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ViroFind: A novel target-enrichment deep-sequencing platform reveals a complex JC virus population in the brain of PML patients.

Abstract Deep nucleotide sequencing enables the unbiased, broad-spectrum detection of viruses in clinical samples without requiring an a priori hypothesis for the source of infection. However, its use in clinical research applications is limited by low cost-effectiveness given that most of the sequencing information from clinical samples is related to the human genome, which renders the analysis of viral genomes challenging. To overcome this limitation we developed ViroFind, an in-solution target-enrichment platform for virus detection and discovery in clinical samples. ViroFind comprises 165,433 viral probes that cover the genomes of 535 selected DNA and RNA viruses that infect humans or could cause zoonosis. The ViroFind probes are used in a hybridization reaction to enrich viral sequences and therefore enhance the detection of viral genomes via deep sequencing. We used ViroFind to detect and analyze all viral populations in the brain of 5 patients with progressive multifocal leukoencephalopathy (PML) and of 18 control subjects with no known neurological disease. Compared to direct deep sequencing, by using ViroFind we enriched viral sequences present in the clinical samples up to 127-fold. We discovered highly complex polyoma virus JC populations in the PML brain samples with a remarkable degree of genetic divergence among the JC virus variants of each PML brain sample. Specifically for the viral capsid protein VP1 gene, we identified 24 single nucleotide substitutions, 12 of which were associated with amino acid changes. The most frequent (4 of 5 samples, 80%) amino acid change was D66H, which is associated with enhanced tissue tropism, and hence likely a viral fitness advantage, compared to other variants. Lastly, we also detected sparse JC virus sequences in 10 of 18 (55.5%) of control samples and sparse human herpes virus 6B (HHV6B) sequences in the brain of 11 of 18 (61.1%) control subjects. In sum, ViroFind enabled the in-depth analysis of all viral genomes in PML and control brain samples and allowed us to demonstrate a high degree of JC virus genetic divergence in vivo that has been previously underappreciated. ViroFind can be used to investigate the structure of the virome with unprecedented depth in health and disease state.
PMID
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Authors

Mayor MeshTerms
Keywords
Journal Title plos one
Publication Year Start




PMID- 29360822
OWN - NLM
STAT- MEDLINE
DCOM- 20180206
LR  - 20180206
IS  - 1932-6203 (Electronic)
IS  - 1932-6203 (Linking)
VI  - 13
IP  - 1
DP  - 2018
TI  - ViroFind: A novel target-enrichment deep-sequencing platform reveals a complex JC
      virus population in the brain of PML patients.
PG  - e0186945
LID - 10.1371/journal.pone.0186945 [doi]
AB  - Deep nucleotide sequencing enables the unbiased, broad-spectrum detection of
      viruses in clinical samples without requiring an a priori hypothesis for the
      source of infection. However, its use in clinical research applications is
      limited by low cost-effectiveness given that most of the sequencing information
      from clinical samples is related to the human genome, which renders the analysis 
      of viral genomes challenging. To overcome this limitation we developed ViroFind, 
      an in-solution target-enrichment platform for virus detection and discovery in
      clinical samples. ViroFind comprises 165,433 viral probes that cover the genomes 
      of 535 selected DNA and RNA viruses that infect humans or could cause zoonosis.
      The ViroFind probes are used in a hybridization reaction to enrich viral
      sequences and therefore enhance the detection of viral genomes via deep
      sequencing. We used ViroFind to detect and analyze all viral populations in the
      brain of 5 patients with progressive multifocal leukoencephalopathy (PML) and of 
      18 control subjects with no known neurological disease. Compared to direct deep
      sequencing, by using ViroFind we enriched viral sequences present in the clinical
      samples up to 127-fold. We discovered highly complex polyoma virus JC populations
      in the PML brain samples with a remarkable degree of genetic divergence among the
      JC virus variants of each PML brain sample. Specifically for the viral capsid
      protein VP1 gene, we identified 24 single nucleotide substitutions, 12 of which
      were associated with amino acid changes. The most frequent (4 of 5 samples, 80%) 
      amino acid change was D66H, which is associated with enhanced tissue tropism, and
      hence likely a viral fitness advantage, compared to other variants. Lastly, we
      also detected sparse JC virus sequences in 10 of 18 (55.5%) of control samples
      and sparse human herpes virus 6B (HHV6B) sequences in the brain of 11 of 18
      (61.1%) control subjects. In sum, ViroFind enabled the in-depth analysis of all
      viral genomes in PML and control brain samples and allowed us to demonstrate a
      high degree of JC virus genetic divergence in vivo that has been previously
      underappreciated. ViroFind can be used to investigate the structure of the virome
      with unprecedented depth in health and disease state.
FAU - Chalkias, Spyros
AU  - Chalkias S
AUID- ORCID: 0000-0002-6254-1158
AD  - Division of NeuroImmunology, Center for Virology and Vaccine Research, Department
      of Medicine, Beth Israel Deaconess Medical Center, Harvard Medical School,
      Boston, Massachusetts, United States of America.
AD  - Department of Genetics, Harvard Medical School, Boston, Massachusetts, United
      States of America.
FAU - Gorham, Joshua M
AU  - Gorham JM
AD  - Department of Genetics, Harvard Medical School, Boston, Massachusetts, United
      States of America.
FAU - Mazaika, Erica
AU  - Mazaika E
AD  - Department of Genetics, Harvard Medical School, Boston, Massachusetts, United
      States of America.
FAU - Parfenov, Michael
AU  - Parfenov M
AD  - Department of Genetics, Harvard Medical School, Boston, Massachusetts, United
      States of America.
FAU - Dang, Xin
AU  - Dang X
AD  - Division of NeuroImmunology, Center for Virology and Vaccine Research, Department
      of Medicine, Beth Israel Deaconess Medical Center, Harvard Medical School,
      Boston, Massachusetts, United States of America.
AD  - Department of Neurological Sciences, Rush University Medical Center, Chicago,
      Illinois, United States of America.
FAU - DePalma, Steve
AU  - DePalma S
AD  - Department of Genetics, Harvard Medical School, Boston, Massachusetts, United
      States of America.
FAU - McKean, David
AU  - McKean D
AD  - Department of Genetics, Harvard Medical School, Boston, Massachusetts, United
      States of America.
FAU - Seidman, Christine E
AU  - Seidman CE
AD  - Department of Genetics, Harvard Medical School, Boston, Massachusetts, United
      States of America.
FAU - Seidman, Jonathan G
AU  - Seidman JG
AD  - Department of Genetics, Harvard Medical School, Boston, Massachusetts, United
      States of America.
FAU - Koralnik, Igor J
AU  - Koralnik IJ
AD  - Division of NeuroImmunology, Center for Virology and Vaccine Research, Department
      of Medicine, Beth Israel Deaconess Medical Center, Harvard Medical School,
      Boston, Massachusetts, United States of America.
AD  - Department of Neurological Sciences, Rush University Medical Center, Chicago,
      Illinois, United States of America.
LA  - eng
GR  - U01-HL098188/NH/NIH HHS/United States
GR  - U01-HL098147/NH/NIH HHS/United States
GR  - U01-HL098153 /NH/NIH HHS/United States
GR  - U01-HL098163 /NH/NIH HHS/United States
GR  - U01-HL098123 /NH/NIH HHS/United States
GR  - U01-HL098162/NH/NIH HHS/United States
PT  - Journal Article
PT  - Research Support, N.I.H., Extramural
DEP - 20180123
PL  - United States
TA  - PLoS One
JT  - PloS one
JID - 101285081
SB  - IM
MH  - Brain/*virology
MH  - Genes, Viral
MH  - High-Throughput Nucleotide Sequencing/*methods
MH  - Humans
MH  - JC Virus/genetics/*isolation & purification
MH  - Leukoencephalopathy, Progressive Multifocal/*virology
PMC - PMC5779639
EDAT- 2018/01/24 06:00
MHDA- 2018/02/07 06:00
CRDT- 2018/01/24 06:00
PHST- 2017/06/04 00:00 [received]
PHST- 2017/10/10 00:00 [accepted]
PHST- 2018/01/24 06:00 [entrez]
PHST- 2018/01/24 06:00 [pubmed]
PHST- 2018/02/07 06:00 [medline]
AID - 10.1371/journal.pone.0186945 [doi]
AID - PONE-D-17-21386 [pii]
PST - epublish
SO  - PLoS One. 2018 Jan 23;13(1):e0186945. doi: 10.1371/journal.pone.0186945.
      eCollection 2018.