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There is an urgent need to identify efficient antiviral compounds to combat existing and emerging RNA virus infections, particularly those related to seasonal and pandemic influenza outbreaks. While inhibitors of the influenza viral integral membrane proton channel protein (M2), neuraminidase (NA), and cap-dependent endonuclease are available, circulating influenza viruses acquire resistance over time. Thus, the need for the development of additional anti-influenza drugs with novel mechanisms of action exists. In the present study, a cell-based screening assay and a small molecule library were used to screen for activities that antagonized influenza A non-structural protein 1 (NS1), a highly conserved, multifunctional accessory protein that inhibits the type I interferon response against influenza. Two potential anti-influenza agents, compounds 157 and 164, were identified with anti-NS1 activity, resulting in the reduction of A/PR/8/34(H1N1) influenza A virus replication and the restoration of IFN-β expression in human lung epithelial A549 cells. A 3D pharmacophore modeling study of the active compounds provided a glimpse of the structural motifs that may contribute to anti-influenza virus activity. This screening approach is amenable to a broader analysis of small molecule compounds to inhibit other viral targets. [ABSTRACT FROM AUTHOR]

Background: Pandemic coronavirus disease 2019 (COVID-19) disease represents a challenge for healthcare structures. The molecular confirmation of samples from infected individuals is crucial and therefore guides public health decision making. Clusters and possibly increased diffuse transmission could occur in the context of the next influenza season. For this reason, a diagnostic test able to discriminate severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) from influenza viruses is urgently needed.Methods: A multiplex real-time reverse-transcription polymerase chain reaction (PCR) assay was assessed using 1 laboratory protocol with different real-time PCR instruments. Overall, 1000 clinical samples (600 from samples SARS-CoV-2-infected patients, 200 samples from influenza-infected patients, and 200 negative samples) were analyzed.Results: The assay developed was able to detect and discriminate each virus target and to intercept coinfections. The limit of quantification of each assay ranged between 5 and 10 genomic copy numbers, with a cutoff value of 37.7 and 37.8 for influenza and SARS-CoV-2 viruses, respectively. Only 2 influenza coinfections were detected in COVID-19 samples.Conclusions: This study suggests that multiplex assay is a rapid, valid, and accurate method for the detection of SARS-CoV-2 and influenza viruses in clinical samples. The test may be an important diagnostic tool for both diagnostic and surveillance purposes during the seasonal influenza activity period. [ABSTRACT FROM AUTHOR]

Abstract: Antiviral monitoring of influenza viruses circulating in Italy has been carried out since 2007 by the National Influenza Centre (NIC), using both phenotypic and sequence-based assays. Here, we report results of the susceptibility evaluation to neuraminidase (NA) inhibitors (NAIs, zanamivir and oseltamivir) and adamantanes of nearly 300 influenza type A and B seasonal viruses isolated in Italy during six recent seasons, together with over 30 pandemic (H1N1) 2009 virus strains. The present work is the first such study conducted in Italy, aimed to develop national data on antiviral drug profile and to establish a nationwide surveillance programme on antiviral susceptibility. Sequencing of the NA gene was undertaken either to confirm the phenotypic findings or to identify any NA change, in potentially resistant viruses (outliers), which might be associated with reduced susceptibility to NAIs. The 50% inhibitory concentration values (IC50s) showed slightly different sensitivities of the seasonal Italian isolates to the two NAI drugs, depending on the specific NA subtype. We found mean zanamivir IC50s of 0.74, 1.33 and 7nM, and oseltamivir IC50s of 0.67, 2.34 and 30.1nM for the N2, N1 and B NAs, respectively. The pandemic (H1N1) 2009 viruses showed IC50values overall comparable to the seasonal N1 viruses from previous years, showing mean zanamivir IC50s of 1.02nM and mean oseltamivir IC50s of 2.82nM. Oseltamivir resistance was found in a total of 19 seasonal N1viruses of 2007/2008 and 2008/2009, and in three pandemic (H1N1) 2009 strains. A gradual increase of resistance to adamantanes was observed among the N2 viruses isolated in recent seasons; no resistant viruses were found among the seasonal N1 strains, whereas all the pandemic (H1N1) 2009 isolates analysed were resistant to the M2 blockers. [Copyright &y& Elsevier]

The European Paediatric Influenza Analysis (EPIA) project is a multi-country project that was created to collect, analyse and present data regarding the paediatric influenza burden in European countries, with the purpose of providing the necessary information to make evidence-based decisions regarding influenza immunisation recommendations for children. The initial approach taken is based on existing weekly virological and age-specific influenza-like illness (ILI) data from surveillance networks across Europe. We use a multiple regression model guided by longitudinal weekly patterns of influenza virus to attribute the weekly ILI consultation incidence pattern to each influenza (sub)type, while controlling for the effect of respiratory syncytial virus (RSV) epidemics. Modelling the ILI consultation incidence during 2002/2003–2008 revealed that influenza infections that presented for medical attention as ILI affected between 0.3% and 9.8% of children aged 0–4 and 5–14 years in England, Italy, The Netherlands and Spain in an average season. With the exception of Spain, these rates were always higher in children aged 0–4 years. Across the six seasons analysed (five seasons were analysed from the Italian data), the model attributed 47–83% of the ILI burden in primary care to influenza virus infection in the various countries, with the A(H3N2) virus playing the most important role, followed by influenza viruses B and A(H1N1). National season averages from the four countries studied indicated that between 0.4% and 18% of children consulted a physician for ILI, with the percentage depending on the country and health care system. Influenza virus infections explained the majority of paediatric ILI consultations in all countries. The next step will be to apply the EPIA modelling approach to severe outcomes indicators (i.e. hospitalisations and mortality data) to generate a complete range of mild and severe influenza burden estimates needed for decision making concerning paediatric influenza vaccination. [ABSTRACT FROM AUTHOR]

Two major lineages of influenza B viruses have circulated in humans, the B/Yamagata/16/88 (Yam88)-lineage and the B/Victoria/2/87 (Vic87)-lineage. Throughout most of the last decade, Yam88-lineage viruses predominated. During the 2001–2002 influenza season, however, Vic87-lineage viruses emerged outside Asia and spread worldwide. Virological surveillance in Italy during this season highlighted a predominance of influenza B viruses, most of which were antigenically related to the B/Sichuan/379/99 vaccine strain, belonging to the Yam88-lineage, together with a smaller number of B viruses antigenically similar to a recent Vic87-lineage variant B/Hong Kong/330/01. Vic87-lineage viruses were exclusively isolated in the subsequent 2002–2003 epidemic season. Phylogenetic analyses of the hemagglutinin (HA) and neuraminidase (NA) genes of representative Italian B isolates from the 2001–2002 and 2002–2003 seasons showed that, during the first season, the NA as well as the HA genes separated into two distinct clades, the Yam88- and the Vic87-lineages and showed no evidence of reassortment. On the contrary, all the B viruses isolated in the 2002–2003 epidemic season were “Vic87 HA–Yam88 NA” reassortants similar to those isolated in other parts of the world, showing that these reassortants became well-established in the human population. [Copyright &y& Elsevier]

A response by Simona Puzelli, Livia Di Trani, Concetta Fabiani, and Laura Campitelli to a letter to the editor about their article "Serological Analysis of Serum Samples From Humans Exposed to Avian H7 Influenza Viruses in Italy Between 1999 and 2003," in the 2005 issue.

Abstract: In view of the increasing interest in mucosal vaccination, we investigated whether type I IFN could act as adjuvant of an intranasally administered influenza vaccine. A single intranasal administration of IFNαβ-adjuvanted vaccine in anesthetized C3H/HeN mice was capable of protecting the totality of animals against virus challenge, while vaccine alone was only partially effective. To mimic intranasal vaccine administration in man and to limit vaccine delivery strictly to nasal mucosa, we used a second method of vaccination based on vaccine fractionation in six doses and intranasal instillation in non-anesthetized mice. By using this vaccination schedule, IFNαβ-adjuvanted vaccine also prevented mice from disease development and induced an efficient long lasting immune response. Further experiments showed that IFNαβ increased the percentage of antigen-associated phagocytes in the nasal mucus layer, thus suggesting a new possible mechanism of action for type I IFN as an adjuvant. [Copyright &y& Elsevier]

Since the “bird flu” incident in Hong Kong SAR in 1997, several studies have highlighted the substantial role of domestic birds, such as turkeys and chickens, in the ecology of influenza A viruses. Even if recent evidence suggests that chickens can maintain several influenza serotypes, avian influenza viruses (AIVs) circulating in domestic species are believed to be introduced each time from the wild bird reservoir. However, so far the direct precursor of influenza viruses from domestic birds has never been identified. In this report, we describe the antigenic and genetic characterization of the surface proteins of H7N3 viruses isolated from wild ducks in Italy in 2001 in comparison to H7N3 strains that circulated in Italian turkeys in 2002–2003. The wild and domestic avian strains appeared strictly related at both phenotypic and genetic level: homology percentages in seven of their genes were comprised between 99.8% (for PB2) and 99.1% (for M), and their NA genes differed mainly because of a 23-aminoacid deletion in the NA stalk. Outside this region of the molecule, the NAs of the two virus groups showed 99% similarity. These findings indicate that turkey H7N3 viruses were derived “in toto” from avian influenza strains circulating in wild waterfowl 1 year earlier, and represent an important step towards the comprehension of the mechanisms leading to interspecies transmission and emergence of potentially pandemic influenza viruses. [Copyright &y& Elsevier]