规模化鸡场中产CMY-2大肠杆菌耐药基因与毒力基因的调查及共转移研究

调查规模化鸡场中产CMY-2大肠杆菌耐药基因与毒力基因的流行现状及共转移情况,对549株来源于鸡场粪样和苍蝇的大肠杆菌进行blaCMY-2PCR检测;K-B法检测耐药表型;PCR扩增19种相关耐药基因和14种毒力基因;接合试验和质粒复制子分型研究耐药基因与毒力基因的共转移.结果显示:6.0%(33/549)大肠杆菌呈CMY-2阳性,且均为多重耐药;检测到15种耐药基因和4种毒力基因,其中floR,blaTEM-1,sul2,sul1,traT,VagC检出率较高;63.6%产CMY-2大肠杆菌中blaCMY-2基因可通过lncA/C或lncI1质粒与耐药基因和(或)毒力基因共转移.本研究表明规模化鸡场粪样和苍蝇源产CMY-2大肠杆菌已成为耐药基因和毒力基因的重要储存库,携带二者质粒的转移导致耐药与毒力的传播扩散.     

宁夏地区162株大肠杆菌临床分离株对磺胺类药物的耐药性分析

分析宁夏地区大肠杆菌临床分离株对磺胺类药物的耐药性,为指导临床合理用药提供参考.用PCR技术和药敏实验对宁夏地区临床分离的162株大肠杆菌进行了磺胺类药物耐药基因sul1,sul2,sul3的分子检测和耐药性研究.结果表明,在162株大肠杆菌临床分离株中,127株检测出了耐药基因,阳性率为78.4%.其中,sul1基因的阳性率最高,为65.4%,sul2,sul3基因的阳性率分别为48.1%,1.9%;106株对磺胺类药物的耐药率为65.4%,耐药菌株中有5株未检测到sul1,sul2,sul3耐药基因.药敏实验结果与基因检测结果的符合率为95.3%.宁夏地区大肠杆菌临床分离株对磺胺类药物具有较高的耐药性,应予以足够重视.     

产β-内酰胺酶伤寒杆菌与痢疾杆菌blaTEM耐药基因研究

目的:研究新余市产β-内酰胺酶伤寒杆菌与痢疾杆菌blaTEM-1D型耐药基因流行特征。方法:收集新余新钢中心医院、新余市人民医院、新余市中医院、新余市妇幼保健院2008年1月—2010年6月分离出的伤寒杆菌产酶株6株和痢疾杆菌产酶株13株共19株,用头孢哨噻吩滤纸片法做β-内酰胺酶的定性检测,确定为产酶株后,采用PCR技术扩增质粒blaTEM-1D耐药基因,用Mark标记量出扩增子的大小(bp),统计blaTEM-1D耐药基因阳性率。结果:伤寒杆菌、痢疾杆菌产酶株blaTEM-1D耐药基因阳性率分别为66.7%(4/6)、76.9%(10/13)。结论:blaTEM-1D型耐药基因在本地区各家医院间存在水平传播和医院内垂直传播。     

鸽源大肠杆菌分离鉴定、毒力基因检测及药敏试验

为了确定疑似细菌感染鸽的病原菌,以病死鸽为研究对象,进行病原菌的分离培养、生化鉴定、16S rDNA序列分析、血清型检测、动物致病性试验、毒力基因检测以及药敏试验。结果表明:此次分离的1株优势菌株,命名为QH-1,是短杆状、中等大小的革兰氏染色阴性菌,且其形态特征和生化鉴定特性与大肠杆菌相符;经16S rDNA序列分析进一步证实菌株QH-1为大肠杆菌;血清型检测结果显示,分离菌株QH-1为O26血清型分离株;动物致病性试验结果显示,分离菌株QH-1能引起鸽和小白鼠感染发病,且试验鸽发生与自然感染病例相似症状;毒力基因检测结果显示,大肠杆菌QH-1携带irp2,FyuA毒力基因;药敏试验结果显示,大肠杆菌QH-1对头孢曲松敏感;对阿莫西林、环丙沙星等耐药。疑似细菌感染的鸽病例为大肠杆菌的感染,可以选择头孢曲松进行防控。     

β－淀粉酶基因在黄单胞菌中的克隆及表达

广泛寄主范围载体ｐＫＴ２１０可在辅助质粒ｐＲＫ２０１３的协助下转移进入黄单胞菌（革兰氏阴性）中并能稳定存在；来源于枯草杆菌的β－淀粉酶基因与载体ｐＫＴ２１０形成重组质粒ｐＹＬ１，并以其转化大肠杆菌；通过三亲本接合将ｐＹＬ１引入带有利福平抗性的黄单胞菌ＮＫ０１－Ｒ中，通过抗性选择接合子，并测定接合子的淀粉酶水解能力及产胶能力．本文获得一株黄原胶产量比出发菌株提高２０％，且淀粉酶水解能力显著提高的工程菌株．     

养禽场周围水环境中磺胺类抗性菌、抗性基因分布

为进行养禽场周围水环境中磺胺类抗性菌、抗性基因分布的研究,从3家养禽场多个区域水环境中分离的56株大肠杆菌(Escherichia coli,E.coli),应用肉汤微量稀释法分析磺胺类药物敏感性,应用PCR方法检测磺胺类药物抗性基因。结果有18株(32.14%)磺胺类抗性菌,对利福平的耐受性最弱,对氯霉素的耐受能性最强;共检测到64个抗性基因,三个养禽场分别为:19(29.69%)、29(45.31%)、16(25.00%);分别为sul1 18(28.12%)、sul2 13(20.32%)、sul3 7(10.94%)、Int1 18(28.12%)、Int2 8(12.50%)。含一种抗性基因的菌株为1株、含2种抗性基因的菌株为1株、含三种抗性基因的菌株为6株、含四种抗性基因的菌株为5株、含五种抗性基因的菌株为4株,而1株无抗生素抗性的菌株检出抗性基因。结果表明在养禽场场内检测到抗性基因和抗性菌株数量较高、养禽场周围水环境中存在磺胺类抗性菌株,且具有多重抗性,可能对生态环境的产生一定的不良影响。     

β—淀粉酶基因在黄单胞菌中的克隆及表达

广泛寄主范围载体ｐＫＴ２１０可在辅助质粒ｐＲＫ２０１３的协助下转移进入黄单胞菌（革兰氏阴性）中并能稳定存在；来源于枯草杆菌的β－淀粉酶基因与载体ｐＫＴ２１０形成重组质粒ｐＹＬ１，并以其转化大肠杆菌；通过三亲本接合将ｐＹＬ１引入带有利福平抗性的黄单胞菌ＮＫ－０１－Ｒ中，通过抗性选择接合子，并测定接合子的淀粉酶水解能力及产胶能力。本文获得一株黄原胶产量比出发菌株提高２０％，且淀粉酶水解能力显著提高的工     

肠球菌临床耐药性分析及耐药基因vanM在屎肠球菌中的检测

目的了解肠球菌临床分离株耐药性及vanM基因在屎肠球菌的分布情况,为临床合理用药及探讨耐药机制提供依据.方法采用VITEK60全自动微生物分析仪对肠球菌进行鉴定及药敏试验,按NCCLS/CLSI标准判断并统计分析结果;PCR法检测耐万古霉素、替考拉宁菌株中的耐药基因vanM,并对阳性基因进行测序,分析耐药基因与耐药性的关系.结果临床分离肠球菌共506株,其中屎肠球菌277株,粪肠球菌229株.尿液标本来源比例最高,为167株（33%）;粪便标本114株（22.5%）;痰液标本78株（15.4%）;胆汁标本57株（11.3%）;脓液标本54株（10.7%）;其他来源36株（7.1%）.药敏结果显示：肠球菌对大部分临床抗菌药高度耐药,屎肠球菌对β-内酰胺类抗菌药物（氨苄西林）、喹诺酮类（左氧氟沙星）、糖肽类（万古霉素、替考拉宁）的耐药率明显高于粪肠球菌（P〈0.05）.vanM基因在耐万古霉素菌株中的检出率为100%,耐替考拉宁菌株检出率为90%.结论屎肠球菌可引起临床各类感染,屎肠球菌对大多数抗生素的耐药率明显高于粪肠球菌,且呈多重耐药趋势.vanM基因可能在糖肽类药物万古霉素、替考拉宁耐药机制中发挥重要作用.     

苏云金芽孢杆菌高效菌株cry基因分析及多价基因组合的研究

对本室筛选的高活性Ｂ．ｔ．野生株进行了ｃｒｙ 基因检测,Ｓｏｕｔｈｅｒｎ杂交证明ｃｒｙＩＡｃ和ｃｒｙ１Ｃ位于质粒上．利用接合转移和电穿孔转化等方法对这些菌株进行了遗传改良研究．结果表明：高效野生株７４０４、ＨＤ－１－Ｘ和９５１０ 不易获得并表达外源ｃｒｙ 基因,而高效野生株Ｂｔｉ． ｔ－１８９７ 的电转化频率较高,并获得以Ｂｔｉｔ－１８９７ 为受体,对甜菜夜蛾、棉铃虫、黏虫和淡色库蚊均有毒力,具有ｃｒｙＩＡｃ、ｃｒｙＩＣ、ｃｒｙＩＶ及ｃｙｔ多价基因的广谱工程菌株．     

多重PCR筛选临床细菌耐药整合子基因

目的：建立多重PCR方法并对临床菌株进行扩增及整合子分类。方法：采用多重PCR对21株来自临床的耐药菌株进行了第一、二和三类整合酶基因（int Ⅰ）筛选。结果：16株携带第一类整合酶基因;1株携带第一和第二类整合酶基因;2株含第二类整合酶基因;1株含第三类整合酶基因;1株不含第一、二和三类整合酶基因。结论：筛选细菌耐药整合子基因分类的多重PCR方法简便可靠;整合子广泛存在于临床耐药细菌中。     

Genetic variations of glycinin subunit genes among cultivated and wild type soybean species

Glycinin is a predominant storage protein in most soybean accessions. It is a hexamer constituted by five major subunits, which can be classified into two groups. Group I contains G1, G2 and G3, and Group II contains G4 and GS. The genes encoding these subunits have been designated from Gyl to Gy5, respectively. In the present study, Gyl genomic fragments were cloned from wild accessions of subgenera Glycine glycine, Glycine soja and a cultivar of Glycine max. Their sequences and the deduced amino acid sequences were compared. The residues critical for assembling of G1 subunits from the wild perennial accession were conservative. The Gy4 fragments were cloned from two wild perennial accessions and compared with that from subgenus Soja. The intron 3 of Gy4 had abundant variations between the subgenera G. soja and G. glycine as well as within the subgenus G. glycine. Abundant variations existed in the disordered regions 3 and 4 of G4 subunits from two wild perennial accessions. The genomic organization of glycinin genes was analyzed in 19 accessions from subgenera Soja and Glycine. The hybridization patterns were identical among the accessions of subgenus Soja. On the contrary, abundant polymorphisms existed between the accessions from subgenus Glycine. These results indicated that glycinin genes have high degree of conservation within subgenus Soja but more variations within subgenus Glycine.     

Genetic variations of glycinin subunit genes among cultivated and wild type soybean species

Glycinin is a predominant storage protein in most soybean accessions. It is a hexamer constituted by five major subunits, which can be classified into two groups. Group I contains G1, G2 and G3, and Group II contains G4 and G5. The genes encoding these subunits have been designated from Gy1 to Gy5 , respectively. In the present study, Gy1 genomic fragments were cloned from wild accessions of subgenera Glycine glycine, Glycine soja and a cultivar of Glycine max . Their sequences and the deduced amino acid sequences were compared. The residues critical for assembling of G1 subunits from the wild perennial accession were conservative. The Gy4 fragments were cloned from two wild perennial accessions and compared with that from subgenus Soja . The intron 3 of Gy4 had abundant variations between the subgenera G. soja and G. glycine as well as within the subgenus G. glycine. Abundant variations existed in the disordered regions 3 and 4 of G4 subunits from two wild perennial accessions. The genomic organization of glycinin genes was analyzed in 19 accessions from subgenera Soja and Glycine . The hybridization patterns were identical among the accessions of subgenus Soja. On the contrary, abundant polymorphisms existed between the accessions from subgenus Glycine . These results indicated that glycinin genes have high degree of conservation within subgenus Soja but more variations within subgenus Glycine .     

Lamins and their genes

The nuclear lamina is an important nuclear structure. The studies on its peptides lamins and genes have made substantial progress in recent years. In this paper, the new achievements in studies of new lamin members and their functions, lamin genes and their origin and differentiation, are reviewed and discussed. Some research areas that deserve to be investigated further are put forward.     

Origin and evolution of new genes

Organisms have variable gcnome sizes and contain different numbers of genes. This difference demonstrates that new gene origination is a fundamental process in evolutionary biology. Though the study of the origination of new genes dated back more than half a century ago, it is not until the 1990s when the first young genejingwei was found that empirical investigation of the molecular mechanisms of origination of new genes became possible. In the recent years, several young genes were identified and the studies on these genes have greatly enriched the knowledge of this field. Yet more details in a general picture of new genes origination are to be clarified. We have developed a systematic approach to searching for young genes at the genomic level, in the hope to summarize a general pattern of the origination and evolution of new genes, such as the rate of new gene appearance, impact of new genes on their host genomes, etc.     

Structural relationship of the SB beta-chain gene to HLA-D-region genes and murine I-region genes

The major histocompatibility complex (MHC) regulates several aspects of the immune response. Class II antigens of the MHC control cellular interactions between lymphocytes. In man, at least three class II antigens (DR, DC and SB), consisting of distinct alpha- and beta-chains, are encoded in the HLA complex. Sequence analysis has established that the DR and DC antigens are the respective structural counterparts of the murine I-E and I-A antigens. Molecular cloning of the SB beta-chain gene has now enabled us to define its relationship to other class II genes. The DR, DC and SB beta genes have diverged from each other to the same extent. In murine DNA and in cloned genes from the I region, the best hybridization of SB beta DNA is with the E beta 2 sequence. E beta 2 may belong to a complete gene (E' beta) because first domain sequences were found adjacent to it.     

松属编码区微卫星特征和相应基因功能分析

从GenBank下载453 892条松属EST序列,序列组装后得到20 886条contig。用Sputnik软件从这些contig中查找了2 678个微卫星,其中3碱基重复微卫星占的比例最高,为59.2%,而其他重复长度的微卫星都相对较低,比例分别为:2碱基重复微卫星占12.0%,4碱基重复微卫星占13.3%,5碱基重复微卫星占15.5%。3碱基重复微卫星变化引起的基因读码框改变最小,松属树种基因区3碱基重复微卫星的富集显示了强烈的密码子选择效应。此次研究还对查找到的微卫星进行了引物设计和扩增分析。实验结果显示,设计的微卫星引物在云南松中的扩增成功率是72.9%。从扩增成功的引物中进一步选取了155对引物,对14个松属树种和1个黄杉属树种进行了引物通用性实验分析,结果显示155对引物在14个松属树种间的通用性在71.0%以上,而在黄杉属树种中的通用性只有25.2%。对松属树种中含有微卫星的基因进行了功能分类研究,结果显示基因在是否保留微卫星序列方面有显著分化,微卫星参与了如细胞成分分类的共质体组成、病毒颗粒及病毒颗粒组成、生物节律调控, 以及生长素转运蛋白等生物学过程。