Mitigating the risk of transfusion-transmitted infections with vector-borne agents solely by means of pathogen reduction.

BACKGROUND: This study evaluated whether pathogen reduction technology (PRT) in plasma and platelets using amotosalen/ultraviolet A light (A/UVA) or in red blood cells using amustaline/glutathione (S-303/GSH) may be used as the sole mitigation strategy preventing transfusion-transmitted West Nile (WNV), dengue (DENV), Zika (ZIKV), and chikungunya (CHIKV) viral, and Babesia microti, Trypanosoma cruzi, and Plasmodium parasitic infections. METHODS: Antibody (Ab) status and pathogen loads (copies/mL) were obtained for donations from US blood donors testing nucleic acid (NAT)-positive for WNV, DENV, ZIKV, CHIKV, and B. microti. Infectivity titers derived from pathogen loads were compared to published PRT log10 reduction factors (LRF); LRFs were also reviewed for Plasmodium and T. cruzi. The potential positive impact on donor retention following removal of deferrals from required questioning and testing for WNV, Babesia, Plasmodium, and T. cruzi was estimated for American Red Cross (ARC) donors. RESULTS: A/UVA and S-303/GSH reduced infectivity to levels in accordance with those recognized by FDA as suitable to replace testing for all agents evaluated. If PRT replaced deferrals resulting from health history questions and/or NAT for WNV, Babesia, Plasmodium, and T. cruzi, 27,758 ARC donors could be retained allowing approximately 50,000 additional donations/year based on 1.79 donations/donor for calendar year 2019 (extrapolated to an estimated 125,000 additional donations nationally). CONCLUSION: Pathogen loads in donations from US blood donors demonstrated that robust PRT may provide an opportunity to replace deferrals associated with donor questioning and NAT for vector-borne agents allowing for significant donor retention and likely increased blood availability.

Patterns of Antibody Response to Severe Acute Respiratory Syndrome Coronavirus 2 Among 1.6 Million Blood Donors: Impact of Vaccination, United States, December 2020-June 2021.

From December 2020 to June 2021, 1654487 blood donors were tested for antibodies to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) S1 protein, and 1028547 (62.17%) were reactive. A rapid increase in prevalence was due to vaccination. Among a subset of 1567446 donors, 729771 (46.56%) reported SARS-CoV-2 vaccination, of whom 633769 (86.84%) were S1-antibody reactive only in response to vaccination and 68269 (9.35%) were reactive to both S1 and nucleocapsid in response to prior infection; the remainder were not reactive to either antibody. Among the 837675 (53.44%) donors who did not report vaccination, 210022 (25.07%) had reactivity to both antibodies and 29446 (3.52%) to S1 only.

Characteristics of US Blood Donors Testing Reactive for Antibodies to SARS-CoV-2 Prior to the Availability of Authorized Vaccines.

In the United States, many blood collection organizations initiated programs to test all blood donors for antibodies to SARS-CoV-2, as a measure to increase donations and to assist in the identification of potential donors of COVID-19 convalescent plasma (CCP). As a result, it was possible to investigate the characteristics of healthy blood donors who had previously been infected with SARS-CoV-2. We report the findings from all blood donations collected by the American Red Cross, representing 40% of the national blood supply covering 44 States, in order to characterize the seroepidemiology of SARS-CoV-2 infection among blood donors in the United States, prior to authorized vaccine availability. We performed an observational cohort study from June 15th to November 30th, 2020 on a population of 1.531 million blood donors tested for antibodies to the S1 spike antigen of SARS-CoV-2 by person, place, time, ABO group and dynamics of test reactivity, with additional information from a survey of a subset of those with reactive test results. The overall seroreactivity was 4.22% increasing from 1.18 to 9.67% (June 2020 - November 2020); estimated incidence was 11.6 per hundred person-years, 1.86-times higher than that based upon reported cases in the general population over the same period. In multivariable analyses, seroreactivity was highest in the Midwest (5.21%), followed by the South (4.43%), West (3.43%) and Northeast (2.90%). Seroreactivity was highest among donors aged 18-24 (Odds Ratio 3.02 [95% Confidence Interval 2.80-3.26] vs age >55), African-Americans and Hispanics (1.50 [1.24-1.80] and 2.12 [1.89-2.36], respectively, vs Caucasian). Group O frequency was 51.5% among nonreactive, but 46.1% among seroreactive donors (P< .0001). Of surveyed donors, 45% reported no COVID-19-related symptoms, but 73% among those unaware of testing. Signal levels of antibody tests were stable over 120 days or more and there was little evidence of reinfection. Evaluation of a large population of healthy, voluntary blood donors provided evidence of widespread and increasing SARS-CoV-2 seroprevalence and demonstrated that at least 45% of those previously infected were asymptomatic. Epidemiologic findings were similar to those among clinically reported cases.

Qualification of the Geenius HIV 1/2 supplemental assay for use in the HIV blood donation screening algorithm.

BACKGROUND: A single, simplified approach for human immunodeficiency virus (HIV)-1/HIV-2 antibody confirmation/differentiation is needed for the HIV blood donation supplemental algorithm used in the United States. A clinical evaluation of the Geenius assay was performed-the same assay used for HIV diagnostic confirmation/differentiation in the United States since 2014. STUDY DESIGN AND METHODS: Well-characterized unlinked donation samples classified as HIV negative, false positive, or confirmed positive were included in the study: 200 antibody-nonreactive, 200 HIV-1 immunofluorescence assay (IFA) confirmed-positive, and 100 antibody-screen false-positive donations, equally divided between serum and plasma. Samples were retrieved from a repository, relabeled, and tested by an immunochromatographic test (Geenius HIV 1/2 Supplemental Assay, Bio-Rad). Comparator testing involved parallel US Food and Drug Administration (FDA)-licensed HIV-1 IFA or HIV-2 enzyme immunoassay (EIA) supplemental testing for any sample missing those results as part of routine testing (otherwise test-of-record results were used). Samples with discordant results were further tested with a rapid test (Multispot HIV-1/HIV-2 Rapid Test, Bio-Rad) to provide final sample interpretations. Testing volume reductions with the Geenius were estimated from screening performed by the American Red Cross from September 2016 to April 2019. RESULTS: Clinical results were 100% sensitivity and specificity with an indeterminate rate of 4.0% to 5.0%. From 2016 to 2019, sole use of the Geenius would reduce testing complexity for 5265 antibody repeat-reactive donations including 95.7% (5028) false positives, eliminating approximately 5000 unnecessary tests. CONCLUSION: Geenius FDA licensure (August 26, 2019) adding the HIV-1/HIV-2 differentiation/confirmation donation supplemental claim will enable replacement of previously used FDA-licensed supplemental assays while maintaining comparable sensitivity, avoiding thousands of unnecessary HIV-1-IFA, western blot, and HIV-2-EIA tests.

Expanded feasibility of ferritin testing: stability of ferritin stored as whole blood and validation of plastic tubes.

BACKGROUND: Ferritin testing is a recommended strategy to mitigate iron depletion in blood donors. A barrier for some testing platforms is a requirement to complete sample management and testing within a temporal window incompatible with the logistics of many blood collectors. The ability to delay separation of plasma/serum from red cells and subsequent testing would enhance the feasibility of ferritin testing on a broader scale. STUDY DESIGN AND METHODS: Thirty blood donors provided a research donation of 12, 4-mL sample tubes of whole blood. Six pairs of serum and K2 -EDTA-plasma tubes were centrifuged and samples tested in triplicate on day of collection and on each of the next 5 days following storage at 4°C. Comparison of ferritin values for serum versus K2 -EDTA-plasma at baseline was performed to validate plastic EDTA-containing tubes. Variation of ferritin values during storage was assessed for direction and strength of any detectable changes. RESULTS: Ferritin values were comparable between EDTA-plasma and serum, with baseline values from EDTA-plasma samples 7% lower on average than serum (p < 0.0001 by paired t-test). Variability over five storage days was within approved parameters in the manufacturer's instructions. Within-run precision averaged 2% to 3% for each test day and within-subject precision across all samples averaged less than 5% for both serum and EDTA-plasma. Repeated measures showed no difference in changes during storage by tube type or day of testing. CONCLUSION: These results support flexible testing procedures, expanding the opportunity for blood centers to adopt this measure for assessing donor iron status.

Investigational Testing for Zika Virus among U.S. Blood Donors.

BACKGROUND: Because of the potential severe clinical consequences of Zika virus (ZIKV) infection, the large numbers of asymptomatic travelers returning from ZIKV-active areas, the detection of ZIKV nucleic acid in blood, and reports of transmission of ZIKV through transfusion, in 2016 the Food and Drug Administration released recommendations for individual-unit nucleic acid testing to minimize the risk of transmission of ZIKV through blood transfusions. METHODS: The American Red Cross implemented investigational screening of donated blood for ZIKV RNA by means of transcription-mediated amplification (TMA). Confirmatory testing of reactive donations involved repeat TMA, TMA testing in exploratory minipools, real-time reverse-transcriptase polymerase chain reaction, IgM serologic testing, and red-cell TMA. Viral loads in plasma and red cells were estimated by means of end-point TMA. The costs of interdicting a donation that was confirmed to be positive were calculated for the 15-month period between June 2016 and September 2017. RESULTS: Of the 4,325,889 donations that were screened, 393,713 (9%) were initially tested in 24,611 minipools, and no reactive donations were found. Of the 3,932,176 donations that were subsequently tested individually, 160 were initially reactive and 9 were confirmed positive (a 1:480,654 confirmed-positive rate overall; positive predictive value, 5.6%; specificity, 99.997%). Six (67%) of the confirmed-positive donations were reactive on repeat TMA, of which 4 were IgM-negative; of these 4, all 3 that could be tested were reactive on minipool TMA. Two confirmed-positive donors had infections that had been transmitted locally (in Florida), 6 had traveled to ZIKV-active areas, and 1 had received an experimental ZIKV vaccine. ZIKV RNA levels in red cells ranged from 40 to 800,000 copies per milliliter and were detected up to 154 days after donation, as compared with 80 days of detection in plasma at levels of 12 to 20,000 copies per milliliter. On the basis of industry-reported costs of testing and the yield of the tests in our study, the cost of identifying 8 mosquito-borne ZIKV infections through individual-unit nucleic acid testing was $5.3 million per ZIKV RNA-positive donation. CONCLUSIONS: Screening of U.S. blood donations for ZIKV by individual-donation TMA was costly and had a low yield. Among the 9 confirmed ZIKV-positive donations, only 4 were IgM-negative; of these donations, all 3 that were tested were reactive on minipool TMA. (Funded by the American Red Cross and Grifols Diagnostic Solutions.).

Two human immunodeficiency virus Type 2 cases in US blood donors including serologic, molecular, and genomic characterization of an epidemiologically unusual case.

BACKGROUND: Blood donation screening for human immunodeficiency virus Type 2 (HIV-2) has been in place in the United States since 1992. However, only three HIV-2 antibody-positive donors have been reported to date, all detected via HIV-1 cross-reactivity. STUDY DESIGN AND METHODS: Here we identify two additional HIV-2-positive donors by routine anti-HIV-1 and anti-HIV-2 screening, including a first-time male donor living in Georgia having recently immigrated to the United States from West Africa (from a 1998 donation) and a Taiwanese female repeat donor (nurse) living in California with no travel outside of Taiwan or apparent connections to West Africa (from a 2015 donation). Neither donor acknowledged any risk factors, and both remained asymptomatic through follow-up. The second donor was further investigated by serologic, molecular, and genomic assays because of her unusual demographics. She was documented to harbor HIV-2 RNA, albeit sporadically by HIV-2-specific nucleic acid tests (35%-100% of replicates) and at very low levels (<9.6 IU/mL). Metagenomic next-generation sequencing (mNGS) confirmed the identification of a Group B HIV-2 strain, with recovered reads covering 46.9% of the predicted genome. CONCLUSIONS: The estimated frequency of an HIV-2-positive blood donor in the United States is one in 57 million donations. Due to the low frequency and low pathogenicity of HIV-2, public health and blood donation screening efforts must focus on HIV-1 detection and prevention. However, detection of HIV-2 infection in a donor with no apparent link to West Africa suggests that the United States must remain vigilant for HIV-2 virus infections. Ultradeep mNGS may be useful in the future for comprehensive identification of rare transfusion-transmissible agents.

Comparative analysis of triplex nucleic acid test assays in United States blood donors.

BACKGROUND: This study assessed the clinical sensitivity of three fully automated, human immunodeficiency virus (HIV), hepatitis C virus (HCV), and hepatitis B virus (HBV) triplex nucleic acid test (NAT) assays by individual donation (ID-NAT) and at operational minipool (MP-NAT) sizes used worldwide. STUDY DESIGN AND METHODS: MPX, Ultrio, and Ultrio Plus were used to test 2222 pedigreed, marker-positive samples with varying viral loads, each from a unique US blood donor. NAT-positive, seronegative yield samples (16 HBV, 156 HCV, and 23 HIV) were tested in replicates of three; undiluted; and in 1:6, 1:8, and 1:16 dilutions (MP6, MP8, and MP16), simulating various MP sizes. Seropositive samples (1276 HBV, 488 HCV, and 263 HIV) were tested by ID-NAT in singlet. RESULTS: MPX-MP6 and Ultrio Plus-MP16 had equivalent HCV sensitivity. Although Ultrio Plus-MP16 for HIV trended toward lesser sensitivity, this was not corroborated in a large substudy of low-viral-load samples in which Ultrio Plus-MP8/MP16 showed 100% reactivity. MPX-ID and Ultrio Plus-ID HBV clinical sensitivity were identical, but MPX-MP6 was significantly more sensitive than Ultrio Plus-MP16; the differential yield projected to one HBV NAT yield per 4.72 million US donations. Ultrio Plus HBV sensitivity did not increase at MP8 versus MP16. Ultrio Plus versus Ultrio sensitivity was significantly increased in HBV-infected donors with early acute, late acute or chronic, and occult infections. No difference in sensitivity was noted for any virus for MPX-MP6 versus Ultrio Plus-ID. CONCLUSIONS: Our data support US donation screening with MPX-MP6 or Ultrio Plus-MP16 since the HBV DNA detection of Ultrio Plus was significantly enhanced (vs. Ultrio) without compromising HIV or HCV RNA detection.

The value of screening signal-to-cutoff ratios for hepatitis C virus antibody confirmation.

A licensed recombinant immunoblot assay (RIBA) has been an integral component of the algorithm for confirmation of reactivity for antibody to hepatitis C virus in screening assays. This test may become unavailable in the future, so we have investigated the potential value of screening test signal values to support confirmation. Depending on the screening test used (Ortho enzyme-linked immunosorbent assay or Abbott PRISM chemiluminescent assay), signal-to-cutoff ratios of 5.00 and 3.20, respectively, were shown to have positive predictive values of 89.1 and 95%, with sensitivities of 93.1 and 88.7% relative to RIBA. However, additional steps will have to be added to assure equivalent performance to RIBA.

Sensitivity comparison of two Food and Drug Administration-licensed, triplex nucleic acid test automated assays for hepatitis B virus DNA detection and associated projections of United States yield.

BACKGROUND: There have been no comparisons of the relative sensitivity of the two Food and Drug Administration-licensed multiplex (MPX) nucleic acid test (NAT) systems (Procleix Ultrio [Gen-Probe], TIGRIS platform [Novartis]; and cobas TaqScreen MPX [Roche Molecular Systems], cobas s 201 platform [Roche Instrument Center]) for detecting hepatitis B virus (HBV)-infected donors in minipool sizes (MP) used in the United States. STUDY DESIGN AND METHODS: Routine blood samples from Thailand were obtained from plasma units from 129 hepatitis B surface antigen (HBsAg)-negative, HBV NAT-yield donations. Blinded US testing included antibody to hepatitis B core antigen (anti-HBc), NAT using both manufacturers' systems (undiluted-individual donation [ID], in singlet and diluted 1:6 and 1:16 in triplicate), quantitative antibody to hepatitis B surface antigen, HBV DNA viral loads, and HBV genotyping. HBV yields in the United States were estimated using the incidence/window period (WP) model and compared to the calculated assay sensitivities. RESULTS: Eighty samples were classified as occult HBV (anti-HBc reactive) and 49 as WP (anti-HBc nonreactive). For US pool sizes, MPX detected significantly more samples than Ultrio (MPX MP6 vs. Ultrio MP16; p < 0.0001 for occult and WP). Ultrio MP16 results were not statistically different from Ultrio MP6 (p = 0.68 for occult; p = 0.42 for WP). There was no difference between platforms for MP sizes used in most of the world (MPX MP6 vs. Ultrio ID; p = 0.70 for occult and p = 0.34 for WP). Viral loads were higher in WP samples. Modeled yield estimates were consistent with measured assay sensitivity on the Thai donor samples. CONCLUSIONS: As used in the United States, MPX MP6 is more sensitive than Ultrio MP16, but the impact of this difference is mitigated by low numbers of HBV WP infections.

Human T-lymphotropic virus antibody screening of blood donors: rates of false-positive results and evaluation of a potential donor reentry algorithm.

BACKGROUND: Blood donor screening with enzyme immunoassays (EIAs) for antibodies to human T-lymphotropic virus (HTLV)-I, and later to HTLV-I/II, has led to the unnecessary deferral of tens of thousands of individuals. The licensure of the Abbott PRISM HTLV-I/HTLV-II chemiluminescent immunoassay (ChLIA) may permit the reinstatement of historically deferred donors. STUDY DESIGN AND METHODS: The efficacy of a reentry algorithm involving a follow-up sample from EIA-deferred donors testing HTLV-I/II ChLIA nonreactive was evaluated using 386 serologic confirmed-positive samples archived since the inception of anti-HTLV donor screening. Reactivity of the 386 samples by the ChLIA, when coupled with the package insert sensitivity data, may be used to demonstrate efficacy of the reentry algorithm. Donor incidence was also examined from 2008 through 2009 to evaluate changes to the existing HTLV screening policy. RESULTS: From January 1, 1995, to April 28, 2008, a total of 64,052 donors to the American Red Cross were deferred solely because of HTLV EIA false positivity, representing more than 130,000 US donors. HTLV ChLIA identified 386 confirmed-positive donations from 386 randomly selected donors representing reactivity to both the bioMérieux and the Abbott HTLV-I/II EIAs (95% confidence interval [CI], 99.2%-100%); both EIAs have since been discontinued. This is comparable to the detection of 843 of 843 confirmed-positive samples during the ChLIA clinical trials (95% CI, 99.48%-100%). Incident HTLV infections occurred primarily among female repeat donors during 2008 throughout 2009. CONCLUSIONS: Donors testing falsely positive by historic EIAs since 1988 should be considered for reinstatement if a contemporary sample tests ChLIA nonreactive. Changes to the existing screening algorithm seem unlikely since new HTLV infections were detected among repeat donors.

Integration of nucleic acid amplification test results into hepatitis C virus supplemental serologic testing algorithms: implications for donor counseling and revision of existing algorithms.

BACKGROUND: The routine use of hepatitis C virus (HCV) nucleic acid amplification testing (NAT) donor screening assays has provided an opportunity for revision of the current HCV supplemental testing algorithm, which requires that recombinant immunoblot assay (RIBA) be performed on every HCV enzyme immunoassay (EIA)-repeat-reactive donation. The FDA has approved variance requests to use a new algorithm that eliminates the need to perform RIBA when HCV NAT results are reactive. Data are provided in support of this new algorithm. STUDY DESIGN AND METHODS: HCV EIA (including signal-to-cutoff optical density ratio), RIBA, and NAT data were compiled from 33.2 million donations screened over an approximately 4-year period by the American Red Cross and Blood Systems Laboratories. Further, donations having specific combinations of HCV EIA, RIBA, and minipool (MP) NAT results were evaluated, with more sensitive individual-donation (ID) NAT, to construct improved counseling messages for donors. RESULTS: Of 47,041 EIA-repeat-reactive donations, 49.3 percent were RIBA-positive, 17.1 percent RIBA-indeterminate, and 33.5 percent RIBA-negative. NAT-reactive rates were 79.2, 2.5, and 0.18 percent for RIBA-positive, -indeterminate, and -negative donations, respectively. The new algorithm classified an additional 1 percent of donations as HCV-infected while at the same time detecting all infections classified as HCV-infected under the current algorithm. An additional 2.4 percent of RIBA-positive, MP NAT-nonreactive donations were reactive when a frozen-thawed aliquot was retested by ID NAT. CONCLUSION: Integrating HCV NAT results with RIBA results for purposes of donor notification allows more appropriate counseling messages to be given to EIA-repeat-reactive donors. The new HCV supplemental algorithm is an acceptable alternative to the current algorithm because it provides equivalent or superior accuracy in formulating donor counseling messages and may also result in reduced costs and more timely notification of infected donors.

West Nile virus among blood donors in the United States, 2003 and 2004.

BACKGROUND: West Nile virus first appeared in the United States in 1999 and has since spread throughout the contiguous states, resulting in thousands of cases of disease. By 2002, it was clear that the virus could be transmitted by blood transfusion, and by the middle of 2003, essentially all blood donations were being tested for West Nile virus RNA with the use of investigational nucleic acid amplification tests; testing was performed on individual samples or on "minipools" of up to 16 donations. METHODS: We analyzed data from the West Nile virus testing program of the American Red Cross for 2003 and 2004 to identify geographic and temporal trends. In areas with a high incidence of infection, individual donations were tested to increase the sensitivity of testing. Donors with reactive results participated in follow-up studies to confirm the original reactivity and to assess the natural history of infection. RESULTS: Routine testing in 2003 and 2004 identified 540 donations that were positive for West Nile virus RNA, of which 362 (67 percent) were IgM-antibody-negative and most likely infectious. Of the 540 positive donations, 148 (27 percent) were detectable only by testing of individual donations, but only 15 of the 148 (10 percent) were negative for IgM antibody. The overall frequencies of RNA-positive donations during the epidemic periods were 1.49 per 10,000 donations in 2003 and 0.44 per 10,000 in 2004. In 2004, 52 percent of the positive donations were from donors in four counties in southern California. CONCLUSIONS: Rapid implementation of a nucleic acid amplification test led to the prospective identification of 519 donors who were positive for West Nile virus RNA and the removal of more than 1000 potentially infectious related components from the blood supply of the Red Cross. No cases of transfusion-transmitted infection were confirmed among recipients of the tested blood.

Impact of HCV 3.0 EIA relative to HCV 2.0 EIA on blood-donor screening.

BACKGROUND: In 1996, the Ortho HCV Version 3.0 ELISA Test System (HCV 3.0 EIA) was licensed in the United States for donor screening but was neither mandated nor universally implemented. Data from two studies comparing the differential performance of HCV 3.0 EIA and HCV 2.0 EIA are presented. The first study evaluated the differential performance in a cross-section of screened whole-blood donors after implementation of HCV 3.0 EIA; the second study evaluated the differential performance of HCV 3.0 EIA in plasma donors acutely infected with HCV, identified during routine Abbott HCV 2.0 EIA and HCV NAT (using Roche Ampliscreen plate assay) donor screening. STUDY DESIGN AND METHODS: The first study evaluated HCV 3.0 EIA repeat-reactive donations from four US blood centers, identified during the first 5 months of HCV 3.0 EIA implementation. HCV EIA repeat-reactive donations confirmed by RIBA HCV 3.0 SIA were retested using both Ortho HCV Version 2.0 ELISA Test System and Abbott HCV 2.0 EIA. All EIA-discordant donations were tested by polymerase chain reaction (PCR). In the second study, Abbott HCV 2.0 EIA-nonreactive, HCV PCR-positive donors were enrolled in a follow-up study in which the index and follow-up samples were re-evaluated by HCV 3.0 EIA. RESULTS: In the first study, of 292,459 donations, 501 (0.17%) confirmed HCV 3.0 EIA-reactive donations were identified; 15 (0.005%) were nonreactive by Ortho HCV 2.0 EIA and were all HCV RNA negative. In the second study, Ortho HCV 3.0 EIA retesting of Abbott HCV 2.0 EIA-nonreactive, RNA-positive index donations identified 16 (23%) as 3.0 EIA reactive. In 42 panels with a discordant time of seroconversion, HCV 3.0 EIA sero-conversion preceded HCV 2.0 EIA in all cases (p < 0.001). Two donors with HCV 3.0 EIA-reactive index donations never seroconverted by HCV 2.0 EIA during 160 to 180 days of follow-up. CONCLUSION: These studies demonstrate that HCV 3.0 EIA compared to HCV 2.0 EIA can better detect 1) remote nonviremic HCV infections, 2) acute infection, and 3) HCV antibodies in cases of atypical seroconversion.