Investigating the Role of Anti-TPO Antibodies in HIV-Associated Thrombocytopenia before and after Initiation of HAART: A Case-Control Longitudinal Study.

This longitudinal, case-control study aimed to investigate the role of thrombopoietin (TPO) and anti-TPO antibodies in HIV-associated thrombocytopenia, focusing on the changes seen before and after the initiation of highly active antiretroviral therapy (HAART). Patients were assessed before and at least six months after the initiation of HAART. In total, 75 PLWHIV (age/sex-matched and randomized at 2:1, according to thrombocytopenia status) were included in this study. The baseline assessment revealed significantly higher TPO levels in thrombocytopenic patients (140.45 vs. 106.8 mg/mL, p = 0.008). Furthermore, anti-TPO-positive patients displayed lower platelet counts (109,000 vs. 139,000/L, p = 0.002) and TPO levels (114.7 vs. 142.7 mg/mL, p = 0.047). Longitudinally, HAART initiation reduced the frequency of thrombocytopenia from 75.47% to 33.96% (p < 0.001) and elevated the median platelet counts from 131,000 to 199,000 (p < 0.001). No significant difference in median platelet counts was found post-HAART among the anti-TPO subgroups (p = 0.338), a result contrasting with pre-HAART findings (p = 0.043). Changes in anti-TPO status corresponded with significant platelet count alterations (p = 0.036). Notably, patients who became anti-TPO negative showed a median increase of 95,000 platelets (IQR: 43,750-199,500). These marked differences between subgroups underscore the potential role of anti-TPO antibodies in modulating the hematological response to HAART. Further research is needed to elucidate the complex interplay between HIV infection, HAART, and thrombocytopenia.

Detection of Virulence-Associated Genes among Brucella melitensis and Brucella abortus Clinical Isolates in Greece, 2001-2022.

Brucellosis remains an important zoonotic disease in several parts of the world; in Greece, although it is declining, it is still endemic, affecting both the financial and public health sectors. The current study was undertaken to investigate the presence and distribution of virulence-associated genes among Brucella spp. clinical strains isolated during 2001-2022. Species identification was performed using conventional methodology and Bruce-ladder PCR. The presence of the virulence genes mviN, manA, wbkA, perA, omp19, ure, cbg and virB was investigated using PCR. During the study period, a total of 334 Brucella isolates were identified, of which 328 (98.2%) were detected from positive blood cultures; 315 (94.3%) of the isolates were identified as B. melitensis, whilst the remaining 16 (4.8%) and 3 (0.9%) were identified as B. abortus and B. suis, respectively. Notably, two of the B. melitensis were assigned to the REV-1 vaccine strain type. The presence of the omp19, manA, mviN and perA genes was confirmed in all 315 B. melitensis isolates, while ure, wbkA, cbg and virB genes were detected in all but 9, 2, 1 and 1 of the isolates, respectively. All eight virulence genes were amplified in all B. abortus and B. suis isolates. The detection rate of virulence genes did not differ significantly among species. In conclusion, brucellosis is still considered a prevailing zoonotic disease in Greece, with the majority of the isolates identified as B. melitensis. The eight pathogenicity-associated genes were present in almost all Brucella isolates, although the ure gene was absent from a limited number of B. melitensis isolates.

Serum ß-Defensin 2, A Novel Biomarker for the Diagnosis of Acute Infections.

BACKGROUND: Defensins are natural antimicrobial peptides that the human body secretes to protect itself from an infection. Thus, they are ideal molecules to serve as biomarkers for infection. This study was conducted to evaluate the levels of human ß-defensins in patients with inflammation. METHODS: CRP, hBD2 and procalcitonin were measured in 423 sera of 114 patients with inflammation and healthy individuals using nephelometry and commercial ELISA assays. RESULTS: Levels of hBD2 in the serum of patients with an infection were markedly elevated compared to those of hBD2 in patients with inflammation of non-infectious etiology (p < 0.0001, t = 10.17) and healthy individuals. ROC analysis demonstrated that hBD2 showed the highest detection performance for infection (AUC 0.897; p < 0.001) followed by PCT (AUC 0.576; p = ns) and CRP (AUC 0.517; p = ns). In addition, analysis of hBD2 and CRP in patients' sera collected at different time points showed that hBD2 levels could help differentiate inflammation of infectious and non-infectious etiology during the first 5 days of hospitalization, while CRP levels could not. CONCLUSIONS: hBD2 has the potential to serve as a diagnostic biomarker for infection. In addition, the levels of hBD2 may reflect the efficacy of antibiotic treatment.

Comparative Characterization of Human Antibody Response Induced by BNT162b2 Vaccination vs. SARS-CoV-2 Wild-Type Infection.

Humoral immunity after SARS-CoV-2 immunization or natural infection is thought to be evanescent. In our study, we aimed to longitudinally characterize the kinetics of antibody titers after dual BNT162b2 immunization or wild-type infection. Vaccinated and recovered individuals displayed distinct antibody kinetics, as convalescents had detectable RBD-, S1-specific, and neutralizing IgG antibody titers two weeks post-infection that gradually increased longitudinally, while RBD-, S1-specific, and neutralizing IgG were detected in vaccinees after the first dose, increased significantly 3 weeks post the second dose and decreased significantly 4-5 months thereafter. Neutralizing IgG was significantly higher initially in convalescent individuals; however, vaccines displayed significantly higher neutralizing antibodies 4-5 months post the second dose. In both groups, there was a strong negative association between elapsed time and antibody levels. The avidity of anti-RBD antibody titers increased significantly in patients longitudinally, while in vaccinees initially increased, with subsequent decrease, remaining however higher than antibody avidity of recovered individuals at all time-points. Anti-RBD antibodies were strongly correlated with neutralizing and anti-S1 antibodies in both groups at all time-points. This study facilitates our further understanding of immune response to SARS-CoV-2 and vaccines.

Development of an Enzyme-Linked Immunosorbent Assay (ELISA) for Accurate and Prompt Coronavirus Disease 2019 (COVID-19) Diagnosis Using the Rational Selection of Serological Biomarkers.

Prompt COVID-19 diagnosis is urgently required to support infection control measures. Currently available serological tests for measuring SARS-CoV-2 antibodies use different target antigens, although their sensitivity and specificity presents a challenge. We aimed to develop an "in-house" serological ELISA to measure antibodies against SARS-CoV-2 by combining different protein antigens. Sera (n = 44) from COVID-19-confirmed patients were evaluated against different SARS-CoV-2 protein antigens and all potential combinations using ELISA. Patients' sera were also evaluated against commercially available ELISA diagnostic kits. The mixture containing RBD 2.5 µg/mL, S2 1 µg/mL and N 1.5 µg/mL was found to be the most potent. Plates were incubated with patients' sera (1:100), and goat anti-human alkaline phosphatase-conjugated IgG, ΙgM and IgA antibody was added. The cut-off value for each assay was determined using the mean optical density plus two standard deviations of pre-pandemic controls. The "in-house" ELISA displayed 91% sensitivity and 97% specificity for IgG antibodies, whereas its sensitivity and specificity for IgM and IgA were 75% and 95% and 73% and 91%, respectively. The "in-house" ELISA developed here combined three SARS-CoV-2 antigens (RBD, S2 and N) as capture antigens and displayed comparable and even higher sensitivity and specificity than otherwise quite reliable commercially available ELISA diagnostic kits.

Diagnostic performance of rapid antigen tests (RATs) for SARS-CoV-2 and their efficacy in monitoring the infectiousness of COVID-19 patients.

The most widely used test for the diagnosis of SARS-CoV-2 infection is a PCR test. PCR has very high sensitivity and is able to detect very low amounts of RNA. However, many individuals receiving a positive test result in a context of a PCR-based surveillance might be infected with SARS-CoV-2, but they are not contagious at the time of the test. The question arises regards if the cost effective, portable rapid antigen tests (RATs) have a better performance than PCR in identification of infectious individuals. In this direction, we examined the diagnostic performance of RATs from 14 different manufacturers in 400 clinical samples with known rRT-PCR cycles threshold (cT) and 50 control samples. Substantial variability was observed in the limit of detection (LOD) of different RATs (cT = 26.8-34.7). The fluorescence-based RAT exhibited a LOD of cT = 34.7. The use of the most effective RATs leads to true positive rates (sensitivities) of 99.1% and 90.9% for samples with cT ≤ 30 and cT ≤ 33, respectively, percentages that can guarantee a sensitivity high enough to identify contagious patients. RAT testing may also substantially reduce the quarantine period for infected individuals without compromising personal or public safety.

Distribution of MICA alleles and haplotypes associated with HLA-B in Greek population.

INTRODUCTION: The Major Histocompatibility Complex Class I-related chain A gene (MICA) is a highly polymorphic functional gene located close to the HLA-B locus. Certain MICA alleles have been related to inflammatory and autoimmune diseases while MICA antibodies have been implicated in organ allograft rejection or graft-versus-host disease (GVHD). AIM: The aim of this study was to identify the frequencies of MICA alleles and MICA ~ HLA-B haplotypes in the Greek population since, as far as we know, these data are still limited. METHODS: DNA was obtained from 277 unrelated healthy Greek individuals of Caucasian origin, volunteer donors of blood stem cells. HLA-B* and MICA* genotyping was performed by reverse PCR-SSOP. RESULTS: A total of 18 MICA alleles were defined in the present study. The five most frequent alleles in the Greek population were MICA*008 (24.6%), MICA*009 (22.36%), MICA*018 (16.03%), MICA*002 (8.02%) and MICA*004 (7.17%) which altogether account for 77.8% of all alleles. The most common MICA ~ HLA-B haplotypes were MICA*018 ~ B*18 (12.5%) and MICA*009 ~ B*51(11.5%). CONCLUSIONS: The five most frequent MICA alleles in the Greek population were *008, *009, *018, *002, *004. In other Caucasian populations, two of these alleles (*008, and *004) were observed in similar frequencies. MICA*002 was observed less frequently (8.02%) in the Greek population compared to other Caucasian groups (frequencies > 15%). Also, MICA*009 and MICA*018 had elevated frequencies (above 15%) whereas in other Caucasian populations they were found around 10% or less. These data may be important for the elucidation of the role that MICA polymorphisms play in organ and stem cell transplantation and to identify the relation of certain MICA with susceptibility to specific diseases.

Major vault protein/lung resistance related protein: a novel biomarker for rheumatoid arthritis.

OBJECTIVES: Rheumatoid arthritis (RA) can lead to joint destruction and early institution of effective treatment can preserve joint function. Biomarkers can establish early diagnosis and predict effect of treatment. Vault particles, large cytoplasmic ribonucleoprotein particles that participate in inflammation, might serve as biomarkers. The aim of this study was to assess the diagnostic and the prognostic value of major vault protein (MVP) and their antibodies in RA. METHODS: Serum samples from 159 RA patients, 26 early RA (ERA) patients, 21 patients with osteoarthritis (OA) and 30 healthy individuals were tested for MVP, anti-cyclic citrullinated peptide (anti-CCP) and C-reactive protein (CRP) using enzyme-linked immunosorbent assays (ELISA). Rheumatoid factor (RF) was tested by nephelometry, and anti-MVP antibodies were detected by anti-MVP peptide ELISA using an in-house protocol. RESULTS: MVP levels were higher in RA and ERA, compared to OA and healthy controls (p<0.00001). A combination of MVP with RF or anti-CCP showed an improved diagnostic accuracy compared to RF or anti-CCP alone in RA and ERA. MVP exhibited similar AUC levels to anti-CCP and RF in RA whereas in ERA, MVP exhibited the same or slightly higher AUC levels, compared to anti-CCP and RF, respectively. High MVP levels were associated with lack of response to treatment. Levels of anti-MVP peptide 2 antibodies were significantly higher in RA compared to healthy controls (t= 2.73, p=0.007). CONCLUSIONS: MVP and autoantibodies against MVP may have the potential to serve as diagnostic and prognostic biomarkers in RA.

Development of a new spectrophotometric assay for rapid detection and differentiation of KPC, MBL and OXA-48 carbapenemase-producing Klebsiella pneumoniae clinical isolates.

The increased prevalence of carbapenemase-producing Enterobacteriaceae (CPE) has made essential the design of quicker tests for CPE detection. In the present study, a simple and rapid assay was developed based on measurement of the hydrolytic activity of imipenem at a final concentration of 65 µg/mL (100 µM) through ultraviolet-visible (UV-Vis) spectrophotometry. All measurements were conducted at 297 nm. A total of 83 carbapenem-non-susceptible CPE, consisting of Klebsiella pneumoniae clinical isolates and genotypically characterised as KPC-, VIM-, NDM- or OXA-48-producers, were tested. For comparison, 30 carbapenem-non-susceptible clinical isolates, consisting of Escherichia coli and K. pneumoniae and genotypically confirmed as non-CPE, were also examined. The spectrophotometric assay enabled efficient discrimination of CPE from non-CPE isolates even in 45 min (P < 0.0001). Moreover, the presence of phenylboronic acid (PBA) or ethylene diamine tetra-acetic acid (EDTA) in the reaction mixture was able to inhibit the hydrolytic capacity of KPC- or metallo-ß-lactamase (MBL)-producers, respectively, while the hydrolytic activity of OXA-48-producing strains was not affected by the presence of these inhibitors (P < 0.001). The newly developed assay presented 100% sensitivity and specificity to detect and differentiate KPC-, MBL- and OXA-48-producers compared with genotypic characterisation. Thus, the proposed spectrophotometric method can be considered as an easy, fast, accurate and cost-effective diagnostic tool for screening carbapenem-non-susceptible K. pneumoniae isolates in the clinical laboratory.

Novel Protein-Based Pneumococcal Vaccines: Assessing the Use of Distinct Protein Fragments Instead of Full-Length Proteins as Vaccine Antigens.

Non-serotype-specific protein-based pneumococcal vaccines have received extensive research focus due to the limitations of polysaccharide-based vaccines. Pneumococcal proteins (PnPs), universally expressed among serotypes, may induce broader immune responses, stimulating humoral and cellular immunity, while being easier to manufacture and less expensive. Such an approach has raised issues mainly associated with sequence/level of expression variability, chemical instability, as well as possible undesirable reactogenicity and autoimmune properties. A step forward employs the identification of highly-conserved antigenic regions within PnPs with the potential to retain the benefits of protein antigens. Besides, their low-cost and stable construction facilitates the combination of several antigenic regions or peptides that may impair different stages of pneumococcal disease offering even wider serotype coverage and more efficient protection. This review discusses the up-to-date progress on PnPs that are currently under clinical evaluation and the challenges for their licensure. Focus is given on the progress on the identification of antigenic regions/peptides within PnPs and their evaluation as vaccine candidates, accessing their potential to overcome the issues associated with full-length protein antigens. Particular mention is given of the use of newer delivery system technologies including conjugation to Toll-like receptors (TLRs) and reformulation into nanoparticles to enhance the poor immunogenicity of such antigens.

Evaluation of Protective Efficacy of Selected Immunodominant B-Cell Epitopes within Virulent Surface Proteins of Streptococcus pneumoniae.

Four previously identified immunodominant B-cell epitopes, located within known virulent pneumococcal proteins CbpD, PhtD, PhtE, and ZmpB, had shown promising in vivo immunological characteristics, indicating their potential to be used as vaccine antigens. In this study, we further evaluated the opsonophagocytic activity of antibodies against these epitopes and their capacity to protect mice from pneumococcal sepsis. An opsonophagocytic killing assay (OPKA) revealed that OPKA titers of human anti-peptide antibodies against pneumococcal serotypes 1, 3, and 19A were significantly higher (P < 0.001) than those of the control sera, suggesting their functional potential against virulent clinical isolates. Data obtained from mice actively immunized with any of the selected epitope analogues or with a mixture of these (G_Mix group) showed, compared to controls, enhanced survival against the highly virulent pneumococcal serotype 3 (P < 0.001). Moreover, passive transfer of hyperimmune serum from G_Mix to naive mice also conferred protection to a lethal challenge with serotype 3, which demonstrates that the observed protection was antibody mediated. All immunized murine groups elicited gradually higher antibody titers and avidity, suggesting a maturation of immune response over time. Among the tested peptides, PhD_pep19 and PhtE_pep40 peptides, which reside within the zinc-binding domains of PhtD and PhtE proteins, exhibited superior immunological characteristics. Recently it has been shown that zinc uptake is of high importance for the virulence of Streptococcus pneumoniae; thus, our findings suggest that these epitopes deserve further evaluation as novel immunoreactive components for the development of a polysaccharide-independent pneumococcal vaccine.

In silico analysis and in vitro evaluation of immunogenic and immunomodulatory properties of promiscuous peptides derived from Leishmania infantum eukaryotic initiation factor.

It is generally considered as imperative the ability to control leishmaniasis through the development of a protective vaccine capable of inducing long-lasting and protective cell-mediated immune responses. In this current study, we demonstrated potential epitopes that bind to H2 MHC class I and II molecules by conducting the in silico analysis of Leishmania infantum eukaryotic Initiation Factor (LieIF) protein, using online available algorithms. Moreover, we synthesized five peptides (16-18 amino acids long) which are part of the N-terminal portion of LieIF and contain promising MHC class I and II-restricted epitopes and afterwards, their predicted immunogenicity was evaluated in vitro by monitoring peptide-specific T-cell responses. Additionally, the immunomodulatory properties of these peptides were investigated in vitro by exploring their potential of inducing phenotypic maturation and functional differentiation of murine Bone-Marrow derived Dendritic Cells (BM-DCs). It was revealed by our data that all the synthetic peptides predicted for H2 alleles; present the property of immunogenicity. Among the synthetic peptides which contained T-cell epitopes, the peptide 52-68 aa (LieIF_2) exhibited immunomodulatory properties with the larger potential. LieIF_2-pulsed BM-DCs up-regulated the expression of the co-stimulatory surface molecules CD80 and CD86, as well as the production of the proinflammatory cytokine TNF-α and of the Th1-polarizing cytokines IL-12 and IFN-γ. The aforementioned data suggest that selected parts of LieIF could be used to develop innovative subunit protective vaccines able to induce effective immunity mediated by MHC class I-restricted as well as class II-restricted T-cell responses.

Discovery of Immunodominant B Cell Epitopes within Surface Pneumococcal Virulence Proteins in Pediatric Patients with Invasive Pneumococcal Disease.

The identification of immunodominant B cell epitopes within surface pneumococcal virulence proteins in pediatric patients with invasive pneumococcal disease (IPD) is a valuable approach to define novel vaccine candidates. To this aim, we evaluated sera from children with IPD and age-matched controls against 141 20-mer synthetic peptides covering the entire sequence of major antigenic fragments within pneumococcal virulence proteins; namely, choline-binding protein D (CbpD), pneumococcal histidine triad proteins (PhtD and PhtE), pneumococcal surface protein A (PspA), plasminogen and fibronectin binding protein B (PfbB), and zinc metalloproteinase B (ZmpB). Ten immunodominant B cell epitopes were identified: CbpD-pep4 (amino acids (aa) 291-310), PhtD-pep11 (aa 88-107), PhtD-pep17 (aa 172-191), PhtD-pep19 (aa 200-219), PhtE-pep32 (aa 300-319), PhtE-pep40 (aa 79-98), PfbB-pep76 (aa 180-199), PfbB-pep79 (aa 222-241), PfbB-pep90 (aa 484-503), and ZmpB-pep125 (aa 431-450). All epitopes were highly conserved among different pneumococcal serotypes, and four of them were located within the functional zinc-binding domain of the histidine triad proteins PhtD and PhtE. Peptides CbpD-pep4, PhtD-pep19, and PhtE-pep40 were broadly recognized by IPD patient sera with prevalences of 96.4%, 92.9%, and 71.4%, respectively, whereas control sera exhibited only minor reactivities (<10.7%). Their specificities for IPD were 93.3%, 95%, and 96.7%; their sensitivities were 96.4%, 92.9%, and 71.4% and their positivity likelihood ratios for IPD were 14.5, 18.6, and 21.4, respectively. Furthermore, purified antibodies against CbpD-pep4, PhtD-pep19, and PhtE-pep40 readily bound on the surfaces of different pneumococcal serotypes, as assessed by FACS and immunofluorescence analysis. The identified immunodominant B cell epitopes provide a better understanding of immune response in IPD and are worth evaluation in additional studies as potential vaccine candidates.

Assessment of antibody responses in local and immigrant residents of areas with autochthonous malaria transmission in Greece.

Greece has been officially malaria free since 1974. However, from 2009 to 2012, several locally acquired, cases of Plasmodium vivax malaria were detected, in immigrants and in Greek citizens. In this study, the antibody (Ab) response of Greeks and immigrants with documented malaria was initially assessed, followed by an Ab screening of Greeks and immigrant residents of local transmission areas. Of the 38 patients tested, 10.5% of Greeks and 15.7% of immigrants were positive 5-7 months after infection. Of the 1,019 individuals from various areas of Greece, including those of autochthonous transmission, 85 of the 721 (11.8%) immigrants were positive, whereas all 298 Greeks were negative. The rapid Ab titer decline observed is reasonable, given the non-endemic epidemiological setting. The seroepidemiological findings indicate that the local Greek population remains malaria naive and that at this point Greeks are unlikely to serve as reservoir for the infection of local mosquitoes.

Synthetic peptides for efficient discrimination of anti-enterovirus antibodies at the serotype level.

Enteroviruses are important human pathogens, causing a broad spectrum of diseases from minor common colds to fatal myocarditis. However, certain disease syndromes are caused by one or few serotypes. Serotype identification is difficult due to the laborious neutralization tests that lack of sensitivity, while in commercial ELISAs homotypic antibodies' activities are largely masked by the recognition of genera-specific epitopes by heterotypic antibodies. In the present study homotypic assays were developed with the ability to discriminate different enterovirus serotypes. Seventy-three children sera, positive for IgM antibodies against enterovirus genus and 49 healthy children were examined for the presence of antibodies against 14 synthetic peptides derived from a non-conserved region of the VP1 protein of coxsackieviruses B2, B3, B4, B5, A9, A16, A24, echoviruses 6, 7, 9, 11, 30, enterovirus 71 and parechovirus 1. 50% of the anti-enterovirus IgM positive sera (>150 BU) reacted with the peptides with the majority of them to preferentially recognize one of them, supporting the homotypic nature of our assay. Inhibition studies yielded homologous inhibition rates 67-95% suggesting that specific peptide recognition actually occurred. The diagnostic value of our assay was tested in blood samples drawn over a 1.5-year period from a 5-year old patient. The anti-enterovirus reactivity was clearly attributed to echovirus serotype 11. The IgM/IgG antibody ratio was reversed 4 months later and subsequently IgM antibodies dropped below the cutoff point. In this paper we demonstrate that our assay can be used to discriminate between antibodies targeting different enterovirus serotypes.

Malignant lymphoma in primary Sjögren's syndrome: an update on the pathogenesis and treatment.

OBJECTIVES: Sjögren's syndrome (SS), a chronic autoimmune disorder, particularly compromises the function of exocrine glands. Its association with lymphoma is well documented. Our aim was to systematically review the molecular, clinical, histopathologic, and therapeutic aspects of these SS-related malignant lymphoproliferations. METHODS: The literature was searched for original articles published between 1968 and 2012 focusing on the risk factors for lymphoma development in Sjögren's syndrome using MEDLINE and PubMed. The search terms we used were "Sjögren's syndrome," "lymphoma," and "risk factors." All papers identified were English-language, full-text papers. RESULTS: A low-grade marginal-zone lymphoma related to mucosa-associated lymphoid tissue is the commonest lymphoid neoplasia in SS. The majority of SS-associated lymphomas are characterized by localized stage, indolent clinical course, and recurrence in other extranodal sites. Although the transition from a chronic inflammatory condition to malignant lymphoma is a multistep process that is yet poorly understood, there is increasing evidence that chronic antigenic stimulation by an exoantigen or autoantigens plays an essential role in the development of SS-associated lymphoproliferation. CONCLUSIONS: This review discusses the pathogenetic aspects of lymphomagenesis in SS. Recent advances in the treatment of lymphoma in SS are also stated.

Fine epitope specificity of anti-erythropoietin antibodies reveals molecular mimicry with HIV-1 p17 protein: a pathogenetic mechanism for HIV-1-related anemia.

BACKGROUND: Circulating autoantibodies to endogenous erythropoietin (anti-Epo) are detected in human immunodeficiency virus type 1 (HIV-1)-infected patients and represent a risk factor for anemia. The aim of this study was to map the B-cell epitopes on the Epo molecule. METHODS: Serum samples from HIV-1-positive patients and healthy individuals were tested against overlapping peptides covering the entire sequence of Epo. RESULTS: Serum samples from anti-Epo-positive patients exhibited significant binding to Epo epitopes spanning the following sequences: amino acids 1-20 (Ep1), amino acids 54-72 (Ep5), and amino acids 147-166 (Ep12). Structural analysis of erythropoietin revealed that the immunodominant epitopes, Ep1 and Ep12, comprise the interaction interface with Epo receptor (EpoR). Autoantibodies binding to this specific region are anticipated to inhibit the Epo-EpoR interaction, resulting in blunted erythropoiesis; this phenomenon is indicated by the significantly higher Epo levels and lower hemoglobin levels of anti-Ep1-positive patients compared with anti-Ep1-negative individuals. The region corresponding to the Ep1 epitope exhibited a 63% sequence homology with the ³4LVCASRELERFAVNPGLLE5² fragment of the HIV-1 p17 matrix protein. CONCLUSIONS: These results suggest that the main body of anti-Epo is directed against a functional domain of Epo, and that the presence of anti-Epo can be considered to be a result of a molecular mimicry mechanism, which is caused by the similarity between the Ep1 region and the p17 protein.

Seroepidemiological study of pandemic influenza H1N1 following the 2009-2010 wave in Greece.

Knowledge of seroprevalence rates against 2009 pandemic H1N1 virus will assist vaccination recommendations and the preparation of the health-care system during subsequent years. This study was conducted in Greece during June-August 2010 to estimate the seroprevalence rate against pandemic H1N1 virus. Persons presenting in 29 health-care facilities across the country were studied. Seroprevalence was estimated employing a virus-free ELISA that specifically recognizes 2009 H1N1 virus antibodies in human sera. Sera collected from 2005 to April 2009 were also used to estimate pre-pandemic seroprevalence rates. A total of 954 persons were studied. The overall seroprevalence rate was 28.5% (95% confidence interval=25.6-31.3%). Age-specific rates were 34.2% in persons 0-4 years, 36.3% in persons 5-19 years, 25.0% in persons 20-39 years, 23.4% in persons 40-59 years, and 31.8% in persons ≥ 60 years. The highest rates were recorded in the Regions of Ionian Islands (67%) and Epirus (42.9%), while the lowest (8.4%) in the Region of Thessaly. Age-specific attack rates of infection during 2009-2010 were 28.8% in persons 0-4 years, 32.5% in persons 5-19 years, 14.3% in persons 20-39 years, 19.1% in persons 40-59 years, and 14.4% in persons ≥ 60 years. Multivariate analysis revealed that Region of residence and caring for children <5 years were associated with increased risk for seropositivity. Urbanity, personal and family characteristics, working in a health-care facility or in a school, history of pandemic H1N1 vaccination or history of influenza-like illness during 2009-2010 were not associated with increased risk for seropositivity.

Estimation of seroprevalence of the pandemic H1N1 2009 influenza virus using a novel virus-free ELISA assay for the detection of specific antibodies.

The pandemic H1N1 2009 influenza A emerged in April 2009 and spread rapidly all over the world. In Greece, the first case of the pandemic H1N1 was reported on May 18, 2009, while a considerable increase in the number of cases was noticed at the beginning of July 2009. The need for surveillance of the immune status of the Greek population led us to develop a virus-free ELISA that specifically recognizes pandemic H1N1 2009 influenza virus antibodies in human sera. The method is based on the use of synthetic peptides (H1-pep and N1-pep) that are derived from the hemagglutinin and neuraminidase of the 2009 pandemic strain, respectively, and differentiate the swine-origin influenza A/California/14/2009 (H1N1) from the seasonal influenza A viruses. Serum samples were obtained from 271 healthy blood donors during May, November, and December 2009. Among sera collected during May, November, and December, IgG antibodies against the peptide H1-pep were detected in 7.4, 13.8, and 19.3% of the donors, respectively, while IgG antibodies against the peptide N1-pep were detected in 5.3, 9.6, and 16.9% of the donors, respectively. The application of the immunoassay indicated a time-dependent increase of the prevalence of anti-H1-pep and anti-N1-pep IgG antibodies during the pandemic H1N1 outbreak in Greece. The method could be also indicative for the discrimination of immune persons from those susceptible to infection with the pandemic H1N1 strain, as well as for the establishment of effective vaccination programs.