title: Cryptosporidiosis in two regions of Saudi Arabia
Creators: name: Areeshi, Mohammed
Publisher: name: University of Liverpool Repository
Year: 2023

description: This is the first study on human and animal infections with Cryptosporidillm spp. in I Gizan and Maddina regions of Saudi Arabia. Between March 2005 and September 2006, stool Isamples from 1641 people (predominantly children) were screened for Cryptosporidillm spp. , These included 454 patients with diarrhoea and 62 controls in Gizan and 1125 patients from IMaddina. In addition, 279 animal samples were examined from Gizan, including cows, goats and i sheep. j The human samples from Gizan were initially screened by miGroscopy using modified j Ziehl-Neelsen (ZN) and Safranin Methylene Blue staining techniques, while the animal samples ; were only screened using a modified ZN staining technique. The control samples were all i negative by both methods. ZN identified Cryptosporidium oocysts in 21 (4.6%) samples and i SM-B identified oocysts in 26 (5.7%) out of454 human samples from Gizan. ZN only identified i Cryptosporidium oocysts in one sheep sample. Stools from Maddina were not routinely examined j by microscopy unless positive by other means. . All 1920 faecal samples (human or animal) were evaluated using an Enzyme Immunoassay (EIA), which detected Cryptosporidium Specific Antigen in 104 (5.4 %) ofthe samples (103 human and one animal). Comparison ofthe detection rates between the diagnostic methods, Le. microscopy and EIA, using a subgroup of454 samples from Gizan, showed 100% agreement in specificity between the two methods, but EIA was more sensitive (5.7% vs 9.9%). In both Gizan and Maddina regions, cryptosporidiosis was more common in children less than two years ofage and the maximum prevalences were observed during the cooler months. All 104 samples (46 from Gizan, 58 from Maddina) containing Cryptosporidium oocysts were initially amplified by I8S rRNA-based PCR. The 101 (97.1%) samples positive for Cryptosporidium were further genotyped by an 18S rRNA-based PCR-Restriction Fragment Length Polymorphism (RFLP) technique. The 104 samples were also amplified using PCR for GP60 and HSP70 genes, for multilocus sub-genotyping. Among the 101 samples which were positive by 18S rRNA PCR, 79 (78.2 %) isolates were subsequently classified by RFLP as C. parvum, 13 (12.9 %) as C. hominis and one (1 %) was C.fe/is (from a sheep in the Gizan area). 8 human samples from the Gizan area contained mixed infections with C. parvum and C. hominis, as determined using RFLP of I8S rRNA gene. 95 (91.3%) and 88 (84.6%) ofthe 104 samples containing Cryptosporidium oocysts were successfully amplified for sub-genotype studies, using PCR for the GP60 and HSP70 genes respectively. Sequence analysis of study isolates ofthese two loci in general confirmed the species identification obtained by thel8S rRNA gene PCR- RFLP analysis. In total, allele groups IIa, IIc and IId of C. parvum and allele groups Ib and Ie of C. hominis were obtained by GP60 sequencing. HSP70 sequence analysis was less successful and did not add much to discrimination. Cryptosporidiosis was common in children, and screening of stools by EIA was twice as sensitive as light microscopy after special staining. The predominant species in humans was zoonotic C. parvum, with some C. parvum subtypes associated with anthroponotic spread also identified by multilocus sequencing. PCR amplification of both GP60 and HSP70 genes was successful, but GP60 was the most useful additional tool, showing extensive genetic heterogeneity in Cryptosporidium spp. at the subtype level. Although this may correlate with origin of the infection, studies in local animals were unsuccessful in confirming possible animal sources.