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Infect Immun. Apr 1997; 65(4): 1431–1439.
PMCID: PMC175150

The Bacteroides fragilis toxin fragilysin disrupts the paracellular barrier of epithelial cells.


Bacteroides fragilis is a member of the normal colonic microflora of most mammals and is the most commonly isolated anaerobe from human clinical specimens. Some strains produce a toxin (fragilysin, a zinc-metalloproteinase) implicated as a cause of diarrheal disease in farm animals and humans. Studies in our laboratory confirm that the proteolytic activity of this toxin is responsible for the fluid secretion and tissue damage observed in vivo. In this study, we investigated the effects of fragilysin on the paracellular barrier of epithelial cells. Researchers suggest that, since the toxin rapidly intoxicates HT-29 cells, it may be internalized. However, we could not prevent cell rounding by using inhibitors of receptor-mediated endocytosis, which indicates that the toxin may act outside the cell. Based on these observations, we studied the effects of the highly purified B. fragilis fragilysin on the barrier function of cultured epithelial cells. Fragilysin rapidly increased the permeability of the paracellular barrier of epithelial cells to ions (decrease in electrical resistance across monolayers) and to larger molecules (increase in mannitol flux across monolayers). We tested a human colon cell line and cell lines from the lung and the kidney; the human colon cell line was most sensitive, but all three were affected in the same manner. Our studies show that B. fragilis fragilysin alters the barrier function of the epithelial lining, possibly by degrading the tight junction proteins, such as ZO-1. The proteolytic activity is required to cause this effect. The toxin's action has been assumed to be limited to the intestine; however, our studies show that fragilysin could also contribute to the pathogenesis of B. fragilis in extraintestinal infections.

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Selected References

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  • Azghani AO. Pseudomonas aeruginosa and epithelial permeability: role of virulence factors elastase and exotoxin A. Am J Respir Cell Mol Biol. 1996 Jul;15(1):132–140. [PubMed]
  • Azghani AO, Gray LD, Johnson AR. A bacterial protease perturbs the paracellular barrier function of transporting epithelial monolayers in culture. Infect Immun. 1993 Jun;61(6):2681–2686. [PMC free article] [PubMed]
  • Bejarano PA, Langeveld JP, Hudson BG, Noelken ME. Degradation of basement membranes by Pseudomonas aeruginosa elastase. Infect Immun. 1989 Dec;57(12):3783–3787. [PMC free article] [PubMed]
  • Bode W, Gomis-Rüth FX, Stöckler W. Astacins, serralysins, snake venom and matrix metalloproteinases exhibit identical zinc-binding environments (HEXXHXXGXXH and Met-turn) and topologies and should be grouped into a common family, the 'metzincins'. FEBS Lett. 1993 Sep 27;331(1-2):134–140. [PubMed]
  • Border M, Firehammer BD, Shoop DS, Myers LL. Isolation of Bacteroides fragilis from the feces of diarrheic calves and lambs. J Clin Microbiol. 1985 Mar;21(3):472–473. [PMC free article] [PubMed]
  • Collins JE, Bergeland ME, Myers LL, Shoop DS. Exfoliating colitis associated with enterotoxigenic Bacteroides fragilis in a piglet. J Vet Diagn Invest. 1989 Oct;1(4):349–351. [PubMed]
  • Donelli G, Fabbri A, Fiorentini C. Bacteroides fragilis enterotoxin induces cytoskeletal changes and surface blebbing in HT-29 cells. Infect Immun. 1996 Jan;64(1):113–119. [PMC free article] [PubMed]
  • Duimstra JR, Myers LL, Collins JE, Benfield DA, Shoop DS, Bradbury WC. Enterovirulence of enterotoxigenic Bacteroides fragilis in gnotobiotic pigs. Vet Pathol. 1991 Nov;28(6):514–518. [PubMed]
  • Gomis-Rüth FX, Kress LF, Kellermann J, Mayr I, Lee X, Huber R, Bode W. Refined 2.0 A X-ray crystal structure of the snake venom zinc-endopeptidase adamalysin II. Primary and tertiary structure determination, refinement, molecular structure and comparison with astacin, collagenase and thermolysin. J Mol Biol. 1994 Jun 17;239(4):513–544. [PubMed]
  • Gumbiner B, Simons K. A functional assay for proteins involved in establishing an epithelial occluding barrier: identification of a uvomorulin-like polypeptide. J Cell Biol. 1986 Feb;102(2):457–468. [PMC free article] [PubMed]
  • Harrington DJ. Bacterial collagenases and collagen-degrading enzymes and their potential role in human disease. Infect Immun. 1996 Jun;64(6):1885–1891. [PMC free article] [PubMed]
  • Häse CC, Finkelstein RA. Bacterial extracellular zinc-containing metalloproteases. Microbiol Rev. 1993 Dec;57(4):823–837. [PMC free article] [PubMed]
  • Hecht G, Koutsouris A, Pothoulakis C, LaMont JT, Madara JL. Clostridium difficile toxin B disrupts the barrier function of T84 monolayers. Gastroenterology. 1992 Feb;102(2):416–423. [PubMed]
  • Hecht G, Pothoulakis C, LaMont JT, Madara JL. Clostridium difficile toxin A perturbs cytoskeletal structure and tight junction permeability of cultured human intestinal epithelial monolayers. J Clin Invest. 1988 Nov;82(5):1516–1524. [PMC free article] [PubMed]
  • Howarth AG, Stevenson BR. Molecular environment of ZO-1 in epithelial and non-epithelial cells. Cell Motil Cytoskeleton. 1995;31(4):323–332. [PubMed]
  • Just I, Selzer J, Wilm M, von Eichel-Streiber C, Mann M, Aktories K. Glucosylation of Rho proteins by Clostridium difficile toxin B. Nature. 1995 Jun 8;375(6531):500–503. [PubMed]
  • Kamata R, Matsumoto K, Okamura R, Yamamoto T, Maeda H. The serratial 56K protease as a major pathogenic factor in serratial keratitis. Clinical and experimental study. Ophthalmology. 1985 Oct;92(10):1452–1459. [PubMed]
  • Kress LF, Kurecki T, Chan SK, Laskowski M., Sr Characterization of the inactive fragment resulting from limited proteolysis of human alpha1-proteinase inhibitor by Crotalus adamanteus proteinase II. J Biol Chem. 1979 Jun 25;254(12):5317–5320. [PubMed]
  • Kress LF, Catanese JJ. Identification of the cleavage sites resulting from enzymatic inactivation of human antithrombin III by Crotalus adamanteus proteinase II in the the presence and absence of heparin. Biochemistry. 1981 Dec 22;20(26):7432–7438. [PubMed]
  • Nath SK, Desjeux JF. Human intestinal cell lines as in vitro tools for electrolyte transport studies with relevance to secretory diarrhoea. J Diarrhoeal Dis Res. 1990 Dec;8(4):133–142. [PubMed]
  • Madara JL. Intestinal absorptive cell tight junctions are linked to cytoskeleton. Am J Physiol. 1987 Jul;253(1 Pt 1):C171–C175. [PubMed]
  • Madara JL. Loosening tight junctions. Lessons from the intestine. J Clin Invest. 1989 Apr;83(4):1089–1094. [PMC free article] [PubMed]
  • Madara JL, Moore R, Carlson S. Alteration of intestinal tight junction structure and permeability by cytoskeletal contraction. Am J Physiol. 1987 Dec;253(6 Pt 1):C854–C861. [PubMed]
  • Maruyama M, Sugiki M, Yoshida E, Shimaya K, Mihara H. Broad substrate specificity of snake venom fibrinolytic enzymes: possible role in haemorrhage. Toxicon. 1992 Nov;30(11):1387–1397. [PubMed]
  • Meisel-Mikołajczyk F, Sebald M, Torbicka E, Rafałowska K, Zielińska U. Isolation of enterotoxigenic Bacteroides fragilis strains in Poland. Acta Microbiol Pol. 1994;43(3-4):389–392. [PubMed]
  • Moncrief JS, Obiso R, Jr, Barroso LA, Kling JJ, Wright RL, Van Tassell RL, Lyerly DM, Wilkins TD. The enterotoxin of Bacteroides fragilis is a metalloprotease. Infect Immun. 1995 Jan;63(1):175–181. [PMC free article] [PubMed]
  • Moore WE, Cato EP, Holdeman LV. Some current concepts in intestinal bacteriology. Am J Clin Nutr. 1978 Oct;31(10 Suppl):S33–S42. [PubMed]
  • Myers LL, Collins JE, Shoop DS. Ultrastructural lesions of enterotoxigenic Bacteroides fragilis in rabbits. Vet Pathol. 1991 Jul;28(4):336–338. [PubMed]
  • Myers LL, Shoop DS, Collins JE. Rabbit model to evaluate enterovirulence of Bacteroides fragilis. J Clin Microbiol. 1990 Jul;28(7):1658–1660. [PMC free article] [PubMed]
  • Myers LL, Shoop DS, Collins JE, Bradbury WC. Diarrheal disease caused by enterotoxigenic Bacteroides fragilis in infant rabbits. J Clin Microbiol. 1989 Sep;27(9):2025–2030. [PMC free article] [PubMed]
  • Myers LL, Shoop DS. Association of enterotoxigenic Bacteroides fragilis with diarrheal disease in young pigs. Am J Vet Res. 1987 May;48(5):774–775. [PubMed]
  • Myers LL, Shoop DS, Byars TD. Diarrhea associated with enterotoxigenic Bacteroides fragilis in foals. Am J Vet Res. 1987 Nov;48(11):1565–1567. [PubMed]
  • Myers LL, Shoop DS, Stackhouse LL, Newman FS, Flaherty RJ, Letson GW, Sack RB. Isolation of enterotoxigenic Bacteroides fragilis from humans with diarrhea. J Clin Microbiol. 1987 Dec;25(12):2330–2333. [PMC free article] [PubMed]
  • Myers LL, Shoop DS, Firehammer BD, Border MM. Association of enterotoxigenic Bacteroides fragilis with diarrheal disease in calves. J Infect Dis. 1985 Dec;152(6):1344–1347. [PubMed]
  • Myers LL, Firehammer BD, Shoop DS, Border MM. Bacteroides fragilis: a possible cause of acute diarrheal disease in newborn lambs. Infect Immun. 1984 May;44(2):241–244. [PMC free article] [PubMed]
  • Obiso RJ, Jr, Lyerly DM, Van Tassell RL, Wilkins TD. Proteolytic activity of the Bacteroides fragilis enterotoxin causes fluid secretion and intestinal damage in vivo. Infect Immun. 1995 Oct;63(10):3820–3826. [PMC free article] [PubMed]
  • Pantosti A, Piersimoni C, Perissi G. Detection of Bacteroides fragilis enterotoxin in the feces of a child with diarrhea. Clin Infect Dis. 1994 Oct;19(4):809–810. [PubMed]
  • Pantosti A, Cerquetti M, Colangeli R, D'Ambrosio F. Detection of intestinal and extra-intestinal strains of enterotoxigenic Bacteroides fragilis by the HT-29 cytotoxicity assay. J Med Microbiol. 1994 Sep;41(3):191–196. [PubMed]
  • Peterson BT, Collins ML, Gray LD, Azghani AO. Aerosolized Pseudomonas elastase and lung fluid balance in anesthetized sheep. J Appl Physiol (1985) 1992 May;72(5):1927–1933. [PubMed]
  • Sack RB, Albert MJ, Alam K, Neogi PK, Akbar MS. Isolation of enterotoxigenic Bacteroides fragilis from Bangladeshi children with diarrhea: a controlled study. J Clin Microbiol. 1994 Apr;32(4):960–963. [PMC free article] [PubMed]
  • Sack RB, Myers LL, Almeido-Hill J, Shoop DS, Bradbury WC, Reid R, Santosham M. Enterotoxigenic Bacteroides fragilis: epidemiologic studies of its role as a human diarrhoeal pathogen. J Diarrhoeal Dis Res. 1992 Mar;10(1):4–9. [PubMed]
  • Saidi RF, Sears CL. Bacteroides fragilis toxin rapidly intoxicates human intestinal epithelial cells (HT29/C1) in vitro. Infect Immun. 1996 Dec;64(12):5029–5034. [PMC free article] [PubMed]
  • San Joaquin VH, Griffis JC, Lee C, Sears CL. Association of Bacteroides fragilis with childhood diarrhea. Scand J Infect Dis. 1995;27(3):211–215. [PubMed]
  • Simon GL, Gorbach SL. Intestinal flora in health and disease. Gastroenterology. 1984 Jan;86(1):174–193. [PubMed]
  • Stöcker W, Grams F, Baumann U, Reinemer P, Gomis-Rüth FX, McKay DB, Bode W. The metzincins--topological and sequential relations between the astacins, adamalysins, serralysins, and matrixins (collagenases) define a superfamily of zinc-peptidases. Protein Sci. 1995 May;4(5):823–840. [PMC free article] [PubMed]
  • Van Tassell RL, Lyerly DM, Wilkins TD. Purification and characterization of an enterotoxin from Bacteroides fragilis. Infect Immun. 1992 Apr;60(4):1343–1350. [PMC free article] [PubMed]
  • Weikel CS, Grieco FD, Reuben J, Myers LL, Sack RB. Human colonic epithelial cells, HT29/C1, treated with crude Bacteroides fragilis enterotoxin dramatically alter their morphology. Infect Immun. 1992 Feb;60(2):321–327. [PMC free article] [PubMed]
  • Wells CL, van de Westerlo EM, Jechorek RP, Feltis BA, Wilkins TD, Erlandsen SL. Bacteroides fragilis enterotoxin modulates epithelial permeability and bacterial internalization by HT-29 enterocytes. Gastroenterology. 1996 May;110(5):1429–1437. [PubMed]

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