Predicting the potential distribution of Amblyomma americanum (Acari: Ixodidae) infestation in New Zealand, using maximum entropy-based ecological niche modelling.

Although currently exotic to New Zealand, the potential geographic distribution of Amblyomma americanum (L.), the lone star tick, was modelled using maximum entropy (MaxEnt). The MaxEnt model was calibrated across the native range of A. americanum in North America using present-day climatic conditions and occurrence data from museum collections. The resulting model was then projected onto New Zealand using both present-day and future climates modelled under two greenhouse gas emission scenarios, representative concentration pathways (RCP) 4.5 (low) and RCP 8.5 (high). Three sets of WorldClim bioclimatic variables were chosen using the jackknife method and tested in MaxEnt using different combinations of model feature class functions and regularization multiplier values. The preferred model was selected based on partial receiver operating characteristic tests, the omission rate and the lowest Akaike information criterion. The final model had four bioclimatic variables, Annual Mean Temperature (BIO1), Annual Precipitation (BIO12), Precipitation Seasonality (BIO15) and Precipitation of Driest Quarter (BIO17), and the projected New Zealand distribution was broadly similar to that of Haemaphysalis longicornis Neumann, New Zealand's only livestock tick, but with a more extensive predicted suitability. The climate change predictions for the year 2050 under both low and high RCP scenarios projected only moderate increases in habitat suitability along the mountain valleys in the South Island. In conclusion, this analysis shows that given the opportunity and license A. americanum could and would successfully establish in New Zealand and could provide another vector for theileriosis organisms.

Apparent lack of efficacy of toltrazuril against Eimeria species affecting brown kiwi (Apteryx mantelli) at a captive rearing facility.

AIM: To assess the efficacy of toltrazuril against the Eimeria spp. affecting brown kiwi (Apteryx mantelli). METHODS: Droppings were collected from three brown kiwi, aged <6 months old, at a captive rearing facility in the North Island of New Zealand, between 22 February and 20 April 2017, on 14 sampling dates. Only droppings (n=30) that were excreted between 03:00 and 07:00, as determined using video surveillance, were included for analysis, reflecting the peak time for shedding of coccidial oocysts for brown kiwi. Oocysts were quantified in each sample and Eimeria species identified on the basis of oocyst morphology. All samples were collected between 2 and 10 days after the birds had been treated with 25 mg/kg toltrazuril. RESULTS: Eimeria spp. oocysts were identified in 28/30 individual samples and on 14/14 sampling dates. Oocyst counts varied from 0 to 328,080 oocysts per gram (opg), and at least one oocyst count >10,000 opg was measured on 12/14 sampling dates. Three species of Eimeria were observed, with Eimeria apteryxii and E. kiwii most commonly encountered, whereas only one sample contained E. paraurii. CONCLUSIONS AND CLINICAL RELEVANCE: In the three birds monitored at this research site, there was a high abundance of E. apteryxii and E. kiwii oocysts in droppings despite recent administration of toltrazuril. These results suggest that the populations of Eimeria spp. affecting brown kiwi at this location appear to possess an ability to survive exposure to toltrazuril. Toltrazuril is widely used at captive rearing facilities to limit the effects of coccidiosis in juvenile kiwi. If a lack of efficacy is confirmed, it will be necessary to investigate alternative treatment regimens alongside broader environmental management strategies.

Associations between Theileria orientalis Ikeda type infection and the growth rates and haematocrit of suckled beef calves in the North Island of New Zealand.

AIMS: The principle aim of this study was to examine the association between infection with Theileria orientalis Ikeda type and growth rates of suckled beef calves on four beef farms. In addition, associations between calf sex, sampling time, and individual farm and T. orientalis Ikeda type infection intensity and haematocrit (HCT) were investigated. METHODS: The study was a prospective longitudinal study in which 240 calves from four purposively selected beef farms in the North Island of New Zealand were blood sampled and weighed in late spring, mid-summer and early autumn. Two farms were from high-risk (A and B) and two from low-risk (C and D) tick areas. Blood samples were analysed to determine HCT, and the number of T. orientalis Ikeda type organisms/µL of blood (infection intensity) using a quantitative PCR assay. A calf was defined as infected if >415 organisms/µL were detected in a blood sample. Linear mixed models were used to examine associations between infection intensity, mean daily liveweight gain (MDG), HCT, calf sex and time of sampling on the four farms. RESULTS: On Farms A and B nearly all calves were infected at each sampling time, on Farm C <30% were infected at any sampling and on Farm D infection prevalence increased from 32 to 79% between late spring and early autumn. On Farms C and D, from mid-summer to early autumn, mean MDG was 0.127 (95% CI=0.072-0.183) kg/day less for infected than uninfected calves (p<0.001). On all farms MDG was negatively associated with infection intensity for mid-summer and early autumn sampling times (p=0.037). The relationship between time of sampling and infection intensity varied between farms (p<0.001), and between male and female calves (p=0.018). Females had a higher infection intensity than males at the mid-summer and early autumn samplings. The association between HCT and infection intensity varied with sampling time and farm (p=0.018). There was a strong negative association between infection intensity and HCT at the late spring sampling, but in mid-summer there was no association, and in early autumn only a weak association. CONCLUSIONS AND CLINICAL RELEVANCE: This study has shown that beef farmers in the North Island of New Zealand should be concerned about the welfare effects and economic impacts of T. orientalis Ikeda type infection in suckled beef calves.

The circadian variation of oocyst shedding of Eimeria spp. affecting brown kiwi (Apteryx mantelli).

Captive rearing of wild brown kiwi (Apteryx mantelli) is widely carried out to assist in the recovery of this declining species. As a consequence, high densities of immunologically naïve kiwi are commonly housed in semi-captive conditions, with the potential to result in substantial morbidity and mortality from coccidiosis caused by multiple species of Eimeria. Previous research has described circadian variation in oocyst shedding across multiple avian host species. The aim of this research was to describe any circadian variation in oocyst shedding in brown kiwi. Droppings were collected from brown kiwi (n = 4) at a single captive rearing facility using video surveillance to determine the time of excretion, and oocyst counts were undertaken. Results show that two of the Eimeria spp. affecting brown kiwi exhibit a peak in oocyst shedding between 03.00 and 07.00 with few or no oocysts shed between 08.00 and midnight. These results are not able to be explained by the current hypotheses theorising the evolutionary forces behind the development of this adaptive trait. Our findings increase the current understanding of the biology of the Eimeria spp. affecting brown kiwi and have important implications for the management of captive-reared kiwi, in particular for the accurate interpretation of faecal oocyst counts.

Clinical haematology and biochemistry profiles of cattle naturally infected with Theileria orientalis Ikeda type in New Zealand.

AIMS: To present the haematology and biochemistry profiles for cattle in New Zealand naturally infected with Theileria orientalis Ikeda type and investigate if the results differed between adult dairy cattle and calves aged <6 months. METHODS: Haematology and biochemistry results were obtained from blood samples from cattle which tested positive for T. orientalis Ikeda type by PCR, that were submitted to veterinary laboratories in New Zealand between October 2012 and November 2014. Data sets for haematology and biochemistry results were prepared for adult dairy cattle (n=62 and 28, respectively) and calves aged <6 months (n=62 and 28, respectively), which were matched on the basis of individual haematocrit (HCT). Results were compared between age groups when categorised by HCT. Selected variables were plotted against individual HCT, and locally weighted scatterplot smoothing (Loess) curves were fitted to the data for adult dairy cattle and calves <6 months old. RESULTS: When categorised by HCT, the proportion of samples with HCT <0.15 L/L (severe anaemia) was greater for adult dairy cattle than for beef or dairy calves, for both haematology (p<0.002) and biochemistry (p<0.001) submissions. There were differences (p<0.05) between adult dairy cattle and calves aged <6 months in the relationships between HCT and red blood cell counts, mean corpuscular volume, mean corpuscular haemoglobin, mean corpuscular haemoglobin concentrations, lymphocyte and eosinophil counts, and activities of glutamate dehydrogenase and aspartate aminotransferase. In both age groups anisocytosis was frequently recorded. The proportion of blood smears showing mild and moderate macrocytosis was greater in adults than calves (p=0.01), and mild and moderate poikilocytosis was greater in calves than adults (p=0.005). CONCLUSIONS AND CLINICAL RELEVANCE: The haematology and biochemistry changes observed in cattle infected with T. orientalis Ikeda type were consistent with extravascular haemolytic anaemia. Adult dairy cattle were more likely to be severely anaemic than calves. There were differences in haematology and biochemistry profiles between adult dairy cattle and calves, but most of these differences likely had a physiological rather than pathological basis. Overall, the haematological changes in calves aged <6 months appeared less severe than in adult dairy cattle.

Description of four new species of coccidia (Apicomplexa: Eimeriidae) from brown kiwi, Apteryx mantelli, in New Zealand.

This study used morphological techniques to describe and name four new species of coccidia from the brown kiwi (Apteryx mantelli). Four distinct eimerian oocyst species were recovered that we describe as new species. The largest of these, Eimeria paraurii n. sp. measured 32.2 × 19.8 µm and is morphologically similar to gametocytes previously described histologically in colorectal polyps (Morgan et al. in Parasitol Res 111(4):1689-1699, 2012). Eimeria apteryxii n. sp. measured 23.9 × 14.9 µm and is similar to renal oocysts described histologically in brown, rowi (A. rowii) and Haast tokoeka kiwi (A. australis "Haast") (Morgan et al. in Avian Pathol 42(2):137-146, 2013). Eimeria kiwii n. sp. measured 14.8 × 13.9 µm and resembled gametocytes described previously in kiwi intestinal epithelial cells in brown kiwi (Morgan et al. in Parasitol Res 111(4):1689-1699, 2012). Eimeria mantellii n. sp. measured 17.8 × 10.7 µm and did not appear similar to any coccidia previously described in histological studies in kiwi. These are the first species of Eimeria to be described and named from brown kiwi. Because the morphological descriptions in the present study were determined from a limited number of kiwi droppings from two geographical locations, it is likely that these represent only a portion of Eimeria species present in other populations of both brown kiwi and other Apteryx species from around New Zealand.

Cluster analysis of the clinical histories of cattle affected with bovine anaemia associated with Theileria orientalis Ikeda type infection.

AIM To determine the most commonly used words in the clinical histories of animals naturally infected with Theileria orientalis Ikeda type; whether these words differed between cases categorised by age, farm type or haematocrit (HCT), and if there was any clustering of the common words in relation to these categories. METHODS Clinical histories were transcribed for 605 cases of bovine anaemia associated with T. orientalis (TABA), that were submitted to laboratories with blood samples which tested positive for T. orientalis Ikeda type infection by PCR analysis, between October 2012 and November 2014. χ2 tests were used to determine whether the proportion of submissions for each word was similar across the categories of HCT (normal, moderate anaemia or severe anaemia), farm type (dairy or beef) and age (young or old). Correspondence analysis (CA) was carried out on a contingency table of the frequency of the 28 most commonly used history words, cross-tabulated by age categories (young, old or unknown). Agglomerative hierarchical clustering, using Ward's method, was then performed on the coordinates from the correspondence analysis. RESULTS The six most commonly used history words were jaundice (204/605), lethargic (162/605), pale mucous membranes (161/605), cow (151/605), anaemia (147/605), and off milk (115/605). The proportion of cases with some history words differed between categories of age, farm type and HCT. The cluster analysis indicated that the recorded history words were grouped in two main clusters. The first included the words weight loss, tachycardia, pale mucous membranes, anaemia, lethargic and thin, and was associated with adult (p<0.001), severe anaemia (p<0.001) and dairy (p<0.001). The second cluster included the words deaths, ill-thrift, calves, calf and diarrhoea, and was associated with young (p<0.001), normal HCT (p<0.001), beef (p<0.001) and moderate anaemia (p<0.001). CONCLUSIONS AND CLINICAL RELEVANCE Cluster analysis of words recorded in clinical histories submitted with blood samples from cases of TABA indicates that two potentially different disease syndromes were associated with T. orientalis Ikeda type infection. One was consistent with the affected cattle suffering from a severe regenerative extravascular haemolytic anaemia, the second displaying as ill thrift and diarrhoea, particularly in young beef cattle.

Benzimidazole resistance in Nematodirus spathiger and N. filicollis in New Zealand.

AIM: To determine the prevalence of benzimidazole resistance in Nematodirus spathiger and N. filicollis from a sample of New Zealand farms. METHODS: The efficacy of albendazole (ABZ) against Nematodirus spp. was assessed by faecal nematode egg count reduction (FECR) tests undertaken in lambs aged 3-8 months old on 27 sheep farms throughout New Zealand. On each farm, groups of 10-16 lambs were either treated with ABZ (4.75 mg/kg) or remained as untreated controls. Faecal samples were collected from all animals at the time of treatment and 7-10 days later. Faecal nematode egg counts (FEC) were performed using a modified McMaster technique. Larvae were cultured from pooled faecal samples, collected 7-10 days after treatment from each group, by incubation at 20°C for 6 weeks, 4°C for 26 weeks then 13°C for 2 weeks. The resulting third stage larvae were identified to species using a multiplex PCR assay, that identified species-specific sequences in the second internal transcribed spacer region of ribosomal DNA. The efficacy of ABZ for N. spathiger and N. filicollis was calculated from the proportion of the two species in culture and the group mean FEC before and after treatment. Only farms with a mean of 10 epg for each species in untreated samples were included for analysis. Resistance was defined as an efficacy <95%. RESULTS: On farms that met the threshold of 10 epg in faecal samples, benzimidazole resistance was found on 20/21 (95%) farms for N. spathiger compared with 4/10 (40%) farms for N. filicollis (p<0.05). In samples collected following treatment, a mean of 83 (min 46, max 100)% of Nematodirus spp. larvae recovered from the untreated groups were N. spathiger, compared with 94 (min 45, max 100)% in the ABZ treated groups (p=0.03). This change in percentage was not influenced by the overall efficacy of treatment based on the FECR test (p=0.324). CONCLUSION: The results confirm the high level of resistance in N. spathiger in New Zealand and that benzimidazole resistance was more common in N. spathiger than N. filicollis. While resistance to benzimidazole anthelmintics has been reported previously in New Zealand, this is the first report of N. filicollis being resistant to benzimidazole anthelmintics.

Using weather data to predict the presence of Lucilia spp. on sheep farms in New Zealand.

Flystrike remains an important animal health issue on New Zealand sheep farms. To date no useful predictive tool to assist farmers to develop control options has been available. The aim of this study was to use National Institute of Water and Atmospheric Research (NIWA) virtual climate station data in New Zealand to develop a weather-based model to accurately predict the presence of Lucilia spp. on sheep farms throughout New Zealand. Three LuciTrap® baited fly traps were positioned on each of eight sheep farms throughout New Zealand (5 in the North Island and 3 in the South Island). The traps were put out for both the 2018-2019 and 2019-2020 seasons. They were emptied each week and the flies morphologically identified; with the counts of Lucilia cuprina and L. sericata combined as Lucilia spp. The count data for Lucilia spp. for each week of trapping was transformed into a binary outcome and a generalised linear mixed effects models fitted to the data, with farm as a random effect. The dependent variable was Lucilia spp. flies caught, yes or no, and the independent variables were mean weekly climate variables from the nearest NIWA virtual climate station to that farm. The model was trained on the 2018-2019 catch data and tested on the 2019-2020 catch data. A cut point was identified which maximised the model's ability to correctly predict whether Lucilia spp. were present or not for the 2019-2020 catch data, and the sensitivity, specificity, accuracy, and area under the curve (AUC) of the model calculated. The final model included just 3 significant variables, mean weekly 10 cm soil temperature, mean weekly soil moisture index, and mean weekly wind speed at 10 m. Mean weekly 10 cm soil temperature accounted for 64.7% of the variance explained by the model, mean weekly soil moisture index 34.7% and mean weekly wind speed at 10 m only 0.6%. The results showed that the predictive model had a sensitivity of 0.93 (95% CI = 0.80-0.98) and a specificity of 0.75 (95% CI = 0.62-0.85), using a cut point for the probability of Lucilia spp. being present on farm = 0.383. This model provides New Zealand farmers with a tool which will allow them to know when Lucilia spp. flies will likely be present and thus more accurately plan their interventions to prevent flystrike.

Enteric coccidiosis in the brown kiwi (Apteryx mantelli).

Enteric coccidiosis may cause significant morbidity and mortality in juvenile brown kiwi (Apteryx mantelli). Morphology of sporulated oocysts indicates that at least two Eimeria species are able to infect the brown kiwi. A histological study of the endogenous stages of coccidia was undertaken in the intestinal tracts of ten naturally infected young kiwi. Sequential sectioning of the entire intestinal tract allowed identification and recording of the distribution of the various coccidial life stages. Macromeronts measuring 268 × 162 µm when mature were found mainly within the lamina propria of the proximal one third of the small intestine. A smaller form of lamina propria meront was also identified (8.7 × 6.4 µm) with a similar distribution to the macromeronts. Small meronts (4.4 × 3.8 µm) were also identified in mucosal epithelial cells, with the overall peak in distribution within the intestinal tract being distal to the lamina propria meronts. Three morphologically distinctive gametocytes were identified. Type A gametocytes contained within epithelial cells shared the same distribution as the epithelial meronts. Polyps containing large numbers of type B gametocytes within the distal intestinal tract were found in two cases, and type C gametocytes were identified throughout the entire intestinal tract in one case only. The observational nature of this study precludes complete knowledge of the parasite life cycles using histology alone. However, it is likely that each of the three morphologically distinct gametocytes represents a separate species of enteric coccidia.

Retrospective cohort study of an outbreak of cryptosporidiosis caused by a rare Cryptosporidium parvum subgenotype.

The occurrence of a gastrointestinal illness among a class of 96 undergraduate veterinary students in New Zealand prompted laboratory and questionnaire-based investigations. Cryptosporidium parvum was the only enteropathogen identified in 4/7 faecal specimens analysed. The C. parvum isolates carried a rare IIa GP60 allele, indicating a point-source outbreak. The infection source could not be microbiologically traced, but the investigation suggested contact with calves during a practical class as the most likely exposure. A total of 25/80 respondents to a questionnaire were defined as cases using a clinical case definition (31% attack rate). The inferred median incubation period was 5 days (range 0-11 days), and the median illness duration was 5-6 days (range 2-23 days), corroborating previous observations in experimental cryptosporidiosis. Disease was self-limiting, characterized by abdominal discomfort, diarrhoea, and in some cases, vomiting. Originating from a rural area and having had previously handled ruminants were associated with a significant risk reduction in males. All the three students who reported chronic use of steroid inhalers for treatment of asthma were cases. This case highlighted, once again, the potential hazard for explosive outbreaks of cryptosporidiosis.

The role of sheep (Ovis aries) in maintaining Theileria orientalis Ikeda type infection.

Theileria orientalis is a tick‒borne intracellular parasite of red blood cells that causes severe and mild infections in various ruminants worldwide. To date there have been 11 types identified within this species, of which 4 types are presently found in New Zealand cattle. Since 2012, New Zealand has suffered a substantial epidemic of infectious bovine anaemia in both dairy and beef cattle associated with the Ikeda type. The speed at which the disease spread through the North Island suggested that other species could have been involved in transmission. The aim of a series of related experiments was to test the null hypothesis that sheep cannot maintain T. orientalis Ikeda type infection or infect ticks that feed on them. Several studies were conducted over 2 years to address this hypothesis which together showed that sheep can have detectable levels of T. orientalis Ikeda type infection in both the acute and chronic phase and that Haemaphysalis longicornis larvae can become infected when feeding on sheep. No anaemia, weight loss or clinical disease was recorded in the sheep in the acute phase of infection. The levels of infection recorded in the sheep were much lower than those found in cattle, consistent with the sheep being asymptomatic carriers of T. orientalis Ikeda type infection.

A survey of gastrointestinal nematode species in red deer (Cervus elaphus) farms in New Zealand using PCR.

Gastrointestinal nematodes are recognised as an animal health issue for farmed red deer. The aim of this study was to explore the range of species infecting farmed deer herds and their farm-level prevalence in New Zealand. Faecal samples were collected from 12-24-month-old deer (n = 6-26; mean 19) on 59 farms located in the North (n = 25) and South (n = 34) Islands. Sub-samples of faeces were pooled by farm and cultured to recover third stage larvae. Twenty four larvae were randomly selected and identified to species using a multiplex PCR (total = 1217 larvae). At farm-level the most prevalent nematodes were Oesophagostomum venulosum 83% (n = 49) and the deer-specific nematodes in the subfamily Ostertagiinae (=Ostertagia-type) including, Spiculoptera asymmetrica 73% (n = 43), Ostertagia leptospicularis 47% (n = 28), Spiculoptera spiculoptera 47% (n = 28). The recently identified Trichostrongylus askivali was present on 32% (n = 19) of the farms and Oesophagostomum sikae on 17% (n = 10). In the analysis of the total number of larvae identified, the proportion was in similar order, 45% (n = 548) were O. venulosum, 14% (n = 173) S. asymmetrica, 10% (n = 124) S. spiculoptera, 9% (n = 114) O. leptospicularis, T. askivali, 3% (n = 40) and only 2% were O. sikae (n = 20). This study is the first to show the farm-level prevalence of nematode species in deer in New Zealand and the first to use PCR as a diagnostic tool. It provides data consistent with cross-infection from sheep/cattle to deer, and provided tentative insights into the proportions of the main GIN species across the deer population including O. sikae and T. askivali which have only recently been identified in New Zealand.

The efficacy of toltrazuril treatment for reducing the infection intensity of Theileria orientalis Ikeda type in dairy calves.

The aim of this study was to test the hypothesis that toltrazuril administered at 4 weeks post-turnout reduces the infection intensity of Theileria orientalis Ikeda type in dairy calves and so prevents serious clinical disease in these animals at 2-3 months of age. Two groups of 40 dairy calves on two separate dairy farms in the Waikato were followed for 16 weeks post-turnout onto pasture. On each farm, 20 calves were randomly selected and orally treated with toltrazuril (15 mg/kg) at 4 weeks post-turnout, whilst the remaining 20 calves were left untreated. All 40 calves were blood sampled and weighed at 2, 4, 6, 8, 12, and 16 weeks post-turnout i.e. 6 samplings per calf. A random subset of 10 calves from each treatment group on each farm were faecal sampled at each visit. The blood samples were used to estimate the T. orientalis Ikeda type infection intensity and haematocrit for each calf and the faecal samples were used to estimate the number of coccidia oocysts per gram of faeces. Three linear mixed effects models, to evaluate the effect of toltrazuril treatment on infection intensity, haematocrit (HCT) and weight respectively were fitted to the data. No calves on either farm developed clinical theileriosis or coccidiosis and the three mixed effects linear models, controlling for the effect of farm and days from turnout, showed that there was no effect of treatment on infection intensity (p = 0.81), on HCT (p = 0.99) and on weight gain (p = 0.79). In conclusion, this study showed no evidence supporting the use of toltrazuril to control T. orientalis Ikeda type infection levels and prevent disease.

Effects of Theileria orientalis Ikeda type infection on libido and semen quality of bulls.

There is an epidemic in New Zealand of infectious bovine anaemia associated with Theileria orientalis Ikeda type, an obligate intracellular protozoan parasite. To establish whether T. orientalis Ikeda type infection adversely affects fertility of bulls used for natural mating, a randomised controlled experimental study was conducted. Ten of 17 2-year-old Friesian bulls that had not been previously infected with T. orientalis were infected with T. orientalis Ikeda type and then evaluations occurred during a 20-week period. There were semen and libido evaluations every 2 weeks, starting 4 weeks before the date of infection. In addition, there were blood collections, for haematocrit and infection intensity evaluations, rectal temperatures recorded, and bulls weighed three times weekly for 13 weeks after infection and then once weekly until completion of the study. Physical activity meters were also attached from Days 9-60 and 65-124 post-infection. The ten bulls were successfully infected with T. orientalis Ikeda type and this resulted in a decrease in HCT to about 0.25 by 70 days post-infection. There were no effects of infection on semen quality; however, during the acute phase of infection, when the infection intensity was rapidly increasing, the infected bulls took a longer time period for repeated mounting of females, and were less dominant in the herd social heiracrchy. In conclusion, although the transitory effects on libido could reduce conception rates, the overall effects of T. orientalis Ikeda type infection on bull fertility will probably be little.

A longitudinal study of the effect of Theileria orientalis Ikeda type infection on three New Zealand dairy farms naturally infected at pasture.

The aims of this study were to monitor the change in Theileria orientalis Ikeda type infection intensity, haematocrit, milk production and reproduction on three New Zealand spring calving dairy herds, over the 2014-2015 milking season. Three spring calving dairy farms, A, B and C, from high risk (endemically stable), low risk (endemically unstable), and zero risk (disease-free) tick areas respectively were followed through the 2014-2015 milking season. On Farms, A and B, 100 cows were randomly selected at the first visit, and the same cows blood sampled every month thereafter, whilst on Farm C, the whole herd was blood sampled bimonthly (140 cows). Blood samples were tested for haematocrit, by centrifugation, and Ikeda infection intensity, using qPCR. Animals that were Ikeda type PCR positive at the first sampling were described as prevalence cases and cows that were negative at the first sampling and became PCR positive during the sampling period were described as incidence cases. Production and reproduction data were accessed through LIC MINDA® and milk production data was standardised to energy corrected milk (ECM). In addition, the effect of buparvaquone (BPQ) treatment on milk production was estimated on Farm B. The prevalence of infection at the first sampling was 100 % on Farm A, 57 % on Farm B and 26 % on Farm C. The incidence risk of infection over the sampling period on Farms B and C was 25 % and 2 % and the incident rate was 0.026 and 0.002 cases per cow-month respectively. The average infection intensity for prevalence cases on all farms was low throughout the milking season, <7000 Ikeda organisms/µL however, cases of anaemia still occurred. There was no direct effect of infection intensity on milk production or from being a prevalence case compared to an uninfected cow on milk production, across all farms. However, on Farm B there was a loss of 266 kg (95 % CI 82 ̶ 450) ECM (∼20 kg milk solids) for incidence cases and a loss of 458 kg (95 % CI 211 ̶ 710) of ECM for buparvaquone treated cows, compared to uninfected cows. No significant effect of Ikeda infection on reproduction could be shown for Farms B and C, reproductive data for Farm A was not available. The effect of T. orientalis Ikeda type infection on production and reproduction appears to be minimal once animals have passed through the acute phase of infection and reached the chronic, asymptomatic carrier phase of infection.

Humoral immune response to Klebsiella spp. In New Zealand sea lions (Phocarctos hookeri) and the passive transfer of immunity to pups.

During the 2001-02 and 2002-03 breeding seasons, epizootics of Klebsiella pneumoniae resulted in a dramatic increase of pup mortality in New Zealand sea lions (Phocarctos hookeri; NZSLs) on Enderby Island (Auckland Islands). To estimate the prevalence of infection in the NZSL population, a serologic test was developed using a Western blot and a polysaccharide antigen derived from a K. pneumoniae isolate from a NZSL pup. All archived serum samples collected between 1997 and 1998 and 2004 and 2005 at Sandy Bay Beach rookery, Enderby Island, were tested (314 pups and 302 adult females). Anti-Klebsiella antibodies were detected throughout this period, but overall, only 16% of NZSL pups between birth and 5 mo of age were seropositive compared with 95.7% of adults. There was no apparent change in antibody prevalence as a result of the two epizootics. A method to determine total immunoglobulin G (IgG) levels in sea lion serum also was developed to investigate passive immunoglobulin transfer to neonates and development of an acquired immune response. The IgG concentration was significantly lower in pups (median 2.1 mg/ml) than in adult females (median 80 mg/ml). Based on serologic results, it was not possible to determine whether K. pneumoniae was an endemic or a novel pathogen to the NZSL population because the test was not able to discriminate between Klebsiella species. However, this study suggested that the transfer of passive immunity to neonates was very low in the NZSL, especially for anti-Klebsiella antibodies.

The effect of month, farm type and latitude on the level of anaemia associated with Theileria orientalis Ikeda type infection in New Zealand cattle naturally infected at pasture.

Commencing in 2012, an epidemic of infectious bovine anaemia associated with Theileria orientalis Ikeda type has been present in New Zealand. The aims of this study were to analyse the temporal and spatial effects of T. orientalis Ikeda type infection on the sample submission rates and haematocrits of infected cattle over the first two years of the New Zealand epidemic. The data were collected from 30/08/2012 to 28/11/2014 and included all samples that met the case definition for Theileria associated bovine anaemia (TABA) and tested positive for T. orientalis Ikeda type by PCR. The sample submission rates by month and farm type were highly seasonal with dairy farm submissions peaking in September a month before beef farm submissions peaked. A second lesser peak of dairy farm submissions in April was absent for beef farms. A mixed effects model was fitted to the data and showed a significant interaction between farm production type (dairy or beef) and month of sampling (p=0.006) and between latitude and month of sampling (p=0.024). The estimated haematocrit, adjusted for month and latitude, for dairy cattle=0.125 (95%CI 0.121-0.129) and for beef cattle=0.151 (95% CI 0.138-0.165), p<0.0001. This research shows that infected beef animals tend to be less severely affected than dairy animals and that the month of sampling and latitude of the sampled farm have significant and interacting effects on the level of anaemia associated with T. orientalis Ikeda type infection.

Experimental infection of Friesian bulls with Theileria orientalis (Ikeda) and effects on the haematocrit, live weight, rectal temperature and activity.

Since 2012, New Zealand has suffered from an epidemic of infectious bovine anaemia associated with T. orientalis (Ikeda), an obligate intracellular protozoan parasite of cattle. Despite widespread agreement that T. orientalis (Ikeda) infection has impacted beef and dairy farming in New Zealand there is very little quantitative data to support this conclusion. A randomised controlled experimental study of the effect of T. orientalis (Ikeda) infection on the live weight, haematocrit (HCT), temperature and activity of 2-year-old Friesian bulls was conducted at a Massey University Research farm, Palmerston North. Ten out of seventeen 2-year-old Friesian bulls were injected intravenously with 30 mL whole blood from 2 clinical cases of Theileria-associated bovine anaemia and then followed over a period of 20 weeks. The bulls were blood sampled, had rectal temperature recorded and weighed 3 times weekly for 13 weeks and then once weekly thereafter until the end of the trial. Infection intensity was monitored using qPCR. All 10 inoculated bulls were successfully infected with T. orientalis (Ikeda). The results showed that the live weight response to infection was varied and the bulls could be divided into two groups based on this response. Four infected bulls showed a significant weight reduction of 41.5 kg (p < 0.0001), a financial loss of around NZ$112 per bull, compared with the other 6 bulls in the infected group, which were not different to the 7 uninfected controls. The live weight of the 4 poor growing bulls was significantly lower than the other 6 infected bulls from Day 71 post infection (p < 0.05). All ten infected bulls showed a similar decrease in HCT, with the lowest HCT reached around Day 60 to 80 post-infection, however the four infected bulls that grew poorly did have a significantly elevated HCT for the first 1 to 3 weeks post infection (p < 0.05). The 4 infected bulls which grew poorly also had a significantly higher infection intensity than the other infected bulls from Day 27 to Day 60 post-infection (p < 0.05). There was no pyrexia recorded in the infected group or control groups, instead there was a tendency for the infected group to have a lower rectal temperature from Day 5 to 70 post infection. The infected bulls walked on average 239 steps per day less than the control bulls, although this difference was not significant (p = 0.35). Overall the study clearly showed, by controlling infection date and infectious dose, that a proportion of cattle infected with T. orientalis (Ikeda) have significantly decreased live weight gains.

Prevalence studies for Theileria orientalis conducted during the early stages of the 2012 New Zealand epidemic of Theileria associated bovine anaemia.

Since 1982 there have been two epidemics of bovine anaemia associated with Theileria orientalis infection (TABA) in New Zealand. The latest and more serious epidemic started in 2012 and is associated with the T. orientalis (Ikeda) type. In 2013 the Ministry for Primary Industries (MPI) carried out several epidemiological investigations on the TABA epidemic, which included a series of on-farm herd prevalence studies. The aims of this paper are to present and summarise the results of these herd prevalence studies. The data were collected from March 2013 to September 2013 and comprised 26 farms. For 21/26 of these farms, a T. orientalis (Ikeda) positive index case of TABA precipitated the follow-up herd prevalence study. Each herd prevalence study involved a random sample of 10 cows for haematocrit, for piroplasm counts (number of Theileria infected RBCs per 1000 RBCs) and for PCR molecular diagnosis. Animals were diagnosed anaemic if the haematocrit ≤0.24. The results showed that for the index cases, the average haematocrit = 0.10 L/L and the average piroplasm count = 10.9 Theileria infected RBCs per 1000 RBCs, and for the herd prevalence studies the average haematocrit = 0.26 and the average piroplasm count = 6.9. The average herd prevalence of T. orientalis (Ikeda) PCR positive cattle and of anaemic cattle was 87% and 26% respectively. For dairy herds (n = 7 herds) it took, on average, 80 days for the within herd prevalence of anaemia to return to zero after the initial diagnosis. In conclusion the prevalence studies showed that infection rates and anaemia rates were already high in herds when the index case of TABA was observed. At the time, these prevalence studies significantly added to our understanding of the epidemiology of the evolving T. orientalis (Ikeda) epidemic and furthermore met the New Zealand MPI objective of being rapid and applied, fit for purpose and completed at a relatively low cost.