Wild birds play an important role as reservoir hosts and vectors for zoonotic arboviruses and foster their spread. Usutu virus (USUV) has been circulating endemically in Germany since 2011, while West Nile virus (WNV) was first diagnosed in several bird species and horses in 2018. In 2017 and 2018, we screened 1709 live wild and zoo birds with real-time polymerase chain reaction and serological assays. Moreover, organ samples from bird carcasses submitted in 2017 were investigated. Overall, 57 blood samples of the live birds (2017 and 2018), and 100 organ samples of dead birds (2017) were positive for USUV-RNA, while no WNV-RNA-positive sample was found. Phylogenetic analysis revealed the first detection of USUV lineage Europe 2 in Germany and the spread of USUV lineages Europe 3 and Africa 3 towards Northern Germany. USUV antibody prevalence rates were high in Eastern Germany in both years. On the contrary, in Northern Germany, high seroprevalence rates were first detected in 2018, with the first emergence of USUV in this region. Interestingly, high WNV-specific neutralizing antibody titers were observed in resident and short-distance migratory birds in Eastern Germany in 2018, indicating the first signs of a local WNV circulation.

Proventricular dilatation disease (PDD) is a fatal infectious disease of birds that primarily affects psittacine birds. Although a causative agent has not been formally demonstrated, the leading candidate is a novel avian bornavirus (ABV) detected in post-mortem tissue samples of psittacids with PDD from the USA, Israel and, recently, Germany. Here we describe the presence of ABV in a parrot with PDD as well as in clinically normal birds exposed to birds with PDD. In two ABV-positive post-mortem cases, the tissue distribution of ABV was investigated by quantitative real-time reverse transcription-polymerase chain reaction. Viraemia was observed in a PDD-affected bird whereas a restriction of ABV to nerve tissue was found in the non-PDD-affected bird. Healthy birds from the same aviary as the affected birds were also found to harbour the virus; 19/59 (32.2%) birds tested positive for ABV RNA in cloacal swabs, providing the first evidence of ABV in clinically healthy birds. In contrast, 39 birds from the same geographic area, but from two different aviaries without PDD cases in recent years, had negative cloacal swabs. ABV RNA-positive, clinically healthy birds demonstrated the same serological response as the animal with confirmed PDD. These results indicate that ABV infection may occur without clinical evidence of PDD and suggest that cloacal swabs can enable the non-invasive detection of ABV infection.

West Nile virus (WNV) and Usutu virus (USUV) are important flaviviruses circulating in Germany. While USUV was first reported more than 10 years ago, WNV has only reached the country in 2018. Wild birds are important amplifying hosts for both viruses. Therefore, we have been monitoring the bird population in different regions of Germany by a previously established network for many years. This report summarizes the results of molecular and/or serological methods of 2345 blood samples from birds of 22 different orders and over 2900 bird carcasses from 2019 and 2020. USUV RNA circulation was found in different regions of Germany, with emphasis on USUV lineages Europe 3 and Africa 3. Increased evidence of USUV lineage Europe 2 was detected in eastern Germany. WNV RNA was found only in birds from the eastern part of the country. The seroprevalence for USUV was between 3.11% and 7.20% in all three regions investigated, whereas the WNV seroprevalence spanned from 14.77% to 16.15% in eastern Germany, with a noticeable tendency for a westward and southward expansion in both years. Thus, wild bird monitoring for WNV and USUV can serve as an early warning system for a human exposure risk.

The paper described a novel technique for semen collection in large psittacines (patent pending), a procedure which was not routinely possible before. For the first time, a large set of semen samples is now available for analysis as well as for artificial insemination. Semen samples of more than 100 psittacine taxa were collected and analysed; data demonstrate large differences in the spermatological parameters between families, indicating an ecological relationship with breeding behaviour (polygamous versus monogamous birds). Using semen samples for artificial insemination resulted in the production of offspring in various families, such as Macaws and Cockatoos, for the first time ever. The present technique represents a breakthrough in species conservation programs and will enable future research into the ecology and environmental factors influencing endangered species.

Psittaciform 1 bornavirus (PaBV) has already been shown to be the aetiologic agent of proventricular dilatation disease, a significant disease of birds. However, the pathogenesis of PaBV infection has not yet been resolved and valid data regarding the pathogenicity of different PaBV species are lacking. Thus, the present study was aimed to characterize the influence of two different PaBV species on the course of disease. Eighteen cockatiels were inoculated intracerebrally (i.c.) or intravenously (i.v.) with a PaBV-2 isolate under the same conditions as in a previous study using PaBV-4. Birds were surveyed and sampled for 33 weeks to analyse the course of infection and disease in comparison to that of PaBV-4. Similar to PaBV-4, PaBV-2 induced a persistent infection with seroconversion (from day 6 p.i. onwards) and shedding of viral RNA (from day 27 p.i. onwards). However, in contrast to PaBV-4, more birds displayed clinical signs and disease progression was more severe. After PaBV-2 infection, 12 birds exhibited clinical signs and 10 birds revealed a dilated proventriculus in necropsy. After PaBV-4 infection only four birds revealed clinical signs and seven birds showed a dilatation of the proventriculus. Clinically, different courses of disease were observed after PaBV-2 infection, mainly affecting the gastrointestinal tract. This had not been detected after PaBV-4 infection where more neurological signs were noted. The results provide evidence for different disease patterns according to different PaBV species, allowing the comparison between the infection with two PaBV species, and thus underlining the role of viral and individual host factors for disease outcome.

Avian bornavirus (ABV) has been identified as the cause of proventricular dilatation disease in birds, but the virus is also found in healthy birds. Most studies of ABV have focused on captive birds. We investigated 86 free-ranging psittacine birds in Brazil and found evidence for natural, long-term ABV infection.