Isolation and Genetic Characterization of a Novel Adenovirus Associated with Mass Mortality in Great Cormorants (Phalacrocorax carbo).

High mortality in great cormorants (Phalacrocorax carbo) was registered on the Alakol Lake in eastern Kazakhstan in 2021 when about 20% of juveniles died. High-throughput sequencing revealed the presence of a putative novel cormorant adenovirus significantly divergent from known aviadenoviruses. We suggest that this cormorant adenovirus can be considered an emerging threat to the health and conservation of this species.

Aislamiento y caracterización genética de un nuevo adenovirus asociado con la mortalidad masiva en cormoranes grandes (Phalacrocorax carbo). En 2021 se registró una alta mortalidad de cormoranes grandes (Phalacrocorax carbo) en el lago Alakol, en el este de Kazajstán, cuando murieron alrededor del 20% de las aves jóvenes. La secuenciación de alto rendimiento reveló la presencia de un supuesto nuevo adenovirus de cormorán significativamente divergente de los aviadenovirus conocidos. Sugerimos que este adenovirus de cormorán puede considerarse una amenaza emergente para la salud y conservación de esta especie.

A citizen science-based survey of avian mortality focusing on haemosporidian infections in wild passerine birds.

BACKGROUND: Haemosporidioses are common in birds and their manifestations range from subclinical infections to severe disease, depending on the involved parasite and bird species. Clinical haemosporidioses are often observed in non-adapted zoo or aviary birds, whereas in wild birds, particularly passerines, haemosporidian infections frequently seem to be asymptomatic. However, a recent study from Austria showed pathogenic haemosporidian infections in common blackbirds due to high parasite burdens of Plasmodium matutinum LINN1, a common parasite in this bird species, suggesting that virulent infections also occur in natural hosts. Based on these findings, the present study aimed to explore whether and to what extent other native bird species are possibly affected by pathogenic haemosporidian lineages, contributing to avian morbidity. METHODS: Carcasses of passerine birds and woodpeckers were collected during a citizen science-based survey for avian mortality in Austria, from June to October 2020. Tissue samples were taken and examined for haemosporidian parasites of the genera Plasmodium, Haemoproteus and Leucocytozoon by nested PCR and sequencing the mitochondrial cytb barcode region, histology, and chromogenic in situ hybridization applying genus-specific probes. RESULTS: From over 160 dead bird reportings, 83 carcasses of 25 avian species were submitted for investigation. Overall haemosporidian infection rate was 31%, with finches and tits prevailing species counts and infections. Sequence analyses revealed 17 different haplotypes (4 Plasmodium, 4 Haemoproteus, 9 Leucocytozoon), including 4 novel Leucocytozoon lineages. Most infected birds presented low parasite burdens in the peripheral blood and tissues, ruling out a significant contribution of haemosporidian infections to morbidity or death of the examined birds. However, two great tits showed signs of avian malaria, suggesting pathogenic effects of the detected species Plasmodium relictum SGS1 and Plasmodium elongatum GRW06. Further, exo-erythrocytic tissue stages of several haemosporidian lineages are reported. CONCLUSIONS: While suggesting generally little contribution of haemosporidian infections to mortality of the investigated bird species, the findings indicate a possible role of certain haemosporidian lineages in overall clinical manifestation, either as main causes or as concurrent disease agents. Further, the study presents new data on exo-erythrocytic stages of previously reported lineages and shows how citizen science can be used in the field of haemosporidian research.

DIPLOTRIAENA OBTUSA (NEMATODA: DIPLOTRIAENIDAE) FROM BARN SWALLOWS (HIRUNDO RUSTICA) AND CLIFF SWALLOWS (PETROCHELIDON PYRRHONOTA) COLLECTED DURING MORTALITY EVENTS IN THE UPPER MIDWEST, USA.

Several mortality events involving barn swallows (Hirundo rustica) and cliff swallows (Petrochelidon pyrrhonota) were reported in the Upper Midwestern states in 2017 and 2018. Barn swallow mortality followed unseasonal cold snaps, with the primary cause of death being emaciation with concurrent air sac nematodiasis. Lesions in cliff swallows were consistent with blunt force trauma from suspected car impacts. Examination of air sac nematodes from both bird species revealed morphological characters consistent with Diplotriaena obtusa. Sequence analysis of the partial 18S rRNA gene indicated the samples clustered with other species in the genus Diplotriaena. These nematodes provide a link between morphological specimens and DNA sequence data for D. obtusa.

SARCOCYSTOSIS IN A CAPTIVE FLOCK OF THICK-BILLED PARROTS (RHYNCHOPSITTA PACHYRHYNCHA) FROM 2005 TO 2016: MORBIDITY, MORTALITY, DIAGNOSTICS, AND MANAGEMENT STRATEGIES.

Sarcocystosis was diagnosed in a captive flock of thick-billed parrots (Rhynchopsitta pachyrhyncha) at the Wildlife Conservation Society's Queens Zoo. Since the index case in 2005, 45% of mortalities in birds over 30 days of age were due to sarcocystosis. Sarcocystis falcatula was repeatedly identified as the causative agent. The disease predominantly affected younger adult parrots. Administration of antiparasitic medications prior to development of respiratory signs prolonged life in infected birds, but disease was fatal until utilization of a three-drug combination (pyrimethamine, trimethoprim-sulfamethoxazole, and ponazuril). This protocol may require in excess of 6 mo of therapy to achieve clinical resolution of active disease. Plasma creatine kinase activity was found to be the most useful test in diagnosing infection and monitoring response to therapy. Polymerase chain reaction (PCR) for apicomplexan organisms on antemortem whole blood, blood smears, or dried blood spots helped confirm suspected cases, but due to the poor sensitivity was sometimes misleading when assessing response to therapy or resolution of clinical disease. Preventive measures, focusing on exclusion and removal of Virginia opossums (Didelphis virginiana) from zoo grounds failed to curtail the occurrence of sarcocystosis in the flock. Other preventative steps, such as modification of feeding stations to exclude potential arthropod paratenic hosts and prophylaxis trials with diclazuril, appeared to successfully mitigate new infections. Given the diagnostic and therapeutic challenges, prevention of exposure to S. falcatula is essential to ex-situ conservation efforts for thick-billed parrots.

Emergence and molecular characterization of pigeon Paramyxovirus-1 in non-native Eurasian collared doves (Streptopelia decaocto) in California, USA.

Eurasian collared doves (Streptopelia decaocto) were introduced into Florida in the 1980s and have since established populations throughout the continental United States. Pigeon paramyxovirus-1 (PPMV-1), a species-adapted genotype VI Avian orthoavulavirus 1, has caused periodic outbreaks among collared doves in the U.S. since 2001 with outbreaks occasionally involving native doves. In California, PPMV-1 mortality events were first documented in Riverside County in 2014 with subsequent outbreaks in 23 additional counties from southern to northern California between 2015 and 2019. Affected collared doves exhibited torticollis and partial paralysis. Pale kidneys were frequently visible on gross necropsy (65.4%; 51/78) while lymphoplasmacytic interstitial nephritis often with acute tubular necrosis (96.0%; 24/25) and pancreatic necrosis (80.0%; 20/25) were common findings on histopathology. In total, PPMV-1 was confirmed by rRT-PCR and sequence analysis from oropharyngeal and/or cloacal swabs in 93.0% (40/43) of the collared doves tested from 16 California counties. In 2017, Avian orthoavulavirus 1 was confirmed in a native mourning dove (Zenaida macroura) found dead during a PPMV-1 outbreak in collared doves by rRT-PCR from formalin-fixed paraffin-embedded (FFPE) tissues, after the initial rRT-PCR from swabs failed to detect the virus. Molecular sequencing of the fusion protein of isolates collected from collared doves during outbreaks in 2014, 2016, and 2017 identified two distinct subgenotypes, VIa and VIn. Subgenotype VIn has been primarily isolated from collared doves in the southern U.S., while VIa has been isolated from mixed avian species in the northeastern U.S., indicating two independent introductions into California. While populations of collared doves are not expected to be substantially impacted by this disease, PPMV-1 may pose a threat to already declining populations of native columbids. This threat could be assessed by monitoring native and non-native columbids for PPMV-1. Based on our study, swab samples may not be sufficient to detect infection in native columbids and may require the use of non-traditional diagnostic approaches, such as FFPE tissues, to ensure virus detection.

Anticoagulant Rodenticides in Red-Tailed Hawks (Buteo jamaicensis) from New York City, New York, USA, 2012-18.

Anticoagulant rodenticides (AR), principally difethialone, brodifacoum, and bromadiolone, were detected in the livers of 89% of 72 Red-tailed Hawks (Buteo jamaicensis) from New York City, New York, US examined for cause of death over a 7-yr period (January 2012-December 2018). Fatal hemorrhage likely attributable to AR exposure was diagnosed in 41% (30/74) of cases, and 46% (18/39) of the cases analyzed with no gross evidence of AR-mediated hemorrhage had liver concentrations of AR that overlapped those with an AR-poisoning diagnosis. Although urban areas like New York City can support surprisingly dense populations of Red-tailed Hawks, the threat posed by extensive use of AR can be large.

COMPARISON BETWEEN STANDARD ZOO QUARANTINE PRACTICES AND RISK-BASED MANAGEMENT OF ANIMAL TRANSFERS: A RETROSPECTIVE ANALYSIS OF AVIAN ACQUISITION MORBIDITY AND MORTALITY (2013-2018).

Beginning 01 January 2016, for all animals transferred into the collection, Disney's Animal Kingdom used a risk-based process that relies on pathogen hazard identification and then assessment and mitigation of the risk for each identified pathogen hazard. Data on morbidity, mortality, and disease introduction was compared for birds transferred in pre- and post-risk-based processes used during the period 2013-2018. When assessing morbidity within 3 mo of acquisition, logistic regression revealed that birds entering "straight to collection" had lower morbidity rates than those entering with either "standard quarantine" or "risk-based standard quarantine" methods (P = 0.032). Mortality within 3 mo of acquisition was not significantly different between straight-to-collection and standard quarantine or risk-based standard quarantine methods (P = 0.40). In addition, no transmissible pathogens of concern were introduced with acquired birds using either method. Implementation of a risk-based approach to animal transfers between zoos and aquariums may be an alternative method that does not pose a greater risk to animal morbidity or mortality while still protecting the collection from disease introduction.

Mortality cost of sex-specific parasitism in wild bird populations.

Sex-specific mortality is frequent in animals although the causes of different male versus female mortalities remain poorly understood. Parasitism is ubiquitous in nature with widespread detrimental effects to hosts, making parasitism a likely cause of sex-specific mortalities. Using sex-specific blood and gastrointestinal parasite prevalence from 96 and 54 avian host species, respectively, we test the implications of parasites for annual mortality in wild bird populations using phylogenetic comparative methods. First, we show that parasite prevalence is not different between adult males and females, although Nematodes showed a statistically significant but small male-biased parasite prevalence. Second, we found no correlation between sex-biased host mortalities and sex-biased parasite prevalence. These results were consistent in both blood and gastrointestinal parasites. Taken together, our results show little evidence for sex-dependent parasite prevalence in adults in wild bird populations, and suggest that parasite prevalence is an unlikely predictor of sex difference in adult mortalities, not withstanding sampling limitations. We propose that to understand causes of sex-biased mortalities, more complex analyses are needed that incorporate various ecological and life history components of animals life that may include sex differences in exposure to predators, immune capacity and cost of reproduction.

A retrospective analysis of mortality in captive Magellanic penguins (Spheniscus magellanicus) in the United States, 2008-2018.

Mortality data for Magellanic penguins (Spheniscus magellanicus) housed in zoos and aquariums in the United States has not previously been published. Necropsy and histopathology records were examined for Magellanic penguins housed at 12 Association of Zoos and Aquariums institutions from 2008 through 2018. If birds lived through the first year, the mean longevity was found to be 18.9 years of age (standard deviation: 7.9). Prefledge chicks and geriatric penguins experienced the highest mortality rates. Aspergillosis was a major cause of death in this species. There was no significant difference in mortality between males and females. Based on these data, recommendations for the husbandry and veterinary care of captive Magellanic penguins can be made.

MYONECROSIS AND DEATH DUE TO PRESUMED MICROCYSTIN TOXICOSIS IN AMERICAN WHITE PELICANS (PELECANUS ERYTHRORHYNCOS).

Over a period of 5 mo, seven out of eight American white pelicans (Pelecanus erythrorhynchos) housed on a spring-fed pond at a zoo died or were euthanized. Clinical signs included inability to stand, anorexia, and weight loss. Clinicopathologic findings included heterophilic leukocytosis and elevated creatine kinase and aspartate aminotransferase. Histopathologic findings on all pelicans demonstrated severe, chronic, diffuse rhabdomyofiber degeneration and necrosis, making vitamin E deficiency a differential diagnosis despite routine supplementation. Based on tissue and pond water assays for the cyanobacterial toxin, microcystin, toxicosis is suspected as the inciting cause of death in these cases. We hypothesize that vitamin E exhaustion and resultant rhabdomyodegeneration and cardiomyopathy were sequelae to this toxicosis.

An assessment of the impacts of litter treatments on the litter quality and broiler performance: A systematic review and meta-analysis.

OBJECTIVE: The choice of the most suitable litter treatment should be based on scientific evidence. This systematic review assessed the effectiveness of litter treatments on ammonia concentration, pH, moisture and pathogenic microbiota of the litter and their effects on body weight, feed intake, feed conversion and mortality of broilers. METHODS: The systematic literature search was conducted using PubMed (Medline), Google Scholar, ScienceDirect and Scielo databases to retrieve articles published from January 1998 to august 2019. Means, standard deviations and sample sizes were extracted from each study. The response variables were analyzed using the mean difference (MD) or standardized mean difference (SMD), (litter treatment minus control group). All variables were analyzed using random effects meta-analyses. RESULTS: Subgroup meta-analysis revealed that acidifiers reduce pH (P<0.001), moisture (P = 0.002) ammonia (P = 0.011) and pathogenic microbiota (P <0.001) of the litter and improves the weight gain (P = 0.019) and decreases the mortality rate of broilers (P<0.001) when compared with controls. Gypsum had a positive effect on ammonia reduction (P = 0.012) and improved feed conversion (P = 0.023). Alkalizing agents raise the pH (P = 0.035), worsen feed conversion (P<0.001), increase the mortality rate (P <0.001), decrease the moisture content (P<0.001) and reduce the pathogenic microbiota of the litter (P<0.001) once compared to controls. Superphosphate and adsorbents reduce, respectively, pH (P<0.001) and moisture (P = 0.007) of the litter compared to control groups. CONCLUSION: None of the litter treatments influenced the feed intake of broilers. Meta-analyses of the selected studies showed positive and significant effects of the litter treatments on broiler performance and litter quality when compared with controls. Alkalizing was associated with worse feed conversion and high mortality of broilers.

Causes of Fatal Cyathostomiasis in Brown Booby (Sula leucogaster) from Brazil: Identification of Pathogen and Implications for Management.

Fatal infection by Cyathostoma (Cyathostoma) phenisci (Nematoda: Syngamidae), was identified in 2 of 52 brown boobies (Sula leucogaster) collected on beaches in the state of Rio de Janeiro, Brazil, and admitted to the veterinary clinic for rehabilitation. Both infected birds were in poor physical condition, with atrophied pectoral muscles, and died soon after starting treatment. The parasitological and pathological examination of the carcasses revealed the presence of C. (C.) phenisci in the trachea, resulting in tracheitis, as well as severe parasitic granulomatous bronchopneumonia caused by eggs deposited in the lungs. In our opinion, these serious pathological changes were the primary cause of chronic respiratory illness. This is the first description of fatal cyathostomiasis in a fish-eating avian host caused by infection by a member of the subgenus Cyathostoma (Cyathostoma). Therefore, it is reasonable to consider C. (C.) phenisci to be a real threat to a wide range of their definitive hosts, and cyathostomiasis should be considered in the differential diagnosis for fish-eating marine birds, even in cases without respiratory signs. This is also the first record of the genus Cyathostoma in Brazil.

RETROSPECTIVE REVIEW OF MORTALITY IN CAPTIVE PINK PIGEONS (NESOENAS MAYERI) HOUSED IN EUROPEAN COLLECTIONS: 1977-2018.

The Mauritian pink pigeon (Nesoenas mayeri) is vulnerable, with only 400 individuals remaining in the free-living population. A European captive population was established in 1977 and a European Endangered Species Program (EEP) in 1992. The EEP long-term management plan recommends integrating the EEP and free-living Mauritius populations through pigeon transfers. A retrospective mortality review of the captive population was performed as part of a disease risk assessment process and to inform infectious disease screening prior to exporting captive birds to Mauritius. Six hundred pink pigeons from 34 institutions died from 1977 to 2018. Each individual was categorized according to age at time of death. Records from 404 individuals were categorized according to cause of death. Neonatal mortality (39%) and juvenile mortality (10.8%) were most commonly caused by noninfectious diseases (52% and 54.4%, respectively), including parental neglect and failure to thrive in neonates and nutritional secondary hyperparathyroidism in juveniles. Trauma (43.1%) was the most common cause of mortality in adults, with significantly higher mortality in males from interspecific aggression and in females due to intraspecific aggression. Yersinia pseudotuberculosis, Mycobacterium avium, and Escherichia coli were the most common infectious causes of adult mortality, and E. coli was the most common infectious cause in neonates. The following infectious diseases were identified as priorities for pre-export disease risk analysis, though not all caused mortality: Y. pseudotuberculosis, M. avium, Trichomonas spp., Chlamydia psittaci, and Coccidia spp. Husbandry changes have been made over the years to mitigate many of the noninfectious causes of mortality. These include alterations to nest sites to reduce neonatal trauma and abandonment, ultraviolet light supplementation and diet optimization to reduce metabolic disorders, improving enclosure design to reduce impact trauma, allowing females rest periods during breeding season, and avoiding housing with certain species.

Experimental Usutu Virus Infection in Domestic Canaries Serinus canaria.

Usutu virus (USUV) is a neurotropic flavivirus closely related to West Nile virus (WNV). Its enzootic cycle mainly involves mosquitoes and birds. Human infection can occur with occasional, but sometimes severe, neurological complications. Since its emergence and spread in Europe over the last two decades, USUV has been linked to significant avian outbreaks, especially among Passeriformes, including European blackbirds (Turdus merula). Strikingly, no in vivo avian model exists so far to study this arbovirus. The domestic canary (Serinus canaria) is a passerine, which is considered as a highly susceptible model of infection by WNV. Here, we experimentally challenged domestic canaries with two different doses of USUV. All inoculated birds presented detectable amounts of viral RNA in the blood and RNA shedding via feathers and droppings during the early stages of the infection, as determined by RT-qPCR. Mortality occurred in both infected groups (1/5 and 2/5, respectively) and was not necessarily correlated to a pure neurological disease. Subsequent analyses of samples from dead birds showed histopathological changes and virus tropism mimicking those reported in naturally infected birds. A robust seroconversion followed the infection in almost all the surviving canaries. Altogether, these results demonstrate that domestic canaries constitute an interesting experimental model for the study of USUV pathogenesis and transmission.

Avian Gastric Yeast (Macrorhabdus ornithogaster) and Mycobacterium genavense Infections in a Zoo Budgerigar (Melopsittacus undulatus) Flock.

Over a 4-mo period, a Michigan zoo had 32 budgerigars, Melopsittacus undulatus, from their flock die. Whole animals or formalin-fixed tissues were submitted to Michigan State University Veterinary Diagnostic Laboratory for diagnosis. Avian gastric yeast infection, Macrorhabdus ornithogaster, was diagnosed in seven birds. There was atrophy of breast musculature and no subcutaneous or coelomic fat stores in six necropsied birds. Only two birds had proventricular dilatation grossly. Histologic examination of the proventriculus of all seven birds revealed abundant 3 × 50-µm septate, parallel-walled, nonbranching yeast organisms morphologically consistent with Macrorhabdus ornithogaster. Mycobacteriosis was diagnosed in three budgerigars, two of which were necropsied. Both necropsied birds had hepatomegaly and one also had splenomegaly. No acid-fast bacilli were found in the livers of either bird but were found in splenic macrophages of the bird with splenomegaly and in the intestine of the other bird. Mycobacterium species were cultured from the enlarged spleen and identified by DNA sequence as Mycobacterium genavense. Pulmonary granulomas with acid-fast bacilli were found in the bird submitted as fixed tissues. None of the budgerigars had a dual infection. The remainder of the budgerigars died from hepatitis, nephrosis, oviductal prolapse, exclusion from food and water by flock mates, or tumors.

RETROSPECTIVE STUDY OF CAUSES OF ANIMAL MORTALITY IN LJUBLJANA ZOO 2005-2015.

Zoos harbor large collections of diverse species, aiding in both conservation and education, as well as research in multiple scientific fields. However, the most common causes of death in zoo animals around the world remain unclear because few extensive reports or reviews are published on this topic. This information could greatly improve preventive veterinary medicine in zoologic gardens. This study provides a retrospective overview of the causes of death of animals from the Ljubljana Zoo in the years 2005-2015. During this period, a total of 353 animals were submitted for necropsy, of which 244 were mammals, 85 were birds, and 25 were reptiles. The causes of deaths were divided into infectious diseases (38%), dysfunctions of individual organs (20%), traumas (13%), parasitosis (7%), reproductive disorders (6%), metabolic disorders (3%), neoplastic disease (4%), and intoxications (4%). In some cases, the cause of death was unable to be determined (7%), most commonly because of autolysis of the body. The results of this retrospective study bring a general overview of the epizootiologic situation in the Ljubljana Zoo over an 11-yr period and valuable information to other zoos to optimize preventative plans and diagnostics.

Different dynamics of Usutu virus infections in Austria and Hungary, 2017-2018.

Usutu virus (USUV), a mosquito-borne flavivirus closely related to West Nile virus, emerged in Austria in 2001, when it caused a considerable mass-mortality of Eurasian blackbirds. Cases in birds increased until 2003 and quickly declined thereafter, presumably due to developing herd immunity. Since 2006, no further cases were recorded, until two blackbirds were tested positive in 2016. In Hungary, USUV first appeared in 2005 and has caused only sporadic infections since then. Initially, the only genetic USUV lineage found across both countries was Europe 1. This changed in 2015/2016, when Europe 2 emerged, which has since then become the prevalent lineage. Due to dispersal of these strains and introduction of new genetic lineages, USUV infections are now widespread across Europe. In 2009, the first cases of USUV-related encephalitis were described in humans, and the virus has been frequently detected in blood donations since 2016. To monitor USUV infections among the Austrian wild bird population in 2017/2018, 86 samples were investigated by RT-PCR. In 67 of them, USUV nucleic acid was detected (17 in 2017, 50 in 2018). The majority of succumbed birds were blackbirds, found in Vienna and Lower Austria. However, the virus also spread westwards to Upper Austria and southwards to Styria and Carinthia. In Hungary, 253 wild birds were examined, but only six of them were infected with USUV (five in 2017, one in 2018). Thus, in contrast to the considerable increase in USUV-associated bird mortality in Austria, the number of infections in Hungary declined after a peak in 2016. Except for one case of USUV lineage Africa 3 in Austria in 2017, Europe 2 remains the most prevalent genetic lineage in both countries. Since USUV transmission largely depends on temperature, which affects vector populations, climate change may cause more frequent USUV outbreaks in the future.