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Transmissibility of B19 by transfusion: evaluation of donor viremia prior to performing a linked donor-recipient study. M Busch, representing REDS-II Parvo B19 protocol team. RADAR Repository. Established from 2000-2003 in seven US cities NHLBI and CDC funded

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M Busch, representing REDS-II Parvo B19 protocol team

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Transmissibility of B19 by transfusion: evaluation of donor viremia prior to performing a linked donor-recipient study

M Busch, representing REDS-II

Parvo B19 protocol team


RADAR Repository

  • Established from 2000-2003 in seven US cities

  • NHLBI and CDC funded

  • Full enrollment for recipients included a pre (or peri) tx specimen and a follow-up specimen at 6-12 months

  • RADAR units (mostly RBC) were targeted to surgical patients on specific clinical services at specific hospitals

  • Enrolled recipients got both RADAR and non-RADAR units


Contents of RADAR repository

  • Donation specimens

    • Linked to fully enrolled recipients (n=13,201)

    • Unlinked(n=99.906)

  • Recipients

    • Fully enrolled (n=3575)

    • Initially enrolled but without f/u (n=1402)

  • There are two plasma aliquots (1.8 ml in first, up to 1.8 ml in second) and one 1.5 ml frozen whole blood aliquot for each specimen


Parvovirus B19 study hypotheses

  • Phase 1:To determine the rate of low, moderate, and high titre viremia and its coexistence with IgM and IgG antibody in blood donors.

    • Assay development

    • Donor viremia study

    • Recipient seroprevalance (for susceptibility)

    • Statistical evaluation

  • Phase 2: To determine the rate of B19 transmission by DNA-positive units and correlate it with donor viral load and antibody status


  • Testing methods

    • Qualitative PCR: magnetic bead capture kinetic Taqman assay- originally developed by Chiron and adapted by BSRI

    • Quantitative PCR: exact same assay - run in duplicate on an independent aliquot with quantitative standards on same run

    • Commercial EIAs (Biotrin) for IgM and IgG antibody: run in singlicate and repeated for equivocal results


    Sensitivity Panel Testing

    Results of 3 Sensitivity Panels tested before amplification of donor samples

    Cut-off: 40 Cycle Threshold


    Sensitivity Panel Results


    TESTING ALGORITHM

    PARVO B-19 PCR

    CT > 45,

    Undetected

    40 <CT ≤45

    CT≤40

    NEGATIVE

    INDETERMINATE

    REACTIVE

    Internal Control

    CT ≤45

    CT > 45, Undetected

    NEGATIVE

    INVALID

    CONFIRMATORY QUANTITATIVE TESTING , in duplicate

    CT: Cycle Threshold


    Control Chart, 100 IU/mL

    Mean+2SD

    Mean

    Mean-2SD

    Mean+/-2SD = 33.36+/-2.96; data for 56 plates (107 controls)


    PCR results: initial and repeat testing

    • Total tested: 5020

    • Initially reactive: 113 (2.25%)

    • Initially indeterminate: 26 (0.52%)

    • Initially invalid: 56 (1.12%)

    • Total Completed:4982

    • Confirmed positive: 44 (0.88%)

    • Indeterminate: 5 (0.10%)

    • Invalids pending conf: 38

    Low viremia Prevalence= 0.88%; 95% CI, 0.64% - 1.2%


    PCR and antibody results on confirmed positives

    Viral load#CPNo abIgM and IgGIgG only

    <20 21 1(5%) 2 (10%) 18 (86%)

    20-<102 11 1(9%) 2 (18%) 8 (73%)

    102-<103 9 0 5 (56%) 4 (44%)

    103-<104 3 0 3 (100%) 0

    Total 44 2(5%) 12 (27%) 30 (68%)

    • Of 23 donations with >20 IU/ML, median was 105 IU/mL,

      (42-481); highest was 1,869 IU/mL


    • PHASE II

    Exposed Recipients

    Non-exposed Recipients

    Test all (13,201) RADAR donation specimens linked to recipients for B19 DNA

    Random Sample

    Exposed Recipients tested for:

    Enrollment specimens: B19 IgG

    6M+ specimens: B19 DNA, IgM and IgG

    230 non-exposed recipients tested for: enrollment and 6M+ specimens: B19 DNA, IgM and IgG

    Tests Interpretation

    Tests Interpretation

    Newly acquired infection probably associated with transfusion of an infected RADAR donor unit

    =

    [RADAR + baseline] transmission rate

    Newly acquired infection not associated with transfusion of an infected RADAR donor unit

    =

    Baseline transmission rate

    Recipient infected pre-transfusion, not infected, or non-evaluable

    Recipient infected pre-transfusion, not infected, or non-evaluable

    RADAR transmission rate

    =

    {[RADAR + baseline] transmission rate} – {baseline transmission rate}


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