Detection of lymphoproliferative disease virus in Iowa Wild Turkeys (Meleagris gallopavo): Comparison of two sections of the proviral genome.

An accurate diagnostic test is an essential aspect of successfully monitoring and managing wildlife diseases. Lymphoproliferative Disease Virus (LPDV) is an avian retrovirus that was first identified in domestic turkeys in Europe and was first reported in a Wild Turkey (Meleagris gallopavo) in the United States in 2009. It has since been found to be widely distributed throughout North America. The majority of studies have utilized bone marrow and PCR primers targeting a 413-nucleotide sequence of the gag gene of the provirus to detect infection. While prior studies have evaluated the viability of other tissues for LPDV detection (whole blood, spleen, liver, cloacal swabs) none to date have studied differences in detection rates when utilizing different genomic regions of the provirus. This study examined the effectiveness of another section of the provirus, a 335-nucleotide sequence starting in the U3 region of the LTR (Long Terminal Repeat) and extending into the Matrix of the gag region (henceforth LTR), for detecting LPDV. Bone marrow samples from hunter-harvested Wild Turkeys (n = 925) were tested for LPDV with the gag gene and a subset (n = 417) including both those testing positive and those where LPDV was not detected was re-tested with LTR. The positive percent agreement (PPA) was 97.1% (68 of 70 gag positive samples tested positive with LTR) while the negative percent agreement (NPA) was only 68.0% (236 of 347 gag negative samples tested negative with LTR). Cohen's Kappa (κ = 0.402, Z = 10.26, p<0.0001) and the McNemar test (OR = 55.5, p<0.0001) indicated weak agreement between the two gene regions. We found that in Iowa Wild Turkeys use of the LTR region identified LPDV in many samples in which we failed to detect LPDV using the gag region and that LTR may be more appropriate for LPDV surveillance and monitoring. However, neither region of the provirus resulted in perfect detection and additional work is necessary to determine if LTR is more reliable in other geographic regions where LPDV occurs.

Prion protein polymorphisms in Michigan white-tailed deer (Odocoileus virginianus).

Chronic Wasting Disease (CWD), a well-described transmissible spongiform encephalopathy of the Cervidae family, is associated with the aggregation of an abnormal isoform (PrPCWD) of the naturally occurring host prion protein (PrPC). Variations in the PrP gene (PRNP) have been associated with CWD rate of infection and disease progression. We analysed 568 free-ranging white-tailed deer (Odocoileus virginianus) from 9 CWD-positive Michigan counties for PRNP polymorphisms. Sampling included 185 CWD-positive, 332 CWD non-detected, and an additional 51 CWD non-detected paired to CWD-positives by sex, age, and harvest location. We found 12 polymorphic sites of which 5 were non-synonymous and resulted in a change in amino acid composition. Thirteen haplotypes were predicted, of which 11 have previously been described. Using logistic regression, consistent with other studies, we found haplotypes C (OR = 0.488, 95% CI = 0.321-0.730, P < 0.001) and F (OR = 0.122, 95% CI = 0.007-0.612, P < 0.05) and diplotype BC (OR = 0.340, 95% CI = 0.154-0.709, P < 0.01) were less likely to be found in deer infected with CWD. As has also been documented in other studies, the presence of a serine at amino acid 96 was less likely to be found in deer infected with CWD (P < 0.001, OR = 0.360 and 95% CI = 0.227-0.556). Identification of PRNP polymorphisms associated with reduced vulnerability to CWD in Michigan deer and their spatial distribution can help managers design surveillance programmesand identify and prioritize areas for CWD management.

Precision of cementum annuli method for aging male white-tailed deer.

The most common method used to estimate ages of harvested white-tailed deer (Odocoileus virginianus) and other cervids is a criterion based on tooth replacement-and-wear (TRW). Previous studies have shown this method is prone to considerable error because TRW is partially subjective. A presumably more accurate, but more labor intensive and expensive, method to estimate age involves the counting of cementum annuli (CA) of cross-sectioned incisors. Quantifying rate of error of the CA aging method is not possible without known-aged specimens, but precision of duplicate CA age estimates for two teeth may be related to accuracy if identical factors influence both CA accuracy and precision. The objective of this research was to identify and assess factors affecting precision of paired CA ages as well as evaluate congruence between TRW and CA age estimates. We obtained paired CA age estimates from a laboratory specializing in CA aging for 473 adult (≥ 1 year old), male white-tailed deer harvested in Iowa (USA; 2014-2018). Not all CA age estimates of paired incisors agreed with one another and probability of agreement between the paired CA ages decreased as the certainty level of the CA ages provided by the laboratory decreased and was dependent upon the batches in which they were aged by the laboratory. We also estimated the age of 1,292 adult, male deer using both TRW and CA methods and compared the congruence between the TRW and CA age estimates. Congruence rates of CA and TRW ages differed among age classes (80% congruence in yearling TRW age classification, 65% with 2-year-olds, 78% with ≥3-year-olds). Our results showed that CA aging is imperfect and that the certainty level is an important factor to consider with CA ages, as shown in previous research, as is the batch in which the teeth were aged. We also confirmed previous studies' findings that CA and TRW ages for adult deer are not always congruent, particularly in age classes other than the yearling age class. Our results suggest managers are best served by using TRW to age adult deer as yearlings or ≥2-years-old. If additional age classes are required, CA aging is likely to be a better tool than TRW.

Survival of white-tailed deer fawns in central Iowa.

Understanding demographic parameters such as survival is important for scientifically sound wildlife management. Survival can vary by region, sex, age-class, habitat, and other factors. White-tailed deer fawn survival is highly variable across the species' range. While recent studies have investigated fawn survival in several Midwestern states, there have been no published estimates from Iowa for 30 years. We radio-collared 48 fawns in central Iowa from 2015-2017 to estimate survival, home range size, and habitat composition and identity causes of mortality. Estimated fawn survival (± SE) was similar to other Midwest studies at 30 (0.78 ± 0.07)) and 60 days (0.69 ± 0.08), but considerably lower at 7 months (0.31 ± 0.02). Survival was positively associated with woodland habitat through 30 and 60 days, but not related to habitat at 7 months. Female fawns avoided agricultural habitat in their home ranges. Fawn 95% kernel density home ranges were smaller than in other studies in the Midwest (21.22 ± 2.74 ha at 30 days, 25.47 ± 2.87 ha at 60 days, and 30.59 ± 2.37 ha at 7 months). The large amount of woodland and grassland (>90%) in our study area meant that fawns did not have to travel far to find suitable cover, which may explain their small home ranges. We recorded 21 mortalities, the leading cause of which was disease (n = 9; 56% epizootic hemorrhagic disease [EHD]) followed by suspected predation (4) and harvest (3). The mortality associated with an outbreak of EHD in 2016, all of which occurred after 60 days post-capture, is the most likely explanation for our low survival estimate at 7 months. While predation, usually early in life, is the leading cause of mortality in most studies, sporadic diseases like EHD can be a major source of mortality in older fawns in some years.

Detection of Mycoplasma ovipneumoniae in Pneumonic Mountain Goat ( Oreamnos americanus) Kids.

We documented bronchopneumonia in seven mountain goat ( Oreamnos americanus) kid mortalities between 2011 and 2015 following a pneumonia epizootic in bighorn sheep ( Ovis canadensis) and sympatric mountain goats in the adjacent East Humboldt Range and Ruby Mountains in Elko County, Nevada, US. Gross and histologic lesions resembled those described in bighorn lambs following all-age epizootics, and Mycoplasma ovipneumoniae was detected with real-time PCR in the lower and upper respiratory tracts of all kids. Mannheimia haemolytica, with one isolate being leukotoxigenic, was cultured from the upper respiratory tract of five kids, and in one kid, a leukotoxigenic strain of Mannheimia glucosida was isolated from both upper and lower respiratory tracts. During this same period, 75 mountain goats within the two populations were marked and sampled for respiratory pathogens, and M. ovipneumoniae, leukotoxigenic Bibersteinia trehalosi, and Mannheimia haemolytica were identified. The M. ovipneumoniae recovered from the kid mortalities shared the same DNA sequence-based strain type detected in the adult goats and sympatric bighorn sheep during and after the 2009-10 pneumonia outbreak. Clinical signs in affected kids, as well as decreased annual kid recruitment, also resembled reports in bighorn lambs from some herds following all-age pneumonia-associated die-offs. Mycoplasma ovipneumoniae, Pasteurellaceae spp., and other respiratory bacterial pathogens should be considered as a cause of pneumonia with potential population-limiting effects in mountain goats.

Evaluating the role of wild songbirds or rodents in spreading avian influenza virus across an agricultural landscape.

BACKGROUND: Avian influenza virus (AIV) infections occur naturally in wild bird populations and can cross the wildlife-domestic animal interface, often with devastating impacts on commercial poultry. Migratory waterfowl and shorebirds are natural AIV reservoirs and can carry the virus along migratory pathways, often without exhibiting clinical signs. However, these species rarely inhabit poultry farms, so transmission into domestic birds likely occurs through other means. In many cases, human activities are thought to spread the virus into domestic populations. Consequently, biosecurity measures have been implemented to limit human-facilitated outbreaks. The 2015 avian influenza outbreak in the United States, which occurred among poultry operations with strict biosecurity controls, suggests that alternative routes of virus infiltration may exist, including bridge hosts: wild animals that transfer virus from areas of high waterfowl and shorebird densities. METHODS: Here, we examined small, wild birds (songbirds, woodpeckers, etc.) and mammals in Iowa, one of the regions hit hardest by the 2015 avian influenza epizootic, to determine whether these animals carry AIV. To assess whether influenza A virus was present in other species in Iowa during our sampling period, we also present results from surveillance of waterfowl by the Iowa Department of Natural Resources and Unites Stated Department of Agriculture. RESULTS: Capturing animals at wetlands and near poultry facilities, we swabbed 449 individuals, internally and externally, for the presence of influenza A virus and no samples tested positive by qPCR. Similarly, serology from 402 animals showed no antibodies against influenza A. Although several species were captured at both wetland and poultry sites, the overall community structure of wild species differed significantly between these types of sites. In contrast, 83 out of 527 sampled waterfowl tested positive for influenza A via qPCR. DISCUSSION: These results suggest that even though influenza A viruses were present on the Iowa landscape at the time of our sampling, small, wild birds and rodents were unlikely to be frequent bridge hosts.

Mercury Exposure in Bald Eagles (Haliaeetus leucocephalus) Admitted for Rehabilitation in Iowa, USA.

To gain insight into mercury exposure in Bald Eagles ( Haliaeetus leucocephalus ) in Iowa, US we collected blood from 22 rehabilitation eagles in 2012-13. The geometric mean blood mercury level was 0.144 µg/g (95% confidence interval: 0.066-0.314) and was at the lower end of the range of levels reported elsewhere.

Association between agricultural land use and West Nile virus antibody prevalence in Iowa birds.

In the Plains states of the central United States, research suggests that the prevalence of West Nile virus (WNV) disease in humans is higher in agricultural areas than in nonagricultural areas. In contrast, there is limited information about WNV exposure in birds, the primary amplifying host of WNV, in agriculturally dominated landscapes. We evaluated whether exposure to WNV in peridomestic birds sampled in central Iowa varied with the proportion of land use devoted to agriculture. Over the summers of 2009 and 2010, we captured birds in sites comprising gradients of agricultural, urban, and natural land uses, and tested their sera for antibodies to WNV. Overall, WNV antibody prevalence was low (2.3%). Our results suggest that agricultural land use had minimal influence on WNV exposure in birds. We conclude that birds are not likely to be useful indicators of WNV activity in agricultural areas in the Plains states despite human risk being highest in those areas. Antibody prevalence for WNV, however, was higher in American Robins, Mourning Doves, and Northern Cardinals than in other species, making these species potentially useful for monitoring WNV activity in the US Plains states.

Efficacy of wildlife rehabilitation centers in surveillance and monitoring of pathogen activity: a case study with West Nile virus.

Surveillance is critical for identifying and monitoring pathogen activity in wildlife populations, but often is cost- and time-prohibitive and logistically challenging. We tested the hypothesis that wildlife rehabilitation centers are useful for monitoring pathogen activity using West Nile virus (WNV) as a case study. We hypothesized that birds submitted to wildlife rehabilitation centers would have a similar prevalence of antibody to WNV as free-ranging birds. From 2008 to 2010, we collected sera from peridomestic birds submitted to the Wildlife Care Clinic (WCC), a wildlife rehabilitation center in central Iowa, and tested them for antibodies to WNV. We also collected and tested sera from free-ranging peridomestic birds in the area from which approximately 50% of WCC submissions historically originated. Prevalences of WNV antibodies in free-ranging birds and in peridomestic WCC birds were 2.3% (44/1,936) and 2.8% (2/72), respectively. However, none of the birds submitted to the WCC from the area where we captured free-ranging birds had antibodies (0/29). Our results indicate that rehabilitation facilities are not likely to be useful for monitoring WNV activity at small spatial scales or over short-time periods due to the low endemic prevalence of WNV, and low and variable submission rates. However, at larger spatial scales (ca. nine Iowa counties) WNV antibody prevalence in peridomestic birds submitted to the WCC was similar to that of free-ranging birds. Although limitations to using rehabilitation birds to monitor WNV must be considered, testing these birds could be useful for monitoring WNV activity regionally, especially with many states limiting surveillance due to budgetary constraints.

Effects of chronic wasting disease on reproduction and fawn harvest vulnerability in Wisconsin white-tailed deer.

Chronic wasting disease (CWD) is a fatal, transmissible spongiform encephalopathy that affects free-ranging and captive North American cervids. Although the impacts of CWD on cervid survival have been documented, little is known about the disease impacts on reproduction and recruitment. We used genetic methods and harvest data (2002-04) to reconstruct parentage for a cohort of white-tailed deer (Odocoileus virginianus) fawns born in spring 2002 and evaluate the effects of CWD infection on reproduction and fawn harvest vulnerability. There was no difference between CWD-positive and CWD-negative male deer in the probability of being a parent. However, CWD-positive females were more likely to be parents than CWD-negative females. Because our results are based on harvested animals, we evaluated the hypothesis that higher parentage rates occurred because fawns with CWD-positive mothers were more vulnerable to harvest. Male fawns with CWD-positive mothers were harvested earlier (>1 mo relative to their mother's date of harvest) and farther away from their mothers than male fawns with CWD-negative mothers. Male fawns with CWD-positive mothers were also harvested much earlier and farther away than female fawns from CWD-positive mothers. Most female fawns (86%) with CWD-positive mothers were harvested from the same section as their mothers, while almost half of male and female fawns with CWD-negative mothers were farther away. We conclude that preclinical stages of CWD infection do not prohibit white-tailed deer from successfully reproducing. However, apparently higher harvest vulnerability of male fawns with CWD-positive mothers suggests that CWD infection may make females less capable of providing adequate parental care to ensure the survival and recruitment of their fawns.

Occurrence and molecular analysis of Campylobacter in wildlife on livestock farms.

Wildlife harbor a variety of Campylobacter spp. and may play a significant role in the transmission of Campylobacter to livestock. Although studies have been conducted on wildlife-associated Campylobacter isolates from farms in other countries, there are little data available for livestock farms in the United States. In addition, the critical questions of whether wildlife harbor Campylobacter that is pathogenic to ruminants and/or antibiotic-resistant Campylobacter have yet to be addressed. We captured wild small mammals (n=142) and small birds (n=188) at livestock farms in central Iowa and sampled them for thermophilic Campylobacter during autumn 2009, spring 2010, and autumn 2010. Overall prevalence was 4.79%, with isolates found only in wild birds. Molecular typing revealed four multilocus sequence types (STs), three of which are novel. The remaining ST (ST-806) was found in two house sparrows and is an ST previously associated with ruminant abortion cases. Further analysis of ST-806 wild bird and ruminant abortion isolates by pulsed-field gel electrophoresis, resistance gene location, and antibiotic susceptibility tests indicated that the isolates are nearly identical. This is the first account of isolation of Campylobacter types from wild birds that are known to be pathogenic to ruminants. Furthermore, these same two wild bird isolates are resistant to the antibiotic fluoroquinolone. Our results indicate there is an overall low prevalence of Campylobacter in selected wildlife in Iowa, but suggest that wildlife may play a role in the epidemiology of pathogenic Campylobacter for domestic livestock, and may also serve as a reservoir for antibiotic-resistant Campylobacter.

Genetic susceptibility to chronic wasting disease in free-ranging white-tailed deer: complement component C1q and Prnp polymorphisms.

The genetic basis of susceptibility to chronic wasting disease (CWD) in free-ranging cervids is of great interest. Association studies of disease susceptibility in free-ranging populations, however, face considerable challenges including: the need for large sample sizes when disease is rare, animals of unknown pedigree create a risk of spurious results due to population admixture, and the inability to control disease exposure or dose. We used an innovative matched case-control design and conditional logistic regression to evaluate associations between polymorphisms of complement C1q and prion protein (Prnp) genes and CWD infection in white-tailed deer from the CWD endemic area in south-central Wisconsin. To reduce problems due to admixture or disease-risk confounding, we used neutral genetic (microsatellite) data to identify closely related CWD-positive (n=68) and CWD-negative (n=91) female deer to serve as matched cases and controls. Cases and controls were also matched on factors (sex, location, age) previously demonstrated to affect CWD infection risk. For Prnp, deer with at least one Serine (S) at amino acid 96 were significantly less likely to be CWD-positive relative to deer homozygous for Glycine (G). This is the first characterization of genes associated with the complement system in white-tailed deer. No tests for association between any C1q polymorphism and CWD infection were significant at p<0.05. After controlling for Prnp, we found weak support for an elevated risk of CWD infection in deer with at least one Glycine (G) at amino acid 56 of the C1qC gene. While we documented numerous amino acid polymorphisms in C1q genes none appear to be strongly associated with CWD susceptibility.

Landscape genetics and the spatial distribution of chronic wasting disease.

Predicting the spread of wildlife disease is critical for identifying populations at risk, targeting surveillance and designing proactive management programmes. We used a landscape genetics approach to identify landscape features that influenced gene flow and the distribution of chronic wasting disease (CWD) in Wisconsin white-tailed deer. CWD prevalence was negatively correlated with genetic differentiation of study area deer from deer in the area of disease origin (core-area). Genetic differentiation was greatest, and CWD prevalence lowest, in areas separated from the core-area by the Wisconsin River, indicating that this river reduced deer gene flow and probably disease spread. Features of the landscape that influence host dispersal and spatial patterns of disease can be identified based on host spatial genetic structure. Landscape genetics may be used to predict high-risk populations based on their genetic connection to infected populations and to target disease surveillance, control and preventative activities.

TB-infected deer are more closely related than non-infected deer.

Identifying mechanisms of pathogen transmission is critical to controlling disease. Social organization should influence contacts among individuals and thus the distribution and spread of disease within a population. Molecular genetic markers can be used to elucidate mechanisms of disease transmission in wildlife populations without undertaking detailed observational studies to determine probable contact rates. Estimates of genealogical relationships within a bovine tuberculosis-infected white-tailed deer (Odocoileus virginianus) population indicated that infected deer were significantly more closely related than non-infected deer suggesting that contact within family groups was a significant mechanism of disease transmission. Results demonstrate that epidemiological models should incorporate aspects of host ecology likely to affect the probability of disease transmission.

Spatial epidemiology of chronic wasting disease in Wisconsin white-tailed deer.

Chronic wasting disease (CWD) is a fatal, emerging disease of cervids associated with transmissible protease-resistant prion proteins. The potential for CWD to cause dramatic declines in deer and elk populations and perceived human health risks associated with consuming CWD-contaminated venison have led wildlife agencies to embark on extensive CWD control programs, typically involving culling to reduce deer populations. We characterized the spatial distribution of CWD in white-tailed deer (Odocoileus virginianus) in Wisconsin to facilitate CWD management. We found that CWD prevalence declined with distance from a central location, was locally correlated at a scale of 3.6 km, and was correlated with deer habitat abundance. The latter result is consistent with patterns expected for a positive relationship between density and prevalence of CWD. We recommend management activities focused on culling in geographic areas with high prevalence to have the greatest probability of removing infected individuals. Further research is needed to elucidate the factors involved in CWD spread and infection rates, especially the role of density-dependent transmission.

Persistence of Pasteurella multocida in wetlands following avian cholera outbreaks.

Avian cholera, caused by Pasteurella multocida, affects waterbirds across North America and occurs worldwide among various avian species. Once an epizootic begins, contamination of the wetland environment likely facilitates the transmission of P. multocida to susceptible birds. To evaluate the ability of P. multocida serotype-1, the most common serotype associated with avian cholera in waterfowl in western and central North America, to persist in wetlands and to identify environmental factors associated with its persistence, we collected water and sediment samples from 23 wetlands during winters and springs of 1996-99. These samples were collected during avian cholera outbreaks and for up to 13 wk following initial sampling. We recovered P. multocida from six wetlands that were sampled following the initial outbreaks, but no P. multocida was isolated later than 7 wk after the initial outbreak sampling. We found no significant relationship between the probability of recovery of P. multocida during resampling and the abundance of the bacterium recovered during initial sampling, the substrate from which isolates were collected, isolate virulence, or water quality conditions previously suggested to be related to the abundance or survival of P. multocida. Our results indicate that wetlands are unlikely to serve as a long-term reservoir for P. multocida because the bacterium does not persist in wetlands for long time periods following avian cholera outbreaks.

Multi-species patterns of avian cholera mortality in Nebraska's Rainwater Basin.

Nebraska's Rainwater Basin (RWB) is a key spring migration area for millions of waterfowl and other avian species. Avian cholera has been endemic in the RWB since the 1970s and in some years tens of thousands of waterfowl have died from the disease. We evaluated patterns of avian cholera mortality in waterfowl species using the RWB during the last quarter of the 20th century. Mortality patterns changed between the years before (1976-1988) and coincident with (1989-1999) the dramatic increases in lesser snow goose abundance and mortality. Lesser snow geese (Chen caerulescens caerulescens) have commonly been associated with mortality events in the RWB and are known to carry virulent strains of Pasteurella multocida, the agent causing avian cholera. Lesser snow geese appeared to be the species most affected by avian cholera during 1989-1999; however, mortality in several other waterfowl species was positively correlated with lesser snow goose mortality. Coincident with increased lesser snow goose mortality, spring avian cholera outbreaks were detected earlier and ended earlier compared to 1976-1988. Dense concentrations of lesser snow geese may facilitate intraspecific disease transmission through bird-to-bird contact and wetland contamination. Rates of interspecific avian cholera transmission within the waterfowl community, however, are difficult to determine.