Genomic Variation in IbA10G2 and Other Patient-Derived Cryptosporidium hominis Subtypes

• Published in: Journal of clinical microbiology Vol. 55; no. 3; pp. 844 - 858
• Main Authors: Sikora, Per, Andersson, Sofia, Winiecka-Krusnell, Jadwiga, Hallström, Björn, Alsmark, Cecilia, Troell, Karin, Beser, Jessica, Arrighi, Romanico B G
• Format: Journal Article
• Language: English
• Published: United States American Society for Microbiology 01.03.2017
• Subjects: clinical parasitology; cryptosporidiosis; Cryptosporidiosis - parasitology; Cryptosporidium - classification; Cryptosporidium - genetics; Cryptosporidium - isolation & purification; Feces - parasitology; Genetic Variation; genome sequencing; Genome, Protozoan; Genomics; Genotype; Humans; immunomagnetic separation; Iowa; Parasite Load; Parasitology; Phylogeny; Sequence Analysis, DNA; Synteny; immunomagnetic separation; genome sequencing; clinical parasitology; cryptosporidiosis
• ISSN: 0095-1137; 1098-660X; 1098-660X
• DOI: 10.1128/jcm.01798-16

In order to improve genotyping and epidemiological analysis of spp., genomic data need to be generated directly from a broad range of clinical specimens. Utilizing a robust method that we developed for the purification and generation of amplified target DNA, we present its application for the successful isolation and whole-genome sequencing of 14 different patient specimens. Six isolates of subtype IbA10G2 were analyzed together with a single representative each of 8 other subtypes: IaA20R3, IaA23R3, IbA9G3, IbA13G3, IdA14, IeA11G3T3, IfA12G1, and IkA18G1. Parasite burden was measured over a range of more than 2 orders of magnitude for all samples, while the genomes were sequenced to mean depths of between 17× and 490× coverage. Sequence homology-based functional annotation identified several genes of interest, including the gene encoding oocyst wall protein 9 (COWP9), which presented a predicted loss-of-function mutation in all the sequence subtypes, except for that seen with IbA10G2, which has a sequence identical to the reference Iowa II sequence. Furthermore, phylogenetic analysis showed that all the IbA10G2 genomes form a monophyletic clade in the tree as expected and yet display some heterogeneity within the IbA10G2 subtype. The current report validates the aforementioned method for isolating and sequencing directly from clinical stool samples. In addition, the analysis demonstrates the potential in mining data generated from sequencing multiple whole genomes of from human fecal samples, while alluding to the potential for a higher degree of genotyping within epidemiology.