Population Structure of Enterocytozoon bieneusi in Captive Macaques

Enterocytozoon bieneusi is a microscopic eukaryote classified in the phylum Microsporidia. It is the most common cause of human microsporidiosis. Over 300 diarrheic or soft fecal samples from captive macaques were screened by immunofluorescence for the presence of E. bieneusi spores and by PCR for E. bieneusi DNA. Fifty-three samples were identified as positive by both methods. Positive animals were significantly younger, but the infection did not negatively affect growth. With the goal of understanding the parasite’s population structure, we analyzed the E. bieneusi internal transcribed spacer (ITS) genotype. Classification of a total of 1,031,089 curated sequence contigs obtained from 53 samples showed the presence of sequences pertaining to three of the previously described eleven ITS genotypic groups. Indicative of mixed parasite populations, most E. bieneusi samples from individual macaques were characterized by ITS sequences belonging to several genotypic groups. Single-genotype isolates were the exception. The analysis of multiple samples from five macaques revealed a statistically highly significant clustering of ITS genotype by animal. The frequent occurrence of genotypically mixed ITS genotypes underscores the need for multiple curated genome sequences to support the unambiguous interpretation of sequences data and advance the study of this enigmatic pathogen.