Isolation and phylogenetic analysis of Mucambo virus (Venezuelan equine encephalitis complex subtype IIIA) in Trinidad

Abstract

In the 1950s and 1960s, alphaviruses in the Venezuelan equine encephalitis (VEE) antigenic complex were the most frequently isolated arboviruses in Trinidad. Since then, there has been very little research performed with these viruses. Herein, we report on the isolation, sequencing, and phylogenetic analyses of Mucambo virus (MUCV; VEE complex subtype IIIA), including 6 recently isolated from Culex (Melanoconion) portesi mosquitoes and 11 previously isolated in Trinidad and Brazil. Results show that nucleotide and amino acid identities across the complete structural polyprotein for the MUCV isolates were 96.6 – 100% and 98.7 – 100%, respectively, and the phylogenetic tree inferred for MUCV was highly geographically- and temporally- structured. Bayesian analyses suggest the sampled MUCV lineages have a recent common ancestry of approximately 198 years (with a 95% highest posterior density (HPD) interval of 63 – 448 years) prior to 2007, and an overall rate of evolution of 1.28 × 10−4 substitutions/site/yr.

Isolation and Characterization of Sylvatic Mosquito-Borne Viruses in Trinidad: Enzootic Transmission and a New Potential Vector of Mucambo Virus

“…Most virus isolates were from mosquitoes collected during the rainy season (June-September), with a peak in October ( Figure 1 In a previous report, we showed strong geographic structuring in an MUCV phylogeny based on 18 sequences from Brazil (n = 4) and Trinidad (n = 14) and temporal structuring within Trinidad, which showed at least three major clades. 8 The ML phylogeny generated in the present study, which includes 25 additional sequences from Trinidad, also supports the existence of at least three clades, with sequences from Trinidad and Brazil in separate clades and two temporally defined subclades in Trinidad ( Figure 2 ). The phylogeny also suggests that there is geographic structure within Trinidad because the sequence from the 2008 BBF isolate (Tri5991), although closely related, was distinct from all of the 2007 and 2008 sequences from the ASSR.…”

“…We recently reported the detection of Mucambo virus (MUCV, a member of the Venezuelan equine encephalitis [VEE] virus complex) and Guama group and group C orthobunyaviruses in mosquitoes from the ASSR, none of which were identified to species. 8 We report additional surveillance in the ASSR (total = 21 months), 7 months at the BBF, and 1 month at the HOA, and further identification and characterization of the viruses isolated.…”

“…Trapped mosquitoes were sorted into pools of 1-50 individuals according to species and sex and assayed for viruses by cytopathic effect assays on Vero cell monolayers as described. 8 If cytopathic effect was detected, the virus was re-isolated in Vero cells for confirmation, and the cell culture supernatant was collected and stored at −70°C for further characterization by serologic analysis and viral sequencing.…”

“…All isolated alphaviruses were subsequently identified as MUCV based on nucleotide homology. Amplification primers (26S and 6kE1), sequencing primers (8432F, 8638R, 8811F, 9132R), and polymerase chain reaction conditions were used as described by Auguste and others 8 to amplify and sequence a 2,079-nucleotide fragment specifying envelope glycoproteins E3 and E2 for further phylogenetic analysis. All nucleotide sequences derived in this study were submitted to GenBank (accession nos.…”

Serological Evidence of Flaviviruses and Alphaviruses in Livestock and Wildlife in Trinidad

“…These studies resulted in the identification and characterization of hundreds of arboviral isolates, primarily from the Bush Bush Forest, which is an approximately 0.85-sq km region within the Nariva swamp in southeastern Trinidad (Aitken et al 1968c;Jonkers et al 1968aJonkers et al , 1968bWorth et al 1968). With the exception of the aforementioned studies, there have been only a few serological studies in the 1970s and early 1980s (Price 1978a(Price , 1978bTikasingh et al 1983), and more recently, mosquito surveillance studies and opportunistic sampling of carcasses of Alouatta seniculus monkeys 1 School of Veterinary Medicine, confirmed the continued circulation of VEE subytpe IIIA viruses (Mucambo virus; MUCV), several orthobunyaviruses, and Yellow fever virus (YFV) (Auguste et al 2009(Auguste et al , 2010a(Auguste et al , 2010b.…”