Blood screening by nucleic acid amplification technology: current issues, future challenges.

BACKGROUND: Nucleic acid amplification technology (NAT) is presently being evaluated in US clinical trials to determine the safety and efficacy of mini-pool testing for human immunodeficiency virus (HIV) and hepatitis C virus (HCV) RNA in the blood-donor population. Although the risk for transfusion-transmitted HIV and HCV infection is extremely low, there is still a small chance that blood donated by infected individuals before seroconversion can escape detection by current antibody-based assays. METHODS: This report describes the amplification technologies being used and reviews several issues surrounding NAT-based blood screening. The performance features of NAT and current enzyme immunoassay technologies are compared, and the benefits of NAT in reducing transfusion-transmitted infections are discussed. CONCLUSIONS: The current US clinical trials of mini-pool NAT testing for HIV and HCV RNA have successfully identified preseroconversion infectious blood units. Although the current NAT-based screening systems are semiautomated, mini-pool testing represents an unprecedented innovation among government and nongovernment agencies in the highly regulated blood transfusion industry. Despite cost-effectiveness issues, based on the public perception of infectious diseases acquired through blood transfusion, NAT-based screening of the blood supply is expected to become a standard in transfusion medicine.     

Screening of HIV infection: role of molecular and immunological assays

Due to technical improvements and new developments of immunological assays, the reliability of serological laboratory diagnosis of HIV infection has improved considerably and the residual risk, due to the diagnostic window for transfusion-transmitted HIV, has been reduced significantly. Through the addition of nucleic acid amplification tests (NAT) to blood donor screening, the residual risk can de further decreased by up to 50%, depending on the sensitivity of the NAT protocol and whether individual or pooled blood donations are screened. In-house and commercially available NAT have been implemented in blood banks as HIV only or multiplexed HIV and hepatitis B or C virus assays. As an alternative to separate antigen and antibody screening, combined fourth-generation assays have been developed in 1997, and have achieved a high degree of sensitivity and specificity. Thus, they can replace stand-alone antigen and third-generation antibody assays. While they are used in the routine diagnostics of HIV infection in many countries throughout the world, they probably represent no alternative for NAT in blood-donor screening in industrialized countries. In the next few years, technical improvements will further simplify NAT screening. While there is still some potential to improve the detection threshold of NAT, the sensitivity of the antigen module of fourth-generation assays (a lowest concentration of 3-5 pg of p24 antigen) is probably very close to its technical limit.     

HIV Screening in the Health Care Setting: Status, Barriers, and Potential Solutions

Thirty years into the human immunodeficiency virus (HIV) epidemic in the United States, an estimated 50,000 persons become infected each year: highest rates are in black and Hispanic populations and in men who have sex with men. Testing for HIV has become more widespread over time, with the highest rates of HIV testing in populations most affected by HIV. However, approximately 55% of adults in the United States have never received an HIV test. Because of the individual and community benefits of treatment for HIV, in 2006 the Centers for Disease Control and Prevention recommended routine screening for HIV infection in clinical settings. The adoption of this recommendation has been gradual owing to a variety of issues: lack of awareness and misconceptions related to HIV screening by physicians and patients, barriers at the facility and legislative levels, costs associated with testing, and conflicting recommendations concerning the value of routine screening. Reducing or eliminating these barriers is needed to increase the implementation of routine screening in clinical settings so that more people with unrecognized infection can be identified, linked to care, and provided treatment to improve their health and prevent new cases of HIV infection in the United States.     

Pneumocystis jirovecii pneumonia in non-HIV-infected patients in the era of novel immunosuppressive therapies

In human immunodeficiency virus (HIV)-infected patients, Pneumocystis jirovecii pneumonia (PCP) is a well-known opportunistic infection, and its management has been established. However, PCP is an emerging threat to immunocompromised patients without HIV infection, such as those receiving novel immunosuppressive therapeutics for malignancy, organ transplantation, or connective tissue diseases. Clinical manifestations of PCP are quite different between patients with and without HIV infections. In patients without HIV infection, PCP rapidly progresses, is difficult to diagnose correctly, and causes severe respiratory failure with a poor prognosis. High-resolution computed tomography findings are different between PCP patients with HIV infection and those without. These differences in clinical and radiologic features are the result of severe or dysregulated inflammatory responses that are evoked by a relatively small number of Pneumocystis organisms in patients without HIV infection. In recent years, the usefulness of PCR and serum β-d-glucan assay for rapid and noninvasive diagnosis of PCP has been revealed. Although corticosteroid adjunctive to anti-Pneumocystis agents has been shown to be beneficial in some populations, the optimal dose and duration remain to be determined. Recent investigations revealed that Pneumocystis colonization is prevalent, and that asymptomatic carriers are at risk for developing PCP and can serve as the reservoir for the spread of Pneumocystis by person-to-person transmission. These findings suggest the need for chemoprophylaxis in immunocompromised patients without HIV infection, although its indication and duration are still controversial. Because a variety of novel immunosuppressive therapeutics have been emerging in medical practice, further innovations in the diagnosis and treatment of PCP are needed.     

用VIDAS~ HIV DUO Ultra方法进行人类免疫缺陷病毒感染筛查

目的寻找一种更快速、简便的人类免疫缺陷病毒（HIV）感染的筛查方法。方法对门诊和住院手术患者标本以及疾病控制中心质控标本，采用VIDAS HIV DUO Ultra方法（简称VIDAS）测定HIV p24抗原和HIV抗体，并与第3代酶联免疫吸附试验（ELISA）以及明胶粒子凝集法（PA）测定结果进行比较。结果对50份标本采用VIDAS进行检测，其敏感性和特异性分别为91．7％和92．1％。结论VIDAS可作为一种更简便的HIV检测方法，在生物安全控制方面较其他2种方法更安全有效。     

Immunosuppressive drugs as an adjuvant to HIV treatment

Although highly active antiretroviral therapy (HAART) has dramatically changed the epidemiological impact of HIV infection, many problems with currently used antiretroviral therapy have underscored the urgent need for additional therapeutic approaches. Structured treatment interruption trials, which can be considered an immune-based therapy with an autologous virus, have failed to control viral replication in most chronically HIV-1-infected patients. Alternative approaches could be the use of immunosuppressive drugs to enhance the control of viral replication mediated by their immune and antiviral properties. The use of immunosuppressive drugs may reduce the number of activated CD4 cells that support massive virus production and may prevent sequestration of CD4 T cells into lymphoid tissue, which is the place of antigen presentation and productive HIV infection. The strategy of using drugs that interfere with the HIV life-cycle, acting on the target cells of HIV rather than on viral enzymes, offers the advantage of avoiding the development of antiretroviral drug-resistant HIV mutants. However, it is not known if these approaches will clinically benefit long-term infection, by establishing a new immunological set-point that may affect the rate of disease progression. Caution is required when using HAART in combination with cytostatic drugs in HIV-1 infection until their impact and long-term safety have been investigated further in larger clinical trials.     

The History and Challenges of Blood Donor Screening in China

Since the establishment of People's Republic of China in 1949, the Chinese government has encountered several catastrophes related to transfusion transmitted diseases. The government's increasing attention to blood safety has prompted the initiation of a series of policies and measures that have enhanced the level of safety for the blood supply and met the basic clinical demands of blood for 1.3 billion people in the country.Blood donation screening strategies in China predominantly comprise donor screening and donor testing. Donor screening includes selection of low-risk blood donors by the use of a donor history questionnaire, predonation physical examination, and initial rapid donor testing. Donor testing includes direct pathogen detection and serology tests. The year 1998 marked the most transformative change in blood donor selection and screening policies in China. Before 1998, paid donation was the predominant mode of blood donation. Donor screening and donor testing were conducted before donation, and only those who were eligible were allowed to donate. To ensure the safety of blood, donor testing was performed again after donation. After the implementation of the Blood Donation Law in 1998, to promote voluntary and unpaid donation, predonation donor testing was eliminated to reduce the amount of waiting time and to provide a more convenient donation experience for blood donors. However, it is the national requirement that donated blood should undergo 2 rounds of testing using different equipment or reagents, conducted by different personnel. Donor selection has transitioned from paid donation and obligatory donation to voluntary donation with fixed volunteer groups, as the latter mode of donation provides the lowest risks. Donations are currently screened for syphilis, hepatitis C virus, HIV, and hepatitis B virus (HBV). Units, previously typed only for ABO, are now routinely tested for both ABO and Rh(D). Innovations in testing technologies and methods have also brought changes to screening parameters. For instance, screening for HBV pathogens evolved from the early use of hemagglutination method to the later use of radioimmunoassay, independent enzyme-linked immunosorbent assay, and now the widespread application of nucleic acid test (NAT). Since 2010, the Chinese government has established NAT capacity in several blood centers; and in 2015, the government invested 900 million RMB on the nationwide expansion of NAT. Although the Chinese government has worked to enhance blood safety, many challenges remain. Concern exists for rising rates of HIV infection. The existence of occult HBV infection and the transmission of emerging blood-borne diseases continue to challenge the safety of the blood supply.     

Acute seroconversion of HIV infection in the ambulatory care setting

Patients frequently visit ambulatory care settings with acute human immunodeficiency virus (HIV) seroconversion illness, but the illness is often misdiagnosed. This acute viral syndrome, or seroconversion illness, occurs after initial exposure to the HIV virus; it is often resolved before the development of HIV-specific antibodies. Primary HIV infection refers to the 12 months following infection; it includes an acute time period after exposure when routine HIV antibody testing is negative. Primary HIV infection is recognized with the help of a detailed screening history. Diagnosis is confirmed through laboratory tests that detect virus presence. The accurate diagnosis of primary HIV infection can have a beneficial effect on the patient's clinical course and also on public health prevention efforts.     

Mathematic modeling of the risk of HBV,HCV, and HIV transmission by window-phase donations not detected by NAT

BACKGROUND : Blood transfusion centers around the world have introduced minipool NAT to reduce the risk of HBV, HCV, and HIV transmission by blood donations drawn in the infectious window phase. What would be the reduction in the residual risk when minipool NAT would be replaced by single‐donation NAT? STUDY DESIGN AND METHODS : A mathematic model was developed to estimate the probability of virus transmission by blood transfusion when NAT screening methods are used for virologic safety testing. The major assumptions used are threefold: 1) The viral nucleic acid concentrations in the early window phase of infection double in 2.8 (HBV), 0.74 (HCV), and 0.90 (HIV) days. 2) The detectability of low copy numbers of viral DNA or RNA by the screening assay can be described with a probit model. 3) The probability of infection depends linearly on the logarithm of the administered dose, with 50‐percent infectivity rates at 10 (HBV and HCV) or 1000 (HIV) viral nucleic acid copies per transfusion unit (estimates based on NAT studies with samples of known infectivity in chimpanzees). RESULTS : A reasonably simple equation was obtained that allows studying the effect of the sensitivity of the NAT assay and of the pool size used for screening on the residual risk of transfusion‐transmitted infection. The computations are illustrated by using observed sensitivity estimates of various NAT methods. By using epidemiologic data among European donors over 1997 as baseline, the calculations predict that the incidence of virus transmission per 10‐million RBC transfusions reduces with the following numbers when lowering the test pool size from 96 to 1 (single‐donation testing): HBV from 11 to 13 to 3.3 to 5.1, HCV from 1.7 to 2.0 to 0.5 to 0.8, and HIV from 0.47 to 0.62 to 0.010 to 0.045 (ranges for the different NAT screening methods). CONCLUSION : A proper mathematic model for the calculation of residual infection risk by blood transfusion helps understand the impact of introducing new NAT methods for blood safety testing.     

Human immunodeficiency virus: the initial physician-patient encounter

Human immunodeficiency virus (HIV) infection is increasingly becoming a disease managed by HIV specialists. However, all primary care physicians have an important role that can affect the epidemic in the United States. These physicians must be able to appropriately identify patients at risk, screen for and diagnose HIV, provide counsel, and refer those who are infected to specialists. The primary care physician will often continue to provide medical care in collaboration with an HIV specialist. The patient will receive optimal care when the primary care physician is knowledgeable regarding HIV and the evaluation of the newly diagnosed patient. Through appropriate screening, evaluation, diagnosis, and counseling, the primary care physician will not only improve the care of the individual patient but also potentially decrease the spread of HIV. This article answers some of the questions that primary care physicians are likely to have when evaluating an adult with newly diagnosed HIV infection.     

浙江省金华市无偿献血人群艾滋病病毒感染现状分析

目的 了解浙江省金华市无偿献血者人类免疫缺陷病毒(human immunodeficiency virus,HIV)感染的状况及流行特征,以确定低危献血人群,降低输血传播HIV的风险。 方法 对2006年1月至2012年7月266 844例无偿献血者利用酶联免疫吸附试验进行抗-HIV检测,结果采用免疫蛋白印迹(WB)确证,并对抗-HIV确证阳性者进行流行病学调查与统计分析。 结果 抗-HIV抗体初筛阳性共406例,WB法确证阳性29例,阳性率为0.01%,不确定5例,其余确证为阴性。经流行病学调查,异性传播19例,占65.5%。同性传播9例,占31%。 结论 结合本地区献血人群的HIV感染状况、流行特征,应从无偿献血招募、体检咨询工作方面采取积极有效的措施,需加强与社会互动,开展对献血者艾滋病相关知识的宣教,从而达到保证血液质量与安全。     

Needlestick and sharps injury prevention

Every day while caring for patients, nurses are at risk to exposure to bloodborne pathogens potentially resulting in infections such as HIV or hepatitis B and C. These exposures, while preventable, are often accepted as being a part of the job. In the United States, needlestick injuries have begun to decrease from an estimated one million exposures per year in 1996 to 385,000 per year in 2000. This decline has resulted from the protections afforded by the Occupational Safety and Health Administration's (OSHA) Bloodborne Pathogens Standard. Reasons for the success in decreasing needlestick and sharps injuries may be attributed to the elimination of needle recapping and the use of safer needle devices, sharps collection boxes, gloves and personal protective gear, and universal precautions. The prevention of needlestick injuries has made slow progress over the past 20 years since the HIV epidemic drew attention to the deadly nature of health care work and to protection of health care worker health and safety. In Africa, where the AIDS virus originated and where the prevalence of the human immunodeficiency virus (HIV) among hospitalized patients is highest in the world, attention has been directed only recently at protecting health care workers. Nurses, especially those infected from a preventable exposure, have been at the forefront of advocacy for prevention. This article includes a review about the hazard of exposure to bloodborne pathogens and epidemiology of occupational infection. The author discusses how to apply standard methods of occupational health and industry hygiene using the hierarchy of controls framework to prevent exposure to blood, and discusses evidence-based prevention and efficacy of particular control measures. Legislative progress and implementation of enforceable policy to protect health care workers is outlined.     

Prevalence and trends of human immunodeficiency virus, hepatitis B virus, and hepatitis C virus among blood donors in Iran, 2004 through 2007

BACKGROUND: Evaluation and monitoring the prevalence of transfusion-transmissible viral infections in blood donors is a valuable index of donor selection and blood safety. This study analyzed the trends of blood-borne infections among Iranian blood donations during 4 years.
STUDY DESIGN AND METHODS: Viral screening results of 6,499,851 allogeneic donations from 2004 through 2007 were analyzed. All donations were screened for hepatitis B virus (HBV), hepatitis C virus (HCV), human immunodeficiency virus (HIV), and syphilis. The prevalence of HBV, HCV, and HIV infections per 100,000 donations and 95% confidence interval was calculated. The p value was estimated by chi-square test.
RESULTS: The prevalences of HBV, HCV, and HIV decreased during the 4-year study from 2004 through 2007. The overall prevalence was 0.56% for HBV, 0.004% for HIV, and 0.13% for HCV. There was a significant and impressive decrease in hepatitis B surface antigen prevalence from 0.73% in 2004 to 0.41% in 2007. The prevalence of HIV appeared to have decreased from 0.005% in 2004 to 0.004% in 2007 although the decrease was not significant. HCV prevalence showed a slight decline in blood donations from 0.14% in 2005 to 0.12% in 2007.
CONCLUSION: The trends of transfusion-transmitted infection prevalence in Iranian blood donations suggest that most of the safety measures employed in recent years in Iran have been effective.     

Donation deferral criteria for human immunodeficiency virus positivity among blood donors in northern Thailand

Background: The purpose of this study was to develop human immunodeficiency virus (HIV) infection donation deferral criteria for blood donors in an HIV-epidemic area of northern Thailand, where the predominant means of transmission of HIV is through heterosexual contact. Study Design and Methods: In a preliminary study, 2242 blood donors were interviewed, and their blood was tested for HIV antibodies between September 1993 and April 1994. The risk factors associated with HIV positivity were identified. Criteria to identify HIV-positive persons on the basis of a logistic equation were developed and applied to another group of 5769 prospective blood donors. Results: A multivariate analysis showed the following odds ratios (OR) for traits that were independently associated with HIV positivity: younger age (OR = 0.93 for each additional year of age), male gender (OR = 2.41), having no more than a primary school education (OR = 2.00), being in the military (OR = 1.78), being unsure of one's own blood safety (OR = 2.00), history of injecting drug use (OR = 5.36), diagnosis of syphilis or positive syphilis serologic test in the past 12 months (OR = 2.67), and genital ulcer in the past 12 months (OR = 4.56). On the basis of the model, with a limit of <10 percent loss of uninfected donors, predicted probabilities of HIV positivity alone or of markers of infection with HIV, hepatitis B virus, or Treponema pallidum were calculated. With a cutoff of 6.5-percent estimated probability of HIV infection, derived from the logistic equation, the donor deferral criteria have 33.6-percent sensitivity and 8.3-percent positive predictive value for HIV positivity and 15.5-percent sensitivity and 18.4-percent positive predictive value for markers of infection with one of the three pathogens. Conclusion: The proposed donor deferral system provides a more flexible, sensitive, and predictive tool for averting donation by those who, though HIV antibody-negative, are at a higher risk of being infected with HIV.     

Intrarectal transmission, systemic infection, and CD4+ T cell depletion in humanized mice infected with HIV-1

Intrarectal infection between men who have sex with men represents a predominant form of human immunodeficiency virus (HIV) transmission in developed countries. Currently there are no adequate small animal models that recapitulate intrarectal HIV transmission. Here we demonstrate that human lymphocytes generated in situ from hematopoietic stem cells reconstitute the gastrointestinal tract of humanized mice with human CD4(+) T cells rendering them susceptible to intrarectal HIV transmission. HIV infection after a single intrarectal inoculation results in systemic infection with depletion of CD4(+) T cells in gut-associated lymphoid tissue and other pathologic sequela that closely mimics those observed in HIV infected humans. This novel model provides the basis for the development and evaluation of novel approaches aimed at immune reconstitution of human gut-associated lymphoid tissue and for the development, testing, and implementation of microbicides to prevent intrarectal HIV-1 transmission.     

Donor testing and risk: current prevalence, incidence, and residual risk of transfusion-transmissible agents in US allogeneic donations

Over the past 20 years, there has been a major increase in the safety of the blood supply, as demonstrated by declining rates of posttransfusion infection and reductions in estimated residual risk for such infections. Reliable estimates of residual risk have been possible within the American Red Cross system because of the availability of a large amount of reliable and consistent data on donations and infectious disease testing results. Among allogeneic blood donations, the prevalence rates of infection markers for hepatitis C virus (HCV) and hepatitis B virus have decreased over time, although rates for markers of human immunodeficiency virus (HIV) and human T-cell lymphotropic virus did not. The incidence (/100?000 person-years) of HIV and HCV among repeat donors showed apparent increases from 1.55 and 1.89 in 2000 through 2001 to 2.16 and 2.98 in 2007 through 2008. These observed fluctuations confirm the need for continuous monitoring and evaluation. The residual risk of HIV, HCV, and human T-cell lymphotropic virus among all allogeneic donations is currently below 1 per 1 million donations, and that of hepatitis B surface antigen is close to 1 per 300?000 donations.     

Current status of clinical laboratory tests for the human immunodeficiency virus

The predictive values of positive and negative test results for human immunodeficiency virus (HIV) antibody are extremely high in laboratories that have good quality control and high performance standards and use licensed FDA-approved enzyme immunoassay (EIA) and Western blot standardized tests. With a carefully designed protocol, the false-positive rate of combined EIA and Western blot has been reported to be as low as 1 in 10(5). When results of HIV confirmatory antibody tests are indeterminate, other tests such as culture and nucleotide probe methods for HIV DNA or RNA may help resolve false-reactive screening EIA tests. Improvements are constantly in progress for HIV laboratory tests with the use of recombinant DNA-derived antigens and synthetic polypeptides. With the use of new-generation synthetic polypeptide antigens, specific assays to identify HIV-1 and HIV-2 have been developed. Recently, assays for the HIV regulatory gene products have been studied for their predictive potential. Antibodies to nef protein, a regulator of HIV-1 replication, may be an early indicator of HIV infection.     

The East Africa Consortium for human papillomavirus and cervical cancer in women living with HIV/AIDS

The East Africa Consortium was formed to study the epidemiology of human papillomavirus (HPV) infections and cervical cancer and the influence of human immunodeficiency virus (HIV) infection on HPV and cervical cancer, and to encourage collaborations between researchers in North America and East African countries. To date, studies have led to a better understanding of the influence of HIV infection on the detection and persistence of oncogenic HPV, the effects of dietary aflatoxin on the persistence of HPV, the benefits of antiretroviral therapy on HPV persistence, and the differences in HPV detections among HIV-infected and HIV-uninfected women undergoing treatment for cervical dysplasia by either cryotherapy or LEEP. It will now be determined how HPV testing fits into cervical cancer screening programs in Kenya and Uganda, how aflatoxin influences immunological control of HIV, how HPV alters certain genes involved in the growth of tumours in HIV-infected women. Although there have been challenges in performing this research, with time, this work should help to reduce the burden of cervical cancer and other cancers related to HIV infection in people living in sub-Saharan Africa, as well as optimized processes to better facilitate research as well as patient autonomy and safety.

KEY MESSAGES

• The East Africa Consortium was formed to study the epidemiology of human papillomavirus (HPV) infections and cervical cancer and the influence of human immunodeficiency virus (HIV) infection on HPV and cervical cancer.
• Collaborations have been established between researchers in North America and East African countries for these studies.
• Studies have led to a better understanding of the influence of HIV infection on the detection and persistence of oncogenic HPV, the effects of dietary aflatoxin on HPV detection, the benefits of antiretroviral therapy on HPV persistence, and the differences in HPV detections among HIV-infected and HIV-uninfected women undergoing treatment for cervical dysplasia by either cryotherapy or LEEP.

     

Neopterin levels during the early phase of human immunodeficiency virus, hepatitis C virus, or hepatitis B virus infection

BACKGROUND: A study was conducted to assess the diagnostic sensitivity of neopterin screening of blood donors with regard to the detection of window-phase specimens of human immunodeficiency virus (HIV), hepatitis C virus (HCV), and hepatitis B virus (HBV) infection. STUDY DESIGN AND METHODS: In total, 1002 diagnostic window-phase specimens from 98 seroconversion panels (29 HIV-1, 52 HCV, and 17 HBV) were analyzed with viral antigen detection, viral nucleic acid amplification testing (NAT), and neopterin quantitation assays. The study was completed by the analysis of 92 anti-hepatitis B core antigen (HBc)-reactive and 103 alanine aminotransferase (ALT)-elevated blood donor specimens. RESULTS: A significant association between elevated neopterin concentrations and the very early phase of HIV-1 infection was found. No significant correlation could be observed between neopterin levels and the early phase of HCV or HBV infection. Neopterin concentration was not increased in specimens from blood donors with anti-HBc reactivity or ALT elevation. CONCLUSIONS: Neopterin screening of blood donors may identify window-phase cases of HIV, but not of HCV or HBV infection. The diagnostic sensitivity of neopterin screening during the HIV window phase is similar to that of the p24 antigen test. With the introduction of viral NATs in blood screening, there is no additional benefit of neopterin screening with regard to the three blood-borne viruses HIV, HCV, and HBV. Acute phases of other infectious agents, however, have been reported to be detected by neopterin enzyme-linked immunosorbent assays.