AABB Chapter 08

C h a p t e r

8

Infectious Disease Screening 䊱 Eberhard W. Fiebig, MD, and Michael P. Busch, MD, PhD

H E P A S T T W O decades have seen tremendous improvement in infectious disease screening of blood donations, which has resulted in a measurably safer blood supply.1 However, the ultimate goal of eliminating infectious disease transmission by transfusion has not yet been achieved, and concerns remain about residual risks from “classical” transfusion-transmitted pathogens such as human immunodeficiency virus (HIV), hepatitis B (HBV), and hepatitis C (HCV), as well as other viruses, bacteria, and parasites. In addition, a growing list of so-called emerging infectious threats to the blood supply, including West Nile virus, (WNV) Chagas’ disease, and variant Creutzfeldt-Jakob disease (vCJD), are also of concern.

This chapter briefly describes the types of infectious pathogens that are known or have the potential to be transmissible by transfusion. Discussion then focuses on the systems and practices designed to prevent these agents from entering blood components and plasma derivatives. Also addressed are supplemental testing for donor counseling, look-back, in-

T

fectious risk of plasma derivatives, and pathogen reduction and inactivation methods for blood components and derivatives.2 TR A N SF USI O N-TR A N SM IS SI BL E PA TH O GE N S Viruses Table 8-1 lists profiles of common pathogens and agents that have the potential to be transmitted by infusible blood products. For ease of discussion, these pathogens are grouped according to taxonomic classifications and properties related to transmissibility by transfusion.

Classical Transfusion-Transmitted Viruses Viruses included in this group are the bloodborne pathogens HIV types 1 and 2 (HIV-1, HIV-2), HBV, and HCV. Certain features of these viruses place them in a category of special concern for recipients of blood and infusible blood products: 1) perennial low-level prevalence among potential blood donors

Eberhard W. Fiebig, MD, Associate Professor of Clinical Laboratory Medicine, University of California, San Francisco (UCSF), Chief, Laboratory Medicine Service, San Francisco General Hospital, and Michael P. Busch, MD, PhD, Professor of Laboratory Medicine, UCSF, and Vice President, Research, Blood Systems Research Institute, San Francisco, California The authors have disclosed no conflicts of interest.

241 Copyright © 2008 by the AABB. All rights reserved.

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TABLE 8-1. Infectious Pathogens and Risk of Transmission by Transfusion Classification and Description

Epidemiology

Disease Manifestation

Estimated US Transmission Risk per Unit Transfused

Hepatitis B virus (HBV)

Member of Hepadnaviridae family. Double-stranded DNA virus with lipid envelope. Six genotypes (A-F) have been described.

Parenteral, sexual, and vertical (pregnant mother to infant) transmission routes. Seroprevalence in US population is about 5.6%; in some parts of the world, it is >50%.

Majority of infections are asymptomatic; jaundice may be present in 30%, and fulminant hepatitis with fatal outcome may be seen in 0.5%-1%. Chronic infections are common in infancy and childhood, but they occur in 5% or less of adults.

1:220,0001

Hepatitis C virus (HCV)

Member of Flaviviridae family, genus Hepacivirus. Small, enveloped, single-stranded RNA virus. Six genotypes (16) have been described.

Parenterally transmitted primarily through illicit drug use, transfusions, and clotting factor use before availability of sensitive screening tests and effective pathogen reduction and inactivation methods. Sexual and mother-to-infant transmission is uncommon, though increased in those co-infected with HIV.

Majority of infections are asymptomatic; nonspecific symptoms and jaundice are present in up to 20%-30% of cases. Approximately 80% of infections become chronic, many lifelong with late risk of liver cirrhosis and a small percentage risk of malignancy.

1:1,800,0003

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AABB TECHNICAL MANUAL

Classical Transfusion-Transmitted Viruses

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Pathogen

Human immunodeficiency virus (HIV-1,-2)

Parenteral transmission by means of the usual routes; sexual and vertical transmission. HIV-1 is found worldwide with varying subtype distribution. Subtype B is most common in North America and Europe; other subtypes dominate elsewhere. HIV-2 is found mainly in West Africa with some spread to Europe.

Acute (flu-like) retroviral syndrome in approximately 40% of cases within a few weeks to months of infection. This reaction is followed by long (up to 10 years or longer) asymptomatic period during which the immune system is progressively weakened. Untreated HIV is usually fatal from opportunistic infections and other HIV-related complications.

1:2,300,0003

Parenteral transmission.

Vertical transmission thought to be rare, but there is a relatively high risk (20%-30%) of transmission from mother to infant through breastfeeding.

Infected persons are frequently asymptomatic. HTLV-I infection is associated with increased risk of an unusual malignancy (T-cell leukemia/lymphoma), neurologic (HTLV-associated myelopathy/ tropical spastic paraparesis), and immunologic disease. HTLV-II infection is associated with an increase in common infections, suggesting immune suppression.

1:2,993,000.4 This estimate may be artificially low as a result of changes in sensitivity of the confirmatory test interpretation. However, this bias is probably offset by near universal implementation of leukocyte reduction, which greatly reduces risk of HTLV transmission.

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Member of Retroviridae family. Enveloped, single-stranded RNA viruses. There is >50% sequence homology between HIV-1 and HIV-2 and common serologic cross-reactivity.

HIV-1 is divided into main and outlier groups with at least nine subtypes (A1) in the main group.

Cell-Associated Transfusion-Transmitted Viruses Member of Retroviridae family. HTLV-I and HTLV-II have 65% nucleotide sequence identity and significant serologic cross-reactivity.

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HTLV-I is endemic in many populations throughout the world,

Infectious Disease Screening

Human T-cell lymphotropic virus (HTLV-I,-II)

(Continued)

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244 䊴

TABLE 8-1. Infectious Pathogens and Risk of Transmission by Transfusion (Continued) Classification and Description

Human T-cell lymphotropic virus (HTLV-I,-II) (Continued)

CMV [aka human herpesvirus 5 (HHV-5)]

Epidemiology

Disease Manifestation

Estimated US Transmission Risk per Unit Transfused

with relative sparing of Europe.5 HTLV-II is most common in indigenous populations in the Americas and among injection drug users in the United States.

Large, double-stranded, enveloped DNA virus; member of the human Herpesviridae family.

Found throughout all geographic locations and socioeconomic groups.

Infects between 50% and 85% of the United States population by 40 years of age.

CMV is the virus most frequently transmitted to a developing child before birth, causing birth defects and congenital infections. For most healthy people who acquire CMV after birth, there are few symptoms. Immunocompromised people may experience severe disease with multiple organ involvement.

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Unknown. Presumed to be rare, given widespread leukocyte reduction of cellular blood products and absence of transmission from plasma products.

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Pathogen

Epstein-Barr virus (EBV) [aka human herpesvirus 4 (HHV4)]

Double-stranded, enveloped DNA virus; member of the human Herpesviridae family.

Ubiquitous virus with worldwide distribution.

Prevalence is near 100% in developing countries; slightly lower in western industrialized nations.

Unknown (rare) Most infections are asymptomatic, followed by latent infection.

Causative agent of infectious mononucleosis. EBV infection is also associated with several malignancies, most notably Burkitt’s lymphoma and nasopharyngeal carcinoma (in Asia).

Human herpesvirus 8 (HHV8) [aka Kaposi’s sarcomaassociated herpesvirus (KSHV)]

Enveloped, double-stranded gamma herpesvirus; member of human Herpesviridae family.

Uncommon in the general United States and northern European populations, but frequently (>40%) found in African countries and among homosexual men.

Causes asymptomatic presumably lifelong latent infections in immunocompetent persons. Associated with Kaposi’s sarcoma, primary effusion lymphomas, and Castleman’s disease in immunocompromised persons, particularly those with HIV infection.

Unknown (rare)

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Nonenveloped, positive, single-stranded RNA virus of the Picornaviridae family, genus Hepatovirus.

Enterovirus transmitted from person to person by the fecal-oral route, resulting in transient viremia. Approximately 30% of US population has evidence of past infection.

Gastrointestinal symptoms; hepatitis with transient viremia without carrier status. May be asymptomatic in one-quarter of infections.

Adults are more likely to have symptomatic disease than children.

Unknown, but presumably <1:1 million (occasional transfusion- transmitted cases have been described from asymptomatic viremic donors).

Hepatitis E virus (HEV)

Nonenveloped, positive, single-stranded RNA virus of the Hepeviridae family.

Similar to HAV above, more common in developing, tropical countries (seroprevalence in US population <2%).

Similar to HAV disease but may be more severe with overall 2% fatality rate (higher in pregnant women).

Rare case reports of transfusion transmissions outside the US have been published.6-8

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Hepatitis A virus (HAV)

Infectious Disease Screening

Transfusion-Transmitted Viruses with Low Transmission Rates or Lack of Disease Associations

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TABLE 8-1. Infectious Pathogens and Risk of Transmission by Transfusion (Continued)

Pathogen

Classification and Description

Epidemiology

Disease Manifestation

Estimated US Transmission Risk per Unit Transfused ä&#x160;´

Enveloped, positive, singlestranded RNA virus in the Flaviviridae family with 3 species (A, B, C).

GBV-A and GBV-B are found in new world primates. GBV-C appears to be the only human variety with prevalence of about 2% in the US blood donor population (higher among intravenous drug users and multitransfused patients).

No disease manifestations have been identified.

Transfusion transmission is well documented but no disease risk has been established.

Parvovirus B19

Small, single-stranded, nonenveloped DNA virus. Member of the Parvoviridae family, genus Erythrovirus.

Common human virus with evidence of past infection in approximately 50% of the adult US population.

Causative agent of fifth disease, a common childhood illness. Other disease manifestations are arthritis in adults, red cell aplastic crisis, and hydrops fetalis caused by fetal anemia.

Viremia rate is 1:20,000 to 1:50,000 with substantially higher rates during epidemic years.9 Documented transmissions by transfusion are limited, as most infections are asymptomatic and the seroprevalence rate in recipients is high.

SEN virus

Small, nonenveloped negative, single-stranded DNA virus in the Circoviridae family.

No disease manifestations have Present in approximately been identified.

3% of US blood donors with 10-fold higher prevalence in transfusion recipients.

Transfusion transmission is well documented, but no disease risk has been established.

Torque Teno virus (TTV)

Similar to SEN virus.

No disease manifestations have Depending on the methbeen identified. odology used, TTV is found frequently (>5% to >50%) among blood donors in the US and elsewhere.

Transfusion transmission is well documented, but no disease risk has been established.

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GB viruses

Emerging Transfusion-Transmitted Viruses and Viruses with Potential for Transfusion Transmission Usually self-limiting febrile illness with joint and muscle pain and a discrete macular or maculopapular rash. Life-threatening complications are dengue hemorrhagic fever and dengue shock syndrome.

Important risk factors for these complications include the strain of the infecting virus, age and prior dengue infection history of the patient.

So far, two cases of transfusiontransmitted Dengue virus have been reported from Hong Kong and Singapore, but dengue viremia has been documented in approximately 1:1000 blood donors in Puerto Rico, which underscores the potential risk of transfusion transmission in the United States.10 Transmissions of dengue virus by marrow transplantation and needle-stick injuries have occurred.

Lymphocytic choriomeningitis virus (LCMV)

Enveloped, negative, singlestranded RNA virus in the Arenoviridae family.

Natural reservoir of LCMV is rodents. Humans can become infected through contact with infectious rodent body fluids. Seroprevalence in the US population is approximately 5%.

LCMV infection may cause meningoencephalitis.

To date, no transmissions of LCMV by transfusion have been documented. However, fatal LCMV infection has been associated with organ transplantions.11

Severe acute respiratory syndrome (SARS)

Enveloped, positive, singlestranded RNA virus in the Coronaviridae family.

Coronaviruses infect a wide range of mammals and birds and occur worldwide. Disease transmission is by the fecaloral and respiratory routes.

Most coronavirus infections cause mild illness, but severe respiratory tract infections have occurred, exemplified by the 2002-2003 SARS outbreak.

To date, no human transmission of SARS coronovirus by transfusion has been documented.

ä&#x160;ł

Dengue virus is the most common arthropodborne viral disease with an estimated 50 million cases worldwide each year. Primary vector is the Aedes aegypti dengue mosquito.

Infectious Disease Screening

Enveloped, positive, singlestranded RNA virus in the Flaviviridae family, with four distinct serotypes (Den-1 to -4).

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Dengue virus

(Continued)

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248

Pathogen

Classification and Description

Epidemiology

The virus is primarily Enveloped, single-stranded RNA virus of the Retroviridae found in African nonhufamily (closely related to HIV). man primates and other mammals who seem to be carriers (ie, show no pathogenic effects). There is also evidence of human infection in a small percentage of animal handlers and veterinarians.

West Nile virus (WNV)

Enveloped, positive, singlestranded RNA virus in the Flaviviridae family (related to dengue virus).

Predominantly found in birds and other animals.

Transmission to humans occurs through bites from infected mosquitoes.

No disease manifestations have been identified.

Estimated US Transmission Risk per Unit Transfused To date, no human transmissions of SFV by transfusion have been documented.

In most cases (80%), WNV infec- Rare breakthrough transmissions tion is aymptomatic. In the remain- may occur despite WNV screening der, a fever and rash illness occur. by NAT.12 Encephalitis and meningitis have been observed in 0.7% of infected persons, with severe illness and fatal outcomes in some and with significant, long-term, neurologic sequelae in many survivors.

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Simian foamy or spumavirus (SFV)

Disease Manifestation

ä&#x160;´

TABLE 8-1. Infectious Pathogens and Risk of Transmission by Transfusion (Continued)

Bacteria Coagulase-negative Staphylo- Gram-positive organisms. coccus spp, Staphylococcus aureus, Streptococcus spp, Bacillus cereus

Treponema pallidum

Helically coiled, gramnegative bacterium (order spirochaetales).

Present in the environment or in the bloodstream of otherwise asymptomatic blood donors.

As above. Listed gram-negatives were associated with 9 of 13 (69%) platelet transfusion-related septic fatalities and 7 of 8 (88%) RBC transfusions associated with fatal septic reactions.14

Platelets: Septic reaction in at least 1:75,000 and fatality in at least 1:500,000 transfused platelet components.13

Currently, relatively rare organism in the US population (annual rate of new infections approximately 0.03%); transmitted from person to person by sexual contact.

Causative agent of syphilis.

Remote (no new transfusion-transmitted cases reported in >30 years).

RBCs: Septic reaction in at least 1:500,000 and fatality in 1:10,000,000 transfused RBC components.15

RBCs: Septic reaction in at least 1:500,000 and fatality in 1:10,000,000 transfused RBC components.15

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Infectious Disease Screening

Gram-negative organisms.

Platelets: Septic reaction in at least 1:75,000 and fatality in at least 1:500,000 transfused platelet components.13

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Serratia liquifaciens, Yersinia enterocolitica, Acinetobacter spp, Enterobacter spp, Escherichia coli, Pseudomonas spp, Providencia rettgeri

May cause septic, potentially fatal reactions in transfusion recipients.

Listed gram-positives accounted for 4 of 13 (31%) fatalities associated with bacterial contamination of platelets, compiled from studies in the United States, United Kingdom, and France.14 Coagulase-negative Staphylococci accounted for 1 of 8 (13%) fatalities associated with contaminated RBC transfusions in the same studies.14

Frequently found on human skin or in the natural environment.

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TABLE 8-1. Infectious Pathogens and Risk of Transmission by Transfusion (Continued) Classification and Description

Epidemiology

Disease Manifestation

Estimated US Transmission Risk per Unit Transfused

Babesia spp.

Protozoan parasite of the Babesiaceae family (order Piroplasmidales).

Found worldwide in erythrocytes of wild and domestic animals, transmitted to humans through an Ixodes (hard tick) vector.

Babesia infection may cause mild, self-limited disease to severe illness (usually in immunocompromised or splenectomized patients).

Symptoms are flu-like, but characteristically include hemolysis with evidence of intraerythrocytic parasites.

Unknown, increased risk in endemic areas such as Connecticut in the United States where the projected risk per RBC unit transfused has been estimated as 1:1800.16

Borrelia burgdorferi

Spirochete bacterium.

Primary reservoirs are rodents and deer. Transmission to humans occurs by means of an Ixodes (hard tick) vector.

Lyme disease has many signs and symptoms, including a rash, arthritic pain, and neurologic symptoms. Similar to syphilis, symptoms frequently seem to resolve, yet the disease progresses.

Unknown, no cases of transfusion transmission have been reported.

Ehrlichia chaffeensis and Anaplasma phagocytophila

Gram-negative obligate intracellular parasites of the Ehrlichiaceae family (order Rickettsiales).

Natural reservoirs are wild animals (deer, rodents).

Transmission to humans by means of tick vectors.

Febrile illness with flu-like symptoms. E. chaffeensis is the causative agent of human monocytic ehrlichiosis; A. phagocytophila causes human granulocytic ehrlichiosis.

Unknown, rare (a single case of transfusion-transmitted human ehrlichiosis has been reported).17

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Vector-Borne Bacteria and Parasites

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Pathogen

Transfusion-transmitted leishmaniasis has been described in endemic areas, but so far, no cases have been described in the United States18

Flagellated protozoan parasites of the Trypanosomaceae family.

Cutaneous, muco-cutaneous, and The organisms are visceral infections, known as kalaendemic in many vertebrates in tropical and sub- azar or black fever. tropical countries.

Transmission to humans occurs by means of sandflies.

Plasmodium spp.

(P. falciparum, P. malariae, P. ovale, P. vivax)

Protozoan parasites of the Plasmodiidae family (order Haemosporida).

The life cycle of the human parasite involves sequential infection of liver and red cells.

Human-to-human spread occurs by means of infected Anopheles mosquitoes.

Causative organism of malaria.

1:4 million19

Rickettsia

Gram-negative obligate intracellular bacterium of the Rickettsiaceae family (order Rickettsiales).

Natural reservoirs are wild animals (deer, rodents).

Transmission to humans by means of Ixodes (hard tick) vector.

Causative agent of Rocky Mountain spotted fever, a severe flu-like illness with characteristic petechial rash.

Unknown, rare (a single case of transfusion-transmitted Rocky Mountain spotted fever was described).17

Trypanosoma cruzi

Flagellate protozoan parasite of the order Trypanosomatida).

Prevalent in Central and South America; transmitted by the feces of reduviid bugs.

Causative agent of Chagas’ disease.

Unknown. At least 9 cases of transfusion-transmitted Chagas’ disease have been reported in North America.

Prions represent a new form of infectious agent that has not been classified among the traditional infectious pathogens.

Inherited and sporadic forms have been described.

Causative agent of transmissible spongiform encephalopathy, a progressive form of encephalitis in animals (scrapie) and humans (Creutzfeldt-Jakob disease and variants).

Remote. However, as of May 2007, four likely cases of transmission of variant Creutzfeldt-Jakob disease by transfusion have been described in the United Kingdom.20

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Leishmania spp.

Infectious Disease Screening

Prions 䊳

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worldwide, 2) relatively long viremic periods during which affected individuals remain asymptomatic, 3) efficient transmission through contaminated blood components and products, and 4) high risk of serious disease manifestation in recipients. Because these viruses may be circulating in the bloodstream for weeks to months, to even years, without causing overt symptoms, infected persons may unknowingly present as blood donors, and their donations may be accepted. In these cases, the only barriers that currently prevent contamination of the blood supply are effective blood donor screening tests. Fortunately, sensitive tests for selection of donors are available, and their use is mandated whenever blood is collected for the purpose of transfusion or manufacture of infusible products.21 As a result, residual transmission risks of blood donations screened with state-of-the-art tests are minimal (Table 8-1). Studies that predated availability of screening tests demonstrated that these viruses are transmitted efficiently through whole blood, all blood components, and many plasma derivatives.22,23 Furthermore, there is evidence that even components with very low viral loads, which are undetectable by currently available screening tests, can still transmit disease.24,25 Therefore, presence of these viruses at any level in an infusible product must be considered potentially infectious. Although routine blood banking procedures (freezing of plasma, blood storage for more than 26 days,26 and simple component manipulations such as washing or leukocyte reduction of cellular blood components27) may reduce transmissibility of HIV, they do not prevent transmission. The best prospect for eventual elimination of the threat from classical transfusion-transmitted viruses is through a combination of careful donor selection, screening of each donation with sensitive tests, and application of pathogen reduction and inactivation methods, where available, safe, and feasible.28 Brief individual

descriptions of the viruses in this group are given in Table 8-1.

Cell-Associated Transfusion-Transmitted Viruses The most prominent members of this heterogeneous group of viruses are human T-cell lymphotropic virus, types I and II (HTLV-I, -II) belonging to the delta-retrovirus genus, and cytomegalovirus (CMV), a member of the beta herpes virus subfamily. The natural mode of transmission is from person to person through contact with infected blood and body fluids. Both virus groups are found worldwide, but HTLV has more selective distribution and is usually present in <10% of affected populations, even in endemic areas.

CMV, by contrast, is a common pathogen with homogenous global distribution. The seroprevalence rate of CMV averages 50% to 80% in the adult US population with generally higher prevalence rates among lower socioeconomic groups.

Disease manifestations of CMV and HTLV are summarized in Table 8-1. Most relevant from a transfusion medicine standpoint is the propensity of CMV to cause severe, even life-threatening, disease in immunocompromised individuals and in very low birthweight infants and the ability of both viruses to cause lifelong latent infections in immunocompetent persons. Because latent infections with CMV or HTLV are usually clinically silent, except for a small percentage of HTLVinfected individuals who develop rare T-cell malignancies or debilitating neurologic conditions,5 blood donors may be latently infected and thus be the source of transmissions. Replication-competent virus is found in infected leukocyte populations and their progenitors. Presence of cell-free infectious HTLV has not been reliably demonstrated, but plasma viremia may occur in CMV disease during primary infection,29 reactivation of latent virus, and in rare cases of second-strain infections. Transfusion-associated

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CHAPTER 8

infectivity, however, is considered limited to whole blood and cellular blood components, with no confirmed reports of HTLV or CMV transmission through “acellular” blood components (frozen plasma, cryoprecipitate) or plasma derivatives.

The risk of transmitting HTLV by means of infected blood has been projected at 10% to 30%. For CMV, the risk of transfusionassociated transmission depends strongly on the status of the recipient’s immune system.

In immunocompromised CMV seronegative recipients, transmission rates of up to 50% have been reported from whole blood and non-leukocyte-reduced cellular blood products with associated significant disease manifestations, particularly in premature infants as well as progenitor cell and solid organ transplant patients.30 In contrast, the risk in immunocompetent recipients appears to be negligible with respect to both transmission rates and associated symptoms.

Prevention of transfusion transmission of HTLV rests on serologic testing of prospective blood donors, with leukocyte reduction as an additional safeguard. General blood donor screening for evidence of latent CMV infection and rejection of seropositive donors is not feasible, given the high prevalence of past CMV exposure in the adult population. In lieu of deferral of CMV seropositive blood donors, transmission to susceptible populations is limited through selection of “CMVreduced-risk” cellular blood components for at-risk recipients. CMV-reduced-risk blood components are 1) whole blood or components from seronegative donors, 2) leukocytereduced cellular blood components containing <5 × 106 residual leukocytes, or 3) both. Both methods are effective in limiting disease transmission (with some controversy over equivalency of the approaches), but neither offers complete prevention of CMV transmission.31 The AABB recommends that each institution review its internal policies for blood use in patients vulnerable to CMV

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infection.32 If leukocyte reduction is chosen as a means to reduce the risk of CMV transmission, then components prepared by prestorage leukocyte reduction should be selected because leukocyte removal by this method is more reliable than bedside filtration. Although it has not been proven that a combination of both methods provides greater safety for patients at highest risk for CMV transmission and disease, this approach has intuitive appeal and was the preferred recommendation for highest risk patients by most panelists participating in the Canadian consensus conference on prevention of transfusion-associated CMV transmission.30 Epstein-Barr virus (EBV), a gamma herpes virus, rarely causes transfusion-transmitted infections. Most adults (>90%) have evidence of previous exposure, and only isolated cases of transfusion-transmitted EBV presenting as infectious mononucleosis syndrome have been described in both immunocompetent recipients and in immunosuppressed patients following solid organ transplantation.33 Aggressive EBV-associated lymphoproliferative disorders have been observed in patients with weakened immune systems following cordblood stem-cell transplantation but have not been documented following transfusion. Because B lymphocytes are the likely source of transfusion-transmitted EBV infection, leukocyte reduction of cellular blood components, which efficiently depletes B cells, is an attractive strategy to prevent infection in transfusion recipients. This expectation has not yet been proven in clinical trials, though.

Human herpes virus type 8 (HHV-8), another gamma herpes virus, has tropism to lymphocytes and monocyte-macrophages. Viral infection can cause uncontrolled cell proliferation and has been associated with a variety of malignancies, including Kaposi’s sarcoma and primary effusion lymphoma. HHV-8 is transmitted primarily through sexual contact, but transmission through kidney transplantation has been documented.34 The possibility of

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transmission by transfusion in the United States has been raised35; definitive evidence is lacking, however. The seroprevalence of HHV-8 among US blood donors was recently estimated at 3% to 3.5%, and no donor viremia was found.36 In contrast, there is evidence of HHV-8 transmission by transfusion in subSaharan Africa where seroprevalence of HHV8 among blood donors is 40% and where transfusion of fresh, non-leukocyte-reduced blood is common practice.37 It can be speculated, but has not been proven, that leukocyte reduction prevents transfusion transmission of HHV-8.

Transfusion-Transmitted Viruses with Low Transmission Rates or Lack of Disease Associations Viruses discussed in this category include parvovirus, enterically transmitted hepatitis viruses A and E (HAV, HEV), and several viruses originally discovered during the search for the elusive infectious agent of non-A-E hepatitis, now identified as GB, TT, and SEN viruses (GBV-C, TTV and SEN-V), respectively.

Parvovirus B19, a non-enveloped DNA virus, has been known to be transmitted by transfusion for decades. It has recently attracted more attention because of the realization that viremia is relatively common in blood donors and inactivation of the virus is not easy or reliable, which raised concern for pooled blood components and plasma derivatives. Parvovirus as detected by its DNA can be expected to be present in 1:50,000 to 1:20,000 blood and plasma donors with substantially (100-fold) higher incidence during epidemic periods.9 However, neutralizing antibodies are frequently present in the donor, recipient, or both. Only rare transmissions of parvovirus caused by single-donor components have been reported.

Before routine parvovirus screening of plasma donations intended for manufacture of clotting factor concentrates and other deriva-

tives, the virus was nearly always present in the large plasma pools that are the source material for these products. Resistance to common inactivation methods allowed infectious virus to persist in the final products.

Although the presence of neutralizing antibodies in the plasma pool provides some protection, this presence does not guarantee that recipients will not be infected. Earlier prospective studies in previously untreated patients with hemophilia, who received virus-attenuated factor concentrates, demonstrated a persistent 40% risk of parvovirus infection. Fortunately, these patients in general did not suffer serious or long-term hematologic sequelae regardless of HIV antibody status. More recently, source plasma screening by nucleic acid amplification testing (NAT) methods has been implemented to exclude high-level viremic donations from use in manufacturing and, together with implementation of enhanced virus removal methods, has dramatically reduced the risk of parvovirus infection from manufactured blood products.38 HAV and HEV are predominantly transmitted through the fecal-oral route, and parenteral transmissions are unusual but well documented. The incubation period of HAV averages 28 days with a peak viremic period occurring 2 weeks before onset of jaundice or elevation of liver enzymes. Correspondingly, transfusion-transmitted HAV infection can usually be traced to a recently infected person.39 The virus is relatively resistant to heating and other pathogen reduction procedures.

Transmission by clotting factor concentrates treated with the solvent/detergent pathogen process has been reported.40 Rare cases of transfusion-transmitted HEV infection have only recently been published.6-8 GBV-C, TTV, and SEN-V have seroprevalence rates of up to 10% or higher in the general population worldwide and are transmitted by blood transfusions. No definitive disease associations have been identified, however, and consequently no screening is warranted.41

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CHAPTER 8

Emerging Transfusion-Transmitted Viruses Viruses discussed in this group have recently come to attention either because of increased disease activity with documented new or rising transmissions by means of blood and blood products or because they were associated with notable outbreaks that raised concern that the responsible virus might spread to the blood donor base.

West Nile virus (WNV) infection has quickly become a major focus of transfusion safety concerns because large outbreaks occurred recently in the United States and Canada. The primary reservoir for WNV is birds. Humans and other mammals, particularly horses, are incidental hosts, with transmission occurring through bites of infected mosquitoes. Peak transmission coincides with the mosquito season in the late summer and early fall. The chief opportunity for transmission by transfusion appears to be the brief asymptomatic viremic period of 2 weeks or less that occurs in most newly infected persons. There were no cases of WNV infection in North America (or the entire Western Hemisphere) before a WNV outbreak in New York City in summer 1999. Since then, the virus has reappeared annually in the United States and Canada during the summer months. WNV has caused large epidemics that were initially centered in the eastern part of both countries but rapidly spread westward. Twenty-three transfusion-transmitted WNV infections were first documented in 2002, which prompted implementation of blood donor screening with NAT for WNV genetic sequences in summer 2003. Blood screening has been very effective, with only a few cases of transfusion-transmitted WNV subsequently reported in the United States42 and none reported in Canada. Despite establishment of sensitive screening strategies using NAT for WNV on pools of blood donor samples, with triggering of individual donation testing when local disease activity exceeds

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preset thresholds, a case of transfusion-transmitted WNV neurologic disease in an immunosuppressed patient in 2006 showed that rare breakthrough transmissions may still occur in the United States.12 A growing list of other viruses is recognized as a potential or emerging threat to the blood supply, although definitive proof of transmissibility by transfusion is lacking for at least some of the viruses discussed here. These viruses include the causative agents of 1) severe acute respiratory syndrome (SARS), 2) dengue fever, 3) lymphocytic choriomeningitis (LCMV), and 4) simian foamy (spuma) virus (SFV) infection, to name some.

With the exception of dengue virus, which is transmitted by means of a mosquito vector from person to person, these viruses have in common that they are usually found in animals, which has raised concern for an apparent increase in species-to-species transmission of pathogenic viruses. This phenomenon termed “zoonotic infection” can be expected to continue and perhaps even increase in the future, as a result of 1) shrinking natural habitats worldwide; 2) growing human activities around wildlife; and 3) rising international travel, trade, and migration.

Viremic periods,11,43-45 or presence of viral sequences in cases of human SFV infection,46 were documented transiently following acute infection for each of the four viruses mentioned above and are likely to occur in other zoonotic infections. An additional unknown but potential threat is that zoonotic viruses acquire new mutations as they adapt to the human host, which could change their mode of transmission and could alter other properties that could increase transmissibility and pathogenicity in general.47 Another cause for concern is well-known human viruses that cause common communicable diseases such as seasonal enteroviruses48 and influenza.49 Although these viruses may cause mild self-limited infection in healthy adults, they can cause severe symp-

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toms in vulnerable populations such as infants, immunosuppressed patients, and the elderly. This danger has led to concerns with respect to blood safety and studies to evaluate the frequency of asymptomatic viremia in blood donors. For example, seasonal fluctuations involving enterovirus species have been reported in approximately 1 in 4000 Scottish blood donors.48 The significance of this finding for disease transmission and the need for routine donor screening or other preventive measures to protect the blood supply from these ubiquitous viruses have yet to be established.

Bacteria Bacterial contamination is an obvious concern for blood components that are directly infused into the bloodstream. Serious, even fatal septic reactions may result, and overall morbidity and mortality associated with transfusion-associated infection likely surpass those of viral transmissions.50 The usual sources of bacterial contamination are donor skin and blood and, less likely, contaminated disposables and the environment. Platelets are by far the most frequent component causing septic transfusion reactions because of room temperature storage, which promotes bacterial growth. Before large-scale prospective screening with culture methods in the United States, the incidence of bacterial contamination in allogeneic platelets based on culture positivity was estimated at 1 in 1000 to 3000 units.50 Experience since implementation of screening for bacterial contamination in 2004 suggests a lower rate of approximately 1:5000 units.51,52 Gram-positive skin saprophytes account for most of the significant organisms contaminating platelet concentrates, with the remaining attributed to gram-negative organisms associated with occult bacteremia. Fatal reactions can be caused by endotoxin associated with gram-negative bacteria. In a series

of 61 septic reactions associated with platelet transfusions compiled from studies in the United States, United Kingdom, and France, the gram-negative bacteria Escherichia coli, Providencia rettgeri, Klebsiella species, and Serratia species accounted for 9 of 13 (69%) fatal reactions.14 Moreover, although the bacterial content in a contaminated component at the time of collection may be exceedingly small, bacteria proliferate during component storage (unlike viruses), resulting in a significant increase in concentration and, in some species, the release of endotoxin and other toxic metabolites. The minimal bacterial doses in platelet concentrates leading to morbidity or mortality is not known precisely and is probably variable. However, experience at a single institution over a 13-year period showed that 13 transfused platelet units associated with septic transfusion reactions had a mean bacterial count of 2.2 × 106 colony-forming units (CFUs), compared to 19 contaminated units not associated with reactions, which had mean counts of 1.6 × 104 CFUs.53 Detection methods for bacterial contamination of platelets, broadly implemented in the United States from 2003 to 2004, significantly reduced the risk of this complication by intercepting contaminated units before they were transfused to patients, but those methods have not eliminated this concern. In the 2004 to 2006 period, the American Red Cross (ARC) documented a residual risk of clinically relevant septic reactions of at least 1 in 74,807 and a fatality rate of at least 1 in 498,711 platelet components that were distributed after undergoing routine bacteria detection by a culture method.13 This level of risk represents an approximately 50% reduction within the ARC system compared to a 10-month period in 2003 before implementation of bacterial screening. Because these figures are based on reported clinical reactions, they represent a low estimate of actually occurring septic reactions that may be overlooked by clinical providers or attributed to

Copyright © 2008 by the AABB. All rights reserved.

CHAPTER 8

other causes.54 Despite only partial success of current bacteria detection methods and strategies in preventing septic reactions, the Food and Drug Administration (FDA) considers individual applications from collection facilities for extension of platelet storage from 5 to 7 days. The agency has granted facilities with a track record of successful implementation of bacteria detection systems permission to extend platelet storage by 2 days under specific conditions, including aerobic and anaerobic culture and postmarket surveillance.55 Refrigerated storage limits growth and viability of most bacteria in Red Blood Cell (RBC) products. In a prospective study, bacterial contamination of RBCs was found at a rate of 1 in 31,385 units transfused.56 Septic reactions and fatalities associated with RBC components that are based on clinical recognition and reporting were documented at rates of 1 in 500,000 and 1 in 10 million in the United States, respectively.15 Because of likely underrecognition and underreporting, others have estimated the risk of septic reaction caused by RBC components at 1:250,000.57 Notably, septic reactions have occurred following autologous RBC transfusions, a finding that may be explained by 1) generally poorer health of autologous donors; 2) higher rates of unrecognized bacteremia; 3) application of less stringent donor selection criteria in autologous donation; and, 4) on average, longer storage of the RBCs.58 Microorganisms isolated from contaminated RBC units include Staphylococcus epidermidis, Serratia liquifaciens, Pseudomonas species, and Yersinia enterocolitica. The latter, a gram-negative endotoxin-producing organism, is remarkable for its preference to grow at colder temperatures in iron-enriched environments.59

Vector-Borne Bacteria and Parasites These pathogens have in common that they are transmitted to humans by a variety of

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arthropod vectors, most commonly mosquitoes and ticks. Prevalence rates of infectious organisms among blood donors depend on local vector activity.

Ticks are an increasingly recognized important vector of bacterial zoonoses; examples of infections transmitted to humans include Lyme disease, ehrlichioses, and rickettsioses. Reports of tick-borne diseases, which also include parasitic infections (among them babesiosis), rose sharply in the United States in recent years. This increase may, in part, be a reflection of newly instituted reporting requirements of tick-borne infections to state health agencies, but it seems to also represent a true increase caused by factors such as the widening geographic range of tick vectors in the United States, the spread of residential areas to tick habitats, and the increase in popularity of outdoor sports and leisure activities that bring humans in contact with the vector.

Transfusion transmission of bacterial tickborne diseases appears to be rare, however.17 No cases of transfusion-transmitted Lyme disease have been confirmed to date, and only single instances of transfusion-transmitted ehrlichioses and rickettsioses were reported in the 1970s and 1990s, respectively. In both cases, the transmitting blood units had not been leukocyte-reduced. Because the causative agents are obligatory intracellular bacteria, recent widespread adoption of leukocyte reduction of cellular blood components can be expected to have lowered the risk of transfusion transmission of these organisms.

With the exception of babesiosis, which is found in temperate climates, parasitic infections that may be transmitted by transfusion are primarily a concern for tropical and subtropical countries where a significant proportion of prospective blood donors are afflicted. However, increasing international travel, tourism, and human migration from countries where transfusion-transmissible parasitic infections are endemic have raised con-

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cern about transfusion-transmitted parasitic disease in areas that are not naturally affected.

Malaria, a mosquito-borne illness, occurs in the United States in travelers, military personnel, and immigrants from countries with malaria-endemic areas. Occasional cases result from mosquito transmission, blood transfusion, or organ transplantation. Approximately three transfusion-associated malaria cases occur per year in the United States with a reported incidence of 0 to 0.2 cases per million units transfused during the 5-year period from 1993 to 1998.19 Malarial parasites, Plasmodium falciparum, P. vivax, P. ovale, and P. malariae, maintain viability in RBCs stored at 4 C, in platelet concentrates stored at room temperature, and after RBC cryopreservation and thawing. Malaria is not transmitted by RBC-free blood products such as Fresh Frozen Plasma and plasma derivatives, with the possible exception of cryoprecipitate.60 In malaria-endemic countries, where the vast majority of donors and recipients have been previously infected, blood donors may be preselected according to degree of malaria risk, screened for presence of parasites with Giemsa-stained blood smears, or assessed with rapid diagnostic tests. In addition, transfusion recipients may be routinely administered inexpensive prophylactic medication before transfusion.60 Prevention of transfusion-transmitted malaria in countries where the disease is not endemic currently relies on deferral of blood donors emigrating or returning from malaria-endemic regions, an imperfect strategy that misses some infected donors and excludes many people from donations that are not infected. Manual laboratory tests that are currently used in malariaendemic areas are generally not suited for areas where malaria is not endemic because they are impractical, insensitive, or too nonspecific for large-scale blood donor screening in low-risk settings. Commercially available antibody tests are already in use in some European countries and Australia for acceler-

ated reentry of blood donors following deferral for travel to areas where malaria is endemic, an approach that is also being considered in the United States.

Babesiosis is a malaria-like zoonosis in which humans are infected incidentally, usually through the bite of an infected tick of the genus Ixodes. Babesia infections are usually asymptomatic or associated with mild flu-like symptoms. However, immunocompromised or splenectomized individuals are at risk for lifethreatening disease. At least 50 cases of transfusion-transmitted babesiosis associated with either RBC or platelet transfusions have been reported in the United States, most of them during the past 20 years.16,61 Persons with a history of babesiosis are deferred from donating blood, but no blood donor screening test or other effective means exist currently to detect asymptomatic carriers of the parasite.

The etiologic agent of American trypanosomiasis, or Chagasâ&#x20AC;&#x2122; disease, is the flagellate protozoan parasite, Trypanosoma cruzi.

The disorder is limited to the Western Hemisphere where it is widespread in continental Latin America from Mexico to the lower half of the South American continent. The parasite is transmitted to humans through bites from T. cruzi-infected insects of the Reduviidae family. Infected persons maintain a lowlevel, intermittent parasitemia that usually persists for life; treatment is most effective early in infection. The lifetime risks of severe heart or intestinal problems in infected individuals averages about 30% (range of 10% to 40%, depending on a variety of factors) and usually occur many years after the initial infection. The prevalence of T. cruzi infection (which is based on detection of antibodies to T. cruzi antigens) among US blood donors varies and depends on the proportion of residents with Latin American backgrounds because virtually all seropositive donors are immigrants from Central and South America.

Nationwide estimates before implementation of broad blood donor screening suggested a

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CHAPTER 8

rate of 1 in 25,000 donors with three or four times higher rates in communities with a large Latin American immigrant population.61 Although only nine cases of transfusion-associated Chagas’ disease have been documented in immunocompromised patients in the United States and Canada, overall transmission rates are most certainly higher, because infections in immunocompetent recipients were likely missed as a result of the nonspecific nature and delayed onset of symptoms and the general unfamiliarity of physicians with T. cruzi infection and Chagas’ disease.62 Transmissions are most likely with whole blood and platelet transfusions. The combined yield of two published look-back studies in the United States and Mexico suggests a transmission rate (which is based on recipient seropositivity) of nearly 40% (5 of 13 recipients). According to these findings and a continued demographic trend of immigration from Central and South America, transfusion transmission of T. cruzi is seen as a rising concern in the United States.

Consequently, screening for infected blood donations was broadly implemented in January 2007, shortly after licensing of the first screening test in the United States. The prevalence of confirmed positive blood donors after 3 months of routine screening was approximately 1:21,000, thus confirming widespread Chagas’ disease infection in the United States blood donor population.63 By mid-January 2008, 1,112 repeat antibody-positive blood donors were identified, of which 336 were confirmed by radioimmune precipitation assay (RIPA), 744 were nonreactive by RIPA, and the remaining test results are pending.64 Rare reports of human transfusion-transmitted trypanosomiasis (African sleeping sickness), leishmaniasis, toxoplasmosis, and microfilariasis have been reported. With the exception of toxoplasmosis, which is essentially ubiquitous, all have occurred in areas where these organisms are endemic.18,65

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Prions Prions, short for proteinaceous infectious particles, are abnormally structured forms of cellular proteins that are able to convert normal protein molecules on contact into an abnormal structure. This unique property allows prions to act like an infectious agent that transmits disease by transforming normal cellular proteins into insoluble precipitates.66 Prion diseases affect primarily brain tissue, causing severe, progressive dementia with generally fatal outcome in affected persons and equivalent disease in animals. The best-known prion disease in humans is socalled classical Creutzfeldt-Jakob disease (CJD).67 Evidence to date suggests that this disease is not transmitted by transfusion.68 A variant form of the disease (vCJD), however, is likely transmissible by transfusion, based on animal model studies and four cases in the United Kingdom,20where recipients of blood from donors later diagnosed with vCJD acquired the disease or evidence of infection.68 Given our limited experience, the full potential of vCJD transmission by transfusion is not presently known.

Like classical CJD, vCJD is a fatal, degenerative neurologic disease. It occurs in younger persons and has distinctive clinical, histopathologic, and biochemical features, including the presence of readily detectable prion protein in non-central-nervous-system lymphoreticular tissues such as the appendix, spleen, tonsils, and lymph nodes.69 In contrast to classical CJD, vCJD disease is new, with the first reports from the United Kingdom occurring in 1996. The etiologic agent of vCJD (presumably a prion) is the same agent that causes bovine spongiform encephalopathy (BSE). Transmission of BSE prions to humans presumably occurred by consumption of beef and perhaps direct exposure to cattle products of the rendering industry, including tallow, gelatin, and bone meal.67 The outbreak of vCJD in the United Kingdom, which resulted

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in 166 documented cases as of January 2008, followed a massive epidemic of BSE that peaked in the 1990s and has now subsided. It appears that the number of new cases of vCJD in the United Kingdom has decreased accordingly in recent years.69 A significant number of vCJD cases (38 as of December 2007) have been diagnosed outside the United Kingdom, mostly in France and several other European countries.

Even before cases of likely transfusiontransmitted vCJD became known, precautionary measures were implemented to reduce the risk of prions entering the blood supply.

Prevention focuses on the deferral of donors with connection to the United Kingdom and parts of Europe. Because no practical or reliable tests for vCJD are yet available, prions are not inactivated by blood storage, and they resist most pathogen inactivation procedures, active efforts are under way to develop a screening test to detect prions and filters to remove them from donated blood.70 ST R A TE G I E S TO PRE VEN T IN FE C T I O U S D I SE A SE TR A N S M I SS I ON BY BL O O D TR A N S F U SI O N Donor Selection Selection of eligible blood donors is a critically important step to protect not only the donor’s health but also the safety and integrity of the blood supply. From the viewpoint of the latter objective, selection of healthy donors by interviews and limited physical examinations represent the first of several protective layers against infectious disease transmission. Deferral of donors with certain past or current medical conditions, life styles, immigration backgrounds, travel histories, or specific physical findings serves to minimize the risk of collecting blood or plasma contaminated with a transmissible agent. Selection of low-risk blood donors by this process thus

provides a strategy against rare, false-negative screening test results or pathogen inactivation failures. In addition, it lowers the risk that a donor may present during the window period of an infectious disease when screening tests are unable to detect newly infected persons.

Blood Donor Screening for TransfusionTransmitted Diseases Testing of each blood donation represents the most effective protective barrier against disease transmission for which laboratory screening tests are available and implemented (Table 82). Blood donor screening tests were introduced and improved incrementally over decades. The first screening test that was mandated was syphilis testing in the 1950s; this test is still required today. Hepatitis B surface antigen, the second required blood screening test, was introduced in 1971. Most current blood donor screening tests in the United States were implemented since the 1980s: testing for anti-HIV-1 in 1985 (combination testing for anti-HIV-1/2 in 1992), antihepatitis B core as a surrogate test for non-A, non-B hepatitis in 1986-1987 (licensed in 1991 to prevent HBV transmission in the transition period between waning hepatitis B surface antigen levels and appearance of protective anti-HBs), testing for anti-HTLV in 1988 (anti-HTLV-I/II in 1997) and anti-HCV in 1990. In 1995, HIV-1 p24 antigen testing was implemented as an interim measure to reduce the number of so-called HIV-1 window-period donations, namely, potentially infectious blood donated in the interval between new HIV infection and the appearance of detectable antibodies in the bloodstream. The test was discontinued in 2002 when NAT assays for HIV-1 were licensed. NAT assays for HIV-1 and HCV were introduced in 1999; screening of blood donors for WNV was begun by NAT technology in 2003; evaluation for bacterial contamination of platelets started in 2004; and the latest test, screening

Copyright © 2008 by the AABB. All rights reserved.

TABLE 8-2.

Pathogen

Routine US Blood Donor Screening and Confirmatory Tests Test

Methodology

Confirmatory Test

Comments

Hepatitis B surface antigen (HBsAg)

Enzyme immunoassay (EIA)/ chemiluminescence immunoassay (ChLIA)

Neutralization test [HBsAg testing is repeated after addition of anti-HBV to donor serum. Presence of HBsAg is confirmed if the test is now nonreactive (ie, neutralizable).]

Donors confirmed HBsAg positive by neutralization have to be considered acutely or chronically infected (see Figs 8-1, 8-2, and Table 8-3), and must be permanently deferred. Deferral is also required if HBsAg reactivity is not neutralizable but if the donor sample is repeat reactive for antibodies to hepatitis B core antigen (antiHBc), indicating acute or past infection; a very small subset of these donors may also have detectable HBV DNA. Donors with nonneutralizable (unconfirmed) HBsAg reactivity and nonreactivity for anti-HBc may be retested a minimum of 8 weeks after the initial HBsAg reactivity and may be reinstated if the HBsAg test is now nonreactive.

Anti-HBc

EIA/ChLIA

No confirmatory test has been licensed.

Reactivity for anti-HBc suggests acute or chronic HBV infection, unless the test result is false reactive (Table 8-3). Donors are deferred if they are reactive on more than one occasion or on a second anti-HBc assay.

Viruses Hepatitis B (HBV)

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261

(Continued)

Pathogen

Routine US Blood Donor Screening and Confirmatory Tests (Continued) Methodology

Confirmatory Test

Comments

HBV nucleic acid amplification testing (NAT)

Polymerase chain reaction (PCR) or transcription-mediated amplification (TMA)

Resolve NAT reactive pool by individual donation testing and confirm NAT reactivity by alternative NAT method or follow-up.

HBV NAT assays have been licensed by the FDA in the United States, but have not been widely implemented. The incremental benefit of HBV minipool NAT over a sensitive HBsAg assay may be minimal,71,72 although at least one study predicted the increment in safety relative to HBV serologic screening to be similar to that for hepatitis C virus (HCV) and in excess of that for human immunodeficiency virus (HIV).73 Individual donation NAT can be expected to detect an incremental yield of 13-15 infectious units per 107 donations compared with a sensitive HBsAg assay or HBV minipool (MP) NAT.71

Anti-HCV

EIA/ChLIA

Licensed multiantigen test (ie, recombinant immunoblot assay or RIBA) or HCV RNA

Donors with repeat-reactive anti-HCV screen results must be deferred, and in-date blood components must be quarantined. Results of confirmatory testing (RIBA) and NAT are helpful in donor counseling and referral.74 RIBA- or NAT-negative donors may be eligible for reentry if specific requirements for follow-up testing are met.

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AABB TECHNICAL MANUAL

Test

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Hepatitis C (HCV)

262

TABLE 8-2.

HCV NAT

Initial step in confirming NAT reactivity on a pooled sample (MP NAT) is repeat NAT on individual samples (ID NAT). Samples reactive by multiplex HIV/HCV NAT should be confirmed by discriminatory NAT. With respect to the efficacy of adding MP NAT to anti-HCV screening, it has been estimated that addition of combination MP NAT for HIV and HCV prevented an additional 56 cases of transfusion-transmitted HCV infection and five cases of HIV transmission in the United States annually at a cost of $1.5 million to $4.3 million per quality-adjusted year of life.75

EIA. Sensitivity of licensed EIAs may vary, depending on HIV strains, showing optimal performance on clade B strains most commonly found in the United States. Sensitivity for other clades, particularly group O HIV, has been suboptimal in the past.

Based on geographic concentration of group O HIV infections, the FDA recommended indefinite deferral of

HIV-1 Western blot or immunofluoresence assay (IFA), HIV-2 EIA, HIV-2 Western blot or IFA, HIV RNA

Donors with repeat reactive HIV-1/2 screen results results must be deferred (reentry may be considered if confirmatory tests are negative and specific requirements for follow-up testing have been met). Confirmatory testing should be performed for donor notification and counseling purposes. Donors with confirmed positive HIV-1 or -2 by Western blot or IFA must be indefinitely deferred and are not eligible for reentry. Results of NAT are helpful in donor counseling and referral. Figure 8-3 shows a decision tree for anti-HIV-1/2 testing of blood donors.

Infectious Disease Screening

Discriminatory NAT (uses specific primers to identify which virus caused reactivity in multiplex NAT), additional NAT (using different primers, amplification methods, or both) or demonstration of seroconversion on followup.

CHAPTER 8

Human immuno- Anti-HIV-1/2 deficiency virus (HIV-1, -2)

PCR or TMA

ä&#x160;ł

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263

(Continued)

264

TABLE 8-2.

Routine US Blood Donor Screening and Confirmatory Tests (Continued)

Pathogen

Test

Comments

blood and plasma donors who were born, resided, or traveled in certain West African countries since 1977 or had sexual contact with someone identified by these criteria. Since then, at least one EIA has been licensed for blood donor screening in the United States with documented sensitivity for HIV group O isolates, allowing discontinuation of deferral based on connection of the donor to West Africa.

HIV-1 NAT

PCR or TMA

As above for HCV NAT

The same principles apply as for HCV NAT above.

Anti-HTLV-I/II

EIA/ChLIA

No specific confirmatory test has been licensed by the FDA. Results of a second different HTLV-I/II screening test may be used. Unlicensed tests or protocols are available (confirmatory immunoblot, immunoflurescent assays, NAT).

Blood donations with repeat reactivity on HTLV-I/II screening EIA/ChLIA must not be used for transfusion. In-date products from this donor must be quarantined, but there is no requirement for look-back on previous donations from this donor. Donors must be indefinitely deferred if results by a second different HTLV-I/II screening EIA/ChLIA on the same donation are repeat reactive or if the donor tests repeat reactive on a subsequent donation.

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AABB TECHNICAL MANUAL

Human T-cell lymphotropic viruses, types I and II (HTLV-I, -II)

Confirmatory Test

ä&#x160;´

HIV-1, -2 (Continued)

Methodology

West Nile virus (WNV)

WNV NAT

PCR or TMA

Repeat NAT, IgM anti-WNV and follow-up for seroconversion.

WNV NAT-reactive donations must not be used for transfusion. In-date components from WNV NAT-positive donors should be quarantined.

Such donors are deferred from blood donation for at least 120 days. To optimize sensitivity in case of regionally increased disease activity, WNV NAT on pooled samples is converted to individual donation testing on the basis of defined trigger thresholds.76

Culture or alternative bacteria detection methods.

Culture methods rely on detection of CO2 production or O2 consumption during bacterial growth. Alternative bacterial detection methods use fluorescent labeling or gram staining coupled with microscopic examination, or multireagent strip testing for evidence of bacteria metabolism.

Nucleic acid-based and cell wallbased detection methods are under development.

Bacterial culture, isolation, and identification methods.

Donor notification and management in cases of initially positive bacteria detection tests are complex issues that may require careful further investigation, including consideration of public health concerns.77

Bacteria CHAPTER 8

All species

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265

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266 ä&#x160;´

TABLE 8-2. Routine US Blood Donor Screening and Confirmatory Tests (Continued) Test

Methodology

Confirmatory Test

Comments

Syphilis

Nontreponemal (Reagin) or treponemal tests

Nontreponemal EIAs, Treponemal tests: fluorescent treponemal antibody absorption (FTA-Abs), Treponema pallidum immobilization (TPI), T. pallidum hemagglutination (TPHA).

Treponemal tests may serve as confirmatory tests for nontreponemal tests. A specific treponemal test such as FTA may serve as confirmatory test for a treponemal hemagglutination test.

Blood donations that are reactive or indeterminate on a nontreponemal test must not be used (and the donor deferred) unless a qualified confirmatory test is nonreactive. In the latter case, the unit must be labeled reactive by a screening test for syphilis, and must be negative by FTA or specified confirmatory test; the donor may be reentered. Donors with confirmed positive results can be reinstated after 12 months with documentation of treatment.

Anti-T. cruzi

EIA

No specific confirmatory tests have been licensed, but testing by radioimmunoprecipitation assay (RIPA) or indirect fluorescence assay (IFA) is generally used for confirmation of repeat-reactive EIA results.

Blood donations repeat reactive by EIA must not be used for transfusion. The donors should be indefinitely deferred and notified of their deferral. Confirmatory testing by RIPA is recommended for donor counseling. Because confirmatory tests have not been licensed, the result of confirmatory testing cannot be used for donor reentry.62

Parasites T. cruzi

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Pathogen

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267

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ä&#x160;ł

FIGURE 8-1. Typical serologic course of acute hepatitis B virus (HBV) infection with recovery. Hepatitis B surface antigen (HBsAg) is the first marker to appear in acute infection, usually 30 to 60 days after exposure (ranging from 1-2 weeks to 11-12 weeks). In persons who recover from acute infection, this marker disappears from serum within 6 months from exposure. Hepatitis B e antigen (HBeAg) is generally detectable during acute infection. Presence of this marker correlates with higher titers of HBV and greater infectivity. Detection of immunoglobulin M (IgM) class antibody to hepatitis B core antigen (anti-HBc) demonstrates acute HBV infection. Its appearance coincides with onset of clinical symptoms and remains detectable for approximately 6 months. Total antiHBc remains detectable for years. Antibody to hepatitis B surface antigen (anti-HBs) replaces anti-HBc during convalescence and indicates recovery and immunity from reinfection. Courtesy of the Centers for Disease Control and Prevention.

268 ä&#x160;´ AABB TECHNICAL MANUAL

FIGURE 8-2. Typical serologic course of progression to chronic hepatitis B virus (HBV) infection. In contrast to acute HBV infection with recovery, hepatitis B

surface antigen (HBsAg), variably accompanied by hepatitis B e antigen (HBeAg), persists for >6 months, and anti-HBs never appears. Antibodies to hepatitis B core antigen (anti-HBc) and e antigens (anti-HBe) follow the same course as in acute HBV infection with recovery, except that anti-HBe may appear much later. Courtesy of the Centers for Disease Control and Prevention.

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CHAPTER 8

Infectious Disease Screening

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TABLE 8-3. Interpretation of the Hepatitis B Test Panel Tests

Results

HBsAg

Negative

Anti-HBc

Negative

Anti-HBs

Negative

HBsAg

Negative

Anti-HBc

Positive

Anti-HBs

Positive

HBsAg

Negative

Anti-HBc

Negative

Anti-HBs

Positive

HBsAg

Positive

Anti-HBc

Positive or negative

IgM anti-HBc

Positive or negative

Anti-HBs

Negative

HBsAg

Positive

Anti-HBc

Positive

IgM anti-HBc

Negative

Anti-HBs

Negative

HBsAg

Negative

Anti-HBc

Positive

Anti-HBs

Negative

Interpretation

Susceptible

Immune because of natural infection

Immune because of hepatitis B vaccination

Acutely infected

Chronically infected

Four interpretations possible*

*Four interpretations: 1. Might be recovering from acute HBV infection 2. Might be distantly immune and test not sensitive enough to detect very low level of anti-HBs in serum 3. Might be susceptible with a false-positive anti-HBc 4. Might be undetectable level of HBsAg present in the serum and in the person who is actually chronically infected Courtesy of the Centers for Disease Control and Prevention.

Anti-HBc = hepatitis B core antibody; anti-HBs = hepatitis B surface antibody; HBsAg = hepatitis B surface antigen; IgM = immunoglobulin M.

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FIGURE 8-3. Decision tree for human immunodeficiency virus, types 1 and 2, (anti-HIV-1, -2) testing of blood donors. If enzyme immunoassay (EIA) testing is nonreactive, nucleic acid amplification testing must also be nonreactive before release of a donation.

IFA = immunofluorescence assay; WB = Western blot.

for Chagas’ disease by antibody testing, was widely implemented in 2007.

General Principles of Screening Blood donor screening tests must have high sensitivity (proportion of true reactives that are correctly identified by the test) to be effective, but the tests must also be sufficiently specific (specificity is the proportion of true negatives that are correctly identified) to prevent unnecessary rejection of suitable donors and to ensure adequate blood availability.

Most relevant for blood donor counseling is the positive predictive value (PPV) or precision rate, ie, the proportion of donors with reactive test results who are correctly identified. The PPV is calculated by multiplying the sensitivity of the test with the disease prevalence to obtain the number of true reactives.

This value is then divided by the sum of true reactives plus false reactives (false reactives are calculated as 1– specificity × 1– prevalence).

For example, current tests for HIV-1/2 antibody screening achieve close to 100% sensitivity with approximately 99.8% specificity (virtually all donors with antibodies to HIV are detected with these tests, but 2 in 1000 screened donors are false reactives). Because the current prevalence of HIV disease among US blood donors is approximately 0.01%, the PPV is low (~5%), ie, only 1 in 20 identified as repeat reactive by the test do have antibodies to HIV. To ensure safety of transfusable blood components, US blood donor screening tests must be licensed by the FDA74 or used in the context of a protocol approved by the agency.

Similar to all clinical laboratory testing, testing for the purpose of blood donor screening

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CHAPTER 8

must meet applicable regulatory requirements and quality assurance expectations.78 Testing is required for each donation and must use a concurrently drawn blood sample. Serologic tests are initially performed singly; nonreactive results are interpreted as negative, allowing release of the component for distribution.

Reactive results are repeated in duplicate; if both repeat results are nonreactive, then the test result is interpreted as negative. If one or both repeat results are reactive, then the test result is considered “repeat reactive,” and the associated blood donation cannot be used for transfusion. Supplemental tests are performed on repeat-reactive samples primarily for donor notification and counseling purposes. NAT testing and bacteria detection follow the same general principle of initial testing designed to screen for the presence of a pathogen for the purpose of preventing the release of a possibly infectious donation for transfusion, which is followed in case of a reactive screening result with follow-up testing to confirm or refute the result.

Screening for Transfusion-Transmitted Viruses Screening for viruses depends primarily on serologic methods, generally in the format of enzyme immunoassays (EIAs) or chemiluminescence immunoassays (ChLIAs), which allow sensitive and specific detection of viral antigens or antibodies against viral antigens in blood donor samples. In the previous decade, NAT methods were added, mainly to detect infectious blood donations given during the window period of HIV and HCV infection and, in some countries, for more sensitive detection also of HBV. More recently, NAT was successfully used without an additional serologic test to detect blood donations contaminated with WNV.42,79,80 Serologic methods have the advantage of simplicity, low cost, and high throughput, which is important for rapid release of cellular blood components. Both antigen and anti-

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body tests have been greatly improved over time through development of highly specific monoclonal antibodies, recombinant proteins, and synthetic peptide antigen assays, which use optimized testing formats (socalled third-generation EIAs).71,81,82 However, the ability to detect infectious donations does not depend solely on assay performance. For a test to identify the presence of a viral marker of infection, the infection has to be present in the sample. Because viral infections naturally progress through stages during which viral markers incrementally appear in, and sometimes subsequently disappear from, the bloodstream (Fig 8-4),83 infectious donations may be missed unless the assay or assay combinations used are capable of detecting the targeted markers at the same time as blood becomes infectious.

HIV and hepatitis C virus (HCV) antibody tests, for example, detect infections only after seroconversion when the humoral immune response has begun to produce detectable antibodies. By engineering the antibody test to detect an early IgM antibody component (as opposed to only IgG antibody), HIV antibody tests became significantly more sensitive to detect infection, but the tests will still miss infectious donations in the preseroconversion stage. This limitation is important because infectious HIV (or HCV) is usually present in high concentrations in these samples, and blood donors are highly infectious during this time. Antigen tests that directly target viral antigens are capable of detecting viral infections earlier, but those tests require a minimum level of antigen to be present before detection becomes possible.

NAT assays that target viral genetic sequences and that signal their presence through amplification methods can identify viruses in concentrations that are several orders of magnitude lower than antigen tests and are, therefore, capable of detecting infectious samples earlier than serologic tests.75 As a result, HIV1 p24 antigen testing could be discontinued in

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FIGURE 8-4. A schematic, semiquantitative display of the progression of HIV markers. From top to bottom: WB = Western blot; Ab = HIV antibody; RNA = HIV RNA; LS-Ab = HIV antibody determined by sensitive/less sensitive enzyme immunoassay (EIA) testing strategy; p24 Ag = HIV p24 antigen. Time line depicts the time of exposure (day 0) through the first 200 days of infection. As each of the markers appears in the bloodstream, the infection is assigned a new stage from 0, characterized by undetectable viral markers in blood samples, except for possible transient low-level concentration of virus (“viral blips”),86 through Stage I (definitive HIV RNA viremia), Stage II (p24 antigenemia), Stage III (HIV EIA antibody reactive), Stage IV (WB indeterminate, “I”), Stage V (WB positive without p31 band, “P*”), and Stage VI (WB positive, “P” with p31 band). Stages I-VI were derived from analysis of 435 serial samples from 51 plasma donors with new HIV-1 infection. Incorporation of LS Ab EIA testing would allow further characterization of Stage VI samples as representing recent vs early chronic infection, ie, infection that occurred within vs beyond approximately 6 months from antibody seroconversion by an immunoglobulin M-sensitive EIA. The standardized optical density (OD) cutoff for the LS Ab EIA may be varied with recommended cutoffs from 0.5 to 1.0. Shown in the figure are the results for an OD cutoff of 0.75, as chosen in the original publication. Cutoffs of 0.5 and 1.0 would result in average demarcations of recent from early chronic infection at 124 and 186 days, respectively. Reproduced with permission from Fiebig et al.83

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the United States without loss in sensitivity when HIV-1 NAT assays were licensed for blood donor screening. Similarly, a test for HCV core antigen did not offer greater sensitivity in blood donor screening once HCV NAT had been established,84 and it was not implemented in the United States but could be useful in countries with limited resources where NAT may be difficult to set up and maintain.

Replacement of serologic tests with NAT assays only is unlikely in the near future. NAT assays have on rare occasions failed to detect viral sequences in people who are truly infected with HCV, for example, while antibody tests correctly demonstrated exposure to the virus.85 Another issue with NAT assays for blood donor screening is that they are currently generally performed on so-called minipools of 6 to 24 donor samples (MP NAT).75 This approach lowers costs and speeds up testing but reduces test sensitivity, as was demonstrated by rare cases in which blood donations that transmitted HIV and HCV, respectively, were missed by MP NAT but were detectable by individual donation (ID) NAT.24,25 Even ID NAT may not be able to detect all infectious donations because of sampling restrictions during the donation process. Detection of very low viral concentrations in a 300-mL blood component may not be possible, even with single copy detection sensitivity.86 NAT assays for HBV blood donor screening have been licensed in the United States, but they have not been widely implemented because of already high sensitivity of current HBsAg assays in combination with anti-HBc testing and relatively low morbidity and mortality from transfusion-transmitted HBV infections, compared with HIV and HCV disease.87 Despite test limitations, the overall performance of current blood donor screening is excellent, as evidenced by the low residual risk of transfusion-transmitted infections from

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screened donations (Table 8-1). Risk estimations for viral transmissions are based on calculations according to the incidence-windowperiod model,88 which assumes that testing and sampling failures occur at a negligible rate and that the main contributing factor for residual risk are blood donations given during the window period of infection. According to the model, the residual transmission risk from screened donations is directly related to the rate of new infections in the blood donor population and the length of the infectious window period, ie, the average time interval between infectiousness of a blood donation and detectability of the infection by the screening test (or tests) used. Originally developed to project residual risk of HIV transmission by screened blood donations in the United States, the principles of the model have been applied to other blood-borne viral infections worldwide and have been generally accepted as yielding meaningful and reliable estimates of the true risk.

Syphilis testing relies on serologic detection of antibodies to antigens associated with Treponema pallidum, the causative agent of the disease. Screening with standard serologic test for syphilis (STS) lacks sensitivity, because primary spirochetemia precedes appearance of detectable antibodies.89 Specificity is also limited because most positive STS results 1) represent so-called biological false positives (antibodies are directed against nontreponemal antigens), 2) reflect inadequately treated noninfectious syphilis, or 3) reflect remaining serologic reactivity after effective treatment.

Given the lack of demonstrable transmission of syphilis in our day (the last reported case of transfusion-transmitted syphilis in the United States occurred in 1968), the utility of serologic syphilis testing has been questioned, which in turn has limited initiatives to develop sensitive and specific NAT assays for Treponema pallidum in blood donor screening.

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Screening for Bacterial Contamination Screening for bacterial contamination of platelet donations uses time-consuming culture methods or less effective but more rapid detection strategies. Bacterial staining or measurement of glucose and pH as surrogate markers for bacteria metabolism14 are used in some hospitals as a release test while more effective methods are in development. Figure 8-5 shows a flow diagram for bacteria detection in platelet units, outlining recommended actions on the part of blood suppliers, transfusion services, and clinicians that are based on initial test results.

Screening for Vector-Borne Bacteria and Parasites, as well as Prions Given the low risk of transfusion transmission, lack of suitable screening tests, or both, blood donor screening for vector-borne bacteria and parasites as well as prions has not yet been implemented, with the notable exception of T. cruzi. Screening for this parasite, which has been in effect for years in South American countries where the disease is endemic,61 is now being implemented by most US blood suppliers.

Malaria is the other parasitic disease for which some form of blood donor screening by laboratory tests is being performed in endemic areas. However, the merits of this approach for areas where malaria is not endemic is being debated.60 Supplemental or Confirmatory Testing The purpose of supplemental blood donor testing is confirmation of repeat-reactive or preliminary positive screening results (see “General Principles of Screening” above), primarily for donor notification and counseling and, less commonly and only where applicable, to support reinstatement of donors who were deferred on the basis of obvious false-positive results. Testing algorithms, technical aspects, and interpretation of results of

supplemental assays are detailed and often complex, particularly for immunoblot assays.

Furthermore, they are to some degree subjective, which generates on occasion false-positive results and more commonly ambiguous (indeterminate) results in a minority of donors.90-92 Licensed supplemental assays are also not available for all blood donor screening tests, most notably HTLV-I/II and antiHBc tests, which complicates the counseling of affected donors. Nevertheless, supplemental assays achieve satisfactory resolution of screening test results in most cases, which is important in notifying blood donors with abnormal test results because these results frequently cause confusion and emotional upset.93 In the United States, supplemental testing must use FDA-approved reagents and algorithms and, in the case of HIV testing, must rule out both HIV-1 and HIV-2 infection. Available supplemental tests and their use are summarized in Table 8-2. Still under development are supplemental testing strategies for NAT screening results, which currently rely on repeat NAT testing and donor follow-up for confirmation.

Look-Back The current donation from a blood donor who is found repeatedly reactive on one of the routinely performed screening assays for transfusion-transmissible diseases is withheld from distribution and destroyed. Prior donations from donors who were newly identified as infected with HIV or HCV require quarantine of untransfused components and lookback to identify and notify the recipients of these previous potentially infectious units so they can be tested and counseled.94,95 Lookback has also been recommended for all prior donations from donors identified as confirmed positive by recently implemented Chagas’ disease screening.62 The rationale for look-back is that previous donations may have been donated during the window period

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Platelet unit AABB compliant testing

Unit cleared for transfusion

Negative result

Culture-based test

Nonculture-based test; Gram’s staining, pH, glucose test

Negative result

Unit cleared for transfusion

Initial positive or abnormal results

Quarantine unit and co-components.

Confirm with a second culture-based test, if the initial test was nonculture-based test. Identify the organism.

Trace the source (skin flora or donor bacteremia).

Donor notification may be required.

Positive result and platelet unit not transfused.

Positive result and a cocomponent has already been transfused.

Negative result

Release unit and co-components from quarantine.

Discard platelet unit and co-components.

Posttransfusion notification: • Notify transfusing facility, hospitals, transfusing physician or a combination immediately.

• Communicate regarding type of microorganism species and antimicrobial susceptibilities.

Clinician response: • Culture residual component, if available.

• Culture transfusion recipient (ie, blood culture).

• Retain isolates.

• Communicate results to medical director of transfusion services.

FIGURE 8-5. Flow diagram summarizing bacteria detection steps and recommended actions to be taken by blood suppliers, transfusion services, and clinicians according to testing results. Reproduced with permission from Rao et al.54

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of infection and could have been infectious despite no reactivity on the screening assay.

Rules for the look-back process are complex, considering many variables and special situations. In general, look-back begins with a search by the blood collection facility for the most recent nonreactive donation from the implicated donor and all units donated during the preceding 12 months. This research assumes that any prior donations that could have been infectious but were not detected because of the window-period phenomenon would have been contained in this time frame. The information on any identified unit (or units) is forwarded to the transfusing facility, which searches its records for the recipient (or recipients). The final step is recipient notification, preferably through the recipient’s physician to allow testing and counseling as appropriate. In case of HIV and HCV, notification of relatives or legal representatives is mandatory if the recipient is deceased or judged incompetent.

Look-back was most effective after implementation of blood donor testing for HIV in the mid-1980s.96 Retrospective HCV lookback in the late 1990s, which was intended to identify recipients of blood components that were donated before implementation of HCV screening in 1990, yielded relatively few recipients who could be contacted and who benefited from the notification.97-99 Nevertheless, look-back did reach some transfusion recipients who did not know that they were infected and who would not have otherwise benefited from testing, counseling, adopting a healthier lifestyle, and undergoing antiviral therapy.

Testing of Autologous Blood Donations Specific federal rules apply to infectious disease testing of autologous units and notification of donor-patients and the hospital staff in cases with positive test results. Autologous units that are transfused within the collection facility are not required to undergo infectious

disease screening. Those that are shipped for transfusion outside the collection facility must be tested with the same infectious disease marker assays that are required for allogeneic blood donations. These tests must be performed before shipping on at least the first unit collected during each 30-day period.21(p39) If significant positive results are obtained, then the patient’s physician, the patient, and the receiving facility must be informed. A controversial issue is whether transfusing facilities have the right to refuse acceptance of autologous units that tested positive for infectious disease markers. Because asymptomatic HIV infection has been recognized as a qualifying condition under the Americans with Disabilities Act (ADA), it can be argued that not accommodating HIV-infected persons who wish to donate and receive autologous blood might constitute a violation of the ADA. It appears that in clinical practice there is no unified position on this issue. Most blood centers collect autologous blood from infected donors, but hospitals or clinics may restrict acceptance of infectious blood units according to self-assessment of risks and benefits for all parties involved and unique local circumstances. The AABB has not taken a position on this issue, but strongly recommends that each institution that collects or transfuses autologous blood carefully weigh the implications of restricting or refusing this service.100 Safeguarding Plasma Derivatives from Contamination with Infectious Pathogens Infectious Risk Potential Current infectious risks of plasma derivatives, namely manufactured products that use human plasma as raw material, are minimal as a result of incremental improvements in blood and plasma donor screening over time and addition of pathogen removal and inactivation methods to the manufacturing process.

In understanding current and historical infec-

Copyright © 2008 by the AABB. All rights reserved.

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tious risk potentials associated with plasma derivatives, several basic facts need to be considered. First, in large-scale manufacture of plasma derivatives, thousands of plasma donations are combined in a single pool, which creates an opportunity for pathogen contamination and spread throughout the pool, even if only a single infectious donation is present. Second, pathogens differ in their susceptibility to removal and inactivation methods. None of the currently available systems can guarantee complete removal or inactivation of non-enveloped viruses. Third, the traditional fractionation process for preparation of plasma derivatives in itself achieves some reduction of contaminating viruses, if present. However, this effect is uneven and more pronounced for albumin and immune globulin protein fractions compared with coagulation factors. Fourth, the stability of various plasma protein fractions is different, which allowed the incorporation of a pasteurization step in the fractionation of albumin early on whereas more sophisticated antiviral measures had to be developed in the manufacture of other derivatives.101 The latter two reasons, plus additional presence of protective antibodies in case of immune globin products, explain why albumin and immune globulins102 have historically been very safe products as far as infectious risks are concerned (with notable exception of HCV transmissions from immune globulin products in the 1970s and 1990s).

Coagulation factor concentrates, however, have in the past transmitted HIV, infectious hepatitis viruses (HAV, HBV, and HCV), and parvovirus to large cohorts of recipients, primarily patients with hemophilia.103 In response to this tragedy, blood and plasma donor screening was incrementally improved as described in this chapter, and additional measures such as NAT screening of plasma donations for HAV and parvovirus B19 and NAT testing of manu-facturing pools were implemented.104 Major improvements were

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also made in the development of pathogen reduction and inactivation methods, including heating; solvent/detergent treatment; and, most recently, nanofiltration.104 These methods are now routinely used, often in combination, in the manufacturing of plasma derivatives. As a result, there have been no reported transmissions of HIV from manufactured clotting factor products in the United States since 1987 and no cases of HAV, HBV, or HCV since 1998.105 Pathogen Reduction and Inactivation Methods Pathogen reduction and inactivation methods (PRIMs) add another layer of safety to donor selection and screening by reducing the pathogenic capabilities of a broad range of organisms, including viruses, bacteria, protozoa, and fungi, with notable exception of prions, which resist most existing PRIMs but may be removable by specially designed leukocyte reduction filters.70 PRIMs have long been used in the manufacture of plasma derivatives and (in some instances) Fresh Frozen Plasma equivalents, because the relative stability of plasma proteins made it comparatively easy to use heat or chemical processes to inactivate infectious agents in the product without damaging the therapeutic ingredient.101 More challenging is treatment of blood components containing living cells or labile plasma proteins.

Many different PRIMs have been developed.2 Among the earliest methods are heat and solvent/detergent treatment. Solvent/ detergent treatment is highly effective against lipid-enveloped viruses, including HIV and HCV, but it is relatively ineffective against non-enveloped viruses; the most important are HAV and parvovirus B19. Alternative PRIMs for treatment of plasma are photochemical treatment in which photosensitive compounds such as methylene blue (MB) psoralens, or riboflavin (vitamin B2) are added and activated by exposure to light of an

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appropriate wavelength. Pathogen inactivation occurs through oxidation of MB or binding to (psoralens and riboflavin) DNA. The photochemical methods have the advantage of being suitable for single-unit treatment, avoiding the problems with pooling, especially the concern of spreading infectious agents that are not inactivated or removed throughout the pool. Some loss of coagulation factor activity occurs, the significance of which is being studied. MB-treated plasma has been available for years in European countries. Psoralens-UV light treatment has also been approved in Europe, and the riboflavin-based method is in earlier stages of investigation. Psoralens- and riboflavin-based photochemical methods are suitable for treatment of platelets, and the former method has gained approval for clinical use in Europe.

Adoption of photochemical principles for pathogen inactivation of RBC components is complicated by absorption of longwave UV light by hemoglobin. New chemical systems

known as frangible anchor-linker effector molecules and PEN-110, which do not require light activation, are being developed and studied for possible clinical use in the future.

PRIMs are currently not routinely applied to blood components prepared in the United States. Solvent/detergent-treated plasma was introduced in the United States in 1998 but was never universally adopted and was withdrawn from distribution in 2002 following a series of deaths from excessive bleeding and thrombotic complications associated with its use in patients with severe liver disease and liver transplants. Since then, no new component products treated with PRIMs have been made available for clinical use in the United States. Nevertheless, a recent international consensus conference backed by US advisory groups recommended accelerated development and broad implementation of pathogen inactivation when a feasible and safe method is available.106

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