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

Authors: SARZHANOV, FAKHRIDDIN - Gazi University; DOGRUMAN-ALL, FUNDA - Gazi University; Santin-Duran, Monica; Maloney, Jenny; GURESER, AYSE - Hitit University; KARASARTOVA, DJURSUN - Hitit University; TAYLAN-OZKAN, AYSEGUL - Hitit University

Submitted to: PLOS Neglected Tropical Diseases
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 8/30/2021
Publication Date: 10/6/2021
Citation: Sarzhanov, F., Dogruman-All, F., Santin, M., Maloney, J.G., Gureser, A.S., Karasartova, D., Taylan-Ozkan, A. 2021. Investigation of neglected protists Blastocystis and Dientamoeba fragilis in immunocompetent and immunodeficient diarrheal patients using both conventional and molecular methods. PLOS Neglected Tropical Diseases. 15(10):e0009779. https://doi.org/10.1371/journal.pntd.0009779.
DOI: https://doi.org/10.1371/journal.pntd.0009779

Interpretive Summary: Blastocystis and Dientamoeba fragilis are single-cell parasites of the human intestine which are common worldwide and reported in cases with gastrointestinal symptoms. However, the role of Blastocystis and D. fragilis in patients with gastrointestinal symptoms is still controversial because their presence is not always associated with symptoms. Intestinal parasitic infections are among the most important pathogens that can cause infections in immunocompromised individuals and need careful consideration. However, testing for Blastocystis and D. fragilis is not routinely carried out by most laboratories and clinicians. Therefore, this study aimed to determine the prevalence of Blastocystis and D. fragilis in immunocompetent and immunosuppressed patients with diarrhea by conventional and molecular methods. Both Blastocystis and D. fragilis were detected more frequently by quantitative polymerase chain reaction than by conventional methods. Next generation sequencing was used to characterize the prevalence of Blastocystis subtypes and mixed subtypes in patients in Turkey. Five Blastocystis subtypes (ST1, ST2, ST3, ST4 and ST6) were detected. ST3 was the most frequent subtype in both immunocompetent and immunosuppressed patients. Mono-infections were more common than mixed subtype infections. Our findings showed that Blastocystis and D. fragilis are commonly present in immunocompetent and immunodeficient patients with diarrhea. Additionally, these data contribute to our understanding of the epidemiology of Blastocystis subtype infections and can be used to shape future studies which aim to better characterize the transmission pathways and health outcomes of Blastocystis infections. This study provides beneficial information to other scientists, physicians, and public health agencies concerned with these parasites.

Technical Abstract: The clinical significance of Blastocystis 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 in human patients in Turkey are not well characterized. This study aimed to determine the prevalence of Blastocystis and D. fragilis in the diarrheic stool samples of immunodeficient and immunocompetent patients using conventional and molecular methods and to identify Blastocystis subtypes using next generation sequencing. Individual stool specimens were collected from 245 immunodeficient and 193 immunocompetent diarrheic patients at the Gazi University Training and Research Hospital in Ankara, Turkey. Samples were screened for Blastocystis and D.fragilis by conventional and molecular methods. Next generation sequencing was used to identify Blastocystis subtypes. The prevalence of Blastocystis and D. fragilis was 16.7% and 11.9%, respectively as measured by qPCR. The prevalence of Blastocystis 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 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 subtypes between immunocompetent and immunodeficient patients. Our findings demonstrated that Blastocystis 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 mixed subtypes and intra-subtype variability in clinical samples in Turkey.