Surveillance for Selected Pathogens and Parasites of Northern Bobwhite (Colinus virginianus) from Western Oklahoma, USA, 2018-20.

The Northern Bobwhite (Colinus virginianus) has been undergoing a range-wide population decline. Potential causes for declines across its historic range have been investigated for decades and include habitat loss and fragmentation and a variety of parasitic and infectious diseases. Although there have been studies on bobwhite ecology in Oklahoma, USA, relatively little is known about parasites and pathogens in the region. We evaluated the health of free-ranging bobwhites from nine sites in western Oklahoma. From 2018 to 2020, 206 bobwhites were evaluated for gross and microscopic lesions and tested for selected pathogens. In general, bobwhites were in good nutritional condition with ample muscle mass and fat stores. No significant gross lesions were observed in any bobwhite and no significant histologic lesions were detected in a subset. There was no evidence of infection with or exposure to reticuloendotheliosis virus, West Nile virus, respiratory Mycoplasmataceae species, Pasteurella multocida, intestinal Eimeria spp., or oral Trichomonas spp. Several pathogens of potential concern were detected, including avian adenovirus (8.6%), Toxoplasma gondii (2.3%), and haemosporidians (a Haemoproteus sp. (1.5%), Leucocytozoon schoutedeni (1.5%), and Plasmodium homopolare haplotype 2 [lineage LAIRI01; 3.6%]). Physaloptera sp. (12%) and Sarcocystis sp. (1%) were detected in the breast muscle. Low intraspecific genetic diversity was noted for Physaloptera sp., and sequences were most similar to Physaloptera sequences from bobwhites and grasshoppers (Orthoptera) in Texas. Low intensities of chewing lice, chiggers, and ticks were observed. A subset of bobwhites had evidence of exposure to selected toxicants and heavy metals; a small number had low levels of iron, manganese, zinc, molybdenum, and copper, which were not considered diagnostically relevant. In general, bobwhites from western Oklahoma appeared to be in good health with a low diversity of pathogens detected, but future work is needed to understand potentially changing disease risks for this population.

Using Opportunistic Samples to Monitor West Nile Virus Infection Status in Greater Sage-grouse (Centrocercus urophasianus) in Wyoming, USA (2020-22).

The frequency of arthropod-borne viral disease in naïve hosts is subject to change based on complex interactions among vector, host, virus, and external factors (e.g., climate). Thus, continual monitoring for both disease susceptibility and host infection dynamics is needed, especially for viruses that have proven detrimental to the health of wildlife hosts of conservation concern. The Greater Sage-grouse (Centrocercus urophasianus) is a gamebird of ecological and economic importance in the western United States for which population declines have led to a Near Threatened conservation status. Although these declines have mainly been attributed to habitat loss, West Nile virus (WNV) also poses a threat, with regional transmission potentially fueled by anthropogenic landscape alterations that may facilitate mosquito breeding. With limited WNV monitoring in Greater Sage-grouse after recognition of high susceptibility to mortality early after its initial detection in the western United States, the potential long-term impacts of WNV on this species are poorly understood. We used the plaque reduction neutralization test of filter paper strip-eluted sera to assess for anti-WNV antibodies, indicating prior infection, in opportunistically collected samples. From 2020 to 2022, 85 Greater Sage-grouse in Wyoming were sampled; none had anti-WNV antibodies. This result corroborates findings of previous studies documenting low seroprevalence. With the tenuous conservation status of the species, all potential population health risks should be considered in future management strategies, especially in the face of changing climate and landscapes.

Storage time and temperature of filter paper strips affect anti-West Nile virus antibody detection in 2 galliform species.

The effects of filter paper strip (FPS) storage time and temperature on antibody detection are poorly understood despite widespread use in wildlife research. We collected sera and FPSs from 23 wild turkeys (Meleagris gallopavo) and 20 northern bobwhites (Colinus virginianus) experimentally infected with West Nile virus (WNV) to compare FPS storage methods with WNV plaque reduction neutralization test (PRNT) sensitivity. FPS storage methods included: immediate elution after drying, and storage at -20°C for 3 mo, -20°C for 6 mo, room temperature (RT) for 3 mo, and RT for 6 mo prior to elution. FPS eluates and sera were co-titrated to determine endpoint antibody titers, which were compared between FPS sera eluted immediately and sera, and among FPS eluates that underwent different storage conditions. Antibody titers were ~4-fold less in FPS sera eluted immediately versus sera, and dropped more frequently below PRNT detection threshold in northern bobwhites, which had ~10-fold lower serum antibody titers than wild turkeys. Antibody titers were lower in FPS samples stored at RT and for 6 mo. WNV serologic surveys may result in falsely low seroprevalence estimates if FPSs are stored at RT for ≥ 3 mo before elution.

Comparison of Pathology and Immunohistochemistry in Natural and Experimental West Nile Virus Infections in Ruffed Grouse (Bonasa umbellus).

Ruffed Grouse (Bonasa umbellus) populations have declined in much of the eastern US in recent decades. Research suggests that West Nile virus (WNV) contributed to these declines, based on decreasing population indices temporally correlated to WNV introduction into North America, high morbidity and mortality in experimentally infected Ruffed Grouse, and relatively low statewide seroprevalence concurrent with high WNV vector indices. We describe lesions and relevant diagnostic findings in six, free-ranging Ruffed Grouse that directly or indirectly died of natural WNV infection and compare results to experimentally infected Ruffed Grouse. All naturally infected grouse were found moribund or dead from August to December 2015-18 in the northeastern US; 4/6 grouse were subadults. Necropsy, histopathology, immunohistochemistry, and/or ancillary tests contributed to the diagnosis of WNV-associated disease in all cases. The most common lesions in naturally infected grouse were myocardial inflammation, degeneration, and/or necrosis, consistent with lesions in experimentally infected grouse. Evidence of trauma in 6/6 naturally infected grouse suggests the possibility of WNV-associated morbidity as a predisposing factor. Diagnostic findings in 3/6 naturally infected grouse were consistent with experimentally infected grouse with severe disease 7-8 d postinoculation; the remaining naturally infected birds had similar findings to experimentally infected subclinical grouse. Our results further support the notion that WNV contributes to mortality of free-ranging Ruffed Grouse and may be used to improve surveillance strategies and population-level management approaches.

Susceptibility of wild turkeys (Meleagris gallopavo) to experimental West Nile virus infection.

Since the introduction of West Nile virus (WNV) to North America in 1999, WNV is estimated to have contributed to population-level declines in numerous avian species. However, the potential impacts of this virus on many free-ranging upland game bird species, including the wild turkey (Meleagris gallopavo), which is undergoing regional population declines, remain unknown. Herein, two age groups (∼5 to 6 weeks and ∼15 to 16 weeks post-hatch) of juvenile wild turkeys were subcutaneously inoculated with WNV, sampled daily from 1 to 7 days post-inoculation (dpi), and euthanized on 14 dpi. No clinical signs and minimal gross lesions were attributable to WNV infection. Peak viraemia titres were similar between age groups (<101.7 to 104.6 plaque-forming units [PFU]/ml), but the duration of viraemia was longer in the old group (3-4 days) than in the young group (0-3 days). Intermittent oral and/or cloacal viral shedding from 2 to 7 dpi was detected in both age groups. No infectious virus was detected in the heart, brain, kidney, skeletal muscle, spleen, and feathers from WNV-inoculated turkeys euthanized on 14 dpi. All WNV-inoculated birds seroconverted by 14 dpi, as well as two co-housed sham-inoculated birds. The most consistent microscopic lesions among all WNV-inoculated birds were mild lymphoplasmacytic myocarditis and encephalitis. Minimal immunohistochemical labelling was detected in tissues in addition to scant macrophages within the blood, spleen, and bone marrow. These data suggest WNV is unlikely to pose a significant risk to wild turkey populations, although the possibility remains that WNV may indirectly decrease fitness or predispose wild turkeys to other health stressors.RESEARCH HIGHLIGHTS Clinical disease was not observed in wild turkeys experimentally infected with WNV.Pathology attributed to WNV was mild and included brain and heart inflammation.Viraemias suggest WNV-infected wild turkeys do not play a role in WNV transmission.No age-associated differences in WNV clinical disease or pathology were observed.

Notoedric mange (Notoedres centrifera) in two species of free-ranging rabbits from Florida, USA.

Mange is a contagious skin disease caused by different mite species affecting numerous domestic and wild animals, worldwide. This report details notoedric mange in an eastern cottontail (Sylvilagus floridanus) and in a marsh rabbit (Sylvilagus palustris) from Florida, USA. Clinical examination revealed similar gross lesions including poor nutritional condition, multifocal alopecia and hyperkeratosis. Skin scrapings from both rabbits revealed numerous subcutaneous mites identified as Notoedres centrifera, a species previously only associated with rodents, primarily squirrels. Mites from both rabbits were identified based on morphology and confirmed by sequencing the internal transcribed spacer-2 (ITS-2) region. These cases emphasize the need for continued surveillance and accurate diagnostic evaluation to determine the cause and characterization of the skin disease, while distinguishing it from other potential pathogens that may manifest similarly in rabbits, such as Notoedres cati, Sarcoptes scabiei or Psoroptes cuniculi.

Listeriosis with viral coinfections in 8 gray foxes, 8 wild turkeys, and 2 young cervids in the southeastern United States.

Listeria monocytogenes is a bacterium that can cause disease in many species, including humans, livestock, and wildlife. Increased interactions via shared habitats may promote pathogen transmission among these groups. Our objectives were to evaluate the Southeastern Cooperative Wildlife Disease Study diagnostic data to characterize and compare L. monocytogenes-induced lesions and comorbidities in gray foxes and wild turkeys, and to describe cases of listeriosis in 2 cervids. From 1991-2020, 8 gray foxes, 8 wild turkeys, a neonatal elk, and a white-tailed deer fawn from several eastern states in the United States were diagnosed with listeriosis. All 8 foxes had hepatitis and/or hepatic necrosis with intralesional gram-positive bacilli, and concurrent canine distemper virus (CDV) infection; 2 of the foxes had been vaccinated recently for CDV. L. monocytogenes was cultured from the liver (6 of 8) or lung (2 of 8) of foxes. Lesions in wild turkeys included hepatocellular necrosis (3 of 8), heterophilic hepatitis (1 of 8), heterophilic granulomas (1 of 8), intrasinusoidal gram-positive bacilli without hepatic lesions (1 of 8), granulomatous dermatitis (1 of 8), and/or granulomatous myocarditis (2 of 8). Lymphoproliferative disease viral DNA was detected in 5 of 6 turkeys tested; reticuloendotheliosis viral DNA was detected in 2 of 3 turkeys tested. Both cervids had systemic listeriosis, with L. monocytogenes isolated from liver. Immunohistochemistry for Listeria spp. on select cases revealed immunolabeling in affected organs. Listeriosis was thus established as a cause of morbidity and mortality in 3 wildlife species, which often suffered from concurrent infections and likely immunosuppression.

West Nile Virus Infections in an Urban Colony of American White Ibises (Eudocimus albus) in South Florida, USA.

West Nile virus (WNV) is pathogenic in a wide range of avian hosts and is endemic in much of North America. This virus is responsible for population declines of some Passeriformes. We describe a WNV-associated mortality event in American White Ibis (Eudocimus albus) nestlings. This is a species, inherent to the Everglades ecosystem, which has recently begun nesting in urban areas. An urban colony in south Florida was monitored from March-July in 2020 as part of an ongoing study. Nestling carcasses were collected opportunistically and sent to the Southeastern Cooperative Wildlife Disease Study, University of Georgia within 24 h for diagnosis. Three ibis nestling deaths were confirmed to be caused by WNV infection based on histopathology, immunohistochemistry, and reverse transcription PCR. Serial plasma samples collected weekly from 36 healthy chicks of the same urban rookery were tested for WNV-neutralizing antibodies via plaque reduction neutralization test; four chicks were seropositive. Antibody titers in three seropositive chicks from which serial samples were collected waned over time, suggesting maternal antibody transfer. Ibis mortalities were consistent with a spike of WNV activity in this region of Florida. West Nile virus infection may be an important seasonal cause of mortality for wading bird nestlings.

Experimental West Nile Virus Infection in Northern Bobwhite Quail (Colinus virginianus).

West Nile virus (WNV) has been implicated in regional declines of numerous North American bird species, although its potential impact upon many species, including some game birds, remains unknown. Specifically, information about susceptibility to infection and infection outcome are crucial to assessing health risks. Northern bobwhite quail (Colinus virginianus) are a popular and common game bird across much of the United States, as well as in captive breeding programs and as backyard birds. Two age groups of bobwhites were subcutaneously inoculated with WNV and euthanatized on 15 days postinoculation (DPI). Three of 10 inoculated 5-wk-old and 4/10 inoculated 15-wk-old birds developed detectable viremia titers during 1-5 DPI, with low peak titers (101.7-103.0 plaque-forming units [PFU]/ml). Three of 10 inoculated 5-wk-old and 1/10 inoculated 15-wk-old birds shed low viral titers (peak 100.7-101.8 PFU/swab) either orally or cloacally or both for limited periods from 2 to 6 DPI. All inoculated birds (n = 20) remained apparently healthy and seroconverted by 15 DPI. No infectious virus was detected in select tissues: heart, kidney, brain, skeletal muscle, spleen (15-wk-old group only), and feathers from any of the bobwhites. No sham-inoculated, contact control birds (n = 8) became viremic or had virus isolated from tissues or swabs. The most consistent microscopic lesion was minimal to mild, lymphoplasmacytic myocarditis (6/10 in 5-wk-olds; 5/10 in 15-wk-olds). Immunohistochemical labeling was most often in macrophages in spleen and bone marrow, likely reflective of clearance of infection. There were no statistically significant differences in the peak viremia and shedding titers between age groups and no differences in the development of WNV-associated lesions between the two age groups. These results suggest that WNV is unlikely to pose a health risk to bobwhites and that bobwhites likely are an incompetent reservoir host species in WNV transmission.

Infección experimental por el virus del Nilo occidental en codornices de Virginia (Colinus virginianus). El virus del Nilo Occidental (con las siglas en inglés) se ha visto implicado en la disminución regional de numerosas especies de aves en América del Norte, aunque se desconoce su impacto potencial sobre muchas especies, incluidas algunas aves de caza. Específicamente, la información sobre la susceptibilidad a la infección y el resultado de la infección es crucial para evaluar los riesgos para la salud. La codorniz de Virginia (Colinus virginianus) es un ave de caza popular y común en gran parte de los Estados Unidos, así como en programas de cría en cautiverio y como aves de traspatio. Dos grupos de codornices de diferentes edades fueron inoculadas por vía subcutánea con el virus del Nilo Occidental y fueron sacrificadas 15 días después de la inoculación (DPI). Tres de diez aves inoculadas de cinco semanas y cuatro de diez aves inoculadas de 15 semanas desarrollaron títulos de viremia detectables durante uno a cinco días después de la inoculación, con bajos títulos durante la cima de la curva (101.7­103.0 unidades formadoras de placa [PFU]/ml). Tres de diez aves inoculadas de cinco semanas de edad y una de diez aves inoculadas a las 15 semanas de edad mostraron títulos virales bajos (pico 100.7­101.8 PFU/hisopo) ya sea por vía oral, cloacal o ambas durante períodos limitados de dos a seis días después de la inoculación. Todas las aves inoculadas (n = 20) permanecieron aparentemente sanas y mostraron seroconversión a los 15 días después de la inoculación. No se detectó ningún virus infeccioso en tejidos seleccionados: corazón, riñón, cerebro, músculo esquelético, bazo (solo en el grupo de 15 semanas de edad) y plumas de cualquiera de las codornices. Ninguna ave no inoculada o de control por contacto (n = 8) se volvió virémica o mostró aislamiento viral de los tejidos o hisopos. La lesión microscópica más consistente fue miocarditis linfoplasmocítica de mínima a leve (6/10 en aves de cinco semanas; 5/10 en aves de 15 semanas). La tinción inmunohistoquímica se observó con mayor frecuencia en macrófagos del bazo y la médula ósea, lo que probablemente refleja la eliminación de la infección. No hubo diferencias estadísticamente significativas en el pico de viremia y los títulos de eliminación entre los grupos de edad y no hubo diferencias en el desarrollo de lesiones asociadas con el virus del Nilo Occidental entre los dos grupos de edad. Estos resultados sugieren que es poco probable que el virus del Nilo Occidental represente un riesgo para la salud de las codornices de Virginia y que estas aves probablemente sean una especie hospedadora reservorio incompetente en la transmisión del virus del Nilo Occidental.

Serologic Evidence of Arthropod-Borne Virus Infections in Wild and Captive Ruminants in Ontario, Canada.

Arthropod-borne viruses (arboviruses) are globally widespread, and their transmission cycles typically involve numerous vertebrate species. Serologic testing of animal hosts can provide a routine surveillance approach to monitoring animal disease systems, can provide a surveillance alternative to arthropod testing and human case reports, and may augment knowledge of epizootiology. Wild and captive ruminants represent good candidate sentinels to track geographic distribution and prevalence of select arboviruses. They often are geographically widespread and abundant, inhabit areas shared by humans and domestic animals, and are readily fed on by various hematophagous arthropod vectors. Ontario, Canada, is home to high densities of coexisting humans, livestock, and wild cervids, as well as growing numbers of arthropod vectors because of the effects of climate change. We collected blood samples from 349 livestock (cattle/sheep) and 217 cervids (wild/farmed/zoo) in Ontario (2016-2019) to assess for antibodies to zoonotic and agriculturally important arboviruses. Livestock sera were tested for antibodies to bluetongue virus (BTV) and epizootic hemorrhagic disease virus (EHDV). Sera from cervids were tested for antibodies to BTV, EHDV, West Nile virus (WNV), eastern equine encephalitis virus (EEEV), Powassan virus (POWV), and heartland virus (HRTV). Fifteen (9.0%) cattle were seropositive for EHDV-serotype 2. Nine (4.2%) cervids were seropositive for arboviruses; three confirmed as WNV, three as EEEV, and one as POWV. All animals were seronegative for BTV and HRTV. These results reveal low seroprevalence of important agricultural, wildlife, and zoonotic pathogens and underline the need for continued surveillance in this and other regions in the face of changing environmental conditions.

Prevalence of avian haemosporidian parasites is positively related to the abundance of host species at multiple sites within a region.

Parasite prevalence is thought to be positively related to host population density owing to enhanced contagion. However, the relationship between prevalence and local abundance of multiple host species is underexplored. We surveyed birds and their haemosporidian parasites (genera Plasmodium and Haemoproteus) at multiple sites across eastern North America to test whether the prevalence of these parasites in a host species at a particular site is related to that host's local abundance. Prevalence was positively related to host abundance within most sites, although the effect was stronger and more consistent for Plasmodium than for Haemoproteus. In contrast, prevalence was not related to variation in the abundance of most individual host species among sites across the region. These results suggest that parasite prevalence partly reflects the relative abundances of host species in local assemblages. However, three nonnative host species had low prevalence despite being relatively abundant at one site, as predicted by the enemy release hypothesis.

Local host specialization, host-switching, and dispersal shape the regional distributions of avian haemosporidian parasites.

The drivers of regional parasite distributions are poorly understood, especially in comparison with those of free-living species. For vector-transmitted parasites, in particular, distributions might be influenced by host-switching and by parasite dispersal with primary hosts and vectors. We surveyed haemosporidian blood parasites (Plasmodium and Haemoproteus) of small land birds in eastern North America to characterize a regional parasite community. Distributions of parasite populations generally reflected distributions of their hosts across the region. However, when the interdependence between hosts and parasites was controlled statistically, local host assemblages were related to regional climatic gradients, but parasite assemblages were not. Moreover, because parasite assemblage similarity does not decrease with distance when controlling for host assemblages and climate, parasites evidently disperse readily within the distributions of their hosts. The degree of specialization on hosts varied in some parasite lineages over short periods and small geographic distances independently of the diversity of available hosts and potentially competing parasite lineages. Nonrandom spatial turnover was apparent in parasite lineages infecting one host species that was well-sampled within a single year across its range, plausibly reflecting localized adaptations of hosts and parasites. Overall, populations of avian hosts generally determine the geographic distributions of haemosporidian parasites. However, parasites are not dispersal-limited within their host distributions, and they may switch hosts readily.

Host immune responses to experimental infection of Plasmodium relictum (lineage SGS1) in domestic canaries (Serinus canaria).

Understanding the complexity of host immune responses to parasite infection requires controlled experiments that can inform observational field studies. Birds and their malaria parasites provide a useful model for understanding host-parasite relationships, but this model lacks a well-described experimental context for how hosts respond immunologically to infection. Here, ten canaries (Serinus canaria) were infected with the avian malaria parasite Plasmodium relictum (lineage SGS1) in a controlled laboratory setting with ten uninfected (control) birds. A suite of immunological blood parameters, including the concentration of four white blood cell types, the concentration of the acute phase protein haptoglobin, and the bacteria-killing ability of blood plasma, were repeatedly measured over a 25-day period covering the acute phase of a primary infection by P. relictum. Three infected and one control bird died during the course of the experiment. A multivariate statistical analysis of the immune indices revealed significant differences between infected and uninfected individuals between 5 and 14 days postinfection (dpi). Group differences corresponded to reduced concentrations of lymphocytes (5 dpi), heterophils (8 dpi), and monocytes (11 and 14 dpi), and an increase in haptoglobin (14 dpi), in infected birds relative to uninfected controls, and no change in bacteria-killing. Upon re-running the analysis with only the surviving birds, immunological differences between infected and control birds shifted to between 11 and 18 dpi. However, there were no clear correlates relating immune parameters to the likelihood of surviving the infection. The results presented here demonstrate the dynamic and complex nature of avian immune function during the acute phase of malaria infection and provide a context for studies investigating immune function in wild birds.

The ecology of host immune responses to chronic avian haemosporidian infection.

Host responses to parasitism in the wild are often studied in the context of single host-parasite systems, which provide little insight into the ecological dynamics of host-parasite interactions within a community. Here we characterized immune system responses to mostly low-intensity, chronic infection by haemosporidian parasites in a sample of 424 individuals of 22 avian host species from the same local assemblage in the Missouri Ozarks. Two types of white blood cells (heterophils and lymphocytes) were elevated in infected individuals across species, as was the acute-phase protein haptoglobin, which is associated with inflammatory immune responses. Linear discriminant analysis indicated that individuals infected by haemosporidians occupied a subset of the overall white blood cell multivariate space that was also occupied by uninfected individuals, suggesting that these latter individuals might have harbored other pathogens or that parasites more readily infect individuals with a specific white blood cell profile. DNA sequence-defined lineages of haemosporidian parasites were sparsely distributed across the assemblage of hosts. In one well-sampled host species, the red-eyed vireo (Vireo olivaceus), heterophils were significantly elevated in individuals infected with one but not another of two common parasite lineages. Another well-sampled host, the yellow-breasted chat (Icteria virens), exhibited no differences in immune response to different haemosporidian lineages. Our results indicate that while immune responses to infection may be generalized across host species, parasite-specific immune responses may also occur.