Incidence Of Virulence Determinants Research Articles

Incidence of virulence determinants among Streptococcus agalactiae isolated from pregnant women and association with their serotypes

ABSTRACT Colonization of pregnant women by Streptococcus agalactiae (S. agalactiae) is the main reason for intrauterine infection or transmission during parturition. This study investigated the prevalence, antimicrobial susceptibility, serotypes, virulence traits of S. agalactiae in pregnant women across three hospitals in Dakahlia Governorate, Egypt. Isolates were identified by culturing onto Hichrome Sterp B selective medium, latex agglutination and biochemical tests. The antimicrobial susceptibility was determined by Kirby-Bauer disc diffusion method. Capsular serotypes were determined by latex agglutination test. Hemolysin production and biofilm formation were assessed quantitively. Virulence genes were detected by polymerase chain reaction (PCR). Among 290 samples, 31.7% were S. agalactiae. All isolates were sensitive to penicillin G, ampicillin, cefotaxime, and vancomycin, but notably resistant to tetracycline (83.6%), erythromycin (68.4%), and clindamycin (54.3%). Serotype III (26.08%) was most common. Strong biofilm was produced by 72.8% of isolates. High hemolytic activity was seen (80.4%). Prevalent virulence genes included fsbA (97.8%), fbsB (96.7%), and scpB (96.7%). rib and alp4 were frequent surface protein genes (60.8% and 57.6%, respectively). Serotypes were significantly correlated to clindamycin resistance, biofilm production (Ia, III, VI serotypes), hemolytic activity (serotype V), and several virulence genes and surface proteins. Our study found a high prevalence of S. agalactiae isolates in pregnant women with diverse resistance patterns, high virulence indicating potential for increased pathogenicity.

Genotypic and phenotypic insights into virulence factors of nosocomial Stenotrophomonas maltophilia isolates collected in Bulgaria (2011-2022).

The present study aimed to explore the virulence characteristics in 221 Bulgarian nosocomial Stenotrophomonas maltophilia isolates (2011-2022) via screening for the presence of virulence genes, their mutational variability, and the corresponding enzyme activity. PCR amplification, enzymatic assays, whole-genome sequencing (WGS), and biofilm quantification on a polystyrene plate were performed. The incidence of virulence determinants was as follows: stmPr1 (encoding for the major extracellular protease StmPr1) 87.3%, stmPr2 (minor extracellular protease StmPr2) 99.1%, Smlt3773 locus (outer membrane esterase) 98.2%, plcN1 (non-hemolytic phospholipase C) 99.1%, and smf-1 (type-1 fimbriae, biofilm-related gene) 96.4%. The 1621-bp allele of stmPr1 was most frequently found (61.1%), followed by the combined allelic variant (17.6%), stmPr1-negative genotype (12.7%), and 868-bp allele (8.6%). Protease, esterase, and lecithinase activity was observed in 95%, 98.2%, and 17.2% of the isolates, respectively. The WGS-subjected isolates (n = 9) formed two groups. Five isolates possessed only the 1621-bp variant of stmPr1, higher biofilm formation ability (Optical Density at λ = 550 nm (OD550): 1.253-1.789), as well as a low number of mutations in the protease genes and smf-1. Three other isolates had only the 868-bp variant, weaker biofilm production (OD550: 0.788-1.108), and higher number of mutations within these genes. The only weak biofilm producer (OD550 = 0.177) had no stmPr1 alleles. In conclusion, the similar PCR detection rates did not allow differentiation of the isolates. In contrast, WGS permitted stmPr1 alleles-based differentiation. To the best of our knowledge, this is the first Bulgarian study presenting genotypic and phenotypic insights into virulence factors of S. maltophilia isolates.

Incidence of virulence determinants and antibiotic resistance in lactic acid bacteria isolated from food products.

Background: Lactic acid bacteria (LAB) confer beneficial health effects in humans. However, the safety of these bacteria and their potential to spread resistance in the environment must be evaluated. Materials & methods: Fifty-three LAB were isolated from different food samplesand assessed for the prevalence of virulence determinants and antibiotic resistance profile. Results: Multiple resistance was reported for Lactobacillus brevis MIM04, having revealed phenotypic resistance to vancomycin (MIC >128μg/ml), ampicillin, cefotaxime, oxacillin and gentamicin. Virulence traits (cylA, gelE, esp and agg) were detected using specific primers. Enterococcus faecium CHE32, Lactobacillus plantarum CHE37 and E. faecium MLK68lack virulence genes, possess antimicrobial activity andsurvive in low pH and bile salt conditions. Conclusion: Isolated LAB revealed probiotic properties.

Characterization of virulence determinants and phylogenetic background of multiple and extensively drug resistant Escherichia coli isolated from different clinical sources in Egypt

Escherichia coli is a multifaceted microbe since some are commensals, normally inhabiting the gut of both humans and animals while others are pathogenic responsible for a wide range of intestinal and extra-intestinal infections. It is one of the leading causes of septicemia, neonatal meningitis, urinary tract infections (UTIs), cystitis, pyelonephritis, and traveler’s diarrhea. The present study aims to survey the distribution and unravel the association of phylotypes, virulence determinants, and antimicrobial resistance of E. coli isolated from different clinical sources in Mansoura hospitals, Egypt. One hundred and fifty E. coli isolates were collected from different clinical sources. Antimicrobial resistance profile, virulence determinants, and virulence encoding genes were detected. Moreover, phylogenetic and molecular typing using ERIC-PCR analysis was performed. Our results have revealed that phylogroup B2 (26.67%) with the greatest content in virulence traits was the most prevalent phylogenetic group. Different virulence profiles and varying incidence of virulence determinants were detected among tested isolates. High rates of resistance to different categories of antimicrobial agents, dramatic increase of MDR (92.67%), and emergence of XDR (4%) were detected. ERIC-PCR analysis revealed great diversity among tested isolates. There was no clustering of isolates according to resistance, virulence patterns, or phylotypes. Our research has demonstrated significant phylogenetic diversity of E. coli isolated from different clinical sources in Mansoura hospitals, Dakahlia governorate, Egypt. E. coli isolates are equipped with various virulence factors which contribute to their pathogenesis in human. The elevated rates of antimicrobial resistance and emergence of MDR and XDR mirror the trend detected globally in recent years.Key points• Clinical E. coli isolates exhibited substantial molecular and phylogenetic diversity.• Elevated rates of antimicrobial resistance and emergence of XDR in pathogenic E. coli.• B2 Phylogroup with the highest VS was the most prevalent among pathogenic E. coli.

Virulence determinants and biofilm formation in clinical isolates of Enterococcus: A cross-sectional study

Objective: To evaluate the relationship between biofilm formation and incidence of virulence determinants in clinical isolates of Enterococcus. Methods: In this cross-sectional study, the clinical isolates of Enterococcus strains were collected from the university teaching hospitals in Ahvaz, Iran from June 2017 to June 2018. Then, the prevalence of Enterococcus species, antibiotic resistance, virulence factors, and biofilm-producing ability were determined. Results: Of the 119 tested isolates, 17 (14.3%) were Enterococcus faecalis, 72 (60.5%) were Enterococcus faecium and 30 (25.2%) were other Enterococcus spp. Gelatinase was detected in 97 (81.5%) isolates, enterococcal surface protein in 41 (34.5%) isolates, serine protease in 39 (32.8%) cases, accessory colonization factor in 111 (93.3%) cases and pathogenicity islands in 17 (14.3%) cases. The biofilm formation ability was observed in 75 (63.0%) of all isolates and the association between the presence of enterococcal surface protein gene and biofilm formation was statistically significant. Higher resistance to vancomycin, gentamycin, and teicoplanin was indicated in Enterococcus faecium with 81.8%, 58.4%, and 85.7% resistance rate, respectively. All Enterococcus faecalis isolates were sensitive to teicoplanin and vancomycin. Conclusions: The presence of antibiotic-resistance with several virulence factors in Enterococcus spp has become a concern. High prevalence of enterococcal surface protein gene among biofilm- producing isolates suggests a potential relation between biofilm formation and the enterococcal surface protein gene, and further studies are needed to identify the mechanism of biofilm inhibition.

Incidence of Virulence Determinants Among Enterococcal Clinical Isolates in Egypt and Its Association with Biofilm Formation.

Background: Although Enterococci compromise an essential part of normal gut microbiota of both animals and humans, they have emerged as a leading opportunistic pathogen causing infections. The pathogenesis of enterococci is attributed to an array of virulence determinants. Objectives: This study aims to explore the prevalence and characteristics of enterococcal clinical isolates collected from Mansoura University Hospitals, Egypt, assess their ability to form biofilm, and the correlation with virulence determinants and antimicrobial resistance. Materials and Methods: A total of 70 Enterococcal clinical isolates were collected from different clinical sources between June and December 2016. Biofilm formation capacity was assessed, and characterization of virulence factors and antibiotic susceptibility was performed. Clonal relatedness between isolates was assessed using enterobacterial repetitive intergenic consensus-PCR (ERIC-PCR) approach. Results and Conclusion: The molecular analysis demonstrated high genetic diversity among enterococcal clinical isolates. The gelE was the most frequently detected gene (91.4%), followed by asa1 (70%), esp (65.7%), and cylA (17.1%), while hyl was not detected in any isolate. Gelatinase activity was detected in 35.7%, while hemolysin and lipase activity was detected in 12.9% and 78.5%, respectively. Most of the enterococcal isolates were biofilm producers, of which 67.1% were strong/moderate biofilm producers. All linezolid-resistant isolates exhibited strong/moderate biofilm formation capacity. Strong/moderate biofilm formation was more frequently observed among esp-positive (esp+) and gelatinase nonproducing (gelatinase-) enterococcal isolates. Multiple regression analysis denoted that esp (odds ratio [OR] 5.371, p = 0.003) and gelatinase production (OR 0.264, p = 0.015) were associated with strong/moderate biofilm formation capacity. These findings suggest that esp gene positivity and gelatinase production may affect biofilm formation capacity among enterococcal clinical isolates.

Incidence of virulence determinants in clinical Enterococcus faecalis and Enterococcus faecium isolates collected in Bulgaria

ObjectivesTo evaluate the prevalence of some virulence genes among 510 clinical Enterococcus spp. isolates and to assess the association of those genes with the species, infection site, and patient group (inpatients/outpatients). MethodsAdhesins genes (aggregation substances agg and asa1 of Enterococcus faecalis and Enterococcus faecium, respectively), enterococcal surface protein (esp), endocarditis-specific antigen A (efaA), collagen-binding proteins (ace/acm)); invasins (hyaluronidase (hyl) and gelatinase (gelE)); cytotoxines (activation of cytolysin (cylA) in E. faecalis); and modulators of the host immunity and inflammation (enhanced expression pheromone (eep) in E. faecalis) were detected by polymerase chain reaction. ResultsThe overall prevalence was: esp – 44.3%, agg/asa1 – 38.4%, ace/acm – 64.3%, efaA – 85.9%, eep – 69.4%, gelE – 64.3%, hyl – 25.1%, and cylA – 47.1%. E. faecalis isolates had significantly higher frequency of adhesin genes (esp and agg/asa1) and gelatinase in comparison to E. faecium. Multiple virulence genes in E. faecalis were significantly more prevalent than in E. faecium isolates. Domination of E. faecium with or without only one gene compared to the isolates of E. faecalis were found. Enterococcus spp. isolates obtained from outpatients compared to inpatients isolates had significantly higher frequency of agg/asa1, eep, gelE and cylA. Some adhesins genes (esp, agg/asa1 and efaA) had higher prevalence among the non-invasive Enterococcus spp. isolates compared to those causing invasive bacteremia, while ace/acm revealed higher dissemination in isolates causing invasive infections compared to non-invasive isolates. ConclusionMost E. faecalis attaches to abiotic surfaces in hospital environment, which correlates with higher prevalence of gene encoding for virulence factors involved in biofilm formation, such as enterococcal surface protein, aggregation substance, and gelatinase. The intestinal tract is an important reservoir for opportunistic enterococcal pathogens and allows them to access infectious sites through different virulence factors, demonstrated in outpatient isolates in this study.

Assessment of Safety of Enterococci Isolated throughout Traditional Terrincho Cheesemaking: Virulence Factors and Antibiotic Susceptibility

Enterococci account for an important fraction of the adventitious microflora of traditional cheeses manufactured in Mediterranean countries from small ruminants' raw milk and play an important role in the development of suitable organoleptic characteristics of the final product. It has been suggested that animals used for food or animals that supply edible products are a reservoir of antibiotic-resistant enterococci. The main purpose of this research effort was thus to identify, to the species level, a total of 73 enterococci with high tolerance to acidic pH and bile salts (as prevailing environmental conditions in the first portion of the gastrointestinal tract), which were previously isolated from the milk feedstock to the final product of Terrincho cheesemaking, and to determine their profiles of antibiotic susceptibility, coupled with the occurrence of specific virulence factors (especially in those that might eventually be claimed to exhibit suitable probiotic and technological performances). Isolates, identified by both API 20 STREP and PCR methods, were found to belong to the following Enterococcus species: E. casseliflavus, E. durans, E. faecalis, E. faecium, and E. gallinarum. Susceptibility of those isolates was observed to most antibiotics tested, whereas none harbored aminoglycoside resistance genes. PCR screenings for cytolysin genes (cylLL, cylLs, cylM, cylB, and cylA), surface adhesin genes (efaAfs, efaAfm, and esp), the aggregation protein gene (agg), and the extracellular metalloendopeptidase gene (gelE) were performed. All isolates proved negative for cylLL, cylM, cylB, and agg genes. Both E. faecalis strains were positive for the cell wall–associated protein Esp and the cell wall adhesin efaAfs, whereas the cell wall adhesin efaAfm was detected in 11 of the 12 E. faecium strains. Only one strain possessed the cylLs determinant, and another possessed the cylA gene. Incidence of virulence determinants was thus very low; hence, the enterococcal adventitious microflora tested is essentially safe.

VanA-Type Enterococci from Humans, Animals, and Food: Species Distribution, Population Structure, Tn 1546 Typing and Location, and Virulence Determinants

VanA-type human (n=69), animal (n=49), and food (n=36) glycopeptide-resistant enterococci (GRE) from different geographic areas were investigated to study their possible reservoirs and transmission routes. Pulsed-field gel electrophoresis (PFGE) revealed two small genetically related clusters, M39 (n=4) and M49 (n=13), representing Enterococcus faecium isolates from animal and human feces and from clinical and fecal human samples. Multilocus sequence typing showed that both belonged to the epidemic lineage of CC17. purK allele analysis of 28 selected isolates revealed that type 1 was prevalent in human strains (8/11) and types 6 and 3 (14/15) were prevalent in poultry (animals and meat). One hundred and five of the 154 VanA GRE isolates, encompassing different species, origins, and PFGE types, were examined for Tn1546 type and location (plasmid or chromosome) and the incidence of virulence determinants. Hybridization of S1- and I-CeuI-digested total DNA revealed a plasmid location in 98% of the isolates. Human intestinal and animal E. faecium isolates bore large (>150 kb) vanA plasmids. Results of PCR-restriction fragment length polymorphism and sequencing showed the presence of prototype Tn1546 in 80% of strains and the G-to-T mutation at position 8234 in three human intestinal and two pork E. faecium isolates. There were no significant associations (P>0.5) between Tn1546 type and GRE source or enterococcal species. Virulence determinants were detected in all reservoirs but were significantly more frequent (P<0.02) among clinical strains. Multiple determinants were found in clinical and meat Enterococcus faecalis isolates. The presence of indistinguishable vanA elements (mostly plasmid borne) and virulence determinants in different species and PFGE-diverse populations in the presence of host-specific purK housekeeping genes suggested that all GRE might be potential reservoirs of resistance determinants and virulence traits transferable to human-adapted clusters.

Incidence of virulence determinants in enterococcal strains of probiotic and clinical origin

Enterococci are gram-positive bacteria that are normal inhabitants of the alimentary tract of humans and other animals. They are often used as starter or protection cultures or food supplements as well as probiotics. Virulence genes pattern might be considered an important criterion for evaluation of the strain potential for pathogenicity. Enterococcal strains used as starter cultures should be tested for the presence of virulence determinants for safety considerations.

Enterococci from Tolminc cheese: Population structure, antibiotic susceptibility and incidence of virulence determinants

Microbiological analysis of ripened artisanal Tolminc cheese revealed the presence of an enterococcal population in numbers of up to 10 6 per g. All colonies, isolated from the citrate azide tween carbonate (CATC) enterococcal selective medium were Gram positive and coccal-shaped and were analysed with PhenePlate™ FS system. This system discriminated 10 PhP clusters among the 90 enterococcal isolates. From each cluster the most representative isolate for that particular type was selected for further study. The 10 representative enterococci were catalase negative and grew in the presence of NaCl (2%, 4% and 6.5%) and bile salts (0.06%). Genus specific primers confirmed all 10 enterococcal representatives as Enterococcus members, while species specific primers determined them further as strains of Enterococcus faecalis species. PCR for vanA and vanB genes detection, respectively, amplified no PCR products. The absence of van genes was confirmed with both disc and E-test, as isolates were susceptible to vancomycin according to the National Committee for Clinical Laboratory Standards (NCCLS). The results of disc tests with other antimicrobial agents (ampicillin, vancomycin, kanamycin, penicillin, erythromycin, neomycin, chloramphenicol, clindamycin, rifampin) did not differ much among the tested enterococci: they were all very resistant to clindamycin only. The incidence of enterococcus virulence determinants was as expected: all of the 10 E. faecalis strains tested possessed multiple determinants (between 7 and 11).

Incidence of virulence determinants in clinical Enterococcus faecium and Enterococcus faecalis isolates collected in Sardinia (Italy).

Enterococci are widely distributed in the environment; within the human body, they are normal commensals of the oral cavity, gastrointestinal tract and vagina. In recent years, enterococci have become one of the most frequent causes of acquired nosocomial infections worldwide. The molecular mechanism of virulence of these bacteria is still not completely understood. The aims of this work were to characterize phenotypically 47 isolates of Enterococcus faecalis and Enterococcus faecium collected in Sardinia (Italy) by their abilities to adhere to different epithelial cell lines (Vero and Caco-2 cells) and to associate their phenotypes with the presence of known virulence genes detected within their genomes by PCR. The following genes were amplified: AS (aggregation substance), esp (surface protein gene), ace (accessory colonization factor), efaA (E. faecalis endocarditis antigen) and gelE (gelatinase). The virulence genes were detected in E. faecalis isolates only, with the exception of esp, which was found in both species. The phenotypic and genotypic results were also compared with the susceptibility of isolates to various antibiotics.