Dientamoeba Research Articles

722. Parasitic and Non-Parasitic Causes of Eosinophilia in Children Presenting to a Tertiary Care Center in the United States

Abstract Background Peripheral eosinophilia can be caused by many underlying conditions, including infectious pathogens, allergic disorders, neoplastic disorders, and immunological disorders. However, uncovering the cause of eosinophilia in children can be challenging due to the wide differential diagnosis. Methods To identify causes and risk factors of eosinophilia in children, we performed a retrospective chart review of children presenting to Texas Children’s Hospital in Houston, TX with peripheral eosinophilia from January 1, 2011 to December 31, 2019. Eosinophilia was classified as mild (absolute eosinophilia count or AEC >500 and < 1500), moderate (AEC >1500 and < 5000) and severe (AEC > 5000). Demographic information in addition to travel history, co-morbid conditions, and diagnostic workup were collected. Results A total of 773 patients under 18 years of age were evaluated. The most common cause of eosinophilia in children was atopy (N=343, 44%), followed by eosinophilia of unknown etiology (N=227, 29%). Infectious pathogens were the etiology in 8% of cases of which all were parasitic pathogens. Helminthic (n=48, 79% of parasitic diseases in total) pathogens were more common than protozoan (n=13, 21% of parasitic diseases in total) and patients with parasitic infections had higher median eosinophilia compared to other subgroups (median = 1661, range 512-17,501) with the majority having moderate to severe eosinophilia (57%). The most common helminth was Toxocara (n=31), followed by Strongyloides (n=15), and, more rarely, pinworms (n=4). The most common protozoa identified was Dientamoeba fragilis (n=9), followed by Giardia species (n=5), Endolimax nana (n=4), Blastocystis species (n=3), and Iodamoeba butschlii (n=1). Many unknown cases had limited workup that did not include investigations for common parasites. Conclusion While atopy and unknown etiology were the most common diagnoses for children presenting with eosinophilia, parasites were relatively common and should be considered in the differential when investigating etiologies for peripheral eosinophilia. Disclosures Joud Hajjar, MD, MS, Baxalta (Grant/Research Support)Horizon (Advisor or Review Panel member)Pharming (Advisor or Review Panel member)

Presence and significance of intestinal unicellular parasites in a morbidly obese population.

Obesity is a chronic disease whose pathogenesis has been related to changes in the intestinal microbiota. Yet, the role of protozoa and other unicellular eukaryotic parasites in this microenvironment is still largely unknown. Their presence within the gut ecosystem in obese subjects warrants further study, as well as their influence on the host metabolism and comorbidities. Herein, a single center, cross-sectional study of 104 obese individuals was performed to assess the presence of six intestinal unicellular parasites in stool using a commercially available kit, and to evaluate its relationship with the presence of abdominal symptoms, metabolic comorbidities, variations in body composition and nutritional deficiencies. The overall parasitic colonization rate was 51%, with Blastocystis sp., identified as the most frequent (44.2%), followed by Dientamoeba fragilis (11.5%) and Giardia intestinalis (8.7%), and significantly related to the consumption of ecological fruits and vegetables. Contrary to what previous studies pointed out, colonization with parasites species was significantly associated with fewer abdominal symptoms and depositions per day. The presence of parasites did not correlate with any nutritional deficiencies nor differences in body composition, while it did with significant lower HOMA-IR levels and a lower trend towards metabolic syndrome. Obese subjects frequently harbor unicellular enteric parasites, apparently without clinical nor nutritional harm. This evidence suggests that carrying these microorganisms, from an endocrinological perspective, has a beneficial effect, especially on insulin resistance and possibly on the development of related comorbidities.

Investigation of neglected protists Blastocystis sp. and Dientamoeba fragilis in immunocompetent and immunodeficient diarrheal patients using both conventional and molecular methods.

IntroductionThe clinical significance of Blastocystis sp. and Dientamoeba fragilis in patients with gastrointestinal symptoms is a controversial issue. Since the pathogenicity of these protists has not been fully elucidated, testing for these organisms is not routinely pursued by most laboratories and clinicians. Thus, the prevalence of these organisms and the subtypes of Blastocystis sp. in human patients in Turkey are not well characterized. This study aimed to determine the prevalence of Blastocystis sp. and D. fragilis in the diarrheic stool samples of immunodeficient and immunocompetent patients using conventional and molecular methods and to identify Blastocystis sp. subtypes using next generation sequencing.Material and methodsIndividual stool specimens were collected from 245 immunodeficient and 193 immunocompetent diarrheic patients between March 2017 and December 2019 at the Gazi University Training and Research Hospital in Ankara, Turkey. Samples were screened for Blastocystis sp. and D. fragilis by conventional and molecular methods. Molecular detection of both protists was achieved by separate qPCRs targeting a partial fragment of the SSU rRNA gene. Next generation sequencing was used to identify Blastocystis sp. subtypes.ResultsThe prevalence of Blastocystis sp. and D. fragilis was 16.7% and 11.9%, respectively as measured by qPCR. The prevalence of Blastocystis sp. and D. fragilis was lower in immunodeficient patients (12.7% and 10.6%, respectively) compared to immunocompetent patients (21.8% and 13.5%, respectively). Five Blastocystis sp. subtypes were identified and the following subtype distribution was observed: ST3 54.4% (n = 37), ST2 16.2% (n = 11), ST1 4.4% (n = 3), ST6 2.9% (n = 2), ST4 1.5% (n = 1), ST2/ST3 11.8% (n = 8) and ST1/ST3 8.8% (n = 6). There was no statistically significant difference in the distribution of Blastocystis sp. subtypes between immunocompetent and immunodeficient patients.Conclusion and recommendationOur findings demonstrated that Blastocystis sp. and D. fragilis are commonly present in immunocompetent and immunodeficient patients with diarrhea. This study is the first to use next generation sequencing to address the presence of Blastocystis sp. mixed subtypes and intra-subtype variability in clinical samples in Turkey.

Fecal screening of Dientamoeba fragilis: relative efficacy of permanent smear and culture

Abstract Dientamoeba fragilis (D. fragilis) is an enteric trichmonad protozoan parasite that remains obscure and neglected. The aim of this study is to detect D. fragilis as a neglected pathogen in children aged 6-12 years old complaining of gastrointestinal illness by stool culture and light microscopy with comparison between the results of both techniques. A total of 100 fresh stool samples were included in this current study. All specimens were subjected to microscopic examination using iron- hematoxylin stained stool smears and stool culture using a Loeffler’s culture medium. Culture detected 2 positive stool samples (2%) while microscopy detected (1%). Sensitivities of culture and microscopy were 100% and 50% respectively. Specificity of culture and microscopy were 100% and 95% respectively. There is a moderate agreement between culture and microscopy (K = 0.4). In conclusion, culture had a high performance compared to microscopy in the diagnosis of D. fragilis infection. Culture can be applied for routine diagnosis for the detection of D. fragilis in clinical samples.

Prevalence of diarrhoeal pathogens among children under five years of age with and without diarrhoea in Guinea-Bissau.

BackgroundChildhood diarrhoea, a major cause of morbidity and mortality in low-income regions, remains scarcely studied in many countries, such as Guinea-Bissau. Stool sample drying enables later qPCR analyses of pathogens without concern about electricity shortages.MethodsDried stool samples of children under five years treated at the Bandim Health Centre in Bissau, Guinea-Bissau were screened by qPCR for nine enteric bacteria, five viruses, and four parasites. The findings of children having and not having diarrhoea were compared in age groups 0–11 and 12–59 months.ResultsOf the 429 children– 228 with and 201 without diarrhoea– 96.9% and 93.5% had bacterial, 62.7% and 44.3% viral, and 52.6% and 48.3% parasitic pathogen findings, respectively. Enteroaggregarive Escherichia coli (EAEC; 60.5% versus 66.7%), enteropathogenic E. coli (EPEC; 61.4% versus 62.7%), Campylobacter (53.2% versus 51.8%), and enterotoxigenic E. coli (ETEC; 54.4% versus 44.3%) were the most common bacterial pathogens. Diarrhoea was associated with enteroinvasive E. coli (EIEC)/Shigella (63.3%), ETEC (54.4%), astrovirus (75.0%), norovirus GII (72.6%) and Cryptosporidium (71.2%). The only pathogen associated with severe diarrhoea was EIEC/Shigella (p<0.001). EAEC was found more frequent among the infants, and EIEC/Shigella, Giardia duodenalis and Dientamoeba fragilis among the older children.ConclusionsStool pathogens proved common among all the children regardless of them having diarrhoea or not.

Diagnostic and Therapeutic Considerations Towards Dientamoeba fragilis in Children: A Survey Amongst General Practitioners and Pediatricians in the Netherlands.

This survey was undertaken to obtain insight in the attitude of Dutch physicians towards pathogenicity, diagnostic- and therapeutic approach towards Dientamoeba fragilis in children. Physicians were invited by e-mail for a questionnaire. A total of 211 of 450 physicians (46.9%) completed the questionnaire, including 67 general practitioners (GPs) and 144 pediatricians. Of all respondents, 175 of 211 (82.9%) considered D fragilis a "potential pathogen", when other causes of gastro-intestinal complaints are ruled out. Only 16 of 211 (7.6%) performed diagnostic tests regularly. Diagnostic tests were performed by 162 of 211 (77%) of respondents in children with diarrhea and abdominal pain in consideration of duration of symptoms. Fecal polymerase chain reaction (PCR) was diagnostic modality of preference. Eighty-nine of 142 (62.7%) prescribed metronidazole as antibiotic of first choice. This study shows heterogeneity in clinical practice amongst Dutch physicians regarding diagnostic- and therapeutic approach of D fragilis in children. Different attitude towards pathogenicity and inconsistent guidelines could be causative factors.

Improved diagnosis of gastrointestinal infections using a semi-automated multiplex real-time PCR for detection of enteropathogens.

Introduction. The identification of enteropathogens is critical for the clinical management of patients with suspected gastrointestinal infection. The FLOW multiplex PCR system (FMPS) is a semi-automated platform (FLOW System, Roche) for multiplex real-time PCR analysis.Hypothesis/Gap Statement. FMPS has greater sensitivity for the detection of enteric pathogens than standard methods such as culture, biochemical identification, immunochromatography or microscopic examination.Aim.The diagnostic performance of the FMPS was evaluated and compared to that of traditional microbiological procedures.Methodology. A total of 10 659 samples were collected and analysed over a period of 7 years. From 2013 to 2018 (every July to September), samples were processed using standard microbiological culture methods. In 2019, the FMPS was implemented using real-time PCR to detect the following enteropathogens: Shigella spp., Salmonella spp., Campylobacter spp., Giardia intestinalis, Entamoeba histolytica, Blastocystis hominis, Cryptosporidum spp., Dientamoeba fragilis, adenovirus, norovirus and rotavirus. Standard microbiological culture methods (2013-2018) included stool culture, microscopy and immunochromatography.Results. A total of 1078 stool samples were analysed prospectively using the FMPS from July to September (2019): bacterial, parasitic and viral pathogens were identified in 15.3, 9.71 and 5.29 % of cases, respectively. During the same period of 6 years (2013-2018), the proportion of positive identifications using standard microbiological methods from 2013 to 2018 was significantly lower. A major significant recovery improvement was observed for all bacteria species tested: Shigella spp./enteroinvasive Escherichia coli (EIEC) (P <0.05), Salmonella spp. (P <0.05) and Campylobacter spp. (P <0.05). Marked differences were also observed for the parasites G. intestinalis, Cryptosporidium spp. and D. fragilis.Conclusion. These results support the value of multiplex real-time PCR analysis for the detection of enteric pathogens in laboratory diagnosis with outstanding performance in identifying labile micro-organisms. The identification of unsuspected micro-organisms for less specific clinical presentations may also impact on clinical practice and help optimize patient management.

Prevalence of Intestinal Parasitic Infections among Children in Europe over the Last Five Years.

While the prevalence of intestinal parasitic infections (IPI) has been most commonly studied in African and Asian populations, less is known about the prevalence rates of IPI in European children, as well as the potential risk factors that favor the spread of parasites. We aimed to review published evidence on the prevalence rates of IPI in children residing in Europe, and to quantitatively synthesize the results of published studies. We searched Medline from 1 January 2015 to 1 April 2021 to address the most recently published prevalence patterns of IPI in European children. Random-effects meta-analyses were performed by type of IPI infection, age group and sex, depending on data availability. Of the 967 potentially relevant articles, eight eligible cross-sectional studies were included in this analysis, yielding a sample of 3376 children (0–19 years). The overall prevalence rate was 5.9% for any IPI in children residing in European countries. Blastocystis hominis was the most commonly detected parasite yielding a prevalence rate of 10.7%. Other parasites included Entamoeba coli, Endolimax nana, and Blastocystis hominis. Studies focusing on specific types of parasites showed prevalence rates ranging from 1.3% for Cryptosporidium to 68.3% for Dientamoeba fragilis. Despite the scarce literature, the present review showed relatively low prevalence rates of IPI in Europe. Future studies accounting for proper diagnostic methods used for the detection of parasites and including information on potential sociodemographic factors, such as travelling history and history of immigration, are needed to guide clinicians about which children to test, as well as when and how to test children for IPI.

Investigation of &lt;i&gt;Dientamoeba fragilis&lt;/i&gt; Frequency in Faecal Samples of Patients with &lt;i&gt;Enterobius vermicularis&lt;/i&gt; Infection by Polymerase Chain Reaction

Dientamoeba fragilis (D. fragilis) is a flagellated protozoan with an amoeba-like morphology, located in the gastrointestinal tract. The hypothesis was that the parasite was transported by Enterobius vermicularis (E. vermicularis) eggs. This study aimed to determine the association of D. fragilis and E. vermicularis with the genotypes of the identified strain of D. fragilis. Results of trichrome staining were compared with those of polymerase chain reaction (PCR), which is widely used in the diagnosis of D. fragilis. A total of 391 samples were obtained. The stool and cellophane slide samples were sent together to the Parasitology Department Laboratory, Faculty of Medicine, Aydın Adnan Menderes University, between 1 October 2017 and 1 October 2018. Stool samples of all patients with E. vermicularis (n=74) and without E. vermicularis (n=74) infection were used. All samples were examined for the presence of D. fragilis by trichrome staining and PCR. The 18S ribosomal RNA region of D. fragilis isolates was sequenced. Demographic characteristics and clinical findings of the patients were evaluated. D. fragilis was detected in 42 (28.37%) of 148 samples; 28 (66.6%) of them were detected in patients with E. vermicularis infection. The coexistence of two parasites was significant (p<0.05). All isolates sequenced were genotype 1. No significant relationship was found between the presence of parasites and clinical findings, living area and gender (p>0.05). D. fragilis is frequently associated with E. vermicularis, so the presence of D. fragilis should be also considered in affected patients. The use of high-sensitivity molecular methods such as PCR is important in preventing false results. Dientamoeba fragilis (D. fragilis), amip benzeri morfolojiye sahip, gastrointestinal yerleşimli, kamçılı bir protozoondur. Parazitin Enterobius vermicularis (E. vermicularis) yumurtalarıyla taşındığı hipotezi kabul görmektedir. Çalışmamızda D. fragilis ve E. vermicularis birlikteliğini incelemek, bulunan D. fragilis’lerin genotiplerini belirlemek ve D. fragilis tanısında yaygın olarak kullanılan trikrom boyama ile polimeraz zincir reaksiyon (PZR) yöntemlerini karşılaştırmak amaçlanmıştır. Çalışmamızda Aydın Adnan Menderes Üniversitesi Tıp Fakültesi, Parazitoloji Laboratuvarı’na 1 Ekim 2017-1 Ekim 2018 tarihleri arasında dışkı ve selofan lam örneği birlikte gönderilmiş toplam 391 olgu örneği incelenmiştir. Selofanlı lam örneklerinde E. vermicularis saptanan tüm gönüllü olguların (74 olgu) dışkı örneği ile E. vermicularis negatif 74 olgunun dışkı örneği çalışılmıştır. Tüm dışkılar trikrom boyama ve PZR yöntemleri ile D. fragilis varlığı açısından incelenmiştir. Saptanan D. fragilis izolatlarının 18S ribozomal RNA bölgesi sekanslanmıştır. Olguların demografik özellikleri ve kliniği değerlendirilmiştir. Toplam 148 olgunun 42’sinde (%28,37) D. fragilis saptanmıştır. D. fragilis pozitif olan 42 olgunun %66,6’sını E. vermicularis pozitif olgular oluşturmuş ve iki parazitin birlikteliği anlamlı bulunmuştur (p<0,05). Sekanslanan tüm izolatlar genotip 1 olarak saptanmıştır. Klinik bulgular, yaşanılan bölge ve cinsiyet ile parazit varlığı arasında anlamlı bir ilişki saptanamamıştır (p>0,05). Araştırmamızda D. fragilis’in sıklıkla E. vermicularis ile birliktelik gösterdiği ve bu olgularda D. fragilis varlığına ayrıca dikkat edilmesi gerektiği vurgulanmıştır. Yanlış sonuçları engellemede, yüksek duyarlılığa sahip PZR gibi yöntemlerin önemi bir kez daha görülmüştür.

Prevalence of intestinal parasitic infection in food handlers of Iran: A systematic review and meta‐analysis

Food handlers regardless of whether preparing or serving food, play key roles in the transmission of food‐borne infections. This study aimed to evaluate the prevalence of intestinal parasitic infections in food handlers in Iran. In the present study, a comprehensive literature search was carried out in electronic databases, including PubMed, Scopus, Google Scholar, Science Direct, Magiran, Scientific Information Database (SID), Iran Medex and Iran Doc, to identify all the published studies from 2000 to 31st April 2019. A total of 25 articles from different regions of Iran were identified and fulfilled our eligibility criteria. Totally, 140,447 cases were examined and 1163 cases were infected with intestinal parasites. Of all cases, 19,516 were male and 5901 were female with 1163 and 652 infected cases, respectively. The overall prevalence of intestinal parasitic infections was evaluated 14.0% [95% CI: 11.0‐17.0%]. It is revealed that protozoan, such as Giardia lamblia, with prevalence of 41.0% [95% CI: 25.0‐59.0%], Blastosystis hominis with 28.0% [95% CI: 15.0‐44.0%] and Entamoeba coli with 22.0% [95% CI: 16.0‐29.0%] had the highest prevalence while, Dientamoeba fragilis 5.0% [95% CI: 4.0‐7.0%], Iodamoeba bütschlii 5.0% [95% CI: 2.0‐8.0%], Chilomastix mesnili 5.0% [95% CI: 2.0‐9.0%] and Endolimax nana with 3.0% [95% CI: 1.0‐7.0%], were less prevalent. Infection with Ascaris lumbricoides7.0% [95% CI: 0.0‐29.0%] was more prevalent helminth followed with Enterobius vermicularis 3.0% [95% CI: 1.0‐5.0%], Hymenolepis nana 2.0% [95% CI: 1.0‐3.0%], Taenia spp. 2.0% [95% CI: 0.0‐7.0%] and Trichuris trichiura 1.0% [95% CI: 0.0‐1.0%]. The high prevalence of commensal parasites, such as Entamoeba coli, which does not need cure is indicating the importance of personal hygiene in food handlers.Our results revealed the high prevalence of intestinal parasitic infection in food handlers in Iran. Monitoring programs to prevent and controlling of transmission to individuals are needed.

The high co-existence rate of Blastocystis and Dientamoeba fragilis in human faecal samples and the analysis of demographic and clinical findings

Aim: Blastocystis and Dientamoeba fragilis (D. fragilis) are among the most common protozoon species in human faecal samples. The cross-sectional studies have reported the frequencies in a variety of populations. However, we have very limited information about the co-existence rate of those protozoans. The study aimed to compare D. fragilis frequency in Blastocystis positive and negative faecal samples in order to determine the co-existence rate. The secondary objective was to analyse demographic characteristics and gastrointestinal (GI) symptoms in relation to both infections.&#x0D; Material and Method: In the present study, we defined a study group that included 100 Blastocystis positive faecal samples and a control group that included 100 Blastocystis negative samples. The frequency of D. fragilis in samples was determined with a PCR assay specific to the small-subunit ribosomal RNA (SS rRNA) gene. A positive control of D. fragilis was used and the samples with amplification of the expected size (863 bp) were considered as positive. In addition to the statistical comparison of frequencies, the descriptive and clinical findings of cases were analysed retrospectively with Pearson chi-square or ANOVA tests. &#x0D; Results: The frequency of D. fragilis was 21% in Blastocystis positive group and it was 10% in Blastocystis negative group. There was statistically significant difference in terms of D. fragilis positivity between the groups (p 0.05). &#x0D; Conclusion: Our study highlighted the high co-existence of D. fragilis and Blastocystis in human faecal samples. A possible explanation for this finding may be the faecal-oral transmission of these protozoans. In addition, analysis of clinical findings was supported common asymptomatic colonisation of Blastocystis and D. fragilis.

Epidemiology and clinical features of intestinal protozoan infections detected by Real-time PCR in non-native children within an Italian tertiary care children's hospital: A cross-sectional study

BackgroundEnteric parasite infections are underestimated due to the limited sensitivity and specificity of microscopy, which remains the diagnostic gold standard in routine clinical practice. This could be a major problem in high-income countries, where the burden of parasitic diseases is low. In recent years, Multiplex Real-Time polymerase chain reaction (RT-PCR) based methods have been implemented. Therefore, the aim of this study was to evaluate the prevalence of four enteric protozoan species detected by RT-PCR in non-native children in Italy, and to describe their clinical characteristics. MethodsAdopted and immigrant children, evaluated for migration health assessment between 2017 and 2020 in a tertiary care children's hospital in Italy, were enrolled. Molecular analysis for Giardia lamblia, Dientamoeba fragilis, Blastocystis hominis, and Entamoeba histolytica, was conducted by in-house RT-PCR. ResultsOverall, 209 children were enrolled and 70% of them resulted positive by RT-PCR for at least one enteric parasite. B. hominis (47.8%) was the most commonly identified protozoa, followed by D. fragilis (44.5%). Co-infections with multiple pathogens were detected in 35.4% of the samples. Almost 80% of parasite-positive children were asymptomatic and the most common symptom was flatulence (60.7% of symptomatic children). Eosinophils were significantly increased in RT-PCR positive children compared to the negative ones and children with D. fragilis presented the highest eosinophils count. ConclusionsThe In-house Multiplex RT-PCR assay provides a valid molecular detection system for selected enteric parasites. This novel and accurate diagnostic method can help in increasing the detection rate of parasite infection, especially in high-risk population.

Parasitic Intestinal Protists of Zoonotic Relevance Detected in Pigs by Metabarcoding and Real-Time PCR.

Several parasite species are shared between humans and pigs. We explored the application of next-generation sequencing-based metabarcoding supplemented with real-time PCR to fecal DNAs from 259 samples from 116 pigs in Denmark to detect and differentiate single-celled intestinal parasites of zoonotic relevance. Enterocytozoon bieneusi, Balantioides coli, and Giardia duodenalis were observed in 34/37 (92%), 148/259 (57%), and 86/259 (33%) samples, respectively. Entamoeba polecki ST1, E. polecki ST3, and Entamoeba hartmanni were detected in 104/259 (40%), 161/259 (62%), and 8/259 (3%) samples, respectively. Metabarcoding and real-time PCR detected Cryptosporidium in 90/259 (35%) and 239/259 (92%) of the samples, respectively, with Cryptosporidium suis and Cryptosporidium scrofarum observed in nearly equal proportions. Blastocystis subtypes 1, 3, 5, and 15 were found in 72 (28%), 6 (2%), 176 (68%), and 36 (14%) of 259 samples, respectively. Iodamoeba was identified in 1/259 samples (<1%), while none of 37 tested samples was positive for Dientamoeba fragilis. Our results illustrate how metabarcoding exemplifies a ‘one-fits-many’ approach to detecting intestinal single-celled parasites in feces supplemented with real-time PCR for selected parasites. Using metabarcoding with pathogen-specific assays may help detect emerging and previously underdetected pathogens and further elucidate the role of micro-eukaryotic parasites in human and animal health and disease.

The effect of Blastocystis sp. and Dientamoeba fragilis on psychological symptom severity in a sample of clinically diverse males and females

Health outcomes associated with Blastocystis sp. and Dientamoeba fragilis are disparate and controversial, ranging from health benefits, to years of asymptomatic carriage, through to severe illness. Evidence that Blastocystis sp. and D. fragilis are commensal members of the gut microbiota is growing. Despite this, little to no research exists investigating the potential effect of these protozoa on psychological symptom expression. As such, the aim of this retrospective cross-sectional study was to be the first to investigate the effect of protozoan carriage on severity of Depressive, Neurocognitive, Stress and Anxiety, and Sleep and Fatigue symptoms, and whether this effect changes as a function of sex. The prevalence of D. fragilis was significantly higher in females compared to males, however there were no sex differences in prevalence for Blastocystis sp. (data used in the current study contained ST1, ST3, and Blastocystis ST unspecified) or co-carriage of the two. Females reported significantly more severe symptoms across all four psychological domains compared to males. There was no significant interaction between sex and Blastocystis sp. carriage on psychological symptom severity, and no significant main effect of Blastocystis sp. on symptom severity compared to those who tested negative for protozoa. When investigating the sexes separately, there was no effect of protozoan carriage on psychological symptom expression in either males or females. These findings add weight to the argument that Blastocystis sp. and D. fragilis are not necessarily pathogenic and are likely to be part of a diverse gut (which is typically associated with better health outcomes). Further research is required given that protozoan members of the gut microbiota have been largely ignored in brain-gut-microbiota axis research.

One Health Approach to Zoonotic Parasites: Molecular Detection of Intestinal Protozoans in an Urban Population of Norway Rats, Rattus norvegicus, in Barcelona, Spain.

Rattus norvegicus, the brown or Norway rat, is the most abundant mammal after humans in urban areas, where they live in close proximity to people. Among rodent-borne diseases, the reservoir role of Norway rats of zoonotic parasites in cities has practically been ignored. Considering the parasitic diseases in the One Health approach, we intended to identify and quantify the zoonotic intestinal protozoans (ZIP) in an urban population of R. norvegicus in the city of Barcelona, Spain. We studied the presence of ZIP in 100 rats trapped in parks (n = 15) as well as in the city’s sewage system (n = 85) in the winter of 2016/17. The protozoans were molecularly identified by means of a multiplex PCR (AllplexTM Gastrointestinal Panel-Parasite Assay). We also investigated the presence of co-infections among the species found. Four ZIP were identified, presenting significant prevalences in sewers, specifically Blastocystis (83.5%), Giardia duodenalis (37.7%), Cryptosporidium spp. (34.1%), and Dientamoeba fragilis (14.1%). Several co-infections among the detected ZIP were also detected. The reservoir role of ZIP that Norway rats play in cities as well as the role rats may play as sentinels of zoonotic parasites affecting humans in urban areas are strongly backed up by our findings. The increasing worldwide urbanization, climate change, and the COVID-19 pandemic are factors that are producing an increase in human–rat interactions. Our results should be considered a warning to the authorities to intensify rat control and surveillance in public health interventions.

Investigation of Intestinal Protozoon Prevalence in Immunocompromised Patients at a University Hospital

Immunocompromised patients are at a greater risk of developing intestinal parasite infections. In this study, we examined the presence of Enterocytozoon bieneusi, Encaphalitozoon intestinalis and other intestinal protozoa in stool samples of immunosuppressed patients. A total of 100 stool samples were obtained from patients receiving chemotherapy because of solid organ tumour with haematological malignancies and those receiving immunosuppressive treatment because of rheumatic diseases, organ transplant patients and patients receiving treatment for HIV-related infections. Stool samples were examined by using the native-lugol method in which the stool concentration, modified Kinyoun acid-fast and trichrome staining methods and parasite presence were analysed. The stool samples were also examined for the presence of Enterocytozoon bieneusi and Encephalitozoon intestinalis using an indirect fluorescent antibody method. Intestinal parasites were detected in 12% of all patients. The distribution of intestinal parasites in patients were 7% Blastocystis spp., 2% Blastocystis spp. + Dientamoeba fragilis, 1% Blastocystis spp. + Entamoeba coli, 1% Blastocystis spp. + Giardia intestinalis and 1% G. intestinalis. Microsporidia spp. were detected in 4% of all patients by the IFAT method and in 8% of all patients by calcoflour staining method. In our study, the most prevalent parasite detected in the immunosuppressed patients was Blastocystis spp. The pathogenesis of Blastocystis spp. remains to be controversial, and their role in immunocompromised patients continues to remain unknown. Although these rates detected in our study are similar to the prevalence in the normal population, it is important to study these microorganisms in immunocompromised patients in terms of the associated decreasing morbidity and mortality rates.

Molecular detection and genetic diversity of Dientamoeba fragilis and Enterocytozoon bieneusi in fecal samples submitted for routine parasitological examination

Objective: To detect Dientamoeba (D.) fragilis and Enterocytozoon (E.) bieneusi and to assess their genetic characteristics in stool samples submitted for routine examination in a clinical laboratory in Southeastern Brazil. Methods: In this survey, 348 stool samples from female and male individuals with age ranging from 0 to ≥55 years were analyzed by PCR amplifying and sequencing based on the small subunit ribosomal RNA (SSU rRNA) gene of D. fragilis and the internal transcribed spacer of E. bieneusi. Results: D. fragilis and E. bieneusi isolates were observed in 2.29% (8/348) and 4.59% (16/348) of the samples, respectively. These parasites were detected in stool samples from individuals of both genders, including young children under nine until adults over 55 years old. No statistically significant differences were found. All D. fragilis isolates were classified as genotype 1 and E. bieneusi isolates included genotypes D (n=15) and A (n=1). Conclusions: The findings provide relevant findings on occurrence and genetic diversity of D. fragilis and E. bieneusi, pointing to the need for the diagnosis of these parasites in routine examinations in clinical laboratories. In addition to sensitive diagnostic methods, it is mandatory that these parasites be considered relevant for physicians and laboratory staff.

Prevalence and genetic characterization of Dientamoeba fragilis in asymptomatic children attending daycare centers.

ABSTRACTIn order to provide additional data on the prevalence and genetic diversity of Dientamoeba fragilis in human populations, we conducted a study in children from low-income communities in Sao Paulo State, Brazil. Fecal samples from daycare center attendees up to 6 years old (n=156) and staff members (n=18) were submitted to PCR and sequencing of D. fragilis as well as to microscopic examination for the presence of other intestinal parasites. All children assessed were asymptomatic and 10.3% (16/156) were positive for D. fragilis. No worker was found to be positive. An association between Dientamoeba and coinfection with other intestinal parasites was observed. Concerning the genetic diversity, 14 and only two isolates were genotype 1 and genotype 2, respectively. Our findings outline interesting aspects: (1) asymptomatic children as carriers of Dientamoeba in communities in which environmental conditions ensure parasite transmission and, (2) association between Dientamoeba infection in young children and coinfection with other enteric parasites, reinforcing its transmission via the fecal–oral route.

Diagnosis of intestinal protozoan infections in patients in Cuba by microscopy and molecular methods: advantages and disadvantages

Microscopy is the gold standard for diagnosis of intestinal parasitic diseases in many countries, including Cuba, although molecular approaches often have higher sensitivity as well as other advantages.Fecal samples from 133 patients were analyzed by light microscopy and also real-time multiplex qPCR targeting Giardia duodenalis, Cryptosporidium spp., and Entamoeba histolytica, and, separately, Dientamoeba fragilis. Microscopy revealed G. duodenalis occurred most commonly (17 patients), followed by Blastocystis spp. (12 patients). In a few patients, Entamoeba histolytica/E. dispar, Cryptosporidium spp., and Cyclospora cayetanensis were identified.Molecular analysis identified 4 more G. duodenalis infections and 2 more Cryptosporidium spp. infections; concordance between microscopy and PCR showed almost perfect agreement for G. duodenalis (κ = 0.88) and substantial agreement for Cryptosporidium (κ = 0.74). PCR indicated that E. dispar, rather than E. histolytica, had been identified by microscopy. Additionally, 16 D. fragilis infections were detected using molecular methods.Although both microscopy and molecular techniques have a place in parasitology diagnostics, for parasites such as D. fragilis, where microscopy can underestimate occurrence, molecular techniques may be preferable, and also essential for distinguishing between morphologically similar microorganisms such as E. histolytica and E. dispar. Although in resource-constrained countries such as Cuba, microscopy is extremely important as a diagnostic tool for intestinal parasites, inclusion of molecular techniques could be invaluable for selected protozoa.

Intestinal Microbiota in Children With Symptomatic Dientamoeba fragilis Infection: A Case-control Study.

Dientamoeba fragilis in children has been associated with gastrointestinal symptoms, like abdominal pain and diarrhea. The mechanism underlying these symptoms in children with D. fragilis remains unclear. We hypothesized that concomitant microbial alterations, which have been described in other parasitic infections, may be associated with gastrointestinal symptoms in D. fragilis. In this case-control study performed in 2 centers, 19 children referred to a pediatrician because of gastrointestinal symptoms and with a positive fecal PCR for D. fragilis were included as cases. We included 19 healthy children as controls and matched for age and gender, selected from an existing cohort of 63 children. A PCR for D. fragilis was performed on fecal samples of the 19 controls to assess D. fragilis carriership in this asymptomatic group. Microbiota was analyzed with the IS-pro technique, and the intestinal microbiota composition and diversity were compared between the 2 groups. Microbiota of children with D. fragilis and gastrointestinal symptoms did not significantly differ in terms of composition and diversity compared with controls, both on phylum and species level. In the asymptomatic controls, a positive fecal PCR for D. fragilis was found in 16 of 19 (84.2%). Intestinal microbiota does not seem to play a key role in the presence of clinical symptoms in children with D. fragilis. The pathogenicity of D. fragilis and pathophysiologic pathways underlying the development of gastrointestinal symptoms remains yet to be clarified.