title: DNA from Dust: Comparative Genomics of Large DNA Viruses in Field Surveillance Samples.
abstract: The intensification of the poultry industry over the last 60 years facilitated the evolution of increased virulence and vaccine breaks in Marek's disease virus (MDV-1). Full-genome sequences are essential for understanding why and how this evolution occurred, but what is known about genome-wide variation in MDV comes from laboratory culture. To rectify this, we developed methods for obtaining high-quality genome sequences directly from field samples without the need for sequence-based enrichment strategies prior to sequencing. We applied this to the first characterization of MDV-1 genomes from the field, without prior culture. These viruses were collected from vaccinated hosts that acquired naturally circulating field strains of MDV-1, in the absence of a disease outbreak. This reflects the current issue afflicting the poultry industry, where virulent field strains continue to circulate despite vaccination and can remain undetected due to the lack of overt disease symptoms. We found that viral genomes from adjacent field sites had high levels of overall DNA identity, and despite strong evidence of purifying selection, had coding variations in proteins associated with virulence and manipulation of host immunity. Our methods empower ecological field surveillance, make it possible to determine the basis of viral virulence and vaccine breaks, and can be used to obtain full genomes from clinical samples of other large DNA viruses, known and unknown.IMPORTANCEDespite both clinical and laboratory data that show increased virulence in field isolates of MDV-1 over the last half century, we do not yet understand the genetic basis of its pathogenicity. Our knowledge of genome-wide variation between strains of this virus comes exclusively from isolates that have been cultured in the laboratory. MDV-1 isolates tend to lose virulence during repeated cycles of replication in the laboratory, raising concerns about the ability of cultured isolates to accurately reflect virus in the field. The ability to directly sequence and compare field isolates of this virus is critical to understanding the genetic basis of rising virulence in the wild. Our approaches remove the prior requirement for cell culture and allow direct measurement of viral genomic variation within and between hosts, over time, and during adaptation to changing conditions.
authors.affiliation: Department of Biochemistry and Molecular Biology, Center for Infectious Disease Dynamics, and the Huck Institutes of the Life Sciences, Pennsylvania State University, University Park, Pennsylvania, USA.
authors.forename: Utsav
authors.initials: U
authors.lastname: Pandey
authors.affiliation: Center for Infectious Disease Dynamics, Departments of Biology and Entomology, Pennsylvania State University, University Park, Pennsylvania, USA.
authors.forename: Andrew S
authors.initials: AS
authors.lastname: Bell
authors.affiliation: Department of Biochemistry and Molecular Biology, Center for Infectious Disease Dynamics, and the Huck Institutes of the Life Sciences, Pennsylvania State University, University Park, Pennsylvania, USA.
authors.forename: Daniel W
authors.initials: DW
authors.lastname: Renner
authors.affiliation: Center for Infectious Disease Dynamics, Departments of Biology and Entomology, Pennsylvania State University, University Park, Pennsylvania, USA.
authors.forename: David A
authors.initials: DA
authors.lastname: Kennedy
authors.affiliation: Department of Biochemistry and Molecular Biology, Center for Infectious Disease Dynamics, and the Huck Institutes of the Life Sciences, Pennsylvania State University, University Park, Pennsylvania, USA.
authors.forename: Jacob T
authors.initials: JT
authors.lastname: Shreve
authors.affiliation: Center for Infectious Disease Dynamics, Departments of Biology and Entomology, Pennsylvania State University, University Park, Pennsylvania, USA.
authors.forename: Chris L
authors.initials: CL
authors.lastname: Cairns
authors.affiliation: Center for Infectious Disease Dynamics, Departments of Biology and Entomology, Pennsylvania State University, University Park, Pennsylvania, USA.
authors.forename: Matthew J
authors.initials: MJ
authors.lastname: Jones
authors.affiliation: Department of Veterinary and Biomedical Sciences, Pennsylvania State University, University Park, Pennsylvania, USA.
authors.forename: Patricia A
authors.initials: PA
authors.lastname: Dunn
authors.affiliation: Center for Infectious Disease Dynamics, Departments of Biology and Entomology, Pennsylvania State University, University Park, Pennsylvania, USA.
authors.forename: Andrew F
authors.initials: AF
authors.lastname: Read
authors.affiliation: Department of Biochemistry and Molecular Biology, Center for Infectious Disease Dynamics, and the Huck Institutes of the Life Sciences, Pennsylvania State University, University Park, Pennsylvania, USA.
authors.forename: Moriah L
authors.identifier: 0000-0001-9859-1678
authors.initials: ML
authors.lastname: Szpara
keywords: Marek’s disease virus; genomics; herpesviruses; polymorphism; virulence