Resequencing and comparison of whole mitochondrial genome to gain insight into the evolutionary status of the Shennongjia golden snub‐nosed monkey (SNJ R. roxellana)

Shennongjia Rhinopithecus roxellana (SNJ R. roxellana) is the smallest geographical population of R. roxellana. The phylogenetic relationships among its genera and species and the biogeographic processes leading to their current distribution are largely unclear. To address these issues, we resequenced and obtained a new, complete mitochondrial genome of SNJ R. roxellana by next‐generation sequencing and standard Sanger sequencing. We analyzed the gene composition, constructed a phylogenetic tree, inferred the divergence ages based on complete mitochondrial genome sequences, and analyzed the genetic divergence of 13 functional mtDNA genes. The phylogenetic tree and divergence ages showed that R. avunculus (the Tonkin snub‐nosed monkey) was the first to diverge from the Rhinopithecus genus ca. 2.47 million years ago (Ma). Rhinopithecus bieti and Rhinopithecus strykeri formed sister groups, and the second divergence from the Rhinopithecus genus occurred ca. 1.90 Ma. R. roxellana and R. brelichi diverged from the Rhinopithecus genus third, ca. 1.57 Ma. SNJ R. roxellana was the last to diverge within R. roxellana species in 0.08 Ma, and the most recent common ancestor of R. roxellana is 0.10 Ma. The analyses on gene composition showed SNJ R. roxellana was the newest geographic population of R. roxellana. The work will help to develop a more accurate protection policy for SNJ R. roxellana and facilitate further research on selection and adaptation of R. roxellana.     

Population Structure and Ranging Patterns of <Emphasis Type="Italic">Rhinopithecus roxellana</Emphasis> in Zhouzhi National Nature Reserve,Shaanxi, China

We describe the population structure and ranging patterns of a troop of Sichuan snub-nosed monkeys (Rhinopithecus roxellana) based on a study conducted between November 2002 and November 2003 in Zhouzhi National Nature Reserve, Shaanxi Province, China. The troop comprised several 1-male units and an all-male unit. Opportunistic censuses revealed that there were ≥112 individuals in the troop. The adult sex ratio (male vs. female) was 1:3.7. The ratios of adults to immatures and infants to adult females were 1:0.7 and 1:2, respectively. Via a grid system, we estimated the home range of the troop to be 18.3 km², of which 7.4 km² was the core area. The subjects exhibited distinct seasonal ranging patterns. Their movement across the home range was extensive in spring and restricted in autumn. In addition, reuse of quadrats was highest in winter and lowest in spring in comparison with other seasons. The daily path length (DPL) varied from .75 to 5 km, with a mean of 2.1 km. Seasonal analysis showed that DPL is significantly shorter in winter than in spring or summer; however, there is no significant difference between the DPLs of spring and summer or those of spring and autumn. The monkeys occupied elevations 1500–2600 m above sea level; the annual mean of altitudinal range is 2137 m. Contrary to early studies that reported Rhinopithecus roxellana migrates to lower elevations in winter, we found no evidence supporting a seasonal altitudinal shift. Using the highest troop count and home range estimate, and considering the extent of range overlap between neighboring troops, we calculated the population density and biomass of Rhinopithecus roxellana to be 7.2 individuals/km² and 68.3 kg/km², respectively. The temporal and spatial distribution of food resources may be the most important determinant of ranging behavior in Rhinopithecus roxellana, though understanding the relationship between resource distribution and seasonal range use may require further investigation.     

Isolation and characterization of 11 microsatellite loci for the Sichuan snub-nosed monkey,Rhinopithecus roxellana

We report on the isolation and characterization of 11 microsatellite loci in the Sichuan snub-nosed monkey (Rhinopithecus roxellana) from AC-enriched genomic libraries. 20 microsatellite loci were screened from the libraries, of which 11 were polymorphic. The number of observed alleles among 32 samples of snub-nosed monkey ranged from three to nine. The observed and expected heterozygosity were 0.071–0.815 and 0.201–0.819, respectively. The high variability revealed in this study should be useful tools for further study on social structure and population genetics of snub-nosed monkey in the future.     

Identification of 13 Human Microsatellite Markers via Cross-species Amplification of Fecal Samples from Rhinopithecus bieti

Yunnan snub-nosed monkeys (Rhinopithecus bieti) are 1 of 3 snub-nosed monkey species endemic to China. Only ca. 1500 individuals remain in high-altitude forests 3000–4500 m above sea level on the Tibetan Plateau, making them the nonhuman primate living at the highest known elevation. It is one of the most endangered 25 primate species in the world. Proper knowledge of the population genetics and social system of Rhinopithecus bieti will contribute to more appropriate conservation management decisions. Cross-species amplification of human microsatellite loci has facilitated analysis of the population genetics and reproductive strategies of various primate species. We screened 72 human-derived markers to assess their utility in Yunnan snub-nosed monkeys. Thirteen of them produced reliable results and exhibited moderate levels of polymorphism.     

Sleeping cluster patterns and retiring behaviors during winter in a free-ranging band of the Sichuan snub-nosed monkey

Little information is available on the sleeping cluster pattern and retiring behavior of Sichuan snub-nosed monkeys (Rhinopithecus roxellana). Here, we provide observational data on a provisioned free-ranging band in the Qinling Mountains, central China. The results suggest that winter night activity of R. roxellana is a compromise between antipredator and thermoregulatory strategies and an adaptation to ecological conditions of their temperate habitat. Monkeys retired between 1804 and 1858 h in winter. In support of the antipredation hypothesis, all monkeys slept in trees at night, whereas 18.8% of individuals slept on the ground during the day. Also, the study band was more spatially cohesive at night than in daytime, with shorter distances between one-male units. Keeping warm is critical for survival in freezing temperatures. Monkeys often slept in the lower stratum of the tree canopy, avoiding the upper canopy where it is cold and windy. They formed larger sleeping clusters at night than in daytime. The most common types of night-sleeping clusters were adult females and juveniles, followed by adult females with other adult females. These accounted for 60.2% of the total records. The frequency of female–male clusters is two times greater, and that of adult male–juvenile clusters was four times less at night than during the day. The variations in composition of sleeping clusters suggest affiliative patterns at night-sleeping sites differ from those during the day.     

Isolation and characterization of microsatellite loci in <Emphasis Type="Italic">Pedicularis verticillata</Emphasis> L. using PCR-based isolation of microsatellite arrays (PIMA)

Pedicularis verticillata L. is a highly valuable herb for traditional Chinese medical treatment. In this report, 11 microsatellite loci from P. verticillata were isolated. The simple sequence repeat (SSR) markers were screened in 23 samples of wild populations of P. verticillata, and eight samples from its sister P. ikomai. The number of alleles ranged from 4 to 13, and value of expected (H _E) and observed (H ₀) heterozygosity was 0.62609–0.89662 and 0–0.95652, respectively. All loci were significantly deviated from Hardy–Weinberg expectations due to the heterozygote deficiency, indicating a dramatic loss of genetic polymorphisms in the restrictedly distributed species. The markers amplified well in the two species are useful for examining genetic diversity and population genetic structure, which, in turn, can provide information for establishing conservation strategy for these endangered species.     

Genetic population structure of the Fire-bellied toad <Emphasis Type="Italic">Bombina bombina</Emphasis> in an area of high population density: implications for conservation

In this study, we report the genetic population structure of the Fire-bellied toad Bombina bombina in Brandenburg (East Germany) in the context of conservation. We analysed 298 samples originating from 11 populations in Brandenburg using mitochondrial control region sequences and six polymorphic microsatellite loci. For comparison, we included one population each from Poland and Ukraine into our analysis. Within Brandenburg, we detected a moderate variability in the mitochondrial control region (19 different haplotypes) and at microsatellite loci (9–12 alleles per locus). These polymorphisms revealed a clear population structure among toads in Brandenburg, despite a relatively high overall population density and the moderate size of single populations (100–2000 individuals). The overall genetic population structure is consistent with a postglacial colonization from South East-Europe and a subsequent population expansion. Based on genetic connectivity, we infer Management Units (MUs) as targets for conservation. Our genetic survey identified MUs, within which human infrastructure is currently preventing any genetic exchange. We also detect an unintentional translocation from South East to North West Brandenburg, presumably in the course of fish stocking activities. Provided suitable conservation measures are taken, Brandenburg should continue to harbor large populations of this critically endangered species.     

Comparison of masticatory morphology between Rhinopithecus bieti and R. roxellana

The masticatory apparatus for two endemic species of golden monkey in China, Rhinopithecus bieti and Rhinopithecus roxellana, were compared with those of macaques, Macaca and leaf monkeys, Presbytis. Multivariate analyses demonstrated that the two golden monkey species are distinct. Interspecies allometric analyses revealed that golden monkeys differ in their masticatory apparatus from both macaques and leaf monkeys. The prominent symphysial fusion, corpus, and sagittal condylar dimension of R. roxellana may produce efficient biting force on the incisal and posterior canine teeth, with the heavy reaction force born on the temporomandibular joint. However, the well-developed bizygamatic width and mandibular height in R. bieti suggest that posterior canine function is similarly prominent in R. roxellana, while incisal function is not. © 1995 Wiley-Liss, Inc.     

Phylogeny of Snub-Nosed Monkeys Inferred from Mitochondrial DNA,Cytochrome B,and 12S rRNA Sequences

Snub-nosed monkeys (Rhinopithecus spp.) are confined to isolated mountainous regions in China and North Vietnam. Their systematic classification and phylogenetic relationship has been controversial. The structures of mitochondrial DNA cytochrome b and 12S rRNA show that the 4 species of Rhinopithecus are quite different from other colobines. It is reasonable to regard them as an independent genus, as determined by external features, morphometric characters and behavior. However, whether or not there should be a subdivision between the Vietnamese and Chinese species at the subgeneric level remains to be clarified; more evidence from a large range of Asian colobine species is needed. The Guizhou species, Rhinopithecus brelichi, is a valid species, which is more closely related to Pygathrix than the other species ( R. roxellana, R. bieti and R. avunculus) are. Results also indicate that 3 species—Rhinopithecus roxellana, R. bieti and R. avunculus—might have diverged from R. brelichi, but the phylogenetic relationship of R. avunculus is not clear.     

Small effective population sizes of two remnant ocelot populations (<Emphasis Type="Italic">Leopardus pardalis albescens</Emphasis>) in the United States

Threatened populations are vulnerable to the effects of genetic drift and inbreeding, particularly when gene flow is low and the effective population size is small. Estimates of effective population size (N _e ) provide important information on the status of endangered populations that have experienced severe fragmentation and serve as indicators of genetic viability. Genetic data from microsatellite loci were used to estimate N _e for the 2 remaining populations of the endangered ocelot (Leopardus pardalis albescens) occurring in the United States. Several methods were used to calculate N _e , resulting in estimates ranging from N _e  = 8.0 (95% CI: 3.2–23.1) to 13.9 (95% CI: 7.7–25.1) for the population located at the Laguna Atascosa Wildlife Refuge in Cameron County, Texas. The ocelot population in Willacy County, Texas, had N _e estimates of 2.9 (95% CI: 1.7–5.6) and 3.1 (95% CI: 1.9–13.5), respectively. Estimates of N _e in both populations were below the critical value recommended for short-term viability.     

MTHFR 677C>T and 1298A>C polymorphisms and male infertility risk: a meta-analysis

Epidemiological studies have evaluated the association between MTHFR 677C>T and 1298A>C polymorphisms and risk of male infertility. However, the results from the published studies on the association between these two MTHFR polymorphisms and male infertility risk are conflicting. To derive a more precise estimation of association between the MTHFR polymorphisms and risk of male infertility, we performed a meta-analysis. A comprehensive search was conducted to identify all case–control studies of MTHFR polymorphisms and male infertility risk. We used odds ratios (ORs) with 95% confidence intervals (CIs) to assess the strength of the association. Overall, we found that both 677C>T and 1298A>C polymorphisms were not significantly associated with male infertility risk. However, in stratified analysis by ethnicity, we found that the 677C>T polymorphism was significantly associated with the risk of male infertility in Asian population (TT vs. CC: OR = 1.57, 95% CI: 1.05–2.37, P = 0.03; TT vs. TC + CC: OR = 1.40, 95% CI: 1.05–1.86, P = 0.02; TT + TC vs. CC: OR = 1.34, 95% CI: 1.01–1.77, P = 0.04). Although some modest bias could not be eliminated, this meta-analysis suggested that the MTHFR 677T allele might be a low-penetrant risk factor for male infertility, especially in Asian population.     

Reproductive parameters in Guizhou snub‐nosed monkeys (Rhinopithecus brelichi)

In this study, we present data on reproductive parameters and birth seasonality of Guizhou snub‐nosed monkeys (Rhinopithecus brelichi). Our analyses are based on data from a small captive population collected over 15 years and on 5 years of observations of free‐ranging snub‐nosed monkeys. Captive females (n=4) mature at an age of 70.8±6.7 months and reproduce for the first time at 103.4±7.5 months. The mean interbirth interval was 38.2±4.4 months if the infant survived more than 6 months, which is longer than that in R. roxellana and R. bieti. In the wild and in captivity, births are very seasonal and occur only in a period from the end of March to the end of April. Our data suggest that population growth in Guizhou snub‐nosed monkeys is slow compared with the other two Chinese snub‐nosed monkey species. The risk of extinction is therefore particularly high in this species, given the small overall population size and slow population recovery potential. Am. J. Primatol. 71:266–270, 2009. © 2008 Wiley‐Liss, Inc.     

The men of Nelson's navy: a comparative stable isotope dietary study of late 18th century and early 19th century servicemen from Royal Naval Hospital burial grounds at Plymouth and Gosport, England

The Guizhou snub‐nosed monkey (Rhinopithecus brelichi) is a primate species endemic to the Wuling Mountains in southern China. With a maximum of 800 wild animals, the species is endangered and one of the rarest Chinese primates. To assess the genetic diversity within R. brelichi and to analyze its genetic population structure, we collected fecal samples from the wild R. brelichi population and sequenced the hypervariable region I of the mitochondrial control region from 141 individuals. We compared our data with those from the two other Chinese snub‐nosed species (R. roxellana, R. bieti) and reconstructed their phylogenetic relationships and divergence times. With only five haplotypes and a maximum of 25 polymorphic sites, R. brelichi shows the lowest genetic diversity in terms of haplotype diversity (h), nucleotide diversity (π), and average number of pairwise nucleotide differences (Π). The most recent common ancestor of R. brelichi lived ～0.36 million years ago (Ma), thus more recently than those of R. roxellana (～0.91 Ma) and R. bieti (～1.33 Ma). Phylogenetic analysis and analysis of molecular variance revealed a clear and significant differentiation among the three Chinese snub‐nosed monkey species. Population genetic analyses (Tajima's D, Fu's F_s, and mismatch distribution) suggest a stable population size for R. brelichi. For the other two species, results point in the same direction, but population substructure possibly introduces some ambiguity. Because of the lower genetic variation, the smaller population size and the more restricted distribution, R. brelichi might be more vulnerable to environmental changes or climate oscillations than the other two Chinese snub‐nosed monkey species. Am J Phys Anthropol, 2012. © 2011 Wiley Periodicals, Inc.     

川金丝猴秦岭与岷山社群社会结构与空间分布差异性比较

对不同地区川金丝猴社会结构和空间分布的比较有助于了解其种群与环境的相互应答关系。在秦岭山脉和岷山山脉的不同地区选取3个观察群,对比它们之间社群结构和空间分布的差异。研究结果表明,尽管同属于仰鼻猴属川金丝猴种,由于栖息地环境的不同,导致社群结构和空间分布既存在相同点又存在一定的差异。对于社会结构,扎如沟群,大坪峪群与玉皇庙群表现出同样的最高进化水平的重层社会体系;但是构成这一社会体系的基本单位一雄多雌（One-male unit,OMU）,其成员整体数量上没有差异,但是OMU内雌雄比例不仅在3个观察群间存在差异,而且与同属其他物种相比也存在差异;对于空间分布,由于食物竞争压力和天敌捕食压力在不同的栖息地内影响不同,导致3个观察群空间分布呈梯度变化关系。研究结果不仅为川金丝猴种群动态,种群与环境的相互应答关系以及种群进化提供基础资料,而且希望能藉此推进我国相关领域研究的开展,为灵长类社会与行为生态学研究者启发思路,并为保护这些濒危物种提供科学依据。     

Historical geographic dispersal of the golden snub-nosed monkey (Rhinopithecus roxellana) and the influence of climatic oscillations

Current understanding of historic climate oscillations that have occurred over the past few million years has modified scientific views on evolution. Major climatic events have caused local and global extinction of plants and animals and have impacted the spatial distribution of many species. The endangered golden snub‐nosed monkey (Rhinopithecus roxellana) currently inhabits three isolated regions of China: the Sichuan and Gansu provinces (SG), the Qinling Mountains in Shaanxi province (QL), and the Shennongjia Forestry District in Hubei province (SNJ). However, considerable uncertainty still exists about their historical dispersal routes under the influence of environment change. To date, two dispersal routes have been proposed: (1) the QL and SNJ populations originated from the SG population; and (2) the SG population recolonized from the QL and SNJ populations. We used the mitochondrial DNA complete control region to perform statistical assessments of the relative probability of alternative migration scenarios and the role of environmental change on the geographic dispersal of Rhinopithecus roxellana. Thirty haplotypes were identified from the three geographic regions and a high degree of genetic structure was observed. The most recent common ancestor among the mitochondrial DNA haplotypes was estimated to live around 0.47–1.88 million years ago and five notable haplotype clusters were found. Phylogenetic analysis and historical gene flow estimates suggested that the QL and SNJ populations originated from the SG population, with at least two dispersal events from the SG population occurring during the Pleistocene (1.17±0.70 and 0.53±0.30 Ma). Composite dispersal history of the golden snub‐nosed monkey can be explained by both environmental change inducing global climate change and the influence of the Tibetan Plateau uplift. Such range shifts involved considerable demographic changes, as revealed in the dramatic decreases in population size during the last 25,000 years.     

Report on the distribution,population, and ecology of the yunnan snub-nosed monkey (Rhinopithecus bieti)

The Yunnan snub-nosed monkey (Rhinopithecus bieti), an endangered species in China, has received more protection in theory than in practice. Therefore it is on the very verge of extinction. The population of the species was estimated less than 2,000 individuals spread in 19 distinct groups. It was confirmed that the monkey was confined to the Yunling Mountain System, the area between the Yangtze River (Changjiang, aka Jinshajiang) to the east and the Mekong River (Lancangjiang) to the west. We further concluded that a lowland belt to the east, about 100 km long and 20 – 30 km wide was not suitable habitat for the monkeys, and appeared to serve as the natural ecogeologic barrier for the species. Our results indicated that the southern limit of the distribution was at Longma (26°14′N), and that the northern limit of the distribution was at Xiaochangdu (29°20′N). The distribution area of the species was substantially smaller than previously estimated. There were substantial ecological differences between the southern and northern parts of the species range. The monkey was found only in fir-larch forest.     

Climate change and recent genetic flux in populations of <Emphasis Type="Italic">Drosophila robusta</Emphasis>

Background  

Studied since the early 1940's, chromosomal polymorphisms in the deciduous woods species Drosophila robusta have been characterized by well-defined latitudinal, longitudinal, and elevational clines, but – until at least ten years ago – stable, local population frequencies. Recent biogeographical analyses indicate that D. robusta invaded North America from southeast Asia and has persisted in eastern temperate forests for at least 20–25 my without speciating. The abundant chromosome polymorphisms found across the range of D. robusta are thus likely to be relatively ancient, having accumulated over many well known climatic cycles in North America. Sufficient long-term data are now available such that we can now gauge the rate of these evolutionary changes in natural populations due to environmental change.     

<Emphasis Type="Italic">MUTYH</Emphasis> Tyr165Cys, <Emphasis Type="Italic">OGG1</Emphasis> Ser326Cys and <Emphasis Type="Italic">XPD</Emphasis> Lys751Gln polymorphisms and head neck cancer susceptibility: a case control study

In the present study we investigated the association between three polymorphisms of the MUTYH (Tyr165Cys, rs34612342), the OGG1 (Ser326Cys, rs1052133) and the XPD (Lys751Gln, rs13181) genes with head and neck cancer risk. Genotypes were determined in DNA from peripheral blood lymphocytes of 265 patients with head and neck squamous cell carcinoma (HNSCC) as well as 280 cancer-free controls by PCR-restriction fragment lenght polymorphisms. We found an association between HNSCC and the Ser326Cys (OR 1.69; 95% CI 1.19–2.45) as well as Cys326Cys (OR 4.56; 95% CI 2.07–10.05) variants of the OGG1 gene. The gene–gene interaction between MUTYH and OGG1 as well as OGG1 and XPD polymorphic variants may contribute to higher prevalence of HNSCC. We also found an association between Ser326Cys and Cys326Cys variants of OGG1 gene and smoking status in HNSCC patients (OR 1.97; 95% CI 1.25–3.11), (OR 3.54; 95% CI 1.39–9.04), respectively. Moreover, we also observed a protective association between Tyr165Cys variant of the MUTYH gene and non-smoking status in HNSCC (OR 0.34; 95% CI 0.17–0.66). We also found a link between gene–gene interaction (MUTYH and OGG1 or OGG1 and XPD) and smoking (ORs 2.17–4.20 and 2.18–5.23) or non-smoking status (ORs 0.11 and 7.61) in HNSCC patients, respectively. In conclusion our data showed that the Ser326Cys polymorphism of the OGG1 gene may modify the risk of HNSCC associated with smoking. Finally we suggested that this polymorphism might be used as predictive factor for head and neck cancer in Polish population.     

The association of polymorphisms in DNA base excision repair genes <Emphasis Type="Italic">XRCC1</Emphasis>, <Emphasis Type="Italic">OGG1</Emphasis> and <Emphasis Type="Italic">MUTYH</Emphasis> with the risk of childhood acute lymphoblastic leukemia

The aim of this study was to evaluate the association of polymorphisms in genes encoding three key proteins of DNA base excision repair (BER): the OGG1 Ser326Cys, the MUTYH Tyr165Cys and the XRCC1 Arg399Gln with the risk of childhood acute lymphoblastic leukemia (ALL). Our study included 97 children patients with ALL (mean age 5.4 ± 2.5) and 131 healthy children (mean age 6.2 ± 2.8) used as controls. Genetic polymorphisms in BER pathway genes were examined using PCR and restriction fragment length polymorphism (RFLP). We have demonstrated that the OGG1 Cys/Cys genotype increases the risk of ALL (OR 5.36) whereas the Ser/Ser genotype variant strongly reduces the risk of this cancer among Polish children (OR 0.45). Although we did not observe the differences in single nucleotide polymorphisms (SNPs) in MUTYH and XRCC1 genes between control group and children with ALL, we have shown that the combined genotypes of examined genes can modulate the risk of childhood ALL in Polish population. We found that the combined genotype Arg/Gln–Cys/Cys of XRCC1/OGG1 (OR 3.83) as well as the Cys/Cys–Tyr/Tyr of OGG1/MUTYH (OR 6.75) increases the risk of ALL. In contrast, the combined genotype Arg/Arg–Ser/Ser of XRCC1/OGG1 (OR 0.40) as well as the Ser/Ser–Tyr/Tyr of OGG1/MUTYH (OR 0.43) played a protective role against this malignant disease. In conclusion, we suggest that polymorphisms of BER genes may be used as an important predictive factor for acute lymphoblastic leukemia in children.