荧光染色法和KOH湿片法检测浅部真菌感染的效果比较

doi:10.3760/cma.j.issn.0412-4030.2019.05.004

摘要/Abstract

摘要： 目的对比荧光染色法和KOH湿片法检测浅部真菌感染的效果。方法收集2017年7月至2018年2月同济大学附属第十人民医院皮肤科门诊600例临床拟诊为浅部真菌感染及102例拟诊为马拉色菌感染（包括花斑糠疹54例，马拉色菌毛囊炎48例）的标本，分别采用荧光染色法和KOH湿片法直接镜检，比较检出阳性率及平均阅片时间。以培养为金标准，比较两种方法的漏诊情况。计数资料的比较采用卡方检验或Fisher精确检验，计量资料的比较采用配对t检验。结果荧光染色法与KOH湿片法检测600例浅部真菌感染标本时检出阳性分别为546例（91.00%）和489例（81.50%），阳性率差异有统计学意义（χ2 = 22.83，P < 0.05），平均阅片时间分别为（73.67 ± 13.56） s和（87.12 ± 15.83） s（t = 14.60，P < 0.05）；54例花斑糠疹标本检出阳性分别为51例（94.44%）、50例（92.59%）（校正χ2 ＝ 0，P > 0.05），平均阅片时间分别为（38.36 ± 8.79） s、（41.25 ± 15.67） s（t = 1.14，P > 0.05）；48例马拉色菌毛囊炎检出阳性分别为43例（89.58%）、11例（22.92%）（χ2 = 43.34，P < 0.05），平均阅片时间分别为（42.14 ± 12.61） s、（103.56 ± 9.48） s（t = 17.83，P < 0.05）。600例浅部真菌感染标本中，479例培养阳性，其中荧光染色法检测阳性476例，阴性3例（0.63%），KOH湿片法检测阳性465例，阴性14例（2.92%），漏诊率差异有统计学意义（χ2 = 7.25，P < 0.05）。结论荧光染色法相比于KOH湿片法可以提高检出阳性率，减少漏诊，缩短阅片时间。

关键词: 癣, 花斑癣, 马拉色霉菌属, 临床实验室技术, 荧光染色法, KOH湿片法

Abstract: 【Abstract】 Objective To compare the diagnostic value of fluorescent staining versus KOH wet-mount microscopy in detecting superficial fungal infection. Methods Totally, 600 specimens from cases of clinically diagnosed superficial fungal infections and 102 from cases of clinically diagnosed Malassezia infection （including 54 cases of pityriasis versicolor and 48 cases of Malassezia folliculitis） were collected from the dermatology clinic of Tenth People′s Hospital of Tongji University between July 2017 and February 2018. These specimens were subjected to fluorescent staining and KOH wet mount separately followed by direct microscopy, and the positive rate and average review time were compared between the two methods. Culture served as the gold standard method, and the missed diagnosis rate was compared between the two methods. Statistical analysis was carried out using chi-square test or Fisher′s exact test for comparing enumeration data, and paired t test for comparing emeasurement data. Results Of the 600 specimens from clinically diagnosed superficial fungal infection cases, fungi were detected in 546 （91.00%） and 489 （81.50%） by fluorescent staining and KOH wet-mount microscopy respectively （χ2 = 22.83, P < 0.05）. Fluorescent staining showed significantly shorter average review time （73.67 ± 13.56 s） compared with KOH wet-mount microscopy （87.12 ± 15.83 s, t = 14.60, P < 0.05）. Among the 54 specimens from pityriasis versicolor cases, fluorescent staining and KOH wet-mount microscopy positive results in 51 （94.44%） and 50 （92.59%） specimens respectively （adjusted χ2 ＝ 0, P > 0.05）, with the average review time being （38.36 ± 8.79） s and （41.25 ± 15.67）s respectively （t = 1.14, P > 0.05）. Of the 48 specimens from Malassezia infection cases, 43 （89.58%） and 11 （22.92%） specimens were detected to be positive for fungi by fluorescent staining and KOH wet-mount microscopy respectively （χ2 = 43.34, P < 0.05）, and fluorescent staining showed shorter average review time （42.14 ± 12.61 s） compared with KOH wet-mount microscopy （103.56 ± 9.48 s, t = 17.83, P < 0.05）. Among the 600 specimens from superficial fungal infection cases, culture yielded fungi in 479. Moreover, 476 specimens were found positive by fluorescent staining, and 3 were found negative （0.63%）, while KOH wet-mount microscopy showed 465 positive results and 14 negative results （2.92%）. There was a significant difference in the missed diagnosis rate between the two methods （χ2 = 7.25, P < 0.05）. Conclusion Compared with KOH wet-mount microscopy, fluorescent staining can increase the detection rate, reduce missed diagnosis rate and shorten review time.

Key words: Tinea, Tinea versicolor, Malassezia, Clinical laboratory techniques, Fluorescent staining, KOH method

余菁许辉刘芝翠马越娥史玉玲. 荧光染色法和KOH湿片法检测浅部真菌感染的效果比较[J]. 中华皮肤科杂志, 2019, 52(5): 314-318.

Yu Jing, Xu Hui, Liu Zhicui, Ma Yue′e, Shi Yuling. Comparison of fluorescent staining versus KOH wet-mount microscopy for detection of superficial fungal infection[J]. Chinese Journal of Dermatology, 2019, 52(5): 314-318.

参考文献 15

［1］	张思平, 王娟. 操作技能直接观察评估方法在皮肤真菌镜检教学中的探索［J］. 中华医学教育杂志, 2017,37（1）:94⁃98. doi: 10.3760/cma.j.issn.1673⁃677X.2017.01.020.
［2］	Vázquez María B, Amodeo Martín R, Bianchinotti María V. Fungal biomass estimation in soils from southwestern Buenos Aires province （Argentina） using calcofluor white stain［J］. Rev Argent Microbiol, 2016,48（3）:252⁃258. doi: 10.1016/j.ram.2016. 05.006.
［3］	Prakash R, Prashanth HV, Ragunatha S, et al. Comparative study of efficacy, rapidity of detection, and cost⁃effectiveness of potassium hydroxide, calcofluor white, and Chicago sky blue stains in the diagnosis of dermatophytoses［J］. Int J Dermatol, 2016,55（4）:e172⁃e175. doi: 10.1111/ijd.13037.
［4］	Gupta MK, Chandra A, Prakash P, et al. Fungal keratitis in north India; spectrum and diagnosis by Calcofluor white stain［J］. Indian J Med Microbiol, 2015,33（3）:462⁃463. doi: 10.4103/0255⁃0857.158609.
［5］	杨通, 谭亚夏. 活检组织中真菌Calcofluor White荧光染色法的改良［J］. 临床与病理杂志, 2015,35（5）:767⁃771. doi: 10. 3978/j.issn.2095⁃6959.2015.05.015.
［6］	陈锐君, 佟韬, 李超, 等. 荧光染色法和KOH湿片法在诊断浅部真菌感染中的效果比较［J］. 中国真菌学杂志, 2016,11（6）:378⁃380.
［7］	韩德忞, 刘原志, 朱均昊, 等. 荧光染色法与KOH湿片法在真菌直接镜检中的应用比较［J］. 中国真菌学杂志, 2016,11（4）:240⁃242. doi: 10.3969/j.issn.1673⁃3827.2016.04.014.
［8］	罗云鹏, 高仰敏, 金云, 等. 特敏增强荧光染色法在浅部真菌病诊断中的应用［J］. 临床检验杂志, 2017,35（10）:736⁃738. doi: 10.13602/j.cnki.jcls.2017.10.04.
［9］	胡记妹, 曾军荣, 粱海燕, 等. 甲真菌病的3种实验诊断方法研究［J］. 中国医药导报, 2010,7（13）:29⁃30. doi: 10.3969/j.issn.1673⁃7210.2010.13.012.
［10］	Yadav S, Saxena AK, Capoor MR, et al. Comparison of direct microscopicmethods using potassium hydroxide, periodic acid Schiff, and calcofluorwhite with culture in the diagnosis of onychomycosis［J］. Indian J Dermatol Venereol Leprol, 2013,79（2）:242⁃243. doi: 10.4103/0378⁃6323.107649.
［11］	Bonifaz A. Comparison of direct microscopy, culture and calcofluor whitefor the diagnosis of onychomycosis［J］. Rev Iberoam Micol, 2013,30（2）:109⁃111. doi: 10.1016/j.riam.2012. 07.001.
［12］	Prohic A, Jovovic Sadikovic T, Krupalija⁃Fazlic M, et al. Malassezia species in healthy skin and in dermatological conditions［J］. Int J Dermatol, 2016,55（5）:494⁃504. doi: 10.1111/ ijd.13116.
［13］	徐明, 张弘, 邓劲松, 等. 马拉色菌显微镜检查的方法学比较［J］. 福建医药杂志, 2011,33（1）:70⁃72. doi: 10.3969/j.issn.1002⁃2600.2011.01.033.
［14］	刘钟, 李东升, 陈金波, 等. 芝加哥天蓝与KOH涂片镜检对马拉色菌诊断的比较［J］. 中国皮肤性病学杂志, 2016,30（5）:534⁃536. doi: 10.13735/j.cjdv.1001⁃7089.201509083.
［15］	Ramos L, Mellado S, Ramadan S, et al. The use of calcofluor white for studying Malassezia species by direct microscopy［J］. Rev Argent Microbiol, 2006,38（1）:4⁃8. PMID: 16784125.

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