MHC polymorphism in Caribbean African green monkeys

African green monkeys (AGM) are among the most widely used nonhuman primate models used in various fields of medical research. One species of AGM that originated from West Africa, Chlorocebus sabaeus, was introduced three centuries ago in the Caribbean islands. We present here a systematic study of the major histocompatibility complex (MHC) polymorphism of Caribbean AGM which is currently frequently used as an animal model. We studied 54 animals originated from Barbados (N?=?25) or Saint Kitts (N?=?29). The MHC polymorphism was characterized by means of 17 MHC microsatellites spread across MHC and DRB genotyping by DGGE sequencing. We defined nine frequent MHC haplotypes of which two were found in the two insular populations suggesting either past exchanges between the two populations or a common origin of the founders of the two populations. By the analysis of a previously described EST library, we characterized 38 MHC cDNA sequences (17 class I and 21 class II). In conclusion, we characterized for the first time the MHC polymorphism of Barbados and Saint Kitts AGM. We found a restricted polymorphism due to a founding effect, which is responsible for a strong bottleneck. The poorness of MHC polymorphism observed in the Caribbean AGM populations is similar to that observed in the Mauritian cynomolgus macaque population.     

Induction of AIDS by simian immunodeficiency virus from an African green monkey: species-specific variation in pathogenicity correlates with the extent of in vivo replication.

Previous studies suggested that simian immunodeficiency viruses isolated from African green monkeys (SIVagm) are relatively nonpathogenic. The report describes the isolation and biologic and molecular characterization of a pathogenic SIVagm strain derived from a naturally infected African green monkey. This virus induced an AIDS-like syndrome characterized by early viremia, frequent thrombocytopenia, severe lymphoid depletion, opportunistic infections, meningoencephalitis, and death of five of eight macaques within 1 year after infection. An infectious clone derived from this isolate reproduced the immunodeficiency disease in pig-tailed (PT) macaques, providing definitive proof of the etiology of this syndrome. Although the virus was highly pathogenic in PT macaques, no disease was observed in experimentally infected rhesus macaques and African green monkeys despite reproducible infection of the last two species. Whereas infection of PT macaques was associated with a high viral load in plasma, peripheral blood mononuclear cells, and tissues, low-level viremia and infrequent expression in lymph nodes of rhesus macaques and African green monkeys suggest that differences in pathogenicity are associated with the extent of in vivo replication. The availability of a pathogenic molecular clone will provide a useful model for the study of viral and host factors that influence pathogenicity.     

Chemotherapy-induced hemolytic uremic syndrome: Description of a potential animal model

Hemolytic uremic syndrome (HUS) is an uncommon complication of chemotherapy that contributes to the morbidity of oncology and bone marrow transplant patients. The pathogenesis is not well understood and no established clinical animal model exists. We studied four rhesus monkeys (RM) that developed fatal HUS following high-dose chemotherapy. Microangiopathic hemolytic anemia (pre-Hct 40% and day 5–8 Hct 31 % (P <.05), increased BUN (168 mg/dl), creatinine (8.2 mg/dl), and lactate dehydrogenase (1458 IU/L) (mean day 5–8 measurements) were observed. Platelets counts decreased to 39±15 × 10⁹/l from a mean of 397±31 × 10⁹/L (P < .0001). vWF, AT_III, thrombin:anti-thrombin complex (T:AT) and prothrombin fragment F1.2 levels were not different from a control group (N = 2). The data presented describe chemotherapy-induced HUS with typical clinical and laboratory features which may provide an animal model for the study of this important syndrome.     

Molecular characterization of simian lentiviruses from east African green monkeys

Asymptomatic infection with simian lentiviruses (also called simian immunodeficiency viruses, or SIV) is common among feral African green monkeys. To characterize the range of SIV genetic diversity among infected African green monkeys, we have determined nucleotide sequences from complete or partial molecular clones of four distinct SIVagm isolates from Kenya and Ethiopia. The nucleotide and amino acid variability we observed among the SIVagm isolates was greater than the variability within any other group of primate lentiviruses. These data suggest that: a) African green monkeys have been infected with simian lentiviruses for many years; and b) novel and uncharacterized primate lentiviruses may exist in the feral African green monkey population in other parts of Africa.     

Molecularly cloned simian immunodeficiency virus SIVagm3 is highly divergent from other SIVagm isolates and is biologically active in vitro and in vivo.

Simian immunodeficiency viruses have been isolated from African green monkeys originating from Ethiopia. A molecular clone, termed SIVagm3, was found to be highly divergent from SIVagmTYO-1 in terms of its restriction map and partial nucleotide sequence. A premature stop codon present in the transmembrane protein of SIVagm TYO-1 was absent in SIVagm3. SIVagm3 was biologically active in vitro and in vivo and displayed characteristics reminiscent of the wild-type virus. Biological activity was demonstrated by seroconversion of juvenile African green monkeys and Macaca nemestrina after inoculation. In contrast to antibody reactivity mainly directed against env proteins in naturally infected African green monkeys. African green monkeys and M. nemestrina infected with the cloned virus showed antibody reactivity directed against all major proteins as demonstrated by immunoblot analysis. The availability of a biologically fully competent molecular clone of SIVagm allows us now to address various pertinent questions in an animal model system which should help to understand features of human immunodeficiency virus infection in human beings.     

Molecular clones of SIVsm and SIVagm: experimental infection of macaques and African green monkeys

We have investigated the ability of biologically-active proviral molecular clones of SIVsm and SIVagm to infect rhesus macaques, pig-tail macaques, and African green monkeys. Two clones of SIVsm were individually inoculated into four rhesus and four pig-tail macaques. All eight macaques became infected, and two have experienced a significant decline in absolute numbers of circulating CD4+ cells. None of three African green monkeys were infected by an SIVsm molecular clone. However, one of four African green monkeys did become infected by SIVsm after receiving lymphocytes directly from an SIVsm-infected rhesus macaque. A molecular clone of SIVagm infected three of four macaques and three of three African green monkeys. None of the three infected macaques had a significant decline in circulating CD4+ cells. Interestingly, infection of pig-tail macaques (but not rhesus macaques) with uncloned SIVagm induced a significant drop in circulating CD4+ cells. These data suggest that molecular clones of SIVsm and SIVagm can be used in experimental models of AIDS for the evaluation of viral gene functions and for the study of in vivo genetic variation.     

The biological functions of polyamine oxidation products by amine oxidases: Perspectives of clinical applications

Summary. The polyamines spermine, spermidine and putrescine are ubiquitous cell components. If they accumulate excessively within the cells, due either to very high extracellular concentrations or to deregulation of the systems which control polyamine homeostasis, they can induce toxic effects. These molecules are substrates of a class of enzymes that includes monoamine oxidases, diamine oxidases, polyamine oxidases and copper containing amine oxidases. Polyamine concentrations are high in growing tissues such as tumors. Amine oxidases are important because they contribute to regulate levels of mono- and polyamines. These enzymes catalyze the oxidative deamination of biogenic amines and polyamines to generate the reaction products H₂O₂ and aldehyde(s) that are able to induce cell death in several cultured human tumor cell lines. H₂O₂ generated by the oxidation reaction is able to cross the inner membrane of mitochondria and directly interact with endogenous molecules and structures, inducing an intense oxidative stress. Since amine oxidases are involved in many crucial physiopathological processes, investigations on their involvement in human diseases offer great opportunities to enter novel classes of therapeutic agents.     

Growth/differentiation factor 7 is preferentially expressed in the primary motor area of the monkey neocortex

We applied a differential display PCR technique to isolate molecules that are area-specific in expression in the primate neocortex, and found that growth/differentiation factor 7 (GDF7), a member of the bone morphogenetic protein (BMP)/transforming growth factor (TGF) beta super-family, is preferentially expressed in the primary motor area of African green monkeys (Cercopithecus aethiops). We proved that GDF7 is 10 times more abundant in the motor cortex than in the visual cortex by northern blotting and quantitative RT-PCR. When we examined the neocortex of closely related rhesus monkeys (Macaca mulatta), GDF7 was also most abundant in the motor cortex, although the regional difference was reduced to 3-fold. This differential expression pattern was observed in both newborn and infant rhesus monkeys. We found that several type I/II receptors of BMP, candidates of the receptors for GDF7, are uniformly expressed in the mature neocortex. The unique expression pattern of GDF7 suggests that it may play an active role in the motor area of the primate neocortex.     

Relation between phylogeny of African green monkey CD4 genes and their respective simian immunodeficiency virus genes

Abstract: An apparent species-specific relatedness of SIV_agm suggests a coevolution with their natural hosts. However, the exact species or subspecies classification of African green monkeys, AGM, is uncertain because current classification schemes rely on phenotype markers, while more definitive genetic data are lacking. In this study, the CD4 protein involved in tissue type recognition was genetically cloned and sequenced from PBMC RNA from all AGM species, including Barbados green monkeys (BGM). Phylogenetic trees were constructed that also included genomic CD4 nucleotide sequences from patas, sooty mangabeys, rhesus and pig-tail macaques, chimpanzees, and humans. Chimpanzees and humans consistently clustered together. Monkeys within the Cercopithecus genus formed a separate cluster which included pata monkeys, supporting its grouping as a member of Cercopithecus. Surprisingly, sooty mangabeys were genetically more closely related to Asian macaques than to other African species, which might explain why macaques are more susceptible to infection by the SIV_sm group than to infection by SIV_agm or HIV-1 and why patas, on the other hand, are highly susceptible to SIV_agm infection. Based on CD4 genetic data, tantalus, vervets, grivets, and sabaeus formed separate subgroups with BGM grouping closely with vervets. The branching order of the AGM species was related to that of their respective SIV_agm env sequences. The study suggests a strong correlation between CD4 phylogeny and the susceptibility of the host species to infection by a specific lentivirus and supports the assumption of a coevolution of SIV_agm and AGM. CD4 sequencing is suggested as a relevant method for genetic determination of primate species.     

Species-specific diversity among simian immunodeficiency viruses from African green monkeys

The prevalence, natural history, and genetic characteristics of simian immunodeficiency virus (SIV) infections in most feral African monkey species are presently unknown, yet this information is essential to elucidate their origin and relationship to other simian and human immunodeficiency viruses. In this study, a combination of classical and molecular approaches were used to identify and characterize SIV isolates from West African green monkeys (Cercopithecus sabaeus) (SIVagm isolates). Four SIVagm viruses from wild-caught West African green monkeys were isolated and analyzed biologically and molecularly. Amplification, cloning, and sequencing of a 279-bp polymerase fragment directly from uncultured peripheral blood mononuclear cells was facilitated by the use of nested polymerase chain reaction. The results indicated that West African green monkeys are naturally infected with SIVs which are closely related to East African SIVagm isolates. However, structural, antigenic, and genetic differences were observed which strongly suggest that the West African green monkey viruses comprise a phylogenetically distinct subgroup of SIVagm. These findings support our previous hypothesis that SIVagm viruses may have evolved and diverged coincident with the evolution and divergence of their African green monkey host. In addition, this study describes a polymerase chain reaction-based approach that allows the identification and molecular analysis of divergent SIV strains directly from primary monkey tissue. This approach, which does not depend on virus isolation methods, should facilitate future studies aimed at elucidating the origins and natural history of SIVs in feral African green monkey populations.     

Genetic polymorphism of alpha 1-antitrypsin in green monkeys studied by isoelectric focusing and family analysis

24 variants of alpha 1-antitrypsin (alpha 1-AT) were recognized in sera of 120 wild and capture-born African green monkeys by isoelectrofocusing in Ampholine PAG-plates (pH 4-6.5) and western blotting with antihuman alpha 1-AT serum. All variants had much more cathodal position than human alpha 1-AT and revealed very high microheterogeneity which was slightly different from the observed in human alpha 1-AT. The alpha 1-AT banding pattern allowed to postulate existence of 10 codominant alleles in the Pi locus of African green monkeys. The reality of 8 alleles was proved by family analysis which included 45 monkey birth cases. Two other alleles were absent in the parents available. Thus, alpha 1-AT is the most polymorphic among the known serum proteins of African green monkeys. The latter can be useful for molecular systematics of these primates.     

A species comparison of low density lipoprotein heterogeneity in nonhuman primates fed atherogenic diets

Six male cynomolgus monkeys and five male African green monkeys were fed dietary cholesterol to induce hypercholesterolemia. The two groups studied had equivalent total plasma cholesterol concentrations. Low density lipoproteins (LDL) were isolated from whole plasma by ultracentrifugation and separated from other lipoprotein classes by agarose column chromatography. LDL were further subfractionated by density gradient ultracentrifugation in a VTi-50 vertical rotor. The material within five density regions was pooled from each sample and molecular weight, electrophoretic mobility, apoprotein heterogeneity, and percentage composition were determined for each subfraction. In general, cynomolgus monkey LDL were larger and more polydisperse than African green monkey LDL, and the LDL subfractions of cynomolgus monkeys were generally of lower densities although molecular weights at any density were in the same range for both species. ApoB-100 was the major apoprotein in each subfraction. ApoE was frequently present in the less dense subfractions while apoA-I was often seen in the more dense subfractions. Cynomolgus monkey LDL appeared to contain more apoE than African green monkey LDL. Over the entire spectrum of LDL, the percentage composition of the particles at any given density was indistinguishable between the species. In general, the average cynomolgus monkey LDL was larger, more polydisperse, less dense, and appeared to contain more apoE than the average African green monkey LDL. One or all of these differences might help explain the increased susceptibility to diet-induced atherosclerosis seen in cynomolgus monkeys.     

Nonclassical mucosal antibodies predominate in genital secretions of HIV-1 infected chimpanzees

To identify mucosal immunity in HIV-infected chimpanzees, IgG, IgA, and IgM from plasma, saliva, rectal swabs, vaginal washes, semen, and urethral washes were tested from four male and three female HIV-1_IIIB infected chimpanzees. The level of HIV infections in the seven chimpanzees were classified as high, intermediate and low depending on the number of HIV-1 infected cells per 10⁷ peripheral blood mononuclear cells (PBMC). One male chimpanzee had a relatively high viral load, two males and two females had moderate viral loads and one male and one female had low levels of infection. All seven animals had plasma antibody. The principal finding was that nonclassical mucosal antibodies of the IgG isotype were the predominant antibody in the saliva, rectal swabs, vaginal washes, semen, and urethral washes of infected animals. All plasma and mucosal samples were negative for IgM antibodies. The results show that HIV-1 specific IgG responses and not sIgA predominate at mucosal surfaces of HIV-1_IIIB infected chimpanzees. A trend was observed in which high viral loads correlated with high plasma IgG, IgA and sIgA titers. An overall correlation between relatively high virus loads and high amounts of mucosal IgG was also found.     

Magnetic resonance imaging in vivo monitoring of T2 relaxation time: Quantitative assessment of primate brain maturation

This work describes quantitative MRI assessment of primate brain maturation. Nine young baboons were followed from the age of one to 30 months. Assessment of myelination was based on the gray/white matter contrast on MR images and the evolution of T2 relaxation time respectively. The brain maturation began in the posterior fossa and progressed to the olfactory bulbs corresponding to decreasing white matter T2 values. Relaxation parameters provide new opportunities to trace the myelination process in vivo.     

A restraining system for rhesus monkeys used in space research

A new chronic restraining system has been designed specifically for male rhesus monkeys (8–13 kg) housed in weightlessness for scientific purposes. The restraining system consists of a coat (jacket and skirt) and a chair. The system separates the thorax from the lower part of the body, and allows large movements of the upper part of the body. The macaque can feed and groom itself, and can also sleep in close-to-normal position.     

Encephalomyocarditis virus infection in African green and squirrel monkeys: comparison of pathologic effects

This study compared the pathological effects of experimental encephalomyocarditis virus infection in African green monkeys (Cercopithecus aethiops) and squirrel monkeys (Saimiri sciureus). African green monkeys died within 5 days post-inoculation and significant changes were limited to moderate to severe multifocal nonsuppurative necrotizing myocarditis. Squirrel monkeys were less severely affected and died or were euthanized 4 to 41 days post-inoculation. Myocardial and central nervous system changes in squirrel monkeys were minimal to moderate and variable in distribution, while pancreatic changes were more severe and included multifocal necrosis, subacute inflammation and atrophy of exocrine cells.     

Quantitation of a lentivirus in its natural host: simian immunodeficiency virus in African green monkeys.

We have examined the viral load in the peripheral blood of simian immunodeficiency virus (SIV)-infected African green monkeys with a view to the unexplained apathogenicity of African green monkey SIV (SIVagm) in its natural host. By using polymerase chain reaction, viral DNA was detected in fresh peripheral blood mononuclear cells (PBMC) of each of nine seropositive animals. The virus DNA load was variable among the monkeys tested, ranging from 5 to 50 (mean = 15) copies per 10(5) PBMC, which is comparable to that of human immunodeficiency virus type 1 (HIV-1) in humans. The level of infectious SIVagm in PBMC was measured by endpoint dilution cultures. SIVagm was recovered from PBMC from 14 of 17 antibody-positive monkeys (82%), and the mean SIVagm titer in PBMC of seropositive African green monkeys was 10 tissue culture infectious doses per 10(6) cells, similar to the titer shown for HIV in asymptomatic carriers. Free infectious virus was isolated from the plasma of 4 of 17 monkeys (24%), and SIVagm expression in peripheral blood in vivo, as demonstrated by in situ hybridization, was detectable only in those animals which were viremic. SIVagm replication is therefore not totally suppressed in vivo, and SIVagm has a viral load equivalent to that seen for HIV-1 in asymptomatic humans.     

Serological survey for two simian retroviruses in macaques and African green monkeys

Colonies of nonhuman primates at the Bowman Gray School of Medicine (BGSM) were tested for antibodies to two retroviruses associated with immunodeficiency by indirect immunofluorescence (IFA) and western blot. A total of 471 cynomolgus macaques (Macaca fascicularis), 144 rhesus monkeys (M. mulatta) and 67 stumptail monkey M. arctoides) were tested for SRV-1, and 152 African green monkeys (Cercopithecus aethiops) were tested for SIV. Of the macaques tested, 170 (36%) cynomolgus, 5 (3%) rhesus and 8 (12%) stumptails were positive for SRV-1 antibodies by IFA. Of the African green monkeys, 54 (36%) were IFA positive for SIV antibodies. A total of 143 African green monkeys tested by IFA also were tested by western blot. In the African green monkeys, the IFA had a positive predictive value of 98% and a negative predictive value of 96%. Of 176 IFA positive macaque sera tested by western blot, 49 (28%) were positive, 55 (31%) were considered equivocal (only one band, usually to p27 core protein), and 72 (41%) were negative.     

Mitochondrial Diversity and Distribution of African Green Monkeys (Chlorocebus Gray, 1870)

African green monkeys (Chlorocebus) represent a widely distributed and morphologically diverse primate genus in sub‐Saharan Africa. Little attention has been paid to their genetic diversity and phylogeny. Based on morphological data, six species are currently recognized, but their taxonomy remains disputed. Here, we aim to characterize the mitochondrial (mt) DNA diversity, biogeography and phylogeny of African green monkeys. We analyzed the complete mitochondrial cytochrome b gene of 126 samples using feces from wild individuals and material from zoo and museum specimens with clear geographical provenance, including several type specimens. We found evidence for nine major mtDNA clades that reflect geographic distributions rather than taxa, implying that the mtDNA diversity of African green monkeys does not conform to existing taxonomic classifications. Phylogenetic relationships among clades could not be resolved suggesting a rapid early divergence of lineages. Several discordances between mtDNA and phenotype indicate that hybridization may have occurred in contact zones among species, including the threatened Bale monkey (Chlorocebus djamdjamensis). Our results provide both valuable data on African green monkeys’ genetic diversity and evolution and a basis for further molecular studies on this genus. Am. J. Primatol. 75:350‐360, 2013. © 2013 Wiley Periodicals, Inc.