month 12. Difference in measurable HIV DNA was statistically significant only when comparing seminal plasma to sperm fractions at week 0 (p=0.02). HIV RNA was detected in only 1/5 seminal plasma specimens with measurable HIV DNA at month 6 and in 0/3 seminal plasma specimens with measurable HIV DNA at month 12. Conclusions: HIV DNA was measurable in seminal plasma fractions at least to month 12 after ART initiation while HIV RNA became undetectable in the same specimens. Further study is necessary to characterize semen as a HIV reservoir and determine reactivation potential and transmissibility. 13 Memory CD4+ T cells expressing HLA-DR contribute to HIV persistence during prolonged ART E. Lee 1,2 , B. Hiener 1,2 , P. Bacchetti 3 , W. Shao 4 , E. Boritz 5 , D. Douek 5 , R. Fromentin 6 , T. Liegler 7 , S.G. Deeks 7 , F.M. Hecht 7 , J. Milush 7 , N. Chomont 6 , S. Palmer 1,2 1 The Westmead Institute for Medical Research, Centre for Virus Research, Westmead, Australia, 2 University of Sydney, Sydney Medical School, Sydney, Australia, 3 University of California, Department of Epidemiology and Biostatistics, San Francisco, United States, 4 Leidos Biomedical Research, Inc, Frederick National Laboratory for Cancer Research, Frederick, United States, 5 National Institutes of Allergy and Infectious Diseases, National Institutes of Health, Human Immunology Section, Vaccine Research Center, New York, United States, 6 Université de Montréal, Department of Microbiology, Infection and Immunology, Montreal, Canada, 7 University of California, Department of Medicine, San Francisco, United States Background: Most measurements of HIV reservoirs are performed using “resting” CD4+ T cells depleted for cells expressing HLA-DR, which is considered an activation marker. However, little is known about the possible role of these cells in HIV persistence during ART. Here, we examine the contribution of memory CD4+ T cells expressing HLA-DR to HIV persistence after prolonged ART (≥15 yrs). Methods: Using LTR-specific qPCR, HIV RNA and DNA were quantified in memory CD4+ T cells expressing HLA-DR or not from 6 participants (P1-P6) on ART (≥15 yrs). Using single-genome/proviral sequencing, we characterized HIV-RNA/DNA (p6-RT) sequences from HLA-DR+ and HLA-DR- memory T cells, central (CM), transitional (TM) and effector (EM) memory T cells. Clonal expansions were defined as ≥2 identical sequences in phylogenetic trees. Results: CD4+ HLA-DR+ and HLA-DR- memory cells contained a median of 3400 and 1000 HIV-RNA copies/million cells; and 36 and 60 HIV-DNA copies/million cells respectively, indicating the HIV transcriptional activity of HLA-DR+ cells is 6-fold higher than HLA-DR-. In participants treated during chronic infection, proportions of HIV-DNA clonal expansions were similar between HLA-DR+ and HLA-DR- memoryT cells but HIV-DNA sequences from EM were more often identical to HIV-DNA sequences from HLA-DR+ memory cells (43% P1, 88% P2) than CM and TM (8–18% P1; 8–44% P2; p<0.0001–0.03). For one participant treated during acute infection, approximately 50% of the sequenced HIV-DNA from HLA-DR- memory T cells, CM, TM, and EM was defective (hypermutated and/or containing stop codons) and 26% of HIV-RNA from HLA-DR- memory T cells was defective. However, the amount of defective HIV-DNA and RNA from HLA-DR+ memory T cells was much lower (21% and 0% respectively). Conclusions: These findings demonstrate that memory CD4+ T cells expressing HLA-DR contain persistent HIV-DNA and measurable HIV-RNA. Furthermore, these cells appear to contain more intact virus in the region sequenced than other cellular subsets. Therefore, measurements of the HIV reservoir during ART should include memory cells expressing activation markers, including HLA-DR. HLA-DR+ and EM cells had clonal expansions of genetically identical HIV-DNA, suggesting these cell subsets replenish the HIV reservoir through proliferation during prolonged ART. 14 Effect of substances of abuse on HIV DNA decay during antiretroviral therapy started during early HIV infection A. Chaillon, C. Anderson, S.J. Little, M.F. Oliveira, R.J. Ellis, D.M. Smith, S.R. Morris, S. Gianella University of California San Diego, La Jolla, United States Background: Substances of abuse are widely used among HIV- infected men who have sex with men (MSM), and are associated with T-cell activation, greater HIV transcription, and HIV disease progression. We hypothesize that drug use might influence HIV DNA decay after early initiation of antiretroviral therapy (ART). Methods: We investigated 552 peripheral blood mononuclear cell samples from 79 recently HIV-infected MSM from the San Diego Primary Infection Cohort. Participants started ART within a median of 4 months from the estimated date of infection (EDI) (IQR: 2–11) and achieved suppressed HIV RNA within a median of 3 months (IQR: 2–6). Participants were followed for a median of 29 months (IQR: 4–38) while on suppressive ART. Levels of CD4-associated HIV DNA and RNA (unspliced and multiply spliced) were measured by (dd)PCR. Substance use was defined as self-reported drug use within the last 3 months at enrollment (including methamphetamine, γ-hydroxybutyrate [GHB], cocaine, alkyl-nitrite [‘popper’], ecstasy, hallucinogens, opiates). Linear mixed-effect regression analysis was used to evaluate associations between use of drugs and changes in log10 HIV DNA levels or cellular HIV RNA transcription during ART. Additional tested variables included: age, nadir and current CD4 + T-cell counts, peak HIV RNA level, pre-ART HIV DNA level, time from EDI to ART initiation, time from ART initiation to virologic suppression. Results: Drug use was self-reported in 41 participants (52%) and included: methamphetamine (N=20), poppers (N=24), opiates (N=22), ecstasy (N=11), cocaine (N=6), and GHB (N=13). The median peak HIV RNA level was 5.6 log10 copies/mL (IQR: 5–6.3) and the median nadir CD4 + T-cell count/μl was 420 (IQR: 297–520) with no difference between groups. Following ART initiation, self-reported recreational use of poppers was the only variable associated with a significantly slower decay rate of log10HIV DNA (p=0.027). There was no difference in cellular HIV RNA transcription between popper users and non-users. Conclusions: Recreational use of poppers was frequent in our cohort and was the only factor associated with a slower decline of HIV DNA during suppressive ART. The mechanism underlying this association is unclear, but it is likely not attributable to less effective viral suppression on ART in users of poppers. 15 Is the lung a site of productive HIV infection that persists through ART? D. Russell 1 , D. Gludish 1 , K. Jambo 2 , H. Mwandumba 2 1 Cornell University, Ithaca, United States, 2 Malawi-Liverpool- Wellcome Research Program, Blantyre, Malawi Background: HIV persistence is maintained by viral reservoir(s) discrete from the peripheral viral population. These viral reservoirs are particularly significant during the rebound of peripheral viremia following failure of anti-retroviral therapy (ART). Using Fluorescent in situ Hybridization (FISH)-based detection system we demonstrated previously that chronically HIV-infected, ART-naïve individuals have virus in their lungs that is present predominantly in alveolar macrophages (AM). AM are long-lived cells that could represent an under-appreciated reservoir for HIV. 13 Journal of Virus Eradication 2016; 2 (Supplement 2): 8–30 POSTER PRESENTATIONS