New World Monkeys and Color

The visual worlds of most primates are rich with potential color signals, and many representatives of the order have evolved the biological mechanisms that allow them to exploit these sources of information. Unlike the catarrhines, platyrrhines typically have sex-linked polymorphic color vision that provides individuals with any of several distinct types of color vision, including both trichromatic and dichromatic variants. In recent years, this polymorphism has been the target of an expanding range of research efforts. As a result, researchers now reasonably understand the proximate biology underlying the polymorphisms, and a number of ideas have emerged as to their evolution. Progress has also been made in illuminating how color vision capacities may be related to the particular visual tasks that New World monkeys face.     

结核分枝杆菌抗原检测研究进展

结核病仍然是一个严重的全球性公共卫生问题,有效控制结核病的障碍在于缺乏早期、准确的诊断方法。机体受到结核分枝杆菌感染后,体内首先出现的是结核分枝杆菌特异性抗原。因此,结核分枝杆菌抗原检测作为结核病早期诊断的方法可能具有很高的诊断价值。我们简要综述了结核分枝杆菌抗原检测的相关研究进展。     

Biomechanical scaling of mandibular dimensions in New World Monkeys

Previous studies show that folivorous Old World monkeys have shorter, deeper mandibles and shorter, wider condyles than frugivorous ones. These morphologies have been related to leaf mastication in colobines and ingestion of large, tough fruits in cercopithecines. This study examines New World monkeys in order to determine whether they exhibit similar adaptations to diet. New World monkeys have relatively long, transversely thin mandibles and somewhat deep mandibles and narrow condyles. Except for their deep mandibles, folivorous New World monkeys (i.e., Alouatta) do not exhibit the mandibular and condylar specializations typical of cercopithecid folivores. Reliance on comparatively nonfibrous foods plus alterations in masticatory muscle ratios among New World monkeys partially accounts for observed differences between folivorous New and Old World monkeys. In addition, adaptations for howling in Alouatta appear to have a significant effect on mandibular morphology. A biomechanical interpretation of craniofacial scaling patterns suggests that the mandibles of New World monkeys are subjected to lower condylar loads and considerably less twisting of the mandibular corpus than those of comparable Old World monkeys.     

Widespread Environmental Contamination with Mycobacterium tuberculosis Complex Revealed by a Molecular Detection Protocol

Environmental contamination with Mycobacterium tuberculosis complex (MTC) has been considered crucial for bovine tuberculosis persistence in multi-host-pathogen systems. However, MTC contamination has been difficult to detect due to methodological issues. In an attempt to overcome this limitation we developed an improved protocol for the detection of MTC DNA. MTC DNA concentration was estimated by the Most Probable Number (MPN) method. Making use of this protocol we showed that MTC contamination is widespread in different types of environmental samples from the Iberian Peninsula, which supports indirect transmission as a contributing mechanism for the maintenance of bovine tuberculosis in this multi-host-pathogen system. The proportion of MTC DNA positive samples was higher in the bovine tuberculosis-infected than in presumed negative area (0.32 and 0.18, respectively). Detection varied with the type of environmental sample and was more frequent in sediment from dams and less frequent in water also from dams (0.22 and 0.05, respectively). The proportion of MTC-positive samples was significantly higher in spring (p<0.001), but MTC DNA concentration per sample was higher in autumn and lower in summer. The average MTC DNA concentration in positive samples was 0.82 MPN/g (CI₉₅ 0.70–0.98 MPN/g). We were further able to amplify a DNA sequence specific of Mycobacterium bovis/caprae in 4 environmental samples from the bTB-infected area.     

Validation of qPCR Methods for the Detection of Mycobacterium in New World Animal Reservoirs

Zoonotic pathogens that cause leprosy (Mycobacterium leprae) and tuberculosis (Mycobacterium tuberculosis complex, MTBC) continue to impact modern human populations. Therefore, methods able to survey mycobacterial infection in potential animal hosts are necessary for proper evaluation of human exposure threats. Here we tested for mycobacterial-specific single- and multi-copy loci using qPCR. In a trial study in which armadillos were artificially infected with M. leprae, these techniques were specific and sensitive to pathogen detection, while more traditional ELISAs were only specific. These assays were then employed in a case study to detect M. leprae as well as MTBC in wild marmosets. All marmosets were negative for M. leprae DNA, but 14 were positive for the mycobacterial rpoB gene assay. Targeted capture and sequencing of rpoB and other MTBC genes validated the presence of mycobacterial DNA in these samples and revealed that qPCR is useful for identifying mycobacterial-infected animal hosts.     

Climatic Niche Evolution in New World Monkeys (Platyrrhini)

Despite considerable interest in recent years on species distribution modeling and phylogenetic niche conservatism, little is known about the way in which climatic niches change over evolutionary time. This knowledge is of major importance to understand the mechanisms underlying limits of species distributions, as well as to infer how different lineages might be affected by anthropogenic climate change. In this study we investigate the tempo and mode climatic niche evolution in New World monkeys (Platyrrhini). Climatic conditions found throughout the distribution of 140 primate species were investigated using a principal component analysis, which indicated that mean temperature (particularly during the winter) is the most important climatic correlate of platyrrhine geographical distributions, accounting for nearly half of the interspecific variation in climatic niches. The effects of precipitation were associated with the second principal component, particularly with respect to the dry season. When models of trait evolution were fit to scores on each of the principal component axes, significant phylogenetic signal was detected for PC1 scores, but not for PC2 scores. Interestingly, although all platyrrhine families occupied comparable regions of climatic space, some aotid species such as Aotus lemurinus, A. jorgehernandezi, and A. miconax show highly distinctive climatic niches associated with drier conditions (high PC2 scores). This shift might have been made possible by their nocturnal habits, which could serve as an exaptation that allow them to be less constrained by humidity during the night. These results underscore the usefulness of investigating explicitly the tempo and mode of climatic niche evolution and its role in determining species distributions.     

Genetic Diversity and Transmission Characteristics of Beijing Family Strains of Mycobacterium tuberculosis in Peru

Beijing family strains of Mycobacterium tuberculosis have attracted worldwide attention because of their wide geographical distribution and global emergence. Peru, which has a historical relationship with East Asia, is considered to be a hotspot for Beijing family strains in South America. We aimed to unveil the genetic diversity and transmission characteristics of the Beijing strains in Peru. A total of 200 Beijing family strains were identified from 2140 M. tuberculosis isolates obtained in Lima, Peru, between December 2008 and January 2010. Of them, 198 strains were classified into sublineages, on the basis of 10 sets of single nucleotide polymorphisms (SNPs). They were also subjected to variable number tandem-repeat (VNTR) typing using an international standard set of 15 loci (15-MIRU-VNTR) plus 9 additional loci optimized for Beijing strains. An additional 70 Beijing family strains, isolated between 1999 and 2006 in Lima, were also analyzed in order to make a longitudinal comparison. The Beijing family was the third largest spoligotyping clade in Peru. Its population structure, by SNP typing, was characterized by a high frequency of Sequence Type 10 (ST10), which belongs to a modern subfamily of Beijing strains (178/198, 89.9%). Twelve strains belonged to the ancient subfamily (ST3 [n = 3], ST25 [n = 1], ST19 [n = 8]). Overall, the polymorphic information content for each of the 24 loci values was low. The 24 loci VNTR showed a high clustering rate (80.3%) and a high recent transmission index (RTI_n−1 = 0.707). These strongly suggest the active and on-going transmission of Beijing family strains in the survey area. Notably, 1 VNTR genotype was found to account for 43.9% of the strains. Comparisons with data from East Asia suggested the genotype emerged as a uniquely endemic clone in Peru. A longitudinal comparison revealed the genotype was present in Lima by 1999.     

Genetic Diversity in Oxytocin Ligands and Receptors in New World Monkeys

Oxytocin (OXT) is an important neurohypophyseal hormone that influences wide spectrum of reproductive and social processes. Eutherian mammals possess a highly conserved sequence of OXT (Cys-Tyr-Ile-Gln-Asn-Cys-Pro-Leu-Gly). However, in this study, we sequenced the coding region for OXT in 22 species covering all New World monkeys (NWM) genera and clades, and characterize five OXT variants, including consensus mammalian Leu⁸-OXT, major variant Pro⁸-OXT, and three previously unreported variants: Ala⁸-OXT, Thr⁸-OXT, and Phe²-OXT. Pro⁸-OXT shows clear structural and physicochemical differences from Leu⁸-OXT. We report multiple predicted amino acid substitutions in the G protein-coupled OXT receptor (OXTR), especially in the critical N-terminus, which is crucial for OXT recognition and binding. Genera with same Pro⁸-OXT tend to cluster together on a phylogenetic tree based on OXTR sequence, and we demonstrate significant coevolution between OXT and OXTR. NWM species are characterized by high incidence of social monogamy, and we document an association between OXTR phylogeny and social monogamy. Our results demonstrate remarkable genetic diversity in the NWM OXT/OXTR system, which can provide a foundation for molecular, pharmacological, and behavioral studies of the role of OXT signaling in regulating complex social phenotypes.     

痰涂片标本用于结核分枝杆菌耐药性检测的研究

目的 研究抗酸染色结核分枝杆菌(简称结核杆菌)阳性痰涂片标本直接用于耐药性检测的方法。方法 对18株临床分离培养的结核杆菌用利福平进行药敏试验。分别提取菌株DNA和与之对应的痰涂片标本的菌体DNA,用聚合酶链反应(PcR)扩增ropB基因后进行固相杂交和核酸测序检测结核杆菌的耐药性。结果 18株结核杆菌中有12株对利福平耐药。经PCR扩增的ropB片段与探针杂交后,敏感菌株未发现rpoB基因的突变,自耐药菌株提取的DNA中rpoB突变体的检出率为100％(12／12),痰涂片提取DNA的检出率为91．7％(11／12)。所有耐药菌株DNA与痰涂片DNA核酸测序结果相吻合,都有rpoB基因核心区域碱基突变。结论 抗酸染色痰涂片阳性标本可直接用于检测结核杆菌利福平耐药基因rpoB突变体,是一种值得临床实验室推广使用的耐药菌诊断方法。     

Genetic Diversity of Mycobacterium tuberculosis in Peru and Exploration of Phylogenetic Associations with Drug Resistance

Background

There is limited available data on the strain diversity of M tuberculosis in Peru, though there may be interesting lessons to learn from a setting where multidrug resistant TB has emerged as a major problem despite an apparently well-functioning DOTS control programme.

Methods

Spoligotyping was undertaken on 794 strains of M tuberculosis collected between 1999 and 2005 from 553 community-based patients and 241 hospital-based HIV co-infected patients with pulmonary tuberculosis in Lima, Peru. Phylogenetic and epidemiologic analyses permitted identification of clusters and exploration of spoligotype associations with drug resistance.

Results

Mean patient age was 31.9 years, 63% were male and 30.4% were known to be HIV+. Rifampicin mono-resistance, isoniazid mono-resistance and multidrug resistance (MDR) were identified in 4.7%, 8.7% and 17.3% of strains respectively. Of 794 strains from 794 patients there were 149 different spoligotypes. Of these there were 27 strains (3.4%) with novel, unique orphan spoligotypes. 498 strains (62.7%) were clustered in the nine most common spoligotypes: 16.4% SIT 50 (clade H3), 12.3% SIT 53 (clade T1), 8.3% SIT 33 (LAM3), 7.4% SIT 42 (LAM9), 5.5% SIT 1 (Beijing), 3.9% SIT 47 (H1), 3.0% SIT 222 (clade unknown), 3.0% SIT1355 (LAM), and 2.8% SIT 92 (X3). Amongst HIV-negative community-based TB patients no associations were seen between drug resistance and specific spoligotypes; in contrast HIV-associated MDRTB, but not isoniazid or rifampicin mono-resistance, was associated with SIT42 and SIT53 strains.

Conclusion

Two spoligotypes were associated with MDR particularly amongst patients with HIV. The MDR-HIV association was significantly reduced after controlling for SIT42 and SIT53 status; residual confounding may explain the remaining apparent association. These data are suggestive of a prolonged, clonal, hospital-based outbreak of MDR disease amongst HIV patients but do not support a hypothesis of strain-specific propensity for the acquisition of resistance-conferring mutations.     

Monomorphic Region of the Serotonin Transporter Promoter Gene in New World Monkeys

Genetic variation in the human serotonin system has long been studied because of its functional consequences and links to various neuropsychiatric and behavior‐related disorders. Among non‐human primates, the common marmosets (Callithrix jacchus) and tufted capuchins monkeys (Cebus apella) are becoming increasingly used as models to study the effects of genes, environments, and their interaction on physiology and complex behavior. In order to investigate the independent functions of and potential interactions between serotonin‐related genes, anxiety and neuropsychiatric disorders, we analyzed the presence and variability of the serotonin transporter gene‐linked polymorphic region (5‐HTTLPR) in marmoset and capuchin monkeys. By PCR and using heterologous primers from the human sequence, we amplified and then sequenced the corresponding 5‐HTT region in marmosets and capuchins. The resulting data revealed the presence of a tandem repeat sequence similar to that described in humans, but unlike humans and other Old World primates, no variable length alleles were detected in these New World monkeys, suggesting that if serotonin transporter is involved in modulating behavior in these animals it does so through different molecular mechanisms. Am. J. Primatol. 74:1028‐1034, 2012. © 2012 Wiley Periodicals, Inc.     

Two New Rapid SNP-Typing Methods for Classifying Mycobacterium tuberculosis Complex into the Main Phylogenetic Lineages

There is increasing evidence that strain variation in Mycobacterium tuberculosis complex (MTBC) might influence the outcome of tuberculosis infection and disease. To assess genotype-phenotype associations, phylogenetically robust molecular markers and appropriate genotyping tools are required. Most current genotyping methods for MTBC are based on mobile or repetitive DNA elements. Because these elements are prone to convergent evolution, the corresponding genotyping techniques are suboptimal for phylogenetic studies and strain classification. By contrast, single nucleotide polymorphisms (SNP) are ideal markers for classifying MTBC into phylogenetic lineages, as they exhibit very low degrees of homoplasy. In this study, we developed two complementary SNP-based genotyping methods to classify strains into the six main human-associated lineages of MTBC, the "Beijing" sublineage, and the clade comprising Mycobacterium bovis and Mycobacterium caprae. Phylogenetically informative SNPs were obtained from 22 MTBC whole-genome sequences. The first assay, referred to as MOL-PCR, is a ligation-dependent PCR with signal detection by fluorescent microspheres and a Luminex flow cytometer, which simultaneously interrogates eight SNPs. The second assay is based on six individual TaqMan real-time PCR assays for singleplex SNP-typing. We compared MOL-PCR and TaqMan results in two panels of clinical MTBC isolates. Both methods agreed fully when assigning 36 well-characterized strains into the main phylogenetic lineages. The sensitivity in allele-calling was 98.6% and 98.8% for MOL-PCR and TaqMan, respectively. Typing of an additional panel of 78 unknown clinical isolates revealed 99.2% and 100% sensitivity in allele-calling, respectively, and 100% agreement in lineage assignment between both methods. While MOL-PCR and TaqMan are both highly sensitive and specific, MOL-PCR is ideal for classification of isolates with no previous information, whereas TaqMan is faster for confirmation. Furthermore, both methods are rapid, flexible and comparably inexpensive.     

聚合酶链式反应检测结核杆菌的研究

以人型结核杆菌基因组DNA为模板,合成二段引物各20个碱基进行聚合酶链式反应(PCR)。经琼脂糖凝胶电泳证实,获得一条245bp扩增带。PCR检测的敏感性染色体基因组DNA为1pg,菌悬液为13个活菌／ml。在特异性试验中,人型结核杆趋,牛型结核杆菌、BCG可见此扩增带。被试的其它14种扰酸菌以及变铅青链霉菌、大肠杆菌质粒Puc19、星状诺卡氏菌、红球菌均未见该扩增带。54例肺结核痰标本3种方法检查的阳性率分别为：萋尼氏抗酸染色16．7％,培养法14．8％,PCR 37．0％。前2种检查方法分别与PCR比较,经统计学处理均有显著性差异(P<0．01)。12例非结核性肺部疾患痰标本抗酸染色和PCR均为阴性。结果表明,PCR技术是快速、敏感、特异诊断结核病的方法。     

Mhc-DRB Genes and the Origin of New World Monkeys

The major histocompatibility complex (Mhc) is a family of loci characterized by its relatively rapid evolutionary turnover, large genetic distances between genes, and long persistence of allelic lineages effected by balancing selection. These features render the Mhc highly suitable for answering questions concerning speciation and adaptive radiation. The aim of the present study was to use Mhc-DRB genes to make inferences about the founding population of the Platyrrhini. Three segments, each approximately 300 base pairs in length, of the platyrrhine DRB genes were amplified by the polymerase chain reaction and sequenced. The segments were derived from intron 2, exon 3, and exon 6 of DRB genes from different species of New World monkeys. The results of the study have revealed that on a phylogenetic tree, all of the tested platyrrhine genes appear to form a single cluster, while all catarrhine DRB genes form a distinct cluster, although the bootstrap values fail to provide statistically significant support for the separation of these two clades. This observation suggests that the multiple platyrrhine genes originated from a single ancestral gene after the divergence of the Platyrrhini and Catarrhini and thus contradicts the results of an earlier study in which some exon 2 DRB sequences appeared to predate the split of the two primate groups. The inconsistency in the DRB gene phylogeny can be explained by postulating convergent evolution for the peptide-binding region of the DRB exon 2 sequences. The phylogeny of the platyrrhine DRB genes (except for exon 2) is relatively "shallow"; the distances between genes are relatively short (in comparison to the catarrhine DRB genes), and there is a tendency for sequences of individual species to cluster together. The phylogeny of the platyrrhine DRB genes is consistent with the postulate that a small population founded the group and that there is an ongoing adaptive radiation from small, relatively isolated founding populations.     

Simian Varicella in Old World Monkeys

Simian varicella virus (SVV) causes a natural erythematous disease in Old World monkeys and is responsible for simian varicella epizootics that occur sporadically in facilities housing nonhuman primates. This review summarizes the biology of SVV and simian varicella as a veterinary disease of nonhuman primates. SVV is closely related to varicella–zoster virus, the causative agent of human varicella and herpes zoster. Clinical signs of simian varicella include fever, vesicular skin rash, and hepatitis. Simian varicella may range from a mild infection to a severe and life-threatening disease, and epizootics may have high morbidity and mortality rates. SVV establishes a lifelong latent infection in neural ganglia of animals in which the primary disease resolves, and the virus may reactivate later in life to cause a secondary disease corresponding to herpes zoster. Prompt diagnosis is important for control and prevention of epizootics. Antiviral treatment for simian varicella may be effective if administered early in the course of infection.Abbreviations: FEAU, 1-(2′-deoxy-2′-flouro-β-D-arabinofuranosyl)-5-iodouracil, IE, immediate early, ORF, open reading frame, PBL, peripheral blood lymphocyte, SVV, simian varicella virus, VZV, varicella–zoster virusSimian varicella is a natural erythematous disease of Old World primates (Superfamily Cercopithecoidea, Subfamily Cercopithecinae), involving particularly patas (Erythrocebus patas), African green or vervet (Chlorocebus aethiops), and various species of macaque (Macaca spp.) monkeys. Epizootics of simian varicella occur sporadically in facilities housing nonhuman primates. These outbreaks are sometimes associated with high morbidity and mortality and the loss of valuable research animals. Simian varicella virus (SVV; Cercopithecine herpesvirus 9), a primate herpesvirus, is the etiologic agent of the disease. SVV is antigenically and genetically related to varicella–zoster virus (VZV; Human herpesvirus 3), the cause of human varicella (chickenpox) and herpes zoster (shingles). The clinical similarities between simian and human varicella and the relatedness of SVV and VZV, indicate that SVV infection of nonhuman primates is a useful model for study of varicella pathogenesis and development of antiviral therapies. A previous comprehensive review emphasized simian varicella as an experimental model for VZV infections.²² This review focuses on simian varicella as a veterinary disease of nonhuman primates. Simian varicella outbreaks and their epidemiology are considered, and the etiologic agent, clinical manifestations, pathogenesis, diagnosis, treatment, and control of the disease are discussed.     

Rapid Detection of Mycobacterium tuberculosis by Recombinase Polymerase Amplification

Improved access to effective tests for diagnosing tuberculosis (TB) has been designated a public health priority by the World Health Organisation. In high burden TB countries nucleic acid based TB tests have been restricted to centralised laboratories and specialised research settings. Requirements such as a constant electrical supply, air conditioning and skilled, computer literate operators prevent implementation of such tests in many settings. Isothermal DNA amplification technologies permit the use of simpler, less energy intensive detection platforms more suited to low resource settings that allow the accurate diagnosis of a disease within a short timeframe. Recombinase Polymerase Amplification (RPA) is a rapid, low temperature isothermal DNA amplification reaction. We report here RPA-based detection of Mycobacterium tuberculosis complex (MTC) DNA in <20 minutes at 39°C. Assays for two MTC specific targets were investigated, IS6110 and IS1081. When testing purified MTC genomic DNA, limits of detection of 6.25 fg (IS6110) and 20 fg (IS1081)were consistently achieved. When testing a convenience sample of pulmonary specimens from suspected TB patients, RPA demonstrated superior accuracy to indirect fluorescence microscopy. Compared to culture, sensitivities for the IS1081 RPA and microscopy were 91.4% (95%CI: 85, 97.9) and 86.1% (95%CI: 78.1, 94.1) respectively (n = 71). Specificities were 100% and 88.6% (95% CI: 80.8, 96.1) respectively. For the IS6110 RPA and microscopy sensitivities of 87.5% (95%CI: 81.7, 93.2) and 70.8% (95%CI: 62.9, 78.7) were obtained (n = 90). Specificities were 95.4 (95% CI: 92.3,98.1) and 88% (95% CI: 83.6, 92.4) respectively. The superior specificity of RPA for detecting tuberculosis was due to the reduced ability of fluorescence microscopy to distinguish Mtb complex from other acid fast bacteria. The rapid nature of the RPA assay and its low energy requirement compared to other amplification technologies suggest RPA-based TB assays could be of use for integration into a point-of-care test for use in resource constrained settings.     

Simple Detection of the IS6110 Sequence of Mycobacterium tuberculosis Complex in Sputum,Based on PCR with Graphene Oxide

Graphene oxide (GO) has proven to be a satisfactory DNA-sensor platform for applications in enzyme-free signal amplification, fluorescence-based amplification, and nanoparticle-based platforms because of its excellent electrical, thermal, and optical properties. In this study, we designed a novel platform for the fluorescence detection of biomolecules, using a fluorescent dye-labeled primer and GO. We applied this system for the detection of the IS6110 insertion sequence of the Mycobacterium tuberculosis complex (MTB) and evaluated its feasibility for use in molecular diagnostics. Fifty-four sputum specimens were collected at our institution from October 2010 to March 2012. To detect MTB in the samples, we performed PCR amplification of the IS6110 DNA sequence using FAM-labeled primers, after which the PCR amplicon was incubated with GO and the fluorescence was measured. The results were compared with those obtained by conventional real-time quantitative PCR (RQ-PCR). The fluorescence intensity observed increased in a concentration-dependent manner with the FAM-labeled IS6110 amplicon. The results of the PCR-GO system for detecting IS6110 DNA were in good agreement with those obtained with conventional RQ-PCR (kappa statistic = 0.925). The PCR-GO system detected MTB DNA in 23 of 25 RQ-PCR-positive sputum samples (92.0%; 95% CI, 75.0–98.0%), but not in 29 of 29 RQ-PCR-negative sputum samples (100%; 95% CI, 88.1–100.0%). These results indicate the utility of the PCR-GO system in molecular diagnostics.     

Identification of New Genomospecies in the Mycobacterium terrae Complex

Members of the Mycobacterium terrae complex are slow-growing, non-chromogenic acid-fast bacilli found in the natural environment and occasionally in clinical material. These genetically closely-related members are difficult to differentiate by conventional phenotypic and molecular tests. In this paper we describe the use of whole genome data for the identification of four strains genetically similar to Mycobacterium sp. JDM601, a newly identified member of the M. terrae complex. Phylogenetic information from the alignment of genome-wide orthologous genes and single nucleotide polymorphisms show consistent clustering of the four strains together with M. sp. JDM601 into a distinct clade separate from other rapid and slow growing mycobacterial species. More detailed inter-strain comparisons using average nucleotide identity, tetra-nucleotide frequencies and analysis of synteny indicate that our strains are closely related to but not of the same species as M. sp. JDM601. Besides the 16S rRNA signature described previously for the M. terrae complex, five more hypothetical proteins were found that are potentially useful for the rapid identification of mycobacterial species belonging to the M. terrae complex. This paper illustrates the versatile utilization of whole genome data for the delineation of new bacterial species and introduces four new genomospecies to add to current members in the M. terrae complex.