Observations on the transmission of Dientamoeba fragilis and the cyst life cycle stage.

Little is known about the life cycle and mode of transmission of Dientamoeba fragilis. Recently it was suggested that fecal­oral transmission of cysts may play a role in the transmission of D. fragilis. In order to establish an infection, D. fragilis is required to remain viable when exposed to the pH of the stomach. In this study, we investigated the ability of cultured trophozoites to withstand the extremes of pH. We provide evidence that trophozoites of D. fragilis are vulnerable to highly acidic conditions. We also investigated further the ultrastructure of D. fragilis cysts obtained from mice and rats by transmission electron microscopy. These studies of cysts showed a clear cyst wall surrounding an encysted parasite. The cyst wall was double layered with an outer fibrillar layer and an inner layer enclosing the parasite. Hydrogenosomes, endoplasmic reticulum and nuclei were present in the cysts. Pelta-axostyle structures, costa and axonemes were identifiable and internal flagellar axonemes were present. This study therefore provides additional novel details and knowledge of the ultrastructure of the cyst stage of D. fragilis.

Detection and genotyping of Dientamoeba fragilis from symptomatic patients: New insights from Italy into a little-known gastrointestinal protozoan.

Dientamoeba fragilis (D. fragilis) represents a common protozoan in both high and low income countries. Despite this, epidemiological data on dientamoebiasis are still limited, and it is possible that the actual prevalence rates of D. fragilis have been underestimated due to the challenges in its detection and identification. In the present study, symptomatic patients from Rome (Central Italy) were surveyed for two years to determine D. fragilis percentage of infection and genotypes. Stool samples collection was performed over 864 patients, DNA extracted, and RT-PCR performed by the SeeGene Allplex™ Gastrointestinal Parasite Panel Assays. Seventy-nine resulted positive for D. fragilis (9.1%). Co-infections were detected in 22 isolates: 21 displayed Blastocystis sp. + D. fragilis (27.8%). Based on the sequence of a central fragment of the SSU rRNA gene, only genotype 1 was identified. These findings are among the few available data regarding genetic diversity of D. fragilis in Italy. Large-scale human and animal research are required to enhance our knowledge of prevalence, host range, genetic variability and zoonotic transmission of this little-known intestinal protozoan.

A Cross-Sectional Study on the Occurrence of the Intestinal Protist, Dientamoeba fragilis, in the Gut-Healthy Volunteers and Their Animals.

Dientamoeba fragilis is a cosmopolitan intestinal protist colonizing the human gut with varying prevalence depending on the cohort studied and the diagnostic methods used. Its role in human health remains unclear mainly due to the very sporadic number of cross-sectional studies in gut-healthy populations. The main objective of this study was to expand knowledge of the epidemiology of D. fragilis in gut-healthy humans and their animals. A total of 296 stool samples from humans and 135 samples from 18 animal species were analyzed. Using qPCR, a prevalence of 24% was found in humans in contrast to conventional PCR (7%). In humans, several factors were found to influence the prevalence of D. fragilis. A more frequent occurrence of D. fragilis was associated with living in a village, traveling outside Europe and contact with farm animals. In addition, co-infection with Blastocystis spp. was observed in nearly half of the colonized humans. In animals, D. fragilis was detected in 13% of samples from eight species using qPCR. Our molecular phylogenies demonstrate a more frequent occurrence of Genotype 1 in gut-healthy humans and also revealed a likely a new protist species/lineage in rabbits related to D. fragilis and other related organisms.

Should Dientamoeba fragillis be looked for in pediatric digestive pathology of an unknown cause? A proposed pilot case-control study.

INTRODUCTION: Dientamoeba (D.) fragilis is a common intestinal protozoan with an unresolved clinical significance. The association between D. fragilis and the etiology of gastrointestinal symptoms in children is unclear. Metronidazole is often used for treatment. The aims of this study are to clarify the clinical relevance of D. fragilis in children with gastrointestinal symptoms, and to determine the clinical and microbiological efficacy of metronidazole in D. fragilis-infected children with gastrointestinal complaints. METHODS: A prospective case-control study was performed from October 2017 to February 2019. A total of 106 individuals aged 1-17 were included. Out of the 106; 59 showed gastrointestinal symptoms (case group), and 47 were without gastrointestinal symptoms (control group). We excluded 2 patients from the case group. D. fragilis was diagnosed by real-time PCR in stool samples. A 10-day course of oral Metronidazole was prescribed in D. fragilis positive children with GI symptoms. Clinical data before and after the treatment as well as peripheral eosinophilia in previous blood samples, were recorded. RESULTS: Of the 104 participants, D. fragilis was found in 17 (29.8%) children from the case group, whereas in the control group the parasite was detected in 11 patients (23.4%) with an odds ratio (OR) of 1.39 (IC 95% 0.53-3.75, p=0.46). The most prevalent clinical manifestation was abdominal pain (46/57, 80.7%). Seventeen cases with a positive PCR received anti-parasitic treatment according to the established protocol, although during the collection period we received only 11 stool samples to perform the post-treatment follow-up. The PCR of the D. fragilis remained positive in 3 patients (3/11, 27.27%). Despite achieving the eradication of the parasite, 4/8 patients (50%) continued with digestive symptoms. CONCLUSIONS: According to our study there were no differences between the D. fragilis infection in children with or without gastrointestinal symptoms. No relation was found between the clinical and microbiological responses after said D. fragilis treatment. Therefore, we conclude that it is not justified to look specifically for D fragilis in pediatric patients with abdominal symptoms.

Should Dientamoeba fragillis be looked for in pediatric digestive pathology of an unknown cause? A proposed pilot case–control study / Se debe investigar la infección por Dientamoeba fragilis en niños con enfermedad digestiva de causa desconocida? Estudio piloto caso-control

Introduction: Dientamoeba (D.) fragilis is a common intestinal protozoan with an unresolved clinical significance. The association between D. fragilis and the etiology of gastrointestinal symptoms in children is unclear. Metronidazole is often used for treatment. The aims of this study are to clarify the clinical relevance of D. fragilis in children with gastrointestinal symptoms, and to determine the clinical and microbiological efficacy of metronidazole in D. fragilis-infected children with gastrointestinal complaints. Methods: A prospective case–control study was performed from October 2017 to February 2019. A total of 106 individuals aged 1–17 were included. Out of the 106; 59 showed gastrointestinal symptoms (case group), and 47 were without gastrointestinal symptoms (control group). We excluded 2 patients from the case group. D. fragilis was diagnosed by real-time PCR in stool samples. A 10-day course of oral Metronidazole was prescribed in D. fragilis positive children with GI symptoms. Clinical data before and after the treatment as well as peripheral eosinophilia in previous blood samples, were recorded. Results: Of the 104 participants, D. fragilis was found in 17 (29.8%) children from the case group, whereas in the control group the parasite was detected in 11 patients (23.4%) with an odds ratio (OR) of 1.39 (IC 95% 0.53–3.75, p=0.46). The most prevalent clinical manifestation was abdominal pain (46/57, 80.7%). Seventeen cases with a positive PCR received anti-parasitic treatment according to the established protocol, although during the collection period we received only 11 stool samples to perform the post-treatment follow-up. The PCR of the D. fragilis remained positive in 3 patients (3/11, 27.27%). Despite achieving the eradication of the parasite, 4/8 patients (50%) continued with digestive symptoms.(AU)

Introducción: Dientamoeba (D.)fragilis es un protozoo intestinal muy común pero con una relevancia clínica incierta. El papel etiopatogénico de la D. fragilis en sintomatología gastro-intestinal no está claramente establecido. El metronidazol es con frecuencia el fármaco de elección. El objectivo de nuestro estudio fue determinar la relación entre D. fragilis y la presencia de clínica digestiva en población pediátrica así como valorar la eficacia del metronidazol en la resolución de los síntomas. Material y métodos: Estudio prospectivo caso-control realizado entre octubre 2017 y febrero 2019. Se incluyeron un total de 106 pacientes entre 1-17 años. 59 pacientes presentaban síntomas digestivos (grupo caso) y 47 pacientes asintomáticos (grupo control). Se excluyeron 2 pacientes del grupo caso. La presencia de D. fragilis se determinó por técnica de PCR real-time(PCR-RT) en muestra de heces. En los pacientes del grupo caso con PCR-RT positiva se realizó tratamiento con metronidazol durante 10 dias. Se recogieron datos clínicos antes y después del tratamiento asi como presencia de eosinofilia periférica. Resultados: De los 104 participantes, se detectó D. fragilis en el 17/59 (29,8%) en el grupo caso y en 11/47 pacientes del grupo control (23,4%) con una odds ratio (OR) de 1.39 (IC95% 0.53-3.75, p=0.46). El síntoma más frecuente fue el dolor abdominal (46/57, 80,7%). Los 17 pacientes del grupo caso recibieron metronidazol según protocolo y se realizó PCR-TR D. fragilis post-tratamiento en 11/17 pacientes. La PCR-RT persistió positiva en 3 pacientes (3/11, 27,27%). De los 8 pacientes que negativizaron PCR-RT, 4 (50%) continuaron con dolor abdominal. Conclusión: Según nuestro estudio no se encontró relación significativa entre la presencia de infección por D.fragillis y la existencia de sintomatología digestiva. Tampoco se observó mejoría clínica significativa en aquellos pacientes infectados que recibieron tratamiento.(AU)

First detection and molecular characterization of Dientamoeba fragilis in cattle.

Dientamoeba fragilis is a flagellated protozoan with amoeba-like morphology that inhabits the human gastrointestinal tract. It is endemic in a vast geography around the world, including developed countries. There are limited studies on non-human hosts of the parasite, and suitable hosts have not been clarified. The parasite has been detected in non-human primates, pigs, cats, dogs and rats. There is no study in the literature investigating and detecting the presence of this parasite in cattle. In this study, stool samples taken from 163 different cattle and calves from 11 different farms between March 2017 and May 2022 were examined for the detection of D. fragilis via PCR. Trichrome staining was performed on all PCR-positive samples. The isolates with the expected amplicon size were sequenced using the 18S ribosomal RNA region, and their genotypes were determined by BLAST analysis. Sequences were analysed with the most similar and reference sequences in the literature, forming a phylogenetic tree. We detected D. fragilis in 31 (19.01%) of the 163 stool samples. D. fragilis cysts/trophozoites were detected by trichrome staining method in six of 31 samples. All PCR products selected for molecular analysis from positive samples had the same nucleotide sequence. As a result of BLAST analysis, all sequences were determined to belong to D. fragilis genotype 1. This study determined for the first time that cattle are suitable hosts for D. fragilis. Furthermore, the parasite subtype we detected belongs to genotype 1, which is the most common type in humans, suggesting that the parasite may have a zoonotic character. Our result is important in terms of the epidemiology of the parasite, as the mode of transmission is controversial, and available data on its suitable hosts are limited.

First report and genotyping of Dientamoeba fragilis in pet budgerigars (Melopsittacus undulatus), with zoonotic importance.

The protozoan Dientamoeba fragilis is one of the most common parasites in the digestive system of humans worldwide. The host range and transmission routes of D. fragilis, including the role of animals, are still ambiguous with few reports from non-human primates, sheep, rodents, pigs, a cat and a dog. In this study, we used microscopic and TaqMan qPCR analyses to investigate D. fragilisin 150 faecal samples from pet budgerigars (Melopsittacus undulatus) in the Central Anatolia Region of Turkey. Dientamoeba fragilis DNA was detected in 32 samples, resulting in a mean prevalence of 21.3%. In microscopic examination, trophozoites/cysts of D. fragilis were detected in 13 of 32 qPCR-positive samples. SSU rRNA sequence analyses of the qPCR-positive isolates identified genotype 1 of D. fragilis as predominant in budgerigars. Phylogenetic analyses of the SSU rRNA gene region clustered D. fragilis genotypes, as well as other trichomonads, in separate monophyletic clusters with bootstrap values ≥79.0. Our study provides the first evidence for the natural host status of pet budgerigars for D. fragilisand contributes to the knowledge of the epidemiology of this parasite. The high prevalence of genotype 1 of D. fragilis suggests that pet budgerigars are suitable reservoirs for zoonotic transmission. Our findings contribute to an increased awareness and knowledge of D. fragilis infections in the context of a one-health approach.

Investigation of Dientamoeba fragilis and Blastocystis in patients from Turkey with ulcerative colitis and irritable bowel syndrome: Any relation with genotypes?

Blastocystis sp. and Dientamoeba fragilis are two most common protists worldwide, whose pathogenic potentials are a matter of debate since their discovery. This study aims to investigate the relationship between the activation of ulcerative colitis (UC) and irritable bowel syndrome (IBS) with these protists. A total of 100 patients (35 IBS, 35 active UC, and 30 remittent UC), diagnosed at Hacettepe University Adult Hospital (Ankara, Turkey), were screened for D. fragilis and Blastocystis sp. with microscopic examination using the methods of wet mount, trichrome staining, conventional PCR, nested PCR, real-time PCR and genotyping. Eight patients (4 IBS, 2 active, and 2 remittent UC patients) were found to be D. fragilis positive. 18S rRNA region of the parasite was amplified in four of the patients, whereas cathepsin L-like cysteine peptidase; clan Sc, family S9, serine peptidase; and clan MH, family M20 metallopeptidase in six different patients. All isolates were Genotype 1. Sequence results showed very limited diversity. A total of nine patients (3 IBS, 5 active UC, 1 remittent UC) were found to be positive for Blastocystis sp., all of which were Subtype 3. One active UC and one IBS patient were found to be positive for both parasites. No statistically significant difference was detected between the patient groups in means of parasite detection. D. fragilis was found to be related to older age (p=0,045). In our study, no significant correlation was identified between D. fragilis and Blastocystis sp., and the activation of UC and IBS. More studies are needed on the host-parasite relationship, including the role of gut microbiota, together with transcriptomic and metabolomic assessments to unveil the pathogenicity of both protists.

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

ABSTRACT: 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.

Investigation of Dientamoeba fragilis Frequency in Faecal Samples of Patients with Enterobius vermicularis Infection by Polymerase Chain Reaction

Objective: 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. Methods: A total of 391 samples were obtained. The stool and cellophane slide samples were sent together to the Parasitology Department Laboratory, Faculty of Medicine, Aydin 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. Results: 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). Conclusion: 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. Amaç: Dientamoeba fragilis (D. fragilis), amip benzeri morfolojiye sahip, gastrointestinal yerlesimli, kamçili bir protozoondur. Parazitin Enterobius vermicularis (E. vermicularis) yumurtalariyla tasindigi hipotezi kabul görmektedir. Çalismamizda D. fragilis ve E. vermicularis birlikteligini incelemek, bulunan D. fragilis'lerin genotiplerini belirlemek ve D. fragilis tanisinda yaygin olarak kullanilan trikrom boyama ile polimeraz zincir reaksiyon (PZR) yöntemlerini karsilastirmak amaçlanmistir. Yöntemler: Çalismamizda Aydin Adnan Menderes Üniversitesi Tip Fakültesi, Parazitoloji Laboratuvari'na 1 Ekim 2017-1 Ekim 2018 tarihleri arasinda diski ve selofan lam örnegi birlikte gönderilmis toplam 391 olgu örnegi incelenmistir. Selofanli lam örneklerinde E. vermicularis saptanan tüm gönüllü olgularin (74 olgu) diski örnegi ile E. vermicularis negatif 74 olgunun diski örnegi çalisilmistir. Tüm diskilar trikrom boyama ve PZR yöntemleri ile D. fragilis varligi açisindan incelenmistir. Saptanan D. fragilis izolatlarinin 18S ribozomal RNA bölgesi sekanslanmistir. Olgularin demografik özellikleri ve klinigi degerlendirilmistir. Bulgular: Toplam 148 olgunun 42'sinde (%28,37) D. fragilis saptanmistir. D. fragilis pozitif olan 42 olgunun %66,6'sini E. vermicularis pozitif olgular olusturmus ve iki parazitin birlikteligi anlamli bulunmustur (p<0,05). Sekanslanan tüm izolatlar genotip 1 olarak saptanmistir. Klinik bulgular, yasanilan bölge ve cinsiyet ile parazit varligi arasinda anlamli bir iliski saptanamamistir (p>0,05). Sonuç: Arastirmamizda D. fragilis'in siklikla E. vermicularis ile birliktelik gösterdigi ve bu olgularda D. fragilis varligina ayrica dikkat edilmesi gerektigi vurgulanmistir. Yanlis sonuçlari engellemede, yüksek duyarliliga sahip PZR gibi yöntemlerin önemi bir kez daha görülmüstür.

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

In 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.

Impact of Metronidazole Treatment and Dientamoeba Fragilis Colonization on Gut Microbiota Diversity.

OBJECTIVES: The intestinal parasite Dientamoeba fragilis is a common colonizer of children in Denmark. Metronidazole has been used to reduce gastrointestinal symptoms in children colonized with D fragilis. We aimed to identify gut microbiota changes associated with D fragilis carrier status and metronidazole treatment of D fragilis-positive children. METHODS: The fecal microbiota of 275 fecal samples from children treated with metronidazole (n = 48) or placebo (n = 48) were characterized by ribosomal DNA sequencing. Samples collected before (T1), 2 weeks after (T2), and 8 weeks (T5) after treatment were included. Seventy fecal samples from 70 age-matched parasite-negative children served as controls. RESULTS: The abundance of 24 bacterial genera differed significantly according to D fragilis carrier status, with Flavonifractor being remarkably more abundant in children testing negative for D fragilis. Eight bacterial genera changed significantly in abundance in children losing versus keeping D fragilis after metronidazole treatment. Of these, 7 returned to pretreatment (T1) levels at T5. Meanwhile, the abundance of Flavonifractor continued to differ at T5, whereas for Ruminococcus the abundance only remained high in children who were D fragilis-negative at T2 and T5. Increases in Hungatella, Sutterella, and Streptococcus abundances observed at T2 were specific to metronidazole exposure and hence independent of D fragilis colonization. CONCLUSIONS: This study revealed that specific bacterial genera were associated with D fragilis colonization. Metronidazole treatment had a short-term impact on the abundance of some bacterial genera, with most of these reverting to pretreatment levels 8 weeks after completed treatment.

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

BACKGROUND: 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. METHODS: 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. RESULTS: 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%). CONCLUSION: 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.

Dientamoeba fragilis in the North-East of Italy: Prevalence study and treatment.

Dientamoeba fragilis is an intestinal protozoan, an inhabitant of the human gastrointestinal tract, with a worldwide distribution. The reported prevalence of D. fragilis varies worldwide in different populations between 0.3% and 82.9%, and its role as a pathogen is still unclear. The parasite has been identified in the faeces of asymptomatic patients and with different acute and chronic symptoms, like abdominal pain, diarrhoea, flatulence, nausea and vomiting. The aims of this study were to evaluate the prevalence of D. fragilis in the North-East of Italy, and the clinical improvement of symptoms after recommended treatment with paromomycin or metronidazole. Furthermore, a literature review of D. fragilis prevalence studies in Italy was carried out to show the Italian situation. Of 575 enrolled people, 85 (14.8%) were positive for D. fragilis. The most prevalent symptoms were abdominal pain 28.2%, anal itching 27.1%, watery diarrhoea 18.8%, meteorism 16.5% and nausea/lack of appetite 14.1%. The high rate of anal itching was unexpected, because it wasn't a common symptom. 32 patients were co-infected with B. hominis (37.7%) and three with G. lamblia (3.5%). Our study showed paromomycin had a high efficacy for treatment of D. fragilis infections 100.0% (45/45), while caution must be used when using metronidazole 53.3% (24/40). We recommend paromomycin for empirical treatment, given its great effectiveness in our population.

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.

Dientamoeba fragilis Infection in Patients with Digestive and Non-Digestive Symptoms: A Case-Control Study.

In most developing countries, Dientamoeba fragilis infection is an obscure protozoan infection. We aimed to determine a frequency and clinical importance of D. fragilis infection in Taif, Saudi Arabia. A 1-year case control study included patients with gastrointestinal (cases, n=114) or non-gastrointestinal symptoms (controls, n=90). The fecal samples were examined with the classical parasitological methods for intestinal protozoa, and by real time PCR for D. fragilis. The infection by D. fragilis was detected in 5.8% by PCR and in 4.4% patients by microscopy. The infection was identified more in control group (n=9) than in cases (n=3); a sole infection in 11 patients and mixed with Giardia in 1 patient. The other enteric parasites detected were Blastocystis sp. (8.3%), Giardia sp. (5.3%), Cryptosporidium sp. (2.9%), Entamoeba histolytica (1.4%), Entamoeba coli (0.9%) and Hymenolepis nana (0.4%). Our results tend to reinforce the need to increase awareness of D. fragilis infection in Saudi Arabia.

Dientamoeba fragilis in children: a systematic review on diagnostic considerations and efficacy of treatment.

Introduction: The presence of D. fragilis in feces is characterized by an asymptomatic carrier ship to a spectrum of gastrointestinal symptoms. However, a causal relationship remains to be elucidated. In this systematic review, we aimed to evaluate the relationship between the eradication of D. fragilis and symptoms to establish the strength of evidence that D. fragilis in symptomatic children warrants antibiotic treatment.Areas covered: This systematic review covers a challenge in daily clinical practice. Is it necessary to test for D. fragilis in children with gastrointestinal symptoms and does a positive fecal PCR test warrant treatment?Expert opinion: Testing for D. fragilis seems justified in a selection of children with persistent unexplained chronic abdominal pain and diarrhea. Treatment of D. fragilis should be withhold until other causes like celiac disease have been excluded. Both microscopic and Real Time-PCR methods (or a combination of the two) can be used for diagnosis. Paromomycin or clioquinol are antibiotics of choice based on their small spectrum of activity, fewer side effects, and better eradication rates than metronidazole. Future randomized studies, with strict inclusion criteria, appropriate diagnostic testing, and doses of antibiotics based on bodyweight are warranted.

Fecal calprotectin as a factor that supports the pathogenicity of Dientamoeba fragilis.

Calprotectin is a protein that is mostly released from neutrophils, monocytes, macrophages and submucosal epithelial cells. Fecal calprotectin (f-CP) is a marker of intestinal inflammation. There are some discussions about the pathogenicity of D. fragilis in the gastrointestinal tract. In this study, we investigated whether f-CP level is a factor supporting the pathogenicity of D. fragilis. The f-CP levels were evaluated in patients with only D. fragilis positive in comparison with healthy controls. Moreover, the levels of f-CP were investigated in fecal samples of D. fragilis negative patients with gastrointestinal complaints. The fecal samples were collected from three groups. Three groups of fecal samples were examined directly microscopy, trichrome staining, cultivation, enzyme immunoassay (EIA) and real-time PCR assay. In the first group (Group 1, n = 34), patient stool samples with gastrointestinal symptoms (without other pathogens) found only with D. fragilis were included. In the second group (Group 2, n = 31), there were patients' stool samples with gastrointestinal symptoms that D. fragilis was negative (but there may be other pathogenic agents). In the control group (Group 3, n = 23), we used fecal samples collected from healthy volunteers without any infection or gastrointestinal complaints. The collected fecal samples were stored at -20 °C until analysis. Levels of f-CP were determined by using human calprotectin ELISA kits. Total of 88 patients were enrolled in three different groups. We obtained f-CP levels as follows: 33.40 ng/mg protein in the group 1, 15.99 ng/mg protein in the group 2 and 1.54 ng/mg protein in the group 3. Statistically significant difference in f-CP levels of the group 1 and the group 2 were obtained when compared with healthy controls (p < 0.0001). However, the f-CP levels of the group 1 were not significantly different from the group 2 (p > 0.99). In conclusion, increased levels of f-CP are shown as a marker of an inflammatory disease of the lower gastrointestinal tract in infected humans. There is continues controversy about the pathogenicity of D. fragilis in symptomatic and asymptomatic patients. The findings of this study contribute to the ongoing debate about the pathogenicity of D. fragilis. In our study, the potential pathogenicity of D. fragilis is associated with increased f-CP concentrations with parasite detection in the fecal samples and therefore we assume that the parasite is not only a harmless commensal. In summary, higher levels of f-CP found in D. fragilis positive patients suggest the importance of researches that support the pathogenicity of indicated parasite.

Distribution and relevance of Dientamoeba fragilis and Blastocystis species in gastroenteritis: results from a case-control study.

The actual role of Dientamoeba fragilis and Blastocystis in patients with gastrointestinal symptoms is still under debate. A multicenter case-control study was performed in The Netherlands to elucidate the clinical relevance of molecular diagnostics results in gastroenteritis (GE). Samples from this case-control study were used to perform a detailed analysis on the presence of D. fragilis and Blastocystis in relation to gastrointestinal symptoms. In the present study, a real-time PCR for Blastocystis was performed on 1374 case samples and 1026 control samples from the multicenter gastroenteritis case-control study previously tested for D. fragilis. Prevalence of both micro-organisms was highest in children under 20 years of age and lowest in the oldest age group. A significantly lower overall detection of D. fragilis and Blastocystis was found in cases (both 25.8%) as compared to controls (37.6% and 40.0%, respectively). The difference for D. fragilis was statistically significant for subjects above 20 years of age. For Blastocystis, the difference was statistically significant in all age groups, except in children less than 5 years of age. A negative relation between D. fragilis-positive cases and diarrhea was found in this study population. More GE symptoms were reported in cases without D. fragilis or Blastocystis. In the present study, prevalence of both D. fragilis and Blastocystis is lower in cases with gastroenteritic symptoms than in controls. Besides, in cases with D. fragilis or Blastocystis, no association is shown between any of the GE symptoms. Interestingly, this suggests that the presence of these protozoans may be considered characteristic of a healthy intestinal microbiome.

Paromomycin is superior to metronidazole in Dientamoeba fragilis treatment.

Dientamoeba fragilis is a trichomonad parasite of the human intestine that is found worldwide. However, the biological cycle and transmission of this parasite have yet to be elucidated. Although its pathogenic capacity has been questioned, there is increasing evidence that clinical manifestations vary greatly. Different therapeutic options with antiparasitic drugs are currently available; however, very few studies have compared the effectiveness of these drugs. In the present longitudinal study, we evaluate 13,983 copro-parasitological studies using light microscopy of stools, during 2013-2015, in Terrassa, Barcelona (Spain). A total of 1150 (8.2%) presented D. fragilis. Of these, 739 episodes were finally analyzed: those that involved a follow-up parasitology test up to 3 months later, corresponding to 586 patients with gastrointestinal symptoms (53% under 15 years of age). Coinfection by Blastocystis hominis was present in 33.6% of the subjects. Our aim was to compare therapeutic responses to different antiparasitic drugs and the factors associated with the persistence of D. fragilis post-treatment. Gender, age, and other intestinal parasitic coinfections were not associated with parasite persistence following treatment. Metronidazole was the therapeutic option in most cases, followed by paromomycin: 65.4% and 17.5% respectively. Paromomycin was found to be more effective at eradicating parasitic infection than metronidazole (81.8% vs. 65.4%; p = 0.007), except in children under six years of age (p = 0.538). Although Dientamoeba fragilis mainly produces mild clinical manifestations, the high burden of infection means we require better understanding of its epidemiological cycle and pathogenicity, as well as adequate therapeutic guidelines in order to adapt medical care and policies to respond to this health problem.