Assessment of the safety and efficacy of low pathogenic avian influenza (H9N2) virus in inactivated oil emulsion vaccine in laying hens.

In Korea, several outbreaks of low pathogenic AI (H9N2) viral infections leading to decreased egg production and increased mortality have been reported on commercial farms since 1996, resulting in severe economic losses. To control the H9N2 LPAI endemic, the Korea Veterinary Authority has permitted the use of the inactivated H9N2 LPAI vaccine since 2007. In this study, we developed a killed vaccine using a low pathogenic H9N2 AI virus (A/chicken/Korea/ADL0401) and conducted safety and efficacy tests in commercial layer farms while focusing on analysis of factors that cause losses to farms, including egg production rate, egg abnormality, and feed efficiency. The egg production rate of the control group declined dramatically 5 days after the challenge. There were no changes in feed consumption of all three groups before the challenge, but rates of the control declined afterward. Clinical signs in the vaccinated groups were similar, and a slight decline in feed consumption was observed after challenge; however, this returned to normal more rapidly than the control group and commercial layers. Overall, the results of this study indicate that the safety and efficacy of the vaccine are adequate to provide protection against the AI field infection (H9N2) epidemic in Korea.

The current epidemiological status of infectious coryza and efficacy of PoulShot Coryza in specific pathogen-free chickens.

Infectious coryza (IC) is an infectious disease caused by Avibacterium (Av.) paragallinarum. IC is known to cause economic losses in the poultry industry via decreased egg production in layers. Between 2012 and 2013, Av. paragallinarum was isolated from seven chicken farms by Chungbuk National University. We identified Av. paragallinarum, the causative pathogen of IC by polymerase chain reaction (PCR) and serovar serotype A, by multiplex PCR. Antibiotic sensitivity tests indicated that a few field-isolated strains showed susceptibility to erythromycin, gentamicin, lincomycin, neomycin, oxytetracycline, spectinomycin, and tylosin. A serological survey was conducted to evaluate the number of flocks that were positive for Av. paragallinarum by utilizing a HI test to determine the existence of serovar A. Serological surveys revealed high positivity rates of 86.4% in 2009, 78.9% in 2010, 70.0% in 2011, and 69.6% in 2012. We also challenged specific pathogen-free chickens with isolated domestic strains, ADL121286 and ADL121500, according to the measured efficacy of the commercial IC vaccine, PoulShot Coryza. We confirmed the effectiveness of the vaccine based on relief of clinical signs and a decreased re-isolation rate of ADL121500 strain. Our results indicate IC is currently prevalent in Korea, and that the commercial vaccine is effective at protecting against field strains.

Pathological Evaluation of Natural Cases of a Highly Pathogenic Avian Influenza Virus, Subtype H5N8, in Broiler Breeders and Commercial Layers in South Korea.

Outbreaks of highly pathogenic avian influenza (HPAI) virus, subtype H5N8, were observed in two different flocks of local broiler breeder farms and a commercial layer farm in South Korea. Clinically, the cases were characterized by a gradual increase in mortality, slow transmission, and unrecognizable clinical signs of HPAI. Gross observations in both cases included hemorrhagic or necrotic lesions in internal organs, such as serosal and mucosal membranes, spleen, and pancreas. Both cases exhibited similar histopathologic lesions, including multifocal malacia in the brain and multifocal or diffuse necrosis in the spleen and pancreas. Immunohistochemical results indicated that neurons and glial cells in the brain, myocytes in the heart, acinar cells in the pancreas, and mononuclear phagocytic cells in several visceral organs were immunopositive for avian influenza viral antigen. To experimentally reproduce the low pathogenicity and the mortality observed in these two cases, 18 specific-pathogen-free chickens and 18 commercial layers were divided into an H5N8 virus-inoculated group and a contact-exposed group. The mortality of the chickens in the inoculation group was 50%-100%, whereas the mean time to death was delayed or death did not occur in the contact-exposed group. The distributions of the viral antigens and histopathologic lesions in the experimental study were similar to those observed in the field cases. These findings suggest that the H5N8 virus induces a different pattern of pathobiology, including slow transmission and low mortality, compared with that of other HPAI viruses. This is the first pathologic description of natural cases of H5N8 in South Korea, and it may be helpful in understanding the pathobiology of novel H5N8 HPAI viruses.

Genetic characterization of three novel chicken parvovirus strains based on analysis of their coding sequences.

Chicken parvovirus (ChPV) is one of the causative agents of viral enteritis. Recently, the genome of the ABU-P1 strain of ChPV was fully sequenced and determined to have a distinct genomic composition compared with that of vertebrate parvoviruses. However, no comparative sequence analysis of coding regions of ChPVs was possible because of the lack of other sequence information. In this study, we obtained the nucleotide sequences of all genomic coding regions of three ChPVs by polymerase chain reaction using 13 primer sets, and deduced the amino acid sequences from the nucleotide sequences. The non-structural protein 1 (NS1) gene of the three ChPVs showed 95.0 to 95.5% nucleotide sequence identity and 96.5 to 98.1% amino acid sequence identity to those of NS1 from the ABU-P1 strain, respectively, and even higher nucleotide and amino acid similarities to one another. The viral proteins (VP) gene was more divergent between the three ChPV Korean strains and ABU-P1, with 88.1 to 88.3% nucleotide identity and 93.0% amino acid identity. Analysis of the putative tertiary structure of the ChPV VP2 protein showed that variable regions with less than 80% nucleotide similarity between the three Korean strains and ABU-P1 occurred in large loops of the VP2 protein believed to be involved in antigenicity, pathogenicity, and tissue tropism in other parvoviruses. Based on our analysis of full-length coding sequences, we discovered greater variation in ChPV strains than reported previously, especially in partial regions of the VP2 protein.

Eggshell apex abnormalities associated with Mycoplasma synoviae infection in layers.

Eggs exhibiting eggshell apex abnormalities (EAA) were evaluated for changes in shell characteristics such as strength, thickness, and ultrastructure. Mycoplasma synoviae (MS) infection was confirmed by serological assay along with isolation of MS from the trachea and oviduct. Changes in eggshell quality were shown to be statistically significant (p < 0.01). We also identified ultrastructural changes in the mammillary knob layer by Scanning Electron Microscopy. While eggs may seem to be structurally sound, ultrastructural evaluation showed that affected eggs do not regain their former quality. In our knowledge, this is the first report describing the occurrence of EAA in Korea.

Evaluation of neuraminidase antigen based competitive enzyme-linked immunosorbent assay in chickens vaccinated with avian influenza inactivated vaccine.

A competitive enzyme-linked immunosorbent assay (c-ELISA) was developed as a serologic diagnostic tool to detect antibodies against NA subtype 3 of avian influenza virus (AIV). The NA antigen used in this c-ELISA was obtained by pronase treatment of allantoic fluid of specific-pathogen-free (SPF) eggs infected with AIV. The NA specific monoclonal antibodies were produced from purified NA. The N3 c-ELISA was carried out on serum samples collected from both SPF chickens and commercial layers to confirm whether the N3 c-ELISA was capable of detecting specific N3 antibodies. The positive cutoff percentage inhibition value was 6.13%. The sensitivity and specificity of the N3 c-ELISA were 83.7% and 95.6%, respectively, which indicated that N3 c-ELISA can detect the antibodies from SPF chickens or commercial chickens vaccinated with H9N3 subtype of AIV.

Outbreaks of hydropericardium syndrome and molecular characterization of Korean fowl adenoviral isolates.

Outbreaks ofhydropericardium syndrome (HPS), caused by fowl adenovirus serotype 4 (FAdV-4), have occurred in Korea and caused severe economic loss due to mortality and weight loss. From these outbreaks, several adenoviruses were isolated and identified in samples from broilers, layers, breeders, and native Korean fowl. In pathologic examinations, hydropericardium and multifocal hepatic necrosis, with an intranuclear inclusion body in hepatocytes, were observed. Specific adenovirus particles were also observed in the nucleus of hepatocytes, by electron microscopic examination. Polymerase chain reaction (PCR) analysis of the hexon gene identified all of the isolates as FAdV, serotype 4 and genotype C. To reproduce FAdV-4 field cases, 8- and 52-week-old specific pathogen free (SPF) chicks were infected intramuscularly with the field isolate CBU070244. The mortality rate of infected chicks ranged from 10%-40%, and specific pathologic lesions, such as swollen livers and hydropericardium, were observed. Further studies to determine the prevalence of infection, and analysis of the economic impact to the poultry industry, are needed in the near future.