作为重要的陆地碳（C）库，脆弱敏感的西藏高寒草原在全球变化和过度放牧的影响下沙化问题日益突出。草地沙化可能会加剧地上植物群落和地下微生物群落的养分限制状态，进而影响高寒草原土壤C储量。土壤氮（N）和磷（P）作为植物生长和微生物代谢的重要养分来源，然而，目前对沙化过程中植物和土壤微生物群落受N和P限制情况及其作用机理仍不清楚。本研究利用生态酶化学计量的方法评估了高寒草原五个不同沙化阶段（包含未退化、轻度、中度、重度和极度沙化）下植物和土壤微生物的养分限制状态。研究结果表明:（1）土壤微生物代谢主要受C和P限制，随沙化程度的加深，植物N限制和微生物C限制有所加剧，微生物P限制得到缓解；（2）植物-土壤-微生物相互作用对微生物C和P限制有显著的影响, 解释度分别为72%和61%。具体而言，沙化通过调节土壤pH、土壤养分和植物N限制来影响微生物代谢限制；（3）微生物C限制进一步降低了微生物的C利用效率（CUE），从而不利于沙化土壤中有机C的保留。本研究揭示了在植物-微生物相互作用下的微生物代谢限制是影响土壤微生物CUE的关键驱动因素，并为推进微生物调节养分循环和C固存提供了新的见解。

Agricultural application studies, including field experiments and acute toxicity tests, were conducted for lipopeptides secreted by marine-derived Bacillus marinus B-9987.  Benefiting from commercially available scaled-up lipopeptide purification, the sample of impurities (isolated from target lipopeptides), raw extracted sample (purity: 9.08%), partially purified sample (purity: 20.86%), and highly purified sample (purity: 87.51%) were prepared from B. marinus B-9987 fermentation broth, and used in lab-scale antagonism tests, field experiments, swarming motility tests, and acute toxicity tests.  Operations and conditions in field experiments were consistent with the Pesticide-Guidelines for the Field Efficacy Trials (GB/T 17980.28-2000), and acute toxicity tests were executed according to Toxicological Test Methods of Pesticides for Registration (GB 15670-1995).  In agar diffusion tests in vitro and pot tests in vivo, all lipopeptide samples with different purities significantly inhibited Botrytis cinerea; meanwhile the sample of impurities isolated from target lipopeptides were not effective against B. cinerea.  Results of lab-scale tests showed that the target lipopeptides were effective substances against B. cinerea.  Thus, partially purified and raw extracted samples were used in field experiments instead of the highly purified sample for cost saving.  In the field experiments against rose grey mold, biological control efficacy of 500 mg L^–1 lipopeptides reached 67.53%, slightly lower than 74.05% reached by the agrochemical pyrimethanil.  However, pyrimethanil severely suppressed B. marinus B-9987, whereas the lipopeptides promoted swarming motility and biocontrol efficacy of Bacillus biomass.  Lipopeptides at 87.51% purity were tested for systemic acute toxicity and confirmed as low-toxicity substances.  In conclusion, low-toxicity lipopeptides were potential alternatives to agrochemicals, and they also performed good promotion when combined with homologous biological control microorganism.  There were 2 breakthroughs in this research: (1) marine-derived bacterial lipopeptides inhibited grey mold caused by B. cinerea in field experiments; and (2) purified bacterial lipopeptides (sample purity: >87.51%) were determined to be low-toxicity substances by systemic acute toxicity tests, satisfying the strict requirement of pesticide registration in China (required purity: >85%).  This study provides support for using extracellular Bacillus-derived lipopeptides commercially similar to Bacillus-based biological control agents.  

    The transcription activator-like effector nuclease (TALEN) technique combined with the somatic cell nuclear transfer (SCNT) method has been successfully applied for creating genetically modified pigs. However, methods for isolating cells with biallelic indels requires further improvement because of the relatively low enrichment efficiency of mutated somatic cells. Moreover, little is known regarding the off-target effects of the TALEN system and the heredity of TALEN-modified pigs. In this study, an efficient method to increase the enrichment efficiency of TALEN-mediated biallelic knockout (KO) cells was established, and corresponding genetically modified pigs with the expected genotype were generated whose off-target effect, fertility and heredity characteristics were aslo evaluated. Two TALEN pairs were constructed to target the porcine α-1,3-galactosyltransferase (GGTA1) gene locus. TALEN mRNA was transfected into the ear ?broblasts followed by the enrichment of α-Gal null cells of minipigs using isolectin B4 (IB4) lectin and magnetic beads. A total of 115 cell colonies were formed and validated to be GGTA1 KO cells by sequencing and 10 biallelic KO cell colonies were used as nuclear donors for SCNT. Thirty GGTA1 biallelic KO piglets were successfully delivered and grew normally. Seventeen potential off-target sites were investigated, and no off-target events were detected in the live piglets. To determine the fertility and heredity characteristics of TALEN-modified pigs, 10 mature founders were mated with each other and the mutations were determined to be transmitted to the F1 piglets. We established a robust and safe technology for developing genetically modified pig lines with expected genotypes for agricultural breeding and biomedical application.

The objective of this study was to evaluate the effects of low ambient temperature (LAT) and dietary vitamin C (VC) supplementation on pulmonary vascular remodeling (PVR) and the relative expression of hypoxia inducible factor-1α (HIF-1α), vascular endothelial growth factor (VEGF) and its receptor 2 (VEGFR-2) mRNA of lungs in 21-d-old broilers. 400 1-d-old male Cobb broilers were assigned randomly to 4 treatments as follows for 21 d: 1) LAT and a basal diet; 2) LAT and a basal diet supplemented with 1 000 mg kg–1 VC (LAT+VC); 3) normal ambient temperature (NAT) and a basal diet; 4) NAT and a basal diet supplemented with 1 000 mg kg–1 VC (NAT+VC). Each treatment was composed of 10 replicates of 10 birds per replicate. Samples of lung were collected after the broilers were killed at d 21. LAT increased the ratio of vessel wall area to vessel total area (WA/TA, %) and mean media thickness in pulmonary arterioles (mMTPA, %) (P<0.05). Dietary VC supplementation decreased mMTPA (P<0.05), but had no effect on the WA/TA. LAT increased (P<0.05) the relative mRNA expression of HIF-1α, VEGF and VEGFR-2, while adding VC to the diet could decrease (P<0.05) their relative mRNA expression. A significant positive correlation existed between the level of VEGF mRNA expression and the value of WA/WT (P<0.05) or mMTPA (P<0.05). These results suggested LAT resulted in pulmonary vascular remodeling, and the increase of HIF-1α, VEGF and VEGFR-2 mRNA expression, and dietary VC supplementation can alleviate pulmonary vascular remodeling in broiler by affecting these gene expression.

Grain size is one of the most important agronomic components of grain yield. Grain length, width and thickness are controlled by multiple quantitative trait loci (QTLs). To understand genetic basis of large grain shape and explore the beneficial alleles for grain size improvement, we perform QTL analysis using an F2 population derived from a cross between the japonica variety Beilu 129 (BL129, wide and thick grain) and the elite indica variety Huazhan (HZ, narrow and long grain). A total number of eight major QTLs are detected on three different chromosomes. QTLs for grain width (qGW), grain thickness (qGT), brown grain width (qBGW), and brown grain thickness (qBGT) explained 77.67, 36.24, 89.63, and 39.41% of total phenotypic variation, respectively. The large grain rice variety BL129 possesses the beneficial alleles of GW2 and qSW5/ GW5, which have been known to control grain width and weight, indicating that the accumulation of the beneficial alleles causes large grain shape in BL129. Further results reveal that the rare gw2 allele from BL129 increases grain width, thickness and weight of the elite indica variety Huazhan, which is used as a parental line in hybrid rice breeding. Thus, our findings will help breeders to carry out molecular design breeding on rice grain size and shape.

Construction of a food-grade expression vector for application to lactic acid bacteria (LAB) is of importance for dairy fermentation system. An α-galactosidase (aga) gene encoding an enzyme degrading melibiose was amplified by PCR from the plasmid pRAF800 of Lactococcus lactis NZ9000. The aga gene was introduced into pMG36e to substitute the primary antibiotic selectable marker of pMG36e, resulting in construction of a new food-grade expression vector pMG36-aga. To testify the expression efficiency of exogenous gene in pMG36-aga, a 1.5 kb long α-amylase (amy) gene from Bacillus licheniformis was cloned by PCR and introduced into the plasmid pMG36-aga. The resultant plasimd pMG36-aga-amy was transformed into L. lactis ML23 by electroporation. The positive clones were selected with the medium containing melibiose as the sole carbon source. The selection efficiency of aga was 8.71×103 CFU with a standard deviation of 9.1×102 CFU mg-1 DNA of pMG36-aga. Furthermore, the SDS-PAGE analysis showed that the pMG36-aga-amy expressed a 56.4 kDa protein which was the same as the putative molecular weight of α-amylase. The starch plate assay also indicated that L. lactis ML23 displayed high activity of α-amylase by expressing of amy gene of pMG36-aga-amy.

Changes in drought trends and its relationship with climate change in China were examined in this study. The temperature vegetation drought index (TVDI) datasets recorded during 2001 to 2010 in China were constructed by using the moderateresolution imaging spectroradiometer (MODIS) data, and the temporal and spatial variations in drought were analyzed. In addition, the mean temperature, mean precipitation, mean relative humidity, and mean sunshine duration data collected from 557 local weather stations in China were analyzed. The relationships between drought and these climate factors were also analyzed by using correlation analysis and partial correlation analysis. Changes in drought tendency were shown to differ among four regions throughout the country. The fluctuations in climate in the regions of northeastern China, Qinling-Huaihe, and central Qinghai were caused by the increase in soil moisture, and that in southern Tibet was caused by the intensification of drought. Meteorological factors exhibited varied effects on drought among the regions. In southern China, the main influential factor was temperature; other factors only showed minimal effects. That in the northern and northwestern regions was sunshine duration, and those in the Qinghai-Tibet Plateau were relative humidity and temperature.

Biochar is increasingly proposed as a soil amendment, with reports of benefits to soil physical, chemical and biological properties. In this study, different biochars were produced from 6 feedstocks, including straw and poultry manure, at 3 pyrolysis temperatures (200, 300 and 500°C) and then added separately to a calcareous soil. Their effects on soil properties and maize growth were evaluated in a pot experiment. The biochars derived from crop straw had much higher C but smaller N concentrations than those derived from poultry manure. Carbon concentrations, pH and EC values increased with increasing pyrolysis temperature. Biochar addition resulted in increases in mean maize dry matter of 12.73% and NPK concentrations of 30, 33 and 283%, respectively. Mean soil pH values were increased by 0.45 units. The biochar-amended soils had 44, 55, 254 and 537% more organic C, total N, Olsen-P and available K, respectively, than the control on average. Both feedstocks and pyrolysis temperature determined the characteristics of the biochar. Biochars with high mineral concentrations may act as mineral nutrient supplements.

Quantitative assessment of development sustainability could be a challenge to regional management and planning, especially for areas facing great risks of water shortage. Surface-water decline and groundwater over-pumping have caused serious environmental problems and limited economic development in many regions all around the world. In this paper, a framework for quantitatively evaluating development sustainability was established with water-related eco-environmental carrying capacity (EECC) as the core measure. As a case study, the developed approach was applied to data of the Haihe River Basin, China, during 1998 through 2007. The overall sustainable development degree (SDD) is determined to be 0.39, suggesting that this rate of development is not sustainable. Results of scenario analysis revealed that overshoot, or resource over- exploitation, of the Basin’s EECC is about 20% for both population and economy. Based on conditions in the study area in 2007, in order to achieve sustainable development, i.e., SDD>0.70 in this study, the EECC could support a population of 108 million and gross domestic product (GDP) of 2.72 trillion CNY. The newly developed approach in quantifying eco- environmental carrying capacity is anticipated to facilitate sustainable development oriented resource management in water- deficient areas.

The β-1,3-glucan from yeast has been extensively examined for its immuno-enhancing effects in animals. However, investigation on the relationship among β-glucan, gut microbiota and immune-modulating effects remains limited particularly in pigs. Considering the critical roles of gut methanogens in the microbial fermentation, energy metabolism and disease resistance, we investigated the phylogenetic diversity of methanogens from fermented cultures of porcine colonic digesta with (G) or without (N) yeast β-glucan based on sequences of the archaeal 16S rRNA gene. A total of 145 sequences in the G library were assigned into 8 operational taxonomic units (OTUs) with the majority of sequences (114/145) related to strains Methanobrevibacter millerae or Methanobrevibacter gottschalkii with high identities ranging from 97.9 to 98.6%, followed by 23 sequences to Methanobrevibacter ruminantium, 2 sequences to Methanobrevibacter smithii and one sequence to Methanobrevibacter wolinii. The 142 sequences in the N library were assigned to 2 OTUs with most sequences (127/142) related to strains M. millerae or M. gottschalkii with sequence identities ranging from 97.9 to 98.5%, and 15 sequences related to M. gottschalkii with 97.9% identity. Shannon diversity index showed that the G library exhibited significantly higher archaeal diversity (P<0.05) and Libshuff analysis indicated the differences in the community structure between the two libraries were significant (P<0.0001). In conclusion, the current study provides evidence that addition of yeast β-glucan significantly increased the diversity of methanogens in in vitro fermented porcine colonic digesta.

The reuse of treated wastewater in agricultural systems could partially help alleviate water resource shortages in developing countries. Treated wastewater differs from fresh water in that it has higher concentrations of salts, Escherichia coli and presence of dissolved organic matter, and inorganic N after secondary treatment, among others. Its application could thus cause environmental consequences such as soil salinization, ammonia volatilization, and greenhouse gas emissions. In an incubation experiment, we evaluated the characteristics and effects of water-filled pore space (WFPS) and N input on the emissions of nitrous oxide (N2O) and carbon dioxide (CO2) from silt loam soil receiving treated wastewater. Irrigation with treated wastewater (vs. distilled water) significantly increased cumulative N2O emission in soil (117.97 μg N kg-1). Cumulative N2O emissions showed an exponentially increase with the increasing WFPS in unamended soil, but the maximum occurred in the added urea soil incubated at 60% WFPS. N2O emissions caused by irrigation with treated wastewater combined with urea-N fertilization did not simply add linearly, but significant interaction (P<0.05) caused lower emissions than the production of N2O from the cumulative effects of treated wastewater and fertilizer N. Moreover, a significant impact on cumulative CO2 emission was measured in soil irrigated with treated wastewater. When treated wastewater was applied, there was significant interaction between WFPS and N input on N2O emission. Hence, our results indicated that irrigation with treated wastewater should cause great concern for increasing global warming potential due to enhanced emission of N2O and CO2.

Five commonly-used reference genes: ACT (actin), UBE (ubiquitin-conjugating enzyme), RPL2 (ribosomal protein L2), BRP II (RNA polymerase II subunit), and NADH (nicotinamide adenine dinucleotide) were examined using geNorm software as reference genes for RT-qPCR. Among the tested reference genes, ACT and UBE were the most stable in all samples. In parallel, expression analysis of nitrilases in Brassica juncea var. tumida, was performed to preliminarily investigate the molecular interactions between nitrilase and clubroot development at 10, 15, 20, 25, 30, and 40 d postinoculation (dpi) with a suspension of resting spores of Plasmodiophora brassicae. The results showed that different gene expressions of nitrilases were regulated during the initial periods of clubroot development. The expression level of BjNIT1 increased sharply from 20 to 40 dpi in infected roots while there were no remarkable changes in healthy roots. From 15 to 30 dpi, the expression levels of BjNIT2 and BjNIT4 in infected roots were lower than those in non-infected roots. Finally, BjNIT2 in treatment was down approximately to control at 40 dpi. Our results suggest that BjNIT1, which promoted overproductions of auxin, might be involved in P. brassicae infection of B. juncea.