Genetic differentiation between baboon subspecies: Relevance for biomedical research

The use of common names which may encompass a number of subspecies or species is pervasive in the biomedical literature. Failure to identify the complete taxonomic classification of research subjects presents a source of error for scientists attempting to evaluate results or to repeat experiments. This paper examines the problem in a common animal model, the baboon. Analyses of the genetic distances among five baboon subspecies (Papio hamadryas anubis, P.h. cynocephalus, P.h. papio, P.h. ursinus, and P.h. hamadryas) based on blood marker information from nine polymorphic protein loci (ADA, APRT, C3, CA1, CA2, GPI, MPI, PEPB, and PGD) available for baboons resident at the Southwest Foundation for Biomedical Research are presented. Statistical tests on the distances showed that significant genetic differences exist among the subspecies. A comparison of P.h. anubis and P.h. cynocephalus revealed that these two subspecies also differ significantly for biomedically relevant lipoprotein cholesterol levels, as can be predicted from the genetic distances. The results emphasize the pitfalls of using different types of baboons interchangeably in experimental protocols.     

Adenine phosphoribosyl transferase polymorphism in baboons

Two allelic isozymes of adenine phosphoribosyl transferase (APRT) were detected by starch gel electrophoresis of baboon hemolysates. Extensive family data verified autosomal codominant inheritance. The gene frequencies of five subspecies of baboons differed significantly. The activity of erythrocyte APRT is sufficiently high to enable the use of this enzyme as a sensitive marker for assessing chimerism in research involving bone marrow transplantation.This research was supported in part by NIH Grants HL28972, HG00336, and HV53030.     

Cranial ontogeny of Papio baboons (Papio hamadryas)

Cranial form in subspecies of Papio baboons (Papio hamadryas) varies in relation to size, geography, and sex. However, knowledge about this variation is based mainly on adults, precluding direct assessments of the evolutionary factors that are ultimately responsible for adult shape variation. Consequently, this study tests hypotheses about the development of size and shape differences among subspecies of Papio baboons, anticipating limited evolutionary divergences in the ontogenetic pathways leading to adult endpoints. Geometric morphometric and bivariate allometric analyses are used to explore developmental size and shape variation. Allometric scaling in adult Papio baboons occurs because both sexes and all subspecies follow similar developmental pathways to a variety of adult forms. However, complex allometry contributes to form differences, producing potentially important shape differences that emerge during development. Modest shape differences that are statistically independent of size distinguish chacma baboons (P. h. ursinus) from other forms. A small-headed subspecies, the Kinda baboon (P. h. kindae), also presents a distinctive ontogeny, and may provide insights into the evolution of size change in this species. Variation among subspecies that is statistically independent of size involves the rostrum, zygomatic breadths, and cranial flexion. These features may be related to diet, but the precise biomechanical correlates of baboon form variation remain unclear.     

Dental eruption schedules of wild and captive baboons

Dental eruption schedules previously used to age wild baboons have in the past derived from studies of captive animals housed under standard conditions and fed standard laboratory diets. This paper reports for the first time eruption schedules derived from wild baboons, the yellow baboons (Papio hamadryas cynocephalus) of Mikumi National Park, Tanzania, and compares these schedules with those of other baboon subspecies inhabiting both similar and dissimilar environments. Eighteen males and twelve females from the Viramba groups, ranging in age from 21 to 103 months, were trapped, and dental impressions and notes were made of the state of eruption of each tooth. Eruption of all teeth were delayed at Mikumi relative to the baboon standards derived from the captive animals at the Southwest Foundation for Biomedical Research, San Antonio, Texas. Teeth of the canine-premolar 3 complex and third molars were most delayed, erupting up to a year and a half later than their counterparts from captive animals. Comparison with data on hamadryas baboons from Erer-Gota in Ethiopia revealed that both the hamadryas and yellow subspecies of baboons, with different genetic backgrounds and living under markedly different environmental conditions, followed the same schedule. This constancy of developmental schedules suggests that these Mikumi data may reasonably be used as standards for other wild baboon populations and that acceleration of dental maturation, as well us maturation of other somatic systems in captivity, is another manifestation of the short-term adaptive plasticity of the baboon species as a whole.     

Evolutionary implication of heterogeneity of the nontranscribed spacer region of ribosomal DNA repeating units in various subspecies of Mus musculus

Genetic variability of the nontranscribed spacer (NTS) region within ribosomal DNA repeating units in the various subspecies of Mus musculus was determined. Mice belonging to several laboratory mouse strains were examined by means of Southern blot hybridization with a mouse ribosomal DNA probe. This probe encompasses the 3' end of the 28S ribosomal RNA (rRNA) gene and the following spacer. Restriction enzyme digestions of the liver DNAs from various wild mice revealed that each of the subspecies has a unique pattern in the spacer encompassing a distance approximately 10 kb downstream from the ribosomal gene. These restriction patterns permit the classification of mouse subspecies and also provide insights into the origin of the laboratory mouse strains.     

Hematology and blood biochemistry in infant baboons (Papio hamadryas)

Although published normative reference standards for hematologic and clinical chemistry measures are available for adult baboons, their applicability to infants has not been addressed. We analyzed these measures in 110 infant baboons (55 females and 55 males) from a large breeding colony at the Southwest Regional Primate Research Center in San Antonio, Texas. The sample consists of olive baboons and olive/yellow baboon hybrids, 1 week to 12 months of age. We produced cross-sectional reference values and examined the effects of age, sex, and subspecies on these variables. Hematology reference ranges for infant baboons are similar to, but wider than, those for adults. Reference ranges for blood biochemistry measures are generally more dissimilar to adults, indicating that for many variables, reference ranges for adult baboons are not adequate for infants. Although sex and subspecies differences are rare, age accounts for more than 10% of the variance in many of the variables.     

Cyclic oscillations in melanin composition within hairs of baboons.

The wild-type agouti-banding pattern for hair is well characterized in lower mammals such as mice. The switch between eumelanin and pheomelanin in bands in the hair results from the interaction of alpha-melanocyte stimulating hormone and agouti signal protein through the melanocortin 1 receptor on melanocytes. However, such banding patterns have not been described to date in higher mammals. We now report such 'agouti'-banding patterns that occur in several subspecies of baboons, and characterize those hairs using chemical and immunohistochemical methods. Hair and skin samples were obtained from the dorsa of adult male baboons of different subspecies (Papio cynocephalus hamadryas (PCH) and Papio cynocephalus anubis (PCA)). The hairs were excised with scissors into the gray and the white bands of the PCH subspecies and into the black and the yellow bands of the PCA subspecies, and were analyzed for total melanin, eumelanin, and pheomelanin by spectrophotometric and chemical methods. Hairs in the PCA subspecies oscillate between a eumelanic band (with high melanin content) and a pheomelanic band, while hairs in the PCH subspecies oscillate between a eumelanic band (with low melanin content) and a non-pigmented band. Those chemical data are consistent with the histological appearance of the hair bulbs stained by the Fontana-Masson technique. The difference in the melanin content between PCH and PCA subspecies is most likely related to tyrosinase levels, as suggested by the presence of unpigmented muzzle in the PCH subspecies compared with the black muzzle in the PCA subspecies.     

Characterization of the subunit composition of HGPRTase from human erythrocytes and cultured fibroblasts

Hypoxanthine-guanine phosphoribosyltransferase is a ubiquitous human enzyme, the inherited deficiency of which leads to a specific metabolic-neurological syndrome. Native acrylamide isoelectric focusing revealed that the human enzyme consists of different numbers of isoenzymes depending on the tissue of origin. The erythrocytic enzyme has the most isoenzymes while the enzyme from cultured fibroblasts has only a single isoenzyme. The isoenzyme pattern of the erythrocytic enzyme changes on storage of the crude hemolysate at 4 C. Treatment of the stored crude hemolysate with 4.5 m urea and 0.35 mm -mercaptoethanol results in an isoenzyme pattern similar to that of the fresh crude extract. Thus the additional isoenzymes are generated on storage not by covalent modification of the enzyme but probably by binding of small molecules to the enzyme or to association of the enzyme molecules. Hypoxanthine-guanine phosphoribosyltransferase has been purified to 80% homogeneity in three steps, DEAE Sephadex chromatography, heat treatment at 85 C for 5 min, and hydroxylapatite chromatography. Denaturing two-dimensional gel electrophoresis of the erythrocytic enzyme revealed that the erythrocytic enzyme is composed of three major types of subunits (1–3) with the same molecular weight but different isoelectric points. In contrast, the fibroblast enzyme is composed of only a single type of subunit, which comigrates with subunit 1 of the erythrocytic enzyme. Since there is a single genetic locus in humans for HGPRTase (the enzyme is X linked) (Nyhan et al., 1967), the observed subunit modification of the erythrocyte enzyme appears to be the result of posttranslational modification. These findings provide a simple explanation for the observed electrophoretic properties of human HGPRTase. A patient with 0.5% of HGPRTase activity in his erythrocytes was found to have small amounts (> 0.5% but < 5% of normal) of the erythrocytic HGPRTase subunits.This work was supported by a grant from NIAMDD, National Institutes of Health, United States Public Health Service. L. J. G. was supported by a fellowship from the National Institute of Child Health and Human Development. D. W. M. is an Investigator, Howard Hughes Medical Institute.     

Cyclic Oscillations in Melanin Composition within Hairs of Baboons

The wild‐type agouti‐banding pattern for hair is well characterized in lower mammals such as mice. The switch between eumelanin and pheomelanin in bands in the hair results from the interaction of α‐melanocyte stimulating hormone and agouti signal protein through the melanocortin 1 receptor on melanocytes. However, such banding patterns have not been described to date in higher mammals. We now report such ‘agouti’‐banding patterns that occur in several subspecies of baboons, and characterize those hairs using chemical and immunohistochemical methods. Hair and skin samples were obtained from the dorsa of adult male baboons of different subspecies (Papio cynocephalus hamadryas (PCH) and Papio cynocephalus anubis (PCA)). The hairs were excised with scissors into the gray and the white bands of the PCH subspecies and into the black and the yellow bands of the PCA subspecies, and were analyzed for total melanin, eumelanin, and pheomelanin by spectrophotometric and chemical methods. Hairs in the PCA subspecies oscillate between a eumelanic band (with high melanin content) and a pheomelanic band, while hairs in the PCH subspecies oscillate between a eumelanic band (with low melanin content) and a non‐pigmented band. Those chemical data are consistent with the histological appearance of the hair bulbs stained by the Fontana‐Masson technique. The difference in the melanin content between PCH and PCA subspecies is most likely related to tyrosinase levels, as suggested by the presence of unpigmented muzzle in the PCH subspecies compared with the black muzzle in the PCA subspecies.     

Polymorphisms, Linkage and Mapping of Four Enzyme Loci in the Fish Genus Xiphophorus (Poeciliidae)

Electrophoretic variants at four additional enzyme loci--two esterases (Est-2, Est-3), retinal lactate dehydrogenase (LDH-1) and mannose phosphate isomerase (MPI)--among three species and four subspecies of fish of the genus Xiphophorus were observed. Electrophoretic patterns in F1 hybrid heterozygotes confirmed the monomeric structures of MPI and the esterase and the tetrametric structure of LDH in these fishes. Variant alleles of all four loci displayed normal Mendelian segregation in backcross and F2 hybrids. Recombination data from backcross hybrids mapped with Haldane's mapping function indicate the four loci to be linked as Est-2--0.43--Est3--0.26--LDH-1--0.19--MPI. Significant interference was detected and apparently concentrated in the Est-3 to MPI region. No significant sex-specific differences in recombination were observed. This group (designated linkage group II) was shown to assort independently from the three loci of linkage group I (adenosine deaminase, glucose-6-phosphate dehydrogenase, and 6-phosphogluconate dehydrogenase) and from glyceraldehyde-3-phosphate dehydrogenase and two isocitrate dehydrogenase loci. Evidence for conservation of the linkage group, at least in part, in other vertebrate species is presented.     

Mitochondrial phylogeny and systematics of baboons (Papio)

Baboons (Papio, s.s.) comprise a series of parapatric allotaxa (subspecies or closely related species) widely distributed in sub-Saharan Africa. Despite extensive studies of their ecology, morphology, and behavior, disagreement about their phylogenetic relationships continues, as expressed in the current coexistence of at least three major, competing taxonomic treatments. To help resolve this situation, we sequenced approximately 900 bases of mitochondrial DNA of 40 individuals from five of the widely recognized "major" allotaxa. Total sequence diversity (>5%) is high compared to most primate species. Major mitochondrial clades correspond to recognized allotaxa, with the important exception that haplotypes from yellow and olive baboons form a single, monophyletic clade within which the two allotaxa do not comprise mutually exclusive clusters. The major clades fall unambiguously into the pattern: (chacma (Guinea (hamadryas (yellow + olive)))). This phylogeny does not support taxonomies that oppose hamadryas to all other baboons ("desert" vs. "savanna"), but is compatible with the view that all definable allotaxa should be recognized as coordinates, either as "phylogenetic" species or "biological" subspecies. The close relationship and unsegregated distribution of haplotypes from Kenyan and Tanzanian yellow and olive baboons are unexplained, but may reflect introgression across the documented hybrid zone. The overall phylogeny, when combined with paleontological data, suggests a southern African origin for extant Papio baboons, with all extant lineages sharing a common mitochondrial ancestor at approximately 1.8 Ma.     

Group housing of hamadryas baboons: a new cage design based upon field studies of social organization

The hamadryas subspecies, which is utilized extensively for laboratory research, has a social structure unique among baboons. Field studies have shown that small one male units are the basic grouping in a complex multi-level social system. A housing design for a research colony of hamadryas baboons was conceived to reflect this unique social organization and related behavioral patterns. The outcome of combining such ethological considerations with practical dictates for housing large laboratory primates has been assessed in terms of reproductive performance, animal health and utilization for research. Various benefits of this approach to cage design for hamadryas baboons are identified in this report.     

A-B-O groups of Chacma baboons (Papio ursinus, Kerr) in South Africa

The blood and saliva groups of 51 Chacma baboons (Papio ursinus, Kerr) were determined. The ABO blood group frequencies differed significantly from another subspecies (P. u. ursinus) but they gave a close agreement with the Hardy-Weinberg estimates. Thus the data are compatible with the hypothesis that the saliva groups of baboons are the products of two allelic genes A and B that are inherited in a similar manner to the A and B genes of humans.     

当归水提液和醇提液体外抗脂质过氧化和红细胞溶血作用

研究当归水提液和醇提液对小鼠肝组织自发性过氧化酯质分解产物丙二醛（malondialdehyde,MDA）的生成和对红细胞膜脂质过氧化及红细胞溶血作用的影响。采用TBA比色法测定肝组织匀浆MDA生成,分光光度法测定过氧化氢诱导红细胞膜脂质过氧化和溶血。实验分为空白组、对照组、加药组。加药组分为25、50、100和200mg/mL四个浓度组。当归水提液和醇提液均在25～200mg／mL的浓度范围内,能够明显抑制小鼠肝组织匀浆自发性MDA的生成,具有抑制过氧化氢诱导红细胞膜脂质过氧化和溶血的作用,抑制效果随当归水提液和醇提液浓度的增大而逐渐增强,抑制率与药物浓度成良好的量效关系。当归水提液和醇提液具有抗脂质过氧化和红细胞溶血的作用。     

The heterogeneity of protein kinase C in various rat tissues

Expression of multiple subspecies of protein kinase C (PKC) was studied in various rat tissues. Three types of the enzyme designated type I, II, and III were analyzed, which have the structures of gamma-, beta- (beta I- and beta II-), and alpha-sequence, respectively. Type I enzyme was found only in the central nervous tissue, whereas type III enzyme appeared to be commonly present in various tissues such as liver, spleen, lung, testis, heart, and kidney. Type II enzyme was also found in these tissues. However, immunoblot and biochemical analysis indicated that type II enzyme of lung and heart was distinct from that of other tissues. The tissue-specific expression of PKC suggests that each subspecies of this enzyme has a defined function in processing and modulating tissue responses to external stimuli.     

Purification and characterization of the thyrotropin-releasing hormone (TRH)-degrading serum enzyme and its identification as a product of liver origin.

Previous biochemical studies have indicated that the membrane-bound thyrotropin-releasing hormone (TRH)-degrading enzyme (TRH-DE) from brain and liver and the serum TRH-DE are derived from the same gene. These studies also suggested that the serum enzyme is of liver origin. The present study was undertaken to verify these hypotheses. In different species, a close relationship between the activities of the serum enzyme and the particulate liver enzyme was noticed. The activity of the serum enzyme decreased when rats were treated with thioacetamide, a known hepatotoxin. With hepatocytes cultured in a sandwich configuration, release of the TRH-DE into the culture medium could also be demonstrated. The trypsin-solubilized particulate liver TRH-DE and the serum TRH-DE were purified to electrophoretic homogeneity. Both enzymes and the brain TRH-DE were recognized by a monoclonal antibody generated with the purified brain enzyme as antigen. Lectin blot analysis indicated that the serum enzyme and the liver enzyme are glycoproteins containing a sugar structure of the complex type, whereas the brain enzyme exhibits an oligomannose/hybrid glycostructure. A molecular mass of 97 000 Da could be estimated for all three enzymes after deglycosylation and SDS/PAGE followed by Western blotting. Fragment analysis of the serum TRH-DE revealed that the peptide sequences correspond to the cDNA deduced amino-acid sequences of the membrane-bound brain TRH-DE, whereby two peptides were identified that are encoded by exon 1. These data strongly support the hypothesis that the TRH-DEs are all derived from the same gene, whereby the serum enzyme is generated by proteolytic cleavage of the particulate liver enzyme.     

Differential microRNA response to a high-cholesterol, high-fat diet in livers of low and high LDL-C baboons

ABSTRACT: BACKGROUND: Dysregulation of microRNA (miRNA) expression has been implicated in molecular geneticevents leading to the progression and development of atherosclerosis. We hypothesized thatmiRNA expression profiles differ between baboons with low and high serum low-densitylipoprotein cholesterol (LDL-C) concentrations in response to diet, and that a subset of thesemiRNAs regulate genes relevant to dyslipidemia and risk of atherosclerosis. RESULTS: Using Next Generation Illumina sequencing methods, we sequenced hepatic small RNAlibraries from baboons differing in their LDL-C response to a high-cholesterol, high-fat(HCHF) challenge diet (low LDL-C, n = 3; high LDL-C, n = 3), resulting in 517 baboonmiRNAs: 490 were identical to human miRNAs and 27 were novel. We compared miRNAexpression profiles from liver biopsies collected before and after the challenge diet andobserved that HCHF diet elicited expression of more miRNAs compared to baseline (chow)diet for both low and high LDL-C baboons. Eighteen miRNAs exhibited differentialexpression in response to HCHF diet in high LDL-C baboons compared to 10 miRNAs in low LDL-C baboons. We used TargetScan/Base tools to predict putative miRNA targets;miRNAs expressed in high LDL-C baboons had significantly more gene targets thanmiRNAs expressed in low LDL-C responders. Further, we identified miRNA isomers andother non-coding RNAs that were differentially expressed in response to the challenge diet inboth high LDL-C and low LDL-C baboons. CONCLUSIONS: We sequenced and annotated baboon liver miRNAs from low LDL-C and high LDL-Cresponders using high coverage Next Gen sequencing methods, determined expressionchanges in response to a HCHF diet challenge, and predicted target genes regulated by thedifferentially expressed miRNAs. The identified miRNAs will enrich the database for noncodingsmall RNAs including the extent of variation in these sequences. Further, weidentified other small non-coding RNAs differentially expressed in response to diet. Ourdiscovery of differentially expressed baboon miRNAs in response to a HCHF diet challengethat differ by LDL-C phenotype is a fundamental step in understating the role of non-codingRNAs in dyslipidemia.     

Thioredoxin reductase is essential for the survival of Plasmodium falciparum erythrocytic stages

The human malaria parasite Plasmodium falciparum poses an increasing threat to human health in the tropical regions of the world, and the validation and assessment of possible drug targets is required for the development of new antimalarials. It has been shown that the erythrocytic stages of the parasites, which are responsible for the pathology of the disease in humans, are under enhanced oxidative stress and are particularly vulnerable to exogenous challenges by reactive oxygen species. Therefore it is postulated that the disruption of the antioxidant and/or redox systems of the parasite is a feasible way to interfere with their development during erythrocytic schizogony. In order to test this suggestion thioredoxin reductase (TrxR), an enzyme heavily involved in maintenance of redox homeostasis and antioxidant defense, was knocked out in P. falciparum. It was impossible to generate parasites with a disrupted trxR gene suggesting that TrxR is essential for P. falciparum erythrocytic stages. Technical problems were excluded by transfecting a 3' replacement construct, which recombined correctly and transfectants did not show any phenotypic alterations. In order to prove that the trxR knockout was responsible for the lethal phenotype of the null mutants, a co-transfection with both the knockout construct and a construct containing the trxR coding region under the control of the calmodulin promoter was conducted. Despite the disruption of the trxR gene, parasites were viable. In a Southern blot analysis a complicated restriction pattern was obtained, but it was shown by pulse field gel electrophoresis and field inverse gel electrophoreses that only the trxR gene locus on chromosome 9 was targeted by the constructs. It was found that the co-transfected constructs form concatemeric structures prior to integration into the trxR gene locus, which is further supported by plasmid rescue followed by restriction analyses of the plasmids. Northern and Western blot analyses proved that the co-transfectants highly overexpress TrxR from the introduced gene. Our results demonstrate that TrxR is essential for the survival of the erythrocytic stages of P. falciparum.     

Structural analysis of genes participating in melibiose fermentation and isocitrate lyase production in Yersinia pestis strains of main and non main subspecies

Comparative analysis of nucleotide sequences of genes participating in melibiose fermentation and isocitrate lyase production was conducted in 90 natural Yersinia pestis strains of main and non main subspecies. It was ascertained that the lack of the ability to utilize disaccharide melibiose in strains of the main subspecies is caused by integration of the insertion sequence IS285 at 73 bp from the beginning of the structural gene melB that encodes the transport protein galactoside permease. In contrast, strains of non main subspecies (caucasica, altaica, and ulegeica) contain the intact gene melB and are capable of fermenting melibiose. Differences in the manifestation of the other differential trait, production of isocitrate lyase, are connected with the presence of mutation (insertion of two nucleotides +CC) in the regulatory gene iclR encoding repressor protein of the acetate operon, which is the reason for constitutive synthesis of this enzyme. Strains of non main subspecies do not contain mutations in gene iclR, and this correlates in these strains with their capacity for inducible synthesis of isocitrate lyase.     

Distances électrophorétiques entre les cinq espèces de babouins du genre Papio basées sur les mobilités des protéines et enzymes sériques

The baboons of the Papio genus have for a long time been divided into five distinct species, P. papio, P. anubis, P. cynocephalus, P. ursinus and P. hamadryas, with the latter being classed apart from the other four. This classification has been based on morphological, sociological and biogeographical grounds. Recently, however, the five forms have been considered as merely different sub-species of a polytypic species. This view has been supported by calculations of genetic distances between four of the species, the data being based on the electrophoretic polymorphism of the erythrocytic enzymes. The overall distances calculated are not greater than those obtained for the subspecies of other vertebrate genera. The present article shows that P. papio and P. anubis are indistinguishable in this respect, but that differences between the four species are observed by measurements of the electrophoretic mobilities of at least seven proteins and serum enzymes, namely: albumin, transferrine, esterases C₁ to C₄ and alkaline phosphatase. The apparent contradiction between these results may be explained in two ways. Firstly, the genetic distances correspond to a relatively low level of individual heterozygosity. The second, and principal reason, is that the proteins and serum enzymes present in the various tissues have evolved much more rapidly with time than the intracellular erythrocytic enzymes. Seven proteins can therefore be used to distinguish between the four species. The differential electrophoretic mobility data have been treated by the classical methods of multidimensional analysis to give a planar projection of the distances between the species. This shows P. hamadryas to be the most distinct form. The results obtaind agree with those derived from studies of the immunology of the transferris, the isoantigens, and from DNA hybridization experiments.