International review of blood donation screening for anti-HBc and occult hepatitis B virus infection.

BACKGROUND: Hepatitis B core antibody (anti-HBc) screening has been implemented in many blood establishments to help prevent transmission of hepatitis B virus (HBV), including from donors with occult HBV infection (OBI). We review HBV screening algorithms across blood establishments globally and their potential effectiveness in reducing transmission risk. MATERIALS AND METHODS: A questionnaire on HBV screening and follow-up strategies was distributed to members of the International Society of Blood Transfusion working party on transfusion-transmitted infectious diseases. Screening data from 2022 were assimilated and analyzed. RESULTS: A total of 30 unique responses were received from 25 countries. Sixteen respondents screened all donations for anti-HBc, with 14 also screening all donations for HBV DNA. Anti-HBc prevalence was 0.42% in all blood donors and 1.19% in new donors in low-endemic countries; however, only 44% of respondents performed additional anti-HBc testing to exclude false reactivity. 0.68% of anti-HBc positive, HBsAg-negative donors had detectable HBV DNA. Ten respondents did universal HBV DNA screening without anti-HBc, whereas four respondents did not screen for either. Deferral strategies for anti-HBc positive donors were highly variable. One transfusion-transmission from an anti-HBc negative donor was reported. DISCUSSION: Anti-HBc screening identifies donors with OBI but also results in the unnecessary deferral of a significant number of donors with resolved HBV infection and donors with false-reactive anti-HBc results. Whilst confirmation of anti-HBc results could be improved to reduce donor deferral, transmission risks associated with anti-HBc negative OBI donors must be considered. In high-endemic areas, highly sensitive HBV DNA testing is required to identify infectious donors.

Occult Hepatitis B Virus Infection and Liver Fibrosis in Chinese Patients.

BACKGROUND: The impact of hepatitis B surface antigen (HBsAg)-negative/hepatitis B virus (HBV) DNA-positive occult HBV infection (OBI) on the severity of liver fibrosis remains unclear. METHODS: A total of 1772 patients negative for HBsAg but positive for antibody to hepatitis B core antigen (HBcAg), stratified by the presence or absence of OBI, were selected for long-term carriage leading to elevation of ≥2 of 4 liver fibrosis indexes-hyaluronic acid (HA), laminin, type III procollagen peptide (PCIII), and type IV collagen (CIV)-at testing in a Chinese hospital. Patients were tested for serum viral load, HBV markers, and histopathological changes in liver biopsy specimens. RESULTS: OBI was identified in 148 patients with liver fibrosis (8.4%), who had significantly higher levels of HA, laminin, PCIII, and CIV than 1624 fibrotic patients without OBI (P < .05). In 36 patients with OBI who underwent liver biopsy, significant correlations were observed between OBI viral load and serum HA levels (P = .01), PCIII levels (P = .01), and pathological histological activity index (HAI) scores (P < .001), respectively; HAI scores and PCIII levels (P = .04); HBcAg immunohistochemical scores and HA levels (P < .001); and HBcAg immunohistochemical scores and PCIII levels (P = .03). Positive fluorescent in situ hybridization results were significantly more frequent in patients with OBIs (80.6% vs 37.5% in those without OBIs). Among patients with OBIs, HBcAg was detected in the liver tissue in 52.8% and HBsAg in 5.6%. CONCLUSIONS: OBI status appears to be associated with liver fibrosis severity.

Monitoring viral genomic sequences in transfusion-transmitted viruses.

BACKGROUND AND OBJECTIVES: Monitoring genomic sequences of blood-borne viruses infecting blood donors enables blood operators to undertake molecular epidemiology, confirm transfusion transmission and assess and characterize molecular and serological screening assays. The purpose of the study was to determine how blood operators globally value viral diversity surveillance and to assess its impact. MATERIALS AND METHODS: An electronic questionnaire was developed and circulated to members of the International Society of Blood Transfusion-transmitted infectious diseases working party. Responses were compiled and complete data sets were analysed. RESULTS: Ninety-seven percent of respondents agreed that monitoring viral genomic sequences was important to blood operators and the transfusion community. However, only 47% of respondents are currently doing this monitoring. The main limitations reported were a lack of financial resources and expertise. Sequencing techniques, primarily next-generation sequencing and also Sanger sequencing, were considered most appropriate, with the preferred option for testing being regional or national reference centres. Respondents agreed that engagement with public health authorities needs to be enhanced. CONCLUSION: Monitoring genomic sequences of blood-borne viruses is widely considered important by the transfusion community because of its direct applications for transfusion safety, and beyond for public health in general. Therefore, there is a need to strengthen collaboration between blood operators and public health authorities. While national and regional reference centres may be the most appropriate structure for such testing, international collaborations should not be overlooked. Overcoming financial barriers will be an important hurdle for many.

Anti-HBc-nonreactive occult hepatitis B infections with HBV genotypes B and C in vaccinated immunocompetent adults.

Absence of anti-HBc reactivity with detectable anti-HBs was observed in blood donors with occult hepatitis B virus (HBV) infection (OBI). The prevalence and mechanisms underlying this uncommon condition were investigated over time in Chinese blood donors with OBI. Isolated anti-HBs OBI status was identified from 466,911 donors from Dalian, China, and monitored in follow-up (range: 2.6-84.3 months). HBV vaccination status was documented, and infecting viral strains were characterized. Of 451 confirmed OBIs (1:1035), 43 (9.5%; 1:10,858) had isolated anti-HBs as only serological marker. Isolated anti-HBs OBIs differed from anti-HBc-reactive OBIs by significantly younger age (median 24 years), higher HBV DNA (median: 20 IU/ml) and anti-HBs (median 60.5 IU/L) levels, paucity of mutations in HBV Core and S proteins, and high vaccination rate (72%). Vaccinated isolated anti-HBs OBIs (n = 31) differed from unvaccinated (n = 11) by significantly younger age (22 vs 38 years), higher anti-HBs level at index (48% vs 9% with anti-HBs >100 IU/L) and higher frequency of anti-HBs immune response (44% vs 20%). Of 15 vaccinated and 5 unvaccinated OBIs follow-up, 65% (8 vaccinated and 5 unvaccinated) became HBV DNA negative suggesting aborted recent infection, while 35% (7 vaccinated) had low persistent viraemia 2 to 65 months post index. In conclusion, isolated anti-HBs OBI in Chinese blood donors appears associated with young, vaccinated, adults exposed to HBV who predominantly develop low level aborted infection revealed by transient HBV DNA and immune anti-HBs response. However, a subset of individuals still experienced low but persistent viral replication whose clinical outcome remains uncertain.

Role of core protein mutations in the development of occult HBV infection.

BACKGROUND & AIMS: Occult HBV infection (OBI) is associated with transfusion-transmitted HBV infection and hepatocellular carcinoma. Studies on OBI genesis have concentrated on mutations in the S region and the regulatory elements. Herein, we aimed to determine the role of mutations in the core region on OBIs. METHODS: An OBI strain (SZA) carrying 9 amino acid (aa) substitutions in the core protein/capsid (Cp) was selected by sequence alignment and Western blot analysis from 26 genotype B OBI samples to extensively explore the impact of Cp mutations on viral antigen production in vitro and in vivo. RESULTS: A large panel of 30 Cp replicons were generated by a replication-competent pHBV1.3 carrying SZA or wild-type (WT) Cp in a 1.3-fold over-length of HBV genome, in which the various Cp mutants were individually introduced by repairing site mutations of SZA-Cp or creating site mutations of WT-Cp by site-directed mutagenesis. The expression of HBcAg, HBeAg, and HBsAg and viral RNA was quantified from individual SZA and WT Cp mutant replicons in transfected Huh7 cells or infected mice, respectively. An analysis of the effect of Cp mutants on intracellular or extracellular viral protein production indicated that the W62R mutation in Cp had a critical impact on the reduction of HBcAg and HBeAg production during HBV replication, whereas P50H and/or S74G mutations played a limited role in influencing viral protein production invivo. CONCLUSIONS: W62R and its combination mutations in HBV Cp might massively affect HBcAg and HBeAg production during viral replication, which, in turn, might contribute to the occurrence of OBI. LAY SUMMARY: Occult hepatitis B virus infections (OBIs) have been found to be associated with amino acid mutations in the S region of the HBV, but the role of mutations in the core protein (Cp) remains unclear. In this study, an OBI strain (SZA) carrying 9 amino acid substitutions in Cp has been examined comprehensively in vitro and in vivo. The W62R mutation in Cp majorly reduces HBcAg and HBeAg production during HBV replication, potentially contributing to the occurrence of OBI.

Follow-up program for blood donors with unconfirmed screening results reveals a high false-positive rate in Dalian, China.

BACKGROUND: Chinese blood donors with unconfirmed serological and/or molecular screening results are deferred permanently. This study investigated the implementation and performance of a follow-up program aiming to improve the notification and management of deferred donors in Dalian, China. STUDY DESIGN AND METHODS: From January 2013 to February 2018, 411,216 donations were tested for HBsAg, anti-HCV, anti-HIV/HIV antigen, and antibodies to Treponema pallidum. HBV, HCV, and HIV nucleic acid testing (NAT) was performed in mini-pools of six or in individual donations (IDs). Reactive donations were evaluated further with alternative serological assays and ID-NAT re-testing. A follow-up procedure was developed to evaluate a subset of deferred donors that were either potential NAT yield cases, serology non-reactive and NAT non-repeated reactive (NRR), or serology NRR irrespective of NAT result. RESULTS: Serological and molecular routine, plus supplemental testing, identified HBV, HCV, HIV, and TP in 503 (0.12%), 392 (0.09%), 156 (0.04%), and 2041 (0.49%) donations, respectively. Overall, 683 of 4156 (16.4%) eligible donors and 205 donors deferred prior 2013 participated in the program. They included 664 serology NRR and 224 NAT yield cases, and 58.8% repeat donors. All markers combined, follow-up documented false reactivity, primary acute infections, and OBI in 61.9% (550/888), 3.3% (29/888), and 12.8% (114/888) of these donors, respectively. Isolated anti-HBc or anti-HBs reactivity was observed in 22% of cases. CONCLUSION: Follow-up testing refined infection status in 78.0% (693/888) of deferred donors with unconfirmed screening results. This high false-positive rate encouraged to reevaluate the current screening strategies and to consider donor reentry.

High rate of hepatitis C virus and human immunodeficiency virus false-positive results in serologic screening in sub-Saharan Africa: adverse impact on the blood supply.

BACKGROUND: False positivity in blood screening may cause unnecessary deferral of healthy donors and exacerbate blood shortages. An international multicenter study was conducted to estimate the frequency of HCV and HIV false seropositivity in seven African countries (Burundi, Cameroon, Democratic Republic of Congo, Madagascar, Mali, Mauritania, and Niger). STUDY DESIGN AND METHODS: Blood donations were tested for hepatitis C virus (HCV) and human immunodeficiency virus (HIV) with rapid detection tests (RDTs), third-generation enzyme immunoassays (EIAs), or fourth-generation EIAs. HCV (456/16,613 [2.74%]) and HIV (249/16,675 [1.49%]) reactive samples were then confirmed with antigen/antibody assays, immunoblots, and nucleic acid testing. Partial viral sequences were analyzed when possible. RESULTS: The HCV reactivity rate with RDTs was significantly lower than with EIAs (0.55% vs. 3.52%; p < 0.0001). The HIV reactivity rate with RDTs was lower than with third-generation EIAs (1.02% vs. 2.38%; p < 0.0001) but similar to a fourth-generation assay (1.09%). Only 16.0% (57/357) and 21.5% (38/177) of HCV and HIV initial reactive samples, respectively, were repeatedly reactive. HCV and HIV infections were confirmed in 13.2% and 13.7%, respectively, of repeated reactive donations. The predominant HCV genotype 2 and 4 strains in West and Central Africa showed high genetic variability. HIV-1 subtype CRF02_AG was most prevalent. CONCLUSION: High rates (>80%) of unconfirmed anti-HCV and anti-HIV reactivity observed in several sub-Saharan countries highlights the need for better testing and confirmatory strategies for donors screening in Africa. Without confirmatory testing, HCV and HIV prevalence in African blood donors has probably been overestimated.

Metagenomic analysis of potential pathogens from blood donors in Guangzhou, China.

OBJECTIVES: This study aimed to identify the emerging/reemerging pathogens in blood donation samples. BACKGROUND: A metagenomic analysis has previously been used to look for pathogens but in this study, the relationship with aminotransferase (ALT) is described. METHODS/MATERIALS: Excluding samples reactive to hepatitis B virus, hepatitis C virus, human immunodeficiency syndrome virus or syphilis and plasma samples were stratified into three groups of ALT levels (IU/L): A ≤ 50, B 51 to 69 and C ≥ 70, respectively. Each group was mixed in a pool of 100 samples, from which DNA and cDNA libraries were established for next generation sequencing and analysis. Pathogens of interest were identified by immunoassays, nested-polymerase chain reaction, phylogenetic analysis and pathogen detection in follow-up donors. RESULTS: Several new or reemerging transfusion-transmitted pathogens were identified; Streptococcus suis, Babesia species and Toxoplasma gondii were found in the three ALT groups, Epstein-Barr virus (EBV) only in group C. Ten S. suis nucleic acid positive samples were detected, all closely phylogenetically related to reference strains. A donor in group A carried both S. suis genome and specific IgM in follow-up samples. This strain was identified as nontoxic S. suis. Five samples contained a short fragment of Babesia species SpeI-AvaI gene, while T. gondii was identified in 20 samples as a short fragment of 18S rDNA gene. In group C, two samples contained EBV genome. CONCLUSIONS: Blood donations that contained S. suis, Babesia species and T. gondii sequences might represent potential transfusion risks. EBV, a potential cause of elevated ALT, was detected. Metagenomic analysis might be a useful technology for monitoring blood safety.

Multiple HBV transfusion transmissions from undetected occult infections: revising the minimal infectious dose.

OBJECTIVE: HBV infection by blood components is currently prevented in most developed countries by combining sensitive HBV surface antigen (HBsAg) assays, nucleic acid testing (NAT) and in a few of them antibodies against the HBV core antigen (anti-HBc) screening. HBV transmissions by blood components from three repeat donors tested negative for HBsAg and HBV DNA with a highly sensitive screening test (limit of detection (LOD): 3.4 IU/mL) were investigated. DESIGN: 30 of the 47 recipients of components produced from these three donors were examined. Transfusion transmission was confirmed by phylogenetic analysis of viral sequences obtained from recipients and donors following viral particle concentration. RESULTS: 9 of 31 (29%) recipients were infected: 7 infections were related to 200 mL of fresh frozen plasma and 2 infections to red blood cells containing 20 mL plasma. Transfusion transmission was confirmed by >99% identity of donor/recipient sequences in five cases, probable in three and possible in one. HBV active infection remained unsuspected for 24-57 months in three recipients. Five non-infected recipients carried anti-HBs when transfused. Six patients transfused with platelet concentrates treated with a pathogen reduction method were not infected. These data enabled to revise previous estimate of the minimal infectious dose from approximately 100 to 16 copies (or 3 IU) of HBV DNA. CONCLUSIONS: HBV transfusion transmission from occult HBV infection carrying extremely low viral loads is related to plasma volume transfused and possibly prevented by anti-HBs. HBV blood safety could be further improved by either anti-HBc screening, HBV DNA NAT with a LOD of 0.8 copies/mL (0.15 IU/mL) or pathogen reduction of blood components.

Molecular characterization of hepatitis B virus in blood donors from Burkina Faso: Prevalence of quasi-subgenotype A3, genotype E, and mixed infections.

Burkina Faso is a highly endemic area for Hepatitis B virus (HBV) which remains a major challenge for blood safety with >13% of candidate blood donors being chronically infected. However, little is known about the molecular epidemiology of the viral strains currently circulating. In this study, 99 HBV strains from HBsAg positive candidate blood donors in Ougadougou were genetically characterized by sequencing the pre-S/S region of the viral genome. Phylogenetic analyses revealed a 25% prevalence of HBV quasi-subgenotype A3 (A3QS ) co-circulating with the confirmed dominant HBV genotype E (72%). HBV/A3QS sequences formed a sub-cluster closely related to West-African sequences previously characterized, and showed a low intra-group genetic diversity (0.75%) suggesting a relatively recent spreading of HBV/A3QS strains in Burkina Faso. Low genetic diversity of genotype E strains compared to A3QS was confirmed. Mixed infections with the two genotypes were identified in 3% of the donors tested and contributed to artifacts during PCR amplification of the viral genome leading to erroneous apparent intergenotype recombinant sequences. While the co-circulation of two HBV genotypes in a restricted area may favor the emergence of intergenotype recombinant variants, strictly controlled molecular experimental procedures should be used to accurately characterize HBV circulating recombinant forms. J. Med. Virol. 88:2145-2156, 2016. © 2016 Wiley Periodicals, Inc.

Poor efficacy of nucleic acid testing in identifying occult HBV infection and consequences for safety of blood supply in Italy.

BACKGROUND & AIMS: In Italy, DNA screening of blood donations for hepatitis B virus (HBV) was introduced to prevent the transmission of window period and occult HBV infection. Anti-HBc screening is not recommended in order to avoid shortage of the blood supply. To contain costs, donor samples are generally pooled before testing. We evaluated the safety of this national policy using a prospective repository of donors/recipient pairs. METHODS: We used highly sensitive nucleic acid testing (NAT) assays to test repository and follow-up samples from donors who were initially classified as negative by minipool NAT assays (6-MP), but were later found to carry occult HBV DNA. When available, we also analysed recipients' pre- and post-transfusion samples, collected in the context of a repository financed by the European Commission (the BOTIA project). RESULTS: Between 2008 and 2011 6-MP NAT assays identified 18 carriers of occult HBV infection among 12,695 donors; 28 samples from previous donations were available from 13 of these carriers. Highly sensitive HBV DNA detection methods showed that 6-MP HBV DNA screening failed to identify 14/28 (50%) viraemic donations, that were released for transfusion. HBV marker testing of such blood product recipients revealed two cases of transfusion transmitted HBV infection, documented by donor-recipient sequence identity. CONCLUSIONS: Viraemic blood donations from occult HBV infection carriers remain undetected by current minipool HBV DNA screening, and transfusion transmission of HBV continues to occur in susceptible patients. More effective individual HBV DNA screening and/or tests for antibodies to HBV core antigen should be considered to improve blood safety.

Nucleic acid testing to detect HBV infection in blood donors.

BACKGROUND: The detection of hepatitis B virus (HBV) in blood donors is achieved by screening for hepatitis B surface antigen (HBsAg) and for antibodies against hepatitis B core antigen (anti-HBc). However, donors who are positive for HBV DNA are currently not identified during the window period before seroconversion. The current use of nucleic acid testing for detection of the human immunodeficiency virus (HIV) and hepatitis C virus (HCV) RNA and HBV DNA in a single triplex assay may provide additional safety. METHODS: We performed nucleic acid testing on 3.7 million blood donations and further evaluated those that were HBV DNA-positive but negative for HBsAg and anti-HBc. We determined the serologic, biochemical, and molecular features of samples that were found to contain only HBV DNA and performed similar analyses of follow-up samples and samples from sexual partners of infected donors. Seronegative HIV and HCV-positive donors were also studied. RESULTS: We identified 9 donors who were positive for HBV DNA (1 in 410,540 donations), including 6 samples from donors who had received the HBV vaccine, in whom subclinical infection had developed and resolved. Of the HBV DNA-positive donors, 4 probably acquired HBV infection from a chronically infected sexual partner. Clinically significant liver injury developed in 2 unvaccinated donors. In 5 of the 6 vaccinated donors, a non-A genotype was identified as the dominant strain, whereas subgenotype A2 (represented in the HBV vaccine) was the dominant strain in unvaccinated donors. Of 75 reactive nucleic acid test results identified in seronegative blood donations, 26 (9 HBV, 15 HCV, and 2 HIV) were confirmed as positive. CONCLUSIONS: Triplex nucleic acid testing detected potentially infectious HBV, along with HIV and HCV, during the window period before seroconversion. HBV vaccination appeared to be protective, with a breakthrough subclinical infection occurring with non-A2 HBV subgenotypes and causing clinically inconsequential outcomes. (Funded by the American Red Cross and others.).

Specific amino acid substitutions in the S protein prevent its excretion in vitro and may contribute to occult hepatitis B virus infection.

Occult hepatitis B virus (HBV) infection (OBI) is defined as low plasma level of HBV DNA with undetectable HBV surface antigen (HBsAg) outside the preseroconversion window period. The mechanisms leading to OBI remain largely unknown. The potential role of specific amino acid substitutions in the S protein from OBI in HBsAg production and excretion was examined in vitro. HBsAg was quantified in culture supernatants and cell extracts of HuH-7 cells transiently transfected with plasmids containing the S gene of eight HBsAg(+) controls and 18 OBI clones. The intracellular (IC)/extracellular (EC) HBsAg production ratio was ∼1.0 for the majority of controls. Three IC/EC HBsAg patterns were observed in OBI strains clones: pattern 1, an IC/EC ratio of 1.0, was found in 5/18 OBI clones, pattern 2, detectable IC but low or undetectable EC HBsAg (IC/EC, 7.0 to 800), was found in 6/18 OBIs, and pattern 3, low or undetectable IC and EC HBsAg, was found in 7/18 clones. Intracellular immunofluorescence staining showed that in pattern 2, HBsAg was concentrated around the nucleus, suggesting retention in the endoplasmic reticulum. The substitution M75T, Y100S, or P178R was present in 4/6 pattern 2 OBI clones. Site-directed-mutagenesis-corrected mutations reversed HBsAg excretion to pattern 1 and, when introduced into a control clone, induced pattern 2 except for Y100S. In a control and several OBIs, variants of a given quasispecies expressed HBsAg according to different patterns. However, the P178R substitution present in all cloned sequences of two OBI strains may contribute significantly to the OBI phenotype.

Comparison of seven hepatitis B virus (HBV) nucleic acid testing assays in selected samples with discrepant HBV marker results from United States blood donors.

BACKGROUND: Sensitive triplex nucleic acid tests (NATs) are implemented for blood donation screening worldwide. Assays have variable ability to detect low-level hepatitis B virus (HBV) DNA. At borderline DNA detection levels, where Poisson distribution impacts results, distinguishing true-positive from false-positive results is challenging. Algorithms are needed to confirm such low-level HBV DNA-positive samples. STUDY DESIGN AND METHODS: A total of 135 blood donor samples reactive by one or more HBV markers that provided discrepant results were tested undiluted with four commercial NATs: Ultrio, Ultrio Plus, MPX, and a quantitative assay (SuperQuant). To further explore discrepancies, three additional in-house NATs including real-time polymerase chain reaction (PCR) and nested PCR and sequencing were performed. RESULTS: The numbers reactive of these 135 "difficult" samples by four commercial NATs were as follows: 39 of 107 (36%) with SuperQuant, 40 (30%) with Ultrio, 100 (74%) with Ultrio Plus, and 102 (76%) with MPX. Of the seven NATs, 109 (81%) samples were reactive by at least two assays and thus considered confirmed positive of which 67 (50%) generated a sequence. Ultrio Plus and MPX performed similarly as above (80%-85% detected of 109 and 81%-90% of 67, respectively). Older (median, 49 years), HBV core antibody-reactive donors carried predominantly Genotype A (58%) with high-frequency amino acid substitutions in the major hydrophilic region of the S-protein. Younger (median, 24 years) hepatitis B surface antigen-positive donors carried wild-type strains predominantly Genotype B (32%) and E (24%), the latter in an apparent cluster. CONCLUSIONS: Highly sensitive NATs require new confirmatory algorithms as presented optimally using different genomic regions or sequence generation. The introduction of immigration-related HBV genotypes may impact HBV epidemiology in the United States.

Head-to-head comparison of two transcription-mediated amplification assay versions for detection of hepatitis B virus, hepatitis C virus, and human immunodeficiency virus Type 1 in blood donors.

BACKGROUND: The second triplex transcription-mediated amplification (TMA) assay version (Ultrio Plus, Novartis Diagnostics) uses an additional reagent enhancing the disruption of hepatitis B virus (HBV) particles and release of DNA for the target capture probe. This study compares the performance of this new assay version with the previous one (Ultrio). STUDY DESIGN AND METHODS: For analytical sensitivity assessment the World Health Organization HBV, hepatitis C virus (HCV), and human immunodeficiency virus (HIV) international standards and various genotype dilution panels were used. Individual donations (IDs) from 9980 first-time donors were screened simultaneously by serology and both TMA assay versions. RESULTS: The 50 and 95% limits of detection (LODs) for HBV using Ultrio Plus were 0.8 (0.6-1.0) and 4.6 (3.2-7.2) IU/mL, respectively, 2.4 (1.4-4.8)-fold more sensitive than Ultrio. The TMA assay versions had comparable LODs for HIV-1 and HCV. The improvement factors on analytical sensitivity panels of HBV Genotypes A to G ranged from 1.3 to 7.3 and 50% LODs (95% confidence interval) reduced from 12.5 (10-15) to 3.8 (3.2-4.4) copies/mL. One Ultrio Plus HBV Genotype D yield sample missed by the Ultrio assay in the donor screening study was detected with ninefold higher sensitivity. The specificities of ID nucleic acid test (ID-NAT) and serologic testing in a similar repeat test algorithm were 100 and 99.41%, respectively. CONCLUSION: More efficient target capture chemistry in the new TMA assay version significantly improved sensitivity and diminished variability in detecting HBV strains of various genotypes. We recommend a triplicate ID-NAT repeat test strategy to eliminate discriminatory tests on false-non-repeat-reactive (anti-HBc-nonreactive) donations.

Infectivity of blood products from donors with occult hepatitis B virus infection.

BACKGROUND: Occult hepatitis B virus (HBV) infection (OBI) is identified in 1:1000 to 1:50,000 European blood donations. This study intended to determine the infectivity of blood products from OBI donors. STUDY DESIGN AND METHODS: Recipients of previous donations from OBI donors were investigated through lookback (systematic retrieval of recipients) or traceback (triggered by clinical cases). Serologic and genomic studies were undertaken on consenting donors and recipients. Multiple variables potentially affecting infectivity were examined. RESULTS: A total of 45 of 105 (42.9%) donor-recipients pairs carried antibodies to HBV core (anti-HBc) as evidence of previous HBV infection. Subtracting 15% of anti-HBc population background, the adjusted transmission rate was 28%. Anti-HBc prevalence increased to 28 of 44 (63.8%) in unvaccinated recipients receiving anti-HBs-negative OBI blood products. In contrast, four of 26 (15.4%) recipients of anti-HBs-positive products were anti-HBc positive. Transmission with anti-HBs-negative products depended on volume of plasma transfused (85%-100% with 200 mL of fresh frozen plasma [FFP], 51% with 50 mL in platelet concentrates [PCs], and 24% with 20 mL in red blood cells [RBCs], p < 0.0001 FFP vs. RBCs). The 50% minimum infectious dose of OBI HBV DNA was estimated at 1049 (117-3441) copies. Donor and recipient strains sequence homology of at least 99% confirmed transfusion-transmitted infection in 10 cases and excluded it in one case. CONCLUSION: Blood products from donors with OBI carry a high risk of HBV transmission by transfusion. This risk is dependent on presence of anti-HBs and viral dose. This may justify safety measures such as anti-HBc and HBV nucleic acid test screening depending on epidemiology.

Occult hepatitis B infection in blood donors from South East Asia: molecular characterisation and potential mechanisms of occurrence.

OBJECTIVE: To investigate the molecular basis of occult hepatitis B virus (HBV) infection (OBI) in Asian blood donors. DESIGN: OBI donors from Hong Kong, Malaysia, Singapore, Taiwan and Thailand were tested for HBV serological markers, and strains were molecularly characterised. RESULTS: Among 138 confirmed OBI carriers (median age 47 years), HBV genotypes B and C were dominant (60% and 34%, respectively) in agreement with the genotype distribution in chronically infected donors in the region. Viral load ranged between unquantifiable and 3670 IU/ml (median 11 IU/ml). Eleven per cent of OBIs showed an unusual anti-HBs-only serological profile without evidence of past vaccination for most of these individuals. Occult HBV strains showed a higher genetic diversity than strains from matched hepatitis B surface antigen (HBsAg)+ donors, irrespective of genotype. No unique genetic signature or evidence of reduced replication competence was found. Mutations in the vicinity of the pre-S2/S splice donor site were common in OBI(B) (44%) and OBI(C) (36%) strains. S regions from four OBI cases were transfected in HuH7 cells. Results showed limited HBsAg secretion and suggested that mutations disrupting the splice donor site structure may affect pre-S2/S mRNA splicing. CONCLUSIONS: There is indirect evidence that incomplete immune control is involved in the occurrence of OBI in Asian blood donors infected with genotypes B and C as observed in Europe with genotype A2 but to a lower extent than with genotype D. A post-transcriptional mechanism may play a role in HBsAg expression in some OBIs irrespective of HBV genotype.

No Hepatitis Delta Virus Seropositivity among Blood Donors with Overt and Occult Hepatitis B Infection in Dalian, Liaoning Province, China.

Hepatitis delta virus (HDV) is an obligate satellite of hepatitis B virus (HBV). Dual HDV/HBV infection is associated with down-regulated HBV replication and fast progression to severe liver disease. Although HDV is transmissible through exposure to infected blood, data about HDV infection in blood donors remain scarce. Between 2011 and 2021, 869,633 donations were collected from prequalified donors in Dalian, China. In total, 1060 (0.12%) were confirmed HBsAg and/or HBV DNA-reactive. Subsequently, anti-HDV IgG was tested in 2175 donations, including 65 that tested HBsAg+ pre donation, 1017 confirmed HBV-positive (507 HBsAg+/HBV DNA+, 33 HBsAg+/DNA-, 477 HBsAg-/DNA+ (451 occult (OBI) and 26 acute infections)), 327 viral DNA non-repeated-reactive, 397 anti-HBc-only, and 369 anti-HBs-only. Two (0.09%) samples tested anti-HDV IgG weakly reactive but were unconfirmed by IgM and IgG repeat testing with alternative assays, suggesting an initial false reactivity. In addition, HDV testing in a subgroup of confirmed OBI donors, comprising 451 donors from Dalian and 126 archived samples of OBI donors from around the world, showed only one non-Chinese donor to be repeatedly anti-HDV-reactive, suggesting that HDV/HBV coinfection does not play a significant role in the genesis of OBI. The overall data suggested an extremely low prevalence of HDV infection among blood donors in Liaoning province, Northeast China.