Epidemiologic and Genomic Evidence for Zoonotic Transmission of SARS-CoV-2 among People and Animals on a Michigan Mink Farm, United States, 2020.

Farmed mink are one of few animals in which infection with SARS-CoV-2 has resulted in sustained transmission among a population and spillback from mink to people. In September 2020, mink on a Michigan farm exhibited increased morbidity and mortality rates due to confirmed SARS-CoV-2 infection. We conducted an epidemiologic investigation to identify the source of initial mink exposure, assess the degree of spread within the facility's overall mink population, and evaluate the risk of further viral spread on the farm and in surrounding wildlife habitats. Three farm employees reported symptoms consistent with COVID-19 the same day that increased mortality rates were observed among the mink herd. One of these individuals, and another asymptomatic employee, tested positive for SARS-CoV-2 by real-time reverse transcription PCR (RT-qPCR) 9 days later. All but one mink sampled on the farm were positive for SARS-CoV-2 based on nucleic acid detection from at least one oral, nasal, or rectal swab tested by RT-qPCR (99%). Sequence analysis showed high degrees of similarity between sequences from mink and the two positive farm employees. Epidemiologic and genomic data, including the presence of F486L and N501T mutations believed to arise through mink adaptation, support the hypothesis that the two employees with SARS-CoV-2 nucleic acid detection contracted COVID-19 from mink. However, the specific source of virus introduction onto the farm was not identified. Three companion animals living with mink farm employees and 31 wild animals of six species sampled in the surrounding area were negative for SARS-CoV-2 by RT-qPCR. Results from this investigation support the necessity of a One Health approach to manage the zoonotic spread of SARS-CoV-2 and underscores the critical need for multifaceted public health approaches to prevent the introduction and spread of respiratory viruses on mink farms.

FOOT INJURIES IN MICHIGAN, USA, GRAY WOLVES (CANIS LUPUS), 1992-2014.

The range of gray wolves (Canis lupus) in the contiguous US is expanding. Research and monitoring to support population recovery and management often involves capture via foothold traps. A population-level epidemiologic assessment of the effect of trap injuries on wolf survival remains needed to inform management. We describe the baseline rate, type, and severity of foot injuries of wolves born 1992-2013 in Michigan's Upper Peninsula, evaluate the reliability of field-scoring trap-related injuries, and the effect of injuries on wolf survival. We assessed foot injuries by physical and radiographic exam at postmortem and/or time of capture for 351 wolves using the International Organization for Standardization 10990-5 standard and the effects of injuries, sex, age, previous capture and body condition on survival using proportional hazards regression. We used ordinal regression to evaluate epidemiologic associations between sex, age, previous capture, body condition, cause of death and injury severity. Most wolves (53%) experienced no physically or radiographically discernable foot injuries over their lifetimes. Among those wolves that did experience injuries, 33% scored as mild. Foot injuries had little epidemiologically discernable effect on survival rates. Wolves with higher foot trauma scores did experience an increased risk of dying, but the magnitude of the increase was modest. Most limb injuries occurred below the carpus or tarsus, and scoring upper-limb injuries added little predictive information to population-level epidemiologic measures of survival and injury severity. There was little association between injury severity and cause of death. Based on necropsy exams, previous trap injuries likely contributed to death in only four wolves (1.1%). Our results suggest that injuries resulting from foothold traps are unlikely to be a limiting factor in recovery and ongoing survival of the Michigan gray wolf population.

Chronic Wasting Disease Diagnostic Discrepancies: The Importance of Testing Both Medial Retropharyngeal Lymph Nodes.

Chronic wasting disease (CWD) of white-tailed deer (Odocoileus virginianus) is a fatal neurologic disease that is spreading across North America. A common surveillance protocol for CWD currently involves screening with an enzyme-linked immunosorbent assay (ELISA) followed by confirmatory testing with immunohistochemistry (IHC). Medial retropharyngeal lymph nodes (MRPLN) are the tissue of choice to diagnose CWD in free-ranging white-tailed deer. We examined left and right MRPLN from 101 ELISA-positive deer harvested from 2015 to 2019 to determine the prevalence of cases in which prion protein was not detected by IHC as well as differences in IHC labeling between contralateral lymph nodes. Prion protein was not detected using IHC in either MRPLN in 5.9% (6/101) of cases. There was a significant but weak positive relationship between the number of IHC-positive follicles and ELISA optical density values (R2=0.08, P=0.039). Mean optical density values in IHC-positive MRPLN were higher than in IHC-negative MRPLN; however, this was not statistically significant (P=0.260). Failure to confirm ELISA diagnoses with IHC may have been because the methods tested different areas of MRPLN, or that there were differences in test sensitivity or antibody affinity. An additional 5.9% (6/101) of cases had one IHC-positive MRPLN, whereas the contralateral MRPLN was IHC negative. Therefore, testing a single MRPLN for CWD may lead to false-negative results, regardless of methodology, which highlights the importance of collecting and testing both MRPLN.

Evaluation of Real-Time Quaking-Induced Conversion, ELISA, and Immunohistochemistry for Chronic Wasting Disease Diagnosis.

Chronic wasting disease (CWD) is a transmissible prion disorder, primarily affecting free-ranging and captive cervids in North America (United States and Canada), South Korea, and Europe (Finland, Norway, and Sweden). Current diagnostic methods used in the United States for detection of CWD in hunter harvested deer involve demonstration of the causal misfolded prion protein (PrPCWD) in the obex or retropharyngeal lymph nodes (RLNs) using an antigen detection ELISA as a screening tool, followed by a confirmation by the gold standard method, immunohistochemistry (IHC). Real-time quaking-induced conversion (RT-QuIC) assay is a newer approach that amplifies misfolded CWD prions in vitro and has facilitated CWD prion detection in a variety of tissues, body fluids, and excreta. The current study was undertaken to compare ELISA, IHC, and RT-QuIC on RLNs (n = 1,300 animals) from white-tailed deer (WTD) in Michigan. In addition, prescapular, prefemoral and popliteal lymph nodes collected from a small subset (n = 7) of animals were tested. Lastly, the location of the positive samples within Michigan was documented and the percentage of CWD positive RLNs was calculated by sex and age. ELISA and RT-QuIC detected PrPCWD in 184 and 178 out of 1,300 RLNs, respectively. Of the 184 ELISA positive samples, 176 were also IHC positive for CWD. There were seven discordant results when comparing IHC and ELISA. RT-QuIC revealed that six of the seven samples matched the IHC outcomes. One RLN was negative by IHC, but positive by ELISA and RT-QuIC. RT-QuIC, IHC, and ELISA also detected PrPCWD in prescapular, prefemoral and popliteal lymph nodes. CWD infection heterogeneities were observed in different age and sex groups, with young males having higher CWD prevalence. All, except one, CWD positive RLNs analyzed were from ten Counties geographically located in the West Michigan region of the Lower Peninsula. Taken together, we show evidence that the RT-QuIC assay is comparable to ELISA and IHC and could be helpful for routine CWD detection in surveillance programs. RT-QuIC also demonstrated that CWD prions are distributed across lymph nodes in a variety of anatomic locations. A multi-laboratory validation on blinded sample panels is underway and is likely to help to provide insight into the variability (lab-to-lab), analytical sensitivity, and specificity of gold standard diagnostics vs. RT-QuIC assay.

Genetic polymorphism of Baylisascaris procyonis in host infrapopulations and component populations in the Central USA.

Baylisascaris procyonis is a nematode of significant concern to public and domestic animal health as well as wildlife management. The population genetics of B. procyonis is poorly understood. To gain insights into patterns of genetic diversity within (infrapopulation level) and among (component population level) raccoon (Procyon lotor) hosts, and specifically to assess the relative importance of indirect and direct transmission of the parasite for explaining observed population structure, we collected 69 B. procyonis from 17 wild raccoons inhabiting five counties in Missouri and Arkansas, USA. Informative regions of mitochondrial (CO1, CO2) and nuclear (28S, ITS2) genes were amplified and the distribution and genetic variability of these genes were assessed within and across raccoons. Concatenation of the CO1 and CO2 mtDNA sequences resulted in 5 unique haplotypes, with haplotype diversity 0.456 ±â€¯0.068. The most common haplotype occurred in 94% of raccoons and 72.5% of B. procyonis. Sequences for 28S rDNA revealed four unique nuclear genotypes, the most common found in 100% of raccoons and 82.6% of B. procyonis. ITS2 genotypes were assessed using fragment analysis, and there was a 1:1 correspondence between 28S and ITS-2 genotypes. Infrapopulation variation in haplotypes and genotypes was high and virtually all hosts infected with multiple sequenced nematodes also harbored multiple haplotypes and genotypes. There was a positive relationship between the size of the analyzed infrapopulation (i.e., the number of nematodes analyzed) and the number of haplotypes identified in an individual. Collectively this work emphasizes the importance of indirect transmission in the lifecycle to this parasite.

MENINGEAL WORM ( PARELAPHOSTRONGYLUS TENUIS) AS A CAUSE OF MORTALITY IN THE RESTORED ELK ( CERVUS CANADENSIS) POPULATION IN MISSOURI, USA.

: Meningeal worm ( Parelaphostrongylus tenuis) is an important cause of mortality of elk ( Cervus canadensis) in populations in the eastern US and has been implicated in the failure of several restoration attempts. From 2011 to 2013, the Missouri Department of Conservation translocated 108 adult and yearling elk from Kentucky (US) to southern Missouri (US) to establish a free-ranging population. From release in spring 2011 through August 2015, we monitored 167 elk (adult, yearling, and calf) to determine causes of mortality. Of 78 mortalities, 26 (33%) were linked to meningeal worm based on necropsy results and/or observed behavior; this group included 19 elk with confirmed or suspected cases of meningeal worm infection that died of other proximate causes. Other important mortality sources included euthanasia ( n=11, 14%), emaciation ( n=7, 9%), and predation ( n=5, 6%). Eleven of the 26 (42%) meningeal worm-related mortalities were adults, and 22 (85%) were female. Meningeal worm was an important cause of mortality during the restoration of Missouri elk, potentially contributing to the loss of 16% of the monitored individuals. Greater mortality in adult female elk could reduce initial population growth by limiting reproductive output in the restored herd, especially given that females were disproportionately affected in Missouri. Because translocated Missouri elk undoubtedly were exposed to meningeal worm in Kentucky, our results could be explained by exposure to a different genetic strain of meningeal worm once in Missouri, loss of immune response due to translocation, increased dose of larval worms, or some unquantified factor.

Transmission of brucellosis from elk to cattle and bison, Greater Yellowstone area, U.S.A., 2002-2012.

Bovine brucellosis has been nearly eliminated from livestock in the United States. Bison and elk in the Greater Yellowstone Area remain reservoirs for the disease. During 1990-2002, no known cases occurred in Greater Yellowstone Area livestock. Since then, 17 transmission events from wildlife to livestock have been investigated.