Cryptosporidium species

Wei, Zilin, Liu, Qiang, Zhao, Wei, Jiang, Xiumei, Zhang, Ying, Zhao, Aiyun, Jing, Bo, Lu, Gang & Qi, Meng, 2019, Prevalence and diversity of Cryptosporidium spp. in bamboo rats (Rhizomys sinensis) in South Central China, International Journal for Parasitology: Parasites and Wildlife 9, pp. 312-316 : 313-314

3.2. Cryptosporidium species /genotypes

Sequence analysis of the SSU rRNA products of nine Cryptosporidium isolates, four Cryptosporidium species, or genotypes including C. parvum (2; 22.2%), C. occultus (1; 11.1%) and two new genotypes named Cryptosporidium bamboo rat genotype I (5; 55.5%) and Cryptosporidium bamboo rat genotype II (1; 11.1%) were identified ( Table 1). Cryptosporidium bamboo rat genotype I was dominant in bamboo rats, and was identified in two farms from the Yongzhou, Hunan Province. A single C. parvum isolate was found in the Luoding farm, whilst another was identified in Yongzhou farm 2. C. occultus was found in a single bamboo rat also from Yongzhou farm 2 ( Table 1).

At the SSU rRNA locus, two sequences of C. parvum (rats 139 and 416) were identical and had 100% similarity with (KC885893) from bamboo rats in Sichuan of China. The sequence of C. occultus revealed 100% similarity with MG 699177 in Rattus norvegicus from the Czech Republic. Five sequences of Cryptosporidium bamboo rat genotype I were identical and shared 97.7% identity with the MH794165 sequence of C. ubiquitum in sheep from Xinjiang of China. The Sequence of the Cryptosporidium bamboo rat genotype II isolate (bamboorat 421; Genebank number MK731962) have not been previously reported and had a homology of 99.3% to the MK241967 isolate in cattle from India.

Two C. parvum -positive and the new genotypes isolates were further subtyped by sequence analysis of the gp60 gene, and the two C. parvum and the Cryptosporidium bamboo rat genotype II were successfully amplified. However, the five isolates of Cryptosporidium bamboo rat genotype I were amplification failure. Phylogenetic analysis of the gp60 sequences suggested that they belong to three subtype families ( Fig. 1 ). Sequences of the two C. parvum isolates had 100% similarity with subtype IIpA9 (KC885904) which was isolated from a bamboo rat from Sichuan, and IIdA15G1 (KJ917586) which was found in a monkey from Shaanxi of China, respectively. The gp60 sequence of the Cryptosporidium bamboo rat genotype II (Genebank number MK731966) had a maximum nucleotide identity of 84.1% to C. parvum subtype IIaA15G2R1 (KX507244), which had extensive sequence polymorphisms in the non-repeat regions.

None of the five isolates of Cryptosporidium bamboo rat genotype I were successfully amplified at the COWP gene but successfully amplified at the HSP70 gene, and the five HSP70 gene sequences (MK731968) of Cryptosporidium bamboo rat genotype I were identical to each other and they had 96.4% similarity with that of C. parvum from human in Canada (DQ389176). Meanwhile, the Cryptosporidium bamboo rat genotype II-positive isolate was successfully amplified at the both COWP and HSP70 genes, which were not been reported previously based on BLAST search results. The COWP gene sequence (MK731967) of isolate of Cryptosporidium bamboo rat genotype II has nine nucleotide substitutions with C. parvum in mouse from China (AB469366). The HSP70 gene sequence (MK731969) of Cryptosporidium bamboo rat genotype II had 96.1% similarity with that of C. parvum from human in Canada (DQ389176). For actin gene only two bamboo rat genotype I isolates were successfully amplified, and the two isolates shared a same sequence which had 95.5% similarity with that of C. ubiquitum from human in USA (XM_029019099). Phylogenetic analysis of the actin, gp60, HSP70, COWP and SSU rDNA gene sequences showed in Fig. 1 .

Figures

Fig. 1. Phylogenetic relationship of the five gene loci of Cryptosporidium species/genotypes. The Panel A to E are represent the actin, gp60, HSP70, COWP and SSU rDNA gene, respectively. The numbers on the branches are percent bootstrapping values from 1000 replicates. Each sequence is identified by its accession number and Cryptosporidium species/genotypes designation. The triangle filled in black indicate the subtypes identified in this study.