Hepatitis-hydropericardium Syndrome Research Articles

Knob domain of Fiber 2 protein provides full protection against fowl adenovirus serotype 4

Highly pathogenic fowl adenovirus serotype 4 (FAdV-4) is an acute infectious disease with severe economic impact, causing chicken hepatitis hydropericardium syndrome (HHS) and high mortality. In the present study, we evaluated the immunogenicity of the recombinant Fiber2-knob protein (F2-Knob) as an FAdV-4 candidate subunit vaccine in 14-day-old SPF chickens. The knob domain is the functional region of the viral surface protein Fiber2. The protein was expressed in Escherichia coli and was administered a single immunization with different vaccine doses. The protective efficacy was evaluated by mortality, clinical symptoms, virus shedding and histopathological examinations after challenged with the FAdV-4. The results showed that the level of ELISA antibodies of the chickens immunized with Fiber2-knob protein was significantly higher than that of the chickens immunized with an inactivated vaccine against FAdV-4. The antibody value of the immunized Fiber2-knob protein was positively correlated with the increase in immunization dose. The challenge experiment showed that the F2-Knob protein provided full protection against virulent FAdV-4 challenge and significantly reduced viral shedding. These results suggest that F2-Knob protein could be a novel vaccine candidate provide insights to control FAdV-4.

Design and Identification of a Novel Antiviral Affinity Peptide against Fowl Adenovirus Serotype 4 (FAdV-4) by Targeting Fiber2 Protein.

Outbreaks of hydropericardium hepatitis syndrome caused by fowl adenovirus serotype 4 (FAdV-4) with a novel genotype have been reported in China since 2015, with significant economic losses to the poultry industry. Fiber2 is one of the important structural proteins on FAdV-4 virions. In this study, the C-terminal knob domain of the FAdV-4 Fiber2 protein was expressed and purified, and its trimer structure (PDB ID: 7W83) was determined for the first time. A series of affinity peptides targeting the knob domain of the Fiber2 protein were designed and synthesized on the basis of the crystal structure using computer virtual screening technology. A total of eight peptides were screened using an immunoperoxidase monolayer assay and RT-qPCR, and they exhibited strong binding affinities to the knob domain of the FAdV-4 Fiber2 protein in a surface plasmon resonance assay. Treatment with peptide number 15 (P15; WWHEKE) at different concentrations (10, 25, and 50 μM) significantly reduced the expression level of the Fiber2 protein and the viral titer during FAdV-4 infection. P15 was found to be an optimal peptide with antiviral activity against FAdV-4 in vitro with no cytotoxic effect on LMH cells up to 200 μM. This study led to the identification of a class of affinity peptides designed using computer virtual screening technology that targeted the knob domain of the FAdV-4 Fiber2 protein and may be developed as a novel potential and effective antiviral strategy in the prevention and control of FAdV-4.

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.

Fowl Adenovirus Infection – Potential Cause of a Suppressed Humoral Immune Response of Broilers to Newcastle Disease Vaccination

Abstract Fowl adenovirus infections have a significant economic impact, especially in the production of broilers. It is considered the leading cause of three syndromes: adenoviral gizzard erosions and ulcerations, inclusion body hepatitis, and hepatitis-hydropericardium syndrome. A critical feature of this virus is its immunosuppressive effect, via suppressing humoral and cellular immunity. In this study, we examined the humoral immune response after administration of the Newcastle disease vaccine in broiler flocks with previously confirmed seroconversion against Fowl adenovirus. The study was conducted on 5 farms. A total of 220 chickens, five weeks of age, showing no clinical signs of the disease, were included in this study. The control group consisted of 20 chickens from a negative farm. Chickens were vaccinated with commercially available live NDV vaccines between 11 and 13 days of life. ELISA determined the presence of specific antibodies against FAdV in a total of 130/200 (65%) blood sera. Depending on the farm, seroprevalence ranged from 30-100%. The presence of specific antibodies against NDV was determined three weeks after vaccination using the hemagglutination inhibition assay. A positive hemagglutination inhibition (HI) titer (≥ 16) was found in 41/200 (20.5%) sera, which was significantly less compared to the control farm, where a positive HI titer was found in 20/20 (100%) sera. The results of our study indicate the immunosuppressive effect of FAdV in subclinically infected birds and highlight the need for its diagnosis, prevention, and control.

An Inactivated Novel Trivalent Vaccine Provides Complete Protection against FAdV-4 Causing Hepatitis-Hydropericardium Syndrome and FAdV-8b/-11 Causing Inclusion Body Hepatitis

Outbreaks of hepatitis-hydropericardium syndrome (HHS) caused by fowl adenovirus serotype 4 (FAdV-4) and inclusion body hepatitis (IBH) related to FAdV-8b and FAdV-11 have been increased in chickens in China since 2015. Clinical concurrent infections of FAdV-4, FAdV-8b, and FAdV-11 are quite common, yet there are no commercially available trivalent vaccines against infection by these three serotypes. In our previous study, a bivalent vaccine based on a recombinant FAdV-4, of which fiber-1 was replaced with the fiber of FAdV-8b, has been developed. In this study, a novel recombinant rFAdV-4-fiber/8b + 11 was constructed by inserting FAdV-11 fiber gene into the 1966-bp deletion region of rFAdV-4-fiber/8b genome. The in vitro replication ability of the rFAdV-4-fiber/8b + 11 was similar to the parental FAdV-4. One dose immunization with the inactivated rFAdV-4-fiber/8b + 11 vaccine generated robust immune responses against FAdV-4, FAdV-8b, and FAdV-11, and provided efficient clinical protection against FAdV-4, FAdV-8b, and FAdV-11 challenge. This study provides a novel strategy for developing potential trivalent vaccines for the prevention and control of HHS and IBH.

Pathogenicity of duck circovirus and fowl adenovirus serotype 4 co-infection in Cherry Valley ducks

Duck circovirus (DuCV) is one of the most prevalent infectious viruses in the duck industry in China. Although the clinical symptoms vary, it often causes immunosuppression in the host and leads to secondary infection with other pathogens. Fowl adenovirus serotype 4 (FAdV-4) mainly infects chickens and causes hydropericardium hepatitis syndrome. However, the incidence of infection in ducks has increased in recent years, and the phenomenon of mixed infection with DuCV is very common, resulting in more severe clinical morbidity. However, there is no systematic study evaluating the presence of mixed infection. To explore the synergistic pathogenicity of DuCV and FAdV-4 co-infection in Cherry Valley ducks, a comparative experiment was established between DuCV and FAdV-4 co-infection and single infection animal models. It was found that DuCV and FAdV-4 co-infected ducks showed more pronounced clinical signs of pericardial effusion, hepatitis and immunosuppression; more severe tissue damage in target organs; and more significant levels of viral load, biochemical indicators and immune indicators in various organs compared with Cherry Valley ducks infected with just one virus. The results showed that co-infection with DuCV and FAdV-4 may promote greater viral replication, causing more severe tissue damage and immunosuppression than infection with just one virus. Therefore, the monitoring and prevention of the two viruses should be strengthened clinically, with a particular focus on the potential harm of DuCV as it carries the highest infection rate.

A Comprehensive Review on Angara Disease in Poultry

Hepatitis-hydropericardium syndrome (HHS) or Angara disease is an immunosuppression and contagious disease that often occurs in broiler poultry farms with a high mortality rate and sudden death. Furthermore, weight loss and sudden death have a significant and apparent economic impact on poultry farms, especially in fast-growing countries. Fowl adenovirus (FAdV) is a causative agent of HHS found in various animals, such as chicken. It causes rapid death and outbreak in young broiler farms, with an accumulation of jelly-like and straw-colored liquid in the heart pericardium, swollen and pale liver, heart muscles petechial and ecchymosis hemorrhages, swollen and pale kidneys, and atrophy thymus seen in gross pathology. Age, host, farm management, biosecurity plan, co-infectious, and level of immunity are risk factors for HHS. Angara is mainly found in Asian countries in Central and South America. The disease is transmitted vertically, like egg-transmitted, and horizontally by oral-fecal route. Rapid diagnostics and active monitoring systems in poultry farms can prevent and control Angara disease in farms. Vaccination and farm-specific biosecurity strategies reduce the initiation of outbreaks in farms. Treatment is based on activating humoral immunity against the virus, similar to vaccination.

Inclusion Body Hepatitis in commercial broiler chickens of an organised poultry farm – A case study

Adenoviruses cause a wide range of diseases among all avian species. Even though most of them cause mild disease, some are associated with specific clinical conditions. Fowl adenoviruses (FAdVs) are the etiological agents of two important diseases; Inclusion body hepatitis (IBH) and Hepatitis hydropericardium syndrome (HHS) in poultry. Ten dead birds of 45 days old were brought for necropsy to the post-mortem facility available at the Division of Pathology, ICAR-IVRI, Izatnagar. The percentage of mortality was nearly 15 per cent and affected birds were dull, depressed with pallor of comb and ruffled feathers. On post-mortem examination, it was observed that the liver was swollen and yellow with fibrinous deposits over the surface. On microscopic examination, large basophilic intranuclear inclusion bodies were present in hepatocytes which was suggestive of IBH. Diagnosis was further confirmed by Polymerase Chain Reaction (PCR).

Síndrome da hepatite-hidropericárdio por Aviadenovirus e em reprodutores de corte

ABSTRACT: Fowl aviadenovirus (FAdV) is an important pathogen in the global poultry industry and the etiology of inclusion body hepatitis-hydropericardium syndrome (HHS) in chickens. Since the 1990s, several outbreaks of HHS have occurred in poultry producing areas, including South America. The coinfection of FAdV and chicken anemia virus (CAV) may markedly impact the incidence of HHS. This study describes an outbreak of HHS in coinfection with CAV in industrial broiler breeders and characterizes the FAdV isolate. The three-week-old male broiler breeders had pale bone marrow, enlarged and yellowish liver, splenomegaly, and atrophied thymus; one chicken was also found with hydropericardium. Virus isolation was performed in SPF chicken embryos of liver and thymus. Tissues of the naturally infected chickens and the inoculated embryos were evaluated by PCR and histopathology. All affected chickens and inoculated embryos were positive for FAdV and CAV. The inoculated embryos had enlarged, greenish and hemorrhagic livers, and 30% died within 7 days of inoculation. Phylogenetic analysis of the FAdV isolate hexon gene partial sequence enabled grouping with E species. The E species has recently become a relevant species in several countries. The association of FAdV with CAV in breeders is of further concern due to both being capable of vertical transmission. Within the last decade, a worldwide upsurge of HHS in broiler breeders owing to failed biosecurity has occurred. In this episode, the failure on biosecurity may have enabled challenge with both FAdV and CAV, with pathological synergism. The CAV-impaired adaptive immunity may have benefited the FAdV infection.

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.

Fiber-1 of serotype 4 fowl adenovirus mediates superinfection resistance against serotype 8b fowl adenovirus.

In recent years, hepatitis-hydropericardium syndrome (HHS) and inclusion body hepatitis (IBH) caused by serotype 4 fowl adenovirus (FAdV-4) and serotype 8b fowl adenovirus (FAdV-8b), respectively, are widely prevalent in China, causing huge economic losses to the poultry industry. Numerous studies have revealed the mechanism of the infection and pathogenesis of FAdV-4. However, little is known about the mechanism of infection with FAdV-8b. Among the major structural proteins of fowl adenoviruses, fiber is characterized by the ability to recognize and bind to cellular receptors to mediate the infection of host cells. In this study, through superinfection resistance analysis and an interfering assay, we found that Fiber-1 of FAdV-4, rather than hexon, penton, and fiber of FAdV-8b, conferred efficient superinfection resistance against the infection FAdV-8b in LMH cells. Moreover, truncation analysis depicted that the shaft and knob domains of FAdV-4 Fiber-1 were responsible for the inhibition. However, knockout of the coxsackie and adenovirus receptor (CAR) in LMH cells inhibited the replication of FAdV-8b only at early time points, indicating that CAR might not be the key cell receptor for FAdV-8b. Overall, our findings give novel insights into the infection mechanism of FAdV-8b and provide a new target for the prevention and control of both FAdV-4 and FAdV-8b.

Molecular Diagnosis of Adeno Virus Associated with Hydropericardium Hepatitis Syndrome of the Broiler Chickens in Nineveh Province, Iraq

Molecular Diagnosis of Adeno Virus Associated with Hydropericardium Hepatitis Syndrome of the Broiler Chickens in Nineveh Province, Iraq

Phylogenetic and pathogenic characterization of current fowl adenoviruses in China.

In recent years, fowl adenoviruses (FAdVs) continue to outbreak and cause huge economic losses to the poultry industry in China. The homologous recombination accounts for the diversity serotypes of adenovirus. However, the prevalence, recombination and pathogenicity of current FAdVs remain unclear. Herein, the prevalence, phylogenetic feature and pathogenicity of FAdVs in China in 2019 were characterized. Our findings showed that multiple species and serotypes of FAdVs currently circulate in China, including A, C, D and E species, and 1, 2, 4, 8a and 8b serotypes. Notably, the recombination occurred between FAdV-8a and FAdV-8b, and the recombination regions included Hexon, Fiber, ORF19 and ORF20. All five FAdVs replicated effectively in various chicken tissues, and viral shedding peaked at 4-8 dpi. Except CH/GDSZ/1905(FAdV-1/A), the remaining FAdVs caused obvious inclusion body hepatitis (IBH) in 3-week-old specific-pathogen-free (SPF) chickens, of which CH/JSXZ/1905(FAdV-4/C) caused hydropericardium-hepatitis syndrome (HHS) with a mortality rate of 62.5%. Taken together, our findings illustrate the prevalence, recombination and pathogenicity of current FAdVs in China and strengthen surveillance and further pathogenicity studies of FAdVs are extremely urgent.

Local cellular immune response plays a key role in protecting chickens against hepatitis-hydropericardium syndrome (HHS) by vaccination with a recombinant fowl adenovirus (FAdV) chimeric fiber protein.

Fowl adenovirus (FAdV)-induced diseases hepatitis-hydropericardium syndrome (HHS) and inclusion body hepatitis (IBH) have been affecting the poultry industry with increasing severity in the last two decades. Recently, a subunit vaccine based on a chimeric fiber protein with epitopes from different fowl adenovirus serotypes (named crecFib-4/11) has been shown to confer simultaneous protection against both HHS and IBH. However, the underlying immune mechanisms in chickens are still enigmatic, especially because of frequently absent neutralizing response despite high levels of protection. In this study, we investigated the kinetics of the humoral and cellular immune responses in specific pathogen-free chickens after vaccination with crecFib-4/11 and/or challenge with a HHS-causing strain, on a systemic level, as well as locally in target and lymphoid organs. The humoral response was assessed via enzyme-linked immunosorbent assay (ELISA) and virus neutralization test in serum, while the cellular immune response was determined by phenotyping using flow cytometry. Although vaccination induced serum antibodies, as confirmed by ELISA, such antibodies exhibited no pre-challenge neutralizing activity against FAdV-4. Nevertheless, immunized birds experienced a significant B cell increase in the liver upon challenge, remaining high throughout the experiment. Furthermore, vaccination stimulated the proliferation of cytotoxic T lymphocytes, with earlier circulation in the blood compared to the challenge control and subsequent increase in liver and spleen. Overall, these findings imply that protection of chickens from HHS after crecFib-4/11 vaccination relies on a prominent local immune response in the target organs, instead of circulating neutralizing antibodies.

Epidemiology, pathology, prevention, and control strategies of inclusion body hepatitis and hepatitis-hydropericardium syndrome in poultry: A comprehensive review.

Infection with fowl adenoviruses (FAdVs) can result in a number of syndromes in the production of chicken, including inclusion body hepatitis (IBH), hepatitis-hydropericardium syndrome (HHS), and others, causing enormous economic losses around the globe. FAdVs are divided into 12 serotypes and five species (A–E; 1–8a and 8b−11). Most avian species are prone to infection due to the widespread distribution of FAdV strains. The genus aviadenovirus, which is a member of the adenoviridae family, is responsible for both IBH and HHS. The most popular types of transmission are mechanical, vertical, and horizontal. Hepatitis with basophilic intranuclear inclusion bodies distinguishes IBH, but the buildup of translucent or straw-colored fluid in the pericardial sac distinguishes HHS. IBH and HHS require a confirmatory diagnosis because their clinical symptoms and postmortem abnormalities are not unique to those conditions. Under a microscope, the presence of particular lesions and inclusion bodies may provide clues. Traditional virus isolation in avian tissue culture is more delicate than in avian embryonated eggs. Additionally, aviadenovirus may now be quickly and precisely detected using molecular diagnostic tools. Preventive techniques should rely on efficient biosecurity controls and immunize breeders prior to production in order to protect progeny. This current review gives a general overview of the current local and global scenario of IBH, and HHS brought on by FAdVs and covers both their issues and preventative vaccination methods.

Genetic evolution of fowl adenovirus serotype 4 and its pathogenicity to Cherry Valley ducks in China

Hydropericardium hepatitis syndrome (HHS), a novel poultry disease, is caused by fowl adenovirus 4 (FAdV-4). It mainly infects 3–5-week-old broilers. In July 2015, the first outbreak of HHS occurred in the broilers in east China, which caused great economic losses to the poultry industry. In June 2019, infectious disease was detected with suspected HHS symptoms on a duck farm in Linyi City, Shandong Province. The main necropsy lesions included pericardial effusion and hepatitis. In this study, we isolated a strain of FAdV-4 from naturally infected ducks and named it SDLY190604, and the hexon gene sequence was amplified and analyzed using polymerase chain reaction (PCR). In order to study the effect of FAdV-4 on Cherry Valley ducks, we inoculated three-week-old ducks with 0.2 ml of FAdV-4 virus fluid (TCID50 of 10-6.3/0.1 ml) by orally, subcutaneously and intramuscularly. Clinical signs, gross lesions and histopathological changes, cytokines and viral load were detected and recorded within 15 days after infection. The results showed that ducks in the experimental groups exhibited typical symptoms of hydropericardium and hepatitis. The histopathological sections showed multiple-organ damage, including serious liver and kidney damage with elevated levels of inflammatory cytokines, probably due to the infection and innate immune response. Later, immunosuppression occurred, resulting in decreased levels of cytokines. The viral load indicated that the virus could be present in several organs of the ducks, with the highest viral DNA found in the liver, followed by the kidney. Compared to the subcutaneous and oral groups, the intramuscular group exhibited the highest viral load. In summary, this study can increase our understanding of the pathogenicity of FAdV-4 in ducks and provide a basis for further understanding of the virus, imparting new insights into disease research.

Pathogenicity Assessment and Vaccine Efficacy of Fowl Adenovirus Serotype 4 and 11 Responsible for Inclusion Body Hepatitis Hydropericardium Syndrome in Broilers

Thisstudy aimed to assess the pathogenicity and vaccine efficacy of fowl adenovirus serotypes 4 and 11, causing inclusion body hepatitis hydropericardium syndrome in broilers. The study was carried out on 144-day-old Cobb broiler chicks divided into six experimental groups, each of 24 birds. Group, I birds served as a control without any vaccine or viral challenge, while birds of Group II were vaccinated with commercial vaccine on 6th day. Group III and IV birds were non-vaccinated and challenged with 107 TCID50 of FAdV serotype 4 and 11, respectively, on the 27th day. In contrast, Group V and VI birds were vaccinated on the 6th day and challenged with 107 TCID50 of FAdV serotype 4 and 11 on the 27th day. The challenge of FAdV serotype 4 led to 66.66% mortality, and serotype 11 produced 5.00% mortality in the experimental groups. The challenge of the virus led to the production of clinical signs such as depression, huddling, reduced feed intake, reluctance to move, and terminal gasping before death. Characteristic gross lesions of IBH-HPS were observed in birds that died after the challenge of serotypes 4 and 11 of FAdV. The liver was enlarged with focal areas of necrosis, subcapsular hemorrhages, and mild fatty changes. The heart was flabby, congested, and showed hydropericardium with an accumulation of clear watery to a yellow jelly-like fluid in the pericardial sac. The kidneys were enlarged and hemorrhagic. On microscopic examination, the liver showed multifocal areas of necrosis with infiltration of mononuclear cells, mild fatty changes and large basophilic intranuclear inclusion bodies. The birds that died after the challenge of the virus were positive for the presence of FAdV upon PCR. The vaccine efficacy study indicated that the vaccine provided satisfactory protection against Fowl Adenovirus challenge in experimental groups V and VI.

Hsp70 Inhibits the Replication of Fowl Adenovirus Serotype 4 by Suppressing Viral Hexon with the Assistance of DnaJC7.

Fowl adenovirus serotype 4 (FAdV-4) infection results in serious hepatitis-hydropericardium syndrome (HHS) in broilers, which has caused great economic losses to the poultry industry; however, the specific host responses to FAdV-4 remain unknown. In this study, we identified 141 high-confidence protein-protein interactions (PPIs) between the main viral proteins (Hexon, Fiber 1, Fiber 2, and Penton bases) and host proteins via a liquid chromatography-tandem mass spectrometry (LC-MS/MS) assay. We found that heat shock protein 70 (Hsp70), the protein with the highest score, and its cofactor DnaJ heat shock protein 40 family member C7 (DnaJC7) could negatively regulate the replication of FAdV-4. Furthermore, the nucleotide binding domain (NBD) of Hsp70 and the J domain of DnaJC7 were necessary for inhibiting FAdV-4 replication. We verified that DnaJC7 as a bridge could bind to Hsp70 and Hexon, assisting the indirect interaction between Hsp70 and Hexon. In addition, we found that FAdV-4 infection strongly induced the expression of autophagy proteins and cellular Hsp70 in a dose-dependent manner. Blockage of Hexon by Hsp70 overexpression was significantly reduced when the autophagy pathway was blocked by the specific inhibitor chloroquine (CQ). Our results showed that Hsp70 was co-opted by DnaJC7 to interact with viral Hexon and inhibited Hexon through the autophagy pathway, leading to a considerable restriction of FAdV-4 replication. IMPORTANCE FAdV-4, as the main cause of HHS, has quickly spread all over the world in recent years, seriously threatening the poultry industry. The aim of this study was to identify the important host proteins that have the potential to regulate the life cycle of FAdV-4. We found that Hsp70 and DnaJC7 played crucial roles in regulating the amount of viral Hexon and extracellular viral titers. Moreover, we demonstrated that Hsp70 interacted with viral Hexon with the assistance of DnaJC7, followed by suppressing Hexon protein through the autophagy pathway. These results provide new insight into the role of the molecular chaperone complex Hsp70-DnaJC7 in FAdV-4 infection and suggest a novel strategy for anti-FAdV-4 drug development by targeting the specific interactions among Hsp70, DnaJC7 and Hexon.

The First Occurrence of Hepatitis-Hydropericardium Syndrome in Iran and Effective Applied Control Measures in the Affected Commercial Broiler Flock.

Fowl adenoviruses cause three economically important diseases in broiler chicken flocks: hepatitis-hydropericardium syndrome (HHS), inclusion body hepatitis (IBH), and adenoviral gizzard erosion. IBH has not been considered a serious threat in northeast Iran since the last decade, because no major effect on flock performance has been noticed along with a low mortality rate. During this period, all the sporadic IBH outbreaks have also been investigated for HHS without finding any confirmed case. In March 2021, a 15-day-old commercial broiler flock in northeast Iran showed a 50% mortality rate, and birds underwent postmortem examination, histopathology, molecular testing, and phylogenetic analysis for possible disease agents. Typical gross lesions of HHS were observed postmortem that included hydropericardium with an unusual accumulation of jelly-like and straw-colored fluid in the pericardial sac (without right ventricular failure); petechial or ecchymotic hemorrhages on the myocardium, myocardial valves, and endocardium; and discolored and mottled liver along with small white foci and petechial or ecchymotic hemorrhages. Histopathologic analysis showed necrosis of hepatocytes and basophilic inclusion bodies in the livers. The molecular tests performed for detection of fowl adenovirus (FAdV), H5 avian influenza virus, Newcastle disease virus, avian infectious bronchitis virus (IBV), H9N2, chicken infectious anemia virus (CIAV), infectious bursal disease (IBD) virus, Marek's disease virus, Ornithobacterium rhinotracheale, Mycoplasma gallisepticum, and Mycoplasma synoviae turned out positive for FAdV, CIAV, IBD vaccine virus, and IBV serotypes 793B and variant I. The phylogenetic tree based on the hexon gene loop 1 demonstrated a FAdV serotype 4 (FAdV-4) that was identical to Pakistani isolate PARC-1/98. Because it was the first detection of a FAdV-4 in Iran, the stamping out program was applied immediately on the basis of HHS gross lesions and positive PCR reaction on pericardial jelly-like fluid. It seems that this eradication strategy was successful because no outbreaks were noticed for 2 mo after the initial outbreak. It was concluded that the use of gross pathologic baselines, quick diagnosis of disease, and close collaboration between governmental and private sectors were the critical factors that helped locally control the first occurrence of HHS in Iran.

Oral immunization of recombinant Saccharomyces cerevisiae expressing fiber-2 of fowl adenovirus serotype 4 induces protective immunity against homologous infection

Hydropericardium-hepatitis syndrome (HHS) caused by fowl adenovirus (FAdV) serotype 4 strains is a highly contagious disease that causes significant economic loss to the global poultry industry. However, subunit vaccine against FAdV-4 infection is not yet commercially available to date. This study aims to explore the potential for oral immunization of recombinant Saccharomyces cerevisiae expressing Fiber-2 of FAdV-4 as a subunit vaccine. Here, we constructed recombinant S. cerevisiae (ST1814G/Fiber-2) expressing recombinant Fiber-2 (rFiber-2), which was displayed on the cell surface. To evaluate the immune response and protective effect of live recombinant S. cerevisiae, chickens were orally immunized with the constructed live ST1814G/Fiber-2, three times at 5-day intervals, and then challenged with FAdV-4. The results showed that oral administration of live ST1814G/Fiber-2 could stimulate the production of humoral immunity, enhance the body's antiviral activity and immune regulation ability, improve the composition of gut microbiota, provide protection against FAdV-4 challenge, reduce viral load in the liver, and alleviate the pathological damage of heart, liver, and spleen for chicken. In addition, we found the synergistic effect in combining the ST1814G/Fiber-2 yeast and inactivated vaccine to trigger stronger humoral immunity and mucosal immunity. Our results suggest that oral live ST1814G/Fiber-2 is a potentially safer auxiliary preparation strategy in controlling FAdV-4 infection.