SOURCE AND SEASONALITY OF EPIZOOTIC MYCOPLASMOSIS IN FREE-RANGING PRONGHORN (ANTILOCAPRA AMERICANA).

Mycoplasma bovis is an economically important bacterial pathogen of cattle (Bos taurus) and bison (Bison bison) that most commonly causes pneumonia, polyarthritis, and mastitis. It is prevalent in cattle and ranched bison; however, infections in other species are rare. In early 2019, we identified M. bovis in free-ranging pronghorn (Antilocapra americana) in northeastern Wyoming. Here, we report on additional pronghorn mortalities caused by M. bovis, in the same approximately 120-km2 geographic region 1 yr later. Genetic analysis by multilocus sequence typing revealed that the mortalities were caused by the same M. bovis sequence type, which is unique among all sequence types documented thus far in North America. To explore whether pronghorn maintain chronic infections and begin assessing M. bovis status in other sympatric species, we used PCR testing of nasal swabs to opportunistically survey select free-ranging ungulates. We found no evidence of subclinical infections in 13 pronghorn sampled from the outbreak area (upper 95% binomial confidence limit [bCL], ∼24.7%) or among 217 additional pronghorn (upper 95% bCL, ∼1.7%) sampled from eight additional counties in Wyoming and 10 in Montana. All mule deer (Odocoileus hemionus; n=231; upper 95% bCL, ∼1.6%) sampled from 11 counties in Wyoming also were PCR negative. To assess the potential for environmental transmission, we examined persistence of M. bovis in various substrates and conditions. Controlled experiments revealed that M. bovis can remain viable for 6 h in shaded water and 2 h in direct sunlight. Our results indicate that environmental transmission of M. bovis from livestock to pronghorn is possible and that seasonality of infection could be due to shared resources during late winter. Further investigations to better understand transmission dynamics, to assess population level impacts to pronghorn, and to determine disease risks among pronghorn and other ungulate taxa appear warranted.

DICTYOCAULUS CERVI-LIKE LUNGWORM INFECTION IN A ROCKY MOUNTAIN ELK (CERVUS CANADENSIS NELSONI) FROM WYOMING, USA.

Dictyocaulus spp. infections are common in North American cervids, with Dictyocaulus viviparus described as most common. A Rocky Mountain elk (Cervus canadensis nelsoni) was found dead in Wyoming, US with significant bronchitis and pneumonia. In the bronchi and trachea, numerous large nematodes were found and grossly identified as Dictyocaulus spp. lungworms. Macroscopic alterations, such as distended interlobular septa and edema with foam and mucus observed on cut surface and in trachea and bronchi, were consistent with those commonly described in D. viviparus infections. Female lungworms were identified to Dictyocaulus spp. level via morphologic examination and molecular analyses based on mitochondrial cyclooxygenase 1 and 18S ribosomal RNA genes. A phylogenetic analysis was conducted employing the maximum likelihood method. Based on both morphologic and genetic assays, the isolated lungworms were most likely a strain of Dictyocaulus cervi. Within the female adult worms, free first stage larvae were observed besides worm eggs, which had not been described for Dictyocaulus spp. Phylogenetic analysis revealed that our parasites clustered closely with D. cervi, forming a subclade with that species within a larger clade that includes Dictyocaulus eckerti. While the elk tested positive for chronic wasting disease, it is assumed that significant pathology in the present case was caused directly by infection with the D. cervi-like lungworm, not previously described in North America.

Mycoplasma bovis Infections in Free-Ranging Pronghorn, Wyoming, USA.

Mycoplasma bovis is 1 of several bacterial pathogens associated with pneumonia in cattle. Its role in pneumonia of free-ranging ungulates has not been established. Over a 3-month period in early 2019, ¼60 free-ranging pronghorn with signs of respiratory disease died in northeast Wyoming, USA. A consistent finding in submitted carcasses was severe fibrinosuppurative pleuropneumonia and detection of M. bovis by PCR and immunohistochemical analysis. Multilocus sequence typing of isolates from 4 animals revealed that all have a deletion in 1 of the target genes, adh-1. A retrospective survey by PCR and immunohistochemical analysis of paraffin-embedded lung from 20 pronghorn that died with and without pneumonia during 2007-2018 yielded negative results. These findings indicate that a distinct strain of M. bovis was associated with fatal pneumonia in this group of pronghorn.

Infectious keratoconjunctivitis in free-ranging mule deer in Wyoming: a retrospective study and identification of a novel alphaherpesvirus.

We describe the clinicopathologic findings, relative prevalence, and pathogens associated with infectious keratoconjunctivitis in mule deer ( Odocoileus hemionus) in Wyoming. Seventeen cases with ocular lesions were identified among 1,036 mule deer postmortem submissions (1.6%) in an ~16 y period. Sixteen cases were observed in winter and most were in male (15 cases) and juvenile (13 cases) deer. Blindness was the most commonly reported clinical sign (10 cases). A herpesvirus was detected only in the 4 cases of bilateral necrotizing bulbar conjunctivitis. Phylogenetic analysis of glycoprotein amino acid sequences consistently identified this virus as a novel alphaherpesvirus. In 2 of these herpesvirus-positive cases, Actinomyces sp. and Moraxella ovis were also identified. Trueperella pyogenes was identified in 4 cases of unilateral ulcerative keratitis, keratoconjunctivitis, and panophthalmitis. M. ovis was cultured from 3 cases of bilateral conjunctivitis and keratoconjunctivitis. In the remaining cases, isolates included Moraxella bovis (1 case), Staphylococcus sp. and Streptococcus sp. (2), Flavobacterium sp. and Pseudomonas sp. (2), Escherichia coli and Enterobacter sp. (1), and bovine viral diarrhea virus 1 (1). No pathogens were identified in 2 cases. The relative prevalence of keratoconjunctivitis in mule deer in Wyoming appears to be low, and this disease is most commonly associated with infection by a novel alphaherpesvirus, T. pyogenes, and M. ovis.

Whole-genome sequences of Odocoileus hemionus deer adenovirus isolates from deer, moose and elk are highly conserved and support a new species in the genus Atadenovirus.

We present the first complete genome sequence of Odocoileus hemionus deer adenovirus 1 (OdAdV-1). This virus can cause sporadic haemorrhagic disease in cervids, although epizootics with high mortality have occurred in California. OdAdV-1 has been placed in the genus Atadenovirus, based on partial hexon, pVIII and fibre genes. Ten field isolates recovered from naturally infected mule deer (Odocoileus hemionus), white-tailed deer (Odocoileus virginiana) and moose (Alces alces) from Wyoming, black-tailed deer (Odocoileus hemionus columbianus) from California, and Rocky Mountain elk (Cervus elaphus nelsoni) from Colorado and Washington state were sequenced. The genome lengths ranged from 30 620 to 30 699 bp, contained the predicted proteins and gene organization typical of members of genus Atadenovirus, and had a high percentage of A/T nucleotides (66.7 %). Phylogenic analysis found that the closest ancestry was with ruminant atadenoviruses, while a divergence of the hexon, polymerase and penton base proteins of more than 15 % supports classification as a new species. Genetic global comparison between the 10 isolates found an overall 99 % identity, but greater divergence was found between those recovered from moose and elk as compared to deer, and a single variable region contained most of these differences. Our findings demonstrate that OdAdV-1 is highly conserved between 10 isolates recovered from multiple related cervid species, but genotypic differences, largely localized to a variable region, define two strains. We propose that the virus type name be changed to cervid adenovirus 1, with the species name Cervid atadenovirus A. Sequence data were used to develop molecular assays for improved detection and genotyping.

Declining mortality in American crow (Corvus brachyrhynchos) following natural West Nile virus infection.

The American crow (Corvus brachyrhynchos) is known to suffer 100% mortality from infection with the New York 1999 strain of West Nile virus (WNV). Following the initial detection of WNV in North America in 1999, we measured prevalence of WNV-reactive antibodies ("seroprevalence") in free-ranging American and fish crows (Corvus ossifragus) of central New Jersey after each transmission season through 2005. In 2002, seroprevalence in American crow juveniles increased to 14% from the 5% of the previous year, potentially indicating increased survival in this species. Using the annual seroprevalence measurements and the number of human West Nile neuroinvasive disease cases as a surrogate for WNV transmission intensity, we developed a model to estimate the annual WNV-associated mortality rates among both of these crow species. Our model supports the hypothesis that mortality is changing over time; the WNV-associated mortality rate declined over time by 1.5% for American crow and by 1.1% for fish crow. The probability that the trend in mortality was negative was 90% for the American crow and 60% for the fish crow.

DNA vaccine for West Nile virus infection in fish crows (Corvus ossifragus).

A DNA vaccine for West Nile virus (WNV) was evaluated to determine whether its use could protect fish crows (Corvus ossifragus) from fatal WNV infection. Captured adult crows were given 0.5 mg of the DNA vaccine either orally or by intramuscular (IM) inoculation; control crows were inoculated or orally exposed to a placebo. After 6 weeks, crows were challenged subcutaneously with 105 plaque-forming units of WNV (New York 1999 strain). None of the placebo inoculated-placebo challenged birds died. While none of the 9 IM vaccine-inoculated birds died, 5 of 10 placebo-inoculated and 4 of 8 orally vaccinated birds died within 15 days after challenge. Peak viremia titers in birds with fatal WNV infection were substantially higher than those in birds that survived infection. Although oral administration of a single DNA vaccine dose failed to elicit an immune response or protect crows from WNV infection, IM administration of a single dose prevented death and was associated with reduced viremia.

A baiting system for delivery of an oral plague vaccine to black-tailed prairie dogs.

Laboratory and field studies were conducted between July and October 1999 to identify bait preference, biomarker efficacy, and bait acceptance rates for delivering an oral plague vaccine to black-tailed prairie dogs (Cynomys ludovicianus). Twenty juvenile captive prairie dogs were offered alfalfa baits containing either alfalfa, alfalfa and 5% molasses, or alfalfa, 5% molasses and 4% salt. Based on the results of these trials we selected a bait containing alfalfa, 7% molasses, and 1% salt for field trials to determine bait acceptance rates by free-ranging animals. The biomarkers DuPont Blue dye, iophenoxic acid, and tetracycline hydrochloride were orally administered to captive prairie dogs to determine their efficacy. Only tetracycline proved effective as a biomarker. Two field trials were conducted at separate prairie dog colonies located at the Buffalo Gap National Grassland (Pennington County, South Dakota, USA). In Trial 1, three baits containing tetracycline were distributed around each active burrow entrance and an additional bait was placed inside the burrow (1,276 baits total). In Trial 2, baits were distributed at the same density per burrow as Trial 1, but along transects spaced 10 m apart (1,744 baits total). Trapping began 3 days after bait distribution, and 30 prairie dogs then were captured at each site to determine the percentage of animals marked. In Trial 1, 67% of the prairie dogs captured had tetracycline deposits indicative of bait consumption. In Trial 2, 83% of the prairie dogs had ingested a bait. Approximately 15% of the animals in both trials ate more than one bait. Fleas (Opisocrostis hirsutus) were found on 64 of 70 (91%) of the prairie dogs captured during this study.