Broad spectrum antiviral fractions from the lichen Ramalina farinacea (L.) Ach.

BACKGROUND: An ethylacetate-soluble fraction (ET4) from the lichen Ramalina farinacea has previously been shown to inhibit the infectivity of lentiviral and adenoviral vectors, as well as wild-type HIV-1. We now determined the antiviral activity of ET4 against other wild-type viruses, including the herpes simplex virus type 1 (HSV-1) and the respiratory syncytial virus (RSV). METHODS: Wild-type HIV-1, HSV-1 or RSV were pre-incubated with various concentrations of ET4 for 30 min at 37 degrees C before adding to P4CCR5 indicator cell line (HIV-1), ELVIS TM indicator cell line (HSV-1) or HEp2 cell line (RSV) in 96-well microtitre plates. Controls contain virus alone without ET4. The anti-HIV and anti-HSV activities were quantified by estimating beta-galactosidase expression of the respective indicator cell lines while the anti-RSV activity was determined via an immunofluorescent technique, employing monoclonal mouse antibody against the P-protein of RSV. Toxicity of ET4 to cell lines was evaluated in parallel using either the BrdU incorporation method or the MTT method. The effect of ET4 on the enzymatic activity of HIV-1 reverse transcriptase was also evaluated using a chemiluminescent reverse transcriptase assay. Bioassay-guided fractionation of the whole methanol extract of R. farinacea involved sequential screening of HPLC fractions using a vector-based assay technique. RESULTS: ET4 inhibited HSV-1 and RSV potently (IC(50)=6.09 and 3.65 microg/ml, respectively). Time-of-addition studies suggest that both entry and post-entry steps of the HIV-1 replication cycle and the entry step of the RSV replication cycle are targeted. Furthermore, ET4 inhibited HIV-1 reverse transcriptase with an IC(50) of 0.022 microg/ml. Bioassay-guided fractionation of ET4 led to the identification sub-fraction rfO, with activity against lentiviral vector and HIV-1 (RNA viruses) but not against HSV-1 (DNA virus) and sub-fraction rfM, with activity against HSV-1 but not against the lentiviral vector. CONCLUSIONS: ET4 represents a novel fraction from the lichen R. farinacea with broad spectrum antiviral activity against DNA viruses (adenovirus and HSV-1) and RNA viruses (HIV-1 and RSV). The effect against DNA and RNA viruses is mediated by different sub-fractions within R. farinacea.

Natural infection of wild-born mandrills (Mandrillus sphinx) with two different types of simian immunodeficiency virus.

We found a novel primate lentivirus in mandrill (Mandrillus sphinx). To clarify the evolutionary relationships and transmission patterns of human/simian immunodeficiency virus (HIV/SIV), we screened blood samples from 30 wild-born healthy Cameroonian mandrills. Five (16.7%) of them were seropositive for SIV. Three SIV strains were isolated from the five seropositive mandrills by cocultivation of their peripheral blood mononuclear cells (PBMCs) with PBMCs of rhesus macaques, a human T cell line (M8166), and/or a cynomolgus macaque T cell line (HSC-F). One of the newly isolated SIV strains was intravenously inoculated into two rhesus macaques and resulted in chronic infection. In the SIV-infected macaques at 45 weeks after inoculation, we observed a mild decline in the number of peripheral CD4(+) lymphocytes, lymphadenopathy, and blastic follicular dendritic cells with mild follicular hyperplasia in the peripheral lymph nodes. A phylogenetic analysis based on the pol sequence showed that the newly found SIVs from Cameroonian mandrills did not cluster with SIVmndGB1, which is the former representative strain of SIVmnd. The SIVmnds from Cameroon formed a new, independent lineage that branched before the root of the HIV-1/SIVcpz lineage with 996 of 1000 bootstrap replications. They clustered host specifically, and exhibited about 16.9% diversity at the level of nucleotide sequence among Cameroonian SIVmnd strains. These results indicate that the SIVmnds isolated in Cameroon are a novel type of SIVmnd and have infected Cameroonian mandrills for a long time. We therefore designated the Cameroonian SIVmnd as SIVmnd type 2 and redesignated SIVmndGB1 as SIVmnd type 1. To date, M. sphinx is the only primate species other than humans that is naturally infected with two different types of SIV.