金黄色葡萄球菌一氧化氮合酶生理功能分析

为了研究金黄色葡萄球菌一氧化氮合酶的功能和作用,克隆该基因的全长序列,在一氧化氮合酶基因缺陷菌株的基础上,构建一氧化氮合酶过表达菌株,进行表型验证,研究NO对细菌表型的影响。结果显示:一氧化氮合酶缺失使细菌自溶速率加快,抗氧化杀伤能力减弱,生物被膜生成能力降低、缺失和过表达均会影响细菌正常的生长周期和速率。外源性NO对野生菌株、敲除菌株和过表达菌株生物被膜有不同影响显示,金黄色葡萄球菌一氧化氮合酶还参与了外源性NO对生物被膜的调控。一系列表型结果表明:金黄色葡萄球菌内源性NO作用不仅仅阻断Fenton反应、增强过氧化氢酶活性,还可以作为一种信号分子调控细菌基因的转录和表达。     

H2S和NO在大白菜抵抗高温胁迫中的作用

结合药理学和植物生理生化的方法,通过外源施加供体或清除剂,并观察幼苗生长的变化,意在研究H_2S和NO两种气体信号分子在大白菜抵抗高温胁迫中的生理作用及相互影响。实验结果表明:高温胁迫明显抑制大白菜幼苗的生长,导致植株叶片明显萎蔫卷曲。胁迫条件下,植株可溶性糖、可溶性蛋白、脯氨酸含量以及相对电导率均显著升高,内源H_2S和NO含量明显上升,外源生理浓度H_2S和NO处理可明显缓解高温胁迫对幼苗生长的影响,并且两种信号分子存在相互作用,外源H_2S缓解内源NO含量的上升,而外源NO对内源H_2S含量变化并没有显著影响。综上,气体信号分子H_2S和NO交叉互作参与大白菜抵御高温胁迫。     

混合盐碱胁迫对柳树幼苗生理指标的影响

为研究混合盐碱胁迫对柳树幼苗叶片含水量、光合色素质量分数和抗氧化酶活性的影响,以盐柳1号(Salix psammophila‘Yanliu NO.1’)无性系幼苗为材料,对其进行12组不同浓度和组分的混合盐碱胁迫处理。研究结果表明:盐柳1号幼苗的叶片含水量下降的趋势随着盐浓度的增高和pH值的增大而明显加剧。B处理组的盐柳1号幼苗的叶片叶绿素a质量分数在盐浓度高于150 mmol/L时显著下降(P<0.05),A、C处理组的盐柳1号幼苗的叶片叶绿素a、叶绿素b质量分数均无显著变化(P>0.05)。在小于150 mmol/L的混合盐碱胁迫下,随着盐浓度的增高,各处理组盐柳1号幼苗的叶片SOD、POD、CAT活性增强或显著增强,在盐浓度高于150 mmol/L时,盐柳1号幼苗的叶片SOD、POD、CAT活性下降,其中,C处理组盐柳1号幼苗的叶片在盐浓度大于150 mmol/L时全部枯黄。综合各项结果,盐浓度在150 mmol/L以下,pH小于9.51时,盐柳1号幼苗具有较好的耐受能力,可以抵御一定的盐碱胁迫。     

拉萨市牦牛肉源金黄色葡萄球菌的鉴定及其耐药性分析

[目的]明确西藏拉萨市牦牛肉源金黄色葡萄球菌常见肠毒素(SEs)基因分布特征及其多重耐药性,为牦牛肉源产SEs金黄色葡萄球菌的快速鉴定及其防控提供科学依据.[方法]通过Baird-Parker显色培养筛选及Nuc基因扩增鉴定从拉萨市牦牛肉样品中分离获得金黄色葡萄球菌,利用PCR鉴定分析金黄色葡萄球菌中9种常见SEs基因的分布特征,并采用K-B纸片扩散法对产SEs金黄色葡萄球菌进行多重耐药性分析.[结果]从100份牦牛肉样品中分离获得29株金黄色葡萄球菌,其SEs基因鉴定结果表明,9种SEs基因检测出6种(SEB、SEC、SEE、SEG、SHE和SEI),有3种(SEA、SED和SEJ)未检出.其中,SEG基因检出率79.31%,SEC和SEI基因检出率均为68.97%,SEB基因检出率55.17%,SHE基因检出率51.72%,SEE基因检出率3.45%.29株金黄色葡萄球菌对青霉素(P)、氨苄西林(AMP)、克拉霉素(CLR)、红霉素(E)和磺胺甲恶唑(SMZ)的耐药率较高,分别为79.31%、65.52%、58.62%、51.73%和31.04%;对阿米卡星(AK)、替卡西林(TIM)、苯唑西林(OX)、环丙沙星(CIP)和四环素(TE)的耐药率较低,分别为10.35%、6.90%、3.45%、3.45%和3.45%;对头孢西丁(FOX)、头孢他啶(CAZ)、头孢噻肟(CTX)、庆大霉素(CN)和氯霉素(C)尚未产生耐药性.29株金黄色葡萄球菌共表现出13种耐药谱,多重耐药率高达93.1%(27/29),其优势耐药谱为AMP/P、SMZ/CLR/E和AMP/P/CLR/E.[结论]西藏拉萨市牦牛肉源金黄色葡萄球菌含有SEG、SEC、SEI、SEB、SEH和SEE等多种SEs基因,且对青霉素类、大环内酯类和磺胺类等多种抗菌药物已产生较强的耐药性.     

金黄色葡萄球菌蛋白A单克隆抗体的制备

为获得稳定性好、特异性强、效价较高的抗金黄色葡萄球菌蛋白A单克隆抗体,用热灭活的金黄色葡萄球菌wol-04菌体抗原免疫BALB/c小鼠,取免疫小鼠脾细胞与小鼠骨髓瘤细胞(SP2/0)融合,以金黄色葡萄球菌蛋白A(SPA)为筛选抗原进行克隆筛选,并对各株单抗的生物学特性进行分析和鉴定.结果表明:试验获得了3株稳定分泌SPA单克隆抗体的杂交瘤细胞,其中单克隆抗体3B11、4F7为SPA表面抗原表位的单克隆抗体,与金黄色葡萄球菌wol-04菌体和SPA有较强的免疫反应性,免疫球蛋白亚类分别为IgG1和IgG3,具有稳定性好、特异性强、效价较高的特点.     

牛肉中金黄色葡萄球菌的复核鉴定研究

[目的]采用2种方法复核检出牛肉中金黄色葡萄球菌阳性结果,确保检测结果的准确性。[方法]按照GB 4789.10—2010《食品微生物学检验金黄色葡萄球菌检验》方法前增菌、选择性分离培养、革兰氏染色和血浆凝固酶试验鉴定金黄色葡萄球菌,应用SN/T1870—2007《食品中致病菌检测方法实时PCR法》对疑似阳性结果进行了进一步验证和复核。[结果]经过2种方法验证可疑菌落,该批牛肉中检出金黄色葡萄球菌。[结论]分离并鉴定牛肉中金黄色葡萄球菌可疑样品时,可采用不同方法进行验证和复核,该方法可提高检测结果的准确性,可用于畜禽肉中致病菌的检测。     

四川兔源金黄色葡萄球菌毒力检测及分子分型

【目的】研究四川部分区域兔源金黄色葡萄球菌Staphylococcus aureus的基因型总体结构特征、遗传变异以及毒力因子的分布情况。【方法】从四川地区分离41株兔源金黄色葡萄球菌,鉴定fem B基因特异性,进行耐甲氧西林金黄色葡萄球菌(Methicillin-resistant S.aureus,MRSA)筛选,并通过PCR法检测13种常见的毒力基因,采用多位点序列分型(Multilocus sequence typing,MLST)和脉冲场凝胶电泳(Pulsed field gel electrophoresis,PFGE)确定基因型特征。【结果】41株金黄色葡萄球菌中共检测出MRSA 31株,检出率为75.61%;共检出9种毒力基因,其中nuc、hla、eta和clf A在所有菌株中均存在,而sea、sec、see、hlb和PVL的阳性检出率分别为9.7%、85.4%、80.5%、90.2%和7.3%。MLST分型结果显示,41株金黄色葡萄球菌只存在2种序列型(ST398、ST3320)和1个克隆群CC398,其中ST398为优势序列型,所占比例为97.6%。PFGE将41株金黄色葡萄球菌分为18个基因型,但不同区域间的基因型条带差异较小。【结论】四川调查区域兔源金黄色葡萄球菌毒力因子携带率较高,其对家兔的养殖业存在较大的安全威胁;分型分析说明四川部分区域金黄色葡萄球菌的主要流行菌株遗传变异程度小,菌株间亲缘关系较近。     

Toxic shock syndrome and lysogeny in Staphylococcus aureus

Lysogeny, or the presence of temperate bacteriophage, was demonstrated, by means of two Staphylococcus aureus indicator strains, in 11 of 12 strains of S. aureus isolated from patients with toxic shock syndrome. Only 1 of 18 strains of S. aureus that were not associated with toxic shock syndrome showed the presence of bacteriophage. A laboratory strain of S. aureus was lysogenized by bacteriophage from two of the toxic shock-associated strains. These results add support to the theory that lysogeny by one or more bacteriophage in certain strains of S. aureus may be responsible for the pathogenesis of toxic shock syndrome.     

Genetic characterization of antimicrobial resistance in Staphylococcus aureus isolated from bovine mastitis cases in Northwest China

Staphylococcus aureus is the most common etiological pathogen of bovine mastitis. The resistant strains make the disease difficult to cure. The aim of this study was to characterize the genetic nature of the antimicrobial resistance in S. aureus cultured from bovine mastitis in Northwest China in 2014. A total of 44 S. aureus were isolated for antimicrobial resistance and resistance-related genes. Antimicrobial resistance was determined by disc diffusion and the corresponding resistance genes were detected by PCR. Phenotype indicated that S. aureus isolates were resistant to penicillin(84.09%), erythromycin(20.45%), tetracycline(15.91%), gentamicin(9.09%), tobramycin(6.82%), kanamycin(6.82%) and methicillin(2.27%). 9.09% of the S. aureus isolates were classified as multidrug resistant. In addition, genotypes showed that the isolates were resistant to rifampicin(100%, rpo B), penicillin(95.45%, bla Z), tetracycline(22.73%, tet K, tet M, alone or in combination), erythromycin(22.73%, erm B or erm C), gentamicin/tobramycin/kanamycin(2.27%, aac A-aph D), methicillin(2.27%, mec A) and vancomycin(2.27%, van A). Resistance to tetracycline was attributed to the genes tet K and tet M(r=0.558, P 0.001). This study noted high-level geno- and phenotypic antimicrobial resistance in S. aureus isolates from bovine mastitis cases in Northwest China.     

白杨素对金黄色葡萄球菌肠毒素SEA和SEB表达的影响

为研究白杨素对金黄色葡萄球菌的抗菌活性以及该化合物对金黄色葡萄球菌肠毒素SEA和SEB表达的影响,应用肉汤微量稀释法检测了白杨素对金黄色葡萄球菌的最小抑菌浓度,采用TNF-α释放试验、蛋白免疫试验以及荧光定量PCR等方法分别从蛋白表型、蛋白水平和基因水平3个层面上考察了白杨素对金黄色葡萄球菌肠毒素SEA和SEB分泌的影响。结果表明:白杨素几乎无抗金黄色葡萄球菌活性,其MIC1 024μg/m L。低浓度白杨素(2~16μg/m L)即可抑制金黄色葡萄球菌肠毒素SEA和SEB的分泌,且这一抑制作用呈现剂量依赖性。     

牛角地黄汤对金葡菌外毒素分泌的影响及调控作用

牛角地黄汤是用于清热解毒、凉血化瘀的基础方剂,由水牛角、地黄、赤芍、丹皮等主要成分组成。【目的】本研究旨在探讨牛角地黄汤对金黄色葡萄球菌外毒素分泌的影响。【方法】将亚抑菌剂量的牛角地黄汤水煎液与金黄色葡萄球菌共同培养,用Western blotting检测培养液上清中外毒素蛋白的表达,用RT-q-PCR法检测细菌相关基因mRNA表达水平。【结果】牛角地黄汤水煎液在40.0mg/mL以上剂量对金黄色葡萄球菌的增殖有明显抑制作用。亚抑菌剂量的牛角地黄汤水煎液能抑制金葡菌毒素蛋白HLα、SEA、SPA的转录和蛋白分泌。同时抑制金葡菌agr系统和SarA调控系统相关基因的表达。【结论】牛角地黄汤在亚抑菌剂量下能抑制金葡菌毒力因子的转录和蛋白表达。其作用机制可能与牛角地黄汤剂抑制金葡菌agr二元调控系统及SarA蛋白家族中毒素相关的调控子转录表达有关。     

野菊花提取物抑菌活性

利用琼脂扩散法和琼脂平板稀释法分别测定野菊花提取物对供试菌的抑菌活性和最小抑菌浓度MIC(Minimal inhibitory concentration)。通过测定不同培养时间后培养液的OD560值。活菌数和电导率研究野菊花提取物对金黄色葡萄球菌的生长曲线的影响。结果表明,野菊花提取物原液对金黄色葡萄球菌、大肠杆菌和沙门氏菌的抑菌活性分别为(27.02±0.45)mm、(20.56±0.43)mm、(17.82±0.36)mm,MIC分别为原液浓度的12.5%(v/v)、25%(v/v)、50%(v/v)。与对照组相比,野菊花提取物作用于金黄色葡萄球菌后,菌体的生长繁殖受到影响,细胞渗透性改变,而且作用方式与麦迪霉素对金黄色葡萄球菌的作用方式存在差异。     

奶牛乳房炎金黄色葡萄球菌凝固酶基因型研究

 【目的】了解上海和贵州地区引起奶牛乳房炎的金黄色葡萄球菌凝固酶基因型情况,为奶牛乳房炎的防治提供理论依据。【方法】利用16S rRNA保守序列设计引物,PCR扩增鉴定金黄色葡萄球菌,并利用凝固酶基因及其酶切产物多态性对分离鉴定出的金黄色葡萄球菌进行了凝固酶基因分型。【结果】共鉴定出78株金黄色葡萄球菌,有74株金黄色葡萄球菌扩增凝固酶基因片段,分为5个基因型和两个亚型。PCR 1型为贵州地区优势基因型,PCR 3型为上海地区优势基因型。【结论】两地区各基因型菌株分布比例有显著的地域性差异,这与两地区地理环境和养殖水平差异对病原流行传播的影响有关。     

Genetic analysis of a high-level vancomycin-resistant isolate of Staphylococcus aureus

Vancomycin is usually reserved for treatment of serious infections, including those caused by multidrug-resistant Staphylococcus aureus. A clinical isolate of S. aureus with high-level resistance to vancomycin (minimal inhibitory concentration = 1024 microg/ml) was isolated in June 2002. This isolate harbored a 57.9-kilobase multiresistance conjugative plasmid within which Tn1546 (vanA) was integrated. Additional elements on the plasmid encoded resistance to trimethoprim (dfrA), beta-lactams (blaZ), aminoglycosides (aacA-aphD), and disinfectants (qacC). Genetic analyses suggest that the long-anticipated transfer of vancomycin resistance to a methicillin-resistant S. aureus occurred in vivo by interspecies transfer of Tn1546 from a co-isolate of Enterococcus faecalis.     

泰安郊区肉鸡金黄色葡萄球菌耐药性分析

对肉鸡金黄色葡萄球菌耐药性进行分析,结果表明,该菌株对中药黄连和抗生素头孢唑啉、替米考星药敏纸片表现为极度敏感;对中药黄芩、大青叶和抗生素恩诺沙星、氯雾素、卡那霉素、痢特灵药敏纸片高度敏感;对链霉素和环丙沙星药敏纸片中度敏感;对板蓝根、四环素、庆大霉素以及白头翁、红霉素、青霉素等表现为低度敏感或耐药。     

金黄色葡萄球菌TRAP基因的克隆与序列分析

参考Genbank已经发表的关于金黄色葡萄球菌TRAP蛋白的基因序列,设计合成1对引物,从本实验室保存的金黄色葡萄球菌Newman株中提取细菌DNA,对TRAP蛋白基因进行PCR扩增,产物经琼脂糖凝胶电泳分析,呈现1条约500bp的条带,回收纯化后,将其克隆至pMD18-T质粒载体中,进行核苷酸序列分析,然后与报道的TRAP蛋白基因进行比较。结果表明:所扩增的基因序列与报道的TRAP核苷酸的同源性高达100/,证实为金黄色葡萄球菌Newman株TRAP蛋白基因。     

海洋动物中微生物的分离与抗金黄色葡萄球菌活性筛选

为了从海洋环境中寻找新的抗金黄色葡萄球菌抗生素,分离并筛选了广西北海70个海洋动物表皮或肠道中的微生物.样品处理后,分别用M1、马丁和高氏培养基分离细菌、真菌及放线菌,以卡那霉素作标准曲线对分离菌株发酵上清测活.分离出250株细菌,64株真菌,6株放线菌.产生抗金黄色葡萄球菌活性物质的细菌为28株,占细菌总数的11.2 %;能产生抗金黄色葡萄球菌活性物质的真菌为12株,占真菌总数的18.8%;能产生抗金黄色葡萄球菌活性物质的放线菌为2株.实验证明海洋动物表皮及肠道存在丰富的抗金黄色葡萄球菌微生物,且部分未见报道.     

Detection and Analysis on Staphylococcal Enterotoxins in Raw Cow Milk from Different Regions of Jinan,China

[Objective] This study was conducted to investigate the pollution caused by staphylococcal enterotoxins (SE) in raw milk and the safety of dairy products in Jinan,and to provide a scientific basis for food safety risk analysis.[Method] A total of 130 raw milk samples were collected from different regions of Jinan,and detected for Staphylococcus aureus by referring to GB 4789.10-2010.Then,enzyme-linked immunosorbent assay was performed to detect whether the S.aureus strains produced enterotoxins,and the enterotoxin type was identified using colloidal gold-based immunochromatographic test strips.[Result] Fiftyseven of the raw milk samples were polluted by S.aureus,so the detection rate of S.aureus was 43.85％;and 11 of the strains produced enterotoxins.Among the 11 enterotoxin-producing strains,seven produced SEB,only one produced SEC,and the SE type of other three strains was not identified.[Conclusion] Enzyme-linked immunosorbent assay and colloidal gold-based immunochromatographic test strips can be used in combination to rapidly detect staphylococcal enterotoxins and identify enterotoxin type,although there are some limitations.SEB is the main type of staphylococcal enterotoxin causing pollution in milk of Shandong Province.     

Effect of Nitric Oxide on Alcoholic Fermentation and Qualities of Chinese Winter Jujube During Storage

This article investigates the effects of nitric oxide （NO） on alcoholic fermentation and the qualities of Chinese Winter Jujube during storage, and explores the action mechanisms of browning and softening of fruits to provide theoretical proofs for using NO in the storage of Chinese Winter Jujube. Chinese Winter Jujube fruits were fumigated with different concentrations of NO gas （0, 10, 20, 30 μL L^-1） under anaerobic conditions and stored at 22±1℃ and 4± 1℃. The changes in appearance qualities, the contents of pyruvate, ethanol, acetaldehyde, and the activities of alcohol dehydrogenase （ADH） and lactate dehydrogenase （LDH） were investigated. The contents of pyruvate, ethanol, and acetaldehyde were significantly reduced, and the peak of pyruvate content was delayed by 20 μL L^-1 NO. The activities of ADH and LDH in 20 μL L^-1 NO treated fruits were also significantly inhibited. However, the alcoholic fermentation and softening of Chinese Winter Jujube fruits were promoted by 30 μL L^-1 NO during storage. The results indicated that 20 μL L^-1 NO could mitigate the injury of ethanol on Chinese Winter Jujube and effectively delay the browning and softening of fruits during storage.     

Observations of ozone formation in power plant plumes and implications for ozone control strategies

Data taken in aircraft transects of emissions plumes from rural U.S. coal-fired power plants were used to confirm and quantify the nonlinear dependence of tropospheric ozone formation on plume NO(x) (NO plus NO(2)) concentration, which is determined by plant NO(x) emission rate and atmospheric dispersion. The ambient availability of reactive volatile organic compounds, principally biogenic isoprene, was also found to modulate ozone production rate and yield in these rural plumes. Differences of a factor of 2 or greater in plume ozone formation rates and yields as a function of NO(x) and volatile organic compound concentrations were consistently observed. These large differences suggest that consideration of power plant NO(x) emission rates and geographic locations in current and future U.S. ozone control strategies could substantially enhance the efficacy of NO(x) reductions from these sources.