结核病实验室检测与图解

章结核病概述一、结核病的历史 .........................................................................................................1 二、结核病的流行病学及宿主易感因素 .....................................................................2 三、结核病相关的疫苗研究 .........................................................................................3 四、结核病的临床表现 .................................................................................................4 五、结核病的诊断 .........................................................................................................5 六、结核病的治疗 .........................................................................................................5 第二章结核病实验室安全防护一、实验室存在的危险 .................................................................................................7 二、实验室设施和设备要求 .........................................................................................8 三、实验室设计原则及基本要求 .................................................................................8 四、个人防护 .................................................................................................................9 五、进入实验室的要求 .................................................................................................9 六、实验室操作要求 .....................................................................................................9 七、实验室常用设备的正确使用和意外处理 ...........................................................10 八、常用消毒杀菌方法 ................................................................................................11 第三章分枝杆菌特性节结核分枝杆菌的生物学特性 ...................................................................................13 一、染色与形态 ...........................................................................................................14 二、培养特性 ...............................................................................................................15 三、生化反应 ...............................................................................................................16 四、变异性 ...................................................................................................................16 五、抵抗力与消毒 .......................................................................................................17 第二节非结核分枝杆菌的生物学特性 ...............................................................................17 一、分类 .......................................................................................................................18 二、形态 .......................................................................................................................18 三、培养特性 ...............................................................................................................18 四、致病性 ...................................................................................................................19 五、药物敏感性 ...........................................................................................................19 六、常见非结核分枝杆菌 ...........................................................................................19 第四章临床送检标本采集、运送和保存一、呼吸道标本的采集 ...............................................................................................23 二、肺外标本的采集 ...................................................................................................25 三、标本采集注意事项、运送及保存 .........................................................................26 第五章抗酸杆菌涂片、染色及显微镜检查节抗酸杆菌涂片处理方法及局限性分析 ...................................................................28 一、概述 .......................................................................................................................28 二、涂片镜检的优缺点 ...............................................................................................28 三、直接涂片法 ...........................................................................................................29 四、漂浮集菌法 ...........................................................................................................30 五、离心沉淀法 ...........................................................................................................31 六、消化后离心沉淀法 ...............................................................................................31 七、涂片注意事项 .......................................................................................................35 第二节抗酸染色及显微镜检查 ............................................................................................36 一、萋—尼抗酸染色 .....................................................................................................36 二、改良萋—尼抗酸染色 .............................................................................................37 三、 Kinyoun抗酸染色.................................................................................................37 四、潘本汉（Pappenheim）抗酸染色 ..........................................................................38 五、弱抗酸染色 ...........................................................................................................38 六、卡贝托（Gabbet）抗酸染色 ..................................................................................39 七、改良I—K抗酸染色 ................................................................................................39 八、显微镜镜检 ...........................................................................................................40 九、结果判断 ...............................................................................................................40 十、结果判读 ...............................................................................................................40 十一、痰涂片抗酸杆菌浓度与涂阳概率 ...................................................................40 十二、读片时间与镜检阳性率的关系 .......................................................................41 十三、不同离心力对涂片镜检及培养结果的影响 ...................................................41 十四、标本中不同抗酸杆菌抗酸染色形态汇总 .......................................................41 第三节荧光染色及显微镜检查 ............................................................................................45 一、原理及特点 ...........................................................................................................45 二、 LED荧光显微镜与传统荧光显微镜比较 ...........................................................46 三、金胺O染色 ...........................................................................................................46 四、金胺O—罗丹明染色 ..............................................................................................47 五、镜检 .......................................................................................................................47 六、结果判断 ...............................................................................................................47 七、结果判读 ...............................................................................................................47 八、荧光染色优越性 ...................................................................................................48 九、荧光染色局限性 ...................................................................................................48 十、标本中不同抗酸杆菌荧光染色形态汇总 ...........................................................49 第四节质量控制 .....................................................................................................................54 一、室内质控 ...............................................................................................................54 二、室间比对 ...............................................................................................................55 三、注意事项 ...............................................................................................................55 第五节抗酸杆菌涂片假阳性、假阴性原因分析及处理措施 ...........................................55 一、假阳性原因分析及预防措施 ...............................................................................55 二、假阴性原因分析及预防措施 ...............................................................................56 第六章分枝杆菌培养节分枝杆菌培养概述 ....................................................................................................59 一、概述 .......................................................................................................................59 二、不同化学物质对分枝杆菌的生长影响 ...............................................................59 第二节临床标本前处理方法 ................................................................................................61 一、概述 .......................................................................................................................61 二、分枝杆菌培养去污染方法 ...................................................................................62 第三节分枝杆菌培养基种类及介绍 ...................................................................................67 一、概述 .......................................................................................................................67 二、商品化快速培养、药敏检测系统 .........................................................................67 三、液体变色培养基 ...................................................................................................68 四、固体培养基 ...........................................................................................................68 五、双相培养基 ...........................................................................................................69 六、常用培养基成分介绍 ...........................................................................................69 第四节分枝杆菌固体培养操作程序 ...................................................................................71 一、试剂、培养基与耗材 .............................................................................................71 二、标本前处理 ...........................................................................................................72 三、标本接种 ...............................................................................................................73 四、结果观察 ...............................................................................................................74 五、报告方式 ...............................................................................................................74 六、注意事项 ...............................................................................................................74 七、室内质控 ...............................................................................................................75 八、罗氏培养基上结核分枝杆菌、非结核分枝杆菌、诺卡菌及常见污染杂菌菌落及镜检形态汇总 ..........................................................................76 第五节 BACTEC MGIT 960 System全自动分枝杆菌检测/ 药敏系统操作程序 ....................................................................................................89 一、概况及检测原理 ...................................................................................................89 二、仪器硬件系统 .......................................................................................................89 三、实验准备 ...............................................................................................................90 四、标本前处理 ...........................................................................................................90 五、标本接种 ...............................................................................................................91 六、结果报告 ...............................................................................................................91 七、质量控制 ...............................................................................................................91 八、仪器状态检查及保养 ...........................................................................................92 九、检测局限性 ...........................................................................................................92 十、 BACTEC MGIT 960 System培养结核分枝杆菌、非结核分枝杆菌、诺卡菌及常见污染杂菌菌落及镜检形态汇总 ........................................93 第六节 BACTEC 9120全自动血培养仪 ..........................................................................98 一、原理 .......................................................................................................................98 二、标本培养检测的操作步骤 ...................................................................................98 三、报警及错误处理 ...................................................................................................99 四、仪器报阳瓶处理 .................................................................................................101 五、结果报告 .............................................................................................................101 六、质量控制 .............................................................................................................101 七、注意事项 .............................................................................................................102 八、 BACTEC 9120培养结核分枝杆菌、非结核分枝杆菌镜检形态汇总 ..............103 第七节分枝杆菌培养报阳后标本的处理 .........................................................................105 一、 BACTEC MGIT 960报阳后标本的处理 ...........................................................105 二、血培养报阳后标本的处理 .................................................................................106 三、罗氏培养阳性菌株转种方法推荐 .....................................................................106 四、罗氏培养阳性菌株涂片方法推荐 .....................................................................107 第八节分枝杆菌培养结果影响因素分析 .........................................................................107 一、假阳性原因分析及处理措施 .............................................................................107 二、假阴性原因分析及处理措施 .............................................................................108 三、分枝杆菌培养注意事项 .....................................................................................109 四、不同培养基培养分枝杆菌敏感性差异比较 .....................................................109 五、提高分枝杆菌培养检出率的方法 .....................................................................109 第七章分枝杆菌菌种鉴定节传统鉴定方法及局限性介绍 ..................................................................................111 一、结核分枝杆菌与非结核分枝杆菌的区别 ..........................................................111 二、结核分枝杆菌和牛分枝杆菌鉴定 ......................................................................113 三、偶发分枝杆菌、龟分枝杆菌和脓肿分枝杆菌鉴定 ............................................114 四、分枝杆菌鉴定流程图 ..........................................................................................115 第二节分枝杆菌免疫学检查 ...............................................................................................115 一、 PPD结核菌素皮肤试验介绍及局限性分析 ......................................................115 二、 γ干扰素释放试验（IGRA） ..................................................................................116 三、结核抗原检测及局限性 .....................................................................................121 四、四、结核抗体检测及局限性 .....................................................................................123 第八章分枝杆菌核酸检测及局限性分析节结核分枝杆菌复合群分子鉴定 .............................................................................125 一、用于结核分枝杆菌复合群鉴定的DNA序列 ...................................................125 二、用于结核分枝杆菌复合群鉴定的技术 .............................................................126 第二节非结核分枝杆菌菌种分子鉴定 .............................................................................128 一、用于非结核分枝杆菌菌种分子鉴定的DNA序列 ...........................................128 二、用于非结核分枝杆菌菌种分子鉴定的技术 .....................................................129 第三节结核分枝杆菌耐药性分子检测 .............................................................................130 一、用于结核分枝杆菌耐药性分子检测的DNA序列 ...........................................130 二、用于结核分枝杆菌耐药性分子检测的技术 .....................................................131 第九章 MALDI—TOF MS技术在分枝杆菌鉴定中的应用节 MALDI Biotyper在分枝杆菌鉴定中的应用.....................................................142 一、基本原理与流程 .................................................................................................142 二、在分枝杆菌鉴定中的应用 .................................................................................144 第二节 VITEK MS在分枝杆菌鉴定中的应用 ................................................................148 一、方法及原理 ........................................................