Vance Vorndam

Texas Lifestyle Limits Transmission of Dengue Virus

Urban dengue is common in most countries of the Americas, but has been rare in the United States for more than half a century. In 1999 we investigated an outbreak of the disease that affected Nuevo Laredo, Tamaulipas, Mexico, and Laredo, Texas, United States, contiguous cities that straddle the international border. The incidence of recent cases, indicated by immunoglobulin M antibody serosurvey, was higher in Nuevo Laredo, although the vector, Aedes aegypti, was more abundant in Laredo. Environmental factors that affect contact with mosquitoes, such as air-conditioning and human behavior, appear to account for this paradox. We conclude that the low prevalence of dengue in the United States is primarily due to economic, rather than climatic, factors.

Molecular evolution and phylogeny of dengue type 4 virus in the Caribbean

We sequenced the E gene and adjacent prM/M and NS1 junctions (1940 bp) of 48 Dengue-4 (DEN-4) isolates collected between 1981 and 1999 from 8 Caribbean islands and from 7 South and Central American countries. Phylogenetic analysis confirms a single introduction in the early 1980s and a high degree of gene flow resulting in a pattern of evolution defined more by time period than geographic origin, especially within the Caribbean basin. A modern Caribbean clade consisting of four distinct lineages has arisen, comprised of isolates from Caribbean islands and nearby regions of South America. This clade is defined by three amino acid substitutions in the E (aa 163 and 351) and NS1 (aa 52) proteins. These findings highlight the importance of migration and gene flow in dengue viral change and suggest that efforts to understand disease dynamics in the Caribbean basin need to focus at regional, rather than local scales.

Molecular and biological characterization of a non-glycosylated isolate of St Louis encephalitis virus.

The glycosylation patterns of the envelope (E) glycoprotein of several naturally occurring strains of St Louis encephalitis (SLE) virus were investigated. SLE viruses were found that contained both glycosylated and non-glycosylated E proteins, and one isolate (Tr 9464) that lacks N-linked glycosylation sites on its E protein was identified. SLE virus monoclonal antibodies that define E protein B cell epitopes and demonstrate biological activities reacted essentially to the same extent with glycosylated and non-glycosylated virions. These results indicate that glycosylation is not essential for epitope conformation or recognition. However, failure to glycosylate the E protein was associated with possible morphogenetic differences as manifested by reduced virus yields and differences in specific infectivity.

Diagnostic value of dengue virus-specific IgA and IgM serum antibody detection.

The diagnostic value of dengue virus (DV)-specific immunoglobulin A (IgA) serum antibody detection, by an indirect immunofluorescence assay (IFA) was evaluated. For this study, the kinetics of DV-specific IgA serum antibodies was analysed in two experimentally immunised macaques, paired samples from 35 patients suspected of a primary or secondary DV infection, paired sera from patients with high levels of IgA specific antibodies against influenza virus (n = 15), sera from patients with other viral infections (n = 40) and healthy blood donors (n = 10), which served as controls. The presence of DV-specific IgA serum antibodies in humans and in monkeys was compared with that of DV-specific IgM demonstrated in a capture enzyme-linked immunosorbent assay (ELISA). The development of DV-specific IgA and IgM antibodies in macaques proved to be similar to that observed in humans with a DV infection. In sera obtained from suspected primary DV patients during the acute phase and convalescent phase, DV-specific IgA was detected in 1/6 (17%) and 6/6 (100%), whereas IgM was detected in 4/6 (67%) and 5/6 (83%), respectively. In sera from suspected secondary DV patients during the acute phase and convalescent phase, DV-specific IgA was detected in 18/29 (62%) and 28/29 (97%), whereas IgM was detected in 20/29 (69%) and 28/29 (97%), respectively. The control group consisted of five paired serum samples from yellow fever vaccinated individuals and a patient with acute tick-borne encephalitis, 15 paired serum samples from patients with high levels of IgA antibodies specific for influenza virus and 40 serum samples from patients with specific IgM antibodies against other viruses. Ten serum samples from healthy blood donors were included. Among the control serum samples, in one patient, both DV-specific IgA and IgM antibodies were present, and in three sera DV-specific IgM antibodies could be demonstrated. These data suggest that detection of DV-specific IgA serum antibodies by IFA may have additional value for the diagnosis of DV infection.

A PCR-restriction enzyme technique for determining dengue virus subgroups within serotypes

The polymerase chain reaction (PCR) and restriction enzyme analysis were used to develop a rapid and simple procedure for identifying geographic subgroups of dengue virus within serotypes for epidemiologic investigations. The entire structural protein region of dengue viruses was amplified and the products were digested with the endonucleases AluI or DdeI. By comparing the restriction fragment length polymorphisms (RFLPs), we recognized dengue-2 and dengue-3 subgroups that corresponded to those previously determined by oligonucleotide fingerprinting or genomic sequencing. This procedure can be performed in 2 days without the use of radioisotopes, and results can be interpreted without computer analysis. For those analyses which require only subgroup affiliations, this is a useful tool for rapidly screening multiple virus isolates.