Eastern Equine Encephalitis in Moose (Alces americanus) in Northeastern Vermont

Abstract

During fall 2010 21 moose (Alces americanus) sera collected in Northeastern Vermont were screened for eastern equine encephalitis virus (EEEV) antibodies using plaque reduction neutralization. Six (29%) were antibody positive. This is the first evidence of EEEV activity in Vermont, and the second report of EEEV antibodies in moose.

Serologic evidence suggests that moose (Alces americanus Linnaeus) are exposed to a wide variety of arboviruses endemic to North America (e.g., Trainer and Jochim 1969; Zarnke et al., 1983). However, only one study (Carstensen et al., 2007) has reported evidence that moose are infected by eastern equine encephalitis virus (EEEV). Although EEEV has been isolated or detected in all surrounding states and provinces (Morris 1988; Armstrong et al., 2008), EEEV activity has not been detected in Vermont (VT).

In 2009, we demonstrated that screening serum of free-ranging white-tailed deer (Odocoileus virginianus) was a sensitive method for detecting EEEV activity and that serosurveys of deer could be used as a tool to map EEEV activity (Mutebi et al., 2011). The VT Department of Health, the VT Agency of Agriculture, Food and Markets and the US Centers for Disease Control and Prevention (CDC) conducted serosurveys of VT white-tailed deer 6 October to 14 November 2010 and collected 489 serum samples (Berl et al., unpubl. data). Additionally, in northeastern VT, during these serosurveys, blood samples were collected from 21 harvested moose carcasses. Our objective was to screen the moose sera for EEEV antibodies.

On 16 October 2010, moose blood samples were collected from moose carcasses brought for harvest registration in Essex, Lamoille, Caledonia, and Orleans Counties (Fig. 1, Table 1). Blood samples were collected from pools in body cavities using disposable plastic pipettes. The samples were collected into 7.5 ml vacutainer tubes, stored on ice for 24–48 hr, and centrifuged (Beckman AccuSpin FR, Beckman Coulter, Inc., Brea, California, USA) at 80 × g for 15 min. to separate the serum at the VT Agency of Agriculture Laboratory. Sera were frozen at −20 C and shipped on dry ice to the CDC laboratories in Fort Collins, Colorado for antibody screening.

Figure 1.

Distribution of eastern equine encephalitis virus antibody-positive and antibody-negative moose (Alces americanus) collected in Vermont, USA, 2010.

Table 1.

Plaque reduction neutralization test (PRNT) results of eastern equine encephalitis virus antibodies for 21 positive moose (Alces americanus) sera collected in Vermont, USA, 2010. The six positive sera are highlighted in bold print.

Moose no.	Township	County	Sexa	Weight (kg)b	Age (yr)	Serum PRNT80
1	Newark	Caledonia	F	305.7	7	<10
2	Averill	Essex	F	249.5	5	<10
3	Averill	Essex	M	329.3	4	<10
4	Lewis	Essex	M	360.2	8	<10
5	Averill	Essex	M	122.9	<1	80
6	Avery’s	Essex	F	181.9	1	<10
7	Bloomfield	Essex	M	284.4	3	20
8	Brighton	Essex	M	372.9	7	<10
9	Brighton	Essex	F	174.6	1	<10
10	Brighton	Essex	M	254.9	2	<10
11	Brighton	Essex	M	286.2	2	20
12	East Haven	Essex	M	193.7	1	<10
13	Ferdinand	Essex	M	199.1	1	<10
14	Ferdinand	Essex	F	172.8	3	<10
15	Lemington	Essex	M	288.9	4	640
16	Lemington	Essex	F	136.5	4	<10
17	Brunswik	Essex	M	324.3	6	<10
18	Victory	Essex	M	290.8	3	≥2560
19	Eden	Lamoille	F	97.52	<1	160
20	Westmore	Orleans	M	236.8	1	<10
21	Morgan	Orleans	M	260.8	2	<10

^aM = male, F = female.

^bdressed weight.

Serum samples diluted 1:10 were screened for EEEV-neutralizing antibodies by plaque-reduction neutralization tests (PRNT; Beaty et al., 1995). Positive samples were retested and titrated in duplicate for confirmation. Serum samples were considered positive for EEEV antibodies if they neutralized 80% of a challenge dose of ≈100 plaque-forming units of EEE-Sindbis chimeric virus (Wang et al., 2007). To ensure that neutralization was specific to EEEV and not resulting from antibody cross-reactivity, samples with low neutralizing titers were screened for Highlands J virus antibodies. The Highlands J virus strain used for these PRNT was MW8–5AD, which was isolated from a mosquito pool in Maryland in 1968, obtained from the CDC virus Bank in Fort Collins.

Twenty-one moose serum samples were collected from four counties in northeastern VT; 17 samples (81%) were from Essex County, 1 (5%) was from Lamoille County, 1 (5%) was from Caledonia County, and 2 (10%) were from Orleans County (Fig. 1). Six (29%) samples were positive for EEEV antibodies (Table 1). Two of the positive animals were calves suggesting recent infections (Table 1). In Essex County, 5 of the 17 (28%; Table 1) moose sera had EEEV antibodies and were collected in different townships (Table 1, Fig. 1). Sample sizes from the other counties were too small for robust estimates of EEEV activity, but the single moose sample collected from Lamoille County was EEEV antibody-positive (Table 1).

The percentage of moose with serologic evidence of EEEV infection in VT (29%) is among the highest reported for wild ungulate populations (range 0.5–31%; Bigler et al., 1975; Hoff et al., 1973; Tate et al., 2005; Mutebi et al., 2011). Recently, (Lubelcyck et al., unpubl. data) detected EEEV antibodies in moose in northern Maine suggesting that moose are widely exposed to EEEV in northern New England. Carstensen et al. (2007) reported 4% EEEV antibody-positive moose sera in northwestern Minnesota suggesting that moose are exposed to EEEV in the northern Midwestern US. Moose commonly graze on hydrophytes in wooded wetlands, marshes, and swamps, which are breeding sites for the EEEV enzootic mosquito vector Culiseta (Climacura) melanura (Coquillett) and the bridge vectors Coquillettidia (Coquillettidia) perturbans (Walker), Aedes (Ochlerotatus) canadensis (Theobald), Aedes (Ochlerotatus) sollicitans (Walker), which may increase the chances of exposure to EEEV. Blood meal analysis has not detected moose DNA in engorged field-collected Cs. melanura (Molaei et al., 2006), but studies have not been conducted in areas with large moose populations such as northern New England.

Moose have home ranges of usually 20–30 km² (Leptich and Gilbert 1989; Morris 1999; Van Dyke et al., 1995); therefore, the presence of EEEV antibodies in moose populations suggests localized EEEV transmission and that EEEV is endemic in VT. Additionally, the white-tailed deer serosurveys conducted in 2010, Berl et al. (unpubl.) detected EEEV antibodies in free-ranging deer from large areas of the state suggesting widespread EEEV activity in VT. The outbreak of EEEV on an emu (Dromaius novaehollandiae) farm in Rutland County, VT, in September 2011 (Berl et al. unpubl.) provides additional documentation of EEEV activity in VT.

We thank wildlife biologists from the VT Department of Fish and Wildlife for help in collecting serum samples on youth weekend. We thank the many volunteers that helped to collect serum samples. Also, we thank students from the University of VT, Green Mountain College and Paul Smiths College and staff from the VT Department of Health; the VT Agency of Agriculture, Food and Markets; the VT Department of Forest, Parks and Recreation; the VT Center for Ecostudies and the United States Department of Agriculture, Animal, and Plant Health Inspection Service. This study was funded in part by the VT Agency of Agriculture, the VT Department of Health and the CDC.