RESUMO
There have been few reports on extra-enteric infections by Blastocystis STs and none have been molecularly identified in samples from human reproductive organs. We report for the first time the identification of 3 different subtypes of Blastocystis (ST1-3) in vaginal and sperm samples, from patients infected with Trichomonas vaginalis. Blastocystis STs were identified by PCR-sequencing and by phylogenetic inferences using 28 vaginal swab samples and 7 sperm samples from patients trichomoniasis. Blastocystis STs were identified in 6 of 28 vaginal swabs (21.4%) and in 3 of 7 sperm samples (42.8%). In both biological samples, STs 1-3 were found; one vaginal sample showed subtype co-infection with ST1 and ST3. High genetic variation was observed in the sequences obtained and no specific clustering in the phylogenetic trees was detected. Most of the haplotypes identified were placed far from the main dispersal centers. Our finding suggested that incorrect cleaning of the genital area or a contamination by combination of anal and vaginal intercourse.
Assuntos
Infecções por Blastocystis , Blastocystis , Coinfecção , Trichomonas vaginalis , Blastocystis/genética , DNA de Protozoário/genética , Fezes , Feminino , Variação Genética , Humanos , Masculino , Filogenia , Sêmen , Espermatozoides , Trichomonas vaginalis/genéticaRESUMO
Extra-enteric infections by Blastocystis spp. have rarely been documented. Here, we report a case of extra-enteric blastocystosis in a patient with minimal cervicitis symptoms. A 47-year-old Hispanic female patient was attended in a primary health centre in Michoacan state, Mexico, for her routine gynaecological medical examination. As only symptom, she referred to a slight vaginal itching. The presence of several vacuolar-stages of Blastocystis spp. were identified by Papanicolaou staining; molecular identification was attempted by culture-PCR sequencing of a region of 18S gene from cervical and faecal samples obtained 2 months after cytological examination, even when patient declared that she tried self-medicating with vaginal ovules. Blastocystis ST1 was identified only in the faecal sample. The presence of Blastocystis spp. in the cervix of a patient with scarce symptomatology, demonstrates the extraordinary flexibility of this microorganism to adapt to new environments and niches.
Assuntos
Infecções por Blastocystis/parasitologia , Blastocystis/isolamento & purificação , Colo do Útero/parasitologia , Cervicite Uterina/parasitologia , Blastocystis/genética , Fezes/parasitologia , Feminino , Genes de Protozoários , Humanos , Pessoa de Meia-Idade , Teste de Papanicolaou , Reação em Cadeia da Polimerase , RNA Ribossômico 18SRESUMO
Blastocystis is a parasite commonly found in the gut of humans and animals; there are 22 known subtypes (STs). STs 1-9 and 12 have been found in humans. This parasite has a faecal-oral route of transmission; its high infection prevalence in developing countries is due to poor hygiene practices, exposure to infected animals, and intake of contaminated water or food. Its pathogenicity has not been established, because it has been found in symptomatic and asymptomatic patients. The goal of this study was to analyze the frequency of Blastocystis and its subtypes (1, 2, 3, 4, 5, and 7), and assess the relationship between these subtypes and abdominal pain and distension. 202 university students participated in this study. A questionnaire was applied to assess the gastrointestinal symptoms, and subsequently the students were asked to provide faecal samples. The presence of parasites was determined by optical microscopy. Blastocystis-positive samples had their DNA extracted and end-point PCR was performed to corroborate the presence of Blastocystis and determine its subtypes. Among the samples, 47.03% were positive according to PCR analysis. The most prevalent subtypes were ST3 (29.79%), ST4 (16.84%), and ST1 (14.89%). We found a relationship between ST1 and abdominal pain (OR = 0.196; CI = 0.0533-0.7318; p = 0.015), and between ST4 and abdominal distension (OR = 0.2928; CI = 0.1017-0.8429; p = 0.023). However, the presence of this parasite and the probable relationship with gastrointestinal symptoms suggest the need to determine its role within intestinal microbiota in order to confirm whether its eradication is really necessary or not.
RESUMO
BACKGROUND: Blastocystis spp. are the most prevalent intestinal eukaryotes identified in humans, with at least 17 genetic subtypes (ST) based on genes coding for the small-subunit ribosomal RNA (18S). It has been argued that the 18S gene should not be the marker of choice to discriminate between STs of these strains because this marker exhibits high intra-genomic polymorphism. By contrast, pyruvate:ferredoxin oxidoreductase (PFOR) is a relevant enzyme involved in the core energy metabolism of many anaerobic microorganisms such as Blastocystis, which, in other protozoa, shows more polymorphisms than the 18S gene and thus may offer finer discrimination when trying to identify Blastocystis ST. Therefore, the objective of the present study was to assess the suitability of the PFOR gene as an additional marker to discriminate among Blastocystis strains or subtypes from symptomatic carrier children. METHODS: Faecal samples from 192 children with gastrointestinal symptoms from the State of Mexico were submitted for coprological study. Twenty-one of these samples were positive only for Blastocystis spp.; these samples were analysed by PCR sequencing of regions of the 18S and PFOR genes. The amplicons were purified and sequenced; afterwards, both markers were assessed for genetic diversity. RESULTS: The 18S analysis showed the following frequencies of Blastocystis subtypes: ST3 = 43%; ST1 = 38%; ST2 = 14%; and ST7 = 5%. Additionally, using subtype-specific primer sets, two samples showed mixed Blastocystis ST1 and ST2 infection. For PFOR, Bayesian inference revealed the presence of three clades (I-III); two of them grouped different ST samples, and one grouped six samples of ST3 (III). Nucleotide diversity (π) and haplotype polymorphism (θ) for the 18S analysis were similar for ST1 and ST2 (π = ~0.025 and θ = ~0.036); remarkably, ST3 showed almost 10-fold lower values. For PFOR, a similar trend was found: clade I and II had π = ~0.05 and θ = ~0.05, whereas for clade III, the values were almost 6-fold lower. CONCLUSIONS: Although the fragment of the PFOR gene analysed in the present study did not allow discrimination between Blastocystis STs, this marker grouped the samples in three clades with strengthened support, suggesting that PFOR may be under different selective pressures and evolutionary histories than the 18S gene. Interestingly, the ST3 sequences showed lower variability with probable purifying selection in both markers, meaning that evolutionary forces drive differential processes among Blastocystis STs.
Assuntos
Infecções por Blastocystis/parasitologia , Blastocystis/classificação , Variação Genética , Enteropatias Parasitárias/parasitologia , Piruvato Sintase/genética , Adolescente , Teorema de Bayes , Blastocystis/enzimologia , Blastocystis/genética , Criança , Pré-Escolar , Fezes/parasitologia , Feminino , Haplótipos , Humanos , Lactente , Masculino , México , Filogenia , Polimorfismo Genético , Proteínas de Protozoários/genéticaRESUMO
Blastocystis is a human common enteric protist that may colonize a large variety of non-human hosts linked to symptoms and diseases such as abdominal pain, constipation, diarrhea, urticaria, flatulence and irritable bowel syndrome (IBS). Blastocystis exhibits remarkable genetic diversity and multiple subtypes (STs) within the genus with no absolute associations with clinical symptomatology. Here we analyzed fecal samples from Argentinean patients (n=270) belonging to symptomatic (urticaria and non-specific gastrointestinal symptoms, n=39) and asymptomatic control (n=28). Those patients infected with Blastocystis (n=67) were submitted for morphological analysis, DNA extraction, 18S PCR, sequencing and STs identification according to DNA barcoding. Blastocystis vacuolar forms were the predominant morphotype (75%), ameboid-like forms were evidenced in 1.5% of samples. Blastocystis ST3 was detected in 71.6% (n=48), of which 71.4%, (n=35) and 28.6% (n=14) belonged to symptomatic and asymptomatic respectively. Other subtypes identified were ST1 (14.9%), ST6 (7.5%) and ST2 (5.9%). Blastocystis 18S barcoding evidenced in non-urticaria symptomatic patients and asymptomatic control group the presence of allele 134 (ST3) (p<0.0001), while allele 34 (ST3) was detected in 85.7% (18/21) of symptomatic uricaria as compared with control group (1/21) (p<0.0001). The presence of a particular allele (a34) significantly associated with urticaria patients was detected and the clinical implications of these findings are herein discussed.