HA222 polymorphism in Influenza A(H1N1) 2009 isolates from Intensive Care Units and ambulatory patients during three influenza seasons.

Amino acid substitutions which can affect the receptor binding specificity of the influenza virus, like the substitution of aspartic acid with glycine in position 222 of the haemagglutinin (HA) of influenza virus A(H1N1) 2009, have been associated with increased viral pathogenicity and increased tropism for the lower respiratory tract. In this paper, the polymorphic site 222 and the site 223 of the HA1 polypeptide of H1N1 2009 viruses were analyzed in order to better clarify the role of these substitutions in H1N1 2009 virus virulence. Viral strains included in this study were collected in Tuscany during 3 different influenza seasons from patients with severe as well as with mild forms of influenza caused by A(H1N1) 2009 virus. In addition, the oseltamivir resistance of the H1N1 2009 strains circulating during the same seasons was monitored with the aim to evaluate whether these changes in the HA and in neuraminidase (NA) tend to be linked and to influence each other. Altogether, the results indicate that in severe forms of influenza viral population is more variable than in mild influenza, as regards the site 222. The frequency of such substitutions varied among the three seasons, it was highest in the season 2010-2011 and very low in the season 2012-2013. However these differences were not significant.

Investigation of an imported case of Middle East Respiratory Syndrome Coronavirus (MERS-CoV) infection in Florence, Italy, May to June 2013.

On 31 May 2013, the first case of Middle East Respiratory Syndrome Coronavirus (MERS-CoV) infection in Italy was laboratory confirmed in a previously healthy adult man, who developed pneumonia with moderate respiratory distress after returning from a holiday in Jordan. Two secondary cases were identified through contact tracing, among family members and colleagues who had not previously travelled abroad. Both secondary cases developed mild illness. All three patients recovered fully.

Molecular surveillance of pandemic influenza A(H1N1) viruses circulating in Italy from May 2009 to February 2010: association between haemagglutinin mutations and clinical outcome.

Haemagglutinin sequences of pandemic influenza A(H1N1) viruses circulating in Italy were examined, focusing on amino acid changes at position 222 because of its suggested pathogenic relevance. Among 169 patients, the D222G substitution was detected in three of 52 (5.8%) severe cases and in one of 117 (0.9%) mild cases, whereas the D222E mutation was more frequent and evenly distributed in mild (31.6%) and severe cases (38.4%). A cluster of D222E viruses among school children confirms reported human-to-human transmission of viruses mutated at amino acid position 222.

A FRET based melting curve analysis to detect nucleotide variations in HA receptor-binding site of H5N1 virus.

Outbreaks of highly pathogenic H5N1 influenza A virus represent a major public health problem because of the possibility of direct transmission of these viruses from avian species to humans. For influenza H5N1 hemagglutinin, a switch from SA-a-2, 3-Gal to SA-a-2, 6-Gal receptor specificity is a critical step that could lead to inter-human transmission. The monitoring of the receptor-binding preference of H5N1 viruses represents an instrument to detect a potential pandemic virus. The aim of this study was to develop a method based on the fluorescence resonance energy transfer (FRET) technology and melting peaks analysis for rapid screening of pandemic H5N1 influenza A virus. Three selected probes corresponding to a 23bp nucleotide sequence of the avian receptor-binding site were used in a real-time RT-PCR to detect nucleotide variations. Five strains of avian influenza A viruses isolated from avian species and two synthesized HA gene were tested. The results showed that the melting peaks analysis is a reliable screening method for detecting the variability of the H5N1 receptor-binding site.

Effectiveness of nanofiltration in removing small non-enveloped viruses from three different plasma-derived products.

The objective of this study was to assess the ability of nanofiltration of albumin solution, prothrombin complex (PTC) and factor IX (FIX) to remove two small, non-enveloped DNA viruses, parvovirus B19 (B19V) and torque teno virus (TTV). Virus removal was investigated with down-scale experiments performed with sequential steps of 35-nm and 15-nm nanofiltrations of products spiked with virus DNA-positive sera. Viral loads were determined by real-time PCRs. The 15-nm nanofiltration removed more than 4.0 B19V log from all the products, TTV was reduced of more than 3.0 log from albumin solution and FIX by 35-nm and 15-nm nanofiltrations, respectively, being viral DNA undetectable after these treatments. Traces of TTV were still found in PTC after the 15-nm nanofiltration. In conclusion, nanofiltration can be efficacious in removing small naked viruses but, since viruses with similar features can differently respond to the treatment, a careful monitoring of large-scale nanofiltration should be performed.

Human parvovirus B19 experimental infection in human fibroblasts and endothelial cells cultures.

With the aim to detect what kind of cells, in addition to erythroid progenitors, could be involved in the pathogenesis of B19 infection in some connective tissue diseases, primary cultures of human fibroblasts (HF) and endothelial cells (HUVEC) were exposed to a B19 positive serum (350 genome copies/cell). The presence of NS1 and VP1 mRNA, in both HF and HUVEC cultures 1, 2 and 6 days after the exposure, indicated infection by B19 virus. However, no significant increase of B19 DNA level in the infected HF and HUVEC cultures was detectable through the entire incubation period of 6 days. It is possible that HF and HUVEC are not permissive for B19 virus replication or, alternatively, that few cells only get infected by B19 virus. HF and HUVEC stimulation with different growth factors or cytokines could be required for a B19 productive infection to occur.