title: Limited Variation in BK Virus T-Cell Epitopes Revealed by Next-Generation Sequencing.
abstract: 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 <5%. When normalized for length, there was no statistical difference in the median number of variants across all genes. Similarly, the predominant virus population within samples harbored T-cell epitopes similar to the reference BKV strain that was matched for the BKV genotype. Despite the conservation of epitopes, low-level variants in T-cell epitopes were detected in 77.7% (14/18) of patients. Understanding epitope variation across the whole genome provides insight into the virus-immune interface and may help guide the development of protocols for novel immunologic-based therapies.
authors.affiliation: Department of Pathology, Stanford University School of Medicine, Stanford, California, USA.
authors.forename: Malaya K
authors.initials: MK
authors.lastname: Sahoo
authors.affiliation: Department of Medicine, Division of Infectious Diseases and Geographic Medicine, Stanford University School of Medicine, Stanford, California, USA.
authors.forename: Susanna K
authors.initials: SK
authors.lastname: Tan
authors.affiliation: Department of Pediatrics, Division of Infectious Diseases, Stanford University School of Medicine, Stanford, California, USA.
authors.forename: Sharon F
authors.initials: SF
authors.lastname: Chen
authors.affiliation: Department of Pathology, Stanford University School of Medicine, Stanford, California, USA.
authors.forename: Beatrix
authors.initials: B
authors.lastname: Kapusinszky
authors.affiliation: Department of Pathology, Stanford University School of Medicine, Stanford, California, USA.
authors.forename: Katherine R
authors.initials: KR
authors.lastname: Concepcion
authors.affiliation: Pediatric Renal Transplant Program, Lucile Packard Children's Hospital at Stanford, Stanford, California, USA.
authors.forename: Lynn
authors.initials: L
authors.lastname: Kjelson
authors.affiliation: Department of Pathology, Stanford University School of Medicine, Stanford, California, USA.
authors.forename: Kalyan
authors.initials: K
authors.lastname: Mallempati
authors.affiliation: Department of Pathology, Stanford University School of Medicine, Stanford, California, USA.
authors.forename: Heidi M
authors.initials: HM
authors.lastname: Farina
authors.affiliation: Department of Pathology, Stanford University School of Medicine, Stanford, California, USA.
authors.forename: Marcelo
authors.initials: M
authors.lastname: Fernández-Viña
authors.affiliation: Department of Pathology, Stanford University School of Medicine, Stanford, California, USA.
authors.forename: Dolly
authors.initials: D
authors.lastname: Tyan
authors.affiliation: Department of Pediatrics, Division of Nephrology, Stanford University School of Medicine, Stanford, California, USA.
authors.forename: Paul C
authors.initials: PC
authors.lastname: Grimm
authors.affiliation: Department of Pathology, Stanford University School of Medicine, Stanford, California, USA.
authors.forename: Matthew W
authors.initials: MW
authors.lastname: Anderson
authors.affiliation: Department of Surgery, Division of Abdominal Transplantation, Stanford University School of Medicine, Stanford, California, USA.
authors.forename: Waldo
authors.initials: W
authors.lastname: Concepcion
authors.affiliation: Department of Pathology, Stanford University School of Medicine, Stanford, California, USA Department of Medicine, Division of Infectious Diseases and Geographic Medicine, Stanford University School of Medicine, Stanford, California, USA bpinsky@stanford.edu.
authors.forename: Benjamin A
authors.initials: BA
authors.lastname: Pinsky
mesh_terms: D000293:Adolescent; D001739:BK Virus / Q000235:genetics* / Q000276:immunology* / Q000302:isolation & purification; D002648:Child; D002675:Child, Preschool; D017124:Conserved Sequence; D004279:DNA, Viral / Q000737:chemistry / Q000235:genetics; D018984:Epitopes, T-Lymphocyte / Q000235:genetics* / Q000276:immunology*; D005260:Female; D014644:Genetic Variation; D016679:Genome, Viral; D006801:Humans; D008297:Male; D017422:Sequence Analysis, DNA; D055815:Young Adult
chemical_list: D004279:DNA, Viral; D018984:Epitopes, T-Lymphocyte