Western Equine Encephalitis Research Articles

Medically important arboviruses of the United States and Canada

Of more than 500 arboviruses recognized worldwide, 5 were first isolated in Canada and 58 were first isolated in the United States. Six of these viruses are human pathogens: western equine encephalitis (WEE) and eastern equine encephalitis (EEE) viruses (family Togaviridae, genus Alphavirus), St. Louis encephalitis (SLE) and Powassan (POW) viruses (Flaviviridae, Flavivirus), LaCrosse (LAC) virus (Bunyaviridae, Bunyavirus), and Colorado tick fever (CTF) virus (Reoviridae, Coltivirus). Their scientific histories, geographic distributions, virology, epidemiology, vectors, vertebrate hosts, transmission, pathogenesis, clinical and differential diagnoses, control, treatment, and laboratory diagnosis are reviewed. In addition, mention is made of the Venezuelan equine encephalitis (VEE) complex viruses (family Togaviridae, genus Alphavirus), which periodically cause human and equine disease in North America. WEE, EEE, and SLE viruses are transmitted by mosquitoes between birds; POW and CTF viruses, between wild mammals by ticks; LAC virus, between small mammals by mosquitoes; and VEE viruses, between small or large mammals by mosquitoes. Human infections are tangential to the natural cycle. Such infections range from rare to focal but are relatively frequent where they occur. Epidemics of WEE, EEE, VEE, and SLE viruses have been recorded at periodic intervals, but prevalence of infections with LAC and CTF viruses typically are constant, related to the degree of exposure to infected vectors. Infections with POW virus appear to be rare. Adequate diagnostic tools are available, but treatment is mainly supportive, and greater efforts at educating the public and the medical community are suggested if infections are to be prevented.

Analysis of the Hemagglutination Activity Domains of the Venezuelan Equine Encephalomyelitis and Eastern Equine Encephalomyelitis Viruses

The hemagglutination (HA) domains of the Venezuelan equine encephalomyelitis (VEE) and the eastern equine encephalomyelitis (EEE) viruses providing the interaction of virions and red blood cells were studied with the use of a panel of 17 hemagglutination inhibition (HI) monoclonal antibodies (MAbs). A highly conserved domain (C domain) forming alphavirus-group-reactive MAbs was identified in the E2 protein of the VEE and EEE viruses. These MAbs inhibited HA of the western equine encephalomyelitis, Semliki Forest, Sindbis, Getah, Aura, Chikungunya and Pixuna viruses. The involvement of amino acid residues 59 and 232 in the formation of the C region was demonstrated by sequencing the gene encoding the E2 protein of three escape variants of the VEE virus.

A Comparison of the Nucleotide Sequences of Eastern and Western Equine Encephalomyelitis Viruses with Those of Other Alphaviruses and Related RNA Viruses

The complete nucleotide sequence of a 1982 Florida strain of eastern equine encephalomyelitis (EEE) virus, and partial sequence of the nonstructural protein genes of western equine encephalomyelitis (WEE) virus, were determined. The EEE virus genome was 11,678 nucleotides in length, excluding the cap nucleotide and poly(A) tail, and the nucleotide composition was 28% A, 24% G, 25% C, and 23% U. The organization of both EEE and WEE virus genomes was like that of other alphaviruses and included a termination codon between the nsP3 and nsP4 genes. Codon usage for 10 of 20 amino acids was nonrandom in the EEE genome, and dinucleotide CpG-containing codons were underutilized in both genomes. The slight CpG deficiency was similar to that seen in other alphaviruses and plant viruses in the alphavirus-like group, but less than that of poliovirus and yellow fever virus. This slight deficiency may reflect adaptation for replication in both CpG-deficient vertebrates, as well as insects which do not have CpG-deficient genomes. Phylogenetic analyses using nonstructural protein amino acid sequences indicated that alphaviruses evolved from a common ancestor which existed a few thousand years ago. An intercontinental introduction of an ancestral virus from the Old to New World, or vice versa, probably resulted in two main extant groups; one includes New World (EEE and Venezuelan equine encephalitis) viruses, while the other includes Old World (Sindbis, Middelburg, O'nyong-nyong, Ross River, and Semliki Forest) viruses. The position of WE E virus in the phylogenetic trees indicated that, in addition to its capsid gene (C. S. Hahn et al. (1988) Proc. Natl. Acad. Sci. USA 85, 5997-6001), WEE virus acquired its nonstructural genes from an EEE-like ancestor during recombination.

Immunogens of encephalitis viruses

The equine encephalitis viruses are members of the genus Alphavirus, in the family Togaviridae. Three main virus serogroups represented by western (WEE), eastern (EEE) and Venezuelan equine encephalitis (VEE) viruses cause epizootic and enzootic infection of horses throughout the western hemisphere. All equine encephalitis viruses are transmitted through the bite of an infected mosquito. The first equine encephalitis virus vaccines were produced by virus inactivation. Problems with inadequate inactivation, which may have caused a major epidemic/epizootic of VEE in central America and Texas in the 1970s, led to the development of a live attenuated VEE virus vaccine (TC-83) derived by cell culture passage. Inactivated vaccine are still used to prevent equine infections with WEE and EEE viruses. Alphaviruses are small single stranded, positive sense RNA viruses. The 12000 nucleotide genome is enclosed in an icosahedral nucleocapsid composed of multiple copies of the capsid (C) protein. The virion is enveloped. The membrane is modified by the insertion of heterodimers of two glycoproteins: E1 and E2. Monoclonal antibody analysis of the surface glycoproteins have provided a detailed understanding of important protective antigens. Recent studies comparing gene sequences from virulent and avirulent VEE viruses have begun to delineate mechanisms of alphavirus attenuation.

Weather factors in the prediction of western equine encephalitis epidemics in Manitoba

Cases of western equine encephalitis in horses in 1987 in western USA and Manitoba, Canada were examined by backward trajectory analysis of winds. Culex tarsalis mosquitoes infected with western equine encephalitis virus could have been carried on southerly winds from Texas and Oklahoma to northern USA and from there to Manitoba. The presence of the Polar front over North Dakota and Minnesota at the end of July would have led to the landing of Cx. tarsalis in Montana and Wisconsin and prevented further carriage into Manitoba. Temperatures in southern Texas during the winter months (average daily maximum temperatures 19.7 degrees C and higher) would have permitted continuous transmission of western equine encephalitis virus by Cx. tarsalis in this area. Weather factors involved in outbreaks from 1975-88 were analysed to see if epidemics in Manitoba (23 or more cases in horses) could be predicted. The conditions for epidemics could be defined as follows: (a) the number of cases in horses in USA was 98 or more, (b) winds were southerly with speeds 45 kmh-1 or higher, and (c) counts of Cx. tarsalis females/light trap per day were 3.2 or higher. There were 3 or fewer cases in Manitoba, when the number of cases in USA was 27 or less, even when Cx. tarsalis counts were higher than 3.2. With Cx. tarsalis counts below 3 and/or unsuitable winds, or the Polar front further south, the number of cases in Manitoba was between 0 and 17, even when the number of cases in USA was from 38-172. Without information on the extent of infection further south, the weather variables would probably be more useful in excluding the possibility of an epidemic in Manitoba than in predicting one.

Western Equine Encephalomyelitis: Virulence Markers and Their Epidemiologic Significance

Comparative studies are described on the virulence of the western equine encephalomyelitis (WEE) complex viruses for mice. Three epizootic WEE virus strains (McMillan, Cba 87, and Cba CIV 180) and five enzootic WEE complex viruses (Highlands J [HJ], Y62-33, Aura, Fort Morgan [FM], and WEE AG80-646) were examined. The neurovirulence and the neuroinvasiveness of these viruses for adult mice were established and correlated with viremia and virus replication in brain tissue. Adult mice inoculated intraperitoneally showed differential responses that corresponded to the epidemiologic attributes of WEE viruses. Viruses associated with equine epizootics were neurovirulent and neuroinvasive, whereas enzootic viruses were neither neuroinvasive nor neurovirulent. In North America, HJ virus appears to be an antigenic link with an intermediate virulence between epizootic WEE virus and the enzootic FM virus. The HJ virus has been associated with rare cases of sporadic equine and human diseases. In South America, no virus with intermediate virulence characteristics has been described. We speculate that epizootics may arise from nonpathogenic strains such as AG80-646 maintained in enzootic transmission cycles.

Circulation of eastern equine encephalitis, western equine encephalitis, Ilhéus, Maguari and Tacaiuma viruses in equines of the Brazilian Pantanal, South America.

Neutralizing antibodies to EEE (6.7%), WEE (1.2%), ILH (26.6%), MAG (28.2%) and TCM (15.7%) viruses were found in sera of 432 equines of the Brazilian Pantanal, area where undiagnosed horse deaths are frequently observed. A 4-fold rise in CF titer to EEE virus was detected in acute and convalescent sera of an encephalitis horse sacrificed in 1992. Antibodies to EEE, ILH, MAG and TCM viruses were detected in horses less than 2 years old indicating recent circulation of these viruses in the Pantanal. The evidence of recent equine encephalitis associated with rising CF titer to EEE warrants a more intensive study with attempts to isolate virus from horses with clinical manifestations of encephalitis.

Distribution of western equine encephalomyelitis virus in the alimentary tract of Culex tarsalis (Diptera: Culicidae) following natural and artificial blood meals.

The distribution of western equine encephalomyelitis (WEE) virus in the alimentary tract of Culex tarsalis (Coquillett), following blood feeding, was examined using autoradiography. Following engorgement on chicks circulating radiolabeled virus, WEE virus in all mosquitoes was concentrated at the margins of the abdominal midgut, adjacent to the epithelium. Thirty percent of mosquitoes also contained small amounts of virus in the foregut, ventral diverticulum, and/or thoracic midgut. Mosquitoes imbibing labeled virus in hanging drops containing washed blood cells and sucrose showed an altered distribution of WEE virus. Virus was not concentrated adjacent to the abdominal midgut epithelium, and all mosquitoes contained large amounts of WEE virus in most regions of the thoracic alimentary tract. These findings may help explain the increased thresholds of infection associated with artificial feeding techniques and indicate that artificial blood meals may alter the sites of initial arbovirus infection within the alimentary tract of mosquito vectors.

Effect of temperature on the transmission of western equine encephalomyelitis and St. Louis encephalitis viruses by Culex tarsalis (Diptera: Culicidae).

The extrinsic incubation rate (inverse of the time in days from infection to median transmission) of western equine encephalomyelitis (WEE) and St. Louis encephalitis (SLE) viruses by laboratory strains of Culex tarsalis Coquillett increased as a linear function of incubation temperatures from 10 to 30 degrees C. The estimated temperatures for zero transmission thresholds (intercept of the X axis) were 10.9 and 14.9 degrees C, and the number of degree days above these thresholds required for median transmission (inverse of the slope) was 67.6 and 115.2, respectively. Although the bodies of most Cx. tarsalis females remained infected and the WEE viral titer did not decrease significantly throughout the incubation periods at all temperatures, transmission rates by females incubated at 20 to 30 degrees C decreased markedly after peaking at 7-10 d after infection. In contrast, midgut escape and salivary gland infection barriers limited the transmission rates of SLE virus at all temperatures, but these rates did not decrease markedly as a function of incubation time, indicating that virus modulation did not occur. Degree-day models were used to calculate monthly changes in the duration of the extrinsic incubation period for WEE and SLE viruses in the San Joaquin and Coachella valleys based on mosquito temperatures estimated by combining nocturnal air and diurnal resting site temperatures. Temperatures in the San Joaquin Valley averaged 5 degrees C cooler than in the Coachella Valley, proportionately shortening the duration of the potential transmission season for WEE virus from 10 to 8 mo and for SLE virus from 8 to 5 mo, respectively.

Arbovirus surveillance in northern Colorado, 1987 and 1991.

Arbovirus surveillance was conducted during an epizootic of western equine encephalitis (WEE) during 1987 and during a nonepizootic year, 1991, in the same area in northern Colorado. Mosquitoes were collected in Larimer County, CO, during weeks 33-37 (10 August to 7 September) in 1987 and during weeks 26-35 (24 June to 26 August) in 1991. In total, 13,099 mosquitoes in 694 pools collected during 1987 and 8,672 mosquitoes in 242 pools collected during 1991 were tested for virus. WEE virus was isolated in both years from Culex tarsalis Coquillett and from Cx. pipiens L. in 1987. Infection rates and population levels of Cx. tarsalis were not significantly different in the 2 yr during weeks 33, 34, and 35 (12-26 August). St. Louis encephalitis virus was isolated in 1987 from Cx. tarsalis. Other viruses isolated included Hart Park, Turlock, and Jerry Slough, a variety of Jamestown Canyon virus.

Evaluation of vero cell lysate antigen for the ELISA of flaviviruses

The vero cell lysate antigen for the enzyme-linked immunosorbent assay (ELISA) of flaviviruses was evaluated for sensitivity, specificity including cross-reactions, and background by comparing with the standard ELISA. Human sera, in serial dilutions, were taken from subjects 14, 35, and 210 days postvaccination with 17D antigen. Early after injection, high sensitivity (82.9%) was shown by the cell lysate antigen method. Late after infection, high sensitivity was achieved by the standard method (96.2% and 94%), with significant difference (P = 0.0001). However, sensitivity achieved by the cell lysate antigen method was also acceptable (91.7% & 88.9%). The cell lysate antigen method showed high specificity and low cross reactivity early after infection. At 35 days postvaccination, no difference in specificity was observed between the two methods, but higher cross-reactions were observed for the standard method. This pattern continued at 210 days postvaccination, with significantly higher cross-reactions with the standard ELISA. The optical density differences by the two methods did not show significant relationship with the serial dilutions of human sera. No difference was observed in early and late infections in the background values of the negative control (Western equine encephalitis) between the two methods. The ELISA by the cell lysate antigen, within the limits of the experiments done, was found to be a good replacement for the ELISA by the standard method.

Relationship of cliff swallows, ectoparasites, and an Alphavirus in west-central Oklahoma.

Approximately 250 isolates of a newly recognized virus, related to western equine encephalitis virus (family Togaviridae, genus Alphavirus), were obtained from cimicid bugs, Oeciacus vicarius; Cliff Swallows, Hirundo pyrrhonata; and House Sparrows, Passer domesticus in a study area in west-central Oklahoma at Buggy Creek and Caddo Canyons. Antigenicity of the virus strains varied slightly from isolate to isolate. This paper summarizes the ecology of the area by describing in general the flora and fauna there.

Pathologic changes in the midgut of Culex tarsalis following infection with Western equine encephalomyelitis virus.

Midguts of two strains of the mosquito vector Culex tarsalis were examined by light and electron microscopy following infection with western equine encephalomyelitis virus. Infection of the highly susceptible Knight's Landing strain with high-titered blood meals resulted in pathologic changes in the midgut epithelium after 2-4 days of incubation; lesions included sloughing of epithelial cells into the lumen and necrosis of cells in situ. Infection of Knight's Landing strain mosquitoes with low-titered blood meals and infection of the less susceptible Fort Collins strain with high-titered blood meals did not result in a significant increase in detached luminal cells, with respect to uninfected controls. Sloughing of infected cells into the midgut lumen may contribute to modulation of the mosquito infection. Lesions in the midgut of Cx. tarsalis are inconsistent with traditional views that regarded arbovirus infections of mosquito vectors as non-pathologic. These findings demonstrate that mosquito pathology is not an oddity limited to the previously described interaction between Culiseta melanura and eastern equine encephalomyelitis virus, and suggest that alphaviruses in general may adversely affect their mosquito vectors in nature.

Experimental inoculation of three arboviruses in black-bellied whistling ducks (Dendrocygna autumnalis).

Wild-caught, immature black-bellied whistling ducks (Dendrocygna autumnalis) were inoculated with eastern equine encephalitis (EEE), St. Louis encephalitis (SLE), or western equine encephalitis (WEE) virus. Susceptibility, duration and titer of viremia, and antibody response to these arboviruses were determined. Birds from all inoculated groups became viremic. Higher virus titers occurred in the EEE group but overall mean titers were not significantly different among experimental groups. All birds inoculated with EEE and SLE viruses developed antibodies, and six of seven ducks receiving WEE virus were seropositive. All seropositive ducks had antibodies for at least 59 days, when the study was terminated. The EEE group had significantly more seropositive ducks during more days than the WEE and SLE groups. Geometric mean antibody titers were significantly smaller in the WEE group when compared to the EEE and SLE groups. Control ducks did not develop viremia or antibodies. Gross and histopathologic lesions compatible with viral encephalitis were absent in all of nine ducks necropsied. Black-bellied whistling ducks can develop low and short-term levels of viremia sufficient to infect mosquitoes, but probably cannot contribute significantly to the transmission of EEE and SLE. They may serve as good indicators of virus activity.

Transmission of western equine encephalomyelitis virus by Argentine Aedes albifasciatus (Diptera: Culicidae).

Aedes albifasciatus (Macquart) has been considered a potential vector of western equine encephalitis (WEE) virus in Argentina because it has been found naturally infected, it is susceptible to infection per os, and its distribution and feeding habits are compatible with those of an epizootic and epidemic vector. Ae. albifasciatus females collected in the vicinity of Cordoba, Argentina, were highly susceptible to WEE virus perorally (ID50 less than 0.5 Vero cell plaque-forming units) when fed on viremic chicks. Virus transmission trials were done 9 to 16 d after infection by feeding potentially infected mosquitoes individually on chicks. Among 31 mosquitoes that survived the incubation period, six refed on susceptible chicks. Virus transmission by bite was demonstrated by five of six (83%) mosquitoes. Therefore, the final piece of evidence incriminating Ae. albifasciatus as a natural vector of WEE virus has been obtained. This is the first mosquito species to be incriminated as a vector of WEE virus in South America.

Serologic survey for selected arboviruses and other potential pathogens in wildlife from Mexico.

During 1988 and 1989, a serologic survey of wildlife was conducted in northeastern Mexico to determine the presence, prevalence, and distribution of arboviruses and other selected disease agents. Eighty mammal specimens were tested. Antibodies to vesicular stomatitis-Indiana, Venezuelan equine encephalitis-Mena II, Rio Grande virus, and vesicular stomatitis-New Jersey were detected predominantly in small mammals. Deer and mouflon (Ovis musimon) had antibodies to bluetongue and epizootic hemorrhagic disease. Two species had serologic evidence of recent exposure to Francisella tularensis. A white-tailed deer (Odocoileus virginianus) had antibodies to Anaplasma marginale. All specimens tested for antibodies against Yersinia pestis and Brucella abortus were negative. Sera from 315 birds were tested for antibody against five equine encephalitis viruses and six avian pathogens. During 1988, antibodies to Venezuelan equine encephalitis-Mena II, Venezuelan equine encephalitis-TC83, St. Louis encephalitis, eastern equine encephalitis, and western equine encephalitis were detected in birds of several species. Antibodies to Pasteurella multocida and Newcastle disease virus were also detected. Birds from five species presented antibodies to Mycoplasma meleagridis. Specimens tested for M. gallisepticum, M. synoviae, and Chlamydia psittaci were negative. To the best of our knowledge, this survey represents the first serologic evidence of bluetongue, Cache Valley virus, epizootic hemorrhagic disease, Jamestown Canyon virus, vesicular stomatitis-Indiana, vesicular stomatitis-New Jersey, Rio Grande virus, and tularemia reported among wildlife in Mexico.

Mosquito and arbovirus ecology in southeastern California, 1986-1990.

Mosquito abundance and western equine encephalomyelitis (WEE) and St. Louis encephalitis (SLE) virus activity were monitored in five valleys in southeastern California from June 1986 through April 1990 to study virus overwintering and possible dissemination from south to north along geographically defined corridors. Culex tarsalis Coquillett predominated in CO2 trap collections and was the only species repeatedly infected with WEE and SLE viruses. Abundance peaked during April-May and August-October. WEE virus infections in Cx. tarsalis generally were detected after the spring peak and were followed approximately 1 mo later by seroconversions in sentinel chickens. SLE virus infections occurred later in the summer but before the fall peak in Cx. tarsalis abundance. Peak Cx. tarsalis abundance occurred when monthly temperatures averaged 25 degrees C, whereas virus infections generally were detected most frequently when temperatures exceeded 29 degrees C. Although the spring increase in Cx. tarsalis abundance occurred earlier in southern valleys, the onset of virus activity was variable among valleys and did not follow a south to north progression.

Evaluation of Domestic Pigeons as Sentinels for Detecting Arbovirus Activity in Southern California

Flocks of sentinel domestic pigeons (Columbia livia) detected increases in St. Louis encephalitis (SLE) and western equine encephalomyelitis (WEE) virus activity in southern California concurrently with flocks of sentinel chickens. However, occasional low-titered, transient seroconversions to both WEE and SLE viruses also occurred in pigeons during periods when virus activity was not detected by seroconversions in sentinel chickens, by virus isolation from Culex mosquitoes, or by human disease. Moreover, SLE virus seroconversions detected in pigeons by a hemagglutination inhibition (HI) test frequently could not be confirmed either by a plaque-reduction neutralization test (PRNT) on the same sera, or by an HI test on the next monthly serum sample from the same bird. Experimental infection studies, in which pigeons were inoculated subcutaneously with SLE (SOUE 16-84) virus, confirmed that pigeons developed low-titered and transient HI antibodies that were detectable infrequently by PRNT. In contrast, experimental infection with WEE (BFS 1703) virus produced elevated antibody responses that were detectable by HI for 8-12 weeks and by PRNT for at least 25 weeks. Pigeons infected with SLE virus rarely developed detectable viremias, whereas most birds infected with WEE virus developed viremias on postinfection day 1 that persisted for two or three days. Host-preference studies indicated that pigeons were less attractive as bait in lard can traps to host-seeking Culex mosquitoes than were chickens, and that blood-engorged Culex females collected near sentinel locations fed more frequently upon galliform than columbiform birds. Collectively, these results indicated that sentinel pigeons would not provide an adequate replacement for sentinel chickens to monitor WEE or SLE viruses, and would be a dead-end host for SLE virus.

Epidemiologia das encefalites por arbovírus na amazônia brasileira

An overview of ecological, epidemiological and clinical findings of potential arthropod-borne encephalitis viruses circulating in the Amazon Region of Brazil are discussed. These viruses are the Eastern Equine Encephalitis (EEE), Western Equine Encephalitis (WEE), St. Louis Encephalitis (SLE), Mucambo (MUC) and Pixuna (PIX). These last two are subtypes (III and IV) of Venezuelan Equine Encephalitis virus. The areas of study were the highways and projects of development, as well as places where outbreaks of human diseases caused by arboviruses had been detected. These viruses are widespread in all Amazonia, and at least four of them, EEE, WEE, SLE and MUC are pathogenic to man. EEE and WEE infections were detected by serology, while SLE and MUC by either serology and virus isolation. The PIX virus has the lowest prevalence and, it was isolated in only a few cases, one being from a laboratory infection. Wild birds are the main hosts for all these viruses, except MUC, whose major hosts are rodents. The symptoms presented by infected people were generally a mild febrile illness. Although, jaundice was observed in two individuals from whom SLE was isolated. A comparison of the clinical symptoms presented by the patients in the Amazon Region and other areas of America, especially in the USA is made. In Brazilian Amazon region epidemics have not been detected although, at least, one EEE epizootic was recorded in Bragança, Para State, in 1960. At that time, of 500 horses that were examined 61% were positive to EEE by HI and of them 8.2% died. On the other hand, SLE has caused four epizootics in a forest near Belem. Wild birds and sentinel monkeys were infected, but no human cases were reported.

An Assessment of the Biological Capacity of a Sacramento Valley Population of Aedes Melanimon to Vector Arboviruses

Daily survivorship, duration of the gonotrophic cycle, absolute abundance and season-long relative abundance were estimated for Aedes melanimon in the Sacramento Valley of California in 1987 and 1988 using mark-release-recapture (MRR) techniques and by monitoring changes in the abundance and parity rate of the native population. One objective of these studies was to determine the extent to which A. melanimon was biologically capable of serving as a horizontal arbovirus vector. Daily survivorship was estimated to be 0.90 and 0.84 in MRR studies conducted in September 1987 and August 1988, 0.89 based on changes in the parity state and abundance of the native population in August 1988 and 0.82 using summer-long parity data in 1988. Gonotrophic cycle length (GCL) was estimated to be five days in three studies. Aedes melanimon densities were estimated to be approximately 1,000,000 and 15,000 females per hectare in September 1987 and August 1988 respectively. Parous A. melanimon females were collected on each sampling occasion from April to November 1988, suggesting that A. melanimon maintained a continuous presence in the study area throughout the summer. The results of these studies suggest that A. melanimon has the potential to be an efficient horizontal vector of western equine encephalomyelitis (WEE), based on high adult survivorship, short GCL, high abundance and a continuous presence across the summer. This supports the concept of a WEE transmission cycle in the Sacramento Valley involving Ae. melanimon as an important vector. Aedes melanimon also can be an efficient horizontal vector of California encephalitis virus (CE), though the importance of horizontal transmission to the maintenance of CE virus is unclear.