Mitochondrial DNA phylogeny of two morphologically enigmatic fishes in the subfamily Schizothoracinae (Teleostei : Cyprinidae) in the Qinghai-Tibetan Plateau

The complete mitochondrial DNA (mtDNA) cytochrome b gene (1140 bp) was sequenced in Herzenstein macrocephalus and Gymnocypris namensis and in 13 other species and sub-species (n = 22), representing four closely related genera in the subfamily Schizothoracinae. Conflicting taxonomies of H. macrocephalus and G. namensis have been proposed because of the character instability among individuals. Parsimony, maximum likelihood and Bayesian methods produced phylogenetic trees with the same topology and resolved several distinctive clades. Previous taxonomic treatments, which variously placed these two species of separate genera or as sub-species, are inconsistent with the mtDNA phylogeny. Both H. macrocephalus and G. namensis appear in a well-supported clade, which also includes nine species of Schizopygopsis, and hence should be transferred to the genus Schizopygopsis. Morphological changes are further illustrated, and their adaptive evolution in response to the local habitat shifts during the speciation process appears to be responsible for conflicting views on the systematics of these two species and hence the contrasting taxonomic treatments. These species are endemic to the Qinghai-Tibetan Plateau, a region with a history of geological activity and a rich diversity of habitats that may have result in the parallel and reversal evolution of some morphological characters used in their taxonomies. Our results further suggest that speciation and morphological evolution of fishes in this region may be more complex than those previously expected. (c) 2007 The Authors Journal compilation (c) 2007 The Fisheries Society of the British Isles.

Patterns of genetic variation in Swertia przewalskii, an endangered endemic species of the Qinghai-Tibet Plateau

Swertia przewalskii Pissjauk. (Gentianaceae) is a critically endangered and endemic plant of the Qinghai-Tibet Plateau in China. RAPD and ISSR analyses were carried out on a total of 63 individuals to assess the extent of genetic variation in the remaining three populations. Percentage of polymorphic bands was 94% (156 bands) for RAPD and 96% (222 bands) for ISSR. A pairwise distance measure calculated from the RAPD and ISSR data was used as input for analysis of molecular variance (AMOVA). AMOVA indicated that a high proportion of the total genetic variation (52% for RAPD and 56% for ISSR) was found among populations; pairwise Phi(ST) comparisons showed that the three populations examined were significantly different (p < 0.001). Significant genetic differentiation was found based on different measures (AMOVA and Hickory theta(B)) in S. przewalskii (0.52 on RAPD and 0.56 on ISSR; 0.46 on RAPD and 0.45 on ISSR). The differentiation of the populations corresponded to low average gene flow (0.28 based on RAPD and 0.31 based on ISSR), whereas genetic distance-based clustering and coalescent-based assignment analyses revealed significant genetic isolation among populations. Our results indicate that genetic diversity is independent of population size. We conclude that although sexual reproduction and gene flow between populations of S. przewalskii are very limited, they have preserved high levels of genetic diversity. The main factors responsible for the high level of difference among populations are the isolation and recent fragmentation under human disturbance.

Enrichment of stable carbon and nitrogen isotopes of plant populations and plateau pikas along altitudes

Stable carbon and nitrogen isotope ratios were measured in plant populations and plateau pikas to determine enrichment in stable isotopes of three alpine meadow ecosystems at different elevations in the Qinghai-Tibet Plateau. The results indicated that stable carbon isotope signatures of plant populations at the three locations showed significant variations, delta C-13 of plant populations showed an enrichment of 0.86 parts per thousand per 1000 in over the linear proportion of the altitudinal response, while stable nitrogen isotopes showed no apparent difference. The stable nitrogen isotopes of plateau pikas became significantly isotopically heavier along altitudinal gradients and showed an enrichment of 3.17 parts per thousand/km. Stable carbon isotopes showed no significance, however, and the enrichment of 0.448 parts per thousand/km. delta C-13 and delta N-15 in plateau pikas were not significantly correlated. There appeared to be segregation between the metabolism of stable carbon and nitrogen isotopes of plateau pikas. Variances in metabolic rate at lower water availability and temperatures are presumed to be the main cause of enrichment of stable nitrogen isotopes along altitudinal gradients. Attention should be paid to construct trophic positions and to trace food chain information based oil an isotopic enrichment model.

Diversities in hepatic HIF-1, IGF-I/IGFBP-1, LDH/ICD, and their mRNA expressions induced by CoCl2 in Qinghai-Tibetan plateau mammals and sea level mice

Ochotona curzoniae and Microtus oeconomus are the native mammals living on the Qinghai-TibetanPlateau of China. The molecular mechanisms of their acclimatization to the Plateau-hypoxia remain unclear. Expressions of hepatic hypoxia-inducible factor (HIF)-1 alpha, insulin-like growth factor-I (IGF-I)/IGF binding protein (BP)-1(IGFBP-1; including genes), and key metabolic enzymatic genes [lactate dehydrogenase (LDH)-A/isocitrate dehydrogenase (ICD)] are compared in Qinghai-Tibetan- Plateau mammals andsea- level mice after injection of CoCl2 (20, 40, or 60 mg/ kg) and normobaric hypoxia (16.0% O-2, 10.8% O-2, and 8.0% O-2) for 6 h, tested by histochemistry, Western blot analysis, ELISA, and RT-PCR. Major results are CoCl2 markedly increased 1) HIF-1 alpha only in mice, 2) hepatic and circulatory IGF-I in M. oeconomus, 3) hepatic IGFBP-1 in mice and O. curzoniae, and 4) LDH-A but reduced ICD mRNA in mice (CoCl2 20 mg/kg) but were unchanged in the Tibetan mammals. Normobaric hypoxia markedly 1) increased HIF-1 alpha and LDH-A mRNA in mice and M. oeconomus (8.0% O-2) not in O. curzoniae, and 2) reduced ICD mRNA in mice and M. oeconomus (8.0% O-2) not in O. curzoniae. Results suggest that 1) HIF-1 alpha responsiveness to hypoxia is distinct in lowland mice and plateau mammals, reflecting a diverse tolerance of the three species to hypoxia; 2) CoCl2 induces diversities in HIF-1, IGF-I/IGFBP-1 protein or genes in mice, M. oeconomus, and O. curzoniae. In contrast, HIF-1 mediates IGFBP-1 transcription only in mice and in M. oeconomus (subjected to severe hypoxia); 3) differences in IGF-I/IGFBP-1 expressions induced by CoCl2 reflect significant diversities in hormone regulation and cell protection from damage; and 4) activation of anaerobic glycolysis and reduction of Krebs cycle represents strategies of lowland-animals vs. the stable metabolic homeostasis of plateau- acclimatized mammals.

CO2 fluxes of alpine shrubland ecosystem on the north-eastern Tibetan plateau

We measured ecosystem CO2 fluxes for an alpine shrubland on the north-eastern Tibetan Plateau, Qinghai, China. The study is to understand (1) the seasonal variation of CO2 flux and (2) how environmental factors affect the seasonality of CO2 exchange in the alpine ecosystem. Daytime ecosystem respiration was extrapolated from the relationship between temperature and nighttime CO2 fluxes under high turbulent conditions.Seasonal patterns of gross ecosystem production, ecosystem respiration and net ecosystem CO2 exchange followed highly the seasonal change of aboveground biomass in the alpine shrubland. The net ecosystem CO2 exchange was mainly controlled by the variation of photosynthetic photon flux density, while the ecosystem respiration was closely correlated to the soil temperature at 5-cm depth. Integrated values of gross ecosystem production, ecosystem respiration and net ecosystem CO2 exchange for the period from November 1, 2002 to October 31 2003 were estimated to be 1418, 1155 and 222 g CO2 m(-2) yr(-1), respectively.

Blood-gas properties of plateau zokor (Myospalax baileyi)

Plateau zokor (Myospalax baileyi) is one of the blind subterranean mole rats that spend their life solely underground in scaled burrows. It is one of the special species of the Qinghai-Tibet plateau. In their burrows, oxygen is low and carbon dioxide is high and their contents fluctuate with the change of seasons, soil types, rain and depth of burrows. However, plateau zokors show successful adaptation to that extreme environment. In this study, their adapting mechanisms to the hypoxic hypercapnic environment were analyzed through the comparison of their blood-gas properties with that of pikas (Ochotona curzniae) and Sprague-Dawley rats. The results indicated that plateau zokors had higher red blood corpuscle counts (8.11 +/- 0.59 (10(12)/L)) and hemoglobin concentrations (147 +/- 9.85 g/L), but hematocrit (45.9 +/- 3.29%) and mean corpuscular volume (56.67 +/- 2.57 fL) were lower than the other rodents. Their arterial blood and venous blood pH were 7.46 +/- 0.07 and 7.27 +/- 0.07. Oxygen pressure in arterial blood of plateau zokors was about 1.5 times higher than that of pikas and rats, and it was 0.36 and 0.26 times in their venous blood. Partial pressure for carbon dioxide in arterial and venous blood of plateau zokors was 1.5-fold and 2.0-fold higher, respectively, than in rats and pikas. Oxygen saturation of plateau zokors was 5.7 and 9.3 times lower in venous blood than that of pikas and rats, respectively. As result, the difference of oxygen saturation in arterial blood to venous blood was 2- and 4.5-fold higher in plateau zokors as that of pikas and rats, respectively. In conclusion, plateau zokors had a high tolerance to pH changes in tissues, together with strong capabilities to obtain oxygen from their hypoxic-hypercapnic environment. (c) 2006 Published by Elsevier Inc.

Mitochondrial cytochrome b sequence variation and phylogenetics of the highly specialized schizothoracine fishes (Teleostei : Cyprinidae) in the Qinghai-Tibet Plateau

The complete 1140 bp mitochondial cytochrome b sequences were obtained from 39 individuals representing five species of all four genera of highly specialized schizothoracine fishes distributed in the Qinghai-Tibet plateau. Sequence variation of the cytochrome b gene was surveyed among the 39 individuals as well as three primitive schizothoracines and one outgroup. Phylogenetic analysis suggested that the group assignment based on 1140 bp of the cytochrome b sequence is obviously; different from previous assignments, and the highly specialized schizothoracine fishes (Schizopygopsis pylzovi, Gymnocypris przewalskii, G. eckloni, Chuanchia lablosa, and Platypharodon extremus) form a monophyletic group that is sister to the clade formed by the primitive schizothoracine fishes (Schizothorax prenanti, S. pseudaksaiensis, and S. argentatus). The haplotypes of Schizopygopsis pylzovi and G. przewalskii were paraphyletic based on cytochrome b data, which most likely reflected incomplete sorting of mitochondrial DNA lineages. The diploid chromosome numbers of Schizofhoracinae were considered in phylogenetic analysis and provided a clear pattern of relationships. Molecular dating estimated for highly specialized schizothoracine fishes suggested that the highly specialized schizothoracine fishes diverged in the late Miocene Pliocene to Pleistocene (4.5x10(4)-4.05x10(6) Years BP). The relationship between the cladogenesis of highly specialized schizothoracine fishes and geographical events of the Qinghai-Tibet plateau is discussed.

Diurnal, seasonal and annual variation in net ecosystem CO2 exchange of an alpine shrubland on Qinghai-Tibetan plateau

Thus far, grassland ecosystem research has mainly been focused on low-lying grassland areas, whereas research on high-altitude grassland areas, especially on the carbon budget of remote areas like the Qinghai-Tibetan plateau is insufficient. To address this issue, flux of CO2 were measured over an alpine shrubland ecosystem (37 degrees 36'N, 101 degrees 18'E; 325 above sea level [a. s. l.]) on the Qinghai-Tibetan Plateau, China, for 2 years (2003 and 2004) with the eddy covariance method. The vegetation is dominated by formation Potentilla fruticosa L. The soil is Mol-Cryic Cambisols. To interpret the biotic and abiotic factors that modulate CO2 flux over the course of a year we decomposed net ecosystem CO2 exchange (NEE) into its constituent components, and ecosystem respiration (R-eco). Results showed that seasonal trends of annual total biomass and NEE followed closely the change in leaf area index. Integrated NEE were -58.5 and -75.5 g C m(-2), respectively, for the 2003 and 2004 years. Carbon uptake was mainly attributed from June, July, August, and September of the growing season. In July, NEE reached seasonal peaks of similar magnitude (4-5 g C m(-2) day(-1)) each of the 2 years. Also, the integrated night-time NEE reached comparable peak values (1.5-2 g C m(-2) day(-1)) in the 2 years of study. Despite the large difference in time between carbon uptake and release (carbon uptake time < release time), the alpine shrubland was carbon sink. This is probably because the ecosystem respiration at our site was confined significantly by low temperature and small biomass and large day/night temperature difference and usually soil moisture was not limiting factor for carbon uptake. In general, R-eco was an exponential function of soil temperature, but with season-dependent values of Q(10). The temperature-dependent respiration model failed immediately after rain events, when large pulses of R-eco were observed. Thus, for this alpine shrubland in Qinghai-Tibetan plateau, the timing of rain events had more impact than the total amount of precipitation on ecosystem R-eco and NEE.

Seasonal thermogenesis and body mass regulation in plateau pikas (Ochotona curzoniae)

Changes in photoperiod, ambient temperature and food availability trigger seasonal acclimatization in physiology and behavior of many animals. In the present study, seasonal adjustments in body mass and in several physiological, hormonal, and biochemical markers were examined in wild-captured plateau pikas (Ochotona curzoniae) from the Qinghai-Tibetan plateau. Our results showed that plateau pikas maintained a relatively constant body mass throughout the year and showed no seasonal changes in body fat mass and circulating levels of serum leptin. However, nonshivering thermogenesis, cytochrome c oxidase activity, and mitochondrial uncoupling protein 1 (UCP1) contents in brown adipose tissues were significantly enhanced in winter. Further, serum leptin levels were positively correlated with body mass and body fat mass while negatively correlated with UCP1 contents. Together, these data suggest that plateau pikas mainly depend on increasing thermogenic capacities, rather than decreasing body mass, to cope with cold, and leptin may play a potential role in their thermogenesis and body mass regulation.

Seasonal regulations of energetics, serum concentrations of leptin, and uncoupling protein 1 content of brown adipose tissue in root voles (Microtus oeconomus) from the Qinghai-Tibetan plateau

Survival of small mammals in winter requires proper adjustments in physiology, behavior and morphology. The present study was designed to examine the changes in serum leptin concentration and the molecular basis of thermogenesis in seasonally acclimatized root voles (Microtus oeconomus) from the Qinghai-Tibetan plateau. In January root voles had lower body mass and body fat mass coupled with higher nonshivering thermogenesis (NST) capacity. Consistently, cytochrome c oxidase activity and mitochondrial uncoupling protein-1 (UCP1) protein contents in brown adipose tissues were higher in January as compared to that in July. Circulating level of serum leptin was significantly lower in winter and higher in July. Correlation analysis showed that serum leptin levels were positively related with body mass and body fat mass while negatively correlated with UCP1 protein contents. Together, these data provided further evidence for our previous findings that root voles from the Qinghai-Tibetan plateau mainly depend on higher NST coupled with lower body mass to enhance winter survival. Further, fat deposition was significantly mobilized in cold winter and leptin was potentially involved in the regulation of body mass and thermogenesis in root voles. Serum leptin might act as a starvation signal in winter and satiety signal in summer.

Hypoxia-inducible factor 1 alpha cDNA cloning and its mRNA and protein tissue specific expression in domestic yak (Bos grunniens) from Qinghai-Tibetan plateau

Adaptation to hypoxia is regulated by hypoxia-inducible factor 1 (HIF-1), a heterodimeric transcription factor consisting of an oxygen-regulated a-subunit and a constitutively expressed beta-subunit. How animals living on Qinghai-Tibetan plateau adapt to the extreme hypoxia environment is known indistinctly. In this study, the Qinghai yak which has been living at 3000-5000 m attitude for at least two millions of years was selected as the model of high hypoxia-tolerant adaptation species. The HIF-1 alpha ORFs (open reading frames) encoding for two isoforms of HIF-1 alpha have been cloned from the brain of the domestic yak. Its expression of HIF-1 alpha was analyzed at both mRNA and protein levels in various tissues. Both its HIF-1 alpha mRNA and protein are tissue specific expression. Its HIF-1 alpha protein's high expression in the brain, lung, and kidney showed us that HIF-1 alpha protein may play an important role in the adaptation to hypoxia environment. (c) 2006 Elsevier Inc. All rights reserved.

Effect of spatial variation on areal evapotranspiration simulation in Haibei, Tibet plateau, China

Quantification of areal evapotranspiration from remote sensing data requires the determination of surface energy balance components with support of field observations. Much attention should be given to spatial resolution sensitivity to the physics of surface heterogeneity. Using the Priestley-Taylor model, we generated evapotranspiration maps at several spatial resolutions for a heterogeneous area at Haibei, and validated the evapotranspiration maps with the flux tower data. The results suggested that the mean values for all evapotranspiration maps were quite similar but their standard deviations decreased with the coarsening of spatial resolution. When the resolution transcended about 480 m, the standard deviations drastically decreased, indicating a loss of spatial structure information of the original resolution evapotranspiration map. The absolute values of relative errors of the points for evapotranspiration maps showed a fluctuant trend as spatial resolution of input parameter data layers coarsening, and the absolute value of relative errors reached minimum when pixel size of map matched up to measuring scale of eddy covariance system. Finally, based on the analyses of the semi-variogram of the original resolution evapotranspiration map and the shapes of spatial autocorrelation indices of Moran and Geary for evapotranspiration maps at different resolutions, an appropriate resolution was suggested for the areal evapotranspiration simulation in this study area.

Temperature and biomass influences on interannual changes in CO2 exchange in an alpine meadow on the Qinghai-Tibetan Plateau

Three years of eddy covariance measurements were used to characterize the seasonal and interannual variability of the CO2 fluxes above an alpine meadow (3250 m a.s.l.) on the Qinghai-Tibetan Plateau, China. This alpine meadow was a weak sink for atmospheric CO2, with a net ecosystem production (NEP) of 78.5, 91.7, and 192.5 g C m(-2) yr(-1) in 2002, 2003, and 2004, respectively. The prominent, high NEP in 2004 resulted from the combination of high gross primary production (GPP) and low ecosystem respiration (R-e) during the growing season. The period of net absorption of CO2 in 2004, 179 days, was 10 days longer than that in 2002 and 5 days longer than that in 2003. Moreover, the date on which the mean air temperature first exceeded 5.0 degrees C was 10 days earlier in 2004 (DOY110) than in 2002 or 2003. This date agrees well with that on which the green aboveground biomass (Green AGB) started to increase. The relationship between light-use efficiency and Green AGB was similar among the three years. In 2002, however, earlier senescence possibly caused low autumn GPP, and thus the annual NEP, to be lower. The low summertime R-e in 2004 was apparently caused by lower soil temperatures and the relatively lower temperature dependence of R-e in comparison with the other years. These results suggest that (1) the Qinghai-Tibetan Plateau plays a potentially significant role in global carbon sequestration, because alpine meadow covers about one-third of this vast plateau, and (2) the annual NEP in the alpine meadow was comprehensively controlled by the temperature environment, including its effect on biomass growth.

Carbon dioxide dynamics and controls in a deep-water wetland on the Qinghai-Tibetan Plateau

To initially characterize the dynamics and environmental controls of CO2, ecosystem CO2 fluxes were measured for different vegetation zones in a deep-water wetland on the Qinghai-Tibetan Plateau during the growing season of 2002. Four zones of vegetation along a gradient from shallow to deep water were dominated, respectively by the emergent species Carex allivescens V. Krez., Scirpus distigmaticus L., Hippuris vulgaris L., and the submerged species Potamogeton pectinatus L. Gross primary production (GPP), ecosystem respiration (Re), and net ecosystem production (NEP) were markedly different among the vegetation zones, with lower Re and GPP in deeper water. NEP was highest in the Scirpus-dominated zone with moderate water depth, but lowest in the Potamogeton-zone that occupied approximately 75% of the total wetland area. Diurnal variation in CO2 flux was highly correlated with variation in light intensity and soil temperature. The relationship between CO2 flux and these environmental variables varied among the vegetation zones. Seasonal CO2 fluxes, including GPP, Re, and NEP, were strongly correlated with aboveground biomass, which was in turn determined by water depth. In the early growing season, temperature sensitivity (Q(10)) for Re varied from 6.0 to 8.9 depending on vegetation zone. Q(10) decreased in the late growing season. Estimated NEP for the whole deep-water wetland over the growing season was 24 g C m(-2). Our results suggest that water depth is the major environmental control of seasonal variation in CO2 flux, whereas photosynthetic photon flux density (PPFD) controls diurnal dynamics.

Leptin cDNA cloning and its mRNA expression in plateau pikas (Ochotona curzoniae) from different altitudes on Qinghai-Tibet Plateau

Leptin, an adipocyte-derived hormone, plays all important role in body energy homeostasis. Plateau pika (Ochotona curzoniae), an endemic and keystone species living only at 3000-5000 in above sea level oil Qinghai-Tibet Plateau, is a typically high hypoxia and low temperature tolerant mammal with high resting metabolic rate (RMR), non-shivering thermogenesis (NST), and high ratio of oxygen utilization to cope with harsh plateau environment. To explore the molecular mechanism of ecological acclimation in plateau pika, we first cloned pika leptin cDNA and compared its mRNA expression in different altitudes (3200 and 3900 in) using real-time RTPCR (Taqman probe) technology. The full-length pika leptin cDNA was 3015 with 504 bp open-reading frame encoding the precursor peptide of 167 amino acids including 21 residues of signal peptide. Pika leptin was 70-72% homologous to that of other species and was of similarly structural characteristics with other species. The pika-specific genetic diversity in leptin sequence occurred at twenty sites. With the increase in attitude, there were larger fat store and high level of ob gene expression in plateau pika. Our results indicated that leptin is sensitive to cold and hypoxia plateau environment and may play one of important roles in pika's ecological adaptation to harsh plateau environment. (c) 2006 Elsevier Inc. All rights reserved.