Intranasal vaccination with lipid-conjugated immunogens promotes antigen transmucosal uptake to drive mucosal and systemic immunity.

To combat the HIV epidemic and emerging threats such as SARS-CoV-2, immunization strategies are needed that elicit protection at mucosal portals of pathogen entry. Immunization directly through airway surfaces is effective in driving mucosal immunity, but poor vaccine uptake across the mucus and epithelial lining is a limitation. The major blood protein albumin is constitutively transcytosed bidirectionally across the airway epithelium through interactions with neonatal Fc receptors (FcRn). Exploiting this biology, here, we demonstrate a strategy of "albumin hitchhiking" to promote mucosal immunity using an intranasal vaccine consisting of protein immunogens modified with an amphiphilic albumin-binding polymer-lipid tail, forming amph-proteins. Amph-proteins persisted in the nasal mucosa of mice and nonhuman primates and exhibited increased uptake into the tissue in an FcRn-dependent manner, leading to enhanced germinal center responses in nasal-associated lymphoid tissue. Intranasal immunization with amph-conjugated HIV Env gp120 or SARS-CoV-2 receptor binding domain (RBD) proteins elicited 100- to 1000-fold higher antigen-specific IgG and IgA titers in the serum, upper and lower respiratory mucosa, and distal genitourinary mucosae of mice compared to unmodified protein. Amph-RBD immunization induced high titers of SARS-CoV-2-neutralizing antibodies in serum, nasal washes, and bronchoalveolar lavage. Furthermore, intranasal amph-protein immunization in rhesus macaques elicited 10-fold higher antigen-specific IgG and IgA responses in the serum and nasal mucosa compared to unmodified protein, supporting the translational potential of this approach. These results suggest that using amph-protein vaccines to deliver antigen across mucosal epithelia is a promising strategy to promote mucosal immunity against HIV, SARS-CoV-2, and other infectious diseases.

Antibody Light Chains: Key to Increased Monoclonal Antibody Yields in Expi293 Cells?

When constructing isogenic recombinant IgM-IgG pairs, we discovered that µ heavy chains strongly prefer partnering with λ light chains for optimal IgM expression in transiently cotransfected Expi293 cells. When µ chains were paired with κ light chains, IgM yields were low but increased by logs-up to 20,000 X-by using λ chains instead. Switching light chains did not alter epitope specificity. For dimeric IgA2, optimal expression involved pairing with λ chains, whereas light-chain preference varied for other immunoglobulin classes. In summary, recombinant IgM production can be drastically increased by using λ chains, an important finding in the use of IgM for mucosal immunoprophylaxis.

Cooperation Between Systemic and Mucosal Antibodies Induced by Virosomal Vaccines Targeting HIV-1 Env: Protection of Indian Rhesus Macaques Against Low-Dose Intravaginal SHIV Challenges.

A virosomal vaccine inducing systemic/mucosal anti-HIV-1 gp41 IgG/IgA had previously protected Chinese-origin rhesus macaques (RMs) against vaginal SHIVSF162P3 challenges. Here, we assessed its efficacy in Indian-origin RMs by intramuscular priming/intranasal boosting (n=12/group). Group K received virosome-P1-peptide alone (harboring the Membrane Proximal External Region), Group L combined virosome-rgp41 plus virosome-P1, and Group M placebo virosomes. Vaccination induced plasma binding but no neutralizing antibodies. Five weeks after boosting, all RMs were challenged intravaginally with low-dose SHIVSF162P3 until persistent systemic infection developed. After SHIV challenge #7, six controls were persistently infected versus only one Group L animal (vaccine efficacy 87%; P=0.0319); Group K was not protected. After a 50% SHIV dose increase starting with challenge #8, protection in Group L was lost. Plasmas/sera were analyzed for IgG phenotypes and effector functions; the former revealed that protection in Group L was significantly associated with increased binding to FcγR2/3(A/B) across several time-points, as were some IgG measurements. Vaginal washes contained low-level anti-gp41 IgGs and IgAs, representing a 1-to-5-fold excess over the SHIV inoculum's gp41 content, possibly explaining loss of protection after the increase in challenge-virus dose. Virosomal gp41-vaccine efficacy was confirmed during the initial seven SHIV challenges in Indian-origin RMs when the SHIV inoculum had at least 100-fold more HIV RNA than acutely infected men's semen. Vaccine protection by virosome-induced IgG and IgA parallels the cooperation between systemically administered IgG1 and mucosally applied dimeric IgA2 monoclonal antibodies that as single-agents provided no/low protection - but when combined, prevented mucosal SHIV transmission in all passively immunized RMs.

A particulate saponin/TLR agonist vaccine adjuvant alters lymph flow and modulates adaptive immunity.

Saponins are potent and safe vaccine adjuvants, but their mechanisms of action remain incompletely understood. Here, we explored the properties of several saponin formulations, including immune-stimulatory complexes (ISCOMs) formed by the self-assembly of saponin and phospholipids in the absence or presence of the Toll-like receptor 4 agonist monophosphoryl lipid A (MPLA). We found that MPLA self-assembles with saponins to form particles physically resembling ISCOMs, which we termed saponin/MPLA nanoparticles (SMNP). Saponin-containing adjuvants exhibited distinctive mechanisms of action, altering lymph flow in a mast cell­dependent manner and promoting antigen entry into draining lymph nodes. SMNP was particularly effective, exhibiting even greater potency than the compositionally related adjuvant AS01B in mice, and primed robust germinal center B cell, TFH, and HIV tier 2 neutralizing antibodies in nonhuman primates. Together, these findings shed new light on mechanisms by which saponin adjuvants act to promote the immune response and suggest that SMNP may be a promising adjuvant in the setting of HIV, SARS-CoV-2, and other pathogens.

Localization of infection in neonatal rhesus macaques after oral viral challenge.

Vertical transmission of human immunodeficiency virus (HIV) can occur in utero, during delivery, and through breastfeeding. We utilized Positron Emission Tomography (PET) imaging coupled with fluorescent microscopy of 64Cu-labeled photoactivatable-GFP-HIV (PA-GFP-BaL) to determine how HIV virions distribute and localize in neonatal rhesus macaques two and four hours after oral viral challenge. Our results show that by four hours after oral viral exposure, HIV virions localize to and penetrate the rectal mucosa. We also used a dual viral challenge with a non-replicative viral vector and a replication competent SHIV-1157ipd3N4 to examine viral transduction and dissemination at 96 hours. Our data show that while SHIV-1157ipd3N4 infection can be found in the oral cavity and upper gastrointestinal (GI) tract, the small and large intestine contained the largest number of infected cells. Moreover, we found that T cells were the biggest population of infected immune cells. Thus, thanks to these novel technologies, we are able to visualize and delineate of viral distribution and infection throughout the entire neonatal GI tract during acute viral infection.

Cooperation Between Systemic IgG1 and Mucosal Dimeric IgA2 Monoclonal Anti-HIV Env Antibodies: Passive Immunization Protects Indian Rhesus Macaques Against Mucosal SHIV Challenges.

Understanding the interplay between systemic and mucosal anti-HIV antibodies can provide important insights to develop new prevention strategies. We used passive immunization via systemic and/or mucosal routes to establish cause-and-effect between well-characterized monoclonal antibodies and protection against intrarectal (i.r.) SHIV challenge. In a pilot study, for which we re-used animals previously exposed to SHIV but completely protected from viremia by different classes of anti-HIV neutralizing monoclonal antibodies (mAbs), we made a surprise finding: low-dose intravenous (i.v.) HGN194-IgG1, a human neutralizing mAb against the conserved V3-loop crown, was ineffective when given alone but protected 100% of animals when combined with i.r. applied HGN194-dIgA2 that by itself had only protected 17% of the animals. Here we sought to confirm the unexpected synergy between systemically administered IgG1 and mucosally applied dIgA HGN194 forms using six groups of naïve macaques (n=6/group). Animals received i.v. HGN194-IgG1 alone or combined with i.r.-administered dIgA forms; controls remained untreated. HGN194-IgG1 i.v. doses were given 24 hours before - and all i.r. dIgA doses 30 min before - i.r. exposure to a single high-dose of SHIV-1157ipEL-p. All controls became viremic. Among passively immunized animals, the combination of IgG1+dIgA2 again protected 100% of the animals. In contrast, single-agent i.v. IgG1 protected only one of six animals (17%) - consistent with our pilot data. IgG1 combined with dIgA1 or dIgA1+dIgA2 protected 83% (5/6) of the animals. The dIgA1+dIgA2 combination without the systemically administered dose of IgG1 protected 67% (4/6) of the macaques. We conclude that combining suboptimal antibody defenses at systemic and mucosal levels can yield synergy and completely prevent virus acquisition.

Mucosal AIDS virus transmission is enhanced by antiviral IgG isolated early in infection.

OBJECTIVE: Antibody-dependent enhancement (ADE) affects host-virus dynamics in fundamentally different ways: i) enhancement of initial virus acquisition, and/or ii) increased disease progression/severity. Here we address the question whether anti-HIV-1 antibodies can enhance initial infection. While cell-culture experiments hinted at this possibility, in-vivo proof remained elusive. DESIGN: We used passive immunization in nonhuman primates challenged with simian-human immunodeficiency virus (SHIV), a chimera expressing HIV-1 envelope. We purified IgG from rhesus monkeys with early-stage SHIV infection - before cross-neutralizing anti-HIV-1 antibodies had developed - and screened for maximal complement-mediated antibody-dependent enhancement (C'-ADE) of viral replication with a SHIV strain phylogenetically distinct from that harbored by IgG donor macaques. IgG fractions with maximal C'-ADE but lacking neutralization were combined to yield enhancing anti-SHIV IgG (enSHIVIG). RESULTS: We serially enrolled naive macaques (Group 1) to determine the minimal and 50% animal infectious doses required to establish persistent infection after intrarectal SHIV challenge. The first animal was inoculated with a 1 : 10 virus-stock dilution; after this animal's viral RNA load was >104copies/ml, the next macaque was challenged with 10x less virus, a process repeated until viremia no longer ensued. Group 2 was pretreated intravenously with enSHIVIG 24 h before SHIV challenge. Overall, Group 2 macaques required 3.4-fold less virus compared to controls (P = 0.002). This finding is consistent with enhanced susceptibility of the passively immunized animals to mucosal SHIV challenge. CONCLUSION: These passive immunization data give proof of IgG-mediated enhanced virus acquisition after mucosal exposure - a potential concern for antibody-based AIDS vaccine development.

PET/CT targeted tissue sampling reveals virus specific dIgA can alter the distribution and localization of HIV after rectal exposure.

Human immunodeficiency virus (HIV) vaccines have not been successful in clinical trials. Dimeric IgA (dIgA) in the form of secretory IgA is the most abundant antibody class in mucosal tissues, making dIgA a prime candidate for potential HIV vaccines. We coupled Positron Emission Tomography (PET) imaging and fluorescent microscopy of 64Cu-labeled, photoactivatable-GFP HIV (PA-GFP-BaL) and fluorescently labeled dIgA to determine how dIgA antibodies influence virus interaction with mucosal barriers and viral penetration in colorectal tissue. Our results show that HIV virions rapidly disseminate throughout the colon two hours after exposure. The presence of dIgA resulted in an increase in virions and penetration depth in the transverse colon. Moreover, virions were found in the mesenteric lymph nodes two hours after viral exposure, and the presence of dIgA led to an increase in virions in mesenteric lymph nodes. Taken together, these technologies enable in vivo and in situ visualization of antibody-virus interactions and detailed investigations of early events in HIV infection.

Combined PET and whole-tissue imaging of lymphatic-targeting vaccines in non-human primates.

Antigen accumulation in lymph nodes (LNs) is critical for vaccine efficacy, but understanding of vaccine biodistribution in humans or large animals remains limited. Using the rhesus macaque model, we employed a combination of positron emission tomography (PET) and fluorescence imaging to characterize the whole-animal to tissue-level biodistribution of a subunit vaccine comprised of an HIV envelope trimer protein nanoparticle (trimer-NP) and lipid-conjugated CpG adjuvant (amph-CpG). Following immunization in the thigh, PET imaging revealed vaccine uptake primarily in inguinal and iliac LNs, reaching distances up to 17 cm away from the injection site. Within LNs, trimer-NPs exhibited striking accumulation on the periphery of follicular dendritic cell (FDC) networks in B cell follicles. Comparative imaging of soluble Env trimers (not presented on nanoparticles) in naïve or previously-immunized animals revealed diffuse deposition of trimer antigens in LNs following primary immunization, but concentration on FDCs in pre-immunized animals with high levels of trimer-specific IgG. These data demonstrate the capacity of nanoparticle or "albumin hitchhiking" technologies to concentrate vaccines in genitourinary tract-draining LNs, which may be valuable for promoting mucosal immunity.

Immunoglobulin M: An Ancient Antiviral Weapon - Rediscovered.

Recent discoveries have shed new light onto immunoglobulin M (IgM), an ancient antibody class preserved throughout evolution in all vertebrates. First, IgM - long thought to be a perfect pentamer - was shown to be asymmetric, resembling a quasi-hexamer missing one monomer and containing a gap. Second, this gap allows IgM to serve as carrier of a specific host protein, apoptosis inhibitor of macrophages (AIM), which is released to promote removal of dead-cell debris, cancer cells, or pathogens. Third, recombinant IgM delivered mucosally by passive immunization gave proof-of-concept that this antibody class can prevent mucosal simian-human immunodeficiency virus transmission in non-human primates. Finally, IgM's role in adaptive immunity goes beyond being only a first defender to respond to pathogen invasion, as long-lived IgM plasma cells have been observed predominantly residing in the spleen. In fact, IgM produced by such cells contained somatic hypermutations and was linked to protection against lethal influenza virus challenge in murine models. Importantly, such long-lived IgM plasma cells had been induced by immunization 1 year before challenge. Together, new data on IgM function raise the possibility that vaccine strategies aimed at preventing virus acquisition could include this ancient weapon.

Correction: ALVAC-HIV B/C candidate HIV vaccine efficacy dependent on neutralization profile of challenge virus and adjuvant dose and type.

[This corrects the article DOI: 10.1371/journal.ppat.1008121.].

Immunotherapy with DNA vaccine and live attenuated rubella/SIV gag vectors plus early ART can prevent SIVmac251 viral rebound in acutely infected rhesus macaques.

Anti-retroviral therapy (ART) has been highly successful in controlling HIV replication, reducing viral burden, and preventing both progression to AIDS and viral transmission. Yet, ART alone cannot cure the infection. Even after years of successful therapy, ART withdrawal leads inevitably to viral rebound within a few weeks or months. Our hypothesis: effective therapy must control both the replicating virus pool and the reactivatable latent viral reservoir. To do this, we have combined ART and immunotherapy to attack both viral pools simultaneously. The vaccine regimen consisted of DNA vaccine expressing SIV Gag, followed by a boost with live attenuated rubella/gag vectors. The vectors grow well in rhesus macaques, and they are potent immunogens when used in a prime and boost strategy. We infected rhesus macaques by high dose mucosal challenge with virulent SIVmac251 and waited three days to allow viral dissemination and establishment of a reactivatable viral reservoir before starting ART. While on ART, the control group received control DNA and empty rubella vaccine, while the immunotherapy group received DNA/gag prime, followed by boosts with rubella vectors expressing SIV gag over 27 weeks. Both groups had a vaccine "take" to rubella, and the vaccine group developed antibodies and T cells specific for Gag. Five weeks after the last immunization, we stopped ART and monitored virus rebound. All four control animals eventually had a viral rebound, and two were euthanized for AIDS. One control macaque did not rebound until 2 years after ART release. In contrast, there was only one viral rebound in the vaccine group. Three out of four vaccinees had no viral rebound, even after CD8 depletion, and they remain in drug-free viral remission more than 2.5 years later. The strategy of early ART combined with immunotherapy can produce a sustained SIV remission in macaques and may be relevant for immunotherapy of HIV in humans.

Modified Adenovirus Prime-Protein Boost Clade C HIV Vaccine Strategy Results in Reduced Viral DNA in Blood and Tissues Following Tier 2 SHIV Challenge.

Designing immunogens and improving delivery methods eliciting protective immunity is a paramount goal of HIV vaccine development. A comparative vaccine challenge study was performed in rhesus macaques using clade C HIV Envelope (Env) and SIV Gag antigens. One group was vaccinated using co-immunization with DNA Gag and Env expression plasmids cloned from a single timepoint and trimeric Env gp140 glycoprotein from one of these clones (DNA+Protein). The other group was a prime-boost regimen composed of two replicating simian (SAd7) adenovirus-vectored vaccines expressing Gag and one Env clone from the same timepoint as the DNA+Protein group paired with the same Env gp140 trimer (SAd7+Protein). The env genes were isolated from a single pre-peak neutralization timepoint approximately 1 year post infection in CAP257, an individual with a high degree of neutralization breadth. Both DNA+Protein and SAd7+Protein vaccine strategies elicited significant Env-specific T cell responses, lesser Gag-specific responses, and moderate frequencies of Env-specific TFH cells. Both vaccine modalities readily elicited systemic and mucosal Env-specific IgG but not IgA. There was a higher frequency and magnitude of ADCC activity in the SAd7+Protein than the DNA+Protein arm. All macaques developed moderate Tier 1 heterologous neutralizing antibodies, while neutralization of Tier 1B or Tier 2 viruses was sporadic and found primarily in macaques in the SAd7+Protein group. Neither vaccine approach provided significant protection from viral acquisition against repeated titered mucosal challenges with a heterologous Tier 2 clade C SHIV. However, lymphoid and gut tissues collected at necropsy showed that animals in both vaccine groups each had significantly lower copies of viral DNA in individual tissues compared to levels in controls. In the SAd7+Protein-vaccinated macaques, total and peak PBMC viral DNA were significantly lower compared with controls. Taken together, this heterologous Tier 2 SHIV challenge study shows that combination vaccination with SAd7+Protein was superior to combination DNA+Protein in reducing viral seeding in tissues in the absence of protection from infection, thus emphasizing the priming role of replication-competent SAd7 vector. Despite the absence of correlates of protection, because antibody responses were significantly higher in this vaccine group, we hypothesize that vaccine-elicited antibodies contribute to limiting tissue viral seeding.

ALVAC-HIV B/C candidate HIV vaccine efficacy dependent on neutralization profile of challenge virus and adjuvant dose and type.

The ALVAC-HIV clade B/AE and equivalent SIV-based/gp120 + Alum vaccines successfully decreased the risk of virus acquisition in humans and macaques. Here, we tested the efficacy of HIV clade B/C ALVAC/gp120 vaccine candidates + MF59 or different doses of Aluminum hydroxide (Alum) against SHIV-Cs of varying neutralization sensitivity in macaques. Low doses of Alum induced higher mucosal V2-specific IgA that increased the risk of Tier 2 SHIV-C acquisition. High Alum dosage, in contrast, elicited serum IgG to V2 that correlated with a decreased risk of Tier 1 SHIV-C acquisition. MF59 induced negligible mucosal antibodies to V2 and an inflammatory profile with blood C-reactive Protein (CRP) levels correlating with neutralizing antibody titers. MF59 decreased the risk of Tier 1 SHIV-C acquisition. The relationship between vaccine efficacy and the neutralization profile of the challenge virus appear to be linked to the different immunological spaces created by MF59 and Alum via CXCL10 and IL-1ß, respectively.

Mucosal Antibodies: Defending Epithelial Barriers against HIV-1 Invasion.

The power of mucosal anti-HIV-1 envelope immunoglobulins (Igs) to block virus transmission is underappreciated. We used passive immunization, a classical tool to unequivocally prove whether antibodies are protective. We mucosally instilled recombinant neutralizing monoclonal antibodies (nmAbs) of different Ig classes in rhesus macaques (RMs) followed by mucosal simian-human immunodeficiency virus (SHIV) challenge. We gave anti-HIV-1 IgM, IgG, and dimeric IgA (dIgA) versions of the same human nmAb, HGN194 that targets the conserved V3 loop crown. Surprisingly, dIgA1 with its wide-open, flat hinge protected 83% of the RMs against intrarectal R5-tropic SHIV-1157ipEL-p challenge, whereas dIgA2, with its narrow hinge, only protected 17% of the animals-despite identical epitope specificities and in vitro neutralization curves of the two dIgA isotypes (Watkins et al., AIDS 2013 27(9):F13-20). These data imply that factors in addition to neutralization determine in vivo protection. We propose that this underlying protective mechanism is immune exclusion, which involves large nmAb/virion aggregates that prevent virus penetration of mucosal barriers. Future studies need to find biomarkers that predict effective immune exclusion in vivo. Vaccine development strategies against HIV-1 and/or other mucosally transmissible pathogens should include induction of strong mucosal Abs of different Ig classes to defend epithelial barriers against pathogen invasion.

Lymphocytes upregulate CD36 in adipose tissue and liver.

CD36 is a multifunctional scavenger receptor and lipid transporter implicated in metabolic and inflammatory pathologies, as well as cancer progression. CD36 is known to be expressed by adipocytes and monocytes/macrophages, but its expression by T cells is not clearly established. We found that CD4 and CD8 T cells in adipose tissue and liver of humans, monkeys, and mice upregulated CD36 expression (ranging from ~5-40% CD36+), whereas little to no CD36 was expressed by T cells in blood, spleen, and lymph nodes. CD36 was expressed predominantly by resting CD38-, HLA.DR-, and PD-1- adipose tissue T cells in monkeys, and increased during high-fat feeding in mice. Adipose tissue and liver promote a distinct phenotype in resident T cells characterized by CD36 upregulation.

Adaptation of an R5 Simian-Human Immunodeficiency Virus Encoding an HIV Clade A Envelope with or without Ablation of Adaptive Host Immunity: Differential Selection of Viral Mutants.

Simian-human immunodeficiency virus (SHIV) infection in rhesus macaques (RMs) resembles human immunodeficiency virus type 1 (HIV-1) infection in humans and serves as a tool to evaluate candidate AIDS vaccines. HIV-1 clade A (HIV-A) predominates in parts of Africa. We constructed an R5 clade A SHIV (SHIV-A; strain SHIV-KNH1144) carrying env from a Kenyan HIV-A. SHIV-A underwent rapid serial passage through six RMs. To allow unbridled replication without adaptive immunity, we simultaneously ablated CD8+ and B cells with cytotoxic monoclonal antibodies in the next RM, resulting in extremely high viremia and CD4+ T-cell loss. Infected blood was then transferred into two non-immune-depleted RMs, where progeny SHIV-A showed increased replicative capacity and caused AIDS. We reisolated SHIV-KNH1144p4, which was replication competent in peripheral blood mononuclear cells (PBMC) of all RMs tested. Next-generation sequencing of early- and late-passage SHIV-A strains identified mutations that arose due to "fitness" virus optimization in the former and mutations exhibiting signatures typical for adaptive host immunity in the latter. "Fitness" mutations are best described as mutations that allow for better fit of the HIV-A Env with SIV-derived virion building blocks or host proteins and mutations in noncoding regions that accelerate virus replication, all of which result in the outgrowth of virus variants in the absence of adaptive T-cell and antibody-mediated host immunity.IMPORTANCE In this study, we constructed a simian-human immunodeficiency virus carrying an R5 Kenyan HIV-1 clade A env (SHIV-A). To bypass host immunity, SHIV-A was rapidly passaged in naive macaques or animals depleted of both CD8+ and B cells. Next-generation sequencing identified different mutations that resulted from optimization of viral replicative fitness either in the absence of adaptive immunity or due to pressure from adaptive immune responses.

Anti-HIV IgM protects against mucosal SHIV transmission.

OBJECTIVE: Worldwide, most new HIV infections occur through mucosal exposure. Immunoglobulin M (IgM) is the first antibody class generated in response to infectious agents; IgM is present in the systemic circulation and in mucosal fluids as secretory IgM. We sought to investigate for the first time the role of IgM in preventing AIDS virus acquisition in vivo. DESIGN: Recombinant polymeric monoclonal IgM was generated from the neutralizing monoclonal IgG1 antibody 33C6-IgG1, tested in vitro, and given by passive intrarectal immunization to rhesus macaques 30 min before intrarectal challenge with simian-human immunodeficiency virus (SHIV) that carries an HIV-1 envelope gene. RESULTS: In vitro, 33C6-IgM captured virions more efficiently and neutralized the challenge SHIV with a 50% inhibitory molar concentration (IC50) that was 1 log lower than that for 33C6-IgG1. The IgM form also exhibited significantly higher affinity and avidity compared with 33C6-IgG1. After intrarectal administration, 33C6-IgM prevented viremia in four out of six rhesus macaques after high-dose intrarectal SHIV challenge. Five out of six rhesus macaques given 33C6-IgG1 were protected at a five times higher molar concentration compared with the IgM form; all untreated controls became highly viremic. Rhesus macaques passively immunized with 33C6-IgM with breakthrough infection had notably early development of autologous neutralizing antibody responses. CONCLUSION: Our primate model data provide the first proof-of-concept that mucosal IgM can prevent mucosal HIV transmission and have implications for HIV prevention and vaccine development.