Post-transfusion Samples Research Articles

Transfusion transmission of highly prevalent commensal human viruses

Anellovirus species Torque teno virus (TTV), Torque teno mini virus (TTMV), and Torque teno midi virus (TTMDV) and flavivirus GBV-C are highly prevalent and genetically diverse chronic human viral infections that have not yet been associated with disease. To determine if these commensal viruses are transmitted by blood transfusions, we genetically analyzed viral species in cryopreserved samples from blood donors and corresponding pre- and posttransfusion samples from recipients enrolled in the Transfusion-Transmitted Viruses Study cohort. All 24 individuals in 12 donor-recipient pairs were infected with TTV, while 16 were infected with TTMV, 15 with TTMDV, and four with GBV-C. None of the 12 informative cases of TTV transfusion or eight cases of TTMV transfusion, where the donor and recipient viruses could be genetically differentiated, resulted in detectable transmissions in which the donor viruses were detected in the recipient by direct sequencing of the polymerase chain reaction products. Of the five informative cases of TTMDV transfusion, including two cases of transfusion into TTMDV-negative recipients, one case of superinfection was seen with both the recipient and the donor viral variants detected in the transfusion recipient for at least 11 days posttransfusion. Three donor-recipient pairs were informative for GBV-C transmission with only one transfusion into a GBV-C-negative recipient resulting in a transiently detected infection. Transmission of the common commensal anelloviruses and GBV-C during transfusion was detected in 2 of 12 already infected or uninfected recipients. Underestimation of the true rate of viral transmission may be due to limitations in detecting donor viral variants present as minority variants in the already infected transfusion recipients.

Screening for G6PD Deficiency in Blood Donor Population

Dear Editor, Glucose-6-phosphate dehydrogenase deficiency is the most common enzyme deficiency worldwide. Approximately 400 million people are said to be affected worldwide. According to world health organization 7.5% of world population are carriers of glucose-6-phosphate dehydrogenase deficiency and 2.9% were G6PD deficient [1]. It causes spectrum of diseases including neonatal hyperbilirubinemia, acute hemolytic and chronic hemolytic anemia. Usually the affected individuals are asymptomatic unless they are provoked by oxidative stress [2]. This enzyme deficiency is very prevalent in individuals of Africa, America, Mediterranean, and East Asia. In India the incidence of G6PD has been variably reported as 0–37% in different castes and communities. Higher incidence of G6PD deficiency is seen in north and west India (15%) as compared to south India (1–2%) [3]. It implies that this inherited metabolic disorder is an important health problem in India and it is necessary to carryout large scale screening in the whole population. Severe manifestations of G6PD deficiency can be prevented if necessary precautions are taken, hence this study was done to determine the incidence of G6PD deficiency in asymptomatic population represented by healthy blood donors and to characterize the demographic profile and laboratory profile of such group. The study was conducted on 2005 healthy blood donor samples. This screening program included 2000 male and 5 female blood donors who came to blood bank for donation of blood. These donors were from different part of India. The G6PD activity was assessed in their red cells using anticoagulated (EDTA) venous blood. Sixteen blood donors out of 2005 screened by methaemoglobin reduction test were found to be G6PD deficient. All of them were male. These individuals were clinically normal and had no significant past history. The incidence of G6PD deficiency was 0.8% in the sampled population. Among this 16 G6PD deficient subjects two blood donors were from West Bengal, one from Kerala and the rest of the 13 subjects were from Karnataka. The mean Haemoglobin values of the normal donor population vs. G6PD deficient donor population were compared using unpaired t test. The difference in the mean Hb value was not statistically significant (P value > 0.5). Sixteen units of blood were transfused to patients. There was two [12.5%] transfusion related reaction among the patients who received the G6PD deficient blood. One of the patients, who was transfused following upper Gastro intestinal bleeding, had itching and rashes all over the body after transfusion of 50 ml blood. The reaction workup done ruled out haemolytic reaction. A probability of allergic reaction was considered. As in this patient transfusion was stopped and he did not receive further transfusions for next 48 h, his Hb fell from 9.0 to 7.7 gm/dl. The second patient was a neonate undergoing surgery for a ruptured meningomyelocoele. The blood was transfused during surgery following which the baby developed mottling and passed red colored urine. A hemolytic transfusion was suspected. The transfusion reaction workup showed haemolysis in the post transfusion sample and hemoglobinuria. But blood bag showed no hemolysis. However as pretransfusion sample also showed haemolysis, a hemolytic reaction could not be conclusively reported. Hemoglobin dropped from 12.2 to 8.0 gm/dl. The beneficial effects of blood transfusion, for the typical 70 kg human each unit of whole blood should increase the hematocrit by 3–5% or hemoglobin by 1–1.5 gm/dl [4].Testing for G6PD deficiency is not routinely practiced for donors units [2]. Various studies have reported a mild increase in bilirubin, LDH, and decrease in hemoglobin following the transfusion of G6PD deficient blood. This is especially in neonates (preterm). Transfusion of G6PD deficient red cells to the premature infants has been associated with hemolytic and severe hyperbilirubinemia requiring exchange transfusion [5]. In our study also one neonate developed suspected hemolytic transfusion reaction following transfusion of G6PD deficient blood. Under normal circumstances the transfusion of G6PD deficient blood is harmless for most of the individuals until they are exposed to certain oxidative drugs. This study was an attempt to find out the prevalence of G6PD deficiency in our population. This will help us to quantitate the magnitude of the problem in the population and initiate measures to screen the population or a selected group in the population. The prevalence of G6PD being very low in this study (0.8%) and as many people remain asymptomatic unless challenged with oxidative drugs or infection, screening the whole population for G6PD deficiency is not warranted. As one of the neonates receiving G6PD deficient blood developed suspected hemolytic reaction, screening of blood to be transfused for select population like neonates is advisable.

An unusual febrile nonhemolytic reaction occurred after transfusion in a thalassemia major patient with asymptomatic Plasmodium falciparum infection

Febrile nonhemolytic transfusion reactions occur in 0.12% of transfusions, usually during transfusion or within 4 to 6 hours after transfusion and are not medically dangerous. A patient with thalassemia from Togo with asymptomatic malaria in which the infection became clinically manifest only after blood transfusion, mimicking a febrile nonhemolytic transfusion reaction, is presented. Thirty-two hours after transfusion of 2 O D- red blood cell (RBC) units, the patient (phenotype A(2) D+) developed fever and multiorgan failure and was admitted to the intensive care unit. Direct and indirect antiglobulin tests were negative on posttransfusion samples. Blood cultures and infectious diseases testing were negative. No malaria parasites were found at thick blood smear microscopic examination on Days 1 and 2 and the malaria rapid diagnostic test gave inconsistent results. Plasmodium total antibodies were detected in the serum at high levels. On Day 5, routine microscopic examination of blood smear revealed the presence of parasites in a very small number of RBCs. This finding was almost simultaneous to the availability of polymerase chain reaction testing results that were positive for P. falciparum. The sequential agglutination with anti-A antiserum allowed patient's and donors' RBCs to be separated and revealed that the parasitized cells were almost exclusively those of donors (14.4% vs. 0.029%). Malaria infection in implicated donors was excluded. In this patient with thalassemia with asymptomatic malaria, the infusion of two normal RBC units provided a favorable environment for a rapid parasite replication leading to a dramatic acute malaria attack.

An Unusual Case of ‘Delayed‘ Febrile Non Hemolytic Transfusion Reaction in a Thalassemia Major Patient with Asymptomatic Plasmodium Falciparum Infection.

Abstract 4186We report the case of a thalassemic patient with asymptomatic malaria in which the infection became clinically manifest only after blood transfusion, mimicking a febrile non haemolytic transfusion reaction. The patient was a 19 yr old thalassemic girl from Togo regularly and uneventfully transfused every 60-90 days since the age of 5 months and splenectomised at 13 yrs. Two malaria episodes in 1995 and 1997 were treated with quinine and halofantrin respectively. In February 2008 the transfusion interval shortened (15 days) and blood transfusions were constantly followed after 30-40 hours by high fever (39-4028C) and hypotension. The patient was referred to our Hospital to clarify and overcome the transfusion problems. Malaria attacks were excluded by the referring clinic.At admission, Hb was 5.9 g/dL, Hct 18.3%, MCV 71.8 fL, MCH 23.1 pg, MCHC 32.2%, reticulocytes 0.82%. WBC 13.1 × 109/L platelet count 953 × 109/L erythroblasts 12.7% of nucleated cells. Molecular analysis of β and α genes revealed double heterozygosity for β IVS1-1 G>A and β IVS2-849 A>G, associated with deletion 3.7 (-α3.7) at the heterozygous state. G6PD 6.29 IU/g Hb (ref.values 5.97±1), Pyruvate kinase 25.1 IU/gHb (ref. v. 11.9-16.1). Red cell osmotic fragility was decreased. The cytofluorimetric analysis of red cells labelled with eosin 5 maleimide gave normal results. Direct antiglobulin test was negative and red cell alloantibodies were not detected. The patient's blood group assessed by molecular biology testing was: genotype O1A2, phenotype A2; ccDee; K-k+; Jk(a+b-); Fy(a-b+); M+N+S+s-. The patient was given 2 crossmatch negative, filtered O neg units: the post-transfusion Hb values were 9.9 g/dL. Thirty-two hours later the patient developed fever (40°C) and hypotension followed by disseminated intravascular coagulation, metabolic acidosis and anuria. Direct and indirect antiglobulin tests were negative on a post-transfusion sample. Because of the rapid worsening of clinical conditions she was admitted to the Intensive Care Unit of our Hospital. Septic shock was excluded on the basis of negative blood cultures and infectious diseases testing. Microscopic examination of thick blood smear was negative for malaria parasites on 2 consecutive days and the malaria rapid diagnostic test (RDT) (Core Malaria Pan-PV-PF®, Core Diagnostics, U.K.-Effegiemme, Nerviano, Milano, Italy) gave inconsistent results. Anti Plasmodium total antibodies (Malaria EIA New Market Laboratories U.K.- Bio-Rad Italia) were detected in the serum at high levels (odd sample/cut off 22). Vital functions were supported by mechanical ventilation, amine administration and continuous venous-venous hemofiltration (CVVH). Broad spectrum antibiotic therapy was started. Fresh frozen plasma and two units of PRC were transfused on day 1 and 2 without increment of Hb level. Three days later, the examination of the peripheral blood smear for counting schistocytes revealed the presence of parasites in a very small number of red cells. This finding was almost simultaneous to the availability of PCR testing results that were positive for P. Falciparum. The differential agglutination with anti A antibody performed on a blood sample collected at admission to ICU allowed to separate patient's and donors' red cell. Counting the number of parasitized red cells in total blood and in the donor fraction blood revealed that the parasitized cells were almost exclusively those of donors (14.4 % vs 0.029%).Treatment with quinine chlorhydrate 500 mg i.v. every 8 hr for 3 days followed by oral quinine sulphate 500 mg every 8 hr for 3 days was successfully undertaken without significant side effects leading to dramatic improvement of clinical conditions and to eradication of P. falciparum. Two RBC units were effectively transfused without any reaction, and in the following two weeks the patient maintained stable haemoglobin values. After discharge, the transfusion interval returned to the previous values (60-90 days) without any post-transfusion reaction.We therefore think that the transfusion of normal, non malaria-resistant red cells fuelled the P. falciparum infection causing fever, DIC and acidosis giving rise to a very severe, atypical febrile non haemolytic transfusion related reaction. Disclosures:No relevant conflicts of interest to declare.

Probiotic‐associated high‐titer anti‐B in a group A platelet donor as a cause of severe hemolytic transfusion reactions

Hemolytic transfusion reactions (HTRs) can occur with transfusion of platelets (PLTs) containing ABO-incompatible plasma. Reported cases have involved group O donors. Two cases of PLT-mediated HTRs associated with the same group A plateletpheresis component, collected from a donor taking high doses of probiotics are reported. Case 1 was a 40-year-old 69-kg group B stem cell transplant patient who received one-half of a group A plateletpheresis component. Severe back pain occurred 10 minutes into the transfusion, accompanied by anemia and hyperbilirubinemia. Case 2 was a 5-year-old 26-kg group B male with aplastic anemia who received the other half of the same plateletpheresis component, volume reduced to 37 mL. Syncope occurred immediately after the transfusion, with laboratory evidence of hemolysis a few hours later. Serologic investigation of posttransfusion samples from both patients revealed positive direct antiglobulin tests: C3d only for Case 1 and immunoglobulin (Ig)G and C3d for Case 2; the eluates contained anti-B. The group A donor's anti-B titer was 16,384 at saline and IgG phases. Donor lookback revealed that the donor had donated 134 apheresis PLTs over many years. For 3 years, he had intermittently taken probiotics; 3 weeks before the index donation, he began taking three tablets of probiotics every day. Lookback of prior group B recipients uncovered a case of acute hemolysis that was not recognized at the time. The solubilized probiotic inhibited anti-B in vitro. Non-group O PLT donors can have high-titer anti-A or anti-B that might mediate HTRs, and probiotic ingestion in blood donors represents a novel mechanism of stimulating high-titer anti-B.

Fetal Hydrops and Other Variables Associated with the Fetal Hematocrit Decrease after the First Intrauterine Transfusion for Red Cell Alloimmunization

Objective: To evaluate the influence of fetal hydrops and other variables on fetal hematocrit (Hct) decrease after the first intrauterine transfusion (IUT) in alloimmunized pregnancies. Methods: From 1996 to 2006, the data of all alloimmunized pregnancies submitted to IUT were assessed. Exclusion criteria included: fetuses submitted to intraperitoneal transfusion; pregnancies complicated by other fetal abnormalities; pregnancies submitted to only one IUT, and cases in which posttransfusion or pretransfusion blood samples were not obtained. Linear regression models were implemented to assess the relationship between the rate of Hct fall after the first IUT and the following variables: fetal hydrops; antibody titer; gestational age at the first IUT; number of days between the first and second IUT; pretransfusion and posttransfusion fetal Hct values. Results: Fifty fetuses fulfilled the study criteria. The fetal Hct decrease after the first IUT was 1.21 (range 0.18–2.3) %/day. The variables independently associated with the fetal Hct drop after the first IUT were the fetal hydrops (p = 0.000), the pretransfusion fetal Hct (p = 0.001) and the posttransfusion fetal Hct (p = 0.016). Conclusion: Fetal hydrops, pretransfusion fetal Hct and posttransfusion fetal Hct seem to influence the fetal Hct decrease between the first and second IUT. These findings may be helpful for estimating the rate of fetal Hct drop and programming the following IUT.

Hyperhemolysis syndrome in sickle cell disease: case report (recurrent episode) and literature review

Hyperhemolysis syndrome (HS) has been well described both in sickle cell disease (SCD) and non-SCD patients. The pathogenesis remains unclear. The possible mechanisms include bystander hemolysis, suppression of erythropoiesis, and destruction of red cells (RBCs) due to contact lysis via activated macrophages. This study reports a child with SCD who presented with recurrent episode of HS. In the first episode, the hemoglobin (Hb) level decreased to 4.1 g per dL. He was treated with intravenous immunoglobulin (IVIG) and oral steroids, and transfusion was avoided. Six months later he had another episode of HS. The nadir Hb level decreased to 3.2 g per dL, and further transfusions were given with IVIG-IV methylprednisolone cover. RBC alloantibodies were not identified in pre- and posttransfusion samples in patient's serum in both episodes. HLA antibodies were also not detected. This is the second reported case of recurrent HS in a child. Recent studies have shown that the adhesion molecules expressed on RBC erythroid precursor cells and reticulocytes can interact with macrophages and can cause hemolysis. Because RBC alloantibodies and HLA antibodies were not identified in this case, it is believed that the patient's cells and transfused cells were destroyed by macrophages either by direct contact lysis or by erythrophagocytosis. The possible mechanism of IVIG and steroids on suppression of macrophages resulting in cessation of hemolysis is discussed. Our case illustrates the danger of recurrent HS and the difficulty of balancing this against the need for transfusions in patients presented with severe hemolysis.

Increased thrombin generation and fibrinogen level after therapeutic plasma transfusion: Relation to bleeding

In a clinical setting, fresh frozen plasma (FFP) is transfused to diluted patients with complicated surgery or trauma, as guided by prolonged conventional coagulation times or low fibrinogen levels. However, the limited sensitivity of these coagulation tests may restrict their use in measuring the effect of transfusion and hence predicting the risk of perioperative bleeding. We used the more sensitive, calibrated automated thrombogram (CAT) method to evaluate the result of therapeutic FFP transfusion to 51 patients with dilutional coagulopathy. Thrombin generation was measured in pre- and post-transfusion plasma samples in the presence of either platelets or phospholipids. For all patients, the transfusion led to higher plasma coagulation factor levels, a shortened activated partial thromboplastin time, and a significant increase in thrombin generation (peak height and endogenous thrombin potential). Interestingly, thrombin generation parameters and fibrinogen levels were higher in post-transfusion plasmas from patients who stopped bleeding (n = 32) than for patients with ongoing bleeding (n = 19). Plasmas from 15 of the 19 patients with ongoing bleeding were markedly low in either thrombin generation or fibrinogen level. We conclude that the thrombin generation method detects improved haemostatic activity after plasma transfusion. Furthermore, the data suggest that thrombin generation and fibrinogen are independent determinants of the risk of perioperative bleeding in this patient group.

Highly Accurate Rhesus and Kell Blood Group Phenotyping of Autologous Red Cells after Allogeneic Blood Transfusion Using a Simplified Method of Microhematocrit Centrifugation and Automation with the DiaMed Classic ID-Gel Station and Diaclon 'Rh Subgroups+K' Gel Cards.

The presence of residual donor red cells in transfused patients impairs the ability to obtain a reliable phenotype for subsequent allogeneic blood transfusions. Post transfusion phenotyping is particularly important in transfused patients developing multiple or weak blood group antibodies. However, separation of autologous and transfused red cells from recently transfused patients is necessary for the proper identification of any suspected red cell alloantibody or autoantibody. The standard technique for the separation of autologous from transfused red cells involves microhematocrit centrifugation with phytate ester solution, a laborious and time consuming method. We have examined the effectiveness of a simplified microhematocrit technique, omitting the use of esters, to separate autologous and transfused cells by performing Rhesus (Rh) and Kell (K) blood group phenotyping, both manually and by automation, pre and post transfusion. The Rh phenotype and K type were recorded on pre- and post-transfusion peripheral blood samples from 100 newly transfused operating theatre patients, both manually and by automation using the DiaMed Classic ID-Gel Station and Diaclon 'Rh subgroups+K' gel cards. On post transfusion blood samples, separation of autologous from transfused cells was performed using the Haematospin 1300, based on the difference in the specific gravity between reticulocytes and mature red cells where the autologous cells concentrate in the top layer of the microcapillary tube. This separation was performed on blood samples taken three days post transfusion, based upon the normal reticulocyte maturation time of three days. On all samples tested, there was 100% correlation between manual and automated ID-GEL Station results. On Rh phenotyping of post transfusion blood samples prior to cell separation, 46 of the 100 patients had a mixed field (Mf) reaction. Repeat Rh phenotyping of day three post transfusion samples, following separation of autologous cells using the microhematocrit centrifugation technique, showed complete correlation with pre transfusion Rh phenotyping results in all but two cases, where a Mf reaction persisted. These two cases had had massive blood transfusions (40 and 36 units of red cells respectively) compared to the amount of red cells (mean 3.3 units, range 1–15) transfused to the other 98 patients. One week after initial Rh phenotyping, allowing sufficient time for autologous reticulocyte recovery in these two patients, repeat Rh phenotyping of separated autologous cells correlated with that of the pre transfusion results. 6 of the 100 patients were K positive on pre transfusion testing. Repeat testing of post transfusion blood samples prior to cell separation showed a Mf reaction for the K antigen in two of these 6 cases, suggesting transfusion of K antigen negative red cell units. Once cell separation was complete, both patients were confirmed as K positive. In conclusion, the simplified microhematocrit centrifugation technique and automated ID-Gel station are reliable, sensitive and less hazardous in providing an accurate Rh phenotype and K type on patient blood samples taken three days post transfusion and could be easily performed in the standard blood transfusion laboratory. The only limitation to this technique is an inability to separate autologous and transfused cells from patients who have undergone massive transfusions within a short space of time.

Incidence of transfusion‐induced platelet‐reactive antibodies evaluated by specific assays for the detection of human leucocyte antigen and human platelet antigen antibodies

The aim of this work was to study the incidence of transfusion-induced platelet-reactive antibodies in a selective patient population and evaluate different methodologies for platelet antibody screening (PAS). The patients were retrospectively selected and divided into three separate groups: haematological malignancies (Group 1: n = 33); cardiac and orthopaedic patients (Group 2: n = 31) and a control group (Group 3: n = 23) selected with the same diagnoses of Group 2. PRE- and POST-transfusion samples were tested for PAS by the following tests: PIFT (platelet immunofluorescence test), MAIPA (monoclonal antibody immobilization of platelet antigen), Flow PRA(R) and LCT (lymphocytotoxicity test). There was not a 100% concordance among the methodologies used. PIFT, MAIPA and Flow PRA presented very similar results whereas that of LCT differed from the other methods. A high rate of positive results (32%) was found in the PRE samples followed by an increase of almost 50% after blood transfusion (POST samples: 42.5% of positivity), but there was a statistical difference (P < 0.05) between the PRE and POST transfusion sample only for the Flow PRA(R) technique tested on Group 2. Human leucocyte antigen (HLA) class I antibodies were present on 97.4% of POST positive samples, 5.4% presented anti-human platelet antigen (HPA)-1b antibodies and 8.1% presented a mix of pan-reactive antibodies against glycoprotein IIbIIIa, IaIIa and IbIX. Blood transfusion did not increase the rate of alloimmunization in our haematological patients (Group 1); however, the patients were already admitted with a high rate of alloimmunization (12%). Group 2 patients are being immunized and the impact of this procedure remains to be studied as these patients may eventually undergo further hospitalization and receive more blood transfusion.

Medicolegal assessment of blood transfusion errors—An interdisciplinary challenge

Given a current total incidence of erroneously administered blood transfusions of 1:12,000–1:36,000 (AB0 incompatible 1:38,000), the percentage of lethal outcomes ranges between 2 and 5%; i.e. the sole fact of an erroneous transfusion does not mandatorily result in a causal connection with lethal outcome, which can give rise to problems in the medicolegal assessment. We report on the conception and results of a novel interdisciplinary approach to assess the lethal significance of blood transfusion errors. Besides autopsy, histological investigation and immunohistochemical detection of AB0 incompatible foreign red blood cells in autopsy specimens, transfusion medicine investigations offer the opportunity to assess several immunohaematologic features. We assessed the immunohaematologic gel card (“ microcolumn”) technique for suitability in the forensic assessment of an AB0 incompatible transfusion incident in a septic patient, who had had no history of previous blood transfusions, with lethal outcome. After such an erroneous transfusion had been simulated in vitro, pre-transfusion and cadaver patient blood samples (p.m. interval: 3 days) were analysed. Amongst other things, IgG-loaded erythrocytes were detected in pre- and post-transfusion samples; the presence of irregular antibodies directed against blood group antigens and anti-A or anti-B isoagglutinins, respectively, especially in the pre-transfusion sample was ruled out. Besides the demonstration of AB0 incompatible red blood cells in the cadaver blood sample, blood group incompatibilities other than AB0 were excluded. With regard to the cause of death, in synopsis with autopsy findings and clinical symptoms, the results did not allow for a final discrimination between the impact of the pre-existing septic inflammatory response syndrome and sepsis, respectively, and potential lethal effects of a (haemolytic) transfusion reaction. Besides pre- and post-transfusion compatibility testing in clinical transfusion medicine as required by German National Guidelines, the reported immunohaematologic investigations offer an important supportive tool for the forensic assessment of lethal erroneous transfusions and investigation of blood samples of survivors of transfusion incidents as well. Besides established morphological techniques, they allow for a certain evaluation of the pathophysiological impact of transfusion incidents as well as a diversified assessment of immunohaematologic features beyond the AB0 system.

Selection of different 5′ untranslated region hepatitis C virus variants during post‐transfusion and post‐transplantation infection

Hepatitis C virus (HCV) translation is initiated in a cap-independent manner by an internal ribosome entry site (IRES) located within the 5' untranslated region (5'UTR). Sequence changes in this region could affect translation efficiency and presumably viral replication. To determine translation efficiency of 5'UTR variants developing during post-transfusion hepatitis C in two immunocompetent subjects and in two immunosuppressed liver recipients with recurrent HCV. Sequential samples were screened for 5'UTR changes by single-strand conformation polymorphism followed by cloning and sequencing whenever band pattern suggested sequence changes. 5'UTR variants were tested for IRES activity using a bicistronic dual luciferase expression plasmid transfected into HepG2 and Huh7 cell-lines. In the transfused patients, translation efficiency of 5'UTR variants from early post-transfusion samples was 5.1- to 13.7-fold higher than that of predominant variants found in late follow-up samples. Post-transplant variants in the other two patients had 2.6- to 5.9-fold higher translation efficiency than those present only in pretransplant samples. In the immunocompetent host there may be selection of low translation efficiency HCV variants over the course of infection. However, in immunosuppressed subjects the opposite seems to be true as low translation efficiency variants are superseded by high translation efficiency variants.

Hazards of Neonatal Blood Transfusion

After completing this article, readers should be able to: 1. Describe acute immune-mediated transfusion reactions seen in neonates, their treatment, and their prevention. 2. Describe acute nonimmune-mediated transfusion reactions seen in neonates, their treatment, and their prevention. 3. Describe delayed transfusion complications. 4. Delineate the steps involved in the blood donor screening process in the United States. The safest blood transfusion is the one not administered. When a transfusion is needed, it is important for both physicians and nursing staff to be aware of its potential acute and delayed adverse effects. Adverse consequences may be minimized through early recognition and prompt therapeutic intervention. Noninfectious complications may be either acute (occurring within hours of transfusion) or delayed. Acute reactions may be categorized further according to their pathogenesis: immunologic versus nonimmunologic. Patients should be monitored closely during the transfusion. For all suspected transfusion reactions, the ordering physician and the nursing staff should be trained to stop the transfusion immediately, keep the intravenous line open, check that the blood component was given to the correct patient, and assess and treat symptoms. A posttransfusion blood specimen and the blood bags should be sent to the Transfusion Service (TS) along with a transfusion reaction report form that includes patient diagnosis, history of previous reactions, date and time of reaction with associated symptoms, and vital signs before and after the transfusion. The TS technologist performs a clerical check, inspects posttransfusion blood samples for hemolysis, and repeats ABO/Rh testing on both the transfused unit and the patient’s sample along with a direct antiglobulin (Coombs) test. Further testing is guided by the nature of the patient’s symptoms. Suspected delayed complications of transfusion also should be reported to the TS so the appropriate investigation may be initiated. ### Acute Immune-mediated Transfusion Reactions Acute hemolytic transfusion reactions are the second most common cause of transfusion-related fatality in adult patients, …

Intravenous immunoglobulin given to lymphoma patients with recurrent haemolytic transfusion reactions after transfusion of compatible blood

Accelerated destruction of red cells after transfusion of compatible blood has been reported in both sickle cell disease (SCD) and non-SCD patients. We report three patients with lymphoma, all of whom had recurrent haemolytic transfusion reactions after receiving compatible red cell units. The direct antiglobulin test (DAT) was negative and there were no detectable red cell alloantibodies in either pre-transfusion or post-transfusion samples. As there was no evidence of red cell antibody-mediated haemolysis and response to oral steroids, a trial of intravenous immunoglobulin (IVIg) was given. Immediate cessation of haemolysis with sustained haemoglobin level was achieved in all cases. The response to IVIg in these cases suggests that IVIg should be tried when recurrent non-antibody mediated haemolytic transfusion reactions occur in patients with a lymphoid malignancy.

Severe Acute Hemolytic Transfusion Reaction Following ABO-Mismatched Platelet Transfusion: Should ABO-Mismatched Platelet Transfusion Policy Be Evaluated.

Transfusion of ABO-mismatched platelets is common practice. Although a unit of single donor apheresis platelet (SDP) contains as much plasma and passive anti-A or/and anti-B antibodies (allo-agglutinins) as a unit of fresh frozen plasma, ABO- mismatched platelet transfusions are considered safe and hemolytic transfusion reactions have only rarely been reported. We report a case of a 67-year old male, blood group A, with mantle cell lymphoma who received 1 SDP for chemotherapy induced thrombocytopenia. Several prior platelet transfusions had been uneventful. After 150 ml of irradiated, leukoreduced, SDP from blood group O donor had been infused, he developed nausea, light-headiness, chills, and back pain. He became bradycardic and hypotensive. Platelet transfusion was discontinued and intravenous dexamethasone and rapid saline infusion was administered. Later, his hemoglobin dropped from 10.6 to 7.4 g/dl and total serum bilirubin increased by 9 fold. A direct antiglobulin test (DAT) on post-transfusion sample demonstrated 1+ reactivity at room temperature. An elution test performed on the patient's circulating red blood cells contained anti-A antibodies. These results are consistent with a severe acute hemolytic transfusion reaction. The donor anti-A and anti-B antibodies titers at room temperature and at 37°C were 512 and 2048 with A1 cells and 128 and 256 with B cells, respectively.This case illustrates the rare possibility of hemolytic transfusion reaction occurring as a result of ABO-mismatched platelets from group O donor with high-titer anti-A antibodies. SDP (in contrast to pooled platelets) offers the advantages of reduced risk of infection transmission and alloimmunization by limiting exposure to one donor only. However, it increases the opportunity for hemolytic transfusion reaction to occur if the donor has high-titer antibodies which would have been diluted if pooled platelets were used. As SDP use becomes more widespread, the risk of clinically significant hemolytic reactions is likely to increase, especially in seriously ill patients who tolerate these reactions particularly poorly. Moreover, increasingly, apheresis platelets are being donated by a small group of highly motivated donors, raising the chances of repeated transfusions from the same donors. We propose that the safety of ABO-mismatched platelet transfusions be re-evaluated. Effectiveness (and costs) of potential preventive strategies such as screening all SDP units for high-titer antibodies and/or plasma volume reduction of all mismatched SDP units should be studied.

The Use of Platelet Antibody Testing at a Tertiary Medical Center.

Detection of platelet antibodies is often helpful in the diagnosis of platelet transfusion refractoriness (PR), neonatal alloimmune thrombocytopenia (NAIT), and post-transfusion purpura (PTP), but is controversial in idopathic thrombocytopenia purpura (ITP). We performed a retrospective chart review to determine the utilization and value of a solid-phase ELISA assay consisting of 6 wells coated with platelet glycoproteins (GP), and one well with HLA class I antigens (GTI PakPlus, Brookfield WI) at our medical center. For a 3 year period between June 2001 and June 2004, 183 tests for 163 patients were ordered for the following diagnostic questions: 151 tests for ITP, 19 for PR, 6 for NAIT, 2 for transfusion-related acute lung injury (TRALI), 5 for unclear reasons, and none for PTP. The mean patient age was 55 (range 12 – 94), 55% were male, 50% were inpatients, and the mean platelet count (excluding mothers with NAIT and a patient with TRALI) was 60x109/l (range 5–215 x 109/l). There were 48 patients who had 51 positive tests (28%), of which 61% were inpatients. The mean platelet count was 47 x 109/l (range 5–115 x 109/l). Only one or more GP wells were positive in 57% of positive tests, only the HLA class I well was positive in 27%, and ≥one GP well plus the HLA well were positive in 16%. To determine how results were used, we reviewed the medical records associated with each positive test. Out of 51 positive tests, no mention was made of the test order or result in 21 (41%). For 6 tests (12%), the order was noted, but no results were mentioned. In the other 24 tests, the result was used to make a diagnosis in 12 cases (3 NAIT, 3 PR, 6 ITP), and in 12 the result was used to confirm an already known diagnosis (1 NAIT, 4 PR, 7 ITP). Therapy was started before the test was completed on 17 occasions (4 PR, 13 ITP). Eighteen patients had repeat testing (38 tests). The interval of time between tests was 112 days on average (range 8–365), including 5 sets within 28 days. The results were unchanged in 11 patients (61%); in 5 patients GP wells changed from positive to negative; and in 2 patients the HLA well changed, including a patient with pre- and post-transfusion samples for evaluation of TRALI, and another in whom the test went from positive to negative. In 3 mothers evaluated for NAIT with positive results (1 HPA-1a, 2 HPA-5b), confirmation was obtained at a reference laboratory (Blood Center for Southeastern Wisconsin). A total of fifteen patients were evaluated for PR with 19 samples. Seventeen samples underwent parallel testing (HLA lymphocytotoxic antibody screen alone, or with platelet antibody detection at the reference laboratory); 13 results (76%) were confirmed. All four mismatches were between the HLA ELISA well and the HLA antibody screen: in 2 samples the PakPlus was positive but the HLA antibody screen was negative; in the other 2 a negative HLA well was accompanied by a high PRA (63%, 80%). These results demonstrated that 1) the ELISA assay was routinely negative in patients with platelet counts >115 x 109/l; 2) 53% of positive results were ignored in the medical records; 3) the test had a moderate reproducibility (61%), but could have been related to clinical changes in the patients; and 4) the yield of clinically useful information for patients with ITP (151 tests, 13 positive results utilized for clinical management) was low. For PR, the PakPlus is reliable but only qualitative, compared to the quantitative HLA antibody screen. The test appears most useful for rapid antibody identification in NAIT and is of limited clinical utility for ITP.

Short tandem repeat (STRs) and sex specific Amelogenin analysis of blood samples from neurosurgical female transfused patients

Blood transfusion and its effects on DNA profiles of an individual is an important issue in the forensic context. In the present study, the effects of unfiltered whole male blood transfusion in seven female subjects were investigated by PCR-based assays on serial post-transfusion blood samples. PCR analysis of specific short tandem repeat (STR) primer pair to CSF1PO, TPOX, TH01, F13A01, FESFPS, vWA and sex specific Amelogenin loci for banding pattern of alleles generated from the pre- and post-transfusion of seven female recipients and their corresponding donors were studied. Pre- and post-transfusion DNA profiles of all the seven female recipients were found to be consistent with no evidence of the male and female donor genetic material. It is concluded that currently used PCR-based STRs and sex specific Amelogenin loci DNA profiling techniques in forensic science are reliable and informative for paternity and identity purposes in situations of transfusion of 1 or 2 U of unfiltered whole male and female blood (>72 h old) to female recipient up to 24 h post-transfusion.

Hyperhaemolysis Syndrome in a Patient with Myelofibrosis

Hyperhaemolysis syndrome is well recognised in patients with sickle cell disease and β-thalassaemia major, but has not been described in patients with other haematological diseases. We describe a case of fatal post-transfusion hyperhaemolysis occurring in a lady with myelofibrosis evolving into acute myeloid leukaemia. No free antibodies were identified in either pre- or post-transfusion serum samples. Since her haemoglobin (Hb) level after the reaction was very much lower, than of would have been the case if only the transfused red cells had haemolysed, it seems likely that this severe haemolytic transfusion reaction was an example of the so-called hyperhaemolysis syndrome.

Differentiation of autologous ABO, RHD, RHCE, KEL, JK, and FY blood group genotypes by analysis of peripheral blood samples of patients who have recently received multiple transfusions.

After multiple transfusions, the serologic typing of autologous blood group phenotypes is difficult, because of mixed RBC populations. The genotyping of ABO, Rh, Kell, Kidd, and Duffy systems could be used to determine autologous blood group antigen status. Blood samples from patients and donors were analyzed before and after 26 multiple-transfusion events. An average of 6.9 non-WBC-reduced RBC units with an average age of 5.9 days were administered per transfusion event. The average period of blood sampling after transfusions was 5.3 days. All samples were serologically phenotyped for ABO, Rh, Kell, Kidd, and Duffy. Pretransfusion, posttransfusion, and buccal samples from patients were genotyped for the corresponding alleles by a uniform PCR sequence-specific primer protocol that allowed their simultaneous determination within 3 hours. All posttransfusion samples exhibited mixed-cell populations of various blood group systems on serologic testing. Genotyping from peripheral blood produced results identical to the autologous blood group phenotypes, regardless of the amount of blood transfused or of the length of the sampling period after transfusion. A fast and reliable PCR-sequence-specific primer DNA genotyping assay for simultaneous determination of autologous ABO, Rh, Kell, Kidd, and Duffy blood groups can be performed on peripheral blood samples, even though the patients have recently received multiple transfusions.

Evaluation of a new enzyme immunoassay for hepatitis C virus (HCV) core antigen with clinical sensitivity approximating that of genomic amplification of HCV RNA

The aim of this study was to analyze the clinical performance of a new enzyme immunoassay (EIA) for hepatitis C virus (HCV) core antigen in comparison with the reverse transcription polymerase chain reaction (RT-PCR). A total of 310 patients with acute or chronic hepatitis C, and 132 HCV-negative controls were studied. Chemiluminescence EIA with monoclonal anti-HCV core antigen was used, and qualitative and quantitative commercial RT-PCRs and an in-house nested RT-PCR were performed. Compared with nested RT-PCR, the core antigen assay showed 97% sensitivity and 100% specificity in 75 patients with chronic hepatitis C and 132 controls. HCV core antigen was positive in 16 (94%) of 17 patients with acute hepatitis C at initial consultation. In 3 persons prospectively followed, core antigen was detected in the first available (1-3 weeks) post-transfusion sample. In 167 anti-HCV–positive individuals, 129 (77%) were viremic; core antigen was detected in 126 (98%) compared with 129 (100%) for nested RT-PCR and 121 (94%) for the commercial RT-PCR. In 48 patients with chronic hepatitis C treated with interferon alfa, the concentration of core antigen before treatment was significantly (P < .002) lower in patients with sustained response than in nonresponders. All responders had a sustained loss of core antigen, whereas all nonresponders remained core antigen positive. The concentrations of HCV core antigen and HCV RNA correlated significantly (n = 48, r = .627, P < .001). In conclusion, the HCV core antigen assay is useful for the diagnosis of acute and chronic hepatitis C, and for predicting and monitoring the effect of interferon alfa treatment. (Hepatology 2000;32:388-393.)