Publications by authors named "Preston A Marx"

92 Publications

CD4 receptor diversity represents an ancient protection mechanism against primate lentiviruses.

Proc Natl Acad Sci U S A 2021 Mar;118(13)

Lukuru Wildlife Research Foundation, Tshuapa-Lomami-Lualaba Project, BP 2012, Kinshasa, Democratic Republic of the Congo.

Infection with human and simian immunodeficiency viruses (HIV/SIV) requires binding of the viral envelope glycoprotein (Env) to the host protein CD4 on the surface of immune cells. Although invariant in humans, the Env binding domain of the chimpanzee CD4 is highly polymorphic, with nine coding variants circulating in wild populations. Here, we show that within-species CD4 diversity is not unique to chimpanzees but found in many African primate species. Characterizing the outermost (D1) domain of the CD4 protein in over 500 monkeys and apes, we found polymorphic residues in 24 of 29 primate species, with as many as 11 different coding variants identified within a single species. D1 domain amino acid replacements affected SIV Env-mediated cell entry in a single-round infection assay, restricting infection in a strain- and allele-specific fashion. Several identical CD4 polymorphisms, including the addition of -linked glycosylation sites, were found in primate species from different genera, providing striking examples of parallel evolution. Moreover, seven different guenons ( spp.) shared multiple distinct D1 domain variants, pointing to long-term trans-specific polymorphism. These data indicate that the HIV/SIV Env binding region of the primate CD4 protein is highly variable, both within and between species, and suggest that this diversity has been maintained by balancing selection for millions of years, at least in part to confer protection against primate lentiviruses. Although long-term SIV-infected species have evolved specific mechanisms to avoid disease progression, primate lentiviruses are intrinsically pathogenic and have left their mark on the host genome.
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http://dx.doi.org/10.1073/pnas.2025914118DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8020793PMC
March 2021

Introduction.

J Med Primatol 2020 10;49(5):221

Tulane National Primate Research Center, Covington, LA, USA.

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http://dx.doi.org/10.1111/jmp.12488DOI Listing
October 2020

DNA analysis reveals non-falciparum malaria in the Democratic Republic of the Congo.

Acta Trop 2020 Dec 29;212:105557. Epub 2020 May 29.

Department of Tropical Medicine, School of Public Health and Tropical Medicine. Tulane University, New Orleans, LA, United States; Division of Microbiology, Tulane National Primate Research Center, Covington, LA, United States. Electronic address:

Background: The World Health Organization (WHO) attributes the entirety of malaria infection and transmission in the Democratic Republic of the Congo (DRC) to Plasmodium falciparum, one of the several species of malaria known to infect humans. Recent studies have put forth some evidence that transmission of Plasmodium vivax may also be occurring in the DRC. As interventions and treatments differ between malaria species, it is crucial to maintain the most accurate understanding of malaria species diversity in each region.

Methods: Blood samples were taken from aymptomatic children 0-5 years old living in three regions of the DRC in 2014. For this study, samples were taken from a larger pool of samples, collected as part of a population-based survey in three regions. Plasmodium infection was screened for using nested polymerase chain reaction (PCR) assays and species were confirmed by cloning and DNA sequencing.

Results: Of 336 samples screened by PCR, 62.2% (n=209) initially tested positive for P. falciparum and 14.6% (n=49) initially tested positive for P. vivax. Sanger sequencing was performed on PCR-positive Plasmodium samples to confirm identity of Plasmodium species. Sequencing showed Plasmodium malariae in one blood sample and Plasmodium ovale in another sample. Plasmodium vivax was detected in 12/65 cases (18.5%). Overall, 14/65 sequenced cases (21.5%) were infected with a non-falciparum malaria. 330bp 18s P. vivax DNA sequences were obtained.

Conclusions: This study reveals Plasmodium vivax and other non-falciparum malaria across several regions of the DRC, and enforces the importance of further testing and more precise diagnostics when testing for and treating malaria in the DRC. Here, we find a higher proportion of cases of P. vivax malaria than found in previous studies. This is the most robust DNA sequencing of Plasmodium vivax in the DRC to date.
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http://dx.doi.org/10.1016/j.actatropica.2020.105557DOI Listing
December 2020

Safety and immunogenicity of a recombinant vaccine against Trypanosoma cruzi in Rhesus macaques.

Vaccine 2020 06 13;38(29):4584-4591. Epub 2020 May 13.

Texas Children's Hospital Center for Vaccine Development, Department of Pediatrics and National School of Tropical Medicine, Baylor College of Medicine, Houston, TX, USA.

Chagas disease, caused by the protozoan parasite Trypanosoma cruzi is one of the most important neglected parasitic diseases in the Americas. Vaccines represent an attractive complementary strategy for the control of T. cruzi infection and pre-clinical studies in mice demonstrated that trypomastigote surface antigen (TSA-1) and the flagellar calcium-binding (Tc24) parasite antigens are promising candidates for vaccine development. We performed here the first evaluation of the safety and immunogenicity of two recombinant vaccine antigens (named TSA1-C4 and Tc24-C4) in naïve non-human primates. Three rhesus macaques received 3 doses of each recombinant protein, formulated with E6020 (Eisai Co., Ltd.), a novel Toll-like receptor-4 agonist, in a stable emulsion. All parameters from blood chemistry and blood cell counts were stable over the course of the study and unaffected by the vaccine. A specific IgG response against both antigens was detectable after the first vaccine dose, and increased with the second dose. After three vaccine doses, stimulation of PBMCs with a peptide pool derived from TSA1-C4 resulted in the induction of TSA1-C4-specific TNFα-, IL-2- and IFNγ-producing CD4 in one or two animals while stimulation with a peptide pool derived from Tc24-C4 only activated IFNγ-producing CD4T cells in one animal. In two animals there was also activation of TSA1-C4-specific IL2-producing CD8 T cells. This is the first report of the immunogenicity of T. cruzi-derived recombinant antigens formulated as an emulsion with a TLR4 agonist in a non-human primate model. Our results strongly support the need for further evaluation of the preventive efficacy of this type of vaccine in non-human primates and explore the effect of the vaccine in a therapeutic model of naturally-infected Chagasic non-human primates, which would strengthen the rationale for the clinical development as a human vaccine against Chagas disease.
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http://dx.doi.org/10.1016/j.vaccine.2020.05.010DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7310587PMC
June 2020

Co-immunization of DNA and Protein in the Same Anatomical Sites Induces Superior Protective Immune Responses against SHIV Challenge.

Cell Rep 2020 05;31(6):107624

Division of Comparative Pathology, Tulane National Primate Research Center, Covington, LA 70433, USA.

We compare immunogenicity and protective efficacy of an HIV vaccine comprised of env and gag DNA and Env (Envelope) proteins by co-administration of the vaccine components in the same muscles or by separate administration of DNA + protein in contralateral sites in female rhesus macaques. The 6-valent vaccine includes gp145 Env DNAs, representing six sequentially isolated Envs from the HIV-infected individual CH505, and matching GLA-SE-adjuvanted gp120 Env proteins. Interestingly, only macaques in the co-administration vaccine group are protected against SHIV CH505 acquisition after repeated low-dose intravaginal challenge and show 67% risk reduction per exposure. Macaques in the co-administration group develop higher Env-specific humoral and cellular immune responses. Non-neutralizing Env antibodies, ADCC, and antibodies binding to FcγRIIIa are associated with decreased transmission risk. These data suggest that simultaneous recognition, processing, and presentation of DNA + Env protein in the same draining lymph nodes play a critical role in the development of protective immunity.
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http://dx.doi.org/10.1016/j.celrep.2020.107624DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7329227PMC
May 2020

Introduction.

J Med Primatol 2019 10;48(5):267

Tulane National Primate Research Center, Covington, LA, USA.

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http://dx.doi.org/10.1111/jmp.12442DOI Listing
October 2019

Repeated semen exposure decreases cervicovaginal SIVmac251 infection in rhesus macaques.

Nat Commun 2019 08 21;10(1):3753. Epub 2019 Aug 21.

The Wistar Institute, Philadelphia, PA, USA.

Semen is the vehicle for virion dissemination in the female reproductive tract (FRT) in male-to-female HIV transmission. Recent data suggests that higher frequency semen exposure is associated with activation of anti-HIV mechanisms in HIV negative sex workers. Here, we use a non-human primate (NHP) model to show that repeated vaginal exposure to semen significantly reduces subsequent infection by repeated low-dose vaginal SIVmac251 challenge. Repeated semen exposures result in lower CCR5 expression in circulating CD4+ T-cells, as well as higher expression of Mx1 (in correlation with IFNε expression) and FoxP3 in the cervicovaginal mucosa, and increased infiltration of CD4+ T-cells. Establishing in vivo evidence of competing effects of semen on transmission impacts our basic understanding of what factors may determine HIV infectivity in humans. Our results clearly indicate that repeated semen exposure can profoundly modulate the FRT microenvironment, paradoxically promoting host resistance against HIV acquisition.
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http://dx.doi.org/10.1038/s41467-019-11814-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6704120PMC
August 2019

Introduction.

J Med Primatol 2018 10;47(5):285

Tulane National Primate Research Center, Covington, LA, USA.

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http://dx.doi.org/10.1111/jmp.12384DOI Listing
October 2018

Preadaptation of Simian Immunodeficiency Virus SIVsmm Facilitated Env-Mediated Counteraction of Human Tetherin by Human Immunodeficiency Virus Type 2.

J Virol 2018 09 29;92(18). Epub 2018 Aug 29.

Institute of Molecular Virology, Ulm University Medical Center, Ulm, Germany

The host restriction factor tetherin inhibits virion release from infected cells and poses a significant barrier to successful zoonotic transmission of primate lentiviruses to humans. While most simian immunodeficiency viruses (SIV), including the direct precursors of human immunodeficiency virus type 1 (HIV-1) and HIV-2, use their Nef protein to counteract tetherin in their natural hosts, they fail to antagonize the human tetherin ortholog. Pandemic HIV-1 group M and epidemic group O strains overcame this hurdle by adapting their Vpu and Nef proteins, respectively, whereas HIV-2 group A uses its envelope (Env) glycoprotein to counteract human tetherin. Whether or how the remaining eight groups of HIV-2 antagonize this antiviral factor has remained unclear. Here, we show that Nef proteins from diverse groups of HIV-2 do not or only modestly antagonize human tetherin, while their ability to downmodulate CD3 and CD4 is highly conserved. Experiments in transfected cell lines and infected primary cells revealed that not only Env proteins of epidemic HIV-2 group A but also those of a circulating recombinant form (CRF01_AB) and rare groups F and I decrease surface expression of human tetherin and significantly enhance progeny virus release. Intriguingly, we found that many SIVsmm Envs also counteract human as well as smm tetherin. Thus, Env-mediated tetherin antagonism in different groups of HIV-2 presumably stems from a preadaptation of their SIVsmm precursors to humans. In summary, we identified a phenotypic trait of SIVsmm that may have facilitated its successful zoonotic transmission to humans and the emergence of HIV-2. HIV-2 groups A to I resulted from nine independent cross-species transmission events of SIVsmm to humans and differ considerably in their prevalence and geographic spread. Thus, detailed characterization of these viruses offers a valuable means to elucidate immune evasion mechanisms and human-specific adaptations determining viral spread. In a systematic comparison of rare and epidemic HIV-2 groups and their simian SIVsmm counterparts, we found that the ability of Nef to downmodulate the primary viral entry receptor CD4 and the T cell receptor CD3 is conserved, while effects on CD28, CD74, and major histocompatibility complex class I surface expression vary considerably. Furthermore, we show that not only the Env proteins of HIV-2 groups A, AB, F, and I but also those of some SIVsmm isolates antagonize human tetherin. This finding helps to explain why SIVsmm has been able to cross the species barrier to humans on at least nine independent occasions.
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http://dx.doi.org/10.1128/JVI.00276-18DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6146695PMC
September 2018

Gag and env conserved element CE DNA vaccines elicit broad cytotoxic T cell responses targeting subdominant epitopes of HIV and SIV Able to recognize virus-infected cells in macaques.

Hum Vaccin Immunother 2018 12;14(9):2163-2177. Epub 2018 Jul 12.

a Human Retrovirus Pathogenesis Section, Center for Cancer Research, National Cancer Institute at Frederick , Frederick , MD , USA.

HIV sequence diversity and the propensity of eliciting immunodominant responses targeting inessential variable regions are hurdles in the development of an effective AIDS vaccine. We developed a DNA vaccine comprising conserved elements (CE) of SIV p27 and HIV-1 Env and found that priming vaccination with CE DNA is critical to efficiently overcome the dominance imposed by Gag and Env variable regions. Here, we show that DNA vaccinated macaques receiving the CE prime/CE+full-length DNA co-delivery booster vaccine regimens developed broad, potent and durable cytotoxic T cell responses targeting conserved protein segments of SIV Gag and HIV Env. Gag CE-specific T cells showed robust anamnestic responses upon infection with SIV which led to the identification of CE-specific cytotoxic lymphocytes able to recognize epitopes covering distinct CE on the surface of SIV infected cells in vivo. Though not controlling infection overall, we found an inverse correlation between Gag CE-specific CD8 T cell responses and peak viremia. The T cell responses induced by the HIV Env CE immunogen were recalled in some animals upon SIV infection, leading to the identification of two cross-reactive epitopes between HIV and SIV Env based in sequence homology. These data demonstrate that a vaccine combining Gag and Env CE DNA subverted the normal immunodominance patterns, eliciting immune responses that included subdominant, highly conserved epitopes. These vaccine regimens augment cytotoxic T cell responses to highly conserved epitopes in the viral proteome and maximize response breadth. The vaccine-induced CE-specific T cells were expanded upon SIV infection, indicating that the predicted CE epitopes incorporated in the DNA vaccine are processed and exposed by infected cells in their natural context within the viral proteome.
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http://dx.doi.org/10.1080/21645515.2018.1489949DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6183272PMC
May 2019

Introduction.

J Med Primatol 2017 08;46(4):119

Tulane National Primate Research Center, Covington, LA, USA.

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http://dx.doi.org/10.1111/jmp.12302DOI Listing
August 2017

Memorial - Andrew A. Lackner, D.V.M., Ph.D. 1960-2017.

Authors:
Preston A Marx

J Med Primatol 2017 08;46(4):120

Tulane National Primate Research Center, Covington, LA, USA.

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http://dx.doi.org/10.1111/jmp.12303DOI Listing
August 2017

Introduction.

J Med Primatol 2016 10;45(5):211

Tulane National Primate Research Center, Covington, LA, USA.

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http://dx.doi.org/10.1111/jmp.12246DOI Listing
October 2016

Th17 Cells Are Preferentially Infected Very Early after Vaginal Transmission of SIV in Macaques.

Cell Host Microbe 2016 Apr;19(4):529-40

Department of Cellular and Molecular Biology, Feinberg School of Medicine, Northwestern University, Chicago, IL 60611, USA. Electronic address:

The difficulty in detecting rare infected cells immediately after mucosal HIV transmission has hindered our understanding of the initial cells targeted by the virus. Working with the macaque simian immunodeficiency virus (SIV) vaginal challenge model, we developed methodology to identify discrete foci of SIV (mac239) infection 48 hr after vaginal inoculation. We find infectious foci throughout the reproductive tract, from labia to ovary. Phenotyping infected cells reveals that SIV has a significant bias for infection of CCR6+ CD4+ T cells. SIV-infected cells expressed the transcriptional regulator RORγt, confirming that the initial target cells are specifically of the Th17 lineage. Furthermore, we detect host responses to infection, as evidenced by apoptosis, cell lysis, and phagocytosis of infected cells. Thus, our analysis identifies Th17-lineage CCR6+ CD4+ T cells as primary targets of SIV during vaginal transmission. This opens new opportunities for interventions to protect these cells and prevent HIV transmission.
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http://dx.doi.org/10.1016/j.chom.2016.03.005DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4841252PMC
April 2016

Derivation and Characterization of a CD4-Independent, Non-CD4-Tropic Simian Immunodeficiency Virus.

J Virol 2016 05 29;90(10):4966-4980. Epub 2016 Apr 29.

Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, USA

Unlabelled: CD4 tropism is conserved among all primate lentiviruses and likely contributes to viral pathogenesis by targeting cells that are critical for adaptive antiviral immune responses. Although CD4-independent variants of human immunodeficiency virus (HIV) and simian immunodeficiency virus (SIV) have been described that can utilize the coreceptor CCR5 or CXCR4 in the absence of CD4, these viruses typically retain their CD4 binding sites and still can interact with CD4. We describe the derivation of a novel CD4-independent variant of pathogenic SIVmac239, termed iMac239, that was used to derive an infectious R5-tropic SIV lacking a CD4 binding site. Of the seven mutations that differentiate iMac239 from wild-type SIVmac239, a single change (D178G) in the V1/V2 region was sufficient to confer CD4 independence in cell-cell fusion assays, although other mutations were required for replication competence. Like other CD4-independent viruses, iMac239 was highly neutralization sensitive, although mutations were identified that could confer CD4-independent infection without increasing its neutralization sensitivity. Strikingly, iMac239 retained the ability to replicate in cell lines and primary cells even when its CD4 binding site had been ablated by deletion of a highly conserved aspartic acid at position 385, which, for HIV-1, plays a critical role in CD4 binding. iMac239, with and without the D385 deletion, exhibited an expanded host range in primary rhesus peripheral blood mononuclear cells that included CCR5(+) CD8(+) T cells. As the first non-CD4-tropic SIV, iMac239-ΔD385 will afford the opportunity to directly assess the in vivo role of CD4 targeting on pathogenesis and host immune responses.

Importance: CD4 tropism is an invariant feature of primate lentiviruses and likely plays a key role in pathogenesis by focusing viral infection onto cells that mediate adaptive immune responses and in protecting virions attached to cells from neutralizing antibodies. Although CD4-independent viruses are well described for HIV and SIV, these viruses characteristically retain their CD4 binding site and can engage CD4 if available. We derived a novel CD4-independent, CCR5-tropic variant of the pathogenic molecular clone SIVmac239, termed iMac239. The genetic determinants of iMac239's CD4 independence provide new insights into mechanisms that underlie this phenotype. This virus remained replication competent even after its CD4 binding site had been ablated by mutagenesis. As the first truly non-CD4-tropic SIV, lacking the capacity to interact with CD4, iMac239 will provide the unique opportunity to evaluate SIV pathogenesis and host immune responses in the absence of the immunomodulatory effects of CD4(+) T cell targeting and infection.
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http://dx.doi.org/10.1128/JVI.02851-15DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4859711PMC
May 2016

Zoonotic Potential of Simian Arteriviruses.

J Virol 2016 01 11;90(2):630-5. Epub 2015 Nov 11.

Department of Pathology and Laboratory Medicine, University of Wisconsin-Madison, Madison, Wisconsin, USA Wisconsin National Primate Research Center, Madison, Wisconsin, USA

Wild nonhuman primates are immediate sources and long-term reservoirs of human pathogens. However, ethical and technical challenges have hampered the identification of novel blood-borne pathogens in these animals. We recently examined RNA viruses in plasma from wild African monkeys and discovered several novel, highly divergent viruses belonging to the family Arteriviridae. Close relatives of these viruses, including simian hemorrhagic fever virus, have caused sporadic outbreaks of viral hemorrhagic fever in captive macaque monkeys since the 1960s. However, arterivirus infection in wild nonhuman primates had not been described prior to 2011. The arteriviruses recently identified in wild monkeys have high sequence and host species diversity, maintain high viremia, and are prevalent in affected populations. Taken together, these features suggest that the simian arteriviruses may be "preemergent" zoonotic pathogens. If not, this would imply that biological characteristics of RNA viruses thought to facilitate zoonotic transmission may not, by themselves, be sufficient for such transmission to occur.
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http://dx.doi.org/10.1128/JVI.01433-15DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4702702PMC
January 2016

Introduction.

J Med Primatol 2015 Oct;44(5):223

Tulane National Primate Research Center.

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http://dx.doi.org/10.1111/jmp.12197DOI Listing
October 2015

Antigenic requirement for Gag in a vaccine that protects against high-dose mucosal challenge with simian immunodeficiency virus.

Virology 2015 Feb 12;476:405-412. Epub 2015 Jan 12.

Yale University School of Medicine, New Haven, CT, United States. Electronic address:

We reported previously on a vaccine approach that conferred apparent sterilizing immunity to SIVsmE660. The vaccine regimen employed a prime-boost using vectors based on recombinant vesicular stomatitis virus (VSV) and an alphavirus replicon expressing either SIV Gag or SIV Env. In the current study, we tested the ability of vectors expressing only the SIVsmE660 Env protein to protect macaques against the same high-dose mucosal challenge. Animals developed neutralizing antibody levels comparable to or greater than seen in the previous vaccine study. When the vaccinated animals were challenged with the same high-dose of SIVsmE660, all became infected. While average peak viral loads in animals were slightly lower than those of previous controls, the viral set points were not significantly different. These data indicate that Gag, or the combination of Gag and Env are required for the generation of apparent sterilizing immunity to the SIVsmE660 challenge.
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http://dx.doi.org/10.1016/j.virol.2014.12.027DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4323353PMC
February 2015

Transmitted/founder simian immunodeficiency virus envelope sequences in vesicular stomatitis and Semliki forest virus vector immunized rhesus macaques.

PLoS One 2014 31;9(10):e109678. Epub 2014 Oct 31.

Division of Microbiology, Tulane National Primate Research Center, Tulane University, Covington, Louisiana, United States of America.

Identification of transmitted/founder simian immunodeficiency virus (SIV) envelope sequences responsible for infection may prove critical for understanding HIV/SIV mucosal transmission. We used single genome amplification and phylogenetic analyses to characterize transmitted/founder SIVs both in the inoculum and in immunized-infected rhesus monkeys. Single genome amplification of the SIVsmE660 inoculum revealed a maximum diversity of 1.4%. We also noted that the consensus sequence of the challenge stock differed from the vaccine construct in 10 amino acids including 3 changes in the V4 loop. Viral env was prepared from rhesus plasma in 3 groups of 6 immunized with vesicular stomatitis virus (VSV) vectors and boosted with Semliki forest virus (SFV) replicons expressing (a) SIVsmE660 gag-env (b) SIVsmE660 gag-env plus rhesus GM-CSF and (c) control influenza hemagglutinin protein. Macaques were immunized twice with VSV-vectors and once with SFV vector and challenged intrarectally with 4000 TCID50. Single genome amplification characterized the infections of 2 unprotected animals in the gag-env immunized group, both of which had reduced acute plasma viral loads that ended as transient infections indicating partial immune control. Four of 6 rhesus were infected in the gag-env + GM-CSF group which demonstrated that GM-CSF abrogated protection. All 6 animals from the control group were infected having high plasma viral loads. We obtained 246 full-length envelope sequences from SIVsmE660 infected macaques at the peak of infection and determined the number of transmitted/founder variants per animal. Our analysis found that 2 of 2 gag-env vaccinated but infected macaques exhibited single but distinct virus envelope lineages whereas rhesus vaccinated with gag-env-GM-CSF or HA control exhibited both single and multiple env lineages. Because there were only 2 infected animals in the gag-env vaccinated rhesus compared to 10 infected rhesus in the other 2 groups, the significance of finding single env variants in the gag-env vaccinated group could not be established.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0109678PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4215841PMC
September 2015

Introduction.

J Med Primatol 2014 Oct;43(5):293

Tulane National Primate Research Center, Covington, LA, USA.

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http://dx.doi.org/10.1111/jmp.12143DOI Listing
October 2014

Infection dynamics of sylvatic dengue virus in a natural primate host, the African Green Monkey.

Am J Trop Med Hyg 2014 Oct 4;91(4):672-6. Epub 2014 Aug 4.

Department of Biology, New Mexico State University, Las Cruces, New Mexico; Laboratory of Infectious Diseases, National Institutes of Allergy and Infectious Diseases, Bethesda, Maryland; Center for Tropical Diseases, Department of Pathology and Center for Biodefense and Emerging Infectious Diseases, University of Texas Medical Branch, Galveston, Texas; Institute for Human Infections and Immunity and Center for Tropical Diseases, University of Texas Medical Branch, Galveston, Texas; Tulane National Primate Research Center, Tulane University, Covington, Louisiana.

The four serotypes of mosquito-borne dengue virus (DENV-1, -2, -3, and -4) that circulate in humans each emerged from an enzootic, sylvatic cycle in non-human primates. Herein, we present the first study of sylvatic DENV infection dynamics in a primate. Three African green monkeys were inoculated with 10(5) plaque-forming units (pfu) DENV-2 strain PM33974 from the sylvatic cycle, and one African green monkey was inoculated with 10(5) pfu DENV-2 strain New Guinea C from the human cycle. All four monkeys seroconverted (more than fourfold rise in 80% plaque reduction neutralization titer [PRNT80]) against the strain of DENV with which they were inoculated; only one (33%) of three monkeys infected with sylvatic DENV showed a neutralizing antibody response against human-endemic DENV. Virus was detected in two of three monkeys inoculated with sylvatic DENV at low titer (≤ 1.3 log10pfu/mL) and brief duration (≤ 2 days). Clinical signs included rash and elevated aspartate aminotransferase (AST) and alanine aminotransferase (ALT) levels.
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http://dx.doi.org/10.4269/ajtmh.13-0492DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4183386PMC
October 2014

Comparison of the vaginal environment of Macaca mulatta and Macaca nemestrina throughout the menstrual cycle.

Am J Reprod Immunol 2014 Apr 13;71(4):322-9. Epub 2014 Feb 13.

Tulane National Primate Research Center, Covington, LA, USA.

Problem: Pigtail macaques, Macaca nemestrina (PT), are more susceptible to vaginal transmission of simian immunodeficiency virus (SIV) and other sexually transmitted diseases (STD) than rhesus macaques (RM). However, comparative studies to explore the reasons for these differences are lacking.

Method Of Study: Here, we compared differences in hormone levels and vaginal mucosal anatomy and thickness of RM and PT through different stages of the menstrual cycle. Concentrations of plasma estradiol (E2) and progesterone (P4) were determined weekly, and vaginal biopsies examined at days 0 and 14 of the menstrual cycle.

Results: Consistent changes in vaginal epithelial thickness occurred at different stages of the menstrual cycle. In both species, the vaginal epithelium was significantly thicker in the follicular than in luteal phase. Keratinized epithelium was strikingly much more prominent in RM, especially during the luteal phase. Further, the vaginal epithelium was significantly thinner, and the P4:E2 ratio was higher in PT during luteal phase than RM.

Conclusions: Striking anatomic differences in the vaginal epithelium between rhesus and pigtail macaques combined with differences in P4:E2 ratio support the hypothesis that thinning and less keratinization of the vaginal epithelium may be involved in the greater susceptibility of pigtail macaques to vaginal transmission of SIV or other STD.
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http://dx.doi.org/10.1111/aji.12201DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3954453PMC
April 2014

Serial cervicovaginal exposures with replication-deficient SIVsm induce higher dendritic cell (pDC) and CD4+ T-cell infiltrates not associated with prevention but a more severe SIVmac251 infection of rhesus macaques.

J Acquir Immune Defic Syndr 2014 Apr;65(4):405-13

*Department of Immunology, The Wistar Institute, Philadelphia, PA; †Caribbean Primate Research Center and Animal Resources Center, University of Puerto Rico (UPR), San Juan, PR; ‡University of Minnesota Medical School, Minneapolis, MN; §Division of Biostatistics and Epidemiology, University of Massachusetts, Amherst, MA; ‖Tulane National Primate Research Center, Covington, LA; Departments of ¶Microbiology and #Internal Medicine, UPR Medical School, San Juan, PR; and **School of Biological Sciences and ††Nebraska Center for Virology, University of Nebraska, Lincoln, NE.

Objective: Intravaginal exposure to simian immunodeficiency virus (SIV) acutely recruits interferon-alpha (IFN-α) producing plasmacytoid dendritic cells (pDC) and CD4 T-lymphocyte targets to the endocervix of nonhuman primates. We tested the impact of repeated cervicovaginal exposures to noninfectious, defective SIV particles over 72 hours on a subsequent cervicovaginal challenge with replication competent SIV.

Methods: Thirty-four female Indian Rhesus macaques were given a 3-day twice-daily vaginal exposures to either SIVsmB7, a replication-deficient derivative of SIVsmH3 produced by a T lymphoblast CEMx174 cell clone (n = 16), or to CEM supernatant controls (n = 18). On the fourth day, animals were either euthanized to assess cervicovaginal immune cell infiltration or intravaginally challenged with SIVmac251. Challenged animals were tracked for plasma viral load and CD4 counts and euthanized at 42 days after infection.

Results: At the time of challenge, macaques exposed to SIVsmB7, had higher levels of cervical CD123 pDCs (P = 0.032) and CD4 T cells (P = 0.036) than those exposed to CEM control. Vaginal tissues showed a significant increase in CD4 T-cell infiltrates (P = 0.048) and a trend toward increased CD68 cellular infiltrates. After challenge, 12 SIVsmB7-treated macaques showed 2.5-fold greater daily rate of CD4 decline (P = 0.0408), and viral load rise (P = 0.0036) as compared with 12 control animals.

Conclusions: Repeated nonproductive exposure to viral particles within a short daily time frame did not protect against infection despite pDC recruitment, resulting instead in an accelerated CD4 T-cell loss with an increased rate of viral replication.
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http://dx.doi.org/10.1097/QAI.0000000000000047DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3943721PMC
April 2014

Comparative characterization of transfection- and infection-derived simian immunodeficiency virus challenge stocks for in vivo nonhuman primate studies.

J Virol 2013 Apr 13;87(8):4584-95. Epub 2013 Feb 13.

AIDS and Cancer Virus Program, SAIC-Frederick, Inc., Frederick National Laboratory for Cancer Research, Frederick, Maryland, USA.

Simian immunodeficiency virus (SIV) stocks for in vivo nonhuman primate models of AIDS are typically generated by transfection of 293T cells with molecularly cloned viral genomes or by expansion in productively infected T cells. Although titers of stocks are determined for infectivity in vitro prior to in vivo inoculation, virus production methods may differentially affect stock features that are not routinely analyzed but may impact in vivo infectivity, mucosal transmissibility, and early infection events. We performed a detailed analysis of nine SIV stocks, comprising five infection-derived SIVmac251 viral swarm stocks and paired infection- and transfected-293T-cell-derived stocks of both SIVmac239 and SIVmac766. Representative stocks were evaluated for (i) virus content, (ii) infectious titer, (iii) sequence diversity and polymorphism frequency by single-genome amplification and 454 pyrosequencing, (iv) virion-associated Env content, and (v) cytokine and chemokine content by 36-plex Luminex analysis. Regardless of production method, all stocks had comparable particle/infectivity ratios, with the transfected-293T stocks possessing the highest overall virus content and infectivity titers despite containing markedly lower levels of virion-associated Env than infection-derived viruses. Transfected-293T stocks also contained fewer and lower levels of cytokines and chemokines than infection-derived stocks, which had elevated levels of multiple analytes, with substantial variability among stocks. Sequencing of the infection-derived SIVmac251 stocks revealed variable levels of viral diversity between stocks, with evidence of stock-specific selection and expansion of unique viral lineages. These analyses suggest that there may be underappreciated features of SIV in vivo challenge stocks with the potential to impact early infection events, which may merit consideration when selecting virus stocks for in vivo studies.
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http://dx.doi.org/10.1128/JVI.03507-12DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3624367PMC
April 2013

DNA-based HIV vaccines do not induce generalized activation in mucosal tissue T cells.

Hum Vaccin Immunother 2012 Nov 30;8(11):1648-53. Epub 2012 Oct 30.

Department of Microbiology; Perelman School of Medicine; University of Pennsylvania; Philadelphia, PA USA.

HIV preferentially infects activated T cells, and activated mucosal CD4+ T cells are the primary sites of viral replication. One potential explanation for increased HIV acquisition rates in the STEP study is that vaccination with adenoviral (Ad) vectors increased CD4+ T cell activation levels at the site of infection, a concept that others and we continue to explore. Whether vaccination with HIV vaccine platforms increases the activation state of CD4+ T cells within peripheral tissues, such as the gastro-intestinal (GI) mucosa, is exceptionally important to determine as a vaccine safety measure, given the susceptibility of activated CD4+ T cells to HIV infection. In this study we examined whether vaccination with DNA plasmids and chemokine adjuvants alter the activation state of T cells within the GI mucosa, inguinal LN, and peripheral blood. T cell activation state was measured by expression of CD25, CD69, and HLA-DR over the course of the prime/boost study. DNA plasmid vaccination did not increase expression of any of these markers in the 3 tissues studied. Addition of the gut-homing chemokine TECK during DNA plasmid vaccination did not alter activation levels of CD4+ T cells at any of these sites. These findings indicate that DNA vaccines do not elicit generalized mucosal T cell activation. Thus, DNA platforms may be especially suitable for HIV vaccine development, where bystander activation could promote increased HIV transmission.
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http://dx.doi.org/10.4161/hv.22247DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3601139PMC
November 2012

Increased cellular immune responses and CD4+ T-cell proliferation correlate with reduced plasma viral load in SIV challenged recombinant simian varicella virus - simian immunodeficiency virus (rSVV-SIV) vaccinated rhesus macaques.

Virol J 2012 Aug 13;9:160. Epub 2012 Aug 13.

Division of Microbiology, Tulane National Primate Research Center, Tulane University School of Medicine, Covington, LA 70433, USA.

Background: An effective AIDS vaccine remains one of the highest priorities in HIV-research. Our recent study showed that vaccination of rhesus macaques with recombinant simian varicella virus (rSVV) vector - simian immunodeficiency virus (SIV) envelope and gag genes, induced neutralizing antibodies and cellular immune responses to SIV and also significantly reduced plasma viral loads following intravenous pathogenic challenge with SIVMAC251/CX1.

Findings: The purpose of this study was to define cellular immunological correlates of protection in rSVV-SIV vaccinated and SIV challenged animals. Immunofluorescent staining and multifunctional assessment of SIV-specific T-cell responses were evaluated in both Experimental and Control vaccinated animal groups. Significant increases in the proliferating CD4+ T-cell population and polyfunctional T-cell responses were observed in all Experimental-vaccinated animals compared with the Control-vaccinated animals.

Conclusions: Increased CD4+ T-cell proliferation was significantly and inversely correlated with plasma viral load. Increased SIV-specific polyfunctional cytokine responses and increased proliferation of CD4+ T-cell may be crucial to control plasma viral loads in vaccinated and SIVMAC251/CX1 challenged macaques.
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http://dx.doi.org/10.1186/1743-422X-9-160DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3485174PMC
August 2012

The adjuvanticity of an O. volvulus-derived rOv-ASP-1 protein in mice using sequential vaccinations and in non-human primates.

PLoS One 2012 17;7(5):e37019. Epub 2012 May 17.

Lindsley F. Kimball Research Institute, New York Blood Center, New York, New York, United States of America.

Adjuvants potentiate antigen-specific protective immune responses and can be key elements promoting vaccine effectiveness. We previously reported that the Onchocerca volvulus recombinant protein rOv-ASP-1 can induce activation and maturation of naïve human DCs and therefore could be used as an innate adjuvant to promote balanced Th1 and Th2 responses to bystander vaccine antigens in mice. With a few vaccine antigens, it also promoted a Th1-biased response based on pronounced induction of Th1-associated IgG2a and IgG2b antibody responses and the upregulated production of Th1 cytokines, including IL-2, IFN-γ, TNF-α and IL-6. However, because it is a protein, the rOv-ASP-1 adjuvant may also induce anti-self-antibodies. Therefore, it was important to verify that the host responses to self will not affect the adjuvanticity of rOv-ASP-1 when it is used in subsequent vaccinations with the same or different vaccine antigens. In this study, we have established rOv-ASP-1's adjuvanticity in mice during the course of two sequential vaccinations using two vaccine model systems: the receptor-binding domain (RBD) of SARS-CoV spike protein and a commercial influenza virus hemagglutinin (HA) vaccine comprised of three virus strains. Moreover, the adjuvanticity of rOv-ASP-1 was retained with an efficacy similar to that obtained when it was used for a first vaccination, even though a high level of anti-rOv-ASP-1 antibodies was present in the sera of mice before the administration of the second vaccine. To further demonstrate its utility as an adjuvant for human use, we also immunized non-human primates (NHPs) with RBD plus rOv-ASP-1 and showed that rOv-ASP-1 could induce high titres of functional and protective anti-RBD antibody responses in NHPs. Notably, the rOv-ASP-1 adjuvant did not induce high titer antibodies against self in NHPs. Thus, the present study provided a sound scientific foundation for future strategies in the development of this novel protein adjuvant.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0037019PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3355165PMC
January 2013

Viral vectored granulocyte-macrophage colony stimulating factor inhibits vaccine protection in an SIV challenge model: protection correlates with neutralizing antibody.

Vaccine 2012 Jun 23;30(28):4233-9. Epub 2012 Apr 23.

Yale University School of Medicine, New Haven, CT 06510, USA.

In a previous vaccine study, we reported significant and apparently sterilizing immunity to high-dose, mucosal, simian immunodeficiency virus (SIV) quasi-species challenge. The vaccine consisted of vectors based on vesicular stomatitis virus (VSV) expressing simian immunodeficiency virus (SIV) gag and env genes, a boost with propagating replicon particles expressing the same SIV genes, and a second boost with VSV-based vectors. Concurrent with that published study we had a parallel group of macaques given the same doses of vaccine vectors, but in addition, we included a third VSV vector expressing rhesus macaque GM-CSF in the priming immunization only. We report here that addition of the vector expressing GM-CSF did not enhance CD8 T cell or antibody responses to SIV antigens, and almost completely abolished the vaccine protection against high-dose mucosal challenge with SIV. Expression of GM-CSF may have limited vector replication excessively in the macaque model. Our results suggest caution in the use of GM-CSF as a vaccine adjuvant, especially when expressed by a viral vector. Combining vaccine group animals from this study and the previous study we found that there was a marginal but significant positive correlation between the neutralizing antibody to a neutralization resistant SIV Env and protection from infection.
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http://dx.doi.org/10.1016/j.vaccine.2012.04.046DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3367070PMC
June 2012

Evaluation of inhaled cidofovir as postexposure prophylactic in an aerosol rabbitpox model.

Antiviral Res 2012 Jan 29;93(1):204-8. Epub 2011 Nov 29.

Division of Microbiology, Tulane National Primate Research Center, Covington, LA, USA.

Smallpox is considered a biological threat based upon the possibility of deliberate reintroduction into the population, creating an urgent need for effective antivirals. The antiviral drug cidofovir (Cr) has shown to be effective against poxviruses, although route-specific nephrotoxicity has hampered its development for emergency post-exposure prophylaxis (PEP). In this study, we use a micronized dry powder formulation of pharmaceutical-grade Cr (NanoFOVIRTM; Nf) to treat rabbits exposed to aerosolized rabbitpox virus (RPXV) to further evaluate the effectiveness of direct drug delivery to the lung. Naïve rabbits were infected with RPXV by aerosol; three subsets received aerosolized Nf at 0.5, 1.0 or 1.75mg/kg daily for 3days post-exposure, positive and negative control groups received intravenous (IV) Cr treatments and no treatment, respectively. Nf groups showed an antiviral-dose associated survival of 50% (0.5mg/kg), 80% (1.0mg/kg) and 100% (1.75mg/kg). All animals (100%) from the IV-Cr treatment group and none (0%) from the untreated controls survived. Nf (1.75) protected rabbits from RPX at approximately 10% of the equivalent IV-Cr dose. A dose-related effect was observed in clinical development of RPX disease in Nf groups. Significant reduction of RPX-induced pathological changes was observed in Nf (1.75) and IV-Cr groups. Results suggest that Nf may be a viable antiviral for emergency post-exposure prophylaxis and should be evaluated in other models of poxviral disease.
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http://dx.doi.org/10.1016/j.antiviral.2011.11.013DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3505885PMC
January 2012

Symposium on Nonhuman Primate Models for AIDS. Introduction.

J Med Primatol 2011 Aug;40(4):205

Tulane National Primate Research Center, Covington, LA, USA.

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http://dx.doi.org/10.1111/j.1600-0684.2011.00493.xDOI Listing
August 2011