HEMATOLOGY AND PLASMA BIOCHEMISTRY REFERENCE INTERVALS FOR FREE-LIVING ADULT CHIMANGO CARACARA (MILVAGO CHIMANGO) DURING THE NONBREEDING SEASON.

Hematological and blood biochemistry values are useful in assessing the physiological, nutritional, and overall health status of captive and free-living wildlife. For the chimango caracara (Milvago chimango), the most common raptor species in Argentina, reference intervals (RIs) for hematology and blood biochemistry are lacking. For this study, 86 chimango caracaras were captured and studied in Mar del Plata and neighboring areas (Buenos Aires, Argentina) during winter (April-July) in 2018 and 2019. This is the first study to present RIs for 33 blood parameters in a large number of free-living chimango caracaras during the nonbreeding season. In addition, the variability of blood parameters according to sex and calendar year was analyzed. Overall, values for the studied parameters were similar to those described for other raptor species. There were significant differences between years for absolute monocyte counts, relative eosinophils, monocyte counts, glucose, phosphorus, and alanine aminotransferase. Only the relative count of eosinophils, aspartate aminotransferase, and calcium showed significant differences between the sexes. The values of absolute monocyte counts and the relative count of eosinophils and monocytes, glucose, phosphorus, and alanine aminotransferase were higher in 2019 than in 2018, whereas mean corpuscular hemoglobin and mean corpuscular hemoglobin concentration were higher in 2018. Relative eosinophil counts were higher for males than for females, and aspartate aminotransferase activity and calcium concentration were significantly higher for females than for males. The RIs for hematology and plasma biochemistry from this large number of chimango caracaras is of clinical relevance not only for chimango caracaras under medical care in rehabilitation centers but also in ecological studies aimed to investigate the physiological responses of this species to natural and anthropogenic changes.

Neutralization of "Chaco eagle" (Buteogallus coronatus) serum on some activities of Bothrops spp. venoms.

Several species of reptiles and mammals have components in their sera that can neutralize toxic components present in snake venoms. In this manuscript, we studied the neutralizing capacity of Chaco eagle's (Buteogallus coronatus) serum. This South American bird of prey eats snakes as a regular part of its diet and has anatomical features that protect from snakes' bites. The neutralizing potency of the Chaco eagle's serum was tested on lethal, hemorrhagic, procoagulant, and phospholipase activities of the venom of "yarará grande" (Bothrops alternatus) and on phospholipase activity of "yarará ñata" (Bothrops ammodytoides) venom; both snakes are known to be the prey of Chaco eagle. Sera of crested caracara (Caracara plancus-a scavenger, omnivorous pan-American bird of prey), secretary bird (Saggitarius serpentarius-an omnivorous bird of prey from Africa that can include venomous snakes in its diet), common hen (Gallus gallus), rat (Rattus norvegicus), mouse (Mus musculus), horse (Equus caballus), and dog (Canis lupus familiaris) were also tested to compare the inhibitory capacity of neutralization. To test isologous and xenologous neutralization, sera from Bothrops alternatus and white-eared opossum (Didelphis albiventris), respectively, were used due to their known inhibitory activity on Bothrops venoms. As a control for the neutralization activity, antibothropic antivenom was used. Chaco eagle's serum neutralized hemorrhagic and phospholipasic activity and slightly neutralized the coagulation and the lethal activity of Bothrops spp. venom. The neutralizing capacity was present in the non-immunoglobulin fraction of the serum, which showed components of acidic characteristics and lower molecular weight than IgY, in correspondence with the characteristics of PLA2s and SVMPs inhibitors described in sera from some snakes and mammals. These studies showed that Chaco eagle's serum neutralizes all toxic activities tested at a higher level than sera from animal species in which inhibitors of snake venoms have not been described (p < 0.05), while it is lower or similar in neutralizing capacity to white-eared opossum and B. alternatus sera.

Emerging Pathogenic Gammaproteobacteria Wohlfahrtiimonas chitiniclastica and Ignatzschineria Species in a Turkey Vulture (Cathartes aura).

New World vultures, such as turkey vultures (Cathartes aura), are obligate scavengers with large geographic ranges. In a preliminary characterization of the turkey vulture (TV) gastrointestinal microbiome in Southern California, we identified 2 recently described emerging bacterial pathogens not previously known to be associated with this avian species. High-throughput sequencing of broad-range 16S rRNA gene amplicons revealed sequences from TV cloacal swabs that were related closest to Wohlfahrtiimonas chitiniclastica and Ignatzschineria species, both Gammaproteobacteria considered by the United States Centers for Disease Control and Prevention as emerging zoonotic pathogens. None of these bacterial sequence types have been previously identified from samples obtained from the turkey vulture gastrointestinal microbiome. With the use of bioinformatics workflows previously established by our research group, we designed specific and sensitive polymerase chain reaction primer sets that represent novel diagnostic assays for the genera Wohlfahrtiimonas and Ignatzschineria. These primer sets were validated by Sanger sequence confirmation from complex TV samples. Because the genera Wohlfahrtiimonas and Ignatzschineria are both known to have dipteran hosts, the molecular diagnostic tools we present here should be useful for better understanding the role of flies, vultures, and other scavengers in the ecology and epidemiology of the genera Wohlfahrtiimonas and Ignatzschineria from a One Health perspective.

Air Sac Nematode ( Serratospiculum tendo) Infection in an Austral Peregrine Falcon ( Falco peregrinus cassini) in Argentina.

We report a case of air sac nematode ( Serratospiculum tendo) infection in an adult male Austral Peregrine Falcon ( Falco peregrinus cassini) admitted to a rehabilitation center in Mendoza Province, Argentina, in September 2017. This case of air sac nematodes reported in an Argentine raptor is only the second report of S. tendo in South America. We recommend examination of all raptors, especially those falcon species that include insects in their diet and inhabit open lands and those in rehabilitation centers or kept for falconry, education, or captive breeding. Fecal analysis and microscopic examination of oral swabs for evidence of parasites are simple noninvasive diagnostic procedures that allow easy detection of these parasites under field and captive circumstances.

ANALYSIS OF HISTORICAL MEDICAL RECORDS OF CALIFORNIA CONDORS ( GYMNOGYPS CALIFORNIANUS) ADMITTED FOR LEAD EXPOSURE TO THE LOS ANGELES ZOO AND BOTANICAL GARDENS BETWEEN 1997 AND 2012: A CASE SERIES STUDY.

Lead toxicosis remains the primary cause of death in wild populations of California condors ( Gymnogyps californianus). Many condors require medical intervention with lead chelation therapy, among other conservation measures, to survive in the wild. An analysis of historical medical records of California condors admitted for lead exposure to the Los Angeles Zoo and Botanical Gardens (LA Zoo) between 1997 and 2012 was performed to investigate clinical presentation, radiographic findings, and treatment along with the potential impact of these factors on recovery and successful release back into the wild. Of 100 individual condors presented to the LA Zoo for suspected lead toxicosis during this period, 84 condors had records for initial laboratory blood lead levels. For these 84 condors (40 [47.6%] females and 44 [52.4%] males), 277 initial laboratory blood lead levels in total were recorded because of repeated admissions. Thirty-two (38%) condors were admitted once to the zoo and 62% were admitted two or more times. Clinical signs were not observed in 87.5% of the condors admitted with median blood lead concentrations of 26 µg/dl. Of the radiographs taken on initial presentation, 65% did not show evidence of gastrointestinal metallic foreign bodies. Various treatments protocols with edetatum calcium disodium, dimercaptosuccinic acid, and lactated Ringer's solution were documented in the medical records. Of the 277 admissions, 154 admissions had a recorded outcome posttreatment of which 140 (91%) admissions were released back to the wild. This study contributes to a better understanding of how intervention and therapeutic approaches have been essential for the recovery, release, and sustainability of these birds as a wild population. Consistency in data collection of California condors admitted for diagnosis and treatment of lead toxicosis is needed to better assess the impacts of medical interventions.

West Nile and st. Louis encephalitis viruses antibodies surveillance in captive and free-ranging birds of prey from Argentina.

We evaluated the prevalence of WNV and SLEV neutralizing antibodies in captive and free-ranging raptors from Argentina by plaque-reduction neutralization test. Eighty plasma samples from 12 species were analyzed. Only one captive adult Crowned Eagle (Harpyhaliaetus coronatus) was WNV seropositive (prevalence: 1.25%; antibody titer of 1:80). Two captive Crowned Eagles were SLEV seropositive (prevalence: 2.50%; antibody titers: 1:80 and 1:40).These findings expand the geographic distribution of WNV and SLEV and confirm their activity in central and northeastern Argentina. West Nile virus activity in Argentina may represent a potential threat to Crowned Eagles and other endangered raptors in this country.

Evaluation of Multidrug Therapy With Azithromycin, Rifampin, and Ethambutol for the Treatment of Mycobacterium avium subsp avium in Ring-neck Doves (Streptopelia risoria): An Uncontrolled Clinical Study.

An uncontrolled clinical study was conducted to evaluate the efficacy of a multidrug protocol in 16 ring-neck doves ( Streptopelia risoria ) from a flock naturally infected with Mycobacterium avium subsp avium. The doves were considered infected on the basis of a high prevalence of infection in a group of 29 birds that were euthanatized from the same flock and clinical signs and pathologic results consistent with infection in the remaining birds. All birds were treated with azithromycin (43 mg/kg), rifampin (45 mg/kg), and ethambutol (30 mg/kg) administered orally once daily for 180 days. Five birds died during treatment and were confirmed positive for mycobacteriosis on postmortem examination. Of the remaining 11 birds, infection and disease were present in 9 (81.8%) at the end of the treatment. Postmortem investigation showed that 2 mycobacterial isolates were resistant to ethambutol, intermediately sensitive to rifampin, and sensitive to azithromycin. Microscopic examination of liver sections equivalent of those that would be taken for biopsy showed that biopsy as a method of monitoring birds for treatment success had poor sensitivity. Toxicity associated with drug therapy was not observed in these doves nor in 6 outwardly healthy ring-neck doves exposed to the same treatment. The results of this study showed that this protocol of azithromycin, rifampin, and ethambutol has poor efficacy when administered for 180 days for treatment of doves infected with M avium subsp avium.

A DNA vaccine expressing ENV and GAG offers partial protection against reticuloendotheliosis virus in the prairie chicken (Tympanicus cupido).

Recurring infection of reticuloendotheliosis virus (REV), an avian oncogenic gammaretrovirus, has been a major obstacle in attempts to breed and release the endangered Attwater's prairie chicken (Tympanicus cupido attwateri). The aim of this study was to develop a DNA vaccine that protects the birds against REV infection. A plasmid was constructed expressing fusion proteins of REV envelope (env) and VP22 of Gallid herpesvirus 2 or REV gag and VP22. Birds vaccinated with these recombinant plasmids developed neutralizing antibodies; showed delayed replication of virus; and had significantly less infection of lymphocytes, specifically CD4+ lymphocytes. Although the vaccine did not prevent infection, it offered partial protection. Birds in field conditions and breeding facilities could potentially benefit from increased immunity when vaccinated.

Association of the chicken MHC B haplotypes with resistance to avian coronavirus.

Clinical respiratory illness was compared in five homozygous chicken lines, originating from homozygous B2, B8, B12 and B19, and heterozygous B2/B12 birds after infection with either of two strains of the infectious bronchitis virus (IBV). All chickens used in these studies originated from White Leghorn and Ancona linages. IBV Gray strain infection of MHC homozygous B12 and B19 haplotype chicks resulted in severe respiratory disease compared to chicks with B2/B2 and B5/B5 haplotypes. Demonstrating a dominant B2 phenotype, B2/B12 birds were also more resistant to IBV. Respiratory clinical illness in B8/B8 chicks was severe early after infection, while illness resolved similar to the B5 and B2 homozygous birds. Following M41 strain infection, birds with B2/B2 and B8/B8 haplotypes were again more resistant to clinical illness than B19/B19 birds. Real time RT-PCR indicated that infection was cleared more efficiently in trachea, lungs and kidneys of B2/B2 and B8/B8 birds compared with B19/B19 birds. Furthermore, M41 infected B2/B2 and B8/B8 chicks performed better in terms of body weight gain than B19/B19 chicks. These studies suggest that genetics of B defined haplotypes might be exploited to produce chicks resistant to respiratory pathogens or with more effective immune responses.

Taking a rational approach in the treatment of avian mycobacteriosis.

Treatment for avian mycobacteriosis is still in its infancy and based on extrapolations from human medicine. The optimum drug choice, dose, or length of treatment has yet to be determined for most exotic animal species. Treatment should include multiple drugs for extended periods of time with appropriate monitoring of both drug levels and overall animal health. Risk to owners and handlers needs to be minimized through appropriate identification of the species of mycobacteri causing disease. More research is necessary on the pharmacokinetics of these drugs in other animal species and antibiotic resistance. Currently, euthanasia remains the most common action in the face of active mycobacteriosis.

Diagnosis of mycobacterial infections in the exotic pet patient with emphasis on birds.

The term "mycobacteriosis" encompasses a variety of infectious diseases of animals caused by bacteria of the genus Mycobacterium, which are chronic and debilitating diseases. More than 35 Mycobacterium spp can cause mycobacteriosis. The wide range of possible clinical signs and physical exam findings can make the antemortem diagnosis inconsistent and challenging. Proper sample collection and test modality in relation to the state of the disease process are essential. Clinicians can determine a presumptive diagnosis of mycobacteriosis, but the definitive etiologic diagnosis of mycobacteriosis relies on the correct identification of the mycobacteria through microbiological and molecular diagnostic methods.

Comparison of sampling methods, culture, acid-fast stain, and polymerase chain reaction assay for the diagnosis of mycobacteriosis in ring-neck doves (Streptopelia risoria).

Even when different diagnostic modalities are available, mycobacteriosis is difficult to diagnose in a live bird. To investigate the diagnostic value of sampling different tissues and using different diagnostic methods, we evaluated results of mycobacterial culture, Ziehl-Neelsen (ZN) staining, and single-amplification polymerase chain reaction assay (PCR) of 18 ring-neck doves (Streptopelia risoria) with confirmed natural infection with Mycobacterium avium avium. Results of testing liver biopsy, duodenal aspirate, and bone marrow aspirate samples and liver and spleen samples collected at necropsy were compared. Results showed the use of one single technique did not allow identification of all infected birds. In liver biopsy and bone marrow aspirate samples, culture had the highest sensitivity, whereas PCR assay and ZN staining had low sensitivity, and their combination was less sensitive than culture alone. Examination of ZN staining of the intestinal aspirate samples failed to detect infection in most birds. More splenic lesions contained acid-fast organisms than did liver lesions, suggesting that splenic biopsy may have the greatest potential for diagnosis of mycobacterial infection antemortem. Sensitivity was higher for postmortem examination of multiple liver sections than of a single biopsy section; therefore, obtaining multiple liver biopsy sections may increase detection of mycobacteria. Examination of multiple tissues and the use of several different diagnostic techniques significantly increases the probability of diagnosis of mycobacteriosis.

Mycobacteriosis in naturally infected ring-neck doves (Streptopelia risoria): investigation of the association between feather colour and susceptibility to infection, disease and lesions type.

Prevalence of infection and disease, the degree of organ involvement and the nature of the lesions were investigated in 11 white and 18 non-white ring-neck doves coming from a flock naturally infected with Mycobacterium avium subsp. avium. Lesions were common in the liver, spleen, lung, kidney, intestines, ovary and bone marrow. Overall, 18 out of 29 (62%) birds were considered infected with a sequevar of M. avium subsp. avium that contains serotypes 2, 3, 4 and 9. The prevalence of infection in the white doves (36.4%) was significantly lower than in the non-white morphs (77.7%). White doves had on average fewer organs affected (mean =3.1) than the non-white doves (mean =5.9). A diffuse pattern of inflammation in the liver and spleen was observed mainly in non-white doves. Focal or multifocal granulomatous inflammation of the liver and spleen was predominant in white doves. Genetic mechanisms of immunity to mycobacteriosis may be contributing or determining these differences. There are three basic colour morphs in ring-neck doves--dark or wild type, blond and white--and the alleles coding for colour are sex-linked and located on the sex (Z) chromosome. Female's single sexual chromosomed (ZW) and homozygous males (ZZ) can be white if they carry the white alleles. It is very probable that the gene or genes modulating the immune response to M. avium subsp. avium infection in these doves could be associated to these loci or at least located in the same (Z) chromosome, as the association with white colour suggests.

Gross and microscopic findings and investigation of the aetiopathogenesis of mycobacteriosis in a captive population of white-winged ducks (Cairina scutulata).

The white-winged duck (Cairina scutulata) is critically endangered. Breeding collections of this duck are established in the United Kingdom and the USA. Infection with Mycobacterium avium avium serotype 1 is a major cause of mortality in the UK collection. In this study, the aetiopathogenesis of deaths occurring in the US collection was studied. All ducks (n=21) that died over a 21-month period were examined. Mycobacteriosis was diagnosed in 20 ducks, killing 19 of them. Multifocal to diffuse granulomatous lesions, often with abundant intralesional organisms, were seen in all 20 ducks. Unusual manifestations of this disease were the extensive involvement of the respiratory system and the absence of multinucleated giant cells. Sequence analysis showed that the ducks were infected with a sequevar of M. a. avium that contains serotypes 2, 3, 4, and 9. Given that the long-term ingestion of metals affects immune function, we measured an array of such elements in the liver of six ducks. Concentrations were undetectable or low. The disseminated nature of the disease, high concentration of mycobacteria and absence of multinucleated giant cells within lesions suggest that these ducks were unable to effectively kill the mycobacteria and point to a possible defect or inhibition in cell mediated immunity. Taken together with previously reported UK data, these results suggest that captive white-winged ducks are highly susceptible to at least two sequevars of M. a. avium and that mycobacteriosis is a major threat to ex situ breeding. We hypothesize that the minimal heterozygosis previously shown in these ducks could be contributing to an apparently ineffective immune response.

First detection of Bacillus anthracis in feces of free-ranging raptors from central Argentina.

Prevalence of anthrax spores in feces of raptors was determined from samples collected in November-December 2000 and April-May 2001 in an agricultural region of Santa Fé province, Argentina. Feces were tested from 48 birds of six raptor species. One of 14 chimango caracaras (Milvago chimango) and one of eight road-side hawks (Buteo magnirostris) tested positive. The prevalence of Bacillus anthracis spores in feces for the six species was 4% (n=48). The prevalence was 7% (n=14) for chimango caracaras, 13% for road-side hawks (n=8), and 0% for the remaining species (Burrowing owl [Speotyto cunicularia] [n=17], Swainson's hawk [Buteo swainsoni] [n=3], Aplomado falcon [Falco femoralis] [n=2], and American kestrel [Falco sparverius] [n=4]). Grouped by their feeding habits, prevalence for scavenger species was not significantly different than for predators (7% vs. 3%, P>0.999). This study provides evidence that in central Argentina scavenger and non-scavenger raptors may have a role in the epidemiology of anthrax. Long-term studies to determine the extent of this potential involvement in the epidemiology of anthrax in central Argentina are required.

Malaria in an eastern screech owl (Otus asio).

Owls are frequent carriers of blood parasites but clinical malaria infections are rare. Various stages of Plasmodium subpraecox were seen in 90% of the erythrocytes of an Eastern screech owl (Otus asio) showing symptoms consistent with malaria 1 wk after admission for traumatic injuries. An additional unidentified blood parasite, either a Plasmodium or a Haemoproteus spp. was found in small numbers of red blood cells on blood films examined at admission and at day 7 postadmission. Combined infestation, trauma-induced stress, and iatrogenic corticosteroid administration are possible factors that could have induced disease. Oral treatment with mefloquine at 30 mg/kg, repeated after 12, 24, and 48 hr, proved successful in eliminating both organisms and signs of clinical disease.

Pathologic and immunohistochemical findings in goshawks (Accipiter gentilis) and great horned owls (Bubo virginianus) naturally infected with West Nile virus.

The carcasses of 25 great horned owls and 12 goshawks were investigated for West Nile virus (WNV) infection by immunohistochemistry (IHC) performed on various organs, including brain, spinal cord, heart, kidney, eye, bone marrow, spleen, liver, lungs, pancreas, intestine, and proventriculus, using a WNV-antigen-specific monoclonal antibody and by WNV-specific reverse transcriptase-polymerase chain reaction (RT-PCR), performed on fresh brain tissue only. WNV infection was diagnosed by IHC in all owls and all goshawks. WNV-specific RT-PCR amplified WNV-RNA in the brain of all goshawks but only 12 owls (48%). Cachexia was a common macroscopic finding associated with WNV infection in owls (76%). Myocarditis was occasionally macroscopically evident in goshawks (33%). Microscopically, inflammatory lesions, including lymphoplasmacytic and histiocytic encephalitis, myocarditis, endophthalmitis, and pancreatitis were present in both species but were more common and more severe in goshawks than in owls. The most characteristic brain lesion in owls was the formation of glial nodules, in particular in the molecular layer of the cerebellum, while encephalitis affecting the periventricular parenchyma of the cerebral cortex was common in the goshawks. In owls, WNV-antigen-positive cells were present usually only in very small numbers per organ. Kidney (80%), heart (39%), and cerebellum (37%) were the organs that most commonly contained WNV antigen in owls. WNV antigen was frequently widely distributed in the organs of infected goshawks, with increased amounts of WNV antigen in the heart and the cerebrum. Spleen (75%), cerebellum (66%), heart (58%), cerebrum (58%), and eye (50%) were often WNV-antigen positive in goshawks. In contrast with the goshawks, WNV antigen was not present in cerebral and retinal neurons of owls. WNV infection appears to be capable of causing fatal disease in great horned owls and goshawks. However, the distribution and severity of histologic lesions, the antigen distribution in the various organs, and the amount of antigen varied among both species. Therefore, the diagnostician may choose organs for histology and immunohistochemistry as well as RT-PCR depending on the investigated species in order to avoid false-negative results.