Gizzard erosions in broiler chickens in Sweden caused by fowl adenovirus serotype 1 (FAdV-1): investigation of outbreaks, including whole-genome sequencing of an isolate.

The present paper describes the investigation of the first outbreaks of adenoviral gizzard erosions (AGE) in Sweden, in five broiler flocks. The investigation included whole viral genome sequencing and investigation of genomic organization and sequence relationships with other adenoviruses. All five flocks had a history of decreased growth and uneven size of birds since 9-10 days of age. Macroscopically, lesions consistent with AGE (detached koilin layers, discolouration, bleeding, erosions) were identified in gizzards in all five flocks. In four flocks histology was performed, and degeneration and inflammation of the koilin layer and gizzard mucosa were identified in all four. In one flock, intranuclear inclusion bodies typical for fowl adenovirus (FAdV) were detected in trapped epithelial cells in the koilin layer. In four flocks in situ hybridization was performed, and cells positive for FAdV serotype 1 (FAdV-1) were demonstrated in the koilin layer and gizzard mucosa. FAdV species A (FAdV-A) was detected in gizzard, liver, caecal tonsils and bursa of Fabricius by polymerase chain reaction (PCR) and sequencing. Ten out of ten examined parent flocks of the affected chickens were seropositive for FAdV, indicating former or on-going infection. However, FAdV was not detected in embryos from seropositive parent flocks and thus vertical transmission was not demonstrated. The entire nucleotide sequence of one sample was determined and found to be 43,856 base pairs (bp) in length. The genome sequence and organization were found to be similar to that of the reference apathogenic avian adenovirus "chicken embryo lethal orphan" (CELO). RESEARCH HIGHLIGHTSAGE in Swedish broilers: necropsy, histopathology, ISH, PCR, whole-genome sequencing.Whole FAdV-genome analysed: 43,856 bp, found to be most similar to CELO (U46933.1).Multiple point mutations, site insertions and deletions identified compared to CELO.Paper adds knowledge about European disease situation and pathogenic FAdV-strains.

Spotlight on avian pathology: fowl adenovirus (FAdV) in chickens and beyond - an unresolved host-pathogen interplay.

Fowl adenovirus (FAdV) infections in chickens have undergone substantial changes in recent decades, driven by host and pathogen factors. Based on the pathogenesis of inclusion body hepatitis (IBH) and hepatitis-hydropericardium syndrome (HHS), modern broilers are much more inclined to have difficulties keeping the metabolic homeostasis, whereas adenoviral gizzard erosion (AGE) is noticed equally in broilers and egg-layers. Defining the importance of certain serotypes for specific FAdV diseases is a major achievement of recent years but the isolation of viruses from clinically healthy birds remains unexplained, as virulence factors are hardly known and continue to be a "black box". Together with further studies on pathogenesis of FAdV-induced diseases, such knowledge on virulence factors would help to improve protection strategies, which presently mainly concentrate on autogenous vaccines of breeders to prevent vertical transmission.

Feed-borne Bacillus cereus exacerbates respiratory distress in chickens infected with Chlamydia psittaci by inducing haemorrhagic pneumonia.

Chlamydia psittaci is an important zoonotic pathogen and its oral route of infection plays an important role in the transmission and persistence. Bacillus cereus (B. cereus) strain, a common contaminant of animal feed and feedstuffs, can cause severe diarrhoea and malnutrition in poultry. In our previous study, a B. cereus strain (Dawu C), isolated from the haemorrhagic lungs of infected chickens, was shown to harbour two virulence genes (hblC and cytk) and was able to induce haemorrhagic lesions in the lungs, as well as gizzard erosion and ulceration (GEU) syndrome in broilers. In the present study, we tested the hypothesis that B. cereus-induced GEU would aggravate C. psittaci infection. Our results showed that SPF chickens exposed to B. cereus developed a severe GEU syndrome. More interestingly, prior infection with B. cereus facilitated C. psittaci infection, and aggravated GEU and respiratory distress, which were accompanied by high chlamydial loads in the lungs and severe lesions in respiratory organs. Moreover, levels of local inflammatory cytokines were elevated and T cell responses were impaired in the infected birds. In conclusion, GEU caused by B. cereus may facilitate chlamydial transmission from the ventriculus to the lungs.RESEARCH HIGHLIGHTS Bacillus cereus contributes to the gizzard erosion and ulceration syndrome in chickens.Exposure to Bacillus cereus exacerbates pneumonia in birds following chlamydial infection.Bacillus cereus facilitates persistent chlamydial infection and exacerbates immune responses.

Nosema apis and Nosema ceranae Tissue Tropism in Worker Honey Bees (Apis mellifera).

The microsporidia Nosema apis and Nosema ceranae are major honey bee pathogens that possess different characteristics in terms of the signs they produce, as well as disease development and transmission. Although the ventricular epithelium is generally considered the target tissue, indirect observations led to speculation that N. ceranae may also target other structures, possibly explaining at least some of the differences between these 2 species. To investigate the tropism of Nosema for honey bee tissues, we performed controlled laboratory infections by orally administering doses of 50 000 or 100 000 fresh mature spores of either species. The fat body was isolated from the infected bees, as well as organs from the digestive (esophagus, ventriculus, ileum, rectum), excretory (Malpighian tubules), circulatory (aorta, heart), respiratory (thoracic tracheas), exocrine (hypopharyngeal, mandibular and labial, cephalic, thoracic salivary glands), and sensory/nervous (brain, eyes and associated nerve structures, thoracic nerve ganglia) systems. Tissues were examined by light and electron microscopy at 7, 10, and 15 days postinfection. Both Nosema species were found to infect epithelial cells and clusters of regenerative cells in the ventriculus, and while the ileum and rectum contained spores of the microsporidia in the lumen, these structures did not show overt lesions. No stages of the parasites or cellular lesions were detected in the other organs tested, confirming the high tropism of both species for the ventricular epithelium cells. Thus, these direct histopathological observations indicate that neither of these 2 Nosema species exhibit tropism for honey bee organs other than the ventriculus.

Anatomopathological changes, quantification and distribution of Libyostrongylus spp. in regions of the proventriculus and ventriculus of naturally- and experimentally-infected ostriches.

Nematodes of the genus Libyostrongylus parasitize ostriches, causing high mortality rates. These nematodes are found in the proventriculus and ventriculus of ostriches, but little is known about their distribution and the possible anatomopathological changes they cause in the various regions of these organs. This paper describes the distribution and quantification of Libyostrongylus and pathological changes found in regions of the proventriculus and ventriculus of ostriches with high and low levels of both natural and experimental infection. Ostriches were necropsied and tissue samples from the distinct regions of both organs were analysed based on nematode counts and histopathology after staining with haematoxylin and eosin, Masson's trichrome or Alcian blue/PAS. The cranial and glandular regions of the proventriculus were the most parasitized. The ventriculus contained more nematodes in the caudal region. No macro- or microscopic pathological changes were observed in either of these organs of experimentally-infected birds. However, naturally-infected birds with high levels of infection presented proventriculus with macroscopic lesions and heterophilic infiltrates surrounding nematodes. In the glandular region of this organ, nematodes were located in the adenomeres of the secretory ducts, causing altered architecture and erosions and ulcerative lesions with damaged epithelium. Nematode eggs were found in the koilin layer of the middle and caudal regions of the ventriculus only of these birds. The pH of the regions assessed by Alcian blue/PAS staining changed from acidic in the proventriculus to more alkaline in the caudal region of the ventriculus. These data add knowledge to the biology of Libyostrongylus. RESEARCH HIGHLIGHTS The most parasitized areas were the cranial and glandular regions of the proventriculus. Naturally-infected birds with high levels of infection presented macro lesions in the proventriculus and damaged epithelium. Nematode eggs were found in the ventriculus. The proventriculus had an acidic pH, which turned alkaline towards the ventriculus.

Erosive Enteritis and Intestinal Obstructions Caused by Decomposed Granite in a Flock of Lesser Flamingos (Phoeniconaias minor).

Decomposed granite is a material commonly used in zoos as a base for flamingo exhibits. In 2012, the artificial turf over the decomposed granite in the Fort Worth Zoo flamingo exhibit was removed and pressure washed, the granite was replaced, and the turf was then restretched in place. Approximately 8 months later, 3 lesser flamingos (Phoeniconaias minor) showed signs of lethargy and hematochezia. Results of physical examination revealed poor body condition in all birds, with variable hematologic abnormalities, including anemia, hypoproteinemia, and electrolyte disturbances. Radiographs of birds showed large amounts of radiopaque material in the ventriculus and dilated intestinal loops. Two birds died either during treatment or surgery, and 1 bird was euthanatized. Postmortem examination of all 3 birds revealed large amounts of fine, granite-like material in the ventriculus and intestines, severe congestion of intestinal mucosa, and obstruction of intestinal lumen by caseous plugs surrounded by fecal material. Microscopic evaluation of the material removed from the ventriculus showed birefringent crystals that corresponded to microscopic silica grains, consistent with the decomposed granite from the exhibit. Histologic examination of tissues showed marked chronic erosive colitis, mild erosive proventriculitis, and lesions in the muscular layer of the ventriculus. Subsequently, 82 remaining flamingos in the flock were radiographed. Results showed the same material in the ventriculus of most birds, but no intestinal obstructions were identified. Different treatment techniques were attempted unsuccessfully to remove the material from the flamingos that had the largest amount of decomposed granite in their gastrointestinal tracts. Ultimately, all birds were treated with oral gastrointestinal protectants, adsorbents, laxatives, and lubricants, along with antibiotics administered in drinking water. No further mortalities occurred once medical treatments were implemented.

Fowl aviadenovirus serotype 1 confirmed as the aetiological agent of gizzard erosions in replacement pullets and layer flocks in Great Britain by laboratory and in vivo studies.

An investigation into the aetiology and pathogenesis of adenoviral gizzard erosion has been conducted following three natural outbreaks affecting one flock of 6-week-old replacement pullets and two consecutive placements of free range layers at the age of 21 and 23 weeks. Affected flocks showed increased mortality (0.12-0.30% per week), and gizzard lesions were consistent with fowl aviadenovirus (FAdV) involvement. To substantiate the initial findings, a selection of archived formalin-fixed paraffin-embedded gizzard samples from another 12 pullet and layer flocks, for which macroscopic and histopathological diagnosis of the disease were recorded in Great Britain during the period 2009-2016, were also investigated. In situ hybridization (ISH), virology and/or PCR confirmed the presence of FAdV species-A, serotype-1 (FAdV-A, FAdV-1) DNA in gizzard samples of all 15 cases investigated. Co-infections with additional FAdV serotypes including FAdV-8a were detected by serology and/or virology in two of the pullet flocks. However, species-specific in situ hybridization revealed that pathological changes of affected gizzards were only associated with the detection of FAdV-A. A subsequent in vivo study infecting 21-day-old SPF pullets with FAdV-1 or FAdV-8a strains isolated from the 6-week-old replacement pullets revealed characteristic pathomorphological changes only in the gizzards from birds infected with FAdV-1. While infection with FAdV-8a was confirmed by virology and serology, infected SPF birds did not develop pathomorphological changes. Therefore, the aetiological involvement of the isolated FAdV-8a in the development of adenoviral gizzard erosion in commercial pullets has been ruled out.

Heat stress as a predisposing factor for necrotic enteritis in broiler chicks.

Heat stress is a physical environmental stressor, which can affect performance, health and welfare of poultry. The present study investigates the effect of cyclic acute heat stress as a predisposing factor for necrotic enteritis in broiler chicks in an experimental challenge model. Two hundred and forty broiler chicks were randomly allocated to four treatment groups, as follows: group A served as negative control (25°C), group B birds were subjected to cyclic acute heat stress (35°C), group C birds were challenged and group D birds were both subjected to heat stress and challenged. From each bird, the intestine, gizzard and liver were collected and scored for gross lesions. The intestinal digesta was collected for pH and viscosity determination. One caecum from each bird was taken for microbiological analysis. The statistical analysis and evaluation of data revealed that the heat stress in challenged birds showed a relative trend to increase the severity and the incidence of necrotic enteritis lesions, although it was not considered as statistically significant (Ρ = 0.077). Additionally, the heat stress induced necrotic enteritis outbreak in unchallenged birds; the challenge of birds as well as its combination with the heat stress affected significantly (Ρ ≤ 0.05) the pH and viscosity of intestinal digesta and the caecal Clostridium perfringens counts. The study provides evidence that cyclic acute heat stress is an environmental stressor, which can significantly affect necrotic enteritis and gut health, and thus should be taken into account in warmer areas of the world where poultry farming becomes a major industry.

Clinical signs and progression of lesions in the gizzard are not influenced by inclusion of ground oats or whole wheat in the diet following experimental infection with pathogenic fowl adenovirus serotype 1.

In the present study the effects of dietary gizzard stimulation on the development and severity of adenoviral gizzard erosion were investigated. For this purpose, specific pathogen-free broilers were divided into six groups, investigating the influence of an oat-containing diet with higher fibre content, a whole wheat-containing diet and a control diet of nearly identical composition, but containing ground wheat. For each feed administered, one group of birds was experimentally infected on the 10th day of age by the oral route with virulent fowl adenovirus serotype 1 (FAdV-1), recently proven to induce gizzard erosions, while the respective negative control groups remained uninfected. Experimental feed was administered from 2 days post-infection onwards. No significant differences on gizzard health or in weight gain could be detected between uninfected control groups or between FAdV-1 infected groups that received different experimental feed. However, independent of the supplied diet, a significantly reduced weight gain was noted from 7 days post-infection onwards in FAdV-1 infected broilers compared to uninfected birds that received the same diet. Macroscopically, discolouration and erosion of the koilin layer and inflammation of the gizzard mucosa were observed in all FAdV-1 infected groups. Histologically, necrosis, degeneration of gizzard epithelial cells and multiple basophilic intranuclear inclusion bodies were observed. In summary, after experimental infection with FAdV-1 development of gizzard erosion in chickens was not influenced by the feeding regimes investigated. Therefore, it is unlikely that dietary gizzard stimulation influences the outcome of adenoviral gizzard erosion in vertically infected broilers.

High mortality in laying hen pullets caused by crop and gizzard impactions associated with ingestion of bale net wrap.

High mortality was observed in young replacement layers. Balls of bale net wrap strings were found in the crop and/or gizzard of birds causing impaction and traumatic injury. Some birds experienced loss of portions or the entire tongue secondary to ischemic necrosis. Mortality stopped with the removal of strings from the environment.

Mortalités élevées chez des poulettes pondeuses causées par des impactions du jabot et du gésier associées à l'ingestion de cordes d'attache de ballots. Une mortalité élevée a été observée chez les jeunes pondeuses de remplacement. Des balles de ficelle de cordes de ballots ont été trouvées dans le jabot et/ou le gésier des oiseaux causant ainsi une impaction et des traumatismes. Quelques oiseaux ont perdu des portions ou la totalité de la langue en raison d'une nécrose ischémique. Les mortalités ont cessé après l'enlèvement des ficelles de l'environnement.(Traduit par Isabelle Vallières).

Temporary feed restriction partially protects broilers from necrotic enteritis.

The objective of this study was to investigate the effect of feed restriction on the intestinal ecosystem and on the pathogenesis of experimental necrotic enteritis in broiler chicks. To induce subclinical necrotic enteritis, an experimental challenge model using a specific diet formulation, Gumboro vaccination, oral inoculation of broilers with a 10-fold dose of attenuated anticoccidial vaccine and multiple oral inoculations with a specific strain of Clostridium perfringens was adopted. Two hundred and forty 1-day-old Cobb 500 broilers were randomly allocated to four groups: feed restricted, challenged, both feed restricted and challenged, and negative control. At 21, 22, 23 and 24 days of age, the intestines, gizzard and liver were collected from 15 birds in each group and scored for gross lesions. The intestinal digesta was collected for pH and viscosity determination. One caecum from each bird was taken for microbiological analysis. The application of feed restriction in birds challenged with C. perfringens reduced the necrotic enteritis lesion score significantly (P ≤ 0.05) and feed restriction significantly reduced (P ≤ 0.05) pH in the small intestine, the viscosity of the jejunum digesta as well as the C. perfringens counts in the caeca compared with the controls. In conclusion, feed restriction of broilers has a positive effect on the intestinal ecosystem and a significant protective effect against necrotic enteritis in the subclinical experimental model.

Gizzard Erosion and Ulceration Syndrome in Moroccan Poultry Flocks and Molecular Characterization of Fowl Adenoviruses (FAdV).

Gizzard erosion and ulceration syndrome (GEUS) is caused by a fowl adenovirus serotype 1 (FAdV-1) and was first reported in laying hens in Japan in 1993. This syndrome has emerged as an epizootic in Morocco since 2014, causing significant economic losses for the poultry industry, but no involvement of a FAdV has been confirmed. Thus, the objective of this work was to assess GEUS cases that occurred in the country and to determine the role of FAdVs in their occurrence. Investigations were based on a retrospective reassessment of tissue sections and paraffin blocks of gizzards and livers from GEUS cases between 2014 and 2021 coupled with a prospective search of cases in 2022. Gizzards and livers were fixed in 10% neutral buffered formalin for histopathologic examinations according to standard methods and stored at -20 C for molecular analysis. After deparaffinizing, 10-µm-thick tissue sections along with fresh organs were subjected to DNA extraction using a commercial kit. A primer pair specific for the Hexon gene of FAdVs was used in conventional PCR; in contrast, for real-time PCR, a primer pair targeting the 52K gene was employed. In total, 24 flock cases with characteristic GEUS were assessed between 2014 and 2022. They were nine broiler cases aged between 11 and 39 days, 11 layer cases with an age between 17 and 29 wk, two cases in meat-type breeders aged 10 and 27 wk, and two flock cases of turkey poults aged 22 and 23 days. In most cases, microscopic lesions were consistent with an ulcerative and lymphoplasmocytic ventriculitis, and pathognomonic viral intranuclear inclusion bodies within degenerate epithelial cells were identified in four broiler flock cases, four layer cases, and one case in breeders and hence were highly suggestive of a FAdV infection. Among these nine cases that were positive at the histopathologic examination, six cases were found to be FAdV-PCR positive; another four cases were negative to histology but FAdV-PCR positive. Furthermore, a sequencing analysis was conducted, providing the initial evidence of the implication of FAdV-1 from species A as the cause of GEUS in Moroccan poultry. Additionally, a phylogenetic analysis was executed to facilitate a comparison between the strains investigated in this study and those identified in diverse geographic regions and across various time periods.

Síndrome de erosión y ulceración de la molleja en parvadas avícolas marroquíes y caracterización molecular de los adenovirus aviares (FAdV). El síndrome de erosión y ulceración de la molleja (GEUS) es causado por un adenovirus del pollo de serotipo 1 (FAdV-1) y se reportó por primera vez en gallinas de postura en Japón en 1993. Este síndrome se ha convertido en una epizootia en Marruecos desde 2014, causando importantes pérdidas económicas a la industria avícola, pero no se ha confirmado la participación de ningún adenovirus del pollo. Por lo tanto, el objetivo de este trabajo fue evaluar los casos de GEUS ocurridos en el país y determinar el papel de adenovirus del pollo en su presentación. Las investigaciones se basaron en una reevaluación retrospectiva de secciones de tejido y bloques de parafina de mollejas e hígados de casos de GEUS entre 2014 y 2021, junto con una investigación prospectiva de casos en el año 2022. Las mollejas y los hígados se fijaron en formalina al 10% amortiguada y neutra para exámenes histopatológicos de acuerdo con métodos estándar y se almacenaron a -20 C para análisis moleculares. Después de la desparafinación, las secciones de tejido de 10 µm de espesor junto con órganos frescos se sometieron a extracción de ADN utilizando un estuche comercial. Para realizar un método de PCR convencional, se utilizó un par de iniciadores específicos para el gene de hexon de los adenovirus del pollo, mientras que, para el método de PCR en tiempo real, se empleó un par de iniciadores dirigidos al gene 52K. En total, se evaluaron 24 casos de parvadas con la presentación característica del síndrome de erosión y ulceración de la molleja entre los años 2014 y 2022. Se trató de nueve casos de pollos de engorde con edades comprendidas entre los 11 y 39 días, 11 casos de ponedoras con una edad de entre 17 y 29 semanas, dos casos en reproductoras pesadas de 10 y 27 semanas, y dos lotes de pavitos de 22 y 23 días. En la mayoría de los casos, las lesiones microscópicas fueron consistentes con una ventriculitis ulcerativa y linfoplasmocítica, y se identificaron cuerpos de inclusión intranucleares virales patognomónicos dentro de células epiteliales degeneradas en cuatro casos de parvadas de pollos de engorde, cuatro casos de ponedoras y un caso en reproductoras y por lo tanto, fueron altamente sugestivos de la infección por adenovirus de pollo. Entre los nueve casos que fueron positivos en el examen histopatológico, se encontró que seis casos fueron positivos para adenovirus de pollo mediante PCR; otros cuatro casos fueron negativos a la histología pero positivos a la presencia de adenovirus del pollo mediante PCR. Además, se realizó un análisis de secuenciación que proporcionó la evidencia inicial del papel de adenovirus del pollo especie A como causante del síndrome de erosión y ulceración de la molleja en la avicultura de Marruecos. Además, se realizó un análisis filogenético para facilitar una comparación entre las cepas investigadas en este estudio y las identificadas en diversas regiones geográficas y en varios períodos de tiempo.

Quantity of virulent fowl adenovirus serotype 1 correlates with clinical signs, macroscopical and pathohistological lesions in gizzards following experimental induction of gizzard erosion in broilers.

In the present study day-old specific-pathogen-free (SPF) and commercial broilers with maternally derived fowl adenovirus serotype 1 (FAdV-1) antibodies were orally infected with a European "pathogenic" FAdV-1, isolated from broilers showing signs of gizzard erosion. During the experiment, broilers were observed and weighed daily up to 17 days post infection (dpi). Clinically, both infected groups showed significant decrease of weight compared to respective negative control groups. Birds were examined by necropsy at 3, 7, 10, 14 and 17 dpi. Pathological changes in the gizzards were noticed in both experimentally infected groups from 7 dpi onwards. Macroscopically, erosion of the koilin layer and inflammation or ulceration of the gizzard mucosa were observed. Histologically, presence of FAdV-1 in intranuclear inclusion bodies of degenerated glandular epithelial cells was demonstrated by in-situ hybridization and inflammatory cell infiltration of the lamina propria, submucosa and muscle layer was detected. Tissue samples were investigated by a recently developed real-time PCR and the viral DNA load was calculated from gizzard, liver, spleen and cloacal swabs with the highest amounts of FAdV-1 DNA found in the gizzard. For the first time, successful reproduction of clinical signs in broilers as well as pathological lesions in the gizzard were achieved with a European FAdV-1 isolate displaying some genetic differences to so far reported virulent FAdV-1 from Japan. Furthermore, highest viral load in gizzards could be linked with macroscopical and histological lesions. Therefore, the conducted analyses provide important insights into the pathogenesis of adenoviral gizzard erosion.

The role of an attenuated anticoccidial vaccine on the intestinal ecosystem and on the pathogenesis of experimental necrotic enteritis in broiler chickens.

The objective of the present study was to investigate the effect of an attenuated anticoccidial vaccination on the intestinal ecosystem and on the pathogenesis of experimental necrotic enteritis (NE) in broiler chickens. Two hundred and forty 1-day-old Cobb 500 broiler chickens were randomly allocated to four treatment groups according to the following experimental design: control Group N; Group PN, where birds were vaccinated with anticoccidial vaccine; Group M, where birds were challenged with Clostridium perfringens and with Eimeria maxima; and Group PM, where birds were both vaccinated and challenged. From each bird, the intestine, gizzard and liver were scored for gross NE lesions. Intestinal digesta were collected for pH and viscosity determination. Samples from the gastrointestinal tract and liver were taken for microbiological analysis. Evaluation of the experimental data revealed that Group M had significantly higher overall mean NE intestinal lesions compared with Group PM. Viscosity values of jejunum digesta as well as pH values of the duodenum, jejunum and ileum digesta in Group M were significantly lower compared with Group PM. C. perfringens counts in the caeca of Group PM were significantly lower compared with Group M. The milder decrease of pH and viscosity values of intestinal content and the reduction of C. perfringens counts in the caeca in challenged and vaccinated birds may explain the lower score of NE gross intestinal lesions and may suggest a positive effect on intestinal ecosystem and a significant protective effect of attenuated anticoccidial vaccination against NE in a subclinical experimental model.

Adenoviral gizzard erosion in broiler chickens in Germany.

Avian adenovirus infections cause important disease complexes in chickens, but many of the viruses also infect chickens without resulting in overt disease. Previously several outbreaks of gizzard erosions caused by a fowl adenovirus A serotype-1 (FAdV-1) were reported from Japan. Here we report an outbreak of gizzard erosions in 12 broiler flocks in Germany in 2011. Chickens had a reduced daily weight gain and a higher total mortality rate of up to 8%. The birds showed a severe detachment of the koilin layer and ulcerative to necrotizing lesions of the underlying mucosa. Histopathologically, necrotizing ventriculitis with basophilic, intranuclear inclusion bodies in epithelial cells was diagnosed. Immunohistochemistry, egg culture, and electron microscopic examination revealed adenovirus-like particles in the samples. No concurrent infectious agent could be identified. The virus was genotyped as FAdV-1 by PCR and subsequent sequencing. Phylogenetic analysis of the hexon loop L1 gene yielded 100% sequence identity to the chicken embryo lethal orphan strain. These findings suggest that outbreaks of adenoviral gizzard erosion can lead to significant economic losses in Germany and may be caused by an unusual virulent FAdV-1 strain.

Vertical transmission and clinical signs in broiler breeders and broilers experiencing adenoviral gizzard erosion.

The present report documents an outbreak of adenoviral gizzard erosion in 22 broiler flocks in Germany. The clinical picture was characterized by uneven growth of affected broilers that resulted in considerably lower than average weight at slaughtering. Fowl adenovirus serotype 1 (FAdV-1) was isolated from gizzard lesions and histological examinations demonstrated FAdV-1-positive intranuclear inclusion bodies in gizzard epithelial cells of affected broilers by in-situ hybridization. Birds from all affected flocks originated from one broiler breeder farm. During production of affected birds, broiler breeders were between 27 and 32 weeks old. Enzyme-linked immunosorbent assay and specific virus neutralization assay of sera from parent birds demonstrated an acute FAdV-1 infection within the first 5 weeks of the production cycle. Clinically, broiler breeders exhibited a moderate fall in the hatchability of their chicks, while egg production remained normal. No further clinical signs could be observed. Genetically identical FAdV-1 strains were isolated from gizzards of embryos at the lowest point of hatchability and from affected broiler flocks raised on independent farms. For the first time, direct detection of viable FAdV-1 from gizzards of embryos and progenies of one FAdV-1-seropositive broiler breeder farm in the course of an outbreak of adenoviral gizzard erosion could be demonstrated, highlighting the importance of vertical transmission of this disease. Additionally, growth retardation and subsequent reduced average weight at the time of slaughter of broiler chickens underline the economic impact of adenoviral gizzard erosion for poultry production.

Non-soluble fibres and narasin reduce spontaneous gizzard erosion and ulceration in broiler chickens.

The prevalence and severity of gizzard erosion and ulceration syndrome (GEU) in broilers is affected by a number of predisposing and preventive factors. Two broiler experiments with almost identical basal diets were conducted to investigate the effect of dietary oat hulls, access to litter and the antimicrobial compound narasin on GEU. The effects on particle size of duodenal digesta, ileal starch concentration, caecal Clostridium perfringens counts, necrotic enteritis and production performance were also examined. Oat hulls reduced GEU severity and starch levels in the ileum in both experiments. Access to litter reduced GEU scores when oat hulls were included in the feed. Access to litter also improved feed efficiency and reduced C. perfringens counts. Narasin reduced GEU severity when the feed was supplemented with oat hulls. Oat hulls were associated with improved feed efficiency in Experiment 1 and impaired feed efficiency in Experiment 2. The inconsistent effect of oat hulls on production performance appeared to be related to an association between oat hulls and high C. perfringens counts in Experiment 2; an association that was absent in Experiment 1. In general, oat hulls interacted with litter access and narasin in exerting a positive effect on gizzard health. However, the association between oat hulls and necrotic enteritis detected in Experiment 2 suggests that the positive effect of oat hulls on GEU occasionally may be outweighed by a negative effect on gut health. In conclusion, our findings demonstrate that the severity of GEU in broilers was influenced by non-soluble fibres and narasin.

Outbreak of gizzard erosion associated with fowl adenovirus infection in Korea.

The pathogenicity of a fowl adenovirus serotype-1 (FAdV-1, K181 strain) isolated from a case of gizzard erosion in layer chickens was investigated in specific-pathogen-free (SPF) chicks. One-week-old SPF chicks were inoculated orally or intramuscularly with the isolate of FAdV-1 and euthanized for necropsy at 7, 14, and 21 d postinoculation. Although there were no clinical signs after inoculation, gizzard erosions were observed grossly and the virus was recovered from the gizzards in the inoculated chickens. Histologically, in the chickens that were infected orally, the lesions found in the gizzard consisted of severe degeneration and necrosis of glandular epitheliums and eosinophilic inclusion bodies. These results indicate that the Korean FAdV-1 isolate could induce gizzard lesions in chickens. Moreover, the present investigation reproduced an outbreak of gizzard erosion caused by FAdV-1 infection and, for the first time, described the isolation of FAdV-1 from chickens in Korea. These findings provide important information on the epidemiology and pathogenesis of FAdV-1 infection in chickens.

High Phenotypic Variation between an In Vitro-Passaged Fowl Adenovirus Serotype 1 (FAdV-1) and Its Virulent Progenitor Strain despite Almost Complete Sequence Identity of the Whole Genomes.

Adenoviral gizzard erosion is an emerging disease with negative impact on health and production of chickens. In this study, we compared in vitro and in vivo characteristics of a fowl adenovirus serotype 1 (FAdV-1), attenuated by 53 consecutive passages in primary chicken embryo liver (CEL) cell cultures (11/7127-AT), with the virulent strain (11/7127-VT). Whole genome analysis revealed near-complete sequence identity between the strains. However, a length polymorphism in a non-coding adenine repeat sequence (11/7127-AT: 11 instead of 9) immediately downstream of the hexon open reading frame was revealed. One-step growth kinetics showed delayed multiplication of 11/7127-AT together with significantly lower titers in cell culture (up to 4 log10 difference), indicating reduced replication efficiency in vitro. In vivo pathogenicity and immunogenicity were determined in day-old specific pathogen-free layer chicks inoculated orally with the respective viruses. In contrast to birds infected with 11/7127-VT, birds infected with 11/7127-AT did not exhibit body weight loss or severe pathological lesions in the gizzard. Virus detection rates, viral load in organs and virus excretion were significantly lower in birds inoculated with 11/7127-AT. Throughout the experimental period, these birds did not develop measurable neutralizing antibodies, prevalent in birds in response to 11/7127-VT infection. Differences in pathogenicity between the virulent FAdV-1 and the attenuated strain could not be correlated to prominently discriminate genomic features. We conclude that differential in vitro growth profiles indicate that attenuation is linked to modulation of viral replication during interaction of the virus with the host cells. Thus, hosts would be unable to prevent the rapid replication of virulent FAdV leading to severe tissue damage, a phenomenon broadly applicable to further FAdV serotypes, considering the substantial intra-serotype virulence differences of FAdVs and the variation of diseases.

Mixed infection with Libyostrongylus dentatus and Libyostrongylus douglassii induces a heterophilic inflammatory infiltrate in the proventriculus of ostriches.

Libyostrongylus dentatus and Libyostrongylus douglassii are haematophagous nematodes found in the proventriculus and the ventriculus of ostriches. Pathological damage leading to bird death has been attributed to L. douglassii. However, histopathology of the mixed infection has not been reported. The aim of the present work was to characterize the cellular inflammatory infiltrate found in the proventriculus of ostriches with a mixed infection. Analysis of the collected nematodes confirmed a mixed infection in the proventriculus of examined birds. Histopathological examination of the proventriculus showed an inflammatory infiltrate composed of granular cells in close proximity to the nematodes. The granulocyte infiltrate was composed mainly of heterophils identified by the lack of peroxidase and presence of fusiform granules.