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

Abstract

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.

Multilocus fragment analysis of Cryptosporidium parvum from pre-weaned calves in Colombia

“…Previous studies based on sequence analysis of the gp60 gene, which is the most popular marker for Cryptosporidium subtyping, have shown the genetic richness of the protozoan in this area. A wide range of subtypes belonging to a single gp60 allelic family have been reported in Brazil (IIaA14G2R1, IIaA14G2R2, IIaA15G2R1, IIaA16G3R2, IIaA17G2R1, IIaA18G1R1, IIaA18G2R2, IIaA19G2R1, IIaA19G2R2, IIaA20G1R1; IIaA20G2R1, IIaA20G2R2) (Meireles et al, 2011;Paz e Silva et al, 2013;do Couto et al, 2014;Heckler et al, 2015;Toledo et al, 2017), Argentina (IIaA16G1R1, IIaA17G1R1, IIaA18G1R1, IIaA19G1R1, IIaA20G1R1, IIaA21G1R1, IIaA22G1R1, IIaA23G1R1) (Tomazic et al, 2013;Del Coco et al, 2014) and Chile (IIaA15G4R1, IIaA16G4R1, IIaA17G4R1) (Mercado et al, 2015). The fact that many of these subtypes were novel allelic variants or were not shared among different countries has been related to the geographic isolation of the protozoan in cattle farms (Avendaño et al, 2018).…”

“…However, the impact of bovine cryptosporidiosis in this continent is not well documented and data on the genetic diversity of C. parvum are limited. Modest numbers of specimens from calves have been characterised by gp60 sequencing in Brazil, Argentina or Chile (Meireles et al, 2011;Paz e Silva et al, 2013;Tomazic et al, 2013;Del Coco et al, 2014;do Couto et al, 2014;Heckler et al, 2015;Mercado et al, 2015;Toledo et al, 2017) and no previous work with VNTR markers has so far been conducted. In Colombia, the single genotyping study reporting the distribution of Cryptosporidium species and gp60 subtypes among pre-weaned calves highlighted this protozoan as a common and widespread pathogen in the central area of the country (Avendaño et al, 2018).…”

Waterborne Giardia and Cryptosporidium: contamination of human drinking water by sewage and cattle feces

“…In a study performed in Sweden, C. parvum subtype IIa had a significant association with the aqueous feces of calves, compared to subtype IId, in addition to a higher oocyst count in the hosts (SILVERLÄS et al, 2013). In Brazil, the few studies on subtyping of bovines have identified zoonotic subtypes (MEIRELES et al, 2011;COUTO et al, 2014;HECKLER et al, 2015;TOLEDO et al, 2017). Identification of the same C. parvum subtype in humans and cattle may indicate a zoonotic infection, but zoonotic subtypes may propagate among the human population in the same way as anthroponotic diseases (ROBERTSON et al, 2014).…”

“…Although there are no documented data to prove the occurrence of cryptosporidiosis and giardiasis outbreaks and waterborne transmission in Brazil, the presence of these parasites in treated and untreated water made available for human consumption was reported by studies conducted in different regions (HACHICH et al, 2004;HELLER et al, 2004;DIAS et al, 2008;NISHI et al, 2009aNISHI et al, , 2009bMACHADO et al, 2009;RAZZOLINI et al, 2010;TOLEDO et al, 2017). As the main source of contamination of public water supply include the discharge of sanitary sewage and agricultural waste, the occurrence and concentration of these parasites in water bodies in developing countries as well as in rural areas are expected to be greater (HELLER et al, 2004).…”

“…Nishi et al (2009b) identified the presence of these protozoa in water samples collected from the river, springs, and artesian wells in the indigenous lands near the Ivaí River, in the central region of Paraná, Brazil, and attributed their presence to human and animal fecal matter contamination. Toledo et al (2017) identified cysts of Giardia spp. and C. parvum oocysts in the spring water used for human consumption in dairy farms in the western central region of Paraná, where animals were carriers of these parasites.…”