Antibody-mediated depletion of select leukocyte subsets in blood and tissue of nonhuman primates.

Understanding the immunological control of pathogens requires a detailed evaluation of the mechanistic contributions of individual cell types within the immune system. While knockout mouse models that lack certain cell types have been used to help define the role of those cells, the biological and physiological characteristics of mice do not necessarily recapitulate that of a human. To overcome some of these differences, studies often look towards nonhuman primates (NHPs) due to their close phylogenetic relationship to humans. To evaluate the immunological role of select cell types, the NHP model provides distinct advantages since NHP more closely mirror the disease manifestations and immunological characteristics of humans. However, many of the experimental manipulations routinely used in mice (e.g., gene knock-out) cannot be used with the NHP model. As an alternative, the in vivo infusion of monoclonal antibodies that target surface proteins on specific cells to either functionally inhibit or deplete cells can be a useful tool. Such depleting antibodies have been used in NHP studies to address immunological mechanisms of action. In these studies, the extent of depletion has generally been reported for blood, but not thoroughly assessed in tissues. Here, we evaluated four depleting regimens that primarily target T cells in NHP: anti-CD4, anti-CD8α, anti-CD8ß, and immunotoxin-conjugated anti-CD3. We evaluated these treatments in healthy unvaccinated and IV BCG-vaccinated NHP to measure the extent that vaccine-elicited T cells - which may be activated, increased in number, or resident in specific tissues - are depleted compared to resting populations in unvaccinated NHPs. We report quantitative measurements of in vivo depletion at multiple tissue sites providing insight into the range of cell types depleted by a given mAb. While we found substantial depletion of target cell types in blood and tissue of many animals, residual cells remained, often residing within tissue. Notably, we find that animal-to-animal variation is substantial and consequently studies that use these reagents should be powered accordingly.

cMPL-Based Purification and Depletion of Human Hematopoietic Stem Cells: Implications for Pre-Transplant Conditioning.

The transplantation of gene-modified autologous hematopoietic stem and progenitor cells (HSPCs) offers a promising therapeutic approach for hematological and immunological disorders. However, this strategy is often limited by the toxicities associated with traditional conditioning regimens. Antibody-based conditioning strategies targeting cKIT and CD45 antigens have shown potential in mitigating these toxicities, but their long-term safety and efficacy in clinical settings require further validation. In this study, we investigate the thrombopoietin (TPO) receptor, cMPL, as a novel target for conditioning protocols. We demonstrate that high surface expression of cMPL is a hallmark feature of long-term repopulating hematopoietic stem cells (LT-HSCs) within the adult human CD34+ HSPC subset. Targeting the cMPL receptor facilitates the separation of human LT-HSCs from mature progenitors, a delineation not achievable with cKIT. Leveraging this finding, we developed a cMPL-targeting immunotoxin, demonstrating its ability to selectively deplete host cMPLhigh LT-HSCs with a favorable safety profile and rapid clearance within 24 hours post-infusion in rhesus macaques. These findings present significant potential to advance our understanding of human hematopoiesis and enhance the therapeutic outcomes of ex vivo autologous HSPC gene therapies.

Do organisms need an impact factor? Citations of key biological resources including model organisms reveal usage patterns and impact.

Research resources like transgenic animals and antibodies are the workhorses of biomedicine, enabling investigators to relatively easily study specific disease conditions. As key biological resources, transgenic animals and antibodies are often validated, maintained, and distributed from university based stock centers. As these centers heavily rely largely on grant funding, it is critical that they are cited by investigators so that usage can be tracked. However, unlike systems for tracking the impact of papers, the conventions and systems for tracking key resource usage and impact lag behind. Previous studies have shown that about 50% of the resources are not findable, making the studies they are supporting irreproducible, but also makes tracking resources difficult. The RRID project is filling this gap by working with journals and resource providers to improve citation practices and to track the usage of these key resources. Here, we reviewed 10 years of citation practices for five university based stock centers, characterizing each reference into two broad categories: findable (authors could use the RRID, stock number, or full name) and not findable (authors could use a nickname or a common name that is not unique to the resource). The data revealed that when stock centers asked their communities to cite resources by RRID, in addition to helping stock centers more easily track resource usage by increasing the number of RRID papers, authors shifted from citing resources predominantly by nickname (~50% of the time) to citing them by one of the findable categories (~85%) in a matter of several years. In the case of one stock center, the MMRRC, the improvement in findability is also associated with improvements in the adherence to NIH rigor criteria, as determined by a significant increase in the Rigor and Transparency Index for studies using MMRRC mice. From this data, it was not possible to determine whether outreach to authors or changes to stock center websites drove better citation practices, but findability of research resources and rigor adherence was improved.

The impact of IdeS (imlifidase) on allo-specific, xeno-reactive, and protective antibodies in a sensitized rhesus macaque model.

BACKGROUND: Highly sensitized patients face many barriers to kidney transplantation, including higher rates of antibody-mediated rejection after HLA-incompatible transplant. IdeS, an endopeptidase that cleaves IgG nonspecifically, has been trialed as desensitization prior to kidney transplant, and successfully cleaves donor-specific antibody (DSA), albeit with rebound. METHODS: IdeS was generated and tested (2 mg/kg, IV) in two naïve and four allosensitized nonhuman primates (NHP). Peripheral blood samples were collected at regular intervals following IdeS administration. Total IgG, total IgM, and anti-CMV antibodies were quantified with ELISA, and donor-specific antibody (DSA) and anti-pig antibodies were evaluated using flow cytometric crossmatch. B cell populations were assessed using flow cytometry. RESULTS: IdeS successfully cleaved rhesus IgG in vitro. In allosensitized NHP, robust reduction of total, DSA, anti-pig, and anti-CMV IgG was observed within one day following IdeS administration. Rapid rebound of all IgG antibody populations was observed, with antibody levels returning to baseline around day 14 post-infusion. Total IgM level was not affected by IdeS. Interestingly, a comparable reduction in antibody populations was observed after the second dose of IdeS. However, we have not observed any significant modulation of B cell subpopulations after IdeS. CONCLUSIONS: This study evaluated efficacy of IdeS in the allosensitized NHP in IgG with various specificities, mirroring antibody kinetics in human patients. The efficacy of IdeS on preexisting anti-pig antibodies may be useful in clinical xenotransplantation. However, given the limitation of IdeS on its durability as a monotherapy, optimization of IdeS with other agents targeting the humoral response is further needed.

SIV clearance from neonatal macaques following transient CCR5 depletion.

Treatment of people with HIV (PWH) with antiretroviral therapy (ART) results in sustained suppression of viremia, but HIV persists indefinitely as integrated provirus in CD4-expressing cells. Intact persistent provirus, the "rebound competent viral reservoir" (RCVR), is the primary obstacle to achieving a cure. Most variants of HIV enter CD4 + T cells by binding to the chemokine receptor, CCR5. The RCVR has been successfully depleted only in a handful of PWH following cytotoxic chemotherapy and bone marrow transplantation from donors with a mutation in CCR5 . Here we show that long-term SIV remission and apparent cure can be achieved for infant macaques via targeted depletion of potential reservoir cells that express CCR5. Neonatal rhesus macaques were infected with virulent SIVmac251, then treated with ART beginning one week after infection, followed by treatment with either a CCR5/CD3-bispecific or a CD4-specific antibody, both of which depleted target cells and increased the rate of plasma viremia decrease. Upon subsequent cessation of ART, three of seven animals treated with CCR5/CD3-bispecific antibody rebounded quickly and two rebounded 3 or 6 months later. Remarkably, the other two animals remained aviremic and efforts to detect replication-competent virus were unsuccessful. Our results show that bispecific antibody treatment can achieve meaningful SIV reservoir depletion and suggest that functional HIV cure might be achievable for recently infected individuals having a restricted reservoir.

Antibody-mediated depletion of select T cell subsets in blood and tissue of nonhuman primates.

Understanding the immunological control of pathogens requires a detailed evaluation of the mechanistic contributions of individual cell types within the immune system. While knockout mouse models that lack certain cell types have been used to help define the role of those cells, the biological and physiological characteristics of mice do not necessarily recapitulate that of a human. To overcome some of these differences, studies often look towards nonhuman primates (NHPs) due to their close phylogenetic relationship to humans. To evaluate the immunological role of select cell types, the NHP model provides distinct advantages since NHP more closely mirror the disease manifestations and immunological characteristics of humans. However, many of the experimental manipulations routinely used in mice (e.g., gene knock-out) cannot be used with the NHP model. As an alternative, the in vivo infusion of monoclonal antibodies that target surface proteins on specific cells to either functionally inhibit or deplete cells can be a useful tool. Such depleting antibodies have been used in NHP studies to address immunological mechanisms of action. In these studies, the extent of depletion has generally been reported for blood, but not thoroughly assessed in tissues. Here, we evaluated four depleting regimens that primarily target T cells in NHP: anti-CD4, anti-CD8α, anti-CD8ß, and immunotoxin-conjugated anti-CD3. We evaluated these treatments in healthy unvaccinated and IV BCG-vaccinated NHP to measure the extent that vaccine-elicited T cells - which may be activated, increased in number, or resident in specific tissues - are depleted compared to resting populations in unvaccinated NHPs. We report quantitative measurements of in vivo depletion at multiple tissue sites providing insight into the range of cell types depleted by a given mAb. While we found substantial depletion of target cell types in blood and tissue of many animals, residual cells remained, often residing within tissue. Notably, we find that animal-to-animal variation is substantial and consequently studies that use these reagents should be powered accordingly.

Genetic modifications designed for xenotransplantation attenuate sialoadhesin-dependent binding of human erythrocytes to porcine macrophages.

The phenomenon of diminishing hematocrit after in vivo liver and lung xenotransplantation and during ex vivo liver xenoperfusion has largely been attributed to action by resident liver porcine macrophages, which bind and destroy human erythrocytes. Porcine sialoadhesin (siglec-1) was implicated previously in this interaction. This study examines the effect of porcine genetic modifications, including knockout of the CMAH gene responsible for expression of Neu5Gc sialic acid, on the adhesion of human red blood cells (RBCs) to porcine macrophages. Wild-type (WT) porcine macrophages and macrophages from several strains of genetically engineered pigs, including CMAH gene knockout and several human transgenes (TKO+hTg), were incubated with human RBCs and "rosettes" (≥3 erythrocytes bound to one macrophage) were quantified by microscopy. Our results show that TKO+hTg genetic modifications significantly reduced rosette formation. The monoclonal antibody 1F1, which blocks porcine sialoadhesin, significantly reduced rosette formation by WT and TKO+hTg macrophages compared with an isotype control antibody. Further, desialation of human RBCs with neuraminidase before addition to WT or TKO+hTg macrophages resulted in near-complete abrogation of rosette formation, to a level not significantly different from porcine RBC rosette formation on porcine macrophages. These observations are consistent with rosette formation being mediated by binding of sialic acid on human RBCs to sialoadhesin on porcine macrophages. In conclusion, the data predict that TKO+hTg genetic modifications, coupled with targeting of porcine sialoadhesin by the 1F1 mAb, will attenuate erythrocyte sequestration and anemia during ex vivo xenoperfusion and following in vivo liver, lung, and potentially other organ xenotransplantation.

Plasmablast Expansion Following the Tetravalent, Live-Attenuated Dengue Vaccine Butantan-DV in DENV-Naïve and DENV-Exposed Individuals in a Brazilian Cohort.

An effective vaccine against the dengue virus (DENV) should induce a balanced, long-lasting antibody (Ab) response against all four viral serotypes. The burst of plasmablasts in the peripheral blood after vaccination may reflect enriched vaccine-specific Ab secreting cells. Here we characterize the acute plasmablast responses from naïve and DENV-exposed individuals following immunization with the live attenuated tetravalent (LAT) Butantan DENV vaccine (Butantan-DV). The frequency of circulating plasmablasts was determined by flow cytometric analysis of fresh whole blood specimens collected from 40 participants enrolled in the Phase II Butantan-DV clinical trial (NCT01696422) before and after (days 6, 12, 15 and 22) vaccination. We observed a peak in the number of circulating plasmablast at day 15 after vaccination in both the DENV naïve and the DENV-exposed vaccinees. DENV-exposed vaccinees experienced a significantly higher plasmablast expansion. In the DENV-naïve vaccinees, plasmablasts persisted for approximately three weeks longer than among DENV-exposed volunteers. Our findings indicate that the Butantan-DV can induce plasmablast responses in both DENV-naïve and DENV-exposed individuals and demonstrate the influence of pre-existing DENV immunity on Butantan DV-induced B-cell responses.

SOSIP Trimer-Specific Antibodies Isolated from a Simian-Human Immunodeficiency Virus-Infected Monkey with versus without a Pre-blocking Step with gp41.

BG505 SOSIP.664 (hereafter referred to as SOSIP), a stabilized trimeric mimic of the HIV-1 envelope spike resembling the native viral spike, is a useful tool for isolating anti-HIV-1 neutralizing antibodies. We screened long-term SHIV-AD8 infected rhesus monkeys for potency and breadth of serum neutralizing activity against autologous and heterologous viruses: SHIV-AD8, HIV-1 YU2, HIV-1 JR-CSF, and HIV-1 NL4-3. Monkey rh2436 neutralized all viruses tested and showed strong reactivity to the SOSIP trimer, suggesting this was a promising candidate for attempts at monoclonal antibody (MAb) isolation. MAbs were isolated by performing single B-cell sorts from peripheral blood mononuclear cells (PBMC) by FACS using the SOSIP trimer as a probe. An initial round of sorted cells revealed the majority of isolated MAbs were directed to the gp41 external domain portion of the SOSIP trimer and were mostly non-neutralizing against tested isolates. A second sort was performed, introducing a gp41 blocking step prior to PBMC staining and FACS sorting. These isolated MAbs bound SOSIP trimer but were no longer directed to the gp41 external domain portion. A significantly higher proportion of MAbs with neutralizing activity were obtained with this strategy. Our data show this pre-blocking step with gp41 greatly increases the yield of non-gp41-reactive, SOSIP-specific MAbs and increases the likelihood of isolating MAbs with neutralizing activity. IMPORTANCE Recent advancements in the field have focused on the isolation and use of broadly neutralizing antibodies for both prophylaxis and therapy. Finding a useful probe to isolate broad potent neutralizing antibodies while avoiding non-neutralizing antibodies is important. The SOSIP trimer has been shown to be a great tool for this purpose because it binds known broadly neutralizing antibodies. However, the SOSIP trimer can isolate non-neutralizing antibodies as well, including gp41-specific MAbs. Introducing a pre-blocking step with gp41 recombinant protein decreased the percent of gp41-specific antibodies isolated with SOSIP probe, as well as increased the number of neutralizing antibodies isolated. This method can be used as a tool to increase the chances of isolating neutralizing antibodies.

Plasmablast expansion following the tetravalent, live-attenuated dengue vaccine Butantan-DV in DENV-Naïve and DENV-exposed individuals in a Brazilian cohort

An effective vaccine against the dengue virus (DENV) should induce a balanced, long-lasting antibody (Ab) response against all four viral serotypes. The burst of plasmablasts in the peripheral blood after vaccination may reflect enriched vaccine-specific Ab secreting cells. Here we characterize the acute plasmablast responses from naïve and DENV-exposed individuals following immunization with the live attenuated tetravalent (LAT) Butantan DENV vaccine (Butantan-DV). The frequency of circulating plasmablasts was determined by flow cytometric analysis of fresh whole blood specimens collected from 40 participants enrolled in the Phase II Butantan-DV clinical trial (NCT01696422) before and after (days 6, 12, 15 and 22) vaccination. We observed a peak in the number of circulating plasmablast at day 15 after vaccination in both the DENV naïve and the DENV-exposed vaccinees. DENV-exposed vaccinees experienced a significantly higher plasmablast expansion. In the DENV-naïve vaccinees, plasmablasts persisted for approximately three weeks longer than among DENV-exposed volunteers. Our findings indicate that the Butantan-DV can induce plasmablast responses in both DENV-naïve and DENV-exposed individuals and demonstrate the influence of pre-existing DENV immunity on Butantan DV-induced B-cell responses.

Biophysical Evaluation of Rhesus Macaque Fc Gamma Receptors Reveals Similar IgG Fc Glycoform Preferences to Human Receptors.

Rhesus macaques are a common non-human primate model used in the evaluation of human monoclonal antibodies, molecules whose effector functions depend on a conserved N-linked glycan in the Fc region. This carbohydrate is a target of glycoengineering efforts aimed at altering antibody effector function by modulating the affinity of Fcγ receptors. For example, a reduction in the overall core fucose content is one such strategy that can increase antibody-mediated cellular cytotoxicity by increasing Fc-FcγRIIIa affinity. While the position of the Fc glycan is conserved in macaques, differences in the frequency of glycoforms and the use of an alternate monosaccharide in sialylated glycan species add a degree of uncertainty to the testing of glycoengineered human antibodies in rhesus macaques. Using a panel of 16 human IgG1 glycovariants, we measured the affinities of macaque FcγRs for differing glycoforms via surface plasmon resonance. Our results suggest that macaques are a tractable species in which to test the effects of antibody glycoengineering.

Measuring the Impact of Targeting FcRn-Mediated IgG Recycling on Donor-Specific Alloantibodies in a Sensitized NHP Model.

Background: In transplantation, plasmapheresis and IVIg provide the mainstay of treatment directed at reducing or removing circulating donor-specific antibody (DSA), yet both have limitations. We sought to test the efficacy of targeting the IgG recycling mechanism of the neonatal Fc receptor (FcRn) using anti-FcRn mAb therapy in a sensitized non-human primate (NHP) model, as a pharmacological means of lowering DSA. Methods: Six (6) rhesus macaque monkeys, previously sensitized by skin transplantation, received a single dose of 30mg/kg anti-RhFcRn IV, and effects on total IgG, as well as DSA IgG, were measured, in addition to IgM and protective immunity. Subsequently, 60mg/kg IV was given in the setting of kidney transplantation from skin graft donors. Kidney transplant recipients received RhATG, and tacrolimus, MMF, and steroid for maintenance immunosuppression. Results: Circulating total IgG was reduced from a baseline 100% on D0 to 32.0% (mean, SD ± 10.6) on d4 post infusion (p<0.05), while using a DSA assay. T-cell flow cross match (TFXM) was reduced to 40.6±12.5% of baseline, and B-cell FXCM to 52.2±19.3%. Circulating total IgM and DSA IgM were unaffected by treatment. Pathogen-specific antibodies (anti-gB and anti-tetanus toxin IgG) were significantly reduced for 14d post infusion. Post-transplant, circulating IgG responded to anti-FcRn mAb treatment, but DSA increased rapidly. Conclusion: Targeting the FcRn-mediated recycling of IgG is an effective means of lowering circulating donor-specific IgG in the sensitized recipient, although in the setting of organ transplantation mechanisms of rapid antibody rise post-transplant remains unaffected.

Antibody-based CCR5 blockade protects Macaques from mucosal SHIV transmission.

In the absence of a prophylactic vaccine, the use of antiretroviral therapy (ART) as pre-exposure prophylaxis (PrEP) to prevent HIV acquisition by uninfected individuals is a promising approach to slowing the epidemic, but its efficacy is hampered by incomplete patient adherence and ART-resistant variants. Here, we report that competitive inhibition of HIV Env-CCR5 binding via the CCR5-specific antibody Leronlimab protects rhesus macaques against infection following repeated intrarectal challenges of CCR5-tropic SHIVSF162P3. Injection of Leronlimab weekly at 10 mg/kg provides significant but partial protection, while biweekly 50 mg/kg provides complete protection from SHIV acquisition. Tissue biopsies from protected macaques post challenge show complete CCR5 receptor occupancy and an absence of viral nucleic acids. After Leronlimab washout, protected macaques remain aviremic, and adoptive transfer of hematologic cells into naïve macaques does not transmit viral infection. These data identify CCR5 blockade with Leronlimab as a promising approach to HIV prophylaxis and support initiation of clinical trials.

A comparative study of human-and rhesus-specific antithymocyte globulins in Rhesus macaques.

Rabbit antithymocyte globulin (RATG) preparations are widely used in transplantation. They are developed in vivo against thymocytes and contain polyclonal antibodies specific for myriad cellular targets. The rhesus monkey is commonly used as a preclinical transplant model, but the fidelity of commercially available human-specific RATGs to anticipate the effects of RATGs in rhesus has not been established. We therefore developed two rhesus-specific ATGs (rhATG) and compared them to human-specific RATG (huATG, Thymoglobulin® ) in rhesus monkeys, assessing the magnitude and phenotype of depletion peripherally and in lymph nodes. Four primates were assigned to each group and received 20 mg/kg of drug. Depletion, repopulation, and changes in lymphocyte subsets were evaluated in peripheral blood and lymph nodes by flow cytometry over four months. We observed similar qualitative changes in lymphocyte subsets, but a generally more profound depletion with huATG compared to either rhATG. Peripheral homeostatic proliferation rather than thymic output was the major mechanism for repopulation with all RATGs. Repopulation was slower but qualitatively similar when examining RATGs in additional animals receiving concomitant chronic immunosuppression. Depletional induction is similar to human- and rhesus-specific RATGs in rhesus macaques. Both rhesus- and human-specific agents appear appropriate for preclinical modeling of clinical RATG use.

Non-neutralizing Antibodies May Contribute to Suppression of SIVmac239 Viremia in Indian Rhesus Macaques.

The antiviral properties of broadly neutralizing antibodies against HIV are well-documented but no vaccine is currently able to elicit protective titers of these responses in primates. While current vaccine modalities can readily induce non-neutralizing antibodies against simian immunodeficiency virus (SIV) and HIV, the ability of these responses to restrict lentivirus transmission and replication remains controversial. Here, we investigated the antiviral properties of non-neutralizing antibodies in a group of Indian rhesus macaques (RMs) that were vaccinated with vif, rev, tat, nef, and env, as part of a previous study conducted by our group. These animals manifested rapid and durable control of viral replication to below detection limits shortly after SIVmac239 infection. Although these animals had no serological neutralizing activity against SIVmac239 prior to infection, their pre-challenge titers of Env-binding antibodies correlated with control of viral replication. To assess the contribution of anti-Env humoral immune responses to virologic control in two of these animals, we transiently depleted their circulating antibodies via extracorporeal plasma immunoadsorption and inhibition of IgG recycling through antibody-mediated blockade of the neonatal Fc receptor. These procedures reduced Ig serum concentrations by up to 80% and temporarily induced SIVmac239 replication in these animals. Next, we transferred purified total Ig from the rapid controllers into six vaccinated RMs one day before intrarectal challenge with SIVmac239. Although recipients of the hyperimmune anti-SIV Ig fraction were not protected from infection, their peak and chronic phase viral loads were significantly lower than those in concurrent unvaccinated control animals. Together, our results suggest that non-neutralizing Abs may play a role in the suppression of SIVmac239 viremia.

Zika virus infection during pregnancy protects against secondary infection in the absence of CD8+ cells.

While T cell immunity is an important component of the immune response to Zika virus (ZIKV) infection generally, the efficacy of these responses during pregnancy remains unknown. Here, we tested the capacity of CD8 lymphocytes to protect from secondary challenge in four macaques, two of which were depleted of CD8+ cells prior to rechallenge with a heterologous ZIKV isolate. The initial challenge during pregnancy produced transcriptional signatures suggesting complex patterns of immune modulation as well as neutralizing antibodies that persisted until rechallenge, which all animals efficiently controlled, demonstrating that the primary infection conferred adequate protection. The secondary challenge promoted activation of innate and adaptive immune cells, possibly suggesting a brief period of infection prior to clearance. These data confirm that ZIKV infection during pregnancy induces sufficient immunity to protect from a secondary challenge and suggest that this protection is not dependent on CD8 T cells.

Induction of Transient Virus Replication Facilitates Antigen-Independent Isolation of SIV-Specific Monoclonal Antibodies.

Structural characterization of the HIV-1 Envelope (Env) glycoprotein has facilitated the development of Env probes to isolate HIV-specific monoclonal antibodies (mAbs). However, preclinical studies have largely evaluated these virus-specific mAbs against chimeric viruses, which do not naturally infect non-human primates, in contrast to the unconstrained simian immunodeficiency virus (SIV)mac239 clone. Given the paucity of native-like reagents for the isolation of SIV-specific B cells, we examined a method to isolate SIVmac239-specific mAbs without using Env probes. We first activated virus-specific B cells by inducing viral replication after the infusion of a CD8ß-depleting mAb or withdrawal of antiretroviral therapy in SIVmac239-infected rhesus macaques. Following the rise in viremia, we observed 2- to 4-fold increases in the number of SIVmac239 Env-reactive plasmablasts in circulation. We then sorted these activated B cells and obtained 206 paired Ab sequences. After expressing 122 mAbs, we identified 14 Env-specific mAbs. While these Env-specific mAbs bound to both the SIVmac239 SOSIP.664 trimer and to infected primary rhesus CD4+ T cells, five also neutralized SIVmac316. Unfortunately, none of these mAbs neutralized SIVmac239. Our data show that this method can be used to isolate virus-specific mAbs without antigenic probes by inducing bursts of contemporary replicating viruses in vivo.

Vaccine protection against rectal acquisition of SIVmac239 in rhesus macaques.

A prophylactic vaccine against human immunodeficiency virus (HIV) remains a top priority in biomedical research. Given the failure of conventional immunization protocols to confer robust protection against HIV, new and unconventional approaches may be needed to generate protective anti-HIV immunity. Here we vaccinated rhesus macaques (RMs) with a recombinant (r)DNA prime (without any exogenous adjuvant), followed by a booster with rhesus monkey rhadinovirus (RRV)-a herpesvirus that establishes persistent infection in RMs (Group 1). Both the rDNA and rRRV vectors encoded a near-full-length simian immunodeficiency virus (SIVnfl) genome that assembles noninfectious SIV particles and expresses all nine SIV gene products. This rDNA/rRRV-SIVnfl vaccine regimen induced persistent anti-Env antibodies and CD8+ T-cell responses against the entire SIV proteome. Vaccine efficacy was assessed by repeated, marginal-dose, intrarectal challenges with SIVmac239. Encouragingly, vaccinees in Group 1 acquired SIVmac239 infection at a significantly delayed rate compared to unvaccinated controls (Group 3). In an attempt to improve upon this outcome, a separate group of rDNA/rRRV-SIVnfl-vaccinated RMs (Group 2) was treated with a cytotoxic T-lymphocyte antigen-4 (CTLA-4)-blocking monoclonal antibody during the vaccine phase and then challenged in parallel with Groups 1 and 3. Surprisingly, Group 2 was not significantly protected against SIVmac239 infection. In sum, SIVnfl vaccination can protect RMs against rigorous mucosal challenges with SIVmac239, a feat that until now had only been accomplished by live-attenuated strains of SIV. Further work is needed to identify the minimal requirements for this protection and whether SIVnfl vaccine efficacy can be improved by means other than anti-CTLA-4 adjuvant therapy.

Postnatal Zika virus infection of nonhuman primate infants born to mothers infected with homologous Brazilian Zika virus.

Recent data in a nonhuman primate model showed that infants postnatally infected with Zika virus (ZIKV) were acutely susceptible to high viremia and neurological damage, suggesting the window of vulnerability extends beyond gestation. In this pilot study, we addressed the susceptibility of two infant rhesus macaques born healthy to dams infected with Zika virus during pregnancy. Passively acquired neutralizing antibody titers dropped below detection limits between 2 and 3 months of age, while binding antibodies remained detectable until viral infection at 5 months. Acute serum viremia was comparatively lower than adults infected with the same Brazilian isolate of ZIKV (n = 11 pregnant females, 4 males, and 4 non-pregnant females). Virus was never detected in cerebrospinal fluid nor in neural tissues at necropsy two weeks after infection. However, viral RNA was detected in lymph nodes, confirming some tissue dissemination. Though protection was not absolute and our study lacks an important comparison with postnatally infected infants born to naïve dams, our data suggest infants born healthy to infected mothers may harbor a modest but important level of protection from postnatally acquired ZIKV for several months after birth, an encouraging result given the potentially severe infection outcomes of this population.

Travel Surveillance and Genomics Uncover a Hidden Zika Outbreak during the Waning Epidemic.

The Zika epidemic in the Americas has challenged surveillance and control. As the epidemic appears to be waning, it is unclear whether transmission is still ongoing, which is exacerbated by discrepancies in reporting. To uncover locations with lingering outbreaks, we investigated travel-associated Zika cases to identify transmission not captured by reporting. We uncovered an unreported outbreak in Cuba during 2017, a year after peak transmission in neighboring islands. By sequencing Zika virus, we show that the establishment of the virus was delayed by a year and that the ensuing outbreak was sparked by long-lived lineages of Zika virus from other Caribbean islands. Our data suggest that, although mosquito control in Cuba may initially have been effective at mitigating Zika virus transmission, such measures need to be maintained to be effective. Our study highlights how Zika virus may still be "silently" spreading and provides a framework for understanding outbreak dynamics. VIDEO ABSTRACT.