Mammal-related Cryptosporidium infections in endemic reptiles of New Zealand.

New Zealand's endemic reptile fauna is highly threatened and pathogens causing infectious diseases may be a significant risk to already endangered species. Here, we investigate Cryptosporidium infection in captive endemic New Zealand reptiles. We found two mammal-related Cryptosporidium species (C. hominis and C. parvum) and six subtypes from three gp60 families (Ib, Ig and IIa) in 12 individuals of captive endemic Tuatara, Otago and Grand skinks, and Jewelled and Rough geckos. Cryptosporidium serpentis was identified in two Jewelled geckos using 18S. In New Zealand, C. hominis and C. parvum are associated with infections in humans and introduced domestic animals but have also been recently found in wildlife. Our finding of Cryptosporidium infection in endemic reptiles can help inform strategies to monitor the conservation of species and manage potential introductions of pathogens to in-situ and ex-situ populations.

Species and genotypes causing human cryptosporidiosis in New Zealand.

Cryptosporidium is one of the most common causes of diarrhoea around the world. Successful management and prevention of this infectious disease requires knowledge of the diversity of species and subtypes causing human disease. We use sequence data from 2598 human faecal samples collected during an 11-year period (2009-2019) to better understand the impact of different species and subtypes on public health and to gain insights into the variation of human cryptosporidiosis in New Zealand. Human cryptosporidiosis in New Zealand is caused by a high diversity of species and subtypes. Six species cause human disease in New Zealand: C. hominis, C. parvum, C. cuniculus, C. erinacei, C. meleagridis and C. tyzzeri. Sequence analysis of the gp60 gene identified 16 subtype families and 101 subtypes. Cryptosporidium hominis IbA10G2 and C. parvum IIaA18G3R1 were the most frequent causes of human cryptosporidiosis with 27% and 29% of infections, respectively. Cryptosporidium hominis presented a peak of notified human cases during autumn (March-May) whereas most cases of human cryptosporidiosis caused by C. parvum are found during the calving and lambing season in spring (September-November). We also reported some subtypes that have been rarely detected in other countries such as IbA20G2 and IIoA13G1 and a low prevalence of the hypertransmissible and virulent IIaA15G2R1. This study provides insight into the variability of cryptosporidiosis in New Zealand essential for disease management and surveillance to prevent the introduction or spread of new species and subtypes in the country.

Population structure and pathogen interaction of Escherichia coli in freshwater: Implications of land-use for water quality and public health in Aotearoa New Zealand.

Freshwater samples (n = 199) were obtained from 41 sites with contrasting land-uses (avian, low impact, dairy, urban, sheep and beef, and mixed sheep, beef and dairy) and the E. coli phylotype of 3980 isolates (20 per water sample enrichment) was determined. Eight phylotypes were identified with B1 (48.04%), B2 (14.87%) and A (14.79%) the most abundant. Escherichia marmotae (n = 22), and Escherichia ruysiae (n = 1), were rare (0.68%) suggesting that these environmental strains are unlikely to confound water quality assessments. Phylotypes A and B1 were overrepresented in dairy and urban sites (p < 0.0001), whilst B2 were overrepresented in low impact sites (p < 0.0001). Pathogens ((Salmonella, Campylobacter, Cryptosporidium or Giardia) and the presence of diarrhoeagenic E. coli-associated genes (stx and eae) were detected in 89.9% (179/199) samples, including 80.5% (33/41) of samples with putative non-recent faecal inputs. Quantitative PCR to detect microbial source tracking targets from human, ruminant and avian contamination were concordant with land-use type and E. coli phylotype abundance. This study demonstrated that a potential recreational health risk remains where pathogens occurred in water samples with low E. coli concentration, potential non-recent faecal sources, low impact sites and where human, ruminant and avian faecal sources were absent.

First report of novel assemblages and mixed infections of Giardia duodenalis in human isolates from New Zealand.

Giardia duodenalis (syn. G. intestinalis and G. lamblia) is a protozoan parasite that cause disease (giardiasis) in humans and other animals. The pathogen is classified into eight assemblages, further divided into sub-assemblages, based on genetic divergence and host specificities. There are two zoonotic subtypes known as assemblages A and B, whilst assemblages from C to H are mainly found in domesticated animals, rodents and marine mammals. Here, we report for the first time the presence of assemblage E and sub-assemblage AIII in human isolates from the South Island in New Zealand. We identified a > 99% nucleotide similarity of assemblage E and sub-assemblage AIII with sequences of the gdh gene available in GenBank from individual human samples collected in Dunedin and Christchurch, respectively. We also performed a deep sequencing approach to assess intra-host assemblage variation. The sample from Dunedin showed evidence of mixed assemblage E and zoonotic sub-assemblage BIV. The report of two novel assemblages and mixed infections provides insights into the genetic diversity, epidemiology and transmission dynamics of Giardia duodenalis in New Zealand.

High-Yield Purification of Giardia intestinalis Cysts from Fecal Samples.

Giardia is an enteric protozoan parasite that causes gastroenteritis in all classes of vertebrates. It is ranked among the leading causes of death in children under 5 years of age. Giardiasis affects approximately 280 million people worldwide annually, a situation exacerbated by the low availability of effective treatments and the lack of a vaccine. In addition, the parasite is difficult to manipulate in in vitro environments, which hampers the development of effective disease management strategies. This article highlights the development of a method for the purification of viable Giardia cysts from fecal samples, verified by a trypan blue dye exclusion test. This protocol produces a 10-fold increase in yield over current methods. By combining sucrose flotation with gated filtration, the protocol significantly reduces the amount of debris in the purified cysts suspension. Cyst viability is verified by a trypan blue dye exclusion test. The ability to purify large quantities of Giardia from fecal samples could advance the development of effective treatments to target this worldwide prevalent parasite. © 2020 Wiley Periodicals LLC. Basic Protocol: Purification of Giardia cysts from fecal samples Support Protocol: Cyst viability test.

Does land use affect pathogen presence in New Zealand drinking water supplies?

Four microbes (Campylobacter spp., Escherichia coli, Cryptosporidium spp. and Giardia spp.) were monitored in 16 waterways that supply public drinking water for 13 New Zealand towns and cities. Over 500 samples were collected from the abstraction point at each study site every three months between 2009 and 2019. The waterways represent a range from small to large, free flowing to reservoir impoundments, draining catchments of entirely native vegetation to those dominated by pastoral agriculture. We used machine learning algorithms to explore the relative contribution of land use, catchment geology, vegetation, topography, and water quality characteristics of the catchment to determining the abundance and/or presence of each microbe. Sites on rivers draining predominantly agricultural catchments, the Waikato River, Oroua River and Waiorohi Stream had all four microbes present, often in high numbers, throughout the sampling interval. Other sites, such as the Hutt River and Big Huia Creek in Wellington which drain catchments of native vegetation, never had pathogenic microbes detected, or unsafe levels of E. coli. Boosted Regression Tree models could predict abundances and presence/absence of all four microbes with good precision using a wide range of potential environmental predictors covering land use, geology, vegetation, topography, and nutrient concentrations. Models were more accurate for protozoa than bacteria but did not differ markedly in their ability to predict abundance or presence/absence. Environmental drivers of microbe abundance or presence/absence also differed depending on whether the microbe was protozoan or bacterial. Protozoa were more prevalent in waterways with lower water quality, higher numbers of ruminants in the catchment, and in September and December. Bacteria were more abundant with higher rainfall, saturated soils, and catchments with greater than 35% of the land in agriculture. Although modern water treatment protocols will usually remove many pathogens from drinking water, several recent outbreaks of waterborne disease due to treatment failures, have highlighted the need to manage water supplies on multiple fronts. This research has identified potential catchment level variables, and thresholds, that could be better managed to reduce the potential for pathogens to enter drinking water supplies.