Molecular analysis of accessory gene regulator functionality and virulence genes in Staphylococcus aureus derived from pediatric wound infections

Staphylococcus aureus is a major human pathogen causing infections with high morbidity and mortality in both healthcare and community settings. The accessory gene regulator (Agr) is a key genetic element controlling the expression of numerous virulence factors in S. aureus. The significance of a functional Agr system in clinical S. aureus isolates derived from pediatric wound infections is still unclear. Therefore, the present study was conducted to identify virulence genes and determine Agr functionality from this cohort of patients. A total of 48 S. aureus wound isolates were collected from patients referred to Tehran Children's Medical Center Hospital from April 2017 to April 2018. In addition, in vitro antimicrobial susceptibility of the isolates was assessed using the disk diffusion and E-test methods. Conventional PCR was performed for the detection of toxins (tsst-1, hla, hlb, hld, eta, etb, etd, edin-A, edin-B, edin-C) and Agr typing (agrI, agrII, agrIII, agrIV). Agr functionality was assessed by quantitative reverse transcriptase real-time PCR (qRT-PCR). All S. aureus isolates were found to be susceptible to linezolid and vancomycin. The most frequently detected toxin gene was eta (100%), and the most prevalent Agr type was agrIII (56.3%). Importantly, qRT-PCR revealed that Agr was functional in 28 (58%) of wound isolates. Consequently, our data suggests that a functional Agr system may not be required for the development of S. aureus wound infections.</p

Molecular analysis of accessory gene regulator functionality and virulence genes in Staphylococcus aureus derived from pediatric wound infections

Staphylococcus aureus is a major human pathogen causing infections with high morbidity and mortality in both healthcare and community settings. The accessory gene regulator (Agr)is a key genetic element controlling the expression of numerous virulence factors in S. aureus. The significance of a functional Agr system in clinical S. aureus isolates derived from pediatric wound infections is still unclear. Therefore, the present study was conducted to identify virulence genes and determine Agr functionality from this cohort of patients. A total of 48 S. aureus wound isolates were collected from patients referred to Tehran Children's Medical Center Hospital from April 2017 to April 2018. In addition, in vitro antimicrobial susceptibility of the isolates was assessed using the disk diffusion and E-test methods. Conventional PCR was performed for the detection of toxins (tsst-1, hla, hlb, hld, eta, etb, etd, edin-A, edin-B, edin-C)and Agr typing (agrI, agrII, agrIII, agrIV). Agr functionality was assessed by quantitative reverse transcriptase real-time PCR (qRT-PCR). All S. aureus isolates were found to be susceptible to linezolid and vancomycin. The most frequently detected toxin gene was eta (100%), and the most prevalent Agr type was agrIII (56.3%). Importantly, qRT-PCR revealed that Agr was functional in 28 (58%)of wound isolates. Consequently, our data suggests that a functional Agr system may not be required for the development of S. aureus wound infections.</p

Distribution of capsular types and drug resistance patterns of invasive pediatric Streptococcus pneumoniae isolates in Teheran, Iran

Objectives: To explore the serotype distribution and drug resistance patterns of invasive pneumococcal isolates from children under 5 years of age. Methods: During a 32-month period, 585 clinical samples (including blood, cerebrospinal fluid (CSF), and synovial fluid) from children suspected of having meningitis, sepsis, pneumonia, or septic arthritis were analyzed using the BACTEC culture system. Positive cultures were examined using biochemical tests and lytA amplification for the identification of pneumococcal strains. The confirmed pneumococcal isolates were examined to determine capsular types using a modified sequential multiplex PCR and susceptibility to antimicrobial agents. Results: Fifty-three pneumococcal isolates were detected in the 585 clinical samples: 21 (39.6%) blood samples and 32 (60.4%) CSF samples. The most frequent serotype was 23F (24.5%), followed by serotypes 19F (18.9%), 19A (7.5%), and 9V (7.5%). Twenty-one percent of pneumococcal isolates were penicillin-non-susceptible and serotype 19A was significantly associated with resistance to penicillin. Conclusions: This study indicated that the 13-valent pneumococcal conjugate vaccine (PCV13) could cover the majority of the invasive pneumococcal isolates. Drug-resistant and multidrug-resistant Streptococcus pneumoniae strains are circulating in Iran. Therefore, public immunization of infants using PCV13 is recommended to reduce the incidence of pneumococcal disease and pneumococcal-resistant strains in Teheran

High prevalence of Mucosa-Associated extended-spectrum β-Lactamase-producing Escherichia coli and Klebsiella pneumoniae among Iranain patients with inflammatory bowel disease (IBD)

Abstract Background Several pieces of evidence suggest that certain pathobionts belonging to Enterobacterales are associated with the development and progression of inflammatory bowel diseases (IBD). Extended-spectrum β-lactamases (ESBLs) ESBLs are frequently found in the Enterobacterales members, particularly in Escherichia coli and Klebsiella spp., and might trigger antibiotic-induced perturbations of the intestinal microbiota and led to more severe disease activity in IBD. Therefore, the severity of IBD could be influenced by ESBL-producing Enterobacterales, and hence, this study aimed to investigate the presence of ESBLs and carbapenemases among mucosa-associated E. coli and Klebsiella pneumoniae isolated from colonic biopsies of Iranian patients with IBD. Methods In this cross-sectional study, E. coli and K. pneumoniae were isolated from inflamed ileum and/or colon tissue of patients with IBD, including Ulcerative colitis (UC) and Crohn’s disease (CD), during colonoscopy. Demographic data and clinical characteristics were recorded, and UC and CD disease activity and extent were evaluated according to the full Mayo score and Crohn’s disease activity index (CDAI), respectively. Phenotypic and molecular detection of ESBL- and carbapenemase-producing E. coli and Klebsiella pneumoniae were carried out. Disease activity and other clinical and microbial features were compared in patients with and without gut colonization with ESBL producers. Results A total of 83 IBD patients, including 67 UC and 16 CD, were enrolled in the initial analysis. Intestinal colonization with ESBL-producing E. coli and/or Klebsiella pneumoniae was found in 37 (55.2%) of UC and 9 (56.2%) of DC patients – mostly harbored E. coli containing the bla CTX−M and bla TEM genes. UC patients with intestinal colonization with ESBL-producers had more severe disease compared with patients without colonization. Moreover, 10.2% of tested E. coli and 34.8% of K. pneumoniea were recognized as potential carbapenemase producers. Conclusion Intestinal colonization with ESBL producers could arise disease activity in IBD patients. Further large-scale case-control studies should be performed to investigate the possible confounding factors that could contribute to this outcome

Investigation and characterization of human gut phageome in advanced liver cirrhosis of defined etiologies.

BACKGROUND: Liver cirrhosis is a major public health problem, accounting for high rates of morbidity and mortality worldwide. The cirrhosis etiology is a broad and essential step in planning a treatment strategy. Many recent studies have documented that gut microbiome alterations play a vital role in the development and progression of cirrhosis and its complications. Nevertheless, there is insufficient data on the correlation between liver cirrhosis and gut phageome alterations in patients with advanced liver diseases. This study aimed to analyze the taxonomic structure and functional attributes of the gut phageome in six different etiologies of advanced liver cirrhosis. METHODS: We first retrieved metagenomic sequencing data from three datasets of fecal samples taken from cirrhotic patients with various etiologies. Subsequently, several bioinformatics pipelines were used to analyze bacteriophage composition and determine their functionality. RESULTS: A gene catalog of 479,425 non-redundant genes was developed as a reference to measure gene prevalence. The results of the analysis revealed a few significant differences among the cohorts at the phage species level. However, the alternations were more evident as there were more in-depth analyses of the functional resolution of the gut phageome. CONCLUSIONS: Our findings suggest that the functional analysis of the gut phageome would provide meaningful markers to predict the progression of liver cirrhosis and facilitate the development of novel treatment approaches

Evaluation of Biofilm Formation Among Klebsiella pneumoniae Isolates and Molecular Characterization by ERIC-PCR

AbstractBackground: Klebsiella pneumoniae is among the most frequently recovered etiologic agents from nosocomial infections. Thisopportunistic pathogen can generate a thick layer of biofilm as one of its important virulence factors, enabling the bacteria to attach toliving or abiotic surfaces, which contributes to drug resistance. Objectives: The resistance of biofilm-mediated infections to effective chemotherapy has adverse effects on patient outcomes and survival.Therefore, the aim of the present study was to evaluate the biofilm-formation capacity of clinical K. pneumoniae isolates and to perform amolecular characterization using enterobacterial repetitive intergenic consensus-polymerase chain reaction (ERIC-PCR) to determine thedominant biofilm-producing genotype. Patients and Methods: In the present study, 94 K. pneumoniae isolates were obtained from two hospitals in Tehran, Iran. Biofilm formationwas assayed by a modified procedure, then ERIC-PCR was carried out. Results: The distributions of the clinical specimens used in this study were 61.7% from urine, 18.1% from wounds, 11.7% from sputum, and8.5% from blood. Among these isolates, 33% formed fully established biofilms, 52.1% were categorized as moderately biofilm-producing,8.5% formed weak biofilms, and 6.4% were non-biofilm-producers. Genotyping of K. pneumoniae revealed 31 different ERIC types. Biofilmformationability in a special ERIC type was not observed. Conclusions: Our results indicated that an enormous proportion of K. pneumoniae isolated from sputum and surgical-wound swabsproduced fully established biofilms. It is reasonable to assume the existence of a relationship between the site of infection and theformation of biofilm. A high level of genetic diversity among the K. pneumoniae strains was observed

Antibacterial, anti-swarming and anti-biofilm formation activities of Chamaemelum nobile against Pseudomonas aeruginosa

AbstractINTRODUCTION:Chamomile ( Chamaemelum nobile ) is widely used throughout the world, and has anti-inflammatory, deodorant, bacteriostatic, antimicrobial, carminative, sedative, antiseptic, anti-catarrhal, and spasmolytic properties. Because of the increasing incidence of drug-resistant bacteria, the development of natural antibacterial sources such as medical herbs for the treatment of infectious diseases is necessary. Extracts from different plant parts such as the leaves, flowers, fruit, and bark of Combretum albiflorum, Laurus nobilis , and Sonchus oleraceus were found to possess anti-quorum sensing (QS) activities. In this study, we evaluated the effect of C. nobile against Pseudomonas aeruginosa biofilm formationMETHODS:The P. aeruginosa samples were isolated from patients with different types of infection, including wound infection, septicemia, and urinary tract infection. The flowers of C. nobile were dried and the extract was removed using a rotary device and then dissolved in dimethyl sulfoxide at pH 7.4. The microdilution method was used to evaluate the minimum inhibitory concentration (MIC) of this extract on P. aeruginosa , and biofilm inhibition was assayed.RESULTS:Eighty percent of the isolated samples (16/20) could form a biofilm, and most of these were isolated from wound infections. The biofilm inhibitory concentration of the C. nobile extract was 6.25-25mg/ml, whereas the MIC was 12.5-50mg/ml.CONCLUSIONS:The anti-QS property of C. nobile may play an important role in its antibacterial activity, thus offering an additional strategy in the fight against bacterial infections. However, molecular investigation is required to explore the exact mechanisms of the antibacterial action and functions of this phytocompound

Genetic diversity of Helicobacter pylori type IV secretion system cagI and cagN genes and their association with clinical diseases

Abstract A number of cagPAI genes in the Helicobacter pylori genome are considered the most evolved genes under a diversifying selection and evolutionary pressure. Among them, cagI and cagN are described as a part of the two different-operon of cagPAI that are involved in the T4SS machinery, but the definite association of these factors with clinical manifestations is still unclear. A total of 70 H. pylori isolates were obtained from different gastroduodenal patients. All isolates were examined for the presence of primary H. pylori virulence genes by PCR analysis. Direct DNA sequence analysis was performed for the cagI and cagN genes. The results were compared with the reference strain. The cagI, cagN, cagA, cagL, vacA s1m1, vacA s1m2, vacA s2m2, babA2, sabA, and dupA genotypes were detected in 80, 91.4, 84, 91.4, 32.8, 42.8, 24.4, 97.1, 84.3, and 84.3% of the total isolates, respectively. The most variable codon usage in cagI was observed at residues 20–25, 55–60, 94, 181–199, 213–221, 241–268, and 319–320, while the most variable codon usage in CagN hypervariable motif (CagNHM) was observed at residues 53 to 63. Sequencing data analysis of cagN revealed a hypothetical hexapeptide motif (EAKDEN/K) in residues of 278–283 among six H. pylori isolates, which needs further studies to evaluate its putative function. The present study demonstrated a high prevalence of cagI and cagN genes among Iranian H. pylori isolates with gastroduodenal diseases. Furthermore, no significant correlation between cagI and cagN variants and clinical diseases was observed in the present study. However, all patients had a high prevalence of cagPAI genes including cagI, cagN, cagA, and cagL, which indicates more potential role of these genes in disease outcome