In nature, rabies virus (RABV; genus Lyssavirus, family Rhabdoviridae) represents an assemblage of phylogenetic lineages, associated with specific mammalian host species. Although it is generally accepted that RABV evolved originally in bats and further shifted to carnivores, mechanisms of such host shifts are poorly understood, and examples are rarely present in surveillance data. Outbreaks in carnivores caused by a RABV variant, associated with big brown bats, occurred repeatedly during 2001–2009 in the Flagstaff area of Arizona. After each outbreak, extensive control campaigns were undertaken, with no reports of further rabies cases in carnivores for the next several years. However, questions remained whether all outbreaks were caused by a single introduction and further perpetuation of bat RABV in carnivore populations, or each outbreak was caused by an independent introduction of a bat virus. Another question of concern was related to adaptive changes in the RABV genome associated with host shifts. To address these questions, we sequenced and analyzed 66 complete and 20 nearly complete RABV genomes, including those from the Flagstaff area and other similar outbreaks in carnivores, caused by bat RABVs, and representatives of the major RABV lineages circulating in North America and worldwide. Phylogenetic analysis demonstrated that each Flagstaff outbreak was caused by an independent introduction of bat RABV into populations of carnivores. Positive selection analysis confirmed the absence of post-shift changes in RABV genes. In contrast, convergent evolution analysis demonstrated several amino acids in the N, P, G and L proteins, which might be significant for pre-adaptation of bat viruses to cause effective infection in carnivores. The substitution S/T₂₄₂ in the viral glycoprotein is of particular merit, as a similar substitution was suggested for pathogenicity of Nishigahara RABV strain. Roles of the amino acid changes, detected in our study, require additional investigations, using reverse genetics and other approaches. Host shifts of the rabies virus (RABV) from bats to carnivores are important for our understanding of viral evolution and emergence, and have significant public health implications, particularly for the areas where “terrestrial” rabies has been eliminated. In this study we addressed several rabies outbreaks in carnivores that occurred in the Flagstaff area of Arizona during 2001–2009, and caused by the RABV variant associated with big brown bats (Eptesicus fuscus). Based on phylogenetic analysis we demonstrated that each outbreak resulted from a separate introduction of bat RABV into populations of carnivores. No post-shift changes in viral genomes were detected under the positive selection analysis. Trying to answer the question why certain bat RABV variants are capable for host shifts to carnivores and other variants are not, we developed a convergent evolution analysis, and implemented it for multiple RABV lineages circulating worldwide. This analysis identified several amino acids in RABV proteins which may facilitate host shifts from bats to carnivores. Precise roles of these amino acids require additional investigations, using reverse genetics and animal experimentation. In general, our approach and the results obtained can be used for prediction of host shifts and emergence of other zoonotic pathogens.

Cat food left out for feral and domestic cats and bird seed spilled from backyard bird feeders are two common anthropogenic food sources that may attract non-target animals like urban mesocarnivores but no studies have quantified mesocarnivore visitation at these food sources. We used motion-activated video cameras to monitor mesocarnivore use of spilled bird seed below 25 bird feeders maintained by residents in four neighborhoods in Flagstaff, Arizona, June-September 2012 and 2014. During the first five nights of monitoring only seed that spilled naturally below feeders was available. On each of the subsequent five nights, we placed a bowl of commercially available dry cat food below feeders so that both spilled seed and cat food were present. In both years, after cat food was added, the number of visits by striped skunks (Mephitis mephitis), raccoons (Procyon lotor) and domestic cats (Felis cattus) doubled and the number of times two animals were present simultaneously also increased. Aggressive interactions, in the form of displays or contacts, increased for all species combinations but significantly only between skunks in the presence of cat food. These results demonstrate that both spilled bird seed and cat food may be exploited frequently by urban mesocarnivores and that the type of food can elicit different behavioral responses that could have important implications for human-wildlife conflict and disease transmission.

We followed radio-collared striped skunks (Mephitis mephitis) from January 2004– December 2005 in two urban areas of Flagstaff, Arizona, USA to determine seasonal patterns of movement and home-range size. We also used automated cameras to determine the potential for inter- and intraspecific interaction at skunks' diurnal resting sites and nocturnal focal locations. We found no difference between sexes in nightly rates of travel or in size of seasonal home range. Nightly rates of travel were greatest in the postbreeding months (May–July) and smallest from November to February, consistent with larger home ranges (95% kernel estimates) from March– August and smaller home ranges from September–February. Sixty-three percent of monitored males and 38% of monitored females crossed the urban–wildland interface, in at least one direction on at least one night, and some remained outside the urban area for days or weeks, indicating that skunks could act as vectors of disease across the urban–wildland interface. We recorded co-occurrence of skunks with domestic cats (Felis domesticus), raccoons (Procyon lotor), gray foxes (Urocyon cinereoargenteus), and other skunks at focal locations and diurnal retreats used by skunks, suggesting these areas are potential sites for both inter- and intraspecific rabies transmission and could be targeted by wildlife managers during trapping or vaccination programs.