Experimental pigeon paramyxovirus-1 infection in chicken: evaluation of infectivity, clinical and pathological manifestations and diagnostic methods.

Several pigeon paramyxovirus-1 (PPMV-1) outbreaks in feral pigeons were described recently in Switzerland. The potential of PPMV-1 to induce the notifiable Newcastle disease in chickens is discussed controversially. Therefore, in order to study epidemiologically relevant parameters such as the kinetics of PPMV-1 replication and shedding as well as seroconversion after infection, chickens were infected experimentally with a Swiss PPMV-1 isolate. This generated also defined sample material for the comparison of diagnostic tests. The infectivity of the Swiss PPMV-1 isolate for chickens was demonstrated successfully by virus shedding after experimental inoculation. Our data suggest that long-lasting shedding for up to 60 days can occur in chickens infected with PPMV-1. The isolate used here was of low pathogenicity for chickens. Different quantitative reverse transcription PCR assays were evaluated with a set of Swiss PPMV-1 isolates, and various samples from experimentally infected chickens were analysed with respect to their suitability for viral RNA detection. At 14 days post-infection, virus genome was detected mainly in spleen, caecal tonsils, heart, cloacal swabs, liver, proventriculus, duodenum and kidney tissue samples. Overall, the level of virus replication was low. Not all assays used routinely in diagnostics were capable of detecting viral genome from the isolates tested. Possible explanations are the genetic divergence of PPMV-1 and the low level of viral RNA in the samples. In contrast, two methods that are not used routinely proved more suitable for virus-genome detection. Importantly, the collection of material from various different organs is recommended, in addition to the kidney and brain analysed routinely. In conclusion, this study shows that there is a need to reconsider the type of samples and the protocols used for the detection of PPMV-1 RNA in chickens.

Survey on Chlamydiaceae in cloacal swabs from Swiss turkeys demonstrates absence of Chlamydia psittaci and low occurrence of Chlamydia gallinacean.

In Switzerland, domestic turkey meat is a niche product. Turkeys are fattened on mixed family-based farms scattered across the country, with most providing access to an uncovered outdoor pasture for the birds. Swiss fattening turkeys may therefore get infected with Chlamydiaceae via wild birds or their faeces, potentially shedding these bacteria at a later stage. The aim of the present study was to acquire baseline data about the shedding of Chlamydiaceae in clinically unremarkable Swiss fattening turkeys at slaughter, potentially exposing slaughterhouse workers to infection. In this large-scale study, 1008 cloacal swabs of Swiss turkeys out of 53 flocks from 28 different grow-out farms with uncovered outdoor pasture were collected over the course of 14 months and examined for the occurrence of Chlamydiaceae by a family-specific 23S-rRNA real-time PCR. Positive samples were further analyzed by Chlamydia psittaci (C. psittaci)-specific real-time PCR and the Arraymate DNA Microarray for species identification. All samples were negative for C. psittaci, but seven swabs out of one flock were tested positive for Chlamydia gallinacea (0.7%). Although turkeys with access to pasture may have contact with Chlamydiaceae-harbouring wild birds or their faeces, the infection rate in Swiss turkeys was shown to be low.

Resource-Effective Serosurveillance for the Detection of West Nile Virus in Switzerland Using Abattoir Samples of Free-Range Laying Hens.

West Nile virus (WNV) is an important zoonotic pathogen maintained in a natural transmission cycle between mosquitoes and birds as reservoir hosts. In dead-end hosts, such as humans, infection may result in fatal neurologic disease translating into disease and death-related suffering and increased health care costs. In humans, WNV may also be transmitted through blood transfusions and organ transplants. WNV is not present in Switzerland yet, but competent vector species (especially Culex pipiens and Aedes japonicus) are prevalent and an introduction of the virus, likely through wild birds, is expected at any time. Therefore, it is important for Switzerland to be prepared and establish a surveillance system for WNV to initiate increased prevention activities, such as the screening of blood and organ donations and public education activities in case virus circulation is detected. The long-term goal of these surveillance measures would be a reduced infection rate in humans resulting in less suffering and reduced health care costs. To provide the basis for a pragmatic and resource-effective WNV surveillance program, this study used aliquots of serum samples of free-range laying hens taken at the abattoir and collected in the frame of the ongoing Swiss Avian Influenza and Newcastle Disease monitoring program for a 2-year period. All 961 aliquots were analyzed using a commercial competitive WNV enzyme-linked immunosorbent assay (ELISA). The study allowed to set up sampling and laboratory routines as a basis for future WNV surveillance activities. At this stage there is no evidence for circulation of WNV in Switzerland.

Cutaneous lesions in pet rabbits following subcutaneous administration of a novel bivalent vaccine against myxomatosis and rabbit haemorrhagic disease.

BACKGROUND: A novel bivalent vaccine to protect against myxomatosis and rabbit haemorrhagic disease is commercially available for pet rabbits. HYPOTHESIS/OBJECTIVES: To describe the appearance of cutaneous lesions arising in pet rabbits positive for myxoma virus (MV) by RT-PCR evaluation shortly after vaccination. ANIMALS: Four pet rabbits presenting with papular, crusting skin lesions ~10 days after vaccination. METHODS: Histological evaluation of formalin-fixed skin biopsies obtained from lesional skin (case 1). Real-time polymerase chain reaction (RT-PCR) evaluation of paraffin-embedded tissue from skin biopsies (case 1) and crusts obtained from the lesion surface (cases 2-4) for myxoma virus are reported as cycle threshold (Ct ) values. RESULTS: Lesions affecting the ear pinna, dorsal aspect of the nose, vulva and/or conjunctiva are reported. Histopathological findings included severe ulcerative, necrotizing dermatitis and intralesional cytoplasmic inclusion bodies in myxoma cells. DNA was amplified from all the paraffin-embedded skin biopsies (Ct  = 34-35) and crusts (Ct  = 20-24). CONCLUSIONS AND CLINICAL IMPORTANCE: Although a wild virus challenge cannot be definitively excluded, veterinarians and pet-owners should be aware that cutaneous lesions have been observed after vaccination with this novel vaccine in low numbers of rabbits.

Peripheral nerve sheath tumor in a subadult golden eagle (Aquila chrysaetos).

A 5-year-old, female golden eagle (Aquila chrysaetos) was admitted with tetraplegia that progressed to a nonambulatory, spastic tetraparesis after a few days of treatment. Clinical and radiologic examinations, including radiography, computed tomography scan, and myelography, were indicative of neoplasia involving a spinal nerve root. Postmortem magnetic resonance imaging and necropsy findings confirmed the diagnosis of a peripheral nerve sheath neoplasia, not, to our knowledge, previously reported in a raptor.

Development of a new real-time polymerase chain reaction assay to detect Duck adenovirus A DNA and application to samples from Swiss poultry flocks.

Between 2008 and 2012, commercial Swiss layer and layer breeder flocks experiencing problems in laying performance were sampled and tested for infection with Duck adenovirus A (DAdV-A; previously known as Egg drop syndrome 1976 virus). Organ samples from birds sent for necropsy as well as blood samples from living animals originating from the same flocks were analyzed. To detect virus-specific DNA, a newly developed quantitative real-time polymerase chain reaction method was applied, and the presence of antibodies against DAdV-A was tested using a commercially available enzyme-linked immunosorbent assay. In 5 out of 7 investigated flocks, viral DNA was detected in tissues. In addition, antibodies against DAdV-A were detected in all of the flocks.

Comparison of ventriculotomy closure with and without a coelomic fat patch in Japanese quail (Coturnix coturnix japonica).

Removal of foreign bodies from the ventriculus in birds may necessitate ventriculotomy. Complications with this intervention include leakage and adhesion formation. To investigate if the use of a coelomic fat patch and a tension-relieving suture in addition to a simple interrupted pattern would improve the healing process after ventriculotomy, 2 groups of 9 Japanese quail (Coturnix coturnix japonica) underwent ventriculotomy. In group 1, only simple-interrupted and tension-relieving sutures were used for closure of the ventriculotomy. In group 2, a coelomic fat patch from the surrounding adipose tissue was applied to the incision site in addition to the sutures. All quail recovered normally and were considered clinically healthy after surgery. Three birds from each group were euthanatized at days 7, 14, and 21 after surgery. On histologic examination, the suture techniques used for closure of the ventriculotomy led to minimal inflammation of the surrounding tissues in both groups. Serosal inflammation was significantly greater in group 2 birds that had the adipose patch at closure compared with group 1 birds. Therefore, the use of a coelomic fat patch to cover the site of ventriculotomy did not result in an improved healing process and its use is not recommended in quail.

Detection of Avian coronavirus infectious bronchitis virus type QX infection in Switzerland.

Infectious bronchitis, a disease of chickens caused by Avian coronavirus infectious bronchitis virus (IBV), leads to severe economic losses for the poultry industry worldwide. Various attempts to control the virus based on vaccination strategies are performed. However, due to the emergence of novel genotypes, an effective control of the virus is hindered. In 1996, a novel viral genotype named IBV-QX was reported for the first time in Qingdao, Shandong province, China. The first appearance of an IBV-QX isolate in Europe was reported between 2003 and 2004 in The Netherlands. Subsequently, infections with this genotype were found in several other European countries such as France, Italy, Germany, United Kingdom, Slovenia, and Sweden. The present report describes the use of a new set of degenerate primers that amplify a 636-bp fragment within the S1 gene by reverse transcription polymerase chain reaction to detect the occurrence of IBV-QX infection in Switzerland.

Development and validation of a Myxoma virus real-time polymerase chain reaction assay.

To aid in the rapid diagnosis of myxomatosis in rabbits, a real-time polymerase chain reaction (PCR) for the specific detection of Myxoma virus is described. Primers and probe were designed to amplify a 147-bp fragment within the Serp2 gene. The assay was able to detect 23 copies of a synthesized oligo indicating a reliable sensitivity. In addition, the real-time PCR did not detect the Rabbit fibroma virus used in myxomatosis vaccines. The novel PCR was shown to be able to detect Myxoma virus in fresh and paraffin-embedded rabbit tissues originating from myxomatosis cases from various regions in Switzerland.

Emergence and establishment of Usutu virus infection in wild and captive avian species in and around Zurich, Switzerland--genomic and pathologic comparison to other central European outbreaks.

In late summer 2006 considerable mortality in wild and captive Passeriformes and Strigiformes was observed in Zurich, Switzerland. All animals were found in a range of 2 km(2). Observed clinical signs involved depression, ruffled plumage, incoordination, seizures and peracute death. Nutritional status was generally moderate to poor in wild birds, and variable in captive animals. Necropsy showed marked splenomegaly, a mild hepatomegaly, and pulmonary hyperemia in most animals. Histopathologic lesions were very discrete and consisted mainly of neuronal necrosis, leucocytolysis in and around the brain blood vessels, and miliary liver necrosis. The diagnosis Usutu virus (USUV) infection was established by USUV-specific immunohistochemistry and reverse transcription-polymerase chain reaction. Partial nucleotide sequence comparisons revealed>99% identity between the viruses that emerged in Zurich in 2006, in Vienna in 2001, and in Budapest in 2005. Since 2008 a significantly lower mortality was observed in wild Passeriformes, but USUV infection was confirmed for the first time beyond Zurich city limits. Indoor housing and regular treatment against ectoparasites are likely to have prevented acute USUV disease in captive Strigiformes. USUV is a mosquito-borne flavivirus causing fatalities in various avian species. After the initial European outbreaks in Austria in 2001 it appears that the virus has extended its range in Central Europe and has established a transmission cycle between local bird and mosquito species. Further episodes of increased avian mortality in the forthcoming years, with impact on wild and captive bird populations, predominantly Passeriformes and Strigiformes, can be anticipated. Furthermore, the possibility of broader dispersal of USUV in Europe during the next mosquito seasons must be considered and an increased mortality in Passeriformes and Strigiformes must be expected until protective "flock immunity" is established. Collections of valuable and endangered Passeriformes and Strigiformes, especially young of the year, should therefore be housed indoors or treated against ectoparasites at acceptable intervals between July and September each year.

Phenotypic and genetic characterization of Pasteurella multocida and related isolates from rabbits in Switzerland.

Several bacteria belonging to the family Pasteurellaceae are potential pathogens in rabbits. In particular, Pasteurella multocida is considered to be important, and outbreaks caused by this species result in considerable economic losses in rabbitries. However, Pasteurellaceae spp. isolated from rabbits are poorly characterized, and thus, proper identification of P. multocida isolates from these animals is problematic and often unsatisfactory, thereby hampering epidemiological investigations. Therefore, 228 isolates from rabbit populations originating from a breeding and fattening organization with group management and postmortem cases with pasteurellosis from individual owners were phenotypically and genotypically analyzed using biochemical tests and repetitive extragenic palindromic polymerase chain reaction (REP-PCR). Furthermore, 41 samples representing observed phenotypes were selected for phylogenetic analysis using 16S ribosomal RNA and rpoB genes. The REP-PCR typing and phylogenetic analyses correlated well and appeared to be distinct molecular methods for characterization of rabbit isolates. Phenotyping, however, diverged from molecular recognition, reflecting the problematic conventional diagnosis of these strains. The fermentation of sorbitol appeared to be an imprecise indicator for P. multocida subspecies classification. According to REP-PCR and sequencing results, 82% of the isolates were characterized as P. multocida subsp. multocida, 3% as P. multocida subsp. septica, and 5% as P. multocida. Further, 5% were identified as Pasteurella canis. The other 5% represented a homogeneous group of unknown species belonging to the Pasteurellaceae. Samples obtained from individual postmortem cases demonstrated a higher phenotypic and genetic heterogeneity than samples from group management rabbits.

Avian bornaviruses in psittacine birds from Europe and Australia with proventricular dilatation disease.

To determine whether avian bornaviruses (ABVs) were a factor in proventricular dilatation disease (PDD), we used immunohistochemistry, reverse transcription-PCR, and nucleotide sequence analysis to examine paraffin wax-embedded or frozen tissue samples of 31 psittacine birds with this disease. PDD is a fatal disease of psittacine birds associated with nonsuppurative encephalitis and ganglioneuritis of the upper intestinal tract. Tissue samples had been collected from 1999 through 2008 in Austria, Switzerland, Hungary, and Australia. Immunohistochemical demonstration of viral antigen within the brain and vegetative nerve system of the gastrointestinal tract provides strong evidence for a causative role of ABVs in this condition. Partial sequences of nucleoprotein (p40) and matrix protein (gp18) genes showed that virus in most of our cases belonged to the ABV-2 and ABV-4 groups among the 5 genogroups described so far. Viral sequences of 2 birds did not match any of the described sequences and clustered together in a new branch termed ABV-6.

Endocarditis due to Lactobacillus jensenii in a Salvin's Amazon parrot (Amazona autumnalis salvini).

A 30-year-old Salvin's Amazon parrot (Amazona autumnalis salvini) with a history of a lifelong poor diet and inappropriate housing was presented in lateral recumbency to a veterinary teaching hospital for further evaluation. Radiological and ultrasonographic examination revealed a mild proventricular dilatation, mild hepatomegaly, signs of enteritis and airsacculitis. The main laboratory findings included a mild macrocytic hyperchromic anaemia, hypoglobulinaemia, decreased bile acids and increased alkaline phosphatase. In this bird a liver pathology was suspected because of the clinical, laboratory and ultrasonographic findings. The bird was treated with supportive care and metabolic aids. After initial improvement of the clinical signs, the bird's condition deteriorated and it died. Pathological findings revealed an endocarditis and myocarditis due to Lactobacillus jensenii and a bacteraemia. Endocarditis due to Lactobacillus sp. is a rare phenomenon in humans not yet described in animals. It is associated with severe underlying illnesses leading to translocation of otherwise non-pathogenic bacteria in the bloodstream. A similar pattern might be assumed in animals with compromised immunity.

Monitoring of wild birds for Newcastle disease virus in Switzerland using real time RT-PCR.

Wild birds are considered to be the natural reservoir of the Newcastle disease virus (NDV; avian paramyxovirus-1) causing New-castle disease, and are often suspected to be involved in outbreaks in domesticated birds. To assess the epidemiologic status of wild birds living, or overwintering, in Switzerland, 3,049 cloacal swabs covering the period 2003-2006 were screened for NDV, using real time RT-PCR. All samples were negative. This result seems in contrast with previously performed serologic screenings of wild birds.

Humoral immune response to avian influenza vaccination over a six-month period in different species of captive wild birds.

In December 2005, the four major Swiss zoos carried out the vaccination of selected zoo birds with the adjuvant inactivated vaccine H5N2 Nobilis influenza. Pre- and post-vaccination antibody titers were determined either by hemagglutination inhibition (HI) test (non-Galliformes) or by enzyme linked immunosorbent assay (ELISA) (Galliformes) at Week 0, 5, 10, and 26 (Day 0-1, 35-36, 70-71, and 182 respectively) to determine the humoral immune response to H5 antigen. After the first vaccination, the overall geometric mean titer of non-Galliformes was 65 (n = 142), which increased to 187 (n = 139) after booster vaccination and dropped to 74 (n = 65) six months after first vaccination. For the Galliformes group, the mean titers were found to be 2.09 at Week 5 (n = 119), 3.24 at Week 10 (n = 113), and 1.20 at Week 26 (n = 39). Within the non-Galliformes, significant differences in geometric mean titers were found among different species representatives. In general, the flamingos (Phoenicopteriformes) showed a strong response to vaccination, reaching a geometric mean titer of 659 at Week 10, while the Sphenisciformes did not show high antibody titers even after booster vaccination, reaching a maximum geometric mean titer of only 65. Based on the antibody titer profiles of all investigated species, we recommend at least annual revaccination for the species that we investigated.

High resolution typing of Chlamydophila psittaci by multilocus VNTR analysis (MLVA).

A multilocus VNTR analysis (MLVA) system for detection of tandem repeats across the whole genome of Chlamydophila psittaci has been developed. Twenty selected genetic loci were initially tested on 9 avian reference strains including representatives of all major serotypes (A to F). Thereafter, 8 loci were retained for a more complete study performed on over 150 C. psittaci isolates from different bird species and geographical origins. Comparative analysis of the MLVA results and those obtained from currently available methods including serotyping and/or ompA sequencing indicate that the MLVA system provides an additional level of discrimination, with 20 distinct patterns identified to date. The newly developed MLVA system therefore provides a highly sensitive, high resolution test for the differentiation of C. psittaci isolates from different origins that is suitable for molecular epidemiological studies.

The 2005/2006 avian influenza monitoring of wild birds and commercial poultry in Switzerland.

In October 2005, the second Swiss national avian influenza monitoring in wild waterfowl and commercial poultry with free range management started. Cloacal swabs were examined by real-time reverse transcription-polymerase chain reaction for both M gene of influenza A virus and H5 subtype. The monitoring (more than 2000 samples tested) documented the introduction of H5N1 in Swiss wild waterfowl in mid-February 2006. Until the end of March, 29 water bird carcasses were found H5 positive. In the same period, domestic poultry flocks with a permit of free-range management were kept under surveillance, with negative results.

Chlamydiae and atherosclerosis: can psittacine cases support the link?

Atherosclerosis is a common disease in pet birds, particularly in psittacines. Little is known about the role of risk factors predisposing birds to this disease. In our study, we tried to detect chlamydiae in formalin-fixed and paraffin-embedded atherosclerotic tissue from 103 pet birds to clarify their role in atherosclerosis. Methods used were polymerase chain reaction (PCR), sequencing, and immunohistochemistry. Histopathologic examination served to classify the extent of atherosclerotic lesions. In the PCR, 4 (3.9%) of 103 cases, all of them with advanced stages of atherosclerosis, were positive. Subsequent sequence analysis revealed high identities (94%-100%) with Chlamydophila psittaci in three cases. Interestingly, two of these birds came from C. psittaci-infected populations. Because of the low incidence (3.9%), the occurrence only in advanced stages, and the association with C psittaci-infected avian populations, a causal relationship between chlamydiae and atherosclerosis in pet birds is rather improbable.

Possible human-avian transmission of Mycobacterium tuberculosis in a green-winged macaw (Ara chloroptera).

This report describes a case of Mycobacterium tuberculosis infection in a green-winged macaw (Ara chloroptera), confirmed by microbiologic and pathologic diagnostics, and notes a possible human-avian transmission. Clinical signs included cutaneous swellings, profound leukocytosis, and signs of osteomyelitis in the long bones. Proliferation consisted of several nodules with small greenish-caseous foci in cross-section and revealed a severe granulomatous inflammation with intralesional acid-fast rods. Mycobacterium tuberculosis was isolated from subcutaneous nodules and biochemically confirmed. The disease in avian species is of zoonotic importance.