Retrospective immunohistochemical investigation on dolphin morbillivirus infection by comparing the performance of heterologous monoclonal and polyclonal antibodies - Short communication.

Dolphin morbillivirus (DMV) is a pathogen of great concern in free-ranging cetaceans. Confirmation and staging of morbillivirus infections rely on histology and immunohistochemistry (IHC), following molecular detection. As at the present time no specific antibodies (Abs) against DMV are available, two heterologous Abs have been used worldwide for the examinations of morbillivirus infections of cetaceans. One is a monoclonal Ab (MoAb) prepared against the N protein of canine distemper virus (CDV), whereas the other is a polyclonal Ab raised in rabbits against rinderpest virus (RPV). Both Abs are known to show cross-reactivity with DMV. In this study we compared the labelling quality and the neuroanatomical distribution of staining with these two Abs by means of IHC analysis. To this end, serial sections of the target organs from ten free-ranging stranded cetaceans, previously diagnosed as being infected with DMV by PCR and/or serology, were subjected to IHC. The brain, lungs and lymph nodes of one animal were found to be positive with both Abs. From two other animals, the brain and the spleen, respectively, tested positive only with the polyclonal Ab. In the positive brain tissues, multifocal immunostaining was observed, with similar staining location and extent, with the two antibodies tested. Our results suggest that the polyclonal anti-RPV Ab might have a stronger binding activity to DMV than the anti-CDV MoAb. Nevertheless, the elaboration and use of specific anti-DMV Abs might be essential to guarantee conclusive results in diagnostic and pathogenetic investigations.

Epidemiological Features of the Highly Pathogenic Avian Influenza Virus H5N1 in a Densely Populated Area of Lombardy (Italy) during the Epidemic Season 2021-2022.

In the last two years, there have been three major epidemic seasons in the territory of the European Union and the HPAI epizootic in 2021-2022 is the most severe in recent history. In Italy, the disease was introduced to dense poultry areas with serious economic consequences for the entire sector. In Lombardy, the analysis of the risk factors was carried out, also taking into account the density of domestic birds. In the most affected areas, 66.7% of the outbreaks occurred in the areas with the highest poultry density and the likelihood of an outbreak occurring increased with an increase in the density of birds per km2. In cells 10 × 10 km with a density greater than 10,000 birds/km2, the probability of outbreak occurrence was over 66.7%. The provinces involved in the last epidemic were the same involved in previous epidemics and, given the risk factors present in the area, it is plausible that the risk remains high also for future epidemic seasons. Therefore, to avoid the repetition of similar events, certain control measures shall be strengthened and vaccination considered as a complementary tool for the control of HPAI virus in risk areas.

Basic information for the development of a toxicity assay in inactivated bacterial vaccines.

This study dealt with the toxicity of inactivated bacteria intended for veterinary autogenous vaccines toward a suitable control assay. Two in vitro methods were used. The [3-(4, 5 -dimethylthiazol-2-yl) -2,5 -diphenyltetrazolium bromide] (MTT) test, based on the metabolic reaction of a tetrazolium salt in vital cells, was adopted on the basis of previous positive results. The Interleukin (IL)-1 beta release assay on monocyte-derived pig macrophages was carried out for comparative purposes, to evaluate the possible role of the inflammatory response. MTT and IL-1 beta responses showed a significant correlation (P < 0.05) at defined test dilutions of bacterial antigens, whereas no correlation was demonstrated using MTT responses normalized on bacterial cell concentration. Furthermore, the toxic effects shown in the MTT test were positively correlated to the extracellular protein content. On the whole, the above results could be a useful basis for the development of a toxicity assay on inactivated bacterial vaccines. Also, our data point at bacterial autolysis as a major component underlying toxicity.