PCR结合反向斑点杂交法检测石蜡包埋组织中的曲霉感染

目的评价PCR结合反向斑点杂交法检测福尔马林固定、石蜡包埋组织中曲霉感染的可行性。方法选取39例病理证实曲霉感染的患者活检标本（21例为鼻窦感染标本、18例为尸检标本），以1对真菌特有的28SrRNA保守序列结构作为真菌通用引物，以临床常见的4个曲霉菌种：烟曲霉、黄曲霉、黑曲霉、土曲霉的种特异性序列为种特异性探针，与扩增产物进行反向斑点杂交。结果尸检标本阳性率为55．6％（10／18），鼻窦标本阳性率为76．2％（16／21），特异性均为100％。在这些曲霉所致的系统性感染中，烟曲霉是主要的致病真菌。结论该方法能对临床无法培养的石蜡组织块进行回顾性病原学研究，并可以鉴定常见的曲霉菌种，有良好的特异性和敏感性，适用于临床曲霉感染的检测。

Detection and identification of Aspergillus species from formalin-fixed,paraffin-embedded human tissues by PCR and dot blot hybridization

Objective To establish and evaluate a molecular diagnostic method for the detection and identification of Aspergillus species in formalin-fixed, paraffin-embedded human tissues. Methods Thirty-nine samples were obtained from patients with histo-logically diagnosed Aspergillus species infections. Eighteen samples were removed at necropsy. Twenty-one samples were tissue sections of the maxillary sinus samples. PCR amplification with universal fungal primers for 28S ribosomal RNA and identification by hybridization with species-specific probes for Aspergillus fumigatus , Aspergillus flavus , Aspergillus niger, Aspergillus terreus were performed for all samples. Results Positive hybridization could be obtained from 10 samples （55.6%） removed at necropsy, and 16 maxillary sinus samples （76.2%） ,with specificity of 100%. Hybridization showed reliable results for A.fumigatus ,which proved to be the most common agent in formalin-fixed, paraffin-embedded human tissues with histologically diagnosed Aspergillus species infections. OtherAspergillus species were rarely found. Conclusions PCR and dot blot hybridization is sensitive and specific for identification of common Aspergillus spp. This technique may be useful, not only for the retrospective study but also for clinical molecular diagnosis of systemic Aspergillus infection.