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In the present study, we have quantified the effects of transport, relocation and acclimate/adapt to their new surroundings on female squirrel monkey. These responses are measured in blood samples obtained from squirrel monkeys, at different time points relative to their relocation from their old home to their new home. A group of squirrel monkeys we transported, by truck, for approximately 10 hours. Peripheral blood mononuclear cells (PBMCs) were assayed in order to evaluate the phenotype of lymphocyte subsets by flow, mitogen-specific immune responses of PBMCs in vitro, and levels of cytokines at various time points including immediately before transport, immediately upon arrival, and after approximately 150 days of acclimation. We observed significant changes in T cells and subsets, NK and B cells (CD4+, CD8+, CD4+/CD8+, CD16+, and CD20+). Mitogen specific (e.g. PHA, PWM and LPS) proliferation responses, IFN-γ by ELISPOT assay, and cytokines (IL-2, IL-4 and VEGF) significant changes were observed. Changes seen in the serum chemistry measurements mostly complement those seen in the hematology data. The specific goal was to empirically assess the effects of relocation stress in squirrel monkeys in terms of changes in the numbers and functions of various leukocyte subsets in the blood and the amount of time required for acclimating to their new environment. Such data will help to determine when newly arrived animals become available for use in research studies., Competing Interests: The authors have declared that no competing interests exist.

Background: The neotropical, non-human primates (NHP) of the genus Saimiri and Aotus are recommended by the World Health Organization as experimental models for the study of human malaria because these animals can be infected with the same Plasmodium that cause malaria in humans. However, one limitation is the lack of immunological tools to assess the immune response in these models. The present study focuses on the development and comparative use of molecular and immunological methods to evaluate the cellular immune response in Saimiri sciureus., Methods: Blood samples were obtained from nineteen uninfected Saimiri. Peripheral blood mononuclear cells (PBMC) from these animals and splenocytes from one splenectomized animal were cultured for 6, 12, 18, 24, 48, 72 and 96 hrs in the presence of phorbol-12-myristate-13-acetate and ionomycin. The cytokine levels in the supernatant were detected using human and NHP cytometric bead array Th1/Th2 cytokine kits, the Bio-Plex Pro Human Cytokine Th1/Th2 Assay, enzyme-linked immunosorbent assay, enzyme-linked immunospot assays and intracellular cytokine secretion assays. Cytokine gene expression was examined through TaqMan® Gene Expression Real-Time PCR using predesigned human gene-specific primers and probes or primers and probes designed based on published S. sciureus cytokine sequences., Results: The use of five assays based on monoclonal antibodies specific for human cytokines facilitated the detection of IL-2, IL-4 and/or IFN-γ. TaqMan array plates facilitated the detection of 12 of the 28 cytokines assayed. However, only seven cytokines (IL-1A, IL-2, IL-10, IL-12B, IL-17, IFN-β, and TNF) presented relative expression levels of at least 70% of the gene expression observed in human PBMC. The use of primers and probes specific for S. sciureus cytokines facilitated the detection of transcripts that showed relative expression below the threshold of 70%. The most efficient evaluation of cytokine gene expression, in PBMC and splenocytes, was observed after 6-12 hrs of culture, except for LTA in PBMC, whose expression was best analysed after 24 hrs of culture., Conclusions: Real-time PCR facilitates the analysis of a large number of cytokines altered during malaria infection, and this technique is considered the best tool for the evaluation of the cellular immune response in S. sciureus.

Due to many physiological and genetic characteristic similarities to humans, squirrel monkeys provide an ideal animal model specifically for studying malaria, and transmissible spongiform encephalopathies (Creutzfeldt-Jacob disease). While squirrel monkeys three years and older are generally considered adult subjects suitable for use in medical research studies, little is known about the functional properties of lymphocytes in relation to the age of these animals, which could significantly impact the quality and quantity of innate and adaptive immune responses. In this study, we investigated differences in the phenotype and function of lymphocytes subsets of young (3-4 years), adult (8-10 years) and aged (16-19 years) squirrel monkeys. In general, animals in all three age groups exhibited comparable numbers of different lymphocyte subsets except for CD20+ B cells that were significantly lower in aged relative to young animals and T cells subsets expressing both CD4 and CD8 (double positive) were significantly higher in aged relative to young animals. With increasing age, phenotypic differences in central and effector memory T cells subsets were observed, that were more pronounced for the CD8+ T cells. Despite equal proportions of CD3+ T cells among the three age groups, responses of peripheral blood mononuclear cells to T cell mitogens PHA and Con A showed lower IFN-γ producing cells in the aged group than that in the young group. Furthermore, aged animals showed significantly higher plasma levels of inflammatory cytokines IL-6, IFN-γ, TNF-α, IL-10 and IL-12. These findings suggest that while the squirrel monkeys in general share phenotypic and functional similarities of lymphocyte subsets with humans in relation to age, specific differences exist in immune function of lymphocytes between young and old animals that could potentially impact experimental outcomes for which the measurement of immunologic endpoints are critical.

Some biomedical research procedures, such as organ xenotransplantation, usually require intensive hemotherapy. Knowledge of the whole phenotype of blood donor and graft could be useful in the field of xenotransplantation. Human and simian-type categories of blood groups have been established and they can be tested by standard methods used for human blood grouping. The aim of this work was to study the incidence of non-ABO blood group systems in different species of non-human primates, which are employed in biomedical research. The phenotype of Rh, Lewis, Kidd, Kell, MNSs, Lutheran, P and Duffy antigens was investigated in olive baboon (n = 48), chacma baboon (n = 9), Guinea baboon (n = 14), Rhesus macaque (n = 38) and squirrel monkey (n = 30) by using commercial microtyping cards. Kell, Lutheran, Kidd and Duffy antigens have been detected in all species, Rh in squirrel monkey, MNSs in rhesus macaque and squirrel monkey, and Lewis in baboon and rhesus macaque. There were differences in frequency and haemagglutination scores between species regardless of their gender and age. The main differences were found in squirrel monkey when compared with baboons and macaques. This typing system provides a tool to assess the presence of antigens in animals used for experimental procedures, such as xenotransplantation and xenotransfusion.

Foamy virus evolution closely parallels that of the host species, indicating virus-host coadaptation. We studied simian foamy viruses (SFVs) from common marmosets, spider monkeys, and squirrel monkeys, New World monkey (NWM) species that share geographic ranges. The TRIM5alpha protein from each of these NWM species inhibited the replication of at least one of the SFVs associated with the other two species but did not affect the replication of its own SFV. Thus, TRIM5alpha has potentially shaped the evolution of SFVs in NWM hosts. Conversely, SFVs may have influenced the evolution of TRIM5 variants in New World primates.

Splenectomized Aotus lemurinus griseimembra, A. azarae boliviensis, A. nancymaae, A. vociferans, and Saimiri boliviensis monkeys were infected with the Uganda I/CDC strain of Plasmodium malariae. The maximum parasite counts were lower if the animals had been previously infected with Plasmodium vivax. Mosquito infection was concentrated in the 12 days following the rise in count above 1,000/microl. Mosquito infection and parasite counts were highest with A. l. griseimembra. Anopheles freeborni was more readily infected than An. gambiae, which was more readily infected than An. stephensi. Parasite counts and mosquito infection with P. brasilianum were much higher in S. boliviensis monkeys than with the Uganda I strain of P. malariae in this host, suggesting marked differences between the host-parasite-vector relationships and indicating that P. brasilianum in S. boliviensis monkeys may be a better reflection of the relationship of P. malariae in the human host.

Objective: To assess the presence of irregular xenoantibodies against human red blood cells (RBCs) in 6 primate species used in xenotransplantation and other experimental procedures., Materials and Methods: Serum samples from 109 baboons of 4 different species (olive, chacma, sacred, and Guinea), 38 rhesus macaques, and 30 squirrel monkeys were tested for irregular xenoantibodies using an agglutination test using human RBCs of known phenotype for Rh, Kell, Kidd, Lewis, Lutheran, P1, and Duffy antigens, commercially available as RBC I, II, and III., Results: We found hemagglutination for RBC I in 49%, 22%, 100%, 57%, 32%, and 33% of olive, chacma, sacred, and Guinea baboons, rhesus macaques, and squirrel monkey, respectively. The frequency for RBC II was 49%, 50%, 100%, 57%, 37%, and 33%, respectively, and for RBC III was 56%, 37%, 100%, 79%, 34%, and 33%, respectively. There were differences in frequency depending on the sex of the rhesus macaques; all 3 RBCs tested were higher in the females: 44% vs 0%, P = .008; 48% vs 1%, P = .02, and 44% vs 9.1%, P = .04 for RBC I, II, and III, respectively. There were differences due to age in only olive baboons, and a higher frequency in younger animals compared with juvenile, subadult, and adult animals for all 3 human RBCs., Conclusions: Assessment of irregular antibodies in the presence of primate serum should be taken into account during any experimental xenotransplantation protocol.

An outbreak of Toxoplasmosis in a colony of squirrel monkeys (Saimiri sciureus) in Israel is described. Serological, pathological, and molecular findings of monkeys, as well as rodents and pigeons from the vicinity are summarized. Seventy-nine percent (19/24) of monkeys were T. gondii seropositive at titer 1:16 whilst 4% (1/24) were also seropositive at titer 1:64 using the Modified Agglutination Test (MAT). Eighty four percent (21/25) of rats were positive at titer 1:16 and 8% (2/25) of rats were positive at titer 1:32. DNA amplification of a 529bp repeated sequence of T. gondii was detected in the liver and lungs of all monkeys tested, 6/7 in myocardial extractions and 5/6 in brain extractions. Sequence analysis of the SAG2 locus disclosed that T. gondii detected was of Type III genotype. The source of disease was thought to be contamination of feed with infective feline oocysts. As a result of this study, the implementation of a program to capture and remove resident feral cats, to discontinue the feeding of stray cats, and to control rodent populations in the park was introduced.

Saimiri boliviensis monkeys were infected via sporozoites with the Salvador I strain of Plasmodium vivax that had been stored frozen for periods ranging from 12 to 5,312 days. Prepatent periods ranged from 16 to 53 days.

The Saimiri sciureus monkey is a well-established host for experimental studies with human malaria parasites. During the course of iterative inoculations with Plasmodium falciparum parasitised red blood cells (RBC), anti-RBC alloantibodies were detected in the sera of two of eight Saimiri monkeys. These anti-RBC antibodies were further used to investigate RBC phenotypes in 35 colony-reared Saimiri monkeys by flow cytometry. Three RBC phenotypes (named I-III) were observed. Their distribution was I (86%), II (11%) and III (3%). Using the Palo Alto FUP-2 strain, a variant P. falciparum line insensitive to hyperimmune serum and the passive transfer of anti-RBC alloantibodies, a dramatic drop in parasite growth was documented in an incompatible monkey.

Background: The neotropical primate squirrel monkey is used in many areas of biomedical research including neuroendocrinology, immunology and infectious diseases. However, research has been hampered by the lack of immunological tools for this primate., Methods: A series of 67 commercially available monoclonal antibodies to human CD antigens or cytokines were tested on Saimiri mononuclear cells and the specificity was assessed by double staining using flow cytometry., Results: Monoclonal antibodies defining the main mononuclear cells subsets (monocytes, B, T, including CD4 and CD8 T cells) as well as activation markers have been identified. The conditions to specifically identify the various cell subsets using two color flow cytometry and establish their relative proportions have been set-up. We also have established normal values of the main circulating mononuclear cell subsets for adult Saimiri sciureus monkeys from the breeding unit of Institut Pasteur in French Guiana. The distribution between spleen, blood and lymph nodes has been compared., Conclusions: These tools allow documenting the phenotype of most Saimiri mononuclear cell subsets and assessing their activation level. This opens new perspectives for vaccinology and immunopathology research in this experimental non-human primate host, in particular for malaria research.

Little is known about the major histocompatibility complex (MHC) class 1 in squirrel monkeys ( Saimiri sciureus). We cloned, sequenced and characterized two alleles and the cDNA of the coding region of MHC class 1 in these New World monkeys. Phylogenetic analyses showed that these sequences are related to HLA class 1 genes ( HLA-A and HLA-G). The structure and organization of one of the two identified clones was similar to that of a class 1 MHC gene ( HLA-A2). All the exon/intron splice acceptor/donor sites are conserved and their locations correspond to the HLA-A2 gene. The sequences of the newly described cDNAs reveal that they code for the characteristic class 1 MHC proteins, with all the features thought necessary for cell surface expression. Typical sequences for the leader peptide, alpha(1), alpha(2), alpha(3), transmembrane and cytoplasmic domains were found.

If a number of cytokines and growth factors that have been characterized from human cells were investigated in non-human primates, results from such approaches would allow the development of assays to detect and quantitate cytokines in experimental models. Tumor necrosis factor-alpha (TNF-alpha) is an important pluripotent cytokine which plays a crucial role in host defense. As yet, no complete molecular data have been reported for the squirrel monkey TNF-alpha. Polymerase chain reaction (PCR) primers were used to trace introns, by comparing product sizes obtained using cDNA and genomic DNA as templates. The genomic DNA is composed of four exons and three introns with 1793 nucleotides. The corresponding cDNA is 702 nucleotides and phylogenetic analysis showed that the Saimiri sciureus was most closely related to that of the genus Aotus, a new-world primate, compared to old-world primates (genus Macaca and Papio). The deduced TNF-alpha protein consists of 233 amino acids with 82% identity to human, 95% to new-world monkeys and 79% to old-world monkeys. The cloned TNF-alpha cDNA will be useful to quantitate TNF-alpha at the mRNA level.

Primates have long been used as models to study the basic mechanisms of human disease. The squirrel monkey, Saimiri sciureus,is a New World monkey that can be satisfactorily bred in captivity and infected with various human pathogens. The basic immunological parameters of squirrel monkeys, including immunoglobulin subclasses and markers for lymphocyte subsets, have already been characterized. However, immunological reagents and assays specific for the detection and quantification of squirrel monkey cytokines are required to elucidate the immunological and physiopathological changes that occur during experimental infection of these animals with pathogens. We therefore cloned, sequenced, and characterized the cDNAs encoding various squirrel monkey Th1 and Th2 cytokines, including interleukin (IL)-1 beta, IL-2, IL-5, IL-6, IL-10, tumor necrosis factor alpha, and gamma interferon. We found 91.4%-98.1% homology between the nucleotide sequence of the squirrel monkey cytokine genes and published sequences of equivalent human and non-human primate genes. The aligned sequences of cytokines for Saimiriand several New and Old World primates and human are shown, and a phylogenetic analysis of published sequences of selected cytokines in other species and those of non-human primates is given. In addition, we adapted a previously described quantitative, reverse transcriptase, competitive polymerase chain reaction technique to measure the mRNA expression of those cytokines in squirrel monkeys. The primer and competitor plasmids that allowed quantification are described. We showed that the assay is sensitive and reproducible.

In this study, we examined serum anti-ovalbumin (OVA) IgE and IgG antibodies in laboratory-reared squirrel monkeys (Saimiri sciureus), that were fed a boiled quail egg everyday. We found that 36 of 95 monkeys (38%) possessed specific IgE and 44% (42/95) had specific IgG against OVA. These antibody titers seemed to increase with age. There was, however, no apparent correlation between the anti-OVA IgE and IgG antibody titers.

The squirrel monkey, a non-human New World primate, has several endocrine peculiarities, including a 10-fold higher plasma cortisol concentration than Old World primates, such as man. Glucocorticoids are known to have immunomodulatory properties. We therefore measured cytokine levels in supernatants of in vitro cultures of mononuclear cells from the peripheral blood of squirrel monkeys and humans. We stimulated monocytes and lymphocytes with lipopolysaccharide (LPS) and phytohemagglutinin (PHA) in the presence or absence of hydrocortisone. Squirrel monkey monocytes secreted a more than 100-fold lower level of interleukin-1 beta (IL-1 beta) but a four-fold higher level of transforming growth factor beta (TGF-beta) than human monocytes, whereas the secretion of other cytokines, such as tumor necrosis factor alpha (TNF-alpha), TNF-beta and interleukin 2 (IL-2), did not differ between squirrel monkeys and humans. However, in squirrel monkey lymphocytes, the PHA-stimulated secretion of TNF-alpha was much greater than that of TNF-beta. Our results support the view that in squirrel monkeys there are subtle adaptations in some immune functions, particularly linked to the hypothalamic-pituitary-adrenal (HPA) system rather than a global suppression of the immune system.

There are relatively few monoclonal antibodies (mAb) that have been characterized for their applicability in studies on the immune system of various nonhuman primates. In the present study, we identified a large number of mAb that can be used in future immunological studies in three different nonhuman primates, i.e., chimpanzees, rhesus macaques, and squirrel monkeys. The reactivity of 161 anti-human mAb to T-cell antigens and cytokine receptors were tested on peripheral blood mononuclear cells (PBMC) from the three primate species by flow cytometric analysis. A total of 105 (65%), 73 (45%), and 68 (42%) antibodies reacted with PBMC from chimpanzees, rhesus macaques, and squirrel monkeys, respectively. Out of the 161 mAb, 38 reacted with all three species and 112 reacted with one or two of the species. No specific reaction was observed with mAb to receptors to GM-CSF, 4-1BB, FLT3, FLX2, common beta-chain, IL-1 (type I receptor), and IL-8.

Although most strategies of vaccination require immunopotentiation to induce efficient immune responses, the development of new adjuvants for human vaccines is highly limited by safety problems. In order to overcome this problem, we developed a new vaccine formulation based on the covalent linkage of protein or peptide to synthetic microspheres. In previous experiments performed in mice, we demonstrated that these particulate antigens induce strong antigen-specific CD4+ T cell proliferative responses in the absence of adjuvant. In the present study, we analyzed the immunogenicity in primate Saimiri sciureus monkeys of two different proteins linked to synthetic microspheres. Immune responses induced by these particulate proteins administered without adjuvant were compared to those stimulated by the soluble antigens injected with alum. We currently demonstrated that, in monkeys, particulate antigens administered without adjuvant, induced good PBMC proliferative response and antibody production. Furthermore, the analysis of antibody responses using mAbs specific for different Saimiri sciureus immunoglobulins showed that the antibody response profiles were different in monkeys immunized with soluble versus particulate form of antigens. Results of this study demonstrate that particulate form of antigen may stimulate qualitatively different immune responses as compared to alum and therefore suggest that this new antigen formulation could be an attractive candidate for the development of vaccines.

The passive transfer of specific antibodies to a naive splenectomized Saimiri sciureus monkey infected with the Palo Alto FUP/SP strain of Plasmodium falciparum resulted in the emergence of parasites resistant to the transferred antibodies. Molecular typing indicated that the original and resistant parasites were isogenic. Saimiri monkeys primed with original parasites were fully susceptible to a challenge by the resistant ones, and vice versa. This absence of crossprotection indicates that strain-specific determinants would be the major targets of protective immunity developed in these monkeys. Phenotypic analysis showed that the surface of the infected red blood cells differed in both lines. Original parasites formed rosettes, autoagglutinated, presented characteristic knobs at the surface of the infected red blood cell, and did not agglutinate in the presence of a pool of human immune sera. In contrast, the resistant parasites did not form rosettes, did not spontaneously autoagglutinate, presented abnormal flattened knobs, and formed large aggregates in the presence of a pool of human immune sera. The presence of strain-specific determinants at the surface of the resistant parasites was confirmed by surface immunofluorescence and agglutination using homologous Saimiri serum. Neither the original nor the resistant parasites cytoadhered to an amelanotic melanoma cell line, suggesting that cytoadherence and agglutination can be dissociated. These results indicate that parasites that differ by the antigens exposed at the surface of the red blood cell induce strain-specific immunity. Furthermore they show that rosetting and nonrosetting parasites differ in their antigenic properties and do not crossprotect.

The squirrel monkey Saimiri sciureus is an experimental host for a range of human pathogens, and for the assessment of vaccine candidate antigens and vaccine strategies. This experimental host is thus particularly suitable for the follow-up of humoral responses. To understand some of the mechanisms that underlie the defense against experimental pathogens, there is a need of basic knowledge on cellular immune effectors also. The authors report here their experience in characterizing squirrel monkey blood T and B lymphocytes, and in studying in vitro induced activation and proliferation of T and B cells. Particular emphasis is given to the in vitro differentiation of squirrel monkey B cells into immunoglobulin secreting cells, with respect to Plasmodium falciparum antigens.

Blood B lymphocytes obtained from Plasmodium falciparum-immune Saimiri monkeys were assayed for their in vitro differentiation in immunoglobulin-secreting cells upon restimulation with P. falciparum-parasitized Saimiri red blood cells. Selected culture conditions enabled appropriately stimulated blood B cells to secrete 3F11/G10+ IgG, detected in the supernatants by means of a dot immunobinding assay. Primed blood B lymphocytes from P. falciparum-immune Saimiri monkeys were thus able to secrete IgG when restimulated by parasitized red blood cells in the presence of T cell- and monocyte-derived cytokines (recombinant human cytokines). These primed blood B cells, which were able to differentiate, were shown to secrete antibodies reactive with P. falciparum-infected red blood cells, as detected by means of an indirect immunofluorescence assay, and reactive with P. falciparum-infected red blood cell extracts, as detected by means of Western blot analysis. Furthermore, due to the possibility of discriminating between IgG subtypes in the squirrel monkey (3F11/G10+::3A2/G6+ IgG [associated with protection against the blood stages of P. falciparum] vs. 3F11/G10+::3E4/H8+ IgG [usually not functionally associated with protection]), we have attempted to estimate the respective proportions of each IgG subtype. In defined culture conditions, Saimiri monkey blood B cells preferentially secrete 3F11/G10+::3E4/H8+ IgG in response to parasitized red blood cells. We therefore discuss the conditions that would render this assay suitable for the selection, among P. falciparum blood stage antigens, of those that have major B-cell epitopes.

South American Aotus and Saimiri monkeys, which are susceptible to infection with human malarias, have been used to develop models for the testing of human malaria vaccines. Studies indicate that blood-stage and sporozoite vaccines can be tested in these monkeys using appropriate strains of parasites.

In the Saimiri monkey, an experimental host for human malaria, acquired protection against Plasmodium falciparum blood stages depends on the IgG antibody populations developed. In vivo protective anti-falciparum activity of IgG antibodies is correlated with the in vitro opsonizing activity promoting phagocytosis of parasitized red blood cells. In contrast, non protective antibodies inhibit this mechanism by competing at the target level. A similar phenomenon can be observed in human infection. Anti-cytoadherent and anti-rosette antibodies developed by Saimiri and humans prevent the development of physiopathological events like cerebral malaria which can also occur in this experimental host. Furthermore, transfer to protective human anti-falciparum IgG antibodies into infected Saimiri monkeys exerts an anti parasite activity as efficient as that observed when it is transferred into acute falciparum malaria patients, making the Saimiri an even more attractive host. Studies on the role of immunocompetent cells in the protective immune response are still in their infancy, however the existence of a restricted polymorphism of MHC II class molecules in the Saimiri confers additional theoretical and practical importance to this model.

We report here on functional aspects of squirrel monkey T cells isolated either from peripheral blood mononuclear cells or in the presence of accessory cells or accessory-cell-secreted cytokines. The T-cell response triggered by polyclonal activators was measured through proliferation, T-cell growth factor production and transient expression of receptors to growth factors. We also sought to delineate mechanisms by which antimitogenic molecules might prevent lymphocytes from being able to express "activation antigens" and/or from proliferating. Under these conditions, we mainly explored the inducible IL2/IL2R pathway and phenomena occurring in lymphocyte "non-activation" in terms of early and late cellular events; indeed, defects in the IL2/IL2R pathway represent one of the dysregulation events presumed to occur during the acute phase of human malaria, disease for which. Saimiri monkeys have been selected as experimental hosts.

The new-world monkeys Saimiri sciureus (squirrel monkeys) are currently used as a model to test the efficacy of vaccines against human malaria. To improve our knowledge on this model, we tested the susceptibility of S. sciureus B cells to Epstein-Barr virus (EBV) infection. B-lymphoblastoid cell lines were obtained from six of six healthy animals after infection with the B95-8 source of EBV. The frequency distributions of spleen B cells clonally committed to the production of immunoglobulins M and G, as measured by limiting dilution analysis, were from 1 in 179 to 1 in 1,085 and from 1 in 45 to 1 in 60, respectively, in three monkeys naturally infected with Plasmodium brasilianum. In the same three animals, the frequency of spleen B cells committed to the production of P. brasilianum-specific antibody ranged from 1 in 2,211 to 1 in 9,099. One B-lymphoblastoid cell line producing anti-P. brasilianum-specific antibody was cloned twice, and the immunoglobulin G produced was purified. This monoclonal antibody recognized a parasite component of 197 kDa and was specific for Plasmodium malariae and P. brasilianum parasites. These data document that squirrel monkey B cells naturally primed by an infectious agent can be efficiently used to produce monospecific antibodies against the infectious agent.

This study evaluated the immunological, hormonal, and behavioral responses of juvenile squirrel monkeys to repeated social separations of varying length. Following a 3-hr separation, lymphocyte responses to stimulation with the mitogen concanavalin A (Con A) declined significantly, and these alterations were sustained after a 24-hr separation period. The responses to Con A and to a second mitogen, phytohemagglutinin (PHA), were suppressed after 2 days. At the end of a 7-day separation period, immune responses were not significantly different from basal values. Plasma cortisol levels were elevated above basal levels in all animals after the 3-hr, 24-hr, and 2-day separations, but were not elevated after the 7-day separation. While we observed no statistically significant changes in locomotor activity or calling behavior during any of the separations, the monkeys tended to be most active and to call most immediately following separation. Our findings concur with earlier reports indicating that social stressors can influence lymphocyte proliferation in nonhuman primates and that certain cell types might be differentially sensitive to stress, but also indicate that these influences are transient.

Saimiri and Aotus, two neotropical primates, are currently used in different domains of human malaria research. Here we present a simplified and non exhaustive enumeration of different aspects in concern of using these monkeys as experimental hosts, their availability, laboratory-bred animals versus wild-caught animals, their potential as experimental model in evaluating anti-malaria vaccine candidates, and some related activities at the Pasteur Institute.

The objective of this study was to determine whether the squirrel monkey (Saimiri scuireus) is indigenously colonized with black-pigmented bacteroides (BPB) resembling human Bacteroides gingivalis and Bacteroides intermedius (suspected periodontal pathogens) and to determine the usefulness of the squirrel monkey as an in vivo model for studying colonization by putative pathogens. We assayed the subgingival plaques of 138 monkeys of various ages and in four different colonies for the presence of anaerobic BPB microorganisms. We also tested half the animals for the presence of Actinobacillus actinomycetemcomitans. Clinical indices and levels of serum antibody to B. gingivalis were recorded. We detected BPB in 50% of the animals and A. actinomycetemcomitans in 69% of the animals. The presence of BPB was generally associated with increased age, increased gingival index, presence of calculus, and increased levels of serum antibody. These data indicate that the squirrel monkey may be a good model for studying the parameters of natural infection of the gingival crevice with suspected periodontopathogenic BPB microorganisms.

Hemolytic serum complement activity was quantitatively compared in baboons, squirrel monkeys, cebus monkeys, and cotton-top marmosets. Squirrel monkeys showed the highest activity, and marmosets had the lowest activity. The complement level in squirrel monkeys and tenfold greater than marmosets and almost four times higher than that of man. Cebus monkeys had levels most similar to that of man while the baboon exhibited activity almost as low as that of the marmoset.

Nine hybrid clones secreting antibodies to squirrel monkey (Saimiri sciureus) IgM were produced and two of these (1F1G5 and 5H11B3) were selected for further studies. These non-precipitating monoclonal antibodies reacted with two distinct repetitive antigenic determinants, probably of the conformational type, only present on the native or SDS-denatured IgM molecule. Reduction of the pentamer with 2-mercaptoethanol led to complete destruction of the corresponding epitopes. 1F1G5 antibodies from ascitic fluids were used in the purification of monkey IgM by affinity chromatography. The characteristics of 1F1G5 and 5H11B3 MAbs permitted the development of a solid-phase two-site immunoradiometric assay for the measurement of IgM levels in serum specimens taken from healthy animal donors of both sexes.

The following study assessed changes in macrophage responsiveness after a 24-h period of psychological disturbance in mother and infant squirrel monkeys. Utilizing a luminol-dependent assay, an 80-min chemiluminescent burst was measured in blood monocytes in response to zymosan stimulation. Cells obtained from stressed mothers and infants showed significant increases in chemiluminescence (CL) as compared to baseline levels. Moreover, the elevated pattern of response persisted for at least 2 weeks after the mothers and infants were reunited. The initial change in CL was associated with increased pituitary-adrenal activity and leukocyte redistribution, but these measures returned to normal levels following reunion. Thus, this study has demonstrated a prolonged change in an immune parameter following a transient alteration in the psycho-endocrine status of the host.

Hypothalamic neurons producing growth hormone-releasing factor (GRF) have been characterized by immunohistochemistry in monkey hypothalamus, using an antiserum raised against hpGRF1-40, a peptide with GRF activity isolated from a human pancreatic tumor. Cell bodies with hpGRF immunoreactivity were found in arcuate and ventromedial nuclei. From these neurons, bundles of fibers innervate median eminence and appear to terminate in contact with portal vessels. In addition to median eminence, hpGRF immunoreactive fibers were found mostly in the anterior hypothalamus and the arcuate and ventromedial nuclei where they give perineuronal endings. These results correlate with earlier physiological data on hypothalamic control of growth hormone secretion and suggest that GRF is also involved in interneuronal relationships related or unrelated to neurohumoral control of pituitary secretions.

Premature separation of rat pups from their mothers, on postnatal Day 15, produced a decreased response of peripheral blood lymphocytes to phytohemagglutinin (PHA) at 40 days of age. A significant lymphopenia was also found in the early weaned animals at 40 days of age although this was accounted for statistically by their lower body weight. These consequences of early maternal separation may have been mediated through the effects of early separation on nutritional state, hypothalamic function, or maturation of the immune system.

The following research assessed the influence of developmental, hormonal, and psychological factors on immunoglobulin and complement protein levels in the squirrel monkey. A cross-sectional life span study established that the developmental pattern of immunoglobulins and complement proteins was similar to that observed in humans. IgG and IgM levels rose progressively with age, while the complement system was mature at birth. In contrast to humans, this species showed a significant sex difference in IgG levels, with higher levels in males during both infancy and adulthood. Males also showed a greater antibody response to viral challenge than did females, and evaluation of gonadectomized subjects suggested that the sex difference in antibody production was testosterone-dependent. The effect of acute and sustained psychological disturbance on IgG levels was also evaluated in infant monkeys. Repeated, brief separations from the mother did not alter IgG levels, but IgG levels were suppressed after a 7-day removal from the mother. Therefore, despite the general view that immunoglobulin levels are relatively stable, these studies have established that immunoglobulin levels can be strongly influenced by hormonal and experiential factors in the squirrel monkey.

The usefulness of nonhuman primates in immunologically relevant research has until now been limited by difficulties in characterizing the major histocompatibility (MHC) gene products of these species. We have now biochemically characterized the MHC-encoded class I molecules from four different species of nonhuman primates using antibodies directed against human MHC class I structures and one-dimensional isoelectric focusing (1-D IEF). We demonstrated the functional relevancy of this technique of MHC typing by generating virus-specific cytotoxic T cells and assaying their cytotoxic activity against a panel of virus-transformed cells that expressed the same or differing class I structures. Only virus-infected cell lines expressing MHC class I antigens identical to those of the cytotoxic T lymphocyte population were lysed. This simple method of MHC class I typing using 1-D IEF will be useful in immunological research involving nonhuman primates and in nonhuman primate colony management.

Serum IgG and IgM levels were measured in domestically bred squirrel monkeys (Saimiri sciureus) ranging in age from 0 days to 42 months, as well as in adult squirrel monkeys from the wild estimated to be 60 months or older. The results indicated that the transplacental transfer of IgG occurs in the squirrel monkey but the transferability is lower in the squirrel monkey than in the cynomolgus monkey. Immune response in the squirrel monkey occurs just after birth, as shown by IgM production.

Characterization and functional aspects of squirrel monkey peripheral blood mononuclear cells (PBMC), and mainly T cells, are described in the present paper; this should enable the study of cellular immune responses in an experimental model for malaria. PBMC were obtained from Ficoll-Hypaque gradient separation and fractionated into T cells and non-T cells by means of E-rosetting techniques and adherence to plastic dishes. PBMC subset phenotypes were characterized by means of monoclonal antibodies (mAb) directed against human leukocyte differentiation antigens (Ag), fluoresceinated lectins, anti-surface Ig (squirrel-monkey-specific) antibodies (Ab) and latex bead ingestion assays. PBMC functions were assayed through lymphoblastic transformation tests (LTT) in the presence of either numerous mitogenic, comitogenic and anti-mitogenic lectins or anti-human leukocyte differentiation Ag mAb. We sought to standardize reference values for lymphocyte phenotypes and functions in normal squirrel monkeys (prior to experimental infection). We also present evidence that splenectomy (generally rendered necessary for experimental human malaria infection) performed six months prior to the present investigation did not modify PBMC numbers and functions in the tested animals.

Three of 275 squirrel monkeys (Saimiri sciureus) demonstrated a positive reaction to Koch's old tuberculin within 72 hours after the test. The animals were again positive when retested 1 week later. Two of the monkeys were killed and necropsied. There was no gross nor histologic evidence of tuberculosis. Organisms were cultured from liver, spleen, and mediastinal lymph node. These were identified as Runyon Group II mycobacteria, Mycobacteria gordoneae.

Antigenicity of IgG was compared among human, the cynomolgus monkey, the African green monkey and the squirrel monkey by the quantitative precipitation test using purified IgG of and rabbit anti-IgG serum to each species. Clear cross-antigenicity was observed between the cynomolgus monkey and the African green monkey and less clear cross-antigenicity between human and the cynomolgus monkey or the African green monkey. The cross-antigenicity observed between the squirrel monkey and the other three species examined was evidently weak.

Humoral and cell-mediated immunity (CMI) to herpesvirus saimiri (HVS), an oncogenic lymphotropic herpesvirus, was studied in squirrel and rhesus monkeys. Natural antibody to HVS was found in five of six squirrel monkeys but there was no evidence of specific CMI directed against HVS. Rhesus monkeys did not show natural antibody or CMI against HVS antigens. Immunization with HVS, however, produced both antibody and specific CMI in the rhesus monkeys, but no CMI developed in the squirrel monkeys. These findings are important in the development of animal models for the treatment of tumors associated with lymphotropic herpesviruses.

The potential for utilization as a contraceptive vaccine of a 60,000 Mr glycoprotein component, purified porcine zona antigen (PPZA), isolated from porcine zonae, was investigated in the squirrel monkey. Immunization resulted in production and maintenance of high antibody titers for at least 1 year. Comparable immune profiles were obtained using either monkey or pig zonae in assay systems, but dose-dependent variations in immune response were not observed. In situ antibody binding to monkey zonae was detected, but significantly fewer ovulated eggs were obtained from immunized monkeys than from controls. Exposure of antibody-pretreated pig, monkey, and human zonae to homologous sperm resulted in total inhibition of sperm attachment for the respective species. Thus, the contraceptive potential of PPZA antibodies in these species is demonstrated.

A method is described for inducing the production of large amounts of ascitic fluid (AF) in the squirrel monkey Saimiri sciureus. The total amount of protein in the induced AF is close to 60% of that in the serum. Electrophoretic analysis of serum and AF samples from the same monkey revealed similar protein patterns, including gamma globulins. Antibody titers against Plasmodium falciparum in infected monkeys, measured by indirect immunofluorescence, were also comparable in serum and AF.

Four monoclonal antibodies (mAb) which recognize distinct epitopes on Saimiri immunoglobulins were successfully used to characterize the protective and non-protective antibodies developed by this experimental host in response to infection by the human malaria parasite Plasmodium falciparum. Two of these mAb, 3F11/G10 and 3E4/H8, were IgG-specific and directed against conformationally conserved epitopes on the intact molecule. mAb 3A2/G6 and 4G3/B5 were specific for epitopes on two distinct L chain types of all Ig. Radioimmunoassay and immunoblots indicated that L-chains defined by 3A2/G6 were present in IgG molecules of type 3F11/G10, while L chains defined by 4G3/B5 were associated with IgG of type 3E4/H8. The use of these four mAb as immunoadsorbents allowed the purification of two IgG and two Ig populations. When assayed in vivo by passive transfer experiments in recipient monkeys previously infected by P. falciparum, protection could be associated with the IgG or Ig populations of type 3F11/G10-3A2/G6. In contrast, recipient animals which received the IgG type 3E4/H8 or Ig type 4G3/B5 presented rapidly evolving parasitemia similar to that in animals which received non-immune IgG or protective immune sera depleted of the above Ig or IgG fractions. The identification of protective and non-protective Ig populations will help in the evaluation of vaccine candidates against P. falciparum.

Monoclonal antibodies against the squirrel monkey Saimiri sciureus IgG have been produced for a more specific analysis of the antibody-related immunological aspects in experimental human or monkey malaria. Two monoclonal antibodies, 3D8/D5 and 3F11/G10, out of 64 reacted with distinct epitopes on the IgG present throughout the complete population without interfering with each other. The 2 monoclonal antibodies were used to develop a highly specific, reliable and sensitive two-site sandwich radioimmunoassay for the measurement of the serum IgG levels in 83 animals. The antibodies also allowed us to produce by a simple immunoabsorbent technique a highly purified IgG standard easy to calibrate and store. The assay permits the detection of IgG levels as low as 0.48 ng/ml. The standard curve is linear between 3.9 and 125 ng protein/ml and allows by a simple mathematical equation an accurate measurement of the serum IgG levels.