PCR与DIFA检测沙眼衣原体的比较研究

用所建立的沙眼衣原体质粒引物聚合酶链反应技术检测１１６例泌尿生殖道沙眼衣原体，并与直接免疫荧光法进行比较。结果ＰＣＲ阳性３８例，ＤＩＦＡ阳性的３２例中有３１例ＰＣＲ阳性。ＣＰＲ敏感性为９６．９％，特异性为９．１７％。ＰＣＲ阳性者于治疗结束１－２财后复诊２１例，其中４例ＰＣＲ仍然阳性。     

不同引物对PCR检测沙眼衣原体结果的影响

为了解不同引物对ＰＣＲ检测泌尿生殖道沙眼衣原体（ＣＴ）感染的影响,作者以直接免疫荧光法（ＤＩＦＡ）为标准,选用引物不同的市售ＰＣＲ试剂盒检测了２０９例泌尿生殖道可疑ＣＴ感染标本。结果引物１ＰＣＲ阳性２９例,引物２ＰＣＲ阳性２８例;在ＤＩＦＡ阳性２５例中,引物１ＰＣＲ阳性２４例,引物２ＰＣＲ阳性２３例;引物１ＰＣＲ敏感性为９６．０％,特异性为９７．８％;引物２ＰＣＲ敏感性为９２．０％,特异性为９８．     

聚合酶链反应检测泌尿生殖道沙眼衣原体

聚合酶链反应检测泌尿生殖道沙眼衣原体莒县皮肤病防治站（２７６５００）关继富临沂市人民医院王家银，关向宏，战雪梅，李雪梅沙眼衣原体（ｃｔ）所致的泌尿生殖道感染为非淋球菌性性传播疾病。我们采用聚合酶链反应（ＰＣＲ）技术对１０８例泌尿生殖道炎症患者的标本进...     

多聚酶链反应快速检测衣原体

应用沙眼衣原体内源性质粒引物和鹦鹉热衣原体１６ｓｒＲＮＡ基因引物，对沙眼衣原体眼部感染和尿道感染进行快速检测，４９份临床诊断为沙眼的标本经ＰＣＲ检测，阳性３１份，阴性１８份；２８份临床诊断为非淋菌性尿道炎的标本，经ＰＣＲ检测，阳性１１份；阴性１７份。以上结果分别与国际公认的免疫荧光试剂盒和酶联免疫试剂盒对同一标本的检测结果进行了比较，表明多聚酶链反应技术远较ＩＦＡ和ＥＬＩＳＡ灵敏，是快速检测沙眼衣     

衣原体感染

抗生素联用丙种球蛋白治疗泌尿生殖道沙眼衣原体感染178例疗效观察——韩武红等（湖北孝感市中心医院432100）；《中国男科学杂志》，2006，20（5）：40—41[目的：探索泌尿生殖道沙眼衣原体（CT）感染满意的治疗途径。方法：对178例泌尿生殖道沙眼衣原体患者进行随机分组治疗，A组91例只用抗生素治疗，B组87例用抗生素加丙种球蛋白治疗。结果：A组有效率84．6％，B组有效率94．3％，两组有效率有显著差别（P〈0．05）。结论：抗生素加丙种球蛋白治疗泌尿生殖道沙眼衣原体感染有一定的辅助治疗作用。]     

衣原体感染

应用超高倍多媒体显微仪检测泌尿生殖道沙眼衣原体——倪少娟等（广西南宁广西壮族自治区妇幼保健院检验科530003）；《中国医疗器械杂志》，2005，29（4）：308．309[应用超高倍多媒体显微仪直接镜检并动态观察衣原体，阳性标本及随机抽取的20例阴性标本与VIDAS CHL法进行比较。镜检法在365例泌尿生殖道标本中检出沙眼衣原体32例；52例标本（镜检法阳性32例，阴性20例）两种检测方法结果差异无统计学意义，P〉0．05。超高倍多媒体显微仪直接镜检法可用于泌尿生殖道沙眼衣原体感染的诊断。]     

热启动多聚酶链反应快速检测泌尿生殖道沙眼衣原体的应用…

应用热启动多聚酶链反应技术，对泌尿生殖道沙眼衣原体感染进行快速检测。共检测８０６例泌尿生殖道炎症病人，其中ＳＴＤ病人４０４例，阳性１０５例，阳性率２６．０％，普通病人４０２例，阳性１０例，阳性率２．５％，经统计学估计，福州市ＳＴＤ患者沙眼衣原体感染率范围为２１．７￣３０．３％。     

幽门螺杆菌感染对胃上皮细胞增殖和凋亡的影响

目的为了探讨幽门螺杆菌感染对胃粘膜上皮细胞动力学的影响。方法应用免疫组织化学和切口末端标记法（ＴＵＮＥＬ），检测了１６例正常胃粘膜者和３１例幽门螺杆菌（Ｈｐ）相关慢性胃炎患者治疗前后胃粘膜上皮细胞增殖细胞核抗原（ＰＣＮＡ）标记指数（ＬＩ％）、细胞凋亡指数（ＡＩ）和表皮生长因子受体（ＥＧＦ－Ｒ）的表达。结果Ｈｐ阳性患者的ＰＣＮＡＬＩ％为１３．９４±１．６４，正常对照组为６．７１±０．９２，差异有非常显著性（Ｐ＜０．０１）；ＥＧＦ－Ｒ表达与ＰＣＮＡＬＩ％呈正相关（ｒ＝０．４４８７，Ｐ＜０．０１）：Ｈｐ阳性患者组的ＡＩ为７．１±１．６，正常对照组为１．３±０．６，差异有非常显著性（Ｐ＜０．０１）；抗Ｈｐ治疗后，２１例Ｈｐ根除者的ＰＣＮＡＬＩ％和细胞ＡＩ分别降至８．２１±１．３２和１．２±０．６，与治疗前相比差异有非常显著性（Ｐ＜０．０１），而１０例Ｈｐ持续阳性者则无明显降低（Ｐ＞０．０５）：ＰＣＮＡＬＩ％、ＥＧＦ－Ｒ表达及细胞ＡＩ与胃粘膜炎症程度无显著相关（Ｐ＞０．０５）。结论上述结果提示，Ｈｐ感Ｈ能引起胃粘膜上皮细胞过度增殖和凋亡。这为Ｈｐ感染胃癌发病中的作用机制提供了一些线索。     

PCR检测淋病患者合并沙眼衣原体、解脲支原体感染的初步探讨

ＰＣＲ检测淋病患者合并沙眼衣原体、解脲支原体感染的初步探讨李凌武建浙江省杭州市第一人民医院（３１０００６）为了探讨淋病患者合并非淋球菌性尿道炎（ＮＧＵ）的感染情况，我们对在本科确诊为淋病的患者用聚合酶联反应对泌尿生殖道沙眼衣原体（ＣＴ）和解脲支原体（...     

热启动多聚酶链反应快速检测泌尿生殖道沙眼衣原体的应用研究

应用热启动多聚酶链反应技术,对泌尿生殖道沙眼衣原体感染进行快速检测。共检测８０６例泌尿生殖道炎症病人,其中ＳＴＤ病人４０４例,阳性１０５例,阳性率２６．０％;普通病人４０２例,阳性１０例,阳性率２．５％。经统计学估计,福州市ＳＴＤ患者沙眼衣原体感染率范围为２１．７～３０．３％。     

不同引物对PCR检测沙眼衣原体结果的影响

为了解不同引物对PCR检测泌尿生殖道沙眼衣原体（CT）感染的影响,作者以直接免疫荧光法（DIFA）为标准,选用引物不同的市售PCR试剂盒检测了209例泌尿生殖道可疑CT感染标本。结果引物1RCR阳性29例,引物2PCR阳性28例;在DIFA阳性25例中,引物1PCR阳性24例,引物2PCR阳性23例;引物1PCR敏感性为96．0％,特异性为97．8％;引物2PCR敏感性为92．0％,特异性为98．4％。与DIFA比较,两者在统计学上无显著差异（P＞0．05）,提示不同引物对PCR检测CT感染无明显影响。     

应用斑点金免疫渗滤法诊断日本血吸虫病

目的：探索一种便捷的血吸虫病诊断方法。方法：在原已建立的ELISA法的基础上，应用3个针对日本血吸虫抗原的不同表位的单克隆抗体标记胶体金，建立检测日本血吸虫病患者血清中血吸虫抗原的斑点金免疫渗滤法。抗原与抗体通过渗滤在硝酸纤维薄膜上进行反应，数分钟用肉眼观察结果。结果：本法检测SEA的敏感度为16ng／ml，在69例慢性日本血吸虫病患者血清的检测中，阳性率为60．8％，特异性为95．2％；同时用dot－ELISA检测137例慢性日本血吸虫病患者，阳性率为54．7％、特异性为94．6％；夹心ELISA检测118例 慢性日本血吸虫病患者，阳性率为61．9％、特异性为95．7％。结论：经数理统计G检验证实斑点金免疫渗滤法的敏感性和特异性同ELISA 相近, 且操作更简便且快速。     

Chlamydia and Borrelia DNA in synovial fluid of patients with early undifferentiated oligoarthritis: results of a prospective study.

OBJECTIVE: More than 50% of patients with synovitis involving 1-4 joints remain classified as having undifferentiated oligoarthritis (UOA) after 1 year of disease. The clinical presentation is often similar to that of reactive arthritis (ReA) and other spondylarthropathies or to Lyme arthritis. We therefore determined how often Chlamydia trachomatis (Ct) and Borrelia burgdorferi (Bb) can be identified in patients with UOA, by using an extensive laboratory approach. METHODS: We prospectively studied 52 patients with UOA who presented at an early synovitis clinic in a region highly endemic for Lyme disease. Patients were examined by standardized clinical and immunoserologic procedures. Synovial fluid was screened for the presence of Ct and Bb DNA by polymerase chain reaction (PCR). Urine was tested for Ct DNA by ligase chain reaction, and serum was tested for Ct antibodies by enzyme-linked immunosorbent assay and Bb antibodies by hemagglutination test and Western blotting. PCR results in the UOA patients were compared with the results in cohorts of patients with definite rheumatoid arthritis (RA), Lyme arthritis, and Chlamydia-induced arthritis (CIA). RESULTS: In the synovial fluid of 9 of 52 patients with UOA (17%), we found Ct DNA, and in 6 of the 52 patients (12%), Bb DNA was found. The frequency of bacteria-specific DNA was 50% (7 of 14) in CIA patients and 69% (11 of 16) in patients with Lyme arthritis. No Bb or Ct DNA was found in the synovial fluid of the 31 RA patients. CONCLUSION: With optimized PCR protocols, it is possible to detect considerable levels of Bb and Ct DNA in the synovial fluid of patients with UOA. Although the presence of bacterial DNA does not unequivocally prove its etiologic significance, we suggest that at least one-third of patients with UOA may have a form of ReA that involves asymptomatic primary infection.     

用DIFA 检测肿瘤患者的弓形虫抗体

目的: 为了解湖北肿瘤患者弓形虫感染概况并探索新技术直接免疫固相凝集试验(DIFA)等3项试验联合筛选诊断弓形虫感染的意义。方法: 使用DIFA检测100 例肿瘤患者的弓形虫IgG 抗体, 阳性者用IgG-IFAT 复查, 再用Toxo-IgG/IgA/IgM-ISAGA 检测。结果: DIFA 检测阳性率为12% (12/100), IgG-IFAT 检测符合率为100%; IgA 可疑1 例; IgM 抗体阳性4 例, 可疑1 例, 其中2 例提示为活动性感染。合并弓形虫感染的12 例分布于肺癌、非何杰金氏淋巴瘤和乳腺癌患者中, 其感染率分别为受检人数的22.2% (6/27)、19.3% (5/26) 及7.1% (1/14)。结论: DIFA 的敏感性和特异性均较佳, 3 种方法的联合应用适宜作弓形虫感染的常规筛选诊断。提示免疫功能降低的肿瘤患者, 特别是肺癌和非何杰金氏淋巴瘤患者容易感染弓形虫病或促使潜伏的弓形虫病活动。     

孕妇与胎儿弓形虫感染状况的研究

目的　探讨孕妇与胎儿弓形虫 (Toxoplasma,TOX)感染的状况与关系。　方法　用酶联免疫吸附法及荧光定量多聚酶链反应技术检测孕妇血中 TOX特异性抗体及 TOX DNA,对 TOX感染孕妇检测羊水或脐血诊断胎儿感染。　结果　 15 6 4例孕妇中血清 TOX- Ig M阳性者 4 1例 (2 .6 2 % ) ,TOX- Ig G阳性 5 9例 (3.77% ) ,TOX DNA阳性 4 5例(2 .88% ) ,4 1例 TOX- Ig M阳性者其 TOX- DNA均为阳性。 4 5例 TOX感染孕妇中羊水或脐血 TOX- DNA阳性 13例(2 8.89% )。 TOX感染胎儿中 2例自然流产 ,1例死胎、1例胎儿生长受限 (Fetal growth restriction,FGR) ,9例出生时无明显症状的新生儿中 ,有 1例生后 1个月患黄疸性肝炎 ,1例有单侧耳聋 ,7例生长发育正常。　结论　孕妇 TOX感染可危害胎儿 ,孕妇 TOX感染后取羊水或脐血检测 TOX DNA是诊断胎儿 TOX感染的有效方法。     

一种莱姆病螺旋体real-time PCR方法的建立及其在鼠标本检测中的应用评价

目的 基于莱姆病螺旋体recA 基因,建立一种检测鼠中莱姆病螺旋体的real-time PCR方法。方法 通过GenBank分析比较莱姆病螺旋体recA 基因,选择其保守序列设计MGB探针及引物并进行方法学评估。并应用建立的real-time PCR方法和nested PCR方法对收集的123份鼠标本进行检测分析。结果 本研究建立的real-time PCR方法仅对莱姆病螺旋体检测阳性,其最小检出浓度为10¹ copies/μL。标准曲线各浓度点Ct值批内、批间平均变异系数(CV)分别为1.56%和2.30%。123份鼠标本中,real-time PCR检测59例阳性,nested PCR检测43例阳性。结论 新建立的real-time PCR方法具有快速、敏感和特异的优点,可用于鼠标本中莱姆病螺旋体的检测。     

实时荧光定量PCR检测布鲁杆菌方法的应用

目的 探讨实时荧光定量PCR(FQ-PCR)用于布鲁杆菌检测的可行性.方法 根据布鲁杆菌BCSP31基因设计合成1对引物和TaqMan探针,以克隆有布鲁杆菌基因片段的PMD18-T质粒作为标准品DNA模板,进行FQ-PCR检测,绘制标准曲线;对所应用的方法分别进行敏感性、特异性及重复性分析;并对临床血液标本进行FQ-PCR,与临床诊断进行比较.结果 用标准品DNA建立的定量标准曲线的循环阈值(Ct)与模板拷贝数呈良好的线性关系(r=0.999);FQ-PCR检测的灵敏度拷贝数为5个/μl,普通PCR为5×102个/μl,FQ-PCR是普通PCR的100倍;用FQ-PCR检测6株布鲁杆菌菌株及5株非布鲁杆菌菌株,布鲁杆菌均检出较强荧光信号,非布鲁杆菌未检出;5×103个拷贝/μl标准品DNA检测后,Ct值批内变异系数(CV)为0.71%,批间CV为7.23%;用FQ-PCR检测临床确诊阳性血标本24份,检出阳性19份,符合率为79%(19/24),阴性对照血标本31份,全部阴性,阴性符合率为100%(31/31),临床症状可疑标本30份,检出阳性2份.结论 用FQ-PCR方法检测布鲁杆菌具有高度敏感性、特异性及良好的重复性,与临床阳性及阴性标本的符合率较高,可用于快速检测各种样本中的微量布鲁杆菌以及作为布鲁杆菌感染的实验室检测方法.     

Chlamydia and Borrelia DNA in synovial fluid of patients with early undifferentiated oligoarthritis: Results of a prospective study

Objective

More than 50% of patients with synovitis involving 1–4 joints remain classified as having undifferentiated oligoarthritis (UOA) after 1 year of disease. The clinical presentation is often similar to that of reactive arthritis (ReA) and other spondylarthropathies or to Lyme arthritis. We therefore determined how often Chlamydia trachomatis (Ct) and Borrelia burgdorferi (Bb) can be identified in patients with UOA, by using an extensive laboratory approach.

Methods

We prospectively studied 52 patients with UOA who presented at an early synovitis clinic in a region highly endemic for Lyme disease. Patients were examined by standardized clinical and immunoserologic procedures. Synovial fluid was screened for the presence of Ct and Bb DNA by polymerase chain reaction (PCR). Urine was tested for Ct DNA by ligase chain reaction, and serum was tested for Ct antibodies by enzyme‐linked immunosorbent assay and Bb antibodies by hemagglutination test and Western blotting. PCR results in the UOA patients were compared with the results in cohorts of patients with definite rheumatoid arthritis (RA), Lyme arthritis, and Chlamydia‐induced arthritis (CIA).

Results

In the synovial fluid of 9 of 52 patients with UOA (17%), we found Ct DNA, and in 6 of the 52 patients (12%), Bb DNA was found. The frequency of bacteria‐specific DNA was 50% (7 of 14) in CIA patients and 69% (11 of 16) in patients with Lyme arthritis. No Bb or Ct DNA was found in the synovial fluid of the 31 RA patients.

Conclusion

With optimized PCR protocols, it is possible to detect considerable levels of Bb and Ct DNA in the synovial fluid of patients with UOA. Although the presence of bacterial DNA does not unequivocally prove its etiologic significance, we suggest that at least one‐third of patients with UOA may have a form of ReA that involves asymptomatic primary infection.

     

PCR技术检测石蜡包埋肝癌、癌旁组织中HBV DNA及与血清学标记之间关系

本文应用先进的聚合酶链反应(PCR),检测了27例原发性肝癌（HCC）患者肝癌及癌旁双份组织中的乙肝病毒DNA（HBV DNA），并探讨了与血清学标记物（HBV－M）之间的关系。检测结果表明：癌组织中HBV DNA检出率较癌旁组织中检出率低，且差异有统计学意义（P＜0．05）。HBsAg阳性合并HBeAg阳性者检出HBV DNA最高，癌旁组织中慢活肝中HBV DNA检出率最高。     

Nationwide Harmonization Effort for Semi-Quantitative Reporting of SARS-CoV-2 PCR Test Results in Belgium

From early 2020, a high demand for SARS-CoV-2 tests was driven by several testing indications, including asymptomatic cases, resulting in the massive roll-out of PCR assays to combat the pandemic. Considering the dynamic of viral shedding during the course of infection, the demand to report cycle threshold (Ct) values rapidly emerged. As Ct values can be affected by a number of factors, we considered that harmonization of semi-quantitative PCR results across laboratories would avoid potential divergent interpretations, particularly in the absence of clinical or serological information. A proposal to harmonize reporting of test results was drafted by the National Reference Centre (NRC) UZ/KU Leuven, distinguishing four categories of positivity based on RNA copies/mL. Pre-quantified control material was shipped to 124 laboratories with instructions to setup a standard curve to define thresholds per assay. For each assay, the mean Ct value and corresponding standard deviation was calculated per target gene, for the three concentrations (10⁷, 10⁵ and 10³ copies/mL) that determine the classification. The results of 17 assays are summarized. This harmonization effort allowed to ensure that all Belgian laboratories would report positive PCR results in the same semi-quantitative manner to clinicians and to the national database which feeds contact tracing interventions.