Correction: Antibiotic-induced microbiome perturbations are associated with significant alterations to colonic mucosal immunity in rhesus macaques.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Antibiotic-induced microbiome perturbations are associated with significant alterations to colonic mucosal immunity in rhesus macaques.

The diverse bacterial communities that colonize the gastrointestinal tract play an essential role in maintaining immune homeostasis through the production of critical metabolites such as short-chain fatty acids (SCFAs) and this can be disrupted by antibiotic use. However, few studies have addressed the effects of specific antibiotics longitudinally on the microbiome and immunity. We evaluated the effects of four specific antibiotics: enrofloxacin, cephalexin, paromomycin, and clindamycin, in healthy female rhesus macaques.

Evidence for persistence of the SHIV reservoir early after MHC haploidentical hematopoietic stem cell transplantation.

Allogeneic transplantation (allo-HCT) has led to the cure of HIV in one individual, raising the question of whether transplantation can eradicate the HIV reservoir. To test this, we here present a model of allo-HCT in SHIV-infected, cART-suppressed nonhuman primates. We infect rhesus macaques with SHIV-1157ipd3N4, suppress them with cART, then transplant them using MHC-haploidentical allogeneic donors during continuous cART.

Intestinal damage precedes mucosal immune dysfunction in SIV infection.

HIV and pathogenic SIV infection are characterized by mucosal dysfunction including epithelial barrier damage, loss of Th17 cells, neutrophil infiltration, and microbial translocation with accompanying inflammation. However, it is unclear how and when these contributing factors occur relative to one another. In order to determine whether any of these features initiates the cycle of damage, we longitudinally evaluated the kinetics of mucosal and systemic T-cell activation, microbial translocation, and Th17 cell and neutrophil frequencies following intrarectal SIV infection of rhesus macaques.

Emerging diagnostic challenges and characteristics of simian betaretrovirus infections in captive macaque colonies.

To better understand Simian betaretrovirus (SRV) seropositivity in virus-negative macaques, we transfused blood from SRV-infected or suspect donors into immunosuppressed naive recipients. Our results do not support typical SRV1-5 infection as the cause, but provide evidence for several possibilities including serological artifact, new/different SRV, or an endogenous virus.

Effects of Fecal Microbial Transplantation on Microbiome and Immunity in Simian Immunodeficiency Virus-Infected Macaques.

Unlabelled: An altered intestinal microbiome during chronic human immunodeficiency virus (HIV) infection is associated with mucosal dysfunction, inflammation, and disease progression. We performed a preclinical evaluation of the safety and efficacy of fecal microbiota transplantation (FMT) as a potential therapeutic in HIV-infected individuals. Antiretroviral-treated, chronically simian immunodeficiency virus (SIV)-infected rhesus macaques received antibiotics followed by FMT.

Enhancement of Microbiota in Healthy Macaques Results in Beneficial Modulation of Mucosal and Systemic Immune Function.

Given the critical role of mucosal surfaces in susceptibility to infection, it is imperative that effective mucosal responses are induced when developing efficacious vaccines and prevention strategies for infection. Modulating the microbiota in the gastrointestinal (GI) tract through the use of probiotics (PBio) is a safe and well-tolerated approach to enhance mucosal and overall health. We assessed the longitudinal impact of daily treatment with the VSL#3 probiotic on cellular and humoral immunity and inflammation in healthy macaques.

The presence of the carboxy-terminal fragment of fibronectin allows maintenance of non-human primate long-term hematopoietic repopulating cells during extended ex vivo culture and transduction.

Objective: Ex vivo expansion of primitive hematopoietic cells remains of interest for gene therapy and transplantation. Previous studies reported loss of repopulating activity following culture of cells for more than 4-7 days in the presence of cytokines or stromal cells. In the current study, we investigated whether prolonged culture and transduction in the presence of the carboxy-terminal portion of fibronectin (FN) could maintain or expand retrovirally transduced repopulating hematopoietic stem cells (HSCs).

Effect of chronic cytokine therapy on clonal dynamics in nonhuman primates.

Hematopoietic cytokines such as filgrastim are used extensively to stimulate granulocyte production or to mobilize hematopoietic progenitors into the circulation; however, their effect on more primitive hematopoietic progenitor and stem cells in vivo is unknown, particularly in large animals or humans. In particular, there is concern that chronic therapy with cytokines could result in stem cell exhaustion or clonal dominance; however, direct assessment of the dynamics of individual stem and progenitor cell clones in vivo has not been previously reported. A number of models can be proposed regarding the mechanisms by which the marrow responds to cytokine stimulation, including recruitment of previously quiescent clones, stimulation of proliferation of already active clones, or prevention of apoptosis of more mature progenitors from all clones.

Retrovirally transduced muscle-derived cells contribute to hematopoiesis at very low levels in the nonhuman primate model.

Recent studies have suggested a remarkable potential of adult stem cells from a variety of organs to give rise to cells of disparate organs, but evidence of such potential at a clonal level is lacking in most if not all studies to date. To assess directly the hematopoietic potential of muscle-derived cells in a relevant large animal, we initiated retroviral-tagging studies in the rhesus macaque to allow tracking at the clonal level by integration site analysis. Four rhesus macaques underwent transplantation with transduced muscle-derived cells after lethal irradiation followed by delayed infusion of an autologous hematopoietic graft.

Direct comparison of RD114-pseudotyped versus amphotropic-pseudotyped retroviral vectors for transduction of rhesus macaque long-term repopulating cells.

Recently, RD114 (feline endogenous retrovirus envelope protein)-pseudotyped retroviral particles have been shown to transduce human NOD/SCID repopulating cells efficiently. In this study, we compared directly transduction of repopulating cells with RD114-pseudotyped vector to that with standard amphotropic vector in the rhesus macaque model. G-CSF/SCF-mobilized CD34(+) rhesus peripheral blood cells were cultured in the presence of SCF, Flt-3 ligand, and MGDF on Retronectin-coated flasks.

Transient in vivo selection of transduced peripheral blood cells using antifolate drug selection in rhesus macaques that received transplants with hematopoietic stem cells expressing dihydrofolate reductase vectors.

One of the main obstacles for effective human gene therapy for hematopoietic disorders remains the achievement of an adequate number of genetically corrected blood cells. One approach to this goal is to incorporate drug resistance genes into vectors to enable in vivo selection of hematopoietic stem cells (HSCs). Although a number of drug resistance vectors enable HSC selection in murine systems, little is known about these systems in large animal models.

Prolonged multilineage clonal hematopoiesis in a rhesus recipient of CD34 positive cells marked with a RD114 pseudotyped oncoretroviral vector.

The ability to efficiently transfer a gene into repopulating hematopoietic stem cells would create many therapeutic opportunities. We have evaluated the ability of particles bearing an alternative envelope protein, that of the feline endogenous virus (RD114), to transduce stem cells in a nonhuman primate autologous transplantation model using rhesus macaques. We have previously shown this pseudotyped vector to be superior to the amphotropic vector at transducing cells in umbilical cord blood capable of establishing hematopoiesis in immunodeficient mice.

Retroviral transduction efficiency of G-CSF+SCF-mobilized peripheral blood CD34+ cells is superior to G-CSF or G-CSF+Flt3-L-mobilized cells in nonhuman primates.

Gene transfer experiments in nonhuman primates have been shown to be predictive of success in human clinical gene therapy trials. In most nonhuman primate studies, hematopoietic stem cells (HSCs) collected from the peripheral blood or bone marrow after administration of granulocyte colony-stimulating factor (G-CSF) + stem cell factor (SCF) have been used as targets, but this cytokine combination is not generally available for clinical use, and the optimum target cell population has not been systematically studied. In our current study we tested the retroviral transduction efficiency of rhesus macaque peripheral blood CD34(+) cells collected after administration of different cytokine mobilization regimens, directly comparing G-CSF+SCF versus G-CSF alone or G-CSF+Flt3-L in competitive repopulation assays.

Retroviral transduction and engraftment ability of primate hematopoietic progenitor and stem cells transduced under serum-free versus serum-containing conditions.

The ability to efficiently transduce hematopoietic stem and progenitor cells under serum-free conditions would be desirable for safety and standardization of clinical gene therapy protocols. Using rhesus macaques, we studied the transduction efficiency and engraftment ability of CD34-enriched SCF/G-CSF mobilized progenitor cells (PBSC) transduced with standard amphotropic marking vectors under serum-free and serum-containing conditions. Supernatants were collected from producer cells 16 hours after serum-free medium or medium containing 10% fetal calf serum was added.