Exacerbated symptoms in Blastocystis sp.-infected patients treated with metronidazole: two case studies.

Blastocystis sp. is a gastrointestinal (GI) protozoan parasite reported to cause non-specific GI symptoms including diarrhea, flatulence, abdominal pain, and nausea. Complete eradication of Blastocystis sp. is rather challenging even with the drug of choice, i.e., metronidazole. Here, we report on two Blastocystis sp.-infected individuals, who presented increased parasite load and exacerbated symptoms upon treatment with the usual recommended dosage and regime of metronidazole. The two studies uniquely demonstrate for the first time a cyst count as high as fivefold more than the original cyst count before treatment and show an exacerbation of GI symptoms despite treatment. The study provides additional support in recognizing metronidazole resistance in Blastocystis sp. and its consequences towards the pathogenicity of the parasite.

PATHOGENICITY OF Blastocystis sp. TO THE GASTROINTESTINAL TRACT OF MICE: RELATIONSHIP BETWEEN INOCULUM SIZE AND PERIOD OF INFECTION / Histopatogenicidade de Blastocystis sp. para o trato gastrointestinal de camundongos: relação com inóculo e tempo de infecção

The pathogenic potential of Blastocystis sp. in experimental models requires further investigation. In this work, the pathogenicity of this parasite in the gastrointestinal tract of male Swiss mice was evaluated according to the inoculum size and period of infection. Animals were infected intragastrically, with 100, 500, 1,000, 5,000 and 10,000 Blastocystis sp. vacuolar forms obtained from a mixture of eight human isolates cultured axenically in Jones' medium. After seven, 14, 21, 28 and 60 days of infection, the animals were sacrificed and fragments of the small intestine (duodenum), large intestine, and cecum were subjected to histopathological analysis. Blastocystis sp. triggered an inflammatory response in the different tissues analyzed, with a predominance of mononuclear cells. The parasite was found in the muscular layer of the cecum, showing its invasive character. Larger inocula triggered inflammatory processes earlier (seven days) than smaller ones (from 21 days). We conclude that, in the proposed model, the pathogenicity of Blastocystis sp. isolates that were studied is related to inoculum size and period of infection.

Pouco é sabido sobre o potencial patogênico de Blastocystis sp. em modelos experimentais. Neste trabalho a patogenicidade desse parasito para o trato gastrointestinal de camundongos Swiss machos foi avaliada de acordo com o inóculo e tempo de infecção. Os animais foram infectados, via intragástrica, com 100, 500, 1.000, 5.000 e 10.000 formas vacuolares de Blastocystis sp. obtidos a partir de uma mistura de oito isolados humanos cultivados axenicamente em meio Jones. Após 7, 14, 21, 28 e 60 dias de infecção os animais foram sacrificados e fragmentos do intestino delgado (duodeno), grosso e ceco foram retirados para análise histopatológica. Blastocystis sp. desencadeou resposta inflamatória nos diferentes tecidos analisados, com predominância de infiltrado mononuclear. No ceco o parasito foi encontrado na túnica muscular mostrando seu caráter invasivo. Inóculos maiores desencadearam processos inflamatórios mais precocemente (7 dias) e inóculos menores mais tardiamente (a partir de 21 dias). Conclui-se que no modelo proposto a patogenicidade dos isolados de Blastocystis sp. estudados tem relação com o inóculo e tempo de infecção.

PATHOGENICITY OF Blastocystis sp. TO THE GASTROINTESTINAL TRACT OF MICE: RELATIONSHIP BETWEEN INOCULUM SIZE AND PERIOD OF INFECTION.

The pathogenic potential of Blastocystis sp. in experimental models requires further investigation. In this work, the pathogenicity of this parasite in the gastrointestinal tract of male Swiss mice was evaluated according to the inoculum size and period of infection. Animals were infected intragastrically, with 100, 500, 1,000, 5,000 and 10,000 Blastocystis sp. vacuolar forms obtained from a mixture of eight human isolates cultured axenically in Jones' medium. After seven, 14, 21, 28 and 60 days of infection, the animals were sacrificed and fragments of the small intestine (duodenum), large intestine, and cecum were subjected to histopathological analysis. Blastocystis sp. triggered an inflammatory response in the different tissues analyzed, with a predominance of mononuclear cells. The parasite was found in the muscular layer of the cecum, showing its invasive character. Larger inocula triggered inflammatory processes earlier (seven days) than smaller ones (from 21 days). We conclude that, in the proposed model, the pathogenicity of Blastocystis sp. isolates that were studied is related to inoculum size and period of infection.

Stress exacerbates infectivity and pathogenicity of Blastocystis hominis: in vitro and in vivo evidences.

BACKGROUND: Stress alters the oxidant-antioxidant state and immune cell responses which disrupts its function to combat infection. Blastocystis hominis, a common intestinal protozoan has been reported to be opportunistic in immunocompromised patients namely cancer. B. hominis infectivity in other altered immune system conditions especially stress is unknown. We aimed to demonstrate the stress effects towards the susceptibility and pathogenicity of B. hominis infection. METHODS/FINDINGS: Three-week-old Wistar rats were divided into four groups: (a)control; (b)stress-induced; (c)B. hominis infected; (d)stress-induced with B. hominis infection; (n = 20 respectively). Stress was induced for an hour daily (30 days) using a Belly Dancer Shaker. Weight gain was monitored, stool samples were collected for B. hominis screening and blood for the determination of differential count, levels of immunoglobulin, oxidative damage, and peripheral blood mononuclear cell (PBMC) proliferation upon induction with solubilized antigen of B. hominis (Blasto-Ag). Group (b) exhibited the highest level of weight gain. Group (d) had higher levels of parasite cyst count in stools, serum IgE, oxidized protein and lipid compared to the group (c). Levels of monocyte and antioxidant in group (d) were decreased and their PBMCs showed highest inhibition of proliferation level when exposed to Blasto-Ag. Monocyte level in Group (b) showed insignificant difference compared to group (a) but was significantly lower compared to group (c). Antioxidant levels in group (c) were generally lower compared to group (a) and (b). Inhibition level exhibited by Blasto-Ag treated PBMCs of group (c) was higher compared to group (a) and (b). CONCLUSION: The pathogenicity and augmentation of B. hominis infection is enhanced when stress is present. Lifestyles today are becoming increasingly stressed and the present findings suggest that the parasite which has been reported to be one of the most common organisms seen in stool surveys, namely in developing countries, may tend to be more pathogenic in stressful situations.

[Investigation of the genetic profile of Blastocystis species in Saint Petersburg residents with gastrointestinal tract diseases in different age groups].

A total of 1500 people, including 1273 children with various gastrointestinal tract diseases and 327 patients with chronic viral hepatitis C, were examined. Microscopy and polymerase chain reaction (PCR) were used to determine Blastocystis in the feces. Blastocysts were detected in 33% of the patients with chronic viral hepatitis C and in 4.9% of the children. Genotyping established that Blastocystis species subtype 3 (antroponous) was encountered relatively rarely (25%) in these patients; there were most common Blastocystis species subtypes 5 (36.1%) and 6 (36.1%). Significant intestinal dyspepsia was noted in all the patients with chronic hepatitis C and Blastocystis invasion. Blastocystis species subtype 3 was prevalent (62.3%) among the examined children. The other subtypes were less frequently detected. These were subtype 1 (29.5%), subtype 2 (24.3%), subtype 4 (1.3%), and subtype 7 (3.8%) whereas subtype 5 and subtype 6 were not found in any case. The comparison of clinical symptoms in children could reveal the following tendency: there were digestive disorders and skin allergic reactions with Blastocystis species subtype 1 and subtype 2, respectively.

Drinking water is a significant predictor of Blastocystis infection among rural Malaysian primary schoolchildren.

Blastocystis infection has a worldwide distribution especially among the disadvantaged population and immunocompromised subjects. This study was carried out to determine the prevalence and the association of Blastocystis infection with the socio-economic characteristics among 300 primary schoolchildren, living in rural communities in Lipis and Raub districts of Pahang state, Malaysia. Stool samples were collected and examined for the presence of Blastocystis using direct smear microscopy after in vitro cultivation in Jones' medium. The overall prevalence of Blastocystis infection was found to be as high as 25.7%. The prevalence was significantly higher among children with gastrointestinal symptoms as compared to asymptomatic children (x2 =4.246; P=0.039). Univariate and multivariate analyses showed that absence of a piped water supply (OR=3.13; 95% CI=1.78, 5.46; P<0.001) and low levels of mothers' education (OR=3.41; 95% CI=1.62, 7.18; P<0.01) were the significant predictors of Blastocystis infection. In conclusion, Blastocystis is prevalent among rural children and the important factors that determine the infection were the sources of drinking water and mothers' educational level. Interventions with provision of clean water supply and health education especially to mothers are required.

New insights on classification, identification, and clinical relevance of Blastocystis spp.

SUMMARY: Blastocystis is an unusual enteric protozoan parasite of humans and many animals. It has a worldwide distribution and is often the most commonly isolated organism in parasitological surveys. The parasite has been described since the early 1900s, but only in the last decade or so have there been significant advances in our understanding of Blastocystis biology. However, the pleomorphic nature of the parasite and the lack of standardization in techniques have led to confusion and, in some cases, misinterpretation of data. This has hindered laboratory diagnosis and efforts to understand its mode of reproduction, life cycle, prevalence, and pathogenesis. Accumulating epidemiological, in vivo, and in vitro data strongly suggest that Blastocystis is a pathogen. Many genotypes exist in nature, and recent observations indicate that humans are, in reality, hosts to numerous zoonotic genotypes. Such genetic diversity has led to a suggestion that previously conflicting observations on the pathogenesis of Blastocystis are due to pathogenic and nonpathogenic genotypes. Recent epidemiological, animal infection, and in vitro host-Blastocystis interaction studies suggest that this may indeed be the case. This review focuses on such recent advances and also provides updates on laboratory and clinical aspects of Blastocystis spp.

Phenotypic and genotypic characterisation of Blastocystis hominis isolates implicates subtype 3 as a subtype with pathogenic potential.

Despite frequent reports on the presence of Blastocystis hominis in human intestinal tract, its pathogenicity remains a matter of intense debate. These discrepancies may be due to the varying pathogenic potential or virulence of the isolates studied. The present study represents the first to investigate both phenotypic and genotypic characteristics of B. hominis obtained from symptomatic and asymptomatic individuals. Symptomatic isolates had a significantly greater size range and lower growth rate in Jones' medium than asymptomatic isolates. The parasite cells of symptomatic isolates exhibited rougher surface topography and greater binding affinity to Canavalia ensiformis (ConA) and Helix pomatia (HPA). The present study also identifies further phenotypic characteristics, which aided in differentiating the pathogenic forms from the non-pathogenic forms of B. hominis. Blastocystis subtype 3 was found to be correlated well with the disease.

Clinical significance of Blastocystis hominis: experience from a medical center in northern Taiwan.

BACKGROUND AND PURPOSE: Blastocystis hominis is an intestinal protozoan. The pathogenic role of this organism in human beings is still controversial and has varied among reports from different geographic areas. The purpose of this study was to determine the clinical significance of B. hominis in northern Taiwan. METHODS: A total of 100 patients who had a positive B. hominis stool examination during the period April to December of 2001 were retrospectively identified from Taipei Veterans General Hospital. The demographic and clinical characteristics of these patients were reviewed from the medical records. RESULTS: All of the patients were adults. Fifty nine patients had more than one underlying diseases, including malignancies. Twenty one patients presented with fever and 10 patients had gastrointestinal symptoms, including diarrhea and/or abdominal pain. However, all of the patients had other conditions that might have contributed to the clinical presentation, and they improved without specific treatment for B. hominis. Furthermore, there were no significant differences in clinical symptoms and white blood cell count between patients with malignancy or diabetes mellitus and those without. Six patients had hypereosinophilia that could not be attributed to other causes. Among 34 patients who had a further stool examination within one year, B. hominis was undetectable in 31 patients (91.2%), despite their having no specific antiprotozoal treatment. CONCLUSIONS: The association of clinical symptoms and B. hominis could not be delineated from our study, even in immunocompromised patients. All of the patients improved without receiving any specific therapy. More studies from different areas are needed in order to delineate the clinical significance B. hominis.

A possible link between subtype 2 and asymptomatic infections of Blastocystis hominis.

Blastocystis hominis is one of the most common eukaryotic organisms in the intestinal tract of humans, while its pathogenic potential is still controversial. A total of 286 stool samples obtained from adult and pediatric patients with or without gastrointestinal symptoms in two hospitals in Manisa, Turkey, were cultured to detect B. hominis infection. Forty-one and 51 isolates were obtained from the adults and children, respectively, and these isolates were subjected to subtyping by polymerase chain reaction (PCR) with the known sequence-tagged site primers. The correlation between the genotype and the symptoms was evaluated. PCR subtyping indicated that subtype 3 was the most common genotype in both symptomatic and asymptomatic groups, and the second common genotype was subtypes 1 and 2 in symptomatic and asymptomatic groups, respectively. A significant correlation between subtype 2 and the asymptomatic groups was found among both in pediatric and adult patients (chi(2) (cal) = 4.38, df = 1, p = 0.044). However, there were no significant differences between the other genotypes and the symptomatic or asymptomatic groups, as well as both the age and sex of the patients. The present study suggests that subtype 2 is a non-pathogenic genotype of B. hominis.

Molecular epidemiology of Blastocystis infections in Turkey.

Blastocystis is a very common unicellular intestinal parasite of ubiquitous occurrence. In order to describe the molecular epidemiology of Blastocystis infections in Turkey, 87 isolates from 69 symptomatic and 18 asymptomatic individuals were sequenced. Sequence data were phylogenetically analyzed and statistically tested against unmodifiable risk factors such as gender and age. Blastocystis-positive males were complaining mainly of gastroenteritis, whereas dyspepsia was the chief complaint among Blastocystis-positive females. Blastocystis sp. subtypes detected in the study included subtypes 1, 2, 3 and 4, subtype 3 being the most predominant (75.9%). No association was detected between Blastocystis sp. subtype and symptoms (p>0.365), or between infection intensity and symptoms (p>0.441). There was a tendency of subtype 2 isolates being more common among older study individuals, and subtype 2 isolates were significantly associated with higher parasite abundance (p=0.017). Compared to data from similar studies, the distribution of Blastocystis sp. isolates in Turkey was found to more or less reflect the one seen in other countries, and it was deduced that subtype 3 is generally by far the most common subtype infecting humans, followed by subtypes 1, 2 and 4.

Pathophysiological variability of different genotypes of human Blastocystis hominis Egyptian isolates in experimentally infected rats.

The genotyping of Blastocystis hominis clinical isolates obtained from 28 gastrointestinal symptomatic patients and 16 asymptomatic individuals were identified by polymerase chain reaction using sequenced-tagged site (STS) primers. Then, pathophysiological variability between different B. hominis genotypes was evaluated in experimentally infected rats. Only four B. hominis subtypes (1, 2, 3, and 4) were detected (18.2%, 9.1%, 54.5%, and 18.2%, respectively) in human isolates. In symptomatic isolates, subtypes 1, 3, and 4 were detected in 8 (28.6%), 16 (57.1%), and 4 (14.3%) patients, respectively. In asymptomatic isolates, subtypes 2, 3, and 4 were identified in 4 (25%), 8 (50%), and 4 (25%), respectively. Subtype 3 was the commonest in humans. Different degrees of pathological changes were found among infected rats by symptomatic subtypes compared with asymptomatic subtypes. The moderate and severe degrees of pathological changes were found only in symptomatic subtypes infected rats while mild degree was found only in asymptomatic subtypes infected rats. Only subtype 1 induced mortality rate with 25% among infected rats. On evaluation of the intestinal cell permeability in the Ussing chamber, a prominent increase in short circuit current (DeltaIsc) was found in symptomatic subtype 1 compared to symptomatic subtypes 3 and 4 infected rats. Minimal effects were found in the asymptomatic and control groups. The results proved that subtype 1 was clinically and statistically highly relevant to the pathogenicity of B. hominis while subtype 2 was irrelevant. Also, the results suggest the presence of pathogenic and nonpathogenic strains among subtypes 3 and 4.

Pathogenicity of Blastocystis hominis, a clinical reevaluation.

Blastocystis (B.) hominis was considered to be a member of normal intestinal flora in the past, but in recent years it has been accepted as a very controversial pathogenic protozoan. In this study, 52 individuals whose stool examination revealed B. hominis were evaluated for clinical symptoms. Metronidazole was administered for 2 weeks to the patients infected with B. hominis. After 2 weeks of treatment they were called for a follow-up stool examination. No other bacteriological and parasitological agents were found during stool examination of these patients. The frequency rate of intestinal symptoms was 88.4% in the B. hominis cases. Abdominal pain was the most frequent symptom (76.9%). Diarrhea and distention followed at a rate of 50.0% and 32.6%. Intestinal symptoms may be seen frequently together with the presence of B. hominis and this protozoan may be regarded as an intestinal pathogen, especially when other agents are eliminated.

The effect of Blastocystis hominis on the growth status of children.

BACKGROUND: B. hominis is a protozoan parasite commonly found in the human gastrointestinal tract. The pathogenesis of B. hominis is still controversial, although it is one of the most common parasites found in stool samples. MATERIAL/METHODS: This study was conducted at the Adnan Menderes Medical Faculty between January 2002 and June 2003 to evaluate the relationship between B. hominis and growth status in children in Aydin, Turkey. Healthy children with positive stool samples for B. hominis but negative for other parasites were selected as the case group (n=89). Two controls matched to each case by age and gender were selected by random sampling of children with negative stool samples for any parasite (n=178). RESULTS: The anthropometric measurements and body mass index were significantly lower in the case group than in the control group (p<0.05). CONCLUSIONS: According to this study there is a correlation between the presence of B. hominis and lower anthropometric indexes in children.

Secretory and humoral antibody responses to Blastocystis hominis in symptomatic and asymptomatic human infections.

The study included 3 groups of individuals, in the first 2 groups they had positive stool microscopic examinations only for B. hominis indicating blastocystosis, with and without gastrointestinal symptoms, respectively, while the last group included apparently healthy individuals with no parasites in stool. Stool and serum samples of these individuals were subjected to detection of anti-B. hominis fecal and serum IgA and serum IgG antibodies by indirect ELISA, and detection of B. hominis fecal and serum antigens by double sandwich ELISA. In symptomatic B. hominis infections with positive stool microscopy the study recorded first: specific secretory IgA and humoral IgA and IgG antibody responses at a prevalence of 100%, 83.3% and 86.6%, respectively, with an increased significant difference (P<0.001) of each from healthy controls, together with an increase in level of secretory IgA than that of humoral IgA antibody (P<0.001), and second: the presence of specific antigens in stool and serum at a prevalence of 96.6% and 90%, respectively. With an increased significant difference (P<0.001) of each from healthy controls together with the former at a higher level than the latter (P<0.05). In asymptomatic B. hominis infections with positive stool microscopy the study recorded first; absence of each of the studied specific secretory and humoral antibody responses with no significant difference (P>0.05) of each from healthy controls, and second; absence of specific antigens in stool and serum with no significant difference (P>0.05) of each from healthy controls nor from each other. The explanations and implications of these results are discussed.

Protozoon infections and intestinal permeability.

Intestinal permeability (IP) studies using some macromolecules have been assumed to demonstrate the intactness of intestinal mucosa. The aim of the present study is to determine the changes in IP among patients with protozoan infections. Thirty nine patients with protozoan infections and ten healthy controls were enrolled in the study. Protozoa were diagnosed by Native-lugol, Richie and Trichrome staining of faeces. IP was evaluated by diethyl triamine penta acetic acid labeled with 99m Technetium (99mTc labeled DTPA) assay. The IP was found to have increased in patients with protozoan infections compared with control patients (7.20+/-5.52 vs. 4.47+/-0.65%, P=0.0017). The IP values were 9.91+/-10.05% in Giardia intestinalis group, 6.81+/-2.25% in Blastocystis hominis group, 5.78+/-2.84% in Entamoeba coli group. In comparison with the control group, the IP was significantly higher in G. intestinalis and B. hominis patients (P=0.0025, P=0.00037, respectively), but not in E. coli patients. In conclusion, the IP increases in patients with G. intestinalis and B. hominis but not with E. coli infection. This finding supports the view that IP increases during the course of protozoan infections which cause damage to the intestinal wall while non-pathogenic protozoan infections have no effect on IP. The increase in IP in patients with B. hominis brings forth the idea that B. hominis can be a pathogenic protozoan.

Morphology, histochemistry and infectivity of Blastocystis hominis cyst.

Different morphological forms of Blastocystis hominis had been identified in human stool samples. These included both cystic and trophic stages. The latter was induced to encyst by keeping them in potassium dichromate solution for two weeks. Suspected of being the infective stage, cysts were studied in more detail as regards their morphology using both light and electron microscopy. Histochemistry and infectivity studies were also carried out. Light microscopy revealed the cysts to be ovoid or round 5-7 microm with thick cyst wall and a single nucleus. The induced cysts were morphologically similar to the cysts present in the stool. By electron microscope, the cyst wall was evident surrounded by an additional fibrillar coat. The cytoplasm contained one nucleus, many mitochondria, glycogen deposits and a number of variable sized vacuoles. Histochemical studies detected carbohydrates in the cyst wall and fat globules in the cytoplasm. Oral inoculation of albino mice with these cysts led to inflammatory changes in the large and small intestine. The parasite was found at the mucosal epithelium but with no invasion. Different forms of the parasite were detected in the lumen of the intestine.

Blastocystis hominis: parasita intestinal pouco estudado no Brasil / Blastocystis hominis

O objetivo deste artigo é difundir o conhecimento sobre Blastocystis hominis e a doença que produz. O autor apresenta uma revisão começando com os dados históricos e taxonômicos, descrevendo, a seguir, a morfologia, biologia e patogenia do protozoário. Conclui analisando os aspectos clínicos e epidemiológicos, incorporando sua própria experiência no diagnóstico e tratamento desta parasitose intestinal. As considerações realizadas pretendem motivar a realização de pesquisas que permitam determinar as características clínico-epidemiológicas da blastocistose no Brasil