First identification of genotypes of Enterocytozoon bieneusi (Microsporidia) among symptomatic and asymptomatic children in Mozambique

Authors: SALIMO MUADICA, ALY - Instituto De Salud Carlos Iii; MESSA, AUGUSTO JR. - Instituto De Salud Carlos Iii; DASHTI, ALEJANDRO - Instituto De Salud Carlos Iii; Santin-Duran, Monica; MANJATE, FILOMENA - Instituto De Salud Carlos Iii; CHIRINDA, PERCINA - Instituto De Salud Carlos Iii; GARRINE, MARCELINO - Universidade Nova De Lisboa; VUBIL, DELFINO - Instituto De Salud Carlos Iii; ACACIO, SOZINHO - Instituto De Salud Carlos Iii; KOSTER, PAMELA - Instituto De Salud Carlos Iii; BAILO, BEGOÑA - Carlo Besta Neurological Institute; NHAMPOSSA, TACILTA - Eduardo Mondlane University; CALERO-BERNAL, RAFAEL - Complutense University Of Madrid (UCM); MWENDA, JASON - University Of Kisangani; MANDOMANDO, INACIO - Universidade Nova De Lisboa; CARMENA, DAVID - Instituto De Salud Carlos Iii

Submitted to: PLOS Neglected Tropical Diseases
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 5/23/2020
Publication Date: 6/30/2020
Citation: Salimo Muadica, A., Messa, A., Dashti, A., Santin, M., Manjate, F., Chirinda, P., Garrine, M., Vubil, D., Acacio, S., Koster, P., Bailo, B., Nhampossa, T., Calero-Bernal, R., Mwenda, J., Mandomando, I., Carmena, D. 2020. First identification of genotypes of Enterocytozoon bieneusi (Microsporidia) among symptomatic and asymptomatic children in Mozambique. PLOS Neglected Tropical Diseases. 14(6):e0008419. https://doi.org/10.1371/journal.pntd.0008419.
DOI: https://doi.org/10.1371/journal.pntd.0008419

Interpretive Summary: Enterocytozoon bieneusi is an obligate intracellular parasite that infects a wide range of vertebrate hosts. It is the most important etiological agent of human microsporidiasis that in immunodeficient immunosuppressed individuals can cause life-threatening chronic diarrhea. Molecular epidemiological studies in humans and animals have revealed that E. bieneusi comprises a very large diversity of genetic variants (genotypes) with marked differences in host specificity and even geographical distribution. In Mozambique, as in many other African countries, the epidemiology of E. bieneusi is completely unknown. Therefore, to identify the occurrence and genetic diversity of this pathogen in Mozambique stool samples were obtained from children, including apparently healthy and symptomatic, in Zambézia and Maputo provinces and tested for E. bieneusi by molecular methods. Results demonstrated the presence of E. bieneusi genotypes for the first time in Mozambique. Four genotypes were identified, three genotypes that have previously been reported in humans (Peru11, Type IV, and S2) and a novel genotype (HhMzEb1). Two of the genotypes Peru11, Type IV have also been frequently identified in animals indicating that potentially zoonotic E. bieneusi genotypes are inadvertently circulating in the surveyed populations. Additional population genetic studies are needed to elucidate the actual extent of the epidemiology and transmission dynamics of E. bieneusi in Mozambique. This study provides beneficial information to other scientists, physicians, and public health agencies concerned with this widespread parasite.

Technical Abstract: Introduction Enterocytozoon bieneusi (Phylum Microsporidia) is a human pathogen with a broad range of animal hosts. Initially, E. bieneusi was considered an emerging opportunistic pathogen in immunocompromised, mainly HIV-infected patients, but it has been increasingly reported in apparently healthy individuals globally. As in other African countries, the molecular epidemiology of E. bieneusi in Mozambique remains completely unknown. Therefore, we undertook a study to investigate the occurrence and genetic diversity of E. bieneusi infections in children with gastrointestinal symptoms as well as in asymptomatic children in Mozambique. Materials and methods Individual stool specimens were collected from 1,247 children aged between 0 and 14 years-old living in urban and rural settings in Zambézia (n = 1,097) and Maputo (n = 150) provinces between 2016 and 2019. Samples were analyzed for E. bieneusi by nested-PCR targeting the internal transcribed spacer (ITS) region of the rRNA gene. All positive amplicons were confirmed and genotyped by Sanger sequencing. Results The overall prevalence of E. bieneusi in this children population was 0.7% (9/1,247). The prevalence was found to be more than 10-fold higher in Maputo (4.0%; 6/150) than in Zambézia (0.3%; 3/1,097). All E. bieneusi-positive samples were from children older than 1-year of age, and most (8/9) from asymptomatic children. Nucleotide sequence analysis of the ITS region revealed the presence of four genotypes, three previously reported (Peru11, n = 1; Type IV, n = 2, and S2, n = 2) and a novel genotype (named HhMzEb1, n = 4). Novel genotype HhMzEb1 was identified in both asymptomatic (75%, 3/4) and symptomatic (25%, 1/4) children from a rural area in Maputo province in southern Mozambique. Genotypes HhMzEb1, Peru11, S2, and Type IV belonged to the Group 1 that includes genotypes with low host specificity and the potential for zoonotic and cross-species transmission. Conclusion and recommendation This study reports the first identification of E. bieneusi genotypes in Mozambique with the identification of three previously reported genotypes in humans as well as a novel genotype (HhMzEb1). Findings highlight the need to conduct additional research to elucidate the epidemiology of E. bieneusi in the country, especially in rural areas where poor hygiene conditions still prevail. Special attention should be paid to the identification of suitable animal and environmental reservoirs of this parasite and to the characterization of transmission pathways.