Immunohistochemical Evaluation of Rabies Virus Dissemination to Peripheral Tissues and Organs in Naturally Infected...

We describe the diagnostic utility of peripheral tissues from naturally infected free-ranging rabid striped skunks (Mephitis mephitis) to explore enhanced rabies surveillance testing using alternative tissues to the brainstem. Fifty-two dead or moribund adult striped skunks were collected by the Larimer County Health Department (Colorado, USA) as part of routine public health rabies surveillance during November 2018-July 2019 throughout Larimer County. Skunk carcasses were submitted to the Colorado State University Diagnostic Laboratory for rabies diagnosis using the direct fluorescent antibody test on the brainstem at the level on the pons and cerebellum. Multiple additional tissues were opportunistically collected from skunk carcasses and fixed in 10% neutral-buffered formalin, including brain, trigeminal ganglion, mandibular and parotid salivary glands, eyes, tongue, lip (including vibrissa), ear pinna, and uterus and fetus if pregnant. Fixed tissues were trimmed for histology, stained with H&E, and immunolabeled for detection of the rabies virus (RV) antigen. We detected RV immunopositive cells in multiple sections of brain, oral and nasal cavities, skin, and eyes as well as in the placenta and skin of fetuses from rabid skunks, suggesting the potential for transplacental transmission. We detected RV in the nerves and ganglia of salivary glands before detection in the salivary parenchymal cells in rabid skunks. The serous demilunes of the mandibular salivary gland appeared to be the first parenchymal cells to be infected with RV. We detected minimal RV immunopositive cells in the serous adenomeres of the parotid salivary gland and the mucous adenomeres of salivary tissue of the tongue. The results suggest that swabs from the oral and nasal cavities and cornea and biopsies of the skin could be of value for enhanced rabies surveillance in cases where the animal's brain has been destroyed or is unavailable.