Optimal titer of anti-HBs in blood components derived from donors with anti-HBc.

BACKGROUND: The ultimate strategy to cope with transfusion-transmitted hepatitis B virus (HBV) infection caused by transfusion of blood from donors with historical HBV infection is to reject all donors having anti-HBV core antigen (anti-HBc). However, this strategy would result in a huge loss of blood donors and subsequent blood inventory collapse. On the other hand, anti-HBc-positive blood is reportedly not infectious when containing more than 100 mIU/mL of anti-HBV surface antigen (anti-HBs). STUDY DESIGN AND METHODS: In Japan, anti-HBc-positive blood has been used for transfusion if it contained 200 mIU/mL or more of anti-HBs. First, to verify the screening policy, clinical outcomes for transfusion of such blood were analyzed for the 2008-2012 period. Second, human hepatocyte-repopulated severe combined immunodeficiency mice were inoculated with HBV preincubated with varying doses of anti-HBs, then viremic status was followed. The effects of anti-HBs across different HBV genotypes were also investigated. RESULTS: Twenty-three transfusion-transmitted HBV infections related to anti-HBc-positive blood components were identified. None of the blood responsible for these cases contained 200 mIU/mL or more of anti-HBs. When 100 µL of plasma containing 104 copies of HBV and 20 mIU of anti-HBs was injected into severe combined immunodeficiency mice, no viremia was detected within 13 weeks. Genotype C anti-HBs was capable of total inhibition of genotype A HBV replication, whereas genotype A anti-HBs inhibited genotype C HBV to a lesser extent. CONCLUSION: Anti-HBc-positive blood containing 200 mIU/mL or more of anti-HBs appears safe as a transfusion component. HBV vaccination seems effective between HBV genotypes A and C.

Seroprevalence of Trypanosoma cruzi infection among at-risk blood donors in Japan.

BACKGROUND: Chagas disease is caused by Trypanosoma cruzi and is endemic in Latin America. In nonendemic countries, including Japan, Chagas disease is primarily a problem in the context of transfusion transmission. Approximately 250,000 immigrants from Latin America reside in Japan, and many of those individuals serve as active blood donors. This study surveyed the seroprevalence of T. cruzi infection among at-risk blood donors in Japan, defined as those who themselves (or whose mothers) were born (or raised) in Latin America, or those with a travel history to Latin America. STUDY DESIGN AND METHODS: Blood samples were obtained from at-risk donors in two periods, 2004-2012 and 2013-2016. Collected samples were tested for T. cruzi antibodies using both an enzyme-linked immunosorbent assay and a chemiluminescent immunoassay. Samples that tested positive in both assays were additionally tested by polymerase chain reaction, and look-back investigation was conducted when necessary. RESULTS: Of 18,484 samples obtained from 18,076 at-risk donors, 3 (1:6,025, 0.017%) donors showed seroreactivity by enzyme-linked immunosorbent assay and chemiluminescent immunoassay. All antibody-positive donors were born in Latin America. One of them also was positive for T. cruzi DNA. Eleven previous donations from this donor were subjected to look-back investigation, and five recipients were tested. All five recipients tested negative for T. cruzi antibodies. CONCLUSION: Seroprevalence of T. cruzi was 0.017% among at-risk donors in Japan. Transfusion-transmitted infection of Chagas disease has not been confirmed to date. Screening for T. cruzi antibodies by targeting at-risk donors is an appropriate strategy for ensuring blood safety in Japan.

Incidence rates of hepatitis C virus infection among blood donors in Japan: a nationwide retrospective cohort study.

BACKGROUND: The classification of many new cases of hepatitis virus infection as overt hepatitis does not reflect the true incidence of infection because the disease takes an asymptomatic course in some cases. In this retrospective cohort study, we aimed to estimate the incidence rates of new hepatitis C virus (HCV) infections among the blood donors. STUDY DESIGN AND METHOD: A 5-year retrospective cohort study was conducted to estimate the incidence rates by using the medical records of the blood donors between 2008 and 2013 for HCV infection. HCV seroconversions were investigated using a chemiluminescent enzyme immunoassay and then confirmed by nucleic acid amplification tests. RESULTS: The incidence rate of HCV infection was 0.40 per 100,000 person-years (95% confidence interval, 0.27-0.57) for HCV RNA seroconversion only and 7.32 per 100,000 person-years (95% confidence interval, 6.73-7.95) if either HCV RNA or anti-HCV seroconversion were taken into consideration. No significant difference of new HCV infections was found between the sexes. CONCLUSION: The incidence rate of HCV infection of this study was lower than that detected in a previous 1994-2004 study in which HCV incidence was 1.86 per 100,000 person-year, which reflects the presence of an effective blood screening system and health strategies targeting hepatitis control and prevention.

Analysis of HTLV-1 proviral load (PVL) and antibody detected with various kinds of tests in Japanese blood donors to understand the relationship between PVL and antibody level and to gain insights toward better antibody testing.

Adult T-cell leukemia/lymphoma (ATL) occurs in approximately 5% of individuals infected with human T-cell leukemia virus type 1 (HTLV-1). A high proviral load (PVL; more than four copies per 100 peripheral blood mononuclear cells (PBMCs) or 1.6 copies per 100 blood leukocytes) and being male are risk factors for ATL development. Whether anti-HTLV-1 antibody level is related to such risk is unknown. Here, PVL and antibody levels were examined using real-time PCR and other tests in 600 HTLV-1 positive screened Japanese blood donors to understand the relationship between PVL and antibody level in asymptomatic carriers and to gain insights toward better antibody testing for HTLV-1 infection. The 430 donors in whom proviral DNA was detected were considered as true positives for HTLV-1 infection. Among donors aged 40 years or older, more males than females had a PVL corresponding to more than 1.6% infected leukocytes, and an antibody titer below the median (P = 0.0018). In antibody tests using an HTLV-1 positive cell line or Env antigens there was a large discrepancy in antibody titer among 13 provirus-positive samples, probably suggesting that antibody-based screening tests should incorporate multiple HTLV-1 antigens, such as Gag and Env antigens.

Genetic Analysis of HIV-1 in Japan: a Comprehensive Analysis of Donated Blood.

In Japan, the number of human immunodeficiency virus (HIV)-1 infections remains relatively low; nevertheless, the annual incidence of HIV-1 infection has not decreased. New infections remain a great concern, and an improved understanding of epidemiological trends is critical for public health. The env C2V3 and pol sequences of HIV-1 RNA from 240 early (1996-2001) and 223 more recent (2010-2012) blood donations were used to compare the distribution of virus subtypes and to generate phylogenetic trees. Subtype B was clearly predominant in both early and more recent donations (both were 88.3%), and CRF01_AE was the second most common subtype. Phylogenetic analysis revealed a peculiar epidemiological transition. Compared to early subtype B isolates from 2 major endemic areas (Tokyo and Osaka), the more recent subtype B isolates formed fewer tight clusters in phylogenetic trees (from 8 to 2 clusters in Tokyo and 5 to zero clusters in Osaka). Furthermore, mixing of HIV-1 infections between these 2 endemic areas appear to increase. Analysis of phylogenetic trees suggested that local outbreaks have become smaller in Japan; however, intermixing of viral types between these 2 areas was more evident in the more recent samples.

Evaluation of the protective ability of plasma from Japanese individuals against mosquito-borne viral infections.

Background: Most of the Japanese population is seropositive for anti-Japanese encephalitis virus (JEV) antibodies because of previous JEV vaccination or natural infection. Because the virological characteristics of JEV are similar to those of West Nile virus (WNV) and dengue virus (DENV), we hypothesized that anti-JEV antibodies can cross-react with WNV and DENV antigens, leading to protection against infection by these viruses. Methods: Using isolated intravenous immunoglobulin (IVIG) from plasma collected in Japan, neutralizing activities against WNV and DENV and antibody-dependent enhancement (ADE) of these viral infections were evaluated using an in vitro assay to determine the potency of immunity against these viruses. Results: The prepared IVIG showed considerable neutralizing activity of 2.57 log10 reduction factor against WNV infection but showed little effect against DENV infection. A strong correlation was observed between the neutralizing activity of individual plasma samples against JEV and WNV (ρ=0.768). Moreover, IVIG showed no significant ADE of WNV infection. Conclusions: Based on these results, we presume that the Japanese population is generally protected from WNV infection. Furthermore, IVIG prepared from plasma donations from Japanese individuals is expected to be an effective therapeutic agent based on its neutralizing activity against JEV and WNV.

First report of the isolation of Lactococcus garvieae from a platelet concentrate in Japan.

BACKGROUND: Bacterial contamination of platelet concentrates (PCs) remains a serious problem in transfusion. We have been conducting sterility tests on all PCs rejected by blood centers or hospitals due to abnormal appearances. We recently experienced a case in which discrepant results were obtained between the methods used to identify a bacterial species isolated from a PC, requiring further analyses. STUDY DESIGN AND METHODS: Bacteria were isolated from a PC using the BacT/ALERT system and plate culture. The species was identified using biochemical tests and molecular analysis. Phylogenetic trees were constructed using sequences of the 16S ribosomal RNA (rRNA) and superoxide dismutase (sodA) genes from the bacterial isolate and related species. In addition, the isolate was cultured at temperatures of 10°C and below to determine its growth activity at low temperatures. RESULTS: Biochemical tests determined that the isolate was Streptococcus alactolyticus, whereas molecular analysis determined that it was Lactococcus garvieae. These two species belonged to different clusters on the phylogenetic tree. Similar to L. garvieae, the isolate could grow at 10°C. CONCLUSIONS: We conclude that the isolate was L. garvieae according to molecular identification and its growth characteristic at 10°C. Molecular analysis enabled the identification of this species, which was difficult to classify by biochemical tests. Blood facilities need to be prepared with multiple techniques, including genetic analysis techniques, for identifying contaminating bacterial species. L. garvieae can grow at 10°C and can contaminate both red blood cell concentrates and PCs; thus, this species should be listed as a cryophilic bacterium that could threaten blood safety.

Sequence analysis of two variable cytomegalovirus genes for distinction between transfusion- and breast milk-transmitted infections in a very-low-birthweight infant.

BACKGROUND: Cytomegalovirus (CMV) infections in very-low-birthweight infants can lead to serious clinical consequences. When CMV-related symptoms occur after transfusion, CMV transmission is often attributed to the transfusion products rather than to breast milk. However, it is sometimes difficult to distinguish between transfusion-transmitted and breast milk-transmitted CMV infections. PATIENT AND METHODS: A patient was born at 27 gestational weeks with a weight of 689 g. He was transfused with leukoreduced red blood cells (LR-RBCs), which were later found to be CMV seropositive and CMV DNA positive. He was also fed with CMV DNA-positive breast milk. Thereafter, he developed CMV disease with thrombocytopenia and jaundice. To determine the route of transmission, we analyzed the sequences of two variable CMV genes, UL139 and UL146, by direct sequence analysis. We also performed deep sequence analysis to determine whether there were polyclonal CMV strains in the LR-RBCs transfused. RESULTS: CMV DNA sequence-matching rates for the LR-RBCs and the patient's blood were 64.6% for the UL139 gene and 68.6% for the UL146 gene. In contrast, the sequences of these genes in the patient's blood were 100% matched with those in the breast milk. Furthermore, by deep sequence analysis, the CMV strain found in the patient's blood was not detected in the LR-RBCs transfused. CONCLUSION: The results indicate that the pathogenic CMV strain was transmitted through breast milk, which is consistent with the claims that transfusion-transmitted CMV infection due to leukoreduced blood products is uncommon.

Late onset post-transfusion hepatitis E developing during chemotherapy for acute promyelocytic leukemia.

We herein report the case of a leukemia patient who developed hepatitis E seven months after undergoing a transfusion with contaminated blood products. The latency period in this case was significantly longer than that of typical hepatitis E. Recently, chronic infection with hepatitis E virus (HEV) genotype 3 has been reported in immunocompromised patients. There is a possibility that our patient was unable to eliminate the virus due to immunosuppression following chemotherapy and the administration of steroids. The prevalence of HEV in healthy Japanese individuals is relatively high and constitutes a critical source of infection via transfusion. Hepatitis E is an important post-transfusion infection, and immunocompromised patients may exhibit a long latency period before developing the disease.

Establishment of culture systems for Genotypes 3 and 4 hepatitis E virus (HEV) obtained from human blood and application of HEV inactivation using a pathogen reduction technology system.

BACKGROUND: It has been demonstrated that the hepatitis E virus (HEV) can be transmitted via blood transfusion, and the risk of HEV transmission via transfusion has become a major global concern. An HEV culture system for blood-derived HEV has been sought to obtain valuable knowledge of the virus and the risk of HEV infection through blood products. STUDY DESIGN AND METHODS: We endeavored to establish an HEV culture system using RNA-positive blood specimens for Genotypes (G) 3 and 4 and applied this system to evaluate tissue culture infectious dose (TCID). We applied this method to investigate the potential of the Mirasol pathogen reduction technology (PRT) system (Terumo BCT) to inactivate live HEV in contaminated platelet samples (PLTs). PLTs were spiked with cultured HEV G3 or G4 and then treated with the Mirasol PRT system. PLTs were examined before and after the treatment for HEV load using TCID titration. RESULTS: We successfully established two strains for HEV production: the JRC-HE3 strain for G3 and the UA1 strain for G4. The Mirasol PRT system expressed more than 3 log inactivation for JRC-HE3 and more than 2 log inactivation for UA1. CONCLUSION: The Mirasol PRT system inactivated greater than 2 to 3 logs of live HEV in PLTs and can potentially be used to lower the possibility of blood-borne HEV transmission. The G3 and G4 HEV inocula identified in this study and the hepatoma cell culture system provide a new means to assess HEV infectious titer and to evaluate other pathogen reduction strategies.

Cytomegalovirus (CMV) seroprevalence in Japanese blood donors and high detection frequency of CMV DNA in elderly donors.

BACKGROUND: The current prevalence of cytomegalovirus (CMV) in Japan and the risk of CMV transfusion transmission are unknown in the era of seronegative leukoreduced blood components. STUDY DESIGN AND METHODS: We measured CMV-specific immunoglobulin (Ig)M and IgG in 2400 samples of whole blood collected from 12 groups of blood donors categorized by sex and age at 10-year intervals from their teens to their 60s. We also tested for CMV DNA using polymerase chain reaction in the cellular fractions of all samples. RESULTS: We found that 76.6% of blood donors were CMV seropositive. The seroprevalences among donors in their 20s and 30s were 58.3 and 73.3%, respectively. We detected CMV DNA in the cellular fraction of 4.3% of samples from donors in their 60s and in 1.0% of samples from donors younger than 60 years. None of the 562 seronegative samples was DNA positive. Furthermore, 14% of DNA-positive samples also contained DNA in the plasma fraction, and two of five such samples were derived from donors in their 60s. Leukoreduced plasma components derived from donations with CMV DNA in plasma samples also contained a relevant amount of CMV DNA. CONCLUSION: The seroprevalence of CMV among Japanese blood donors of child-bearing age has not changed over the past 15 years. Latent CMV becomes reactivated more frequently among elderly donors than among younger donors. A proportion of them have free CMV DNA in their plasma fraction, which could not be diminished by leukoreduction. The risk of transfusion-transmitted CMV infection in blood with plasma CMV DNA should be determined.

Residual risk of transfusion-transmitted hepatitis B virus (HBV) infection caused by blood components derived from donors with occult HBV infection in Japan.

BACKGROUND: Nucleic acid amplification testing (NAT) for hepatitis B virus (HBV) during blood screening has helped to prevent transfusion-transmitted HBV infection (TT-HBV) in Japan. Nevertheless, 4 to 13 TT-HBV infections arise annually. STUDY DESIGN AND METHODS: The Japanese Red Cross (JRC) analyzed repository samples of donated blood for TT-HBV that was suspected through hemovigilance. Blood donations implicated in TT-HBV infections were categorized as either window period (WP) or occult HBV infection (OBI) related. In addition, we analyzed blood from 4742 donors with low antibody to hepatitis B core antigen (anti-HBc) and antibody to hepatitis B surface antigen (anti-HBs) titers using individual-donation NAT (ID-NAT) to investigate the relationship between anti-HBc titer and proportion of viremic donors. RESULTS: Introduction of a more sensitive NAT method for screening minipools of 20 donations increased the OBI detection rate from 3.9 to 15.2 per million, while also the confirmed OBI transmission rate increased from 0.67 to 1.49 per million. By contrast the WP transmission rate decreased from 0.92 to 0.46 per million. Testing repository samples of donations missed by minipools of 20 donations NAT showed that 75 and 85% of TT-HBV that arose from WP and OBI donations, respectively, would have been interdicted by ID-NAT. The ID-NAT trial revealed that 1.94% of donations with low anti-HBc and anti-HBs titers were viremic and that anti-HBc titers and the frequency of viremia did not correlate. CONCLUSIONS: The JRC has elected to achieve maximal safety by discarding all units with low anti-HBc and anti-HBs titers that account for 1.3% of the total donations.

Xenotropic murine leukemia virus-related virus proviral DNA not detected in blood samples donated in Japan.

The xenotropic murine leukemia virus-related virus (XMRV) was first described as a novel human gammaretrovirus in prostate tumor tissues and was reported to be found in blood, suggesting the possibility of XMRV transmission via blood transfusion. The gag and env regions of the XMRV proviral DNA that were detected 1,030 blood samples collected from the greater Tokyo area were examined by real-time PCR analysis. However, XMRV infection was not found in the samples; this suggested that the risk of XMRV transmission via transfusion is very low in Japan.

Prevalence of amino acid mutation in hepatitis C virus core region among Japanese volunteer blood donors.

It is not known whether there is a trend of increasing or decreasing incidence of new hepatitis C virus (HCV) infections in Japan. From the treatment point of view, it is important to verify HCV genotypes and the prevalence of treatment-resistant clones of HCV. At the Japanese Red Cross blood centers, all blood samples obtained from blood donation have been screened using serological methods and the minipool nucleic acid amplification testing. One hundred and fourteen donors have been identified over the past 10 years to be HCV RNA-only positive without detectable anti-HCV and were considered to be in the acute phase of HCV infection. There was a trend of decreasing incidence of such new infections among the blood donors. HCV RNA-only-positive samples were examined further for genotyping and HCV RNA quantitation. Genotype 2 (2a plus 2b) was predominant (78.2%) among them followed by genotype 1b (21.2%). Direct sequencing was carried out to detect the possible treatment-resistant mutant clones 70Q and 91M, clones with amino acid substitutions at positions 70 and 91 of the HCV core protein, respectively. 70Q and 91M were found regularly in donors with genotype 1b, but not in those with other genotypes. No particular endemic areas for the mutant clones were identified. There was no significant difference in the mean viral titer between the 70Q mutant type and the non-70Q wild-type. Even in newly infected people, the mutant clone 70Q was detected frequently.

No viremia of pandemic (H1N1) 2009 was demonstrated in blood donors who had donated blood during the probable incubation period.

BACKGROUND: In the spring of 2009, the novel swine-origin influenza A (pandemic [H1N1] 2009) virus emerged and spread globally. Although no established cases of transfusion-transmitted influenza have been reported, the widespread outbreak of pandemic (H1N1) 2009 caused serious concern regarding the safety of blood products. The Japanese Red Cross Blood Centers have intercepted blood products with accompanying postdonation information indicating possible pandemic (H1N1) 2009 infection. To study the risk of transmission of pandemic (H1N1) 2009 by blood transfusion, we searched for the viral genome in such products using nucleic acid amplification technology. STUDY DESIGN AND METHODS: Between June and December 2009, blood components were collected from 579 blood donors who were diagnosed as or strongly suspected of having pandemic (H1N1) 2009 within 7 days after donation. Viral RNA was extracted from plasma and red blood cell (RBC) products, and RNA samples were subjected to real-time reverse transcription-polymerase chain reaction of the hemagglutinin and matrix genes of the pandemic (H1N1) 2009 virus. RESULTS: A total of 565 plasma and 413 RBC products from the 579 blood donors were available. No viral RNA of the pandemic (H1N1) 2009 was detected in any of the blood samples from the 579 blood donors. CONCLUSION: No viremia of pandemic (H1N1) 2009 was demonstrated in any of the 579 blood donors who had most likely donated blood during the incubation period. It is considered that the risk of transmitting pandemic (H1N1) 2009 by blood transfusion is extremely low.