Nonhuman Primates In Biomedical Research Research Articles

The Impact and Effects of Host Immunogenetics on Infectious Disease Studies Using Non-Human Primates in Biomedical Research.

Understanding infectious disease pathogenesis and evaluating novel candidate treatment interventions for human use frequently requires prior or parallel analysis in animal model systems. While rodent species are frequently applied in such studies, there are situations where non-human primate (NHP) species are advantageous or required. These include studies of animals that are anatomically more akin to humans, where there is a need to interrogate the complexity of more advanced biological systems or simply reflect susceptibility to a specific infectious agent. The contribution of different arms of the immune response may be addressed in a variety of NHP species or subspecies in specific physiological compartments. Such studies provide insights into immune repertoires not always possible from human studies. However, genetic variation in outbred NHP models may confound, or significantly impact the outcome of a particular study. Thus, host factors need to be considered when undertaking such studies. Considerable knowledge of the impact of host immunogenetics on infection dynamics was elucidated from HIV/SIV research. NHP models are now important for studies of emerging infections. They have contributed to delineating the pathogenesis of SARS-CoV-2/COVID-19, which identified differences in outcomes attributable to the selected NHP host. Moreover, their use was crucial in evaluating the immunogenicity and efficacy of vaccines against COVID-19 and establishing putative correlates of vaccine protection. More broadly, neglected or highly pathogenic emerging or re-emergent viruses may be studied in selected NHPs. These studies characterise protective immune responses following infection or the administration of candidate immunogens which may be central to the accelerated licensing of new vaccines. Here, we review selected aspects of host immunogenetics, specifically MHC background and TRIM5 polymorphism as exemplars of adaptive and innate immunity, in commonly used Old and New World host species. Understanding this variation within and between NHP species will ensure that this valuable laboratory source is used most effectively to combat established and emerging virus infections and improve human health worldwide.

Association of Primate Veterinarians Guidelines for Cerebrospinal Fluid Aspiration for Nonhuman Primates in Biomedical Research.

Association of Primate Veterinarians Guidelines for Cerebrospinal Fluid Aspiration for Nonhuman Primates in Biomedical Research.

Optimization of Milk Substitutes for the Artificial Rearing of Chinese Tree Shrews (Tupaia belangeri chinensis).

Simple SummaryThe Chinese tree shrew, a squirrel-like mammal, has been widely used as a laboratory animal in biological research. However, the low survival rate of the pups has seriously hindered the establishment of inbred lines of this species and further limited its wider use. We found a milk substitute appropriate for artificial rearing of Chinese tree shrew pups independent of any obvious adverse effects on their survival, health, and reproductive performance compared to those of the maternally reared pups. The successful optimization of a milk substitute for the artificial rearing of Chinese tree shrew pups may increase the availability of this experimental animal.The Chinese tree shrew (Tupaia belangeri chinensis) has the potential to replace the use of non-human primates in biomedical research. To increase the availability of this species, we have undertaken the ambitious task of establishing inbred lines of the Chinese tree shrew; however, we have been hindered by a low survival rate of inbred pups. Here, we report our artificial rearing (AR) of Chinese tree shrew pups using four different milk substitutes: the formula described by Tsang and Collins (milk TC) and three commercially available milk substitutes intended for possums (milk A and milk C) and for guinea pigs (milk B). We compared the effects of these milk substitutes and maternal milk on the daily milk consumption, growth performance, and survival of the pups. We also assessed the life span and reproductive performance of the F1 individuals given the best milk substitute as compared to the maternally reared (MR) pups. Milk B was found to be appropriate for AR. Pups fed with milk B had a high survival rate at the weaning age compared to those fed with the other milk substitutes. The AR pups fed with milk B had a life span similar to that of MR pups. AR females fed with milk B had an earlier age of the first reproduction, a larger number of litters, and a higher rate of survival of the offspring at the weaning age compared with the MR females. The successful optimization of a milk substitute for AR of Chinese tree shrew pups will undoubtedly facilitate the wide usage of this experimental animal.

Compositional analysis of chondroitin/dermatan sulfate in rhesus monkeys (Macaca mulatta).

Glycosaminoglycans (GAGs) are found in various tissues and are involved in many physiological functions. Since the rhesus monkey (Macaca mulatta) is the most widely used nonhuman primate in biomedical research, an understanding of the compositions of GAGs in their tissues is important. The aim of this study was to determine the content and sulfation pattern of disaccharides contained in several tissues of the rhesus monkey. The chondroitin sulfate (CS)/dermatan sulfate (DS) hybrid chain was extracted from several tissues of female and male rhesus monkeys. Compositional analysis was performed after digestion with chondroitinases ABC and ACI to reveal the sulfation pattern of the CS/DS hybrid chain. This study revealed that the major CS/DS disaccharide units present in the tissues were A and C types. The E and iE types were specifically distributed not only in the tracheal tissue but also in gastrointestinal tissues.

Association of Primate Veterinarians Cranial Implant Care for Nonhuman Primates in Biomedical Research.

Association of Primate Veterinarians Cranial Implant Care for Nonhuman Primates in Biomedical Research.

Effect of Housing Conditions on Cortisol and Body Fat Levels in Female Rhesus Macaques.

Simple SummaryMacaques are highly social animals and commonly used in biomedical research. These studies often require group-living animals to be pair-housed in a controlled environment. This controlled environment likely affects stress and body fat levels. This study investigates the effect of a change from group- to pair-housing on cortisol, as measure for stress, and body fat levels of 32 female rhesus macaques. Besides individual differences, cortisol levels were higher in pair-housing compared to group-housing. Body fat levels did not differ between housing conditions. Accordingly, there was no clear association between cortisol and body fat levels.Macaques are among the most commonly used non-human primates in biomedical research. They are highly social animals, yet biomedical studies often require group-living animals to be pair-housed in a controlled environment. A change in environment causes only short-term stress in adapting individuals, while non-adapting animals may experience long-term stress that can adversely affect study results. Individuals likely differ in their ability to adapt depending on individual characteristics. Changes in cortisol and body fat levels may reflect these different individual responses. Here, we investigate the long-term effect of a change from group- to pair-housing on cortisol and body fat levels in 32 female rhesus macaques, exploring whether age, dominance rank, original cortisol, and body fat levels are related to long-term stress in pair-housing. Hair samples were analyzed for cortisol levels, while anthropometric measurements and computed tomography were performed to quantify body fat. Monkeys served as their own control with a 7.5-month period between the measurements. Cortisol levels increased, while average body fat levels did not differ when individuals were moved from group- to pair-housing. Cortisol and body fat levels were not significantly correlated. Changes in cortisol were independent of age and dominance rank, whereas individual variation in body fat alterations was related to the group-housed body fat level and dominance rank. Although this study did not identify individual characteristics related to long-term stress in pair-housing, the individual variation confirms that some individuals are more resilient to change than others and provides possibilities for future refinement studies.

Generation of intestinal chemosensory cells from nonhuman primate organoids

Several gastrointestinal epithelial cells are involved in taste signal transduction. Although rodent tissues are extensively used as a human gut model, recent studies show that the chemical sensing system in rodents differs from that in humans. Nonhuman primates in biomedical research are valuable animal models to advance our understanding of biological responses in humans. The 3D organoid culture produces functional gastrointestinal epithelial cells in vitro and can be generated from animal and human tissues. Here, we report the generation of intestinal chemosensory cells from nonhuman primates, macaques, using an organoid culture system. We were able to maintain macaque intestinal organoids in the proliferation medium for more than six months. Upon switching to differentiation medium, we observed a drastic change in organoid morphology and chemosensory cell marker protein expression. This switch from proliferation to differentiation was confirmed by transcriptome analysis of the duodenum, jejunum, and ileum organoids. We further observed that the supplementation of culture media with interleukin (IL)-4 or the Notch inhibitor dibenzazepine (DBZ) accelerated terminal cell differentiation into chemosensory cells.Overall, we generated monkey intestinal organoids for the first time. These organoids are suitable for studying the function of primate chemosensory cells.

Reduced meiotic recombination in rhesus macaques and the origin of the human recombination landscape.

Characterizing meiotic recombination rates across the genomes of nonhuman primates is important for understanding the genetics of primate populations, performing genetic analyses of phenotypic variation and reconstructing the evolution of human recombination. Rhesus macaques (Macaca mulatta) are the most widely used nonhuman primates in biomedical research. We constructed a high-resolution genetic map of the rhesus genome based on whole genome sequence data from Indian-origin rhesus macaques. The genetic markers used were approximately 18 million SNPs, with marker density 6.93 per kb across the autosomes. We report that the genome-wide recombination rate in rhesus macaques is significantly lower than rates observed in apes or humans, while the distribution of recombination across the macaque genome is more uniform. These observations provide new comparative information regarding the evolution of recombination in primates.

Ethical and Welfare Implications of Genetically Altered Non-Human Primates for Biomedical Research.

Breakthroughs in gene editing technologies have made it feasible to create genetically altered (GA) non-human primate (NHP) models of disease. This area of research is accelerating, particularly in China, Japan and the USA, and could lead to an increase in NHP use globally. The hope is that genetic models in animal species closely related to humans will significantly improve understanding of neurological diseases and validation of potential therapeutic interventions, for which there is a dire need. However, the creation and use of GA NHPS raises serious animal welfare and ethical issues, which are highlighted here. It represents a step change in how these highly sentient animals are used in biomedical research, because of the large numbers required, inherent wastage and the sum of the harms caused to the animals involved. There is little evidence of these important issues being addressed alongside the rapidly advancing science. We are still learning about how gene editing tools work in NHPS, and significant added scientific and medical benefit from GA NHP models has yet to be demonstrated. Together, this suggests that current regulatory and review frameworks, in some jurisdictions at least, are not adequately equipped to deal with this emerging, complex area of NHP use.

MACSNVdb: a high-quality SNV database for interspecies genetic divergence investigation among macaques.

Macaques are the most widely used non-human primates in biomedical research. The genetic divergence between these animal models is responsible for their phenotypic differences in response to certain diseases. However, the macaque single nucleotide polymorphism resources mainly focused on rhesus macaque (Macaca mulatta), which hinders the broad research and biomedical application of other macaques. In order to overcome these limitations, we constructed a database named MACSNVdb that focuses on the interspecies genetic diversity among macaque genomes. MACSNVdb is a web-enabled database comprising ~74.51 million high-quality non-redundant single nucleotide variants (SNVs) identified among 20 macaque individuals from six species groups (muttla, fascicularis, sinica, arctoides, silenus, sylvanus). In addition to individual SNVs, MACSNVdb also allows users to browse and retrieve groups of user-defined SNVs. In particular, users can retrieve non-synonymous SNVs that may have deleterious effects on protein structure or function within macaque orthologs of human disease and drug-target genes. Besides position, alleles and flanking sequences, MACSNVdb integrated additional genomic information including SNV annotations and gene functional annotations. MACSNVdb will facilitate biomedical researchers to discover molecular mechanisms of diverse responses to diseases as well as primatologist to perform population genetic studies. We will continue updating MACSNVdb with newly available sequencing data and annotation to keep the resource up to date. Database URL: http://big.cdu.edu.cn/macsnvdb/

An IQ Consortium Perspective on The Scientific Committee on Health, Environmental and Emerging Risks Final Opinion on the Need for Nonhuman Primates in Biomedical Research, Production and Testing of Products and Devices (Update 2017).

The recent Scientific Committee on Health, Environmental and Emerging Risks Final Opinion on "The need for nonhuman primates in biomedical research, production and testing of products and devices" (2017 SCHEER) highlights approaches that could significantly contribute to the replacement, reduction, and refinement of nonhuman primate (NHP) studies. Initiatives that have the potential to affect NHP welfare and/or their use are expected to be appropriate, fair, and objective and publicly disseminated information focused on NHPs in biomedical research, which includes toxicologic and pathologic research and testing, should be objectively evaluated by stakeholder scientists, researchers, and veterinarians. Thus, IQ Consortium member companies convened to develop an informed and objective response, focusing on identifying areas of agreement, potential gaps, or missing information in 2017 SCHEER. Overall, the authors agree that many positions in the 2017 SCHEER Opinion generally align with industry views on the use of NHPs in research and testing, including the ongoing need of NHPs in many areas of research. From the perspective of the IQ Consortium, there are several topics in the 2017 SCHEER that merit additional comment, attention, or research, as well as consideration in future opinions.

High rates of aggression do not predict rates of trauma in captive groups of macaques

Socially inflicted traumas are a major concern for the management of captive groups of rhesus macaques. Rhesus macaques are the most commonly used nonhuman primate in biomedical research, and social housing is optimal for promoting psychological well-being. However, trauma is frequent due to a strong reliance on aggression to establish and maintain hierarchical relationships. We studied six captive groups of rhesus macaques (Macaca mulatta) that underwent a variety of social perturbations and explored whether rates of aggression mapped onto rates of trauma using a fine-grained analysis that divided both aggression and trauma variables into specific, behaviorally-relevant categories (e.g., severe aggression by adult males relative to lacerations). Results did not show the expected positive relationship between aggression variables and trauma variables. Instead, rates of trauma (i.e., lacerations, moderate-severe trauma, total trauma) were negatively associated with the rate of impartial interventions (i.e., an intervention directed at both targets during an ongoing conflict) during baseline periods. Additionally, rates of trauma (i.e., lacerations, punctures, moderate-severe trauma, total trauma) were negatively associated with rates of total aggression following temporary knockouts of the individuals who commonly intervene impartially (i.e., conflict policers), and punctures and moderate-severe trauma were negatively associated with rates of severe aggression by adult males following permanent knockout of a high-ranked natal male. These results suggest that under homeostatic conditions, impartial interventions serve as a mechanism to reduce socially inflicted trauma but, following social manipulations of high-ranking males, an imbalance emerges. Our results underscore the importance of developing management strategies for rhesus macaque groups that promote internal social mechanisms of social stability such as maintaining conflict policing individuals (i.e., adult males) in social groups.

Genetic analysis of samples from wild populations opens new perspectives on hybridization between long-tailed (Macaca fascicularis) and rhesus macaques (Macaca mulatta).

In the past decade, many researchers have published papers about hybridization between long-tailed and rhesus macaques. These previous works have proposed unidirectional gene flow with the Isthmus of Kra as the zoogeographical barrier of hybridization. However, these reports analyzed specimens of unknown origin and/or did not include specimens from Thailand, the center of the proposed area of hybridization. Collected specimens of long-tailed and rhesus macaques representing all suspected hybridization areas were examined. Blood samples from four populations each of long-tailed and rhesus macaques inhabiting Thailand, Myanmar, and Laos were collected and analyzed with conspecific references from China (for rhesus macaques) and multiple countries from Sundaic regions (for long-tailed macaques). Ninety-six single nucleotide polymorphism (SNP) markers specifically designed to interrogate admixture and ancestry were used in genotyping. We found genetic admixture maximized at the hybrid zone (15-20°N), as well as admixture signals of varying strength in both directions outside of the hybrid zone. These findings show that the Isthmus of Kra is not a barrier to gene flow from rhesus to long-tailed populations. However, to precisely identify a southernmost barrier, if in fact a boundary rather than simple isolation by distance exists, the samples from peninsular Malaysia must be included in the analysis. Additionally, a long-tailed to rhesus gene flow boundary was found between northern Thailand and Myanmar. Our results suggest that selection of long-tailed and rhesus macaques, the two most commonly used non-human primates for biomedical research, should take into account not only the species identification but also the origin of and genetic admixture within and between the species.

Analysis of Simian Immunodeficiency Virus-specific CD8+ T-cells in Rhesus Macaques by Peptide-MHC-I Tetramer Staining.

Peptide-major histocompatibility complex class I (pMHC-I) tetramers have been an invaluable tool to study CD8+ T-cell responses. Because these reagents directly bind to T-cell receptors on the surface of CD8+ T-lymphocytes, fluorochrome-labeled pMHC-I tetramers enable the accurate detection of antigen (Ag)-specific CD8+ T-cells without the need for in vitro re-stimulation. Moreover, when combined with multi-color flow cytometry, pMHC-I tetramer staining can reveal key aspects of Ag-specific CD8+ T-cells, including differentiation stage, memory phenotype, and activation status. These types of analyses have been especially useful in the field of HIV immunology where CD8+ T-cells can affect progression to AIDS. Experimental infection of rhesus macaques with simian immunodeficiency virus (SIV) provides an invaluable tool to study cellular immunity against the AIDS virus. As a result, considerable progress has been made in defining and characterizing T-cell responses in this animal model. Here we present an optimized protocol for enumerating SIV-specific CD8+ T-cells in rhesus macaques by pMHC-I tetramer staining. Our assay permits the simultaneous quantification and memory phenotyping of two pMHC-I tetramer+ CD8+ T-cell populations per test, which might be useful for tracking SIV-specific CD8+ T-cell responses generated by vaccination or SIV infection. Considering the relevance of nonhuman primates in biomedical research, this methodology is applicable for studying CD8+ T-cell responses in multiple disease settings.

The population genomics of rhesus macaques (Macaca mulatta) based on whole-genome sequences.

Rhesus macaques (Macaca mulatta) are the most widely used nonhuman primate in biomedical research, have the largest natural geographic distribution of any nonhuman primate, and have been the focus of much evolutionary and behavioral investigation. Consequently, rhesus macaques are one of the most thoroughly studied nonhuman primate species. However, little is known about genome-wide genetic variation in this species. A detailed understanding of extant genomic variation among rhesus macaques has implications for the use of this species as a model for studies of human health and disease, as well as for evolutionary population genomics. Whole-genome sequencing analysis of 133 rhesus macaques revealed more than 43.7 million single-nucleotide variants, including thousands predicted to alter protein sequences, transcript splicing, and transcription factor binding sites. Rhesus macaques exhibit 2.5-fold higher overall nucleotide diversity and slightly elevated putative functional variation compared with humans. This functional variation in macaques provides opportunities for analyses of coding and noncoding variation, and its cellular consequences. Despite modestly higher levels of nonsynonymous variation in the macaques, the estimated distribution of fitness effects and the ratio of nonsynonymous to synonymous variants suggest that purifying selection has had stronger effects in rhesus macaques than in humans. Demographic reconstructions indicate this species has experienced a consistently large but fluctuating population size. Overall, the results presented here provide new insights into the population genomics of nonhuman primates and expand genomic information directly relevant to primate models of human disease.

The well-being of laboratory non-human primates.

The well-being of non-human primates in captivity is of joint concern to scientists, veterinarians, colony managers, caretakers, and researchers working with non-human primates in biomedical research. With increased regulatory, accreditation, and research focus on optimizing the use of social housing for laboratory primates, as well as the advent of techniques to assess indices of chronic stress and related measures of well-being, there is no better time to present the most current advances in the field of non-human primate behavioral management. The collective body of research presented here was inspired in part by a 2014 symposium entitled, "Chronic Hormones and Demographic Variables: Center-Wide Studies on Non-Human Primate Well-Being" held at the American Society of Primatologists' 37th Annual Meeting in Decatur, GA. By aiming to target readership with scientific and/or management oversight of captive primate behavioral management programs, this special issue provides badly-needed guidance for implementing social housing programs in a research environment and leverages collaboration across multiple facilities to address key components of behavioral management, explore refinements in how well-being can be measured, and identify the interrelationships between varying indices. Am. J. Primatol. 79:e22520, 2017. © 2016 Wiley Periodicals, Inc.

Tree shrews at the German Primate Center

Abstract. For many years, Tupaia (family Tupaiidae), most commonly known as tree shrews, have been studied almost exclusively by zoologists resulting in a controversial debate on their taxonomic status among mammals. Today, tree shrews are placed in the order Scandentia; they are valuable, widely accepted and increasingly used model animals as an alternative to rodents and non-human primates in biomedical research. After a brief description on how tree shrews entered science and their taxonomic odyssey, the present article describes the history of the tree shrew (Tupaia belangeri) colony at the German Primate Center and selected aspects of our work with special emphasis on the psychosocial stress model in these animals.

Sequencing strategies and characterization of 721 vervet monkey genomes for future genetic analyses of medically relevant traits.

BackgroundWe report here the first genome-wide high-resolution polymorphism resource for non-human primate (NHP) association and linkage studies, constructed for the Caribbean-origin vervet monkey, or African green monkey (Chlorocebus aethiops sabaeus), one of the most widely used NHPs in biomedical research. We generated this resource by whole genome sequencing (WGS) of monkeys from the Vervet Research Colony (VRC), an NIH-supported research resource for which extensive phenotypic data are available.ResultsWe identified genome-wide single nucleotide polymorphisms (SNPs) by WGS of 721 members of an extended pedigree from the VRC. From high-depth WGS data we identified more than 4 million polymorphic unequivocal segregating sites; by pruning these SNPs based on heterozygosity, quality control filters, and the degree of linkage disequilibrium (LD) between SNPs, we constructed genome-wide panels suitable for genetic association (about 500,000 SNPs) and linkage analysis (about 150,000 SNPs). To further enhance the utility of these resources for linkage analysis, we used a further pruned subset of the linkage panel to generate multipoint identity by descent matrices.ConclusionsThe genetic and phenotypic resources now available for the VRC and other Caribbean-origin vervets enable their use for genetic investigation of traits relevant to human diseases.Electronic supplementary materialThe online version of this article (doi:10.1186/s12915-015-0152-2) contains supplementary material, which is available to authorized users.

Whole-genome sequencing of six Mauritian Cynomolgus macaques (Macaca fascicularis) reveals a genome-wide pattern of polymorphisms under extreme population bottleneck.

Cynomolgus macaques (Macaca fascicularis) were introduced to the island of Mauritius by humans around the 16th century. The unique demographic history of the Mauritian cynomolgus macaques provides the opportunity to not only examine the genetic background of well-established nonhuman primates for biomedical research but also understand the effect of an extreme population bottleneck on the pattern of polymorphisms in genomes. We sequenced the whole genomes of six Mauritian cynomolgus macaques and obtained an average of 20-fold coverage of the genome sequences for each individual. The overall level of nucleotide diversity was 23% smaller than that of the Malaysian cynomolgus macaques, and a reduction of low-frequency polymorphisms was observed. In addition, we also confirmed that the Mauritian cynomolgus macaques were genetically closer to a representative of the Malaysian population than to a representative of the Indochinese population. Excess of nonsynonymous polymorphisms in low frequency, which has been observed in many other species, was not very strong in the Mauritian samples, and the proportion of heterozygous nonsynonymous polymorphisms relative to synonymous polymorphisms is higher within individuals in Mauritian than Malaysian cynomolgus macaques. Those patterns indicate that the extreme population bottleneck made purifying selection overwhelmed by the power of genetic drift in the population. Finally, we estimated the number of founding individuals by using the genome-wide site frequency spectrum of the six samples. Assuming a simple demographic scenario with a single bottleneck followed by exponential growth, the estimated number of founders (∼20 individuals) is largely consistent with previous estimates.

Genotyping of non-human primate models: perspectives and challenges for the implementation of the "three R's"

Abstract. Although non-human primates (NHPs) represent only a minor fraction of all animals used in biomedical research, there is a continuous effort to further reduce, refine and replace research with NHPs in accordance with the principles of the three R's. Most of the NHP model species are genetically highly diverse, and significant variation occurs among populations of different geographic origins, particularly in macaques. Since such differences can considerably affect the outcome of biomedical experiments, genotyping represents a promising tool to refine research approaches and to reduce the number of NHPs in biomedical research. Accordingly, the European Primate Network (EUPRIM-Net) developed an anonymous online survey to evaluate possibilities and potential hindrances for the application of genotyping in NHP research. On the one hand, our results point to the importance of genetic variation in NHPs and the need to consider the genetic background for future research approaches. On the other hand, our survey identified several hindrances and limiting factors for the application of genotyping and its incorporation in research, primate husbandry and breeding. We provide some fundamental recommendations on how to meet these challenges and how genotyping can be efficiently used to refine NHP research and to reduce the number of NHPs in biomedical research in the long term.