Limited Variation in BK Virus T-Cell Epitopes Revealed by Next-Generation Sequencing
Limited Variation in BK Virus T-Cell Epitopes Revealed by Next-Generation Sequencing
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DOI：
10.1128/jcm.01385-15
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发表时间：
2015-10-01
影响因子：
9.4
通讯作者：
Pinsky, Benjamin A.
中科院分区：
文献类型：
--
作者：
Sahoo, Malaya K.;Tan, Susanna K.;Pinsky, Benjamin A.
BK病毒（BKV）感染引起的终末器官疾病仍然是造血细胞移植（HCT）和肾移植领域面临的巨大挑战。由于BKV特异性治疗是有限的，基于免疫的治疗可能是一个有前途的和新的治疗选择移植受体与持续BKV感染。在这里，我们描述了一种全基因组、深度测序方法和生物信息学管道，用于识别整个基因组和BKV特异性HLA-A2-、HLA-B 0702-和HLA-B 08-限制性CD 8 T细胞表位的BKV变体。使用具有四个反向引物组的长距离PCR扩增BKV全基因组，并在Ion Torrent Personal Genome Machine（PGM）上对片段化文库进行测序。开发了错误模型和变体调用算法，以准确识别罕见变体。来自18名患有可定量BKV DNA血症的儿童HCT和肾脏接受者的共65份样本进行了全基因组测序。观察到有限的遗传变异。每个样品鉴定的氨基酸变体的中位数为8（范围，2至37;四分位数范围，10），其中大多数变体（77%）以1.5的频率检测。
BK virus (BKV) infection causing end-organ disease remains a formidable challenge to the hematopoietic cell transplant (HCT) and kidney transplant fields. As BKV-specific treatments are limited, immunologic-based therapies may be a promising and novel therapeutic option for transplant recipients with persistent BKV infection. Here, we describe a whole-genome, deep-sequencing methodology and bioinformatics pipeline that identify BKV variants across the genome and at BKV-specific HLA-A2-, HLA-B0702-, and HLA-B08-restricted CD8 T-cell epitopes. BKV whole genomes were amplified using long-range PCR with four inverse primer sets, and fragmentation libraries were sequenced on the Ion Torrent Personal Genome Machine (PGM). An error model and variant-calling algorithm were developed to accurately identify rare variants. A total of 65 samples from 18 pediatric HCT and kidney recipients with quantifiable BKV DNAemia underwent whole-genome sequencing. Limited genetic variation was observed. The median number of amino acid variants identified per sample was 8 (range, 2 to 37; interquartile range, 10), with the majority of variants (77%) detected at a frequency of