Elucidation of the bottleneck to HIV-1 and SIV transmission at mucosal surfaces of the vagina, cervix and rectum, including a precise definition ofthe eariiest virus - host interactions necessary for infection, could be instrumental in the design of effective vaccines. Our laboratories have taken significant steps toward addressing these goals by developing an experimental framework with which to identify transmitted/founder HIV-1 and SIV genomes responsible for productive infection (Keele 2008; Keele 2009; Salazar 2009) and by developing in situ methods to characterize early virus - host cell interactions underlying mucosal transmission (Li 2005; Estes 2006; Estes 2007; Estes 2008; Li 2009a). The current project combines forthe first time ivag, ir and iv transmission studies using naturally-occurring SIVsm transmitted/founder viruses Project objectives are to characterize the rectal, vaginal and cervical bottleneck to SIVsmm transmission, to identify the initial cell types productively infected by transmitted/founder SIVsmm, and to identify early innate Immune responses that act to constrain or facilitate productive viral Infection. By examining the transmissibllity of molecularly-cloned transmitted/founder viruses from SIVsmm strains FTq, D215 and SL92b, along with SIVmac239 and infectious monkey plasmas containing complex SIVsmm quasispecies, we will test the following hypothesis: Distinct barriers to SIVsmm transmission exist at mucosal interfaces of the vagina, cervix and rectum. These barriers are both passive (sieving) and active in nature, the latter selecting for viruses with particular cellular tropism and replication properties necessary for transmission and productive infection. Specific Aims of the project are: (1) To quantify and characterize phylogenetically the bottleneck to rectal versus cervicovaginal versus intravenous transmission by naturally-occurring SIVsmm swarnis and by defined mixtures of moleculariy-cloned transmitted/founder viruses; (ii) To identify by in situ analysis of rectal, vaginal and cervical mucosa and submucosa the initial cell types that are infected by transmitted/founder SIVsmm viruses and to identify eariy innate immune responses that act to constrain or facilitate expansion of these foci of infection leading to productive clinical infection; (iii) To determine by in situ analysis, PCR-based single genome amplification and sequencing, and 454 pyrosequencing of SIVsm in mucosal, submucosal and lymphoreticular tissues and plasma, where in the transmission process the Drincioal Quantitative and qualitative bottlenecks to transmission and eariv virus replication occur.