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In Poland, hepatitis A virus (HAV) RNA screening was performed in plasma for fractionation usually immediately before shipment.Our goal was to study epidemiology, rate of transfusion transmitted HAV during epidemic (2017-2019), and viral characteristics of infected plasma donors.HAV RNA was tested in 1,866,590 donations from 1,210,423 donors using RT-PCR in mini pools of 96 (MP96) or TMA in MP16. Virological characteristics included RNA level (RL), antibody testing, and sequencing.Twenty-one HAV infections were identified (1.13/100,000 donations; 95% confidence interval [95% CI]: 0.74-1.72) and (1.73/100,000 donors; 95% CI: 1.35-2.65). The Blood Transfusion Centers were also informed about three donors, who were hospitalized for hepatitis A soon after their blood donation. In addition, we identified a donor, who had reactive result for HAV after receiving HAV vaccination. He tested positive twice 10 days after receiving the first and the second dose. The highest RL was 16 million IU/ml, mean 1,706,905 IU/ml, and median 220 IU/ml. The longest detectable RL lasted for 113 days. HAV-infected donors were seronegative (36%) or IgM positive (64%). We followed up on 12 HAV contaminated blood components issued for transfusion. In two out of seven identified patients viral transmission was confirmed (28.6%).Based on our results, we propose a 6 month deferral after HAV infection and 14 days post HAV vaccination. The infectivity rate was below 30%. The HAV RNA testing could be considered as an additional safeguard against HAV transmission at the time of increased incidence of HAV infections in the general population.

The risk of hepatitis B virus (HBV) transmission through blood and blood components is markedly reduced through serologic testing of each donation for the presence of HBV infection markers. Testing for hepatitis B virus surface antigen (HBsAg) is obligatory almost everywhere in the world. Several countries have also implemented testing for antibodies to hepatitis B core antigen (anti-HBc) and some perform HBV DNA tests, either in single donations or minipools of up to 96 donations. In countries with obligatory hepatitis B vaccination, the incidence of HBsAg-positive infections has been on the decline. Questions are therefore raised regarding the choice of optimal strategies for blood donor screening. In Poland, screening for hepatitis B virus surface antigen (HBsAg) to identify HBV infected donors was initiated in 1971. Pursuant to the guidelines of the Council of Europe, the analytical sensitivity of 0.13 IU/ml is required for a HBsAg diagnostic test to detect a HBsAg-positive infection. In 2005, HBV-DNA testing was added to the panel of screening tests for further reduction of HBV transmission risk through blood and blood components. The study presents the analysis of the HBV-DNA screening tests used in Poland so far. The analysis is based on the author’s doctoral dissertation and the outcome may largely contribute to the process of determining the effectiveness of the methods used up to date and to development of an optimal panel of HBV screening tests for Polish blood donors.

Transfusion of blood and blood components is one of the widely used medical procedures. The responsibility for provision of blood and blood components lies with Polish blood transfusion centers (CKiK) substantively supervised by the Institute of Hematology and Transfusion Medicine. Hospital based blood banks, hospital wards, immunohematology laboratories are supervised by CKiK. Every year approximately 600 thousand people donate blood, more than 75% of which are regular donors. The annual number of donations is maintained at a constant level of about 1.3 million. The aim of this position paper is to present the current status, achievements and advancement in collection, preparation and testing procedures and methods used to obtain the best possible quality blood components dedicated for clinical use. The aim is also to show the structure of blood transfusion service in Poland. Collaboration of all organizational units involved in transfusion medicine is crucial for providing high quality health care for patients. Polish transfusiologists and blood transfusion officers have largely contributed to transfusion science with numerous publications and significant research work. The implementation of novel methods and ongoing research positions Polish blood transfusion service among those of highly developed countries.

The new coronavirus — SARS-CoV-2 infections have recently become the main epidemiological and clinical challenge worldwide (Poland included). This article is based on the presentation entitled “Hematology and transfusion medicine and COVID-19” given/conferred during the webinar in May 2020. The aim was to introduce the procedure for diagnosing the new coronavirus infection. The procedure highlights the aspects of the pre-analytical and analytical phase that are critical for the quality of the performed tests and affected by both the commissioning and test-performing personnel. On the basis of current literature, further recommendations for the improvement of molecular diagnostics of SARS-CoV-2 are also presented.

Nucleic acid testing (NAT) was implemented in Poland in 1999 for screening of plasma for fractionation and for all blood donors in 2002. To analyze seronegative NAT‐positive samples representing hepatitis C virus (HCV) window‐period (WP) in the years 2000 to 2016 and to determine infection outcome. We analyzed results of 17 502 739 donations screened in minipools (6‐48) or individually. Index samples underwent viral load (VL) quantification, genotyping and Ag, and anti‐HCV re‐testing using chemiluminescence (CMIA), electrochemiluminescence (ECLIA), and fourth‐generation enzyme‐linked immunosorbent assay (IV EIA) assays. HCV‐seronegative infections were identified in 126 donations (7.2/mln donations; 95% confidential intervals, 6.0‐8.6). Frequency of NAT yields was decreasing over time. Of the initial 126 seronegative index cases 106 were retested: 32.1% were reactive in IV EIA, 11.3% in ECLIA, and 1.9% in CMIA. The lowest VL correlated with absent anti‐HCV and HCV Ag, while VL was highest when the antigen was detectable and then it decreased when anti‐HCV appeared at a level detectable by sensitive third generation tests while retesting. The proportion of genotype 1 was 38.9% in samples positive only for HCV RNA and 71.4% in samples that were anti‐HCV reactive in re‐testing. In parallel, genotype 3 frequency was 50% in the former group and 21% in the latter. NAT is an effective measure to limit HCV transmission by transfusion and IV EIA seems to have higher clinical sensitivity than ECLIA. Samples representing likely successive phases of early HCV infection were characterized by different genotype distribution probably due to very early elimination of genotype 3., Highlight NAT is highly effective in prevention of transmission of HCV seronegative infections.In nearly 7% of followed up donors antibodies appeared later than 50 days after index donation.The highest clinical sensitivity was demonstrated by the IVth generation tests.Among the third generation assays, ECLIA showed higher sensitivity than CMIA.Samples representing likely successive phases of early HCV infection demonstrated different genotype distribution.

Background Until now, markers of hepatitis E virus (HEV) infection have not been studied in blood donors throughout Poland, and no acute case of HEV infection has been closely documented or confirmed by HEV RNA detection. The prevalence of HEV infection markers, including HEV RNA in Polish blood donors and virus genotypes was investigated. Study design and methods In total, 12,664 individual donations from 22 Polish blood transfusion centers were tested for HEV RNA by transcription-mediated amplification. In addition, 3079 first-time donors sampled throughout Poland also were screened for antibodies to HEV. HEV RNA and immunoglobulin M-positive donations were confirmed using real-time reverse transcription-polymerase chain reaction and Western blotting, respectively. Results Ten donors were identified as RNA initial reactive (one of 1266 donors), and six (one of 2109) were identified as repeat reactive and confirmed by real-time reverse transcription-polymerase chain reaction or seroconversion. Sequence analysis identified HEV Genotype 3c in one donor and Genotype 3i in two others. On average, 43.5% of donors were immunoglobulin G-positive. Immunoglobulin G seroprevalence ranged from 22.7% to 60.8% in group ages 18 to 27 years and 48 to 57 years, respectively and differed between administrative regions from 28.9% in Podlasie to 61.3% in Wielkopolska. Thirty-nine of the donors were immunoglobulin M-positive, and seven donors were IgM positive only (0.2%). Of 37 immunoglobulin M-reactive samples tested by Western blot, 24 (64.9%) were confirmed. Conclusions The current results indicate a high level of HEV endemicity throughout Poland compared with other countries. There is an urgent need to consider the protection of recipients of blood components against transfusion-transmitted HEV infection.

Blood transfusion in Poland is the safest in history. High virological level of safety has been achieved mainly by improving not only the qualification of donors and methods used for donor screening, but also applying leukoreduction, pathogen reduction technology and grace period for serum.In this article, we discuss the improvement of the epidemic situation among blood donors for hepatitis B virus (HBV) and hepatitis C virus (HCV) and the increasing trend for HIV. Preliminary results of residual risk calculation for these pathogens are presented.Hepatitis E virus (HEV) and Babesia microti were considered as new factors potentially relevant for the safety of blood transfusion in our country. Due to evidence of West Nile virus (WNV) circulation in the ecosystem in Poland, it is also necessary to monitor the infections with this pathogen.In this article, it was emphasized that the reporting of all possible complications associated with transfusion and meticulous implementation of the look-back procedure play a key role for monitoring the risk of transmission of infectious agents by blood. It is especially important in view of the increasing epidemiological problems associated with emerging infectious agents.

International audience

Until now, markers of hepatitis E virus (HEV) infection have not been studied in blood donors throughout Poland, and no acute case of HEV infection has been closely documented or confirmed by HEV RNA detection. The prevalence of HEV infection markers, including HEV RNA in Polish blood donors and virus genotypes was investigated.In total, 12,664 individual donations from 22 Polish blood transfusion centers were tested for HEV RNA by transcription-mediated amplification. In addition, 3079 first-time donors sampled throughout Poland also were screened for antibodies to HEV. HEV RNA and immunoglobulin M-positive donations were confirmed using real-time reverse transcription-polymerase chain reaction and Western blotting, respectively.Ten donors were identified as RNA initial reactive (one of 1266 donors), and six (one of 2109) were identified as repeat reactive and confirmed by real-time reverse transcription-polymerase chain reaction or seroconversion. Sequence analysis identified HEV Genotype 3c in one donor and Genotype 3i in two others. On average, 43.5% of donors were immunoglobulin G-positive. Immunoglobulin G seroprevalence ranged from 22.7% to 60.8% in group ages 18 to 27 years and 48 to 57 years, respectively and differed between administrative regions from 28.9% in Podlasie to 61.3% in Wielkopolska. Thirty-nine of the donors were immunoglobulin M-positive, and seven donors were IgM positive only (0.2%). Of 37 immunoglobulin M-reactive samples tested by Western blot, 24 (64.9%) were confirmed.The current results indicate a high level of HEV endemicity throughout Poland compared with other countries. There is an urgent need to consider the protection of recipients of blood components against transfusion-transmitted HEV infection.

Blood transfusion in Poland is the safest in history. High virological level of safety has been achieved mainly by improving not only the qualification of donors and methods used for donor screening, but also applying leukoreduction, pathogen reduction technology and grace period for serum.In this article, we discuss the improvement of the epidemic situation among blood donors for hepatitis B virus (HBV) and hepatitis C virus (HCV) and the increasing trend for HIV. Preliminary results of residual risk calculation for these pathogens are presented.Hepatitis E virus (HEV) and

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

Blood donor screening of viral markers in Poland is based on serologic testing for anti-HCV, HBsAg, anti-HIV1/2 (chemiluminescence tests) and on nucleic acid testing (NAT) for RNA HCV, RNA HIV-1 and DNA HBV performed in minipools of 6 with real-time PCR (MPX 2.0 test on cobas s201) or with TMA in individual donations (Ultrio Plus or Ultrio Elite). Donors of plasma for anti-D and anti-HBs production are tested for parvovirus B19 DNA. Before implementation tests and equipment are evaluated at the Institute of Hematology and Transfusion Medicine (IHTM). The last 20 years witnessed a decreasing trend for HBsAg in both first time and repeat donors (1%-0.3% and 0.1%-0.02% respectively). Prevalence of anti-HCV repeat reactive results was stable and oscillated around 0.8% for first time donors and 0.2% for repeat donors. Elevated prevalence of seropositive HIV infected donors was recently observed (7.5-9 cases/100,000 donors). Since respective molecular markers implementation HCV RNA was detected on average in 1/119,235 seronegative donations, HIV RNA in 1/783,821 and HBV DNA in 1/61,047. HBV NAT yields were mostly occult hepatitis B (1/80,248); window period cases were less frequent (1/255,146). The efficiency of HBV DNA detection depends on the sensitivity of the HBV DNA screening system.

BACKGROUND The Ultrio Elite assay (Hologic/Grifols) runs on the Panther blood screening system and is comparable to the Ultrio Plus assay apart from the addition of oligonucleotides for human immunodeficiency virus Type 2 (HIV-2) detection. In this multicenter evaluation study the analytical sensitivity and genotype detection efficiency of the two assay versions were compared. STUDY DESIGN AND METHODS The analytical sensitivity and genotype detection efficiency were analyzed by replicate (18-303) testing of 27 hepatitis B virus (HBV), hepatitis C virus (HCV), HIV-1, and HIV-2 standard dilution panels calibrated in international units (IUs) and copies/mL. A wider range of subgenotypes was tested at 25 copies/mL. Specificity was evaluated in 30,756 donor samples. RESULTS The 95% lower limits of detection (LODs) in Ultrio Elite assay on WHO standards were 4.6, 7.3, 23.5, and 23.3 IU/mL for HBV, HCV, HIV-1, and HIV-2, respectively, and ranged from 13 to 44, 7 to 23, 6 to 15, and 9 copies/mL on genotype panels of the respective viruses. Comparable LODs had been previously found on the same panels with the Ultrio Plus assay. The specificity was 99.95% on initial test and 100% in the repeat test algorithm. CONCLUSION The change in the oligonucleotide design of the Ultrio Elite assay to enable HIV-2 detection has not affected the analytical sensitivity for the other viruses regardless of the genotype. Genotype reference panels are instrumental to compare the sensitivity of nucleic acid test assay versions and could serve as an alternative to seroconversion panels.

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

The study assessed the incidence of HBV markers (HBsAg, anti-HBc, anti-HBs) important for determination of the risk of reactivation of infection, with particular interest of occult infection (presence of HBV DNA in the absence of HBsAg) in patients treated at the Institute of Hematology and Transfusion Medicine. Anti-HBc frequency was correlated with the age and sex of patients. HBsAg was detected in 16/468 examined patients, 98/468 (21%) were anti-HBc positive. HBV DNA was detected in 41/98 anti-HBc positives; in 13 simultaneously with HBsAg. 28 patients had occult HBV infection (HBV DNA+/HBsAg). Antibody to HBsAg was detected in 163/430 (38%) patients, 81 out of them on protective level (100 IU/l). It was shown that occult HBV infection occurs in approximately 6% of patients. In most of them the protective levels of anti-HBs are detected.

Since 2004 Polish blood donors have been tested for parvovirus B19 (B19V) DNA. The screening testing has been performed in donors of plasma for fractionation and anti-D and anti-HBs production and donors of erythrocytes used for immunization. AIM is to present methods of the testing, quality control and results in period 2004-2010.Testing was performed in individual donation testing (IDT) in Regional Blood Transfusion Center (RBTC) in Lublin or in pools of 24 in Institute of Haematology and Transfusion Medicine in Warsaw (IHTM). Quantitative testing with real-time PCR was preceded with nucleic acid isolation on silica based methods (Prepito Viral DNA/RNA, Chemagen and QIAamp DNA Mini Kit, QIAGEN). Amplification was performed initially with home made method and later with commercial assay (Artus Parvo B19 RG PCR Kit on Rotor Gene 6 000). In total 17 625 donations were tested: 8 539 in pools and 9 090 individually. Beside routine external quality control programmes in which both laboratories participated (Proficiency Study VQC,Amsterdam, Holand; EQA Programe, Glasgow, Scotland), panel containing negative samples, positive with very high DNA B 19V level and plasma infected with genotype 2 was prepared for RBTC in Lublin.B19V infection frequency was 1:980 donations, low viraemic donations were detected most frequently (1:1 037). It was identified only one donation with DNA load that could cause potential health risk for plasma product recipients (1:17 625). In one of the donors B 19V DNA was observed for 3 years and 3 months. In acute or persistent phase of infection no clinical or laboratory symptoms (morphology of peripheral blood, ALT) were observed. Due to risk of underestimation of viral load connected with viral genome polymorphism all donations with B19V positive result were not allowed to be clinically used.

The results of HCV genotype distribution between 1995 and 2007 in Polish blood donors were analyzed. Special attention was drawn into the analysis of HCV polymorphism isolated from blood donors in the early phase of infection (RNA HCV positive/anti-HCV negative). Such donors identified in Poland since 2000 when the NAT for HCV was introduced are very important for molecular epidemiology analysis. The increasing frequency of 3a and 4 genotypes is observed in comparison to blood donors and patients with anti-HCV antibodies.