接合菌病及实验室诊断

1、接合菌病及诊断第一医院真菌和真菌病研究中心余进Dept. Dermatology, Peking University First Hospital and Research Center for Medical Mycology, Peking University常见致病接合菌根霉Rhizopus克银汉霉瓶霉被孢霉蛙粪霉毛霉Mucor毛霉目Mucorale接合菌Zygomycota虫霉目Entomophthorales根毛霉Rhizomucor共头霉属囊托霉属科克霉犁头霉Lichtheimmiaor Myoclaadius (Absidia)耳霉Dept. Dermatology, Pek

2、ing University First Hospital and Research Center for Medical Mycology, Peking University接合菌比例上升Dept. Dermatology, Peking University First Hospital and Research Center for MedicKontoyiannis DP,cology, Peking University. JInfectDis. 2005;191:1350Incidence per 1000 Patient Days0,7Aspergillus0,210,6Zyg

3、omycetes0,180,50,150,40,120,30,090,20,060,10,03002000 2001 2002 2003 2004Year毛霉目真菌重要的医学真菌常见属：根霉 Rhizopus黄梗霉Lichtheimia(犁头霉Absidia) 根毛霉Rhizomucor毛霉Mucor少见属： Saksenaea， Syncephalastrum，Cunninghamela，CokeromycesDept. Dermatology, Peking University First Hospital and Research Center for Medical Mycology

4、, Peking University毛霉病（mucormycosis）分布于自然界和腐烂植物中条件致病菌：相对罕见，次于念珠菌、曲霉易感因素生前诊断难，治疗效果差，预后差Dept. Dermatology, Peking University First Hospital and Research Center for Medical Mycology, Peking University途径吸入孢子，肺和鼻窦是最常见的部位经皮途径，皮肤创伤，注射器污染，绷带污染食用被孢子污染的蚊虫叮咬Dept. Dermatology, Peking University First Hospital a

5、nd Research Center for Medical Mycology, Peking University免疫抑制(中性粒细胞缺乏,糖皮质激素治疗,移植, HIV)代谢问题 (, 未,去铁胺治疗,慢性酮症酸的)代谢性酸皮肤软组织破损 (烧伤,外伤接种,手术伤口)其它(静脉吸毒,早产儿, 营养不良)Dept. Dermatology, Peking University First Hospital and Research Center for Medical Mycology, Peking University临床类型鼻脑(44-49%)皮肤(1016%) 肺(1011%)播散性

6、(611.6%)消化道 (211%)其他:肾脏等Dept. Dermatology, Peking University First Hospital and Research Center for Medical Mycology, Peking University鼻脑毛霉病rhino-orbital-cerebral mucormycosis最常见70%由根霉属引起，多见于米根霉和小孢根霉一半以上的患者有表现一侧头痛，鼻内疼痛和发热波及上腭，患侧腭部有黑色焦痂，鼻中隔穿孔或硬腭穿孔波及眼眶，引起眶周或鼻周持续性肿胀波及第2、4、6对颅神经时，可出现瞳孔散大、固定、凸眼或上睑下垂，甚至失明

7、波及大时，可在颅内引起栓塞和坏死，伴脑DD软Dept. D化ermatology, Peking University First Hospital and Research Center for Medical Mycology, Peking University肺毛霉病pulmonary mucormycosis恶性血液病37%，脑毛霉病吸入空气中或巴管播散后引起32%，仅次于鼻鼻窦中的孢子，或行、淋非特异性：持续性高热、咳嗽、和胸痛胸片：浸润、多发结节、实变，空洞少见高分辨率CT痰培养Dept. Dermatology, Peking University First Hospital

8、 and Research Center for Medical Mycology, Peking University胃肠毛霉病gastrointestinal mucormycosis多发生于营养不良的婴儿或儿童无特异性，有腹痛、呕血、黑便，严重可形成坏死性胃炎或肠道溃疡等最常见的部位是胃（占该型的95%以上），其次是大肠慢性消化道溃疡是重要的基础疾病胃镜检查可见表浅或深在的溃疡，表面覆以厚层污苔，严重的深部损害可形成穿孔或镜可见到绿色或黑色的中心坏死区；结肠Dept. Dermatology, Peking University First Hospital and Research C

9、enter for Medical Mycology, Peking University皮肤接合菌病cutaneous zygomycosis最轻的一型，急性皮肤率低，预后较好伴有严重基础病发热, 肿胀, 组织坏死, 梗死,黑色焦痂慢性皮肤肉芽肿不伴有严重基础病性肉芽肿胰岛素注射部位 , 导管处, 伤口表面、移植 (肝),骨髓移植, 烧伤,Dept. Dermatology, Peking University First Hospital and Research Center for Medical Mycology, Peking University播散型毛霉病disseminate

10、d mucormycosis可继发于上述4型，累及两个或多个肺毛霉病易于出现脑部播散脑最常见，脾、心和皮肤多发梗死最常见于中性粒细胞减少症或淋巴瘤患者，伏立康唑预防应用时率高，90% ，100%Dept. Dermatology, Peking University First Hospital and Research Center for Medical Mycology, Peking University其它型毛霉病心脏、腹膜、肾、骨髓等Dept. Dermatology, Peking University First Hospital and Research Center for

11、 Medical Mycology, Peking University诊断高危患者，临床表现，使用抗细菌或唑类抗真菌影像学 微生物学（泊沙康唑除外）无效组织病理尤为重要诊断Dept. Dermatology, Peking University First Hospital and Research Center for Medical Mycology, Peking University治疗基础疾病，纠正酸和水电解质紊乱，某些的影响外科手术重要，尽可能采用 可显著降低率 70%10%早期应用有效的抗真菌Dept. Dermatology, Peking University First

12、Hospital and Research Center for Medical Mycology, Peking University抗真菌两性霉素B及其脂泊沙康唑（posaconazole) 伊曲康唑比前二者效差用药 两性霉素B+泊沙康唑 两性霉素B+卡泊芬净 泊沙康唑+卡泊芬净Dept. Dermatology, Peking University First Hospital and Research Center for Medical Mycology, Peking University辅助治疗高压氧铁鳌合剂（非去铁胺）IFN-r, GM-CSFDept. Dermatology

13、, Peking University First Hospital and Research Center for Medical Mycology, Peking University预后与预防鼻脑毛霉病（80%90%） 、肺毛霉病（65%）和播散型毛霉病预后差，率高积极治疗基础疾病，杜绝医源性因素是重要的环节环境：高危人群尽量远离空气中散布大量孢子的环境Dept. Dermatology, Peking University First Hospital and Research Center for Medical Mycology, Peking University接合菌病的诊断D

14、ept. Dermatology, Peking University First Hospital and Research Center for Medical Mycology, Peking University接合菌诊断临床症状高危因素影像学表现体征临床诊断微生物学组织病理学诊断确诊、拟诊或排除Dept. Dermatology, Peking University First Hospital and Research Center for Medical Mycology, Peking University免疫抑制(中性粒细胞缺乏,糖皮质激素治疗,移植, HIV)代谢问题 (,

15、 未,去铁胺治疗,慢性酮症酸的)代谢性酸皮肤软组织破损 (烧伤,外伤接种,手术伤口)其它(静脉吸毒,早产儿, 营养不良)Dept. Dermatology, Peking University First Hospital and Research Center for Medical Mycology, Peking University临床症状和体征病史无特异性某些症状或体征有提示意义患者出现复视颌面部、眶周、鼻旁出现坏死性焦痂中性粒细胞缺乏患者出现胸膜疼免疫受损患者出现坏死性皮疹使用抗细菌 唑除外）无效或唑类抗真菌（泊沙康Dept. Dermatology, Peking Univers

16、ity First Hospital and Research Center for Medical Mycology, Peking University影像学诊断早期CT检查肺、鼻窦常见表现 结节 反向晕征（Reversed halo signs)或晕征（halo signs） 楔形浸润影 胸膜渗出Dept. Dermatology, Peking University First Hospital and Research Center for Medical Mycology, Peking UniversityRHS（反向晕征）由于肺梗死周边毛霉病中更常见重于中心Dept. Derm

17、atology, Peking University First Hospital and Research Center for Medical Mycology, Peking UniversityReversed Halo Sign and Pulmonary Zygomycosis189 确诊或临床诊断真菌性132 肺曲霉病37 肺毛霉病20 肺镰刀菌病8名患者出现RHS(4%)7 名肺毛霉病1名 肺曲霉病Dept. Dermatology, Peking University First Hospital and Research Center for Medical Mycolog

18、y, Peking UniversityHS（晕征）肺梗死周边肺泡周边性坏死无炎症细胞浸润IPA更常见Dept. Dermatology, Peking University First Hospital and Research Center for Medical Mycology, Peking University肺曲霉病与肺接合菌病临床和影像学差异临床差异 鼻窦累及 伏立康唑预防治疗影像学差异 多发结节（>=10个） 胸膜渗出Dept. Dermatology, Peking University First Hospital and Research Center for M

19、edical Mycology, Peking University病原诊断微生物学Dept. Dermatology, Peking University First Hospital and Research Center for Medical Mycology, Peking University微生物学特征-显微镜下形态直接涂片10%KOHDept. Dermatology, Peking University First Hospital and Research Center for Medical Mycology, Peking University微生物学特征-培养特点血培

20、养阳性罕见 组织培养阳性率低 避免组织匀浆 剪碎或切碎组织 35-37C培养可提高阳性率生长快速霉样菌落Dept. Dermatology, Peking University First Hospital and Research Center for Medical Mycology, Peking University鉴定要点出现产孢结构 孢子囊、孢囊梗、假根、囊托-分属 温度试验、孢子囊孢子、孢囊梗长度、厚壁孢子等-分种DNA序列比对Dept. Dermatology, Peking University First Hospital and Research Center for M

21、edical Mycology, Peking UniversityAbsidia,Mucor,Rhizomucor,Rhizopus的孢子囊特征犁头霉:*接合孢子附属丝*孢囊下分隔*无临床致病Lichtheimia(“横梗霉”)*圆锥状囊轴*没有孢囊下分隔根毛霉* 囊托 -* 假根 +根霉 :* 囊托 +* 假根 +毛霉* 囊托 -* 假根 -其他:微生物学特征-组织病理表现HE、PAS、GMS染色均可见典型表现：无分隔或者分隔少的宽大菌丝，壁薄，易折叠，90度分支Dept. Dermatology, Peking University First Hospital and Research

22、 Center for Medical Mycology, Peking UniversityInvasive zygomycosis免疫组化和原位杂交ImmunohistochemicalPASISHDept. Dermatology, Peking University First Hospital and Research Center for Medical Mycology, Peking University新型诊断技术基于DNA的核酸扩增技术基于蛋白质的质谱技术 (MALDI-TOF） 代谢产物 LC-MS (soluble) GC-MS (volatile organic co

23、mpounds VOCs)EIA Galactofuranose Epitope-based cell wall components Other cell wall epitopesDept. Dermatology, Peking University First Hospital and Research Center for Medical Mycology, Peking University诊断-组织新鲜/冰冻组织福尔固定石蜡包埋组织SequencingRFLP提取DNAPCRRLBDept. Dermatology, Peking University First Hospital and Research Center for Medical Mycology, Peking University新鲜/冰冻组织敏感性和特异性较好正确鉴定种属Dept. Dermatology, Peking University First Hospital and Research Center for Medical Mycology, Peking University福尔方法固定石蜡包埋组织 PCR+Sequencing( ITS1) Semi-nested PCR(18SrRNA) Real-time PC