The zoonotic potential of Giardia and Cryptosporidium in Norwegian sheep: a longitudinal investigation of 6 flocks of lambs.

Faecal samples collected from lambs on 6 Norwegian farms on 2 separate occasions during spring/summer 2008 (approximately 550 samples collected at each occasion) were examined for Cryptosporidium oocysts and Giardia cysts by immunofluorescent antibody test. Overall prevalence at the first sampling was approximately 23% for Giardia and 15% for Cryptosporidium, and at the second sampling approximately 31% for Giardia and 24% for Cryptosporidium, with substantial between-farm variation on each sampling occasion. To assess the potential public health significance of these infections, molecular analyses were conducted on 42 Giardia isolates and 42 Cryptosporidium isolates, with PCR targeted at one or both of two genes (Giardia: glutamate dehydrogenase and beta-giardin genes; Cryptosporidium: SSU rRNA and actin genes) for each parasite. Of the Giardia isolates, 41 were Assemblage E (non-zoonotic) and 1 was Assemblage B (zoonotic). Of the Cryptosporidium isolates, 35 were cervine genotype (potentially zoonotic) and 7 C. xiaoi (non-zoonotic). These results suggest that sheep in Norway are unlikely to be an important reservoir of zoonotic Giardia in Norway, but might have some public health significance with respect to Cryptosporidium.

Giardia cysts in sewage influent in Bergen, Norway 15-23 months after an extensive waterborne outbreak of giardiasis.

AIMS: In autumn/winter 2004, a large outbreak of waterborne giardiasis occurred in Bergen, Norway. Over 1 year later, the concentrations and genotypes of Giardia cysts occurring in sewage influent were studied to investigate the impact of the outbreak event on Giardia infections in the community. METHODS AND RESULTS: Sewage influent samples from four sewage treatment works (STW) serving Bergen were analysed for Giardia cysts on four occasions between 15 and 23 months after the outbreak. Cysts genotypes were investigated at one to three genes. Data from influent analysis from one of the STW before the outbreak, and from patient faecal samples analysed during the outbreak, provided baseline comparative data. Relatively high concentrations of Giardia cysts of diverse genotypes, both from Assemblages A and B, were detected at all STW. CONCLUSIONS: Comparison of data suggests that although Giardia cyst concentrations in sewage influent returned to pre-outbreak levels within 18 months after the outbreak peak, the genetic composition of the isolates remained significantly influenced by the Assemblage B isolate associated with the outbreak. SIGNIFICANCE AND IMPACT OF THE STUDY: Genotypes associated with an extensive outbreak of giardiasis continued to occur in Giardia infections in Bergen's population many months after the outbreak was considered to be over.

Molecular characterisation of Giardia isolates from clinical infections following a waterborne outbreak.

OBJECTIVES: During autumn/winter 2004/2005 an outbreak of waterborne giardiasis occurred in Bergen, Norway. Genetic characterisation at 2 genes of Giardia duodenalis isolates from samples from the outbreak peak showed significant variations between isolates. Characterisation of further isolates from patients diagnosed in the subsequent months was conducted to determine whether isolates with particular sequences might predominate, or whether the sequence variation would continue. METHODS: Genetic characterisation was conducted on 63 isolates from patients diagnosed in the 12 months subsequent to the outbreak peak. RESULTS: At the beta-giardin gene and glutamate dehydrogenase gene, particular isolate sequences within Assemblage B, gradually predominated over time. These sequences had not been the most frequently identified amongst 21 isolates from the outbreak peak. Nor were they apparently associated with a particular sequence at the triose phosphate isomerase gene. CONCLUSIONS: The predominance of particular sequences at the beta-giardin gene and glutamate dehydrogenase gene over time suggests that these sequences may be associated with enhanced transmission characteristics such as higher virulence, greater cyst environmental resistance, increased proliferation, or a combination of these factors. Alternatively greater association with clinical disease may have led to increased submission of samples with these sequences. Whether these sequences may be associated with particular symptom characteristics such as overt clinical disease, infection persistence or unresponsiveness to treatment warrants further study.

Fate of Cryptosporidium oocysts and Giardia cysts in the Norwegian aquatic environment over winter.

We investigated the survival of Cryptosporidium oocysts and Giardia cysts during winter in an aquatic environment (approximate temperature measurements between 1 and 7 degrees C) in Norway, using morphology and uptake of dyes as indicators of viability. Previous research has shown that in the terrestrial environment, shear forces caused by freeze and thaw cycles probably cause the parasites to be inactivated. Such forces occurred infrequently in the aquatic environment, as freezing of the water around the parasites was not observed during the study period (although freezing of the water surface did occur). The rate of decline in viability (log(10) N (t)/N (0)) was similar in control and experimental environments for both parasites; no Cryptosporidium oocysts with viable morphology were detected after approximately 20 weeks and no Giardia cysts with apparently viable morphology could be detected after 1 month. These results suggest that infection with these parasites in Norway is not usually from transmission stages that have over-wintered in the Norwegian environment.

Occurrence of Cryptosporidium oocysts and Giardia cysts in sewage in Norway.

Samples of sewage influent from 40 sewage treatment works (STW) throughout Norway were examined for Cryptosporidium oocysts and Giardia duodenalis cysts. Both parasites were detected frequently (80% of STW were Cryptosporidium positive; 93% of STW were Giardia positive) and at maximum concentrations of > 20,000 parasites/liter. The data suggest giardiasis is more widespread, and/or occurs with greater infection intensity, than cryptosporidiosis in Norway. STW serving higher person equivalents were more likely to be positive and had higher parasite concentrations. Parasite concentrations were used to estimate the proportion of contributing populations that could be clinically infected. For Cryptosporidium, the highest estimates were up to 5 per 100,000 individuals for two populations in eastern Norway. For Giardia, the highest estimate was 40 infected per 100,000 persons (approximately five times the usual national annual average) contributing to an STW in western Norway. As this population experienced a large waterborne giardiasis outbreak 6 months after sampling, it can be speculated that regular challenge with Giardia may occur here. Most Giardia isolates in sewage influent were assemblage A, although some assemblage B isolates were detected. There was substantial heterogeneity, but most samples contained isolates similar to genotype A3. Removal efficiencies at two STW with secondary treatment processes were estimated to be approximately 50% for Cryptosporidium and > 80% for Giardia. An STW with minimal treatment had negligible removal of both parasites. Many STW in Norway have minimal treatment and discharge effluent into rivers and lakes, thus, risk of contamination of water courses by Cryptosporidium and Giardia is considerable.

Cryptosporidium parvum infections in Bergen, Norway, during an extensive outbreak of waterborne giardiasis in autumn and winter 2004.

During a large waterborne giardiasis outbreak in Norway, many diarrheic patients were found to have Cryptosporidium infections. Gene sequencing identified these infections as Cryptosporidium parvum infections, although they were not identical. Whether these infections were due to a simultaneous outbreak of waterborne cryptosporidiosis or reflected background levels not normally detected is discussed.

Application of genotyping during an extensive outbreak of waterborne giardiasis in Bergen, Norway, during autumn and winter 2004.

During the autumn and winter of 2004 and 2005, an extensive outbreak of waterborne giardiasis occurred in Bergen, Norway. Over 1,500 patients were diagnosed with giardiasis. Analysis of water from the implicated source revealed low numbers of Giardia cysts, but the initial contamination event probably occurred up to 10 weeks previously. While sewage leakage from a residential area is now considered to be the probable source of contamination, during the episode waste from one particular septic tank was thought to be a possible source. Genotyping of cysts from the septic tank demonstrated that they were assemblage A cysts, although the sequences were not identical to any previously published sequences. For the beta-giardin gene, the closest published subgenotype was subgenotype A3; for the gdh gene, the closest published subgenotype was subgenotype A2. Genotyping of cysts from 21 patient samples revealed that they were assemblage B cysts; thus, the septic tank was unlikely to be the contamination source. Sequencing of the beta-giardin and gdh genes from patient samples and a comparison of the sequences gave complex results. For the beta-giardin gene, three isolates had sequences identical to subgenotype B3 sequences. However, other isolates had between one and four single-nucleotide polymorphisms (SNPs). For the gdh gene, none of the sequences were identical to the sequence published for subgenotype B3, and the sequences had between one and three SNPs. One isolate, which was identical to subgenotype B3 at the beta-giardin gene, was more similar to subgenotype B2 at the gdh gene. Grouping the isolates on the basis of SNPs resulted in different groups for the two genes. The results are discussed in relation to giardiasis in Norway and to other Giardia genotyping studies.

Effects of the Norwegian winter environment on Giardia cysts and Cryptosporidium oocysts.

The structural integrity of Cryptosporidium oocysts and Giardia cysts in the Norwegian winter environment was investigated. During winter 2001/2002, Cryptosporidium oocysts and Giardia cysts were placed in the upper layers of soil in different matrices contained in chambers and exposed to the Norwegian climate. Morphological characteristics and inclusion/exclusion of vital dyes were monitored and compared to refrigerated controls. Reduction in parasite numbers was recorded for all parasites, geographical locations, and matrices. Shear forces generated during freeze-thaw cycles are postulated to have disintegrated the parasites exposed to the Norwegian winter and retrospective laboratory studies support this theory. Increased dye inclusion, possibly indicative of viability loss, was also noted. The refrigerated control parasites exhibited no decline in numbers, and alteration in dye inclusion characteristics for refrigerated parasites was slower. Cryptosporidium oocysts were apparently more robust than Giardia cysts; differences between isolates were also noted. These results suggest Cryptosporidium oocysts and Giardia cysts do not persist in the Norwegian terrestrial environment over winter, and when detected, will have been excreted since the previous winter. Differences in the morphological characteristics, matrix effects, and the possible relationship of the dye data to parasite survival are discussed in relation to further studies.

Pneumonyssoides caninum, the canine nasal mite, reported for the first time in a fox (Vulpes vulpes).

This is the first report describing the finding of the canine nasal mite, Pneumonyssoides caninum, in a silver fox (Vulpes vulpes). It is also the first time P. caninum has been found in a species other than the dog (Canis familiaris). A severely debilitated 10-month-old, male silver fox was euthanised due to suspected renal failure. During autopsy, a female mite matching the description of P. caninum was found in the nasal cavity of the fox. The finding of P. caninum in the fox suggests the existence of a new host, or at the very least a transient host. The fox's role in maintaining and/or propagating canine nasal mite infection needs to be explored.