Avian Adenovirus Research Articles

Novel pathogenic adenovirus in Timneh grey parrot (Psittacus timneh) unveils distinct lineage within Aviadenovirus

Wild birds harbour a vast diversity of adenoviruses that remain uncharacterised with respect to their genome organisation and evolutionary relatedness within complex host ecosystems. Here, we characterise a novel adenovirus type within Aviadenovirus genus associated with severe necrotising hepatitis in a captive Timneh grey parrot, tentatively named as Timneh grey parrot adenovirus 1 (TpAdV-1). The TpAdV-1 genome is 39,867 bp and encodes 46 putative genes with seven hitherto not described ones. Comparative genomics and phylogenetic analyses revealed highest nucleotide identity with psittacine adenovirus 1 and psittacine adenovirus 4 that formed a discrete monophyletic clade within Aviadenovirus lineage suggesting a deep host co-divergent lineage within Psittaciformes hosts. Several recombination breakpoints were identified within the TpAdV-1 genome, which highlighted an ancient evolutionary relationship across the genera Aviadenovirus, Mastadenovirus and Atadenovirus. This study hints towards a host-adapted sub-lineage of avian adenovirus capable of having significant host virulence in Psittaciformes birds augmented with ecological opportunity.

Surveillance for Selected Pathogens and Parasites of Northern Bobwhite (Colinus virginianus) from Western Oklahoma, USA, 2018-20.

The Northern Bobwhite (Colinus virginianus) has been undergoing a range-wide population decline. Potential causes for declines across its historic range have been investigated for decades and include habitat loss and fragmentation and a variety of parasitic and infectious diseases. Although there have been studies on bobwhite ecology in Oklahoma, USA, relatively little is known about parasites and pathogens in the region. We evaluated the health of free-ranging bobwhites from nine sites in western Oklahoma. From 2018 to 2020, 206 bobwhites were evaluated for gross and microscopic lesions and tested for selected pathogens. In general, bobwhites were in good nutritional condition with ample muscle mass and fat stores. No significant gross lesions were observed in any bobwhite and no significant histologic lesions were detected in a subset. There was no evidence of infection with or exposure to reticuloendotheliosis virus, West Nile virus, respiratory Mycoplasmataceae species, Pasteurella multocida, intestinal Eimeria spp., or oral Trichomonas spp. Several pathogens of potential concern were detected, including avian adenovirus (8.6%), Toxoplasma gondii (2.3%), and haemosporidians (a Haemoproteus sp. (1.5%), Leucocytozoon schoutedeni (1.5%), and Plasmodium homopolare haplotype 2 [lineage LAIRI01; 3.6%]). Physaloptera sp. (12%) and Sarcocystis sp. (1%) were detected in the breast muscle. Low intraspecific genetic diversity was noted for Physaloptera sp., and sequences were most similar to Physaloptera sequences from bobwhites and grasshoppers (Orthoptera) in Texas. Low intensities of chewing lice, chiggers, and ticks were observed. A subset of bobwhites had evidence of exposure to selected toxicants and heavy metals; a small number had low levels of iron, manganese, zinc, molybdenum, and copper, which were not considered diagnostically relevant. In general, bobwhites from western Oklahoma appeared to be in good health with a low diversity of pathogens detected, but future work is needed to understand potentially changing disease risks for this population.

Avian adenovirus infections: diversity of pathogens, hazard to poultry industry and problems of immunoprophylaxis (review)

The data on diversity of adenovirus pathogens in nature and the role of the main representatives of the Adenoviridae family in poultry infectious pathology are presented. Special attention is paid to problematic issues of immunoprophylaxis due to lack of cross-immunity between different virus serotypes. There is no single and effective approach in the global strategy of immunoprophylaxis of avian adenoviruses, therefore, improving the means of avian adenovirus disease control is an urgent and important task. Avian adenovirus infections are represented by different nosological units: egg drop syndrome, hydropericardium syndrome, adenoviral gizzard erosion, marbled spleen disease of pheasants, hemorrhagic enteritis of turkeys, inclusion body hepatitis and many unclassified diseases. The paper provides data on the main nosological forms of adenovirus infections that pose a threat to cost-effective poultry farming, and highlights test results obtained by foreign authors on the effectiveness of some vaccines against adenovirus infection. Most vaccines have been developed to prevent avian hydropericardium syndrome, however, occurrence of many virus serotypes requires effective means of prevention and diagnosis in order to control other infections caused by adenoviruses. There is no registered vaccine against adenovirus infections that cause inclusion body hepatitis and adenoviral gizzard erosion. At the same time, inclusion body hepatitis alone accounts for 2.9% of all recorded avian infectious diseases. Vaccines registered in the Russian Federation are not enough to fully control these infections, and that requires a timely solution to the problem. The variety of avian adenoviruses determines the problems of their differential diagnosis and specific prevention.

Variation in the affinity of three representative avian adenoviruses for the cellular coxsackievirus and adenovirus receptor.

According to previous studies, three representative avian adenoviral strains utilize coxsackievirus-adenovirus receptor (CAR) as a receptor and seem to exhibit diverse binding affinities and modes. Thus, further revealing the exact molecular mechanism underlying the interaction between different FAdVs and the attachment receptor CAR is necessary. In this study, we successfully solved the crystal structure of the FAdV-4 fiber1 knob at 1.6 Å resolution. The interaction between the fibre knob and different domains of CAR was verified by confocal microscopy, coimmunoprecipitation and surface plasmon resonance (SPR) analysis. The fibre knobs of the three representative fowl adenoviruses specifically recognized CAR domain 1 (D1), but the recognition of CAR domain 2 (D2) by chicken embryo lethal orphan (CELO) strains was weak. These results provide insights into the differences in adenovirus‒host cell interactions and have important implications for the exploration of viral invasion mechanisms.

Development of a unique sandwich enzyme-linked immunosorbent assay based on monoclonal antibodies for the specific detection of the egg drop syndrome virus

ABSTRACT An abrupt decrease in egg production and quality in laying hens can be brought on by the avian adenovirus known as the egg drop syndrome virus (EDSV). So far, there are no efficient commercial diagnostic approaches for EDSV. In the present study, we first purified two monoclonal antibodies (mAbs 5G4 and 6G6) specifically against the EDSV fibre protein. The mAb 5G4 was then employed as capture antibody. Additionally, the detection antibody 6G6 (HRP-6G6) was conjugated with horseradish peroxidase. Consequently, based on these two mAbs, we developed a sandwich enzyme-linked immunosorbent assay (ELISA) for EDSV detection. Specificity evaluation demonstrated that only EDSV gave a positive result while other avian viruses showed negative results. Sensitivity investigation showed that the limit of detection of EDSV was 102.9 TCID50/ml and the limit of detection of the purified His-Fiber was 5 ng/ml. Furthermore, the screening of samples from infected chickens showed that this method was very effective and could be applied to the detection of EDSV in infected samples. In summary, the sandwich enzyme-linked immunosorbent assay developed here provides an efficient, sensitive, and large-scale method for the specific diagnosis of EDSV. RESEARCH HIGHLIGHTS A sandwich ELISA was developed to detect EDSV using the mAbs 5G4 and HRP-6G6. The sandwich ELISA maintained high specificity and sensitivity. The sandwich ELISA had equivalent consistency with real-time PCR assay.

Structural and antigenic characterization of the avian adeno-associated virus capsid.

AAVs are extensively studied as promising therapeutic gene delivery vectors. In order to circumvent pre-existing antibodies targeting primate-based AAV capsids, the AAAV capsid was evaluated as an alternative to primate-based therapeutic vectors. Despite the high sequence diversity, the AAAV capsid was found to bind to a common glycan receptor, terminal galactose, which is also utilized by other AAVs already being utilized in gene therapy trials. However, contrary to the initial hypothesis, AAAV was recognized by approximately 30% of human sera tested. Structural and sequence comparisons point to conserved epitopes in the fivefold region of the capsid as the reason determinant for the observed cross-reactivity.

Molecular docking and simulation studies of antiviral compounds against to fowl adenovirus type 4 receptor from – Psittacine bird – Melopsittacus undulates

The adenovirus family of adenoviridae includes mammalian adenoviruses (mastadenoviruses) and avian adenoviruses (avi-adenoviruses), which are known to cause cold or flu and HPS. A wide variety of afflicted birds, including chicken, pigeon, and psittacine species, have been reported to carry aviadenoviruses. Fowl adenovirus, which causes hydropericardium syndrome (FAdV). The disease is highly contagious and quickly spreads from one flock to another and from one farm to another using mechanical and horizontal transmission paths as well as contaminated litter. The drug Dehydroepiandrosterone (DHEA) is reported to have a significant binding affinity against the 7W83 receptors (−7.7 kcal/mol) respectively. The study focuses on the therapeutic methodology development of Adenoviral infection. In order to discover useful combinations of the drugs in practice, molecular docking methods were also used to match fowl adenovirus protein with anti-viral compounds. To further support the docking effects, extensive molecular dynamics simulations were also used.

Study on Propagation and Adaptation of EDS-76 Avian Adenovirus in Duck and SPF Primary Embryonic Chicken Cell Culture Comparison to Duck and SPF Embryonated Chicken Eggs.

Egg drop syndrome (EDS) is a major viral infectious poultry disease with severe economic losses in laying hens. The disease is caused by an adenovirus and can be transmitted horizontally and vertically. This study investigated the EDS virus (EDSV) infection in duck embryo fibroblasts (DEF), specific pathogen-free (SPF) embryo fibroblasts, and SPF egg embryos using different methods. The results were compared to the virus culture in duck and SPF chicken eggs. Duck and chicken fibroblast cells were used as the primary cell culture in Dulbecco's Modified Eagle Medium, and the low-pathogenic duck adenovirus was used to infect the ducks and SPF fibroblasts primary cell cultures, as well as the duck and SPF eggs. The titer of the virus was measured by hemagglutination assay, ECID50, plaque-forming unit, and TCID50 methods. The results revealed that EDSV could proliferate in the chorioallantoic membrane of DEF cells and duck eggs, compared to the chorioallantoic membrane of chicken embryo fibroblasts (CEF) and SPF chicken eggs. The findings showed that duck egg embryos and primary DEF cell lines are more appropriate for EDSV replication, compared to CEF and SPF chicken eggs. This suggests that the use of DEF culture for producing avian adenovirus EDS-76 is a suitable alternative for the embryonic egg culture.

Fowl adenovirus serotype 4 enters leghorn male hepatocellular cells via the clathrin-mediated endocytosis pathway

Hepatitis-hydropericardium syndrome (HHS) induced by fowl adenovirus serotype-4 (FAdV-4) has caused large economic losses to the world poultry industry in recent years. HHS is characterized by pericardial effusion and hepatitis, manifesting as a swollen liver with focal necroses and petechial haemorrhage. However, the process of FAdV-4 entry into hepatic cells remains largely unknown. In this paper, we present a comprehensive study on the entry mechanism of FAdV-4 into leghorn male hepatocellular (LMH) cells. We first observed that FAdV-4 internalization was inhibited by chlorpromazine and clathrin heavy chain (CHC) knockdown, suggesting that FAdV-4 entry into LMH cells depended on clathrin. By using the inhibitor dynasore, we showed that dynamin was required for FAdV-4 entry. In addition, we found that FAdV-4 entry was dependent on membrane cholesterol, while neither the knockdown of caveolin nor the inhibition of a tyrosine kinase-based signalling cascade affected FAdV-4 infection. These results suggested that FAdV-4 entry required cholesterol but not caveolae. We also found that macropinocytosis played a role, and phosphatidylinositol 3-kinase (PI3K) was required for FAdV-4 internalization. However, inhibitors of endosomal acidification did not prevent FAdV-4 entry. Taken together, our findings demonstrate that FAdV-4 enters LMH cells through dynamin- and cholesterol-dependent clathrin-mediated endocytosis, accompanied by the involvement of macropinocytosis requiring PI3K. Our work potentially provides insight into the entry mechanisms of other avian adenoviruses.

Development and Evaluation of a Blocking Lateral Flow Assay Strip for Detection of Newcastle Disease Virus Antibodies.

Newcastle disease (ND) is an acute septicemic infectious disease caused by Newcastle disease virus (NDV). Considering that vaccination is currently the main modality for the prevention of ND, it is essential to assess the effectiveness of clinical immunization. In this study, we have developed a blocking lateral flow assay (bLFA) strip for the rapid detection of NDV antibodies using the monoclonal antibody 9C1 against haemagglutinin-neuraminidase (HN), which allows for the determination of an NDV-specific antibody titer within 10 min at room temperature. In addition, the bLFA strip has no cross-reactivity with the positive serum of other avian pathogens including avian influenza subtypes H5, H7, and H9, MD, IBD, IB, EDS, and avian adenovirus. The ability of the bLFA strip for detecting a neutralizing antibody was also estimated. The results showed that the chicken NDV hyperimmunized serum had a complete blocking (100%) titer of 11 log 2, and half-blocking titer of 13 log 2, which are 4 times less than and the same as that of the HI test (13 log 2), and 8 and 2 times less than that of the VN test (14 log 2), respectively. A total of 510 clinical samples were tested for NDV antibodies. The coincidence rate between the results of the bLFA strip and HI test was 97.65%. Therefore, it is an ideal alternative method for assessing the clinical immunity of ND vaccines in the field in real-time.

Pathogenicity of Avian Adenoviruses Type D and Type E Isolated from Field Cases of Inclusion Body Hepatitis–Hydropericardium syndrome (IBH–HPS) in Broiler chickens in Egypt

In this study, 210 one-day-old commercial broiler chicks were used to investigate the pathogenicity of two types of Avian Adenoviruses (type D and type E) recently isolated from field cases. These chicks were divided into four groups; birds of group 1 (G1) and group 2 (G2) were experimentally inoculated with two local isolates of fowl aviadenovirus FAdV D-2 and FAdV D-11, respectively, while chicks of group 3 (G3) were experimentally inoculated with FAdV-E 8a. All birds of these three groups were experimentally inoculated orally at one-day of age with a dose of 0.5mL (107 TCID50/mL). Birds of group 4 (G4) served as uninoculated blank controls. Clinical signs, gross lesions and histopathological changes were examined at different intervals post inoculation. Birds of all infected groups showed hydropericardium, focal hepatic necrosis and ecchymotic hemorrhage of thigh and breast muscles. Hepatitis, glomerulonephritis, pericarditis, myocarditis and lymphoid depletion in the spleen were the main histopathological observations. Mortality rates of 12, 16 and 6% were observed in chicks of G1, G2 and G3, respectively, indicating that FAdV-D isolates were more virulent than FAdV-E. Hemagglutination inhibition test revealed significant decrease in antibodies titers against inactivated NDV vaccine in birds of the three FAdVs infected groups compared to the chicks of the control group. In Egypt, several studies reported Adenoviruses infections in broilers; however, to our knowledge this is the first study in which the pathogenicity of FAdVs isolates has been reported. It was concluded that FAdV-D 2, FAdV-D 11 and FAdV-E 8a are primary agents, in addition to possibility of these isolates to act as neglectable immunosuppressive agents incriminating to vaccination failure in chickens.

Adenoviruses in Avian Hosts: Recent Discoveries Shed New Light on Adenovirus Diversity and Evolution.

While adenoviruses cause infections in a wide range of vertebrates, members of the genus Atadenovirus, Siadenovirus, and Aviadenovirus predominantly infect avian hosts. Several recent studies on avian adenoviruses have encouraged us to re-visit previously proposed adenovirus evolutionary concepts. Complete genomes and partial DNA polymerase sequences of avian adenoviruses were extracted from NCBI and analysed using various software. Genomic analyses and constructed phylogenetic trees identified the atadenovirus origin from an Australian native passerine bird in contrast to the previously established reptilian origin. In addition, we demonstrated that the theories on higher AT content in atadenoviruses are no longer accurate and cannot be considered as a species demarcation criterion for the genus Atadenovirus. Phylogenetic reconstruction further emphasised the need to reconsider siadenovirus origin, and we recommend extended studies on avian adenoviruses in wild birds to provide finer evolutionary resolution.

Pigeon Rotavirus A as the cause of systemic infection in juvenile pigeons (young pigeon disease).

Recent investigations suggested pigeon associated Rotavirus Typ A genotype G18P[17] (RVA) as a causative agent of the classical 'young pigeon disease' (YPD). YPD was first described in the late 1980 s as an acute, mainly seasonally recurring disorder of mostly juvenile domestic pigeons (Columba livia) with clinical signs such as anorexia, dairrhea, vomiting, congested crops, weight loss and occasionally mortality. Various studies in the past indicated a multifactorial nature of YPD. Several pathogens, such as pigeon circovirus 1, avian adenoviruses and Escherichia coli were also suggested, but none of these could reproduce the disease experimentally. However, the impact of other pathogens on the clinical development of YPD cannot be excluded and requires further investigation. This present review summarizes available information on RVA-induced disease in pigeons, its association with YPD, the transmission, and diagnosis of the infection, and on prophylactic strategies to prevent RVA outbreaks.

Epidemiological survey of avian adenovirus in China from 2015 to 2021 and the genetic variability of highly pathogenic Fadv-4 isolates.

The prevalence of poultry adenovirus in China is determined using clinical diagnosis, molecular biological testing, serological testing, and LMH cell virus isolation. These methods can track and test key poultry and waterfowl breeding areas across the country. From 2015 to 2021, 9613 suspected adenovirus samples were collected from 28 provinces. After the first generation of gene sequencing, a total of 2210 hexo gene fragments were obtained. Among them, FAdV-1 type accounted for 7.65%, FAdV-2 type accounted for 5.34%, FAdV-3 type accounted for 2.04%, FAdV-4 type accounted for 38.24%, FAdV-5 type accounted for 2.17%, FAdV-6 type accounted for 0.32%, FAdV-7 type accounted for 0.77%, FAdV-8a type accounted for 10.63%, FAdV-8b type accounted for 11.58%, FAdV-9 type accounted for 0.50%, FAdV-10 type accounted for 8.10%, and FAdV-11 type accounted for 12.67%. A total of 877 FAdV strains were isolated from FAdV suspected samples by seeding LMH cells, and there were 475 FAdV-4 strains among them. A total of 473 isolates were highly pathogenic FAdV-4, and the percentage of amino acid homology with the highly pathogenic FAdV-4 reference strains was >99.1%. Two isolates were non-pathogenic, and the amino acid homology with the ON1 reference strain was >99.6%. Part of the amino acid positions of the hexon gene have mutations, including positions 188, 193, 195, 238, and 240.

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

ABSTRACT 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 HIGHLIGHTS AGE 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.

HYDROPERICARDIUM HEPATITIS SYNDROME IN CHICKENS/ARTICLE REVIEW

Hydropericardium hepatitis syndrome is an acute infectious disease of broiler chickens characterized by high mortality, accumulation of fluids in the pericardial sac and necrosis of hepatocytes, during 1987 the virus was isolated from chickens in Angara near Karachi city in Pakistan and the disease takes it is name of this city. It is reported in China in 2015 in birds of 3-6 weeks in broiler chickens causing sudden death with high mortality reaches up to 75%. Avian adenovirus divided into 3genrea: Atadodenovirus, Siadenovirus, and Aviadenovirus the last one divided in to 5 species A to E subgroup1 strain 4 causing (Hydropericardium hepatitis syndrome). Subgroup 2 causing turkey (hemorrhagic enteritis, marble spleen disease). Subgroup 3 causing (Egg drop syndrome EDS). strain D and E produce inclusion body hepatitis while strain 1 causing (quill bronchitis). outbreaks of the disease producing economic losses for the poultry industry. Both vertical and horizontal transmission have an important role in the spreading of the disease. gross examination of the liver showing friable, enlargement, paleness of the liver. the kidney is swelling and hemorrhage with pale areas of necrosis in the parenchyma, pericardium is filled with clear or jelly fluids but histological changes showing hepatitis, nephritis and presence of basophilic intranuclear inclusion bodies. There are many tests can be used for the diagnosis such as PCR, ELISA, neutralization test. We concluded that the disease has economic importance due to high mortality and high losses of chicken’s meat production, so, biosecurity and vaccination are essential for preventing the infection.

Complete Genome Sequence, Molecular Characterization and Phylogenetic Relationships of a Novel Tern Atadenovirus

Discovery and study of viruses carried by migratory birds are tasks of high importance due to the host’s ability to spread infectious diseases over significant distances. With this paper, we present and characterize the first complete genome sequence of atadenovirus from a tern bird (common tern, Sterna hirundo) preliminarily named tern atadenovirus 1 (TeAdV-1). TeAdV-1 genome is a linear double-stranded DNA molecule, 31,334 base pairs which contain 30 methionine-initiated open reading frames with gene structure typical for Atadenovirus genus, and the shortest known inverted terminal repeats (ITRs) within the Atadenovirus genus consisted of 25 bases. The nucleotide composition of the genome is characterized by a low G + C content (33.86%), which is the most AT-rich genome of known avian adenoviruses within Atadenovirus genus. The nucleotide sequence of the TeAdV-1 genome shows high divergence compared to known representatives of the Atadenovirus genus with the highest similarity to the duck atadenovirus 1 (53.7%). Phylogenetic analysis of the protein sequences of core genes confirms the taxonomic affiliation of the new representative to the genus Atadenovirus with the degree of divergence from the known representatives exceeding the interspecies distance within the genus. Thereby we proposed a novel TeAdV-1 to be considered as a separate species.

Isolation and Molecular Characterization of Fowl Adenoviruses Associated with Inclusion Body Hepatitis-hydropericardium Syndrome in Broiler Chickens in Egypt

Avian adenoviruses are an extremely diversified group of pathogens that recently triggering a variety of problems for poultry production. In particular, Inclusion Body Hepatitis-Hydropericardium Syndrome (IBH-HPS), which has been observed in broiler birds from 3 to 6 weeks of age and is associated with anemia, hemorrhagic disorders, hydropericardium, and high mortality. The disease has been reported worldwide, and recently it was reported in many Egyptian farms, causing severe economic losses. Therefore, the current study aimed to isolate, and genetically type the most common Adenovirus serotypes associated with this syndrome in Egyptian farms. A total of 50 broiler chicken farms (3-6 weeks old) located in different Egyptian governorates were examined. Macroscopically, the diseased flock revealed hydropericardium, enlarged friable livers with ecchymotic hemorrhages, and varying mortality rates (1 to 7.5%). Histopathologically, severe diffuse necrotizing enteritis, hepatitis, pericarditis, and diffuse lymphoid depletion of the spleen were the most prominent lesions. Liver tissues and cloacal swabs were collected from all examined flocks for FAdVs detection by conventional polymerase chain reaction (PCR) targeting the L1 loop in the hexon gene. The PCR products were sequenced for typing of the detected viruses. It was found that 10 out of 50 flocks examined were PCR positive for FAdVs (20%). Phylogenetic analysis of the sequenced genes revealed that the obtained viruses clustered with reference strains belonging to FAdV type D and E serotype 2, 11, and 8a respectively. The isolation of both FAdV type D and FAdV type E were carried out on a primary cell culture chicken embryo liver cell (CEL) and the presence of these viruses was confirmed by PCR after the appearance of cytopathic effect (CPE). From this study, it could be concluded that both FAdVs types D and E are the most common adenoviruses circulating in poultry farms suffering from hydropericardium and inclusion body hepatitis.

Оценка методов инактивирования аденовируса птиц при производстве гриппозных вакцин

Inactivation of influenza virus and other potential contaminants like avian adenoviruses coming from embryonated chicken eggs is a critical step in the production of inactivated influenza vaccines. Inactivation must lead to a guaranteed reduction in contaminant titers by at least 4 lg (PFU)/ml. The aim of this study was to identify an optimum cell line for adenovirus propagation and to estimate a reduction in adenovirus titers in vaccine intermediates after inactivation. In a series of experiments, we identified the optimum conditions and the optimum cell line for the propagation of avian adenovirus (strains CELO and Fontes). The most commonly used inactivation methods were analyzed, including inactivation by β-propiolactone and UV light. Viral titers were measured by plaque assays. After 10 h of inactivation with β-propiolactone, CELO titers fell by 4.12 ± 0.06 lg, whereas Fontes titers, by 4.20 ± 0.19 lg, suggesting that β-propiolactone is an effective inactivating agent. Exposure to UV light led to a reduction in CELO titers by 4.69 ± 0.89 lg and a reduction in Fontes titers by 4.44 ± 1.06 lg after 5 min. N-octyl-β-D-glucopyranoside added at the splitting step reduced CELO titers by 0.93 ± 0.15 lg and Fontes titers by 1.04 ± 0.12 lg, whereas tetradecyltrimethylammonium bromide led to a reduction in CELO and Fontes titers by 1.18 ± 0.17 lg and 1.12 ± 0.38 lg, respectively.

The fowl adenovirus serotype 4 (FAdV-4) induce cellular pathway in chickens to produce interferon and antigen-presented molecules (MHCI/II)

FAdV-4 is the major strain of adenovirus that responsible for hydro-pericardial syndrome (HPS) in poultry. In this study, the virus's specific gene fragments were isolated from clinically suspected cases and amplified by PCR. Finally, after a viral infection to investigate the immune response of the host, the gene expression of MHC (major histo-compatible) molecules (MHCIα, MHCIIβ), Ii (Invariant Chain) gene, inflammatory cytokines (IFN-β, IFN-γ, and IL-1β), and transcription factors (MDA5, STING, IRF7, and NF-kB) were detected by real-time PCR (fluorescence technology). The results of sequence comparison showed that the clinically isolated virus was 100% homologous to a virulent strain of avian adenovirus group C serotype 4 (FAdV-4), which were named AH-FAdV-4. The TCID50 and pathogenicity of the virus were determined that was 106.52/0.1 mL with a mortality rate of 100% in chickens and 0% in ducks. Furthermore, results showed that the expression level of MHCIα, MHCIIβ, and Ii genes in chicken embryo kidney cells significantly (P < 0.01) upregulated (increased) after infection, which was 43, 5.2, and 2.5 times higher than the control group. With the addition of PDTC, an inhibitor of NF-kB, then the expression level of MHCIα, MHCIIβ, and Ii was decreased significantly (P < 0.01) than the control group. The transcription levels of these genes were decreased 0.64, 0.27, and 0.26 respectively. Simultaneously, the expression levels of IFN-β, IFN-γ, and IL-1β were also significantly (P < 0.01) up-regulated (increased) 7.8, 22.7, and 5 times higher than the control group. It was found that up-regulation of STING and NF-κB pathways are directly involved in the regulation of inflammatory cytokines (IFN-β, IFN-γ, and IL-1β), MHC molecules (MHCIα, MHCIIβ), and Ii gene. The results also showed that the gene regulation pathways consecutively increased the expression levels of MDA5, STING, IRF7, and NF-kB. It is conducted that the expression levels of cytokines, MHC molecules, and li gene were increased by STING and NF-kB pathways.