Digestive Diseases and Sciences

, Volume 60, Issue 10, pp 2953–2962 | Cite as

Fecal Microbiota in Patients with Irritable Bowel Syndrome Compared with Healthy Controls Using Real-Time Polymerase Chain Reaction: An Evidence of Dysbiosis

  • Ratnakar Shukla
  • Ujjala Ghoshal
  • Tapan N. Dhole
  • Uday C. Ghoshal
Original Article



Dysbiosis may play a role in irritable bowel syndrome (IBS), hitherto an enigmatic disorder. We evaluated selected fecal microbes in IBS patients and healthy controls (HC).


Fecal 16S rRNA copy number of selected bacteria was studied using qPCR in 47 patients with IBS (Rome III) and 30 HC.


Of 47 patients, 20 had constipation (IBS-C), 20 diarrhea (IBS-D), and seven unclassified IBS (IBS-U). Relative difference in 16S rRNA copy number of Bifidobacterium (P = 0.042) was lower, while those of Ruminococcus productus-Clostridium coccoides (P = 0.016), Veillonella (P = 0.008), Bacteroides thetaiotamicron (P < 0.001), Pseudomonas aeruginosa (P < 0.001), and Gram-negative bacteria (GNB, P = 0.001) were higher among IBS patients than HC. Number of Lactobacillus (P = 0.002) was lower, while that of Bacteroides thetaiotamicron (P < 0.001) and segmented filamentous bacteria (SFB, P < 0.001) was higher among IBS-D than IBS-C. Numbers of Bacteroides thetaiotamicron (P < 0.001), P. aeruginosa (P < 0.001), and GNB (P < 0.01) were higher among IBS-C and IBS-D than HC. Quantity of SFB was higher among IBS-D (P = 0.011) and lower among IBS-C (P = 0.002) than HC. Number of Veillonella species was higher among IBS-C than HC (P = 0.002). P. aeruginosa was frequently detected among IBS than HC (46/47 [97.9 %] vs. 10/30 [33.3 %], P < 0.001). Abdominal distension (n = 34/47) was associated with higher number of Bacteroides thetaiotamicron, Clostridium coccoides, P. aeruginosa, SFB, and GNB; bloating (n = 22/47) was associated with Clostridium coccoides and GNB. Microbial flora was different among IBS than HC on principal component analysis.


Fecal microbiota was different among IBS than HC, and different sub-types were associated with different microbiota. P. aeruginosa was more frequent and higher in number among IBS patients.


Functional gastrointestinal disorder Quantitative real-time polymerase chain reaction Dysbiosis Gastrointestinal microbiota 



The authors wish to thank all the patients and volunteers for taking part in this study. Ratnakar Shukla thanks the Department of Science and Technology for providing his fellowship.

Conflict of interest

There is no financial conflict of interest to declare.


  1. 1.
    Longstreth GF, Thompson WG, Chey WD, Houghton LA, Mearin F, Spiller RC. Functional bowel disorders. Gastroenterology. 2006;130:1480–1491.CrossRefPubMedGoogle Scholar
  2. 2.
    Malinen E, Krogius-Kurikka L, Lyra A, et al. Association of symptoms with gastrointestinal microbiota in irritable bowel syndrome. World J Gastroenterol. 2010;16:4532–4540.PubMedCentralCrossRefPubMedGoogle Scholar
  3. 3.
    Parkes GC, Brostoff J, Whelan K, Sanderson JD. Gastrointestinal microbiota in irritable bowel syndrome: their role in its pathogenesis and treatment. Am J Gastroenterol. 2008;103:1557–1567.CrossRefPubMedGoogle Scholar
  4. 4.
    Ohman L, Simren M. New insights into the pathogenesis and pathophysiology of irritable bowel syndrome. Dig Liver Dis. 2007;39:201–215.CrossRefPubMedGoogle Scholar
  5. 5.
    Pimentel M, Lembo A, Chey WD, et al. Rifaximin therapy for patients with irritable bowel syndrome without constipation. N Engl J Med. 2011;364:22–32.CrossRefPubMedGoogle Scholar
  6. 6.
    Guandalini S, Magazzu G, Chiaro A, et al. Vsl#3 improves symptoms in children with irritable bowel syndrome: a multicenter, randomized, placebo-controlled, double-blind, crossover study. J Pediatr Gastroenterol Nutr. 2010;51:24–30.CrossRefPubMedGoogle Scholar
  7. 7.
    Silk DB, Davis A, Vulevic J, Tzortzis G, Gibson GR. Clinical trial: the effects of a trans-galactooligosaccharide prebiotic on faecal microbiota and symptoms in irritable bowel syndrome. Aliment Pharmacol Ther. 2009;29:508–518.CrossRefPubMedGoogle Scholar
  8. 8.
    Malinen E, Rinttila T, Kajander K, et al. Analysis of the fecal microbiota of irritable bowel syndrome patients and healthy controls with real-time pcr. Am J Gastroenterol. 2005;100:373–382.CrossRefPubMedGoogle Scholar
  9. 9.
    Lee BJ, Bak YT. Irritable bowel syndrome, gut microbiota and probiotics. J Neurogastroenterol Motil. 2011;17:252–266.PubMedCentralCrossRefPubMedGoogle Scholar
  10. 10.
    Kerckhoffs AP, Ben-Amor K, Samsom M, et al. Molecular analysis of faecal and duodenal samples reveals significantly higher prevalence and numbers of pseudomonas aeruginosa in irritable bowel syndrome. J Med Microbiol. 2011;60:236–245.CrossRefPubMedGoogle Scholar
  11. 11.
    Sung J, Morales W, Kim G, et al. Effect of repeated campylobacter jejuni infection on gut flora and mucosal defense in a rat model of post infectious functional and microbial bowel changes. Neurogastroenterol Motil. 2013;25:529–537.CrossRefPubMedGoogle Scholar
  12. 12.
    Balsari A, Ceccarelli A, Dubini F, Fesce E, Poli G. The fecal microbial population in the irritable bowel syndrome. Microbiologica. 1982;5:185–194.PubMedGoogle Scholar
  13. 13.
    Liebregts T, Adam B, Bredack C, et al. Immune activation in patients with irritable bowel syndrome. Gastroenterology. 2007;132:913–920.CrossRefPubMedGoogle Scholar
  14. 14.
    Barbara G, Cremon C, Carini G, et al. The immune system in irritable bowel syndrome. J Neurogastroenterol Motil. 2011;17:349–359.PubMedCentralCrossRefPubMedGoogle Scholar
  15. 15.
    Loh G, Blaut M. Role of commensal gut bacteria in inflammatory bowel diseases. Gut Microbes. 2012;3:544–555.PubMedCentralCrossRefPubMedGoogle Scholar
  16. 16.
    Bellavia M, Damiano G, Gioviale MC, et al. Abnormal expansion of segmented filamentous bacteria in the gut: a role in pathogenesis of chronic inflammatory intestinal diseases? Rev Med Microbiol. 2011;22:45–47.CrossRefGoogle Scholar
  17. 17.
    Hattori T, Fukudo S. Use of Rome III criteria for diagnosing irritable bowel syndrome. Nihon Rinsho. 2006;64:1425–1428.PubMedGoogle Scholar
  18. 18.
    Ghoshal UC, Gwee KA, Chen M, et al. Development, translation and validation of enhanced asian rome iii questionnaires for diagnosis of functional bowel diseases in major asian languages: a Rome foundation-asian neurogastroenterology and motility association working team report. J Neurogastroenterol Motil. 2015;21:83–92.PubMedCentralCrossRefPubMedGoogle Scholar
  19. 19.
    Bartosch S, Fite A, Macfarlane GT, McMurdo ME. Characterization of bacterial communities in feces from healthy elderly volunteers and hospitalized elderly patients by using real-time pcr and effects of antibiotic treatment on the fecal microbiota. Appl Environ Microbiol. 2004;70:3575–3581.PubMedCentralCrossRefPubMedGoogle Scholar
  20. 20.
    Carroll IM, Chang YH, Park J, Sartor RB, Ringel Y. Luminal and mucosal-associated intestinal microbiota in patients with diarrhea-predominant irritable bowel syndrome. Gut Pathog. 2010;2:19.PubMedCentralCrossRefPubMedGoogle Scholar
  21. 21.
    Matsuki T, Watanabe K, Tanaka R, Fukuda M, Oyaizu H. Distribution of bifidobacterial species in human intestinal microflora examined with 16s rRNA-gene-targeted species-specific primers. Appl Environ Microbiol. 1999;65:4506–4512.PubMedCentralPubMedGoogle Scholar
  22. 22.
    Ponnusamy K, Choi JN, Kim J, Lee SY, Lee CH. Microbial community and metabolomic comparison of irritable bowel syndrome faeces. J Med Microbiol. 2011;60:817–827.PubMedCentralCrossRefPubMedGoogle Scholar
  23. 23.
    Qin X, Emerson J, Stapp J, Stapp L, Abe P, Burns JL. Use of real-time PCR with multiple targets to identify pseudomonas aeruginosa and other nonfermenting gram-negative bacilli from patients with cystic fibrosis. J Clin Microbiol. 2003;41:4312–4317.PubMedCentralCrossRefPubMedGoogle Scholar
  24. 24.
    Kubota H, Tsuji H, Matsuda K, Kurakawa T, Asahara T, Nomoto K. Detection of human intestinal catalase-negative, gram-positive cocci by rrna-targeted reverse transcription-PCR. Appl Environ Microbiol. 2010;76:5440–5451.PubMedCentralCrossRefPubMedGoogle Scholar
  25. 25.
    Conte MP, Schippa S, Zamboni I, et al. Gut-associated bacterial microbiota in paediatric patients with inflammatory bowel disease. Gut. 2006;55:1760–1767.PubMedCentralCrossRefPubMedGoogle Scholar
  26. 26.
    Klausegger A, Hell M, Berger A, et al. Gram type-specific broad-range pcr amplification for rapid detection of 62 pathogenic bacteria. J Clin Microbiol. 1999;37:464–466.PubMedCentralPubMedGoogle Scholar
  27. 27.
    Suzuki K, Meek B, Doi Y. Aberrant expansion of segmented filamentous bacteria in iga-deficient gut. Proc Natl Acad Sci USA. 2004;101:1981–1986.PubMedCentralCrossRefPubMedGoogle Scholar
  28. 28.
    Lyra A, Rinttila T, Nikkila J, et al. Diarrhoea-predominant irritable bowel syndrome distinguishable by 16s rRNA gene phylotype quantification. World J Gastroenterol. 2009;15:5936–5945.PubMedCentralCrossRefPubMedGoogle Scholar
  29. 29.
    Kassinen A, Krogius-Kurikka L, Makivuokko H, et al. The fecal microbiota of irritable bowel syndrome patients differs significantly from that of healthy subjects. Gastroenterology. 2007;133:24–33.CrossRefPubMedGoogle Scholar
  30. 30.
    Parkes GC, Rayment NB, Hudspith BN, et al. Distinct microbial populations exist in the mucosa-associated microbiota of sub-groups of irritable bowel syndrome. Neurogastroenterol Motil. 2011;24:31–39.CrossRefPubMedGoogle Scholar
  31. 31.
    Yoon JS, Sohn W, Lee OY, et al. Effect of multispecies probiotics on irritable bowel syndrome: a randomized, double-blind, placebo-controlled trial. J Gastroenterol Hepatol. 2014;29:52–59.CrossRefPubMedGoogle Scholar
  32. 32.
    Kerckhoffs AP, Samsom M, van der Rest ME, et al. Lower bifidobacteria counts in both duodenal mucosa-associated and fecal microbiota in irritable bowel syndrome patients. World J Gastroenterol. 2009;15:2887–2892.PubMedCentralCrossRefPubMedGoogle Scholar
  33. 33.
    Spiller R. Review article: probiotics and prebiotics in irritable bowel syndrome. Aliment Pharmacol Ther. 2008;28:385–396.CrossRefPubMedGoogle Scholar
  34. 34.
    Jeffery IB, Quigley EM, Ohman L, Simren M, O’Toole PW. The microbiota link to irritable bowel syndrome: an emerging story. Gut Microbes. 2012;3:572–576.PubMedCentralCrossRefPubMedGoogle Scholar
  35. 35.
    Ghoshal UC, Shukla R, Ghoshal U, Gwee KA, Ng SC, Quigley EMM. The gut microbiota and irritable bowel syndrome: friend or foe? Int J Inflamm. 2012.Google Scholar
  36. 36.
    Salonen A, de Vos WM, Palva A. Gastrointestinal microbiota in irritable bowel syndrome: present state and perspectives. Microbiology. 2010;156:3205–3215.CrossRefPubMedGoogle Scholar
  37. 37.
    King TS, Elia M, Hunter JO. Abnormal colonic fermentation in irritable bowel syndrome. Lancet. 1998;352:1187–1189.CrossRefPubMedGoogle Scholar
  38. 38.
    Arumugam M, Raes J, Pelletier E, et al. Enterotypes of the human gut microbiome. Nature. 2011;473:174–180.PubMedCentralCrossRefPubMedGoogle Scholar
  39. 39.
    Martens EC, Koropatkin NM, Smith TJ, Gordon JI. Complex glycan catabolism by the human gut microbiota: the bacteroidetes sus-like paradigm. J Biol Chem. 2009;284:24673–24677.PubMedCentralCrossRefPubMedGoogle Scholar
  40. 40.
    Rios-Covian D, Arboleya S, Hernandez-Barranco AM, et al. Interactions between bifidobacterium and bacteroides species in cofermentations are affected by carbon sources, including exopolysaccharides produced by bifidobacteria. Appl Environ Microbiol. 2013;79:7518–7524.PubMedCentralCrossRefPubMedGoogle Scholar
  41. 41.
    Ng SC, Lam EF, Lam TT, et al. Effect of probiotic bacteria on the intestinal microbiota in irritable bowel syndrome. J Gastroenterol Hepatol. 2013;28:1624–1631.PubMedGoogle Scholar
  42. 42.
    Tana C, Umesaki Y, Imaoka A, Handa T, Kanazawa M, Fukudo S. Altered profiles of intestinal microbiota and organic acids may be the origin of symptoms in irritable bowel syndrome. Neurogastroenterol Motil. 2010;22:512–519, e114–515.Google Scholar
  43. 43.
    Liu C, Finegold SM, Song Y, Lawson PA. Reclassification of clostridium coccoides, ruminococcus hansenii, ruminococcus hydrogenotrophicus, ruminococcus luti, ruminococcus productus and ruminococcus schinkii as blautia coccoides gen. Nov., comb. Nov., blautia hansenii comb. Nov., blautia hydrogenotrophica comb. Nov., blautia luti comb. Nov., blautia producta comb. Nov., blautia schinkii comb. Nov. And description of blautia wexlerae sp. Nov., isolated from human faeces. Int J Syst Evol Microbiol. 2008;58:1896–1902.CrossRefPubMedGoogle Scholar
  44. 44.
    Treem WR, Ahsan N, Kastoff G, Hyams JS. Fecal short-chain fatty acids in patients with diarrhea-predominant irritable bowel syndrome: In vitro studies of carbohydrate fermentation. J Pediatr Gastroenterol Nutr. 1996;23:280–286.CrossRefPubMedGoogle Scholar
  45. 45.
    Bourdu S, Dapoigny M, Chapuy E, et al. Rectal instillation of butyrate provides a novel clinically relevant model of noninflammatory colonic hypersensitivity in rats. Gastroenterology. 2005;128:1996–2008.CrossRefPubMedGoogle Scholar
  46. 46.
    Chichlowski M, Hale LP. Bacterial-mucosal interactions in inflammatory bowel disease: an alliance gone bad. Am J Physiol Gastrointest Liver Physiol. 2008;295:G1139–G1149.PubMedCentralCrossRefPubMedGoogle Scholar
  47. 47.
    Dulon S, Leduc D, Cottrell GS, et al. Pseudomonas aeruginosa elastase disables proteinase-activated receptor 2 in respiratory epithelial cells. Am J Respir Cell Mol Biol. 2005;32:411–419.CrossRefPubMedGoogle Scholar
  48. 48.
    Vergnolle N. Clinical relevance of proteinase activated receptors (pars) in the gut. Gut. 2005;54:867–874.PubMedCentralCrossRefPubMedGoogle Scholar
  49. 49.
    Srivastava D, Ghoshal U, Mittal RD, Ghoshal UC. Associations between IL-1RA polymorphisms and small intestinal bacterial overgrowth among patients with irritable bowel syndrome from India. Neurogastroenterol Motil. 2014;26:1408–1416.CrossRefPubMedGoogle Scholar
  50. 50.
    Rodriguez-Fandino O, Hernandez-Ruiz J, Schmulson M. From cytokines to toll-like receptors and beyond—current knowledge and future research needs in irritable bowel syndrome. J Neurogastroenterol Motil. 2010;16:363–373.PubMedCentralCrossRefPubMedGoogle Scholar
  51. 51.
    Zhong W, Zhou Z. Alterations of the gut microbiome and metabolome in alcoholic liver disease. World J Gastrointest Pathophysiol. 2014;5:514–522.PubMedCentralCrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2015

Authors and Affiliations

  • Ratnakar Shukla
    • 1
  • Ujjala Ghoshal
    • 1
  • Tapan N. Dhole
    • 1
  • Uday C. Ghoshal
    • 2
  1. 1.Department of MicrobiologySanjay Gandhi Postgraduate Institute of Medical SciencesLucknowIndia
  2. 2.Departments of GastroenterologySanjay Gandhi Postgraduate Institute of Medical SciencesLucknowIndia

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