Lindsley M D, Hurst S F, Iqbal N J, Morrison C J
Mycotic Diseases Branch, Division of Bacterial and Mycotic Diseases, Centers for Disease Control and Prevention, Atlanta, Georgia, USA.
J Clin Microbiol. 2001 Oct;39(10):3505-11. doi: 10.1128/JCM.39.10.3505-3511.2001.
Specific oligonucleotide probes were developed to identify medically important fungi that display yeast-like morphology in vivo. Universal fungal primers ITS1 and ITS4, directed to the conserved regions of ribosomal DNA, were used to amplify DNA from Histoplasma capsulatum, Blastomyces dermatitidis, Coccidioides immitis, Paracoccidioides brasiliensis, Penicillium marneffei, Sporothrix schenckii, Cryptococcus neoformans, five Candida species, and Pneumocystis carinii. Specific oligonucleotide probes to identify these fungi, as well as a probe to detect all dimorphic, systemic pathogens, were developed. PCR amplicons were detected colorimetrically in an enzyme immunoassay format. The dimorphic probe hybridized with DNA from H. capsulatum, B. dermatitidis, C. immitis, P. brasiliensis, and P. marneffei but not with DNA from nondimorphic fungi. Specific probes for H. capsulatum, B. dermatitidis, C. immitis, P. brasiliensis, P. marneffei, S. schenckii, C. neoformans, and P. carinii hybridized with homologous but not heterologous DNA. Minor cross-reactivity was observed for the B. dermititidis probe used against C. immitis DNA and for the H. capsulatum probe used against Candida albicans DNA. However, the C. immitis probe did not cross-react with B. dermititidis DNA, nor did the dimorphic probe hybridize with C. albicans DNA. Therefore, these fungi could be differentiated by a process of elimination. In conclusion, probes developed to yeast-like pathogens were found to be highly specific and should prove to be useful in differentiating these organisms in the clinical setting.
开发了特异性寡核苷酸探针，以鉴定在体内呈现酵母样形态的医学上重要的真菌。针对核糖体DNA保守区域的通用真菌引物ITS1和ITS4，用于扩增来自荚膜组织胞浆菌、皮炎芽生菌、粗球孢子菌、巴西副球孢子菌、马尔尼菲青霉、申克孢子丝菌、新型隐球菌、五种念珠菌属以及卡氏肺孢子虫的DNA。开发了用于鉴定这些真菌的特异性寡核苷酸探针，以及一种用于检测所有双相性、系统性病原体的探针。PCR扩增产物在酶免疫测定形式中通过比色法进行检测。双相性探针与来自荚膜组织胞浆菌、皮炎芽生菌、粗球孢子菌、巴西副球孢子菌和马尔尼菲青霉的DNA杂交，但不与非双相性真菌的DNA杂交。针对荚膜组织胞浆菌、皮炎芽生菌、粗球孢子菌、巴西副球孢子菌、马尔尼菲青霉、申克孢子丝菌、新型隐球菌和卡氏肺孢子虫的特异性探针与同源而非异源DNA杂交。观察到用于检测粗球孢子菌DNA的皮炎芽生菌探针以及用于检测白色念珠菌DNA的荚膜组织胞浆菌探针存在轻微交叉反应。然而，粗球孢子菌探针不与皮炎芽生菌DNA交叉反应，双相性探针也不与白色念珠菌DNA杂交。因此，这些真菌可以通过排除过程进行区分。总之，发现针对酵母样病原体开发的探针具有高度特异性，并且在临床环境中区分这些微生物时应被证明是有用的。
