Characterization of West Nile virus strains isolated in Italy

West Nile virus (WNV) is a neurotropic flavivirus that is maintained in an enzootic cycle between mosquitoes and birds, but can also infect and cause disease in humans and other vertebrate species. Most of WNV infections in humans are asymptomatic, but approximately 20% of infected people develop clinical symptoms, although severe neurological diseases are observed in less than 1% of them. WNV is the most widely distributed arbovirus in the world and has been recently associated with outbreaks of meningo-encephalitis in Europe, including Italy, caused by different viral strains belonging to distinct lineages 1 and 2. The hypothesis is that genetic divergence among viral strains currently circulating in Italy might reflect on their pathogenic potential and that the rapid spread of WNV with increased pathogenicity within naïve population suggest that epidemic forms of the virus may encode mechanisms to evade host immunity. Infection with WNV triggers a delayed host response that includes a delay in the production of interferon-α (IFN-α). IFNs are a family of immuno-modulatory cytokines that are produced in response to virus infection and serve as integral signal initiators of host intracellular defenses. The increased number of human cases and the lack of data about virulence of European WNV isolates highlight the importance to achieve a better knowledge on this emerging viral infection. In the present study, we investigate the phenotypic and IFN-α-regulatory properties of different WNV lineage 1 and 2 strains that are circulating in Europe/Italy in two cell lines: Vero and 1321N1. We demonstrate that: Vero and 1321N1 cells are capable of supporting WNV replication where different WNV strains show similar growth kinetics; WNV lineage 2 strain replicated in Vero and 1321N1 cells as efficiently as WNV lineage 1 strains; and both lineages 1 and 2 were highly susceptible to the antiviral actions of IFN-α.

Identification and genetic diversity in phytoplasmas associated with diseases of cassava and other agronomic relevant crops in south-east Asia and Latin America

Identification and genetic diversity of phytoplasmas infecting tropical plant species, selected among those most agronomically relevant in South-east Asia and Latin America were studied. Correlation between evolutionary divergence of relevant phytoplasma strains and their geographic distribution by comparison on homologous genes of phytoplasma strains detected in the same or related plant species in other geographical areas worldwide was achieved. Molecular diversity was studied on genes coding ribosomal proteins, groEL, tuf and amp besides phytoplasma 16S rRNA. Selected samples infected by phytoplasmas belonging to diverse ribosomal groups were also studied by in silico RFLP followed by phylogenetic analyses. Moreover a partial genome annotation of a ‘Ca. P. brasiliense’ strain was done towards future application for epidemiological studies. Phytoplasma presence in cassava showing frog skin (CFSD) and witches’ broom (CWB) diseases in Costa Rica - Paraguay and in Vietnam – Thailand, respectively, was evaluated. In both cases, the diseases were associated with phytoplasmas related to aster yellows, apple proliferation and “stolbur” groups, while only phytoplasma related to X-disease group in CFSD, and to hibiscus witches’ broom, elm yellows and clover proliferation groups in CWB. Variability was found among strains belonging to the same ribosomal group but having different geographic origin and associated with different disease. Additionally, a dodder transmission assay to elucidate the role of phytoplasmas in CWB disease was carried out, and resulted in typical phytoplasma symptoms in periwinkle plants associated with the presence of aster yellows-related strains. Lethal wilt disease, a severe disease of oil palm in Colombia that is spreading throughout South America was also studied. Phytoplasmas were detected in symptomatic oil palm and identified as ‘Ca. P. asteris’, ribosomal subgroup 16SrI-B, and were distinguished from other aster yellows phytoplasmas used as reference strains; in particular, from an aster yellows strain infecting corn in the same country.