Two Cases of Anti-D Alloimmunization in D-Negative Thai Patients as a Result of the Asian-Type DEL on Transfused Red Cells.

Introduction: DEL is known to be one of the weakest D variants, which can be detected by the adsorption-elution technique or by molecular study. Currently, in Thailand, we do not routinely test for DEL variants serologically or genetically among serologic RhD-negative blood donors. Case Presentation: We reported 2 cases of alloimmunization after transfused with Rh DEL, RHD*DEL1 allele, in the Thai population. The first case was a 73-year-old male with anemia who presented with post-cardiac arrest and septic shock. The patient was group B, RhD-negative, and was transfused with RhD-negative red blood cells (RBCs). Antibody screening and identification found that the patient developed anti-D and anti-Mia during the admission course. The second case was a 38-year-old woman with pseudomyxoma peritonei who developed anti-D after receiving four units of RhD-negative RBCs during cytoreductive surgery with hyperthermic intraperitoneal chemotherapy. Both patients did not receive anti-D immunoglobulin and had no previous history of anti-D detection. We retrospectively investigated and found two units of RHD*DEL1 among the RBCs transfused to these patients. Discussion: Previous reports of several cases of anti-D alloimmunization in RhD-negative recipients transfused by RHD*DEL1, an Asian-type DEL, are limited only to East Asia. We first identified 2 patients with anti-D alloimmunization after receiving the RHD*DEL1 RBCs in the Thai population. This raises concern about Rh DEL screening among D-negative Thai blood donors and whether to remove DEL units from the D-negative inventory to improve patient safety.

How do we translate gaps and unmet needs of blood management for thalassemia into a collaborative implementation framework?

BACKGROUND AND OBJECTIVE: The mainstay of management for thalassemia is regular blood transfusions. However, gaps and unmet needs of blood services for thalassemia are still not clearly identified and addressed in Thailand, a country prevalent with thalassemia. What can be a collaborative implementation framework that helps advance practices and policies relating to blood management for thalassemia? METHODS: The first Blood & Beyond Roundtable Discussion was held in July 2022 to gather the current situation, gaps, and unmet needs of blood services for thalassemia from multidisciplinary experts and thalassemic patients. The Implementation Guide as suggested by the Centre for Effective Services was applied as a tool to consolidate information from the discussions and construct the collaborative implementation framework. RESULTS: The National Blood Center and hospitals in Thailand followed the missions specified in the National Blood Policy and the standard guidelines to ensure the best practice of blood management for thalassemia. However, there were six gaps and unmet needs identified from the discussions. After all discussion points were mapped onto the framework, an implementation plan comprised of five specific activities became clear and actionable. CONCLUSION: Without the complete information from both experts and patients, the implementation plan would not have been successfully constructed. The method that we employed to translate all information into the framework can be adapted by other countries to develop their own specific framework efficiently.

Erythrocyte Alloimmunity and Genetic Variance: Results from the Collaborative Study of Alloimmunity to Antigen Diversity in Asian Populations (All ADP).

As an East-Asian international study, we evaluated erythrocyte alloimmunity by gender and history of transfusion or pregnancy. In total, data from more than 1,826,000 patients were analyzed, from whom 26,170 irregular erythrocyte antibodies were detected in 22,653 cases. Antibody frequencies in these cases were as follows: anti-E, 26.8%; anti-Lea, 20.0%; anti-P1, 7.1%; anti-M, 6.4%; anti-Mia, 5.6%; anti-c + E, 5.6%; anti-Leb, 4.6%; anti-D, 2.8%; anti-Fyb, 2.6%; anti-Lea+Leb, 2.5%; anti-Dia, 2.0%; and others. For pregnant patients, anti-D (12.7%) was statistically more frequent. For transfused patients, anti-E (37.3%), anti-c + E (9.5%), anti-C + e (3.3%) and anti-Jka (3.1%) were significantly more frequent.

Differences in levels of platelet-derived microparticles in platelet components prepared using the platelet rich plasma, buffy coat, and apheresis procedures.

BACKGROUND: There has been an increased interest in platelet-derived microparticles (PMPs) in transfusion medicine. Little is known about PMP status during the preparation of platelet concentrates for transfusion. AIM: The aim of this study is to compare the PMP levels in platelet components prepared using the buffy coat (BC), platelet-rich plasma platelet concentrate (PRP-PC), and apheresis (AP) processes. METHODS: Platelet components were prepared using the PRP-PC and BC processes. Apheresis platelets were prepared using the Trima Accel and Amicus instruments. The samples were incubated with annexin A5-FITC, CD41-PE, and CD62P-APC. At day 1 after processing, the PMPs and activated platelets were determined using flow cytometry. RESULTS: Both the percentage and number of PMPs were higher in platelet components prepared using the Amicus instrument (2.6±1.8, 32802±19036 particles/µL) than in platelet components prepared using the Trima Accel instrument (0.5±0.4, 7568±5298 particles/µL), BC (1.2±0.6, 12,920±6426 particles/µL), and PRP-PC (0.9±0.6, 10731±5514 particles/µL). Both the percentage and number of activated platelets were higher in platelet components prepared using the Amicus instrument (33.2±13.9, 427553±196965 cells/µL) than in platelet components prepared using the Trima Accel instrument (16.2±6.1, 211209±87706 cells/µL), BC (12.9±3.2, 140624±41003 cells/µL), and PRP-PC (21.1±6.3, 265210±86257 cells/µL). CONCLUSIONS: The study suggests high variability of PMPs and activated platelets in platelet components prepared using different processes. This result may be important in validating the instruments involved in platelet blood collection and processing.

A Prospective Randomized Controlled Trial Comparing Posterolateral Lumbar Fusion With and Without Bone Marrow Concentrate Augmentation in Single-Level Lumbar Spondylolisthesis.

Background: Bone marrow (BM), which is a good source of stem cells and biological factors, has the potential to enhance bone fusion. Simple centrifugation technique is one of the procedures used to concentrate BM aspirate for increasing number of cells. However, there are limited clinical study for using BM concentrate augmentation in spinal fusion. Objective: This study was designed to examine the spinal fusion enhancement effects of bone marrow (BM) concentrate augmentation on poster lateral lumbar fusion (PLF) with autologous local bone graft in terms of both quality and quantity, as compared with a control procedure without BM concentrate augmentation. Material and Method: Twelve patients with L4-L5 spondylolisthesis scheduled for PLF after decompressive laminectomy and pedicle screw instrumentation were included in this study. This prospective randomized controlled trial was conducted at Siriraj Hospital during the 2009 to 2012 study period. Patients were randomly assigned to two groups. One group underwent PLF with local bone graft with BM concentrate augmentation (BM group) and the other group underwent PLF with local bone graft only (non-BM group). Clinical outcomes were evaluated by the Oswestry Disability Index (ODI) preoperatively and at 3 and 6 months after PLF. Bone fusion quality was evaluated by bony bridging on 3D-CT imaging. Fusion mass volumes were measured on quantitative 3D-CT scans at 1 week and 6 months, postoperatively. Results: Clinical outcome scores did not differ between groups. Six-month postoperative 3D-CT imaging showed complete PLF bridging in 58.3% and 100% of patients in the BM and non-BM groups, respectively. PLF mass volumes were decreased at 6 months by 51.1% in the BM group and by 48.5% in the non-BM group. One patient in the BM group had local inflammation at the BM aspiration site. Conclusion: Bone marrow concentrate augmentation in this small randomized controlled trial failed to demonstrate positive effects on autologous local bone graft in posterolateral lumbar fusion relative to both quality and quantity. The high percentage of incomplete bridging should also be noted and further investigated.

Anti-SARS-CoV-2 antibody among SARS-CoV-2 vaccinated vs post-infected blood donors in a tertiary hospital, Bangkok, Thailand.

SARS-CoV-2 virus infection has imposed a significant healthcare burden globally. To contain its spread and decrease infection-related mortality, several vaccines have been deployed worldwide in the past 3 years. We conducted a cross-sectional seroprevalence study to assess the immune response against the virus among blood donors at a tertiary care hospital, Bangkok, Thailand. From December 2021 to March 2022, total of 1,520 participants were enrolled, and their past history of SARS-CoV-2 infection and vaccination was recorded. Two serology test, namely, quantitative IgG spike protein (IgGSP) and qualitative IgG nucleocapsid antibody (IgGNC) were performed. The median age of study participants was 40 years (IQR 30-48) and 833 (54.8%) were men. Vaccine uptake was reported in 1,500 donors (98.7%) and 84 (5.5%) reported the past infection history. IgGNC was detected in 46/84 donors with the past infection history (54.8%) and in 36 out of the rest 1,436 (2.5%) with no past history. IgGSP positivity was observed in 1484 donors (97.6%). When compared to unvaccinated donors (n = 20), IgGSP level was higher in the donors who had received one vaccine dose (p< 0.001) and these antibody levels increased significantly among those with 3rd and 4th vaccine doses. Factors associated with low IgGSP (lowest quartile) by multivariate analysis included: no past infection history, homologous vaccination, < 3 vaccine doses, and > 90 days duration since last vaccination. In conclusion, vaccine uptake among our study donors was high (98.7%) and IgGSP antibody was observed in nearly all the vaccinated donors (97.6%). Previous SARS-CoV-2 infection, use of heterologous vaccination, vaccines ≥ 3 doses, and duration of the last vaccination >90 days affected IgGSP levels. Use of serological assays were found beneficial in the evaluation and differentiation of immune response to vaccination, and natural infection including the identification of previous asymptomatic infections.

Prevalence of G6PD deficiency in Thai blood donors, the characteristics of G6PD deficient blood, and the efficacy of fluorescent spot test to screen for G6PD deficiency in a hospital blood bank setting.

BACKGROUND: Transfusion of blood from glucose-6-phosphate dehydrogenase (G6PD) enzyme deficient donors could cause a potentially unfavorable outcome, especially in newborns and those with hemoglobinopathies. AIMS: To determine the prevalence of G6PD deficiency in Thai blood donors, the characteristics of G6PD deficient blood, and the efficacy of fluorescent spot test (FST) to screen for G6PD deficiency in a hospital blood bank setting. METHODS: Blood samples were obtained from 514 Thai blood donors who donated blood at Siriraj Hospital (Bangkok, Thailand) during December 2020-February 2021. G6PD deficiency status was screened using FST, and in vitro hemolysis of red blood cell parameters of G6PD deficient blood units was compared with those of normal control units at different time points during 35 days of refrigerated storage. RESULTS: The prevalence of G6PD deficiency was 7.59% (35 [8.73%] males, 4 [3.54%] females). The sensitivity of FST was 100% (95% confidence interval [CI]: 90.97-100%), and the specificity was 99.58% (95%CI: 98.49-99.95%). In vitro hemolysis was not significantly different between G6PD deficiency and normal controls. CONCLUSION: The prevalence of G6PD deficiency in this study was 7.59%. FST was demonstrated to be an effective and reliable method for G6PD deficiency screening among Thai blood donors in a hospital blood bank setting.

Mutation of ABO gene in Thai blood donors with A3 phenotype.

The ABO system is the most important of all blood group systems in transfusion practice. The subgroup gives a weak reaction when treated with anti-A or anti-B. The most common subgroup found in Thai blood donors is subgroup A3, which is characterized by mixed-field agglutination when reacted with anti-A and anti-A,B, was caused by mutation in the ABO gene, especially in the exon 7. In the present study mutation in A3 were charactered in exon 7 of the ABO gene in 10 A3 phenotype Thai blood donors from the National Blood Centre, Thai Red Cross Society by PCR amplification and DNA sequencing. Mutations in exon 7 were identified in the A allele of six cases. In four cases, mutations were detected at positions 646T > A, 681G > A, 771C > T and 829G > A. One case showed a double mutations at positions 467C > Tand 745C > T and one case showed a mutation at position 467C > T. Four cases showed wild type exon 7 as A101 allele. These mutations were previously reported in BGMUT database and no novel mutation was identified These data suggest genetic heterogeneity in A3phenotype in Thai blood donors.

Younger age of onset and uveitis associated with HLA-B27 and delayed diagnosis in Thai patients with axial spondyloarthritis.

To identify characteristics associated with HLA-B27, and to identify factors associated with delayed diagnosis in Thai patients with axial spondyloarthritis (axSpA). This cross-sectional study included Thai patients were diagnosed with axSpA by a rheumatologist at Siriraj Hospital. Clinical data were collected. Regression analyses were employed to identify factors associated with study outcomes. Of total 177 patients, 127 (72%) were positive HLA-B27. Uveitis [Odds ratio (OR) 4.0], age at onset of the first musculoskeletal symptom of ≤ 28 years [OR 3.5], female [OR 0.4], and psoriasis [OR 0.4] were significantly associated with HLA-B27 in multiple regression analysis. Those with positive HLA-B27 had less spinal flexibility. Elevated C-reactive protein (p = 0.012) was associated with shorter delay in diagnosis, while uveitis (p < 0.001) and younger age at onset of the first symptom (p = 0.002) were associated with longer delay in diagnosis in multiple regression analysis. Younger age at onset of the first musculoskeletal symptom and uveitis were associated with HLA-B27 and delayed diagnosis in axSpA patients. Young people with musculoskeletal symptom and uveitis should be referred to a rheumatologist to rule out or make a timely diagnosis of axSpA.

HLA-B*27:04 associated with enthesitis and younger age of onset, and HLA-B allele profile in patients with ankylosing spondylitis in Thailand: A cross-sectional study.

AIM: The aims of this study were to estimate human leukocyte antigen (HLA)-B allele frequency, to identify alleles associated with ankylosing spondylitis (AS), and to explore manifestations in various HLA-B*27 in Thai AS patients. METHODS: This was a cross-sectional study. Thai patients older than 18 years with diagnosed AS according to modified New York criteria who visited Siriraj Hospital (Bangkok, Thailand) were consecutively enrolled. HLA-B alleles were determined by reverse sequence-specific oligonucleotide assays, and were assigned at a 4-digit resolution. HLA-B alleles of 334 unrelated healthy Thai donors who participated in a previous phase 2b dengue vaccine clinical trial were included as controls. Odds ratio (OR) and Fisher's exact test were used to estimate association between allele and AS. The P value significance threshold was calculated according to Bonferroni. RESULTS: Among the 88 patients who were recruited, 34 HLA-B alleles were identified, and all patients were heterozygous. The prevalence of HLA-B*27 was 89.8%, and 4 alleles of HLA-B*27 were identified. HLA-B*27:04 (OR: 39.4, P < .0001) and HLA-B*27:05 (OR: 13.8, P = .0011) were associated with AS. In contrast, HLA-B*27:06 was not found to be associated with AS (OR: 0.4, P = .241). AS patients carrying HLA-B*27:04 were more likely to have enthesitis and younger age at onset than those carrying HLA-B*27:05. CONCLUSIONS: HLA-B*27:04 and HLA-B*27:05 were both found to be strongly associated with Thai AS. HLA-B*27:06 showed a neutral allele for Thai AS. AS patients with HLA-B*27:04 had more enthesitis and younger age at onset than those with HLA-B*27:05.

Induction of TNF-alpha in human macrophages by avian and human influenza viruses.

The highly pathogenic avian influenza virus H5N1 is known to induce high level of tumor necrosis factor alpha (TNF-alpha) from primary macrophages. However, it is still unclear whether current H5N1 strains also induce high TNF-alpha production, as most of the data were derived from extinct clade 0 H5N1 strain. Here, we show that current clade 1 and 2 H5N1 strains induce variable levels of TNF-alpha that are not necessarily higher than those induced by seasonal influenza viruses. The result suggests that hyper-induction of TNF-alpha in human macrophages is not always associated with a highly pathogenic phenotype. We further tested the contribution of the NS gene segment from H5N1 isolates to TNF-alpha induction by using reverse genetics. While NS conferred some variation in TNF-alpha induction when incorporated into an H1N1 virus genetic background, it did not affect TNF-alpha induction in an H5N1 virus genetic background, suggesting that other viral genes are involved.

Web interface-supported transmission risk assessment and cost-effectiveness analysis of postdonation screening: a global model applied to Ghana, Thailand, and the Netherlands.

BACKGROUND: The goal of our research was to actively involve decision makers in the economic assessment of screening strategies in their region. This study attempted to accomplish this by providing an easy-to-use Web interface at http://www.bloodsafety.info that allows decision makers to adapt this model to local conditions. STUDY DESIGN AND METHODS: The cost-effectiveness was compared of 1) adding antigen screening to antibody screening for hepatitis C virus (HCV) and human immunodeficiency virus (HIV); 2) adding nucleic acid amplification testing (NAT) on hepatitis B virus (HBV), HCV, and HIV in minipool (pool of 6 [MP6] and 24 [MP24]) to antibody screening and hepatitis B surface antigen (HBsAg) screening; and 3) individual-donation NAT on HBV, HCV, and HIV to antibody screening and HBsAg screening for Ghana, Thailand, and the Netherlands. RESULTS: The combination of HCV antibody-antigen combination (combo) and HIV combo added to antibody screening in Ghana and Thailand was cost-effective according to the WHO criteria. MP24-NAT screening in Ghana was also cost-effective. MP24-NAT on HBV, HCV, and HIV was not cost-effective compared to the other screening strategies evaluated for the Netherlands. Large regional differences in cost-effectiveness were found for Thailand. CONCLUSION: The young transfusion recipient population of Ghana in combination with a high risk of viral transmission yields better cost-effectiveness for additional tests. The advanced age of the transfused population of the Netherlands and a small risk of viral transmission gives poor cost-effectiveness for more sensitive screening techniques. It was demonstrated that a global health economic model combined with a Web interface can provide easy access to risk assessment and cost-effectiveness analysis.

Risk factors of hepatitis C virus infection in blood donors in Thailand: a multicenter case-control study.

OBJECTIVE: To investigate the risk of hepatitis C virus (HCV) infection in healthy blood donors in Thailand MATERIAL AND METHOD: We performed a case-control study of 435 HCV-seropositive blood donors and 894 HCV-seronegative blood donors as controls. The study was done with direct interview regarding demographic characteristics and risk factors. Odds ratios (OR) with 95% confidence intervals (CIs) were calculated by using conditional logistic regression. RESULTS: The final multivariable model included only the following independent HCVrisk factors: intravenous drug user (IDU) (OR = 61.5; 95%CI, 26.6-142.5), previous blood or blood products transfusion (OR = 12.3; 95%CI, 7.6 -19.9), sharing of razors (OR = 2.3, 95%CI, 1.6-3.2),unsafe injection (OR = 3.3, 95%CI, 1.8-5.9), unused condom (OR = 1.6; 95%CI, 1.1, 2.4). No risk was shown for a history of tattoo, ear piercing, or acupuncture and multiple sexual partners. CONCLUSION: The risk factors for HCV infection in healthy blood donors in Thailand are IDU, past history of blood transfusion and unsafe injection.

HLA class II (DRB1, DQA1 and DQB1) allele and haplotype frequencies among HIV-infection discordant Thai couples.

We investigated the association of HLA-DRB1, -DQA1 and -DQB1 alleles and haplotypes in 33 Thai HIV discordant couples. A significantly lower frequencies of DRB1*14 (3.0% vs 11.3%, p = 0.048) and DQA1*0103 (0.0% vs 5.63%, p = 0.042) alleles were found in the seropositive individuals when compared with HIV-negative controls. In contrast, there was no significant difference in HLA-DQB1* allele frequencies. The haplotype analysis revealed that DRB1*1501-DQA1*0102-DQB1*0601 (7.6% vs 0.0%, p = 0.002), DRB1*0405-DQA1*0302-DQB1*0401 (7.6% vs 1.3%, p = 0.024) and DRB1*1401-DQA1*0104-DQB1*05031 (6.1% vs 0.0%, p = 0.007) were found to be significantly higher frequencies when compared between HIV seronegative partners and HIV negative controls, but DRB1*1501-DQA1*0102-DQB1*0502 (0.0% vs 8.1%, p = 0.01) was significantly lower. The DRB1*1602-DQA1*0101-DQB1*0502 (4.6% vs 0.0%, p = 0.024) haplotype was found to be significantly higher frequencies in HIV seropositive individuals when compared to HIV negative controls but the DRB1*1502-DQA1*0101-DQB1*0501 (1.5% vs 8.1%, p = 0.049) haplotype was lower.

Comprehensive investigation of common antibody-dependent cell-mediated cytotoxicity antibody epitopes of HIV-1 CRF01_AE gp120.

The antibody-dependent cell-mediated cytotoxicity (ADCC) mechanism involves both innate and adaptive immune systems. While a number of epitope mapping studies of neutralizing (Nt) antibodies and cytotoxic T lymphocyte (CTL) against a variety of HIV-1 clades have been reported, there has been a paucity of similar studies aimed at identifying epitopes of ADCC-inducing antibodies. Herein we screened 35 sera from HIV-1 CRF01_AE-infected blood donors for ADCC antibody activity against gp120 utilizing an EGFP-CEM-NK(r) flow cytometric assay. Eighteen sera with high ADCC antibody activity were then comprehensively examined for ADCC antibody epitopes using the HIV-1 subtype CRF01_AE TH023 gp120 peptide set consisting of 126 peptides of 15 amino acids in length, overlapping by 11 amino acids. This peptide set was divided into five pools (E1-E5). Each positive peptide pool was further investigated for fine epitope mapping of ADCC antibody activity using a 5 by 5 peptide matrix format. Interestingly, six and three peptides from peptide pools E1 and E2, respectively, responded to at least 33.33% of the tested sera. These nine common ADCC epitopes were localized to the C1 and V2 region of gp120. Furthermore, 5/9 epitopes were also shown to serve as full or partial Nt antibody targets for HIV-1 subtypes B and CRF01_AE. We submit these data on epitope mapping of ADCC or dual ADCC-Nt antibodies against HIV-1 gp120 that should be considered in the formulation of vaccines against HIV-1.