Mosquito saliva causes enhancement of West Nile virus infection in mice.

West Nile virus (WNV) is transmitted to vertebrate hosts primarily by infected Culex mosquitoes. Transmission of arboviruses by the bite of infected mosquitoes can potentiate infection in hosts compared to viral infection by needle inoculation. Here we examined the effect of mosquito transmission on WNV infection and systematically investigated multiple factors that differ between mosquito infection and needle inoculation of WNV. We found that mice infected with WNV through the bite of a single infected Culex tarsalis mosquito exhibited 5- to 10-fold-higher viremia and tissue titers at 24 and 48 h postinoculation and faster neuroinvasion than mice given a median mosquito-inoculated dose of WNV (10(5) PFU) by needle. Mosquito-induced enhancement was not due to differences in inoculation location, because additional intravenous inoculation of WNV did not enhance viremia or tissue titers. Inoculation of WNV into a location where uninfected mosquitoes had fed resulted in enhanced viremia and tissue titers in mice similar to those in mice infected by a single infected mosquito bite, suggesting that differences in where virus is deposited in the skin and in the virus particle itself were not responsible for the enhanced early infection in mosquito-infected mice. In addition, inoculation of mice with WNV mixed with salivary gland extract (SGE) led to higher viremia, demonstrating that mosquito saliva is the major cause of mosquito-induced enhancement. Enhanced viremia was not observed when SGE was inoculated at a distal site, suggesting that SGE enhances WNV replication by exerting a local effect. Furthermore, enhancement of WNV infection still occurred in mice with antibodies against mosquito saliva. In conclusion, saliva from C. tarsalis is responsible for enhancement of early WNV infection in vertebrate hosts.

Enhanced fibroblast-myocyte interactions in response to cardiac injury.

RATIONALE: A critical event in the development of cardiac fibrosis is the transformation of fibroblasts into myofibroblasts. The electrophysiological consequences of this phenotypic switch remain largely unknown. OBJECTIVE: Determine whether fibroblast activation following cardiac injury results in a distinct electrophysiological phenotype that enhances fibroblast-myocyte interactions. METHODS AND RESULTS: Neonatal rat myocyte monolayers were treated with media (CM) conditioned by fibroblasts isolated from normal (Fb) and infarcted (MI-Fb) hearts. Fb and MI-Fb were also plated on top of myocyte monolayers at 3 densities. Cultures were optically mapped after CM treatment or fibroblast plating to obtain conduction velocity and action potential duration (APD(70)). Intercellular communication and connexin43 expression levels were assessed. Membrane properties of Fb and MI-Fb were evaluated using patch clamp techniques. MI-Fb CM treatment decreased conduction velocity (11.1%) compared to untreated myocyte cultures. APD(70) was reduced by MI-Fb CM treatment compared to homocellular myocyte culture (9.4%) and Fb CM treatment (6.4%). In heterocellular cultures, MI-Fb conduction velocities were different from Fb at all densities (+29.8%, -23.0%, and -16.7% at 200, 400, and 600 cells/mm(2), respectively). APD(70) was reduced (9.6%) in MI-Fb compared to Fb cultures at 200 cells/mm(2). MI-Fb had more hyperpolarized resting membrane potentials and increased outward current densities. Connexin43 was elevated (134%) in MI-Fb compared to Fb. Intercellular coupling evaluated with gap fluorescence recovery after photobleaching was higher between myocytes and MI-Fb compared to Fb. CONCLUSIONS: These data demonstrate cardiac injury results in significant electrophysiological changes that enhance fibroblast-myocyte interactions and could contribute to the greater incidence of arrhythmias observed in fibrotic hearts.

Viral pathogenesis in mice is similar for West Nile virus derived from mosquito and mammalian cells.

West Nile virus (WNV) is a mosquito-borne pathogen. During replication, WNV acquires different carbohydrates and lipid membranes, depending on its mosquito or vertebrate hosts. Consequently, WNV derived from mosquito and vertebrate cell lines differ in their infectivity for dendritic cells (DCs) and induction of type I interferon (IFN-alpha/beta) in vitro. We evaluated the pathogenesis of WNV derived from mosquito (WNV(C6/36)) and vertebrate (WNV(BHK)) cell lines in mice. The tissue tropism, infectivity, clinical disease, and mortality did not differ for mice inoculated with WNV(C6/36) or WNV(BHK), and there were only minor differences in viral load and serum levels of IFN-alpha/beta. The replication kinetics of WNV(C6/36) and WNV(BHK) were equivalent in primary DCs and skin cells although primary DCs were more susceptible to WNV(C6/36) infection than to WNV(BHK) infection, suggesting that less virus is produced per infected cell for WNV(C6/36). In conclusion, viral source has minimal effect on WNV pathogenesis in vivo.

Serologic evidence for West Nile virus transmission in Puerto Rico and Cuba.

During the spring of 2004, approximately 1,950 blood specimens were collected from resident and Nearctic-Neotropical migratory birds on the Caribbean islands of Puerto Rico and Cuba prior to northerly spring migrations. Eleven birds and seven birds, collected in Puerto Rico and Cuba, respectively, showed evidence of antibody in a flavivirus enzyme-linked immunosorbent assay. Confirmatory plaque-reduction neutralization test results indicated neutralizing antibodies to West Nile virus in non-migratory resident birds from Puerto Rico and Cuba, which indicated local transmission.