极速实时荧光聚合酶链反应与常规方法对新型布尼亚病毒检测的评价

目的探讨极速实时荧光聚合酶链反应(polymerase chain reaction,PCR)、实时荧光PCR、酶联免疫吸附测定(enzyme-linked immunosorbent assay,ELISA)和胶体金免疫层析法(gold immunochromatography assay,GICA)4种方法检测新型布尼亚病毒的特异度和灵敏度,为发热伴血小板减少综合征的早期诊断提供依据。方法采集2017年6月1日至9月30日山东大学附属济南市传染病医院86例临床诊断为发热伴血小板减少综合征患者的血清样本,分别应用极速实时荧光PCR、实时荧光PCR、ELISA和GICA 4种方法进行检测。统计学分析采用χ^2检验。结果86份患者血清标本中,极速实时荧光PCR、实时荧光PCR、IgM-ELISA、IgG-ELISA、IgM-GICA、IgG-GICA的新型布尼亚病毒阳性分别为82份(95.34%)、79份(91.86%)、41份(47.67%)、8份(9.3%)、19份(22.09%)和3份(3.49%)。极速实时荧光PCR特异度为100%,灵敏度达到1×103拷贝/mL,3次重复扩增试验显示其Ct值变异系数均<2%。在发热伴血小板减少综合征进展的1期、2期、3期病程中,极速实时荧光PCR的阳性检出率为41份(97.62%)、34份(94.44%)、7份(87.50%),实时荧光PCR的阳性检出率为39份(92.86%)、33份(91.67%)、7份(87.50%),在1期和2期两个病程,极速实时荧光PCR阳性检出率略高;IgM-ELISA阳性检出率从1期(28.57%)到3期(87.50%)显著增高,2期、3期与1期相比,差异均有统计学意义(χ^2=8.347、7.561,均P<0.01);IgM-GICA的阳性检出率从1期(14.29%)到2期(33.33%)也有增高,差异有统计学意义(χ^2=3.962,P<0.05),但与其他方法相比,其检出率偏低。1期,实时荧光PCR阳性检出率显著高于ELISA(IgM和IgG)和GICA(IgM和IgG),差异均有统计学意义(χ^2=33.740、55.080、49.010、64.340,均P<0.01)。2期,实时荧光PCR的阳性检出率高于ELISA(IgM和IgG)和GICA(IgM和IgG),差异均有统计学意义(χ^2=7.700、46.720、23.700、50.630,均P<0.01)。3期,极速实时荧光PCR、实时荧光PCR和IgM-ELISA表现出同样高的阳性检出率,远高于IgG-ELISA和GICA(IgM和IgG)。实时荧光PCR阳性检出率和IgG-ELISA、IgM-GICA、IgG-GICA之间差异均有统计学意义(均χ^2=6.250,P<0.05)。结论极速实时荧光PCR在新型布尼亚病毒的早期检测中有更高的灵敏度和特异度,且重复性好、稳定度高,与传统实时荧光PCR相比大大缩短了扩增时间,对发热伴血小板减少综合征的早期快速诊断具有重要价值。     

△LuxS的表型分析及其在GBS毒力调控机制研究中的应用

目的采用LuxS缺失突变株模型对B型链球菌中数量感应相关分子LuxS的功能及与其相关的自诱导因子-2数量感应通路进行研究，以探索其毒力调控机制。方法采用RT-PCR法、菌落印迹分析、生长曲线测定、cAMP因子测定等方法对该缺失突变菌株的表型特性进行了研究。最后应用哈氏弧菌作为报告菌株，通过生物发光测定法分析了突变株对B型链球菌诱导报告菌株中的生物发光活性的影响。结果发现此缺失突变可导致B型链球菌中scpB基因表达的上调；与野生株相比，突变株的生物发光诱导活性比野生株降低了大约两倍。结论本研究结果证实了LuxS在GBS中AI-2数量感应通路中的重要性，并为GBS中毒力调控机制的进一步研究提供了新的线索。     

广州地区无偿献血人群人类疱疹病毒8型血清流行病学调查

目的：了解广州地区无偿献血者HHV-8的感染情况，为制定预防策略提供依据。方法：采用ELISA法检测3135名无偿献血者血浆HHV-8IgG抗体。结果：3135名献血者中6名血浆标本被检出HHV-8IgG抗体阳性，均为汉族、男性献血员，总阳性率为0．19％。不同年龄和性别组HHV-8感染率差异无统计学意义。结论：广州地区无偿献血人群HHV-8的感染率较低。     

Development and evaluation of a new PCR assay for detection of Pseudomonas aeruginosa D genotype

This report describes a new PCR-based assay for the detection of Pseudomonas aeruginosa genotype D in occupational saturation diving systems in the North Sea. This genotype has persisted in these systems for 11 years (1993-2003) and represents 18% of isolates from infections analysed during this period. The new PCR assay was based on sequences obtained after randomly amplified polymorphic DNA (RAPD)-PCR analysis of a group of isolates related to diving that had been identified previously by pulsed-field gel electrophoresis (PFGE). The primer set for the D genotype targets a gene that codes for a hypothetical class 4 protein in the P. aeruginosa PAO1 genome. A primer set able to detect P. aeruginosa at the species level was also designed, based on the 23S-5S rDNA spacer region. The two assays produced 382-bp and 192-bp amplicons, respectively. The PCR assay was evaluated by analysing 100 P. aeruginosa isolates related to diving, representing 28 PFGE genotypes, and 38 clinical and community P. aeruginosa isolates and strains from other species. The assay identified all of the genotype D isolates tested. Two additional diving-relevant genotypes (TP2 and TP27) were also identified, as well as three isolates of non-diving origin. It was concluded that the new PCR assay is a useful tool for early detection and prevention of infections with the D genotype.     

Digestive properties of the venom of the Australian Coastal Taipan, Oxyuranus scutellatus (Peters, 1867).

The digestive properties of Australian elapid snake venoms have not been studied to any great extent. To address this, the in vitro digestive properties of Oxyuranus scutellatus (Australian Coastal Taipan) venom were investigated in a simulation of the in vivo conditions using the parameters reported for the stomach of snakes and representative prey for this species. The amount of soluble protein released was measured over time using a bicinchoninic acid (BCA) assay. Dismembered mouse hindlegs were injected intramuscularly with 0.1 ml O. scutellatus venom (concentration 10 mg/ml) and maintained in a micro-anaerobic, acidic environment (pH approximately 1.2-1.7) at 25 degrees C. The bathing liquid was sampled every 24 h for 7 days, and assayed for soluble protein. Statistical analysis revealed that O. scutellatus venom increased the rate at which proteins were released when compared to a negative control suggesting the potential importance of envenomation in the digestion of whole prey.     

HPLC法测定小柴胡颗粒剂中黄芩苷的含量

目的 :建立小柴胡颗粒剂中黄芩苷的含量测定方法。方法 :采用反相高效液相色谱法。色谱柱 :Inertsil ODS- 3(5 μm,4 .6× 2 5 0 mm)柱 ;流动相 :甲醇 -水 -磷酸 (45∶ 5 5∶ 0 .2 ) ;检测波长 :315 nm。结果 :黄芩苷在 0 .2 4 2 9～ 1.2 14 4 μg/ml范围内呈现良好的线性关系 (r=0 .9999) ,回收率 10 0 .0 % ,RSD=0 .96 %。结论 :本法可用于测定小柴胡颗粒剂中黄芩苷的含量。     

急性脑血管病患者血清髓鞘碱性蛋白含量研究

用酶联免疫吸附法对３０例急性脑血管病（ＣＶＤ）患者及３２位正常人血清髓鞘碱性蛋白（ＭＢＰ）含量进行检测。结果表明：急性ＣＶＤ组患者血清ＭＢＰ含量显著高于正常人组（Ｐ＜０．０１）；血清ＭＢＰ含量与急性ＣＶＤ的严重程度相关。提示检测血清ＭＢＰ含量对急性ＣＶＤ诊断及预后判断有重要价值     

Allergen content in dust from homes and schools in northern Norway in relation to sensitization and allergy symptoms in schoolchildren

Background Previous studies have shown a high prevalence of atopic diseases among school children in the community of Sør-Varanger. Moreover, animal dander followed by pollen und house dust mite, were the most common allergens in skin prick tests. Objective To assess the allergen content in homes (living-rooms and mattresses) and classrooms of children living in an arctic area at 70° north. The presence of allergens in homes and schools and their relationship to atopy was of particular interest. Methods Dust samples from 38 homes and seven schools in northern Norway were collected by vacuum cleaning. The presence of allergens of dog, birch, timothy, Cladosporium herbanun, codfish and hen egg-white was investigated by radio-allergosorbent test (RAST) inhibition and the presence of major allergens of cat Felis domesticus (Fel d I) and house dust mites (HDM) Dermatophagoides pteronyssinus (Derp I) and Dermatophagoides farinae (Derf I) by enzyme-linked immunosorbent assay (ELISA). Results Mattresses contained significantly more dust per unit area than living-rooms and classrooms. No statistically significant differences in allergen content for dog. birch, timothy, Cladosporium, codfish and hen egg-white were seen between HDM-sensitized and non-atopic children. Most dust samples contained dog allergens with the highest allergenic activity found in living-rooms of those keeping dogs. An increased level of Feld I was detected in only one of 38 samples from living-rooms (this family kept a cat) and in 25 of 38 samples from mattresses with ranges from 24 to 84ng/m². The highest concentrations were found in mattresses of children keeping cats. Increased levels ( 25 ng/m²) of Derp I were found only in homes and virtually only in mattresses of HDM-sensitized children. An increased level of Derf] was found in only one case, i.e. in the mattress of an HDM-sensitized child where additionally Der p I and HDMs were demonstrated microscopically. When relating Der p I to HDMsensitization an odds ratio of more than 16 (95% Cl: 1.6–394.3) was found. All extracts from living-rooms included codfish allergens. Low RAST inhibition values were detected for hen egg-white, Cladosporium, birch and timothy pollen in most samples. Furthermore, the study demonstrated that dust from schools was relatively free of allergens. Conclusion Previous findings indicating that the main allergen exposure problem in this geographical area is that of pet allergens were confirmed.     

Effect of periodontal therapy on specific antibody responses to suspected periodontopathogens

The effects of clinically successful periodontal therapy were studied in juvenile periodontitis (JP) and rapidly progressive periodontitis (RP) patients and compared with periodontally healthy subjects (HS). Serum samples were obtained in 35 HS prior to the study and in 12 of these subjects 3-4 years later. Serum samples were obtained from 50 JP patients initially, 9 subjects immediately following surgical therapy and 29 of these subjects 3-4 years later. RP patients provided 46 initial serum samples, 9 following therapy and 27 samples 3-4 years later. Antibody levels were determined utilizing a standardized enzyme-linked immunosorbent assay with Bacteroides gingivalis, B. ochracea, Fusobacterium nucleatum and Actinobacillus actinomycetemcomitans serving as antigens. The JP patients showed an initial rise in antibody levels immediately following therapy followed by a significant decrease in antibody levels 3 to 4 years later. The RP patients did not show an early change in antibody levels but by 3 to 4 years post-therapy, antibody levels had significantly decreased. However, during this study, the antibody levels of JP and RP patients remained significantly higher when compared with HS patients.     

PCR和ELISA法在检测HBV-DNA、HCV-RNA和结核分枝杆菌中的作用

目的:探讨PCR技术和ELISA法在检测HBV-DNA、HCV-RNA和结核分枝杆菌中的作用,以及与直接涂片法的对比结果.方法:分别用两法对临床确诊的感染病例和非感染者同时进行检测,并进行对比分析.结果:HBeAg阳性者HBV-DNA阳性,检出率为96.8%,HCV-RNA阳性检出率为19.4%.不同标本TB-DNA平均阳性检出率为39.98%.结论:PCR法灵敏度和特异度分别高于ELISA法和涂片查菌法.