Comparative transcriptome analysis between rhesus macaques(Macaca mulatta) and crab-eating macaques(M. fascicularis)

Understanding gene expression variations between species is pivotal for deciphering the evolutionary diversity in phenotypes. Rhesus macaques(Macaca mulatta, MMU)and crab-eating macaques(M. fascicularis, MFA) serve as crucial nonhuman primate biomedical models with different phenotypes. To date, however, large-scale comparative transcriptome research between these two species has not yet been fully explored. Here, we conducted systematic comparisons utilizing newly sequenced RNA-seq data from84 samples(41 MFA samples and 43 MMU samples)encompassing 14 common tissues. Our findings revealed a small fraction of genes(3.7%) with differential expression between the two species, as well as 36.5% of genes with tissue-specific expression in both macaques. Comparison of gene expression between macaques and humans indicated that 22.6% of orthologous genes displayed differential expression in at least two tissues. Moreover,19.41% of genes that overlapped with macaque-specific structural variants showed differential expression between humans and macaques. Of these, the FAM220A gene exhibited elevated expression in humans compared to macaques due to lineage-specific duplication. In summary,this study presents a large-scale transcriptomic comparison between MMU and MFA and between macaques and humans. The discovery of gene expression variations not only enhances the biomedical utility of macaque models but also contributes to the wider field of primate genomics.

BARN:Behavior-Aware Relation Network for multi-label behavior detection in socially housed macaques

Quantification of behaviors in macaques provides crucial support for various scientific disciplines,including pharmacology,neuroscience,and ethology.Despite recent advancements in the analysis of macaque behavior,research on multi-label behavior detection in socially housed macaques,including consideration of interactions among them,remains scarce.Given the lack of relevant approaches and datasets,we developed the Behavior-Aware Relation Network(BARN)for multi-label behavior detection of socially housed macaques.Our approach models the relationship of behavioral similarity between macaques,guided by a behavior-aware module and novel behavior classifier,which is suitable for multi-label classification.We also constructed a behavior dataset of rhesus macaques using ordinary RGB cameras mounted outside their cages.The dataset included 65?913 labels for19 behaviors and 60?367 proposals,including identities and locations of the macaques.Experimental results showed that BARN significantly improved the baseline SlowFast network and outperformed existing relation networks.In conclusion,we successfully achieved multilabel behavior detection of socially housed macaques with both economic efficiency and high accuracy.

Seasonal variation and synchronization of sexual behaviors in free-ranging male Tibetan macaques (Macaca thibetana) at Huangshan, China

Although seasonal breeding has been documented in many non-human primates, it is not clear whether sexual behaviors show seasonal variation among male individuals. To test this hypothesis, the focal animal sampling method and continuous recording were used to investigate seasonal variation and synchronization of sexual behaviors in five male Tibetan macaques （Macaca thibetana） at Mt. Huangshan from Oct 2005 to Sept 2006. Both copulatory and sexually motivated behaviors （i.e., sexual chase, grimace, and sexual-inspection）, which were significantly higher in the mating season than non-mating season. Furthermore, seasonal variations of sexual behaviors, including copulatory and sexually motivated behaviors, were synchronized among males. The results shed light on sexual competition and tactics for reproductive success of male M. thibetana and other non-human primates with seasonal breeding.

Superior intestinal integrity and limited microbial translocation are associated with lower immune activation in SIVmac239-infected northern pig-tailed macaques(Macaca leonina)

Microbial translocation is a cause of systemic immune activation in HIV/SIV infection. In the present study, we found a lower CD8+ T cell activation level in Macaca leonina (northern pig-tailed macaques, NPMs) than in Macaca mulatta (Chinese rhesus macaques, ChRMs) during SIVmac239 infection. Furthermore, the levels of plasma LPS-binding protein and soluble CD14 in NPMs were lower than those in ChRMs. Compared with ChRMs, SIV-infected NPMs had lower Chiu scores, representing relatively normal intestinal mucosa. In addition, no obvious damage to the ileum or colon epithelial barrier was observed in either infected or uninfected NPMs, which differed to that found in ChRMs. Furthermore, no significant microbial translocation (Escherichia coli) was detected in the colon or ileum of infected or uninfected NPMs, which again differed to that observed in ChRMs. In conclusion, NPMs retained superior intestinal integrity and limited microbial translocation during SIV infection, which may contribute to their lower immune activation compared with ChRMs.

Interchange between grooming and infant handling in female Tibetan macaques(Macaca thibetana)

In some non-human primates, infants function as a social tool that can bridge relationships among group members. Infants are a desired commodity for group members, and mothers control access to them. The biological market theory suggests that grooming is widespread and represents a commodity that can be exchanged for infant handling. As a limited resource, however, the extent to which infants are interchanged between mothers(females with an infant) and non-mothers(potential handlers,females without an infant) remains unclear. In this study, we collected behavioral data to investigate the relationship between grooming and infant handling in free-ranging Tibetan macaques(Macaca thibetana)at Mt. Huangshan, China. Our results showed that females with infants received more grooming than females without infants. After her infant was handled,mother females received more grooming than they did during daily grooming interactions. However, with the increasing number of infants within the social group, both the grooming that mothers received and the grooming that non-mothers invested for handling infants decreased. We also found that non-mothers invested more time in grooming to gain access to younger infants than older infants. Our results provide evidence that infants are social commodities for both mother and non-mother females. Mothers use infants for obtain grooming and non-mothers use grooming to gain access to infants. The current study implies a bidirectional and complex interchange pattern between grooming and infant handling to compensate for the dyadic grooming disparity in non-human primates.

Influence of dominance rank and affiliation relationships on selfdirected behavior in female Tibetan macaques (Macaca thibetana)

Self-directed behavior(SDB) is characterized as an indicator of anxiety, frustration and stress in nonhuman primates. In this study, we collected self-directed behavior data from one group of free-ranging Tibetan macaques(Macaca thibetana) at Mt. Huangshan, China(September 2012–May 2013) using a combination of behavioral sampling methods including focal animal sampling, behavioral sampling, continuous sampling and instantaneous sampling. Our results showed that females engaged in significantly higher rates of self-directed behavior when they were in proximity to dominant individuals compared to subordinate ones. Conflict losers significantly increased their SDB rates after agonistic episodes, indicating that SDB might also serve as an index of anxiety in M. thibetana. We further found that females significantly increased their SDB rates when focal individual was proximity to weakly affiliation relationship higher rank members than to strongly affiliation relationship higher rank members. If conflicts were not reconciled, the postconflict SDB rates of losers were higher when they stayed with strongly affiliation opponents; if conflicts were reconciled, victims of strongly affiliation relationships opponents engaged in more SDB rates before reconciliation than after reconciliation, while victims of moderately affiliation relationships opponents did not engaged in more SDB rates before reconciliation than after reconciliation. We conclude that both of dominance rank and affiliation relationships might both influence the SDB rates of female Tibetan macaques significantly, suggesting that SDB is not only an index of anxiety in Tibetan macaques, but also can provide a new insight into evaluation of social relationships between individuals.

Male mate choice in Tibetan macaques Macaca thibetana at Mt.Huangshan,China

Though females are generally more selective in mate choice, males may also benefit from mate choice if male reproductive success is limited by factors other than simply the number of female mates, and if females differ in short-term reproductive potential. We studied male mate choice in a free-ranging troop of Tibetan macaques Macaca thibetana at Mt. Huangshan, China, from August 2007 to April 2008. We employed focal animal sampling and all occurrence sampling to record sexual related behaviors. Eight adult females were divided into three female quality categories according to the females＇ age, rank and parity. Using male mating effort as a proxy for male mate choice, we found that males do distinguish female quality and show time-variant mating strategies. Specifically, females with dominant rank, high fecundity, and middle age attracted significantly more males. Our results suggest that female short-term reproductive potential appears to be an important variable in determining male mating effort. Male Tibetan macaques do exercise mate choice for higher quality females as well as reduce useless reproductive cost, which is consistent with the direct benefits theory of mate choice.

Metagenomic comparison of the rectal microbiota between rhesus macaques(Macaca mulatta) and cynomolgus macaques(Macaca fascicularis)

Rhesus macaques(Macaca mulatta) and cynomolgus macaques(Macaca fascicularis) are frequently used in establishing animal models for human diseases. To determine the differences in gut microbiota between these species, rectal swabs from 20 rhesus macaques and 21 cynomolgus macaques were collected, and the microbial composition was examined by deep sequencing of the 16 S rR NA gene. We found that the rectal microbiota of cynomolgus macaques exhibited significantly higher alpha diversity than that of rhesus macaques, although the observed number of operational taxonomic units(OTUs) was almost the same. The dominant taxa at both the phylum and genus levels were similar between the two species, although the relative abundances of these dominant taxa were significantly different between them. Phylogenetic Investigation of Communities by Reconstruction of Unobserved States(PICRUSt) showed significant differences in the functional components between the microbiota of the two species, in particular the lipopolysaccharide(LPS) synthesis proteins. The above data indicated significant differences in microbial composition and function between these two closely related macaque species, which should be taken into consideration in the future selection of these animals for disease models.

Dental Variation Among Asian Colobines, with Specific Reference to the Macaques on the Same Continent

In order to discern dental morphometric variations among the Asian colobines, residuals of the colobines, derived from allometric baselines formed by the Asian macaques （Macaca）, were analyzed with Principal Components Analysis and Euclidean Distances. Results indicated that the widely accepted view that the colobines possess relatively smaller front teeth than the macaques is only the case for the first incisors. The colobines show relatively smaller molars than the macaques. Such profiles may be related to the differences in dietary preferences between the two major groups of the Asian Old World monkeys. The magnitude of such differences is not as great as usullay assumed for the two groups that contain both African and Asian taxa. In other words, the two Asian cercopithecoid groups may have homogenously been shaped by the tectonic modifications and climate alterations in the past five million years. There exist marked differences among the Asian colobines when each of the genera is compared with macaques; the dental profile reflects not only the variation in geographic distribution but also in phylogenetic divergence. Thus, the snub-nosed monkeys （Rhinopithecus） and the gray langurs （Semnopithecus） are characterized by relatively larger molars than the other colobines - larger even than those of the macaques. The differences among Asian colobines, depicted by Euclidean Distances, seems to reflect the relationship of the phylogeny and evolution between colobines and cercopithecines.

Social rank and cortisol among female rhesus macaques(Macaca mulatta)

In animal societies,some stressful events can lead to higher levels of physiological stress.Such stressors,like social rank,also predict an increased vulnerability to an array of diseases.However,the physiological relationship between social rank and stress varies between different species,as well as within groups of a single species.For example,dominant individuals are more socially stressed at times,while at other times it is the subordinate ones who experience this stress.Together,these variations make it difficult to assess disease vulnerability as connected to social interactions.In order to learn more about how physiological rank relationships vary between groups of a single species,cortisol measurements from hair samples were used to evaluate the effects of dominance rank on long-term stress levels in despotic and less stringent female rhesus macaque hierarchal groups.In despotic groups,cortisol levels were found not to be correlated with social rank,but a negative correlation was found between social rank and cortisol levels in less stringent hierarchies.Low ranking monkeys in less stringent groups secreted elevated levels of cortisol compared to higher ranking animals.These data suggest that variations in the strictness of the dominance hierarchy are determining factors in rank related stress physiology.The further consideration of nonhuman primate social system diversity and the linear degree of their hierarchies may allow for the development of valid rank-related stress models that will help increase our understanding and guide the development of new therapeutics for diseases related to human socioeconomic status.

Intestinal ischemia-reperfusion of macaques triggers a strong innate immune response

AIM: To investigate inflammatory injury in the intestinal mucosa after intestinal ischemia-reperfusion (IIR) with Toll-like receptor (TLR)-mediated innate immunity. METHODS: Ten macaques were randomized into control and IIR groups. The distribution and expression level of TLR2, TLR4, MD2, nuclear factor (NF)-kappa B p65 and interferon (IFN)-gamma were measured by immunohistochemical stain and western blotting. The mRNA expression of TLR4, TLR2, MD2, interleukin (IL)-1 beta and tumor necrosis factor (TNF)-alpha were measured by reverse transcriptase-polymerase chain reaction. The cytokine levels in blood and intestinal tissues were measured by ELISA. RESULTS: Obvious hemorrhage and erosion of mucosae were seen in the IIR group. Expression of TLR2, TLR4, MD2, NF-kappa B p65 and IFN-gamma. was significantly higher in the IIR group than in the control group (0.13 +/- 0.04, 0.22 +/- 0.04, 0.16 +/- 0.06, 0.65 +/- 0.12, 0.38 +/- 0.10 vs 0.07 +/- 0.04, 0.08 +/- 0.03, 0.04 +/- 0.02, 0.19 +/- 0.06, 0.14 +/- 0.05, P < 0.05). In addition, the expression of TLR2, TLR4, MD2, IL-1 beta and TNF-alpha mRNA in the IIR group were significantly higher than those of control group(1.52 +/- 0.15, 1.39 +/- 0.06, 1.94 +/- 0.12, 1.48 +/- 0.15, 0.66 +/- 0.08 vs 0.31 +/- 0.05, 0.5 +/- 0.04, 0.77 +/- 0.05, 0.35 +/- 0.08, 0.18 +/- 0.04, P < 0.05). Furthermore, IL-1 beta, IL-6 and TNF-alpha levels in the macaques ileum and plasma were significantly higher than in the control group (plasma: 86.3 +/- 15.2, 1129 +/- 248.3, 77.8 +/- 16.2 vs 29.5 +/- 7.3, 19.8 +/- 8.2, 5.6 +/- 1.7; ileum: 273.4. +/- 44.7, 1636 +/- 168.0, 205.5 +/- 30.7 vs 76.8 +/- 20.5, 663.4 +/- 186.9, 49.0 +/- 9.4; P < 0.05). CONCLUSION: After IIR, general inflammatory injury in the intestinal mucosa is correlated with a strong innate immune response, mediated by activation of the TLR-NF-kappa B-cytokine pathway. (C) 2014 Baishideng Publishing Group Inc. All rights reserved.

Molecular cloning and anti-HIV-1 activities of APOBEC3s from northern pig-tailed macaques(Macaca leonina)

Northern pig-tailed macaques （NPMs, Macaca leonina） are susceptible to HIV-1 infection largely due to the loss of HIV-l-restricting factor TRIM5a. However, great impediments still exist in the persistent replication of HIV-1 in vivo, suggesting some viral restriction factors are reserved in this host The APOBEC3 proteins have demonstrated a capacity to restrict HIV-1 replication, but their inhibitory effects in NPMs remain elusive. In this study, we cloned the NPM A3A-A3H genes, and determined by BLAST searching that their coding sequences （CDSs） showed 99% identity to the corresponding counterparts from rhesus and southern pig-tailed macaques. We further analyzed the anti-HIV-1 activities of the A3A-A3H genes, and found that A3G and A3F had the greatest anti-HIV-1 activity compared with that of other members. The results of this study indicate that A3G and A3F might play critical roles in limiting HIV-1 replication in NPMs in vivo. Furthermore, this research provides valuable information for the optimization of monkey models of HIV-1 infection.

Social rank and cortisol among female rhesus macaques (Macaca mulatta)

In animal societies, some stressful events can lead to higher levels of physiological stress. Such stressors, like social rank, also predict an increased vulnerability to an array of diseases. However, the physiological relationship between social rank and stress varies between different species, as well as within groups of a single species. For example, dominant individuals are more socially stressed at times, while at other times it is the subordinate ones who experience this stress. Together, these variations make it difficult to assess disease vulnerability as connected to social interactions. In order to learn more about how physiological rank relationships vary between groups of a single species, cortisol measurements from hair samples were used to evaluate the effects of dominance rank on long-term stress levels in despotic and less stringent female rhesus macaque hierarchal groups. In despotic groups, cortisol levels were found not to be correlated with social rank, but a negative correlation was found between social rank and cortisol levels in less stringent hierarchies. Low ranking monkeys in less stringent groups secreted elevated levels of cortisol compared to higher ranking animals. These data suggest that variations in the strictness of the dominance hierarchy are determining factors in rank related stress physiology. The further consideration of nonhuman primate social system diversity and the linear degree of their hierarchies may allow for the development of valid rank-related stress models that will help increase our understanding and guide the development of new therapeutics for diseases related to human socioeconomic status.

Multilocus phylogeny suggests a distinct species status for the Nepal population of Assam macaques(Macaca assamensis):implications for evolution and conservation

Phylogenetic relationships within the sinica-group of macaques based on morphological,behavioral,and molecular characteristics have remained controversial.The Nepal population of Assam macaques(Macaca assamensis)(NPAM),the westernmost population of the species,is morphologically distinct but has never been used in phylogenetic analyses.Here,the phylogenetic relationship of NPAM with other congeners was tested using multiple mitochondrial and Ychromosomal loci.The divergence times and evolutionary genetic distances among macaques were also estimated.Results revealed two major mitochondrial DNA clades of macaques under the sinica-group:the first clade included M.thibetana,M.sinica,and eastern subspecies of Assam macaque(M.assamensis assamensis);the second clade included M.radiata together with species from the eastern and central Himalaya,namely,M.leucogenys,M.munzala,and NPAM.Among the second-clade species,NPAM was the first to diverge from the other members of the clade around 1.9 million years ago.Our results revealed that NPAM is phylogenetically distinct from the eastern Assam macaques and closer to other species and hence may represent a separate species.Because of its phylogenetic distinctiveness,isolated distribution,and small population size,the Nepal population of sinica-group macaques warrants detailed taxonomic revision and high conservation priority.

Macaques Contribute to New Breakthroughs in Autism Research

A China-U.S. joint research team has made new breakthroughs in developing nonhuman primate models for autism research. This does much to pave the way for possible drug discovery and gene therapy of the disorder. The research was conducted by scientists from the Shenzhen Institutes of Advanced Technology (SIAT) under the Chinese Academy of Sciences, Massachusetts Institute of Technology (MIT), Sun Yat-Sen University, and South China Agricultural University.

Analysis of immunoglobulin, complements and CRP levels in serum of captive northern pig-tailed macaques （Macaca leonina）

The northem pig-tailed macaque （NPM, Macaca leonina） has become a widely used animal model in biomedical research. In this study, we measured serum immunoglobulin IgG, IgM, IgA, complement C3, C4 and CRP levels in 3-11 year old captive northem pig-tailed macaques using HITACHI 7600-20 automated chemistry analyzer in order to determine the influences of age and gender on these items. The results showed that serum IgA, IgM, C3 and C4 levels were not correlated with age （P〉0.05）, while serum IgG levels increased progressively with age （r=0.202; P=0.045）. Serum IgG, IgA, IgM and C3 levels were higher in females than in males （P〈0.05）. Moreover, serum C3 concentration was both positively and strongly correlated with that of C4 （r=0.700; P〈0.0001）. This study provides basic serum immunoglobulin and complement data of captive northem pig-tailed macaques, which may prove useful for future breeding efforts and biomedical research.

Longitudinal analysis reveals characteristically high proportions of bacterial vaginosis-associated bacteria and temporal variability of vaginal microbiota in northern pig-tailed macaques （Macaca leonina）

The complex and dynamic vaginal microbial ecosystem is critical to both health and disease of the host. Studies focusing on how vaginal microbiota influences HIV-1 infection may face limitations in selecting proper animal models. Given that northern pig-tailed macaques （Macaca leonina） are susceptible to HIV-1 infection, they may be an optimal animal model for elucidating the mechanisms by which vaginal microbiota contributes to resistance and susceptibility to HIV-1 infection. However, little is known about the composition and temporal variability of vaginal microbiota of the northern pig-tailed macaque. Here, we present a comprehensive catalog of the composition and temporal dynamics of vaginal microbiota of two healthy northern pig-tailed macaques over 19 weeks using 454-pyrosequencing of 16S rRNA genes. We found remarkably high proportions of a diverse array of anaerobic bacteria associated with bacterial vaginosis. Atopobium and Sneathia were dominant genera, and interestingly, we demonstrated the presence of Lactobacillus-dominated vaginal microbiota. Moreover, longitudinal analysis demonstrated that the temporal dynamics of the vaginal microbiota were considerably individualized. Finally, network analysis revealed that vaginal pH may influence the temporal dynamics of the vaginal microbiota, suggesting that inter-subject variability of vaginal bacterial communities could be mirrored in inter-subject variation in correlation profiles of species with each other and with vaginal pH over time. Our results suggest that the northern pig-tailed macaque could be an ideal animal model for prospective investigation of the mechanisms by which vaginal microbiota influence susceptibility and resistance to HIV-1 infection in the context of highly polymicrobial and Lactobacillus-dominated states.

Flow cytometric characterizations of leukocyte subpopulations in the peripheral blood of northern pig-tailed macaques （Macaca leonina）

Pig-tailed macaques(Macaca nemistrina group) have been extensively used as non-human primate animal models for various human diseases in recent years, notably for AIDS research due to their sensitivity to HIV-1. Northern pig-tailed macaques(M. leonina) are distributed in China and other surrounding Southeast Asia countries. Although northern pig-tailed macaques have been bred on a large scale as experimental animals since 2012, the reference value of normal levels of leukocytes is not available. To obtain such information, 62 blood samples from male and female healthy northern pig-tailed macaques at different ages were collected. The normal range of major leukocyte subpopulations, such as T lymphocytes, B lymphocytes, natural killer(NK) cells, monocytes, and the expression levels of activation or differentiation related molecules(CD38, HLA-DR, CCR5, CD21, IgD, CD80 and CD86) on lymphocytes were analyzed by flow cytometry. The counts of B cells decreased with age, but those of CD8+ T cells and NK cells and the frequency of CD38+HLA-DR+CD4+ T cells were positively correlated with age. The counts of leukocyte subpopulations were higher in males than those in females except for CD4+ T cells. Males also showed higher expression levels of Ig D and CD21 within B cells. This study provides basic data about the leukocyte subpopulations of northern pig-tailed macaques and compares this species with commonly used Chinese rhesus macaques(M. mulatta), which is meaningful for the biomedical application of northern pig-tailed macaques.

Disease Progression Patterns of SHIV-KB9 in Rhesus Macaques of Chinese Origin in Comparison with Indian Macaques

Objective To develop a model of SHIV-KB9/Chinese origin rhesus （Ch Rh） macaques for vaccine research and to compare the pathogenesis of SHIV-KB9 in Ch Rh macaques with that reported in Indian rhesus （Ind Rh） macaques. Methods Seven mamu-A＊01 negative Ch Rh macaques were inoculated intravenously with 1-10000 MID50 of SHIV-KB9. The monkeys were monitored for viral load, CD4, CDS, SHIV-specific antibody and virus genetic variation. The results were compared with those previously observed in Ind Rh macaques. Results As compared to that observed in Ind Rh macaques, SHIV-KB9 in Ch Rh macaques displayed three identical disease progression patterns. However, the primary pattern was not identical between the two subspecies. The level of plasma viremia differed in SHIV-KB9-infected Ch Rh macaques which exhibited different outcomes from those in Ind Rh macaques. Generally, the values of viral load and the maintenance of CD4^＋ T cells were associated with humoral responses. Otherwise, the viral genetic distances （divergence, diversity） were larger in animals （M419, M425） with their CD4^＋ T cells profoundly depleted. Conclusion The model of SHIV-KB9/Ch Rh macaques displays a relatively slow progression to AIDS compared with Ind Rh macaques, which may more accurately reflect the potential of candidate vaccines in humans.

Parasites may exit immunocompromised northern pig-tailed macaques (Macaca leonina)infected with SIVmac239

Parasites can increase infection rates and pathogenicity in immunocompromised human immunodeficiency virus （HIV） patients. However, in vitro studies and epidemiological investigations also suggest that parasites might escape immunocompromised hosts during HIV infection Due to the lack of direct evidence from animal experiments, the effects of immunocompromised hosts parasitic infections on remain unclear. Here we detected 14 different parasites in six northern pig-tailed macaques （NPMs） before or at the 50th week of simian immunodeficiency virus （SIV） infection by ELISA. The NPMs all carried parasites before viral injection. At the 50th week after viral injection, the individuals with negative results in parasitic detection （i.e., 08247 and 08287） were characterized as the Parasites Exit （PE） group, with the other individuals （i.e., 09203, 09211, 10205, and 10225） characterized as the Parasites Remain （PR） group. Compared with the PR group, the NPMs in the PE group showed higher viral loads, lower CD4＋ T cells counts, and lower CD4/CD8 rates. Additionally, the PE group had higher immune activation and immune exhaustion of both CD4~ and CD8~ T cells. Pathological observation showed greater injury to the liver, cecum, colon, spleen, and mesenteric lymph nodes in the PE group This study showed more seriously compromised immunity in the PE group, strongly indicating that parasites might exit an immunocompromised host.