Subgenotype analysis of Cryptosporidium parvum isolates from humans and animals in Japan using the 60-kDa glycoprotein gene sequences

Abstract

Cryptosporidium parvum is a well-known intestinal parasite which is associated with severe acute diarrhea in humans and animals. This parasite is composed of morphologically identical but genetically different multiple genotypes. In humans, cryptosporidiosis is mainly caused by two C. parvum genotypes, human genotype (previously known as genotype 1 and recently proposed as new species C. hominis) and cattle genotype (previously known as genotype 2). However, recent molecular studies indicate the genetic heterogeneity among the isolates of C. parvum human or cattle genotype. Therefore, identification of the isolates at the subgenotype level is more useful for control of the Cryptosporidium infection or for understanding of the population structure of C. parvum genotypes. In the present study, we identified the subgenotypes of the C. parvum human or cattle genotype isolates from humans and animals in Japan using DNA sequencing analysis of the C. parvum 60-kDa glycoprotein gene (GP60) and showed the new subgenotype in a raccoon dog isolate. This study suggested that C. parvum cattle genotype might be composed of zoonotic and host-specific multiple subgenotypes.

Classification ofCryptosporidiumSpecies from Patients with Sporadic Cryptosporidiosis by Use of Sequence-Based Multilocus Analysis following Mutation Scanning

“…For example, a sample with a sequence consisting of 10 TCA repeats, 3 TCG repeats, and 2 ACA repeats would be identified to the subgenotype as A10G3R2. As there currently is no consensus opinion on the precise definition of a genotype (or the threshold of intragenotypic versus intergenotypic sequence variation), a sample was assigned to a particular genotype based on its relationship with strongly supported clades after the phylogenetic analysis of the p-gp60 sequence data for all samples studied, including published sequences representing all currently recognized gp60 genotypes of C. hominis (Ia to If and Ib2 [7,36,71]) and C. parvum (IIa to IIk [1,4,7,47,58,72,76]) (for accession numbers, see Fig. 3).…”

“…Cryptosporidium parvum IIa (GenBank accession number DQ192502 [78]) was used as the outgroup in the analysis of p-gp60 sequence data for C. hominis. Cryptosporidium hominis Ia (GenBank accession number AB273130 [1]) was used as the outgroup in the analysis of p-gp60 data for C. parvum. Upon the completion of the Bayesian analysis, a 50% majority-rule consensus tree for each species was constructed in Treeview X v.0.5.0 (http://darwin.zoology.gla.ac.uk /ϳrpage/treeviewx/) and enhanced using InkScape 0.45 (www.inkscape.org).…”

“…Given the substantial sequence variability in p-gp60 between C. hominis and C. parvum and the inability to reliably align all sequences to achieve positional homology, the sequence data set for each species was subjected to separate phylogenetic analyses. The analysis of gp60 data showed that each recognized genotype of C. hominis (Ia to If) (7,36,71) and of C. parvum (IIa to IIk (1,4,7,47,58,72,76) resolved as a separate, strongly supported clade (pp of 1.00 for individual genotypic clades). The detailed appraisal of the C. hominis tree (Fig.…”

Multicentric Evaluation of a New Real-Time PCR Assay for Quantification of Cryptosporidium spp. and Identification of Cryptosporidium parvum and Cryptosporidium hominis

“…Higher sensitivity of detection in environmental and clinical samples can be achieved by performing molecular diagnosis of cryptosporidiosis using PCR. The most commonly used targets are the 18S rRNA gene (18), the Cryptosporidium oocyst wall protein (19), the 60-kDa glycoprotein (20), heat shock protein 70, the Laxer locus, and microsatellite loci (reviewed in reference 21). Several studies agree on the higher sensitivity of PCR targeting the 18S rRNA gene, in relation to its copy number (22).…”

Potencial bioterapêutico dos probióticos nas parasitoses intestinais

“…No caso de Cryptosporidium, essas discrepâncias de resultados são particularmente difíceis de serem analisadas, pois a quantidade de dados disponíveis é pequena, os resultados foram obtidos em experimentos envolvendo diferentes espécies ou categorias de animais experimentais e de bactérias probióticas e foram empregados distintos protocolos para a administração dos probióticos. Além disso, atualmente, são admitidas várias espécies e vários genótipos de Cryptosporidium, morfologicamente indistinguíveis de C. parvum, cuja identificação genotípica é importante, inclusive para se avaliar as possibilidades de controle (ABE et al, 2006).…”