Mamu-DQA1 allele and genotype frequencies in a randomly sampled breeding colony of rhesus macaques (Macaca mulatta)

We studied the allelic and genotypic distribution of the major histocompatibility class-II locus DQA1 observed in a random sample of Indian rhesus macaques (Macaca mulatta) from a major breeding facility in the United States. The DNA was isolated from whole blood samples collected between 1991 and 1994 from 65 Indian rhesus monkeys. Polymerase chain reaction-restriction fragment length polymorphism analysis (PCR-RFLP), which involves use of specific amplification of DQA1 exon 2 and subsequent restriction digestion of the 242-base pair fragment, was used to genotype the animals for the 20 known macaque (Mamu)-DQA1 alleles. Frequencies for four alleles (DQA1*240x, *2502, *2503 and *0102) differed significantly from those reported in a smaller sample of rhesus macaques from the German Primate Center. The modest genetic survey of Mamu-DQA1 genotypes presented here will be particularly useful in designing epidemiologic studies that investigate associations between immunogenetic background and disease susceptibility in macaque models of human disease.     

Tracking of dye-labeled lymphocytes in rhesus macaques

Abstract: Lymphocytes were isolated from rhesus monkeys and marked with a fluorescent lipophilic dye to monitor their distribution in vivo. Dye-labeled cells were either monitored by blood draws over a three-month period, or identified within peripheral organs upon autopsy. Lymphocyte labeling conditions were optimized. Dye-labeled lymphocytes could be detected in the circulation for at least 100 days by flow cytometry and fluorescence microscopy. Activated lymphocytes were removed from the circulation more rapidly than lymphocytes that had not been activated.     

A recombinant rhesus cytomegalovirus expressing enhanced green fluorescent protein retains the wild-type phenotype and pathogenicity in fetal macaques

To facilitate identification of rhesus cytomegalovirus (RhCMV)-infected cells, a recombinant virus expressing enhanced green fluorescent protein (EGFP), designated RhCMV-EGFP, was constructed. An expression cassette for EGFP under the control of the simian virus 40 (SV40) early promoter was inserted into the intergenic region between unique short 1 (US1) and US2 of the RhCMV genome by homologous recombination. RhCMV-EGFP exhibited comparable growth kinetics to that of wild-type virus in rhesus fibroblast cultures and retained its pathogenicity in monkey fetuses. Typical neurologic syndromes caused by CMV infection were observed in all fetuses experimentally inoculated with RhCMV-EGFP, as evidenced by sonographic and gross examinations. Systemic RhCMV infections were established in all fetuses, as viral antigen was detected in multiple organs and virus was isolated from fetal blood samples. The engineered viral genome was stable following rapid serial passages in vitro and multiple rounds of replication in vivo. Infected cells could be readily distinguished by green fluorescence both in tissue cultures and in the fetuses. In addition, EGFP expression was detected in various cell types that were permissive to RhCMV infection, consistent with a broad tissue tropism of the SV40 promoter. These results demonstrate that RhCMV can be successfully engineered without loss of wild-type replication and pathogenic potential. Further, the spectrum of cortical anomalies and the distribution of infected cells in the brain tissues indicated that RhCMV may have preferentially targeted immature neuronal cells. The pattern of RhCMV infection in the central nervous system may offer an explanation for the severe developmental outcomes associated with congenital human CMV infection early in gestation.     

Experimental infection of rhesus macaques with Streptococcus pneumoniae: a possible model for vaccine assessment

BACKGROUND: We explored the possibility of using normal adult rhesus macaques for the preclinical assessment of safety, immunogenicity, and efficacy of newly developed vaccines against Streptococcus pneumoniae infection of the lung. METHODS: Our primary objective was to determine whether an intra-bronchial inoculum of at least 10(6)S. pneumoniae colony-forming units, or one as high as 10(8)-10(9) organisms, could detectably survive in rhesus macaques for a period longer than 1-2 weeks. If so, we hypothesized, it would be possible to observe signs of pneumonia commonly observed in humans, and discriminate between vaccinated/protected animals and controls. Infection was detectable in bronchoalveolar lavage fluids 3-5 weeks post-inoculation. RESULTS: The clinical course of disease mimicked aspects of that of human pneumococcal pneumonia. Signs of inflammation typical of the disease in humans, such as elevated concentrations of neutrophils and of pro-inflammatory cytokines in bronchoalveolar lavage fluids were also observed. CONCLUSIONS: These findings underscore the utility of this model to assess the safety, immunogenicity, and efficacy of newly developed S. pneumoniae vaccines.     

Genetic variation in transferrin alleles of rhesus macaques, Macaca mulatta

The rhesus macaque displays an extensive polymorphism at the transferrin locus. A principal components analysis describes the variance and covariance of alleles at the transferrin locus in eight widely dispersed sample populations. Using an eigenvectorial representation of the covariance matrix and systematically approximated geographical locations the distribution of populations and transferrin alleles is compared. Alleles with high variance prove to be the determining factor in the placement of populations in a "genetic map" and provide a means for interpreting the low congruence of genetics and geography found.     

Testis size,epididymis weight,and sperm competition in rhesus macaques

The intensity of sperm competition is often measured using the gonadosomatic index (testes/body weight). But sperm competition could be mediated more by size of the epididymis than by size of the testicles, and little information is available on the relationship between testicular and epididymal size. We found that both organs were positively correlated in size among male rhesus macaques. Body weight accounted for over 70% of the variance in testicle size and volumetric estimates of testicle size accurately reflected testicle weight. We conclude that methods for ascertaining testicle size are accurate, but the covariation in size between testicles and epididymis will hamper understanding of the physiological mechanisms involved in sperm competition in primates. © 1993 Wiley-Liss, Inc.     

Vaccine-induced control of viral shedding following rhesus cytomegalovirus challenge in rhesus macaques

The use of animal models of human cytomegalovirus (HCMV) infection is critical to refine HCMV vaccine candidates. Previous reports have demonstrated that immunization of rhesus monkeys against rhesus cytomegalovirus (RhCMV) can reduce both local and systemic replication of RhCMV following experimental RhCMV challenge. These studies used prime/boost combinations of DNA expression plasmids alone or DNA priming and boosting with either inactivated virion particles or modified vaccinia virus Ankara (MVA) expressing the same antigens. Viral outcomes included reduced RhCMV replication at the site of subcutaneous inoculation and RhCMV viremia following intravenous inoculation. Since shedding of cytomegalovirus from mucosal surfaces is critical for horizontal transmission of the virus, DNA priming/MVA boosting was evaluated for the ability to reduce oral shedding of RhCMV following subcutaneous challenge. Of six rhesus monkeys vaccinated exclusively against RhCMV glycoprotein B (gB), phosphoprotein 65 (pp65), and immediate-early 1 (IE1), half showed viral loads in saliva that were lower than those of control monkeys by 1 to 3 orders of magnitude. Further, there was a strong association of memory pp65 T cell responses postchallenge in animals exhibiting the greatest reduction in oral shedding. These results highlight the fact that a DNA/MVA vaccination regimen can achieve a notable reduction in a critical parameter of viral replication postchallenge. The recently completed clinical trial of a gB subunit vaccine in which the rate of HCMV infection was reduced by 50% in the individuals receiving the vaccine is consistent with the results of this study suggesting that additional immunogens are likely essential for maximum protection in an outbred human population.     

Bioacoustic monitoring of aggression in group-housed rhesus macaques

Many captive primate facilities house rhesus macaques in multimale-multifemale social groups in large enclosures that simulate the natural social and environmental features characteristic of the species, enhancing their reproductive performance as well as their psychological well-being, yet one of the most difficult management problems in socially housed macaques is their propensity for exhibiting spontaneous bouts of deleterious aggression. To address this management problem, an automated bioacoustic monitoring system might be developed that is capable of detecting and forecasting problematic patterns of contact aggression. To evaluate the utility of this approach, this study examined the magnitude of aggression and the co-occurrence of certain vocalization types and aggression in 10 groups of rhesus macaques. The data confirmed aggression as a significant problem in rhesus groups and indicated that certain patterns of vocalizations are indicative of the type or level of aggression. The detection and classification of these vocalization types need further research to eventually design and implement an efficacious bioacoustic system for monitoring aggression in rhesus macaques.     

Characterization of dermatologic changes in geriatric rhesus macaques

Abstract: Nonhuman primates are frequently used for aging studies. We observed a high prevalence of skin disease among a group of geriatric rhesus monkeys (mean age ? 25 years; n ? 9) used in aging behavioral studies. Gross and histopathologic changes in the skin of these geriatric rhesus were compared with skin from control adult monkeys (mean age ? 10; n ? 4) and sun-exposed monkeys (mean age ? 11; n = 4) to characterize age-related skin changes. Biopsy specimens were taken from four specified skin locations (lateral to bridge of nose, ventral midline, dorsal midline, perineal area) and from additional areas where skin lesions were present. Samples were routinely processed and evaluated by light microscopy. Blood samples were collected and tested for estrogen, thyroid-stimulating hormone, triiodothyronine, thyroxine, and Cortisol levels. The axilla was swabbed and samples were obtained for bacterial culturing. All nine of the geriatric monkeys had notable dermal lesions, while one of the control monkeys and one of the sun-exposed monkeys had abnormal findings. Major gross findings included increased areas of erythematous skin, wrinkling, focal skin scaling, thinning of hair, foot calluses, and exudative lesions. Histologic skin changes included subacute dermatitis, acanthotic dermatitis, and a lesion resembling an early solar lentigo in the sun-exposed animal. These changes were not associated with hormonal abnormalities or bacterial pathogens. Histologic changes are compatible with nonspecific skin changes observed in elderly humans. This study establishes a baseline of dermatologic changes of the aging rhesus macaque.     

Fluctuating asymmetry in fetuses of diabetic rhesus macaques

This study examines dental fluctuating asymmetry (FA) in two samples of fetal rhesus monkeys, one composed of 19 fetuses from diabetic mothers (FDM) and the other of 20 fetuses from nondiabetic mothers. Seventeen measurements were taken on the deciduous dentition of right and left mandibles. The degree of FA was assessed by comparing FDM to fetuses of normal mothers by correlation between right and left sides, and analysis of variation differences between right and left sides. Significant FA was found for three traits based on the correlation between right and left sides and for seven traits by the between-treatment ratio of variance between sides. Distal teeth, both within and outside of a morphologic field, exhibit significantly greater FA than mesial teeth. Our results support the hypothesis that developmental instability is detectable by dental FA.     

Male rank,reproductive behavior,and reproductive success in free-ranging rhesus macaques

Paternity assessment through DNA fingerprinting by synthetic oligonucleotide probes was applied to one birth cohort in a social group of free-ranging rhesus macaques (Macaca mulatta) on Cayo Santiago. The 11 group males and 9 males from other groups were observed mating with the females. Paternity was determined for 11 of the 15 infants. Male dominance rank was not associated with reproductive success. High-ranking resident males (N=5) sired 27% of the infants born during a one-year study. Four of the 11 infants of known paternity were sired by males of other social groups. The four infants of unknown paternity were sired either by males not observed mating with the females or the low-ranking male who was not fingerprinted. Male dominance rank was not associated with reproductive activity during conception cycles. These results suggest that the effect of rank on male reproductive success is not a predictable correlation, but a conditional probability.     

Molecular evidence for rhesus lymphocryptovirus infection of epithelial cells in immunosuppressed rhesus macaques

Epstein-Barr virus (EBV) is a human oncogenic herpesvirus associated with epithelial cell and B-cell malignancies. EBV infection of B lymphocytes is essential for acute and persistent EBV infection in humans; however, the role of epithelial cell infection in the normal EBV life cycle remains controversial. The rhesus lymphocryptovirus (LCV) is an EBV-related herpesvirus that naturally infects rhesus macaques and can be used experimentally to model persistent B-cell infection and B-cell lymphomagenesis. We now show that the rhesus LCV can infect epithelial cells in immunosuppressed rhesus macaques and can induce epithelial cell lesions resembling oral hairy leukoplakia in AIDS patients. Electron microscopy, immunohistochemistry, and DNA-RNA in situ hybridization were used to identify the presence of a lytic rhesus LCV infection in these proliferative, hyperkeratotic, or parakeratotic epithelial cell lesions. These studies demonstrate that the rhesus LCV has tropism for epithelial cells, in addition to B cells, and is a relevant animal model system for studying the role of epithelial cell infection in EBV pathogenesis.     

Absence of deleterious effects of inbreeding in rhesus macaques

The limited availability of rhesus macaques through importation and the need for large numbers of them for use in biomedical research in this country have led to large- scale efforts to breed them domestically in captive groups. Since gene flow into the groups is usually restricted, consanguineous inbreeding can lead to inbreeding depression, which is characterized by increases in mortality, declines in fertility, and reductions in traits associated with physiological efficiency. For this reason, it is desirable to determine the level of inbreeding at which inbreeding depression occurs in rhesus macaques. I compared fertility, mortality, and the rate of weight gain in inbred and noninbred rhesus monkeys in three captive groups. There is no evidence of inbreeding depression in rhesus macaques whose inbreeding coefficients are ≤ 0.125, a level at which inbreeding depression is known to occur in some species.     

Leishmania (Leishmania) major-infected rhesus macaques (Macaca mulatta) develop varying levels of resistance against homologous re-infections

Seven rhesus macaques were infected intradermally with 10(7) promastigotes of Leishmania (Leishmania) major. All monkeys developed a localized, ulcerative, self-healing nodular skin lesion at the site of inoculation of the parasite. Non-specific chronic inflammation and/or tuberculoid-type granulomatous reaction were the main histopathological manifestations of the disease. Serum Leishmania-specific antibodies (IgG and IgG1) were detected by ELISA in all infected animals; immunoblot analyses indicated that numerous antigens were recognized. A very high degree of variability was observed in the parasite-specific cell-mediated immune responses [as detected by measuring delayed-type hypersensitivity (DTH) reaction, in vitro lymphocyte proliferation, and gamma interferon (IFN-gamma) production] for individuals over time post challenge. From all the recovered monkeys (which showed resolution of the lesions after 11 weeks of infection), 57.2% (4/7) and 28.6% (2/7) animals remained susceptible to secondary and tertiary infections, respectively, but the disease severity was altered (i.e. lesion size was smaller and healed faster than in the primary infection). The remaining monkeys exhibited complete resistance (i.e. no lesion) to each rechallenge. Despite the inability to consistently detect correlates of cell-mediated immunity to Leishmania or correlation between resistance to challenge and DTH, lymphocyte transformation or IFN-gamma production, partial or complete acquired resistance was conferred by experimental infection. This primate model should be useful for measuring vaccine effectiveness against the human disease.     

Primigravidity,body weight,and costs of rearing first offspring in rhesus macaques

Body size is associated with menarche and ovarian function, but the relationship to first conception is rarely examined. We conducted a longitudinal investigation of rhesus macaques, Macaca mulatta, to determine the effect of differences in body weight on both the age at first conception and survivorship of first progeny. Young females who became pregnant weighed significantly more than peers who remained barren, with weight changes for females who successfully raised offspring no different than those for females who did not produce offspring. Infant mortality among primiparae was not significantly greater than that among multiparae, although nearly twice as high. First-born males tended to have lower survivorship than first-born female offspring. We suggest that reproductive costs encountered by primiparous females are more likely to be modulated by immature neuroendocrine function than by inexperience, small body size, or infant suckling patterns. We conclude that body size influences probability of first conception, socioendocrine factors mediate the likelihood of infant survival, and primiparous production of male progeny seems to exert a greater reproductive cost than does production of female progeny. Am. J. Primatol. 46:135–144, 1998. © 1998 Wiley-Liss, Inc.     

Unprecedented polymorphism of Mhc-DRB region configurations in rhesus macaques

The rhesus macaque is an important model in preclinical transplantation research and for the study of chronic and infectious diseases, and so extensive knowledge of its MHC (MhcMamu) is needed. Nucleotide sequencing of exon 2 allowed the detection of 68 Mamu-DRB alleles. Although most alleles belong to loci/lineages that have human equivalents, identical Mhc-DRB alleles are not shared between humans and rhesus macaques. The number of -DRB genes present per haplotype can vary from two to seven in the rhesus macaque, whereas it ranges from one to four in humans. Within a panel of 210 rhesus macaques, 24 Mamu-DRB region configurations can be distinguished differing in the number and composition of loci. None of the Mamu-DRB region configurations has been described for any other species, and only one of them displays major allelic variation giving rise to a total of 33 Mamu-DRB haplotypes. In the human population, only five HLA-DRB region configurations were defined, which in contrast to the rhesus macaque exhibit extensive allelic polymorphism. In comparison with humans, the unprecedented polymorphism of the Mamu-DRB region configurations may reflect an alternative strategy of this primate species to cope with pathogens. Because of the Mamu-DRB diversity, nonhuman primate colonies used for immunological research should be thoroughly typed to facilitate proper interpretation of results. This approach will minimize as well the number of animals necessary to conduct experiments.