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Cryptosporidiosis of calves is caused by the enteroprotozoan Cryptosporidium spp. The disease results in intense diarrhea of calves associated with substantial economic losses in dairy farming worldwide. The aim of this study was to determine calf, herd, and within-herd Cryptosporidium prevalence and identify Cryptosporidium species and subtypes in calves with diarrhea in intensive dairy herds in central Argentina. A total of 1073 fecal samples were collected from 54 randomly selected dairy herds. Cryptosporidium-oocysts were isolated and concentrated from fecal samples using formol-ether and detected by light microscopy with the modified Ziehl-Neelsen technique. Overall prevalence of oocyst-excreting calves was found to be 25.5% (274/1073) (95% C.I. 22.9; 28.1%). Of the herds studied, 89% (48/54) included at least one infected calf, whereas within-herd prevalence ranged from the absence of infection to 57% (20/35). A highly significant association was found between the presence of diarrhea and C. parvum infection (χ2 = 55.89, p < 0.001). For species determination, genomic DNA isolated from oocyst-positive fecal samples was subjected to PCR-RFLP of the 18S rRNA gene resulting exclusively in Cryptosporidium parvum identification. C. parvum isolates of calves displaying diarrhea and high rate of excretion of oocysts were subtyped by PCR amplification and direct sequencing of the 60 kDa glycoprotein (GP60) gene. Altogether five GP60 subtypes, designated IIaA18G1R1, IIaA20G1R1, IIaA21G1R1, IIaA22G1R1, and IIaA24G1R1 were identified. Interestingly, IIaA18G1R1 and IIaA20G1R1 were predominant in calves with diarrhea and high infection intensity. Notably, IIaA24G1R1 represents a novel, previously unrecognized C. parvum subtype. The subtype IIaA18G1R1, frequently found in this study, is strongly implicated in zoonotic transmission. These results suggest that calves might be an important source for human cryptosporidiosis in Argentina.

Background Cryptosporidium spp. are worldwide protozoan parasites which include species that can lead to cryptosporidiosis in humans. Different animal species can serve as reservoirs and sources of dissemination of the disease, such as rodent species due their potential in transmitting zoonotic pathogens to humans and other animals. In the Canary Islands (Spain), Cryptosporidium parvum and Cryptosporidium hominis have been identified in patients with diarrhea. However, the occurrence of Cryptosporidium spp. in possible reservoirs in this archipelago remains unclear. Considering the zoonotic potential of these protozoans, the aim of the present study was to determine the presence of Cryptosporidium spp. in peridomestic wild rodents and the possible role of these mammals as a source of transmission of these protozoans in Canary Islands. Methods A total of 179 rodents belonging to Rattus rattus and Mus musculus domesticus from four Canary Islands, La Palma, El Hierro, Tenerife and Lanzarote, were analyzed. Feces were screened for Cryptosporidium spp. by nested PCR of the 18S ribosomal RNA fragment and the sequences used for phylogenetic analyses. Results Cryptosporidium spp. were found widely distributed with an overall prevalence of 12.30% in rodents (13.86% for R. rattus and 10.25% for M. m. domesticus ). The overall prevalence by island was 19.60% for Tenerife, 7.14% for La Palma, 5.71% for El Hierro and 0% for Lanzarote. Cryptosporidium tyzzeri , Cryptosporidium meleagridis , Cryptosporidium muris and Cryptosporidium sp. rat genotype I and II/III were successfully identified, in addition to two unidentified Cryptosporidium genotypes. Conclusions This study contributes to the knowledge of the biodiversity and distribution of Cryptosporidium spp. in wild rodents from the Canary Islands, highlighting the presence of three zoonotic species, C. tyzzeri , C. meleagridis and C. muris , being the first detection of these three species in wild rodents in the Canary Islands and the first report of C. meleagridis in R. rattus . Given the results obtained in our study, future studies in non-sampled areas are required to better understand the epidemiology of these protozoans in wild rodents in the archipelago.

Background Little is known on the occurrence and identity of Cryptosporidium species in sheep and goats in Algeria. This study aimed at investigating the occurrence of Cryptosporidium species in lambs and goat kids younger than 4 weeks. Methods A total of 154 fecal samples (62 from lambs and 92 from kid goats) were collected from 13 sheep flocks in Médea, Algeria and 18 goat flocks across Algiers and Boumerdes. They were screened for Cryptosporidium spp. by nested-PCR analysis of a fragment of the small subunit ( SSU ) rRNA gene, followed by restriction fragment length polymorphism and sequence analyses to determine the Cryptosporidium species present. Cryptosporidium parvum and C. ubiquitum were further subtyped by sequence analysis of the 60 kDa glycoprotein gene. Results Cryptosporidium spp. were detected in 17 fecal samples (11.0%): 9 from lambs (14.5%) and 8 from goat kids (8.7%). The species identified included C. parvum in 3 lambs, C. xiaoi in 6 lambs and 6 goat kids, and C. ubiquitum in 2 goat kids. Cryptosporidium infections were detected mostly in animals during the first two weeks of life (7/8 for goat kids and 7/9 for lambs) and in association with diarrhea occurrence (7/17 or 41.2% goat kids and 7/10 or 70.0% lambs with diarrhea were positive for Cryptosporidium spp.). Subtyping of C. parvum and C. ubiquitum isolates identified the zoonotic IIaA13G2R1 and XIIa subtype families, respectively. Minor differences in the SSU rRNA gene sequences were observed between C. xiaoi from sheep and goats. Conclusions Results of this study indicate that three Cryptosporidium species occur in lambs and goat kids in Algeria, including zoonotic C. parvum and C. ubiquitum . They are associated with the occurrence of neonatal diarrhea.

Cryptosporidium is increasingly recognized as one of the major causes of moderate to severe diarrhoea in developing countries. With treatment options limited, control relies on knowledge of the biology and transmission of the members of the genus responsible for disease. Currently, 26 species are recognized as valid on the basis of morphological, biological and molecular data. Of the nearly 20 Cryptosporidium species and genotypes that have been reported in humans, Cryptosporidium hominis and Cryptosporidium parvum are responsible for the majority of infections. Livestock, particularly cattle, are one of the most important reservoirs of zoonotic infections. Domesticated and wild animals can each be infected with several Cryptosporidium species or genotypes that have only a narrow host range and therefore have no major public health significance. Recent advances in next-generation sequencing techniques will significantly improve our understanding of the taxonomy and transmission of Cryptosporidium species, and the investigation of outbreaks and monitoring of emerging and virulent subtypes. Important research gaps remain including a lack of subtyping tools for many Cryptosporidium species of public and veterinary health importance, and poor understanding of the genetic determinants of host specificity of Cryptosporidium species and impact of climate change on the transmission of Cryptosporidium.

Fecal samples of 177 calves of up to 180 days of age with diarrhea from 70 farms in Austria were examined to obtain information on the occurrence of Cryptosporidium species. Initially, all samples were examined by phase-contrast microscopy. Cryptosporidium-positive samples (55.4%; n = 98) were screened by gp60 PCR, resulting in 68.4% (n = 67) C. parvum–positive samples. The remaining 31 gp60-PCR-negative and the phase-contrast microscopy negative samples (n = 79) were screened by PCR targeting a 700 bp fragment of the 18S rRNA gene. Sequencing of the PCR products revealed the presence of C. parvum (n = 69), C. ryanae (n = 11), and C. bovis (n = 7). The latter two species have never been described in Austria. C. parvum–positive samples were genotyped at the gp60 gene locus, featuring four subtypes (IIaA15G2R1, IIaA21G2R1, IIaA19G2R1, IIaA14G1R1). The most frequently detected subtype IIaA15G2R1 (n = 52) was present in calves from 30 different farms. IIaA14G1R1 (n = 5) occurred on a single farm, subtype IIaA21G2R1 (n = 4) on two farms, and subtype IIaA19G2R1 (n = 4) on three farms. The results confirm the widespread occurrence of zoonotic C. parvum in diarrheic calves.

Background Cryptosporidiosis is a gastrointestinal disease with global distribution. It has been a reportable disease in Canada since 2000; however, routine molecular surveillance is not conducted. Therefore, sources of contamination are unknown. The aim of this project was to identify species and subtypes of Cryptosporidium in clinical cases from Ontario, the largest province in Canada, representing one third of the Canadian population, in order to understand transmission patterns. Methods A total of 169 frozen, banked, unpreserved stool specimens that were microscopy positive for Cryptosporidium over the period 2008–2017 were characterized using molecular tools. A subset of the 169 specimens were replicate samples from individual cases. DNA was extracted directly from the stool and nested PCR followed by Sanger sequencing was conducted targeting the small subunit ribosomal RNA (SSU) and glycoprotein 60 ( gp60 ) genes. Results Molecular typing data and limited demographic data were obtained for 129 cases of cryptosporidiosis. Of these cases, 91 (70.5 %) were due to Cryptosporidium parvum and 24 (18.6%) were due to Cryptosporidium hominis . Mixed infections of C. parvum and C. hominis occurred in four (3.1%) cases. Five other species observed were Cryptosporidium ubiquitum ( n = 5), Cryptosporidium felis ( n = 2), Cryptosporidium meleagridis ( n = 1), Cryptosporidium cuniculus ( n = 1) and Cryptosporidium muris ( n = 1). Subtyping the gp60 gene revealed 5 allelic families and 17 subtypes of C. hominis and 3 allelic families and 17 subtypes of C. parvum . The most frequent subtype of C. hominis was IbA10G2 (22.3%) and of C. parvum was IIaA15G2R1 (62.4%). Conclusions The majority of isolates in this study were C. parvum , supporting the notion that zoonotic transmission is the main route of cryptosporidiosis transmission in Ontario. Nonetheless, the observation of C. hominis in about a quarter of cases suggests that anthroponotic transmission is also an important contributor to cryptosporidiosis pathogenesis in Ontario. Graphical Abstract

One of the most common causes of calf diarrhoea is the parasite Cryptosporidium parvum. Two longitudinal studies were carried out on a dairy farm Scotland to determine the prevalence of Cryptosporidium species and subtypes in a group of calves and to determine whether dams were a possible source of calfhood infection. Fecal samples were collected from 25 calves from birth to 12 months in the first year. In the second year, fecal samples were collected from pregnant cows (n = 29) and their calves (n = 30) from birth to 6 months. The samples were tested for Cryptosporidium and speciated. Cryptosporidium parvum-positive samples were subtyped by GP60 fragment analysis. All calves in both studies shed Cryptosporidium during the study period. Cryptosporidium parvum was the predominant species detected in calves ⩽6 weeks of age and at 6 months of age, C. bovis and C. ryanae were detected in calves older than 4 weeks of age but ⩽6 months of age. The prevalence of Cryptosporidium was higher in younger animals than in older animals. GP60 subtyping revealed two subtypes in calves on this farm (IIaA15G2R1 and IIaA19G2R1) that differed in frequency by age. Adult cattle also shed C. parvum, of four gp60 genotypes.

Cryptosporidium and Giardia are globally significant protozoan parasites responsible for severe foodborne and waterborne outbreaks, posing substantial zoonotic and environmental risks. This study aimed to comprehensively assess the prevalence of cryptosporidiosis, giardiasis, and co-infections in Beni-Suef Governorate, Egypt, using an integrated diagnostic approach combining microscopy and molecular techniques. Additionally, it was sought to identify associated risk factors in cattle fecal samples. Microscopical examination of 970 cattle fecal samples revealed an overall infection rate of 67.42% (654/970), with Cryptosporidium detected in 42.68% (414/970), Giardia in 11.96% (116/970), and co-infections in 12.78% (124/970) of cases. In irrigation water, Cryptosporidium oocysts and Giardia cysts were detected in 2/24 (8.33%) and 1/24 (4.16%) of samples, respectively. Molecular and phylogenetic analyses identified Cryptosporidium hominis in cattle and, for the first time in Egypt, Cryptosporidium ubiquitum and Cryptosporidium ryanae in irrigation water, while also proving the presence of Cryptosporidium bovis and Giardia assemblage A in cattle. Risk factors, including sex, age, season, and fecal consistency, significantly influenced infection rates, with higher prevalence in females, calves under two months, spring season, and diarrheic feces. These findings underscore the urgent need for One Health-based control strategies, integrating targeted interventions to mitigate the burden of Cryptosporidium and Giardia infections and environmental contamination.

Background Opportunistic infections are a ubiquitous complication in human immunodeficiency virus (HIV)/acquired immunodeficiency syndrome (AIDS) patients. Cryptosporidium spp., Giardia duodenalis , and Enterocytozoon bieneusi are common opportunistic intestinal pathogens in humans. In China, despite the number of HIV/AIDS patients being extremely large, only a few studies have investigated opportunistic infections caused by intestinal pathogens in this patient population. The aims of this study were to elucidate the occurrence and genetic characteristics of Cryptosporidium spp., G. duodenalis , and E. bieneusi in HIV/AIDS patients. Methods We collected fecal specimens from 155 HIV/AIDS patients (one from each patient). All of the specimens were examined for the presence of the pathogens by genotyping using polymerase chain reaction and sequencing of the small subunit ribosomal RNA gene for Cryptosporidium spp.; the triosephosphate isomerase, β-giardin and glutamate dehydrogenase genes for G. duodenalis ; and the internal transcribed spacer region of the rRNA gene for E. bieneusi . The Cryptosporidium -positive specimens were further subtyped by polymerase chain reacion and sequencing of the 60-kDa glycoprotein gene. Results Six (3.9%), three (1.9%), and eight (5.2%) HIV/AIDS patients were positive for Cryptosporidium spp., G. duodenalis , and E. bieneusi , respectively. No statistical differences were observed in occurrence rate between the groups by gender, clinical symptom (diarrhea), and CD4 + cell count. Four Cryptosporidium species were identified: Cryptosporidium hominis ( n  = 2), Cryptosporidium parvum ( n  = 1), Cryptosporidium meleagridis ( n  = 1), and Cryptosporidium andersoni ( n  = 2). Furthermore, two C. hominis subtypes (IeA12G3T3 and IaA28R4) were detected. Three G. duodenalis -positive specimens were successfully amplified and sequenced at the triosephosphate isomerase and β-giardin loci, which led to the identification of assemblages C and B, respectively. Seven genotypes (D, Type IV, EbpC, Peru11, EbpD, A, and I) were identified in E. bieneusi -positive specimens. Conclusions Our findings should increase awareness of AIDS-related opportunistic intestinal pathogens, and indicate the need for routine examination in clinical practice for the detection of Cryptosporidium spp., G. duodenalis , and E. bieneusi . Homology analyses of the three intestinal pathogens at the nucleotide and/or amino acid levels indicated their zoonotic potential. Graphical Abstract

Cryptosporidium species can infect humans and more than 260 animal species, including 54 rodent species. However, data on the occurrence and genetic characterizations of Cryptosporidium spp. in laboratory rodents are limited. The present study aimed to determine the occurrence rate and genetic characterizations of Cryptosporidium spp. in laboratory mice and rats. We collected 506 fresh combined fecal pellet specimens (457 from mice and 49 from rats) of more than 2,000 laboratory rodents in Heilongjiang Province and Shanghai City, China. Cryptosporidium spp. were identified and subtyped by DNA sequencing of the SSU rRNA and the gp60 genes, respectively. By sequence analysis of the SSU rRNA gene, the occurrence rate of Cryptosporidium spp. was 16.6% (84/506) in combined fecal specimens, with 18.2% (83/457) for mice and 2.0% (1/49) for rats. Cryptosporidium parvum ( n  = 39), C. tyzzeri ( n  = 33), and C. parvum  +  C. tyzzeri ( n  = 11) were identified in mice. Cryptosporidium parvum was only detected in one rat fecal specimen. At the gp60 locus, 71.4% (60/84) of the Cryptosporidium -positive specimens were successfully amplified, and they all came from mice. We identified five C. parvum subtypes (IIaA14G2R1, IIaA16G2R1, IIaA17G1R1, IIaA17G2R1, and IIaA18G2R1) and two C. tyzzeri subtypes (IXaA6R1 and IXbA8). Based on the identification in laboratory mice of C. parvum subtypes that have been reported previously in humans, the mice infected with this species may threaten human health, especially for people who have contact with the animals and their feces. Les espèces de Cryptosporidium peuvent infecter les humains et plus de 260 espèces animales, dont 54 espèces de rongeurs. Cependant, les données sur la présence et les caractérisations génétiques de Cryptosporidium spp. chez les rongeurs de laboratoire sont limitées. La présente étude visait à déterminer le taux de présence et les caractérisations génétiques de Cryptosporidium spp. chez des souris et des rats de laboratoire. Nous avons collecté 506 échantillons de boulettes fécales fraîches combinées (457 de souris et 49 de rats) de plus de 2 000 rongeurs de laboratoire dans la province du Heilongjiang et la ville de Shanghai, en Chine. Les Cryptosporidium spp. ont été identifiés et sous-typés par séquençage de l’ADN des gènes SSU rRNA et gp60 , respectivement. Par analyse de séquence du gène SSU rRNA, le taux de présence de Cryptosporidium spp. était de 16,6% (84/506) dans les échantillons fécaux combinés, avec 18,2 % (83/457) pour les souris et 2,0 % (1/49) pour les rats. Cryptosporidium parvum ( n  = 39), C. tyzzeri ( n  = 33) et C. parvum  +  C. tyzzeri ( n  = 11) ont été identifiés chez la souris. Cryptosporidium parvum n’a été détecté que dans un échantillon fécal de rat. Au locus gp60 , 71,4 % (60/84) des échantillons positifs à Cryptosporidium ont été amplifiés avec succès, et ils provenaient tous de souris. Nous avons identifié cinq sous-types de C. parvum (IIaA14G2R1, IIaA16G2R1, IIaA17G1R1, IIaA17G2R1 et IIaA18G2R1) et deux sous-types de C. tyzzeri (IXaA6R1 et IXbA8). Sur la base de l’identification, chez des souris de laboratoire, de sous-types de C. parvum qui ont déjà été signalés chez l’homme, les souris infectées par cette espèce peuvent menacer la santé humaine, en particulier pour les personnes qui sont en contact avec les animaux et leurs excréments.