In recent times, some common "non-pathogenic" parasites, such as Blastocystis and Dientamoeba fragilis, have been associated to the aetiology of irritable bowel syndrome (IBS), while host pro-inflammatory cytokine gene polymorphisms might have a role in the pathophysiology of the disease. Therefore, Blastocystis subtypes (ST), D. fragilis and gene promoter single nucleotide polymorphisms of interleukin-6 (IL-6) and tumour necrosis factor-alpha (TNF-α) in IBS patients and controls were studied. After giving written consent, 45 patients with symptoms of IBS according to the Rome III criteria and 45 controls were enrolled. DNA was extracted from peripheral blood for SNP analysis at position -174 for IL-6 as well as -238 and -308 for TNF-α. Blastocystis was more common in the IBS group (p = 0.043). Interestingly, D. fragilis was found more frequently in the control group (p = 0.002); Blastocystis ST1 and 3 were most frequent in both groups. Haploview analysis revealed linkage disequilibrium in TNF-α (p < 0.0001); however, none of the SNPs for IL-6 and TNF-α were found to be significantly related with IBS. The clinical and molecular approaches undertaken for the first time in Latin American IBS patients demonstrated an association with Blastocystis that supports a pathogenic role of this parasite in IBS Furthermore, co-infections with ST1 and ST3 were frequent; thus, the genetic diversity proposed within ST polymorphisms does not rule out that particular strains might be associated with disease. In addition, our results do not support a major contribution of IL-6 and TNF-α gene polymorphisms in the susceptibility to IBS.

Irritable bowel syndrome (IBS) is one of the most common gastrointestinal diagnoses seen by primary care providers and gastroenterologists. Proinflammatory cytokines interleukin (IL)-6 and IL-8 have been found increased in IBS patients. Cytokine gene single nucleotide polymorphisms (SNPs) of IL-8 and IL-10 have not been assessed in Mexican IBS patients. DNA was extracted from peripheral blood leucocytes of 45 IBS unrelated patients and 137 controls. Allele, genotype, and haplotype frequencies were determined by analyzing SNPs of IL-8 and IL-10 genes. IL-8 + 396 G allele (P = 0.02), IL-8 + 396(G/G) and IL-8 + 781(C/T) genotypes (P < 0.001), IL-10 - 1082A allele and IL-10 - 1082(A/A) genotype (P < 0.001) were significantly increased in the IBS group. Haplotypes IL-8 ATCC (P = 0.03) and IL-10 ACC (P < 0.001) were associated with susceptibility to develop IBS. An association of certain polymorphisms of IL-8 and IL-10 in IBS patients compared to controls was demonstrated, suggesting a role of these cytokine SNPs in the pathophysiology of IBS.

Blastocystis spp is a common intestinal parasite of humans and animals that has been associated to the etiology of irritable bowel syndrome (IBS); however, some studies have not found this association. Furthermore, many biological features of Blastocystis are little known. The objective of present study was to assess the generation times of Blastocystis cultures, from IBS patients and from asymptomatic carriers. A total of 100 isolates were obtained from 50 IBS patients and from 50 asymptomatic carriers. Up to 50 mg of feces from each participant were cultured in Barret’s and in Pavlova’s media during 48 h. Initial and final parasitological load were measured by microscopy and by quantitative PCR. Amplicons were purified, sequenced and submitted to GenBank; sequences were analysed for genetic diversity and a Bayesian inference allowed identifying genetic subtypes (ST). Generation times for Blastocystis isolates in both media, based on microscopic measures and molecular assays, were calculated. The clinical symptoms of IBS patients and distribution of Blastocystis ST 1, 2 and 3 in both groups was comparable to previous reports. Interestingly, the group of cases showed scarce mean nucleotide diversity (π) as compared to the control group (0.011±0.016 and 0.118±0.177, respectively), whilst high gene flow and small genetic differentiation indexes between different ST were found. Besides, Tajima’s D test showed negative values for ST1-ST3. No statistical differences regarding parasitological load between cases and controls in both media, as searched by microscopy and by qPCR, were detected except that parasites grew faster in Barret’s than in Pavlova’s medium. Interestingly, slow growth of isolates recovered from cases in comparison to those of controls was observed (p<0.05). We propose that generation times of Blastocystis might be easily affected by intestinal environmental changes due to IBS probably because virulent strains with slow growth may be selected, reducing their genetic variability.

The intestinal protozoan parasite Blastocystis is one of the most common parasites worldwide in humans and, although its ability to cause human disease has been questioned, some reports have demonstrated that this microorganism is associated to the development of irritable bowel syndrome (IBS) and to a proinflammatory response, in which the expression of some cytokines is unregulated. Since inflammatory cytokine gene polymorphisms might have a role in the pathophysiology of IBS, we assessed the role of single nucleotide polymorphisms (SNPs) for interleukin (IL)-8 and IL-10, in previously collected DNA samples from IBS patients and controls, with or without Blastocystis infection. IL-8+396(G) and IL-10-1082 (A) alleles (p=0.0437 and p=0.0267, respectively), as well as their homozygous (p<0.0001 and p=0.0039, respectively) and IL-8+781(CT) (p=0.0248) genotypes were significantly overrepresented in patients with IBS in comparison with controls. IL-8+396(GG) genotype was relevant because it was associated to IBS (p<0.0001), to Blastocystis (p=0.0025), and to IBS–Blastocystis (p=0.0272). In the latter binomial association, this genotype presented a high contribution (etiological fraction =0.452) and a risk >fourfold to develop IBS. IL-8+781 (T) and IL-10-592 (C) alleles were also associated to Blastocystis and to IBS–Blastocystis, respectively (p=0.0448 and p=0.0166). Our results suggest that some IL-8 and IL-10 SNPs could change individual susceptibility increasing the relative risk in the development of IBS in Blastocystis carriers.

One hundred and fifteen patients with symptoms suggestive of irritable bowel syndrome (IBS) according to Rome III criteria and 209 patients with gastrointestinal symptoms different from IBS (control) were identified through medical records from the Gastroenterology Clinic of the "Dr. Manuel Gea Gonzalez General Hospital" from January 2008 to March 2010. No statistical differences in IBS data as compared with control groups were observed except in bloating, that was more frequent in the IBS group (P = 0.043). Although the pathogenicity of specific intestinal protozoa could not be demonstrated due to lack of association with the development of gastrointestinal symptoms, Blastocystis spp, in the IBS group, exhibited a trend of association to diarrhoea (odds ratio = 2.73, 95% confidence interval = 0.84-8.80, P = 0.053), while having any parasite and diarrhoea was significant (odds ratio = 3.38, 95% confidence interval = 1.33-8.57, P = 0.008). The association between Blastocystis and diarrhoea in IBS patients although not conclusive is an interesting finding; nonetheless more extensive case-controlled studies are required to clearly define the role of some "non-pathogenic" parasites in intestinal disease and IBS.