COMPARISON OF INTRAMUSCULAR FENTANYL-MIDAZOLAM, FENTANYL-MIDAZOLAM-KETAMINE, AND KETAMINE-MEDETOMIDINE FOR IMMOBILIZATION OF JAPANESE MACAQUES ( MACACA FUSCATA).

The combination of fentanyl and midazolam is commonly used as a sedative in humans. The objective of this study was to evaluate the sedative properties and physiological effects of fentanyl-midazolam and fentanyl-midazolam-ketamine compared with medetomidine-ketamine given intramuscularly in Japanese macaques ( Macaca fuscata). In a randomized crossover design, eight Japanese macaques were hand-injected with either 30 µg/kg fentanyl + 0.3 mg/kg midazolam (FM), 15 µg/kg fentanyl + 0.3 mg/kg midazolam + 5.0 mg/kg ketamine (FMK), or 0.05 mg/kg medetomidine + 5.0 mg/kg ketamine (MedK). Heart rate; indirect systolic, mean, and diastolic arterial pressure; respiratory rate; blood gas concentrations; rectal temperature; and duration of immobilization were recorded. Mixed linear models were used to evaluate the effects of drug treatment on all continuous variables, with a significance level of P < 0.05. Only three of seven animals receiving FM were successfully immobilized. All eight animals in both the FMK and MedK treatment groups had a rapid, smooth induction and were successfully immobilized. Both FMK and MedK treatments resulted in significant hypoxia and the animals required supplemental oxygen via face mask. The mean duration of FMK immobilization was 42 ± 10 min, significantly shorter than the 65 ± 14 min for the animals receiving MedK. Immobilization with MedK resulted in significantly lower heart rates, and significantly higher arterial pressure compared with FMK. Hypoventilation was significantly more pronounced in FMK-treated animals compared with MedK treatments. Immobilization with FMK resulted in a gradual, slow recovery whereas MedK-treated animals woke up more rapidly. Fentanyl-midazolam alone is not a useful sedative in Japanese macaques. A combination of fentanyl and midazolam with ketamine can be used as an alternative to medetomidine-ketamine in this species.

A comparison of West Nile virus surveillance using survival analyses of dead corvid and mosquito pool data in Ontario, 2002-2008.

The aim of this study was to improve understanding of the relative performance of the use of dead wild corvids and mosquito pools infected with West Nile virus (WNv) in surveillance for WNv activity in the environment. To this end, all records on dead corvid submissions and mosquito pools tested in Public Health Units (PHUs) in Ontario, from 2002 to 2008, were explored. Survival analyses were employed using the first-WNv-positive cases detected each year for each PHU, and censored observations for PHUs which did not detect WNv during a given year using each data source (504 observations). Survival analyses were employed to compare the number of surveillance weeks before WNv was detected by either data source, and the influence of temporal, geographic and sociodemographic factors on these data. The outcome measurement for the final accelerated failure time (AFT) model with log-logistic distribution was a time ratio, which represents the ratio of the survival time of one group relative to another. Dead corvid surveillance was faster at detecting WNv than testing mosquito pools during the early years of WNv incursion into Ontario, while mosquito testing found WNv more quickly later in the study period. There was also regional variation in time-to-detection of WNv, by modality, as well as for various types of urban/rural settings. In comparison to mosquito surveillance, West Nile virus was detected more quickly using dead corvid surveillance in sparsely populated regions. These areas may benefit from collection of dead corvids to optimize detection and direct early surveillance efforts. When we compared the time-to-detection of WNv using dead corvids and the onset of human cases in PHUs, we found that dead corvid surveillance was predictive of West Nile activity in health units that reported human cases during the first 3 years of the incursion into Ontario.

Prevalence of Anaplasma phagocytophilum and Babesia microti in Ixodes scapularis from a Newly Established Lyme Disease Endemic Area, the Thousand Islands Region of Ontario, Canada.

Blacklegged ticks (Ixodes scapularis) are vectors for several important human diseases, including Lyme disease, human granulocytic anaplasmosis (HGA), and human babesiosis, caused by Borrelia burgdorferi, Anaplasma phagocytophilum, and Babesia microti, respectively. The continued northward range expansion of blacklegged ticks and associated pathogens is an increasing public health concern in Canada. The Thousand Islands region of eastern Ontario has recently been identified as a new endemic area for Lyme disease in Canada, but the occurrence of other pathogens in ticks in this area has not been fully described. Our objectives were to determine the prevalence of A. phagocytophilum and B. microti in small mammals and questing ticks in the Thousand Islands area and identify the strains of A. phagocytophilum circulating in ticks in the area. Serum and larval ticks were collected from trapped small mammals, and questing ticks were collected via drag sampling from up to 12 island and mainland sites in 2006, 2009, and 2010. A. phagocytophilum was identified by PCR in 3.4% (47/1388) ticks from eight of 12 sites; the prevalence ranged from 8.9% in 2006 to 3% in 2009. All 365 ticks tested for B. microti were negative. Antibodies to A. phagocytophilum were detected in 2.8% (17/611) of white-footed mice (Peromyscus leucopus) at two of 11 sites in 2006, 2009, or 2010. All 34 A. phagocytophilum-positive ticks submitted for strain identification using single-nucleotide polymorphism genotyping assays targeting the 16S rRNA gene were identified as a variant strain (Ap variant-1), which is not commonly associated with human disease. Our findings suggest that people are at low risk of contracting HGA or human babesiosis due to locally acquired tick bites in the Thousand Islands area. However, continued surveillance is warranted as these pathogens continue to expand their ranges in North America.

Effect of glucocorticoids on expression of cutaneous antimicrobial peptides in northern leopard frogs (Lithobates pipiens).

BACKGROUND: Many species of frogs secrete cutaneous antimicrobial peptides that are capable of killing Batrachochytrium dendrobatidis. Some of these species are nonetheless susceptible to chytridiomycosis, suggesting that host factors causing dysregulation of this innate immune response may be important in pathogenesis. Since stresses, such as from environmental perturbations, are a potential cause of such dysregulation, this study investigated the effect of glucocorticoid on cutaneous gene expression of these antimicrobial peptides. RESULTS: Northern leopard frogs (Lithobates pipiens) were injected with either the corticosteroid methylprednisolone or saline every 48 h. Norepinephrine-elicited cutaneous secretions were collected every 8 days for 40 days. Gene expression of antimicrobial peptides (brevinin-1P and ranatuerin-2P) in the cutaneous secretions was measured relative to the reference genes EF1-α and RPL8 using quantitative RT-PCR. Corticosteroid treatment was associated with a significant increase in brevinin-1P gene expression, which was most notable at 24-40 days of corticosteroid administration. Ranatuerin-2P expression followed a similar but non-significant trend. CONCLUSION: This treatment protocol, including corticosteroid-administration and frequent norepinephrine-induced secretion, increased AMP gene expression in the skin of L. pipiens under these experimental conditions. The findings do not support the hypothesis that environmental stress predisposes frogs to chytridiomycosis by causing glucocorticoid-induced suppression of antimicrobial peptide defences.

Factors influencing mortality in a captive breeding population of Loggerhead Shrike, Eastern subspecies (Lanius ludovicianus ssp.) in Canada.

BACKGROUND: The Loggerhead Shrike, Eastern subspecies (Lanius ludovicianus ssp.) (LOSH) is a predatory songbird native to Eastern North America. It is estimated that there are fewer than 55 breeding pairs of this subspecies in North America. Captive breeding plays a critical role in preventing the extirpation of this subspecies from its Canadian range. Unfortunately, high numbers of unexplained deaths among young birds in the captive breeding population threatened the success of this program. This paper describes fledgling mortality in the captive breeding population, and seeks to identify factors associated with fledgling survival and, ultimately, to identify steps to mitigate fledgling mortality. RESULTS: Over the study period (2006-2011) at two breeding sites, 696 LOSH were fledged. Among these, 68 % (n = 474) were released, 10 % (n = 69) were retained in the captive breeding population, and 22 % (n = 155) died. Fledgling survival declined from 99 % in 2006 to 44 % in 2011. The odds of survival were significantly lower for fledglings that were part of a second clutch. As the number of fledglings in a clutch increased, the odds of surviving increased significantly. As the breeding female aged from one to four years of age, there was a marked increase in the odds of a fledgling surviving, which then subsequently declined as females aged further. CONCLUSIONS: Based on our analyses, clutch number (first or second), number of fledglings in the brood, and age of breeding females were significant predictors of fledgling survival. Long-term breeding management decisions will have to balance the need to increase the number of individuals and breeding pairs in the wild by releasing large numbers of young, against the need to maintain a genetically viable captive population, until the wild population is large enough to be self-sustaining.

Serum and hepatic vitamin A levels in captive and wild marine toads (Bufo marinus).

The captive breeding program for the endangered Puerto Rican crested toad (Peltophryne [Bufo] lemur) has been hampered by an undiagnosed condition called "Brown Skin Disease" (BSD). Toads develop widespread skin darkening, skin thickening and abnormal shedding and eventually succumb to a chronic loss of viability. This project evaluated the marine toad (Bufo marinus) as a model for the PRCT, examining vitamin A deficiency as a potential cause of BSD. Wild caught marine toads had significantly higher liver vitamin A concentrations (61.89 ± 63.49 µg/g) than captive born marine toads (0.58 ± 0.59 µg/g); P<0.001). A significant difference in serum vitamin A concentration was found between the captive and wild caught toads (P=0.013) and between the low vitamin A-fed and wild caught toads (P=0.004), when controlling for liver vitamin A concentrations. After captive toads were treated with topical and/or oral vitamin A, their hepatic vitamin A concentrations were similar to those of the wild toads, averaging 48.41 ± 37.03 µg/g. However, plasma vitamin A concentrations pre- and post-vitamin A supplementation did not differ statistically. We concluded that plasma vitamin A concentrations do not provide a linear indication of liver/body vitamin A status, and that both topical and oral supplementation with an oil-based vitamin A formulation can increase liver stores in amphibians. No evidence of BSD or other signs of deficiency were noted in the marine toads, although this feeding trial was relatively short (127 days). To date, clinical, pathological and research findings do not support vitamin A deficiency as a primary factor underlying BSD.

Geography, deer, and host biodiversity shape the pattern of Lyme disease emergence in the Thousand Islands Archipelago of Ontario, Canada.

In the Thousand Islands region of eastern Ontario, Canada, Lyme disease is emerging as a serious health risk. The factors that influence Lyme disease risk, as measured by the number of blacklegged tick (Ixodes scapularis) vectors infected with Borrelia burgdorferi, are complex and vary across eastern North America. Despite study sites in the Thousand Islands being in close geographic proximity, host communities differed and both the abundance of ticks and the prevalence of B. burgdorferi infection in them varied among sites. Using this archipelago in a natural experiment, we examined the relative importance of various biotic and abiotic factors, including air temperature, vegetation, and host communities on Lyme disease risk in this zone of recent invasion. Deer abundance and temperature at ground level were positively associated with tick abundance, whereas the number of ticks in the environment, the prevalence of B. burgdorferi infection, and the number of infected nymphs all decreased with increasing distance from the United States, the presumed source of this new endemic population of ticks. Higher species richness was associated with a lower number of infected nymphs. However, the relative abundance of Peromyscus leucopus was an important factor in modulating the effects of species richness such that high biodiversity did not always reduce the number of nymphs or the prevalence of B. burgdorferi infection. Our study is one of the first to consider the interaction between the relative abundance of small mammal hosts and species richness in the analysis of the effects of biodiversity on disease risk, providing validation for theoretical models showing both dilution and amplification effects. Insights into the B. burgdorferi transmission cycle in this zone of recent invasion will also help in devising management strategies as this important vector-borne disease expands its range in North America.

Validation of a shed skin corticosterone enzyme immunoassay in the African House Snake (Lamprophis fuliginosus) and its evaluation in the Eastern Massasauga Rattlesnake (Sistrurus catenatus catenatus).

This study investigates the use of an enzyme immunoassay to measure keratin glucocorticoid concentrations in reptilian shed skins. Keratin glucocorticoid concentrations were compared to fecal glucocorticoid concentrations during the period of keratin growth in the African House Snake (Lamprophis fuliginosus) and the Eastern Massasauga Rattlesnake (Sistrurus catenatus catenatus). Biochemical validation was performed for the shed skin and fecal corticosterone enzyme immunoassays in the African House Snake. Biological and physiological validations were attempted in the African House Snake. A statistically significant positive association was detected between shed skin corticosterone and the mean fecal corticosterone metabolites from 3 weeks before to 1 week after the previous ecdysis in the African House Snake. A statistically significant difference was not detected between the shed skin corticosterone concentrations of the minimally handled control and the weekly handled (or experimentally stressed) African House Snakes. Adrenocorticotropic hormone stimulation did not result in the physiological validation anticipated for shed skin corticosterone concentrations in the African House Snake.

Alveolar hydatid disease (Echinococcus multilocularis) in the liver of a Canadian dog in British Columbia, a newly endemic region.

An adult dog that lived in central British Columbia was examined because of a history of lethargy and vomiting. Histology, immunohistochemistry, and polymerase chain reaction (PCR) examination of a hepatic mass confirmed the presence of an alveolar hydatid cyst, the first description of Echinococcus multilocularis in British Columbia. We provide recommendations for case management and remind practitioners in endemic areas of western Canada that dogs can serve as definitive and, rarely, intermediate hosts for E. multilocularis.

RésuméHydatidose alvéolaire(Echinococcus multilocularis)dans le foie d'un chien canadien en Colombie-Britannique, une région nouvellement endémique. Un chien adulte habitant dans le centre de la Colombie-Britannique a été examiné en raison d'une anamnèse d'abattement et de vomissements. L'histologie, l'immunohistochimie et l'amplification en chaîne par la polymérase d'une masse hépatique ont tous confirmé la présence d'un kyste hydatique, la première description d'Echinococcus multilocularis en Colombie-Britannique. Nous présentons des recommandations pour la gestion des cas et rappelons aux praticiens dans les régions endémiques de l'Ouest canadien que les chiens peuvent servir d'hôtes définitifs, et rarement, d'hôtes intermédiaires, pour E. multilocularis.(Traduit par Isabelle Vallières).

Antimicrobial resistance in generic Escherichia coli isolates from wild small mammals living in swine farm, residential, landfill, and natural environments in southern Ontario, Canada.

To assess the impacts of different types of human activity on the development of resistant bacteria in the feces of wild small mammals, we compared the prevalences and patterns of antimicrobial resistance and resistance genes in generic Escherichia coli and Salmonella enterica isolates from fecal samples collected from wild small mammals living in four environments: swine farms, residential areas, landfills, and natural habitats. Resistance to antimicrobials was observed in E. coli isolates from animals in all environments: 25/52 (48%) animals trapped at swine farms, 6/69 (9%) animals trapped in residential areas, 3/20 (15%) animals trapped at landfills, and 1/22 (5%) animals trapped in natural habitats. Animals trapped on farms were significantly more likely to carry E. coli isolates with resistance to tetracycline, ampicillin, sulfisoxazole, and streptomycin than animals trapped in residential areas. The resistance genes sul2, aadA, and tet(A) were significantly more likely to be detected in E. coli isolates from animals trapped on farms than from those trapped in residential areas. Three S. enterica serotypes (Give, Typhimurium, and Newport) were recovered from the feces of 4/302 (1%) wild small mammals. All Salmonella isolates were pansusceptible. Our results show that swine farm origin is significantly associated with the presence of resistant bacteria and resistance genes in wild small mammals in southern Ontario, Canada. However, resistant fecal bacteria were found in small mammals living in all environments studied, indicating that environmental exposure to antimicrobials, antimicrobial residues, resistant bacteria, or resistance genes is widespread.

Avian influenza virus H13 circulating in ring-billed gulls (Larus delawarensis) in southern Ontario, Canada.

Avian influenza virus (AIV) was studied in ring-billed gulls (Larus delawarensis) in one breeding colony on Lake Erie in 2000, and two on Lake Ontario in both 2000 and 2004. Antibodies to H13 AIV were detected in 92% of adults in 2000 and 82% in 2004. Antibody prevalence in 3-wk-old chicks was 5%-30% (overall 15%) in 2000 and 21% and 76% (overall 48%) in 2004. In 5-wk-old chicks, antibody prevalence was 23%-75% (overall 53%) in 2000 and 53% and 79% (overall 66%) in 2004. Geometric mean antibody titers at 3 and 5 wk did not differ in 2000, but increased significantly at one colony in 2004. In 2000, overall prevalence of AIV isolation from cloaca in embryonated chicken eggs was 32% (3 wk old), 13% (5 wk old), and 0 (adults), but AIV was also isolated from kidney and lung in a high proportion of tissues cultured from 3-wk-old birds in one colony. Isolates from cloaca were characterized as subtype H13 by serology; all 15 tested for neuraminidase were H13N6. However, three AIV detections considered on the basis of nucleotide sequence to be subtype H16 were among the 28 detected retrospectively by PCR in archived cloacal swabs; the remainder were subtype H13. Outcome of virus isolation was not related to presence of antibody titers in chicks. The presence of antibody to AIV in chicks was associated significantly with inflammation in heart, kidney, pancreas, and liver. AIV was not isolated in 2004. AIV infected chicks annually within the first 3 wk of life, ultimately infecting the majority of birds in most colonies, but did not appear to cause clinical disease.

Paraparesis in a polar bear (Ursus maritimus) associated with West Nile virus infection.

A polar bear (Ursus maritimus) housed at the Toronto Zoo presented with acute-onset, nonambulatory paraparesis. Physical examination 24 hr after onset was otherwise unremarkable, spinal radiographs looked normal, and blood tests indicated mild dehydration. With continued deterioration in its general condition, euthanasia was elected a day later. Necropsy did not reveal a cause for the major presenting clinical signs. Serum collected at the time of initial examination was positive for West Nile virus (WNV) antibodies in a serum neutralization assay and at the time of euthanasia was positive in both a competitive enzyme-linked immunosorbent assay and in a plaque reduction neutralization assay. The major microscopic finding was a mild-to-moderate nonsuppurative meningoencephalomyelitis. WNV was not detected by immunohistochemistry in brain or spinal cord or by real-time reverse transcription-polymerase chain reaction (RT-PCR) and cell culture of brain and kidney, but it was isolated and identified by RT-PCR in second passage cell culture of spleen. Retrospective immunohistochemistry on spleen revealed rare antigen-positive cells, probably macrophages. Prevention of exposure to potentially WNV-infected mosquitoes or vaccination of captive bears against WNV should be considered.

Sarcomatoid carcinoma in the lung of an Egyptian fruit bat (Rousettus aegyptiacus).

A sarcomatoid carcinoma was diagnosed in the lung of a 10-year-old captive Egyptian fruit bat (Rousettus aegyptiacus). Both carcinomatous and sarcomatous cytologic phenotypes were identified histologically. Cells of both types stained positive for pancytokeratin and S-100. Stromal cells stained positively for muscle actin. No staining for vimentin was noted in either neoplastic or normal internal control tissues. To the authors' knowledge, this is the first report of a pulmonary sarcomatoid carcinoma in a bat, and only the third report of sarcomatoid carcinoma outside the human literature.

Risk maps for range expansion of the Lyme disease vector, Ixodes scapularis, in Canada now and with climate change.

BACKGROUND: Lyme disease is the commonest vector-borne zoonosis in the temperate world, and an emerging infectious disease in Canada due to expansion of the geographic range of the tick vector Ixodes scapularis. Studies suggest that climate change will accelerate Lyme disease emergence by enhancing climatic suitability for I. scapularis. Risk maps will help to meet the public health challenge of Lyme disease by allowing targeting of surveillance and intervention activities. RESULTS: A risk map for possible Lyme endemicity was created using a simple risk algorithm for occurrence of I. scapularis populations. The algorithm was calculated for each census sub-division in central and eastern Canada from interpolated output of a temperature-driven simulation model of I. scapularis populations and an index of tick immigration. The latter was calculated from estimates of tick dispersion distances by migratory birds and recent knowledge of the current geographic range of endemic I. scapularis populations. The index of tick immigration closely predicted passive surveillance data on I. scapularis occurrence, and the risk algorithm was a significant predictor of the occurrence of I. scapularis populations in a prospective field study. Risk maps for I. scapularis occurrence in Canada under future projected climate (in the 2020s, 2050s and 2080s) were produced using temperature output from the Canadian Coupled Global Climate Model 2 with greenhouse gas emission scenario enforcing 'A2' of the Intergovernmental Panel on Climate Change. CONCLUSION: We have prepared risk maps for the occurrence of I. scapularis in eastern and central Canada under current and future projected climate. Validation of the risk maps provides some confidence that they provide a useful first step in predicting the occurrence of I. scapularis populations, and directing public health objectives in minimizing risk from Lyme disease. Further field studies are needed, however, to continue validation and refinement of the risk maps.

Pharmacokinetic disposition of the oral iron chelator deferiprone in the white leghorn chicken.

Deferiprone is a bidentate oral iron chelator used for the treatment of transfusional iron overload in people. The purpose of this study was to determine the pharmacokinetic disposition of deferiprone in the white leghorn chicken as a potential model upon which to base therapeutic regimens for the treatment of iron storage disease (hemochromatosis) in affected avian species. A suspension of deferiprone (DFP) was administered orally at a single dose of 50 mg/kg to 10 birds that were iron-loaded (IL-DFP) and 10 non--iron-loaded control birds (NIL-DFP). After a 30-day washout period, 5 birds from the NIL-DFP group were used for a bioavailability study of deferiprone administered intravenously at the same dose. Blood samples were collected at varying intervals over a 24-hour period and were analyzed for deferiprone by high-performance liquid chromatography, then plasma concentration versus time curves were developed. Deferiprone was rapidly absorbed from the gastrointestinal tract of the chicken, with plasma concentrations effective for iron chelation in humans (>20 micromol/L) maintained for at least 8 hours after oral dosing. The half-life (mean +/- SD) of the orally administered deferiprone in the IL-DFP and NIL-DFP groups was 2.91 +/- 0.78 hours and 3.61 +/- 0.90 hours, respectively, and was 2.42 +/- 0.24 hours for deferiprone administered intravenously. The mean oral bioavailability was 93%. Deferiprone is well absorbed and widely distributed in the chicken, with a longer half-life than reported in mammals.

Pharmacokinetic disposition of the oral iron chelator deferiprone in the domestic pigeon (Columba livia).

Deferiprone is a bidentate oral iron chelator used for the treatment of iron overload in people. The purpose of this study was to determine the pharmacokinetic disposition of deferiprone in the domestic pigeon (Columba livia) and to compare the results with a previous study in the white leghorn chicken. Deferiprone (DFP) was administered orally as a suspension at a single dose of 50 mg/kg to 10 iron-loaded (IL-DFP) pigeons and 10 non--iron-loaded controls (NIL-DFP). Six NIL-DFP birds were also administered deferiprone intravenously to determine the bioavailability of the drug after a 30-day washout period. To evaluate if deferiprone induces its own metabolism, the pharmacokinetic disposition of the drug was also studied in the IL-DFP group after oral therapy with deferiprone at a dosage of 50 mg/kg q12h for 30 days. For each phase, collected blood was analyzed for deferiprone by high-performance liquid chromatography to develop a plasma concentration versus time curve. Deferiprone was rapidly absorbed from the gastrointestinal tract, with plasma concentrations effective for iron chelation maintained for at least 8 hours after administration in iron-loaded birds. The half-life (mean +/- SD) for deferiprone given orally to the IL-DFP and NIL-DFP groups was 2.98 +/- 0.85 hours and 3.26 +/- 1.25 hours, respectively, and when intravenously administered was 3.79 +/- 1.23 hours. The half-life after 30 days of treatment was 3.42 +/- 1.18 hours. Oral bioavailability was 44%. This study demonstrated that oral absorption of deferiprone is acceptable, it does not induce its own metabolism, and the drug was widely distributed in the pigeon, as it was in the chicken, with a longer half-life than that reported in mammals. Minor interspecies variations in the pharmacokinetics of deferiprone exist between chickens and pigeons.

Investigating the spatial risk distribution of West Nile virus disease in birds and humans in southern Ontario from 2002 to 2005.

BACKGROUND: The West Nile virus (WNv) became a veterinary public health concern in southern Ontario in 2001 and has continued to threaten public health. Wild bird mortality has been shown to be an indicator for tracking the geographic distribution of the WNv. The purpose of this study was to investigate the latent risk distribution of WNv disease among dead birds and humans in southern Ontario and to compare the spatial risk patterns for the period 2002-2005. The relationship between the mortality fraction in birds and incidence rate in humans was also investigated. METHODS: Choropleth maps were created to investigate the spatial variation in bird and human WNv risk for the public health units of southern Ontario. The data were smoothed by empirical Bayesian estimation before being mapped. Isopleth risk maps for both the bird and human data were created to identify high risk areas and to investigate the potential relationship between the WNv mortality fraction in birds and incidence rates in humans. This was carried out by the geostatistical prediction method of kriging. A Poisson regression analysis was used to model regional human WNv case counts as a function of the spatial coordinates in the east and north direction and the regional bird mortality fractions. The presence of disease clustering and the location of disease clusters were investigated by the spatial scan test. RESULTS: The isopleth risk maps exhibited high risk areas that were relatively constant from year to year. There was an overlap in the bird and human high risk areas, which occurred in the central-west and south-west areas of southern Ontario. The annual WNv cause-specific mortality fractions in birds for 2002 to 2005 were 31.9, 22.0, 19.2 and 25.2 positive birds per 100 birds tested, respectively. The annual human WNv incidence rates for 2002 to 2005 were 2.21, 0.76, 0.13 and 2.10 human cases per 100,000 population, respectively. The relative risk of human WNv disease was 0.72 times lower for a public health unit that was 100 km north of another public health unit. The relative risk of human WNv disease increased by the factor 1.44 with every 10 positive birds per 100 tested. The scan statistic detected disease cluster in the bird and human data. The human clusters were not significant, when the analysis was conditioned on the bird data. CONCLUSION: The study indicates a significant relationship between the spatial pattern of WNv risk in humans and birds.

Toxicity of Clostridium botulinum type E neurotoxin to Great Lakes fish: implications for avian botulism.

Since 1999, large-scale mortalities of fish-eating birds have been observed on the Great Lakes, and more specifically on Lake Erie. Type E botulism has been established as the primary cause of death. The mechanism of type E botulism exposure in fish-eating birds is unclear. Given that these birds are thought to eat live fish exclusively, it seems likely that their prey play a key role in the process, but the role of fish as potential transport vectors of botulinum neurotoxin type E (BoNT/E) to birds has not been adequately investigated. Between June 2003 and April 2004 a methodological model for exposing fish to Clostridium botulinum was developed and used to compare the sensitivity of rainbow trout (Oncorhynchus mykiss), round goby (Neogobius melanostomas), walleye (Stizostedion vitreum), and yellow perch (Perca flavescens) to four doses (0, 800, 1,500, and 4,000 Mouse Lethal Doses) of Clostridium botulinum type E neurotoxin. Each fish species expressed unique changes in both behavior and skin pigmentation prior to death. Yellow perch survived significantly longer (P < 0.05) than the three other species at all toxin treatments. Results of this study suggest that live fish can represent a significant vector for transfer of BoNT/E to birds.

Repeated low-level exposure of the round goby (Neogobius melanostomas) to Clostridium botulinum type E neurotoxin.

In a 4-mo study (June 2004-September 2004), round gobies (Neogobius melanostomas) were dosed orally every 72 hr for up to 21 days with Clostridium botulinum neurotoxin type E (BoNT/E) at one of four doses: 0, 50, 250, and 500 mouse lethal doses (MLD). Fish were observed for changes in pigmentation and behavior for the duration of the experiment. Mortality was observed with all treatments, with the exception of the 0 MLD control. Clinical signs observed were consistent with prior research and appeared to occur in a threshold manner. The mean times to death and percent mortalities were dose dependent. Hazard ratios were determined to have a significant positive (parameter estimate = 0.03) linear relationship with dose. The hazard ratio showed that per one unit dose increase, the instantaneous probability of a fish dying increased 1.02%. Postmortem analysis of experimental fish demonstrated that 11% (3/27) of fish contained detectable BoNT/E in their visceral fraction. The other 89% tested negative for BoNT/E, despite the fact that all fish died as a result of BoNT/E exposure. Therefore, botulism should not necessarily be ruled out as the cause of a fish kill, even if the fish test negative for BoNT/E.