Long-Term Follow-Up of Acute Hepatitis B: New Insights in Its Natural History and Implications for Antiviral Treatment.

Acute hepatitis B infection (AHB) is still a common viral acute hepatitis worldwide. As vaccination, antiviral treatment, and immigration are bound to affect the epidemiological landscape of HBV infections, and some of its aspects need to be investigated: (1) the circulation of vaccine escape mutants and of primary drug resistant strains; (2) the change in HBV genotype prevalence; and (3) the clinical implications of AHB and the probability of chronification. The serological, virological, and clinical parameters of 75 patients, acutely infected by HBV, were gathered for a retrospective study. Long-term follow up, either to complete seroconversion or for up to five years, was possible for 44 patients. Sequence analysis of the reverse transcriptase/HBsAg and precore regions was performed to investigate the molecular epidemiology and pathogenesis of recent infections by HBV. Genotype distribution in AHB in Italian patients was radically different from that of chronic infections, with a dramatic increase of extra-European genotypes (A1, F), suggesting that a proportion of AHBs are currently related to imported strains. None of the documented infections occurred in vaccinated individuals, while HBsAg variants (potentially vaccine escape variants) were rare and less prevalent than in chronic infections. No drug resistant strains were observed. Spontaneous viral clearance occurred in all but three cases. Time to viral clearance was inversely proportional to liver damage, but HBsAg titer on day 28 and, better still, HBsAg decay from day 0 to day 28 after admission, were the best predictors of chronification. They are, thus, potentially useful to guide antiviral treatment to prevent chronic evolution.

In-Depth Analysis of HA and NS1 Genes in A(H1N1)pdm09 Infected Patients.

In March/April 2009, a new pandemic influenza A virus (A(H1N1)pdm09) emerged and spread rapidly via human-to-human transmission, giving rise to the first pandemic of the 21th century. Influenza virus may be present in the infected host as a mixture of variants, referred to as quasi-species, on which natural and immune-driven selection operates. Since hemagglutinin (HA) and non-structural 1 (NS1) proteins are relevant in respect of adaptive and innate immune responses, the present study was aimed at establishing the intra-host genetic heterogeneity of HA and NS1 genes, applying ultra-deep pyrosequencing (UDPS) to nasopharyngeal swabs (NPS) from patients with confirmed influenza A(H1N1)pdm09 infection. The intra-patient nucleotide diversity of HA was significantly higher than that of NS1 (median (IQR): 37.9 (32.8-42.3) X 10(-4) vs 30.6 (27.4-33.6) X 10(-4) substitutions/site, p = 0.024); no significant correlation for nucleotide diversity of NS1 and HA was observed (r = 0.319, p = 0.29). Furthermore, a strong inverse correlation between nucleotide diversity of NS1 and viral load was observed (r = - 0.74, p = 0.004). For both HA and NS1, the variants appeared scattered along the genes, thus indicating no privileged mutation site. Known polymorphisms, S203T (HA) and I123V (NS1), were observed as dominant variants (>98%) in almost all patients; three HA and two NS1 further variants were observed at frequency >40%; a number of additional variants were detected at frequency <6% (minority variants), of which three HA and four NS1 variants were novel. In few patients multiple variants were observed at HA residues 203 and 222. According to the FLUSURVER tool, some of these variants may affect immune recognition and host range; however, these inferences are based on H5N1, and their extension to A(H1N1)pdm09 requires caution. More studies are necessary to address the significance of the composite nature of influenza virus quasi-species within infected patients.

Molecular characterization of hepatitis A outbreak in the province of Rome, Lazio region, Italy, January-July 2013.

Reduced circulation of hepatitis A virus lead to an increase of susceptible individuals, and outbreaks occurred recently. In Northern Italy an outbreak is ongoing, attributed to a monophyletic genotype IA strain, with mixed frozen berries as probable source. From 01/01/2013 to 07/15/2013, 30 cases were diagnosed at National Institute for Infectious Diseases, Rome, Italy, representing about twice the number of cases in whole 2012. Phylogenetic analysis indicated that most, although not all, infections were attributable to the same monophyletic genotype IA strain identified in the contemporary Northern Italy outbreak. This strain is also very similar to previous isolates from Venezuela.

Extent of HCV NS3 protease variability and resistance-associated mutations assessed by next generation sequencing in HCV monoinfected and HIV/HCV coinfected patients.

HCV quasispecies variability represents the background for the selection of mutations and for the development of drug resistance. Natural aminoacid changes in NS3, associated with reduced protease inhibitor susceptibility, have been observed in treatment-naïve patients. Massively parallel sequencing has been used to analyze NS3 quasispecies in patients infected with HCV genotype 1, naive to anti-HCV treatment, with/without HIV-coinfection, to establish the genetic heterogeneity and the presence of amino acid substitutions at positions responsible for drug resistance. Genomes carrying substitutions represented either predominant or minority components of viral quasispecies, and were observed in 85.7% of patients. Multiple substitutions, frequently associated on the same haplotype, were observed in 46.4% of patients. High resistance combinations were not detected, neither on the same genome, nor in the whole quasispecies. Heterogeneity of HCV NS3 was lower in HIV-coinfected as compared to HCV-monoinfected patients, but factors underlying this difference remain to be established. Although the relevance of naturally occurring mutations with respect of resistance development and probability of success of direct acting antivirals is questioned, UDPS may be beneficial to help understanding viral dynamics, providing high resolution view of viral diversity.

Evolutionary pattern of pandemic influenza (H1N1) 2009 virus in the late phases of the 2009 pandemic.

Influenza A( H1N1)v has spread rapidly in all parts of the globe in 2009 as a true pandemic, although fortunately a clinically mild one. The relevant evolutionary steps for the new virus to adapt to human populations occurred very early during the pandemic, before the end of April. Of the several resulting clades or clusters, clade 7 appeared later and proved more successful, substituting all other early clades before the bulk of the worldwide infections occurred.