title: Genomic Variation in IbA10G2 and Other Patient-Derived Cryptosporidium hominis Subtypes.
abstract: In order to improve genotyping and epidemiological analysis ofCryptosporidiumspp., 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 differentCryptosporidium hominispatient 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 encodingCryptosporidiumoocyst 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 theCryptosporidium parvumreference Iowa II sequence. Furthermore, phylogenetic analysis showed that all the IbA10G2 genomes form a monophyletic clade in theC. hoministree as expected and yet display some heterogeneity within the IbA10G2 subtype. The current report validates the aforementioned method for isolating and sequencingCryptosporidiumdirectly from clinical stool samples. In addition, the analysis demonstrates the potential in mining data generated from sequencing multiple whole genomes ofCryptosporidiumfrom human fecal samples, while alluding to the potential for a higher degree of genotyping withinCryptosporidiumepidemiology.
authors.affiliation: Laboratory Medicine Core Facility, Sahlgrenska University Hospital, Gothenburg, Sweden.|Department of Microbiology, Public Health Agency of Sweden, Solna, Sweden.
authors.forename: Per
authors.initials: P
authors.lastname: Sikora
authors.affiliation: Department of Microbiology, Public Health Agency of Sweden, Solna, Sweden.
authors.forename: Sofia
authors.initials: S
authors.lastname: Andersson
authors.affiliation: Department of Microbiology, Public Health Agency of Sweden, Solna, Sweden.
authors.forename: Jadwiga
authors.initials: J
authors.lastname: Winiecka-Krusnell
authors.affiliation: Department of Microbiology, Public Health Agency of Sweden, Solna, Sweden.
authors.forename: Björn
authors.initials: B
authors.lastname: Hallström
authors.affiliation: Department of Microbiology, National Veterinary Institute, Uppsala, Sweden.|Division of Pharmacognosy, Department of Medicinal Chemistry, Uppsala University, Sweden.
authors.forename: Cecilia
authors.initials: C
authors.lastname: Alsmark
authors.affiliation: Department of Microbiology, National Veterinary Institute, Uppsala, Sweden.
authors.forename: Karin
authors.initials: K
authors.lastname: Troell
authors.affiliation: Department of Microbiology, Public Health Agency of Sweden, Solna, Sweden.
authors.forename: Jessica
authors.initials: J
authors.lastname: Beser
authors.affiliation: Department of Microbiology, Public Health Agency of Sweden, Solna, Sweden romanico.arrighi@folkhalsomyndigheten.se.
authors.forename: Romanico B G
authors.initials: RB
authors.lastname: Arrighi
mesh_terms: D003457:Cryptosporidiosis / Q000469:parasitology*; D003458:Cryptosporidium / Q000145:classification* / Q000235:genetics* / Q000302:isolation & purification; D005243:Feces / Q000469:parasitology*; D014644:Genetic Variation; D018503:Genome, Protozoan; D023281:Genomics; D005838:Genotype; D006801:Humans; D007484:Iowa; D059208:Parasite Load; D010802:Phylogeny; D017422:Sequence Analysis, DNA; D026801:Synteny
keywords: clinical parasitology; cryptosporidiosis; genome sequencing; immunomagnetic separation