The risk of sepsis through bacterial transmission is one of the most serious problems in platelet transfusion. In processing platelet concentrates (PCs), several methods have been put into practice ...to minimize the risk of bacterial transmission, such as stringent monitoring by cultivation assays and inactivation treatment by photoirradiation with or without chemical agents. As another potential option, we applied a light-emitting diode (LED) with a peak emission wavelength of 265 nm, which has been shown to be effective for water, to disinfect PCs. In a bench-scale UV-LED exposure setup, a 10-min irradiation, corresponding to an average fluence of 9.2 mJ/cm2, resulted in >2.0 log, 1.0 log, and 0.6 log inactivation (mean, n = 6) of Escherichia coli, Staphylococcus aureus, and Bacillus cereus, respectively, in non-diluted plasma PCs. After a 30-min exposure, platelet counts decreased slightly (18 ± 7%: mean ± SD, n = 7); however, platelet surface expressions of CD42b, CD61, CD62P, and PAC-1 binding did not change significantly (P>0.005), and agonist-induced aggregation and adhesion/aggregation under flow conditions were well maintained. Our findings indicated that the 265 nm UV-LED has high potential as a novel disinfection method to ensure the microbial safety of platelet transfusion.
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DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
During pilot studies to investigate the presence of viral RNA of xenotropic murine leukemia virus (MLV)-related virus (XMRV) infection in sera from chronic fatigue syndrome (CFS) patients in Japan, a ...positive band was frequently detected at the expected product size in negative control samples when detecting a partial gag region of XMRV using a one-step RT-PCR kit. We suspected that the kit itself might have been contaminated with small traces of endogenous MLV genome or XMRV and attempted to evaluate the quality of the kit in two independent laboratories. We purchased four one-step RT-PCR kits from Invitrogen, TaKaRa, Promega and QIAGEN in Japan. To amplify the partial gag gene of XMRV or other MLV-related viruses, primer sets (419F and 1154R, and GAG-I-F and GAG-I-R) which have been widely used in XMRV studies were employed. The nucleotide sequences of the amplicons were determined and compared with deposited sequences of a polytropic endogenous MLV (PmERV), XMRV and endogenous MLV-related viruses derived from CFS patients. We found that the enzyme mixtures of the one-step RT-PCR kit from Invitrogen were contaminated with RNA derived from PmERV. The nucleotide sequence of a partial gag region of the contaminant amplified by RT-PCR was nearly identical (99.4% identity) to a PmERV on chromosome 7 and highly similar (96.9 to 97.6%) to recently identified MLV-like viruses derived from CFS patients. We also determined the nucleotide sequence of a partial env region of the contaminant and found that it was almost identical (99.6%) to the PmERV. In the investigation of XMRV infection in patients of CFS and prostate cancer, researchers should prudently evaluate the test kits for the presence of endogenous MLV as well as XMRV genomes prior to PCR and RT-PCR tests.
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Background
Hepatitis B virus (HBV)‐positive individuals with isolated anti‐HBs are found among HBV vaccine recipients and healthy blood donors with no vaccination history. HBV infectivity from blood ...transfusions derived from such individuals remains unclear.
Case Presentation
A male patient who received transfusion with blood negative for individual donation‐NAT, HBsAg and anti‐HBc but weakly positive for anti‐HBs developed typical transfusion‐transmitted (TT)‐HBV with anti‐HBc response. The responsible blood donor was a frequent repeat donor showing a marked increase in anti‐HBs titer without anti‐HBc response 84 days after index donation. Test results for his past donations showed transient viremia with very low viral load and fluctuating low‐level anti‐HBs. The HBV vaccination history of this donor was unknown.
Discussion
Anti‐HBs and anti‐HBc kinetics of the donor suggest a second antibody response to new HBV challenge, representing a vaccine breakthrough case. On the other hand, transient low‐level viremia and fluctuating anti‐HBs in the test results of past donations suggested chronic occult HBV infection with isolated anti‐HBs.
Conclusion
Whatever the basic infection state, blood donors with isolated weak anti‐HBs may include a small population with a risk of causing TT‐HBV. Identifying individuals harboring such TT‐HBV risk among individuals positive only for anti‐HBs is difficult under current screening strategies. Active surveillance for the occurrence of TT‐HBV with blood positive only for anti‐HBs is necessary.
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SBCE, SBMB, UL, UM, UPUK
Background
Bacterial contamination in platelet concentrates (PCs) is a major problem in transfusion medicine. Contamination with Staphylococcus aureus is occasionally missed, even with cultural ...screening.
Study design and methods
Donors implicated in S. aureus‐contaminated PC were followed up. Skin and nasal swab specimens from six donors and S. aureus isolated from PCs related to these donors were subjected to multilocus sequence typing (MLST) and pulsed‐field gel electrophoresis (PFGE) to determine the identity of bacteria. To evaluate the validity of the screening method using BacT/ALERT 3D, we spiked S. aureus and three other bacterial species as comparisons into PCs and investigated their growth pattern.
Results
S. aureus was isolated from all nasal specimens and from the arm skin specimens of three donors with atopic dermatitis. In all cases, the S. aureus strains isolated from the PC and those from the nasal and skin specimens of the same donor showed concordant results using MLST and PFGE. In the spiking study, S. aureus showed irregular detectability over 24 to 48 h post‐spike periods, whereas the three other bacterial species were detected in all culture bottles after a 24‐h post‐spike period.
Discussion
The strain identity of S. aureus between donor and PC suggests that the contaminants were derived from those colonized in the donor. Furthermore, S. aureus yielded false‐negative results using BacT/ALERT 3D.
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BACKGROUND
Platelet concentrates (PCs) are the most common blood components eliciting nonhemolytic transfusion reactions (NHTRs), such as allergic transfusion reactions and febrile reactions. ...However, the precise mechanisms of NHTRs in PC transfusion remain largely unknown. Previous studies reported that mitochondria‐derived damage‐associated molecular patterns (DAMPs) could be important mediators of innate cell inflammation. Platelets (PLTs) represent a major reservoir of mitochondria in the blood circulation. The aim of this study was to determine the possible involvement of mitochondrial DAMPs in NHTRs.
STUDY DESIGN AND METHODS
The amount of mitochondrial DAMPs was determined as an index of total copy numbers of mitochondrial DNA (mtDNA), including mtDNA itself and free mitochondria, using quantitative real‐time polymerase chain reaction. To examine whether neutrophils, monocytes, and basophils were activated by mitochondrial DAMPs in vitro, an in vitro whole blood cell culture assay was performed.
RESULTS
In blood components associated with NHTRs, the mean total mtDNA concentration was highest in PCs followed in order by fresh‐frozen plasma and red blood cells. The amount of mtDNA in NHTR PCs was higher than that in control PCs without NHTRs. The mitochondrial DAMPs present in NHTR PCs was high enough to activate neutrophils, monocytes, and basophils, when costimulated with N‐formyl‐l‐methionyl‐l‐leucyl‐l‐phenylalanine or HLA antibodies.
CONCLUSION
PLT‐derived mitochondrial DAMPs are candidate risk factors for the onset of NHTRs.
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SBCE, SBMB, UL, UM, UPUK
Background
Transfusion‐transmitted bacterial infections (TTBIs) in Japan have been largely prevented due to a short shelf life of 3.5 days after blood collection for platelet concentrate (PC) and ...washed PCs (WPCs; PC in which 95% plasma is replaced by platelet additive solution).
Case Presentation
Case 1: In January 2018, a woman in her 50s with aplastic anaemia who received WPC transfusion and developed a fever the next day and Streptococcus dysgalactiae subspecies equisimilis (SDSE) was detected in the residual WPC. Case 2: In May 2018, a man in his 60s with a haematologic malignancy who received PC transfusion and developed chills during the transfusion. SDSE was detected in the patient's blood and residual PC.
The contaminated platelet products were both manufactured from blood donated by the same donor. The multi‐locus sequencing typing revealed that SDSE detected in case 1 was identical to that from case 2; however, whole blood subsequently obtained from the donor was culture negative.
Conclusion
WPC and PC produced from two blood donated 106 days apart by the same donor were contaminated with SDSE of the same strain and both caused TTBIs. Safety measures should be considered regarding blood collection from a donor with a history of bacterial contamination.
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Abstract
Tetanus is a fatal disease caused by tetanus neurotoxin (TeNT). TeNT is composed of a light chain (Lc) and a heavy chain, the latter of which is classified into two domains, N-terminus Hn ...and C-terminus Hc. Several TeNT-neutralizing antibodies have been reported, but it remains unclear which TeNT domains are involved in neutralization. To further understand the mechanism of these antibodies, we isolated TeNT-reactive human antibody clones from peripheral blood mononuclear cells. We then analyzed the reactivity of the isolated antibody clones to each protein domain and their inhibition of Hc-ganglioside GT1b binding, which is critical for TeNT toxicity. We also investigated the TeNT-neutralizing ability of isolated antibody clones and showed that an Hn-reactive clone protected strongly against TeNT toxicity in mice. Furthermore, combination treatment of Hn-reactive antibody clones with both Hc-reactive and TeNT mix (the mixture of Hc, Hn, and Lc proteins)–reactive antibody clones enhanced the neutralizing effect. These results indicated that antibody clones targeting Hn effectively neutralized TeNT. In addition, the use of a cocktail composed of Hc-, Hn-, and TeNT mix–reactive antibodies provided enhanced protection compared to the use of each antibody alone.
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IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
Background
In Japan, plasma with a high concentration of Hepatitis B Virus (HBV) antibodies for hepatitis B immunoglobulin (HBIG) is almost entirely imported. We aimed to produce recombinant HBIG by ...isolating immunoglobulin cDNAs against the HBV surface antigen (HBsAg).
Study Design and Methods
B cells expressing HBsAg antibodies were obtained from blood center personnel who had been administered HB vaccine booster and then isolated by either an Epstein–Barr virus hybridoma or an antigen‐specific memory B cell sorting method. Each cDNA of the heavy and light chains of the target antibody was cloned into an IgG1 expression vector and transfected into Expi293F cells to produce a recombinant monoclonal antibody (mAb), which was screened by ELISA and in vitro HBV neutralizing assays. The cross‐reactivity of the mAbs to normal human molecules was evaluated by ELISA and immunohistochemistry.
Results
Antibody cDNAs were cloned from 11 hybridoma cell lines and 204 HBsAg‐bound memory B cells. Three of the resulting recombinant mAbs showed stronger neutralizing activity in vitro than the currently used HBIG. All three bind to the conformational epitope(s) of HBsAg but not to human DNA or cells.
Discussion
We successfully isolated HBV‐neutralizing monoclonal antibodies from B cells collected from healthy plasma donors boosted against the HBV. To obtain an alternative source for HBIG, HBV‐neutralizing monoclonal antibodies from B cells collected from healthy plasma donors boosted against the HBV may be useful.
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BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SBCE, SBMB, UL, UM, UPUK
Background
The severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) essentially affects respiratory organs and tissues. SARS‐CoV‐2 RNAemia is often associated with more severe cases of ...coronavirus disease 2019 (COVID‐19) compared to cases without RNAemia. To determine the impact of the pandemic on transfusion medicine, particularly transfusion‐related infection, we examined the frequency of blood donation with RNAemia, the viral RNA (vRNA) concentration, and any possibility of transfusion‐transmitted infection (TTI) among transfusion recipients.
Study Design and Methods
vRNA was examined in plasma/serum samples from 496 of 513 blood donors who reported having been infected with SARS‐CoV‐2 within 2 weeks of donation among a total of ca. 9.9 million blood donations in Japan between January 15, 2020, and December 31, 2021. The clinical course of patients transfused with the blood component containing vRNA was also examined.
Results
vRNA was detected in 23 of 496 samples. The median period from blood donation to COVID‐19 onset was 1 day in 16 RNAemia‐positive donors. Most samples had vRNA concentrations below the limit of quantification. Three patients were transfused with either a packed red blood cell or platelet concentrate that tested positive for vRNA, showing no COVID‐19 symptoms and testing negative for vRNA in post‐transfusion blood.
Conclusion
The rate of RNAemia was 4.6% among blood donors who were found to be infected with SARS‐CoV‐2 shortly after donation, and vRNA concentrations in their donated blood were extremely low. There was no evidence of TTI in the recipients transfused with RNAemia‐positive blood components. TTI risk in SARS‐CoV‐2 is negligible.
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The potential risk and association of bovine leukemia virus (BLV) with human remains controversial as it has been reported to be both positive and negative in human breast cancer and blood samples. ...Therefore, establishing the presence of BLV in comprehensive human clinical samples in different geographical locations is essential.
In this study, we examined the presence of BLV proviral DNA in human blood and breast cancer tissue specimens from Japan. PCR analysis of BLV provirus in 97 Japanese human blood samples and 23 breast cancer tissues showed negative result for all samples tested using long-fragment PCR and highly-sensitive short-fragment PCR amplification. No IgG and IgM antibodies were detected in any of the 97 human serum samples using BLV gp51 and p24 indirect ELISA test. Western blot analysis also showed negative result for IgG and IgM antibodies in all tested human serum samples.
Our results indicate that Japanese human specimens including 97 human blood, 23 breast cancer tissues, and 97 serum samples were negative for BLV.
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