Prevalence of Giardia duodenalis genotypes in pre-weaned dairy calves

To determine the prevalence of Giardia genotypes in pre-weaned dairy calves, fecal samples were collected from a minimum of 18, 1-7-week-old dairy calves per farm on two farms each in the states of Vermont, New York, Pennsylvania, Maryland, Virginia, North Carolina, and Florida. Samples cleaned of fecal debris and concentrated using CsCl density gradient centrifugation were stained and examined by immunofluorescence microscopy and also subjected to PCR and gene sequence analysis. Prevalence by PCR ranged from 9% on a farm in Pennsylvania to 93% on a farm in Vermont, with an average prevalence for 407 calves on 14 farms of 40%. Gene sequence analysis of the TPI, beta-giardin and 16S rRNA genes revealed 85% of the positive samples to be Assemblage E, while 15% were Assemblage A, although the percentages of these genotypes varied greatly from farm to farm. Some farms had no Assemblage A Giardia. Thus, while a majority of the calves were infected with a genotype that is not known to be infectious for humans, calves on 7 of 14 farms did harbor Assemblage A Giardia. Calves should be considered as a potential source of human infectious cysts in the environment.     

Molecular characterization of Giardia intestinalis and Cryptosporidium parvum from calves with diarrhoea in Austria and evaluation of point-of-care tests

To obtain information about the occurrence and genotype distribution of G. intestinalis and C. parvum in Austrian cattle, faecal samples from diarrhoeic calves younger than 180 days of age originating from 70 farms were examined. Of the 177 faecal samples, 27.1% were positive for Giardia cysts (immunofluorescence microscopy) and 55.4% for Cryptosporidium oocysts (phase-contrast microscopy). Positive samples were characterized by nested PCR for Giardia, 83.3% (triosephosphate isomerase; tpi) and 89.6% (β-giardin; bg) were positive, while the Cryptosporidium nested PCR returned 92.5% (60-kDa glycoprotein) positive results. Sequence analysis revealed one assemblage A-positive sample and 30 (bg) respectively 29 (tpi) assemblage E-positive samples for G. intestinalis. For C. parvum four subtypes within the IIa family (IIaA15G2R1, n = 29; IIaA19G2R2, n = 3; IIaA21G2R1, n = 2; IIaA14G1R1, n = 1) could be differentiated. Validation of two immunochromatographic point-of-care tests resulted in a sensitivity of 29.2% and 77.6%; a specificity of 98.4% and 91.1% for the detection of Giardia intestinalis and Cryptosporidium parvum, respectively. Results confirm the widespread occurrence of both protozoa in diarrhoeic calves in Austria.     

Prevalence and genotypic characterisation of Giardia in dairy calves from Western Australia and Western Canada

In this study, the prevalence of Giardia duodenalis infections was determined in Western Canadian and Western Australian dairy calves. Faecal samples were collected from Holstein calves located on a commercial dairy near Lethbridge, Alta., Canada (N=28) and from calves located on two commercial dairies located near Perth, WA, Australia (N=36). Faecal samples were examined for the presence of Giardia cysts using sucrose gradient centrifugation, followed by immunofluoresence microscopy. DNA was then extracted from Giardia isolates obtained from positive samples. A PCR based method was employed to amplify and sequence a 292bp region of the 16S-rRNA gene. Genetic sequences obtained from Giardia isolates were compared to each other and to previously sequenced isolates. Following a single faecal sample, 58% of Western Australian calves and 57% of Western Canadian calves were positive for Giardia. Geometric mean cyst counts/g of faeces were 839 for Western Australian calves and 3475 for Western Canadian calves, but these values did not differ significantly. Genetic sequences were obtained from 10 calves from Western Canada, while six sequences were obtained from Western Australian calves. Of the Western Canadian isolates, eight aligned with the proposed 'Hoofed livestock' genotype. Of the five isolates obtained from Western Australian calves, four sequences were identical to the 'Hoofed livestock' genotype. Two isolates from the Western Canadian calves and one isolate from the Western Australian calves had the identical genetic sequence to the Genotype (Assemblage) A sequence, a common human genotype. The results of this study demonstrate, for the first time, that Giardia infections occur in Western Australian calves. Also, calves from different geographical locations appear to be primarily infected with a Giardia genotype unique to hoofed livestock. However, calves can shed Giardia cysts potentially infective for humans. Thus, Giardia infections should be considered important to Australian dairy producers, and infections in calves may pose a risk to public health regardless of geographical location.     

Molecular characterization of Cryptosporidium and Giardia isolates from cattle from Portugal

Fecal samples from 291 calves and 176 adult cattle in Northern Portugal were screened for Cryptosporidium and Giardia using a formalin-ethyl acetate concentration method. Acid-fast staining techniques for Cryptosporidium oocyst identification and direct microscopic observation of fecal smears for Giardia cyst identification were performed so as immunofluorescence microscopy examination. Polymerase chain reaction methods were employed to determine the genotype of each isolate. Molecular characterization was performed using amplification and sequencing of the hsp70 and 18SrRNA genes of Cryptosporidium and beta-giardin gene and glutamate dehydrogenase for assemblage determination of Giardia duodenalis. Seventy-four out of 291 calves (25.4%) and 8 out of 176 adult bovines (4.5%) were positive for Cryptosporidium. Forty-one out of 291 calf samples (14.1%) and 1 out of 176 adults samples (0.57%) were positive for Giardia. From the Cryptosporidium positive samples we obtained 63 isolates from calves samples and 7 isolates from adult samples. Additionally, Giardia was isolated in 13 out of 41 positive samples from calves and it was also possible to isolate Giardia from the positive adult sample. Molecular characterization of the Cryptosporidium and Giardia isolates showed us that C. parvum and G. duodenalis assemblage E were the prevalent species. C. parvum may infect humans, representing a potential public health risk. On the other hand, the assemblages B and A2 of Giardia, previously described in humans, were here identified in cattle. Further studies will be needed for determine the importance of cattle as carrier of zoonotic assemblages of G. duodenalis.     

Giardia and Cryptosporidium in dairy calves in British Columbia.

A study was undertaken to determine the prevalence of Giardia infections in dairy calves and to compare Giardia and Cryptosporidium infections in calves of different ages. Fresh fecal samples were collected from 386 male and female Holstein calves (newborn to 24 wk) in 20 dairies located in the lower Fraser river valley area of British Columbia. Giardia intestinalis, Cryptosporidium parvum, and Cryptosporidium muris were enumerated in each sample after concentration by sucrose gradient centrifugation and immunofluorescent staining. Giardia was identified at all farm locations. The overall prevalence of Giardia in calves was 73% with a geometric mean cyst count of 1180 cysts per gram of feces (CI, 41 to 5014). Cryptosporidium parvum and C. muris were identified in 80% and 40% of the farms, respectively. The prevalence of C. parvum was 59%, and the geometric mean for oocysts was 457 oocysts per gram of feces (CI, 18 to 160). The prevalence of C. muris was only 2% and the mean oocyst counts were 54 oocysts per gram of feces. Giardiasis was not age dependent, and approximately 80% of the calves from 2 to 24 wk were infected. In contrast, C. parvum infections were predominant in calves 2 to 4 wk, while C. muris was demonstrated in calves older than 4 wk. Fourty-seven percent of calves with diarrhea had high numbers of Giardia cysts in their feces. Giardia infections are highly prevalent in dairy calves and should be considered in animals with diarrhea or failure to thrive.     

Prevalence of Giardia spp. infection in pre-weaned and weaned calves in relation to management factors

Two hundred and forty calf faecal samples from 16 Malaysian farms were screened by PCR for Giardia spp. The overall prevalence was 12.5% and the overall farm prevalence was 68.8% (11/16 farms). The prevalence in pre-weaned and weaned calves was 16.7% and 8.3%, respectively. Sequence analysis of 25 isolates identified all as G. duodenalis assemblage E. Management factors associated with an increased risk of infection with Giardia spp. included keeping weaned calves in pens with sand floors and calf age. Keeping pre-weaned calves in pens with concrete floors and calving in single cow calving areas decreased the risk.     

Prevalence of Giardia and Cryptosporidium in dairy calves in three areas of Norway

A study was undertaken to determine the prevalences of Cryptosporidium and Giardia in dairy calves less than 6 months of age in Norway. Faecal samples were collected from a total of 1386 calves, between 3 and 183 days of age, in 136 dairy farms from three different areas of Norway. Faecal samples were examined for Cryptosporidium oocysts and Giardia cysts after concentration and immunofluorescent staining.

Giardia was found in 93% (127 out of 136) of the farms and in 49% (679 out of 1386) of the calves. Cryptosporidium was found in 53% (72 out of 136) of the farms and in 12% (167 out of 1386) of the calves. The level of Giardia and/or Cryptosporidium was low in the majority of the infected calves.

Infection peaked in the age group 2–3 months for both Cryptosporidium and Giardia. The prevalences of both parasites were higher in samples taken during winter than in samples taken during summer, and statistically significant differences were found when prevalences in different age groups of calves were compared between the three areas. A significantly lower prevalence of Cryptosporidium was found in calves housed in shared pens that were thoroughly washed more than three times a year than in calves from pens washed less often. For Giardia there was a trend for decreasing intensity of infection with increasing age in the sampled calves. For Cryptosporidium there was a trend for increasing herd prevalence with increasing number of calves in the herd, but this trend was not statistically significant. Other parameters which were investigated such as housing, feeding or management routines were not associated with prevalence or intensity of infection with either parasite.     

Prevalence of Giardia and Cryptosporidium spp in calves from a region in New Zealand experiencing intensification of dairying

AIM: To investigate the prevalence of Giardia and Cryptosporidium spp in calves born during two spring-calving seasons in a rapidly intensifying dairying region in the South Island; to evaluate potential correlations between the prevalence of the organism and age, characteristics of faeces, and animal-housing practices; and to compare the results with those from established dairying regions in the North Island. METHODS: A longitudinal study was conducted in 2005 and 2006 on 10 dairy farms located in the Otago region, South Island, New Zealand. A total of 1,190 faecal samples were collected from calves 1-7 weeks old. Direct immunofluorescent microscopy was used to screen the faecal samples for Giardia cysts and Cryptosporidium oocysts. The prevalence of Giardia and Cryptosporidium spp detected in calves in Otago was compared with that previously measured in calves from dairying regions in the Waikato and Manawatu, in the North Island . RESULTS: On average, Giardia and Cryptosporidium spp were detected in 31% and 2.6% of all samples, respectively. The prevalence of Giardia spp cysts in faeces was higher in calves >or=3 weeks of age in 2005 (por=2 weeks of age in 2006 (p=0.07) than in younger calves. No age-related pattern was observed for Cryptosporidium spp in either year. No correlations were evident between characteristics of faeces or animal housing practices and the prevalence of either organism, which did not differ between the two dairy farming regions. CONCLUSIONS: Prevalence rates of Giardia and Cryptosporidium spp in calves 1-7 weeks old did not differ between the two geographical regions, nor did the regions' distinct climate conditions appear to influence the prevalence of either pathogen. Considering data from both years together, the presence of Giardia spp cysts in faeces appeared to increase in the first week or two after birth, so that, on average, 30-40% of animals from 3-6 weeks of age were affected. CLINICAL RELEVANCE: This is the first study to report the prevalence of Giardia and Cryptosporidium spp in dairy calves in the South Island of New Zealand.     

Prevalence and genetic characterization of Giardia and Cryptosporidium in cats from Italy

One hundred and eighty one cats living in central Italy were tested for the presence of Giardia and Cryptosporidium infection by IFAT test and specific PCRs. Overall eight (4.4%) samples were IFAT-positive for Giardia. All the IFAT-positive samples for Giardia scored positive for the PCRs, and three more samples IFAT-negative generated PCR products leading to a total 6.1% molecular positivity rate for Giardia. All the examined samples were negative for Cryptosporidium. Sequencing of samples molecularly positive to Giardia indicated that three cats harbored the zoonotic Giardia duodenalis Assemblage A, whereas all other positive animals were infected with the feline-specific G. duodenalis Assemblage F. Phylogenetic analysis carried out on the sequences obtained supported the clustering of the isolates within Assemblages A and F. The results here presented provide data on the occurrence of Giardia genotypes in cats living in close contact with humans highlighting the potential importance of this protozoan disease for the public health.     

Prevalence and genetic characterization of Giardia and Cryptosporidium in cats from Italy

One hundred and eighty one cats living in central Italy were tested for the presence of Giardia and Cryptosporidium infection by IFAT test and specific PCRs. Overall eight (4.4%) samples were IFAT-positive for Giardia. All the IFAT-positive samples for Giardia scored positive for the PCRs, and three more samples IFAT-negative generated PCR products leading to a total 6.1% molecular positivity rate for Giardia. All the examined samples were negative for Cryptosporidium. Sequencing of samples molecularly positive to Giardia indicated that three cats harbored the zoonotic Giardia duodenalis Assemblage A, whereas all other positive animals were infected with the feline-specific G. duodenalis Assemblage F. Phylogenetic analysis carried out on the sequences obtained supported the clustering of the isolates within Assemblages A and F. The results here presented provide data on the occurrence of Giardia genotypes in cats living in close contact with humans highlighting the potential importance of this protozoan disease for the public health.     

Giardia and Cryptosporidium infections in neonatal reindeer calves: Relation to the acute phase response

This longitudinal observational study was conducted to investigate the spontaneous effect of Giardia and Cryptosporidium infections on acute phase response (APR) in reindeer calves (Rangifer tarandus tarandus) in Finnish Lapland.Serum (n = 609) and faecal samples (n = 366) were collected from 54 reindeer calves aged zero to 33 days. The samples were analysed for Giardia, Cryptosporidium, acute phase proteins (APP) and γ-globulins.Linear regression models were used to investigate associations of early Giardia infection (before 12 days of life) with the response of APPs and acquiring of passive immunity.Giardia was detected in 100% and Cryptosporidium in 23% of calves. There was a negative association between early Giardia infection and γ-globulin concentrations (p = 0.032) and a positive association with serum amyloid A (SAA) concentrations (p = 0.042). The results suggest a protective effect of colostrum against Giardia infection and that early infection may induce activation of APR.     

Kunjin flaviviral encephalomyelitis in an Arabian gelding in New South Wales, Australia

Flaviviruses, including Kunjin virus, are arboviruses that cause encephalomyelitis in humans and horses. This case report describes an Arabian gelding exhibiting neurological signs of flavivirus encephalomyelitis, the diagnostic investigation and confirmation of an unreported case of Kunjin virus equine encephalomyelitis in Australia.     

Molecular characterization of Cryptosporidium in Brazilian sheep

Feces were collected from 125 sheep between January and December 2007, on ten farms in the State of Rio de Janeiro, Brazil, and examined for the presence of Cryptosporidium. Ninety samples were collected from lambs 2 to 6 months of age, and 35 were from sheep over 12 months of age. All samples were subjected to molecular analysis by polymerase chain reaction (nested PCR) in two steps of the SSU rRNA. Two samples (1.6%) from the lambs were positive, and after sequencing were identified as Cryptosporidium ubiquitum. This species has been reported worldwide and it is considered a zoonotic pathogen since it has been found and in several animal species and humans. However, because of the low frequency of C. ubiquitum found, the risk for public health in this region may not be high.     

Identification of novel Cryptosporidium genotypes in kangaroos from Western Australia

A total of 763 faecal samples were collected from western grey kangaroos (Macropus fuliginosus) in Western Australia and screened for the presence of Cryptosporidium by PCR at the 18S ribosomal RNA (rRNA) locus. Samples that were positive at the 18S locus were also amplified at the actin locus. The overall prevalence was 9.3% (71/763). At the 18S rRNA locus, sequences were obtained for 28 of the 71 positives. Sequence analysis identified four species; Cryptosporidium fayeri in seven isolates, Cryptosporidium marcopodum in four isolates, Cryptosporidium xiaoi in six isolates and a novel genotype (kangaroo genotype I) in eleven isolates. Analysis at the actin locus confirmed the genetic distinctness of the novel genotype. The results of the present study indicate that in addition to C. fayeri and C. marcopodum, kangaroos may be capable of being infected with a wider range of Cryptosporidium species and genotypes including livestock species such as C. xiaoi. The novel genotype identified in the kangaroos most likely represents a cryptic species that requires further analyses to confirm its species status.