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Item Molecular typing of Leptospira spp. in farmed and wild mammals reveals new host-serovar associations in New Zealand.(Taylor and Francis Group, 2024-01-01) Wilkinson DA; Edwards M; Shum C; Moinet M; Anderson NE; Benschop J; Nisa SAIMS: To apply molecular typing to DNA isolated from historical samples to determine Leptospira spp. infecting farmed and wild mammals in New Zealand. MATERIALS AND METHODS: DNA samples used in this study were extracted from urine, serum or kidney samples (or Leptospira spp. cultures isolated from them) collected between 2007 and 2017 from a range of domestic and wildlife mammalian species as part of different research projects at Massey University. Samples were included in the study if they met one of three criteria: samples that tested positive with a lipL32 PCR for pathogenic Leptospira; samples that tested negative by lipL32 PCR but were recorded as positive to PCR for pathogenic Leptospira in the previous studies; or samples that were PCR-negative in all studies but were from animals with positive agglutination titres against serogroup Tarassovi. DNA samples were typed using PCR that targeted either the glmU or gyrB genetic loci. The resulting amplicons were sequenced and typed relative to reference sequences. RESULTS: We identified several associations between mammalian hosts and Leptospira strains/serovars that had not been previously reported in New Zealand. Leptospira borgpetersenii strain Pacifica was found in farmed red deer (Cervus elaphus) samples, L. borgpetersenii serovars Balcanica and Ballum were found in wild red deer samples, Leptospira interrogans serovar Copenhageni was found in stoats (Mustela erminea) and brushtail possums (Trichosurus vulpecula), and L. borgpetersenii was found in a ferret (Mustela putorius furo). Furthermore, we reconfirmed previously described associations including dairy cattle with L. interrogans serovars Copenhageni and Pomona and L. borgpetersenii serovars Ballum, Hardjo type bovis and strain Pacifica, sheep with L. interrogans serovar Pomona and L. borgpetersenii serovar Hardjo type bovis, brushtail possum with L. borgpetersenii serovar Balcanica, farmed deer with L. borgpetersenii serovar Hardjo type bovis and hedgehogs (Erinaceus europaeus) with L. borgpetersenii serovar Ballum. CONCLUSIONS: This study provides an updated summary of host-Leptospira associations in New Zealand and highlights the importance of molecular typing. Furthermore, strain Pacifica, which was first identified as Tarassovi using serological methods in dairy cattle in 2016, has circulated in animal communities since at least 2007 but remained undetected as serology is unable to distinguish the different genotypes. CLINICAL RELEVANCE: To date, leptospirosis in New Zealand has been diagnosed with serological typing, which is deficient in typing all strains in circulation. Molecular methods are necessary to accurately type strains of Leptospira spp. infecting mammals in New Zealand.Item A cross-sectional investigation of Leptospira at the wildlife-livestock interface in New Zealand(PLOS, 2023-09-06) Moinet M; Oosterhof H; Nisa S; Haack N; Wilkinson DA; Aberdein D; Russell JC; Vallée E; Collins-Emerson J; Heuer C; Benschop J; Stevenson BThere has been a recent upsurge in human cases of leptospirosis in New Zealand, with wildlife a suspected emerging source, but up-to-date knowledge on this topic is lacking. We conducted a cross-sectional study in two farm environments to estimate Leptospira seroprevalence in wildlife and sympatric livestock, PCR/culture prevalence in wildlife, and compare seroprevalence and prevalence between species, sex, and age groups. Traps targeting house mice (Mus musculus), black rats (Rattus rattus), hedgehogs (Erinaceus europaeus) and brushtail possums (Trichosurus vulpecula) were set for 10 trap-nights in March-April 2017 on a dairy (A) and a beef and sheep (B) farm. Trapped wild animals and an age-stratified random sample of domestic animals, namely cattle, sheep and working dogs were blood sampled. Sera were tested by microagglutination test for five serogroups and titres compared using a Proportional Similarity Index (PSI). Wildlife kidneys were sampled for culture and qPCR targeting the lipL32 gene. True prevalence in mice was assessed using occupancy modelling by collating different laboratory results. Infection profiles varied by species, age group and farm. At the MAT cut-point of ≥ 48, up to 78% of wildlife species, and 16-99% of domestic animals were seropositive. Five of nine hedgehogs, 23/105 mice and 1/14 black rats reacted to L. borgpetersenii sv Ballum. The sera of 4/18 possums and 4/9 hedgehogs reacted to L. borgpetersenii sv Hardjobovis whilst 1/18 possums and 1/9 hedgehogs reacted to Tarassovi. In ruminants, seroprevalence for Hardjobovis and Pomona ranged 0-90% and 0-71% depending on the species and age group. Titres against Ballum, Tarassovi and Copenhageni were also observed in 4-20%, 0-25% and 0-21% of domestic species, respectively. The PSI indicated rodents and livestock had the most dissimilar serological responses. Three of nine hedgehogs, 31/105 mice and 2/14 rats were carrying leptospires (PCR and/or culture positive). True prevalence estimated by occupancy modelling in mice was 38% [95% Credible Interval 26, 51%] on Farm A and 22% [11, 40%] on Farm B. In the same environment, exposure to serovars found in wildlife species was commonly detected in livestock. Transmission pathways between and within species should be assessed to help in the development of efficient mitigation strategies against Leptospira.Item Whole-Genome Sequencing and Virulome Analysis of Escherichia coli Isolated from New Zealand Environments of Contrasting Observed Land Use(American Society for Microbiology, 2022-05-10) Cookson AL; Marshall JC; Biggs PJ; Rogers LE; Collis RM; Devane M; Stott R; Wilkinson DA; Kamke J; Brightwell G; Elkins CAGeneric Escherichia coli is commonly used as an indicator of fecal contamination to assess water quality and human health risk. Where measured E. coli exceedances occur, the presence of other pathogenic microorganisms, such as Shiga toxin-producing E. coli (STEC), is assumed, but confirmatory data are lacking. Putative E. coli isolates (n = 709) were isolated from water, sediment, soil, periphyton, and feces samples (n = 189) from five sites representing native forest and agricultural environments. Ten E. coli isolates (1.41%) were stx2 positive, 19 (2.7%) were eae positive, and stx1-positive isolates were absent. At the sample level, stx2-positive E. coli (5 of 189, 2.6%) and eae-positive isolates (16 of 189, 8.5%) were rare. Using real-time PCR, these STEC-associated virulence factors were determined to be more prevalent in sample enrichments (stx1, 23.9%; stx2, 31.4%; eae, 53.7%) and positively correlated with generic E. coli isolate numbers (P < 0.05) determined using culture-based methods. Whole-genome sequencing (WGS) was undertaken on a subset of 238 isolates with assemblies representing seven E. coli phylogroups (A, B1, B2, C, D, E, and F), 22 Escherichia marmotae isolates, and 1 Escherichia ruysiae isolate. Virulence factors, including those from extraintestinal pathogenic E. coli, were extremely diverse in isolates from the different locations and were more common in phylogroup B2. Analysis of the virulome from WGS data permitted the identification of gene repertoires that may be involved in environmental fitness and broadly align with phylogroup. Although recovery of STEC isolates was low, our molecular data indicate that they are likely to be widely present in environmental samples containing diverse E. coli phylogroups. IMPORTANCE This study takes a systematic sampling approach to assess the public health risk of Escherichia coli recovered from freshwater sites within forest and farmland. The New Zealand landscape is dominated by livestock farming, and previous work has demonstrated that "recreational exposure to water" is a risk factor for human infection by Shiga toxin-producing Escherichia coli (STEC). Though STEC isolates were rarely isolated from water samples, STEC-associated virulence factors were identified more commonly from water sample culture enrichments and were associated with increased generic E. coli concentrations. Whole-genome sequencing data from both E. coli and newly described Escherichia spp. demonstrated the presence of virulence factors from E. coli pathotypes, including extraintestinal pathogenic E. coli. This has significance for understanding and interpreting the potential health risk from E. coli where water quality is poor and suggests a role of virulence factors in survival and persistence of E. coli and Escherichia spp.Item Transmission dynamics of an antimicrobial resistant Campylobacter jejuni lineage in New Zealand’s commercial poultry network(Elsevier B.V, 2021-12) Greening SS; Zhang J; Midwinter AC; Wilkinson DA; Fayaz A; Williamson DA; Anderson MJ; Gates MC; French NPUnderstanding the relative contribution of different between-farm transmission pathways is essential in guiding recommendations for mitigating disease spread. This study investigated the association between contact pathways linking poultry farms in New Zealand and the genetic relatedness of antimicrobial resistant Campylobacter jejuni Sequence Type 6964 (ST-6964), with the aim of identifying the most likely contact pathways that contributed to its rapid spread across the industry. Whole-genome sequencing was performed on 167C. jejuni ST-6964 isolates sampled from across 30 New Zealand commercial poultry enterprises. The genetic relatedness between isolates was determined using whole genome multilocus sequence typing (wgMLST). Permutational multivariate analysis of variance and distance-based linear models were used to explore the strength of the relationship between pairwise genetic associations among the C. jejuni isolates and each of several pairwise distance matrices, indicating either the geographical distance between farms or the network distance of transportation vehicles. Overall, a significant association was found between the pairwise genetic relatedness of the C. jejuni isolates and the parent company, the road distance and the network distance of transporting feed vehicles. This result suggests that the transportation of feed within the commercial poultry industry as well as other local contacts between flocks, such as the movements of personnel, may have played a significant role in the spread of C. jejuni. However, further information on the historical contact patterns between farms is needed to fully characterise the risk of these pathways and to understand how they could be targeted to reduce the spread of C. jejuni.Item Diverse Epidemiology of Leptospira serovars Notified in New Zealand, 1999-2017(MDPI (Basel, Switzerland), 14/10/2020) Nisa S; Wilkinson DA; Angelin-Bonnet O; Paine S; Cullen K; Wight J; Baker MG; Benschop JLeptospirosis in New Zealand has been strongly associated with animal-contact occupations and with serovars Hardjo and Pomona. However, recent data suggest changes in these patterns, hence, serovar-specific epidemiology of leptospirosis from 1999 to 2017 was investigated. The 19-year average annual incidence is 2.01/100,000. Early (1999-2007) and late (2008-2017) study period comparisons showed a significant increase in notifications with serovar Ballum (IRR: 1.59, 95% CI: 1.22-2.09) in all cases and serovar Tarassovi (IRR: 1.75, 95% CI: 1.13-2.78) in Europeans and a decrease in notifications with serovars Hardjo and Pomona in all cases. Incidences of Ballum peaked in winter, Hardjo peaked in spring and Tarassovi peaked in summer. Incidence was highest in Māori (2.24/100,000) with dominant serovars being Hardjo and Pomona. Stratification by occupation showed meat workers had the highest incidence of Hardjo (57.29/100,000) and Pomona (45.32/100,000), farmers had the highest incidence of Ballum (11.09/100,000) and dairy farmers had the highest incidence of Tarassovi (12.59/100,000). Spatial analysis showed predominance of Hardjo and Pomona in Hawke's Bay, Ballum in West Coast and Northland and Tarassovi in Waikato, Taranaki and Northland. This study highlights the serovar-specific heterogeneity of human leptospirosis in New Zealand that should be considered when developing control and prevention strategies.