Cryptosporidium rubeyi n. sp. (Apicomplexa: Cryptosporidiidae) in multiple Spermophilus ground squirrel species.

Previously we reported the unique Cryptosporidium sp. "c" genotype (e.g., Sbey03c, Sbey05c, Sbld05c, Sltl05c) from three species of Spermophilus ground squirrel (Spermophilus beecheyi, Spermophilus beldingi, Spermophilus lateralis) located throughout California, USA. This follow-up work characterizes the morphology and animal infectivity of this novel genotype as the final step in proposing it as a new species of Cryptosporidium. Analysis of sequences of 18S rRNA, actin, and HSP70 genes of additional Cryptosporidium isolates from recently sampled California ground squirrels (S. beecheyi) confirms the presence of the unique Sbey-c genotype in S. beecheyi. Phylogenetic and BLAST analysis indicates that the c-genotype in Spermophilus ground squirrels is distinct from Cryptosporidium species/genotypes from other host species currently available in GenBank. We propose to name this c-genotype found in Spermophilus ground squirrels as Cryptosporidium rubeyi n. sp. The mean size of C. rubeyi n. sp. oocysts is 4.67 (4.4-5.0) μm × 4.34 (4.0-5.0) μm, with a length/width index of 1.08 (n = 220). Oocysts of C. rubeyi n. sp. are not infectious to neonatal BALB/c mice and Holstein calves. GenBank accession numbers for C. rubeyi n. sp. are DQ295012, AY462233, and KM010224 for the 18S rRNA gene, KM010227 for the actin gene, and KM010229 for the HSP70 gene

Elevation and vegetation determine Cryptosporidium oocyst shedding by yellow-bellied marmots (Marmota flaviventris) in the Sierra Nevada Mountains.

Wildlife are increasingly recognized as important biological reservoirs of zoonotic species of Cryptosporidium that might contaminate water and cause human exposure to this protozoal parasite. The habitat range of the yellow-bellied marmot (Marmota flaviventris) overlaps extensively with the watershed boundaries of municipal water supplies for California communities along the foothills of the Sierra Nevada. We conducted a cross-sectional epidemiological study to estimate the fecal shedding of Cryptosporidium oocysts by yellow-bellied marmots and to quantify the environmental loading rate and determine risk factors for Cryptosporidium fecal shedding in this montane wildlife species. The observed proportion of Cryptosporidium positive fecal samples was 14.7% (33/224, positive number relative to total number samples) and the environmental loading rate was estimated to be 10,693 oocysts animal(-1) day(-1). Fecal shedding was associated with the elevation and vegetation status of their habitat. Based on a portion of the 18s rRNA gene sequence of 2 isolates, the Cryptosporidium found in Marmota flaviventris were 99.88%-100% match to multiple isolates of C. parvum in the GenBank

Elevation and vegetation determine Cryptosporidium oocyst shedding by yellow-bellied marmots (Marmota flaviventris) in the Sierra Nevada Mountains

Wildlife are increasingly recognized as important biological reservoirs of zoonotic species of Cryptosporidium that might contaminate water and cause human exposure to this protozoal parasite. The habitat range of the yellow-bellied marmot (Marmota flaviventris) overlaps extensively with the watershed boundaries of municipal water supplies for California communities along the foothills of the Sierra Nevada. We conducted a cross-sectional epidemiological study to estimate the fecal shedding of Cryptosporidium oocysts by yellow-bellied marmots and to quantify the environmental loading rate and determine risk factors for Cryptosporidium fecal shedding in this montane wildlife species. The observed proportion of Cryptosporidium positive fecal samples was 14.7% (33/224, positive number relative to total number samples) and the environmental loading rate was estimated to be 10,693 oocysts animal-1 day-1. Fecal shedding was associated with the elevation and vegetation status of their habitat. Based on a portion of the 18s rRNA gene sequence of 2 isolates, the Cryptosporidium found in Marmota flaviventris were 99.88%–100% match to multiple isolates of C. parvum in the GenBank

Detection and Prevalence of Cryptosporidium spp. and Giardia spp. from Wild Rodents Adjacent to Produce Production Fields in California

Between 2009 and 2011, fecal samples were collected from ten species of wild rodents in locations adjacent to leafy green blocks in a major produce production region of California. Samples were screened for Cryptosporidium spp. oocysts and Giardia spp. cysts using immunofluorescent microscopy. Five and seven species of wild rodents carried Cryptosporidium spp. and Giardia spp., respectively. In general, 26.0% and 24.2% of the trapped wild rodents were positive for Cryptosporidium spp. and Giardia spp., respectively. Deer mice (Peromyscus manicultus) were the primary trapped species, with 30.3% (63/208) positive for Cryptosporidium spp. and 25.5% (53/208) positive for Giardia spp

Prevalence and Genotypes of Cryptosporidium in Wildlife Populations Co-Located in a Protected Watershed in the Pacific Northwest, 2013 to 2016.

Between October 2013 and May 2016, 506 scat samples were collected from 22 species of wildlife located in a protected watershed of a major municipal water supply in the Pacific Northwest, USA. Overall prevalence of Cryptosporidium in the wildlife scat was 13.8% (70/506), with 15 species of wildlife found positive for Cryptosporidium. Prevalence of Cryptosporidium varied among species of wildlife, with higher prevalences observed in cougars (50.0%), mountain beavers (40.0%), and bobcats (33.3%), but none of these species are riparian-dependent. Genotyping of Cryptosporidium by sequencing PCR amplicons from the 18S rRNA gene were successful for seven species of wildlife, including bobcat, unknown predator, black-tailed deer, deer mouse, snowshoe hare, mountain beaver, and western spotted skunk. BLAST and phylogenetic analyses indicated that multiple species and genotypes of Cryptosporidium were present, with some isolates possibly co-circulating within and between wildlife populations in this protected watershed. Evidence of oocyst exchange between infected prey and their predators was also found. During the study period, several zoonotic Cryptosporidium species and genotypes that are uncommon in humans were detected in bobcat (99.58% identical to Cryptosporidium felis), unknown predator (100% identical to Cryptosporidium canis), snowshoe hare (100% identical to Cryptosporidium sp. skunk genotype), and mountain beaver (100% identical to Cryptosporidium ubiquitum). Novel sequences were also found in mountain beaver. To our knowledge, this is the first published report of a unique genotype or species of Cryptosporidium in mountain beaver (Aplodontia rufa)

Prevalence and Genotypes of Cryptosporidium in Wildlife Populations Co-Located in a Protected Watershed in the Pacific Northwest, 2013 to 2016.

Between October 2013 and May 2016, 506 scat samples were collected from 22 species of wildlife located in a protected watershed of a major municipal water supply in the Pacific Northwest, USA. Overall prevalence of Cryptosporidium in the wildlife scat was 13.8% (70/506), with 15 species of wildlife found positive for Cryptosporidium. Prevalence of Cryptosporidium varied among species of wildlife, with higher prevalences observed in cougars (50.0%), mountain beavers (40.0%), and bobcats (33.3%), but none of these species are riparian-dependent. Genotyping of Cryptosporidium by sequencing PCR amplicons from the 18S rRNA gene were successful for seven species of wildlife, including bobcat, unknown predator, black-tailed deer, deer mouse, snowshoe hare, mountain beaver, and western spotted skunk. BLAST and phylogenetic analyses indicated that multiple species and genotypes of Cryptosporidium were present, with some isolates possibly co-circulating within and between wildlife populations in this protected watershed. Evidence of oocyst exchange between infected prey and their predators was also found. During the study period, several zoonotic Cryptosporidium species and genotypes that are uncommon in humans were detected in bobcat (99.58% identical to Cryptosporidium felis), unknown predator (100% identical to Cryptosporidium canis), snowshoe hare (100% identical to Cryptosporidium sp. skunk genotype), and mountain beaver (100% identical to Cryptosporidium ubiquitum). Novel sequences were also found in mountain beaver. To our knowledge, this is the first published report of a unique genotype or species of Cryptosporidium in mountain beaver (Aplodontia rufa)

Quantitative Shedding of Multiple Genotypes of Cryptosporidium and Giardia by Deer Mice (Peromyscus maniculatus) in a Major Agricultural Region on the California Central Coast

Deer mice (Peromyscus maniculatus) are abundant and widely distributed rodents in North America that occupy diverse habitats, including agricultural landscapes. Giardia and Cryptosporidium are common parasites in wildlife including deer mice, which may play a role in on-farm contamination of produce. An important step in assessing the risk of produce contamination by Cryptosporidium and Giardia shed by deer mice is to determine the prevalence, levels, and genotypes of (oo)cysts in mouse feces. A total of 63 (30.3%) and 53 (25.5%) of 208 deer mice trapped on 12 farms on the California Central Coast were positive for Cryptosporidium and Giardia, respectively. Of these mice, 41 (19.7%) contained both parasites. The odds of Cryptosporidium shedding were 2.5 to 5 times higher for mice trapped in autumn than for mice trapped in summer or spring. Female mice had a higher prevalence and two- to threefold higher levels of Cryptosporidium and Giardia compared with male mice. Female adults and female juveniles had the highest rates of contamination of the environment with Cryptosporidium and Giardia, respectively. We estimated that 20 infected deer mice inhabiting 1 ha of a typical leafy green produce farm in the study region could shed approximately 5.3 × 108 Cryptosporidium and 10.5 × 108 Giardia, respectively, per day into the environment. The small-subunit rRNA gene loci from a subset of protozoan isolates were sequenced and compared with existing sequences in GenBank. Multiple genotypes of Cryptosporidium and Giardia were found, and BLAST analyses suggest that Giardia and the majority of Cryptosporidium genotypes in deer mice circulate within various rodent populations, but some Cryptosporidium isolates possess zoonotic potential

Statewide Cross-Sectional Survey of Cryptosporidium and Giardia in California Cow-Calf Herds

Cryptosporidium spp. and Giardia duodenalis are common protozoal parasites in livestock including beef cattle on rangeland and irrigated pasture. A statewide cross-sectional study was conducted to determine the prevalence, species or genotype, and risk factors for fecal shedding of Cryptosporidium and Giardia by cattle from California cow-calf operations. Species and genotypes of Cryptosporidium and Giardia were determined by molecular fingerprinting. Prevalence of Cryptosporidium (19.8%) and Giardia (41.7%) in fecal samples from calves were approximately twice as high as fecal samples from cows (9.2% and 23.1%, respectively). In addition to age, multivariable logistic regression showed that higher stocking density and a higher number of replacement heifers were positively associated with fecal shedding of Cryptosporidium while longer calving interval, a winter/spring calving season, and higher numbers of replacement heifers were positively associated with shedding of Giardia. The dominant species and genotypes of Cryptosporidium and Giardia in feces from these cow-calf herds were Cryptosporidium ryanae (75%) and assemblage E for Giardia duodenalis (90%), which have low impact on public health compared with other zoonotic species/genotypes of these two parasites. We identified host and potential management practices that can be used to protect cattle health and reduce the risk of surface water contamination with protozoal parasites from cow-calf operations. In addition, this work updated the scientific data regarding the predominance of low zoonotic genotypes of Cryptosporidium and Giardia shed in the feces of commercial cow-calf herds on California rangeland and irrigated pasture