Molecular Characterization of Cryptosporidium spp., Giardia duodenalis,and Enterocytozoon bieneusi in Captive Wildlife at Zhengzhou Zoo,China

Cryptosporidium spp., Giardia duodenalis, and Enterocytozoon bieneusi are common gastrointestinal protists in humans and animals. Two hundred and three fecal specimens from 80 wildlife species were collected in Zhengzhou Zoo and their genomic DNA extracted. Three intestinal pathogens were characterized with a DNA sequence analysis of different loci. Cryptosporidium felis, C. baileyi, and avian genotype III were identified in three specimens (1.5%), the manul, red‐crowned crane, and cockatiel, respectively. Giardia duodenalis was also found in five specimens (2.5%) firstly: assemblage B in a white‐cheeked gibbon and beaver, and assemblage F in a Chinese leopard and two Siberian tigers, respectively. Thirteen genotypes of E. bieneusi (seven previously reported genotypes and six new genotypes) were detected in 32 specimens (15.8%), of which most were reported for the first time. A phylogenetic analysis of E. bieneusi showed that five genotypes (three known and two new) clustered in group 1; three known genotypes clustered in group 2; one known genotype clustered in group 4; and the remaining four genotypes clustered in a new group. In conclusion, zoonotic Cryptosporidium spp., G. duodenalis, and E. bieneusi are maintained in wildlife and transmitted between them. Zoonotic disease outbreaks of these infectious agents possibly originate in wildlife reservoirs.     

Molecular Surveillance of Cryptosporidium spp., Giardia duodenalis, and Enterocytozoon bieneusi by Genotyping and Subtyping Parasites in Wastewater

Background

Despite their wide occurrence, cryptosporidiosis and giardiasis are considered neglected diseases by the World Health Organization. The epidemiology of these diseases and microsporidiosis in humans in developing countries is poorly understood. The high concentration of pathogens in raw sewage makes the characterization of the transmission of these pathogens simple through the genotype and subtype analysis of a small number of samples.

Methodology/Principal Findings

The distribution of genotypes and subtypes of Cryptosporidium spp., Giardia duodenalis, and Enterocytozoon bieneusi in 386 samples of combined sewer systems from Shanghai, Nanjing and Wuhan and the sewer system in Qingdao in China was determined using PCR-sequencing tools. Eimeria spp. were also genotyped to assess the contribution of domestic animals to Cryptosporidium spp., G. duodenalis, and E. bieneusi in wastewater. The high occurrence of Cryptosporidium spp. (56.2%), G. duodenalis (82.6%), E. bieneusi (87.6%), and Eimeria/Cyclospora (80.3%) made the source attribution possible. As expected, several human-pathogenic species/genotypes, including Cryptosporidium hominis, Cryptosporidium meleagridis, G. duodenalis sub-assemblage A-II, and E. bieneusi genotype D, were the dominant parasites in wastewater. In addition to humans, the common presence of Cryptosporidium spp. and Eimeria spp. from rodents indicated that rodents might have contributed to the occurrence of E. bieneusi genotype D in samples. Likewise, the finding of Eimeria spp. and Cryptosporidium baileyi from birds indicated that C. meleagridis might be of both human and bird origins.

Conclusions/Significance

The distribution of Cryptosporidium species, G. duodenalis genotypes and subtypes, and E. bieneusi genotypes in urban wastewater indicates that anthroponotic transmission appeared to be important in epidemiology of cryptosporidiosis, giardiasis, and microsporidiosis in the study areas. The finding of different distributions of subtypes between Shanghai and Wuhan was indicative of possible differences in the source of C. hominis among different areas in China.     

Occurrence of human-pathogenic Enterocytozoon bieneusi,Giardia duodenalis and Cryptosporidium genotypes in laboratory macaques in Guangxi,China

Captive nonhuman primates have been identified as common hosts of Enterocytozoon bieneusi, Giardia duodenalis, Cryptosporidium hominis, and Cyclospora spp., thus are potential reservoirs of some enteric parasites in humans. However, few studies have examined the source and human-infective potential of enteric parasites in laboratory nonhuman primates. In the present work, 205 fecal specimens were collected from three groups of captive Macaca fascicularis kept in different densities in a laboratory animal facility in Guangxi, China, and examined by PCR for E. bieneusi, G. duodenalis, Cryptosporidium spp., and Cyclospora spp. The infection rates of E. bieneusi and G. duodenalis were 11.3% and 1.2% in Group 1 (young animals kept individually; n = 168), 72.2% and 11.1% in Group 2 (young animals kept in groups; n = 18), and 31.6% and 5.3% in Group 3 (adults kept in groups; n = 19), respectively. Sequence analysis of PCR products showed the presence of five E. bieneusi genotypes, with genotype D (in 16/36 genotyped specimens) and a new genotype (in 15/36 genotyped specimens) as the dominant genotypes. All five E. bieneusi genotypes belonged to the zoonotic group (Group 1). The G. duodenalis genotypes (assemblages AII and B) in five specimens and C. hominis subtype (IdA14) in one specimen were also known human-pathogens, although the Cyclospora seen in one animal appeared to be unique to macaque monkeys. The higher infection rate in younger animals reared in groups and common occurrence of zoonotic genotypes indicated that human-pathogenic E. bieneusi could be transmitted efficiently in captive nonhuman primates, and group-housing was a risk factor for transmission of zoonotic pathogens in young nonhuman primates in research facilities.     

Molecular Identification of Cryptosporidium spp., Enterocytozoon bieneusi,and Giardia duodenalis in Captive Pet Birds in Henan Province,Central China

Cryptosporidium spp., Enterocytozoon bieneusi, and Giardia duodenalis are common enteric pathogens that are capable of infecting humans and animals. Total of 1,005 fecal samples from captive pet birds were collected from seven locations in Henan Province, China. The results demonstrated that 9.9% (99/1,005) of the captive birds were infected with one of these three pathogens. Enterocytozoon bieneusi was the most prevalent species among the birds (45/1,005, 4.5%) followed by G. duodenalis (33/1,005, 3.3%) and Cryptosporidium spp. (21/1,005, 2.1%). Five Cryptosporidium species were identified, namely, C. baileyi (10), C. galli (5), C. meleagridis (4), C. andersoni (1), and C. parvum (1). Two known E. bieneusi genotypes were identified: Peru 6 (44) was identified in pigeons (34) and European turtle doves (10); whereas, the genotype PtEb I (1) was only identified in a pigeon. Only G. duodenalis assemblage E (33) was identified in some pet birds. To the best of our knowledge, this study is the first to undertake the molecular identification of G. duodenalis in birds in China. The identification of potentially zoonotic species/genotypes of the pathogens suggests that exposure to the excreta of these birds, either directly or via food and water, may pose a threat to human health.     

Enterocytozoon bieneusi Genotypes in Yaks (Bos grunniens) and Their Public Health Potential

Enterocytozoon bieneusi, the most frequently diagnosed microsporidian species in humans, is also identified in a wide range of animals. To date, few data are available on E. bieneusi in yaks (Bos grunniens). In this study, we examined the occurrence and genotype identity of E. bieneusi in yaks in four counties in Qinghai Province of China. Of 327 fecal specimens examined by nested PCR analysis of the ribosomal internal transcribed spacer, 23 (7.0%) were E. bieneusi‐positive. DNA sequence analysis of the PCR products revealed the presence of five distinct genotypes: three Group 2 genotypes previously reported in cattle as well as humans (BEB4, I and J) and two novel genotypes (CHN11 and CHN12) belonging to the large zoonotic group (Group 1). Data of the study suggest that these animals could be potential reservoirs for human E. bieneusi infection.     

Molecular Characterization of Enterocytozoon bieneusi in Wild Carnivores in Spain

Microsporidia comprises a diverse group of obligate intracellular parasites that infect a broad range of invertebrates and vertebrates. Among Microsporidia, Enterocytozoon bieneusi is the most frequently detected species in humans and animals worldwide bringing into question the possible role of animal reservoirs in the epidemiology of this pathogen. Although E. bieneusi is an emerging zoonotic pathogen able to infect many domestic and wild mammals that could act as reservoir of infection for humans and other animals, only few studies have documented its occurrence in wild carnivores. To determine the occurrence of E. bieneusi in wild carnivores, we examined 190 wild carnivores collected from different locations in Spain. Twenty‐five fecal samples (13.2%) from three host species (European badger, beech marten, and red fox) were E. bieneusi‐positive by PCR. Nucleotide sequence analysis of the ITS region revealed a high degree of genetic diversity with a total of eight distinct genotypes including four known (PtEbIX, S5, S9, and WildBoar3) and four novel (EbCar1‐EbCar4) genotypes identified. Phylogenetic analysis showed that the four novel genotypes (EbCar1‐EbCar4), S5, S9, and WildBoar3 clustered within the previously designated zoonotic Group 1. Our results demonstrate that human‐pathogenic genotypes are present in wild carnivores, corroborating their potential role as a source of human infection and environmental contamination.     

Genotypes of Cryptosporidium spp. and Enterocytozoon bieneusi in Human Immunodeficiency Virus‐Infected Patients in Lagos,Nigeria

Molecular characterization of Cryptosporidium spp. and Enterocytozoon bieneusi has improved our understanding of the transmission of both organisms in humans. In this study, to infer possible infection sources, Cryptosporidium spp. and E. bieneusi in fecal specimens from 90 HIV‐infected patients attending antiretroviral clinics in Lagos, Nigeria were detected and genotyped by PCR and DNA sequencing. Cryptosporidium spp. and E. bieneusi were identified in four and five patients, respectively, including the occurrence of subtype IeA11T3G3 of Cryptosporidium hominis in two patients, subtype IIcA5G3k of Cryptosporidium parvum in one patient, and Type IV of E. bieneusi in four patients. Among the remaining positive patients, one had mixed infection of Cryptosporidium meleagridis and C. hominis and one had mixed E. bieneusi genotypes. These data highlight a possible difference in major transmission routes (anthroponotic vs. zoonotic) between Cryptosporidium spp. and E. bieneusi in HIV+ patients in the study area.     

Genetic characterizations of Cryptosporidium spp. and Giardia duodenalis in humans in Henan, China

Cryptosporidium and Giardia infections are common causes of diarrhea worldwide. To better understand the transmission of human cryptosporidiosis and giardiasis in Henan, China, 10 Cryptosporidium-positive specimens and 18 Giardia-positive specimens were characterized at the species/genotype and subtype levels. Cryptosporidium specimens were analyzed by DNA sequencing of the small subunit rRNA and 60 kDa glycoprotein genes. Among those genotyped, nine belonged to C. hominis and one C. felis, with the former belonging to three subtype families: Ia, Ib, and Id. The three Ib subtypes identified, IbA16G2, IbA19G2, and IbA20G2, were very different from the two common Ib subtypes (IbA9G3 and IbA10G2) found in other areas of the world. The distribution of Giardia duodenalis genotypes and subtypes was assessed by sequence analysis of the triosephosphate isomerase (tpi) gene. The assemblages A (eight belonging to A-I and four A-II) and B (belonging to six new subtypes) were found in 12 and six specimens, respectively. More systematic studies are needed to understand the transmission of Cryptosporidium and G. duodenalis in humans in China.     

Cryptosporidium spp., Giardia duodenalis,Enterocytozoon bieneusi and Other Intestinal Parasites in Young Children in Lobata Province,Democratic Republic of S?o Tomé and Principe

Rare systemic studies concerning prevalence of intestinal parasites in children have been conducted in the second smallest country in Africa, the Democratic Republic of São Tomé and Príncipe. Fecal specimens from 348 children (214 in-hospital attending the Aires de Menezes Hospital and 134 from Agostinho Neto village) in São Tome Island were studied by parasitological and molecular methods. Of the 134 children from Agostinho Neto, 52.2% presented intestinal parasites. 32.1% and 20.2% of these children had monoparasitism and polyparasitism, respectively. Ascaris lumbricoides (27.6%), G. duodenalis (7.5%), T. trichiura (4.5%) and Entamoeba coli (10.5%) were the more frequent species identified in the children of this village. Giardia duodenalis (7.5%) and E. bieneusi (5.2%) were identified by PCR. Nested-PCR targeting G. duodenalis TPI identified Assemblage A (60%) and Assemblage B (40%). The E. bieneusi ITS-based sequence identified genotypes K (57.1%), KIN1 (28.6%) and KIN3 (14.3%). Among the 214 in-hospital children, 29.4% presented intestinal parasites. In 22.4% and 7.0% of the parasitized children, respectively, one or more species were concurrently detected. By microscopy, A. lumbricoides (10.3%) and Trichiuris trichiura (6.5%) were the most prevalent species among these children, and Cryptosporidium was detected by PCR in 8.9% of children. GP60 locus analysis identified 6.5% of C. hominis (subtypes IaA27R3 [35.7%], IaA23R3 [14.3%], IeA11G3T3 [28.6%] and IeA11G3T3R1 [21.4%]) and 2.3% of C. parvum (subtypes IIaA16G2R1 [20.0%], IIaA15G2R1 [20.0%], IIdA26G1 [40.0%] and IIdA21G1a [20.0%]). G. duodenalis and E. bieneusi were identified in 0.5% and 8.9% of the in-hospital children, respectively. G. duodenalis Assemblage B was characterized. The E. bieneusi genotypes K (52.6%), D (26.4%), A (10.5%) and KIN1 (10.5%) were identified. Although further studies are required to clarify the epidemiology of these infectious diseases in this endemic region the significance of the present results highlights that it is crucial to strength surveillance on intestinal pathogens.     

Evidence for Zoonotic Potential of Enterocytozoon bieneusi in Its First Molecular Characterization in Captive Mammals at Bangladesh National Zoo

To determine the occurrence and genotypes of Enterocytozoon bieneusi in captive mammals at Bangladesh National Zoo and to assess their zoonotic significance, 200 fecal samples from 32 mammalian species were examined using a nested PCR and sequencing of internal transcribed spacer (ITS) gene. Enterocytozoon bieneusi was detected in 16.5% (33/200) of the samples. Seven different ITS genotypes were identified, including two known genotypes (D and J) and five new ones (BAN4 to BAN8). Genotype D was the most common genotype being observed in 19 isolates. In phylogenetic analysis, four genotypes (D, BAN4, BAN5, and BAN6), detected in 30 isolates (90.9%), belonged to Group 1 having zoonotic potential. The sequence of genotype J found in a Malayan pangolin was clustered in so‐called ruminant‐specific Group 2. The other two genotypes BAN7 and BAN8 were clustered in primate‐specific Group 5. To our knowledge, this is the first report of molecular characterization of E. bieneusi in Bangladesh, particularly in captive‐bred wildlife in this country. The potentially zoonotic genotypes of E. bieneusi are maintained in zoo mammals that may transmit among these animals and to the humans through environmental contamination or contact.     

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.     

Molecular characterization of Cryptosporidium spp. and Giardia duodenalis in experimental rats in China

Cryptosporidium and Giardia are ubiquitous protozoan parasites that infect a broad range of hosts. The presence of Cryptosporidium spp. and G. duodenalis was detected in 355 fecal samples of laboratory experimental rats from four experimental rat rearing facilities in China by PCR amplification of the small subunit (SSU) rRNA gene. The G. duodenalis positive samples were further characterized in the β-giardin (bg), glutamate dehydrogenase (gdh), and triosephosphate isomerase (tpi) genes. The overall infection rates of Cryptosporidium spp. and G. duodenalis were 0.6% (2/355) and 9.3% (33/355), respectively, with no co-infection. Among the four facilities, only the rats in Zhengzhou1 were found positive for the two pathogens. Undetermined Cryptosporidium genotype was observed in one sample and C. ubiquitum in another sample. Assemblage G was identified in all the 33 G. duodenalis positive isolates at SSU rRNA gene, out of which 19, 20, and 21 isolates were also subtyped as assemblage G at tpi, gdh and bg gens, respectively. To our knowledge, this is the first report of Cryptosporidium and G. duodenalis infections in laboratory experimental rats in China. The infections of these pathogens in laboratory animals should be monitored routinely since they may interfere the biological experiments in these animals.     

Host Specificity and Source of Enterocytozoon bieneusi Genotypes in a Drinking Source Watershed

To assess the host specificity of Enterocytozoon bieneusi and to track the sources of E. bieneusi contamination, we genotyped E. bieneusi in wildlife and stormwater from the watershed of New York City''s source water, using ribosomal internal transcribed spacer (ITS)-based PCR and sequence analyses. A total of 255 specimens from 23 species of wild mammals and 67 samples from stormwater were analyzed. Seventy-four (29.0%) of the wildlife specimens and 39 (58.2%) of the stormwater samples from streams were PCR positive. Altogether, 20 E. bieneusi genotypes were found, including 8 known genotypes and 12 new ones. Sixteen and five of the genotypes were seen in animals and stormwater from the watershed, respectively, with WL4 being the most common genotype in both animals (35 samples) and stormwater (23 samples). The 20 E. bieneusi genotypes belonged to five genogroups (groups 1, 3, 4, and 7 and an outlier), with only 23/113 (20.4%) E. bieneusi-positive samples belonging to zoonotic genogroup 1 and 3/20 genotypes ever being detected in humans. The two genogroups previously considered host specific, groups 3 and 4, were both detected in multiple groups of mammals. Thus, with the exception of the type IV, Peru11, and D genotypes, which were detected in only 7, 5, and 2 animals, respectively, most E. bieneusi strains in most wildlife samples and all stormwater samples in the watershed had no known public health significance, as these types have not previously been detected in humans. The role of different species of wild mammals in the contribution of E. bieneusi contamination in stormwater was supported by determinations of host-adapted Cryptosporidium species/genotypes in the same water samples. Data from this study indicate that the host specificity of E. bieneusi group 3 is broader than originally thought, and wildlife is the main source of E. bieneusi in stormwater in the watershed.     

Enterocytozoon bieneusi in Dairy Cattle in the Northeast of China: Genetic Diversity of ITS Gene and Evaluation of Zoonotic Transmission Potential

Enterocytozoon bieneusi is the most frequently diagnosed microsporidian species in humans. It has been found in a wide range of animals and is considered an important zoonotic pathogen. To date, little information is available on the role that cattle play in the epidemiology of human microsporidiosis caused by E. bieneusi in China. In this study, 133 fecal specimens from dairy cattle were collected in Heilongjiang Province, China. Enterocytozoon bieneusi was identified and genotyped by nested PCR analysis of the internal transcribed spacer (ITS) region of the rRNA gene, with 30.1% positive. Nine ITS genotypes were identified: six known genotypes—O (n = 26), EbpA (n = 2), I (n = 2), J (n = 2), D (n = 1) and BEB4 (n = 1)—and three novel genotypes named as CC‐I to CC‐III (two each). Genotype O was identified in cattle for the first time. The observation of all the six known genotypes here reported previously in humans, and also the fact of all the three novel genotypes (CHN‐DC1 to CHN‐DC3) falling into zoonotic group 1, indicate the possibility of cattle in the transmission of E. bieneusi to humans.     

Molecular Detection,Multilocus Genotyping,and Population Genetics of Enterocytozoon bieneusi in Pigs in Southeastern China

Enterocytozoon bieneusi is an important opportunistic pathogen widely distributed in humans and animals that causes diarrhea or fatal diarrhea in immunocompromised hosts. To examine the infection status and molecular characteristics of E. bieneusi in pigs, 725 fecal samples were collected from pigs in six areas of Fujian Province. The E. bieneusi genotypes were identified based on the internal transcribed spacer (ITS) regions of the ribosomal RNA (rRNA) gene by nested PCR, and its population genetics were analyzed by multilocus sequence typing (MLST). The results showed that the infection rate of E. bieneusi was 24.4% (177/725), and 11 known genotypes (EbpC, EbpA, CHN‐RR2, KIN‐1, CHG7, CHS5, CM11, CHG23, G, PigEBITS, and D) and 2 novel genotypes (FJF and FJS) were identified. All the genotypes were found to be clustered into zoonotic Group 1. Moreover, 52 positive samples were successfully amplified at minisatellite and microsatellite loci and formed 48 distinct multilocus genotypes (MLGs). Further population structure analyses showed strong genetic linkage disequilibrium (LD) and several recombination events (Rm), indicating that E. bieneusi has a clonal population structure. This study is the first to investigate the prevalence and molecular characteristics of E. bieneusi in Fujian Province and could provide baseline data to control E. bieneusi infection in pigs and humans and deepen our understanding of the zoonotic risk of E. bieneusi and its distribution in China.     

Occurrence and genotypes of Giardia isolated from lambs in Spain

Three hundred and eighty six faecal specimens were randomly collected from 1- to 3-month-old lambs from 16 farms in Spain to investigate the presence of different genotypes of Giardia duodenalis. Individual specimens were examined by IFA (Immunofluorescence assay) and β-giardin PCR polymerase chain reaction. Cysts of G. duodenalis were shed by lambs in every flock analyzed, showing a prevalence by farms of 100%. The average prevalence of G. duodenalis for the 386 specimens was 42%, ranging from 8.3 to 80% depending on the farm. β-giardin PCR positive samples were sequenced to determine the genotypes present at each farm and seven new subtypes of β-giardin Assemblage E are reported in this study. In each farm, one to six different β-giardin subtypes were found, showing the high variability of the target. Also, one flock had the zoonotic Assemblage A. This is the first report of Giardia subgenotype A-1 in sheep in Spain.     

Host-Adapted Cryptosporidium spp. in Canada Geese (Branta canadensis)

The prevalence and distribution of Cryptosporidium spp. in the fecal droppings of the free-living waterfowl Canada geese were examined at 13 sites in Ohio and Illinois. On the basis of the analysis of the small-subunit rRNA gene by PCR, followed by restriction fragment length polymorphism analysis and DNA sequencing, 49 (23.4%) of 209 fecal specimens collected from 10 sites (76.9%) were positive for Cryptosporidium spp. The following five Cryptosporidium species and genotypes were identified: Cryptosporidium goose genotype I (in 36 specimens), Cryptosporidium goose genotype II (in 9 specimens), Cryptosporidium duck genotype (in 1 specimen), Cryptosporidium parvum (in 4 specimens), and C. hominis (in 2 specimens). Cryptosporidium goose genotype I was the most prevalent parasite and was found at all five Cryptosporidium-positive sites in Ohio and at four of five positive sites in Illinois, followed by Cryptosporidium goose genotype II, which was found at two of five positive sites in Ohio and at four of five positive sites in Illinois. Cryptosporidium goose genotype II was detected for the first time, and it is phylogenetically related to goose genotype I and the duck genotype. All three genotypes have not so far been reported in humans, and their pathogenicity in geese has not been determined. Only 10.2% of the Cryptosporidium-positive specimens had C. parvum and C. hominis. The results of this study indicate that Canada geese might only serve as accidental carriers of cryptosporidia infectious to humans and probably play a minor role in the animal-to-human transmission cycle of the pathogen.     

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.