Outbreaks of autochthonous Dengue in Lazio region, Italy, August to September 2023: preliminary investigation.

Between August and September 2023, three distinct autochthonous dengue virus transmission events occurred in Lazio, Italy, with the main event in Rome. The events involved three different dengue serotypes. No link with previous imported cases was identified. Here we describe the epidemiological and phylogenetic analysis of the first autochthonous cases and the implemented control actions. The multiple transmission events call for a strengthening of the vector control strategies and future research to better characterise the risk in countries like Italy.

A forecasting model to estimate the drop in blood supplies during the SARS-CoV-2 pandemic in Italy.

OBJECTIVES: To estimate the number of actually Severe acute respiratory syndrome Coronavirus-2 (SARS-CoV-2) infected blood donors applying a statistical forecasting model. BACKGROUND: Following the outbreak of the SARS-CoV-2 epidemic, a drop in blood donation has been observed. It is crucial to determine the actual number of potential SARS-CoV-2-positive donors to define the measures and ensure adequate blood supply. METHODS: The cumulative incidence of SARS-CoV-2 positivity, calculated on the general population, was applied to the donor population by estimating the number of positive subjects. The calculation model was validated by the linear interpolation method. The number of blood units actually discarded based on post-donation information was also taken into account. RESULTS: Three months after the outbreak, 5322 donors were estimated to be positive for SARS-CoV-2 and were therefore potentially excluded from donation. A total of units of blood components were discarded following post donation information. The estimated number of donors deceased (180) and the number of clinically recovered individuals in the same period was also considered. CONCLUSION: This forecasting model can be used to obtain information on blood donors' involvement during future SARS-CoV-2 outbreaks, especially in case of changes concerning epidemiology, incidence by age bracket and geographical distribution and also for new outbreaks of emerging viruses.

Inhibitors in Patients with Congenital Bleeding Disorders Other Than Hemophilia.

The most worrying complication of replacement therapy for severe hemophilia A and B is currently the occurrence of inhibitory alloantibodies against infused factor VIII and factor IX, respectively. Inhibitors compromise the management of hemorrhage in affected patients, with a considerable increase in complications, disability, and costs. While these alloantibodies have been extensively studied in the past years in hemophilia A and B, those occurring in patients with other inherited bleeding disorders are less well characterized and still poorly understood, mostly due to the rarity of these hemorrhagic conditions. This narrative review will deal with inhibitors arising in patients with inherited bleeding disorders other than "classical" hemophilia, focusing in particular on those developing in patients with congenital deficiency of coagulation factor V, factor VII, factor XI, and factor XIII.

Emergency response of four transfusion centers during the last Chikungunya outbreak in Italy.

Between September 15 and November 11, 2017, the area of Rome was subjected to cessation of donation in an area of approximately 1,300,000 inhabitants and with a 5-day quarantine for the rest of metropolitan areas due to Chikungunya virus (CHIKV) outbreak. Quarantine was applied to red blood cell (RBC) and plasma units while platelet concentrates (PCs) were subjected to pathogen inactivation methods (PIMs). Quarantine was based on an active recall of all donors and, in case of declared absence of any symptom or illness, on release of RBC and plasma units after 5 days. Four regional centers in which PIMs were already performed were charged for PIMs for the rest of Rome's area. These centers increased by 236% their previous PIM procedures, producing additional 1425 pathogen-reduced (PR) PCs in 57 days, beside their production (996 PR PCs) for local needs. The emergency support was close to the maximal production capacity of the four centers and was successful only thanks to the limited length of emergency. No adverse events were reported by all centers through a passive hemovigilance approach and by a follow-up of approximately 4 months. None of the donors referred symptoms or illness at recall and none of the blood products were discarded after quarantine. To date, no cases of CHIKV-transmitted infection were reported in transfused patients. This experience has taught some relevant strategic and organizing aspects that should be taken into account when an infectious outbreak must be faced in a large metropolitan area.

The Role of ABO Blood Type in Thrombosis Scoring Systems.

In addition to their major role in transfusion medicine, there is increasing evidence that ABO blood group antigens (complex carbohydrate molecules widely expressed on the surface of red blood cells and several other cell types) are implicated in the development of a wide array of pathologic conditions. In particular, intense research has been dedicated over the last 50 years to the study of the association between non-O blood type and the risk of developing cardiovascular disorders. Several pathways have been hypothesized to explain this relationship, the most reasonable implying the influence of the ABO blood group on circulating plasma levels of von Willebrand factor, factor VIII, and several inflammatory cytokines. This narrative review summarizes the current knowledge on the role of ABO antigens in both venous and arterial thromboses, focusing on their association with clinical scoring systems evaluating thrombotic risk.

Clinical effectiveness of platelets in additive solution treated with two commercial pathogen-reduction technologies.

BACKGROUND: Two noninferiority, randomized, controlled trials were conducted in parallel comparing the safety and efficacy of platelets treated with Intercept or Mirasol pathogen-reduction technologies versus standard platelets. STUDY DESIGN AND METHODS: The primary endpoint was the percentage of hematology patients who developed World Health Organization Grade 2 or greater bleeding. A noninferiority margin of 11% was chosen based on expected Grade 2 or greater bleeding in 20% of controls. The study was closed for financial restrictions before reaching the planned sample size of 828 patients, and an intention-to-treat analysis was conducted on 424 evaluable patients. RESULTS: In the Intercept trial (113 treated vs. 115 control patients), the absolute risk difference in Grade 2 or greater bleeding was 6.1%, with an upper one-sided 97.5% confidence limit of 19.2%. The absolute risk difference in the Mirasol trial (99 treated vs. 97 control patients) was 4.1%, and the upper one-sided 97.5% confidence limit was 18.4%. Neither absolute risk difference was statistically significant. In both trials, posttransfusion platelet count increments were significantly lower in treated versus control patients. Mean blood component use in treated patients versus controls was 54% higher (95% confidence interval, 36%-74%; Intercept) and 34% higher (95% confidence interval, 16%-54%; Mirasol) for platelets and 23% higher (95% confidence interval, 8%-39%; Intercept) and 32% higher (95% confidence interval, 10%-57%; Mirasol) for red blood cells. Unexpected reactions and adverse events were not reported. Mortality did not differ significantly between treated and control patients. CONCLUSION: Although conclusions on noninferiority could not be drawn due to low statistical power, the study provides additional information on the safety and efficacy of pathogen-reduced platelets treated with two commercial pathogen-reduction technologies.

Human T-lymphotropic virus and transfusion safety: does one size fit all?

Human T-cell leukemia viruses (HTLV-1 and HTLV-2) are associated with a variety of human diseases, including some severe ones. Transfusion transmission of HTLV through cellular blood components is undeniable. HTLV screening of blood donations became mandatory in different countries to improve the safety of blood supplies. In Japan and Europe, most HTLV-infected donors are HTLV-1 positive, whereas in the United States a higher prevalence of HTLV-2 is reported. Many industrialized countries have also introduced universal leukoreduction of blood components, and pathogen inactivation technologies might be another effective preventive strategy, especially if and when generalized to all blood cellular products. Considering all measures available to minimize HTLV blood transmission, the question is what would be the most suitable and cost-effective strategy to ensure a high level of blood safety regarding these viruses, considering that there is no solution that can be deemed optimal for all countries.

A survey on the implementation of Patient Blood Management programs in Italy.

BACKGROUND: Since 2012, in line with the World Health Organization (WHO) resolution WHA63.12 of 05/21/2010, the Italian National Blood Center has been promoting patient blood management (PBM). In order to verify the level of PBM implementation nationwide, we submitted a survey to all healthcare providers. MATERIAL AND METHODS: In line with what was proposed in the international scientific literature in the field, a series of indicators were used derived from the four main blocks related to PBM strategies: the management of patient anemia; the optimization of hemostasis; blood conservation strategies; patient-centred decision-making. We also added two blocks containing important information on general PBM management and other PBM-related aspects. RESULTS: The survey showed good implementation of anemia screening programs in accordance with the timelines established by national and international guidelines, and the single unit policy is used in line with national guideline recommendations. However, the survey also revealed limited auditing of PBM programs and reduced monitoring and reporting of clinical outcomes and indicators. DISCUSSION: The first national survey on the level of PBM implementation in Italy shows widespread adoption of diagnostic-therapeutic care pathways aimed at the diagnosis and treatment of anemia in the perioperative setting.

Concise review: stem cell-derived erythrocytes as upcoming players in blood transfusion.

Blood transfusions have become indispensable to treat the anemia associated with a variety of medical conditions ranging from genetic disorders and cancer to extensive surgical procedures. In developed countries, the blood supply is generally adequate. However, the projected decline in blood donor availability due to population ageing and the difficulty in finding rare blood types for alloimmunized patients indicate a need for alternative red blood cell (RBC) transfusion products. Increasing knowledge of processes that govern erythropoiesis has been translated into efficient procedures to produce RBC ex vivo using primary hematopoietic stem cells, embryonic stem cells, or induced pluripotent stem cells. Although in vitro-generated RBCs have recently entered clinical evaluation, several issues related to ex vivo RBC production are still under intense scrutiny: among those are the identification of stem cell sources more suitable for ex vivo RBC generation, the translation of RBC culture methods into clinical grade production processes, and the development of protocols to achieve maximal RBC quality, quantity, and maturation. Data on size, hemoglobin, and blood group antigen expression and phosphoproteomic profiling obtained on erythroid cells expanded ex vivo from a limited number of donors are presented as examples of the type of measurements that should be performed as part of the quality control to assess the suitability of these cells for transfusion. New technologies for ex vivo erythroid cell generation will hopefully provide alternative transfusion products to meet present and future clinical requirements.

Transcriptomic and phospho-proteomic analyzes of erythroblasts expanded in vitro from normal donors and from patients with polycythemia vera.

Erythropoiesis is a tightly regulated process which becomes decoupled from its normal differentiation program in patients with polycythemia vera (PV). Somatic mutations in JAK2 are commonly associated with this myeloid proliferative disorder. To gain insight into the molecular events that are required for abnormally developing erythroid cells to escape dependence on normal growth signals, we performed in vitro expansion of mature erythroblasts (ERY) from seven normal healthy donors and from seven polycythemic patients in the presence of IL3, EPO, SCF for 10, 11, or 13 days. Normal ERYs required exposure to the glucocorticoid dexamethasone (Dex) for expansion, while PV-derived ERYs expanded in the absence of dexamethasone. RNA expression profiling revealed enrichment of two known oncogenes, GPR56 and RAB4a, in PV-derived ERYs along with reduced expression levels of transcription factor TAL1 (ANOVA FDR < 0.05). While both normal and polycythemic-derived ERYs integrated signaling cascades for growth, they did so via different signaling pathways which are represented by their differential phospho-profiles. Our results show that normal ERYs displayed greater levels of phosphorylation of EGFR, PDGFRß, TGFß, and cKit, while PV-derived ERYs were characterized by increased phosphorylation of cytoplasmic kinases in the JAK/STAT, PI3K, and GATA1 pathways. Together these data suggest that PV erythroblast expansion and maturation may be maintained and enriched in the absence of dexamethasone through reduced TAL1 expression and by accessing additional signaling cascades. Members of this acquired repertoire may provide important insight into the pathogenesis of aberrant erythropoiesis in myeloproliferative neoplasms such as polycythemia vera.

A survey on preoperative autologous blood donation policy in bone marrow stem cell donors in Italy.

BACKGROUND: The high safety of homologous blood components, together with the introduction of the Patient Blood Management strategy, has led to the progressive abandonment of preoperative autologous blood donation (PAD) in surgery. Furthermore, recent scientific publications provide evidence about the non-usefulness of PAD in the collection of hematopoietic stem cells (HSC) from bone marrow (BM), also in consideration of harvest procedure safety. Nevertheless, no conclusive studies have been published yet. MATERIALS AND METHODS: Blood Establishments (BE) and Bone Marrow Collection Centers (BMCC) participated in a specific qualitative survey proposed by Italian National Blood and Transplant centers with the support of the relevant Italian Scientific Societies. The survey aimed at evaluating the policy adopted for PAD in related and unrelated adult HSC donors in Italy during the period 2018-2020. RESULTS: Forty-one BE corresponding to 37 BMCC filled in the questionnaire. Of 830 BM donors, 661 (80%) underwent 1063 PAD (mean 1.6 PAD/donor). The remaining 169 donors (20%) underwent BM harvest without PAD. No serious adverse events were reported for either donor group. In the case of ineligibility of donors for the PAD program, due to low hemoglobin values, 7/10 centers shifted donors to peripheral blood stem cell collection and three centers chose a different donor. Remarkably, only 51% of the PAD units requested were eventually transfused during the BM harvest process. Finally, the iron support policy among centers was heterogeneous. DISCUSSION: The results of this survey show that PAD is heterogeneously applied in Italian BMCC, as in other countries. However, all BMCC except two are willing to adopt a Patient Blood Management strategy as an alternative approach to adult related and unrelated BM donor harvests.

Time-course investigation of SAGM-stored leukocyte-filtered red bood cell concentrates: from metabolism to proteomics.

BACKGROUND: Results from recent, highly debated, retrospective studies raised concerns and prompted considerations about further testing the quality of long stored red blood cells from a biochemical standpoint. DESIGN AND METHODS: We performed an integrated mass spectrometry-based metabolomics and proteomics time-course investigation on SAGM-stored red blood cells. In parallel, structural changes during storage were monitored through scanning electron microscopy. RESULTS: We detected increased levels of glycolytic metabolites over the first 2 weeks of storage. From day 14 onwards, we observed a significant consumption of all metabolic species, and diversion towards the oxidative phase of the pentose phosphate pathway. These phenomena coincided with the accumulation of reactive oxygen species and markers of oxidation (protein carbonylation and malondialdehyde accumulation) up to day 28. Proteomics evidenced changes at the membrane protein level from day 14 onwards. Changes included fragmentation of membrane structural proteins (spectrin, band 3, band 4.1), membrane accumulation of hemoglobin, anti-oxidant enzymes (peroxiredoxin-2) and chaperones. While the integrity of red blood cells did not show major deviations at day 14, at day 21 scanning electron microscope images revealed that 50% of the erythrocytes had severely altered shape. We could correlate the scanning electron microscopy observations with the onset of vesiculation, through a proteomics snapshot of the difference in the membrane proteome at day 0 and day 35. We detected proteins involved in vesicle formation and docking to the membrane, such as SNAP alpha. CONCLUSIONS: Biochemical and structural parameters did not show significant alterations in the first 2 weeks of storage, but then declined constantly from day 14 onwards. We highlighted several parallelisms between red blood cells stored for a long time and the red blood cells of patients with hereditary spherocytosis.

Lack of Evidence of Chikungunya Virus Infection among Blood Donors during the Chikungunya Outbreak in Lazio Region, Italy, 2017.

BACKGROUND: The latest European Chikungunya virus (CHIKV) outbreak occurred in Italy in 2017, in the municipalities of Anzio and Rome (Lazio Region), with a secondary outbreak in the Calabrian Region. Most CHIKV infections are symptomatic but about 15% of people who acquire the infection may be asymptomatic. A retrospective study was conducted with the aim of assessing the prevalence of recent/ongoing CHIKV infections on the blood donor population in the Lazio Region, during the 2017 outbreak (including in the period before it was detected). METHODS: The study was conducted on 4595 plasma samples from donors who donated in 14 different Blood Establishments in the Lazio Region, in the period June-November 2017. A total of 389 of these samples were collected in provinces not affected by the outbreak and were used as negative controls. All samples were tested for IgM detection by the use of an ELISA test, and positive samples were tested for confirmation through the use of a PRNT. Molecular tests were performed on sera that were found to be IgM-positive or borderline. RESULTS: A total of 41 (0.89%) blood donors tested positive for IgM. None of these positive IgM ELISA results was confirmed either by PRNT or by molecular tests. CONCLUSIONS: Our study has shown no evidence of recent/ongoing CHIKV infection in blood donors of the affected area.

Plasma-derived clotting factor VIII: heterogeneity evaluation in the quest for potential inhibitory-antibody stimulating factors.

In this study, we investigated the heterogeneity and the purity grade of three commercially available plasma-derived clotting factor VIII (FVIII) concentrates, which highly differ with regard to purification strategies, relative concentrations of stabilizers (von Willebrand factor, with or without albumin) and virus inactivation strategies (solvent/detergent and/or heat/pasteurization treatments). Western blot analyses were used to evaluate product-specific variations from Emoclot(®) , Alphanate(®) and Haemate(®) both in the presence and absence of reducing agents (dithiotreithol). All the plasma-derived concentrates showed a strong heterogeneity, as they all included a significant amount of truncated forms of the full-length (FL) clotting FVIII protein. The intact protein accounted for the 38% of the total FVIII proteins in Haemate(®) and 29 and 23% in Alphanate(®) and Emoclot(®) , respectively. Lower intact FVIII amounts in Emoclot might be mainly due to the low von Willebrand factor dosage and the absence of albumin. Upon addition of thrombin, both the FL and truncated forms of the FVIII protein were almost completely digested. Indeed, after thrombin activation, we could still observe a mixture of B-domain truncated forms of the FL protein along with biologically active digested-A1 forms. Batch-to-batch variation was tested with no evident changes appearing among different batches. Despite the variables in manufacturing processes, inter-product comparisons yielded similar results for all the plasma-derived FVIII considered in this study. However, we could individuate in Emoclot a band that was not digested by thrombin, which we could characterize as the 200 kDa FVIII heavy chain. This investigation prompts new concerns about the strong heterogeneity observed upon thrombin digestion of plasma-derived FVIII, which might contribute to the development of inhibitory antibodies at an early stage of therapy, and to which extent these untoward phenomena could be avoided through direct intervention on routine manufacturing processes.

Peroxiredoxin-2 as a candidate biomarker to test oxidative stress levels of stored red blood cells under blood bank conditions.

BACKGROUND: Several researches on aging red blood cells (RBCs)--performed both in vivo and under blood bank conditions--revealed that RBC membrane proteins undergo a number of irreversible alterations, mainly due to oxidative stress. The individuation of proteins to be used as indicators of irreversible RBC injury and to be proposed as candidate biomarkers of oxidative damage or aging status during blood storage is therefore of great interest. STUDY DESIGN AND METHODS: Based on this purpose we performed proteomic analysis of the membranes of RBCs during various storage periods under blood bank conditions. Changes in protein composition of RBC membranes were monitored as a function of the storage period by means of polyacrylamide gel electrophoresis coupled with immunoblotting and mass spectrometry analyses. RESULTS: During storage, a progressive linkage of typical cytosolic proteins to the membrane was detected, including both antioxidant and metabolic enzymes (such as catalase, peroxiredoxin-2 [Prx2], and 2,3-bisphosphoglycerate-mutase), as well as nonreducible cross-linkings of probably oxidized or denatured hemoglobin. This phenomenon was unequivocally related to oxidative stress, since storage of RBCs under anaerobic conditions showed a suppression of these protein recruitments to the membrane. CONCLUSION: The detailed analysis of these protein associations to the membrane of aged RBCs allowed Prx2 to be suggested as a potential RBC oxidative stress marker for the sake of developing new approaches in quality assurance of blood components.

Proteomic analysis of plasma derived from platelet buffy coats during storage at room temperature. An application of ProteoMiner™ technology.

The present study was aimed at revealing new insights into the analysis of storage-related processes occurring in the supernatants of platelet concentrates (PCs) derived from pooled buffy coats suspended in whole plasma. To reduce the dynamic range of plasma protein concentrations and access low-abundance proteins, we made use of a solid-phase combinatorial peptide ligand library, known under the trade name of ProteoMiner™. Afterwards, two-dimensional electrophoresis (2-DE) was coupled with mass spectrometry (MS) to reveal changes in proteomic profiles. Several storage-induced protein alterations were identified including changes to major plasma proteins. In particular, a precursor of the secretory form of clusterin was shown to accumulate during storage of PC supernatants, together with platelet-derived tropomyosin, suggesting a progressive loss of platelet integrity. Platelet-released proteins following activation have also been detected (alpha-1-B-glycoprotein, kininogen-1, and serpin proteinase inhibitor 8). Moreover, specific protein fragments (vitronectin, plakoglobin, hornerin, and apolipoprotein A-IV) were found to be modulated upon storage, possibly indicating a time-dependent buffy-coat PC deterioration. Globally, our findings provided the disclosure of unique proteins in PC supernatants with respect to previous studies conducted in similar experimental conditions, suggesting ProteoMiner enrichment technology to be a possible complementary tool in the identification of diagnostically relevant proteins as age/quality biomarkers of therapeutic products.

The use of whole blood in traumatic bleeding: a systematic review.

Hemostatic resuscitation is currently considered a standard of care for the management of life-threatening hemorrhage, but in some critical settings the access to high quantities of blood components is problematic. Whole blood (WB) transfusion has been proposed as an alternative modality for hemostatic resuscitation of traumatic major bleeding. To assess the efficacy and safety of WB in trauma-associated massive bleeding, we performed a systematic review of the literature. We selected studies comparing WB transfusions to transfusion of blood components (COMP) in massive trauma bleeding; both randomized clinical trial (RCT) and observational studies were considered. The outcomes were mortality (30-day/in-hospital and 24-h mortality) and adverse events/transfusion reactions. The effect sizes were crude odds ratio (OR), adjusted OR and hazard ratio (HR). The methodological quality of studies was assessed using the Cochrane Risk of Bias tool for RCTs, and the ROBIN-1 tool for observational studies. The overall quality of the available evidence was assessed with the GRADE system. One RCT (2 reports) and 6 cohort studies were included (3642 adult patients; 675 receiving WB, 2967 receiving COMP). Three studies were conducted in military setting, and 4 in civilian setting. In the overall analysis, 30-day/in-hospital and 24-h mortality did not differ significantly between groups (very low quality of the evidence due to high risk of bias, imprecision and inconsistency). After adjustment for baseline covariates in three cohort studies, the OR for mortality was significantly lower in WB recipients compared to COMP (OR 0.22; 95% CIs 0.10/0.45) (moderate grade of evidence). Adverse events and transfusion reactions were overlooked and not consistently reported. The available evidence does not allow to draw definite conclusions on the short-term and long-term efficacy and safety of WB transfusion compared to COMP transfusion. Further well designed research is needed.

West Nile and Usutu viruses co-circulation in central Italy: outcomes of the 2018 integrated surveillance.

BACKGROUND: West Nile (WNV) and Usutu (USUV) are emerging vector-borne zoonotic flaviviruses. They are antigenically very similar, sharing the same life cycle with birds as amplification host, Culicidae as vector, and man/horse as dead-end host. They can co-circulate in an overlapping geographic range. In Europe, surveillance plans annually detect several outbreaks. METHODS: In Italy, a WNV/USUV surveillance plan is in place through passive and active surveillance. After a 2018 WNV outbreak, a reinforced integrated risk-based surveillance was performed in four municipalities through clinical and serological surveillance in horses, Culicidae catches, and testing on human blood-based products for transfusion. RESULTS: Eight WNV cases in eight equine holdings were detected. Twenty-three mosquitoe catches were performed and 2367 specimens of Culex pipiens caught; 17 pools were USUV positive. A total of 8889 human blood donations were tested, and two asymptomatic donors were USUV positive. CONCLUSIONS: Different surveillance components simultaneously detected WNV only in horses and USUV only in humans and mosquitoes. While in endemic areas (i.e. northern Italy) entomological surveillance is successfully used as an early detection warning, this method in central Italy seems ineffective. To achieve a high level of sensitivity, the entomological trapping effort should probably exceed a reasonable balance between cost and performance. Besides, WNV/USUV early detection can be addressed by horses and birds. Further research is needed to adapt the surveillance components in different epidemiological contexts.

COVID-19-associated coagulopathy.

Coronavirus disease 2019 (COVID-19), a viral respiratory illness caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has been recently recognized as a systemic disorder inducing a prothrombotic state. The molecular mechanisms underlying the hypercoagulable state seen in patients with COVID-19 is still incompletely understood, although it presumably involves the close link between inflammatory and hemostatic systems. The laboratory coagulation monitoring of severely ill COVID-19 patients is mandatory to identify those patients at increased thrombotic risk and to modulate thromboprophylaxis accordingly. In this review, we summarize the current understanding on the pathogenesis, epidemiology, clinical and laboratory features and management of coagulopathy associated with COVID-19.