Postnatal pre- and postexposure passive immunization strategies: protection of neonatal macaques against oral simian-human immunodeficiency virus challenge.

Simian-human immunodeficiency viruses (SHIV) allow the evaluation of antiviral strategies that target the envelope glycoproteins of the human immunodeficiency virus 1 (HIV-1) in macaques. We previously protected neonates from oral challenge with cell-free SHIV-vpu+ by passive immunization with synergistic human neutralizing monoclonal antibodies (mAbs) (Baba et al., Nat Med 6:200-206, 2000). mAbs were administered prenatally to pregnant dams and postnatally to the neonates. Here, we used solely postnatal or postexposure mAb treatment, thus significantly reducing the amount of mAbs necessary. All neonatal monkeys were also protected with these abbreviated mAb regimens. Our results are directly relevant for humans because we used mAbs that target HIV-1 envelope glycoproteins. Thus, the large-scale use of passive immunization with neutralizing mAbs may be feasible in human neonates. The mAbs, being natural human proteins, can be expected to have low toxicity. Passive immunization has promise to prevent intrapartum as well as milk-borne virus transmission from HIV-1-infected women to their infants.

Protection of neonatal macaques against experimental SHIV infection by human neutralizing monoclonal antibodies.

Neonatal macaques were completely protected against oral challenge with SHIV-vpu+, a simian-human immunodeficiency virus that encodes the envelope gene of a laboratory-adapted HIV strain, by pre- and post-natal treatment with a triple combination of human neutralizing monoclonal antibodies (mAbs). The mAbs were directed either against the CD4 binding site, a glycosylation-dependent gp120 epitope, or against a linear epitope on gp41. This triple combination was highly synergistic in vitro and neutralized primary HIV completely. Subsequently, oral challenge was performed with pathogenic SHIV89.6P, an animal-passaged variant of a chimeric virus that encodes the envelope gene of the primary, dual-tropic HIV89.6. Only post-natal treatment with a similar triple mAb combination was used. One out of 4 mAb-treated infants was completely protected from infection. In the other 3 treated animals, there was a tendency towards lower peak viral RNA loads compared with untreated controls. Two out of 4 mAb-treated infants maintained normal CD4+ T-cell numbers, in contrast to all controls that had steep declines at 2 weeks post-challenge. We conclude that the triple mAb combination significantly protected the neonates, even against mucosal challenge with pathogenic SHIV89.6P. Passively administered synergistic human mAbs may play a role in preventing mother-infant transmission of HIV, both against intrapartum transmission as well as against infection through breast milk. As passive immunization is a tool to assess correlates of immune protection, we conclude that the epitopes recognized by the mAbs in our combinations are important for AIDS vaccine development. Future passive immunization studies may reveal other important conserved epitopes.

Passive immunization against oral AIDS virus transmission: an approach to prevent mother-to-infant HIV-1 transmission?

To develop immunoprophylaxis regimens against mother-to-child human immunodeficiency virus type 1 (HIV-1) transmission, we established a simian-human immunodeficiency virus (SHIV) model in neonatal macaques that mimics intrapartum mucosal virus exposure (T.W. Baba, J. Koch, E.S. Mittler et al: AIDS Res Hum Retroviruses 10:351-357, 1994). We protected four neonates from oral SHIV-vpu+ challenge by ante- and postpartum treatment with a synergistic triple combination of immunoglobulin (Ig) G1 human anti-HIV-1 neutralizing monoclonal antibodies (mAbs) (T.W. Baba, V. Liska, R. Hofmann-Lehmann et al: Nature Med 6:200-206, 2000), which recognize the CD4-binding site of Env, a glycosylation-dependent gp120, or a linear gp41 epitope. Two neonates that received only postpartum mAbs were also protected from oral SHIV-vpu+ challenge, indicating that postpartum treatment alone is sufficient. Next, we evaluated a similar mAb combination against SHIV89.6P, which encodes env of primary HIV89.6. One of four mAb-treated neonates was protected from infection and two maintained normal CD4+ T-cell counts. We conclude that the epitopes recognized by the three mAbs are important determinants for achieving protection. Combination immunoprophylaxis with synergistic mAbs seems promising to prevent maternal HIV-1 transmission in humans.

Postnatal passive immunization of neonatal macaques with a triple combination of human monoclonal antibodies against oral simian-human immunodeficiency virus challenge.

To develop prophylaxis against mother-to-child human immunodeficiency virus (HIV) transmission, we established a simian-human immunodeficiency virus (SHIV) infection model in neonatal macaques that mimics intrapartum mucosal virus exposure (T. W. Baba et al., AIDS Res. Hum. Retroviruses 10:351-357, 1994). Using this model, neonates were protected from mucosal SHIV-vpu(+) challenge by pre- and postnatal treatment with a combination of three human neutralizing monoclonal antibodies (MAbs), F105, 2G12, and 2F5 (Baba et al., Nat. Med. 6:200-206, 2000). In the present study, we used this MAb combination only postnatally, thereby significantly reducing the quantity of antibodies necessary and rendering their potential use in humans more practical. We protected two neonates with this regimen against oral SHIV-vpu(+) challenge, while four untreated control animals became persistently infected. Thus, synergistic MAbs protect when used as immunoprophylaxis without the prenatal dose. We then determined in vitro the optimal MAb combination against the more pathogenic SHIV89.6P, a chimeric virus encoding env of the primary HIV89.6. Remarkably, the most potent combination included IgG1b12, which alone does not neutralize SHIV89.6P. We administered the combination of MAbs IgG1b12, 2F5, and 2G12 postnatally to four neonates. One of the four infants remained uninfected after oral challenge with SHIV89.6P, and two infants had no or a delayed CD4(+) T-cell decline. In contrast, all control animals had dramatic drops in their CD4(+) T cells by 2 weeks postexposure. We conclude that our triple MAb combination partially protected against mucosal challenge with the highly pathogenic SHIV89.6P. Thus, combination immunoprophylaxis with passively administered synergistic human MAbs may play a role in the clinical prevention of mother-to-infant transmission of HIV type 1.

Hematologic and serum biochemical reference intervals for the chimpanzee (Pan troglodytes) categorized by age and sex.

Normal reference range intervals for hematologic and serum biochemical values in the chimpanzee (Pan troglodytes) have seldom been reported. The few studies that have been conducted either report values on the basis of a small number of animals, report values for all age groups or both sexes combined, or were designed specifically to document the effect of a particular condition on the normal range of hematologic and serum biochemical values. On the basis of data collected from 133 chimpanzees over a 17-year period, empirically based clinical reference ranges were derived to provide a guide for basic diagnostic and clinical care of chimpanzees. For either sex within each of four age groups, there is a table that summarizes serum biochemical and a table that summarizes hematologic values. These values are compared with prior values, and their importance in the care and well being of captive chimpanzee populations is discussed.

Surgical technique for ambulatory management of airsacculitis in a chimpanzee (Pan troglodytes).

PURPOSE: Bacterial infections of the air sac have been reported in many nonhuman primates. Approaches to the management of airsacculitis have included combinations of medical and surgical therapies. These strategies have often required repeated attempts to drain exudate from the affected air sac, as well as necessitating that the animal endure isolation or undergo intensive postoperative care before returning to its social group. METHODS: A stoma was created via deliberate apposition of the air sac lining and skin to allow continuous drainage. Antibiotic therapy based on culture and antimicrobial susceptibility of the air sac contents was administered while the chimpanzee remained in its social group. RESULTS: We were able to attain complete resolution of the infection after a course of oral antibiotic therapy. The stoma closed gradually over a three-week period, and the chimpanzee has remained free of infection since that time. CONCLUSION: Despite the severity of the air sac infection in this chimpanzee, we were able to resolve the infection easily, using a simple surgical technique. This method allowed treatment without interfering with social standing or subjection to repeated anesthetic and treatment episodes. This method could be a simple, useful alternative for managing airsacculitis in nonhuman primates.

Human neutralizing monoclonal antibodies of the IgG1 subtype protect against mucosal simian-human immunodeficiency virus infection.

Although maternal human immunodeficiency virus type 1 (HIV-1) transmission occurs during gestation, intrapartum and postpartum (by breast-feeding), 50-70% of all infected children seem to acquire HIV-1 shortly before or during delivery. Epidemiological evidence indicates that mucosal exposure is an important aspect of intrapartum HIV transmission. A simian immunodeficiency virus (SIV) macaque model has been developed that mimics the mucosal exposure that can occur during intrapartum HIV-1 transmission. To develop immunoprophylaxis against intrapartum HIV-1 transmission, we used SHIV-vpu+ (refs. 5,6), a chimeric simian-human virus that encodes the env gene of HIV-IIIB. Several combinations of human monoclonal antibodies against HIV-1 have been identified that neutralize SHIV-vpu+ completely in vitro through synergistic interaction. Here, we treated four pregnant macaques with a triple combination of the human IgG1 monoclonal antibodies F105, 2G12 and 2F5. All four macaques were protected against intravenous SHIV-vpu+ challenge after delivery. The infants received monoclonal antibodies after birth and were challenged orally with SHIV-vpu+ shortly thereafter. We found no evidence of infection in any infant during 6 months of follow-up. This demonstrates that IgG1 monoclonal antibodies protect against mucosal lentivirus challenge in neonates. We conclude that epitopes recognized by the three monoclonal antibodies are important determinants for achieving substantial protection, thus providing a rational basis for AIDS vaccine development.

Evaluation of cellular immune responses in rhesus monkeys subjected to adenovirus-mediated gene transfer into the cervix.

We reported previously that direct injection of a recombinant adenovirus (rAd), Ad5CMV-beta-gal, into the cervix of the rhesus monkey resulted in efficient beta-galactosidase expression in the cervix within 3 days. In these studies, we also observed the induction of anti-adenovirus (Ad)-specific immunoglobulin G responses after 22 days. In the continuation of evaluating the anti-Ad-specific immune responses resulting from this approach of gene targeting to the cervix, we measured the cellular immune responses. The introduction of Ad5CMV-beta-gal into the cervix by direct injection, but not by the abrasion technique, resulted in the induction of strong proliferative responses against extracts of cells infected with Ad5CMV-beta-gal but not against control uninfected cells. These responses were initially detected at 22 days postinjection and coincided with the abrogation of transgene expression. Significant levels of proliferative responses were maintained for < or =83 days. Multiple injections of rAds had no significant enhancing effect on either the level or longevity of the proliferative responses. At 3 days after the injection of Ad5CMV-beta-gal, when the transgene expression in the cervix was clearly evident, proliferative responses against the rAd were not detectable. However, the production of low but significant amounts of interleukin-10, a cytokine characteristic of T helper type 2 responses that promote humoral immune responses, was observed at the 3-day point in these animals. These results suggest that significant differences exist between the kinetics of transgene expression and the priming of specific host immune responses, and that these differences may be important for devising alternate strategies to improve techniques for Ad-mediated gene therapy of cervical cancer.

Oral transmission of primate lentiviruses.

Oral transmission of human immunodeficiency virus type 1 (HIV-1) is well documented in children who become infected postnatally through breast milk. In contrast, epidemiologic surveys have yielded conflicting data regarding oral HIV-1 transmission among adults, even though case reports have described seroconversion and the development of AIDS in adults whose only risk was oral-genital contact. To study oral virus transmission in primate models, we exposed rhesus macaques of various ages to cell-free simian immunodeficiency virus (SIV), including uncloned and molecularly cloned viruses. In neonates, viremia and AIDS developed after nontraumatic oral exposure to several SIV strains. Furthermore, chimeric simian human immunodeficiency viruses containing the HIV-1 envelope can also cross intact upper gastrointestinal mucosal surfaces in neonates. In adult macaques, infection and AIDS have resulted from well-controlled, nontraumatic, experimental oral exposure to different strains of SIV. These findings have implications for the risks of HIV-1 transmission during oral-genital contact.

Rev/RRE-independent Mason-Pfizer monkey virus constitutive transport element-dependent propagation of SIVmac239 vectors using a single round of replication assay.

In a step toward creating live-attenuated or DNA subunit vaccines for AIDS, the replication of simian immunodeficiency virus (SIV) was studied independently of the Rev and RRE (Rev-responsive element) regulatory system, over a single round. To accomplish this, the env gene of an SIV vector was made defective by the insertion of a SV40 promoter/enhancer hygromycin B phosphotransferase gene cassette. Using this vector as the backbone, molecular clones of SIV were generated that contained a mutated Rev, Rev(-), a deleted RRE, RRE(-), or both, Rev(-)RRE(-). It has been shown recently that human immunodeficiency virus type 1 (HIV-1) Rev and RRE functions can be replaced in vitro by a cis-acting sequence, constitutive transport element (CTE), from simian type D retroviruses. To determine whether such a cis-acting element from Mason-Pfizer monkey virus (MPMV) would substitute for SIV Rev and RRE functions, the MPMV CTE was inserted either into the Nef ORF or at the junction of vpx and vpr of our Rev(-), RRE(-), and Rev(-)RRE(-) SIV molecular clones. Cell-free viral stocks harvested from Cos cells following transfections of these molecular clones revealed that these stocks were infectious over a single round of replication; however, their replication was attenuated 16-fold compared to that of wild-type virus. In addition, our experiments revealed that CTE functions in a position-dependent manner such that its insertion at the junction of vpx and vpr attenuated SIV replication 8- to 12-fold compared to the attenuation observed when it was inserted in the nef region. Our results demonstrate that MPMV CTE is capable of substituting for SIV Rev and RRE functions, resulting in an attenuated ability to produce infectious virus.

Maintenance of genetic variability in a specific pathogen-free breeding colony.

We used 18 genetic loci including blood groups, isozymes, and a serum protein to evaluate our efforts to preserve genetic variability in a specific pathogen-free (SPF) colony of rhesus monkeys. We compared genetic variability in the SPF population to the virally contaminated, non-SPF population from which it was derived. There was no change in the average gene diversity between the SPF and non-SPF populations. However, gene diversity at blood group Q locus increased significantly in the SPF population, while blood group M locus showed an insignificant trend toward decreased gene diversity. Allele frequencies changed significantly at blood group Q locus, although no alleles were lost from the population. We hypothesized that this change was due to extensive overreproduction by a small number of founder males that possessed the initially rare allele, Q1. There was no evidence that this change was associated with genes involved in viral infection.

Genetic structure of three populations of rhesus macaques (Macaca mulatta): Implications for genetic management.

One of the prime concerns at zoos and at primate breeding facilities is to maintain genetic variability. This can be accomplished by avoiding inbreeding. It is relatively easy to assess genetic variability and the level of inbreeding by using pedigree information and genetic markers. In this study we used genetic markers controlled by 6 independent polymorphic loci (GPI, PGD, CA2, MPI, DIA1, Tf) to ascertain genetic variation in two captive and one wild population of rhesus monkeys. Two other loci ADA and NP were also examined and found to be monomorphic in the three populations. F-statistics and contingency chi-square analyses indicated that there was significant genetic differentiation among the populations. We also found that the mean heterozygosities were very similar in the three populations, in spite of the diverse breeding strategies. These data are important because rhesus monkeys are frequently used for biomedical research; and the genetic markers provide useful information for genetic management of captive colonies of nonhuman primates. © 1992 Wiley-Liss, Inc.

Predisposition to invasive pneumococcal illness following parainfluenza type 3 virus infection in chimpanzees.

An outbreak of invasive disease, including pneumococcal bacteremia, meningitis, and pneumonia, involved 17 of 83 (20.5%) chimpanzees at a primate rehabilitation unit. Invasive disease was more common in splenectomized than in nonsplenectomized animals (42.9% vs 18.4%), but the difference was not statistically significant. The outbreak followed closely an outbreak of upper respiratory tract infection (URTI) that occurred with equal frequency in splenectomized and nonsplenectomized chimpanzees. Those with URTI were 5.7 times as likely to develop invasive disease than those without URTI (P less than 0.005). Fourteen of 20 (70%) chimpanzees with recent URTI and serologically examined had a 4-fold or greater rise in titer to parainfluenza type 3 virus. The outbreak of invasive disease occurred despite the fact that most of the chimpanzees had been vaccinated with pneumococcal vaccine. Efficacy of pneumococcal vaccine could not be demonstrated among any segment of the chimpanzee population, and testing of sera from 23 vaccinated chimpanzees against 4 pneumococcal serotypes (3, 6, 8, and 14) failed to show a meaningful immune response. The findings demonstrated that viral URTI can predispose primates to invasive infections and suggested that pneumococcal vaccine is not protective in chimpanzees.

Chimpanzee holding, rehabilitation and breeding: facilities design and colony management.

A multipurpose chimpanzee facility was designed and constructed to provide the unique housing requirements of rehabilitating laboratory-raised, behaviorally-deficient chimpanzees. The housing complex provided quarantine facilities, single or group housing for long-term holding, and semi-free-ranging compounds for established breeding groups. The facilities were designed to accommodate 170 animals of diverse ages and group configurations. Reproductive performance of the chimpanzees resulted in the production of 0.41 live infants per adult female year. Information was also given on the care, diet, maintenance, health status, handling, and rehabilitation of the chimpanzee. Development of this facility has made it practical to house large numbers of chimpanzees economically and conveniently in an enriched environment which promotes breeding and contributes to the survival of this irreplaceable animal model.

Reproductive, gestational, and newborn physiology of the chimpanzee.

The Yerkes Regional Primate Research Center has successfully bred chimpanzees since 1930. Breeding statistics for the last 6 yr and problems of fetal waste and infant mortality are presented here. Experience with a potential advantages of semi-free-ranging breeding systems are presented. This 6-yr study period also produced considerable information on gestational and newborn physiology.

A quantitative and morphological study of the pigmentary system of the chimpanzee with the light and electron microscope.

The epidermal melanocyte system of the chimpaneze was studied by the combined skin-splitting DOPA, and electron microscopic techniques. It is very similar to man. There are DOPA-positive epidermal melanocytes in all body regions regradless of the degree of macroscopic skin pigmentation or hirsutism. Furthermore, as in man, but in contrast to rodents, chimpanzee skin contains a very high level of melanocytes in the epidermis; approximately 3,320+/-350 per square millimeter skin. Chimpanzee melanosomes are long, wide, and fully melanized. In keratinocytes, these organelles are individually dispersed in all body regions, regardless of the degree of skin color, as is true for other mammalian species with large melanosomes.