Virus In Peripheral Blood Mononuclear Cells Research Articles

93. Novel recombinant measles virus vaccine prevents measles virus disease in rhesus macaques

Abstract Background Measles-Mumps-Rubella (MMR) is an effective live-virus vaccine against measles virus (MeV). However, use of MMR is limited by its inability to boost MeV immunity, lack of immunogenicity in infants, and contraindication in pregnant and immunocompromised persons. Methods We evaluated a novel recombinant dimeric MeV hemagglutinin protein vaccine (rMeV) in a rhesus macaque model. Sixteen macaques were primed at day 0 and boosted at day 42 by experimental group: 1) MMR x2; 2) rMeV x2; 3) MMR prime/rMeV boost; 4) control; n=4. Macaques were challenged intratracheally with Bilthoven strain wild type MeV 8 months later. Blood, bone marrow (BM), and lymph node (LN) samples were collected over 3–28 days after challenge. Replication-competent MeV was measured in peripheral blood mononuclear cells (PBMC), BM cells, and LN cells by infectious assay; MeV RNA in PBMC and BM cells was determined by quantitative reverse transcriptase polymerase chain reaction. Plasma was evaluated for MeV-specific IgG and plaque reduction neutralization titer (PRNT). Results Six months after vaccination, mean PRNT was 2,432 in rMeV x2 (standard deviation (SD) 3,840), 3,584 in MMR-prime/rMeV boost (SD 3,072) and 5,120 in MMR x2 groups (SD 3,547). Upon infectious challenge, macaques who received any MeV-containing vaccine developed no clinical signs of measles and had no detectable infectious virus in PBMC, BM cells, or LN cells. All unvaccinated macaques had virus in PBMC that peaked at day 7 (mean 3,162 TCID50/mL, SD 4.1) and resolved by day 14 post challenge, and one macaque developed an extensive rash. Macaques who received any MeV-containing vaccine had no detectable MeV RNA in PBMC or BM cells, whereas all unvaccinated macaques had detectable MeV RNA that peaked at day 7 (1.6e5 copies, SD 10.5) in PBMC. In all experimental groups, MeV-specific IgG titers increased after MeV challenge. Conclusion Macaques who received rMeV and/or MMR were protected from rash, viremia, and detection of MeV RNA in PBMC and BM cells, unlike unvaccinated macaques. These data suggest that rMeV vaccine generates protective immune responses against measles and may be a novel candidate for future measles vaccine strategies. Study of cellular responses after rMeV vaccination and MeV challenge is warranted. Disclosures Jessica Rubens, MD, Mevox: Grant/Research Support Guillaume Stewart-Jones, PhD, Moderna: Inventor of SARS-CoV-2 vaccine sequences|Moderna: Stocks/Bonds Michael Watson, MD, MEVOX Ltd: Board Member|MEVOX Ltd: Ownership Interest|MEVOX Ltd: Stocks/Bonds Barney S. Graham, MD, PhD, BSG: BSG is an inventor on patents for the stabilization of the RSV F protein (WO2014160463A1, Prefusion RSV F proteins and their use).|National Institutes of Health: Inventor on patents for RSV vaccines|National Institutes of Health: inventor on patents for measles and other paramyxovirus vaccines Diane Griffin, MD PhD, Gilead: Grant/Research Support|GlaxoSmithKline: Advisor/Consultant|GreenLight Biosciences: Advisor/Consultant|Merck: Advisor/Consultant|MeVox: Grant/Research Support|Takeda Pharmaceuticals: Advisor/Consultant.

HPLC-Based Purification and Isolation of Potent Anti-HIV and Latency Reversing Daphnane Diterpenes from the Medicinal Plant Gnidia sericocephala (Thymelaeaceae).

Despite the success of combination antiretroviral therapy (cART), HIV persists in low- and middle-income countries (LMIC) due to emerging drug resistance and insufficient drug accessibility. Furthermore, cART does not target latently-infected CD4+ T cells, which represent a major barrier to HIV eradication. The “shock and kill” therapeutic approach aims to reactivate provirus expression in latently-infected cells in the presence of cART and target virus-expressing cells for elimination. An attractive therapeutic prototype in LMICs would therefore be capable of simultaneously inhibiting viral replication and inducing latency reversal. Here we report that Gnidia sericocephala, which is used by traditional health practitioners in South Africa for HIV/AIDS management to supplement cART, contains at least four daphnane-type compounds (yuanhuacine A (1), yuanhuacine as part of a mixture (2), yuanhuajine (3), and gniditrin (4)) that inhibit viral replication and/or reverse HIV latency. For example, 1 and 2 inhibit HIV replication in peripheral blood mononuclear cells (PBMC) by >80% at 0.08 µg/mL, while 1 further inhibits a subtype C virus in PBMC with a half-maximal effective concentration (EC50) of 0.03 µM without cytotoxicity. Both 1 and 2 also reverse HIV latency in vitro consistent with protein kinase C activation but at 16.7-fold lower concentrations than the control prostratin. Both 1 and 2 also reverse latency in primary CD4+ T cells from cART-suppressed donors with HIV similar to prostratin but at 6.7-fold lower concentrations. These results highlight G. sericocephala and components 1 and 2 as anti-HIV agents for improving cART efficacy and supporting HIV cure efforts in resource-limited regions.

Discordance between HIV-1 Population in Plasma at Rebound after Structured Treatment Interruption and Archived Provirus Population in Peripheral Blood Mononuclear Cells.

ABSTRACTAntiretroviral therapy (ART) can sustain the suppression of plasma viremia to below detection levels. Infected individuals undergoing a treatment interruption exhibit rapid viral rebound in plasma viremia which is fueled by cellular reservoirs such as CD4+ T cells, myeloid cells, and potentially uncharacterized cellular sources. Interrogating the populations of viruses found during analytical treatment interruption (ATI) can give insights into the biologically competent reservoirs that persist under effective ART as well as the nature of the cellular reservoirs that enable viral persistence under ART. We interrogated plasma viremia from four rare cases of individuals undergoing sequential ATIs. We performed next-generation sequencing (NGS) on cell-associated viral DNA and cell-free virus to understand the interrelationship between sequential ATIs as well as the relationship between viral genomes in circulating peripheral blood mononuclear cells (PBMCs) and RNA from rebound plasma. We observed population differences between viral populations recrudescing at sequential ATIs as well as divergence between viral sequences in plasma and those in PBMCs. This indicated that viruses in PBMCs were not a major source of post-ATI viremia and highlights the role of anatomic reservoirs in post-ATI viremia and viral persistence.IMPORTANCE Even with effective ART, HIV-1 persists at undetectable levels and rebounds in individuals who stop treatment. Cellular and anatomical reservoirs ignite viral rebound upon treatment interruption, remaining one of the key obstacles for HIV-1 cure. To further examine HIV-1 persistence, a better understanding of the distinct populations that fuel viral rebound is necessary to identify and target reservoirs and the eradication of HIV-1. This study investigates the populations of viruses found from proviral genomes from PBMCs and plasma at rebound from a unique cohort of individuals who underwent multiple rounds of treatment interruption. Using NGS, we characterized the subtypes of viral sequences and found divergence in viral populations between plasma and PBMCs at each rebound, suggesting that distinct viral populations appear at each treatment interruption.

BVDV permissiveness and lack of expression of co-stimulatory molecules on PBMCs from calves pre-infected with BVDV

Bovine viral diarrhea virus (BVDV) has been detected in peripheral blood mononuclear cells (PBMCs) of immunocompetent animals, not being clear whether the development of a specific humoral immune response can prevent BVDV infection. The aim of this study was to evaluate the ability of non-cytopathic BVDV to replicate and produce infectious virus in PBMCs from calves pre-infected with BVDV and to elucidate the immunomodulatory effect of BVDV on these cells in an in vitro model. Quantification of virus was by quantitative PCR, while its replicative capacity and shedding into the extracellular environment was evaluated by viral titration. Apoptosis was assessed by flow cytometry analysis of annexin V and propidium iodide, and by expression of caspase-3/7. Flow cytometry was used to analyze the expression of CD14/CD11b/CD80, CD4/CD8/CD25, MHC-I/MHC-II and B-B2 markers. Our results showed that PBMCs from cattle naturally infected with BVDV were more susceptible to in vitro BVDV infection and showed a more severe apoptosis response than those from naïve animals. Non-cytopathic BVDV in vitro infection also resulted in a lack of effect in the expression of antigen presentation surface markers. All these findings could be related to the immunosuppressive capacity of BVDV and the susceptibility of cattle to this infection.

Evolutional conservation of molecular structure and antiviral function of a type I interferon, IFN-kappa, in poultry

IFN-kappa (IFN-κ) is a type I IFN expressed by keratinocytes, monocytes and dendritic cells with important roles during the innate immune response period. This research was conducted to elaborate the evolution and characteristics of IFN-κ in poultry. Chicken IFN-κ is located on the sex-determining Z chromosome, which is greatly different from mammals. Poultry IFN-κ cluster together in a species-specific manner through positive selection pressure and share only 19–33% homology with mammalian IFN-κ and poultry other type I IFN. Both chicken and duck IFN-κ was constitutively expressed in spleen, skin, lung, and peripheral blood mononuclear cells (PBMC), as well as being significantly induced after treatment with virus in PBMC. Biologically, poultry IFN-κ has antiviral activity against VSV in chicken embryonic fibroblasts and duck embryonic fibroblasts (CEF and DEF) cells, and induces the expression of IFN stimulated genes (ISGs). After treatment with JAK1 inhibitor, the ISGs expression can be down-regulated. Overall, our research on poultry IFN-κ not only enriches the knowledge about IFN-κ but also facilitates further research on the role of type I IFNs in antiviral defense responses in poultry.

Dissecting the contribution of EBNA3C domains important for EBV-induced B-cell growth and proliferation.

Epstein-Barr virus (EBV) is an oncogenic gammaherpes virus which is linked to pathogenesis of several human lymphatic malignancies. The EBV essential latent antigen EBNA3C is critical for efficient conversion of primary human B-lymphocytes to lymphoblastic cell lines and for continued LCL growth. EBNA3C, an EBV latent antigen with oncogenic potential can bind and regulate the functions of a wide range of cellular transcription factors. In our current reverse genetics study, we deleted the full length EBNA3C, and independently the RBP-Jκ and Nm23-H1 binding sites within EBNA3C using BACmid recombinant engineering methodology. Our experiments demonstrated that deletion of the EBV EBNA3C open reading frame (ORF) and more specifically the residues 621-675 which binds Nm23H1 and SUMO-1 showed a significant reduction in the ability of the cells to proliferate. Furthermore, they exhibited lower infectivity of human peripheral blood mononuclear cells (PBMCs). We also showed that recombinant EBV with deletions of the EBNA3C ORF, as well as a recombinant with residues 621-675 within EBNA3C ORF deleted had diminished abilities to activate CD40. Our study also revealed that the full length (1-992) and 621-675 aa deletions of EBNA3C when compared to wild type EBV infected PBMCs had differential expression patterns for the phosphorylation of MAP kinases specifically p38, JNK and ERK. Regulation of β-catenin also differed among wild type and EBNA3C deleted mutants. These temporal differences in signaling activities of these recombinant viruses in PBMCs is likely important in defining their functional importance in EBV-mediated B-cell transformation.

Peripheral blood mononuclear cells from field cattle immune to bovine viral diarrhea virus (BVDV) are permissive in vitro to BVDV.

The aim of this study was to determine the in vitro permissivity of peripheral blood mononuclear cells (PBMCs) from bovine viral diarrhea virus (BVDV)-immune field cattle to homologous and heterologous BVDVs. PBMCs from seventeen BVDV-naïve and sixteen BVDV-immune animals were infected with noncytopathic BVDV-1 or BVDV-2. The immune status of cattle was indicated by the presence of virus neutralizing antibodies, while viral load of PBMCs was determined by real-time RT-PCR. The results revealed that the PBMCs from naïve or immune animals were permissive to either BVDV-1 or BVDV-2, but the viral load was significantly higher for the naïve than for the immune animals. Furthermore, the load of homologous virus in PBMCs from immune animals was lower than that of heterologous virus. Our results provide evidence that the PBMCs from BVDV-immune cattle in field are susceptible to reinfection with homologous or heterologous BVDV, albeit to a lower extent in the former case.

HIV Coreceptor Tropism in Paired Plasma, Peripheral Blood Mononuclear Cell, and Cerebrospinal Fluid Isolates from Antiretroviral-Naïve Subjects

A survey of HIV coreceptor usage in cerebrospinal fluid (CSF) samples, peripheral blood mononuclear cells (PBMCs), and plasma samples from naïve seropositive patients was conducted. One hundred patients were enrolled in this study. Of the 100 patients, 36 had a primary or recent infection (P-RI), 31 had an early chronic infection (>350 CD4 cells) (ECI), and 33 had a late chronic infection (LCI). All 3 compartments were sampled in a subset of 33 participants, while the remaining 67 patients provided plasma samples and PBMCs only. Seventy-seven patients harbored the R5 virus in plasma samples and had a significantly higher median and percentage of CD4(+) T cells than patients with X4 virus (437 and 281 cells/μl, respectively; P = 0.0086; 20.6% and 18.6%, respectively). The X4 strain was detected more frequently in patients with LCI than in patients with P-RI or ECI (39.3%, 19.4%, and 9.6%, respectively; P = 0.0063). PBMC and plasma tropism was concordant in 90 patients, and 73 had the R5 strain. Among patients with discordant results, 4 had the R5 virus in their plasma and the X4 virus in PBMCs; 6 showed the opposite profile. Plasma, PBMC, and CSF tropism determinations were concordant in 26/33 patients (21 patients had R5, and 5 had X4). The tropism was discordant in 5/33 patients, with the X4 virus in plasma and R5 in CSF; the HIV tropism in PBMCs was X4 in 3 patients. The remaining 2/33 patients had the R5 virus in plasma and PBMCs and the X4 virus in CSF; one of these patients had a P-RI. The discordant tropism in CSF and blood may have implications for chemokine (C-C motif) receptor 5 (CCR5) antagonist use in patients with limited response to antiretroviral therapy (ART) or in responding patients evaluated for simplification of treatment.

Various Viral Compartments in HIV-1-Infected Mothers Contribute toIn UteroTransmission of HIV-1

Perinatal HIV transmission occurs in utero or intrapartum. The mechanisms and timing of transmission are not clearly understood. To compare the genetic sequences of the V3 envelope region of infant's plasma HIV to that of the mother's plasma, peripheral blood mononuclear cells (PBMC) and vaginal secretions, and correlate with timing of transmission. All 3 infants had a positive HIV PCR in the first days of life, thus classified as in utero infections. In the first mother-infant pair, two different variants were present in the infant, one correlating with maternal PBMC virus and highly homologous to virus from vaginal secretions and the other identical to sequences in maternal plasma. In the second pair, the infant plasma virus was similar to that of maternal PBMC. In the third pair, the cord blood and infant plasma virus were highly similar to maternal vaginal virus. The presence of more than one HIV variant from the maternal blood and from the vaginal compartment in the cord blood of infants presumably infected in utero could point to more than one episode of transmission or, alternatively, to transmission of PBMC virus.

High frequency of X4/DM-tropic viruses in PBMC samples from patients with primary HIV-1 subtype-B infection in 1996-2007: the French ANRS CO06 PRIMO Cohort Study

To estimate the frequency of viruses with X4 or dual-X4/DM tropism from peripheral blood mononuclear cells (PBMCs) of 390 human immunodeficiency virus type 1 (HIV-1) subtype-B patients diagnosed at the time of primary HIV-1 infection (PHI) between 1996 and 2007 and enrolled in the PRIMO Cohort. V3 loop sequences were amplified from HIV-1-DNA and analysed with a combination of five genotypic rules to predict tropism: (i) the '11/25 rule'; (ii) the net charge rule; (iii) the PSSM(X4/DM) algorithm; (iv) the PSSM(SI/NSI) algorithm; and (v) the SVM(Geno2pheno) algorithm. A high proportion (62/390, 15.9%) of patients harboured X4/DM-tropic viruses. This prevalence was stable over time: 18.1% before 2003 versus 14.8% since 2003. No difference according to HIV tropism was noted in HIV-RNA levels, CD4 cell count, time between infection and enrolment, and HIV infection risk factor. The frequency of X4/DM-tropic virus was similar among patients infected with a resistant virus (12/62, 19.4%) compared with patients harbouring wild-type strains (50/328, 15.2%). This large French epidemiological study evidenced a high proportion of patients (15.9%) harbouring X4/DM-tropic viruses in PBMCs at the time of PHI, suggesting the existence of a cellular X4/DM viral reservoir that could persist for lengthy period of time. Several reports identified that HIV-1 CXCR4 usage was more frequent among patients who developed AIDS and was a powerful predictor of the response to antiretrovirals. Further studies are needed to evaluate the impact of such strains on the outcome of HIV disease, when they are detected at the time of primary infection.

Suppression of HIV Replication In Vitro by CpG and CpG Conjugated to the Non Toxic B Subunit of Cholera Toxin

Administration of oligodeoxynucleotides (ODNs) containing CpG motifs generates a rapid and potent response of CC-chemokines, known as ligands of the HIV-1 co-receptor CCR5, in the murine female genital tract. The present study explored the potential HIV inhibitory activities of different human CpG prototypes either alone or conjugated to the non-toxic subunit of cholera toxin (CTB). Results showed that in vitro replication of both HIV-1 and HIV-2 can be suppressed by different human CpG prototypes. Importantly, the conjugation of CpG ODN to CTB (CTB-CpG) enhanced the antiviral activity of CpG against primary HIV-1 isolates of both R5 and X4 phenotypes in peripheral blood mononuclear cells (PBMC) as well as U87.CD4 co-receptor indicator cells. CTB-CpGs triggered higher amounts of MIP-1alpha, and MIP-1beta in PBMC than the corresponding CpG ODNs, which may explain the superior antiviral effect of CTB-CpG against R5 virus in PBMC. Incubation of PBMC with CpG ODN and CTB-CpG did not alter surface expression of HIV-1 receptors indicating that the observed anti-HIV-1 effect is not mediated through down regulation of HIV-1 receptors on target cells. Further, the enhanced antiviral effect of CTB-CpG was dependent on the presence of phosphorothioate backbone in the ODN, whereas the presence of CpG motif in ODNs was dispensable. These results have implications for the development of novel intervention strategies to prevent HIV infection.

Phylodynamics of HIV-1 in Lymphoid and Non-Lymphoid Tissues Reveals a Central Role for the Thymus in Emergence of CXCR4-Using Quasispecies

BackgroundDuring HIV-1 infection coreceptor switch from CCR5- (R5)- to CXCR4 (X4)-using viruses is associated with disease progression. X4 strains of HIV-1 are highly cytopathic to immature thymocytes. Virtually no studies have evaluated the HIV-1 quasispecies present in vivo within thymic and lymphoid tissues or the evolutionary relationship between R5 and X4 viruses in tissues and peripheral blood.Methodology/Principal FindingsHigh-resolution phylodynamic analysis was applied to virus envelope quasispecies in longitudinal peripheral blood mononuclear cells (PBMCs) and lymphoid and non-lymphoid tissues collected post mortem from therapy naïve children with AIDS. There were three major findings. First, continued evolution of R5 viruses in PBMCs, spleen and lymph nodes involved multiple bottlenecks, independent of coreceptor switch, resulting in fitter quasispecies driven by positive selection. Second, evolution of X4 strains appeared to be a sequential process requiring the initial fixation of positively selected mutations in V1-V2 and C2 domains of R5 variants before the emergence of high charge V3 X4 variants. Third, R5 viruses persisted after the emergence of CXCR4-using strains, which were found predominantly but not exclusively in the thymus.Conclusions/SignificanceOur data indicate that the evolution of X4 strains is a multi-step, temporally structured process and that the thymus may play an important role in the evolution/amplification of coreceptor variants. Development of new therapeutic protocols targeting virus in the thymus could be important to control HIV-1 infection prior to advanced disease.

HIV-1 Variants from a Perinatal Transmission Pair Demonstrate Similar Genetic and Replicative Properties in Tonsillar Tissues and Peripheral Blood Mononuclear Cells

Human immunodeficiency virus type 1 (HIV-1) can be acquired through oropharyngeal tissues in breastfeeding infants. Efforts to better understand the determinants of breast milk transmission are hampered by the lack of a relevant oral human mucosa model and well-defined breast milk-derived viruses. This study used human ex vivo palatine tonsil tissues and peripheral blood mononuclear cells (PBMCs) to characterize the genetic, biological, and replicative properties of HIV-1 variants obtained from a perinatal transmission pair. Unique viral populations from maternal breast milk and infant blood were identified by gp120 V1-V2- and V3-specific heteroduplex tracking assays (HTAs). Full-length infectious recombinant viruses, containing a common HIV-1 NL4-3 genetic background, were generated with V1-V3 gp120 fragments from maternal and infant isolates representing the major viral populations identified in the HTAs. The resulting recombinant viruses used the CCR5 coreceptor, were nonsyncytium forming, and demonstrated replication properties similar to those of parental and control viruses in PBMCs and tonsillar explants. These findings indicate that viruses from breast milk cells and infant blood can infect PBMCs and tonsil tissues. The maternal and infant HIV-1 viruses detailed here will provide useful tools for defining the viral and host factors that contribute to HIV breastfeeding transmission.

In vitro activation of feline immunodeficiency virus in ramified microglial cells from asymptomatically infected cats.

Intravenous infection of cats with feline immunodeficiency virus was used as a model system to study activation of virus replication in brain-resident microglial cells in vitro. Virus release by ramified microglial cells isolated from subclinically infected animals was detectable in cell-free tissue culture supernatant only by reverse transcription and nested PCR of gag-specific RNA sequences and not by virion-associated reverse transcriptase activity. In contrast, cocultivation of in vivo-infected microglial cells with mitogen-activated peripheral blood mononuclear cells (PBMC) regularly allows detection of high virus yields in cell-free tissue culture fluid. Besides uptake and multiplication of microglia-derived virus in PBMC, release of virus from microglia is stimulated by cell contact with PBMC. The data suggest that T lymphocytes patrolling the central nervous system could reactivate the semilatent state of lentiviruses in microglial cells in the course of clinically silent central nervous system infection.

Quantitative assessment of hepatitis C virus RNA in peripheral blood mononuclear cells during therapy with interferon-α2a

A significant number of patients with hepatitis C (HCV) treated with interferon (IFN) will initially clear their serum of HCV RNA, but will then have recurrence of viraemia either during or after therapy. One proposed mechanism for relapse is that HCV may persist in peripheral blood mononuclear cells (PBMCs) and that the PBMCs serve as a 'viral reservoir' that is resistant to IFN. To address this hypothesis, we performed serial, quantitative polymerase chain reaction (PCR) of HCV RNA in serum and PBMCs from 26 consecutive patients treated with IFN-alpha2a. Of the 26 patients, 11 (42%) did not clear virus from their serum during therapy and were termed non-responders. Five patients (19%) had sustained clearance of virus from serum and were termed complete responders. The remaining 10 patients (39%) initially eliminated HCV RNA from their serum, but had relapse of viraemia. They were termed partial responders. In all 10 partial responders HCV RNA was undetectable in PBMCs at the same time that it was undetectable in serum. When virus recurred in serum, it was preceded by or occurred at the same time as the return of virus in PBMCs. The results of our study indicate that PBMCs did not serve as an IFN-resistant 'viral reservoir' during therapy. Partial responders who transiently cleared virus from serum also cleared virus from PBMCs and the presence or titre of HCV RNA in PBMCs at the initiation of therapy did not predict response to therapy.

Quantification of human immunodeficiency virus type 1-infected mononuclear cells in peripheral blood of seropositive subjects by newly developed flow cytometry analysis of the product of an in situ PCR assay

The presence of human immunodeficiency virus type 1 (HIV-1) proviral DNA in peripheral blood mononuclear cells (PBMC) of three groups (group 1, more than 500 CD4+ T cells per microliter; group 2, between 200 and 499 CD4+ T cells per microliter; group 3, fewer than 200 CD4+ T cells per microliter) of HIV-1-infected patients, in different stages of the disease, was determined by using a newly developed flow cytometry analysis of the product of in situ PCR assay and compared with other markers of viral replication (HIV-1 p24 antigenemia and viral isolation). Results showed varied percentages of HIV-1-infected PBMC, ranging from 0.6 to 20%. Patients with more than 500 CD4+ T cells per microliter showed the lowest percentage of HIV-1-infected PBMC (2.1 +/- 1.7), compared with patients with CD4+ T-cell counts of between 200 and 499 per microliter (6.5% +/- 4.1%; P < 0.001) and patients with fewer than 200 CD4+ T cells per microliter (4.9% +/- 4.7%; P < 0.05). The difference in the percentage of HIV-1-infected PBMC between group 2 and group 3 patients may in part reflect the loss of CD4+ T lymphocytes in more advanced stages of the disease. However, the results clearly indicate a striking coincidence between the fall of the CD4+ T-cell count below 400/microliter and the sharp increase in PBMC virus loading and p24 antigenemia. Since the procedure is relatively easy to perform, it could be used to monitor the evolution of HIV-1 infection and may prove a useful adjunct in tailoring therapeutic strategies.

Feline immunodeficiency virus: quantification in peripheral blood mononuclear cells and isolation from plasma of infected cats.

The titer of feline immunodeficiency virus in peripheral blood mononuclear cells (PBMC) and the presence of infectious virus in plasma was investigated over 20 week period in 8 experimentally infected cats, 3 uninfected cats and 2 naturally infected cats by end point dilution cultures using a feline T-lymphoblastoid cell line (MYA-1). FIV was isolated from PBMC of all infected cats, but not from the uninfected cats. FIV was also isolated consistently from 100 microliters plasma from most of the experimentally infected cats, but not from the 2 naturally infected cats. The virus titer in PBMCs in both experimentally and naturally infected cats was comparatively high, ranging from 10 TCID/10(6) PBMC to 14,286 TCID/10(6) PBMC. The titers in PBMC of individual cats remained unchanged or varied only slightly over the 20 week period. In contrast, the titers varied substantially between cats, with significantly higher titers in the youngest litter (4 cats) than in the oldest litter (3 cats). This suggests that there is an age-related factor influencing the level of PBMC virus titers in experimental infection with FIV. A similar age-related susceptibility has been shown with feline leukemia virus. More importantly, the sustained titers in the experimentally infected cats bear close resemblance to infection of children with human immunodeficiency virus. These data reinforce suggestions that age and immune maturity have a fundamental influence on PBMC and plasma titers in lentivirus infections.