Potential sources of the 1995 Venezuelan equine encephalitis subtype IC epidemic.

Venezuelan equine encephalitis viruses (VEEV) belonging to subtype IC have caused three (1962-1964, 1992-1993 and 1995) major equine epizootics and epidemics. Previous sequence analyses of a portion of the envelope glycoprotein gene demonstrated a high degree of conservation among isolates from the 1962-1964 and the 1995 outbreaks, as well as a 1983 interepizootic mosquito isolate from Panaquire, Venezuela. However, unlike subtype IAB VEEV that were used to prepare inactivated vaccines that probably initiated several outbreaks, subtype IC viruses have not been used for vaccine production and their conservation cannot be explained in this way. To characterize further subtype IC VEEV conservation and to evaluate potential sources of the 1995 outbreak, we sequenced the complete genomes of three isolates from the 1962-1964 outbreak, the 1983 Panaquire interepizootic isolate, and two isolates from 1995. The sequence of the Panaquire isolate, and that of virus isolated from a mouse brain antigen prepared from subtype IC strain P676 and used in the same laboratory, suggested that the Panaquire isolate represents a laboratory contaminant. Some authentic epizootic IC strains isolated 32 years apart showed a greater degree of sequence identity than did isolates from the same (1962-1964 or 1995) outbreak. If these viruses were circulating and replicating between 1964 and 1995, their rate of sequence evolution was at least 10-fold lower than that estimated during outbreaks or that of closely related enzootic VEEV strains that circulate continuously. Current understanding of alphavirus evolution is inconsistent with this conservation. This subtype IC VEEV conservation, combined with phylogenetic relationships, suggests the possibility that the 1995 outbreak was initiated by a laboratory strain.

Genetic diversity and relationships among Venezuelan equine encephalitis virus field isolates from Colombia and Venezuela.

During field studies of enzootic Venezuelan equine encephalitis (VEE) viruses associated with epizootic emergence, a large number of virus isolates were made in sylvatic foci of Venezuela and Colombia. To rapidly characterize these isolates, antigenic subtypes were determined by means of immunofluorescence and by single-strand conformational polymorphism (SSCP) analysis by use of an 856-bp fragment from the P62 gene, which we used to distinguish genetic variants. Representative isolates were sequenced to assess the sensitivity of SSCP to detect genetic differences. The SSCP analysis distinguished isolates differing by as little as 1 nucleotide; overall, differences of > or = 1 nucleotide were recognized 89% of the time, and the sensitivity to distinguish strains that differed by only 1 or 4 nucleotides was 17 and 57%, respectively. Phylogenetic analyses of representative sequences showed that all recent isolates from the Catatumbo region of western Venezuela and the middle Magdalena Valley of Colombia were closely related to epizootic subtype IAB and IC strains; strains from Yaracuy and Miranda States were more distantly related. Cocirculation of the same virus genotype in both Colombian and Venezuelan foci indicated that these viruses are readily transported between enzootic regions separated by > 300 km. The SSCP analysis appears to be a simple, fast, and relatively efficient method of screening VEE virus isolates to identify meaningful genetic variants.

Emergence of a new epidemic/epizootic Venezuelan equine encephalitis virus in South America.

One of the most important questions in arbovirology concerns the origin of epidemic Venezuelan equine encephalitis (VEE) viruses; these viruses caused periodic, extensive epidemics/epizootics in the Americas from 1938-1973 (reaching the United States in 1971) but had recently been presumed extinct. We have documented the 1992 emergence of a new epidemic/epizootic VEE virus in Venezuela. Phylogenetic analysis of strains isolated during two outbreaks indicated that the new epidemic/epizootic virus(es) evolved recently from an enzootic VEE virus in northern South America. These results suggest continued emergence of epizootic VEE viruses; surveillance of enzootic viruses and routine vaccination of equines should therefore be resumed.

Venezuelan equine encephalitis virus: comparison of infectivity and virulence of strains V-38 and P676 in donkeys.

Two strains of Venezuelan equine encephalitis (VEE) virus were examined for the ability to replicate in, as well as to produce death among donkeys. One, a low passage strain known as strain P676 was originally isolated from mosquitos in Venezuela. The other, strain V-38 was isolated from a horse brain in 1938 and had undergone an unknown number of laboratory passages; it is used extensively for the preparation of inactivated VEE vaccine. Both strains were found to be approximately equal in their ability to infect donkeys. However, a quantity as small as 50% hamster intraperitoneal infectious units of strain V-38 resulted in fatal infection. On the other hand, as much as 631 million infectious units of strain P676 were nonfatal in one of two donkeys. It appears that strain V-38 is approximately 100 million times more virulent than strain P676 in equine species. One donkey which received strain P676 demonstrated a biphasic pattern of clinical illness and viremia, and there is suggestive evidence that another animal experienced a second and fatal clinical response 3 weeks after virus inoculation.