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Emerg Infect Dis. 2006 August; 12(8): 1274–1277.
PMCID: PMC3291214

Bat-associated Rabies Virus in Skunks

Abstract

Rabies was undetected in terrestrial wildlife of northern Arizona until 2001, when rabies was diagnosed in 19 rabid skunks in Flagstaff. Laboratory analyses showed causative rabies viruses associated with bats, which indicated cross-species transmission of unprecedented magnitude. Public health infrastructure must be maintained to address emerging zoonotic diseases.

Keywords: Rabies, surveillance, bat, skunk, molecular epidemiology, wildlife reservoirs, RT-PCR, phylogenetic analysis, cross-species infection, dispatch

In North America, >90% of cases of rabies in animals occur in wildlife (1); several mammalian taxa harbor characteristic rabies virus variants (RABVV). In Arizona, skunks (Mephitis mephitis) and gray foxes (Urocyon cinereoargenteus) maintain independent rabies enzootic cycles, and in indigenous bats, rabies has been diagnosed in 14 of 28 species (Arizona Department of Health Services, unpub. data). Although skunks live throughout Arizona, until 2001, rabid skunks had been found only in the southeastern quadrant of the state.

In the United States, bat RABVV are a source of infection for humans and other mammals (28). Typically, interspecies infection produces a single fatal spillover event; secondary transmission has rarely been observed. Antigenic typing of rabid carnivores in Arizona from 1996 through 2000 identified bat RABVV in 1 domestic dog and 2 gray foxes. This report describes the largest documented rabies epizootic among terrestrial mammals infected with bat RABVV, with perpetuated animal-to-animal transmission. Coincident with the zoonotic disease significance, this report provides contemporary insight into pathogen evolution (9).

The Study

In January 2001, a homeowner contacted Flagstaff Animal Control about a dead skunk. Although no human had been exposed to the skunk, tissues were submitted to the Arizona State Health Laboratory, where rabies was diagnosed. This skunk was the first rabid terrestrial wild carnivore reported from the area. The Texas Department of State Health Services subsequently identified an RABVV associated with bats in tissues sent for antigenic characterization. From January through April, 14 more skunks, dead or exhibiting abnormal behavior, were found throughout a large residential subdivision within 4 km of the initial case. All were infected with the same bat RABVV. From April through July, 4 more skunks infected with bat RABVV were identified ≈9 km west of the initial focus (Figure 1). Control measures included prohibiting relocation of nuisance skunks, comprehensive public education, pet rabies vaccine clinics, and a 90-day emergency quarantine requiring pets to be leashed or confined and vaccinated (Figure 1). Additionally, 217 urban skunks were vaccinated and marked with ear tags during a 6-month phased program of trap, vaccinate, and release.

Figure 1
Temporal and geographic distribution of rabies outbreak in Flagstaff, Arizona. A) Timeline and control measures. TVR: trap, vaccinate, release program. B) Geographic location of rabid skunks (dark gray dots = subclade 1, light gray dots = subclade 2). ...

In Flagstaff and the surrounding county, during the decade before this epizootic, 2 rabid bats, on average, were reported each year. During the epizootic, 218 animals were submitted for rabies testing (Table). Rabies was confirmed in 19 (13%) of 145 tested skunks and 2 (9%) of 22 tested bats. Although most (18 [95%]) of the rabid skunks were identified and reported by lay citizens, no contact between these skunks and humans or domestic animals was reported.

Table
Animals from Flagstaff submitted for rabies diagnosis, January–July, 2001

Local baseline population estimates were not available to indicate whether skunk demography affected disease attributes. Synchronous with this outbreak, independent epizootic activity caused by well-established skunk RABVV was documented in southern Arizona, which suggests that regional skunk epizootiologic dynamics were similarly affected. Skunks' seasonal behavior may have contributed to transmission events. This epizootic was initially recognized when a dead skunk appeared in a snow-covered backyard, during a season when skunks are in communal dens. Given an incubation period of 2 months, most transmission would have occurred between late autumn (when skunks are in their dens) and late winter (when they are mating).The Flagstaff epizootic peak coincided with nationwide seasonal trends of rabid skunks (1). Enhanced postepizootic surveillance in Flagstaff did not detect additional rabid terrestrial mammals for the next 3 years. However, in 2004, a total of 5 skunks found in the initially affected east Flagstaff neighborhood and 1 fox 28 km south of Flagstaff were infected with the same bat RABVV (10).

Viruses isolated from the rabid skunks exhibited monoclonal antibody patterns similar to RABVV associated with big brown (Eptesicus fuscus) and Myotis bats in the western United States (11). These are among the most abundant bat species in Arizona and often roost in houses and outbuildings; however, no bat colonies were found in association with any of the rabid skunks. Restriction digests of PCR amplicons from the rabid skunks did not match patterns known for RABVV from North American terrestrial reservoirs (12). Phylogenetic analysis of a 300-bp region of the N gene showed that the Flagstaff skunk RABVV was identical (100%) to Arizona bat RABVV (Table A1, Figure 2A), and differed by 22% from skunk and gray fox RABVV. A monophyletic clade (clade A) of 8/8 big brown, 5/14 Myotis, and 1/6 southern yellow (Lasiurus ega) bats shared >95% identity with Flagstaff skunk RABVV. An additional 44 samples, representing 11 bat species, differed by >8% from Flagstaff skunk RABVV.

Figure 2
A) Phylogenetic tree of the 19 rabid skunk isolates and representative samples of known rabies virus variants (RABVV) from Arizona based on 300 bp of the nucleoprotein (N) gene (GenBank accession no. AY170226–304). B) Detailed analyses of clade ...

An analysis of clade A, which incorporates N and G genes, indicated that the Flagstaff skunk RABVV were more closely related to 2 bat RABVV (E. fuscus from Coconino County, M. velifer from Maricopa County) collected in 1999 and 1997 than to the 2 bat RABVV collected locally during the outbreak. In clade B, subclade 1 RABVV were collected from January through early April from the northeastern region of the outbreak, whereas subclade 2 RABVV were collected from early March through July from the southeastern and western regions of the outbreak (Figure 1). However, phylogenetic data do not support a wavelike spread from northeast to west because this would require nesting of subclade 1 within subclade 2. In contrast, both subclades exhibit independently derived mutations. East-to-west epizootic movement of RABVV within subclade 2 (sk16–19 form a monophyletic clade nested within subclade 2) during April is supported by the data and may be related to dispersal of infected skunks along river corridors or translocation by humans. One person reported trapping, moving, and releasing a skunk before the outbreak was known in the community. Alternatively, apparent shifts may be an artifact of intensified public awareness and reporting. Lack of sampling in the uninhabited forest between the eastern and western foci limits our ability to discriminate among these hypotheses.

Conclusions

This is the largest recorded cluster of bat RABVV infection in terrestrial mammals. Investigation of this novel outbreak showed evolution in action with the emergence of an RABVV that successfully adapted from Chiroptera to Carnivora. Previously documented clusters involving 3–4 to terrestrial mammals infected with a single insectivorous bat rabies virus variant did not corroborate sustained transmission (12). Although >1 skunk may have been exposed to a single rabid bat, it is highly unlikely that each skunk was exposed to the same bat or that multiple bat-skunk exposures occurred. We could not ascertain the complete scope of this outbreak or whether it was the index event. Phylogenetic analyses support the evolution of 2 independent lineages, suggesting establishment for months or years. Additionally, virus isolation from salivary glands of 5 affected skunks and the reappearance of rabid skunks with the same RABVV in 2004 support the probability of independent transmission.

The recognition of this epizootic can be credited to a coordinated laboratory-based disease surveillance program to monitor sick and dead wildlife for potential zoonoses (plague, tularemia, rabies) even in situations lacking human or pet exposures. Comprehensive animal disease surveillance provides direct benefits to public health and animal health by promoting early recognition of risk and opportunities for disease control and prevention interventions.

Unpredictable health threats related to emerging zoonoses, especially those involving wildlife reservoirs, pose notable surveillance and control challenges (1315). Recent bioterrorism initiatives emphasize integration of human and animal disease surveillance, and enhanced laboratory capacity, as essential functions in zoonosis detection (13). Rabies surveillance and control programs serve as historic prototypes for effective, long-term, public health programs. Quintessential zoonotic disease programs require innovative and expanded capacities, commitments to public health and veterinary laboratory infrastructure, and appropriate interagency and interdisciplinary coordination and communication.

Acknowledgments

We thank the citizens of Flagstaff and a large multiagency task force, who contributed to managing this outbreak, including Flagstaff City Police Department Animal Control Program, B. Worgess, P. Barbeau, C. Levy, J. Henderson, M. VanDeGriend, D. Bergman, Northern Arizona University, R. Rosatte, Texas Department of State Health Services Rabies Laboratory, M. Niezgoda, L. Orciari, J. Dragoo, Arizona Department of Game and Fish, Coconino County Humane Society, and the Coconino County Board of Supervisors. We also thank Doug Beckner for the timeline graphic.

Biography

• 

Dr Leslie was Arizona's state public health veterinarian during 1995–2002 and currently holds the same position in Washington State. Her work is focused on surveillance, investigation, and control of zoonotic and vectorborne diseases. She chairs the National Association of State Public Health Veterinarian's Compendium of Animal Rabies Prevention and Control Committee.

Table A1

Representative rabies virus variants found in Arizona and included in phylogenetic analyses
CDC IDState IDTaxon nameCommon nameScientific nameCollection dateCollection Site
3629504318Ap1Pallid batAntrozous pallidusJul 1997Navajo
3626498674Ap2Pallid batA. pallidusJul 1997Maricopa
3628500509Ap3Pallid batA. pallidusJul 1997Wickenburg
3925390721Ap4Pallid batA. pallidusSep 1995Coconino
3927735430Ap5Pallid batA. pallidusMar 1997Tucson
489110793Ef1Big brown batEptesicus fuscusSep 1999Yuma
4862947906Ef2Big brown batE. fuscusJun 1999Davis AFB
48679630Ef3Big brown batE. fuscusAug 1999Tucson
485015171Ef4Big brown batE. fuscusOct 1999Tucson
487199026842Ef5Big brown batE..fuscusJun 1999Coconino
488699046548Ef6Big brown batE. fuscusOct 1999Yavapi
54501034778Ef7Big brown batE. fuscusJul 2001Coconino
54421026934Ef8Big brown batE. fuscusJul 2001Coconino
407498007821Em1Spotted batEuderma maculatumSep 1998Maricopa
3057396597Ln1Silver-haired batLasionycteris noctivagansOct 1995Maricopa
3285432420Le6Southern yellow batLasiurus egaMay 1996Coconino
3046264111Le1Southern yellow batL. egaAug 1993Yuma
3870814565Le2Southern yellow batL. egaOct 1997Pima
3284447760Le3Southern yellow batL. egaAug 1996Yuma
3050374106Le4Southern yellow batL. egaJun 1995Yuma
3347395408Le5Southern yellow batL. egaSep 1995Yuma
5422141014Scsk1Striped skunkMephitis mephitisMay 2001Cochise
5423142140Scsk2Striped skunkM. mephitisMay 2001Cochise
49951001810Sk1Striped skunkM. mephitisJan 2001Coconino/Flagstaff
49981004508Sk2Striped skunkM. mephitisJan 2001Coconino/Flagstaff
50791006928Sk3Striped skunkM. mephitisJan 2001Coconino/Flagstaff
54701006403Sk4Striped slunkM. mephitisFeb 2001Coconino/Flagstaff
50741009087Sk5Striped skunkM. mephitisFeb 2001Coconino/Flagstaff
50751010227Sk6Striped skunkM. mephitisFeb 2001Coconino/Flagstaff
50761010229Sk7Striped skunkM. mephitisFeb 2001Coconino/Flagstaff
50771011591Sk8Striped skunkM. mephitisMar 2001Coconino/Flagstaff
50811012600Sk9Striped skunkM. mephitisMar 2001Coconino/Flagstaff
50801014008Sk10Striped skunkM. mephitisMar 2001Coconino/Flagstaff
51001015436Sk11Striped skunkM. mephitisApr 2001Coconino/Flagstaff
51011015687Sk12Striped skunkM. mephitisApr 2001Coconino/Flagstaff
51021015688Sk13Striped skunkM. mephitisApr 2001Coconino/Flagstaff
51031016511Sk14Striped skunkM. mephitisApr 2001Coconino/Flagstaff
51321016704Sk15Striped skunkM. mephitisApr 2001Coconino/Flagstaff
51331016707Sk16Striped skunkM. mephitisApr 2001Coconino/Flagstaff
54401023718Sk17Striped skunkM. mephitisMay 2001Coconino/Flagstaff
54411025813Sk18Striped skunkM. mephitisMay 2001Coconino/Flagstaff
54511036291Sk19Striped skunkM. mephitisJul 2001Coconino/Flagstaff
3858721957Mc1California myotisMyotis californicusDec 1996Pima
3847433224Mc2Western small-footed batM. ciliolabrumMay 1996Holbrook
3848445332Ml1Little brown batM. lucifugusAug 1996Wickenburg
488299043079Mu1Myotis batMyotis sp.Sep 1999Bullhead City
3862800694Mu2Myotis batMyotis sp.Jul 1997Pima
488799048912Mu3Myotis batMyotis sp.Oct 1999Pima
41932455Mu4Myotis batMyotis sp.Apr 1983Mesa
3350457788Mu5Myotis batMyotis sp.Oct 1996Maricopa
3351390759Mu6Myotis batMyotis sp.Sep 1995Yuma
488199042806Mu7Myotis batMyotis sp.Sep 1999Yuma
487399038008Mu8Myotis batMyotis sp.Aug 1999Eager
487299028231Mu9Myotis batMyotis sp.Jun 1999Pima
3855502675Mv1Cave myotisM. veliferJul 1997Maricopa
3852489891My1Yuma batM. yumanensisMay 1997San Carlos
3043259807Ph1Western pipistrellePipistrellus hesperusJul 1993Maricopa
3860747838Ph2Western pipistrelleP. hesperusJun 1997Pima
3044259808Ph3Western pipistrelleP. hesperusJul 1993Maricopa
3924455031Ph4Western pipistrelleP. hesperusSep 1996Navajo
20608543Ph5Western pipistrelleP. hesperusSep 1981Sedona
3863805082Ph6Western pipistrelleP. hesperusAug 1997Pima
3345466665Ph7Western pipistrelleP. hesperusDec 1996Maricopa
3859735181Ph8Western pipistrelleP. hesperusMar 1997Oro Valley
3053391577Ph9Western pipistrelleP. hesperusSep 1995Coconino
41026202Tb1Mexican free-tailed batTadarida brasiliensisMay 1985Riveria
4485646Tb2Mexican free-tailed batT. brasiliensisAug 1982Thatcher
4860943431Tb3Mexican free-tailed batT. brasiliensisMay 1999Tucson
4857821298Tb4Mexican free-tailed batT. brasiliensisDec 1997Tucson
488999057833Tb5Mexican free-tailed batT. brasiliensisDec 1999Phoenix
4863848036Tb6Mexican free-tailed batT. brasiliensisJun 1999Tucson
48689810071Tb7Mexican free-tailed batT. brasiliensisOct 1998Phoenix
4864949009Tb8Mexican free-tailed batT. brasiliensisJun 1999Tucson
4866949396Tb9Mexican free-tailed batT. brasiliensisJun 1999Tucson
4865949010Tb10Mexican free-tailed batT. brasiliensisJun 1999Tucson
484710197Tb11Mexican free-tailed batT. brasiliensisSep 1999Tucson
488499046042Tb12Mexican free-tailed batT. brasiliensisSep 1999Winslow
3344713793Pt1Townsend's big-eared batPlecotus townsendiiOct 1996Pima
3659513855Nm1Big free-tailed batNyctinomops macrotisOct 1997Yavapai

Footnotes

Suggested citation for this article: Leslie, MJ, Messenger S, Rohde RE, Smith J, Cheshier R, Hanlon C, et al. Bat-associated rabies virus in skunks. Emerg Infect Dis [serial on the Internet]. 2006 Aug [date cited]. http://dx.doi.org/10.3201/eid1208.051526

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