The remarkable diversity and mobility of Newcastle disease viruses (NDV) includes virulent viruses of genotype VI. These viruses are often referred to as pigeon paramyxoviruses 1 because they are normally isolated and cause clinical disease in birds from the Columbidae family. Genotype VI viruses occasionally infect, and may also cause clinical disease in poultry. Thus, the evolution, current spread and detection of these viruses are relevant to avian health. Here, we describe the isolation and genomic characterization of six Egyptian (2015), four Pakistani (2015), and two Ukrainian (2007, 2013) recent pigeon-derived NDV isolates of sub-genotype VIg. These viruses are closely related to isolates from Kazakhstan, Nigeria and Russia. In addition, eight genetically related NDV isolates from Pakistan (2014–2016) that define a new sub-genotype (VIm) are described. All of these viruses, and the ancestral Bulgarian (n = 2) and South Korean (n = 2) viruses described here, have predicted virulent cleavage sites of the fusion protein, and those selected for further characterization have intracerebral pathogenicity index assay values characteristic of NDV of genotype VI (1.31 to 1.48). A validated matrix gene real-time RT-PCR (rRT-PCR) NDV test detect all tested isolates. However, the validated rRT-PCR test that is normally used to identify the virulent fusion gene fails to detect the Egyptian and Ukrainian viruses due to mismatches in primers and probe. A new rapid rRT-PCR test to determine the presence of virulent cleavage sites for viruses from sub-genotypes VIg was developed and evaluated on these and other viruses. We describe the almost simultaneous circulation and continuous evolution of genotype VI Newcastle disease viruses in distant locations, suggesting epidemiological connections among three continents. As pigeons are not migratory, this study suggests the need to understand the possible role of human activity in the dispersal of these viruses. Complete genomic characterization identified previously unrecognized genetic diversity that contributes to diagnostic failure and will facilitate future evolutionary studies. These results highlight the importance of conducting active surveillance on pigeons worldwide and the need to update existent rapid diagnostic protocols to detect emerging viral variants and help manage the disease in affected regions.

Our study demonstrates the repeated isolation of vaccine-derived Newcastle disease viruses from different species of wild birds across four continents from 1997 through 2014. The data indicate that at least 17 species from ten avian orders occupying different habitats excrete vaccine-derived Newcastle disease viruses. The most frequently reported isolates were detected among individuals in the order Columbiformes (n = 23), followed in frequency by the order Anseriformes (n = 13). Samples were isolated from both free-ranging (n = 47) and wild birds kept in captivity (n = 7). The number of recovered vaccine-derived viruses corresponded with the most widely utilized vaccines, LaSota (n = 28) and Hitchner B1 (n = 19). Other detected vaccine-derived viruses resembled the PHY-LMV2 and V4 vaccines, with five and two cases, respectively. These results and the ubiquitous and synanthropic nature of wild pigeons highlight their potential role as indicator species for the presence of Newcastle disease virus of low virulence in the environment. The reverse spillover of live agents from domestic animals to wildlife as a result of the expansion of livestock industries employing massive amounts of live virus vaccines represent an underappreciated and poorly studied effect of human activity on wildlife.

The Azov and Black Sea basins are part of the transcontinental wild bird migration routes from Northern Asia and Europe to the Mediterranean, Africa, and Southwest Asia. These regions constitute an area of transit, stops during migration, and nesting for many different bird species. From September 2010 to September 2011, a wild bird surveillance study was conducted in these regions to identify avian influenza viruses. Biological samples consisting of cloacal and tracheal swabs and fecal samples were collected from wild birds of different ecological groups, including waterfowl and sea- and land-based birds, in places of mass bird accumulations in Sivash Bay and the Utlyuksky and Molochniy estuaries. The sampling covered the following wild bird biological cycles: autumn migration, wintering, spring migration, nesting, and postnesting seasons. A total of 3634 samples were collected from 66 different species of birds. During the autumn migration, 19 hemagglutinating viruses were isolated, 14 of which were identified as low pathogenicity avian influenza (LPAI) virus subtypes H1N?, H3N8, H5N2, H7N?, H8N4, H10N7, and H11N8. From the wintering samples, 45 hemagglutinating viruses were isolated, 36 of which were identified as LPAI virus subtypes H1N1, H1N? H1N2, H4N?, H6N1, H7N3, H7N6, H7N7, H8N2, H9N2, H10N7, H10N4, H11N2, H12N2, and H15N7. Only three viruses were isolated during the spring migration, nesting, and postnesting seasons (serotypes H6, H13, and H16). The HA and NA genes were sequenced from the isolated H5 and N1 viruses, and the phylogenetic analysis revealed possible ecological connections between the Azov and Black Sea regions and Europe. The LPAI viruses were isolated mostly from mallard ducks, but also from shellducks, shovelers, teals, and white-fronted geese. The rest of the 14 hemagglutinating viruses isolated were identified as different serotypes of avian paramyxoviruses (APMV-1, APMV-4, APMV-6, and APMV-7). This information furthers our understanding of the ecology of avian influenza viruses in wild bird species.

Despite the existence of 10 avian paramyxovirus (APMV) serotypes, very little is known about the distribution, host species, and ecological factors affecting virus transmission. To better understand the relationship among these factors, we conducted APMV wild bird surveillance in regions of Ukraine suspected of being intercontinental (north to south and east to west) flyways. Surveillance for APMV was conducted in 6,735 wild birds representing 86 species and 8 different orders during 2006 to 2011 through different seasons. Twenty viruses were isolated and subsequently identified as APMV-1 (n= 9), APMV-4 (n= 4), APMV-6 (n= 3), and APMV-7 (n= 4). The highest isolation rate occurred during the autumn migration (0.61%), with viruses isolated from mallards, teals, dunlins, and a wigeon. The rate of isolation was lower during winter (December to March) (0.32%), with viruses isolated from ruddy shelducks, mallards, white-fronted geese, and a starling. During spring migration, nesting, and postnesting (April to August) no APMV strains were isolated out of 1,984 samples tested. Sequencing and phylogenetic analysis of four APMV-1 and two APMV-4 viruses showed that one APMV-1 virus belonging to class 1 was epidemiologically linked to viruses from China, three class II APMV-1 viruses were epidemiologically connected with viruses from Nigeria and Luxembourg, and one APMV-4 virus was related to goose viruses from Egypt. In summary, we have identified the wild bird species most likely to be infected with APMV, and our data support possible intercontinental transmission of APMVs by wild birds.

Here, we report the complete genome sequence of a virus of a putative new serotype of avian paramyxovirus (APMV). The virus was isolated from a white-fronted goose in Ukraine in 2011 and designated white-fronted goose/Ukraine/Askania-Nova/48-15-02/2011. The genomic characterization of the isolate suggests that it represents the novel avian paramyxovirus group APMV 13.