title: Identification and characterization of HIV-1 latent viral reservoirs in peripheral blood.
abstract: Plasma viral load and CD4 counts are effective for clinical monitoring, but they do not give a full representation of HIV-1 quasispecies in cellular reservoirs, the major repository of replication-competent HIV-1 in infected individuals. We sought to develop a diagnostic system that might stimulate the replication-competent HIV-1 reservoirs for enhanced clinical monitoring, including selection of antiretroviral regimens. Whole-blood samples from 45 HIV-infected individuals were collected into 1 ViraStim HIV-1 activation tube and 1 EDTA tube. Samples were tested for viral load and cell type-specific HIV-1 replication. Further, 7 matched activated/nonactivated samples were sequenced using the Trugene HIV-1 genotyping kit. The percentage of patients with replication-competent virus in peripheral blood mononuclear cells (PBMCs) varied, depending on the baseline plasma viral load in the EDTA tubes. Six out of 24 patients with a starting plasma viral load of <20 copies/ml (cp/ml), 6 out of 8 patients with starting viral loads of >20 and <1,000 cp/ml, and 8 out of 13 patients with starting viral loads of >1,000 all showed increases in viral replication of >5-fold. These increases came from cellular reservoirs in blood as determined by simultaneous ultrasensitive subpopulation staining/hybridization in situ (SUSHI). When resistance genotypes in plasma from activation tubes were compared to those from EDTA tubes for 7 patients, all patients showed additional mutations in the activation tube, while 3 patients demonstrated additional genotypic resistance determinants. We show that HIV-1 viral replication can be stimulated directly from infected whole blood. The sequencing results showed that 3 of 7 cases demonstrated additional drug resistance following stimulation.
authors.affiliation: IncellDx, Inc., Menlo Park, California, USA.
authors.forename: Amanda
authors.initials: A
authors.lastname: Chargin
authors.affiliation: IncellDx, Inc., Menlo Park, California, USA.
authors.forename: Fangfang
authors.initials: F
authors.lastname: Yin
authors.affiliation: IncellDx, Inc., Menlo Park, California, USA.
authors.forename: Min
authors.initials: M
authors.lastname: Song
authors.affiliation: IncellDx, Inc., Menlo Park, California, USA.
authors.forename: Srividyabhuvaneswari
authors.initials: S
authors.lastname: Subramaniam
authors.affiliation: IncellDx, Inc., Menlo Park, California, USA.
authors.forename: Grace
authors.initials: G
authors.lastname: Knutson
authors.affiliation: IncellDx, Inc., Menlo Park, California, USA brucep@incelldx.com.
authors.forename: Bruce K
authors.initials: BK
authors.lastname: Patterson
mesh_terms: D023241:Antiretroviral Therapy, Highly Active; D018791:CD4 Lymphocyte Count; D015496:CD4-Positive T-Lymphocytes / Q000378:metabolism / Q000821:virology; D005838:Genotype; D015658:HIV Infections / Q000188:drug therapy / Q000382:microbiology*; D015497:HIV-1 / Q000145:classification / Q000502:physiology*; D006801:Humans; D007963:Leukocytes, Mononuclear / Q000378:metabolism / Q000821:virology*; D016865:Reassortant Viruses; D019562:Viral Load; D017735:Virus Latency; D014779:Virus Replication