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Revello M.G.,Science Ostetricia e Ginecologia | Genini E.,Science Virologia e Microbiologia | Gorini G.,Science Virologia e Microbiologia | Klersy C.,Servizio di Biometria e Epidemiologia Clinica | And 2 more authors.
Journal of Clinical Virology | Year: 2010

Background: The interpretation of a positive IgM antibody result to human cytomegalovirus (HCMV) in a pregnant woman is of major importance for the correct management of the pregnancy. Determination of HCMV-specific IgG avidity is considered an useful approach for distinguishing IgM antibody due to primary HCMV infection from IgM antibody elicited during non-primary infection. Objective: Comparative evaluation of eight commercial HCMV IgG avidity assays currently available in Europe. Study design: A panel of 198 sequential samples collected from 65 pregnant women at 0-90, 91-180, and >180 days after the onset of primary HCMV infection was retrospectively tested by Abbott, BioMérieux, Bio-Rad, DiaSorin, Diesse, Euroimmun, Radim, and Technogenetics HCMV IgG avidity assays according to the manufacturer's instructions. Results: None of the 198 samples tested yielded identical scores by the kits under evaluation. The Euroimmun and Radim assays showed the best correlation with expected results in terms of low (0-90 days), intermediate (90-180 days) and high (>180 days) avidity results, respectively. The best accuracy in diagnosing a recent (<90 days after the onset) or non-recent (>180 days after the onset) primary HCMV infection was shown by Radim followed by Euroimmun and Diesse. The best correlation with a well established in-house developed HCMV IgG avidity assay was shown by Radim. Conclusions: HCMV IgG avidity kits need to be improved and standardized. In the meantime, highly specific IgM assays are preferable for screening purposes in pregnant women. © 2010 Elsevier B.V.

Gerna G.,Laboratori Sperimentali Of Ricerca
Future Microbiology | Year: 2012

Using the murine CMV animal model and the well-established model of Cre-lox-P-mediated green-fluorescence tagging of endothelial cell (EC)-derived mouse CMV to quantify the role of infected ECs in transplantation-associated CMV dissemination (in mice expressing Cre recombinase under the control of either the Tie2 or the Tek promoter selectively expressed in vascular EC-Tie-Cre and Tek-Cre mice), it was shown that EC-derived virus contributed to 50% of the total viral load during primary infection, and there was no preference for dissemination of EC-derived viruses over viruses produced by other cell types. In addition, during secondary viremia, there was only a negligible contribution of EC-derived virus to dissemination to other organs. These results are novel in the methodology employed and are somewhat interesting. However, the data are limited to the mouse model with a short-term follow-up, and the immunodeficient host has not yet been studied. In humans, these conclusions must be taken with caution. First, in primary infection occurring through natural routes, epithelial cells are infected first, then ECs, unless primary infection occurs through blood transfusion, in which case endothelial vascular cells may become infected first. In both cases, the virus transport occurs through the intervention of leukocytes, namely monocytes and polymorphonuclear leukocytes. As monocytes differentiate to macrophages, they become highly susceptible to human CMV replication inside organ tissues, while polymorphonuclear leukocytes are active in virus capturing from infected endothelial vascular cells and transporting to distant sites. © 2012 Future Medicine Ltd.

Piralla A.,S.S. Virologia Molecolare | Baldanti F.,S.S. Virologia Molecolare | Gerna G.,Laboratori Sperimentali Of Ricerca
Journal of Clinical Microbiology | Year: 2011

Human rhinovirus species C (HRV-C) was the second most common HRV species detected in hospitalized patients in Italy with acute respiratory disease during a 1-year surveillance period. Sequencing of the picornavirus VP4/VP2 region allowed molecular typing of HRV-A and HRV-B and provisional typing of HRV-C. Copyright © 2011, American Society for Microbiology. All Rights Reserved.

Kabanova A.,University Svizzera Italiana | Perez L.,University Svizzera Italiana | Lilleri D.,University Svizzera Italiana | Lilleri D.,Laboratori Sperimentali Of Ricerca | And 11 more authors.
Proceedings of the National Academy of Sciences of the United States of America | Year: 2014

The use of neutralizing antibodies to identify the most effective antigen has been proposed as a strategy to design vaccines capable of eliciting protective B-cell immunity. In this study, we analyzed the human antibody response to cytomegalovirus (human cytomegalovirus, HCMV) infection and found that antibodies to glycoprotein (g)B, a surface glycoprotein that has been developed as a HCMV vaccine, were primarily nonneutralizing. In contrast, most of the antibodies to the complex formed by gH, gL, protein (p)UL128, pUL130, and pUL131 (the gHgLpUL128L pentamer) neutralized HCMV infection with high potency. Based on this analysis, we developed a single polycistronic vector encoding the five pentamer genes separated by "self-cleaving" 2A peptides to generate a stably transfected CHO cell line constitutively secreting high levels of recombinant pentamer that displayed the functional antigenic sites targeted by human neutralizing antibodies. Immunization of mice with the pentamer formulated with different adjuvants elicited HCMV neutralizing antibody titers that persisted to high levels over time and that were a hundred- to thousand-fold higher than those found in individuals that recovered from primary HCMV infection. Sera from mice immunized with the pentamer vaccine neutralized infection of both epithelial cells and fibroblasts and prevented cell-to-cell spread and viral dissemination from endothelial cells to leukocytes. Neutralizing monoclonal antibodies from immunized mice showed the same potency as human antibodies and targeted the same as well as additional sites on the pentamer. These results illustrate with a relevant example a general and practical approach of analytic vaccinology for the development of subunit vaccines against complex pathogens. © 2014, National Academy of Sciences. All rights reserved.

Gerna G.,Laboratori Sperimentali Of Ricerca | Lilleri D.,Laboratori Sperimentali Of Ricerca | Comolli G.,Laboratori Sperimentali Of Ricerca | Pellegrini C.,Divisione di Cardiochirurgia | And 3 more authors.
American Journal of Transplantation | Year: 2011

Control of human cytomegalovirus (HCMV) infection during the posttransplant period was investigated in 134 solid-organ transplant recipients by monitoring in parallel virologic and immunologic parameters for at least 1 year of follow-up. Virologic monitoring was achieved by determining HCMV DNAemia with real-time PCR, using the threshold of 300 000 DNA copies/mL blood as a cutoff for starting preemptive therapy. Immunologic monitoring included measurement of HCMV-specific CD4+ and CD8+ T cells by cytokine flow cytometry, using HCMV-infected dendritic cells as a stimulus. HCMV infection was diagnosed in 110 (82%) and required treatment in 49 (36%) patients. At 12 months after transplantation 'protective' immunity (≥0.4 CD4+ and CD8+ HCMV-specific T cells/lL blood) was achieved in 115/129 (89%) patients. During the entire study period, 122 patients reconstituting HCMV-specific CD4+ and CD8+ T-cell immunity at 60 days posttransplant onward were able to control HCMV infection, except for one patient who developed HCMV disease because of a rejection episode. Patients reconstituting HCMV-specific CD8+ only did not control HCMV infection. In conclusion, the presence of both HCMV-specific CD4+ and CD8+ T cells ≥ 0.4/lL blood appears to be protective against HCMV disease. This result does not apply to patients undergoing antirejection treatment, or reconstituting HCMV-specific CD8+ T cells only. © Copyright 2011 The American Society of Transplantation.

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