| Summary: | Microsporidia are unicellular eukaryotic obligate intracellular parasites with a wide range of hosts reported worldwide; however, little is known about the epidemiological data on microsporidia infection in animals from the Canary Islands. Since data on microsporidia infection in hedgehog species are scarce, the aim of this study was to analyze the presence and identity of microsporidia in a group of North African hedgehogs (<i>Atelerix algirus</i>) using microscopic and molecular methods. From December 2020 to September 2021, a total of 36 fecal samples were collected from naturally deceased hedgehogs from Tenerife and Gran Canaria. All samples showed spore-compatible structures (100%; 36/36) under microscopic analysis, of which 61.1% (22/36) were amplified via the nested-polymerase chain reaction (PCR) targeting the partial sequence of the 16S rRNA gene, the internal transcribed spacer (ITS) region, and the partial sequence of the 5.8S rRNA gene. After Sanger sequencing and ITS analysis, <i>Enterocytozoon bieneusi</i> was detected in 47.2% (17/36) of the samples, identifying two novel genotypes (AAE1 and AAE2), followed by the detection of an undetermined species in 8.3% (3/36) and <i>Encephalitozoon cuniculi</i> genotype I in 5.6% (2/36) of the samples. This study constitutes the first report of microsporidia species in <i>Atelerix algirus</i> worldwide, highlighting the high prevalence of zoonotic species.
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