Case-Control Study on the Role of Enterotoxigenic Bacteroides fragilis as a Cause of Diarrhea among Children in Kolkata, India

Abstract

A total of 874 fecal specimens (446 diarrheal cases and 428 controls) from diarrheal children admitted in the Infectious Diseases Hospital, Kolkata and age and sex matched asymptomatic subjects from an urban community were assessed for the prevalence of enterotoxigenic Bacteroides fragilis (ETBF). Isolates of B. fragilis were tested for the presence of enterotoxin gene (bft) by PCR. The detection rate of ETBF was 7.2% (63 of 874 specimens) that prevailed equally in diarrheal cases and controls (7.2% each; 32 of 446 cases and 31 of 428 controls). Male children up to one year age group was significantly (p<0.05) associated with ETBF infection as compared to children > 2 years of age in cases and controls. In 25 ETBF isolates, the bft gene was genotyped using PCR-RFLP and only two alleles were identified with prevalence rate of 40% and 60% for bft-1 and bft-3, respectively. All the ETBF isolates were susceptible for chloramphenicol and imipenem but resistant to clindamycin (48%), moxifloxacin (44%) and metronidazole (32%). Resistance of ETBF to moxifloxacin (44%) and metronidazole is an emerging trend. Pulsed-field gel electrophoresis (PFGE) revealed that majority of the ETBF isolates are genetically diverse. In the dendrogram analysis, two clusters were identified, one with ETBF resistant to 5–8 antimicrobials and the other cluster with metronidazole and moxifloxacin susceptible isolates from diarrheal cases. To our knowledge, this is the first detailed report on ETBF from India indicating its clinical importance and molecular characteristics.

Antimicrobial resistance, prevalence of resistance genes, and molecular characterization in intestinal Bacteroides fragilis group isolates

“…fragilis , Bile‐Esculin (BBE) agar (HiMedia Laboratories Pvt. Ltd, India) plate was used and incubated anaerobically (Whitley Jar Gassing System; Don Whitley Scientific, Bingley, UK) at 37 °C for 72 h. After incubation each of black or gray colony, raised circular colonies from each specimen were subcultured on Colombia blood agar (CBA) plate (bioMérieux, Marcy l'Etoile, France) and incubated anaerobically for 48 h. The identification of anaerobic Gram‐negative bacilli was done using API‐32A system (bioMérieux) .…”

Practical Guidance for Clinical Microbiology Laboratories: Diagnosis of Bacterial Gastroenteritis

“…A meta-analysis of 17 studies that evaluated the association of ETBF with diarrheal disease found that 12 (71%) of the studies demonstrated a higher frequency of ETBF in patients with diarrhea than in controls (211). In contrast, a recent Indian study found no difference in the rate of isolation of ETBF as a sole pathogen from children with and without diarrhea (212). This suggests that other, mitigating factors may play a role in the infective process for ETBF.…”

“…Potential B. fragilis isolates can be recovered from stool on Bacteroides bile esculin agar (Becton Dickinson, Sparks, MD) and then tested for enterotoxigenicity in vitro using PCR for the Bacteroides fragilis toxin gene (bft) (212). Alternatively, the cytopathic effect (CPE) produced by fragilysin on HT29/C1 (human colon) cell lines can be evaluated (211).…”

The prevalence of enterotoxin and antibiotic resistance genes in clinical and intestinal Bacteroides fragilis group isolates in Turkey

“…Enterotoxin producing B. fragilis (ETBF) has been implicated in enteritis, acute diarrheae in pediatric and adult populations and as an inciting agent in inflammatory bowel disease [11]. The prevalence of ETBF in the patients with diarrheae is reported to be 7,2%-15%; the asymptomatic carriage rate in the healthy individuals is reported as 6,5%-12,5% [12][13][14][15]. Interestingly, a recent study suggested yet another intriguing link between ETBF and human disease: in a cohort of Turkish patients, those with sporadic colorectal cancer had a higher prevalence of the bft gene by PCR than control patients-38% versus 12% [16].…”