Abstract
BackgroundData on the genus Sarcocystis in insectivores are limited. The Asian gray shrew Crocidura attenuata is one of the most common species of the insectivore family Soricidae in South Asia and Southeast Asia. To our knowledge, species of Sarcocystis have never been recorded previously in this host.MethodsTissues were obtained from 42 Asian gray shrews caught in 2017 and 2018 in China. Sarcocysts were observed using light microscopy (LM) and transmission electron microscopy (TEM). To describe the parasite life cycle, muscle tissues of the host infected with sarcocysts were force-fed to two beauty rat snakes Elaphe taeniura. Individual sarcocysts from different Asian gray shrews, and oocysts/sporocysts isolated from the small intestines and feces of the experimental snakes, were selected for DNA extraction, and seven genetic markers, namely, two nuclear loci [18S ribosomal DNA (18S rDNA) and internal transcribed spacer region 1 (ITS1)], three mitochondrial genes [cytochrome oxidase subunit 1 (cox1), cox3 and cytochrome b], and two apicoplast genes (RNA polymerase beta subunit and caseinolytic protease C), were amplified, sequenced and analyzed.ResultsSarcocysts were found in 17 of the 42 (40.5%) Asian gray shrews. Under LM, the microscopic sarcocysts showed saw- or tooth-like protrusions measuring 3.3–4.5 μm. Ultrastructurally, the sarcocyst wall contained numerous lancet- or leaf-like villous protrusions, similar to those described for type 9h of the common cyst wall classification. The experimental beauty rat snakes shed oocysts/sporocysts measuring 11.9–16.7 × 9.2–10.6 μm with a prepatent period of 10–11 days. Comparison of the newly obtained sequences with those previously deposited in GenBank revealed that those of 18S rDNA and cox1 were most similar to those of Sarcocystis scandentiborneensis recorded in the tree shrews Tupaia minor and Tupaiatana (i.e., 97.6–98.3% and 100% identity, respectively). Phylogenetic analysis based on 18S rDNA or ITS1 sequences placed this parasite close to Sarcocystis spp. that utilize small animals as intermediate hosts and snakes as the known or presumed definitive host. On the basis of morphological and molecular characteristics and host specificity, the parasite was proposed as a new species, named Sarcocystis attenuati.ConclusionsSarcocysts were recorded in Asian gray shrews, to our knowledge for the first time. Based on morphological and molecular characterization, a new species of parasite is proposed: Sarcocystisattenuati. According to the LM and TEM results, S. attenuati sarcocysts are distinct from those of Sarcocystis spp. in other insectivores and those of S. scandentiborneensis in tree shrews. The 18S rDNA or cox1 sequences of Sarcocystis attenuati shared high similarity with those of Sarcocystisscandentiborneensis, Sarcocystis zuoi, Sarcocystis cf. zuoi in the Malayan field rat, and Sarcocystis sp. in the greater white-toothed shrew. Therefore, we suggest that more research on the relationships of these closely related taxa should be undertaken in the future.Graphical abstract
Highlights
Data on the genus Sarcocystis in insectivores are limited
The examination of fresh samples under light microscopy (LM) revealed that the cyst walls of the sarcocysts had numerous 3.3to 4.5-μm-long [mean = 4.0 ± 0.26 μm (± SD); n = 25 measurements taken from ten sarcocysts] saw- or toothlike protrusions (Fig. 1a)
Phylogenetic analysis Phylogenetic analysis based on 18S 18S ribosomal DNA (rDNA) or internal transcribed spacer region 1 (ITS1) sequences confirmed that the parasite found in the present study belonged to Sarcocystis (Figs. 2, 3)
Summary
The Asian gray shrew Crocidura attenuata is one of the most common species of the insectivore family Soricidae in South Asia and Southeast Asia. Species of Sarcocystis have never been recorded previously in this host. More than 200 species of Sarcocystis have been described in a variety of wild and domesticated animals [1]. Only two species, Sarcocystis booliati and Sarcocystis russuli, have been described which are thought to use insectivorous animals as intermediate hosts [1,2,3]. The Asian gray shrew Crocidura attenuata, a species of the insectivore family Soricidae, is one of the most common species of insectivore found throughout South and Southeast Asia [5]. No species of the genus Sarcocystis have been found and recorded previously in this host
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