On the clinical importance of Dientamoeba fragilis infections in childhood

Clinical and laboratory findings among 123 paediatric patients infected by intestinal protozoa were analysed. Dientamoeba fragilis (D. f) was found in 102 cases. The other patients proved to be carriers of Giardia lamblia or of mixed infections with several protozoa. Acute and recurrent diarrhoea have been found to be the most common symptoms, whereas abdominal pain was most common in children with chronic infections. Peripheral blood eosinophilia was seen in a third of the children with dientamoebiasis. Metronidazole, oxytetracycline, doxycycline, and erythromycin were effective drugs in the treatment of D. f. infections. The therapy coincidentally led to the elimination of protozoal infections as well as the abdominal complaints. These results underline the pathogenic role of D. f. in children with gastrointestinal symptoms.     

Internal transcribed spacer dimorphism and diversity in Dientamoeba fragilis

The internal transcribed spacer (ITS) region of the ribosomal RNA operon is frequently used for detecting sequence variation among closely related species as it is usually homogeneous within strains but evolves more rapidly than ribosomal RNA coding regions. We have studied this region in both genotypes of the human intestinal parasite Dientamoeba fragilis. In contrast to most organisms, we have identified extensive variation between copies of the sequence within the same strain. The ITS occurs in 2 major forms in each genotype but additional heterogeneity is also present within each form. The significance of this finding is unclear, but the only precedent for such variation is in the Apicomplexa, which have multiple dispersed ribosomal RNA operons in contrast to the tandem arrays found in most other eukaryotes.     

Detection of Dientamoeba fragilis in fresh stool specimens using PCR

Dientamoeba fragilis is a trichomonad parasite that causes human gastrointestinal disease. Currently microscopy is considered to be the gold standard for diagnosis of D. fragilis infection. However, this method is time-consuming and relatively insensitive. A PCR assay based on the small-subunit ribosomal RNA gene of D. fragilis for the specific detection of D. fragilis DNA in fresh unpreserved stool samples was developed. The D. fragilis PCR was positive in 29/31 samples with positive microscopy and did not cross-react with other protozoan parasites. The PCR protocol showed a specificity of 100% and a sensitivity of 93.5% and the entire procedure can be performed in one day.     

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.     

Transmission of Dientamoeba fragilis: evaluation of the role of Enterobius vermicularis

The role of Enterobius vermicularis in the transmission of Dientamoeba fragilis has been evaluated in two groups of patients admitted to the Parasitology Laboratory of Celal Bayar University: one group with E. vermicularis infection (n=187, Pinworm Group), and the other with D. fragilis infection (n=126, Dientamoeba Group). The presence of the other parasite, pinworm or Dientamoeba, was investigated with the microscopic examination of cellophane tape and stool samples for three consecutive days. In the Pinworm Group, 9.6% of the patients were found to be coinfected with D. fragilis, while 25.4% of the patients in the Dientamoeba Group were found to be coinfected with pinworms. The coincidence rates of D. fragilis and E. vermicularis, higher than the prevalence of each parasite in similar populations, suggest a common relation between these two parasites, possibly in entering the human body. E. vermicularis infection was found to be significantly more common in younger children (p<0.001), indicating that younger children may also be at higher risk for D. fragilis infection. These findings also raise the question of whether the unrelated symptoms of the pinworm infected patients such as abdominal pain and diarrhea may actually be due to overlooked Dientamoeba infections.     

Dientamoeba fragilis and dientamoebiasis: aspects of clinical parasitology and laboratory diagnosis

The flagellate Dientamoeba fragilis is a potential intestinal pathogen of humans. It is probably undervalued overall because its identification relies on permanent stains directly on fresh fecal specimens (Giemsa) or preserved in PVA or SAF (Trichrome or Iron-hematoxylin). During 1999 we processed stoll samples of 151 subjects and we searched also D. fragilis. We processed 105 cases of enteritis (82 children and 23 adults) and 46 samples of formed stools to investigate only for intestinal parasites (12 children and 34 adults). The microscopical observations were made on smears directly in saline solution, and after Dobell solution, MIF and Giemsa stained. Iron hematoxylin and trichrome stains were performed in some circumstances only. D. fragilis was observed in 11.3% of cases, 7.6% among the group of enteritis and 13.8% among the other subjects. These positive 17 cases were observed more often among adults (76.5% of cases) than among children (23.5%). In our study area D. fragilis seems more frequent than Giardia intestinalis and represents the third important intestinal parasite, following Enterobius vermicularis (children) and Strongyloides stercoralis (elders).     

The ambiguous life of Dientamoeba fragilis: the need to investigate current hypotheses on transmission

Dientamoeba fragilis is an inhabitant of the human bowel and is associated with gastrointestinal illness. Despite its discovery over a century ago, the details of Dientamoeba's life cycle are unclear and its mode of transmission is unknown. Several theories exist which attempt to explain how Dientamoeba may be transmitted. One theory suggests that animals are responsible for the transmission of Dientamoeba. However, reports of Dientamoeba in animals are sporadic and most are not supported by molecular evidence. Another theory suggests that Dientamoeba may be transmitted via the ova of a helminth. Given that the closest relative of Dientamoeba is transmitted via the ova of a helminth, this theory seems plausible. It has also been suggested that Dientamoeba could be transmitted directly between humans. This theory also seems plausible given that other relatives of Dientamoeba are transmitted in this way. Despite numerous investigations, Dientamoeba's mode of transmission remains unknown. This review discusses the strengths and weaknesses of theories relating to Dientamoeba's mode of transmission and, by doing so, indicates where gaps in current knowledge exist. Where information is lacking, suggestions are made as to how future research could improve our knowledge on the life cycle of Dientamoeba.     

子宫内膜异位细胞改良体外培养技术的研究进展

体外细胞培养是子宫内膜异位性疾病研究的重要工具.本文回顾了细胞体外培养技术在分离、纯化环节的改良以及子宫内膜异位性疾病永生化细胞系改造中的研究进展,并总结了近几年来激素及细胞因子诱导培养、细胞共培养及三维培养技术在该疾病体外机制研究中的应用,并对其发展前景进行了展望.     

Dientamoeba fragilis is more prevalent than Giardia duodenalis in children and adults attending a day care centre in Central Italy

Giardia duodenalis is a well recognised enteropathogen, while Dientamoeba fragilis is rarely detected and consequently it is not recognised as an important human pathogen. In 2002-2003, a survey has been carried out on enteroparasites in faecal samples of outpatients attending a day care centre in the town of Perugia (Central Italy). To improve the detection level, at least three samples from each patient were collected at different days and within two hours from defecation. The coproparasitological examination has been carried out by direct microscopic examination, faecal concentration, and Giemsa and modified Ziehl-Nielsen stainings of faecal smears. The genotypes of Giardia duodenalis isolates were determined by PCR of the beta-giardin gene. Of 1,989 enrolled people (966 children, 1,023 adults), 165 persons (8.3%; 153 adults, 15.0%; 12 children, 1.2%), were positive for parasites, but only 1 12 adults (73.2% of those infected) and eight children (66.7% of those infected) harboured D. fragilis and G. duodenalis. Both the Assemblages A and B were detected in 18 G. duodenalis isolates examined at the beta-giardin gene. The higher prevalence of D. fragilis infections than that of G. duodenalis is probably related to the method used, a procedure, which is rarely followed in laboratories for the diagnosis of enteric parasites. These epidemiological data suggest that when faecal samples are examined after a period of time and without Giemsa staining, most D. fragilis infections goes undetected.     

Characterization of the Bacteroides fragilis pathogenicity island in human blood culture isolates

Bacteroides fragilis is an important anaerobic pathogen accounting for up to 10% of bacteremias in adult patients. Enterotoxin producing B. fragilis (ETBF) strains have been identified as enteric pathogens of children and adults. In order to further characterize the B. fragilis pathogenicity island (BfPAI) and using PCR assays for bft- and mpII-metalloprotease genes, we determined the frequency of B. fragilis strains with pattern I (containing the BfPAI and its flanking region), pattern II (lacking both the BfPAI and the flanking region), and pattern III (lacking the BfPAI but containing the flanking region) in 63 blood culture isolates. The results were compared to 197 B. fragilis isolates from different clinical sources. We found 19% of blood culture isolates were pattern I (ETBF), 43% were pattern II (NTBF) and 38% were pattern III (NTBF). Comparatively, B. fragilis isolates from other clinical sources were 10% pattern I, 47% pattern II and 43% pattern III. This suggests that the pathogenicity island and the flanking elements may be general virulence factors of B. fragilis.     

Optimal in-vitro expansion of chondroprogenitor cells in monolayer culture

A continuous production of large quantities of chondroprogenitor cells for the manufacture of engineered cartilage tissue products is required. Expansion of the cell population in vitro has become an essential step in the process of tissue engineering of articular cartilage and the optimization of the culture conditions is a fundamental problem that needs to be addressed. The analysis of both seeding density and passage length was considered crucial in the optimization of expansion processes, and their correct selection should be taken as a requisite to establish culture conditions for monolayer systems. The determination of the optimal seeding density and the corresponding passage length for cell expansion in a serial passaging operation was found to be a compromise between growth kinetics and process time. This optimal determination was carried out using a mathematical approach that led to values of 10(4) cell/cm(2) for seeding density and 73 h for passage length. Additional considerations concerning the running cost of the process were introduced. Although the optimal passage length gave the desired expansion factor in a minimum process time, the selection of an alternative value of 120 h was shown to reduce the cost of the expansion process in more than 60%. The optimization approach presented will contribute to the development of feasible large scale expansion operations of chondroprogenitor cells required by the cartilage tissue engineering industry.     

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.     

Epidemiology of Dientamoeba fragilis Jepps and Dobell, 1918. 3. Further studies on Enterobius transmission through eggs]

In two new attempts to infect human subjects with pin-worms that were infested, at that time, with protozoa Dientamoeba fragilis, presence of these protoza in the intestinal canal was observed in one case for more than 84 days, and in another case for more than 31 days after contamination. The duration of latent periods fixed for Enterobius vermicularis was almost identical (51 and 50 days). New attempts implement and confirm the results of the first successful attempts to transfer the infection (Ockert, 1972b) as well as the hypothesis of the epidemiological interrelation between both parasites (Burrows and Swerdlow 1956).