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Cervid babesiosis, caused by the protozoan hemoparasite Babesia odocoilei and transmitted by the blacklegged tick Ixodes scapularis, is an emerging disease of Canadian cervids. This pathogen has not yet been described in humans. Data are lacking on the role of migratory birds in the adventitious spread of Ba. odocoilei-infected ticks, as well as on the infection status of I. scapularis in environments used by susceptible wildlife hosts. Following a high-mortality outbreak of cervid babesiosis at the Toronto Zoo [TZ], the present study was initiated to investigate Ba. odocoilei and other tick-borne pathogens of veterinary and public health importance (Borrelia burgdorferi sensu stricto (s.s.), Anaplasma phagocytophilum, Borrelia miyamotoi, and Babesia microti) in I. scapularis at three sites in southern Ontario, Canada. Blanket dragging for questing ticks yielded I. scapularis from the three sites evaluated: TZ, Point Pelee National Park, and Long Point Bird Observatory [LPBO]. Babesia odocoilei was identified in I. scapularis collected by dragging at the TZ and at LPBO. Borrelia burgdorferi s.s. was identified in I. scapularis at all three sites. Anaplasma phagocytophilum was identified in I. scapularis collected from the TZ. During the springs of 2016 and 2017, 1102 northward-migrating birds were examined for ticks at LPBO. One or more I. scapularis were found on 3.2% of birds (n = 595) in 2016, and 6.7% (n = 507) of birds in 2017. Overall, across both years, 0.2% and 0.5% of birds carried one or more I. scapularis ticks that tested PCR-positive for Ba. odocoilei and Bo. burgdorferi s.s., respectively. These data indicate that Ba. odocoilei-positive I. scapularis are found in southern Ontario, and suggest that bird-borne ticks have the potential to contribute to range expansion of both Ba. odocoilei and Bo. burgdorferi s.s. in Canada.

Two male inland bearded dragons (Pogona vitticeps) of 5 and 6 yr of age were presented for anorexia with and without lethargy, respectively. In both cases, a firm, spherical, and mobile mass, measuring 2 cm in diameter, was palpated in the cranio-medial coelom. Radiographs were unremarkable. Ultrasonographic evaluation was highly suggestive of cholecystolithiasis. Coeliotomy confirmed the presence of a markedly enlarged gallbladder containing a cholecystolith in both cases, and cholecystectomies were performed. Both dragons resumed eating after 1–7 days and were doing well 3–6 months postoperatively. The gallbladder wall was unremarkable on histology, and the choleliths were composed of protein and calcium carbonate crystals. Cholecystolithiasis in bearded dragons has been the subject of very few case reports, probably due to its low occurrence. The lack of available data on the diagnosis and management may have led to this condition being underdiagnosed in this species. Ultrasonography was essential to the clinical diagnosis of cholecystolithiasis in our two cases and should be recommended as part of the diagnostic approach of bearded dragons with cranio-medial coelomic masses.

A 2-mo-old Vancouver Island marmot ( Marmota vancouverensis), housed at a quarantined breeding facility, presented for acute obtundation and vestibular ataxia. Physical examination revealed poor growth compared with littermates, poor nutritional condition, and mild dehydration. The animal's condition deteriorated over 24 hr, and it was euthanized following the development of generalized seizures. No gross abnormalities were observed upon postmortem evaluation. Histologic evaluation revealed severe, multifocal, granulomatous and lymphoplasmacytic meningoencephalomyelitis and interstitial nephritis, with intralesional, intracytoplasmic spore-filled, parasitophorous vacuoles and segmental, multi-organ, fibrinoid vasculitis (disseminated encephalitozoonosis). The etiologic agent was evident by hematoxylin and eosin and Gram-chromotrope stains, and confirmed as Encephalitozoon cuniculi by polymerase chain reaction on brain tissue. Sequencing of the internal transcribed spacer region of the rRNA gene showed 100% homology with E. cuniculi strain IV, which is a newly described genotype. This is the first report of encephalitozoonosis in this critically endangered species.

Mycobacteriosis was implicated in the deaths of eight Chinese gliding frogs (Rhacophorus dennysi) in a zoo population over a 3 yr period. Clinical signs included nonhealing skin lesions, cloacal prolapse, hind limb weakness, weight loss, and sudden death. Abnormalities on postmortem were proliferative or ulcerative skin lesions in four of eight, pneumonia in three of eight, and gall bladder empyema in two of eight cases. All eight clinical cases had multisystemic granulomas containing acid-fast bacilli. Tissues most commonly affected were lung (seven of eight), liver (six of eight), kidney (six of eight), spleen (five of eight), and heart (five of eight). The remaining eight clinically normal frogs in the population were euthanized: eight of eight had granulomatous lesions, with acid-fast bacilli in three of eight cases. A mycobacterial species was cultured from four of the clinical cases by the Public Health Ontario Laboratory and was initially misidentified as Mycobacterium tuberculosis complex by a commercial line-probe assay (GenoType Mycobacterium CM, Hain Lifesciences, Nehren, Germany). Further diagnostic testing using 16S rRNA gene sequencing ultimately identified the mycobacterial species as Mycobacterium marinum. The correct identification of mycobacterial species is essential in epidemiological investigations at zoological facilities, and in assessing health risks to staff and to other animals in the zoo population.

A 23-yr-old captive-born Przewalski's horse mare (Equus przewalskii) was euthanized at a Canadian zoo because of severe colic resulting from rupture of a jejunal pseudodiverticulum. An incidental finding of an encysted larval cestode within a hepatic granuloma was diagnosed on histopathology. Gel-based polymerase chain reaction (PCR) on liver tissue was positive for Echinococcus granulosus sensu lato, and deoxyribonucleic acid sequencing of the PCR product was 100% homologous with Echinococcus equinus. This appears to be the first molecular confirmation of E. equinus in North America, and the first report of cystic echinococcosis in a Przewalski's horse.

The endangered Puerto Rican crested toad (Peltophryne [Bufo] lemur) has been held and bred in zoos for release into protected areas in Puerto Rico since 1982. In 2004, several cases of a novel syndrome of skin changes in toads were noticed at the Toronto Zoo. A total of 21 toads were found to have similar lesions and the condition has been seen in several other groups of toads in subsequent years. Affected toads show an uncharacteristic sheen of dark-brown leathery skin, followed by recurring dysecdysis, reduced appetite, weight loss, and death from secondary causes. Histologically the condition is characterized by epithelial hyperplasia and hyperkeratosis, ulceration, and the presence of superficial mats of bacterial and fungal agents. No etiology has been identified and to date toads have not permanently responded to treatment with various pharmaceutical and nutritional therapies.

Babesia odocoilei, a tick-borne protozoan hemoparasite of white-tailed deer ( Odocoileus virginianus), is being increasingly recognized as a cause of disease in captive cervids in North America. Historically endemic in white-tailed deer, the natural wildlife reservoir in the southeastern US, B. odocoilei has been recently associated with hemolytic anemia in captive Eurasian tundra reindeer ( Rangifer tarandus tarandus), wapiti ( Cervus canadensis), and woodland caribou ( Rangifer tarandus caribou) in the northcentral and northeastern US and several Canadian provinces. The emergence of B. odocoilei is likely related to the northward expansion of the range of the tick vector, Ixodes scapularis, and possibly to cervid translocations. Following a disease outbreak in reindeer and wapiti at the Toronto Zoo in Ontario, Canada, we utilized a prospective postmortem survey to investigate the prevalence of B. odocoilei in wild, farmed, and zoo cervids in Ontario ( n=270) in 2016-18 by PCR and DNA sequencing of spleen samples. Zoo bovids have been suggested as potential hosts of B. odocoilei in zoos affected by cervid babesiosis, so we also collected postmortem samples from five species of bovids ( n=7) at the Toronto Zoo that died or were euthanized during this time. We detected B. odocoilei in 1% (2/142) of farmed red deer ( Cervus elaphus) as well as in 3% (1/29) of captive wapiti and 4% (3/68) of wild white-tailed deer. Tissues from all zoo bovids and caribou, zoo and wild moose ( Alces alces), and farmed white-tailed deer, wapiti-red deer hybrids, and fallow deer ( Dama dama), tested negative for B. odocoilei. No clinical cases of babesiosis were encountered during this study. These findings suggest that white-tailed deer are a potential natural wildlife reservoir for B. odocoilei in Ontario and that red deer and wapiti could serve as more-localized reservoirs.

Nine hundred and fifty-five pathology cases collected in Ontario between 1992 and 2011 from wild free-ranging Canada geese, trumpeter swans and mute swans were retrospectively evaluated for the pathology associated with avian bornavirus (ABV) infection. Cases were selected based on the presence of upper gastrointestinal impaction, central nervous system histopathology or clinical history suggestive of ABV infection. The proportion of birds meeting at least one of these criteria was significantly higher at the Toronto Zoo (30/132) than elsewhere in Ontario (21/823). Central, peripheral and autonomic nervous tissues were examined for the presence of lymphocytes and plasma cells on histopathology. The presence of virus was assessed by immunohistochemistry and reverse transcriptase-polymerase chain reaction (RT-PCR) on frozen brains and on formalin-fixed paraffin-embedded tissues. Among selected cases, 86.3% (44/51) were considered positive on histopathology, 56.8% (29/51) were positive by immunohistochemistry, and RT-PCR was positive on 88.2% (15/17) of the frozen brains and 78.4% (40/51) of the formalin-fixed paraffin-embedded samples. Histopathological lesions included gliosis and lymphoplasmacytic perivascular cuffing in brain (97.7%), spinal cord (50%), peripheral nerves (55.5%) and myenteric ganglia or nerves (62.8%), resembling lesions described in parrots affected with proventricular dilatation disease. Partial amino acid sequences of the nucleocapsid gene from seven geese were 100% identical amongst themselves and 98.1 to 100% identical to the waterfowl sequences recently described in the USA. Although ABV has been identified in apparently healthy geese, our study confirmed that ABV can also be associated with significant disease in wild waterfowl species.

Isospora greineri sp. n. and Isospora superbusi sp. n. are described from captive superb glossy starlings, Lamprotornis superbus, from the Toronto Zoo succumbing to visceral coccidiosis. Sequence data from nuclear 18S recombinant DNA (rDNA) and mitochondrial cytochrome c oxidase subunit I (COI) loci from sporulated oocysts and infected tissues (liver, lung, or spleen) demonstrated two distinct Isospora sp. genotypes that varied in their relative abundance. In the tissues of one affected bird, as well as its associated fecal sample, two distinct COI sequences (1.7 % divergence) and two distinct 18S rDNA sequences (0.6 % divergence) were found at almost the same abundance; in other specimens, one of the 18S and one of the COI sequences were less abundant than the other. In the tissues of some birds, only a single COI and single 18S sequence were present. In all cases, the same pair of 18S rDNA and COI sequences fluctuated in abundance in parallel, indicating that there were two distinct species present rather than one species with more than one COI or 18S locus. The oocysts of these new species cannot be differentiated morphologically. Sporulated oocysts of both were spherical to subspherical measuring 17.7 ± 0.22 μm by 17.1 ± 0.20 μm with a mean L/W ratio of 1.03 ± 0.004. Sporocysts were ovoid measuring 13.5 ± 0.17 μm by 9.3 ± 0.15 μm with a mean L/W ratio of 1.4 ± 0.02. Sporocysts had a small Stieda body with indistinct sub-Stieda body; each sporocyst had a compact residuum. Two morphologically similar but genetically divergent Isospora species were shown to cause simultaneous enteric and extraintestinal infections in captive superb glossy starlings.

In 2008, avian bornaviruses (ABV) were identified as the cause of proventricular dilatation disease (PDD). PDD is a significant condition of captive parrots first identified in the late 1970s. ABV infection has subsequently been shown to be widespread in wild waterfowl across the United States and Canada where the virus infects 10–20% of some populations of ducks, geese and swans. In most cases birds appear to be healthy and unaffected by the presence of the virus; however, infection can also result in severe non-suppurative encephalitis and lesions similar to those seen in parrots with PDD. ABVs are genetically diverse with seven identified genotypes in parrots and one in canaries. A unique goose genotype (ABV-CG) predominates in waterfowl in Canada and the northern United States. ABV appears to be endemic in North American waterfowl, in comparison to what appears to be an emerging disease in parrots. It is not known whether ABV can spread between waterfowl and parrots. The discovery of ABV infection in North American waterfowl suggests that European waterfowl should be evaluated for the presence of ABV, and also as a possible reservoir species for Borna disease virus (BDV), a related neurotropic virus affecting horses and sheep in central Europe. Although investigations have suggested that BDV is likely derived from a wildlife reservoir, for which the shrew and water vole are currently prime candidates, we suggest that the existence of other mammalian and avian reservoirs should not be discounted.