Behavioural responses by yaks in different physiological states (lactating, dry or replacement heifers), when grazing natural pasture in the spring (dry and germinating) season on the Qinghai-Tibetan plateau

Using heterogeneous vegetation in alpine grassland through grazing is a necessary component of deintensification of livestock systems and conservation of natural environments. However, better understanding of the dynamics of animal feeding behaviour would improve pasture and livestock grazing managements, particularly in the early part of the spring season when forage is scarce. The changes in behaviour may improve the use of poor pastures. Then, enhancing management practices may conserve pasture and improve animal productivity. Grazing behaviour over 24 In periods by yaks in different physiological states (lactating, dry and replacement heifers) was recorded in the early, dry and later, germinating period of the spring season. Under conditions of inadequate forage, the physiological state of yaks was not the primary factor affecting their grazing and ruminating behaviour. Forage and sward state affected yaks' grazing and ruminating behaviour to a greater extent. Generally, yaks had higher intake and spent more time grazing and ruminating during the later part of the spring season, following germination of forage, than during the earlier dry part of the season. However, the live weight of yaks was less during pasture germination than during the early dry part of the season because the herbage mass is low, and the yaks have to expend much energy to seek feed at this particular time. (c) 2007 Elsevier B.V. All rights reserved.

Genetic variation in the endangered Anisodus tanguticus (Solanaceae), an alpine perennial endemic to the Qinghai-Tibetan Plateau

We used random amplified polymorphic DNA markers (RAPDs) to assess genetic variation between- and within-populations of Anisodus tanguticus (Solanaceae), an endangered perennial endemic to the Qinghai-Tibetan Plateau with important medicinal value. We recorded a total of 92 amplified bands, using 12 RAPD primers, 76 of which (P = 82.61%) were polymorphic, and calculated values of H-t and H-sp of 0.3015 and 0.4459, respectively, suggesting a remarkably high rate of genetic variation at the species level. The average within-population diversity also appeared to be high, with P, H-e and H-pop values of 55.11%, 0.1948 and 0.2918, respectively. Analyses of molecular variance (AMOVA) showed that among- and between-population genetic variation accounted for 67.02% and 32.98% of the total genetic variation, respectively. In addition, Nei's coefficient of differentiation (G(ST)) was found to be high (0.35), confirming the relatively high level of genetic differentiation among the populations. These differentiation coefficients are higher than mean corresponding coefficients for outbreeding species, but lower than reported coefficients for some rare species from this region. The genetic structure of A. tanguticus has probably been shaped by its breeding attributes, biogeographic history and human impact due to collection for medicinal purposes. The observed genetic variations suggest that as many populations as possible should be considered in any planned in situ or ex situ conservation programs for this species.

Group size effects on foraging and vigilance in migratory Tibetan antelope

Large group sizes have been hypothesized to decrease predation risk and increase food competition. We investigated group size effects on vigilance and foraging behaviour during the migratory period in female Tibetan antelope Pantholops hodgsoni, in the Kekexili Nature Reserve of Qinghai Province, China. During June to August, adult female antelope and yearling females gather in large migratory groups and cross the Qinghai-Tibet highway to calving grounds within the Nature Reserve and return to Qumalai county after calving. Large groups of antelope aggregate in the migratory corridor where they compete for limited food resources and attract the attention of mammalian and avian predators and scavengers. We restricted our sampling to groups of less than 30 antelopes and thus limit our inference accordingly. Focal-animal sampling was used to record the behaviour of the free-ranging antelope except for those with lambs. Tibetan antelope spent more time foraging in larger groups but frequency of foraging bouts was not affected by group size. Conversely, the time spent vigilant and frequency of vigilance bouts decreased with increased group size. We suggest that these results are best explained by competition for food and risk of predation. (C) 2007 Elsevier B.V. All rights reserved.

Analysis of five pharmacologically active compounds from the Tibetan medicine Elsholtzia with micellar electrokinetic capillary chromatography

A high performance capillary electrophoresis method with diode array detector detection for the determination of five bioactive ingredients in Tibetan medicine Elsholtzia, namely quercetin, rutin, saussurenoside, kaempferol, and oleanolic acid, has been developed. The effects of several factors, such as the acidity, concentration of running buffer, separation voltage, temperature, and SDS concentration were investigated. The optimal conditions were 44 mmol/L boric acid running buffer (pH 8.5), 45 mmol/L SDS, 16 KV voltage, 20 degrees C, and 10.0% (V/V) of acetonitrile. Under the optimum conditions, five components could be separated with a good baseline resolution within 17 min. The calibration curves showed good linear relationship over the concentration range of 5 x 10(-4)similar to 0.1 mg/mL for quercetin, rutin, saussurenoside, kaempferol, and 1 x 10(-3) similar to 0.1 mg/mL for oleanolic acid. The average recoveries of the method and RSD were ( 99.2%, 3.2%) for quercetin, (102.1%, 2.1%) for rutin, (99.4%, 1.5%) for saussurenoside, (98.9%, 1.8%) for kaempferol, and (99.0%, 2.9%) for oleanolic acid, respectively. The detection limits (S/N = 3) were 1.1 x 10(-4) mg/mL for quercetin, 2.6 x 10(-4) mg/mL for rutin, 1.8 x 10(-4) mg/mL for saussurenoside, 2.9 x 10(-4) mg/mL for kaempferol, and 6.3 x 10(-4) mg/mL for oleanolic acid, respectively. The method was simple, rapid, and reproducible and could be applied for the determination of quercetin, rutin, saussurenoside, kaempferol, and oleanolic acid in Tibetan medicine Elsholtzia, and the assay results were satisfactory.

Mitochondrial and chloroplast phylogeography of Picea crassifolia Kom. (Pinaceae) in the Qinghai-Tibetan Plateau and adjacent highlands

The disjunct distribution of forests in the Qinghai-Tibetan Plateau (QTP) and adjacent Helan Shan and Daqing Shan highlands provides an excellent model to examine vegetation shifts, glacial refugia and gene flow of key species in this complex landscape region in response to past climatic oscillations and human disturbance. In this study, we examined maternally inherited mitochondrial DNA (nad1 intron b/c and nad5 intron 1) and paternally inherited chloroplast DNA (trnC-trnD) sequence variation within a dominant forest species, Picea crassifolia Kom. We recovered nine mitotypes and two chlorotypes in a survey of 442 individuals from 32 populations sampled throughout the species' range. Significant mitochondrial DNA population subdivision was detected (G(ST) = 0.512; N-ST = 0.679), suggesting low levels of recurrent gene flow through seeds among populations and significant phylogeographical structure (N-ST > GST, P < 0.05). Plateau haplotypes differed in sequence from those in the adjacent highlands, suggesting a long period of allopatric fragmentation between the species in the two regions and the presence of independent refugia in each region during Quaternary glaciations. On the QTP platform, all but one of the disjunct populations surveyed were fixed for the same mitotype, while most populations at the plateau edge contained more than one haplotype with the mitotype that was fixed in plateau platform populations always present at high frequency. This distribution pattern suggests that present-day disjunct populations on the QTP platform experienced a common recolonization history. The same phylogeographical pattern, however, was not detected for paternally inherited chloroplast DNA haplotypes. Two chlorotypes were distributed throughout the range of the species with little geographical population differentiation (G(ST) = N-ST = 0.093). This provides evidence for highly efficient pollen-mediated gene flow among isolated forest patches, both within and between the QTP and adjacent highland populations. A lack of isolation to pollen-mediated gene flow between forests on the QTP and adjacent highlands is surprising given that the Tengger Desert has been a geographical barrier between these two regions for approximately the last 1.8 million years.

Interannual fluctuations in floral longevity, pollinator visitation and pollination limitation of an alpine plant (Gentiana straminea Maxim., Gentianaceae) at two altitudes in the Qinghai-Tibetan Plateau

Prolonged floral longevity and bumblebees as dominate pollinators in alpine ecosystem have been suggested to overcome pollination limitation of alpine plants arising from the decrease of pollinator activity with increasing altitude. However, this conclusion has never been examined in the Qinghai-Tibetan Plateau (QTP), the highest and largest plateau in the world. In this study, we intended to test year-to-year correlations between floral longevity, visiting frequency and pollen limitation of this species between two populations (at 3200 m and 4000 m) of Gentiana straminea in this plateau. Pollinator exclusion elongated both male and female phases greatly at both sites, and durations of both male and female phases in natural condition varied greatly over three years. The visiting frequency of bumblebees varied greatly at the higher altitude, but seemed to be stable at the lower altitude. Seed production was pollination limited in both populations in most studied years. The floral durations, pollinator frequency and pollination limitation showed no significant and consistent variations with the increase of altitude. The previous hypothesis that the prolonged floral longevity of alpine plants can compensate for low levels of pollinator visitation therefore could not be confirmed, and our results further suggested that in the QTP platform, the altitude shows no consistent effect on the reproductive performance of this species, despite that the fluctuation of visit frequency intensified at the higher altitude.

Net primary productivity and spatial distribution of vegetation in an alpine wetland, Qinghai-Tibetan Plateau

To initially describe vegetation structure and spatial variation in plant biomass in a typical alpine wetland of the Qinghai-Tibetan Plateau, net primary productivity and vegetation in relationship to environmental factors were investigated. In 2002, the wetland remained flooded to an average water depth of 25 cm during the growing season, from July to mid-September. We mapped the floodline and vegetation distribution using GPS (global positioning system). Coverage of vegetation in the wetland was 100%, and the vegetation was zonally distributed along a water depth gradient, with three emergent plant zones (Hippuris vulgaris-dominated zone, Scirpus distigmaticus-dominated zone, and Carex allivescers-dominated zone) and one submerged plant zone (Potamogeton pectinatus-dominated zone). Both aboveground and belowground biomass varied temporally within and among the vegetation zones. Further, net primary productivity (NPP) as estimated by peak biomass also differed among the vegetation zones; aboveground NPP was highest in the Carex-dominated zone with shallowest water and lowest in the Potamogeton zone with deepest water. The area occupied by each zone was 73.5% for P. pectinatus, 2.6% for H. vulgaris, 20.5% for S. distigmaticus, and 3.4% for C. allivescers. Morphological features in relationship to gas-transport efficiency of the aerial part differed among the emergent plants. Of the three emergent plants, H. vulgaris, which dominated in the deeper water, showed greater morphological adaptability to deep water than the other two emergent plants.

Pollinator shift and reproductive performance of the qinghai-tibetan plateau endemic and endangered Swertia przewalskii (Gentianaceae)

Reproductive failure results in many plant species becoming endangered. However, little is known of how and to what extent pollinator shifts affect reproductive performance of endangered species as a result of the artificial introduction of alien insects. In this study we examined breeding systems, visitor species, visiting frequency and seed set coefficients of Swertia przewalskii in two years that had different dominant pollinator species (native vs. alien). Flowers of this species were protandrous and herkogamous and insects were needed for the production of seeds. The stigmatic receptivity of this species was shorter than for other gentians. No significant difference in seed set coefficient was found for hand-pollinated plants between the two years, indicating that pollinator shift only had a minor effect on this plant's breeding system. The commonest pollinators in 2002 were native bumblebees, alien honeybees and occasional solitary bees, however, only alien honeybees were observed in 2004. The flower visitation rate in both years was relatively high, although the total visit frequency decreased significantly in 2004. The control flowers without any treatment produced significantly fewer seed sets in 2004 than in 2002. In the past decade the seed production of this species may have partly decreased due to pollination by alien honeybees, however, we suggest that they might have acted as alternative pollinators ensuring seed production of S. przewalskii when native pollinators were unavailable. The main reason that this plant is endangered is probably the result of habitat destruction, but changes in land use, namely intensified agricultural practice and unfavorable animal husbandry have also contributed to its decline. We recommend that in-situ conservation, including the establishment of a protected area, is the best way to preserve this species effectively.

Grazing behavior of lactating and non-lactating yaks in the summer season of the Qinghai-Tibetan plateau

The effect of the physiological states of lactating vs. non-lactating (dry) on grazing behavior and herbage intake by yaks was examined in the summer season in the Qinghai alpine area under continuous stocking management. Intake rates were estimated over periods of 1 h by weighing the animals before and after grazing, retaining the feces and urine excreted, and applying a correction for insensible weight loss (the 1-h weight changes of yaks when non-eating before or after the intake rate measurement). It is hypothesized that the lactating yaks should eat more and spend more time eating than nonlactating yaks, because they expend more energy. In our experiment, there were no differences in the effect of physiological state (lactating vs. dry) of yaks observed on the rate of insensible weight loss, intake rate, grazing jaw movement rate, bites per grazing jaw movement, or bite mass. The dry yaks tended to eat more and spend more time eating than lactating yaks, but not significantly so. Compared with the dry yaks, the lactating yaks had a significantly lower bite rate and bites per bolus.

Genetic diversity and historical population structure of Schizopygopsis pylzovi (Teleostei : Cyprinidae) in the Qinghai-Tibetan Plateau

1. Complete sequences of 1140 base pair of the cytochrome b gene from 133 specimens were obtained from nine localities including the inflow drainage system, isolated lakes and outflow drainage system in Qinghai-Tibetan Plateau to assess genetic diversity and to infer population histories of the freshwater fish Schizopygopsis pylzovi.2. Nucleotide diversities (pi) were moderate (0.0024-0.0045) in populations from the outflow drainage system and Tuosuo Lake, but low (0.0018-0.0021) in populations from Qiadam Basin. It is probable that the low intra-population variability is related with the paleoenvironmental fluctuation in Qiadam Basin, suggesting that the populations from Qiadam Basin have experienced severe bottleneck events in history.3. Phylogenetic tree topologies indicate that the individuals from different populations did not form reciprocal monophyly, but the populations from the adjacent drainages cluster geographically. Most population pairwise F-ST tests were significant, with non-significant pairwise tests between Tuosu Lake and Tuosuo Lake in the north-west of the Qinghai-Tibetan Plateau. Analysis of molecular variance (AMOVA) indicates that the significant genetic variation was explained at the levels of catchments within and among, not among specific boundaries or inflow and outflow drainage systems.4. The nested clade phylogeographical analysis indicates that historical processes are very important in the observed geographical structuring of S. pylzovi, and the contemporary population structure and differentiation of S. pylzovi may be consistent with the historical tectonic events occurred in the course of uplifts of the Qinghai-Tibetan Plateau. Fluctuations of the ecogeographical environment and major hydrographic formation might have promoted contiguous range expansion of freshwater fish populations, whereas the geological barriers among drainages have resulted in the fragmentation of population and restricted the gene flow among populations.5. The significantly large negative F-s-value (-24.91, P < 0.01) of Fu's F-s-test and the unimodal mismatch distribution indicate that the species S. pylzovi underwent a sudden population expansion after the historical tectonic event of the Gonghe Movement.6. The results of this study indicate that each population from the Qinghai-Tibetan Plateau should be managed and conserved separately and that efforts should be directed towards preserving the genetic integrity of each group.

Separation and determination of the major active components in Tibetan folk medicinal species Swertia franchetiana by HPLC with DAD

A sensitive and specific reversed-phase high performance liquid chromatography (RP-HPLC) method with diode array detection (DAD) was established for the quantitative determination of the nine active components, namely, swertiamarin (SWM, 1), mangiferin (MA, 2), gentipicroside (GE, 3), sweroside (SWO, 4), isoorientin (IS, 5), swertisin (SWS, 6), swertianolin (SWN, 7), 7-O-[alpha-L-rhamnopyranosyl-1 -> 2)-beta-D-xylopyranosyl]-1,8-dihydroxy-3-methoxyxanthone (RX, 8), and bellidifolin (BE, 9) used as the external standard, in Tibetan folk medicinal species Swertia franchetiana. Based on the baseline chromatographic separation of most components from the methanolic extract of Swertia franchetiana on a reversed-phase Eclipse XDB-C8 column with water-acetonitrile-formic acid as mobile phase, the nine components were identified by comparison with standard samples and qualified by using the external standard method with DAD at 254 nm. The correlation coefficients of all the calibration curves were found to be higher than 0.9980. The relative standard deviations (RSDs) of the peak areas and retention times for the nine standards were less than 2.07% and 2.86%, respectively.

Mass spectrometric identification of multihydroxy phenolic compounds in Tibetan herbal medicines

A highly selective and accurate method based on derivatization with dansyl chloride coupled with liquid chromatography-mass spectrometry has been developed for identification of natural pharmacologically active phenolic compounds in extracts of Lomatogonium rotatum plants (Tibetan herbal medicine) obtained by solid-phase extraction. The number of hydroxyl groups on the dansylated phenols was estimated by LC-MS-MS analysis in positive-ion mode. Dansyl derivatization of the compounds introduced basic secondary nitrogen into the phenolic core structures and this was readily ionized when acidic HPLC mobile phases were used. MS fragmentation of the derivatives generated intense protonated molecular ions of m/z [MH](+) (phenol aglycones were transformed into the corresponding free phenols by cleavage of an aglycone bond). Collision-induced dissociation of the protonated molecule generated characteristic product ions of m/z 234 and 171 corresponding to the protonated 5-(dimethylamino)naphthalene sulfoxide and 5 -(dimethylamino) naphthalene moieties, respectively. Selected reaction monitoring based on the m/z [MH](+) to 234 and 171 transitions was highly specific for these phenolic compounds. Characteristic ions with m/z values of [MH - 234](+), [MH 2 x 234](+), and [MH - 3 x 234](+) were of great importance for estimation of the presence of multihydroxyl groups on the phenolic backbone.

Modeling gross primary production of alpine ecosystems in the Tibetan Plateau using MODIS images and climate data

The eddy covariance technique provides measurements of net ecosystem exchange (NEE) Of CO2 between the atmosphere and terrestrial ecosystems, which is widely used to estimate ecosystem respiration and gross primary production (GPP) at a number Of CO2 eddy flux tower sites. In this paper, canopy-level maximum light use efficiency, a key parameter in the satellite-based Vegetation Photosynthesis Model (VPM), was estimated by using the observed CO2 flux data and photosynthetically active radiation (PAR) data from eddy flux tower sites in an alpine swamp ecosystem, an alpine shrub ecosystem and an alpine meadow ecosystem in Qinghai-Tibetan Plateau, China. The VPM model uses two improved vegetation indices (Enhanced Vegetation Index (EVI), Land Surface Water Index (LSWI)) derived from the Moderate Resolution Imaging Spectral radiometer (MODIS) data and climate data at the flux tower sites, and estimated the seasonal dynamics of GPP of the three alpine grassland ecosystems in Qinghai-Tibetan Plateau. The seasonal dynamics of GPP predicted by the VPM model agreed well with estimated GPP from eddy flux towers. These results demonstrated the potential of the satellite-driven VPM model for scaling-up GPP of alpine grassland ecosystems, a key component for the study of the carbon cycle at regional and global scales. (c) 2006 Elsevier Inc. All rights reserved.

Experimental warming, not grazing, decreases rangeland quality on the Tibetan Plateau

We investigated experimental warming and simulated grazing ( clipping) effects on rangeland quality, as indicated by vegetation production and nutritive quality, in winter-grazed meadows and summer- grazed shrublands on the Tibetan Plateau, a rangeland system experiencing climatic and pastoral land use changes. Warming decreased total aboveground net primary productivity ( ANPP) by 40 g . m(-2) . yr(-1) at the meadow habitats and decreased palatable ANPP ( total ANPP minus non- palatable forb ANPP) by 10 g . m(-2) . yr(-1) at both habitats. The decreased production of the medicinal forb Gentiana straminea and the increased production of the non- palatable forb Stellera chamaejasme with warming also reduced rangeland quality. At the shrubland habitats, warming resulted in less digestible shrubs, whose foliage contains 25% digestible dry matter ( DDM), replacing more digestible graminoids, whose foliage contains 60% DDM. This shift from graminoids to shrubs not only results in lower- quality forage, but could also have important consequences for future domestic herd composition. Although warming extended the growing season in non- clipped plots, the reduced rangeland quality due to decreased vegetative production and nutritive quality will likely overwhelm the improved rangeland quality associated with an extended growing season.Grazing maintained or improved rangeland quality by increasing total ANPP by 20 - 40 g . m(-2) . yr(-1) with no effect on palatable ANPP. Grazing effects on forage nutritive quality, as measured by foliar nitrogen and carbon content and by shifts in plant group ANPP, resulted in improved forage quality. Grazing extended the growing season at both habitats, and it advanced the growing season at the meadows. Synergistic interactions between warming and grazing were present, such that grazing mediated the warming- induced declines in vegetation production and nutritive quality. Moreover, combined treatment effects were nonadditive, suggesting that we cannot predict the combined effect of global changes and human activities from single- factor studies.Our findings suggest that the rangelands on the Tibetan Plateau, and the pastoralists who depend on them, may be vulnerable to future climate changes. Grazing can mitigate the negative warming effects on rangeland quality. For example, grazing management may be an important tool to keep warming- induced shrub expansion in check. Moreover, flexible and opportunistic grazing management will be required in a warmer future.

Determination of 30 free fatty acids in two famous Tibetan medicines by HPLC with fluorescence detection and mass spectrometric identification

A simple and sensitive high-performance liquid chromatographic (HPLC) method with fluorescence detection and mass spectrometric identification has been developed for analysis of 30 long-chain and short-chain free Fatty acids (FFAs). The fatty acids were derivatized to their esters with 1-[2-(p-toluenesulfonate)ethyl]-2-phenylimidazole-[4,5-f]-9,10-phenanthrene (TSPP) in N,N-dimethylformamide (DMF) at 90 degrees C with anhydrous K2CO3 as catalyst. A mixture Of C-1-C-30 fatty acids was completely separated within 60 min by gradient elution on a reversed-phase C-8 column. Qualitative identification of the acids was performed by atmospheric-pressure chemical ionization mass spectrometry (APCI-MS) in positive-ion mode. The fluorescence excitation and emission wavelengths were 260 and 380 nm, respectively. Quantitative determination of the 30 acids in two Tibetan medicines Gentiana straminea and G. dahurica was performed. The results indicated that the medicines contained many FFAs. Linear correlation coefficients for the FFA derivatives were > 0.9991. Relative standard deviations (RSDs, n = 6) for the fatty acid derivatives were < 3%. Detection limits (at a signal-to-noise ratio of 3:1) were 3.1-38 fmol. When the fatty acid derivatives were determined in the two real samples results were satisfactory and the sensitivity and reproducibility of the method were good.