The study investigates the ability of subtype A Avian metapneumovirus (AMPV) to accept foreign genes and be used as a vector for delivery of Infectious bronchitis virus (IBV) QX genes to chickens. Initially the GFP gene was added to AMPV at all gene junctions in conjunction with the development of cassetted full length DNA AMPV copies. After recombinant virus had been recovered by reverse genetics, GFP positions supporting gene expression while maintaining virus viability in vitro, were determined. Subsequently, either S1 or nucleocapsid (N) genes of IBV were positioned between AMPV M and F genes, while later a bivalent recombinant was prepared by inserting S1 and N at AMPV MF and GL junctions respectively. Immunofluorescent antibody staining showed that all recombinants expressed the inserted IBV genes in vitro and furthermore, all recombinant viruses were found to be highly stable during serial passage. Eyedrop inoculation of chickens with some AMPV-IBV recombinants at one-day-old induced protection against virulent IBV QX challenge 3 weeks later, as assessed by greater motility of tracheal cilia from chickens receiving the recombinants. Nonetheless evidence of AMPV/IBV seroconversion, or major recombinant tracheal replication, were largely absent.

nomic losses mainly due to respiratory or reproductive problems (Cecchinato et al., 2012), often exacerbated by secondary bacterial infections (Giovanardi et al., 2014).First, aMPV isolations in Europe date back to the second half of the 1980s, when turkey rhinotracheitis (TRT) outbreaks appeared in

The current information on the prevalence of avian metapneumovirus (aMPV) infection in layers is fragmentary and its true impact on egg production often remains unknown or unclear. In order to draw an epidemiologic picture of aMPV presence in layer flocks in Italy, a survey was performed on 19 flocks of pullets and layers based on longitudinal studies or sporadic samplings. aMPV was detected by reverse transcription (RT)-PCR, and blood samples were collected for serology by aMPV ELISA. Occurrences of respiratory signs and a drop in egg production were recorded. Possible involvement of infectious bronchitis (IB) and egg drop syndrome (EDS) viruses that could have caused loss of egg production we ruled out for IB virus by RT-PCR, and EDS virus was ruled out by hemagglutination-inhibition (HI). Only subtype B of aMPV was found in both pullet and layer farms. Surveys of pullets showed that most groups became infected prior to the onset of lay without showing clear respiratory signs. At the point of lay, these groups were serologically positive to aMPV. In two layer flocks, egg drops were observed and could be strongly linked to the presence of aMPV infection. Results were correlated with aMPV vaccination programs applied to the birds in three flocks on the same farm. Only a vaccination program which included two live and one killed vaccines gave complete protection from aMPV infection to the birds, while a single live vaccine application was not efficacious. The current study gives an inside view of field aMPV diffusion in Italy and its control in layers.

No abstract

Avian metapneumovirus (AMPV) infection of poultry causes serious disease in most countries and subtype A reverse-genetic (RG) systems have allowed a generation of viruses of known sequence, and proved useful in developments towards better control by live vaccines. While subtype B viruses are more prevalent, bacterial cloning issues made subtype B RG systems difficult to establish. A molecular comparison of subtype A and B viruses was undertaken to assess whether subtype A RG components could be partially or fully substituted. AMPV subtype A and B gene-end sequences leading to polyadenylation are, to our knowledge, reported for the first time, as well as several leader and trailer sequences. After comparing these alongside previously reported gene starts and protein sequences, it was concluded that subtype B genome copies would be most likely rescued by a subtype A support system, and this assertion was supported when individual subtype A components were successfully substituted. Application of an advanced cloning plasmid permitted eventual completion of a fully subtype B RG system, and proved that all subtype-specific components could be freely exchanged between A and B systems.