[The relationship between mosquito vectors and aquatic birds in the potential transmission of 2 arboviruses]. / Relación entre mosquitos vectores y aves acuáticas en la transmisión potencial de dos arbovirus.

The authors studied for two years the role of the chicks of aquatic birds in the arboviral cycles in coastal lagoons in central Panama in order to determine the relation between Culex (Melanoconion) ocossa and Mansonia (Mansonia) dyari mosquitoes in the transmission and dissemination of the viruses of Saint Louis Encephalitis (SLE) and Venezuelan Equine Encephalitis (VEE). Mosquitoes were captured every fifteen days on two consecutive nights to isolate the virus, using light traps (CDC) and baited traps. The attempts to isolate the virus were made using Vero cell cultures and the determination of antibodies was performed. The results of the serologic tests seem to indicate that four bird species: the ex (?) heron (Bubulcus ibis), the American heron (Casmerodius albus), the spoon-billed duck (Cochlearius cochlearius) and the needle crow (Anhinga anhinga) could function as intermediate hosts in the transmission cycle of SLE. Two species, the ibis (Endocimus albus) and the spoon-billed duck (Cochlearius cochlearius) could also be intermediate hosts of VEE in the coastal lagoons of Panama. The presence of antibodies in chicks could indicate an infection acquired recently, after their birth, in this area. The VEE virus was recovered from blood filled mosquitoes which had fed on a spoon-billed duck probably infected and exposed in a Trinidad #10 trap. No SLE virus was isolated. Other unknown viruses were isolated from mosquitoes selected for these studies, such as C. ocossa and M. dyari. The results obtained with these studies indicate the need for more studies utilizing new field techniques in order to establish a link between SLE and VEE, the vector mosquitoes and the aquatic birds in the coastal lagoons of the area under investigation.

Comparative infections of epizootic and enzootic strains of Venezuelan equine encephalomyelitis virus in Amblyomma cajennense (Acari: Ixodidae).

To compare the potential for an enzootic or an epizootic strain of Venezuelan equine encephalomyelitis (VEE) virus to infect Amblyomma cajennense (F.), larval ticks were fed on guinea pigs (strain 13) inoculated with an enzootic viral strain of variant I-E (68U201) or on guinea pigs inoculated with an epizootic strain of variant I-A (Trinidad donkey). Peak viremias were 10(5.2) plaque-forming units (PFU)/ml and 10(7.3) PFU/ml in guinea pigs infected with enzootic and epizootic viral strains, respectively. Ticks feeding on enzootic- and epizootic-infected hosts had viral titers of 10(2.5) and 10(3.9) PFU per tick, respectively, at drop-off. Although epizootic virus was recovered from 98% (127 of 130) of larval ticks up to 16 d after drop-off, enzootic virus was recovered from 95% (19 of 20) at drop-off (mean titer, 10(2.5) PFU per tick), with recovery rates declining rapidly to 2 of 10 (mean titer, 10(1.4) PFU per tick) by 16 d after drop-off. Transstadially transmitted epizootic virus was found in 0.4% (12 of 2,950) of unfed nymphs (mean titer, 10(2.8) PFU per tick) 63 d after drop-off, 1% (5 of 521) fed nymphs 69 d after drop-off, and 1% (4 of 400) of unfed adults (mean titer, 10(3.6) PFU per tick) 106 d after drop-off. No enzootic virus was recovered from 4,600 unfed nymphs tested 63 d after drop-off.

Pathogenesis of Venezuelan equine encephalitis virus infection in mice and hamsters.

The pathogenesis of Venezuelan equine encephalitis (VEE) virus infection was compared in intraperitoneally inoculated mice (n = 24, 6 to 8 weeks old) and hamsters (n = 9, 90-110 g) using histopathology and immunohistochemical localization of VEE virus antigen. Infected mice developed paralysis, and the majority died by 9 days after inoculation. In contrast, hamsters did not survive beyond 3 days after inoculation, and they did not develop any neurologic signs. VEE virus antigen, demonstrated by immunoperoxidase staining, and pathologic changes were present in extraneural organs of both mice and hamsters. There was more severe involvement in hamsters, particularly in Peyer's patches of the distal small intestine. There was a severe encephalomyelitis in mice, but pathologic changes were not well established in the brains of hamsters before death. VEE virus antigen was widespread in the central nervous system of both mice and hamsters. VEE virus was found to be highly neurotropic in hamsters and had a similar distribution in the brain as in mice, but hamsters died from their extraneural disease before major central nervous system disease developed.

Venezuelan equine encephalitis virus propagation in the olfactory tract of normal and immunized mice.

The role of virus spread in the induction of damage to the central nervous system (CNS) of mice infected with Venezuelan equine encephalitis virus (VEEV) via the respiratory route was studied. The virus concentration in various organs and in the blood, the sensitivity to different doses of virus, and ultrastructural lesions in various tissues were examined. It is concluded that VEEV can enter the CNS of nonimmunized mice both by vascular and by olfactory pathways, whereas in immunized mice the olfactory pathway is the main route.

Nose-only versus whole-body aerosol exposure for induction of upper respiratory infections of laboratory mice.

The effectiveness of two aerosol delivery systems, nose-only and whole-body, were compared using Swiss-Webster mice and two pathogens, Klebsiella pneumoniae and Venezuelan equine encephalitis (VEE) virus. With K. pneumoniae the median lethal dose (LD50) and the mean time to death correlated with the inhaled dose. An LD50 value of 335 colony forming units (cfu) for nose-only exposure was significantly less than the LD50 value of 3741 cfu obtained for whole-body exposure. The LD50 values obtained with VEE virus for nose-only exposure [8 plaque forming units (pfu)] and whole-body exposure (11 pfu) were similar to each other. Following a 10-min nose-only exposure, concentrations of K. pneumoniae approximating 10(4)/g were present after 24 hr in the upper respiratory tract (URT) and lungs. The numbers of bacteria reached a peak at 72 hr, when resolution of the infection began. Detectable levels of bacteria in the blood and tissues were delayed in mice given whole-body exposure, plus there was a decreased concentration of bacteria per gram of tissue. Major pathological lesions induced by K. pneumoniae were mild suppurative rhinitis and minimal suppurative bronchopneumonia. Viremia was greatest at 96 hr following aerosol exposure to VEE. Virus concentrations in the URT, lungs, cerebrum, spleen and mesenteric lymph nodes reached maximum titers earlier for mice exposed by nose-only than for mice exposed to whole-body aerosols.(ABSTRACT TRUNCATED AT 250 WORDS)

[Antiviral factor isolated from infected cell cultures]. / Antivirusnyi faktor, vydeliaemyi iz infitsirovannykh kul'tur kletok.

An antiviral factor of protein nature inhibiting reproduction of VEE, herpes, vaccinia VSV, fowl plague viruses was isolated from infected cell cultures. The factor has no virucidal or prophylactic effect, stable to low pH values and heating at 100 degrees C for 30 min.

Vector competence of Culex (Melanoconion) taeniopus for allopatric and epizootic Venezuelan equine encephalomyelitis viruses.

The vector competence of Culex (Melanoconion) taeniopus was examined in the laboratory for "enzootic" allopatric and "epizootic" strains of Venezuelan equine encephalomyelitis viruses of hemagglutination inhibition subtypes I, II, III, and IV. Following bloodmeals from viremic hamsters, and extrinsic incubation of 20-22 days, mosquitoes were allowed to refeed for transmission attempts. Infection rates never exceeded 50% with oral doses of less than 10(4) chick embryo cell culture plaque forming units (CECPFU), and approached 100% only after ingestion of greater than or equal to 10(5.5) PFU. Transmission was achieved for some "epizootic" subtype IABC and "enzootic" subtype ID strains after bloodmeals containing greater than or equal to 10(3.4) CECPFU; subtypes II, III, and IV were never transmitted despite oral doses up to 10(5.0) CECPFU. These data contrast sharply with those reported previously for sympatric "enzootic" subtype IE Middle American Venezuelan encephalitis viruses.

Arbovirus investigations in Argentina, 1977-1980. I. Historical aspects and description of study sites.

This is the introductory paper to a series on the ecology of arboviruses in Argentina. Epizootics of equine encephalitis have occurred since at least 1908, principally in the Pampa and Espinal biogeographic zones, with significant economic losses; human cases of encephalitis have been rare or absent. Both western equine and eastern equine encephalitis viruses have been isolated from horses during these epizootics, but the mosquitoes responsible for transmission have not been identified. A number of isolations of Venezuelan equine encephalitis (VEE) virus were reported between 1936 and 1958 in Argentina, but the validity of these findings has been seriously questioned. Nevertheless, serological evidence exists for human infections with a member of the VEE virus complex. Serological surveys conducted in the 1960s indicate a high prevalence of infection of humans and domestic animals with St. Louis encephalitis (SLE), and 2 SLE virus strains have been isolated from rodents. Human disease, however, has rarely been associated with SLE infection. Only 7 isolations of other arboviruses have been described (3 of Maguari, 1 of Aura, 2 of Una, and 1 of an untyped Bunyamwera group virus). In 1977, we began longitudinal field studies in Santa Fe Province, the epicenter of previous equine epizootics, and in 1980 we extended these studies to Chaco and Corrientes provinces. The study sites are described in this paper.

Studies on the transmission of Venezuelan equine encephalitis virus by Colombian simuliidae (Diptera).

The ability of Simulium mexicanum and Simulium metallicum to serve as biological or mechanical vectors of an enzootic and an epizootic strain of Venezuelan equine encephalitis (VEE) virus was examined. Guinea pigs were inoculated with the epizootic Cordoba strain or the enzootic RPVP407 strain of VEE virus. Wild-caught adult Simuliidae were fed on the viremic guinea pigs and the virus content of groups of flies was determined at daily intervals post-engorgement to test for viral replication. Flies were refed on suckling mice at greater than or equal to 8 days post-engorgement to test for biological transmission. Other flies were interrupted while feeding on viremic guinea pigs and refed on suckling mice to test for mechanical transmission. Neither S. mexicanum nor S. metallicum appear to be efficient vectors of either strain of VEE virus, although occasional mechanical transmission was obtained. Titers of virus in flies decreased rapidly after engorgement and from 3-12 days post-engorgement virus was detected only in 5%-25% of both species of flies. Although earlier field evidence implicated both S. mexicanum and S. metallicum as vectors of epizootic VEE, we conclude that it is highly unlikely that they play an important role as vectors of the virus in nature.

[Variability of attenuated alphavirus strains in virion aggregate infection]. / Izmenchivost' attenuirovannykh shtammov al'favirusov v usloviiakh infektsii agregatami virionov.

A natural process of infection with multiploid virions was stimulated and the genetic effects occurring in this infection were studied in experiments with homogeneous attenuated strain of Venezuelan equine encephalomyelitis virus producing the smallest plaques. The process of infection with multiploid virions was reproduced by infecting the cells with artificially obtained aggregates. Under these conditions, small- and large-plaque virus was produced which in some cases had a higher virulence than the parental strain. A possible mechanism of this phenomenon and its practical implications for the evaluation of the properties of attenuated alphavirus strains is discussed.

[Morphological study of a mixed alphavirus infection]. / Morfologicheskoe izuchenie smeshannoi infektsii al'favirusami.

In combined paired cultivation of 8 Venezuelan equine encephalomyelitis virus, multiploid virions formed in 17.8% of cases. Five clones with this effect were used in mixed infections with clones of Semliki Forest and Sindbis viruses. In these infections changes in virions of the virus progeny were observed in 20% and 16.6%, respectively. Mixed cultivation of pairs of Sindbis and Semliki Forest viruses resulted in formation of multiploid virions in 42.8%. Besides, in two mixed populations formed upon combined multiplication of the latter viruses virions of unusual shapes were found: rounded, oval, elongated, and triangular designated as polymorphic.

[Use of continuous human lymphoblastoid cell lines (T- and B-origin) to produce persistent tick-borne encephalitis virus and Venezuelan equine encephalomyelitis virus infections]. / Ispol'zovanie perevivaemykh limfoblastoidnykh linii kletok cheloveka (T- i B-proiskhozhdeniia) dlia polucheniia persistentnoi infektsii virusa kleshchegogo éntsefalita i virusa venesuél'skogo éntsefalomielita loshadei.

Persistent infection with tick-borne encephalitis virus (TBE) was established in experimentally infected continuous lymphoblastoid human cell lines Raji, L-101 (of B-origin) and 1387 (T-origin) and with Venezuelan equine encephalomyelitis (VEE) virus in Raji and 1387 lines. The persistently infected lines produced infectious virus, the cells showed specific fluorescence in immunofluorescent tests, and electron microscopic examinations revealed TBE and VEE virions in sections.

[Alphavirus reproduction in cultures of transplantable human lymphoblastic cells in acute infection]. / Reproduktsiia al'favirusov v kul'turakh perevivaemykh limfoblastoidnykh kletok cheloveka pri ostroi infektsii.

The results of investigations of acute infection of continuous human B- and T-cells with typical members of the alphavirus group--Semliki Forest, Sindbis, and Venezuelan equine encephalomyelitis viruses, are presented. Virus amplification was shown to pass through the typical phases: eclipse, logarithmic growth, plateau. Infectious virus production per one cell was from 10 to 10,000 PFU in various cultures. Cell infection results in interferon production. Replication of the viruses under study in lymphoblastoid cell cultures is not accompanied by the active cytocidal effect. The regularities determining the sensitivity of lymphoblastoid cells to viruses in general and alphaviruses in particular are discussed. Proceeding from the results of the study of alphavirus replication in human continuous B- and T-cells it is suggested that this system be used as a model for the analysis of antiviral activity of interferon, its inducers, and chemopreparations in special cells. Lymphoblastoid and fibroblast interferon are as active in B-cells and show no antiviral activity in continuous T-cells, as interferon inducer.

Virion envelope glycoproteins as epidermiological markers of Venezuelan encephalitis virus isolates.

Virion polypeptide compositions of 26 isolates of Venezuelan encephalitis virus were analyzed by a reproducible and comparative technique of discontinuous sodium dodecyl sulfate-polyacrylamide slab gel electrophoresis. Although the molecular weight of the core polypeptide for each isolate was 36,000, numbers and molecular weights of envelope glycoproteins were heterogeneous. Isolates associated with human, but not equine, disease usually had two glycoproteins of 50,000 to 51,000 and 51,000 to 55,000 molecular weight, whereas isolates associated with both human and equine disease usually had an additional, third polypeptide band of either 45,000 to 46,000 or 56,000 to 58,000 molecular weight. The former isolates were in hemagglutination inhibition subtypes I-D, I-E, III, or IV, and the latter were in subtypes I-A, I-B, I-C, or II. Thus virion envelope glycoproteins should be useful markers of Venezuelan encephalitis virus isolates in epidemiological investigations.

Multi-factorial specification of virus-host interactions: studies with strains of Venezuelan equine encephalomyelitis virus in mice.

Mice of different ages were infected i.p. or i.c. by 23 different strains of VEE virus. The course of the virus host interaction was specified in terms of the efficiency of infection, the outcome of infection as lethality or protection and the survival time. These separately quantifiable features all showed several host-maturation events that combine to provide a multifactorial specification of virus-strains and host-responses. This base-line for correlations with the responses of principal hosts (equidae and man) may be expanded to test correlations with the antigenic or in vitro characteristics of virus-strains.

Further observations on infections of guinea pigs with Venezuelan encephalitis virus strains.

Guinea pigs from a Guatemalan colony died after subcutaneous inoculating of moderately small doses of equine-benign strains of Venezuelan encephalitis (VE) virus of hemagglutination-inhibition subtype I-E from enzootic habitats in Mexico and Guatemala. Thus these guinea pigs were unlike English short hair and inbred 13 guinea pigs, which usually survive infections with equine-benign VE strains of subtype I-E. We therefore caution others that not all strains of guinea pigs can be used to evaluate the potential equine virulence of VE viruses.

Venezuelan encephalitis virus: in vivo induction of a chromosomal abnormality in hamster bone marrow cells.

This study reports the induction in vivo of aneuploidy in bone marrow cells of the Syrian hamster after short-term infection with Venezuelan encephalitis virus.