Tissue Tropisms of Avian Influenza A Viruses Affect Their Spillovers from Wild Birds to Pigs.

Wild aquatic birds maintain a large, genetically diverse pool of influenza A viruses (IAVs), which can be transmitted to lower mammals and, ultimately, humans. Through phenotypic analyses of viral replication efficiency, only a small set of avian IAVs were found to replicate well in epithelial cells of the swine upper respiratory tract, and these viruses were shown to infect and cause virus shedding in pigs. Such a phenotypic trait of the viral replication efficiency appears to emerge randomly and is distributed among IAVs across multiple avian species and geographic and temporal orders. It is not determined by receptor binding preference but is determined by other markers across genomic segments, such as those in the ribonucleoprotein complex. This study demonstrates that phenotypic variants of viral replication efficiency exist among avian IAVs but that only a few of these may result in viral shedding in pigs upon infection, providing opportunities for these viruses to become adapted to pigs, thus posing a higher potential risk for creating novel variants or detrimental reassortants within pig populations.IMPORTANCE Swine serve as a mixing vessel for generating pandemic strains of human influenza virus. All hemagglutinin subtypes of IAVs can infect swine; however, only sporadic cases of infection with avian IAVs are reported in domestic swine. The molecular mechanisms affecting the ability of avian IAVs to infect swine are still not fully understood. From the findings of phenotypic analyses, this study suggests that the tissue tropisms (i.e., in swine upper respiratory tracts) of avian IAVs affect their spillovers from wild birds to pigs. It was found that this phenotype is determined not by receptor binding preference but is determined by other markers across genomic segments, such as those in the ribonucleoprotein complex. In addition, our results show that such a phenotypic trait was sporadically and randomly distributed among IAVs across multiple avian species and geographic and temporal orders. This study suggests an efficient way for assessment of the risk posed by avian IAVs, such as in evaluating their potentials to be transmitted from birds to pigs.

Tissue tropisms opt for transmissible reassortants during avian and swine influenza A virus co-infection in swine.

Genetic reassortment between influenza A viruses (IAVs) facilitate emergence of pandemic strains, and swine are proposed as a "mixing vessel" for generating reassortants of avian and mammalian IAVs that could be of risk to mammals, including humans. However, how a transmissible reassortant emerges in swine are not well understood. Genomic analyses of 571 isolates recovered from nasal wash samples and respiratory tract tissues of a group of co-housed pigs (influenza-seronegative, avian H1N1 IAV-infected, and swine H3N2 IAV-infected pigs) identified 30 distinct genotypes of reassortants. Viruses recovered from lower respiratory tract tissues had the largest genomic diversity, and those recovered from turbinates and nasal wash fluids had the least. Reassortants from lower respiratory tracts had the largest variations in growth kinetics in respiratory tract epithelial cells, and the cold temperature in swine nasal cells seemed to select the type of reassortant viruses shed by the pigs. One reassortant in nasal wash samples was consistently identified in upper, middle, and lower respiratory tract tissues, and it was confirmed to be transmitted efficiently between pigs. Study findings suggest that, during mixed infections of avian and swine IAVs, genetic reassortments are likely to occur in the lower respiratory track, and tissue tropism is an important factor selecting for a transmissible reassortant.

Influenza D Virus Infection in Feral Swine Populations, United States.

Influenza D virus (IDV) has been identified in domestic cattle, swine, camelid, and small ruminant populations across North America, Europe, Asia, South America, and Africa. Our study investigated seroprevalence and transmissibility of IDV in feral swine. During 2012-2013, we evaluated feral swine populations in 4 US states; of 256 swine tested, 57 (19.1%) were IDV seropositive. Among 96 archived influenza A virus-seropositive feral swine samples collected from 16 US states during 2010-2013, 41 (42.7%) were IDV seropositive. Infection studies demonstrated that IDV-inoculated feral swine shed virus 3-5 days postinoculation and seroconverted at 21 days postinoculation; 50% of in-contact naive feral swine shed virus, seroconverted, or both. Immunohistochemical staining showed viral antigen within epithelial cells of the respiratory tract, including trachea, soft palate, and lungs. Our findings suggest that feral swine might serve an important role in the ecology of IDV.

Feral Swine in the United States Have Been Exposed to both Avian and Swine Influenza A Viruses.

Influenza A viruses (IAVs) in swine can cause sporadic infections and pandemic outbreaks among humans, but how avian IAV emerges in swine is still unclear. Unlike domestic swine, feral swine are free ranging and have many opportunities for IAV exposure through contacts with various habitats and animals, including migratory waterfowl, a natural reservoir for IAVs. During the period from 2010 to 2013, 8,239 serum samples were collected from feral swine across 35 U.S. states and tested against 45 contemporary antigenic variants of avian, swine, and human IAVs; of these, 406 (4.9%) samples were IAV antibody positive. Among 294 serum samples selected for antigenic characterization, 271 cross-reacted with ≥1 tested virus, whereas the other 23 did not cross-react with any tested virus. Of the 271 IAV-positive samples, 236 cross-reacted with swine IAVs, 1 with avian IAVs, and 16 with avian and swine IAVs, indicating that feral swine had been exposed to both swine and avian IAVs but predominantly to swine IAVs. Our findings suggest that feral swine could potentially be infected with both avian and swine IAVs, generating novel IAVs by hosting and reassorting IAVs from wild birds and domestic swine and facilitating adaptation of avian IAVs to other hosts, including humans, before their spillover. Continued surveillance to monitor the distribution and antigenic diversities of IAVs in feral swine is necessary to increase our understanding of the natural history of IAVs.IMPORTANCE There are more than 5 million feral swine distributed across at least 35 states in the United States. In contrast to domestic swine, feral swine are free ranging and have unique opportunities for contact with wildlife, livestock, and their habitats. Our serological results indicate that feral swine in the United States have been exposed to influenza A viruses (IAVs) consistent with those found in both domestic swine and wild birds, with the predominant infections consisting of swine-adapted IAVs. Our findings suggest that feral swine have been infected with IAVs at low levels and could serve as hosts for the generation of novel IAVs at the interface of feral swine, wild birds, domestic swine, and humans.

Weathered MC252 crude oil-induced anemia and abnormal erythroid morphology in double-crested cormorants (Phalacrocorax auritus) with light microscopic and ultrastructural description of Heinz bodies.

Injury assessment of birds following the Deepwater Horizon (DWH) oil spill in 2010 was part of the Natural Resource Damage Assessment. One reported effect was hemolytic anemia with the presence of Heinz bodies (HB) in birds, however, the role of route and magnitude of exposure to oil is unknown. The purpose of the present study was to determine if double-crested cormorants (Phalacocorax auritis; DCCO) exposed orally and dermally to artificially weathered crude oil would develop hemolytic anemia including HB and reticulocytosis. In the oral experiment, sub-adult, mixed-sex DCCOs were fed control (n = 8) or oil-injected fish with a daily target dose of 5 (n = 9) or 10 (n = 9) ml oil/kg for 21 days. Then, subadult control (n = 12) and treated (n = 13) cormorant groups of similar sex-ratio were dermally treated with approximately 13ml of water or weathered MC252 crude oil, respectively, every 3 days for 6 dosages approximating 20% surface coverage. Collected whole blood samples were analyzed by light (new methylene blue) and transmission electron microscopy. Both oral and dermal treatment with weathered DWH MC252 crude oil induced regenerative, but inadequately compensated, anemia due to hemolysis and hematochezia as indicated by decreased packed cell volume, relative increase in reticulocytes with lack of difference in corrected reticulocyte count, and morphologic evidence of oxidant damage at the ultrastructural level. Hemoglobin precipitation, HB formation, degenerate organelles, and systemic oxidant damage were documented. Heinz bodies were typically <2µm in length and smaller than in mammals. These oblong cytoplasmic inclusions were difficult to see upon routine blood smear evaluation and lacked the classic button appearance found in mammalian red blood cells. They could be found as light, homogeneous blue inclusions upon new methylene blue staining. Ultrastructurally, HB appeared as homogeneous, electron-dense structures within the cytosol and lacked membranous structure. Oxidant damage in avian red blood cells results in degenerate organelles and precipitated hemoglobin or HB with different morphology than that found in mammalian red blood cells. Ultrastructural evaluation is needed to definitively identify HB and damaged organelles to confirm oxidant damage. The best field technique based on the data in this study is assessment of PCV with storage of blood in glutaraldehyde for possible TEM analysis.

Organ weights and histopathology of double-crested cormorants (Phalacrocorax auritus) dosed orally or dermally with artificially weathered Mississippi Canyon 252 crude oil.

A series of toxicity tests were conducted to assess the effects of low to moderate exposure to artificially weathered Deepwater Horizon Mississippi Canyon 252 crude oil on representative avian species as part of the Natural Resource Damage Assessment. The present report summarizes effects of oral exposure (n=26) of double-crested cormorants (DCCO; Phalacrocorax auritus) to 5 or 10ml oil kg-1 day-1 for up to 21 days or dermal application (n=25) of 13ml oil to breast and back feathers every three days totaling 6 applications in 21 days on organ weights and histopathology. Absolute and relative kidney and liver weights were increased in birds exposed to oil. Additionally, gross and/or histopathologic lesions occurred in the kidney, heart, pancreas and thyroid. Clinically significant renal lesions in the orally dosed birds included squamous metaplasia and increased epithelial hypertrophy of the collecting ducts and renal tubules and mineralization in comparison to controls. Gross cardiac lesions including thin walls and flaccid musculature were documented in both orally and dermally dosed birds and myocardial fibrosis was found in low numbers of dermally dosed birds only. Cytoplasmic vacuolation of the exocrine pancreas was noted in orally dosed birds only. Thyroid follicular hyperplasia was increased in dermally dosed birds only possibly due to increased metabolism required to compensate damaged feather integrity and thermoregulate. Gastrointestinal ulceration was found in orally dosed birds only. There were no significant hepatic histopathologic lesions induced by either exposure route. Therefore, hepatic histopathology is likely not a good representation of oil-induced damage. Taken together, the results suggest that oral or dermal exposure of DCCOs to artificially weathered MC252 crude oil induced organ damage that could potentially affect survivability.

Changes in white cell estimates and plasma chemistry measurements following oral or external dosing of double-crested cormorants, Phalacocorax auritus, with artificially weathered MC252 oil.

Scoping studies were designed whereby double-crested cormorants (Phalacocorax auritus) were dosed with artificially weathered Deepwater Horizon (DWH) oil either daily through oil injected feeder fish, or by application of oil directly to feathers every three days. Preening results in oil ingestion, and may be an effective means of orally dosing birds with toxicant to improve our understanding of the full range of physiological effects of oral oil ingestion on birds. Blood samples collected every 5-6 days were analyzed for a number of clinical endpoints including white blood cell (WBC) estimates and differential cell counts. Plasma biochemical evaluations were performed for changes associated with oil toxicity. Oral dosing and application of oil to feathers resulted in clinical signs and statistically significant changes in a number of biochemical endpoints consistent with petroleum exposure. In orally dosed birds there were statistically significant decreases in aspartate amino transferase (AST) and gamma glutamyl transferase (GGT) activities, calcium, chloride, cholesterol, glucose, and total protein concentrations, and increases in plasma urea, uric acid, and phosphorus concentrations. Plasma electrophoresis endpoints (pre-albumin, albumin, alpha-2 globulin, beta globulin, and gamma globulin concentrations and albumin: globulin ratios) were decreased in orally dosed birds. Birds with external oil had increases in urea, creatinine, uric acid, creatine kinase (CK), glutamate dehydrogenase (GLDH), phosphorus, calcium, chloride, potassium, albumin, alpha-1 globulin and alpha-2 globulin. Decreases were observed in AST, beta globulin and glucose. WBC also differed between treatments; however, this was in part driven by monocytosis present in the externally oiled birds prior to oil treatment.

Reprint of: CYP1A protein expression and catalytic activity in double-crested cormorants experimentally exposed to Deepwater Horizon Mississippi Canyon 252 oil.

Double-crested cormorants (Phalacrocorax auritus, DCCO) were orally exposed to Deepwater Horizon Mississippi Canyon 252 (DWH) oil to investigate oil-induced toxicological impacts. Livers were collected for multiple analyses including cytochrome P4501A (CYP1A) enzymatic activity and protein expression. CYP1A enzymatic activity was measured by alkoxyresorufin O-dealkylase (AROD) assays. Activities specific to the O-dealkylation of four resorufin ethers are reported: benzyloxyresorufin O-debenzylase (BROD), ethoxyresorufin O-deethylase (EROD), methoxyresorufin O-demethylase (MROD), and pentoxyresorufin O-depentylase (PROD). CYP1A protein expression was measured by western blot analysis with a CYP1A1 mouse monoclonal antibody. In study 1, hepatic BROD, EROD, and PROD activities were significantly induced in DCCO orally exposed to 20ml/kg body weight (bw) oil as a single dose or daily for 5 days. Western blot analysis revealed hepatic CYP1A protein induction in both treatment groups. In study 2 (5ml/kg bw oil or 10ml/kg bw oil, 21day exposure), all four hepatic ARODs were significantly induced. Western blots showed an increase in hepatic CYP1A expression in both treatment groups with a significant induction in birds exposed to 10ml/kg oil. Significant correlations were detected among all 4 AROD activities in both studies and between CYP1A protein expression and both MROD and PROD activities in study 2. EROD activity was highest for both treatment groups in both studies while BROD activity had the greatest fold-induction. While PROD activity values were consistently low, the fold-induction was high, usually 2nd highest to BROD activity. The observed induced AROD profiles detected in the present studies suggest both CYP1A4/1A5 DCCO isoforms are being induced after MC252 oil ingestion. A review of the literature on avian CYP1A AROD activity levels and protein expression after exposure to CYP1A inducers highlights the need for species-specific studies to accurately evaluate avian exposure to oil.

CYP1A protein expression and catalytic activity in double-crested cormorants experimentally exposed to deepwater Horizon Mississippi Canyon 252 oil.

Double-crested cormorants (Phalacrocorax auritus, DCCO) were orally exposed to Deepwater Horizon Mississippi Canyon 252 (DWH) oil to investigate oil-induced toxicological impacts. Livers were collected for multiple analyses including cytochrome P4501A (CYP1A) enzymatic activity and protein expression. CYP1A enzymatic activity was measured by alkoxyresorufin O-dealkylase (AROD) assays. Activities specific to the O-dealkylation of four resorufin ethers are reported: benzyloxyresorufin O-debenzylase (BROD), ethoxyresorufin O-deethylase (EROD), methoxyresorufin O-demethylase (MROD), and pentoxyresorufin O-depentylase (PROD). CYP1A protein expression was measured by western blot analysis with a CYP1A1 mouse monoclonal antibody. In study 1, hepatic BROD, EROD, and PROD activities were significantly induced in DCCO orally exposed to 20ml/kg body weight (bw) oil as a single dose or daily for 5 days. Western blot analysis revealed hepatic CYP1A protein induction in both treatment groups. In study 2 (5ml/kg bw oil or 10ml/kg bw oil, 21day exposure), all four hepatic ARODs were significantly induced. Western blots showed an increase in hepatic CYP1A expression in both treatment groups with a significant induction in birds exposed to 10ml/kg oil. Significant correlations were detected among all 4 AROD activities in both studies and between CYP1A protein expression and both MROD and PROD activities in study 2. EROD activity was highest for both treatment groups in both studies while BROD activity had the greatest fold-induction. While PROD activity values were consistently low, the fold-induction was high, usually 2nd highest to BROD activity. The observed induced AROD profiles detected in the present studies suggest both CYP1A4/1A5 DCCO isoforms are being induced after MC252 oil ingestion. A review of the literature on avian CYP1A AROD activity levels and protein expression after exposure to CYP1A inducers highlights the need for species-specific studies to accurately evaluate avian exposure to oil.

Clinostomum album n. sp. and Clinostomum marginatum (Rudolphi, 1819), parasites of the great egret Ardea alba L. from Mississippi, USA.

Members of the genus Clinostomum Leidy, 1856, colloquially known as yellow grubs, are cosmopolitan parasites of piscivorous birds, freshwater snails, fish and amphibians. In the southeastern United States, piscivorous birds present a continuous challenge for producers of farm-raised catfish. Ciconiiform birds are common hosts of Clinostomum spp. in North America and are endemic on most commercial catfish operations. The great egret Ardea alba L. is an avian predator often found foraging on commercial catfish operations, but to date the trematode fauna of great egrets preying on catfish ponds remains mostly understudied. Thirteen great egrets were captured from commercial catfish ponds in northeast Mississippi, and examined for trematode infections. Two morphologically distinct Clinostomum spp. were observed in the great egrets sampled, one morphologically consistent with Clinostomum marginatum (Rudolphi, 1819) and one morphologically unique species. These morphological descriptions were supplemented with molecular sequence data (c.4,800 bp of ribosomal DNA and c.600 bp of mitochondrial DNA). Gene sequences confirmed the identification of C. marginatum. However, the second species differed significantly from its congeners in both morphology and DNA sequence. Given these distinct morphological and molecular characters we propose this second species as Clinostomum album n. sp.

Dynamics of virus shedding and antibody responses in influenza A virus-infected feral swine.

Given their free-ranging habits, feral swine could serve as reservoirs or spatially dynamic 'mixing vessels' for influenza A virus (IAV). To better understand virus shedding patterns and antibody response dynamics in the context of IAV surveillance amongst feral swine, we used IAV of feral swine origin to perform infection experiments. The virus was highly infectious and transmissible in feral swine, and virus shedding patterns and antibody response dynamics were similar to those in domestic swine. In the virus-inoculated and sentinel groups, virus shedding lasted ≤ 6 and ≤ 9 days, respectively. Antibody titres in inoculated swine peaked at 1 : 840 on day 11 post-inoculation (p.i.), remained there until 21 days p.i. and dropped to < 1 : 220 at 42 days p.i. Genomic sequencing identified changes in wildtype (WT) viruses and isolates from sentinel swine, most notably an amino acid divergence in nucleoprotein position 473. Using data from cell culture as a benchmark, sensitivity and specificity of a matrix gene-based quantitative reverse transcription-PCR method using nasal swab samples for detection of IAV in feral swine were 78.9 and 78.1 %, respectively. Using data from haemagglutination inhibition assays as a benchmark, sensitivity and specificity of an ELISA for detection of IAV-specific antibody were 95.4 and 95.0 %, respectively. Serological surveillance from 2009 to 2014 showed that ∼7.58 % of feral swine in the USA were positive for IAV. Our findings confirm the susceptibility of IAV infection and the high transmission ability of IAV amongst feral swine, and also suggest the need for continued surveillance of IAVs in feral swine populations.

Seasonal climatic niche and migration movements of Double-crested Cormorants.

Avian migrants are challenged by seasonal adverse climatic conditions and energetic costs of long-distance flying. Migratory birds may track or switch seasonal climatic niche between the breeding and non-breeding grounds. Satellite tracking enables avian ecologists to investigate seasonal climatic niche and circannual movement patterns of migratory birds. The Double-crested Cormorant (Nannopterum auritum, hereafter cormorant) wintering in the Gulf of Mexico (GOM) migrates to the Northern Great Plains and Great Lakes and is of economic importance because of its impacts on aquaculture. We tested the climatic niche switching hypothesis that cormorants would switch climatic niche between summer and winter because of substantial differences in climate between the non-breeding grounds in the subtropical region and breeding grounds in the northern temperate region. The ordination analysis of climatic niche overlap indicated that cormorants had separate seasonal climatic niche consisting of seasonal mean monthly minimum and maximum temperature, seasonal mean monthly precipitation, and seasonal mean wind speed. Despite non-overlapping summer and winter climatic niches, cormorants appeared to be subjected to similar wind speed between winter and summer habitats and were consistent with similar hourly flying speed between winter and summer. Therefore, substantial differences in temperature and precipitation may lead to the climatic niche switching of fish-eating cormorants, a dietary specialist, between the breeding and non-breeding grounds.

EXPERIMENTAL ELUCIDATION OF THE LIFE CYCLE OF DREPANOCEPHALUS SPATHANS (DIGENEA: ECHINOSTOMATIDAE) WITH NOTES ON THE MORPHOLOGICAL PLASTICITY OF D. SPATHANS IN THE UNITED STATES.

The echinostomatid Drepanocephalus spathans (syn. Drepanocephalus auritus) parasitizes the double-crested cormorant Phalacrocorax auritus. In North America, the marsh rams-horn snail Planorbella trivolvis and ghost rams-horn snail Biomphalaria havanensis serve as snail intermediate hosts, both of which inhabit catfish aquaculture ponds in the southeastern United States. Studies have demonstrated D. spathans exposure can be lethal to juvenile channel catfish Ictalurus punctatus. Two studies were undertaken to elucidate the life cycle of D. spathans to establish a developmental time line. In both studies, D. spathans cercariae collected from naturally infected P. trivolvis individuals were used to infect channel catfish fingerlings, which were then fed to double-crested cormorants (DCCOs) that had been pharmaceutically dewormed. In study 1, laboratory-reared P. trivolvis and B. havanensis individuals were placed in aviary ponds with experimentally infected DCCO and examined bi-weekly for release of cercariae. Trematode eggs were observed in the feces of exposed birds 3 days post-infection. Birds were sacrificed 18 days post-exposure (dpe), and gravid adults morphologically and molecularly consistent with D. spathans were recovered. Snails from the aviary pond were observed shedding D. spathans cercariae 18-54 dpe. In study 2, trematode eggs were observed in the feces of exposed DCCOs beginning 8 dpe. Once eggs were observed, birds were allowed to defecate into clean tanks containing naïve laboratory-reared P. trivolvis individuals. Additionally, eggs from experimental DCCO feces were recovered by sedimentation and placed in an aquarium housing laboratory-reared P. trivolvis individuals. Birds in study 2 were sacrificed after 60 days, and gravid D. spathans specimens were recovered. Snails from the experimental DCCO tanks shed D. spathans cercariae 89-97 dpe. Lastly, trematode eggs were isolated and observed for the hatching of miracidia, which emerged on average after 16 days at ambient temperatures. No D. spathans adults were observed in control birds fed non-parasitized fish. This is the first experimental confirmation of the D. spathans life cycle, resolving previously unknown developmental time lines. In addition, the effects of fixation on adult trematode morphology were assessed, clarifying reports of pronounced morphological plasticity for D. spathans.

Inferring seasonal infection risk at population and regional scales from serology samples.

Accurate estimates of seasonal infection risk can be used by animal health officials to predict future disease risk and improve understanding of the mechanisms driving disease dynamics. It can be difficult to estimate seasonal infection risk in wildlife disease systems because surveillance assays typically target antibodies (serosurveillance), which are not necessarily indicative of current infection, and serosurveillance sampling is often opportunistic. Recently developed methods estimate past time of infection from serosurveillance data using quantitative serological assays that indicate the amount of antibodies in a serology sample. However, current methods do not account for common opportunistic and uneven sampling associated with serosurveillance data. We extended the framework of survival analysis to improve estimates of seasonal infection risk from serosurveillance data across population and regional scales. We found that accounting for the right-censored nature of quantitative serology samples greatly improved estimates of seasonal infection risk, even when sampling was uneven in time. Survival analysis can also be used to account for common challenges when estimating infection risk from serology data, such as biases induced by host demography and continually elevated antibodies following infection. The framework developed herein is widely applicable for estimating seasonal infection risk from serosurveillance data in humans, wildlife, and livestock.

Individual-Level Antibody Dynamics Reveal Potential Drivers of Influenza A Seasonality in Wild Pig Populations.

Swine are important in the ecology of influenza A virus (IAV) globally. Understanding the ecological role of wild pigs in IAV ecology has been limited because surveillance in wild pigs is often for antibodies (serosurveillance) rather than IAVs, as in humans and domestic swine. As IAV antibodies can persist long after an infection, serosurveillance data are not necessarily indicative of current infection risk. However, antibody responses to IAV infections cause a predictable antibody response, thus time of infection can be inferred from antibody levels in serological samples, enabling identification of risk factors of infection at estimated times of infection. Recent work demonstrates that these quantitative antibody methods (QAMs) can accurately recover infection dates, even when individual-level variation in antibody curves is moderately high. Also, the methodology can be implemented in a survival analysis (SA) framework to reduce bias from opportunistic sampling. Here we integrated QAMs and SA and applied this novel QAM-SA framework to understand the dynamics of IAV infection risk in wild pigs seasonally and spatially, and identify risk factors. We used national-scale IAV serosurveillance data from 15 US states. We found that infection risk was highest during January-March (54% of 61 estimated peaks), with 24% of estimated peaks occurring from May to July, and some low-level of infection risk occurring year-round. Time-varying IAV infection risk in wild pigs was positively correlated with humidity and IAV infection trends in domestic swine and humans, and did not show wave-like spatial spread of infection among states, nor more similar levels of infection risk among states with more similar meteorological conditions. Effects of host sex on IAV infection risk in wild pigs were generally not significant. Because most of the variation in infection risk was explained by state-level factors or infection risk at long-distances, our results suggested that predicting IAV infection risk in wild pigs is complicated by local ecological factors and potentially long-distance translocation of infection. In addition to revealing factors of IAV infection risk in wild pigs, our framework is broadly applicable for quantifying risk factors of disease transmission using opportunistic serosurveillance sampling, a common methodology in wildlife disease surveillance. Future research on the factors that determine individual-level antibody kinetics will facilitate the design of serosurveillance systems that can extract more accurate estimates of time-varying disease risk from quantitative antibody data.

Potential of Double-crested Cormorants ( Phalacrocorax auritus), American White Pelicans ( Pelecanus erythrorhynchos), and Wood Storks ( Mycteria americana) to Transmit a Hypervirulent Strain of Aeromonas hydrophila between Channel Catfish Culture Ponds.

Aeromonas hydrophila is a Gram-negative bacterium ubiquitous to freshwater and brackish aquatic environments that can cause disease in fish, humans, reptiles, and birds. Recent severe outbreaks of disease in commercial channel catfish ( Ictalurus punctatus) aquaculture ponds have been associated with a hypervirulent Aeromonas hydrophila strain (VAH) that is genetically distinct from less virulent strains. The epidemiology of this disease has not been determined. Given that research has shown that Great Egrets ( Ardea alba) can shed viable hypervirulent A. hydrophila after consuming diseased fish, we hypothesized that Double-crested Cormorants ( Phalacrocorax auritus), American White Pelicans ( Pelecanus erythrorhynchos), and Wood Storks ( Mycteria americana) could also serve as a reservoir for VAH and spread the pathogen during predation of fish in uninfected catfish ponds. All three species, when fed VAH-infected catfish, shed viable VAH in their feces, demonstrating their potential to spread VAH.

Seasonal effects of wind conditions on migration patterns of soaring American white pelican.

Energy and time expenditures are determinants of bird migration strategies. Soaring birds have developed migration strategies to minimize these costs, optimizing the use of all the available resources to facilitate their displacement. We analysed the effects of different wind factors (tailwind, turbulence, vertical updrafts) on the migratory flying strategies adopted by 24 satellite-tracked American white pelicans (Pelecanus erythrorhynchos) throughout spring and autumn in North America. We hypothesize that different wind conditions encountered along migration routes between spring and autumn induce pelicans to adopt different flying strategies and use of these wind resources. Using quantile regression and fine-scale atmospheric data, we found that the pelicans optimized the use of available wind resources, flying faster and more direct routes in spring than in autumn. They actively selected tailwinds in both spring and autumn displacements but relied on available updrafts predominantly in their spring migration, when they needed to arrive at the breeding regions. These effects varied depending on the flying speed of the pelicans. We found significant directional correlations between the pelican migration flights and wind direction. In light of our results, we suggest plasticity of migratory flight strategies by pelicans is likely to enhance their ability to cope with the effects of ongoing climate change and the alteration of wind regimes. Here, we also demonstrate the usefulness and applicability of quantile regression techniques to investigate complex ecological processes such as variable effects of atmospheric conditions on soaring migration.

Potential effects of incorporating fertility control into typical culling regimes in wild pig populations.

Effective management of widespread invasive species such as wild pigs (Sus scrofa) is limited by resources available to devote to the effort. Better insight of the effectiveness of different management strategies on population dynamics is important for guiding decisions of resource allocation over space and time. Using a dynamic population model, we quantified effects of culling intensities and time between culling events on population dynamics of wild pigs in the USA using empirical culling patterns and data-based demographic parameters. In simulated populations closed to immigration, substantial population declines (50-100%) occurred within 4 years when 20-60% of the population was culled annually, but when immigration from surrounding areas occurred, there was a maximum of 50% reduction, even with the maximum culling intensity of 60%. Incorporating hypothetical levels of fertility control with realistic culling intensities was most effective in reducing populations when they were closed to immigration and when intrinsic population growth rate was too high (> = 1.78) to be controlled by culling alone. However, substantial benefits from fertility control used in conjunction with culling may only occur over a narrow range of net population growth rates (i.e., where net is the result of intrinsic growth rates and culling) that varies depending on intrinsic population growth rate. The management implications are that the decision to use fertility control in conjunction with culling should rely on concurrent consideration of achievable culling intensity, underlying demographic parameters, and costs of culling and fertility control. The addition of fertility control reduced abundance substantially more than culling alone, however the effects of fertility control were weaker than in populations without immigration. Because these populations were not being reduced substantially by culling alone, fertility control could be an especially helpful enhancement to culling for reducing abundance to target levels in areas where immigration can't be prevented.

Effects of scale of movement, detection probability, and true population density on common methods of estimating population density.

Knowledge of population density is necessary for effective management and conservation of wildlife, yet rarely are estimators compared in their robustness to effects of ecological and observational processes, which can greatly influence accuracy and precision of density estimates. In this study, we simulate biological and observational processes using empirical data to assess effects of animal scale of movement, true population density, and probability of detection on common density estimators. We also apply common data collection and analytical techniques in the field and evaluate their ability to estimate density of a globally widespread species. We find that animal scale of movement had the greatest impact on accuracy of estimators, although all estimators suffered reduced performance when detection probability was low, and we provide recommendations as to when each field and analytical technique is most appropriately employed. The large influence of scale of movement on estimator accuracy emphasizes the importance of effective post-hoc calculation of area sampled or use of methods that implicitly account for spatial variation. In particular, scale of movement impacted estimators substantially, such that area covered and spacing of detectors (e.g. cameras, traps, etc.) must reflect movement characteristics of the focal species to reduce bias in estimates of movement and thus density.

Optimization of Scat Detection Methods for a Social Ungulate, the Wild Pig, and Experimental Evaluation of Factors Affecting Detection of Scat.

Collection of scat samples is common in wildlife research, particularly for genetic capture-mark-recapture applications. Due to high degradation rates of genetic material in scat, large numbers of samples must be collected to generate robust estimates. Optimization of sampling approaches to account for taxa-specific patterns of scat deposition is, therefore, necessary to ensure sufficient sample collection. While scat collection methods have been widely studied in carnivores, research to maximize scat collection and noninvasive sampling efficiency for social ungulates is lacking. Further, environmental factors or scat morphology may influence detection of scat by observers. We contrasted performance of novel radial search protocols with existing adaptive cluster sampling protocols to quantify differences in observed amounts of wild pig (Sus scrofa) scat. We also evaluated the effects of environmental (percentage of vegetative ground cover and occurrence of rain immediately prior to sampling) and scat characteristics (fecal pellet size and number) on the detectability of scat by observers. We found that 15- and 20-m radial search protocols resulted in greater numbers of scats encountered than the previously used adaptive cluster sampling approach across habitat types, and that fecal pellet size, number of fecal pellets, percent vegetative ground cover, and recent rain events were significant predictors of scat detection. Our results suggest that use of a fixed-width radial search protocol may increase the number of scats detected for wild pigs, or other social ungulates, allowing more robust estimation of population metrics using noninvasive genetic sampling methods. Further, as fecal pellet size affected scat detection, juvenile or smaller-sized animals may be less detectable than adult or large animals, which could introduce bias into abundance estimates. Knowledge of relationships between environmental variables and scat detection may allow researchers to optimize sampling protocols to maximize utility of noninvasive sampling for wild pigs and other social ungulates.