In the present study, we have extended earlier taxonomic, biochemical and experimental investigations to characterize Echinococcus granulosus from various hosts in Iran utilizing DNA regions (designated pcox1 and pnad1) within the cytochrome c oxidase subunit 1 and NADH dehydrogenase 1 mitochondrial genes, respectively. An emphasis was placed on the characterization of E. granulosus isolates (cyst material) from humans, sheep, goats, cattle and camels, and on assessing their genetic relationships. PCR-based SSCP analysis of pcox1 and pnad1 amplicons derived from individual isolates (n=148) of E. granulosus revealed five (pc1-pc5) and nine (pn1-pn9) electrophoretic profiles, respectively. Sequencing of pcox1 and pnad1 amplicons representing unique SSCP profiles demonstrated that each profile was linked unequivocally to a particular sequence and that single point mutations were readily detectable by SSCP. Phylogenetic analyses of pcox1 and/or pnad1 nucleotide sequence data were conducted using Bayesian inference and maximum likelihood tree-building methods. Following the phylogenetic analyses of concatenated pcox1+pnad1 sequence data, including representatives of all presently recognized Echinococcus species/genotypes as well as Taenia saginata (as the outgroup), the majority of cyst isolates (142 of 148; 95.9%) from humans, ruminants (sheep, goats and cattle) and camels were assigned to the G1-G3 complex of E. granulosus (or E. granulosus sensu stricto), whereas some E. granulosus cysts (6 of 19; 31.6%) from camels were assigned to the G6-G10 complex (or E. canadensis). The present study reinforces the advantages of the mutation scanning-sequencing-phylogenetic approach to explore variation in multiple mitochondrial loci within and among Echinococcus populations, which provides a platform for future, detailed studies of the molecular epidemiology of E. granulosus in Iran and other countries. (Note: The sequences determined in the present study have been deposited in the GenBank database under accession numbers: FJ796203-FJ796207 (pcox1) and FJ796208-FJ796216 (pnad1)).

In the present study, we analyzed genetic variation in Cryptosporidium species from humans (n ‫؍‬ 62) with clinical cryptosporidiosis in South Australia. Sequence variation was assessed in regions within the small subunit of nuclear rRNA (p-SSU), the 70-kDa heat shock protein (p-hsp70), and the 60-kDa glycoprotein (p-gp60) genes by employing single-strand conformation polymorphism analysis and sequencing. Based on the analyses of p-SSU and p-hsp70, Cryptosporidium hominis (n ‫؍‬ 38) and Cryptosporidium parvum (n ‫؍‬ 24) were identified. The analysis of p-gp60 revealed eight distinct subgenotypes, classified as C. hominis IaA17R1 (n ‫؍‬ 3), IbA9G3R2 (n ‫؍‬ 14), IbA10G2R2 (n ‫؍‬ 20), and IfA12G1R1 (n ‫؍‬ 1), as well as C. parvum IIaA18G3R1 (n ‫؍‬ 15), IIaA20G3R1 (n ‫؍‬ 6), IIaA22G4R1 (n ‫؍‬ 2), and IIcA5G3R2 (n ‫؍‬ 1). Subgenotypes IaA17R1 and IIaA22G4R1 are new. Of the six other subgenotypes, IbA10G2R2, IIaA18G3R1, IIaA20G3R1, and IIcA5G3R2 were reported previously from the state of Victoria. This is the fourth record in Australia of C. parvum subgenotype IIaA18G3R1 from humans, which, to date, has been isolated only from cattle in other countries. This subgenotype might be a significant contributor to sporadic human cryptosporidiosis and may indicate a greater zoonotic contribution to the infection of humans in the area of study. Comparative analyses revealed, for the first time, the differences in the genetic makeup of Cryptosporidium populations between two relatively close, major metropolitan cities.

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This study explored the genetic composition of Giardia in fecal samples from 284 individual lambs on pasture-based sheep farms in three regions of Victoria, Australia. An approach, combining targeted sequencing, phylogenetic analysis and PCR-coupled restriction endonuclease fingerprinting, was used to identify and genetically categorize Giardia present in 43 (15.1%) of the 284 samples and to infer their zoonotic potential. The specific identity and genetic classification were based on the phylogenetic analysis of sequence data for a portion of the triose-phosphate isomerase gene. Fourteen different sequence variants (including seven sequences that contained between one and five polymorphic sites) representing two distinct assemblages of Giardia (recognized in the current literature) were defined, of which 13 were new records. One dominant sequence type (with accession no. GQ444447, representing a genotype within assemblage A) has been detected previously in humans and is thus considered to be of zoonotic potential. (Nucleotide sequences reported in this article are available in the GenBank database under accession nos. GQ444447-GQ444451 and GQ444454-GQ444462).

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