家蚕脂蛋白基因BmLp-c21的表达分析及蛋白质亚细胞定位

家蚕30 kD脂蛋白家族在家蚕的生长发育过程中具有重要的生理功能。从家蚕蛹cDNA文库中克隆到一条编码30kD蛋白质的基因cDNA序列,该序列全长2 803 bp,ORF为792 bp,编码含263个氨基酸残基的脂蛋白(low molecular 30K lipo-protein pBmHPC-21;GenBank登录号:Q00801),命名为BmLp-c21。将BmLp-c21克隆至原核表达载体pET-28a(+),转化E.coli Rosetta(DE3),IPTG诱导表达后通过亲和层析得到纯化的融合蛋白,并免疫新西兰大白兔制备多克隆抗体。亚细胞定位显示BmLp-c21主要存在于细胞质中,呈点状或者片状分布。通过实时荧光定量PCR和Western blotting方法,分别检测到家蚕不同发育时期和5龄幼虫不同组织中BmLp-c21 mRNA的转录水平与蛋白质的表达水平存在较大差异,在蛹期及5龄幼虫血淋巴中的mRNA转录水平和蛋白质表达水平最高。初步推测BmLp-c21在家蚕蛹的变态发育过程以及蚕体脂质的运输方面具有重要的功能。     

广州近代城市公园的中西合璧特征探析

广州近代城市公园设计精巧,风格独特,在岭南造园史上具有重要地位。结合历史图文资料,运用风景园林史学和风景园林美学相结合的研究方法,归纳分析其造园特征。广州近代城市公园的中西合璧特征主要体现在3个方面:因地制宜的空间布局、中西融合的建筑小品、多姿多彩的植物景观。探析其中西合璧特征,有利于丰富岭南近代园林历史与理论研究,推进对广州近代城市公园的保护与造园智慧的传承。     

Antibiotic resistant Escherichia coli in southeastern Australian pig herds and implications for surveillance

To investigate public health implications of antibiotics to control post‐weaning scours, we surveyed 22 commercial pig herds in southeastern Australia. Fifty faecal samples per herd were collected from pre‐ and post‐weaned piglets. Presumptive Escherichia coli isolates were confirmed by MALDI‐TOF MS. Isolates (n = 325) were screened for susceptibility to 19 veterinary antibiotics using MIC broth microdilution. All 325 E. coli isolates underwent further testing against 27 antibiotics used in human medicine and were screened for ETEC adhesin and enterotoxin genes (F4 (K88), F5 (K99), F6 (987P), F18, F41, STa, STb, Stx2e and LT) by multiplex PCR. Isolates identified as phenotypically resistant to third‐generation cephalosporin (3GC) and aminoglycoside antibiotics were screened by multiplex PCR/reverse line blot to detect common β‐lactam and aminoglycosides resistance genes, confirmed by sequencing. Twenty (6.1%) of the E. coli isolates were resistant to 3GC antibiotics and 24 (7.4%) to the aminoglycoside antibiotic gentamicin. Genetic analysis revealed six different extended spectrum β‐lactamase (ESBL) genes (blaCTX‐M‐1, ‐14, ‐15, ‐27, blaSHV‐12 and blaCMY‐2‐like genes), four of which have not been previously reported in Australian pigs. Critically, the prevalence of 3GC resistance was higher in non‐pathogenic (non‐ETEC) isolates and those from clinically normal (non‐diarrhoeal) samples. This highlights the importance of non‐ETECE. coli as reservoirs of antimicrobial resistance genes in piglet pens. Antimicrobial resistance surveillance in pig production focused on diagnostic specimens from clinically‐affected animals might be potentially misleading. We recommend that surveillance for emerging antimicrobial resistance such as to 3GC antibiotics should include clinically healthy pigs.     

西部12省市农村居民生活消费支出现状的比较研究

选取西部12省市的农村居民和各地区农村居民家庭平均每人生活消费支出的数据,采用面板数据模型分析居民收入结构对生活消费支出的影响实证研究。结果表明：农村居民的工资性收入、家庭经营纯收入、财产性收入和转移性收入对生活消费支出都有显著影响,且财产性收入和转移性收入的边际消费倾向都大于1。     

β-catenin在不同毛色羊驼皮肤中的表达和定位

旨在研究β-catenin在不同毛色羊驼皮肤中的表达和定位,探索其与毛色的关系。以成年白色和棕色羊驼为研究对象,采用实时荧光定量PCR技术、Western blot和免疫组织化学技术,对β-catenin在白色和棕色羊驼皮肤中mRNA、蛋白表达水平和定位进行研究。实时荧光定量PCR结果显示,β-catenin在棕色羊驼皮肤组织中的相对基因表达量是白色羊驼皮肤组织的1.662倍;Western blot结果显示,羊驼皮肤组织粗蛋白提取物中存在分子量约85 ku与兔抗β-catenin多克隆抗体发生免疫阳性反应的蛋白条带,棕色羊驼平均蛋白表达量显著高于白色羊驼;免疫组织化学结果显示,β-catenin在羊驼皮肤的表皮、毛乳头、毛根鞘和皮脂腺中表达,根据光密度值得出,除皮脂腺之外,在表皮、毛乳头和毛根鞘的表达差异极显著(P0.01)。结果提示β-catenin在白色和棕色羊驼皮肤组织中定位以及表达的差异,表明β-catenin参与毛色形成。     

Flow alteration and thermal pollution depress modelled growth rates of an iconic riverine fish,the Murray cod Maccullochella peelii

The serial discontinuity concept (SDC) proposes that hypolimnetic‐releasing impoundments cause major disruptions to the naturally occurring physical, chemical and biological gradients of rivers but that this impact diminishes with distance downstream. Such a gradient in discharge, flow velocity and temperature regime occurs below a large hypolimnetic‐releasing impoundment, the Hume Dam, on the River Murray in south‐eastern Australia. To examine the effects of this disturbance gradient on a warm‐water large‐bodied freshwater fish, the Murray cod (Maccullochella peelii), a bioenergetics model was developed and calibrated to explore energy expended under differing water velocities and temperature regimes. Model simulations predicted negative growth of juveniles directly downstream of the impoundment, due largely to the energetic costs associated with active and, to a lesser extent, standard metabolism outweighing the achievable energetic gains through food consumption. As flow velocity and temperature regimes became more favourable downstream, so did the simulated growth of the species. It was not until +239 km downstream of the impoundment that the model predicted that flow velocity and temperature regimes were suitable for greater weight gains. The modelled growth responses of juvenile Murray cod are consistent with the predictions of the SDC, emphasising that changes in the bioenergetics of individuals are likely to be reflected in reduced growth rates under the changed flow velocity and temperature regimes imposed by disturbance gradients. This research represents a valuable step in the biological understanding of Murray cod within variable riverine environments and emphasises the urgency required to mitigate impacts associated with hypolimnetic impoundments.     

Detection and characterization of viruses of the genus Megalocytivirus in ornamental fish imported into an Australian border quarantine premises: an emerging risk to national biosecurity

This report documents an emerging trend of identification of Megalocytivirus-like inclusions in a range of ornamental fish species intercepted during quarantine detention at the Australian border. From September 2012 to February 2013, 5 species of fish that had suffered mortality levels in excess of 25% whilst in the post-entry quarantine and had Megalocytivirus-like inclusion bodies in histological sections were examined by PCR. The fish had been imported from Singapore, Malaysia and Sri Lanka. Ninety-seven of 111 individual fish from affected tanks of fish tested were positive for the presence of Megalocytivirus by PCR. Sequence analysis of representative PCR products revealed an identical sequence of 621 bp in all cases which was identical to a previously characterized Megalocytivirus (Sabah/RAA1/2012 strain BMGIV48). Phylogenetic analysis of available Megalocytivirus major capsid protein (MCP) sequences confirmed the existence of 3 major clades of Megalocytivirus. The virus detected in this study was identified as a member of Genotype II. The broad host range and pathogenicity of megalocytiviruses, coupled to the documented spread of ornamental fish into the environment, render this a significant and emerging biosecurity threat to Australia.     

犬瘟热病毒H基因的序列分析与表达

以临床疑似犬瘟热病犬外周血总RNA为模板,采用RT-PCR方法扩增犬瘟热病毒(CDV)血凝素(H)基因。序列测定和分析表明该病料犬瘟热病毒H基因序列与国内其他地区分离到的毒株同源性较高(94.6%~99.2%),而与Onderstepoort株、Convac株等疫苗毒株的同源性较低(90.4%~91.2%)。分子遗传进化分析显示所有毒株可分为7个大的分支,且各分支间具有一定的地域性,其中待检病料中CDV与大多数中国分离株一样处于Ⅰ型。进一步以H全基因为模板,截短表达其3′端816 bp、459 bp两个片段,并克隆入原核表达载体pET30 a进行表达。结果显示它们分别能表达大小约为36.4 ku和22.2 ku的融合蛋白。免疫转印表明该纯化蛋白均可与犬瘟热病毒抗血清发生阳性反应,说明重组蛋白具备抗原性,可用于进一步的血清学研究。     

禽多杀性巴氏杆菌C48-1株OmpA基因表达与抗原性鉴定

为了研究禽多杀性巴氏杆菌(Pm)外膜蛋白A(Omp A)的免疫原性,根据C48-1菌株Omp A基因序列设计一对特异性引物,采用PCR方法扩增去信号肽的成熟Omp A基因.将去信号肽的成熟Omp A基因扩增片段双酶切后,克隆到p GEX-6p-1表达载体中,构建重组表达质粒p GEX-Omp A,转化宿主菌BL21并经异丙基硫代半乳糖苷(IPTG)诱导表达.SDS-PAGE的结果显示,融合蛋白GST-Omp A大小约为63 ku,与预期的分子质量相符.免疫印迹分析结果表明,该融合蛋白与鸡抗Pm血清具有明显的免疫反应,说明Omp A具有良好的抗原性,这为禽Pm Omp A在免疫防御研究中的应用奠定了基础.