Background: Some strains of Bacteroides fragilis species are associated with diarrhea as a result of enterotoxin production (bft or fragilysin). Fragilysin is activated by C11 protease (fpn) and together with C10 protease (bfp) play a significant role in its invasiveness. The objectives of this study were to investigate the proportion of clinical isolates from extra-intestinal sources that are toxin producers and characterize the genes mediating toxin production. Clinical isolates submitted to our reference laboratory over the last 13 years were screened for toxin production using PCR technique. All stool isolates were excluded. The isolates were tested for their susceptibility to 8 antimicrobial agents by E test. Carbapenem resistance gene cfiA was detected by PCR.
Results: A total of 421 B. fragilis isolates were viable. Out of these, bft was detected in 210 (49.9%) isolates. Of the 210 bft-positive isolates, 171 (81.4%), 33 (15.7%) and 6 (2.8%) harbored bft-1, bft-2, and bft-3 genes, respectively. Twenty (9.5%) of the bft-positive strains originated from bloodstream infections. Twenty-five, 20 and 9 strains harbored bfp-1, bfp-2 and bfp-3 gene, respectively. Two, 3, 4 bfp isotypes were detected simultaneously in some of strains. The resistance rates against amoxicillin-clavulanic acid was 32%, clindamycin 62%, cefoxitin 26%, imipenem 11%, meropenem 17%, metronidazole 4%, piperacillin 61% and tigecycline 14%. A chromosomally located cfiA gene that encode metallo-β-lactamase was identified in only 34 isolates (16.2%).
Conclusions: The prevalence of enterotoxin-producing B. fragilis was high among the extra-intestinal isolates. Metronidazole was the most active agent against all isolates. There was no statistically significance difference between resistance rates among bft-positive and bft-negative isolates except for clindamycin.
Garrett WS, Onderdonk AB. B. Bacteroides, Prevotella, Porphyromonas and Fusobacterium species (and other medically important anaerobic gram-negative bacilli) In: Mandell GL, Benett JE, Dolin R, editors. Mandell, Douglas and Bennett’s principles and practice of infectious diseases. 8. New York: Churchill Livingston; 2015. pp. 2773–2780.
Jandhyala SM, Talukdar R, Subramanyam C, Vuyyuru H, Sasikala M, Nageshwar RD. Role of the normal gut microbiota. World J Gastroenterol. 2015;21(29):8787–8803. doi: 10.3748/wjg.v21.i29.8787. - DOI - PMC - PubMed
Myers LL, Firehammer BD, Shoop DS, Border MM. Bacteroides fragilis: a possible cause of acute diarrheal disease in newborn lambs. Infect Immunol. 1984;44(2):241–244. doi: 10.1128/IAI.44.2.241-244.1984. - DOI - PMC - PubMed
Sack RB, Myers LL, Almeido-Hill J, Shoop DS, Bradbury WC, Reid R, et al. Enterotoxigenic Bacteroides fragilis: epidemiologic studies of its role as a human diarrhoeal pathogen. J Diarrhoeal Dis Res. 1992;10(1):4–9. - PubMed
Akpinar M, Aktaş E, Cömert F, Külah C, Sümbüloğlu V. Evaluation of the prevalence of enterotoxigenic Bacteroides fragilis and the distribution bft gene subtypes in patients with diarrhea. Anaerobe. 2010;16(5):505–509. doi: 10.1016/j.anaerobe.2010.08.002. - DOI - PubMed
Mundy LM, Sears CL. Detection of toxin production by Bacteroides fragilis: assay development and screening of extraintestinal clinical isolates. Clin Infect Dis. 1996;23(2):269–276. doi: 10.1093/clinids/23.2.269. - DOI - PubMed
Chung GT, Franco AA, Wu S, Rhie GE, Cheng R, Oh HB, et al. Identification of a third metalloprotease toxin gene in extraintestinal isolates of Bacteroides fragilis. Infect Immun. 1999;67(9):4945–9. doi: 10.1128/IAI.67.9.4945-4949.1999. - DOI - PMC - PubMed
Sears CL. The toxins of Bacteroides fragilis. Toxicon. 2001;39(11):1737–1746. doi: 10.1016/S0041-0101(01)00160-X. - DOI - PubMed
Obiso RJ, Jr, Azghani AO, Wilkins TD. The Bacteroides fragilis toxin fragilysin disrupts the paracellular barrier of epithelial cells. Infect Immun. 1997;65(4):1431–1439. doi: 10.1128/IAI.65.4.1431-1439.1997. - DOI - PMC - PubMed
Sarvari KP, Soki J, Ivan M, Miszti C, Latkoczy K, Melegh SZ, et al. Detection of enterotoxin and protease genes among Hungarian clinical Bacteroides fragilis isolates. Anaerobe. 2017;48:98–102. doi: 10.1016/j.anaerobe.2017.07.005. - DOI - PubMed
Delahooke DM, Barclay GR, Poxton IR. A re-appraisal of the biological activity of bacteroides LPS. J Med Microbiol. 1995;42(2):102–112. doi: 10.1099/00222615-42-2-102. - DOI - PubMed
Rotimi VO, Verghese TL, Al-Sweih N, Khodakhast FB, Ahmed K. Influence of five anti-anaerobic antibiotics on endotoxin liberation by gram-negative anaerobes. J Chemother. 2000;12(1):40–47. - PubMed
Prindiville TP, Sheikh RA, Cohen SH, Tang YJ, Cantrell MC, Silva J., Jr Bacteroides fragilis enterotoxin gene sequences in patients with inflammatory bowel disease. Emerg Infect Dis. 2000;6(2):171–174. doi: 10.3201/eid0602.000210. - DOI - PMC - PubMed
Toprak NU, Yagci A, Gulluoglu BM, Akin ML, Demirkalem P, Celenk T, et al. A possible role of Bacteroides fragilis enterotoxin in the aetiology of colorectal cancer. Clin Microbiol Infect. 2006;12(8):782–786. doi: 10.1111/j.1469-0691.2006.01494.x. - DOI - PubMed
Kato N, Kato H, Watanabe K, Ueno K. Association of enterotoxigenic Bacteroides fragilis with bacteremia. Clin Infect Dis. 1996;23(Suppl. 1):S83–S86. doi: 10.1093/clinids/23.Supplement_1.S83. - DOI - PubMed
Choi VM, Herrou J, Hecht AL, Teoh WP, Turner JR, Crosson S, Bubeck, et al. Activation of Bacteroides fragilis toxin by a novel bacterial protease contributes to anaerobic sepsis in mice. Nat Med. 2016;22(5):563–567. doi: 10.1038/nm.4077. - DOI - PMC - PubMed
Sóki J, Edwards R, Hedberg M, Fang H, Nagy E, Nord CE, et al. Examination of cfiA-mediated carbapenem resistance in Bacteroides fragilis strains from a European antibiotic susceptibility survey. Int J Antimicrob Agents. 2006;28(6):497–502. doi: 10.1016/j.ijantimicag.2006.07.021. - DOI - PubMed
Jamal W, Al Hashem G, Rotimi VO. Antimicrobial resistance among anaerobes isolated from clinical specimens in Kuwait hospitals: comparative analysis of 11-year data. Anaerobe. 2015;31:25–30. doi: 10.1016/j.anaerobe.2014.08.012. - DOI - PubMed
Jamal WY, Rotimi VO, Brazier JS, Johny M, Wetieh WM, Duerden BI. Molecular characterization of nitroimidazole resistance in metronidazole-resistant Bacteroides species isolated from hospital patients in Kuwait. Med Princ Pract. 2004;13(3):147–152. doi: 10.1159/000076954. - DOI - PubMed
Jamal W, Shahin M, Rotimi VO. Surveillance and trends of antimicrobial resistance among clinical isolates of anaerobes in Kuwait hospitals from 2002 to 2007. Anaerobe. 2010;16(1):1–5. doi: 10.1016/j.anaerobe.2009.04.004. - DOI - PubMed
Magiorakos AP, Srinivasan A, Carey RB, Carmeli Y, Falagas ME, Giske CG, et al. Multidrug–resistant, extensively drug-resistant and pandrug-resistant bacteria: an international expert proposal for interim standard definitions for acquired resistance. Clin Microbiol Infect. 2012;18(3):268–281. doi: 10.1111/j.1469-0691.2011.03570.x. - DOI - PubMed
Thornton RF, Kagawa TF, O’Toole PW, Cooney JC. The dissemination of C10 cysteine protease genes in Bacteroides fragilis by mobile genetic elements. BMC Microbiol. 2010;10:122. doi: 10.1186/1471-2180-10-122. - DOI - PMC - PubMed
Wexler HM. Bacteroides: the good, the bad and the nitty–gritty. Clin Microbiol Rev. 2007;20(4):593–621. doi: 10.1128/CMR.00008-07. - DOI - PMC - PubMed
Kierzkowska M, Majewska A, Szymanek-Majchrzak K, Sawicka-Grzelak A, Mlynarczyk A, Mlynarczyk G. The presence of antibiotic resistance genes and bft genes as well as antibiotic susceptibility testing of Bacteroides fragilis strains isolated from inpatients of infant Jesus teaching hospital, Warsaw during 2007-2012. Anaerobe. 2019;56:109–115. doi: 10.1016/j.anaerobe.2019.03.003. - DOI - PubMed
Szoke I, Dósa E, Nagy E. Enterotoxigenic Bacteroides fragilis in Hungary. Anaerobe. 1997;3(2–3):87–89. doi: 10.1006/anae.1997.0078. - DOI - PubMed
Scotto d’Abusco AS, Del Grosso M, Censini S, Covacci A, Pantosti A. The alleles of the bft gene are distributed differently among enterotoxigenic Bacteroides fragilis strains from human sources and can be present in double copies. J Clin Microbiol. 2000;38(2):607–612. doi: 10.1128/JCM.38.2.607-612.2000. - DOI - PMC - PubMed
Claros MC, Claros ZC, Tang YJ, Cohen SH, Silva J, Jr, Goldstein EJ, et al. Occurrence of Bacteroides fragilis enterotoxin gene-carrying strains in Germany and the United States. J Clin Microbiol. 2000;38(5):1996–1997. doi: 10.1128/JCM.38.5.1996-1997.2000. - DOI - PMC - PubMed
Nagy E, Urban E, Nord EC, on behalf of the ESCMID Study Group on Antimicrobial Resistance in Anaerobic Bacteria Antimicrobial susceptibility of Bacteroides fragilis group isolates in Europe: 20 years of experience. Clin Microbiol Infect. 2011;17(3):371–379. doi: 10.1111/j.1469-0691.2010.03256.x. - DOI - PubMed
Wang FD, Liao CH, Lin YT, Sheng WH, Hsueh PR. Trends in the susceptibility of commonly encountered clinically significant anaerobes and susceptibilities of blood isolates of anaerobes to 16 antimicrobial agents, including fidaxomicin and rifaximin, 2008-2012, northern Taiwan. Eur J Clin Microbiol Infect Dis. 2014;33(11):2041–2052. doi: 10.1007/s10096-014-2175-y. - DOI - PubMed
Snydman DR, Jacobus NV, McDermott LA, Golan Y, Goldstein EJ, Harrell L, et al. Update on resistance of Bacteroides fragilis group and related species with special attention to carbapenems 2006-2009. Anaerobe. 2011;17(4):147–151. doi: 10.1016/j.anaerobe.2011.05.014. - DOI - PubMed
Gao Q, Wu S, Xu T, Zhao X, Huang H, Hu F. Emergence of carbapenem resistance in Bacteroides fragilis in China. Int J Antimicrob Agents. 2019;53(6):859–863. doi: 10.1016/j.ijantimicag.2019.02.017. - DOI - PubMed
Hurlbut S, Cuchural GJ, Tally FP. Imipenem resistance in Bacteroides distasonis mediated by a novel β-lactamase. Antimicrob Agents Chemother. 1990;34(1):117–120. doi: 10.1128/AAC.34.1.117. - DOI - PMC - PubMed
Sóki J, Fodor E, Hecht DW, Edwards R, Rotimi VO, Kerekes I, et al. Molecular characterization of imipenem-resistant, cfiA-positive Bacteroides fragilis isolates from the USA, Hungary and Kuwait. J Med Microbiol. 2004;53(Pt 5):413–419. doi: 10.1099/jmm.0.05452-0. - DOI - PubMed
Yamazoe K, Kato N, Kato H, Tanaka K, Katagiri Y, Watanabe K. Distribution of the cfiA gene among Bacteroides fragilis strains in Japan and relatedness of cfiA to imipenem resistance. Antimicrob Agents Chemother. 1999;43(11):2808–2810. doi: 10.1128/AAC.43.11.2808. - DOI - PMC - PubMed
Edwards R, Read PN. Expression of the carbapenemase gene (cfiA) in Bacteroides fragilis. J Antimicrob Chemother. 2000;46(6):1009–1012. doi: 10.1093/jac/46.6.1009. - DOI - PubMed
Clinical and Laboratory Standards Institute: CLSI Publication Document M100-S28 . Performance standards for antimicrobial susceptibility testing: Twenty eighth Informational Supplement, Vol. 34. Wayne: Clinical and Laboratory Standards Institute; 2018. p. No.1.
Eitel Z, Sóki J, Urbán E, Nagy E, the ESCMID Study Group on Anaerobic Infection The prevalence of antibiotic resistance genes in Bacteroides fragilis group strains isolated in different European countries. Anaerobe. 2013;21:43–49. doi: 10.1016/j.anaerobe.2013.03.001. - DOI - PubMed