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Although Blastocystis sp. has been classically considered a commensal parasite with limited pathogenicity, recent studies suggest that its pathogenic potential is high. We report the case of a 9-year-old Spanish male who presented with peritonitis secondary to acute appendicitis with abundant intra-abdominal turbid-free fluid. A standard appendectomy was performed, and a sample of the fluid was taken for microbiological culture. Multimicrobial flora was isolated in peritoneal fluid culture. The antibiotic resistance study showed that all the microorganisms were sensitive to meropenem. On the 5th postoperative day, a control blood test showed relative eosinophilia and a persistently elevated C-reactive protein. A stool parasitological study showed abundant cysts morphologically compatible with Blastocystis hominis . The hematoxylin & eosin and Giemsa study identified abundant parasitic cysts in the appendix. The patient evolved favorably and is currently asymptomatic and under follow-up. Regarding acute appendicitis, there is only one report in the literature of peritonitis of appendiceal origin associated with Blastocystis sp. In conclusion, although infrequent, parasitosis should be considered as a potential etiological agent of acute appendicitis, even in nonendemic areas. Relative eosinophilia or persistently elevated acute phase reactants despite adequate antibiotic coverage should help to establish diagnostic suspicion., Competing Interests: The authors have no funding or conflicts of interest to disclose., (Copyright © 2024 Wolters Kluwer Health, Inc. All rights reserved.)

Blastocystis sp. and Dientamoeba fragilis are intestinal protists, which are common worldwide, but the pathogenic role of these organisms in gastrointestinal diseases is still controversial. This study aimed to investigate the frequency of Blastocystis sp. and D. fragilis in stool samples from adult patients with celiac disease (CD) by using conventional and molecular methods. A total of 75 patients with CD and 75 healthy individuals were included in this study. Fresh stool specimens collected from each individual were analyzed by conventional and molecular methods. The overall prevalence of Blastocystis sp. and D. fragilis was 41.3% (31/75) and 24% (18/75) in patients with CD, and 46.7% (35/75) and 13.3% (10/75) in healthy controls, respectively. There was no statistically significant difference in the prevalence of Blastocystis sp. and D. fragilis between CD patients and healthy individuals. Blastocystis sp. subtypes were identified in 20 CD and 16 control patients and the overall subtype distribution was observed as ST1 13.9%, ST2 30.6%, and ST3 55.6%. The prevalence of Blastocystis sp. and D. fragilis in adults with CD is similar to the prevalence of protozoa in healthy adults. In this study, the most prevalent Blastocystis subtype was ST3 and the most frequent allele was a34 in both CD patients and healthy individuals. No significant difference was found between the two groups in terms of the detection rates of Blastocystis sp. and D. fragilis, and it is thought that both protists may be colonisers of the intestinal microbiome., Competing Interests: Declaration of competing interest Not applicable., (Copyright © 2024. Published by Elsevier B.V.)

Introduction and Objective: Intestinal parasitoses are important causes of morbidity and mortality, especially in immunocompromised individuals. In patients with chronic renal insufficiency (CRI), the accumulation of non-excreted metabolites leads to uraemia, which induces a state of immunodeficiency, increasing the incidence of infections. The aim of the study was molecular screening for enteric protozoa in patients with chronic renal insufficiency., Material and Methods: A total of 53 samples were collected in January 2023 from patients undergoing dialysis at Logman Ltd. Nephrodialysis Centre in Košice, Slovakia. Samples were examined by polymerase chain reaction (PCR) for the presence of Cryptosporidium parvum / Cryptosporidium hominis , Giardia intestinalis , Microsporidia spp., and Blastocystis sp., Results: From the 53 samples, the only pathogen identified by PCR was Blastocystis sp., in 13 patients (24.5 %). Sequence analyses confirmed that the most prevalent subtype (ST) among patients was ST 3 (n=9, 69.2%), followed by ST 1 (n=3, 23.1%) and ST 2 (n=1, 7.7%)., Conclusions: Molecular methods for the detection of microscopic enteric parasites are not used as a first-line diagnostic method in Slovakia. In immunocompromised patients, diarrhoea can be caused not only by a chronic disease or therapy but can also be a result of an ongoing underdiagnosed infection. Early diagnosis leads to targeted therapy and subsequent partial improvement of the quality of life. This study also shows the first insights into Blastocystis sp. subtype distribution in humans in Slovakia.

Urticaria is a skin rash with several etiologic factors, including infectious agents. Blastocystis hominis is an intestinal protozoan parasite that has been linked to urticaria and skin lesions. The aim of this work was to investigate the association between B. hominis infection and chronic urticaria. In a case-control study, stool samples were obtained from 94 patients with chronic urticaria as case group and 285 healthy individuals as control group. Urticaria activity score 7 (UAS7) was used to score the severity of urticaria, classified as mild, moderate and intense. All stool samples underwent routine stool examinations, as well as polymerase chain reaction (PCR) for the detection of B. hominis. Molecular detection was carried out using the small subunit ribosomal RNA (SSU-rRNA) gene and the parasite subtypes were determined by sequencing. The rate of B. hominis infection was 21.3% (20 out of 94) and 17.2% (49 out of 285) between the case and control groups, respectively (p = 0.463). Three subtypes of B. hominis, including ST-1, ST-2 and ST-3, were detected in the case and control groups (ST-1 = 30% vs. 8.3%, ST-2 = 40% vs. 25% and ST-3 = 30% vs. 66.6% in the case and control group, respectively), which was statistically significant (p = 0.00001). However, no statistical differences were found between the severity of the urticaria and the B. hominis subtypes (p = 0.533). This study revealed a higher prevalence (but not significant) of B. hominis infection among patients with urticaria than healthy individuals. However, the results did not find a significant association between the subtypes of B. hominis and the severity of urticaria., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Blastocystis spp. is a ubiquitous protozoon in the intestinal tract of human and many animals. Microscopic examination is the main method of clinical diagnosis for Blastocystis spp. , which is prone to false negative. A simple and rapid diagnosis of Blastocystis spp. infection is an important step to prevent and control blastocystosis. Here, a recombinase polymerase amplification-lateral flow dipstick (RPA-LFD) assay was developed for rapid visual detection of Blastocystis spp. DNA amplification could be performed within 18 min at 37°C. The minimum DNA detection limit was 1 pg/μL, and there was no cross-reactivity with 12 other non-target pathogens, which was consistent with the sensitivity of conventional PCR (cPCR). Furthermore, 56 fecal samples from the Third Affiliated Hospital of Xinxiang Medical University were tested using RPA and cPCR methods respectively, and the results were completely consistent. The results show that RPA-LFD method has high accuracy and visual results, which provides a new choice for the differential diagnosis and rapid field detection of Blastocystis spp., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Mei, Su, Xin, Jia, Zhang, Yang, Xiaowei, Zhang and Wang.)

Blastocystis is an intestinal microeukaryote that has raised attention due to its wide distribution in animals and humans. The risk of zoonotic circulation primarily arises from close contact with infected animals. Therefore, the following study aimed to evaluate the diversity and frequency of Blastocystis subtypes in Colombian human and animal samples using complete sequencing of the 18S rRNA gene. For this purpose, 341 human stool samples and 277 animal fecal samples (from cattle, sheep, goat, pigs, cats, and dogs), were collected from different Colombian regions and analyzed using PCR-based detection and full-length 18S SSU rRNA gene Next-Generation Sequencing (NGS). Among the 618 samples from both hosts, humans and animals, the results revealed widespread Blastocystis frequency, with 48.09% (n = 164) in humans and 31.4% (n = 87) detection in animals. Dogs, cats, sheep, pigs, and wild animals tested positive, aligning with global prevalence patterns. Also, 29 human samples and 23 animal samples were sequenced using ONT technology from which 11 long-read unique sequences were generated and cluster with their compared reference sequences. The subtype distribution varied within hosts, detecting ST1 and ST3 in both human and animal samples. Subtypes ST5, ST10, ST14, ST15, ST21, ST24, ST25 and ST26 were limited to animals hosts, some of which are considered to have zoonotic potential. On the other hand, ST2 was found exclusively in human samples from Bolivar region. Mixed infections occurred in both animal and humans, 60.86% and 27.58% respectively. Moreover, to our knowledge, this is the first study in Colombia identifying ST15 in pigs and ST25 in sheep. The subtypes (STs) identified in this study indicate that certain animals may serve as reservoirs with the potential for zoonotic transmission. The identification of zoonotic subtypes highlights the use of Next Generation Sequencing as the depth and resolution of the sequences increases providing insights into STs of medical and veterinarian significance. It also reveals the coexistence of diverse subtypes among hosts. Further research is essential for understanding transmission dynamics, health implications, and detection strategies for Blastocystis occurrence in animals and humans, mainly associated to the role of animals as reservoirs and their close interaction with humans., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023. Published by Elsevier B.V.)

Blastocystis sp. is a common parasite in the intestinal tract of humans and animals. The clinical diagnosis of Blastocystis sp. mainly depends on the microscopic observation of parasite, which can lead to false-negative results. An accurate and convenient diagnostic approach for Blastocystis sp. infection is crucial for effectively preventing and controlling blastocystosis. Herein, we developed a recombinase polymerase amplification (RPA) method for detecting Blastocystis sp. The results showed that the DNA amplification by RPA established in this study could be performed within 5 min at 37°C, with maximum band intensity observed at 30 min. The minimum detection limit of RPA was 100 fg μ L −1 , consistent with conventional polymerase chain reaction (cPCR). Furthermore, the RPA method exhibited no cross-reactivity with 7 other non-target pathogens in the intestinal tract. Next, the newly established RPA method was used to analyse 40 fecal samples collected clinically, and the detection results were consistent with cPCR. These results corroborate that the newly developed RPA method has good sensitivity and specificity and offers the advantage of short detection times, which can be harnessed for differential diagnosis and rapid detection of Blastocystis sp.

Blastocystis sp. is among the most frequent intestinal protists identified in humans globally. However, characterization of Blastocystis subtype diversity in humans is ongoing. We report here the identification of novel Blastocystis subtype ST41 in a Colombian patient undergoing colorectal cancer screening involving colonoscopy and fecal testing (microscopy, culture, PCR). The full-length ssu rRNA gene sequence of the protist was generated using MinION long-read sequencing technology. The validity of the novel subtype was confirmed via phylogenetic and pairwise distance analyses of the full-length ST41 sequence and all other valid subtypes. The study provides reference material essential for conducting subsequent experimental studies., (© 2023 International Society of Protistologists.)

Blastocystis is a polymorphic enteric parasite with a worldwide distribution. It is one of the most common human intestinal protozoans in developing countries. The primary objective of this study was to determine the diagnostic value of microscopy, stool culture, and a polymerase chain reaction (PCR) technique for assessment of Blastocystis prevalence and risk factors. Human stool samples were collected from 110 individuals from Dakahlia governorate, Egypt as a part of a routine check-up or having gastrointestinal tract (GIT) symptoms. These samples were subjected to direct fecal smear microscopy, culture, and PCR for the detection of Blastocystis sp. Positive results for Blastocystis screening among the study population were 36 (32.7%), 41 (37.3%), and 43 (39.1%) by microscopy, PCR, and culture, respectively. Statistical analyses demonstrated that the agreement between the culture and PCR was perfect (Κ=0.925). Compared to culture, the sensitivity of PCR was 95% and the specificity was 97% while the sensitivity of microscopy was 84% and the specificity was 90.5%. We concluded that the in vitro culture and molecular assay have significant diagnostic value for the accurate detection and identification of Blastocystis in stool samples. The pathogenic potential of Blastocystis cannot be ruled out because our results found a link between Blastocystis carriage and gastrointestinal symptoms., (Copyright© by the Egyptian Association of Immunologists.)

Blastocystosis is an infection caused by Blastocystis sp., which colonizes the digestive tract of various hosts, including humans, although its pathogenicity is debated. It is crucial to detect and distinguish the different forms of Blastocystis to understand better its impact on human health and its epidemiological evolution. This study evaluated three diagnostic methods on 105 stool samples: direct examination, culture in Jones' medium, and conventional PCR. PCR is considered the gold standard and revealed a high prevalence of Blastocystis (67.62%) compared to direct examination (20.95%) and culture in Jones' medium (51.43%). Although the sensitivity of direct examination and culture was 31% and 76.1%, respectively, their specificity was 100%. No significant risk factors were identified. A statistically significant association was observed between Blastocystis infection and abdominal pain. Microscopic analysis revealed various morphological forms. Molecular diagnosis is an essential tool to determine the true prevalence of Blastocystis, and studying the different forms of this microorganism will contribute to a better understanding of its biological cycle and, therefore, the impact of this emerging infection on human health., (© M. Boutahar et al., published by EDP Sciences, 2023.)

The enteric protist Blastocystis has a worldwide distribution, however its prevalence in the human population is still underestimated, especially in developing countries where proper diagnosis is not performed in the routine of clinical laboratories. In this study, we aimed to assess the frequency, genetic diversity, and spatial distribution of Blastocystis isolates detected in fecal samples referred to a clinical laboratory for routine examination in inner São Paulo State, Brazil. A total of 348 leftover stool samples available for disposal from female and male individuals with age ranging from 3 months to 88 years were analyzed by both microscopic examination and PCR/sequencing of the SSU rRNA gene. The overall frequency of Blastocystis sp. was 31% (108/348), including 20.1% (70/348) and 31% (108/348) by microscopic examination and PCR/sequencing, respectively. Significant association was found only between Blastocystis infection and age, since the highest rate of positive samples was detected among 5-9 years old individuals (p < 0.0001). In addition, spatial distribution revealed a wide distribution of the positive samples, however they were densely concentrated in more populated areas. Seven subtypes were identified, namely ST1 (40.7%), ST2 (9.2%), ST3 (45.3%), ST4 (0.9%), ST6 (1.8%), ST7 (0.9%) and ST9 (0.9%). The intra-subtype analysis revealed a total of 25 different alleles previously reported. Here, the findings lead us to highlight the following aspects: (1) the identification of a ST9 isolate is a relevant finding since it is considered a very rare subtype in human infections as well as this is the first report in Brazil; (2) the high frequency of Blastocystis in fecal samples submitted for examination in a clinical laboratory points to the need to consider its search in routine parasitological examinations, (3) the spatial distribution of Blastocystis infection was not homogeneous but concentrated in more populated areas where the access for population to diagnostic services in healthcare is likely to be easier and, (4) the genetic variability of Blastocystis isolates suggests exposure of inhabitants living in inner municipalities to different sources of contamination involving anthroponotic and zoonotic transmission pathways., (Copyright © 2022. Published by Elsevier B.V.)

Blastocystis sp. - is the most prevalent anaerobic intestinal protozoan parasite in humans and many animals; from 1 to 2 billion people in the world are colonized by this pathogen. Blastocystis sp. is found both in faecal samples (FS) of healthy people (asymptomatic persons) and - patients (symptomatic persons) with nonspecific symptoms of gastrointestinal tract, skin, joints and other organs lesions. The prevalence of people affected by Blastocystis sp. of both cohorts in the world vary widely (from 0.08% to about 90%) depending on the degree of the country's economic development, sanitary and hygienic conditions, cultural values, etc. Currently, microscopic, cultural, immunological and molecular genetic methods are used for Blastocystis sp. detection in stool samples. Each group of methods of Blastocystis sp. detection/identification in FS has its advantages and disadvantages. The goal of this study was to determine the prevalence of Blastocystis sp. in faecal samples in different cohorts of people (clinically healthy and symptomatic people with symptoms of gastrointestinal lesions) in Kharkiv by microscopic and cultural methods. Cohort of surveyed residents of Kharkiv (n=169) included 72 clinically healthy individuals and 97 symptomatic individuals with gastrointestinal tract diseases. All 169 FSs (their precipitates) were subjected to microscopic examination after the formalin-phosphate-salt buffer (FPBSCS) enrichment (concentration) procedure (pH=7.4) at 500 g for 10 minutes. Blastocystis sp. identification was carried out by means of microscopy of the faecal smears, which were stained by Wheatley's modification trichrome stain (mWTS) and by Heidenhain's iron-hematoxylin stain (HIHS). The inoculated material was a filtered suspension of native FS (200 μl) which was inoculated in 5 ml of liquid media RPMI/IMDMEM (mixture of equal volumes of RPMI and IMDMEM media) with antibiotics and serum. Blastocystis sp. culture growth was carried out under anaerobic conditions at 37 oC for 5 days. The blastocysts final identification was carried out by means of light microscopy of suspensions smears stably stained with mWTS HIHS. It was carried out a comparative evaluation of the effectiveness Blastocystis sp. detection methods as microscopy (smears of enriched faecal material stained with mWTS or HIHS) and cultivation (on RPMI/IMDMEM medium) based on the results of parallel studies of 169 FS from different groups of people by both methods. An insignificant increase (4.1%) of the Blastocystis sp. frequency detection/identification by means of cultural method in comparison with the frequency of microscopic parasites detection in all FS was determined: in FS from asymptomatic individuals (n=72) only by 2.7%, and in FS from symptomatic individuals (n=97) - by 5.2% (p>0.05). From all FS in which Blastocystis sp. was detected microscopically, the growth of these parasite primary cultures was obtained. Among the total results (negative + positive) Blastocystis sp. detection / identification by microscopic and cultural methods in all FS from humans rf reaches +0.92, and for groups FS from asymptomatic and symptomatic individuals - rf=+0.94 and rf=+0.90, respectively. In the sample of only positive results detection / identification of Blastocystis sp. by microscopic and cultural methods, the value of rf is: + 0.59 for all studied FS from humans, + 0.20 - for FS from asymptomatic individuals and + 0.66 - for FS from symptomatic individuals. According to the results of a parallel study of microscopic and cultural methods of 169 FS from different groups of people it was found that the cultural method dominates over microscopic in sensitivity of Blastocystis sp. detection in FS (20.6%) and is characterized by a much higher level of specificity (accuracy of parasite identification), which reaches 100%. The method of in vitro diagnostics helps to increase the efficiency of parasites detection in human FS, can be used for epidemiological studies to establish the population prevalence of protozoa, to determine the sensitivity of Blastocystis sp. cultures to drugs, control of the etiotropic blastocystosis therapy effectiveness, obtaining parasites antigens, study the disease pathogenesis and the virulence potential of pathogen strains of different origin, etc.

Blastocystis is one of the most frequently detected protozoa in the human large intestine. One of the most effective and cheap methods for detecting Blastocystis in faeces is culture on a special medium in anaerobic conditions. Sampling faeces using traditional containers and their transport to the laboratory has certain limitations: a sample taken in this way should reach the laboratory relatively quickly, moreover, some patients are uncomfortable during sampling and protection of material in this way. We propose utilizing a swab for sampling and transportation of the faeces samples to be examined for Blastocystis instead of using traditional containers. We believe this is an excellent method allowing the material to be transported over longer distances without additional, and sometimes expensive, safety measures, and at the same time permitting the possibility of obtaining living cells after a relatively long period of storage.

Studies on the presence of Blastocystis subtypes in water samples are far less numerous compared with stool samples. The main aim of this study was to examine the occurrence of Blastocystis subtypes in 36 natural water bodies in north-western Poland in the period from winter 2009 to autumn 2010. Single PCR with the use of Blast 505-532/Blast 998-1017 set of primers was used to detect Blastocystis DNA in the obtained water samples. Sequencing of the obtained amplicons revealed the presence of ST1 and ST3 subtypes in five of the 36 (13.9%) examined water bodies within 1 year period. Further examinations with the use of new samples are needed in order to check if Blastocystis occurs in the examined water bodies at the present time, however, the risk of infection should be taken into consideration.

Introduction: Blastocystis sp. is a protozoan that colonizes the gastrointestinal tract of humans and many animals and is currently the most common parasite found in human stools. In some developing countries, its prevalence in study populations may exceed 50%. Morphologically, isolates of Blastocystis sp. found in different hosts are very similar. However, these same isolates show a very high genetic diversity between them and no less than 17 subtypes (or genotypes) have already been identified from molecular data. Genotyping studies have been carried out in many countries around the world and in particular in some Mediterranean countries such as France, spain, Italy, Turkey and Egypt. However, very little genotyping data is available in Algeria. To this end, we conducted the present study to identify and genotype Blastocystis in human and animal stool samples., Patients and Methods: One thousand eight hundred and sixty-nine (1,869) stool samples from kitchen staff as part of the periodic medical check-up, from subjects for the provision of a medical certificate required for the processing of a visa file and from patients with gastrointestinal disorders were examined. In addition to human faeces, animal samples, including 10 poultry, 2 cattle and 2 murine animals were examined. All stools were subjected to direct microscopic examination supplemented by concentration techniques and modified Ziehl Neelsen staining. Molecular characterization of 39 human and 14 animal isolates was performed by sequencing and the resulting sequences compared with those available from GenBank. Sequencing was only contributory for 30 human and 9 animal strains., Results: Of all human samples examined 284 were positive (15.19%) with a prevalence of 7.38% for Blastocystis. Of the 30 strains that were molecularly characterized, ST3 was predominant (15/30, 50%) followed by ST1 (10/30, 33.33%) and in third place ST2 (4/30, 13.33%). ST4 was identified in only one patient (1/30, 3.33%). The correlation between clinical status and the subtype of Blastocystis identified showed that the number of ST3 was high in asymptomatic subjects (11/15, 73%) compared to symptomatic subjects (4/15, 26.66%), as well as for the ST1 subtype (7/10, 70% versus 3/10, 30%). Conversely, the number of ST2 was higher in subjects with gastrointestinal disorders (3/4, 75%). In addition to human strains, we genotyped 7 avian, 2 murine and 2 bovine strains. Characterization of the avian strains revealed 5 ST6 (71.42%) and 2 ST7 (28, 57%). The murine and bovine strains are identified as ST7 and ST6 respectively., (Copyright © 2022 SFMTSI.)

Purpose: Blastocystis spp. are parasites of the intestinal tract found in many hosts including humans. This pathogen is commonly found in immunocompetent in asymptomatic individuals and in patients with gastrointestinal and extra-intestinal symptoms. Recently, it has been implicated as an important cause of diarrheal illness in immunocompromised individuals, including HIV-infected patients. At least six life cycle stages have been described in faeces and cultures, namely vacuolar, granular, multi-vacuolar, avacuolar, ameboid and cyst forms. The aim of the present study was to describe the histological findings of Blastocystis infection in an adult HIV-infected patient with gastrointestinal symptoms., Methods: Parasitological techniques and PCR were applied to stool samples. Histological analysis was performed on duodenal biopsy specimens., Results: Standard parasitological methods revealed vacuolar, granular, cyst and multi-vacuolar forms of Blastocystis in faecal samples with the presence of Blastocystis DNA being confirmed by PCR. DNA sequencing revealed Blastocystis subtype ST1. Histological findings in duodenal samples showed an inflammatory infiltrate with plasma cells and lymphocytes. We identified cyst, granular, ameboid and multi-vacuolar forms in the lumen., Conclusion: To our knowledge, there are no previous peer review reports describing these four different forms of Blastocystis in histological sections from the lumen and the brush border of the enterocytes., (© 2021. Witold Stefański Institute of Parasitology, Polish Academy of Sciences.)

Blastocystis is the most commonly found intestinal protist in the world. Accurate detection and differentiation of Blastocystis including its subtypes (arguably species) are essential to understand its epidemiology and role in human health. We compared (i) the sensitivity of conventional PCR (cPCR) and qPCR in a set of 288 DNA samples obtained from stool samples of gut-healthy individuals, and (ii) subtype diversity as detected by next-generation sequencing (NGS) versus Sanger sequencing. Real-time PCR resulted in more positive samples than cPCR, revealing high fecal load of Blastocystis based on the quantification curve in most samples. In subtype detection, NGS was largely in agreement with Sanger sequencing but showed higher sensitivity for mixed subtype colonization within one host. This fact together with use of the combination of qPCR and NGS and obtaining information on the fecal protist load will be beneficial for epidemiological and surveillance studies., (© M. Šloufová et al., published by EDP Sciences, 2022.)

Objective: The aim: To detect the infection rate of Blastocystis hominis in children less than 10 years old with diarrhea in Diyalaby polymerase chain reaction (PCR) method, to determine the subtype of Blastocystis hominis by sequencing the product of the positive result, and to determine the association between Blastocystis hominis infection and different factors such as gender, age, the level of mother education and the presence or absence animals in their houses., Patients and Methods: Materials and methods: A cross-sectional study was conducted on children with diarrhea at Al-Batool Teaching Hospital in Diyala governorate, during the period from November 2020 to April 2021, a total of 100 children 55 males and 45 females, then, stool samples were collected and examined by conventional polymerase chain reaction., Results: Results: The rate of infection with the parasite Blastocystis hominis was 8%, 8 out of 100. The infection was higher among females 62.5% than to males 37.5%, while the positive result was higher in the age group less than two years 75%, the highest percentage occur with patient whose mothers were incomplete primary and primary education was reached 37.5% and 25%; respectively and the study showed the highest percentage was with those who kept animals at homes was 75%., Conclusion: Conclusions: According to the genetic analysis of the sequence of eight samples that were positive for Blastocystis hominis parasite using the conventional polymerase chain reaction and they were back to the subtypes 3.

The present study aimed to evaluate the occurrence of Blastocystis sp. in Brazilian studies over a period of years (2000-2020), as well as point out relevant aspects of this enigmatic organism. We performed a literature search using six sources of international databases. The data were divided into diagnostic by parasitological and molecular techniques, and relevant aspects. After applying the inclusion and exclusion criteria, 52 studies were included in the final analysis. The occurrence of Blastocystis sp. in Brazil ranged from 0.5% to 86.6%, as determined using parasitological techniques. The highest occurrence was in the North (27.3%) and the lowest, in the Midwest region (13.4%). In Brazil, most studies have employed molecular techniques and are concentrated in the Southeast region. The Blastocystis sp. subtype ST3 had the highest average positivity, followed by ST1 and ST2. These findings represent a panorama that reflects the reality of Brazil; thus, we believe that the effectiveness of parasitological diagnosis should be considered with regard to making an appropriate choice of technique for detecting Blastocystis sp. Additionally, we emphasize the importance of further studies in the context of molecular epidemiology with regard to this genus. Blastocystis sp. is not well understood yet, and very little information regarding this genus is available; hence, further research regarding this genus is urgently needed.

Blastocystis sp. is a single-cell microorganism occurring in the gastrointestinal tract of humans and various animals and is distributed worldwide. Blastocystis exhibits extensive genetic diversity of 28 subtypes (STs) based on the small subunit ribosomal RNA (SSU rRNA) gene. In this study, the genetic diversity and zoonotic potential of Blastocystis were evaluated using pig faecal samples from two farms in Slovakia. Blastocystis spp. were detected in pigs intended for distribution and consumption. ST 5 subtype was identified in all positive samples and age categories with a prevalence of 12%. However, the prevalence on one of the farms was up to 28.6%. This is the first study of Blastocystis in pigs carried out in Slovakia. Although a number of samples obtained was small, the identified subtype of ST5 Blastocystis sp. occurs in humans and animals. It may have zoonotic potential and therefore may be a risk factor due to the close contact between humans and pigs on the breeding farms., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Blastocystis spp. is known as a common intestinal protozoan parasite in human and animals. The parasite has a worldwide distribution and is frequently detected in faecal samples in clinical parasitology laboratories. The goal of the study was to compare the sensitivity and specificity of formol-ether technique (FECT), trichrome staining, xenic in vitro culture (XIVC), microscopy of faecal smears, and polymerase chain reaction (PCR) methods for detecting Blastocystis spp. in human stool samples. The prevalence of the parasite in the stool samples referred to educational hospitals was also determined. A total of 575 cases were assessed to detect the parasite. After collecting from patients referring to Urmia educational hospitals, the samples were examined by microscopy of faecal smears, trichrome staining, FECT, XIVC using Jones' medium, and PCR, to evaluate the presence of Blastocystis spp. Microscopy of faecal smears, trichrome staining, FECT, and PCR technique detected 94, 100, 96, and 44 positive cases, with the sensitivity of 71.3%, 74.4%, 74.4%, and 80.4% and the specificity of 99.6%, 99.1%, 100%, and 93.1%, respectively. XIVC method identified the highest number of positive cases (129 cases) among the other methods. Our findings indicates that XIVC technique is more sensitive method for the detection of Blastocystis spp. in human stool, as compared to direct smear, trichrome staining, FECT, and PCR methods.

The symptomatic form of Blastocystis spp. infection not only with mild diarrhea or dysentery-like syndrome, but also with the development of severe ulcerative necrotic lesions of the intestine. Meanwhile, the pathogenicity of these microorganisms should not be exaggerated, due to majority asymptomatic cases or infection transmission with minor impaired bowel function.

Acute gastroenteritis with persistent vomiting, high degree fever and blood streaking stools often suggests bacterial aetiology in children. Authors report a 13-year-old boy presenting with acute watery diarrhoea with persistent vomiting, fever of 103°F, abdominal cramps and blood streaking stools who failed to show any response to parenteral third-generation cephalosporin for 72 hours. The stool examination revealed numerous cystic and amoeboid forms of Blastocystis hominis Metronidazole was started and the boy promptly responded within 24 hours. There was no recurrence of symptoms then onwards. The case highlights the crucial stool examination in case of acute diarrhoeal disease for rare aetiology., Competing Interests: Competing interests: None declared., (© BMJ Publishing Group Limited 2020. No commercial re-use. See rights and permissions. Published by BMJ.)

Objective: Since the identification of Blastocystis subtypes (ST) in the last decade, much has been learned about the genetic diversity of Blastocystis isolates in different populations, except pregnant women. The objective of this study is to investigate the genetic diversity of Blastocystis in pregnant women and analyse some demographic factors., Methods: The faecal samples from 100 pregnant women were collected at an Obstetrics and Gynecology Department in Muğla, Turkey. Thereafter, Blastocystis positivity was detected by direct microscopy and culture. The positive cultures were subjected to DNA isolation, and the Blastocystis barcode region was amplified with polymerase chain reaction. Next, the sequences were queried against GenBank nucleotide and Blastocystis STs (18S) databases., Results: Blastocystis was detected in 14% (14 out of 100) of the faecal samples by culture and 10% (10 out of 100) of the samples by direct microscopy. Nine of Blastocystis isolates (64.4%) were ST3, three (21.4%) were ST1 and two (14.2%) were ST2. Neither the demographic features nor the gastrointestinal symptoms were statistically related to Blastocystis infection., Conclusion: The findings in this study agreed with the most of the previous human studies that found ST3 as the most abundant genotype. This study reported the frequency of Blastocystis in pregnant women and highlighted the importance of comprehensive studies with more cases of Blastocystis during pregnancy.

Blastocystis sp. is a common enteric parasite of humans and animals associated with inadequate sanitation and poor personal hygiene. Over the years, the Malaysian thriving economy has been facilitated largely by migrant workers from developing countries, and there is concern that diseases endemic to their countries may be imported. Therefore, this study aimed to determine the current status of Blastocystis infection as well as subtypes (STs) from fecal samples among migrant workers in Selangor and Kuala Lumpur, Malaysia. Overall, almost a third of the study cohort (30.9%; n = 68/220) screened were infected with Blastocystis sp. predominantly with ST3 (54.5%; n = 12), followed by ST1 (36.4%; n = 8) and ST2 (9.1%; n = 2). Infection levels was almost similar among the different sectors; manufacturing (32.8%), domestic service (32.3%), and food service (27.3%) with common symptoms for infection included stomach and abdominal pain or discomfort and diarrhea (48.5%; n = 33). None of the socio-demographic risk factors evaluated were significant. Therefore, this study warrants continuous monitoring as well as understanding the impact of transmission among the migrant community with the local population especially those involved in food service sector.

Introduction: Blastocystis is a common intestinal parasite of human and animal hosts. The parasite has 17 subtypes, and among those at least nine subtypes (ST1-ST9) are found in human hosts., Objective: The aim of the present study was to investigate the presence of different subtypes of Blastocystis spp. among the patients referred to Velayat hospital of Qazvin province, Iran., Methods: Overall, 864 stool samples were examined by using formalin-ethyl acetate concentration method and Trichrome staining. All specimens were cultured in clotted fetal bovine medium. Later, DNA extraction and PCR amplification of 18S ribosomal RNA gene region was conducted and phylogenetic tree constructed., Results: The results revealed 7.9% (68/864) of the study population were infected with Blastocystis . Intestinal symptoms were observed in 61% (36/59) of individuals positive for Blastocystis , with abdominal pain in 58% (21/36) of cases which was more frequent than other intestinal signs. No significant relationship was observed among the study variables. By molecular and phylogenetic analysis, three subtypes ST1 (45%), ST2 (30%) and ST3 (23%) of parasite were identified., Conclusion: This study showed ST1 subtype was the predominant subtype among the positive specimens, meanwhile the highest haplotype and nucleotide diversity were clarified in ST3 subtype., (© 2020 Delshad A et al.)

Blastocystis has recently been recognized as the most common eukaryotic microbe of the human gut. We investigated the prevalence of Blastocystis and their subtypes in diarrheal and non-diarrheal groups and the associated clinical parameters. A total of 324 stool samples were obtained from 196 diarrheal and 128 non-diarrheal subjects. Blastocystis subtypes were determined by sequencing the small subunit ribosomal DNA (SSU rRNA) gene. Demographic, clinical and laboratory data were collected and analyzed by diarrhea and Blastocystis status. The overall rate of Blastocystis positivity was 9.0% (29/324) but was significantly higher in the non-diarrheal group (18.0% vs. 3.1%, P&lt;0.0001). Of the 6 Blastocystis-positive diarrheal patients, 3 (50.0%), none (0.0%), 2 (33.3%), and 1 (16.7%) were infected with subtypes ST1, ST2, ST3, and multiple subtypes, respectively. Of the 23 Blastocystis-positive non-diarrheal patients, 4 (17.4%), 1 (4.3%), and 18 (78.3%) were infected with subtypes ST1, ST2, and ST3, respectively. Blastocystis was less common in the diarrheal than the non-diarrheal group (odds ratio, 0.144; 95% confidence interval, 0.057-0.365, P&lt;0.001). Of the 3 subtypes, ST3 was more frequently observed in the non-diarrheal than diarrheal group (78.3% vs. 33.3%, P=0.0341). Collectively, Blastocystis was found in both the diarrheal and non-diarrheal groups and ST3 was the most common subtype in Korea.

Blastocystis is a parasite with a worldwide distribution and a varying prevalence in different countries. The pleomorphic nature of the protozoon and the lack of understanding a possible pathogenesis have led to confusion regarding its clinical significance. The aim of the study was to shed light on clinical characteristics of pediatric patients in Swiss children with a positive stool sample for Blastocystis, in order to provide recommendations for a practical approach for the clinician to know whom, when, and how to test. This is a retrospective study of pediatric patients, whose stool has been tested positive for Blastocystis in the last 10 years in northern Switzerland. A total of 4047 stool samples, belonging to 1887 different patients, were analyzed; 240 stool samples (of 160 patients) were tested positive for Blastocystis. On average, 2.2 (CI 1.98-2.35) stool samples per patient were analyzed, of which 1.48 (CI 1.36-1.61) were positive for Blastocystis. In 63% abdominal pain was the leading symptom, while in 17.5% it was an accidental finding without symptoms. There was a high significance in correlation of abdominal pain and chronicity (p < 0.0001) but none in diarrhea (p = 0.082) nor nausea/vomiting or other symptoms and chronicity. Followed by Entamoeba coli (8%), 26.3% of the patients with Blastocystis had a co-infection with another parasite, mostly Endolimax nana (13%).Conclusion: Carriage of Blastocystis is common; therefore, only children/teenagers at risk for a symptomatic Blastocystis infection should be tested. There is a good correlation between Blastocystis and chronic abdominal pain. Children with abdominal symptoms persisting over 4 weeks should have two different stool samples analyzed. No screening after travels/immigration is recommended.What is Known:• Blastocystis has a worldwide distribution.• The clinical significance is unclear.What is New:• Based on retrospective data, we recommend to only test children/teenagers with chronic abdominal pain for Blastocystis.• Two different stool samples should be examined by microscopy; serological investigations are not warranted.

Blastocystis is one of the most common intestinal protozoan parasites worldwide, which is linked to cutaneous lesions and urticaria. In a setting of systematic review, the data on the association of Blastocystis infection with cutaneous lesions were searched in order to summarize the main clinical symptoms, diagnostic methods, treatment, and outcome of the patients. The search identified 28 eligible articles, including 12 cross-sectional studies and 16 case reports/case series (including 23 cases). A diverse spectrum of skin symptoms, mainly urticaria, rash, and itching, was reported from the studies. Of the 23 infected cases with the skin symptoms, gastrointestinal symptoms were reported from the 16 cases, whereas 7 cases with urticaria had asymptomatic infection. The most frequent subtypes were ST1, ST2, and ST3, respectively. Metronidazole, paromomycin, and tinidazole were the most prescribed drugs in patients with single Blastocystis infection. Notably, urticaria and other cutaneous symptoms of all treated patients were resolved after treatment. In conclusion, this study indicates that Blastocystis infection can be a neglected cause of urticaria and skin disorders. Since the treatment of Blastocystis infection is simple, screening and treatment of this infection should be considered in patients with urticaria and other skin disorders.

Blastocystis sp. is a protist commonly found in stool samples of humans and animals. Biological and genetic factors of this organism remain controversial. The present study aimed to develop and implement the Blastocystis in vitro culture of Brazilian human isolates for routine use. The fecal isolates (n = 20) were maintained in our laboratory by several passages in Pavlova's medium. Cultures were monitored every 72 h by light microscopy. Genomic DNA was extracted to identify the subtypes (STs). In most isolates, the vacuolar form was prevalent. The amoeboid, granular and cystic forms were observed during in vitro cultivation. STs 1, 2, 3, 4 and 7 were identified. Our preliminary results show the generation time and forms present in the in vitro culture of Blastocystis subtypes isolated from Brazilian human isolates. Therefore, we emphasize the use of in vitro culture as a tool in future studies for the better understanding of the biological aspects of Blastocystis sp.

Blastocystis species (spp.) is an emerging pathogen. There are several unsolved issues linked to this parasite ranging from its nomenclature, commensal status, standardization of laboratory diagnostic methods, genotypes and treatment. Recently, there has been an increase in reports of Blastocystis spp. from symptomatic cases which provide enough evidence of its pathogenic potential. A range of signs and symptoms, from gastro-intestinal to cutaneous manifestations have been attributed to Blastocystis infection. Few reports have established an association between intestinal infection with Blastocystis spp. and skin manifestations in form of urticaria, palmoplantar pruritus and allergy with complete resolution of cutaneous lesions with eradication of the parasite. In this report, we describe a case of Steven Johnson's syndrome (SJS) in a 6 years old girl along with infection with Blastocystis spp. marked by diarrhea and abdominal pain. Stool examination revealed the presence of all forms of the parasite with subsequent decrease in parasite burden and diarrhea over a period of time. Interestingly, the clearance of Blastocystis spp. from stool was followed by recovery from skin lesions and other symptoms. In this case, the course of SJS was clearly associated with Blastocystis infection. Though skin manifestation with Blastocystis infection has been previously reported, this is the first report of its association with SJS. This report indicates newer insights of the parasite that are less well studied.

Blastocystis is a prevalent parasite that has a wide distribution. In order to design HRM real-time PCR, primers were selected from SSU rRNA gene to amplify specific fragment with different melting temperatures for each subtype of Blastocystis. Subsequently, HRM real-time PCR was performed and melting curve analysis was done by Rotor-Gene Q software. The results of HRM real-time PCR was then compared with sequence results of "barcoding region" of SSU rRNA gene of Blastocystis. To evaluate sensitivity of test, 10-fold serial dilutions of the parasite were prepared from ~ 10 6 to 1 parasite per mL of stool sample and were investigated by HRM real-time PCR. In order to determine specificity of method, HRM real-time PCR was done for some microorganisms and Blastocystis-negative stool samples. In silico analysis showed that all seventeen subtypes of Blastocystis were distinguish. In vitro analysis revealed that the test discriminated subtypes with specific melting temperatures.

Blastocystis is a ubiquitous protozoan with a wide range of hosts. In humans, its presence has been associated with gastrointestinal disorders, although its role as a pathogen still needs to be elucidated. Until now, 17 Blastocystis subtypes (STs) have been identified, with ST1-ST4 the most commonly found in humans. Among domestic animals, the same STs reported in humans have been detected in dogs. An epidemiological survey on dog kennels was carried out to evaluate the prevalence of Blastocystis and the STs involved. Overall, 99 faecal samples were collected from the rescue shelters. Blastocystis detection was performed through conventional barcoding PCR targeting the 1800-bp SSU-rDNA, followed by sequencing and phylogenetic analysis. Blastocystis DNA was found in 21 faecal samples (21.2%), and all samples were successfully sequenced and identified as ST3 in a unique monophyletic group. The presence of Blastocystis was reported for the first time in dogs from Italy, with the identification of ST3, the subtype most commonly found in humans.

Blastocystis spp. is one of the most common protozoa in Turkey and throughout the world; laboratory diagnosis, genetic diversity and clinical features are among the most controversial topics related to the parasite. The aims of the present study were to investigate the subtype distribution of Blastocystis spp. İsolates from Aydin, Turkey, to evaluate the efficiency of some diagnostic methods and to evaluate the relationship between Blastocystis spp. infection with demographic factors and clinical findings. According to the direct microscopy results, 100 stool samples with and without Blastocystis spp. were selected by simple random sampling method. All were directly subjected to DNA isolation and cultured in Jones medium. DNA isolation was also carried out in Blastocystis spp. positive cultures with a different kit. Genomic DNA samples were analysed by PCR targeting the Blastocystis spp. small subunit ribosomal RNA (SSU rRNA) gene and subtypes (ST) were determined according to the sequence analyses. Moreover, the samples with undetected ST were further studied with sequence tagged site-PCR (STS-PCR). In addition, the patients with and without Blastocystis spp. were compared in terms of demographic characteristics (gender, age, residence) and clinical findings (itching, diarrhoea, abdominal pain, dyspepsia, nausea, vomiting, constipation and weight loss)., Among 100 stool positive samples diagnosed with direct microscopic examination 81 (81%) and 86 (86%) were found as positive with culture and PCR, retrospectively. Additionally, among 100 Blastocystis spp. negative stool samples five (5%) and seven (7%) samples were found positive with the same methods, respectively. The results of the analysis of Blastocystis spp. with SSU rRNA gene sequencing and STS-PCR methods revealed the subtype distribution of 95 Blastocystis spp. isolates as follows: ST3 (n= 50, 52.6%), ST2 (n= 21, 22.1%), ST1 (n= 17, 17.9%), ST7 (n= 4, 4.2%), ST2 + ST3 (n= 2, 2.1%) and ST1 + ST3 (n= 1, 1.1%). In addition, a complete accordance was observed in subtype distribution between direct DNA isolation from stools and 35 randomly selected isolates from the culture. In our study, the comparison of 107 Blastocystis spp. positive (by any of the methods) cases and 93 negative cases showed that there was no correlation in terms of demographic characteristics and clinical findings. Similarly, there was no significant relationship between symptoms and subtypes. In conclusion, it is recommended that in addition to direct microscopic examination, the use of additional methods such as culture and PCR will be useful in routine laboratory diagnosis of Blastocystis spp. The distribution of Blastocystis subtype in Aydin is mainly in accordance with the global findings. Lack of a relationship between Blastocystis spp. İnfection and symptoms in our study was supported the idea that Blastocystis spp. infection is mostly asymptomatic in humans and it may be a member of healthy microbiota.

Blastocystis is a common inhabitant of the human gut, colonizing at least one billion people at a prevalence ranging from <10% to 100% in healthy human populations globally. The majority of carriers remain asymptomatic, suggesting that Blastocystis is largely a commensal, though Blastocystis has also been implicated in disease in some people. However, there are no in vivo model systems in which to experimentally test the impact of Blastocystis on mammalian hosts and the gut ecosystem and determine which factors underlie these variable clinical outcomes. We evaluated a rat model for sustaining of a human-derived Blastocystis ST1 and assess colonization success and longevity. Because of the broad host range of Blastocystis, we compared the rat with three other rodent species to establish the reproducibility of our method. Blastocystis was introduced by esophageal gavage and colonization success evaluated by Blastocystis culture. Culture was also used to determine that all animals were negative prior to colonization and negative controls remain Blastocystis-free. In this study, Blastocystis ST1 established in 100% of the outbred rats (Rattus norvegicus) and gerbils (Meriones unguiculatus) challenged. Rats were colonized asymptomatically for more than one year, but Blastocystis ST1 was not transmitted between rats. Mus musculus strain CD1 and Mastomys coucha were not susceptible to Blastocystis ST1. Thus, rats appear to be a suitable in vivo model for studies of Blastocystis ST1, as do gerbils though testing was less extensive. This work lays the foundation for experimental work on the role of Blastocystis in health and disease., (Copyright © 2018 Elsevier Inc. All rights reserved.)

Chronic abdominal pain has many etiologies, one of them being parasites. The aim of this study was to find an association between chronic abdominal pain in children and Blastocystis hominis (Bh). Clinical files of patients with Bh and functional abdominal pain were reviewed. A comparison was made between patients who showed an improvement of their symptoms and those who did not. Out of the 138 patients who had functional abdominal pain and Bh, 37 patients did not receive any treatment (26.8%), while 101 received it and were treated with different antimicrobial agents (73.2%); regarding the improvement of symptoms, a statistically significant difference (p < 0.001) was observed. Chronic abdominal pain in children has different etiologies; however, we have documented through this work that it is appropriate to provide antimicrobial treatment for patients with Bh and chronic abdominal pain.

Blastocystis hominis is an enteric protozoan with many subtypes. It is frequently found in children and may cause chronic diarrhea. This study revealed Blastocystis subtypes among primary school children and comparison of molecular technique and culture method in Blastocystis diagnosis. A total of 141 stools were collected, examined microscopically, selected into the Blastocystis and negative parasite groups, for diagnostic comparison between culture and 18S rRNA polymerase chain reaction (PCR) methods. Positive PCR amplicons were subsequently sequenced for subtyping. The PCR results revealed 89%, 78%, 80% and 88% sensitivity, specificity and positive and negative predictive values, respectively, in comparison with the culture method (McNemar, p > 0.05). Sixteen PCR samples were successfully sequenced and resulted in three Blastocystis subtypes 1, 3 and 4. In conclusion, PCR was sensitive enough and can be used to exclude Blastocystis infection up to 88% of the cases. Subtypes 3 and 1 were the main subtypes found in apparently healthy school children in Jakarta.

Intestinal parasitic infection rate among school-aged children in Thailand has been decreasing. However, certain intestinal parasites remain problematic in some regions. This cross-sectional study was conducted between February and September 2016 in three suburban government primary schools (KK, BR, and HK), Saraburi, Thailand. Stool was collected from 263 asymptomatic subjects (4-15 years old), using simple direct smear, formalin-ether concentration, Boeck and Drbohlav's Locke-Egg-Serum (LES) medium culture, and agar plate culture. A self-administered questionnaire was used to collect data about lifestyle and socioeconomic status. The overall rate of intestinal parasites was 22.1% (15.6% single infection and 6.5% multiple infections). The helminths involving the digestive system found were Strongyloides stercoralis (1.5%) and Opisthorchis viverrini (0.4%). For protozoan infection, the major cause was Blastocystis hominis (17.5%). The other protozoa included Endolimax nana (4.6%), Entamoeba coli (3.4%), Entamoeba histolytica / Entamoeba dispar (1.1%), and Giardia intestinalis (0.8%). The sensitivity for the detection of B. hominis increased with the LES culture technique. The infection rate of each organism was not significantly different among the three schools except for B. hominis which showed the highest prevalence in the HK school ( P = 0.001). This was correlated with the questionnaire results in which the HK school showed the highest risk of drinking contaminated water ( P = 0.004). The present study emphasized the persistent problems of protozoan infections among suburban school-aged children. Lifestyle was still an important factor for intestinal parasitic infections among suburban school-aged Thai children in this study. Health education as well as routine surveillance was necessary to control the infections.

Polymerase chain reaction (PCR) is an effective technique for diagnosis of Blastocystis infection. Notably, DNA isolation procedure is extremely critical for the PCR step. In the present study, a recently described extraction procedure, named as the "sand method" was modified and adapted for isolation of Blastocystis DNA. To evaluate its efficacy, the current method and QIAamp DNA Stool Mini Kit (Qiagen) were applied to fresh human stool samples. Our results indicated that, the mean DNA concentrations obtained by the sand method and the commercial kit were 48 and 55 ng/μl, respectively. Also, no DNA inhibitors were detected in two methods. The sand method was capable of detecting 16 parasites per 50 mg feces. DNA samples extracted by both methods were subjected to PCR. Blastocystis spp. were detected in 11 (31.4%) of 35 samples, and perfect agreement (κ: 1.000) was found between the PCR-sand method and PCR-commercial kit method. The samples that were detected positive by PCR-sand method were successfully sequenced, and Blastocystis subtypes (STs) were identified as ST3, ST2 and ST1. In conclusion, the present study indicates that the sand method provides a simple, rapid and inexpensive procedure for reliable extraction of Blastocystis DNA from stool samples., (Copyright © 2018 Elsevier Inc. All rights reserved.)

The present study was undertaken to investigate the prevalence and staining characteristics of Blastocystis isolated from food animals. Smears of the duodenal and caecal mucosal scrapings, collected from food animals, were stained with Giemsa, Gram's, modified acid-fast and acridine orange. Blastocystis was identified in 295 samples, including faeces and intestinal contents of animals like small ruminants (95), poultry (170) and pigs (30). The prevalence in pigs was found to be high (94.4%) followed by poultry (29.4%) and small ruminants (14%). Various forms of Blastocystis such as vacuolar, granular and amoeboid forms were identified by using different stains. The parasites stained with Giemsa were identified by the presence of eosinophilic nucleus and basophilic cytoplasm. In organisms stained with Gram's stain, the cytoplasm of the vacuolar forms took up the counter stain safranine. Blastocystis appeared as a pink colored cyst against bluish green background with modified acid-fast staining. The study shows that there is a very high prevalence of Blastocystis among the food animals investigated. Simple parasitological procedures, including direct microscopical examination and staining with agents like Giemsa, Gram's and acridine orange can assist identification of the parasites from intestinal contents and faecal material., (Copyright © 2017 Elsevier B.V. All rights reserved.)

The symptomatology of Blastocystis cannot be attributed to any particular subtype, although can be related to a high Blastocystis infection load. One stool sample from each of 217 schoolchildren of Vale de Sol Paranaguá Bay (Paraná, Brazil) was collected. Three milliliters of each fixed stool sample were processed applying the formalin-ether concentration technique. After obtaining the overall prevalence of intestinal parasites, quantification was carried out in Blastocystis positive samples. A total of 75/217 (34.6%) children suffered from intestinal symptoms (abdominal pain and/or persistent diarrhea), of whom 41.3% (31/75) presented moderate/heavy Blastocystis load with a statistically significant risk to present intestinal symptoms (OR 0.039 [0.006-0.15]; p<0.001) Moreover, those symptomatic schoolchildren monoparasitized only by Blastocystis (10/75, 13.3%), and those polyparasitized by Blastocystis with other non-pathogenic species (15/75, 20%) with moderate/heavy loads, also entail a statistically significant risk of intestinal symptomatology, both in monoparasitism (12%, OR 0.10 [0.004-0.63]; p=0.021) and in polyparasitism with a non-pathogenic parasite (18.6%, OR 0.059 [0.002-0.35]; p=0.001). For the first time in Brazil, using data from schoolchildren of Paranaguá Bay, we demonstrated that moderate/ heavy loads of Blastocystis could be related to intestinal symptoms.

AbstractThis study was conducted to investigate the prevalence of Blastocystis spp. and its subtypes (STs) in North Cyprus; and to evaluate the presence of this parasite and its STs with respect to demographic, socioeconomic, and epidemiological factors, as well as gastrointestinal symptoms. Stool samples were collected from 230 volunteers. Each participant also filled out a questionnaire. The samples were examined microscopically by native-Lugol and trichrome methods and further tested by polymerase chain reaction (PCR) and sequencing. Prevalence of Blastocystis spp. infection was found to be 10.5%, 10.5%, and 27.8%, by direct microscopy, trichrome method, and PCR, respectively. No other parasites were detected in the specimens except Giardia spp. ( n = 2; 0.8%) and Entamoeba coli ( n = 1; 0.4%). The most common Blastocystis STs were ST3 (20; 31.2%), ST2 (18; 28.2%), ST1 (8; 12.5%), and ST4 (7; 11%); whereas other STs were identified as ST6 (3; 4.7%), ST7 (2; 3.2%), and non-ST (6; 9.4%). Presence of Blastocystis spp. and its STs was not significantly related to any of the demographic, socioeconomic, and epidemiological factors. Furthermore, no significant association of Blastocystis spp. and its STs with gastrointestinal symptoms was found. This study is the first investigation of the epidemiology of Blastocystis spp. in North Cyprus. Distribution of Blastocystis spp. and its STs among demographic, socioeconomic, and epidemiological factors showed complete homogeneity. Presence of the parasite and its STs was not significantly related with the gastrointestinal symptoms among symptomatic and asymptomatic individuals. These findings suggest that Blastocystis spp. may be part of the intestinal flora in humans.

Blastocystis spp. is the most common enteric protist found in human feces. The pathogenic role of Blastocystis remains controversial and it has been suggested that the symptomatology of Blastocystis is associated with its subtypes (ST). However, only few studies have investigated the relationship between the symptomatology and subtypes of Blastocystis in children. This study aimed to investigate the prevalence of Blastocystis in children aged 3 to 13years with or without gastrointestinal complaints and determine the distribution of the subtypes of Blastocystis. A total of 303 stool samples obtained from symptomatic (n=84) and asymptomatic (n=219) children were included in the study. The presence of Blastocystis was investigated using native-lugol examination, trichrome staining and real-time PCR method. Using the real-time PCR method, 115 samples were found positive for Blastocystis. Subtyping was successfully performed on 46 samples using sequenced-tagged site (STS) primers and PCR. The remaining 69 samples could not be subtyped. The most frequently detected subtype was ST3 (43.4%) followed by ST1 (26.1%), ST4 (10.9%) and ST2 (8.7%). The mixed subtypes were identified in five samples (10.9%) as; ST1+ST3 (n=3), ST1+ST2 (n=1) and ST2+ST3 (n=1). None of the samples had ST5, ST6 or ST7. No statistically significant difference was found between the symptomatic and asymptomatic groups in terms of the Blastocystis positivity and the distribution of subtypes (p>0.05). To our knowledge, this is the first study to investigate the subtype distribution of Blastocystis in children in Turkey and the results are in agreement with the related data available in Turkey., (Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.)

Blastocystis spp. is the most common protozoan detected in human stool samples. In developing countries, infection rates are higher than 20%. The presence of this parasite in the feces of several host species suggests its zoonotic potential. The clinical relevance and the pathogenic role of Blastocystis spp. in the intestinal tract remain unclear. There are several clinical reports that recognize it as the etiologic agent of several intestinal disorders such as diarrhea, inflammatory bowel disease and ulcerative colitis, although the pathogenicity of this parasite has not been proved yet. This wide range of clinical manifestations could be related to the genetic diversity exhibited by this parasite., (Copyright © 2016 Asociación Argentina de Microbiología. Publicado por Elsevier España, S.L.U. All rights reserved.)

Background and Aims: Hepatic encephalopathy (HE) is a serious neuropsychiatric sequela emerging in the advanced stages of cirrhosis. The gut microbiota plays an important role in the development of HE. The aim of the study was to analyze the dynamic interplay between microbiota and Blastocystis in cirrhotic patients with or without encephalopathy., Methods: The study was designed as cross-sectional study. A total of 37 patients from the Ankara city, admitted to the University Hospital within a 6-month period prior to enrolment into the study were included in the study. After the regular health checks, clinical histories, clinical examinations, and Psychometric HE Score (PHES) points, patients' MELD and CTP scores were recorded. The fecal microbiota configurations were characterized by targeting hypervariable regions V3 and V4 of the 16S rRNA gene using Illumina MiSeq System., Results: Blastocystis spp. were detected in 21.6% (n = 8) of all cirrhotic patients. When those were analyzed by subgroups, four of them were subtype 2, three were subtype 3 and one was subtype 1. Blastocystis spp. were not found in any of the patients with HE; however, they were detected in 38.1% of the patients without HE. Also the increase in the bacterial diversity was observed along with the absence of Blastocystis. It was suggested that there was an inverse relationship between Blastocystis spp. and advanced stages of HE and the structure and composition of gut microbiota., Conclusion: The absence of Blastocystis spp. is associated with the HE severity and dysbiosis in the gut microbiota.

Blastocystis hominis provides major challenges for laboratory diagnosis due to its polymorphic nature in wet mounts which can result in confusion with other protozoa, yeast or even fat globules. Studies revealed that simple smears were less sensitive than in vitro cultivation using different media for the detection of B. hominis in stool specimens. Cultures of B. hominis are usually enriched by different types of sera to enhance growth and multiplication of the parasite. The aim of this study is to assess the use of two sera types other than horse serum that is commonly used in culture media for the growth, multiplication and detection of B. hominis in examined stool samples and comparing the results with those obtained using horse serum. Fifty stool samples were collected from patients suffering from different colonic manifestations attending Cairo University Hospitals. The samples were freshly cultured in three different culture media using horse serum (in Jones' medium), donkey serum (as a modification ii Jones' medium) and human plasma (in modified Pavlova's medium) in adequate preparations. Cultures were then left for incubation and examined by direct microscopy to detect Blastocystis hominis. The result showed of 50 stool samples studied,. 18 samples (36%) were positive results for B. hominis. The number of positive results obtained by horse serum, donkey serum and human plasma were 13,18and 11 respectively. Paired comparisons were made between each 2 cultures with each culture set as a reference once to detect the most appropriate one for diagnosis When horse was set as the reference method, donkey serum showed a sensitivity of 100% and specificity of 86.5% with a 90% agreement between the 2 methods. While human plasma showed a sensitivity of 46.2% and specificity of 86.5% with an agreement of 76%. In addition, the vacuolar form was the commonest pattern observed in this study throughout all the three cultures.

Despite Blastocystis being one of the most widespread and prevalent intestinal eukaryotes, its role in health and disease remains elusive. DNA-based detection methods have led to a recognition that the organism is much more common than previously thought, at least in some geographic regions and some groups of individuals. Molecular methods have also enabled us to start categorizing the vast genetic heterogeneity that exists among Blastocystis isolates, wherein the key to potential differences in the clinical outcome of Blastocystis carriage may lie. In this review we summarize some of the recent developments and advances in Blastocystis research, including updates on diagnostic methods, molecular epidemiology, genetic diversity, host specificity, clinical significance, taxonomy, and genomics. As we are now in the microbiome era, we also review some of the steps taken towards understanding the place of Blastocystis in the intestinal microbiota., (Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.)

PCR and DNA sequencing are currently the diagnostic methods of choice for detection of Blastocystis spp. and their suptypes. Fresh or frozen stool samples have disadvantages in terms of several aspects such as transportation, storage, and existence of PCR inhibitors. Filter paper technology may provide a solution to these issues. The aim of the present study was to detect Blastocystis spp. and their subtypes by employing two different preservation methods: conventional frozen stool (FS) and dried stool spots on filter paper (DSSFP). Concentration and purity of DNA, sensitivity of PCR, and DNA sequencing results obtained from the two methods were also compared. A total of 230 fecal samples were included and separated into two parts: one part of the fecal samples were directly frozen and stored at -20 °C. The remaining portion of the specimens were homogenized with saline and spread onto the filter papers as thin layer with a diameter of approximately 3 cm. After air-dried, the filter papers were stored at room temperature. DSSFP samples were collected by scraping from the filter papers. DNA were extracted by EURx Stool DNA Extraction Kit from both samples. Concentration and purity were measured with Nano-Drop, then PCR and sequencing were conducted for detection of Blastocystis spp. and its genotypes. Pure DNA was obtained with a A260/A280 ratio of 1.7-2.2 in both methods. DNA yield from FS was 25-405 ng/μl and average DNA concentration was 151 ng/μl, while these were 7-339 and 122 ng/μl for DSSFP, respectively. No PCR inhibition was observed in two methods. DNA from DSSFP were found to be stable and PCR were reproducible for at least 1 year. FS-PCR- and DSSFP-PCR-positive samples were 49 (21.3 %) and 58 (25.3 %), respectively (p = 0.078). The 43 specimens were concordantly positive by both FS-PCR and DSSFP-PCR. When the microscopy was taken as the gold standard, sensitivity of DSSFP-PCR and FS-PCR was 95.5 and 86.4 %, while specificity of both tests was 99.4 and 98.3 %, respectively. DNA sequencing results of 19 microscopically confirmed cases were strictly identical (concordance 100 %) in both methods, and ST2:6, ST3:8, ST4:3, and ST6:2 were the detected subtypes. Among the 230 fecal samples, the most predominant subtypes were ST3, ST2, ST4, and ST1 by both FS and DSSFP methods. Concordance of DNA sequencing results obtained from the two methods was noted to be 90.7 %. To our knowledge, this is the first study that demonstrates DNA extraction from DSSFP is more sensitive and effective than the FS method for diagnosis of Blastocystis spp. and their subtypes by PCR and DNA sequencing.

Background: Blastocystis spp. are among the most frequently observed intestinal parasites in humans. Despite the discovery of Blastocystis approximately 100 years ago, limited information is available regarding its pathogenesis, genetic diversity, and available treatment options. The aim of this study was to describe the epidemiological and clinical characteristics of patients with Blastocystis sp. infections diagnosed at Vall d'Hebron University Hospital (Barcelona, Spain)., Methods: A retrospective observational study was performed which included all adult patients who attended Vall d'Hebron University Hospital from February 2009 to March 2014 that had Blastocystis sp. detected in their stool., Results: Four hundred eighteen patients were included, the median age was 36 (18-86) years and 236 (56.5 %) were men. Regarding patient symptoms, 234 (56 %) patients were completely asymptomatic, 92 (22 %) patients had symptoms, and 92 (22 %) patients had symptoms that could be attributed to other causes. Of the 92 patients with symptoms not attributable to other etiologies except for Blastocystis infection, the most frequent symptoms were diarrhea (61 patients, 66.3 %) and abdominal pain (34 patients, 37 %). Additionally, nine (9.8 %) patients had cutaneous manifestations. Thirty-one (7.4 %) patients received specific treatment for Blastocystis infection. The clinical response of treated patients was varied. Five patients experienced complete resolution of symptoms, 12 patients reported improvement of clinical symptoms, eight patients described no clinical improvement, and information was unavailable for six patients., Conclusions: Blastocystis infection was detected in 418 patients, most of them foreign-born. Although the vast majority of patients were asymptomatic, 22 % of patients had gastrointestinal symptoms or cutaneous manifestations in the absence of other causes. Despite the scarce information available, given the safety of antiparasitic treatment, and the percentage of patients who experienced resolution or improvement of symptoms, treatment should be considered in patients with chronic symptoms.