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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.

We have applied a panel of monoclonal antibodies against antigens present on the surface of human mononuclear cells to the study of mononuclear cell surface antigens expressed by seven species of New World primates. Antibodies to the sheep erythrocyte receptor (OKT11a) to a thymocyte antigen (OKT10), to the I region of the major histocompatibility locus (OKIa), and to an antigen found on the surface of human monocytes (OKM1) cross-reacted with mononuclear cell surface antigens of most of the species studied. Antibodies to antigens which have been correlated with functional capabilities in the human system (OKT4, OKT5, OKT8, 3A1) were much less reactive with platyrrhine mononuclear cells. These reagents may be quite useful in studies of primate phylogeny and immunology.

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.