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Detection of parvovirus B19 and novel human parvoviruses in high-risk individuals

Detection of parvovirus B19 and novel human parvoviruses in high-risk individuals. Ashleigh Manning 1 , Kate Templeton 2 , Ed. Gomperts 3 , Peter Simmonds 1,2 1 Centre for Infectious Diseases, University of Edinburgh 2 Specialist Virology Laboratory, Royal Infirmary of Edinburgh

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Detection of parvovirus B19 and novel human parvoviruses in high-risk individuals

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  1. Detection of parvovirus B19 and novel human parvoviruses in high-risk individuals Ashleigh Manning1, Kate Templeton2, Ed. Gomperts3, Peter Simmonds1,2 1Centre for Infectious Diseases, University of Edinburgh 2Specialist Virology Laboratory, Royal Infirmary of Edinburgh 3Hospital for Sick Children, Los Angeles

  2. Human parvovirus infections • Human parvovirus B19 • Widespread in human populations • 3 genotypes, limited genetic heterogeneity • Acute, resolving infections, associated with intense viraemia • Recent evidence for long term persistence • Frequent detection in autopsy tissue, despite lack of persistent viraemia • Strong, life-long CTL reactivity to B19 antigens, suggests ongoing low-level replication • Novel human parvoviruses • Genome-based Virus discovery • Based on molecular methods to detect non-host DNA or RNA sequences within samples • Recent description of novel human parvoviruses (Allander et al., 2005; Jones et al., 2005)

  3. Novel Parvoviruses in humans • Parvoviridae • Wide range of diverse viruses infecting mammals • Highly host-specific • Acute resolving infections • Highly transmissible, stable in environment • Human Parvoviruses • Human Erythrovirus (B19) • PARV4 (Jones et al., 2005) • Acute infection syndrome • Little known about epidemiology • Human Bocavirus (Allander et al., 2005)

  4. Study Aims • Human Growth and Development Cohort • NIH-supported prospectively collected cohort • Recipients of non-virally inactivated factor VIII and IX concentrates • 6-monthly assessment and sample archiving. • Prospectively collected samples for > 10 years • Subject to several clinically-based and virological natural history studies • Edinburgh Respiratory Archive • LREC approval for construction of anonymous archives • Clinical and epidemiological information recorded, incapable of identifying specific patient • Sample type and month, donor code, age band, location codes, • Supplied clinical information, Results of other diagnostic tests (viral and bacteriological)

  5. Study Methods • PCR-based Parvovirus Detection • Highly conserved region identified in NS • Nested PCR with B19, PARV4 and HBoV-specific primers • Calibration and Run Controls • NIBSC Run control, calibrated to B19 International Standard • Quantified plasma samples containing PARV4 variant, PARV5 • Cloned, full length pre-quantified HBoV plasmid • All assays detected single copies of target sequence • Virus Screening • Nucleic acid extracted by Qiagen MinElute • 50 ul effective test volume for plasma Primers 1000 100 10 1 0.1 Neg PARV4(5) 8/8 16/16 16/16 8/16 1/16 0/8 HBoV 12/12 12/12 12/12 5/12 1/12 0/25

  6. Haemophilia Screen • Single samples from 59 haemophiliacs • Test sensitivity 20 DNA copies / ml • All sample negative for B19 and HBoV Primers Positive Tested Frequency Parvovirus B19 0 59 0% Human Bocavirus 0 59 0% PARV4/5 2 59 3.4% • Two samples positive for PARV4/5 • One haemophiliac HIV+/HCV+, one HCV+ only • Relatively high viral load, positive in 1st round • Genetic characterisation • One identical to PARV4 over 216 bases • One showed 14 substitutions, all synonymous (6.5%)

  7. Respiratory Screen • 942 respiratory samples from 589 individuals • Human Bocavirus • 53 positive from 37 individuals for HBoV • Almost invariably non-persistent, short period of excretion • Generally confined to infants and young children • Three adults with immunosuppression (transplant) showed persistent infections (2 from 3 with multiple samples), high titres • Parvovirus B19 • 4 positive from 3 individuals for B19 • 1 persistent infection in an immunosuppressed adult • PARV4/5 • All samples negative

  8. HBoV Epidemiology • Closely resembles RSV in epidemiology • Peak incidence December/January • Infections largely confined to < 2 years of age • Strongly associated with lower respiratory tract infections • Frequent HBoV / RSV or adenovirus coinfections • Potential exacerbating role in LRTIs

  9. Detection of parvovirus B19 and novel human parvoviruses in high-risk individuals Centre for Infectious Diseases, University of Edinburgh Ashleigh Manning Peter Simmonds Sick Children’s Hospital Los Angeles Ed Gomperts Specialist Virology Laboratory, Royal Infirmary of Edinburgh Kate Templeton ACKNOWLEDGEMENTS HGDS CoordinatingGroup Sally Baylis Tobias Allander

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