Development and evaluation of a novel loop-mediated isothermal amplification (LAMP) method targeting Theileria parasites infecting Yezo sika deer

The increasing Yezo sika deer (Cervus nippon yesoensis) population is creating a large problem. Yezo sika deer are an important blood meal source, and these deer contribute to the maintenance of tick populations. Theileria spp. infections in Yezo sika deer and T. orientalis infections in cows occur at high frequencies, and the same tick species infests both deer and cows. Therefore, a specific detection method to identify deer Theileria spp. is important. In this study, we establish a novel molecular detection method for identifying Theileria spp. from deer and tick samples using loop-mediated isothermal amplification (LAMP). This method targets a metalloprotease/cell division cycle protein gene homologue. Our LAMP protocol was able to detect deer Theileria and did not show cross reactivity with other closely related protozoan parasites, including T. orientalis. The LAMP method showed sensitivity and specificity equivalent to those of nested PCR performed on the same field samples from deer and ticks. These results demonstrate the applicability of LAMP to field surveys in which the detection of deer Theileria spp. is required. In conclusion, due to its simplicity, specificity, and reliability, we suggest our LAMP protocol as an appropriate method for routine surveys to detect Yezo sika deer and ticks infected with deer Theileria spp. parasites. Additionally, this LAMP method offers great promise as a useful tool to distinguish Yezo sika deer Theileria from related Theileria parasites present in livestock.     

Seroprevalence of fasciolosis in Hokkaido sika deer (Cervus nippon yesoensis) from Hokkaido Prefecture,Japan revealed by ELISA using recombinant cathepsin L1

Fasciolosis, a zoonotic disease caused by liver flukes of the genus Fasciola, has been reported in Hokkaido (Yezo) sika deer (Cervus nippon yesoensis) in Hokkaido Prefecture, Japan; however, the actual seroprevalence in the animal has not been adequately evaluated. The objective of the present study was to analyze the seroprevalence of the disease among Hokkaido sika deer. Recombinant cathepsin L1 (rCatL1) was used as an antigen for an indirect enzyme-linked immunosorbent assay (ELISA) to detect antibodies against Fasciola flukes. The sensitivity and specificity of the ELISA were 84.6% and 100%, respectively. The average seroprevalence in 1109 Hokkaido sika deer from 20 locations in Hokkaido Prefecture was 43.9%. Mature deer showed higher seroprevalence than younger individuals; however, even younger animals may act as a reservoir for the disease. Monitoring infection levels in the Hokkaido sika deer population is important not only for the livestock industry, but also for preventing human fasciolosis.     

Occurrence and molecular characterization of Cryptosporidium spp. and Giardia duodenalis among captive mammals in the Bangladesh National Zoo

Cryptosporidium and Giardia are protozoan parasites capable of causing gastrointestinal illness in humans and animals. The purpose of this research was to determine the occurrence, genetic characteristics, and zoonotic potential of Cryptosporidium spp. and Giardia duodenalis in captive mammals at the Bangladesh National Zoo. A total of 200 fresh fecal samples from 32 mammalian species were collected and examined for Cryptosporidium spp. using nested polymerase chain reaction (PCR) targeting the small subunit (SSU) rRNA gene and G. duodenalis targeting the β-giardin (bg), glutamate dehydrogenase (gdh), and triosephosphate isomerase (tpi) genes. The overall infection rates of Cryptosporidium and G. duodenalis among captive mammals in the zoo were 3.5% (7/200) and 5.5% (11/200), respectively. Five species/genotypes of Cryptosporidium (C. hominis, C. andersoni, C. muris, C. felis, and Cryptosporidium deer genotype) were identified. C. hominis was subtyped as IbA12G3 by sequence analysis of the glycoprotein 60 (gp60) gene. Multilocus genotyping of G. duodenalis revealed assemblages A, B, and D. Mixed infections of assemblages B and D and A and B were found in an Asiatic jackal and a Nilgiri langur, respectively. To our knowledge, this is the first report on the occurrence and genetic identity of the two parasites among zoo animals in Bangladesh. The results suggest that zoonotic Cryptosporidium spp. and G. duodenalis are maintained in and transmitted between captive mammals. Therefore, washing, cleaning, and disinfection measures should be implemented to reduce the spread of Cryptosporidium and G. duodenalis infections.     

Advances in molecular epidemiology of cryptosporidiosis in dogs and cats

The use of molecular tools has led to the identification of several zoonotic Cryptosporidium spp. in dogs and cats. Among them, Cryptosporidium canis and Cryptosporidium felis are dominant species causing canine and feline cryptosporidiosis, respectively. Some Cryptosporidium parvum infections have also been identified in both groups of animals. The identification of C. canis, C. felis and C. parvum in both pets and owners suggests the possible occurrence of zoonotic transmission of Cryptosporidium spp. between humans and pets. However, few cases of such concurrent infections have been reported. Thus, the cross-species transmission of Cryptosporidium spp. between dogs or cats and humans has long been a controversial issue. Recently developed subtyping tools for C. canis and C. felis should be very useful in identification of zoonotic transmission of both Cryptosporidium spp. Data generated using these tools have confirmed the occurrence of zoonotic transmission of these two Cryptosporidium spp. between owners and their pets, but have also shown the potential presence of host-adapted subtypes. Extensive usage of these subtyping tools in epidemiological studies of human cryptosporidiosis is needed for improved understanding of the importance of zoonotic transmission of Cryptosporidium spp. from pets.     

Novel Cryptosporidium Genotype in Wild Australian Mice (Mus domesticus)

A total of 250 mouse fecal specimens collected from crop farms in Queensland, Australia, were screened for the presence of Cryptosporidium spp. using PCR. Of these, 19 positives were detected and characterized at a number of loci, including the 18S rRNA gene, the acetyl coenzyme A gene, and the actin gene. Sequence and phylogenetic analyses identified two genotypes: mouse genotype I and a novel genotype (mouse genotype II), which is likely to be a valid species. Cryptosporidium parvum, which is zoonotic, was not detected. The results of the study indicate that wild Australian mice that are not in close contact with livestock are probably not an important reservoir of Cryptosporidium infection for humans and other animals.     

Spatiotemporal Analysis of Cryptosporidium Species/Genotypes and Relationships with Other Zoonotic Pathogens in Surface Water from Mixed-Use Watersheds

Nearly 690 raw surface water samples were collected during a 6-year period from multiple watersheds in the South Nation River basin, Ontario, Canada. Cryptosporidium oocysts in water samples were enumerated, sequenced, and genotyped by detailed phylogenetic analysis. The resulting species and genotypes were assigned to broad, known host and human infection risk classes. Wildlife/unknown, livestock, avian, and human host classes occurred in 21, 13, 3, and <1% of sampled surface waters, respectively. Cryptosporidium andersoni was the most commonly detected livestock species, while muskrat I and II genotypes were the most dominant wildlife genotypes. The presence of Giardia spp., Salmonella spp., Campylobacter spp., and Escherichia coli O157:H7 was evaluated in all water samples. The greatest significant odds ratios (odds of pathogen presence when host class is present/odds of pathogen presence when host class is absent) for Giardia spp., Campylobacter spp., and Salmonella spp. in water were associated, respectively, with livestock (odds ratio of 3.1), avian (4.3), and livestock (9.3) host classes. Classification and regression tree analyses (CART) were used to group generalized host and human infection risk classes on the basis of a broad range of environmental and land use variables while tracking cooccurrence of zoonotic pathogens in these groupings. The occurrence of livestock-associated Cryptosporidium was most strongly related to agricultural water pollution in the fall (conditions also associated with elevated odds ratios of other zoonotic pathogens occurring in water in relation to all sampling conditions), whereas wildlife/unknown sources of Cryptosporidium were geospatially associated with smaller watercourses where urban/rural development was relatively lower. Conditions that support wildlife may not necessarily increase overall human infection risks associated with Cryptosporidium since most Cryptosporidium genotypes classed as wildlife in this study (e.g., muskrat I and II genotype) do not pose significant infection risks to humans. Consequently, from a human health perspective, land use practices in agricultural watersheds that create opportunities for wildlife to flourish should not be rejected solely on the basis of their potential to increase relative proportions of wildlife fecal contamination in surface water. The present study suggests that mitigating livestock fecal pollution in surface water in this region would likely reduce human infection risks associated with Cryptosporidium and other zoonotic pathogens.     

Tracking Host Sources of Cryptosporidium spp. in Raw Water for Improved Health Risk Assessment

Recent molecular evidence suggests that different species and/or genotypes of Cryptosporidium display strong host specificity, altering our perceptions regarding the zoonotic potential of this parasite. Molecular forensic profiling of the small-subunit rRNA gene from oocysts enumerated on microscope slides by U.S. Environmental Protection Agency method 1623 was used to identify the range and prevalence of Cryptosporidium species and genotypes in the South Nation watershed in Ontario, Canada. Fourteen sites within the watershed were monitored weekly for 10 weeks to assess the occurrence, molecular composition, and host sources of Cryptosporidium parasites impacting water within the region. Cryptosporidium andersoni, Cryptosporidium muskrat genotype II, Cryptosporidium cervine genotype, C. baileyi, C. parvum, Cryptosporidium muskrat genotype I, the Cryptosporidium fox genotype, genotype W1, and genotype W12 were detected in the watershed. The molecular composition of the Cryptosporidium parasites, supported by general land use analysis, indicated that mature cattle were likely the main source of contamination of the watershed. Deer, muskrats, voles, birds, and other wildlife species, in addition to sewage (human or agricultural) may also potentially impact water quality within the study area. Source water protection studies that use land use analysis with molecular genotyping of Cryptosporidium parasites may provide a more robust source-tracking tool to characterize fecal impacts in a watershed. Moreover, the information is vital for assessing environmental and human health risks posed by water contaminated with zoonotic and/or anthroponotic forms of Cryptosporidium.     

Prevalence,Environmental Loading,and Molecular Characterization of Cryptosporidium and Giardia Isolates from Domestic and Wild Animals along the Central California Coast

The risk of disease transmission from waterborne protozoa is often dependent on the origin (e.g., domestic animals versus wildlife), overall parasite load in contaminated waterways, and parasite genotype, with infections being linked to runoff or direct deposition of domestic animal and wildlife feces. Fecal samples collected from domestic animals and wildlife along the central California coast were screened to (i) compare the prevalence and associated risk factors for fecal shedding of Cryptosporidium and Giardia species parasites, (ii) evaluate the relative importance of animal host groups that contribute to pathogen loading in coastal ecosystems, and (iii) characterize zoonotic and host-specific genotypes. Overall, 6% of fecal samples tested during 2007 to 2010 were positive for Cryptosporidium oocysts and 15% were positive for Giardia cysts. Animal host group and age class were significantly associated with detection of Cryptosporidium and Giardia parasites in animal feces. Fecal loading analysis revealed that infected beef cattle potentially contribute the greatest parasite load relative to other host groups, followed by wild canids. Beef cattle, however, shed host-specific, minimally zoonotic Cryptosporidium and Giardia duodenalis genotypes, whereas wild canids shed potentially zoonotic genotypes, including G. duodenalis assemblages A and B. Given that the parasite genotypes detected in cattle were not zoonotic, the public health risk posed by protozoan parasite shedding in cattle feces may be lower than that posed by other animals, such as wild canids, that routinely shed zoonotic genotypes.     

A whole water catchment approach to investigating the origin and distribution of Cryptosporidium species

Aims: Investigating the distribution and origin of Cryptosporidium species in a water catchment affected by destocking and restocking of livestock as a result of a foot and mouth disease epidemic. Methods and Results: Surface water, livestock and wildlife samples were screened for Cryptosporidium and oocysts characterised by sequencing SSU rRNA and COWP loci, and fragment analysis of ML1, ML2 and GP60 microsatellite loci. Oocyst concentrations in water samples (0–20·29 per 10 l) were related to rainfall events, amount of rainfall and topography. There was no detectable impact from catchment restocking. Cryptosporidium spp. found in water were indicative of livestock (Cryptosporidium andersoni and Cryptosporidium parvum) and wildlife (novel genotypes) sources. However, C. andersoni was not found in any animals sampled. Calf infections were age related; C. parvum was significantly more common in younger animals (<4 weeks old). Older calves shared Cryptosporidium bovis, Cryptosporidium ryanae and C. parvum. Wildlife shed C. parvum, Cryptosporidium ubiquitum, muskrat genotype II and deer genotype. Conclusions: Several factors affect the occurrence of Cryptosporidium within a catchment. In addition to farmed and wild animal hosts, topography and rainfall patterns are particularly important. Significance and Impact of the Study: These factors must be considered when undertaking risk‐based water safety plans.     

Cryptosporidium parvum in brown brocket (Mazama gouazoubira) from Brazil: First report of the subtype IIaA16G3R1 in cervids

This research had as objective to evaluate the occurrence and to characterize genetically the infections by Cryptosporidium in Mazama gouazoubira. By a non-invasive harvest methodology using trained sniffer dogs to locate fecal samples of cervids, 642 fecal samples were obtained from six Brazilian localities. The cervids species responsible for the excretion of each fecal sample were identified by the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP), using the mitochondrial cytochrome b target gene (cyst b) and the restriction enzymes Sspl, AflIII and BstN. From this identification, 437 fecal samples of M. gouazoubira were selected for research of Cryptosporidium spp. performed through negative staining with malachite green and polymerase chain reaction (nPCR), with the subunit of 18S rRNA gene, followed by sequencing the amplified products. In the samples that were diagnosed the presence of parasite species with zoonotic potential, genotyping was also performed using nPCR with the subunit of GP60 gene. Statistical analysis consisted of the Fisher exact test to verify the association of the presence of the enteroparasite in relation to the presence of cattle in each locality, and the McNemar tests and Kappa correlation coefficient used to compare the results obtained between the two diagnostic techniques. In the fecal samples of M. gouazoubira the occurrences of Cryptosporidium were diagnosed in 1.6% (7/437) and 1.1% (5/437), respectively, through nPCR and microscopy. Cryptosporidium. parvum was diagnosed in 100% (7/7) of the samples submitted to sequencing (18S gene). The IIaA16G3R1 subtype was diagnosed in five of the C. parvum samples submitted to genotyping (GP60 gene). This is the first world report of C. parvum in M. gouazoubira and subtype IIaA16G3R1 in cervids.     

Molecular detection of Giardia duodenalis and Cryptosporidium spp. from stray dogs residing in monasteries in Bangkok,Thailand

Both Cryptosporidium spp. and Giardia duodenalis are enteric protozoan parasites that infect a wide variety of domestic animals as well as humans worldwide, causing diarrheal diseases. Giardia duodenalis assemblages C and D are specific to canine hosts and zoonotic assemblages A and B are also found in dogs as a reservoir host. In dogs, Cryptosporidium canis is the host-specific species while humans are infected by C. hominis and C. parvum and at least another 16 zoonotic Cryptosporidium species have been reported causing human infections, with C. meleagridis, C. viatorum, and C. ubiquitum being the most frequent. The objective of this study was to determine the prevalence of Cryptosporidium spp. and G. duodenalis from stray dogs in areas of Bangkok and to identify the species and assemblages. Fecal samples (540) were collected from dogs residing in 95 monasteries in 48 districts in the Bangkok metropolitan area. Nested Polymerase Chain Reaction (PCR) was performed using the ssu-rRNA gene for both parasites. In total, 3.0% (16/540) samples were positive for G. duodenalis, with most being G. duodenalis assemblage D (7/16) followed by assemblage C (7/16) and zoonotic assemblage A (2/16). The prevalence of Cryptosporidium spp. was 0.7% (4/540) based on the PCR results and all were the dog genotype C. canis. These results indicated that dogs residing in Bangkok monasteries poses a limited role as source of human giardiosis and cryptosporidiosis.     

High prevalence of Cryptosporidium infection caused by C. scrofarum and C. suis among pigs in Thailand

Cryptosporidium spp. is an important intestinal protozoan causing diarrhea among both healthy and immunocompromised patients especially those with HIV/AIDS. Cryptosporidium spp. can be transmitted via foodborne, waterborne and person-to-person routes. In addition, several Cryptosporidium species are zoonotic. This study aimed to determine the prevalence of Cryptosporidium infection among pigs raised in both smallholder (<50 heads/farm) and large scale farms (50–500 heads/farm) in Chonburi Province, eastern Thailand using nested PCR amplifying the small subunit of the ribosomal RNA (SSU-rRNA) gene. DNA sequencing was also performed to identify the species of Cryptosporidium. A total of 245 fecal samples were collected from 11 pig farms. The overall prevalence of Cryptosporidium infection was 20.8% (51/245) which were found in both smallholder and small large scale pig farms. The prevalence of Cryptosporidium infection among pigs aged ≤6 months was significantly higher than those aged >6 months (p < .001). Among 51 Cryptosporidium positive samples, Cryptosporidium scrofarum (42/51, 82.4%) and Cryptosporidium suis (9/51, 17.6%) were identified. The prevalence of C. scrofarum infection observed among pigs aged ≤6 months was significantly higher when compared with those aged >6 months (20.7% and 2.1%, respectively, p < .001). The high prevalence of C. scrofarum and C. suis infections among pigs could be a potential source of infection to humans.     

Helminthes detected from wild sika deer (Cervus nippon centralis) in Kanto-Chubu region,Japan

The helminth fauna of 105 sika deer (Cervus nippon centralis) captured in Yamanashi, Kanagawa and Nagano Prefectures, Japan was investigated during 2014–2019. As a result, 12 helminthes, i.e. 3 digeneans (Ogmocotyle sikae, Dicrocoelium chinensis and D. dendriticum), 8 nematodes (Gongylonema pulchrum, Dictyocaulus sp., Pygarginema sp., Spiculopteragia houdemeri, Chabaudstrongylus ninhae, Trichuris discolor, Oesophagostomum sikae and Oes. asperum), and 1 cestode (Moniezia sp.) were detected. To our knowledge, this is the first report of Pygarginema sp., Cha. ninhae, and Oes. asperum from sika deer in Japan. Some helminthes detected in the present study can infect livestock. Considering the possibility of the spread of the helminthes to livestock through deer excrement, it is important to promote understanding the parasite fauna in wild deer.     

Immunoenzymatic analysis and genetic detection of Cryptosporidium parvum in lambs from Italy

Cryptosporidiosis is a worldwide-diffused protozoan disease causing important economic losses to animal husbandry and livestock production. Additionally, several species/genotypes of Cryptosporidium have a relevant zoonotic potential and ruminants may be important sources of infection for human beings. Nonetheless, in Europe, little is known of the presence of Cryptosporidium in sheep nor on the species/genotypes involved. To obtain information on the occurrence of cryptosporidiosis in lambs and the potential zoonotic role of the Cryptosporidium isolates, one hundred and forty-nine faecal samples individually collected from lambs in central Italy have been examined for the presence of Cryptosporidium. All faecal specimens were processed with a commercial ELISA kit immunoassay and all ELISA-positive samples were further analyzed genetically. Twenty-six ELISA-positive samples scored positive at the PCR and the sequences obtained displayed 100% identity with the zoonotic Cryptosporidum parvum. This work suggests for the first time that lambs in Italy may shed C. parvum, thus representing a potential public health hazard.     

First Molecular Characterization of Cryptosporidium spp. Infecting Buffalo Calves in Brazil

With the aim of determining the occurrence of Cryptosporidium spp., 222 fecal samples were collected from Murrah buffalo calves aged up to 6 mo. Fecal DNA was genotyped with a nested polymerase chain reaction targeting the 18S rRNA gene and sequencing of the amplified fragment. Nested 18S PCR was positive for 48.2% of the samples. Sequence analysis showed that the most frequent species in these animals was Cryptosporidium ryanae, which was present in buffalo calves as young as 5 d. The zoonotic species Cryptosporidium parvum was detected in one animal. An uncommon Cryptosporidium 18S genotype was found in buffaloes.     

The first reported cases of human cryptosporidiosis caused by Cryptosporidium hominis in Slovak Republic

Cryptosporidiosis belongs to the important parasitic infections with zoonotic potential and the occurrence in European countries is rare. The first cases of cryptosporidiosis caused by Cryptosporidium hominis detected in the Slovak republic were described here. Collection of examined humans consisted of five family members. Faecal specimens were examined by formalin sedimentation, by the Sheather??s sugar flotation and by immunochromatography and visualised by the Ziehl?CNeelsen acid fast stain. A fragment of the Cryptosporidium small subunit ribosomal RNA gene was amplified by nested polymerase chain reaction and species was determined by restriction fragment length polymorphism analysis with the endonucleases SspI and VspI. C. hominis was found in faeces of two immunocompetent siblings (a 7-year-old boy and a 2-year-old girl). The symptoms occurred only in the boy as gastrointestinal disorders lasting 5?days, and manifested by abdominal pain, an elevated body temperature (37.2?°C), mild diarrhoea, accompanied by lassitude, depression and anorexia. Ultrasonic scan revealed enlarged spleen and mezenteric lymph nodes. Microscopic examination of the stool sample revealed numerous Cryptosporidium oocysts. The DNA typing identified C. hominis subtype IbA10G2. Cryptosporidium was also detected in the boy??s sister without any complications and symptoms. Their father, mother and grandmother were parasitologically negative. The source of infection remained unknown. Human cases in present study reflect necessity of systematic attention on intestinal parasites diagnostic inclusive of cryptosporidia.     

Generalist nematodes dominate the nemabiome of roe deer in sympatry with sheep at a regional level

The growth of livestock farming and the recent expansion of wild ungulate populations in Europe favor opportunities for direct and/or indirect cross-transmission of pathogens. Comparatively few studies have investigated the epidemiology of gastro-intestinal nematode parasites, an ubiquitous and important community of parasites of ungulates, at the wildlife/livestock interface. In this study, we aimed to assess the influence of livestock proximity on the gastrointestinal nematode community of roe deer in a rural landscape located in southern France. Using ITS-2 rDNA nemabiome metabarcoding on fecal larvae, we analysed the gastrointestinal nematode communities of roe deer and sheep. In addition, we investigated Haemonchus contortus nad4 mtDNA diversity to specifically test parasite circulation among domestic and wild host populations. The dominant gastrointestinal nematode species found in both the roe deer and sheep were generalist species commonly found in small ruminant livestock (e.g. H. contortus), whereas the more specialised wild cervid nematode species (e.g. Ostertagia leptospicularis) were only present at low frequencies. This is in marked contrast with previous studies that found the nemabiomes of wild cervid populations to be dominated by cervid specialist nematode species. In addition, the lack of genetic structure of the nad4 mtDNA of H. contortus populations between host species suggests circulation of gastrointestinal nematodes between roe deer and sheep. The risk of contact with livestock only has a small influence on the nemabiome of roe deer, suggesting the parasite population of roe deer has been displaced by generalist livestock parasites due to many decades of sheep farming, not only for deer grazing close to pastures, but also at a larger regional scale. We also observed some seasonal variation in the nemabiome composition of roe deer. Overall, our results demonstrate significant exchange of gastrointestinal nematodes between domestic and wild ungulates, with generalist species spilling over from domestic ungulates dominating wild cervid parasite communities.     

Molecular detection of tick-borne protozoan parasites in sika deer (Cervus nippon) from western regions of Japan

The sika deer (Cervus nippon) is one of the most common species of wildlife in Japan. This study aimed to reveal the prevalence of tick-borne protozoan parasites in wild sika deer living in western Japan. We used nested polymerase chain reaction (PCR) to detect the 18S rRNA gene of tick-borne apicomplexan parasites (Babesia, Theileria, and Hepatozoon spp.) from 276 blood and liver samples from sika deer captured in the Yamaguchi, Oita, Kagoshima, Okayama, Ehime, Kochi, and Tokushima Prefectures. In total, 259 samples (259/276; 93.8%) tested positive in the nested PCR screening. Gene sequencing revealed that 99.6% (258/259) of positive samples contained Theileria sp. (sika 1), while Theileria sp. (sika 2), another Theileria species, was detected in only 3 samples. We also found that one sample from a sika deer captured in Kagoshima contained the gene of an unidentified Babesia sp. related to Babesia sp. Kh-Hj42, which was previously collected from tick in western Siberia. In conclusion, we found a high prevalence of piroplasms in sika deer from western Japan, and DNA analysis revealed that Theileria sp. (sika 1) had the highest infection rate.     

Genetic Diversity of Cryptosporidium spp. in Captive Reptiles

The genetic diversity of Cryptosporidium in reptiles was analyzed by PCR-restriction fragment length polymorphism and sequence analysis of the small subunit rRNA gene. A total of 123 samples were analyzed, of which 48 snake samples, 24 lizard samples, and 3 tortoise samples were positive for Cryptosporidium. Nine different types of Cryptosporidium were found, including Cryptosporidium serpentis, Cryptosporidium desert monitor genotype, Cryptosporidium muris, Cryptosporidium parvum bovine and mouse genotypes, one C. serpentis-like parasite in a lizard, two new Cryptosporidium spp. in snakes, and one new Cryptosporidium sp. in tortoises. C. serpentis and the desert monitor genotype were the most common parasites and were found in both snakes and lizards, whereas the C. muris and C. parvum parasites detected were probably the result of ingestion of infected rodents. Sequence and biologic characterizations indicated that the desert monitor genotype was Cryptosporidium saurophilum. Two host-adapted C. serpentis genotypes were found in snakes and lizards.