Celso Bianco

Protecting the Blood Supply From Emerging Pathogens: The Role of Pathogen Inactivation

n n Although the risk of infection by blood transfusion is relatively low, breakthrough infections still occur, Transfusion-related fatalities caused by infections continue to be reported, and blood is not tested for many potentially dangerous pathogens. The current paradigm for increasing the safety of the blood supply is the development and implementation of laboratory screening methods and restrictive donor criteria. When considering the large number of known pathogens and the fact that pathogens continue to emerge, it is clear that the utility of new tests and donor restrictions will continue to be a challenge when considering the cost of developing and implementing new screening assays, the loss of potential donors, and the risk of testing errors. Despite improving the safety of blood components, testing remains a reactive approach to blood safety. The contaminating organisms must be identified before sensitive tests can be developed. In contrast, pathogen inactivation is a proactive strategy designed to inactivate a pathogen before it enters the blood supply. Almost all pathogen inactivation technologies target nucleic acids, allowing for the inactivation of a variety of nucleic acid–containing pathogens within plasma, platelets, or red blood cells thus providing the potential to reduce transfusion-transmitted diseases. However, widespread use of a pathogen inactivation technology can only be realized when proven safe and efficacious and not cost-prohibitive.n n

West Nile Virus Infections Projected from Blood Donor Screening Data, United States, 2003

Routine donor nucleic acid amplification testing is a valuable surveillance screening tool.

The 2003 West Nile virus United States epidemic: the America's Blood Centers experience

BACKGROUND: A detailed assessment of West Nile virus (WNV) yield is needed to evaluate the effectiveness of the WNV nucleic acid amplification technology (NAT) screening implemented in 2003.

Selection Criteria to Protect the Blood Donor in North America and Europe: Past (Dogma), Present (Evidence), and Future (Hemovigilance)

The safety of the blood supply depends on measures to protect not only the transfusion recipient but also the blood donor. Donor selection criteria have been voluntarily adopted or enforced through regulation in different countries, but review of practices in different blood centers reveals wide disparity in the current approaches. Such variability in practice suggests that the criteria for the protection of donor are often arbitrary or reflect deeply engrained precautionary practices and exposes the inherent uncertainty about the best way to minimize risk to the donor. Certain selection criteria introduced years ago have become dogma in some countries but were never subjected to systematic study and persist despite available evidence that the measures do not measurably improve donor safety. Current efforts to define a rational, evidence-based approach are crucial to eliminate practices that lead to the unnecessary deferral of large numbers of blood donors without improving the safety of the donation process. Future prospects to improve the safety of the donation process rest with hemovigilance initiatives to monitor the effectiveness of interventions to minimize the risks to blood donors.

The impact of discontinuation of 7-day storage of apheresis platelets (PASSPORT) on recipient safety: an illustration of the need for proper risk assessments

BACKGROUND: Seven‐day stored apheresis platelets (APs) were withdrawn from the US market after detection of two culture‐positive units from 2571 tested at outdate in the PASSPORT surveillance study. The impact of this discontinuation on recipient safety was explored using mathematical modeling.

Deferral of males who had sex with other males

Donor history questionnaires for the determination of blood donor eligibility are a critical layer of blood safety. Early in the course of the AIDS epidemic in North America homosexual men with multiple partners were identified as one of the segments of the population with the highest risk of infection. Voluntary deferral of this group from blood donation led to a dramatic decrease in transfusion-transmitted HIV even before testing was introduced. In the early 1980s blood donors were deferred in England, the US and other nations, if they were ‘homosexual males with multiple partners’. After the implementation of HIV testing in 1985, the majority of the HIV-positive donors identified revealed ‘men having sex with men’ (MSM) behavior, leading the US Food and Drug Administration (FDA) to recommend indefinite deferral of all men who ‘have had sex with men, even once since 1977’; many other regulators and jurisdictions have enacted similar criteria. Three decades later, despite the recognition of other modes of transmission, MSM donors are still among the population segments with the highest prevalence and incidence of HIV in countries around the world. No other donor eligibility criterion has generated as much controversy or public discourse [1,2]. Proponents for change point out that in many countries other key components of blood safety such as donor testing and blood center process control have improved vastly, reducing the contribution of donor questioning to safety. Gay advocates in particular argue that donor selection policies based on MSM are discriminatory against gay and bisexual men in that they amount to a de facto permanent exclusion on the grounds of sexual preference, and are unfair, as other groups with similar risks of HIV infection are allowed to donate blood after shorter time-period deferrals designed to cover the seroconversion window. On the opposite side of the discussion, recipient advocacy groups and regulators are understandably adverse to any change that is not centered on improving safety. Recipient groups argue that they have suffered greatly due to transmission of HIV and HCV by transfusion, and they will be the bearers of any increase in risk that may result from policy changes. Because both MSM and recipients are vulnerable groups that have suffered in the past, the debate over possible changes in criteria has ethical, societal, and emotional dimensions not seen in discussions concerning other donor selection criteria. Of particular concern to blood operators is the prospect that young eligible donors may be dissuaded from donating blood to institutions that are perceived to act in a discriminatory and unfair fashion. This International Forum seeks to describe approaches to this issue and challenges to the status quo, in a snapshot in time. Since it is extremely difficult to obtaindatatoevaluatethepossibleimpactofpolicy changes made to address concerns expressed by advocacy groups, comparison of international practice is particularly valuable, since we may learn from approaches implemented in other jurisdictions. We received responses from 24 respondents representing countries on six continents. In most, but not all, the MSMpolicy isdetermined atthe national level. The following questions were asked of the respondents:

Dengue and Chikungunya viruses in blood donations: risks to the blood supply?

A rbovirus epidemics are raging in tropical areas. Dengue virus (DENV), dengue shock syndrome (DSS), and dengue hemorrhagic fever (DHF) affect millions of individuals every year and cause significant mortality in Latin America, Africa, and Asia. Chikungunya virus (CHIKV) has caused recurrent epidemics in the Indian subcontinent and recent epidemics in Reunion and other islands in the Indian Ocean, with recent detection in areas of Europe. This issue of TRANFUSION includes two articles on the detection of DENV RNA in blood donors from epidemic areas, one article on the clearance of spiked DENV in the course of the manufacture of plasma derivatives, and one article on statistical modeling of transfusion risk during an epidemic of mosquito-borne CHIKV. The surprising seriousness of recurring epidemics of West Nile virus (WNV) in North America has heightened concerns about the potential for introduction and similar epidemic spread of other arbovirus infections in the United States. Dengue has received particular attention since cases have been recognized in the United States at the border between Texas and Mexico. The mosquito species that transmit DENV, Aedes egypti and Aedes albopictus, are present in the Southern and Southeastern parts of the United States, raising the specter of significant spread within the United States if the virus is introduced and efficiently spreads. Does dengue represent a risk to the safety of the US blood supply? Transmission by transfusion (TT) is often difficult to evaluate in the midst of an epidemic because the infection could have been acquired through a mosquito bite, through a transfusion, or even through a needle stick. Doubts about TT of WNV were eliminated when the virus was transmitted to multiple recipients of organs and, subsequently, multiple recipients of blood components obtained from the blood donors determined to be viremic. Despite the recognition of millions of cases of DENV infection and disease every year, there are very few published reports of transfusion transmission. One report was submitted to the public health agency of Hong Kong and, although accessible online, has not been published in a peer-reviewed journal. The other report describes a recent cluster of TT-DENV in Singapore, which is currently in press (P.A. Tambyah, National University of Singapore, personal communication, 2008). There are also reports of transmission by needle sticks and one case associated with a marrow transplant in Puerto Rico (references are included in the articles being published). Considering the available information, should precautionary measures be considered to prevent TT-DENV in the United States or even more important in countries in which DENV epidemics are occurring at expanding rates? What are the data required for decisions to implement measures that mitigate this potential risk? The articles published in the current issue of TRANSFUSION contribute data that will be needed for policy decisions that may be required in the not so distant future. Linnen and coworkers used a transcriptionmediated amplification (TMA) assay for detection of DENV RNA to screen 13,372 specimens collected from blood donors in Honduras, Brazil, and Australia, countries with ongoing seasonal dengue epidemics. They identified 9 donors in Honduras who were repeat reactive for DENV RNA by TMA. Eight were confirmed by a polymerase chain reaction (PCR) assay that identified three different DENV serotypes, DENV-1, -2, and -4. Infectious virus could be recovered from four of these donors. Three samples from Brazil were repeatedly reactive on TMA. One was typed as DENV-1 and the other as DENV-3. None of the Australian specimens was repeatedly reactive. Samples were also tested by enzyme-linked immunosorbent assay for immunoglobulin M and immunoglobulin G antibodies to DENV and by plaque reduction neutralization assay to determine the existence and serotype of prior DENV infections in the viremic donors. Five of the donors in Honduras and one of the donors in Brazil appeared to have secondary DENV infections. The authors discuss the implications of reinfection with heterotypic subtypes because of the increased risk of DHF in these individuals. The study thus documented the presence of asymptomatic viremic donors in Honduras and Brazil who could theoretically transmit the virus to blood recipients. Mohammed and colleagues screened specimens from 16,521 blood donations made during an 11-week period to the American Red Cross in Puerto Rico, starting 2 weeks after the peak of dengue activity at the end of 2005. Twelve were DENV RNA repeatedly reactive using the same TMA assay used by Linnen and coworkers; 5 of the 12 were reactive in a pool of 16 (the operational pool size for WNV using TMA), and 4 of the 5 were positive on PCR. The four PCR-positive samples had viral loads TRANSFUSION 2008;48:1279-1281.

Detection of West Nile virus RNA and antibody in frozen plasma components from a voluntary market withdrawal during the 2002 peak epidemic.

BACKGROUND: The US West Nile virus (WNV) epidemic in the summer and fall of 2002 included the first documented cases of transfusion‐transmitted WNV infection. In December 2002, the FDA supported a voluntary market withdrawal by the blood banking community of frozen blood components collected in WNV high‐activity areas. At the time, the prevalence of viremia and serologic markers for WNV in the blood supply was undefined.

Technical considerations for the performance of Nucleic acid Amplification Technology (NAT): The NAT Task Force Group

The complexity of Nucleic acid Amplification Technology (NAT(1)), comprising sample preparation, amplification and detection methods, requires specific design considerations for both the laboratory and the procedures utilized in such testing. The purpose of this paper is to establish technical considerations for the performance of NAT. These include the collection, handling and assay of specimens and the design of laboratories to routinely and reliably detect low levels of nucleic acid sequences. The sensitivity of NAT due to the exponential amplification of nucleic acids makes contamination a major concern from specimen collection to sample detection. Therefore, laboratories need to be designed to prevent and control contamination through adequate equipment and appropriate workflow. These technical considerations should provide a basis for establishing a robust and reproducible NAT system.

Infectivity of occult hepatitis B from two different points of view

T wo manuscripts in this issue of TRANSFUSION assess the risk of transmission of occult hepatitis B (OBI) by transfusion and approaches to its prevention in Japan and in Europe. OBI is defined as the presence of circulating hepatitis B virus (HBV) DNA as detected by HBV nucleic acid test (NAT), in the absence of detectable HBV surface antigen (HBsAg), excluding the window period. During the window period of infection, HBV DNA is present but the immune system has not yet had the opportunity to generate antibodies to the several antigens of HBV, particularly HBV core (anti-HBc) and HBV surface (anti-HBs) antigens. In OBI anti-HBc is always detectable. In contrast, successful vaccination induces the formation of anti-HBs, which is protective, and HBV DNA is absent. It should be noted that the classification of an infection as OBI is entirely dependent on the sensitivity of the assays being used to detect HBV antigens, antibodies, and DNA. The name “occult HBV” can and is often misinterpreted because it conveys the impression that the etiologic agent is hidden somewhere in the body of the infected individual and is being missed by diagnostic tests, reminding us of “AIDS without HIV,” a suggestion that caused major panic in the early 1990s and reignited concerns about the tragic epidemic of the early 1980s. While HIV is the etiologic agent of AIDS, HBsAg is not the agent of hepatitis B. It is part of a family of small particles made of proteins and lipids that are produced and released by HBV-infected cells into the blood stream in high concentration at the same time that infectious viral particles are released. HBsAg can circulate with infectious HBV virus particles but can also circulate in the absence of HBV particles; HBsAg consequently is not itself infectious. However, it was the first recognized HBV antigen and became the most prominent and effective target of immunoassays for diagnosis of infection and for screening of blood donors. Until the more recent development of molecular assays, serologic assays for HBsAg have been the most important contributors to the safety of the blood supply with regard to hepatitis B. Taira and colleagues from the Japanese Red Cross (JRC) estimated the residual risk of transmission of HBV by donors with OBI. JRC had been screening blood donations for HBV using hemagglutination for HBsAg. Since 2008 all hemagglutination tests have been replaced by chemiluminescent enzyme immunoassay (CLEIA). JRC rejects donations that are positive for HBsAg and subjects negative donations to further determination of titers of antiHBc and anti-HBs. It accepts donations with high titers of anti-HBs ( 200 IU/L) regardless of the titer of anti-HBc. However, it rejects donations with a high titer of anti-HBc and low titer of anti-HBs (Table 1 in Taira et al.). In addition, all donations are tested for HBV DNA using minipool NAT (currently pools of 20 donations; previously pools of 50). Since four to 13 transfusion-transmitted infections continue to occur annually, the investigators performed retrospective individual-donation HBV NAT (ID-NAT) on a large number of samples from lookback and traceback cases that were stored in their extensive repository. Whenever possible they tested follow-up samples from recipients with suspected HBV transmission by transfusion. The donations tested by ID-NAT came from donors who had screened as negative for HBV DNA by HBV NAT performed in pools of 20 to 50 donations. The authors also determined the impact of changes in their NAT screening algorithm since the introduction of molecular testing for HBV in 1999. When they compared the more sensitive TaqScreen currently performed in pools of 20 donations with the early AmpliNAT performed in pools of 50 donations they observed that the OBI detection rate increased from 3.9 to 15.2 per million and that the yield of window period donations decreased from 13.2 per million to 5.7 per million (Table 2 in Taira et al.). The counterintuitive decline in yield of window phase NATyield donations after the reduction in pool size likely resulted from the introduction of the more sensitive CLEIA test for HBsAg in 2008, although a decline in HBV incidence may have also contributed to the overall reduction in DNA-only window phase infections. The investigators concluded that approximately 2% of the donors with low titers of anti-HBc and anti-HBs presented low-level viremia and that this viremia did not correlate with anti-HBc titers, leading them to suggest that the best strategy to prevent transmission of HBV by donors with OBI requires elimination of all donations with low titers of anti-HBc and anti-HBs. In prior studies, JRC had estimated that the risk of transmission of HBV associated with donations with low titers of anti-HBc was 2.4% to 3.0%, 10 times lower than that of donors in the window period of HBV infection where the infectivity was estimated at 27%. The authors calculate, based on the 3% rate of transfusion transmission attributed to OBI, that there would be 47 OBI transfusion transmissions that ID-NAT would prevent but some infectious units would also be missed due to fluctuating low-level TRANSFUSION 2013;53:1379-1381.