Molecular characterization of Cryptosporidium isolates from calves in Argentina

Tomazic, Mariela Luján; Maidana, Jimena; Dominguez, Mariana Gabriela; Uriarte, Enrique Leopoldo Louge; Galarza, R.; Garro, Carlos; Florín-Christensen, Mónica; Schnittger, Leonhard

doi:10.1016/j.vetpar.2013.09.022

Cited by 24 publications

(25 citation statements)

References 41 publications

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“…In our study, we identified two strains of the IIa family in 16 stool samples: strains IIaA20G1R1 and IIaA17G2R1, which not been reported in studies in this same country. The most common strain in our study, IIaA20G1R1, has been detected in cattle in some regions of the world: Serbia and Montenegro, Sweden, and Argentina [73,74,75,76], but there are no reports of this strain in humans. The second most common strain identified in stool samples, IIaA17G2R1, has been detected in other studies on cattle [77,78] and on humans [79,80].…”

Section: Discussionmentioning

confidence: 61%

Cryptosporidium spp. and Giardia spp. in feces and water and the associated exposure factors on dairy farms

et al. 2017

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The aims of this study were to verify the prevalence of Cryptosporidium spp. and Giardia spp. in animal feces and drinking water on dairy farms and to identify a possible relation between the exposure factors and the presence of these parasites. Fecal samples from cattle and humans and water samples were collected on dairy farms in Paraná, Brazil. Analysis of (oo)cysts in the feces was performed by the modified Ziehl-Neelsen staining and centrifugal flotation in zinc sulfate. Test-positive samples were subjected to nested PCR amplification of the 18SSU ribosomal RNA gene for identification of Cryptosporidium and Giardia and of the gp60 gene for subtyping of Cryptosporidium. Microbiological analysis of water was carried out by the multiple-tube method and by means of a chromogenic substrate, and parasitological analysis was performed on 31 samples by direct immunofluorescence and nested PCR of the genes mentioned above. Identification of the species of Cryptosporidium was performed by sequencing and PCR with analysis of restriction fragment length polymorphisms. The prevalence of Giardia and Cryptosporidium was higher in calves than in adults. Among the samples of cattle feces, Cryptosporidium parvum was identified in 41 (64%), C. ryanae in eight (12.5%), C. bovis in four (6.3%), C. andersoni in five (7.8%), and a mixed infection in 20 samples (31.3%). These parasites were not identified in the samples of human feces. Thermotolerant coliform bacteria were identified in 25 samples of water (45.5%). Giardia duodenalis and C. parvum were identified in three water samples. The gp60 gene analysis of C. parvum isolates revealed the presence of two strains (IIaA20G1R1 and IIaA17G2R2) in the fecal samples and one (IIaA17G2R1) in the water samples. The presence of coliforms was associated with the water source, structure and degradation of springs, rain, and turbidity. The prevalence of protozoa was higher in calves up to six months of age. C. parvum and G. duodenalis were identified in the water of dairy farms, as were thermotolerant coliforms; these findings point to the need for guidance on handling of animals, preservation of water sources, and water treatment.

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Section: Discussionmentioning

confidence: 61%

Cryptosporidium spp. and Giardia spp. in feces and water and the associated exposure factors on dairy farms

et al. 2017

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“…Genotyping enabled the identification of relatively less dominant subtypes (IIaA17G1R1 in the Allier department, IIaA17G3R1 in the Moselle, and IIaA19G1R1 in the Ardèche). The IIaA17G1R1 subtype has previously been described in French beef cattle calves (Follet et al, 2011), as well as in many other countries such as Argentina (Tomazic et al, 2013), Estonia (Santoro et al, 2018), and the USA , for example. Our study is the first to report the presence of IIaA17G3R1 and IIaA19G1R1 subtypes in French calves.…”

Section: Discussionmentioning

confidence: 87%

Molecular characterization of Cryptosporidium isolates from diarrheal dairy calves in France

Mammeri

Chevillot

Chenafi

et al. 2019

Veterinary Parasitology: Regional Studies and Reports

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A B S T R A C TCryptosporidium is an obligate intracellular protist parasite infecting a wide range of vertebrate hosts and causes significant intestinal disease in both animals and humans, as some species are zoonotic. Cattle and especially calves have been identified as one of the most common reservoirs of this protist. However, little is known about the genetics of Cryptosporidium in calves in some regions of France. The aim of this study was to detect and isolate Cryptosporidium spp. in faecal samples from naturally infected pre-weaned calves (≤45 days-old) in France. A total of 35 diarrhoeic pre-weaned calf faecal samples were collected from 26 dairy cattle farms in six departments (French administrative provinces). Cryptosporidium presence was established by microscopically screening samples for oocystes with an immunofluorescent (DFA) staining method. DFA-positive samples were then analysed by PCR-RFLP and 18S rRNA gene sequencing to determine species. Cryptosporidium parvum-positive samples were subtyped via nested PCR analysis of a partial fragment of the 60 kDa glycoprotein (gp60) gene product. Data were then integrated into phylogenetic tree analysis. DFA revealed the presence of Cryptosporidium oocysts in 31 out of 35 (88%) samples. Combined with 18S rRNA gene analysis results, C. parvum was detected in 30 samples. Subtyping analysis in 27/30 samples (90%) of the C. parvum isolates revealed two zoonotic subtype families, IIa (24/27) and IId (3/27). Four subtypes were recognised within the subtype family IIa, including the hypertransmissible IIaA15G2R1 subtype that is the most frequently reported worldwide (21/ 27), IIaA17G3R1 (1/27), IIaA17G1R1 (1/27), and IIaA19G1R1 (1/27). Two subtypes were recognised within the IId subtype family including IIdA22G1 (2/27) and IIdA27G1 (1/27). These findings illustrate the high occurrence of Cryptosporidium in calves in dairy herds and increase the diversity of molecularly characterised C. parvum isolates with the first description of IIaA17G3R1, IIaA19G1R1, and IId subtypes in France. The presence of zoonotic C. parvum subtype families (IIa, IId) in this study suggests that pre-weaned calves are likely to be a significant reservoir of zoonotic C. parvum, and highlights the importance of animal to human cryptosporidiosis transmission risk. Further molecular studies in calves and small ruminants from other French regions are required to better understand the epidemiology of cryptosporidiosis in France.However, due to strong inter-species similarities in microscopic size and shape, species within this genus require additional identification via genetic characterization. Molecular biology techniques have enabled the description of species that are highly host-specific, as well as others that are capable of infecting many hosts. Cryptosporidium parvum is considered to be the most prevalent species worldwide and a major zoonotic transmission risk (Xiao, 2010). Using molecular approaches to genetically characterise Cryptosporidium spp. has facilitated an

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“…Molecular characterization of C. parvum at the subtype level in dairy calves, important to determine zoonotic potential, have been conducted in several countries (Santín et al 2008;Xiao and Fayer 2008;Amer et al 2010;Silverlås et al 2013;Smith et al 2014;Kaupke and Rzeżutka 2015). In Argentina, two previous studies have reported C. parvum as the only species present in 120 preweaned dairy calves and subtypes IIaA16G1R1 IIaA17G1R1, IIaA18G1R1, IIaA19G1R1, IIaA20G1R1, IIaA21G1R1, IIaA22G1R1, and IIaA23G1R1 have been identified (Tomazic et al 2013;Del Coco et al 2014). Whereas an association between subtypes and different clinical manifestations has been demonstrated in humans, few studies have tested the hypothesis that C. parvum-subtypes are associated with diarrhea intensity in dairy calves Insulander et al 2013;Adamu et al 2014).…”

Section: Introductionmentioning

confidence: 99%

Prevalence of Cryptosporidium parvum in dairy calves and GP60 subtyping of diarrheic calves in central Argentina

et al. 2019

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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 (χ 2 = 55.89, p < 0.001). For species determination, genomic DNA isolated from oocystpositive 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.

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Molecular characterization of Cryptosporidium isolates from calves in Argentina

Cited by 24 publications

References 41 publications

Cryptosporidium spp. and Giardia spp. in feces and water and the associated exposure factors on dairy farms

Cryptosporidium spp. and Giardia spp. in feces and water and the associated exposure factors on dairy farms

Molecular characterization of Cryptosporidium isolates from diarrheal dairy calves in France

Prevalence of Cryptosporidium parvum in dairy calves and GP60 subtyping of diarrheic calves in central Argentina

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