Fernando Martínez-Hernández

Blastocystis infection is associated with irritable bowel syndrome in a Mexican patient population

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.

Natural Crossbreeding between Sympatric Species of the Phyllosoma Complex (Insecta: Hemiptera: Reduviidae) Indicate the Existence of Only One Species with Morphologic and Genetic Variations

The nucleotide sequences of the cytochrome B gene and the antennal phenotypes were analyzed for the following triatomine species: Triatoma longipennis, Triatoma pallidipennis, and Triatoma picturata, which belong to the Phyllosoma complex. These species inhabit sympatric areas from Talpa de Allende, Autlan de Navarro, and Teocuitatlan de Corona in Jalisco, Mexico. Molecular marker analysis showed that the sympatric individuals are the natural crossbred descendents of different individuals living in close proximity in these natural areas that resulted in mixed populations. The antennal phenotype results are coincident with these genetic findings, which point to the high similitude between all Phyllosoma complex populations analyzed. These data support the hypothesis that these species are morphotypes with chromatic and genetic varieties, which preserves the possibility of natural breeding with fertile descent. In conclusion, our results strongly support the hypothesis that T. pallidipennis, T. longipennis, and T. picturata are subspecies of the Phyllosoma complex.

Findings related to IL-8 and IL-10 gene polymorphisms in a Mexican patient population with irritable bowel syndrome infected with Blastocystis

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.

Geographical widespread of two lineages of Taenia solium due to human migrations: can population genetic analysis strengthen this hypothesis?

In this paper we discuss, with a new analysis of published data, the phylogenetic hypothesis of two genotypes of Taenia solium previously suggested. Sequences of mitochondrial (co1, cob, nad) and nuclear (18S+ITS1+5.8S, LMWA1 and LMWA2) T. solium DNA from Africa, Asia, Latin America, and Oceania deposited in GenBank international databases were analyzed for diversity and genetic structure. Overall, we found that percentages of polymorphic and informative sites were comparatively less in mitochondrial genes, and minimum or null values of nucleotide diversity and nucleotide polymorphism were also observed. Analysis of co1 populations showed two associations of particular interest: Asia/Latin America and Africa/Latin America; with minimal differentiation between them and a constant genetic flow. Bayesian phylogenetic trees built with the available sequences for co1+cob showed two clusters, one for Asia and another one for Africa/Latin America while with ribosomal sequences only one cluster was obtained that grouped Asian and Latin American populations. The haplotype network tree built using co1+cob showed two major clades, one clustering African and Latin American parasite populations and the other grouping Asian populations, hallmarking Mexican/Peruvian and the Indian populations as dispersion centers, respectively. The haplotype network tree built with ribosomal sequences exhibited Philippines and Peru/Mexico/Colombia as the two major dispersion centers, with several Latin American haplotypes diverging from the latter. Our results suggest that the gene flow within the different T. solium populations has the same dispersion pattern than the main maritime trade routes used between the XV and XIX centuries.

Genetic Variation of North American Triatomines (Insecta: Hemiptera: Reduviidae): Initial Divergence between Species and Populations of Chagas Disease Vector

The triatomines vectors of Trypanosoma cruzi are principal factors in acquiring Chagas disease. For this reason, increased knowledge of domestic transmission of T. cruzi and control of its insect vectors is necessary. To contribute to genetic knowledge of North America Triatominae species, we studied genetic variations and conducted phylogenetic analysis of different triatomines species of epidemiologic importance. Our analysis showed high genetic variations between different geographic populations of Triatoma mexicana, Meccus longipennis, M. mazzottii, M. picturatus, and T. dimidiata species, suggested initial divergence, hybridation, or classifications problems. In contrast, T. gerstaeckeri, T. bolivari, and M. pallidipennis populations showed few genetics variations. Analysis using cytochrome B and internal transcribed spacer 2 gene sequences indicated that T. bolivari is closely related to the Rubrofasciata complex and not to T. dimidiata. Triatoma brailovskyi and T. gerstaeckeri showed a close relationship with Dimidiata and Phyllosoma complexes.

Suitability of internal transcribed spacers (ITS) as markers for the population genetic structure of Blastocystis spp

BackgroundThe purpose of this study was to assess the genetic variation and differentiation of Blastocystis subtypes (STs) recovered from symptomatic children by analysing partial sequences of the small subunit rDNA gene region (SSUrDNA) and internal transcribed spacers (1 and 2) plus the 5.8S region (ITS, ITS1 + 5.8S + ITS2) and comparing with isolates from other countries.FindingsFaecal samples from 47 Blastocystis-infected children with gastrointestinal symptoms and negative for pathogenic enterobacteria were analysed. PCR was performed on DNA from all the samples to identify Blastocystis STs, amplifying a fragment of SSUrDNA and the ITS region. The amplicons were purified and sequenced, and consensus sequences were submitted to GenBank; afterwards, SSUrDNA sequences were analysed for genetic diversity according to geographic area. Regarding the Blastocystis STs found, 51% were ST1, 23% ST2, 19% ST3 and 2% ST7. For ITS, a haplotype network tree and Bayesian inference revealed the presence of two novel variants of ST1, clustering some sequences into ST1A and ST1B. The values of nucleotide diversity (π) and haplotype polymorphism (θ) for ST1, ST2 and ST3 ranged from 0 to 1, whereas the ratio of genetic differentiation (FST)/migration index (Nm) showed the highest differentiation between Libya and Thailand-Philippines for ST2 (0.282/0.63). In contrast, a high flow gene was observed between Czech Republic-Denmark-Holland-Spain and USA-Mexico-Colombia for ST1 (0.003/84).ConclusionOur data on genetic differentiation and gene flow might explain the differences for the prevalence of Blastocystis STs. Moreover, the ITS region could be used as a genetic marker to assess genetic variation in this parasite.

Microsatellite and mini-exon analysis of Mexican human DTU I Trypanosoma cruzi strains and their susceptibility to nifurtimox and benznidazole.

Chagas disease is caused by the protozoan parasite Trypanosoma cruzi, and it affects as many as 10 million people in North and South America, where it represents a major public health problem. T. cruzi is a parasite with high genetic diversity, and it has been grouped into 6 discrete typing units (DTUs), designated as T. cruzi I (TcI) to T. cruzi VI (TcVI). Mexican isolates from humans and from vector insects have been primarily found to be TcI, and these isolates are likely to be the strains that cause the clinical manifestations observed in Mexico. However, genetic characterization and drug susceptibility assays are limited in Mexican TcI strains. In this work, 24 Mexican T. cruzi strains, obtained primarily from humans, were studied with 7 locus microsatellites and mini-exon gene by PCR. Also, drug susceptibility was evaluated by growth and mobility assays. All of the human strains belonged to TcI, and they could be further grouped through microsatellite analysis into 2 subgroups (microsatellite genotypes 1 and 2), which were not related to the host clinical status or biological origin of the strain. Two strains, both from wild mammals, belonged to the TcII-TcVI groups; these strains and the CL Brener strain constituted microsatellite genotype 3. The number of alleles in each locus was lower than reported for South American strains, and a departure from the Hardy-Weinberg equilibrium was observed. The susceptibility of these strains to nifurtimox and benznidazole was heterogeneous. T. cruzi strains characterized as microsatellite genotypes 2 and 3 were significantly more susceptible to benznidazole than strains of microsatellite genotype 1. Only 1 Mexican strain resistant to both drugs was found in this study.

Plasma cytokine levels and cytokine gene polymorphisms in Mexican patients during the influenza pandemic A(H1N1)pdm09

BACKGROUND In Mexico, the initial severe cases of the 2009 influenza pandemic virus A (H1N1) [A(H1N1)pdm09] were detected in early March. The immune mechanisms associated with the severe pneumonia caused by infection with this new virus have not been completely elucidated. Polymorphisms in interleukin genes have previously been associated with susceptibility to infectious diseases due to their influence on cytokine production. OBJECTIVES The present case-control study was performed to compare several immunologic and genetic parameters of patients and controls during the initial phase of the pandemic. STUDY DESIGN Sixty-five patients who were hospitalized due to infection with the influenza A(H1N1)pdm09 virus and 46 healthy controls were studied. A hemagglutination inhibition assay (HIA) was performed to measure anti-influenza antibody titers in these subjects. Protein levels of the cytokines interleukin (IL)-4, IL-6, IL-8, IL-10, tumor necrosis factor-α (TNFα), interferon gamma (IFNγ), transforming growth factor beta (TGFβ)1 and TGFβ2 were quantified in plasma. Single nucleotide polymorphisms in IL6, IL10 and TNFα were also assessed. RESULTS Influenza patients had lower antibody titers and produced significantly higher levels of IL-6, IL-10 and TNFα than healthy controls. The frequencies of the TNFα -308G, IL-10 -592C and IL-10 -1082A alleles and the IL10 -1082(A/A) genotype were associated with susceptibility to severe disease, while the haplotypes TNFα AG and IL-10 GTA and GCA were associated with protection from severe disease [P=0.016, OR (CI)=0.11 (0.01-0.96); P=0.0187, OR (CI)=0.34 (0.13-0.85); P=0.013, OR (CI)=0.39 (0.18-0.83)]. CONCLUSIONS This study demonstrates that the influenza A(H1N1)pdm09 patients and healthy controls have different profiles of immune parameters and that there is an association between IL-10 and TNFα polymorphisms and the outcome of this disease.

Entomological Indices, Feeding Sources, and Molecular Identification of Triatoma phyllosoma (Hemiptera: Reduviidae) One of the Main Vectors of Chagas Disease in the Istmo de Tehuantepec, Oaxaca, Mexico

The purpose of this study was to conduct an entomological analysis, determination of feeding sources, and molecular identification of triatomines in five communities of the Istmo de Tehuantepec, Oaxaca. The only found species in two of five searched communities (San Mateo del Mar and Tehuantepec City) was Triatoma phyllosoma. Colonization indices were high in both communities. In San Mateo del Mar, the insects were found indoors and in Tehuantepec City in peridomestic areas. The Trypanosoma cruzi infection indices were 2.1% in San Mateo del Mar and 39.4% in Tehuantepec City. This difference could be related to the high numbers of triatomine feeding on hens in the former community. In contrast, in Tehuantepec, dogs were the principal triatomine feeding sources. All nymphs and adults that were genetically analyzed belonged to the species T. phyllosoma. Low levels of genetic variation were found between vectors from both communities.

Update of phylogenetic and genetic diversity of Sporothrix schenckii sensu lato.

Sporothrix schenckii sensu lato causes subcutaneous mycosis. In this article we analysed its phylogeny and genetic diversity using calmodulin DNA sequences deposited in GenBank database. Population genetics indices were calculated, plus phylogenetic and haplotype network trees were built. Five clades with high values of posterior probability, 47 haplotypes and high diversity in the complex were found. Analysis of partial calmodulin sequences alignment revealed conserved and polymorphic regions that could be used as reference for taxonomic identification. The use of population genetics analysis allowed understanding the phylogenetic proximity of S. schenckii s. str. and S. brasiliensis; scarce genetic flow among them with low migration index and high ancestry coefficient was found. Similarly, S. globosa, S. mexicana and S. pallida sequences showed highly differentiated species with no genetic exchange. The phylogenetic tree suggests that S. mexicana shared a common ancestor with S. pallida; while S. globosa and S. brasiliensis are more related to S. schenckii s. str. and showed less haplotype diversity and restrictions in geographic distribution. In the haplotype network tree S. schenckii s. str. species displayed worldwide distribution without dispersion centres; while S. brasiliensis and S. globosa, exhibited Brazil and Euro-Asia as dispersion centres, respectively. Our data suggest that S. schenckii complex has been submitted to a divergent evolution process, probably due to the pressure of the environment and of the host. In contrast, S. brasiliensis could have been submitted to purifying selection or expansion process.