肺炎支原体ELISA检测试剂盒的临床应用

目的评价肺炎支原体ELISA检测试剂盒在临床应用的效果。方法用肺炎支原体ELISA检测试剂盒检测呼吸道感染患儿的咽拭子标本,并以肺炎支原体快速检测培养基试剂做同步盲法对照试验,分析该试剂盒的准确性及批内、批间产品的稳定性。结果在100例呼吸道感染患儿咽拭子标本中,肺炎支原体ELISA检测法阳性率为38%,肺炎支原体快速检测培养基法阳性率为37%,两种方法阳性结果符合率为97%。同步盲法试验结果显示,肺炎支原体ELISA检测试剂盒批内、批间产品阳性结果的一致率均为100%。结论该试剂盒具有较好的准确度和特异性,并且操作简便、快速,临床可推广应用。     

2008-2009年南京地区儿童肺炎支原体感染情况分析

目的了解2008年至2009年南京地区儿童呼吸道肺炎支原体（MP）的感染情况。方法应用MP快速培养法对南京地区2008年1月至2009年12月980例急性呼吸道感染患儿的咽拭子标本进行MP培养检测。结果 980例急性呼吸道感染患儿的咽拭子标本中MP培养阳性384例,总阳性率为39.2%。其中〈3岁、35岁、〉5岁各年龄组患儿中MP阳性率分别是36.9%,38.5%和43.3%;不同季度MP感染的检出率分别是：春季（13月）45.7%,夏季（46月）24.8%,秋季（79月）20.4%,冬季（1012月）50.3%,其中冬、春两季MP感染的检出率明显高于其他两季（P〈0.01）。结论 MP为南京地区儿童急性呼吸道感染的重要病原体,各年龄儿童普遍易感;冬、春两季高发。     

咽拭子快速培养在肺炎支原体感染中的临床应用价值

目的:探讨咽拭子快速培养在肺炎支原体感染中的临床应用价值。方法:收集2014年2月~2016年2月期间我院收治的呼吸道感染患儿220例,用肺炎支原体专用液体培养基进行肺炎支原体快速培养,用胶体金法检测肺炎支原体MP-Ig M。比较两种方法的阳性率。结果:咽拭子培养快速培养阳性率与血清MP-Ig M检测阳性率比较,差异无统计学意义(P0.05)。MP-Ig检测显示,≤1岁阳性率最低,其阳性率随年龄增加不断增高(P0.05)。肺炎支原体咽拭子培养显示,≤1岁阳性率最高,2~8岁最低(P0.05)。病程≤7 d患者肺炎支原体咽拭子培养阳性率(34.21%)显著高于肺炎支原体MP-Ig检测阳性率(14.04%)(P0.05)。病程7 d患者肺炎支原体咽拭子培养阳性率(11.32%)显著低于肺炎支原体MP-Ig检测阳性率(52.83%)(P0.05)。肺炎支原体咽拭子培养的灵敏度性以及特异性显著高于肺炎支原体MP-Ig检测,差异具有统计学意义(P0.05)。结论:咽拭子快速培养对肺炎支原体感染的早期诊断有一定临床应用价值,方法简单,无创伤,值得临床进一步研究和应用。     

肺炎支原体P1重组蛋白的提取纯化及应用研究

提取纯化肺炎支原体(Mycoplasma pneumoniae,Mp)P1重组蛋白,建立酶联免疫吸附实验(ELISA)的方法,协助临床肺炎支原体感染的诊断。以GST融合蛋白层析柱提取、纯化Mp P1重组蛋白做抗原,以全肺炎支原体菌体成分做抗原对照,建立间接ELISA实验方法,检测40份正常献血者血清标本和51份疑似MP感染临床血清标本的IgG抗体。重组蛋白经SDS-PAGE可见诱导表达的样品在分子量大约59 ku处有明显条带,经Western blotting可与肺炎支原体免疫血清发生反应。ELISA实验检测51份临床标本,由P1重组蛋白抗原检测阳性31份,阳性率为60.78%。Mp检测阳性20份,阳性率为39.22%。实验精确度检测阳性混合血清的变异系数(CV值)为5.40%,阴性混合血清变异系数为1.10%。用Mp P1重组蛋白抗原建立的ELISA检测方法,其敏感性高于全肺炎支原体抗原,可用于临床肺炎支原体感染的诊断。     

肺炎支原体感染与小儿支气管哮喘急性发作的关系

目的:研究肺炎支原体(MP)感染与支气管哮喘急性发作的相关性。方法:选择我院2012年10月~2013年9月收治的支气管哮喘急性发作和缓解期患儿,作为观察组和对照组,分析两组患儿血清中MP特异性IgM(MP-IgM)和咽拭子MP-DNA的检测结果。结果:观察组MP-IgM和MP-DNA阳性率以及MP-DNA拷贝量均显著高于对照组(P0.05)。结论:MP感染与小儿支气管哮喘急性发作具有密切关系,MP-IgM和MP-DNA检测,可为临床诊疗提供一定的参考依据。     

杭州地区儿童肺炎支原体感染分析

目的调查杭州地区呼吸道感染儿童肺炎支原体（MP）感染情况和流行特点,为临床治疗和防止其爆发流行提供依据。方法随机采集在本院就诊并有呼吸道感染患儿的咽拭子,用FQ-PCR测定标本中MPDNA含量,若结果〉125拷贝为阳性,〈25拷贝为阴性。用统计学软件SPSS10．0分析MP感染和各调查因素之间的关系。结果在1502例呼吸道感染患儿标本中共检出391例MP阳性,阳性率占26．0％（391／1502）。感染机会、性别差异无显著性。7～15岁的儿童更容易感染,感染率占48．0％-62．1％。一年之中以夏秋季感染率最高,其中7月份高达37．3％。结论杭州地区MP引起儿童呼吸道感染的流行特点和南方其他地区的报道基本一致。     

肺炎支原体实验室检测方法研究进展

肺炎支原体(Mycoplosma pneumonia,MP)为人类非典型肺炎的病原体,是引起呼吸道感染的重要病原体。但是支原体肺炎与其他病原体感染的肺炎,在临床症状、影像学上并无特异性差别,且其对一般治疗肺炎、上呼吸道感染的药物有耐药性,因此肺炎支原体及时、准确的实验室检测对于支原体肺炎的诊断治疗显得尤为重要。目前MP的实验室检测方法不断推陈出新,但各种方法均有其优势与不足,临床可选择两种不同的方法同时检测。比如:血清学抗体的检测结合MP快速培养药敏的方法;血清学抗体的检测结合PCR的方法,不同方法相互补充为临床的早期诊断、治疗提供依据。而MP药敏试验的检测和耐药机制的研究对于临床用药方案的选择,减少耐药株的产生和流行具有重要意义。     

儿童肺炎支原体咽拭子快速液体培养结果分析

目的分析儿童肺炎支原体（MP）咽拭子快速液体培养的结果及与患儿年龄以及季节的关系,为临床诊断、治疗肺炎支原体感染提供依据。方法选取深圳市宝安区人民医院1217例急性呼吸道感染的患儿,采用咽拭子快速液体培养基进行筛查,观察其阳性率。结果在1217例患儿中,共检出阳性281例,阳性率为23．09％。〈1岁、1～3岁、3～6岁和6～14岁各年龄组的阳性率分别为15．13％、26．52％、28．31％和18．07％;不同季节MP感染率分别为春23．75％,夏20．11％,秋18．61％,冬29．72％。1～3岁,3—6岁小儿感染率明显高于6—14和〈1岁儿童（P〈0．01）;冬春季阳性率较夏秋季高。结论MP快速液体培养鉴定对MP诊断具有重要的临床诊断价值。肺炎支原体感染全年均可发病,好发于冬季,以3—6岁小儿多见。     

儿科呼吸道疾病中肺炎支原体感染的结果分析

目的探讨儿科肺炎支原体（MP）感染特点,辅助临床医师早期诊断,合理用药。方法测定我院一年来2013例儿科呼吸道疾病患儿的肺炎支原体抗体（IgM）。结果2013例呼吸道感染患儿,检出肺炎支原体抗体（IgM）阳性者769例,占38.2%,769例阳性分别表现为肺炎369例（48%）,支气管炎238例（31%）,咽炎92例（12%）,哮喘70例（9%）。其中,肺炎组与各组相比较,具有统计学意义。MP IgM感染的检出率明显高于其他各组（P〈0.01）。结论MP感染是患儿不可忽视的病原体,检测患儿血清MP抗体能够及早诊断,指导治疗。     

肺炎支原体和沙眼衣原体致儿童急性下呼吸道感染的研究

本文旨在研究儿童社区获得性急性下呼吸道感染(ALRTI)中肺炎支原体(MP)和沙眼衣原体(CT)的感染特征。采用实时荧光定量聚合酶链反应(PCR)检测2006年10月～2008年2月因ALRTI收入复旦大学附属儿科医院的患儿呼吸道标本中MP和CT的DNA,并分析2种病原体感染患儿的临床特征与实验室检查结果。在1312份深部鼻咽分泌物标本中,MP和CT检出率分别为7.85%(103/1312)和2.97%(39/1312)。MP在5岁以上患儿中的检出率为33.33%(30/90),而CT在3个月以内患儿中的检出率为6.28%(31/494)。MP感染后易出现40℃以上高热,较少发生发绀与重症呼吸道感染;白细胞计数常无明显升高,但C反应蛋白升高较多见。CT感染后40℃以上高热和重症呼吸道感染少见,C反应蛋白升高也较少见。结果提示,在5岁以上儿童的社区获得性ALRTI中,MP是重要的病原体;而在3个月以内儿童中,CT为常见病原体之一。     

PCR技术在鼠肺支原体检测中应用的初步研究

目的　探讨PCR技术在鼠肺支原体检测中的应用,希望能建立一种可行、快速、敏感的检测方法。方法　使用支原体通用引物及鼠肺支原体特异性引物对14 份大鼠喉气管拭子洗液和拭子支原体培养液进行PCR扩增,2 % 琼脂糖电泳鉴定。另设M53 和ATCC19612 二株标准鼠肺支原体菌株作阳性对照。结果　通用引物对大鼠喉气管拭子洗液检出率8/14 ,拭子支原体培养液检出率14/14,鼠肺支原体特异引物PCR扩增对大鼠喉气管拭子洗液检出率0/14 ,拭子支原体培养液3/14。通用引物扩增M53 和ATCC19612 二株标准株均呈现阳性,而鼠肺支原体特异引物扩增M53 和ATCC19612,只有M53 呈现阳性。结论　PCR通用引物检测比普通分离培养省时省力,而我们采用国外某学者认为对鼠肺支原体有特异性的引物,是否可用于鼠肺支原体的特异性PCR 检查仍需进一步探讨。     

Detection of Mycoplasma pneumoniae in simulated and true clinical throat swab specimens by nanorod array-surface-enhanced Raman spectroscopy

The prokaryote Mycoplasma pneumoniae is a major cause of respiratory disease in humans, accounting for 20% of all community-acquired pneumonia and the leading cause of pneumonia in older children and young adults. The limitations of existing options for mycoplasma diagnosis highlight a critical need for a new detection platform with high sensitivity, specificity, and expediency. Here we evaluated silver nanorod arrays (NA) as a biosensing platform for detection and differentiation of M. pneumoniae in culture and in spiked and true clinical throat swab samples by surface-enhanced Raman spectroscopy (SERS). Three M. pneumoniae strains were reproducibly differentiated by NA-SERS with 95%-100% specificity and 94-100% sensitivity, and with a lower detection limit exceeding standard PCR. Analysis of throat swab samples spiked with M. pneumoniae yielded detection in a complex, clinically relevant background with >90% accuracy and high sensitivity. In addition, NA-SERS correctly classified with >97% accuracy, ten true clinical throat swab samples previously established by real-time PCR and culture to be positive or negative for M. pneumoniae. Our findings suggest that the unique biochemical specificity of Raman spectroscopy, combined with reproducible spectral enhancement by silver NA, holds great promise as a superior platform for rapid and sensitive detection and identification of M. pneumoniae, with potential for point-of-care application.     

小儿肺炎支原体的分离培养及其药敏试验分析

对356例小儿肺炎患者的咽拭子进行肺炎支原体(MP)分离培养并对9种抗生素进行药敏试验.结果显示,MP阳性55例,阳性率为15.4%,耐药性由高到低为罗红霉素(54.5%)>克拉霉素(32.7%)=阿奇霉素(32.7%)>红霉素(21.8%)=加替沙星(21.8%)>克林霉素(20.0%)>乙酰螺旋霉素(9.1%)>司...     

Trypanosoma cruzi: parasite persistence in tissues in chronic chagasic Brazilian patients

Chagas disease in the chronic phase may develop into cardiac and/or digestive forms. The pathogenesis of the disease is not yet clear and studies have been carried out to elucidate the role of parasite persistence in affected organs. The aim of this study was to detect and quantify Trypanosoma cruzi in paraffin-embedded tissue samples from chronic patients using NPCR (nested polymerase chain reaction) and QPCR (quantitative polymerase chain reaction) methods. These results were correlated to anatomopathological alterations in the heart and gastrointestinal tract (GIT). Of the 23 patients studied, 18 presented the cardiac form and five presented the cardiodigestive form of Chagas disease. DNA samples were randomly isolated from formalin-fixed paraffin-embedded sections of heart and GIT tissue of 23 necropsies and were analyzed through NPCR amplification. T. cruzi DNA was detected by NPCR in 48/56 (85.7%) heart and 35/42 (83.3%) GIT samples from patients with the cardiac form. For patients with the cardiodigestive form, NPCR was positive in 12/14 (85.7%) heart and in 14/14 (100%) GIT samples. QPCR, with an efficiency of 97.6%, was performed in 13 samples (11 from cardiac and 2 from cardiodigestive form) identified previously as positive by NPCR. The number of T. cruzi copies was compared to heart weight and no statistical significance was observed. Additionally, we compared the number of copies in different tissues (both heart and GIT) in six samples from the cardiac form and two samples from the cardiodigestive form. The parasite load observed was proportionally higher in heart tissues from patients with the cardiac form. These results show that the presence of the parasite in tissues is essential to Chagas disease pathogenesis.     

Detection of influenza viruses in throat swab by using polymerase chain reaction

An assay protocol based on exploiting the polymerase chain reaction (PCR) for the direct detection of influenza virus in throat swab is described. By use of the mixture of H1 and H3 primers, it was possible to determine the subtype of the influenza A viruses simultaneously. No visible band was detected after PCR of influenza B or A (H2N2) viruses with a pair of H1 or H3 primers. The dilution experiment showed that the influenza viruses, as few as 1.3-6 plaque-forming units, were sufficient for detecting the HA gene by PCR. All throat swab samples from which influenza viruses had been isolated by conventional method were also positively detected by PCR method.     

荧光定量RT-PCR检测甲型H1N1病毒核酸的临床意义

目的观察荧光定量RT-PCR技术在检测甲型H1N1病毒核酸的临床意义。方法对135例经确诊为甲型H1N1的感染患者的咽式子采用荧光定量PCR技术检测H1N1病毒核酸,同时对40例健康体检者的咽式子做为对照组一同进行甲型H1N1病毒核酸检测。结果 135例确诊为甲型H1N1的患者经荧光定量PCR检测阳性129例,符合率为96.99%。40例健康体检者结果全阴性。结论荧光定量PCR检测甲型H1N1病毒核酸具有快速、特异性高等特点,在采用此方法诊断甲型H1N1时,阳性即可确诊,阴性者要结合临床。     

Evaluation of the NucliSens miniMAG RNA extraction and real-time NASBA applications for the detection of Mycoplasma pneumoniae and Chlamydophila pneumoniae in throat swabs

Nucleic acids were extracted from 215 throat swabs from patients with community-acquired pneumonia by the manual Boom extraction, the NucliSens miniMAG and the Qiagen DNA blood kit and amplified respectively by in-house real-time NASBA, NucliSens EasyQ reagents, and real-time PCR for the detection of Mycoplasma pneumoniae and Chlamydophila pneumoniae. Five out of 215 throat swabs were found to be C. pneumoniae positive by all techniques used. A total of 11 out of 215 throat swabs were positive for M. pneumoniae; 10/215 by Qiagen extraction and PCR amplification and 9/215 by NucliSens miniMAG and NucliSens EasyQ amplification. The NucliSens miniMAG and NucliSens EasyQ applications were successfully coupled to detect both organisms in throat swabs.