儿童呼吸道感染者肺炎支原体及肺炎衣原体特异性抗体检测临床意义

目的 探究儿童呼吸道感染者肺炎支原体及肺炎衣原体特异性抗体检测临床意义。方法 选择我院2018年1月~8月收治的85例呼吸道感染患儿设为研究组,另选择同期在我院进行健康体检的85例儿童设为对照组,对照组采用ELISA法检测肺炎支原体及衣原体,研究组在此基础上行咽拭子培养法检测,比较两组肺炎支原体、衣原体抗体检出率,ELISA法与咽拭子培养法对肺炎支原体、衣原体抗体的阳性检出率,分析研究组呼吸道感染性疾病中肺炎支原体、衣原体抗体阳性分布情况。结果 研究组患儿肺炎支原体抗体MP-IgM抗体、MP-IgA抗体、MP-IgG抗体及肺炎衣原体特异性抗体CP-IgM抗体、CP-IgG抗体的检出率均高于对照组儿童,差异有统计学意义（P＜0.05）;ELISA法对肺炎支原体特异性抗体阳性的检出率为54.12%高于咽拭子培养法的36.47%,组间对比具有统计学意义（P＜0.05）;ELISA法对肺炎衣原体特异性抗体阳性的检出率为58.82%高于咽拭子培养法的37.65%,差异有统计学意义（P＜0.05）;肺炎支原体、肺炎衣原体抗体阳性在各种儿童呼吸道感染性疾病中均有表现,其中,肺炎和支气管肺炎的阳性检出率较咽炎、扁桃体炎均较高。结论 肺炎支原体、肺炎衣原体是引发肺炎和支气管肺炎感染的主要病原菌。     

急性呼吸道感染患儿咽拭子标本的肺炎衣原体套式PCR检测研究

本文应用套式（Nested）PCR技术检测了132例急性呼吸道患儿之咽拭子标本中的肺炎衣原体（CPn）特异性DNA。结果发现．受检标本中有19例检出阳性，即各处呼吸道患儿之咽拭子标本中的CPn－DNA阳性率高达14．4％。提示CPn在我国儿童中有较高的感染率。     

肺炎支原体快速聚合酶链式反应（PCR）法检测

本文根据Ｂｅｒｎｅｉ设计的肺炎支原体ＰＣＲ引物，采用国产的耐热ＤＮＡ聚合酶，建立了双温循环肺炎支原体ＰＣＲ快速检测法。４０例儿童肺炎咽拭子标本中，８例呈阳性结果。     

用PCR技术检测喘息性疾病患儿肺炎支原体的研究

本文应用聚合酶链反应（ＰＣＲ）技术检测了１５６例喘息性疾病患儿咽拭子标本的肺炎支原体（ＭＰ），阳性率为１２．２％，对照组为同期３８例肺炎患儿的咽拭子标本，ＭＰ阳性率为１７．３％，二组ＭＰ感染率无显著性差异（ｘ２＝１．７７，Ｐ＞０．０５）。提示：ＭＰ不仅是肺炎也是喘息性疾病患儿常见的致病微生物。因此，对喘息性疾病患儿应常规用ＰＣＲ技术检测ＭＰ，以提高诊断和治疗水平     

反复呼吸道感染儿肺炎支原体PCR检测及免疫功能研究

本文对１５２例对复呼吸道感和肺炎支原体ＰＣＲ及免疫功能进行检测。结果咽拭子肺炎支原体ＰＣＲ阳性２５例，阳性率１６．５％。     

反复呼吸道感染患儿肺炎支原体PCR检测及免疫功能研究

本文对152例反复呼吸道感染患儿肺炎支原体PCR及免疫功能进行检测。结果咽拭子肺炎支原体PCR阳性25例，阳性率16．5％。血清IgG、IgA明显降低。CD3＋、CD4＋细胞、CD8＋细胞和CD4＋／CD8＋值均较对照组降低，而可溶性白细胞介素－2受体（SIL－2R）水平明显升高。资料表明反复呼吸道感染患儿存在着免疫功能紊乱。     

小儿肺炎支原体肺炎临床特点及治疗(附32例分析)

为了对小儿肺炎支原体做出快速、准确的诊断,实施最佳的治疗方案,我们对32例小儿肺炎支原体肺炎进行了血中肺炎支原体特异性IgM抗体的测定和咽拭子肺炎支原体培养,阳性者分别为24例,21例,治疗上先予红霉素静点,后改为琥乙红霉素口服,总疗程为14～21夭,显示满意的疗效。结果表明,特异性抗体测定和咽拭子培养有助于明确诊断,及时选用大环内酯类抗生素,降低了患儿住院天数,减少院内交叉感染的机率。     

新生儿肺炎患儿人型支原体套式PCR检测研究

本文应用套式ＰＣＲ检测了１５６例新生儿肺炎患儿鼻咽部分泌物中的人型支原体特异性ＤＮＡ，其结果：患儿阳性数１３０例，阳性率８３．３％，而应用人型支原体培养法及单式ＰＣＲ阳性率均比应用套式ＰＣＲ为低，提示了临床必须对人型支原体的感染率及致病性引起足够的重视。同时，通过本文研究也提示了临床对于不同的支原体感染可采用不同的抗生素治疗。目前国内对于人型支原体与新生儿肺炎的相关性报道尚未检索到。     

Th17/Treg失衡在肺炎支原体肺炎患儿中的作用及其机制

目的：探讨Th17/Treg失衡在肺炎支原体肺炎患儿中的作用及其机制。方法：纳入2013年6月至2015年1月我院收治的60例肺炎支原体肺炎急性期患儿和30例同期体检的健康儿童为研究对象。流式细胞术分析两组儿童外周血Th17和Treg的百分率,ELISA分析Th17和Treg特异性细胞因子IL-17和IL-10的分泌水平,定量PCR测定Th17和Treg特异性转录因子RORγt和Foxp3 mRNA的表达水平,以及Notch信号分子(Notch1,Hes1和Hey1)的表达水平。结果：与健康儿童相比,肺炎支原体肺炎急性期患儿外周血的Th17的百分率,IL-17分泌水平以及特异性转录因子RORγt的表达水平均明显增高,差异均具有统计学意义(P<0.05),而急性期患儿外周血的Treg的百分率,IL-10分泌水平以及特异性转录因子Foxp3的表达水平均较健康儿童明显降低,差异均具有统计学意义(P<0.05)。进一步分析显示急性期患儿Notch1、Hes1和Hey1的表达水平均明显高于健康儿童,其差异均具有统计学意义(P<0.05)。结论：Th17/Treg失衡参与儿童支原体肺炎的发生发展,Notch信号通路可能通过调节免疫细胞分化和细胞因子分泌参与Th17/Treg失衡。     

肺炎衣原体及肺炎支原体临床血清学检测试剂的研发及应用

目的 通过实验获得工程重组肺炎支原体和肺炎衣原体的抗原,探讨其临床诊断价值;分析儿童肺炎支原体和肺炎衣原体的发病情况.方法 采集医院收治的社区获得性肺炎患儿的血标本进行肺炎支原体和肺炎衣原体检测,并通过PCR方法得到肺炎衣原体的特异性抗原Cpn-MOMP和Cpn-CPAF和肺炎支原体P1-C和P1-B基因片段,应用工程重组抗原进行肺炎衣原体抗原特异性及灵敏度检测和肺炎支原体的灵敏度实验.结果 肺炎支原体及肺炎衣原体感染的阳性率随着年龄的增长而增加,肺炎衣原体冬季发病率较高.PCR方法可以得到完整的Cpn-MOMP和Cpn-CPAF基因片段,且Cpn-MOMP重组抗原产量高特异性较好.重组肺炎支原体P1-C和P1-B抗原产量较满意.结论 肺炎支原体及肺炎衣原体感染阳性率逐年升高,通过重组的Cpn-MOMP和Cpn-CPAF抗原可用于肺炎衣原体感染的血清学检测,具有临床应用前景.     

Basic and clinical studies on a new method for detecting anti-Mycoplasma pneumoniae antibody, using high density particles

We developed a new method for detecting anti-Mycoplasma pneumoniae antibody, using high density particles (HDP). We characterized the HDP, and also studied the clinical applicability of the HDP agglutination method (HDPA). HDP were uniform, spherical particles, having a specific gravity of 2.0 and a mean particle volume of 18 fl. Adsorption of IgG by HDP was approximately 2 times that by polystyrene latex. The sensitized antigen to the HDP, extracted from the Mycoplasma pneumoniae (Mac strain) bound specifically to the anti-Mycoplasma pneumoniae antibody. With HDPA, the antibody was determined within 30 minutes, which was more rapid than with other routine methods such as complement fixation test, passive hemagglutination test and particle agglutination test (PA). HDPA showed excellent correlation with PA. However, the titers with HDPA were generally 2 to 5 tubes higher than that with PA. A follow-up study on 7 cases of mycoplasma pneumonia revealed remarkably high titers of anti-Mycoplasma pneumoniae antibodies in the acute phase of mycoplasma pneumonia by HDPA, compared to that by PA. HDPA is highly sensitive and specific test for detection of anti-Mycoplasma pneumoniae antibody, and useful for rapid diagnosis of mycoplasma pneumonia.     

Expression of Mycoplasma pneumoniae antigens in Escherichia coli.

A genomic library of Mycoplasma pneumoniae was generated by using bacteriophage lambda EMBL3 as the vector. Screening of the library for the expression of M. pneumoniae protein antigens with adsorbed anti-M. pneumoniae serum revealed strong reactivity from a third of those clones which contained mycoplasma DNA inserts. Three of the most highly reactive clones were analyzed in detail and found to synthesize discrete mycoplasma proteins. Two carried overlapping fragments of mycoplasma DNA which encoded a protein that was readily detected in Escherichia coli after infection with recombinant bacteriophage. The third clone contained a novel mycoplasma DNA fragment which directed the synthesis of two additional mycoplasma proteins. Further screening of the library with antiserum raised against the major M. pneumoniae adhesin protein P1 (165 kilodaltons [kDa]) yielded one clone which produced an immunologically reactive protein of 140 kDa. Adsorption of anti-P1 serum by this clone selected a population of antibodies that were reactive with M. pneumoniae adhesin P1 (165 kDa). These results demonstrate that immunologically active M. pneumoniae proteins are synthesized in E. coli.     

肺炎支原体抗体检测方法比较及临床意义

目的：探讨肺炎支原体（MP）早期诊断的实验室检测方法及血清肺炎支原体抗体滴度在判断儿童肺炎支原体感染病情中的临床价值。方法：对586例疑似支原体感染的患儿同时进行明胶颗粒法、酶联免疫吸附法检测肺炎支原体,评估两种方法的应用价值。回顾性分析374例肺炎支原体感染患儿的临床资料。用双变量Spearman等级相关分析研究应用间接凝集试验法检测所得的血清MP抗体的滴度与MP感染患儿病情的关系。结果：明胶颗粒法和ELISA间接法在检测MP抗体中差异无显著性。374例MP感染患儿中,轻度MP感染175例（46.8%）,中度感染104例（27.8%）,重度感染95例（25.4%）;MP抗体滴度1∶80有140例（37.4%）,滴度1∶160有78例（20.9%）,滴度1∶320有58例（15.5%）,滴度1∶640有29例（7.8%）。滴度1∶1280有69例（18.4%）,经统计学分析,血清MP抗体滴度与MP感染患儿的病情严重程度呈正相关（r=0.689,P〈0.01）。结论：明胶颗粒法更能精确测定抗体滴度,敏感度高,操作简单,MP抗体滴度的测定有助于判断患儿的病情,为临床采取有效的治疗方案提供依据,可在临床推广使用。     

Biological effects of anti-lipid and anti-protein monoclonal antibodies on Mycoplasma pneumoniae.

Monoclonal antibodies directed against Mycoplasma pneumoniae surface components were examined for their ability to block mycoplasma attachment to chicken erythrocytes. Purified preparations of antibodies which recognize the major mycoplasma ligand mediating cytadherence (protein P1, 165 kilodaltons) inhibited attachment by more than 85% of the control values. Monoclonal antibodies reactive with two other surface proteins of 110 and 32 kilodaltons also blocked attachment. Surprisingly, monoclonal antibodies specific for M. pneumoniae lipids (J. Morrison-Plummer, D. H. Jones, and J. B. Baseman, J. Immunol. Methods 64:165-178, 1983) enhanced mycoplasma-erythrocyte binding. All antibodies examined had no effect on thymidine incorporation by M. pneumoniae.     

Detection of autoantibody to mitotic spindle associated antigen (MSA) in sera from patients with positive mycoplasma pneumoniae particle agglutination (PA) test]

Autoantibody to mitotic spindle associated antigen (MSA) was examined in sera from patients with positive mycoplasma pneumoniae particle agglutination (PA) test along with various autoimmune diseases and healthy subjects by means of indirect immunofluorescence (IF) using HEp-2 cells as substrates. Anti-MSA antibody was identified in 9 patients sera including 7 out of 180 sera with positive mycoplasma pneumoniae PA test, one with systemic lupus erythematosus (SLE) and one with scleroderma (PSS) respectively. Five of 9 sera with positive anti-MSA antibody contained anti-nuclear antibody (ANA) including 4 with speckled staining ANA and one with homogeneous + nucleolar staining ANA. Two collagen diseases sera positive for anti-MSA antibody showed negative mycoplasma pneumoniae PA test. In addition, there was no correlation between the presence of anti-MSA antibody and the titers of mycoplasma pneumoniae PA test. Our results indicated that anti-MSA antibody was not limited to such connective tissue diseases as previously described by other workers. It would be speculated that mycoplasma pneumoniae infection might induce anti-MSA antibody production.     

阿奇霉素对肺炎支原体肺炎的疗效及患儿体液免疫功能的影响

目的：探讨阿奇霉素治疗肺炎支原体肺炎患儿的效果及其对患儿体液免疫功能的调节作用。方法：118例肺炎支原体肺炎患儿均常规退热、止咳、平喘和对症治疗，并随机分为实验组和对照组，前者采用阿奇霉素治疗，后者采用红霉素治疗。比较两组治疗有效率、副反应发生情况及两组患儿治疗前后血清IgM、IgG、IgA、补体C3和C4水平。结果：用药14d后，实验组和对照组的总有效率依次为96．55％和总有效为80％，副反应发生率依次为20．69％和60％，差异均有显著性（P〈0．01）。实验组治疗后的IgG、州、C3和C4水平均低于治疗前，差异有统计学意义（P〈0．01～0．05）。结论：阿奇霉素治疗肺炎支原体肺炎效果较好，并可以改善机体的体液免疫水平。     

Kinetics of cellular and humoral response of rabbits immunized with mycoplasma pneumoniae.

The cellular and humoral responses of rabbits immunized with Mycoplasms pneumoniae antigen in saline or incorporated in Freund's complete adjuvant were examined. Peripheral blood leukocytes were used for the leukocyte migration inhibition (LMI) test. Both groups of animals showed significant LMI activity in the presence of M. pneumoniae as well as cross-reacting M. salivarium antigens but response to M. pneumoniae antigen was more pronounced. In the humoral response no such cross-reactivity was observed. Although some of the animals (3/8) demonstrated antibodies to M. salivarium prior to immunization the titers were not influenced by the immunization with M. pneumoniae antigen. Both groups of animals produced antibodies to M. pneumoniae antigen only, but significantly higher titers were observed in the adjuvant group. Cold hemagglutinins in both groups appeared earlier than the specific antibodies to mycoplasma. The adjuvant had no effect on the production of the cold agglutinins.     

Human colostral macrophages as indicator cells in migration inhibition test

The macrophage inhibition test was carried out using colostral macrophages obtained from PPD sensitive and measles and mycoplasma pneumoniae antigen insensitive woman. The macrophage migration was inhibited by PPD but neither by measles nor by mycoplasma pneumoniae antigen. Human colostral macrophages can be used as indicator cells in migration inhibition tests.