EEE Bridge Vector Control
Jeffrey S. Brown
Centers for Disease Control (CDC) and Prevention has published Guidelines
for Arbovirus Surveillance in the Unites States (April 1993) to standardize
the surveillance for mosquito borne viral encephalitis. These guidelines
emphasize predictive, proactive and efficient methods of surveillance whenever
possible (Moore et al. 1993). The national trend in mosquito control has been
to decentralize arbovirus surveillance programs from federally funded state
programs to local county programs. The operational emphasis at the local level
has been placed on incorporating Integrated Pest Management (IPM) practices into
mosquito control operations. This can be difficult to achieve because of the
technical, financial and political constraints placed on mosquito control
activities. Operational mosquito control programs are tasked with managing
mosquito populations to minimize the risk of both nuisance and public health
The CDC defines surveillance as “the organized
monitoring of levels of virus activity, vector populations, infections in
vertebrate hosts, human cases, weather and other factors to detect or predict
changes in the transmission dynamics of arboviruses.” They further suggest “a
sound (arbovirus) surveillance program requires a thorough understanding of the
biology, ecology and interactions of the vertebrate and mosquito hosts (Moore
et al. 1993). Very few local mosquito programs have the financial resources,
laboratory facilities or technical expertise to address the dynamics of
arboviruses. Most focus their energies on managing mosquito vector
populations. Local programs are often a key source for information regarding
historical mosquito populations. Control measures are typically initiated when
a particular predictor exceeds the action threshold (usually determined from
historical data and experience) (Moore et al. 1993). The CDC suggests that
different measures or predictors for epidemic arboviral transmission are
effective at different times of the year. The earliest useful predictors are climatological
factors that influence the size of the early mosquito population. Midseason
predictors usually consist of population estimates of vectors and vertebrate
hosts (especially the young of the year), and evidence of early virus
transmission in the natural cycle. The likelihood of an outbreak is estimated
by comparing current vector and vertebrate host population densities and the
age structures with long-term averages. Late season predictors consist of
evidence of virus spillover to sentinel bird/chicken flocks, epidemic/epizootic
vectors and domestic animals. The likelihood of transmission to humans or
domestic animals becomes more accurate as the virus begins to circulate in
vector and vertebrate host populations (Moore et al. 1993). Local
programs without arbovirus testing capabilities are forced to rely solely on climatological
and mosquito surveillance data.
involves using weekly mosquito surveillance and monitoring information to
create a timeline that can be used to project the potential for increased
arbovirus activity. The timeline will help to identify the best opportunities
for collecting, pooling and testing mosquitoes that may be carrying Eastern
Equine Encephalitis (EEE). Because mosquitoes reside in a constantly changing
environment, recognizing the environmental trends that limit or enhance
mosquito production is essential for successful control operations. Mosquito
production is regulated seasonally by climactic variables, temperature, day
length, groundwater levels and rainfall and other variables such as predators,
suitable oviposition habitat, effects of ongoing control efforts and the
reproductive capacity of the mosquito species in question. Mosquito
surveillance and monitoring are essential for successful control activities.
The timeline (Diagram 1) anticipates arbovirus activity
for EEE based on maintenance vector activity of Culiseta melanura and
associated bridge vector activity. This is not a scientific model per say, but
a surveillance strategy that encourages weekly surveillance and monitoring of
mosquito species and their associated population trends to assess both nuisance
mosquito populations and to project the potential for arbovirus activity. This
can ultimately enhance nuisance mosquito control operations and potentially
minimize the risk of arbovirus transmission
The timeline promotes mosquito and environmental
monitoring to identify potential EEE virus-scenarios. For example, by using
larval dipping and light trapping to document the adult emergence of Cs.
melanura, programs can track the sequence of events that must occur for EEE
transmission. The timeline considers 6 variables that must be in place:
EEE virus activity
Presence of reservoirs in the
Maintenance vector (abundance and
Bridge vector (abundance and
Susceptible mammal population
Suitable environmental conditions
Day 1 EEE virus
Day 1-4 Wild bird
infected, virus amplifies in bird
Day 5-7 Wild bird
Day 5-7 Cs. melanura
bites infected bird
Day 5-17 EEE virus
amplifies in Cs. melanura
Day 18-25 Cs. melanura
capable of vectoring virus
Day 18-20 Cs. melanura
infects a wild bird or sentinel chicken
Day 20-23 Bird host becomes
Day 20-25 Bridge vector
bites infected bird
Day 21-32 Virus amplifies in
competent bridge vector
Day 23-36 Local mosquito
control program could be notified of seroconversion
Day 32-42 Bridge vector
capable of virus transmission to mammals
Day 34-42 Mammal infection
Day 34-42 Virus incubation
period in mammal
Unknown Virus diagnosed by
physician or veterinarian
Unknown Local mosquito
control program is notified
In southeastern North
Carolina, the typical Cs. melanura
adult female lives about 25 days (based on weekly light trap data). The EEE
virus is only infectious in the mosquito for the last week to ten days of its
life because of virus amplification considerations. Some of the questions to
consider when evaluating the EEE potential are listed below.
Is there a new emergence of
competent bridge vectors within the projected 7-day window that Cs. melanura
What is the overlap in habitats
of the bridge vector population and the infectious Cs. melanura?
Is it early in the arbovirus
How significant are the
maintenance and bridge vector populations?
Are environmental conditions
favorable for future populations of Cs. melanura and bridge vector
Interpreting this information can help to identify
the risk to the human population and weekly monitoring allows this risk to be
assessed in real time. Therefore, the timeline is proactive and not of
reactive. In order to apply the timeline in an operational setting, some
fundamental information about EEE and how it interacts with maintenance vectors,
bridge vectors, wild birds, sentinel chickens, horses, humans and the
environment should be reviewed.
Eastern Equine Encephalomyelitis (EEE)
Eastern Equine Encephalitis (EEE) virus is an
arthropod borne virus belonging to the virus family Togaviridae, genus Alphavirus.
The initial isolate of EEE was recovered in 1933 from the brain of a dead horse
in New Jersey by Giltner and Shahan (Eldridge and Edman 2000).
The first mosquito isolate of EEE virus was obtained from Coquillettidia
perturbans in 1948. The first isolation of EEE in the enzootic mosquito
vector, Culiseta melanura, was reported by Chamberlain et al. in
1951 (Scott and Weaver 1988).
America, the principal area of
transmission is the Atlantic Coast from New
Hampshire to Florida and
along the Gulf States to Texas (Scott and Weaver 1988). In the northern part of
this virus range, both human and equine cases occur between July and October.
In Florida, human cases occur throughout the year, but are concentrated between
May and August. The virus or disease is often recognized in wild birds, penned
pheasants, or equines prior to human involvement (Morris 1988). Epidemics
typically occur during August and September and are often preceded (from 2
weeks to 2 months) by epizootics among game birds or
horses (Scott and Weaver 1988). The seasonal changes in Cs. melanura
biology and its relationship to EEE virus transmission vary with geographic
location and its associated climate. In Florida, for example, transmission occurs throughout the
year but peaks during the summer months (Bigler et al. 1976; Wilson et al.
1986)*. Conversely, in temperate areas like Maryland, there is a distinct
transmission season. Virus is not detected until July and can remain active in
the mosquito-bird cycle until the first heavy frost in November or December.
Most virus isolations in Maryland have been recovered in late August through
October (Williams et al. 1972, 1974; Watt et al. 1987; Scott et al. 1987)*.
There seems to be no clear-cut relationship between
epidemics and any known environmental factors. It is likely that a complex of
environmental conditions must simultaneously impact several parameters, such as
vertebrate-host population density, brood size and nutritional status, vector
population density and longevity, and winter survival of both vectors and vertebrate
hosts (Centers for Disease Control 1996). The isolation of EEE from mosquitoes
is very common during epizootics but much less common during inter-epizootic
periods (Morris 1992).
EEE and Culiseta
It is now generally accepted that in North America
EEE is transmitted among passerine birds primarily, if not solely, by Culiseta
melanura in freshwater, forested swamp habitats. (Scott and Weaver 1988).
The disease is rural in distribution and most cases are associated with wooded
areas adjacent to swamps and marshes (Morris 1988).
EEE epidemiology typically concludes that
Cs. melanura is the principal endemic vector.
Wild birds, primarily passerines,
serve as the amplifying hosts.
Epizootics begin in swamps and
move locally outward through viremic birds.
In some cases, migrating birds
introduce the virus into distant “clean” ecosystems.
Humans and equines are dead-end
There is a single most important
epidemic or epizootic vector, non-avian vertebrates are potential but unproven
The over-wintering mechanism is
unknown or an over-wintering mechanism is not required (Morris 1988).
is a swamp mosquito, probably limited in its flight range and nocturnal or
crepuscular habitat. Apparently increased humidity and temperature are
critical factors in determining its range of distribution, population density,
and seasonal prevalence. Developmental rates in nature are unpredictable and
are subject to such variations imposed by weather factors and fluctuations in water
level. In the southern states, breeding may be continual, although retarded
during the winter months. In the northern states, development is interrupted
during the winter; larvae may sustain themselves in protected habitats and
complete their development the following spring. The number of annual
generations ranges from a few in the northern states to several in the south (Siverly
and Schoof 1962).
Population densities of this mosquito are highest
deep in the interior of the swamp habitat, which is also where most enzootic
EEE virus transmission occurs (Williams et al. 1972, 1974). However,
mosquitoes will leave the swamp breeding sites and move to drier upland,
forested habitats, especially during the late summer and fall (Joseph and Bickley
1969; Saugstad et al. 1972; Muul et al. 1975; Morris et al. 1976, 1980; Nasci
and Edman 1984). It is believed that in these upland locations, blood feeding
occurs after engorging female mosquitoes return to the swamp to oviposit
(Morris et al., 1980; Nasci and Edman, 1984).*
The complexity of the swamp ecosystem is increased by
the uneven upbuilding of the swamp floor. Because of the high water table, the
substrate is habitually saturated so that root systems do not penetrate deeply
into the organic soil. Instead, they grow near the surface, spreading widely.
The resulting root buttress brings about a rapid and unequal elevation of the
land, with levels around trees as much as 1 m above the general swamp level.
Thus, the built-up areas associated with tree bases are also well above water
level, while the intervening spaces are be below it. As the water level
recedes, the exposed muck becomes subject to invasion by mosses, liverworts,
and swamp annuals. Tussocks of ferns, uprooted trees and slowly decomposing
fallen trunks provide a second series of rises and falls between the trees,
making foot travel hazardous and slow. This uneven topography, when coupled
with differences in
moisture, temperature, illumination and wind
exposure, creates a wide diversity of niches in the swamp forest environment.
Which microhabitats favor the development of Cs. melanura and influence
the epizootiology of EEE are yet unidentified. The relationship between soil
types and land use has significant impact on the present and future
determinations of the human population at risk to EEE (Morris et al. 1980).
Various observations have been made on the
development of Culiseta melanura. Average periods of development are as
follows: eggs-2 days, first instar larvae-3 days, second instar larvae-2 days,
third instar larvae-2 days, fourth instar larvae-3 days, and pupa-2 days
(Joseph and Bickley 1969). The life span of the adult female includes
emergence, feeding (survival), resting, mating, and oviposition (Siverly and Schoof
1962). These mosquitoes breed in swarms above larval sites (Hayes 1958)*.
Larvae develop in cool, acidic water that is shaded and often retained in
depressions at the base of tree trunks, stumps or uprooted trees (Scott and
Weaver 1988). Cs. melanura breeding and oviposition is usually
concentrated at the edge of the swamps but also occurs in the swamp interior
(Morris 1988). Larvae are most often found in sphagnum bogs, cedar swamps,
or maple swamps (Scott and Weaver 1988).
In temperate regions, Cs. melanura populations
have two or three major peaks of adult abundance, typically one each during
May-June, July-August, and August-October (Morris et al. 1976, 1980; Pierson
and Morris 1982; Nasci and Edman 1984; Watts et al. 1987; Scott et al. 1987)*.
Host-seeking activity, and presumably blood-feeding, is greatest during the
first 2 hours after sunset and then continues at a low but constant level until
sunrise (Hayes 1962b, Nasci and Edman 1981a)*. During sunrise, there is an
increase in activity that is believed to be more closely associated with
location of suitable daytime resting sites than with location of hosts on which
to blood-feed (Nasci and Edman 1981a)*. This mosquito does not usually feed
diurnally. Results from analyses of Cs. melanura distribution within
the forest stratum vary from equal activity at all heights tested to a flight
preference to the canopy (Love and Smith 1958; Bast and Rhen 1963; Main et al.
1966; Joseph and Bickley 1969; Nasci and Edman 1981b)*
and Weaver 1988).
Finally, the longevity of mosquito
encephalitis vectors is of special interest since it has been demonstrated that
once mosquitoes became infective, EEE virus persisted as long as they remained
alive (Davis 1940)*. Cs. melanura appears to have a survival of 3.5 months
under insectary conditions (Love and Goodwin 1961)*. Studies of eastern (EEE)
and western (WEE) equine encephalitis in and around swamplands have
demonstrated that densities of Culiseta melanura are higher within
swamps than the densities found at river and upland boarders of swamps
(Williams et al. 1971a, Saugstad et al. 1972)**, that the largest numbers of
mosquitoes infected with these viruses also occur inside swamps (Williams et
al. 1972)**, and that infection of sentinel birds may occur at twice the rate
within swamps compared to sentinel infection at localities immediately outside
the swamps (Hayes et al. 1960, Dalrymple et al. 1972)**.
The greater number of mosquitoes at the perimeter
reflects the true distribution resulting from an edge effect (Morris et al.
1980). Parity data indicates that Cs. melanura remains in the swamp
when populations are low or during adverse weather conditions but, during
periods of high populations or warm, wet weather, older females, and presumably
older males, spread at least 2 km from the nearest known Cs. melanura
breeding area (Morris et al. 1980). During EEE transmission seasons, Cs.
melanura populations at the perimeter have the greatest vector potential,
but as late season populations increases, vector potential is relatively
greater more distant from the swamp (Morris et al. 1980).
Dalrymple et al. (1972) presented evidence indicating
that the fresh water swamp habitat is the enzootic focus for WEE and EEE in the
eastern U.S. and that under normal weather conditions these
viruses are largely restricted to this habitat. Adult mosquito population
densities are closely linked to environmental conditions and densities are
highest during years with high rainfall. Associations between EEE outbreaks
and excessive rainfall have been observed when there is unusually heavy
rainfall during the summer of the outbreak and during the autumn months of the
preceding year (Hays and Hess 1964). The relative infrequency of EEE or WEE
infection in man in the coastal states seems attributable to the infrequency of
endemic vector, Cs. melanura, feeding on man,
and its greatest densities occurring in swampy habitats away from man. Cs.
melanura seems to be important in endemic transmission because it is ornithophilic,
multivoltine and abundant throughout the summer. Its abundance is attributable
to its unusual breeding habits and because its larvae are able to survive in
the saturated root mat when the water table declines, leaving other breeding
sites dry (Muul et al. 1975). Late season dispersal of Cs. melanura
could carry virus from the swamp to outlying areas and thereby initiate
outbreaks in non-avian vertebrates (Morris et al. 1980). Epidemics of disease
produced by EEE and WEE generally occur during late summer and fall when wild
birds are at their maximum population density and are most mobile. It would
appear logical that these vertebrates could largely be responsible for the
maintenance of annual virus enzootics (Dalrymple et al. 1972).
EEE and Bridge Vectors
A major question in the ecology of EEE is the
identity of the bridging vectors that transfer the virus from the enzootic
cycle to humans and equines. A variety of species serve as vectors, depending
on time of year, environmental conditions, geographic location and population
dynamics (Moore et al. 1993). Among the factors that determine the vector
potential of various mosquito species is their distribution in time and space.
Mosquito larvae are confined to aquatic habitats which can be roughly
correlated with patterns of vegetation, especially those patterns which are
produced as a result of edaphic factors (influenced by the soil rather than the
climate). Adult mosquitoes are more mobile and their distribution in space may
or may not conform to the distribution of the larval stages. Furthermore,
adult distribution varies with time and adult activity cycles. If suitable
larval habitats are present in one type of area, but suitable hosts for adults
are scarce or absent, adult females must migrate from their breeding areas to
other areas in order to obtain the blood necessary for reproduction (Saugstad
et al. 1972).
The endemicity of EEE may vary in intensity from year
to year depending on the size of the population of Cs. melanura, the
timing of virus introduction during the population
growth phase of the endemic vector (late
introductions may not allow sufficient time for massive replication and
proliferation), and the availability of susceptible hosts, especially
non-immunes. An epidemic may occur depending on the population size of the
endemic vector, and the intensity of the endemic transmission cycle,
particularly in the vicinity of human habitations (Muul et al.1975). A
mechanism for “spillover” of virus from the swamp to areas inhabited by man and
domestic animals is probably dependant upon unusually large populations of
“invader” species entering and leaving the swamp during periods of abnormally
high precipitation and temperature (Saugstad et al. 1972). It is important to
emphasize that there is no single epizootic vector species for all foci
or for all hosts within a focus (Morris 1988).
Mosquito survival is one of the most important factors
used to determine the extent in which a mosquito participates in arbovirus
transmission. Mosquito survival affects the vectorial capacity of the mosquito
population, the rate of mosquitoes biting on man and the duration of the virus
extrinsic incubation (EI) period in the mosquito. Mosquito biting rate,
mosquito population size, and the EI period of viruses are also basic
considerations in any analysis of arbovirus transmission (Scott and Weaver
1988). Extrinsic incubation in mosquitoes requires that susceptible mosquitoes
must locate and blood feed on a viremic vertebrate host. These mosquitoes must
then survive an extrinsic incubation period. During which time the virus
replicates in posterior midget epithelial cells, disseminates through hemolymph
to salivary glands, replicates in salivary acinar cells, and is released into
the salivary matrix. During re-feeding, infected mosquitoes transmit virus via
their saliva to susceptible hosts, which may become viremic and infect
additional mosquitoes that imbibe their blood (Scott and Weaver, 1988).
It is important to emphasize that mosquito population
density is not, in itself an indicator of either detectable endemic or epidemic
EEE activity (Morris 1988). For example, Ochlerotatus sollicitans is a
brooded mosquito and in brooded mosquitoes, high populations are usually an
indication of a fresh emergence. Annoyance can be considerable but the adults
are mainly nulliparous, thus, the vector potential is low.
Over time, annoyance declines but the adults become parous
and the vector potential of the biting population increases. The technique of
evaluating vector potential by measuring the number of parous mosquitoes coming
to bite yields the index that New Jersey uses to institute control for the prevention of
human disease. When EEE is known to be circulating in Cs. melanura,
chemical control is directed toward “old” populations of Oc. sollicitans.
This results in spot treatments in designated areas of the state rather than
broad scale chemical coverage during emergency periods (Crans and McCuiston,
In some foci, endemic passerine birds and mosquitoes
are monitored for endemic EEE activity (Crans 1985, Howard 1985)*. Virus
isolations from mosquitoes other than Cs. melanura signal possible
epidemic activity and elude appropriate warnings and recommendations for
increased mosquito control activity (Crans 1985, Maxfield 1982, Howard 1985)*.
Presently these techniques are time consuming and costly. Often the turn
around time from the field to laboratory results is too long for implementing
effective EEE preventative mosquito control.
Komar et al. (1999) permitted Oc. triseriatus,
Ae. aegypti and Ae. albopictus to feed on starlings 2 days after
they were inoculated with virus. Engorged females were incubated for 10 days
and then individually tested for virus by plaque assay. Specimens of all three
species became infected. Although some mosquito species might be involved in
epidemic transmission more often than others, the identification of the
epidemic vectors of EEE virus has been difficult because no single species is
consistently associated with the transmission of virus to horses or humans.
Depending on time of year environmental conditions, geographic location, or
dynamics of respective mosquito populations, various species probably serve as
epidemic vectors (Scott and Weaver 1988).
Mosquito arbovirus collection and pooling for EEE in North Carolina
is conducted by the Department of Environment and Natural Resources’ Public
Health Pest Management Section. This section works closely with county health
officials and operational
mosquito programs to monitor and respond to EEE
activity across the state. They provide technical assistance and facilitate
laboratory testing for arthropod vectors and the diseases they transmit.
The potential EEE bridge vector species in North
Carolina include Aedes albopictus, Aedes vexans, Anopheles crucians,
Anopheles punctipennis, Anopheles quadrimaculatus s.l., Coquillettidia
perturbans, Culex erraticus, Culex pipiens complex, Culex restuans, Culex
salinarius, Ochlerotatus atlanticus/tormentor, Ochlerotatus c. canadensis,
Ochlerotatus infirmatus, Ochlerotatus sollicitans, Ochlerotatus triseriatus,
and Psorophora columbiae. Aedes vexans, Culex restuans, Culex
salinarius, Culiseta melanura, Ochlerotatus atlanticus/tormentor, Ochlerotatus
triseriatus and Psorophora columbiae have tested positive for EEE in
North Carolina (Harrison 2001).
EEE and Wild Birds
As stated earlier, in North America, EEE is
transmitted among passerine birds primarily by Cs. melanura. Blood meal
analysis of engorged Cs. melanura show that this ornithophilic mosquito
feeds almost exclusively on passerines. Nestlings or young birds that have
just left the nest are considered the most important vertebrate hosts for EEE
because they are the most susceptible to infection. Their viremia has greater
magnitude and duration, and they are behaviorally less defensive than adult
birds toward host seeking mosquitoes (Dalrymple et al. 1972; Edman and Scott
1987; Scott et al. 1988a)*
Crans et al. (1994) discussed the cryptic cycle of
EEE that begins with recrudescence of latent virus in previously infected AHY
(after hatch year) birds. In the early spring, reactivation of latent virus
may result from changes in the physiological state of local birds (i.e., summer
residents, residents) related to factors such as stress of migration,
establishment of territory, or other breeding activities. Epiornitic (bird to
bird) cycling requires an influx of newly emerged Cs. melanura. If
recrudescing virus is acquired by Cs. melanura females that have already
completed one gonotrophic cycle, it is unlikely
these females will live long enough for the virus to
replicate and be transmitted to other hosts.
In some years, the virus can become epiornitic by
early July, when conditions are favorable for Cs. melanura. If vector
populations are limited by unfavorable conditions during that critical period,
the virus cycle is either postponed until the August emergence or eliminated
entirely (Crans et al. 1994). Birds that frequent the deep swamp are at
greater risk of infection with EEE virus than wild birds at the river and
upland boarders of the swamp. Similarly, sentinel birds located deep in the
swamp have been found to acquire infections more readily than sentinels located
on its borders (Williams et al. 1972, Dalrymple et al. 1972)*. Thus, the
innermost swamp appears to be a central focus for the transmission of EEE
virus. It is the optimum larval habitat for Cs. melanura, and the
highest densities of adult female Cs. melanura mosquitoes occur there.
Significant movements of Cs. melanura occur between the swamp and
outlying areas (Saugstad et al. 1972)*, perhaps introducing viruses to areas
outside of the swamp. Also, viremic birds may depart the deep swamp during
migration or at other times and convey viruses to new areas.
Characteristic symptoms of EEE infection in birds
begins with fever, weakness, inactivity, ruffed plumage, or stupor. Later,
ataxia, trembling and paralysis may ensue (Scott and Weaver 1988). Komar et al
(1999) suggest EEE viremia in starlings remains sufficiently intense to infect
mosquitoes for three days, compared to just one day for robins. Although
starlings sustained a viremia intense enough to infect mosquitoes for at least
two days, viremias were sustained at this level in other birds for only one
County mosquito programs in North Carolina
do not have the resources to conduct wild bird sampling as a surveillance tool
for monitoring EEE. State personnel conduct wild bird sampling (using mist
nets) on a very limited scale to address specific arbovirus questions.
Historically, North Carolina has relied heavily on their sentinel chicken program
as the primary surveillance tool for monitoring EEE activity.
EEE and Sentinel Chickens
most widely used sentinel animal for arbovirus surveillance is the domestic
chicken. Chickens are attractive hosts for Culex mosquito vectors and
are susceptible to arboviral infections. Additionally, they can tolerate these
infections and produce readily identifiable antibodies. Chickens are hardy and
are easily handled and bled. They are relatively inexpensive to maintain on
farms or in urban-suburban locations by residents or health officials. Eggs
laid by the birds may provide an added incentive and help defray any costs of
maintaining the birds (Moore et al. 1993).
spring, monitoring sites are stocked with 10 to 30 pre-tested, non-immune, individually
banded birds approximately 6 to 8 weeks old. Dispersing smaller groups
throughout an “at risk” area yields a more representative estimate of arbovirus
activity. The choice of monitoring sites should be based on historical records
of virus activity, vector resting sites or flight corridors, and the likelihood
of virus transmission rather than convenience. The chickens are kept in
standard sentinel sheds or similar structures (Moore et al. 1993).
chickens are bled biweekly or monthly from the wing, the jugular vein, or from
the heart throughout the transmission season. Seroconversions may occur 2-3
weeks before the detection of equine or human cases of EEE. If the intent of
surveillance is to monitor season-long transmission, birds that seroconvert are
replaced by non-immune birds, preferably of the same age (Moore et al. 1993).
uses sentinel flocks consisting of 5 white leghorn laying hens per flock to
detect virus EEE activity in specific endemic areas. Although the flocks do
not indicate the degree or distribution of the EEE virus, they do provide
valuable data about current EEE activity. Because the flocks are bled on rigid
schedules, the approximate date of infection can be projected when a bird serocoverts.
Comparing this result against weekly mosquito light trap data provides valuable
information about the maintenance and bridge vector populations and their
potential for disease transmission. Consecutive seroconversions from a
sentinel flock location and elevated mosquito light trap counts generally
indicates a need for increased mosquito control measures.
Carolina has maintained sentinel chickens for EEE since 1984. Their
operational response to a seropositive bird in a sentinel flock is considered
to be a focal in nature. Typically spot spraying around the flock and its
associated mosquito habitat is appropriate. Additionally, local health
departments may issue press releases recommending mosquito avoidance practices
and the use of personal protection measures such as mosquito repellants.
Surveillance for equine cases in areas with
susceptible horse populations may provide the most practical and sensitive tool
for the recognition of a potential public health problem caused by EEE viruses.
This is especially true in areas that lack the resources to monitor virus
activity in birds and mosquitoes. As a result of their high field exposure,
horses are subject to high vector attack rates (Moore et al. 1993). A survey
of 67 equine deaths due to EEE in the US in 1971 showed that the most common signs of illness
were depression, fever above 103o F, ataxia, paralysis, anorexia and
stupor. Other symptoms of note include irregular gate, grinding of teeth, lack
of coordination, circling, staggering, recumbency and hyper-excitability.
Death typically occurs in 1 to 3 days (Morris 1988) and the mortality rate is
80 to 90% (Eldridge and Edman 2000). Most EEE endemic states have a diagnostic
service for veterinarians and physicians to submit blood and organ samples from
suspected EEE cases. Reporting is required in some states and voluntary in
others. These systems are inappropriate for early warning since the turnaround
time for most is too long (Morris 1988). Limitations to using equines include
their vaccination status, movement into and out of the surveillance area, and
lack of prompt reporting of morbidity by attending veterinarians (Moore et. al
al. (2002) reported a 14 month-old gelding quarter horse receiving routine vaccination
for EEE and becoming symptomatic with EEE one week later. This horse case
occurred in California, which is outside the known geographic range of EEE.
Although the source of the EEE infection cannot be conclusively determined, it
may suggest a possible incubation period for the EEE virus before clinical
EEE activity is monitored using passive surveillance for humans and horses. North
Carolina averages 1 to 2 human cases and 10 to15 horse cases of EEE a year.
Passive veterinary surveillance provides county programs in North Carolina with
information about positive horse cases of EEE within their jurisdictions.
Usually, by the time the county receives the laboratory test results it is too
late to implement effective control measures with respect to the mosquito
population causing the initial infection. Responding to a confirmed horse case
of EEE does allow the program to determine if the environmental conditions are
favorable for continued virus transmission. Documenting and mapping the
mosquito production habitats in the area around a verified EEE horse case may
provide information about the potential bridge vector species involved in
future EEE transmission in that area.
Human cases of EEE can offer important insight into
the dynamics of EEE and mosquito populations. An understanding of the etiology
of EEE in humans can assist in determining the initial time of the EEE
infection. It takes 4 to 10 days after a bite for symptoms to develop (Centers
for Disease Control 1996). The illness can be systemic or encephalitic. A
systemic infection is abrupt and characterized by malaise, arthralgia (joint
pain) and myalgia (muscle pain). The patient experiences chills and severe
muscular shaking which may last for a few days. Temperatures reach 100oF
to 104oF and the illness can last 1 to 2 weeks. There is no central
nervous system involvement and recovery is complete (Morris 1988).
The encephalitic form of the disease is characterized
by an abrupt onset in infants, whereas in older children and adults onset of
active encephalitis typically occurs after a few days of indisposition.
Symptoms include fever from 102oF to 106oF, irritability,
restlessness, drowsiness, anorexia, vomiting, diarrhea, headache, cyanosis
(blue skin color due to lack of oxygen), convulsions and coma. Death occurs 2
to 10 days after being bitten (Morris 1988).
During epidemics, the initial mortality approximates
70%. Nearly all that recover have some degree of progressive mental and/or
physical disabilities. In one study, the initial mortality was 74%, but 9
years post-infection total mortality reached 90%, with only a 3% complete
recovery rate. People with the greatest risk of developing EEE are children
under 15 and adults over 55. These age groups make up 70 to 90% of the cases
in any given outbreak (Morris 1988).
The public health response for mosquito transmitted
disease is initiated when there is confirmation of increased arbovirus activity
in an area. At the local level, confirmation is received after a human or
veterinary case is identified and reported to the State Health Department.
This type of surveillance is often equated to closing the barn door after the
horse is already out. The impact of prevention or control measures on the
course of a potential epidemic is diminished by even the smallest delay. By
the time human cases are confirmed (a very accurate predictor), the epidemic
may be waning of its own accord and control measures may have little impact
(Moore et al. 1993).
Most of the
human cases of EEE in North Carolina occur in areas of the state that do not
have strong county operational programs. Incorporating IPM strategies with
mosquito surveillance and monitoring is paramount to protecting the health of the
citizens of North Carolina.
Vectorborne disease surveillance systems seldom
incorporate weather data, partly because of the lack of understanding of the
relationship between weather patterns and disease outbreaks (Eldridge and Edman
2000). The effects of precipitation with regard to mosquito production cannot
be understated. Letson evaluated rainfall patterns in states and locales where
human EEE occurred between 1983 and 1989. He found a significant association
between the occurrence of human cases and excess rainfall in the years when the
cases occurred. The association was stronger with data from local weather
stations than those from statewide rainfall averages (Centers for Disease
Weather Service maintains stations that collect environmental variables such as
daily precipitation, minimum daily temperature, maximum daily temperature,
average daily temperature and wind speed and direction. The average annual
precipitation for southeast North Carolina is 54.3 inches. The average high
temperature is 73.8oF, the average low temperature is 53.0oF
and the average mean is 63.4oF. The weather data for this project
was collected from the Wilmington Weather Forecast office (National Weather
Service). The Wilmington office monitors Bladen, Brunswick, Columbus, New
Hanover, Pender and Robeson Counties in southeastern North Carolina.
Documenting daily precipitation and temperature
within a defined control area using a standardized weekly format that can be
compared to weekly light trap data is essential to reconciling mosquito
production over time, within and across habitat types. Creating and
maintaining a long-term weather monitoring database at the local level can help
to determine the relationship between weather patterns, Cs. melanura
population densities and potential EEE virus amplification (Centers for Disease
environmental variables that can influence mosquito production within the
system throughout the year should be identified. Each habitat type has very
specific environmental influences that affect mosquito production over time.
Each new site should be monitored and compared against existing sites with
similar designations throughout the year. The goal is to identify similar
seasonal trends for mosquito production within each habitat type and to monitor
the effects of fluctuating environmental variables affecting each habitat type.
Pooling Projections for 2000, 2001and 2002
In 1999, this timeline was developed to address EEE
potential at the local mosquito program level. The purpose was to combine and
extrapolate weekly mosquito surveillance and monitoring data across multiple
counties (Map 1) to project the potential
for EEE activity across local and regional mosquito populations. Brunswick
County Mosquito Control has been monitoring Cs. melanura emergence since
1990 using a New Jersey light trap. Using their weekly surveillance data as
the foundation (Graphs 1, 2,
and 3), the EEE timeline was used to project
mosquito pooling dates in 2000, 2001 and 2002. When a pooling date was
selected, each participating county placed CDC miniature light traps baited
with CO2 near their sentinel flocks. These sites were selected
because of their proven history of EEE activity. The light traps were placed
at least ten feet into the tree line edge between the sentinel flock and the
mosquito habitat the flock monitors. Counties with extra resources placed
additional traps at other potential EEE sites. The additional trap sites were
chosen based on a history of Cs. melanura activity from previous light
2000 - September 26 Pooling (Week 39)
Using the EEE timeline, the 2000 pooling date was
selected based on Brunswick County’s weekly light trap data collected during
Week 35 (August 28-September 3, 2000) (Graph 1).
The goal of this pooling effort was to project a pooling date when the bridge
vector species could transmit the EEE virus to humans. Cs. melanura
populations were peaking during the week of August 28. Based on the previous
week’s trap data, it was evident that this brood of Cs. melanura emerged
the week of August 21 (Week 34). Using the timeline and counting August 27
(Week 34) as day 1, 29 days were added and a pooling date was projected
for September 24 (Week 38) in hopes of collecting bridge vector
mosquito species. Note that any date between timeline (TL) day 29 and day 35
could be used as the pooling date. A pooling date of September 26 (Week
39) was chosen and the participating counties were contacted. The county
mosquito programs participating in this multi-county effort included Brunswick,
Bladen, New Hanover, and Pender Counties.
2001 - October 3 Pooling (Week 40)
Following the strategy developed from the 2000
season, Cs. melanura populations were closely monitored at the Brunswick County trap
site (Graph 2). Week 35 (Aug 27-Sept 2) indicated
an increase in the Cs. melanura population from 7 in Week 34 to 26 in
Week 35. A pooling date of Sept 20 (Week 38, TL day 25) was
projected. The goal of this pooling was to assess the Cs. melanura
populations between the woodland pool habitat in Brunswick County and
the flood plain habitat in New Hanover County. It was noted that another brood of Cs. melanura
emerged during Week 38 (Sept 17-23). On September 20, a multi-county pooling
date of October 3 (Week 40) was projected. This pooling would have two
objectives, to collect EEE positive bridge vectors from the initial brood of Cs.
melanura (TL day 38) and to collect EEE positive Cs. melanura from
the second brood of Cs. melanura (TL day 17). The mosquito programs
participating in this multi-county effort included Brunswick,
Bladen, Columbus Onslow, New Hanover, and Pender Counties.
2002 - October 9 Pooling (Week 41)
Following the previous strategy, Cs. melanura
populations were closely monitored at the Brunswick County trap site (Graph 3). Week
37 (Sept 11) indicated an increase in the Cs. melanura
population from 3 in Week 36 to 65 in Week 37. Daily collections of the New Jersey
trap identified an increase in Cs. melanura populations on Sept 11 (Week
37, TL day 6). A pooling date of Oct 8 (Week 41, TL day 33) was projected. The
programs participating in this multi-county effort included Brunswick,
Bladen, Columbus, Onslow, New Hanover, and Pender Counties.
Mosquito Arbovirus Pooling Results 2000, 2001 and 2002
Five mosquito pools tested positive
for an arbovirus from this pooling effort. Bladen County had 2 pools of An. crucians
testing positive for a California group virus. Brunswick County had 1 pool of Ochlerotatus taeniorhynchus test positive for a California group virus. New Hanover County had 1 pool of Ochlerotatus infirmatus test positive for Highlands
J virus. Finally, Pender County had 1 pool of Culex salinarius test positive for EEE. Table 1
provides an overview of the 2000 multi-county pooling results. Graph 4 shows the distribution of mosquito pools by
Table 1. September 26, 2000
Multi-County Pooling Results
melanura of Total
Mosquito Pools Collected
Pools Testing Positive EEE
Pools Testing Positive an Arbovirus
2001 (Week 40)
One mosquito pool tested positive for
an arbovirus from this pooling effort. Pender County collected 1 pool of Culiseta
melanura that tested positive for Highlands J virus. Table 2 provides an
overview of the 2001 multi-county pooling results. Graph
5 shows the distribution of mosquito pools by genus.
Table 2. October 3, 2001
Pooling Results (Week 40)
Total Mosquitoes Collected
Bridge Vectors Collected
Total Cs. melanura Tested
% Cs. melanura of Total
Number of Mosquito Species Collected
Number of Mosquito Pools Collected
Number of Trap Nights
Mosquito Pools Testing Positive EEE
Mosquito Pools Testing Positive an Arbovirus
pools tested positive for an arbovirus from this pooling effort. Pender County
collected 1 pool of Culiseta melanura testing positive for Highlands J
virus. New Hanover County collected 1 pool of Culiseta melanura testing
positive for Highlands J virus. Table 3 provides an overview of the 2002
multi-county pooling results. Graph 6 shows the
distribution of mosquito pools by genus.
Table 3. October 9, 2002 Pooling Results (Week 41)
Cs. melanura Tested
Cs. melanura of Total
of Mosquito Species Collected
of Mosquito Pools Collected
of Trap Nights
Pools Testing Positive EEE
Pools Testing Positive an Arbovirus
Chicken Flock Results from Project Counties 2000-2002
Table 4 shows that the counties in the project area
had at least one bird seroconvert for EEE during the 3-year study period. The
2000 season identified 10 sentinel birds serocoverting for EEE. Two birds seroconverted
in both 2001 and 2002. The table also shows which week each year the birds seroconverted
and which week each year the multi-county pooling effort was projected and
conducted. For example, during the 2000 season, the pooling date of September
26 (Week 39) was selected in Week 34 based on EEE activity in the sentinel
flock in Bladen County and the emergence of a new brood of Cs. melanura
during the same week in Brunswick County (Graph 2). In order to properly interpret the data
it should be noted that a seroconversion in a sentinel bird provides
information about an earlier brood of Cs. melanura. In the 2000
example, the brood of Cs. melanura that caused the seroconversion in
Week 34 emerged in Week 31. The operational information needed when a bird seroconverts
are the flocks bleeding schedule and the sample collection date. By overlaying
this information onto weekly light trap data, the brood of Cs. melanura
most likely involved in EEE transmission to the sentinel bird can be
Table 4. Sentinel Flock Surveillance Supporting the Projected Pooling Dates
Pooling was Projected
Sentinel Flock EEE Positives by County
Brunswick, New Hanover, Onslow
Statewide Passive Human and Veterinary Surveillance
During the 2000 season, 2 human and 17 horse EEE
cases were identified. The 2001 season produced no human cases and 13 horse
cases, in 2002, there were 5 horse cases and 1 human case (Graph
7). The summary of reported human and veterinary surveillance for
the project counties includes 1 EEE positive horse in 2000 (Week 30) from New Hanover County, 1
EEE positive horse in 2001 (Week 38) from Bladen County and 1 EEE horse positive from Onslow County in
2002 (Week 52). There were no positive human EEE cases reported from any of
the project counties during 2000-2002.
Environmental Results 2000-2002
Graph 8 shows the weekly
precipitation amounts for 2000, 2001, 2002 and the average rainfall for
southeastern North Carolina for the last 8 years. Rainfall for 2000 was 0.42
inches below the norm, 2001 was 17.36 inches below the norm and 2002 season was
4.92 inches below the norm. Operationally speaking, annual rainfall amounts
are not nearly as important as which week during the year has significant
During this project, the affects of temperature on Cs.
melanura populations were observed. While this was difficult to measure,
it was apparent that heat indices above 100oF had an impact on the Cs.
melanura populations. The possible explanations for this may be that
temperatures over 100oF caused significant mortality to the Cs.
melanura populations, the high temperatures drove the Cs. melanura
population deeper into the swamp habitat reducing the likelihood of collection,
and finally, exceedingly high temperatures could be initiating summer
aestivation in the existing Cs. melanura population.
Benefits of the Timeline
The EEE timeline gives county programs responsible
for operational mosquito control a virus transmission scenario to look for
within the data they are collecting. Combining mosquito light trap data with
sentinel flock and environmental data to anticipate a potential increase in EEE
activity is a proactive initiative and not a reactive response, which is so
often the case. The timeline also encourages data sharing and inter-agency
communication among city, county, state and federal officials.
Operationally, the timeline shows local programs the
mosquito population situation. This can reduce unneeded mosquito spraying and
increase the effectiveness of their applications by targeting the correct
species at the correct locations at the correct time.
One of the toughest things for a local program to assess
is the need for adulticide control measures based on passive arbovirus
surveillance results. Typical limitations include inadequate reporting by the
medical and veterinary community and the laboratory confirmation process.
Utilizing the timeline, local programs can determine which part of the timeline
they are dealing with when notified of a EEE positive horse, for example.
Additionally, the timeline can be used to support operational control decisions.
The timeline will indicate that the mosquito responsible for infecting a horse
between days 19 and 41 is already dead by the time the lab results come in,
sometimes weeks or even months after the initial infection.
The Centers for Disease Control has suggested that
one of the limitations to in an arbovirus situation is that long-term
surveillance information about local mosquito populations is rarely available.
Encouraging long-term surveillance using the EEE timeline as a regular
component of nuisance mosquito operations will streamline control efforts and
facilitate a public health response to increased arbovirus activity.
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