serogroups b, c, bunyamwera, patois, and minatitlan

Transcription

serogroups b, c, bunyamwera, patois, and minatitlan
Am. J. Trop. Med. Hyg., 32(4), 1983, pp. 877—885
Copyright ©1983 by The American Society of Tropical Medicine and Hygiene
.
IDENTIFICATION
OF
ARBOVIRUSES
HITHERTO
FROM
UNRECOGNIZED
ECUADOR:
MEMBERS
OF
SEROGROUPS B, C, BUNYAMWERA,
PATOIS, AND MINATITLAN
CHARLES H. CALISHER,* ERNESTO GUTIERREZ V.,t D. BRUCE FRANCY,8
ARACELY
ALAVA
A.,t
DAVID
J. MUTH,*
@DJOHN
S. LAZUICK*
*Division of Vector-Borne Viral Diseases, Centers for Disease Control, Fort Collins,
Colorado 80522, and tlnstituto Nacional de Higiene, Guayaquil, Ecuador
Abstract.
Three hundred seventy-nine virus isolates were obtained from mosquitoes cob
lected and sentinel hamsters exposed in coastal Ecuador from 1974 to 1978. These included
four alphaviruses [Venezuelan equine encephalitis lB (1), Venezuelan equine encephalitis 1D
(35), western equine encephalitis (1) and eastern equine encephalitis (4)]; two flaviviruses [St.
Louis encephalitis (3) and Nananjal (6)1; 11 bunyaviruses [Maguari (243), Playas (3), Vinces
(33), Tunbock (2), Abras (5), Babahoyo (3), Acana (2), Guajara (3), San Juan (6), Pueblo Viejo
(3), 18 unspecified Gamboa serogroup viruses, Palestina (7)]; and one vesiculovirus (vesicular
stomatitis
New
Jersey).
All but Venezuelan
equine
encephalitis
virus
were
new to Ecuador,
and Naranjal (scrognoup B), Playas (Bunyamwera serogroup), Vinces (scrognoup C), Abras
and Babahoyo (Patois scrogroup), San Juan and Pueblo Viejo (Gamboa scrogroup) and Pa
lestina (Minatitban serogroup) are newly recognized viruses. These isolates have enabled us
to 1) expand our knowledge of the geographic distribution of recognized viruses, 2) expand
our knowledge of the members of certain scrogroups and 3) establish two new serogroups
(Gamboa and Minatitlan).
In January 1969, an epidemic of Venezuelan
equine encephalitis (VEE) occurred in Guayas
Province, Ecuador, affecting humans and equines.
Retrospective studies suggested that this had been
an extension and expansion of an earlier outbreak
of VEE further inland.' Since the enzootic pncs
ence of epizootic VEE virus subtypes could have
accounted fon periodic appearances of VEE in
coastal Ecuador and since, subsequent to 1969,
VEE spread northward from Ecuador and Cen
tnal America to south Texas,2 a cooperative in
vestigation of arbovirus transmission and ecology
in the coastal lowlands of Ecuador was initiated
to determine whether cnzootic transmission of epi
zootic strains was occurring there. From 1974 to
1978, mosquitoes and other arthropods were col
bected and sentinel hamsters exposed at various
sites,
maximizing
the number
of virus
isolates
ob
tamed.
Ecologic descriptions of the study areas will be
published elsewhere, as will extensive tabulations
of arthropod collections and definitive idcntifica
tions of mosquitoes.3 This article describes the
identifications of virus isolates obtained from an
Accepted 24 November 1982.
thnopods,
brates.
sentinel
Among
hamsters,
the 379 isolates
alphaviruscs,
two flaviviruses,
and one vesiculovirus. These
newly recognized viruses.
MATERIALS
and other
obtained
verte
were four
11 bunyaviruscs,
18 included eight
AND METHODS
Collection and processing of specimens
Adult hamsters were exposed in the field for up
to 10 days, after which they were returned to the
laboratories in Guayaquib and observed for signs
of illness. Sick hamsters were tested for virus by
inoculating clarified 10% suspensions of their tis
sues (whole blood, brain, heart, kidney, skeletal
muscle on liver-lung-spleen
pools) intracnanially
into separate bitters of 2- to 4-day-old suckling mice
(SM). Mice were observed daily for 2 weeks for
signs of illness; the brains of those which appeared
ill were passed as 10% suspensions to other SM
to establish strains for subsequent virus identifi
cations. Other mammals and birds were shot on
live-trapped in the field and their tissues tested for
virus by the same methods used for virus isolation
from sentinel hamsters.
Mosquitoes and other arthropods were collected
877
878
CALISHERET AL.
with CDC light traps4 supplemented with dry ice
(CO2), using methods published previously.5 Ar
thropods were sent frozen on dry ice (—60°C)
to
the laboratories in Fort Collins, Colorado, where
they were sorted, pooled by species, identified and
ground in Medium 199 with heated (56°C/30')20%
fetal bovine serum, using sterile abundum and
chilled mortars and pestles.5 Each of the resulting
suspensions was inoculated onto monolayer cub
tures of continuously propagated Veno and pri
many duck
embryo
cell cultures.
The cell cultures
were overlaycd with a medium containing agar
and examined daily for plaque formation by the
method of Hayes et al.5 When plaques were ob
served, subsequent passage to a litter of SM served
to establish the isolate.
Virus identification
and characterization
dropped onto a monolayer culture of appropriate
cells and the cells were incubated for 45 mm at
37°Cand overhayed with nutrient medium con
taming agar. For viruses producing plaques in 1—
3 days, a single overlay, containing !:40,000 neu
trab red, was used; a double overlay—the second,
containing 1:25,000 neutral red, applied on the
3rd day after inoculation—was used for viruses
which produced plaques 4 or more days after in
oculation of cell cultures.
Virus diluent for passage of tissue suspensions,
harvested plaques, and stock viruses was Medium
199 with 5% heated fetal bovine serum supple
mented with antibiotics.
Stock viruses were prepared as !0% SM brain
suspensions in phosphate-buffered
saline (pH 7.4)
with 4% bovine albumin. For production of an
tibody,
mice
were
hypcrimmunized
according
to
the schedule suggested by Brandt et al. , 12and as
citic fluids were elicited by the method of Tika
singh et al. i:t
@
Complement fixation (CF) tests were performed
by the microtiter adaptation method of Casey,6
using crude, alkaline extracts of SM
When
sucrose-acetone extracted antigens were required,
RESULTS
for example, with crude antigens of low titer, or
when definitive (reference comparison) reactivitics
Nine flavivirus strains were isolated from sen
were needed, extracts were prepared by the meth
tinel hamsters and mosquitoes. Three isolates of
St. Louis encephalitis (SLE) virus were obtained
ods of Clarke and Casals.8 The same sucrose-ace
(76V-! 177, 24684 and 78V-5682), two from Culex
tone extracted antigens also were used for hem
agglutination and hemagglutination-inhibition
(HI)
nigripalpus
mosquitoes
and one from a sentinel
hamster. In addition, six strains of another flavi
teSts.―For preliminary serologic identifications, a
virus were isolated from Culex (Melanoconion)
battery of grouping fluids, prepared by the Ar
bovirus Reference Branch, or obtained from the
adamesi and Cx. (Mel. ) spp. mosquitoes and the
heart and skeletal muscle of a sentinel hamster.
Reference Resources Branch, U.S. National In
stitutes of Health, was used. This battery of im
Strain
25008, isolated
from a sentinel
hamster,
was chosen as the prototype for this virus because
munc mouse ascitic fluids (MAF) represented more
it was isolated, identified, and characterized first.
than 200 recognized viruses. When an antigen was
This virus titered 6. 7 and 7. 1 hog,,,PFU/0. 1 ml and
found to react with one or more of these reagents,
formed 1- and 4-mm indistinct, irregular plaques
additional tests, using MAF for individual virus
in duck embryo and Vcro cells, respectively. Strain
es, were performed.
effects in C6/36 (Aedes
A test of sensitivity to the dchipidizing action of 25008 causes cytopathic
albopictus mosquito) cells 6 days after inoculation
sodium deoxycholatc (SDC) was performed in mice
by the method of Thciler9 and in cell
‘°and kills SM on the 5th day after intracranial in
To determine virulence by the peripheral route of oculation but does not kill weaned or older mice
inoculated intrapenitoneally.
inoculation, 0. ! ml of a 1:100 suspension of virus
Strain 25008 was shown by HI and CF tests to
was inoculated intraperitoncally
into five or six
be a flavivirus and identified as a newly recog
8-week-old mice.
nized flavivirus by N tests (Table 1). Hemagglu
Virus titrations were by plaque assay in cell cul
tination of goose crythrocytes by strain 25008 oc
tures. For definitive virus identifications, a serum
curned at pH 6. 1 to 7.0 with optimum titers at pH
dilution-plaque
reduction neutralization
(N) test
6. 7—6.9.
was used.― Briefly, 200 plaque-forming
units
(PFU) of virus were mixed with an equal 0. 1-mb
Thirty-three
Group C virus strains were iso
volume of antibody. After overnight incubation at bated, three from Cx. (Mel. ) vomerifer mosquitoes
4°C, 0. 1 ml of the serum-virus
mixture was
and 30 from tissues of 12 sentinel hamsters. In CF
IDENTIFICATION
OF ARBOVIRUSES
FROM ECUADOR
879
TABLE!
Results of hemag.glutination-inhibition
(HI), complement-fixation
(CF) and serum dilution-plaque
tralization (N) tests with Ecuadorian strain 25008 and other fiaviviruses
to:25008
NHICFNBussuquara
Antigenorvirus
StrainTiter
HI
CF
of antibody
NBSQSLEHICF
—!60——BeAn4O7880
256
25008160
@2,56016St.
10——160@1,0241,280Rocio
Louis enc.
TBH-28—
——864Yellow SPH-34675!0
fever
17D20
—108Aroa
!680816Ilheus
VENA 1809320
—40864Jutiapa Onig.80
reduction
64
64040256
!6080@!,024
16
32
8
!6
8108—Tamana JG-128!0
154—8—Antigen
bat
Tr 27
to:ROCYFAROAHI
or virus
NHICFNBussuquara
StrainTiter
CFNHI
2500880
—108BeAn
32—40
6420
25610
407880
TBH-2840
SPH-34675640
fever
17D@80
160108Aroa
VENA !809320
—320@‘!,024Ilheus
Onig.1,280
—4032Jutiapa
—10256St.
—1064Rocio
Louis enc.
——64Yellow
—108Tamana
of antibody
CF
—
@‘1,0241,280—
64160
256
6440
51280
JG-!2820
3210
@27!54——8Antigen
bat
Tr
to:ILHJUTTB
CFNBussuquara
or virus
StrainTiter
CFNRI
CFNHI
8St.
8Rocio Louis enc.
4078640
TBH-28160
!6—20
6410
!28—
8Yellow
SPH-34675160
512—
8—tBeAn
25008640
8Aroa
8Ilheus
8Jutiapa
fever
!7D320
VENA !8092,560
Orig.@!0,480
@!,024Tamana
154——S
bat
tested.t
Blank
signifies
JG-!2880
Tr 27
not
—= <10 HI, or N, <8 CF.
6410
6440
@!,02464080
—2,560
of antibody
HI
neu
880
CALISHER ET AL.
TABLE 2
TABLE 3
Results of serum dilution-plaque reduction neutraliza
tion (N) tests with Ecuadorian strain 75V-807 and 12
Results of serum dilution-plaque reduction neutraliza
tion (N) tests with two Bunyamwera serogroup viruses
other Group C viruses
from Ecuador
viruses
and 22 other Bunyamwera
serogroup
75V-807VirusStrainHomologous
75V-3066vi@sStrainHomologous
bodyBruconha75V-807
titerVirusAnti
body75V-3066160160160MaguariE4-34841,28040@eMaguariBeAr
titervi@Anti
80OssaBT-182032020320RestanTr-5!!44@5,120320—“ApeuBeAn
6401,280
77V-148141,280@5,!201,280
727232040—BataiMM-2222@2,560160—BehiefeUGMP-68302,56020—ileshaKO-2@5
84816040—CaraparuBeAn
399416040—MaritubaBeAn
1532040—MurutucuBeAn
974@64040—Gumbo
Valley6V-63316040—Tlacotalpan6!D-24032020—TensawA9-!7
!H160—40OribocaBeAn
LimboFe3-7
17a'64020—ItaquiBeAn
lb64040—BunyamweraRI-!a1,28020—GermistonSAAr-!0502,560——ShokweSAAr-4
12797@640——NepuyoTr-!846280——Madrid40308320——a—
signifies <20.
RosaM2-14931,280——LokernFMS-43322,56040—Northway0234160——Taia
67!320——WyeomyiaOriginal64020—AnhembiSPAr-298464020—SororocaBeAr
tests all 33 strains reacted to within twofold of the
homologous titer (1: 128) of the prototype strain
(75V-807)
and
were
glc Group
C virus.
considered
Strain
as strains
75V-807
of a sin
formed
plaques
in Veno (1 mm, distinct on day 4) but not duck
embryo cells, and was pathogenic for 3- to 4-week
old mice inoculated intraperitoneally;
these mice
died 4—6days after inoculation by this route.
Strain 78V-807 was tested by N with 12 other
Group C viruses (Table 2).
Most of the viruses isolated during these studies
arc members of the Bunyamwera senogroup. Aedes
scapularis and Aedes spp. mosquitoes were the
source of 243 isolates of Maguari virus (topotype
E4-3484), but three Bunyamwera scrogroup iso
hates were obtained from Aedes taeniorhynchus
mosquitoes. These three (75V-3066, 75V-5758 and
75V-5938) had been isolated from mosquitoes cob
hected 8 March 1975. By N tests (results not shown)
all three were identical to each other but distinct
from the prototype Maguari virus (BeAr 7272). For
definitive identifications both E4-3484 (Maguari
virus)
and the second
virus
(prototype
75V-3066)
were tested by N with all recognized Bunyamwena
senogroup viruses (Table 3).
Eight
viruses
related
by CF to members
of the
Patois serogroup were isolated, six from Culex
(Mel. ) paracrybda, one from Cx. (Mel.) adamesi
mosquitoes and one from the blood of a sentinel
hamster.
Since
preliminary
149@2,56020—Main 32
DrainBFS-50151,280——KairiTr-8900320——GuaroaJC-216040—TucundubaBeAr
tests indicated
shared
CF antigens, N tests were performed. First, the
278160——a
sjg@@fles<20.
eight strains were tested among themselves. Re
sults of these tests showed that five viruses, rep
resented by prototype strain 75V-1 183, were iden
tical to each other but distinct from three other
viruses, represented by prototype strain 75V-2858.
Subsequent N tests included the four recognized
Patois senogroup viruses and both Ecuadonian
prototypes. The results showed that both Ecua
dorian viruses were distinct from the four necog
nized Patois serogroup viruses (Table 4).
All eight Ecuadorian Patois serogroup viruses
were tested in SM and Vero cells for pathogenicity
and in vitro characteristics.
The results revealed
similar characteristics in both hosts by senotype;
strain 75V-1 183 and viruses identical to it did not
cause CPE but strain 75V-2858 and viruses iden
tical to it did (days 5—7).The eight field strains
killed SM in 6—10days and titered i0@°to 1064
in Veno cells by plaque assay, with 1-mm plaques
appearing 5—7days after inoculation.
Five isolates, two from Cx. (Mel.) paracrybda,
and one each from Cx. (Aedinus) amazonensis and
IDENTIFICATION
OF ARBOVIRUSES
TABLE
FROM ECUADOR
881
4
Results of serum dilution-plaque reduction neutralization (N) tests with two viruses from Ecuador and four other
Patois serogroup viruses
[email protected]:?.@:c@—*————7SV-28S880@:P40———PatoisBT-497
1——160———Shark
RiverFE4-!R——40@!,280——PahayokeeFE3-S2F————320—ZeglaBT-S0!2————40320a
signifies
<20.
Uranotaenia spp. mosquitoes and the blood of a
sentinel hamster, were shown by CF tests to be
related to members of the Capim senogroup. CrossCF tests revealed such extensive antigen sharing
that N tests were required for definitive identifications. Preliminary N tests showed that strains
strain E4-2535 is a strain of Acara virus and strain
183 15 is a variant of Guajara virus (Table 5). As
strain E4-2535 was serially passed in SM, its vir
ulence for and titer in these animals decreased
until, after six passages, it did not kill SM; its titer
by plaque assay in Vero cells remained stable rc
E4-2535
gardless
and 76V-885 1 were identical
of passage
level.
Strain 71U-344 is most closely related to Bcnfica
virus, and strains 7 1U-253 and 7 1U-350 most
closely related to Guajara virus.
Twenty-seven
virus isolates from Aedeomyia
serognoup
7 1U-
squamipennis
ham-
ed by CF and N tests to the previously
viruses
253, 7 1U-344
@
and distinct
from strains 183 15, 76V-2585 and 76V-3029, which
arc also identical. Therefore, for definitive identifications, N tests were performed with strains
E4-2535 and 18315. In addition, three Capim
from
Guatemala
and 7 1U-350,
(strains
all from sentinel
sters) were received from W. F. Schercr'4 and included in these studies. The results indicated that
bunyavirus,
lishcd
TABLE
Results
ofserum
dilution-plaque
reduction
neutralization
Acara
ACA
2535) (MOR) BSB
BeAn27639
.@2
320
E4-2S3S
640!60
Moriche
TrS7896
80
—320
BEN
344)GJA18315350)353)
—
—
—
—
—640
7!U-344
—
—
—
Guajara
BeAn!06!S
640
320
18315
640
320
7!U-3S0
320
80
640
640
160
320
BeAn !S3S64
Capim
BeAn8S82
S _
= <20.
—
B@S
CAP
—
—
20
160
640
7!U-253
—
JD
—*
BeAn8438!
Benevides
ungrouped
(N) tests with 13 Capim serogroup bunyaviruses
TR 2668
MARU8S63
to be rehat
5
BushBush
JuanDiaz
shown
Gamboa virus. Further studies, pub
revealed that at least two new
Benfica
@
were
Titer ofN antibody
to:(E4-(71U-(71U-(71U-virus
Strain
mosquitoes
—
320
—
—
320
—
—
—
320
320
—
—
—
80
320
20
40
20
—
—
—
640
640
—
640
640
—
640
CALISHER ET AL.
882
TABLE 6
Results
of complement-fixation
(CF) and serum dilution-plaque
reduction
neutralization
(N) tests with strain 76V-
l5o5from Ecuador and Minatitlan and Mirim viruses
to:76v-ls6sMNTMIR
antibody
Antigen or virusStrainTiterof
NCFNCFNMinatitlan
<8
CF,
<20
256
16
840
M67U5
BeAn 7722.@:
Mirim76V-!565
320C
=
CF
—!28
virus
and
three
of Pueblo
Viejo
virus
were identified; 18 isolates were not identified more
specifically than as members of the newly necog
nized Gamboa senogroup.
Seven isolates were shown by both CF and N
tests to be strains of a single virus (76V-1565); three
were isolated from Culex (Mel. ) paracrybda mos
quitocs
256—
—8
N.
ly recognized agents, San Juan and Pueblo Viejo
viruses, were present in Ecuador. Six strains of
San Juan
—
8
2,560
8—@
and four from tissues
of sentinel
Cross-CF tests with a representative
virus of
the six, strain 76V-1565, as well as Minatitlan and
Mirim viruses, showed that strain 76V-1565 cross
reacts significantly with Minatitlan but not Minim
viruses. Additional N tests confirmed the relation
ship of strain 76V-1565 with Minatitlan but not
Minim viruses (Table 6).
A single strain of vesicular stomatitis New Jer
sey (VSNJ)
virus
was isolated
from a pool of Man
sonia indubitans mosquitoes. The virus was iden
tified in CF tests with a vesicular stomatitis
grouping MAF and with viruses and immune MAF
for other vesicular stomatitis senogroup viruses
(Table 7).
hamsters.
In preliminary CF tests using the battery of im
mune MAFs used throughout this study, these iso
lates reacted only with the NIH Polyvalent #2
MAF, which contains antibodies to M.inatitlan,
Jurona, Ahajucla,'5 Gamboa, and Belem viruses.
Additional CF tests showed that the six Ecuador
ian isolates reacted to titer with MAF for Mina
titlan and Minim viruses; weakly with MAF for
Patois, Capim, and Guama viruses; and not at all
with MAF for Junona, Abajucla, Gamboa, and
Belem viruses.
DISCUSSION
The six flavivinus strains shown to be distinct
from SLE virus appear to be isolates of a single
newly recognized virus, for which the name Nan
anjal virus is proposed (prototype strain 25008).
TABLE 7
Results of complement-fixation
viruses
(CF) tests with Ecuadorian
strain 76V-4357 and other vesicular stomatitis
serogroup
of CF antibody
to:76V-4357vSNJvslCHPCOC1SFPIRYvSA@76V-4357!,0241288—t———VS
vims5trainTiter
JerseyHazelhurst1,0241,024—VS
New
strain81,0248Chandipura1-6535
IndianaLab
1481,024CocalTr-402331625616IsfahanOriginal16512PiryBeAn
2423216!@
C
@
@5 (Alagoas):
antiserum
kindly
restricted and was not available
t —signifies <8.
Blank indicates not tested.
provided
to us.
by
Dr.
J. J. Callis,
U.S.
Department
of Agriculture,
Plum
Island,
Greenport,
NY;
use
of live virus
is
IDENTIFICATION
It is most closely related to Bussuquara
OF ARBOVIRUSES FROM ECUADOR
virus but
is cleanly distinct from it by both CF and N tests.
Strain
75V-807,
the prototype
Group
C bun
yavinus from Ecuador,
@
is distinct from all other
recognized
Group C viruses; the name
rus is proposed
for this agent.
Vinces
vi
Bunyamwera serogroup strains 75V-3066, 75V5758, and 75V-5938 are three isolates of a single,
newly recognized virus for which the name Playas
virus is proposed. The prototype, strain 75V-3066,
was easily distinguished from others in the seno
group, including the Ecuadorian topotype of Ma
guam virus which represented 243 of 379 (64.1%)
strains isolated during these studies. Previous
studies of Bunyamwera senognoup viruses in the
Americas, as elsewhere, have shown that they are
isolated from mosquitoes, such as Aedes scapula
ris
@
@
and
other
Aedes
spp.
which
feed
predomi
nantly on mammals. Of some apparent signifi
cance are the accumulated pieces of information
regarding equine pathogenicity of certain mem
bers of this serogroup; Maguari virus has been
isolated from horses in Guyana'6 and Argentina
(M. Sabattini, personal communication, 1978) and
Cache Valley virus from a horse in
and antibody to Bunyamwcna scrogroup viruses
is regularly found in equines distributed through
out the
In a serosurvey of 157 Ecuadorian
equines 106 (67.5%) had antibody to the Ecua
dorian Maguani virus topotype, strain E4-3484 (E.
Gutienrez
V.
and
C.
H.
Calisher,
and
25024)
caused
cytopathic
effects
identical to strain 183 15. Since recombination be
tween and among closely related bunyaviruses is
recognized, such may be an explanation for this
phenomenon; previous studies with the Gamboa
serogroup have shown similar antigen
These speculations notwithstanding,
Schercr (pen
sonal communication, 1982) has suggested that the
area in Guatemala from which these three isolates
were obtained is not likely to provide sufficient
Capim scrognoup virus amplification to support a
milieu in which viral persistence and recombina
tion could take place on a regular basis.
By the tests summarized in Table 5, Juan Diaz
and Benevides viruses arc indistinguishable.
We compared Ecuadonian
strain 76V-1565,
Minatitlan and Mirim viruses by both CF and N
tests. Mirim virus has been shown by R. E. Shope
(personal communication,
1978) to be rebated to
Minatitban virus. In our tests, strain 76V-1565 is
distinct from both Minatitban and Minim viruses;
we suggest the name Palcstina virus for this newly
recognized agent. Furthermore, the identification
of a virus closely related to, but distinct from, the
heretofore ungrouped Minatitlan virus is sufficient
grounds to warrant establishment of the Minati
than scrogroup of bunyaviruses. Minim virus is cx
cluded from those in the Minatitban serognoup be
cause (1) it is not sufficiently close antigenically
and (2) in other studies Minim virus was shown to
be properly classified as a member of the Guama
‘9
unpublished
data).
A comparison of the two prototype Patois scro
group bunyavirus strains (75V-1 183 and 75V-2858)
with the four previously recognized members of
the Patois serogroup showed that both Ecuador
ian viruses arc distinct, newly recognized viruses.
The name Abras virus is suggested for viruses rep
resented by strain 75V-1 183 and Babahoyo virus
for those identical to strain 75V-2858. Interesting
by, the three strains of Babahoyo virus (75V-2858,
76V-440
883
in
Vero cells, whereas the five strains of Abras virus
(75V-1285) did not. The significance, if any, of
such an observation is unknown.
Two Capim serogroup bunyaviruses were iso
lated. Strains E4-2535 and 786V-885 1 are two iso
lates of Acara virus; and strains 18315, 76V-2585
and 76V-3029 arc variants of Guajara virus.
However, while strains 183 15 and 7 1U-350 are
strains of Guajara virus, they are distinct from
each other; and strain 7 1U-253, a subtype of Gua
jara virus, is also a subtype of strain 7 1U-350 but
The isolation of vesicular stomatitis (New Jer
scy) virus, a known pathogen of cattle and hu
mans, has potential significance in Ecuador; this
virus should be considered in future arbovirus Sc
rosurveys of the area. As the pressures of human
populations cause expansion into the Amazon re
gion, a similarly expanding cattle industry will
probably emerge to feed the people necessary for
conducting exploration and mining of minerals and
other resources. Since alterations of the ecosystem
of this area will occur, the natural equilibrium will
be concomitantly altered, and VSNJ and other
viruses currently maintained cnzootically could
result in infection and perhaps disease outbreaks
in settlers in these areas.
A summary of virus isolations from these stud
ics in Ecuador is presented in Table 8. In all, at
least 18 distinct agents were isolated, eight of which
arc newly recognized.
In the absence of serosurvey data on prevalence
of antibodies to these viruses on their isolations
from clinical specimens collected from humans or
884
CALISHER
TABLE
Summary
ofvirus
8
isolationsfrom
Ecuador,
1974 —1978
ET AL.
Viral Diseases, Centers for Disease Control, for
his constant comments and continuing advice.
NumberofSerogroupVirusisolatesA
REFERENCES
equine
encephalitis lB
Venezuelan
1. Gutierrez V. , E. , Monath, T. P. , Alava A. , A.,
Uriguen,
equine
35
encephalitis !D
Western equine
1
encephalitis
Eastern equine
B. D., Arzuba,
R. M., and Chamber
lain, R. W. , 1975. Epidemiologic investigations
of the 1969 epidemic of Venezuelan encephalitis
in Ecuador. Am. J. Epidemiol., 102: 400—4!3.
2. Sudia,
W.
D. , Newhouse,
V.
Miller, D. L., Johnston,
encephalitis
BVenezuelan
St. Louis encephalitis
6BunyamweraMaguari
Naranjal1
4
3
L.
D.,
J. G., Jr., Young,
F. , Beadle,
R.,
Calisher, C. H. , and Maness, K. , 1975. Epidem
ic Venezuelan
equine encephalitis
in North Amer
ica in !97!: Vector studies. Am. J. Epidemiol.,
101: 17—35.
3. Francy,
D. B. Manuscript
in preparation.
4. Sudia,
W. D. , and Chamberlain,
R. W. , 1962.
3CVinces33TurhockTurlock2PatoisAbras
Playas243
Battery-operated light trap, an improved model.
3CapirnAcara
Mosq. News, 22: 126—129.
BabahoyoS
S. Hayes,
Guajara2
Juan
3GamboaSan
Pueblo Viejo
3
18MinatitlanPalestina7Vesicular
Unspecified6
stomatitis
(New Jersey)
stornatitis
TotalsVesicuhar
181
R. 0. , Francy,
D. B. , Lazuick,
Public Health Service Publication
379
J. S. , Smith,
G. C. , and Jones, R. H. , 1976. Arbovirus sur
veiblance in six states during 1972. Am. J. Trop.
Med. Hyg., 25: 463—476.
6. Casey, H. L. , 1965. Standardized diagnostic corn
phemcnt-fixation method and adaptation to micro
test, part H. Adaptation of LBCF method to rni
cro technique. Public Health Monograph No. 74,
No. 1228, U.S.
Govt. Printing Office, Washington, D.C. , pp. 3!—
34.
7. Calisher,C. H., and Maness, K. S. C., 1975. Lab
oratory studies of Venezuelan equine encephalitis
domestic animals, conclusions regarding their in
dividual or collective significance would be pre
mature. However, such a large and diverse viro
logical burden may, indeed, impact upon some
facet of the local vertebrates—be they humans,
animals of domestic importance or wild mammals
or birds. No evidence of such impact is presently
available.
ACKNOWLEDGMENTS
The authors are grateful to many individuals
whose efforts in the field and the laboratory made
this work possible. We would bike to extend par
ticular thanks to the following for their advice,
expert technical assistance and persistence in com
pleting these virus identifications: R. E. Shope,
Yale University School of Medicine, New Haven,
Conn. ; 0.
Garcia
giene, Guayaquih;
P. , National
and
Institute
R. Bohin, W.
of Hy
L. Jakob,
J. France, A. Hunt, D. Inghish, N. Karabatsos,
J. Trimble, and K. Wolff, Centers for Disease
Control, Fort Collins, Cob. We also thank Dr.
T. P. Monath, Director, Division of Vector-Borne
virus in equines, Texas,
biol., 2: 198—205.
8. Clarke,
D. H.,
and
197!.
Casals,
J.,
J. Clin. Micro
1958.
Techniques
for hemaggbutination and hemagglutination-in
hibition with arthropod-borne viruses. Am. J.
Trop. Med. Hyg., 7: 561—573.
9. Theiler,
M. , 1957.
Action
of sodium
deoxychohate
on arthropod-borne viruses. Proc. Soc. Exp. Biol.
Med., 96: 380—382.
10. Calisher,
C. H., and Maness,
K. S. C., 1973.
Use
of the sodium deoxycholate sensitivity test in cell
cultures. Acta Virol., 17: 26!—262.
11. Lindsey, H. S. , Calisher, C. H. , and Mathews, J.
H. , 1976. Serum dilution neutralization test for
California group virus identification and serobo
gy. J. Clin. Microbiol., 4: 503—5
10.
12. Brandt,
W. E. , Buescher,
E. L. , and Hetrick,
F.
M. , 1967. Production and characterization of ar
bovirus antibody in mouse ascitic fluid. Am. J.
Trop. Med. Hyg., 16: 339—347.
13. Tikasingh, E. S. , Aitken, T. H. G. , Worth, C. B.,
Spence, L., and Mongul, F. E., 1965. An out
break of eastern equine encephalomyehitis on the
Courantyne Coast of British Guiana. West In
dian Med. J., 14: 158—166.
14. Ordonez,
J. V. , Scherer,
W.
F. , and
Dickerman,
R. W. , 1976. Studies of possible movement of
Venezuelan encephalitis virus from an enzootic
focus in Guatemala
during
1971—1974. Am. J.
Trop. Med. Hyg., 25: 163—172.
IDENTIFICATION
15. Calisher,
C. H. , La.zuick,
J. S. , Justines,
OF ARBOVIRUSES
G. , Fran
cy, D. B., Monath, T. P., Gutierrez V., E., Sa
battini, M. S. , Bowen, G. S. , and Jakob, W. L.,
198!.
Viruses isolated from Aedeomyia
squami
pennis mosquitoes collected in Panama, Ecuador,
and Argentina:
Establishment
of the Gamboa
serogroup. Am. J. Trop. Med. Hyg., 30: 219—
223.
16. Spence, L. , Jonkers,
A. H. , and Grant, L. S. , 1968.
Arboviruses in the Caribbean Islands. Progr.
Med. Virol., 10: 415—486.
17. McLean, R. G., Parham, G. L., and Calisher, C.
885
FROM ECUADOR
H. Isolation of Cache Valley virus from a horse
in Michigan, 1980. (Manuscript in preparation.)
18. Berge, J. 0., ed., 1975. International Catalogue of
Arboviruses, 2nd ed. D.H.E.W. Publication No.
(CDC) 75-830!, Washington,
19. Calisher,
C. H.,
Coimbra,
D.C.
J. L. M.,
Lopes,
0.
de
S. , Muth, D. J. , Sacchetta, L. de A. , Francy, D.
B. , Lazuick, J. S. , and Monath, T. P. , 1983.
Isolation of new Guama and Group C serogroup
bunyaviruses
and an ungrouped
virus from
southern Brazil. Am. J. Trop. Med. Hyg., 32:
424—43!.