一株乳源松鼠葡萄球菌ExhC基因的克隆及序列分析

松鼠葡萄球菌(Staphylococcus sciuri,S.sciuri)是引起奶牛乳房炎(Cow Mastitis)的病原菌之一,其主要致病因子是该菌产生的脱皮毒素C(Exfoliative Toxin C,ExhC)。为了深入了解该基因的分子特征,本试验对从乳房炎患牛牛乳中分离的一株松鼠葡萄球菌ExhC基因(Genbank登录号:MT845354)进行了克隆及序列分析。按GenBank收录的ExhC基因设计并合成引物,利用PCR对ExhC基因进行扩增,并对扩增后的ExhC基因进行测序及分析。结果表明,松鼠葡萄球菌ExhC基因的开放阅读框序列长度为837bp,共编码278个氨基酸。通过Blast模块对ExhC基因进行同源性分析,得到5株相似序列,与ExhC基因序列相比同源性均为99.04%。系统进化树图指示该基因与其余五株相似序列属于不同的分支,遗传距离较远。利用TMHMM对松鼠葡萄球菌ExhC基因837bp序列跨膜区进行预测,结果显示ExhC蛋白的第1~277位氨基酸在细胞膜外,5~25位氨基酸所在位置为跨膜区。通过ExhC氨基酸序列分析和ExhC蛋白二级结构、三级结构预测,推断第97位氨基酸插入Asn以及其他位点氨基酸的突变可能会导致松鼠葡萄球菌功能区变化和毒株毒力的相对变化,这将给松鼠葡萄球菌致病机理的研究奠定基础。     

Quantitative and qualitative aspects of peroxidases extracted from cladodes of Opuntia ficus indica

Peroxidases (EC 1.11.1.7) were extracted from fresh cladodes harvested from nine ecotypes of the cactus species Opuntia ficus indica Mill. growing as a collection, in Marrakech (South Morocco). Two enzyme fractions were obtained by a progressive solubility method leading to soluble peroxidases (S) and ionically wall-bound peroxidases (I). The preferred substrate for cladode peroxidases was determined to be o-dianisidine over 4-chloro-1-naphtol and guaiacol. Contrarily to roots, no guaiacol-based activity was found in cladodes. The late ecotypes 1‘Haddaouia’ and ‘Moussa’ showed a relatively high peroxidase ratio S/I (units g⁻¹ fresh weight). When subjected to electrophoresis on polyacrylamide gels, S and I peroxidase fractions each exhibited two enzyme forms based on their electric charges in basic and acidic gel media. Acidic peroxidase forms, well separated on basic gels, showed two principal migration zones with great differences in their enzyme activities depending on the fraction types S and I. Acidic ionically wall-bound peroxidases exhibited fast, highly active isoforms with R_f values 0.42–0.58. Basic forms, represented essentially in fractions S and resolved on acidic gels, were typified by slow and fast isoforms with a double banded pattern in most ecotypes. Opuntia ecotypes collected from localities surrounding Marrakech exhibited fast basic soluble isoforms, while distant ecotypes from Marrakech, including late ones, were typified by a fast acidic ionically bound isoperoxidase of R_f 0.58. Possible roles of O. f. indica peroxidases in growth and their evaluation as markers in this cactus species are discussed in this study.     

牛支原体、无乳支原体和丝状支原体丝状亚种小克隆三重PCR检测方法的建立及初步应用

本研究旨在建立一种可一次性区分牛支原体、丝状支原体丝状亚种小克隆和无乳支原体的三重PCR诊断方法,为临床诊断和流行病学调查提供可靠检测技术.根据GenBank发表的上述3种病原的基因组序列保守区域设计3对特异性引物建立三重PCR方法;确定其检测敏感性,以猪支原体、鸡支原体、无乳支原体和丝状支原体丝状亚种小克隆基因作模板检验其特异性;同时和病原分离鉴定结果对比其准确性.结果表明在优化体系和条件下能够同时得到扩增长度为448、549、375 bp 3条特异性片段,未扩增出猪、鸡支原体模板特异性片段;其敏感性(可检测到的最小模板DNA含量)为0.8 ng·μL-1;36份临床样品检测结果显示,三重PCR检测结果与分离培养鉴定方法一致,均能鉴定出牛支原体阳性病料.本研究建立的三重PCR诊断方法能够一次性鉴别3种支原体,具有高敏感性、特异性和准确性,可用于临床诊断和流行病学调查.     

Is the microagglutination test (MAT) good for predicting the infecting serogroup for leptospirosis in Brazil?

Leptospirosis is a zoonotic infection caused by pathogenic members of the genus Leptospira spp. Knowledge of the prevalent serovars and their maintenance hosts is essential to understand the disease. The aim of this study was to evaluate the ability of serology by the microscopic agglutination test (MAT) to predict the serogroups compared with results of identification of leptospires in São Paulo, Brazil. MAT correctly assigned the serogroup of the infecting isolate in 49/52 cases (94.23%). The serogroup Icterohaemorrhagiae was the predominant serogroup (88.46%). This study showed the usefulness of the MAT to correctly identify the infecting serogroup with a good overall agreement between the serologically-identified infecting serogroup and by identification of the isolate and can be used in epidemiological surveys in São Paulo. However, it should be complemented by the identification of Leptospira isolates.     

Potential novel markers to discriminate between active and latent tuberculosis infection in Chinese individuals

Latent tuberculosis infection (LTBI) constitutes the main reservoir for reactivation tuberculosis. The finding of potential biomarkers for differentiating between TB and LTBI is very necessary. In this study, the immunological characteristics and potential diagnostic utility of Rv2029c, Rv2628 and Rv1813c proteins were assessed. These three proteins stimulated PBMCs from ELISPOT-positive LTBI subjects produced higher levels of IFN-γ in comparison with TB patients and ELISPOT-negative healthy subjects (p < 0.05). BCG vaccination and non-TB respiratory disease had little influence on the immunological responses of Rv2029c and Rv2628 proteins (p > 0.05). The LTBI diagnostic performance of Rv2029c was higher than Rv2628 and Rv1813c by ROC evaluation. But Rv2628 had much higher specificity than Rv2029c in active TB patients and uninfected healthy subjects. The IgG level against Rv1813c was higher in the TB group than in LTBI and uninfected healthy subjects (p < 0.05). These results suggest that T cell response to Rv2628 and antibody against Rv1813c might be applicable as biomarkers to distinguish TB from LTBI and uninfected individuals.     

中草药微量元素的研究进展

文章概述了中草药微量元素研究的必要性以及中草药微量元素相关方面的研究,介绍了中草药及微量元素在养殖业中的应用,对存在问题和前景进行了探讨。     

鸭呼肠孤病毒TaqMan与SYBR Green荧光定量检测方法的建立及比较

本研究旨在针对新型鸭呼肠孤病毒(NDRV)建立两种快速、敏感、特异的荧光定量PCR诊断方法并对其临床实用性进行比较.试验根据呼肠孤病毒S1基因序列设计合成1对特异性引物和1条MGB探针,分别建立TaqMan和SYBR GreenI两种荧光定量PCR检测方法,并对两种方法的特异性、灵敏性、重复性进行比较.结果 显示,两种...     

黄翅绢丝野螟气味受体基因GcaeOR10的鉴定及组织表达分析

为明确黄翅绢丝野螟Glyphodes caesalis气味受体基因GcaeOR10(GenBank登录号MW701397)的生物信息学特征及组织表达谱,使用生物信息学软件对其进行序列分析,并通过实时荧光定量PCR(real-time fluorescence quantitative PCR,qPCR)技术检测其在黄翅绢丝野螟成虫不同组织中的表达量。结果显示,黄翅绢丝野螟GcaeOR10基因开放阅读框（open reading frame,ORF）全长为1 200 bp,编码399个氨基酸残基,预测其编码的蛋白质分子量为45.67 kD,等电点为8.65,无信号肽,具有昆虫气味受体家族7个跨膜蛋白（7 transmembrane proteins,7tm-6）超家族的保守结构域,存在26个潜在的磷酸化位点,含有7个跨膜结构域,且N端位于细胞膜内,C端位于细胞膜外。黄翅绢丝野螟GcaeOR10与桑螟Glyphodes pyloalis的GpylOR17亲缘关系最近,其次与松蛀螟Conogethes pinicolalis的CpinOR39、CpinOR36和桃蛀螟Conogethes p...     

赤拟谷盗PH_3抗性与CarE的关系研究

羧酸酯酶(Carboxylesterase,CarE)是昆虫体内代谢杀虫剂重要的水解酶之一。以重要储藏物害虫赤拟谷盗为供试对象,利用微量滴度酶标板法比较研究了PH3抗性与敏感品系成虫、幼虫、蛹阶段CarE的活力及动力学参数等生化指标。结果表明,各供试虫态中,赤拟谷盗各品系成虫期的CarE比活力均最高。与敏感品系相比,赤拟谷盗2个PH3抗性品系CarE的活力及比活力均低于敏感品系。进一步的动力学参数研究表明,各品系中,HSTC-R品系3个虫态的Km和Vmax值均最高,而HYTC-R品系的Km和Vmax值总体上也略高于敏感品系ABTC-S。Km在抗性品系中上升的事实表明CarE在抗性品系中结构发生了改变,这可能是该酶在抗性品系中水解能力下降的原因。     

重庆肉牛腹泻相关病毒的检测及遗传进化分析

为了解重庆市肉牛病毒性腹泻的病原流行情况,本研究对重庆市8个肉牛养殖场的81份腹泻粪便样本中牛病毒性腹泻-黏膜病病毒（bovine viral diarrhea virus,BVDV）、牛冠状病毒（bovine coronavirus,BCV）、牛轮状病毒（bovine rotavirus,BRV）和牛星状病毒（bovine astrovirus,BAstV）4种致腹泻病毒进行了RT-PCR检测,对PCR产物进行测序,用Mega 6.0软件进行系统发育分析。结果显示,BRV、BAstV和BVDV检出率分别为66.7%、8.6%和7.4%,BCV未检出。遗传进化分析结果表明,测序的5个BRV单独聚为一小支,与GenBank中其他VP6序列有明显的遗传距离;BVDV与中国株和丹麦株聚为一支,遗传关系最近;5个BAstV单独聚为一支,与中国香港株遗传关系最近,但仍有明显的遗传距离。本试验结果表明,重庆地区肉牛腹泻主要发生在6月龄以下犊牛,BRV是该地区肉牛腹泻的重要原因,BRV、BAstV和BVDV 3种病毒的遗传多样性值得进一步关注。