Serum concentration and increased temperature alter Mayaro virus RNA and protein synthesis in Aedes albopictus (mosquito)-infected cells

We have previously shown the inhibition of Mayaro virus multiplication in Aedes albopictus-infected cells maintained at a supraoptimal temperature for growth (37§C) and a stimulation of virus production in response to high serum concentrations in the incubation medium. In the present study, we addressed the question of how the effect of continuous heat stress and high serum concentration soon after infection interfere with virus macromolecule synthesis. Cells maintained at 28§C in the presence of 2 percent serum synthesized a viral genomic RNA of 12 kb and a subgenomic RNA of 5.2 kb 6 h post-infection. Analysis of the protein profile showed the presence of the viral nucleocapsid protein of 34 kDa (P34). However, if infected cells were maintained at 37§C, a smear starting immediately below the 5.2-kb RNA was noticed and the viral P34 was not detected by SDS-PAGE. Addition of 10 percent serum to the growth medium of infected cells maintained at 37§C results in a viral RNA profile and proteins synthesis similar to those observed in cultures kept at 28§C, i.e., the smear was not observed and the P34 protein was detected. The results suggest that the inhibition of virus multiplication by temperature may be related to the inhibition of viral nonstructural protein synthesis early during infection. The presence of high serum levels in the incubation medium protects macromolecule synthesis against heat stress

Reduction of the accumulation of viral mRNA and of cellular rRNA synthesis in MHV3-infected macrophages of resistant mice

1. After MHV3 infection, only macrophages from resistant A/J mice partially restricted virus growth compared to those from susceptible BALB/c mice (2 logs of difference in virus titer). 2. Cellular ribosomal ribonucleic acid (rRNA) synthesis by MHV3-infected macrophages was decreased only in A/J mouse macrophages as indicated by accumulation of the 28S rRNA fraction. 3. The accumulation of viral messenger ribonucleic acids (mRNAs) in MHV3-infected macrophages was also reduced in A/J mouse macrophages compared to BALB/c mice. 4. In pulse-chase experiments of viral protein synthesis, the appearance, glycosylation and cleavage of glycoprotein S, as well as the metabolism of nucleoprotein N were delayed in A/J mouse macrophages. 5. These data show that MHV3 infection of A/J mouse macrophages induced an imbalanced accumulation of the 28S fraction of rRNA. Furthermore the synthesis of mRNAs correlated with viral protein synthesis in both A/J and BALB/c macrophages, but was delayed in A/J mice. 6. These results suggest that the partial restriction of MHV3 replication in macrophages of resistant A/J mice may take place during or before the mRNA synthesis, although it is correlated with the appearance, glycosylation, cleavage and metabolism of viral proteins

Fast growth of a Brazilian hepatitis A virus (HAF-203) in a primate cell line

A hepatitis A virus (HAV, HAF-203) isolated in Brazil was submitted to 8 serial passages through fetal Rhesus kidney cells (FRhK-4). The kinetics of replication were monitored by enzyme immunoassay (EIA-HAVAg) and cDNA-RNA dot blot hybridization. The maximum level of RNA, which was observed 21 days post-infection (p.i.) during the 3rd passage, when HAVAg was still undetectable by EIA, served as a basis to establish subsequent passages every 21 days p.i. This schedule of passage resulted in a progressive reduction of time between culture infection and HAVAg and RNA production, together with an enhancement in antigen titer content of cell lysates. During the 7th passage, maximum HAVAg and RNA levels were detected at 7 days. Fourteen days after the 8th passage, clear morphological modifications appeared, suggesting a good adaptation of HAF-203 to FRhK-4 cells. Obtaining a fast-growing Brazilian HAV is very important for the development of vaccines