK 2 system, and polymerase chain reaction (PCR) were utilized to identify Shigella isolates. The broth microdilution method was used to determine antibiotic susceptibility. PMQR genes including qnrA, qnrB, qnrC, qnrD, qnrE, qnrS, qnrVC, qepA, oqxAB, aac(6′)-Ib-cr, and crpP and biofilm formation were investigated in quinolone-resistant isolates by PCR and microtiter plate method, respectively. An enterobacterial repetitive intergenic consensus polymerase chain reaction (ERIC-PCR) technique was used to determine the clonal relatedness of quinolone-resistant isolates.ResultsA total of 95 Shigella isolates including S. sonnei (53, 55.8%), S. flexneri (39, 41.1%), and S. boydii (3, 3.2%) were identified. The highest resistance rates of the isolates were against ampicillin (92.6%, n = 88/95). Overall, 42 of 95 (44.2%) isolates were simultaneously resistant against two or more quinolones including 26 (61.9%) S. sonnei and 16 (38.1%) S. flexneri. All isolates were multidrug-resistant (resistance to more than 3 antibiotics). The occurrence of PMQR genes was as follows: qnrS (52.4%), qnrA and aac(6′)-Ib-cr (33.3%), and qnrB (19.0%). The prevalence in species was as follows: 61.5% and 37.5% (qnrS), 19.2% and 56.3% (qnrA), 38.5% and 25.0 (aac(6′)-Ib-cr), and 19.2% and 18.8% (qnrB) for S. sonnei and S. flexneri, respectively. The other PMQR genes were not detected. In total, 52.8% (28/53) of quinolone-susceptible and 64.3% (27/42) of quinolone-resistant isolates were biofilm producers. Biofilm formation was not significantly different between quinolone-resistant and quinolone-susceptible isolates (P-value = 0.299). Quinolone-resistant isolates showed a high genetic diversity according to the ERIC-PCR.ConclusionIt seems that qnrS, qnrA, and aac(6′)-Ib-cr play a significant role in the quinolone resistance among Shigella isolates in our region. Also the quinolone-resistant S. flexneri and S. sonnei isolates had a high genetic diversity. Hence, antibiotic therapy needs to be routinely revised based on the surveillance findings.

Isolation, identification, antibiotic resistance profile and molecular analysis of Ornithobacterium rhinotracheal isolates from turkeys.

Ornithobacterium rhinotracheal (ORT) infects numerous birds, particularly chickens and turkeys. ORT is an emerging bacterial pathogen of global concern in the poultry industry. As ORT is rapidly spreading throughout commercial poultry, it requires intensive studies of its epidemiology, diagnostic procedures, molecular typing, virulence genes and antimicrobial resistance. The present study was conducted in isolation and identification of ORT from slaughtered turkeys. Cleft palate swabs of 200 were collected from slaughtered turkeys and cultured on blood agar. ORT was characterized using biochemical tests and PCR targeting the ORT 16S rRNA gene. Virulence genes of isolates were determined targeting adenylate kinase (adk), copA and virulence-associated protein D (vapD) genes. Additionally, diversity of ORT isolates was performed by enterobacterial repetitive intergenic consensus (ERIC) and RAPD PCR. Disk diffusion was used to determine the antibiotic sensitivity of the isolates. ORT was identified in 23 (11.5%) samples using both the biochemical tests and PCR. The result of detecting virulence genes showed that all the isolates (23: 100%) had the adk gene, whereas two (8.7%) isolates had the copA gene, and seven (30.43%) isolates had the vapD gene. Molecular typing of isolates revealed 21 different patterns by RAPD PCR assay using M13 primer and 20 distinct patterns by ERIC PCR test. Both ERIC and RAPD PCR were distinctive methods for investigating the genetic diversity of ORT isolates. The antibiotic resistance test showed that 18 (78.26%) isolates were resistant to gentamicin, amikacin, cefazolin, streptomycin and penicillin. All isolates (100%) were resistant to cloxacillin and fosfomycin. This study showed the prevalence of ORT in turkey and high resistance of this bacterium to many common veterinary antibiotics. Moreover, both ERIC and RAPD PCR are distinctive methods for investigating the genetic diversity of ORT isolates. These data may help monitor antibiotic resistance and typing of ORT in epidemiological studies and serve as the foundation for designing region-specific vaccines for future use.

Molecular Epidemiological Characteristics of Carbapenem Resistant Aeromonas from Hospital Wastewater.

Hospital wastewater (HWW) promotes the spread of carbapenem resistance genes (CRGs). Aeromonas carry a large number of CRGs in HWW, they may play a role as a suitable reservoir for CRGs, while resistomes in HWW are still poorly characterized regarding carbapenem resistant Aeromonas. Thus, the aim of the study was to evaluate the molecular epidemiological characteristics of carbapenem resistant Aeromonas in HWW. A total of 33 carbapenem resistant Aeromonas were isolated from HWW. Antimicrobial susceptibility testing and polymerase chain reaction (PCR) were used to assess the antimicrobial resistance profiles. Molecular typing was performed using enterobacterial repetitive intergenic consensus PCR (ERIC-PCR) and multilocus sequence typing (MLST). The horizontal transmission mode of bla KPC was explored through conjugation and transformation experiments. The stability of bla KPC-IncP-6 plasmids was assessed through plasmid stability and in vitro competition test. The PCR mapping method was used to investigate the structural diversity of bla KPC. The detection rates of bla KPC and cphA in Aeromonas were 97.0% and 39.4% respectively. Aeromonas caviae were grouped into 13 clusters by ERIC-PCR and 12 STs by MLST. Aeromonas veronii were grouped into 11 clusters by ERIC-PCR and 4 STs by MLST. 56.3% bla KPC were located on mobilizable IncP-6 plasmids. bla KPC-IncP-6 plasmid showed high stability and low cost fitness. Carbapenem resistant Aeromonas from HWW mainly carried bla KPC, which exhibited great structural diversity. Aeromonas might serve as reservoirs for bla KPC and bla KPC might spread mainly through transformation in HWW.

Multidrug-resistant Escherichia coli strains isolated from swine manure biofertilizer in Brazil.

Escherichia coli is one of the key bacteria responsible for a variety of diseases in humans and livestock-associated infections around the globe. It is the leading cause of mortality in neonatal and weaned piglets in pig husbandry, causing diarrhea and significant harm to the industry. Furthermore, the frequent and intensive use of antimicrobials for the prevention of diseases, particularly gastrointestinal diseases, may promote the selection of multidrug-resistant (MDR) strains. These resistant genotypes can be transmitted through the excrement of animals, including swine. It is common practice to use porcine manure processed by biodigesters as fertilizer. This study aimed to examine the antimicrobial susceptibility, the presence of virulence genes frequently associated with pathotypes of intestinal pathogenic E. coli (InPEC), and antimicrobial resistance genes (ARGs) of 28 E. coli isolates collected from swine manure fertilizers. In addition, the enterobacterial repetitive intergenic consensus-PCR (ERIC-PCR) technique was used to investigate the genetic relationship among the strains. Using disk diffusion, the antimicrobial susceptibility profiles of the strains were determined. Using polymerase chain reaction (PCR), 14 distinct virulence genes associated with the most prevalent diarrhea and intestinal pathogenic E. coli (DEC/InPEC) and five ARGs were analyzed. All isolates tested positive for multidrug resistance. There was no detection of any of the 14 virulence genes associated with InPECs, indicating the presence of an avirulent commensal microbiota. Molecular classification by ERIC-PCR revealed that the majority of isolates (27 isolates) coalesced into a larger cluster with a genetic similarity of 47.7%; only one strain did not cluster in this cluster, indicating a high level of genetic diversity among the analyzed isolates. Thus, it is of the utmost importance to conduct epidemiological surveillance of animal breeding facilities in order to determine their microbiota and formulate plans to reduce the use of antimicrobials and improve animal welfare.

Antimicrobial Activity, Genetic Relatedness, and Safety Assessment of Potential Probiotic Lactic Acid Bacteria Isolated from a Rearing Tank of Rotifers (Brachionus plicatilis) Used as Live Feed in Fish Larviculture.

Aquaculture is a rapidly expanding agri-food industry that faces substantial economic losses due to infectious disease outbreaks, such as bacterial infections. These outbreaks cause disruptions and high mortalities at various stages of the rearing process, especially in the larval stages. Probiotic bacteria are emerging as promising and sustainable alternative or complementary strategies to vaccination and the use of antibiotics in aquaculture. In this study, potential probiotic candidates for larviculture were isolated from a rotifer-rearing tank used as the first live feed for turbot larvae. Two Lacticaseibacillus paracasei and two Lactiplantibacillus plantarum isolates were selected for further characterization due to their wide and strong antimicrobial activity against several ichthyopathogens, both Gram-positive and Gram-negative. An extensive in vitro safety assessment of these four isolates revealed the absence of harmful traits, such as acquired antimicrobial resistance and other virulence factors (i.e., hemolytic and gelatinase activities, bile salt deconjugation, and mucin degradation, as well as PCR detection of biogenic amine production). Moreover, Enterobacterial Repetitive Intergenic Consensus-PCR (ERIC-PCR) analyses unveiled their genetic relatedness, revealing two divergent clusters within each species. To our knowledge, this work reports for the first time the isolation and characterization of Lactic Acid Bacteria (LAB) with potential use as probiotics in aquaculture from rotifer-rearing tanks, which have the potential to optimize turbot larviculture and to introduce novel microbial management approaches for a sustainable aquaculture.

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.

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.

An emerging pathogen found in farm-cultured fourfinger threadfin (Eleutheronema tetradactylum), Edwardsiella piscicida, its genetic profile, virulent genes, and pathogenicity

The diseased fourfinger threadfin (Eleutheronema tetradactylum) exhibited clinical symptoms, such as hemorrhaging in the buccal area and skin. Gross pathological examination revealed hepatic hemorrhage, spleen enlargement, and brain congestion. Histopathological examination revealed multifocal necrosis and the presence of bacteria in various organs, including the heart, liver, spleen, and head kidney. Severe blood vessel congestion was also observed. All bacterial isolates obtained from the liver, spleen, head kidney, and brain were identified as gram-negative, motile, rod-shaped bacilli that displayed α-hemolysis. Utilizing a combination of the API 20 E system and species-specific primer-based polymerase chain reaction assays (EDtA, EDtD, and EDi primer sets), as well as sequencing of the 16S ribosomal DNA (16S rDNA) and iron-cofactor superoxide dismutase (sodB), the findings unequivocally established the identity of all isolates as E. piscicida. Examination of the E. piscicida isolates using transmission electron microscopy revealed the presence of flagella. Furthermore, genetic fingerprinting using enterobacterial repetitive intergenic consensus PCR (ERIC-PCR) demonstrated that the E. piscicida strains isolated from the fourfinger threadfin constituted a distinct genotype separate from the previously identified strains associated with Edwardsiellosis in different fish hosts. The pathogenicity of the representative E. piscicida isolates (E8–E12) was evaluated by subjecting the fourfinger threadfin to intraperitoneal injections. The gross lesions and histopathological alterations observed in experimentally infected fourfinger threadfins closely mirrored those observed in naturally infected fish. This study confirmed that E. piscicida-induced Edwardsiellosis could lead to disease outbreaks in farm-cultured fourfinger threadfin.

High levels of multidrug-resistant isolates of genetically similar Salmonella 1,4, [5],12:I:- from Brazil between 1983 and 2020.

Introduction. Salmonella 1,4, [5],12:i:- strains with different antimicrobial resistance profiles have been associated with foodborne disease outbreaks in several countries. In Brazil, S. 1,4, [5],12:i:- was identified as one of the most prevalent serovars in São Paulo State during 2004-2020.Gap Statement. However, few studies have characterized this serovar in Brazil.Aim. This study aimed to determine the antimicrobial resistance profiles of S. 1,4, [5],12:i:- strains isolated from different sources in Southeast Brazil and compare their genetic diversity.Methodology. We analysed 113 S. 1,4, [5],12:i:- strains isolated from humans (n=99), animals (n=7), food (n=5) and the environment (n=2) between 1983 and 2020. Susceptibility testing against 13 antimicrobials was performed using the disc diffusion method for all the strains. Plasmid resistance genes and mutations in the quinolone resistance-determining regions were identified in phenotypically fluoroquinolone-resistant strains. Molecular typing was performed using enterobacterial repetitive intergenic consensus PCR (ERIC-PCR) for all strains and multilocus sequence typing (MLST) for 40 selected strains.Results. Of the 113 strains, 54.87 % were resistant to at least one antimicrobial. The highest resistance rates were observed against ampicillin (51.33 %), nalidixic acid (39.82 %) and tetracycline (38.05 %). Additionally, 39 (34.51 %) strains were classified as multidrug-resistant (MDR). Nine fluoroquinolone-resistant strains exhibited the gyrA mutation (Ser96→Tyr96) and contained the qnrB gene. The 113 strains were grouped into two clusters using ERIC-PCR, and most of strains were present in one cluster, with a genetic similarity of ≥80 %. Finally, 40 strains were typed as ST19 using MLST.Conclusion. The prevalence of MDR strains is alarming because antimicrobial treatment against these strains may lead to therapeutic failure. Furthermore, the ERIC-PCR and MLST results suggested that most strains belonged to one main cluster. Thus, a prevalent subtype of Salmonella 1,4, [5],12:i:- strains has probably been circulating among different sources in São Paulo, Brazil, over decades.

Molecular typing of clinical and non-clinical &lt;i&gt;Escherichia coli&lt;/i&gt; from Rivers State, Nigeria using amplification-based RAPD and ERIC-PCR typing methods

Due to the diverse nature of Escherichia coli, typing of these organisms is essential. This can provide key epidemiological information not just on the strains in circulation and their relationship, but also on the development and spread of drug resistant clones. Affordable PCR-based typing techniques have resulted in an increase in typing studies particularly in resource limited settings. But information from Nigeria on strain typing is limited. This study was therefore aimed at typing clinical and non-clinical strains of E. coli from Nigeria using the PCR based Random Amplification of Polymorphic DNA (RAPD) and Enterobacterial Repetitive Intergenic Consensus-PCR (ERIC-PCR) typing methods. &#x0D; E. coli was isolated from clinical and non-clinical sources using Eosin Methylene Blue agar. Characteristic green metallic sheen colonies associated with E. coli were purified, identities determined using standard biochemical tests and confirmed using molecular methods. Susceptibility testing was carried out using the modified Kirby-Bauer disc diffusion test and RAPD and ERIC-PCR typing carried out on 48 confirmed E. coli isolates as previously described. ERIC-PCR typing resulted in the generation of 38 unique patterns with a 0.94 diversity. Nine of these were singletons comprised of only a single isolate, while the remaining 29 patterns were grouped to 10 clusters. RAPD gave similar results of 37 patterns, 9 singletons, 0.94 diversity and 13 clusters. For ERIC-PCR, all isolates with identical patterns were from the same source. RAPD, however, had identical patterns present in isolates from different sources. &#x0D; Using the ERIC-PCR typing method, this study was able to identify non-clinical isolates as distinct from the clinical E. coli isolates as evidenced by 100% identical ERIC-PCR profiles of isolates from the same source and a lack of 100% relatedness in isolates from different sources.

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.

Diversity analysis of livestock-associated Staphylococcus aureus (LA-SA) nasal strains between animal and humans

Background: Staphylococcus aureus has emerged as a major public health concern. It is a common pathogen in animal and human medicine. Aim: Enterobacterial Repetitive Intergenic Consensus Polymerase Chain Reaction (ERIC-PCR) was used to fingerprint 10 strains of S. aureus obtained from nasal swabs in domesticated animals and humans to ascertain how comparable the different strains' genetic makeup was. Methods: These isolates were previously identified using standard molecular and microbiological methods. ERIC primers were amplified for all isolates. The dendrogram was generated using PGMA and the Dice similarity coefficient. The strains were genotyped according to the diversity of sample source (human or animal), and the geographic source. Results: Ten S. aureus strains were classified into eight “ERIC” kinds (genotypes) using “ERIC-PCR genotyping”, in which the two most common clones were genotypes 8 and 2, which were represented by one strain from humans, one from cows, and two strains from sheep. Two strains derived from separate geographic areas and from different sample sources (human and cow) were determined to share the same genotype. Another two strains from different geographic areas but from the same sample source (sheep) were categorized under the same genotype. All the remaining strains were classified as a singular genotype. Conclusion: This study supports the possible bacterial transmission from animal to human and from animals themselves that usually happens during live animal marketing. Recognizing the interconnected nature of transmission systems and implementing the required approaches to disease prevention and control is essential for mitigating the risks posed by bacterial pathogens.

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 prevalence, genetic diversity, and 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.