Molecular characterization of common zoonotic protozoan parasites and bacteria causing diarrhea in dairy calves in Ningxia Hui Autonomous Region, China.

Diarrhea caused by zoonotic pathogens is one of the most common diseases in dairy calves, threatening the health of young animals. Humans are also at risk, in particular children. To explore the pathogens causing diarrhea in dairy calves, the present study applied PCR-based sequencing tools to investigate the occurrence and molecular characteristics of three parasites (Cryptosporidium spp., Giardia duodenalis, and Enterocytozoon bieneusi) and three bacterial pathogens (Escherichia coli, Clostridium perfringens, and Salmonella spp.) in 343 fecal samples of diarrheic dairy calves from five farms in Lingwu County, Ningxia Hui Autonomous Region, China. The total positive rate of these pathogens in diarrheic dairy calves was 91.0% (312/343; 95% CI, 87.9-94.0), with C. perfringens (61.5%, 211/343; 95% CI, 56.3-66.7) being the dominant one. Co-infection with two to five pathogens was found in 67.3% (231/343; 95% CI, 62.4-72.3) of investigated samples. There were significant differences (p < 0.05) in the positive rates of Cryptosporidium spp. and diarrheagenic E. coli among farms, age groups, and seasons. Two Cryptosporidium species (C. parvum and C. bovis) and five gp60 subtypes of C. parvum (IIdA15G1, IIdA20G1, IIdA19G1, IIdA14G1, and a novel IIdA13G1) were identified. Two assemblages (assemblage E and zoonotic assemblage A) of G. duodenalis and six ITS genotypes of E. bieneusi (J, Henan-IV, EbpC, I, EbpA, and ESH-01) were observed. Four virulence genes (eaeA, stx1, stx2, and st) of diarrheagenic E. coli and one toxin type (type A) of C. perfringens were detected. Our study enriches our knowledge on the characteristics and zoonotic potential of diarrhea-related pathogens in dairy calves.

Title: Caractérisation moléculaire des protozoaires parasites zoonotiques courants et des bactéries responsables de diarrhée chez les veaux laitiers dans la région autonome Hui du Ningxia, en Chine. Abstract: La diarrhée causée par des agents pathogènes zoonotiques est l'une des maladies les plus courantes chez les veaux laitiers, menaçant la santé des jeunes animaux. Ceci est également un risque pour la santé humaine, en particulier les enfants. Pour explorer les agents pathogènes responsables de la diarrhée chez les veaux laitiers, cette étude a utilisé des outils de séquençage basés sur la PCR pour étudier l'occurrence et les caractères moléculaires de trois parasites (Cryptosporidium spp., Giardia duodenalis et Enterocytozoon bieneusi) et de trois agents pathogènes bactériens (Escherichia coli, Clostridium perfringens et Salmonella spp.) dans 343 échantillons fécaux de veaux laitiers diarrhéiques provenant de cinq fermes du comté de Lingwu, région autonome Hui du Ningxia, en Chine. Le taux total positif de ces pathogènes chez les veaux laitiers diarrhéiques était de 91,0 % (312/343; IC à 95 %, 87,9­94,0), et C. perfringens (61,5 %, 211/343; IC à 95 %, 56,3­66,7) était le plus répandu. Une co-infection avec deux à cinq pathogènes a été trouvée dans 67,3 % (231/343; IC à 95 %, 62,4­72,3) des échantillons étudiés. Il y avait des différences significatives (p < 0,05) dans les taux positifs de Cryptosporidium spp. et d'E. coli diarrhéogènes entre les fermes, les groupes d'âge et les saisons. Deux espèces de Cryptosporidium (C. parvum et C. bovis) et cinq sous-types de gp60 de C. parvum (IIdA15G1, IIdA20G1, IIdA19G1, IIdA14G1 et un nouveau, IIdA13G1) ont été identifiés. Deux assemblages (assemblage E et assemblage zoonotique A) de G. duodenalis et six génotypes ITS d'E. bieneusi (J, Henan-IV, EbpC, I, EbpA et ESH-01) ont été observés. Quatre gènes de virulence (eaeA, stx1, stx2 et st) d'E. coli diarrhéogènes et un type de toxine (type A) de C. perfringens ont été détectés. Notre étude enrichit les connaissances sur les caractères et le potentiel zoonotique des agents pathogènes liés à la diarrhée chez les veaux laitiers.

Epidemiological survey of gastrointestinal helminths and protozoa in Testudines from Sardinia, Italy.

The order Testudines comprises some of the most endangered groups of vertebrates. Under specific circumstances, infectious and parasitic diseases may affect the survival and fecundity of these animals, potentially threatening Testudines populations. In Sardinia, besides the three species of tortoises present in the wild (Testudo hermanni, Testudo graeca and Testudo marginata), many others are kept as pets. However, epidemiological studies on these animals have not been conducted so far. Thus, the aim of the work was to investigate the presence of gastrointestinal parasites in captive and wild tortoises of Sardinia, Italy, with particular regard to those of zoonotic importance. For the 215 animals examined (n = 36 wild caught and n = 179 private-owned), fecal samples were collected and processed by flotation and modified Ziehl-Neelsen technique. An overall prevalence of 81.4% for endoparasites was detected, with oxyurids being the most prevalent (74.4%), followed by Nyctotherus spp. (18.6%), Cryptosporidium spp. (12.6%), Angusticaecum spp. (2.8%), strongyles (0.9%), Balantidium spp. (0.9%), coccidia (0.9%), cestodes (0.5%), and Giardia spp. (0.5%). Data suggest that tortoises are affected by a great variety of endoparasites, and further molecular analysis are required to assess the impact of Cryptosporidium and Giardia species in these hosts. Therefore, regular health screenings are of importance for the management of these animals and for preventing emerging infectious diseases.

A pilot study of intestinal protist detection in humans, animals, and the environment in a slum area in Mymensingh, Bangladesh.

The purpose of this study was to investigate the presence and transmission of Cryptosporidium spp., Blastocystis sp., Giardia intestinalis and Entamoeba histolytica between different hosts and their shared environment in a slum area of Bangladesh. A total of 102 samples were collected from Mymensingh, Bangladesh. This collection encompassed 16 human samples, 35 soil samples and 51 animal samples from various households within the region. The detection of Cryptosporidium spp., Blastocystis sp., G. intestinalis, and E. histolytica was carried out using nested PCR and/or quantitative PCR methodologies. Among the samples, 24 human, animal, and soil out of 102 were found positive for Blastocystis sp., spanning seven subtypes: ST2, ST3, ST7, ST10, ST23, ST24 and ST25. Additionally, eight samples (8/102) including human, animal and soil tested positive for Cryptosporidium spp., including C. parvum, C. baileyi, C. bovis, and C. meleagridis. Furthermore, one soil sample tested positive for G. intestinalis assemblage B, while no samples tested positive for E. histolytica. The detection of Cryptosporidium spp., Blastocystis sp., and G. intestinalis in this study has provided insights into their presence, extending beyond humans. Moreover, these findings highlight the importance of embracing a One Health perspective with an emphasis on specific parasitic microorganisms.

Wastewater-based intestinal protozoa monitoring in Shanghai, China.

Intestinal protozoa Cryptosporidium spp., Giardia duodenalis, and Enterocytozoon bieneusi have been implicated in serious waterborne outbreaks worldwide. Wastewater-based epidemiology (WBE) is a promising approach for evaluating the disease prevalence in a catchment population in that it monitors the contamination level of the intestinal pathogens in wastewater. We collected 48 urban wastewater samples (24 from influents and 24 from effluents) from the Yangpu Wastewater Treatment Plant (YPWTP) in Shanghai, China. We identified Cryptosporidium spp., G. duodenalis, and E. bieneusi by nested polymerase chain reaction (PCR) amplification. Cryptosporidium hominis and subtype IdA14 were identified in two samples by analyzing the sequences of small subunit ribosomal RNA (SSU rRNA) and 60-kDa glycoprotein (gp60) genes, respectively. The G. duodenalis sub-assemblage AII (n = 8) and assemblage C (n = 4) in 12 samples were determined by analyzing triosephosphate isomerase (tpi) gene sequences. The E. bieneusi genotype A was identified in one sample by analyzing the sequence of the internal transcribed spacer (ITS) region of the rRNA gene. These findings suggest that improving wastewater treatment and monitoring the virility of pathogens in effluents is critical. We observed similar prevalence and genotypes/subtypes of the three intestinal protozoa in our wastewater samples as those reported in previous studies, providing evidence that WBE can be used as an effective epidemic management tool.IMPORTANCECryptosporidium spp., Giardia duodenalis, and Enterocytozoon bieneusi are common intestinal protozoa causing diarrhea. The infective oocysts, cysts, and spores released in feces can survive in different environments, including multiple types of water bodies. Humans can acquire these intestinal protozoan infections via the fecal-oral route as in waterborne transmission. Wastewater-based epidemiology can rapidly and reliably detect and monitor the emergence and spread of waterborne diseases. We detected Cryptosporidium spp., G. duodenalis, and E. bieneusi in a wastewater treatment plant in Shanghai, China, reflecting the occurrence and genetic characterizations of the three intestinal pathogens from community members served by the wastewater treatment plant.

Prevalence of Cryptosporidium spp. infection in rodents and chickens in Franceville, Gabon.

Background and Aim: Cryptosporidium spp. members of the phylum Apicomplexa are obligate protozoan parasites capable of infecting various vertebrate hosts, including rodents and chickens. Infection caused by these parasites may lead to zoonotic diseases in humans. The aim of this study was to estimate the prevalence of Cryptosporidium spp. in rodents and domestic chickens sampled in Franceville, Gabon. Materials and Methods: Two hundred and eighty-five samples were collected, of which 185 samples were from rodents and 100 from domestic chickens. Microscopy after modified Ziehl-Neelsen staining and nested polymerase chain reaction targeting the small subunit (SSU) rRNA gene were used to examine Cryptosporidium spp. Results: The overall prevalence of Cryptosporidium oocysts was 55.8%, with a prevalence of 72.4% in rodents and 25.0% in domestic chickens. Molecular analysis showed that Cryptosporidium spp. were present in 4.0% of the samples. No significant correlation was observed between Cryptosporidium spp. carriage and sex or location in this study. These results indicate that Cryptosporidium spp. persist and circulate in the studied animal species in Franceville, Gabon. Conclusion: Infection with Cryptosporidium is very common in rodents and chickens in Franceville. The potential risk of human contamination cannot be ruled out. More research should be conducted to characterize Cryptosporidium species circulating in rodents and chickens in Gabon. Such studies are essential to better understand the epidemiology of this protozoan and its potential impact on public health.

Cryptosporidium spp. in captive snakes from 26 provinces in China: Prevalence, molecular characterization, and symptoms.

Snakes are sometimes regarded as pets and are used in traditional Chinese medicine. Cryptosporidium spp. are frequently identified in snakes, representing an important pathogen and causing gastrointestinal diseases. Current data indicate that risk factors for infection and patterns of clinical symptom presentation may differ among Cryptosporidium spp. To better understand the infection status by Cryptosporidium spp., fecal samples were collected from 603 asymptomatic and 147 symptomatic snakes in 26 provinces of China. These samples came from Elaphe guttata, Elaphe obsoleta, Pituophis melanoleucus, Thamnophis sirtalis, Lampropeltis getulus, and Heterodon nasicus. The partial small subunit (SSU) rRNA gene was amplified using nested polymerase chain reaction (PCR) to investigate the infection rate of Cryptosporidium spp., and to assess evolutionary relationships and genetic characterization. A prevalence of 20% was recorded in asymptomatic snakes, with age identified as a significant risk factor. In contrast, 70% of symptomatic snakes were positive for Cryptosporidium spp., with Cryptosporidium serpentis and Cryptosporidium varanii (syn. C. saurophilum). Further analysis revealed a potential association between C. serpentis and regurgitation, and C. varanii and diarrhea, while neither species was linked to flatulence. To our knowledge, this is the first study to report Cryptosporidium spp. and associated clinical signs in symptomatic snakes in China. This study aims to enhance the understanding of Cryptosporidium infections, risk factors, and clinical manifestations in snakes, providing data crucial for the control and prevention of cryptosporidiosis.

Title: Cryptosporidium spp. chez les serpents captifs de 26 provinces de Chine : prévalence, caractérisation moléculaire et symptômes. Abstract: Les serpents sont parfois considérés comme animaux de compagnie et sont utilisés en médecine traditionnelle chinoise. Des Cryptosporidium spp. sont fréquemment identifiés chez les serpents, ont un rôle d'agent pathogène important et provoquent des maladies gastro-intestinales. Les données actuelles indiquent que les facteurs de risque d'infection et les schémas de présentation des symptômes cliniques peuvent varier en fonction des espèces de Cryptosporidium. Pour mieux comprendre l'état d'infection par Cryptosporidium spp., des échantillons fécaux ont été collectés auprès de 603 serpents asymptomatiques et 147 serpents symptomatiques dans 26 provinces de Chine. Ces échantillons provenaient d'Elaphe guttata, Elaphe obsoleta, Pituophis melanoleucus, Thamnophis sirtalis, Lampropeltis getulus et Heterodon nasicus. Le gène de l'ARNr de la petite sous-unité partielle (SSU) a été amplifié à l'aide d'une réaction en chaîne par polymérase (PCR) imbriquée pour étudier le taux d'infection par Cryptosporidium spp. et évaluer les relations évolutives et la caractérisation génétique. Une prévalence de 20 % a été trouvée chez les serpents asymptomatiques, l'âge étant identifié comme un facteur de risque important. En revanche, 70 % des serpents symptomatiques étaient positifs à Cryptosporidium spp. avec Cryptosporidium serpentis et Cryptosporidium varanii (syn. C. saurophilum). Une analyse plus approfondie a révélé une association potentielle entre C. serpentis et la régurgitation, et C. varanii et la diarrhée, alors qu'aucune des deux espèces n'était liée aux flatulences. À notre connaissance, il s'agit ici de la première étude à signaler la présence de Cryptosporidium spp. et les signes cliniques associés chez des serpents symptomatiques en Chine. Cette étude vise à améliorer la compréhension des infections à Cryptosporidium, des facteurs de risque et des manifestations cliniques chez les serpents, en fournissant des données cruciales pour le contrôle et la prévention de la cryptosporidiose.

Global epidemiology and species/genotype distribution of Cryptosporidium in camels: A systematic review and meta-analysis.

This review analyzed reported data of Cryptosporidium prevalence in camels and the species/genotype distribution. Four databases (PubMed, Web of Science, Scopus, Google Scholar) were screened, and studies published by April 1, 2024, were included. Total estimates and 95% CIs were calculated using a random-effects model. The weighted prevalence of Cryptosporidium spp. in 7372 camels examined from 12 different countries was estimated at 13.8% with a 95% CI of 10.3-18.4%. The sensitivity analysis based on excluding the individual studies did not result in significant statistical changes in the final weighted prevalence. Subgroup prevalence of Cryptosporidium spp. in camels was analyzed by publication year, continent, WHO region, country, camel type, sample size, diagnostic method, age, and gender. A significant publication bias (P < 0.05) was reported in the present study. Limitations encountered in this study encompassed: insufficient study diversity, reliance on single study results, inadequate molecular and serological studies in comparison to microscopic studies, etc., all of which could impact the findings. The study identified eight Cryptosporidium spp. in camels: C. parvum, C. andersoni, C. bovis, C. muris, C. ratti, C. occultus, C. ubiquitum, and C. hominis. The first three species had pooled prevalence rates of 65.5%, 66%, and 19.2%, respectively. Each of the remaining five species was documented using a single dataset/study. Moreover, genotypes IIdA19G1, IIaA15G1R1, If-like-A15G2, IIdA15G1, IIaA15G2R1, IIaA17G2R1, and IIaA18G2R1 (C. parvum), genotype IV (C. ratti), genotype XIIa (C. ubiquitum), and genotype IkA19G1 (C. hominis) have been identified in camels globally. The findings suggest that camels can act as a source of infection for a variety of Cryptosporidium species/genotypes, and can therefore play a key role in disseminating this protozoan to humans and animals.

Molecular characterization and zoonotic risk assessment of Cryptosporidium spp. in children and calves in Bangladesh.

Cryptosporidium is a gastro-intestinal protozoan parasite that has been found to infect both humans and livestock. This study investigated the parasite in 998 fecal samples from Bangladeshi children (n = 299) and calves (n = 699) to determine its prevalence, genetic variation, and zoonotic importance. The nested PCR and sequencing of the SSU rRNA gene in the samples showed a Cryptosporidium infection rate of 2.3% (7/299) in children and 15.7% (110/699) in calves. Statistical analysis revealed insignificant variations in Cryptosporidium infections among children across age, gender, and study area, while in calves, the infection rate significantly differed based on location and breed. Genotyping of seven human isolates of Cryptosporidium confirmed C. hominis (n = 5) and C. parvum (n = 2). After characterizing 110 Cryptosporidium isolates from calves, C. andersoni (n = 55), C. ryanae (n = 29), C. bovis (n = 14), C. parvum (n = 10), C. ubiquitum (n = 1), and C. occultus (n = 1) were identified. Cryptosporidium hominis and C. parvum-positive samples were further subjected to nested PCR and sequencing of the glycoprotein 60 (gp60) gene for subtyping. Four C. hominis subtypes (IaA19R3, IaA23R3, IbA9G3, and IdA15G1) and one C. parvum subtype (IIdA15G1) were observed. In conclusion, Cryptosporidium was prevalent in calves but less common in children in the study locations, and the presence of zoonotic Cryptosporidium species and subtypes in calves raises concerns regarding zoonotic transmission to humans.

Molecular characterizations of Cryptosporidium spp. in brown rat (Rattus norvegicus) from an animal feedlot in Xinjiang, China.

Cryptosporidium infection is a common occurrence in rodents worldwide. In this study, 435 wild brown rats were captured from an animal feedlot in Xinjiang, China, with a fecal sample obtained directly from the rectal contents of each rat. The DNA extracted from these fecal samples was analyzed for Cryptosporidium spp. using PCR targeting the SSU rRNA gene. The prevalence of Cryptosporidium infection in brown rats was found to be 5.5% (24 out of 435). Interestingly, the infection rates varied among different animal enclosures, with rates of 0% in the chicken coop (0/51), cowshed (0/3), and varying rates in other areas including the sheepfold (6.1%, 6/98), the pigsty (7.6%, 10/132), the dovecote (7.0%, 5/71), and outdoor environments (3.8%, 3/80). The study identified three species and one genotype of Cryptosporidium, namely C. occultus (n = 10), C. parvum (n = 4), C. ditrichi (n = 1), and Cryptosporidium rat genotype IV (n = 9). Additionally, two of the C. parvum isolates were successfully subtyped as IIdA19G1 (n = 2) at the gp60 gene. These results offer valuable insights into the prevalence and genetic diversity of Cryptosporidium in brown rats within the region.

Cryptosporidium spp. Infection and Genotype Identification in Pre-Weaned and Post-Weaned Calves in Yunnan Province, China.

BACKGROUND: Cryptosporidium is a globally distributed zoonotic protozoan parasite in humans and animals. Infection is widespread in dairy cattle, especially in calves, resulting in neonatal enteritis, production losses and high mortality. However, the occurrence of Cryptosporidium spp. in pre- and post-weaned calves in Yunnan Province remains unclear. METHODS: We collected 498 fecal samples from Holstein calves on 10 different farms in four regions of Yunnan Province. Nested PCR and DNA sequencing were used to determine the infection, species and genotypes of Cryptosporidium spp. in these animals. RESULTS: The overall occurrence of Cryptosporidium spp. in Holstein calves was 32.9% (164/498), and the prevalence in pre- and post-weaned calves was 33.5% (106/316) and 31.9% (58/182), respectively. Four Cryptosporidium species were identified in these animals, namely C. bovis (n = 119), C. parvum (n = 23), C. ryanae (n = 20) and C. andersoni (n = 2). Based on sequencing analysis of the 60 kDa glycoprotein gene of C. bovis, C. parvum and C. ryanae, six subtypes of C. bovis (XXVIe, XXVIb, XXVIf, XXVIa XXVIc and XXVId), two subtypes of C. parvum (IIdA19G1 and IIdA18G1) and four subtypes of C. ryanae (XXIf, XXId, XXIe and XXIg) were identified. CONCLUSIONS: These results provide essential information to understand the infection rate, species diversity and genetic structure of Cryptosporidium spp. populations in Holstein pre-weaned and post-weaned calves in Yunnan Province. Further, the presence of IIdA18G1 and IIdA19G1 in C. parvum implies significant animal and public health concerns, which requires greater attention and more preventive measures.

Evaluation of molecular-based methods for the detection and quantification of Cryptosporidium spp. in wastewater.

Cryptosporidium poses significant public health risks as a cause of waterborne disease worldwide. Clinical surveillance of cryptosporidiosis is largely underreported due to the asymptomatic and mildly symptomatic infections, clinical misdiagnoses, and barriers to access testing. Wastewater surveillance overcomes these limitations and could serve as an effective tool for identifying cryptosporidiosis at the population level. Despite its potential, the lack of standardized wastewater surveillance methods for Cryptosporidium spp. challenges implementation design and the comparability between studies. Thus, this study compared and contrasted Cryptosporidium wastewater surveillance methods for concentrating wastewater oocysts, extracting oocyst DNA, and detecting Cryptosporidium genetic markers. The evaluated concentration methods included electronegative membrane filtration, Envirocheck HV capsule filtration, centrifugation, and Nanotrap Microbiome Particles, with and without additional immunomagnetic separation purification (except for the Nanotrap Microbiome Particles). Oocyst DNA extraction by either the DNeasy Powersoil Pro kit and the QIAamp DNA Mini kit were evaluated and the impact of bead beating and freeze-thaw pretreatments on DNA recoveries was assessed. Genetic detection via qPCR assays targeting either the Cryptosporidium 18S rRNA gene or the Cryptosporidium oocyst wall protein gene were tested. Oocyst recovery percentages were highest for centrifugation (39-77 %), followed by the Nanotrap Microbiome Particles (24 %), electronegative filtration with a PBST elution (22 %), and Envirocheck HV capsule filtration (13 %). Immunomagnetic separation purification was found to be unsuitable due to interference from the wastewater matrix. Bead-beating pretreatment enhanced DNA recoveries from both the DNeasy Powersoil Pro kit (314 gc/µL DNA) and the QIAamp DNA Mini kit (238 gc/µL DNA). In contrast, freeze-thaw pretreatment reduced DNA recoveries to under 92 gc/µL DNA, likely through DNA degradation. Finally, while both qPCR assays were specific to Cryptosporidium spp., the 18S rRNA assay had a 5-fold lower detection limit and could detect a wider range of Cryptosporidium spp. than the Cryptosporidium oocyst wall protein assay.

Molecular characterization of Cryptosporidium spp. in Bactrian camels (Camelus bactrianus) from Yili Kazak Autonomous Prefecture of Xinjiang, China.

Introduction: Cryptosporidium spp. is a significant zoonotic parasite. The prevalence and infection characteristics of Cryptosporidium spp. in Bactrian camels in Yili Kazak Autonomous Prefecture have yet to be fully understood. Thus, the molecular epidemiology of cryptosporidiosis in camels was investigated in this region. Methods: A total of 1,455 fecal samples were collected from 6 counties in three regions (Altay, Tacheng, and Yili) in Yili Prefecture. Nested PCR targeting the small subunit ribosomal RNA (ssu rRNA) gene was used to identify the species or genotypes of Cryptosporidium infection in camels. For C. parvum positive samples, the subtypes were identified using the 60-kDa glycoprotein (gp60) gene. Results and discussion: The overall infection rate was 8.7% (126/1,455), ranging from 5.6% to 11.7% in different regions, and 4.2% to 15.8% in different counties. A significant difference was observed amongst the counties (p < 0.001). Three species were detected, namely C. andersoni (65.1%, 82/126), C. parvum (34.1%, 43/126), and C. occultus (0.8%, 1/126). Three C. parvum subtypes, If-like-A15G2 (n = 29), IIdA15G1 (n = 4), and IIdA19G1(n = 1) were detected, with If-like-A15G2 being the most prevalent subtype. Camels aged 3-12 months exhibited the highest infection rate (11.4%, 44/387), with no significant difference among age groups (p > 0.05). C. parvum was predominant in camels under 3 months, while C. andersoni prevailed in camels over 3 months. There was an extremely significant difference observed among seasons (p < 0.001), summer had the highest infection rates (16.9%, 61/360). This study collected nearly 1,500 samples and, for the first time, investigated Cryptosporidium spp. infection in camels based on different age groups and seasons. All three Cryptosporidiumspecies identified were zoonotic, posing a potential threat to human health and requiring close attention.

Prevalence and Molecular Characterization of Cryptosporidium spp., Giardia duodenalis, and Enterocytozoon bieneusi in Cattle in Heilongjiang Province, Northeast China.

Crytosporidium spp., Giardia duodenalis, and Enterocytozoon bieneusi are important diarrheal pathogens with a global distribution that threatens the health of humans and animals. Despite cattle being potential transmission hosts of these protozoans, the associated risks to public health have been neglected. In the present study, a total of 1155 cattle fecal samples were collected from 13 administrative regions of Heilongjiang Province. The prevalence of Cryptosporidium spp., G. duodenalis, and E. bieneusi were 5.5% (64/1155; 95% CI: 4.2-6.9), 3.8% (44/1155; 95% CI: 2.7-4.9), and 6.5% (75/1155; 95% CI: 5.1-7.9), respectively. Among these positive fecal samples, five Cryptosporidium species (C. andersoni, C. bovis, C. ryanae, C. parvum, and C. occultus), two G. duodenalis assemblages (E and A), and eight E. bieneusi genotypes (BEB4, BEB6, BEB8, J, I, CHS7, CHS8, and COS-I) were identified. Phylogenetic analysis showed that all eight genotypes of E. bieneusi identified in the present study belonged to group 2. It is worth noting that some species/genotypes of these intestinal protozoans are zoonotic, suggesting a risk of zoonotic disease transmission in endemic areas. The findings expanded our understanding of the genetic composition and zoonotic potential of Cryptosporidium spp., G. duodenalis, and E. bieneusi in cattle in Heilongjiang Province.

Clostridium and Cryptosporidium outbreak linked to a splash pad.

BACKGROUND: . Splash pads for recreational purposes are widespread. Using these pads can pose a health risk if they lack installation regulation and water quality supervision. Our aim was to describe a waterborne disease outbreak caused by Clostridium perfringens and Cryptosporidium spp. in a Barcelona district and the measures taken for its control. METHODS: . On August 2018, 71 cases of acute gastroenteritis were detected, affecting people who used a splash pad or were in contact with a user. Microbiological and environmental investigations were carried out. A descriptive analysis of the sample and Poisson regression models adjusted for age and sex were performed, obtaining frequencies, median values, and adjusted prevalence ratios with their 95% confidence intervals. RESULTS: The median age of the cases was 6.7 years, 27 (38%) required medical care, and three (4.2%) were hospitalized. The greater the number of times a person entered the area, the greater the number of symptoms and their severity. Nineteen (76%) of the 25 stool samples collected from cases showed the presence of one or both pathogens. Environmental investigations showed deficiencies in the facilities and identified the presence of both species in the splash pad. Health education and hygiene measures were carried out, and 14 days after the closure of the facilities, no more cases related to the pad were recorded. CONCLUSIONS: . Specific regulations are needed on the use of splash pads for recreational purposes. Until these regulations are in place, these types of facility should comply with the regulations that apply to swimming pools and spas, including those related to the design of the tanks, water recirculation systems, and adequate disinfection systems.

A molecular survey of zoonotic pathogens of public health importance in rodents/shrews and their ectoparasites trapped in Puducherry, India.

BACKGROUND: Globally, India has a high zoonotic disease burden and lacks surveillance data in humans and animals. Rodents are known reservoirs for many zoonotic diseases and their synanthropic behavior poses a great public health threat. METHODS: In this study, trapped rodents/shrews from randomly selected villages within Puducherry, India, and their ectoparasites were screened for zoonotic pathogens, namely, Orientia tsutsugamushi, other pathogenic rickettsiae, Leptospira spp., Cryptosporidium spp., Coxiella burnetii and methicillin-resistant Staphylococcus aureus (MRSA) using conventional PCR. A total of 58 rodents/shrews were trapped from 11 villages. The species trapped were Suncus murinus (49/58, 84.48%), Rattus rattus (8/58, 13.79%) and Rattus norvegicus (1/58, 1.72%). All ectoparasites collected were identified as mites and its infestation rate was 46.55% (27/58). RESULTS: Real-time PCR targeting the 47 kDa gene of O. tsutsugamushi revealed positivity in one rodent and one shrew (3.45%) and two mite pools (7.41%). Conventional PCR targeting the 56 kDa gene revealed positivity in one shrew and two mite pools and the phylogenetic analysis of all three amplicons indicated the circulation of the Gilliam-related serotype. MRSA was detected in the alimentary tract of a shrew (1/32, 3.13%). Leptospira spp., Rickettsia, Cryptosporidium spp. and Co. burnetii tested negative. CONCLUSIONS: The detection of zoonotic pathogens within reservoir hosts and vectors poses a risk of transmission to humans. This study signifies the need for zoonotic pathogen surveillance in synanthropic rodents/shrews.

Cryptosporidium spp. in large-scale sheep farms in China: prevalence and genetic diversity.

Cryptosporidium spp. are significant zoonotic intestinal parasites that induce diarrhea and even death across most vertebrates, including humans. Previous studies showed that sheep are important hosts for Cryptosporidium and that its distribution in sheep is influenced by geography, feeding patterns, age, and season. Environmental factors also influence the transmission of Cryptosporidium. Molecular studies of Cryptosporidium in sheep have been conducted in only a few regions of China, and studies into the effect of sheep-housing environments on Cryptosporidium transmission are even rarer. To detect the prevalence of Cryptosporidium in large-scale sheep-housing farms, a total of 1241 fecal samples were collected from sheep, 727 environmental samples were taken from sheep housing, and 30 water samples were collected in six regions of China. To ascertain the existence of the parasite and identify the species of Cryptosporidium spp., we conducted nested PCR amplification of DNA extracted from all samples using the small-subunit (SSU) rRNA gene as a target. For a more in-depth analysis of Cryptosporidium spp. subtypes, C. xiaoi-and C. ubiquitum-positive samples underwent separate nested PCR amplification targeting the 60 kDa glycoprotein (gp60) gene. The amplification of the Cryptosporidium spp. SSU rRNA gene locus from the whole genomic DNA of all samples yielded a positive rate of 1.2% (20/1241) in fecal samples, 0.1% (1/727) in environmental samples, and no positive samples were found in water samples. The prevalence of Cryptosporidium spp. infection in large-scale housed sheep was 1.7%, which was higher than that in free-ranging sheep (0.0%). The highest prevalence of infection was found in weaning lambs (6.8%). Among the different seasons, the peaks were found in the fall and winter. The most prevalent species were C. xiaoi and C. ubiquitum, with the former accounting for the majority of infections. The distribution of C. xiaoi subtypes was diverse, with XXIIIc (n = 1), XXIIId (n = 2), XXIIIe (n = 2), and XXIIIl (n = 4) identified. In contrast, only one subtype, XIIa (n = 9), was found in C. ubiquitum. In this study, C. xiaoi and C. ubiquitum were found to be the predominant species, and Cryptosporidium was found to be present in the environment. These findings provide an important foundation for the comprehensive prevention and management of Cryptosporidium in intensively reared sheep. Furthermore, by elucidating the prevalence of Cryptosporidium in sheep and its potential role in environmental transmission, this study deepens our understanding of the intricate interactions between animal health, environmental contamination, and public health dynamics.

Molecular Detection of Cryptosporidium spp. and Microsporidia in Human and Animal Stool Samples.

Cryptosporidium spp. and Microsporidia are opportunistic microorganisms with remarkable zoonotic transmission potential due to their capacity to infect humans and animals. The aim of this study was to evaluate the prevalence of these microorganisms in stool samples of animal and human origin. In total, 369 stool samples (205 from human patients with diarrhea and 164 of animal origin) were included in the study. Cryptosporidium spp. and Microsporidia presence were determined by using multiplex nested PCR. Positive results were analyzed by using Sanger sequencing of the amplicon, utilizing BLASTN and ClustalX software to confirm identification. Cryptosporidium spp. were found in 0.97% and 4.26% of human and animal samples, respectively. Enterocytozoon bieneusi was detected in human and animal stools in 6.82% and 3.05% of the samples, respectively. No associations were found when analyzing the presence of Cryptosporidium spp. and E. bieneusi and the demographic and clinical variables of patients and animals. This study demonstrates the presence of these microorganisms in human and animal samples from different species, and the most interesting findings are the detection of Cryptosporidium spp. in pets (e.g., rodents) that are not usually included in this type of study, and the identification of E. bieneusi in patients with diarrhea without underlying disease.

The prevalence and genotypes of Cryptosporidium spp. in bar-headed goose (Anser indicus) in China.

Cryptosporidium spp. is an important foodborne and waterborne pathogen in humans and animals, causing diarrhoea in humans and respiratory and gastrointestinal diseases in birds. However, reports of Cryptosporidium infection in bar-headed goose are limited. To determine the infection rate and species/genotypes of Cryptosporidium in bar-headed goose in China, a total of 358 fecal samples were collected from 3 regions. Nested PCR was used to amplify Cryptosporidium SSU rRNA regions from the fecal extracted-DNA samples. The total infection rate of Cryptosporidium in bar-headed in China was 3.9 % (14/358), with 4.2 % (5/120) in Aba (Ngawa) Tibetan and Qiang Autonomous Prefect, Sichuan province, 7.6 % (9/119) in Maqu county, Gansu province, and 0.0 % (0/119) in Caohai, Wei ning county, Guizhou province. The differences in prevalence rate by region were statistically significant. All positive samples were identified as Cryptosporidium goose genotype I (n = 14). This is the first systematic investigation of the epidemiological status and dominant species/genotypes of Cryptosporidium in bar-headed goose in China, thereby enhancing our understanding of the epidemiology of Cryptosporidium infection in wild migratory birds.

Molecular Characterization of Giardia intestinalis and Cryptosporidium spp. Detected in Humans in Agri, Türkiye.

Background: We aimed to determine the prevalence of Giardia intestinalis and Cryptosporidium spp. in patients who admitted hospital with diarrhea and to gain information about the transmission of these parasites in Agri, Türkiye. Methods: This study included 184 patients who applied to Agri-Diyadin State Hospital, Türkiye in 2022. The immunochromatographic card test was used for detection of the G. intestinalis and Cryptosporidium spp. Nested PCR-RFLP analysis of the COWP gene and sequence analysis of the gp60 gene were used to genotype and subtype Cryptosporidium spp., whereas Nested PCR and sequence analyses of ß-giardin gene were used genotype G. intestinalis. Results: Of the 184 stool specimens examined, 12 (14.29%) and 7 (3.80%) were positive for G. intestinalis and Cryptosporidium spp., respectively. The Cryptosporidium species were identified as C. parvum belonging to the IId sub-type family. The G. intestinalis were identified assemblages A. Conclusion: Assemblage A, which is associated with diarrhea, is responsible for giardiasis and C. parvum IId subtype, often found in sheep, goats and cattle, is responsible for cryptosporidiosis in Agri, Türkiye.

Prevalence and Genotyping of Water- and Food-Borne Parasitic Protozoans (Giardia duodenalis and Cryptosporidium spp.) in Hospitalized Patients from Northwestern Romania.

Giardia duodenalis and Cryptosporidium spp. are important zoonotic protozoan pathogens that infect the gastro-intestinal tract of numerous vertebrates, including humans, and both parasites are responsible for water- or food-borne outbreaks of disease worldwide. Although, globally, both parasites are highly prevalent, particularly in developing countries, epidemiological data from Romania are scarce, and genotyping has rarely been performed. The aims of the present study were to investigate the occurrence and genetic diversity of G. duodenalis and Cryptosporidium spp. in patients hospitalized in Northwestern Romania in relation to clinical and paraclinical presentation and to identify the relative frequency of non-specific symptoms and potential risk factors. Between June 2022 and January 2024, 426 fecal samples were screened for gastro-intestinal parasites by rapid tests and microscopical examination, further confirmed by PCR and sequencing. Giardia duodenalis was detected and characterized in 12 samples (2.82%), while Cryptosporidium parvum was confirmed in four samples (0.94%). A majority of positive patients were symptomatic and reported nausea and vomiting with a significantly higher frequency compared to negative ones. This study provides new insights into the epidemiological status and clinical implications of gastro-intestinal parasite species and genospecies in Romania that are necessary for an in-depth understanding of the potential zoonotic transmission and improvement of patient care.