______________________________________
Reflections on the Ecology and Epidemiology of
Eastern Equine Encephalitis in the Northeastern
United States
________________________________________________
Theodore G. Andreadis
Center for Vector Biology & Zoonotic Diseases and
Northeast Regional Center for Excellence in
Vector-Borne Diseases
The Connecticut Agricultural Experiment Station
New Haven, CT
Eastern Equine Encephalitis
• Most pathogenic arthropod-borne virus in North
America (Alphavirus: Togaviridae, SSRNA)- ~ 6-7 human cases / year
- 40% case fatality rate
- Neurological impairment in survivors (35%)
- No commercial vaccine or effective treatment
• Activity is most common in and around freshwater
hardwood swamps – highly focal
• Perpetuates in an enzootic cycle involving wild
Passeriformes birds and ornithophilic mosquitoes
• Principal enzootic vectors in the northeastern US
- Culiseta melanura
- Culiseta morsitansRole as “bridge vectors”
unresolved
• 1831 - First equine outbreak of EEE virus in Massachusetts - “horses dying
of a brain disease”
• 1933 - First isolation of EEE virus from horse brain during an outbreak in
coastal areas of Delaware, Maryland, New Jersey and Virginia
• 1934 - Mosquitoes first incriminated as potential vectors in a series of vector
competence studies with species of Aedes, Culex and Coquillettidia
• 1935 - Birds considered as reservoir hosts; 1950 - first isolation of EEE made
from a wild bird
• 1938 - The first human cases confirmed – 35 (25 fatal) human cases, > 300
horse cases in Massachusetts; 38 horse cases in Connecticut
Key EEE Historical Events – Northeastern US
• 1938 - Shown that virus could cause of encephalitis in wild and
domestic pheasants in Connecticut
• 1949 - First isolation of EEE virus from mosquitoes – Cq. perturbans
• 1951 - First isolation of EEE virus from Culiseta melanura
• 1959 - Major outbreak in New Jersey – 33 human cases
• 1971 - EEE discovered in Central New York – 1st human case
Human Cases of EEE in the United States 1964 - 2019
N = 347
NY
NJ
MA
NH
VT
CT RI
ME
0
5
10
15
20
25
19
66
19
68
19
70
19
72
19
74
19
76
19
78
19
80
19
82
19
84
19
86
19
88
19
90
19
92
19
94
19
96
19
98
20
00
20
02
20
04
20
06
20
08
20
10
20
12
20
14
20
16
20
18
< 1 case / yr
Resurgence
4-5 cases / yr
Northward
Expansion
Human Cases
- 1964 – 2002 = 47
- 2003 - 2019 = 79
• 1964 to 2002 –
sporadic outbreaks
with no apparent
pattern
• 2003 – resurgence
and expansion
Resurgence and Expansion of EEE Activity in NE US
Armstrong & Andreadis NE J Med 2013
RI
NY
ME
NJ
NHVT
MA
CT
0
5
10
15
20
25
30
35
40
45
50
20
03
20
04
20
05
20
06
20
07
20
08
20
09
20
10
20
11
20
12
20
13
20
14
20
15
20
16
20
17
20
18
20
19
EEE Veterinary Cases
Northeastern US
2003 - 2019
N = 309
Ave = 18 / yr
Human and Veterinary Cases of EEE in the Northeastern US - 2019
CT MA ME NH NJ NY RI
Veterinary Cases = 42
0
2
4
6
8
10
12
14
CT MA ME NH NJ NY RI
Human Cases = 23
Fatalities = 8 (35%)
NY
NJ
MA
NH
VT
CT
RI
ME
EEE Activity = 30 Counties, 7 States
Human cases = 23 (8 fatalities, 35%)
Veterinary cases = 42
• Reforestation and wetland restoration –
by mid 1800’s much of the forests in the
northeastern US were stripped and
cedar swamps were destroyed
Factors Contributing to the Resurgence of EEE in the Northeastern US
- Increased habitat for Culiseta
melanura
- Proliferation of wetland roosting sites
for birds (e.g. robins, wood thrush)
• Suburban development near critical
wetland mosquito habitat
- Increasingly expose people to the
threat of EEE infection
• Changes in average temperatures and
precipitation events related to climate
change
- Milder winters
- Warmer summers
- Extremes in both precipitation and
drought
- Enhance overwintering survival
- Extend transmission season
- Accelerate generation time
- Increase frequency of blood feeding
- Accelerate virus replication within
mosquito
- Allow mosquitoes to extend northward
range
Komar & Speilman NY Acad Sci 1994
Pre-Season
• Significant EEE activity in the previous
year
• Mild winters with insulating snow cover
• High water table in enzootic swamps
• Above average rainfall in the prior
fall/winter and spring
Historical Risk Factors for EEE in the Northeastern US
In-Season
• Above average Culiseta melanura
populations
• EEE virus isolations from mosquitoes in
June or early July
• Isolations of EEE virus from a mammal-
biting mosquitoes – Cq. perturbans
• Numerous EEE isolations > 30 – 50
• High MIR in Culiseta melanura >1:1000
• EEE activity beyond traditional areas
• Early and above average equine cases
• Infection of a human prior to August
aaa
a
aaaaa
a
a
0
5
10
15
20
0
10
20
30
40
50
60
70
80
901
99
6
199
7
199
8
199
9
200
0
200
1
200
2
200
3
200
4
200
5
200
6
200
7
200
8
200
9
201
0
201
1
201
2
201
3
201
4
201
5
201
6
201
7
201
8
201
9
Me
an
Cs
. m
ela
nu
ra/ tr
ap
EE
E Is
ola
tes
fro
m C
s.
me
lan
ura
Year
No. EEE virus isolations
Mean no. Cs. melanura / trap
EEE isolations and abundance of Cs. melanura – Connecticut, 1996-2019
EEE activity
associated with
Cs. melanura
abundance
0
5
10
15
20
25
30
35
40
45A
ve
. N
o M
osq
uito
es p
er
Lig
ht T
rap
Culiseta melanura AbundanceConnecticut
2019
22 Year Mean
June July August September October
0
40
80
120
7/1 7/8 7/15 7/22 7/29 8/5 8/12 8/19 8/24 9/2 9/9 9/16 9/23 9/30 10/7 10/14 10/21 10/28
No
. E
EE
vir
us
is
ola
tio
ns
Week
1996-2018
2013
Weekly Isolations of EEE virus
from field collected mosquitoes
in Connecticut
0
40
80
120
7/1 7/8 7/15 7/22 7/29 8/5 8/12 8/19 8/24 9/2 9/9 9/16 9/23 9/30 10/7 10/14 10/21 10/28
No
. E
EE
vir
us
is
ola
tio
ns
Week
1996-2018
2019
2013
2019
2019
2019
Weekly Isolations of EEE virus
from field collected mosquitoes
in Connecticut
Earlier EEE virus
amplification in
2019
Culiseta melanura
• Habitat: Densely wooded freshwater swamps
(red maple and white cedar) and sphagnum
bogs
• Development: Develop in subterranean
“crypts” in deep shaded cavities under tree
roots
• Seasonal Distribution: mid-May – October
• Feeding Preference: Primarily birds with
occasional feeding on mammals including
humans
• Number of Generations: 2-3 per year
• Adult Flight Range: > 2 miles
• Overwintering Stage: Larvae (all instars)
0.0
4.0
8.0
12.0
16.0Crypt water temperature
0 C
0.0
0.2
0.4
0.6
0.8
1.0
P L4 L3 L2
* * * ***
adults
Rela
tive p
roport
ion o
f each s
tage
DEC JAN FEB MAR APR MAY
* L1
Phenology of Overwintering Development of Cs. melanura
Andreadis et al JAMCA 2012
Cs. melanura64.1%
Ae. canadensis7.8%
Ae. cinereus4.0%
Ae. vexans3.6%
Cx. salinarius2.9%
Ur. sapphirina2.9%
Cx. pipiens2.4%
Ae. trivittatus2.2%
An. punctipennis1.9%
Minor species (10)7.8%
Species (n = 19) No.
Cs. melanura 264
Ae. canadensis 32
Ae. cinereus 18
Ae. vexans 15
Cx. salinarius 12
Ur. sapphirina 12
Cx. pipiens 10
Ae. trivittatus 9
An. punctipennis 8
Ae. cantator 5
Cs. morsitans 5
Cq. perturbans 4
Others (7 species) 18
EEE Virus Isolations from Mosquitoes in Connecticut 1996 - 2018
Cs. melanura65.6%
Ur. sapphirina5.7%
Cq. perturbans4.0%
Ae. canadensis 4.1%
Cx. salinarius3.3%
Ae. vexans3.3%
Ps. ferox3.3%
An. punctipennis2.5%
Ae. cinereus 1.6%Cx. pipiens
1.6%
Minor species (5)4.1%
EEE Virus Detections from Mosquitoes in CT, MA, NJ and NY - 2019
Connecticut
Cs. melanura45.9%
Cq. perturbans33.4%
Ae. canadensis5.6%
Cx. salinarius10.6%
Ae. vexans1.6%
Cx. pipiens/restuans2.8%
Massachusetts
New Jersey
Cs. melanura74.0% Ae. canadensis
2.7%
Ae. triseriatus2.7%
Ae.albopictus2.7%
Cx. pipiens/restuans17.8%
Cs. melanura88.9%
Cq. perturbans4.8%
Cx. salinarius4.8%
Ae. vexans1.6%
New York
1.0
<0.8
<0.8
1.6
1.5
2.8
>3.0
1.3
0.8
6.6
1.4
1.7
1.7
1.2
2.9
0 1 2 3 4 5 6 7
Ur. sapphirina
Ps. ferox
Ae. trivittatus
Ae. triseriatus
Ae. cantator
Ae. canadensis
Cq. perturbans
Cx. salinarius
Cx. restuans
Cs. melanura
An. walkeri
An. quadrimaculatus
An. punctipennis
Ae. vexans
Ae. cinereus
Mean Log10 PFU’s/mosquito pool
Required for transmission
1,000 – 10 million PFU’s
1Armstrong & Andreadis EID 2010
2Nasci & Mitchell JAMCA 1996
2
• There are major
differences in the quantity
of virus found in EEE
virus-positive, field-
collected mosquitoes
• Cs. melanura appears to
be the only species in
which virus titers are
sufficiently high enough to
support efficient
transmission
• Other species include Ae.
cinereus, Cq. perturbans,
Ae. canadensis
• Important to consider virus
titers when implicating
other mosquito vectors
Mean EEE Virus Titers in Field-Collected Mosquitoes by
Plaque Assay 1
0
20
40
60
80
100
Cs. melanura Cq. perturbans Ae. canadensis Cx. salinarius Ae. cinereus
Bird
Mammal
Bird & Mammal
Host Feeding Patterns of Culiseta melanura and Potential Bridge Vectors of EEE in the Northeastern US (CT, MA, NY, VT)
Perc
ent
0.2% Human
5.1% Human
2.0% Human
3.7% Human
Molaei & Andreadis EID 2006, Molaei et al AJTMH 2006, JME 2008, VBZD 2013
8.4% Human
0 20 40 60 80 100
Connecticut
Massachusetts
New York
Vermont
Mammal
Bird
94.2%
94.0%
98.9%
99.7%
6.0%
5.8%
1.1%
0.3%
Proportion of Avian and Mammalian Derived Blood Meals in Culiseta melanura populations in the Northeastern US
1.3% Human
0.3% Human
Molaei & Andreadis EID 2006, Molaei et al AJTMH 2006, JME 2008, VBZD 2013
3.7% Equine
Wood Thrush
American Robin
Tufted Titmouse
Common Grackle
Chipping Sparrow
Black-capped Chickadee
Northern Cardinal
Red-eyed Vireo
Scarlet Tanager
ConnecticutN = 42 species
Molaei & Andreadis EID 2006, Molaei et al AJTMH 2006, JME 2008, VBZD 2013
18%
14%
13%
Wood Thrush
American Robin
Song Sparrow
Ovenbird
Red-eyed Vireo
Scarlet Tanager
Common Yellowthroat
Baltimore Oriole
Black-Capped
Chickadee
New YorkN = 52 species
24%
9%8%
American Robin
Tufted Titmouse
Black-capped
Chickadee
Scarlet Tanager
Field Sparrow
Northern Cardinal
Wood Thrush
Red-eyed Vireo
Baltimore Oriole
Massachusetts N = 55 species
22%
9%
9%
Green Heron
American RobinCommon
Yellowthroat
Black-capped Chickadee
Savannah Sparrow
Common Grackle
Northern Cardinal
Rose-breasted Grosbeak
Veery
VermontN= 49 species
18%
12%
10%
Culiseta melanuraAvian-Derived Blood Meals
EEE Antibody Prevalence in Wild Birds: Regional Comparisons
0
20
40
60
80
100
Wood Trush American Robin Gray Catbird Ovenbird Song Sparrow
NJ (Crans et al. JME 1994)
MA (Main et al. AJTMH 1988)
NY (Emord and Morris, JME 1984)
NY (Howard et al. JME 2004)
ME (Elias et al. VBZD 2017)
Perc
ent
Hillsborough NH 2005
Merrimack NH 2005
New Haven CT 2006
Rockingham NH 2006
Rockingham NH 2005
Hillsborough NH 2005
Rockingham NH 2006
Rockingham NH 2006
Hillsborough NH 2006
Fairfield CT 2003
New London CT 2004
New London CT 2003
New London CT 2003
New London CT 2004
New London CT 2003
New London CT 2003
Fairfield CT 2003
Windham CT 2003
Fairfield CT 2003
New London CT 2003
Fairfield CT 2003
Fairfield CT 2003
New London CT 2001
New Haven CT 2001
Fairfield CT 2001
New London CT 2001
Middlesex CT 2001
Fairfield CT 2000
New Haven CT 2003
NH
2005-06
CT
2003-04
CT
2001 & 03
CT
2000-01
Oswego NY 2007
Oswego NY 2005
Oswego NY 2004
Oswego NY 2004
Oswego NY 2004
New London CT 2006
New London CT 2007
New London CT 2007
New London CT 2007
New London CT 2006
New London CT 2004
Fairfield CT 2003
New London CT 2003
New London CT 2003
Fairfield CT 2003
New London CT 1998
Fairfield CT 1998
Middlesex CT 1998
New London CT 2003
Georgia 1997
New London CT 2003
New London CT 2000
Hartford CT 2000
Florida 1991
Florida 1982
New London CT 1996
Florida 1993
NY
2004-07
CT
2006-07
CT
2003
CT
2000
• EEE viruses group into
temporally discrete genetically
diverse clades by year -
suggests separate annual
introduction events into the
region
− Migrating viremic birds
• Some strains persist into 2nd
year - provides evidence for
local overwintering
− Vertical transmission in
mosquitoes (Philbrook et al. CDC TR
1961)
− Recrudescence in chronically
infected birds (Crans et al. JME 1994)
− Reptiles or amphibians (?)
• Support for both hypotheses
Tracking Eastern Equine Encephalitis Virus
Perpetuation in the Northeastern United States by
Phylogenetic Analysis
PM Armstrong, TG Andreadis, JF Anderson, JW Stull, CN Mores.
2008. Am. J. Trop. Med. Hyg.
• Sequenced complete genomes of 433
EEEV strains collected within the U.S.
from 1934 to 2014
• EEEV evolves relatively slowly and that
transmission is enzootic in Florida,
characterized by higher genetic diversity
and long-term local persistence
• EEEV in CT, MA and NY were
characterized by lower genetic diversity,
multiple introductions, and shorter local
persistence
• Supports a source-sink model in which FL
is the major source of EEEV
No.
sequences
No.
humancases
160
120
40
80
24
15
7
9
1-4
Journal of Virololgy 92 (12), 2018, 1-18.
Large-Scale Complete-Genome Sequencing and
Phylodynamic Analysis of Eastern Equine Encephalitis
Virus Reveals Source-Sink Transmission Dynamics in
the United States
Tan et al. (19 authors)
Northeastern US EEE Virus Transmission Cycle
Virus
EnzooticCycle
Wild Passerine Bird Reservoir
and Amplifying Hosts
Juneto
October
Culiseta melanura
August to October
Epidemic /
Epizootic
Transmission
“Bridge Vectors”
Coquillettidia perturbans
Aedes canadensis
Culex salinarius
Culiseta melanura
Local
Overwintering
& Annual
Introduction
• High likelihood that the EEE virus will
reemerge
- EEE usually persists after a major outbreak
- Have consistently experienced equine
and/or human cases every year since 2004
• Unlikely that we will experience as
high a level of EEE virus activity
- Herd immunity in reservoir birds – dampen
enzootic transmission
• Remains to be seen how widespread
activity will be
- Will we see further expansion into NH, ME
and VT?
Human case
Veterinary case
Expectations for 2020
0
10
20
30
40
50
60
70
20
04
20
05
20
06
20
07
20
08
20
09
20
10
20
11
20
12
20
13
20
14
20
15
20
16
20
17
20
18
20
19
2019
Human cases
Veterinary cases
2004 - 2019
• Human serosurvey - human exposure ?
- EEE antibodies detected in 0.7% of persons with no history of
encephalitis after 1955 outbreak in Massachusetts (Feemster et
al NEJM 1958)
- Inapparent infections ranged from 3.1% to 7.6% after the
1959 outbreak in New Jersey (Goldfield et al. Am J Epidem 1968)
• Identification of Cs. melanura breeding sites in newly
recognized foci of human and animal infection
• Screening larvae for virus – overwintering
- One reported isolation of EEE virus from Cs. melanura larvae (Philbrook et al CDC Tech Rep 1961)
- Never been duplicated or confirmed
• Pre-season treatment of Cs. melanura breeding sites
• Methoprene has been shown to be an effective larvicide
when applied by fixed wing aircraft (Woodrow et al JAMCA
1995)
• Enhanced mosquito surveillance – in season
Research – Surveillance Priorities
EEE Challenges and Issues
1. Risk assessment and communication
• How do best assess human risk and communicate it to the public
• Analysis and interpretation of surveillance findings
• Triggers for response
2. Sharing samples for genetic analysis and validation
• Virus availability – virus isolation vs PCR detection
3. Delays in laboratory diagnosis of human infection
• Concerns with commercial labs – serology and false negatives
4. Prevention and control
• Personal protective measures – effectiveness?
• Preseason preemptive treatments of Cs. melanura breeding sites with larvicides
• Truck-mounted and aerial adulticides – how do we evaluate effectiveness
• Difficulties with public acceptance – environmental issues
• Delays in implementation and high costs
• What level of control is needed to reduce human risk of infection
Questions?
Alexander Skochkov “Old Mosquito”