Clinical Research Center, Marshfield Clinic Research Foundation, Marshfield Clinic, Marshfield, Wisconsin 54449, USA.
Med Mycol. 2010 Mar;48(2):285-90. doi: 10.1080/13693780903103952.
Blastomycosis is a serious and potentially fatal infection caused by the thermally dimorphic fungus Blastomyces dermatitidis. Polymerase chain reaction (PCR) assays targeting the BAD-1 virulence gene promoter have been developed to aid in the detection of the pathogen in clinical and environmental samples. However, little is known regarding the genetic diversity of B. dermatitidis and how this might affect the performance characteristics of these assays. We explored the genetic relatedness of 106 clinical and environmental isolates of B. dermatitidis using a previously described rDNA PCR restriction fragment length polymorphism (RFLP) assay. In addition, we looked for polymorphisms in the promoter region upstream of BAD-1. RFLP analysis showed that all isolates fell into one of five genotypic groups, designated A through E. Genotypic groups A and B predominated, comprising 50/106 (47.2%) and 51/106 (48.1%) of isolates, respectively. Three of 106 (2.8%) isolates were genotype C. Genotypes D and E represented novel genotypes and were each associated with single clinical isolates. PCR of the BAD-1 promoter revealed significant size differences among amplification products. Fifty-one of 106 isolates (50/50 RFLP genotypic group A and 1/51 genotypic group B) had amplicons of 663-bp, nearly twice the size of the expected product. Sequence analysis of amplification products from 17 representative isolates revealed four haplotypes and showed that the size disparity was due to two large insertions. Because these insertions were present in a high percentage of isolates, the utility of the PCR assays for diagnostic purposes could be affected. However, the novel RFLP genotypes and multiple BAD-1 haplotypes may prove useful as markers in population genetic studies.
芽生菌病是一种由热双相真菌皮炎芽生菌引起的严重且潜在致命的感染。聚合酶链反应（PCR）检测已针对 BAD-1 毒力基因启动子开发，以帮助在临床和环境样本中检测病原体。然而，对于皮炎芽生菌的遗传多样性以及这如何影响这些检测的性能特征知之甚少。我们使用先前描述的 rDNA PCR 限制性片段长度多态性（RFLP）检测法，研究了 106 株临床和环境分离株的遗传相关性。此外，我们还研究了 BAD-1 上游启动子区域的多态性。RFLP 分析表明，所有分离株均属于五个基因型组之一，命名为 A 至 E。基因型组 A 和 B 占主导地位，分别占分离株的 50/106（47.2%）和 51/106（48.1%）。106 个分离株中的 3 个（2.8%）为基因型 C。基因型 D 和 E 代表新的基因型，分别与单个临床分离株相关。BAD-1 启动子的 PCR 显示扩增产物之间存在显著的大小差异。106 个分离株中的 51 个（50/50 RFLP 基因型组 A 和 1/51 基因型组 B）的扩增子大小为 663-bp，几乎是预期产物的两倍。来自 17 个代表性分离株的扩增产物的序列分析显示了四个单倍型，表明大小差异是由于两个大插入所致。由于这些插入存在于高比例的分离株中，因此 PCR 检测在诊断目的中的实用性可能会受到影响。然而，新的 RFLP 基因型和多个 BAD-1 单倍型可能可用作群体遗传学研究中的标记。
