Performing Department
CA Animal Health and Food Safety Laboratory System (CAHFS)
Non Technical Summary
Infectious bronchitis virus (IBV) contributes to respiratory, reproductive and renal disease and in some cases mortality in chickens.Commercial poultry farms have a high prevalence of infectious bronchitis (IB) and experience severe economic losses, which theyattempt to curtail through vaccination programs. However, IBV's high mutation rate allows new variants to evade the host'simmune system and lead to continual emergence of new strains and outbreaks. California harbors significant numbers ofcommercial poultry farms as well as numerous small flocks, often closely situated, leading to possible transmission, exchange, andmaintenance of infectious agents. Backyard or small flock owners are less stringent in biosecurity compared to the commercialfacilities, and do not employ vaccination strategies for IBV. The proposed study is designed to plot the prevalence of IBV inbackyard chickens submitted to the diagnostic laboratories for investigating causes of death and disease in the flocks. Every chickensubmitted for postmortem examinations during three months will be tested for IBV by PCR and immunohistochemistry. Thepathological and histopathological features of IBV will be studied in tissues, and the findings associated with PCR results as well asto the causes of mortality. In PCR positive cases the IBV will be sequenced to genetically characterize the circulating strains and toassociate the backyard strains with the known commercial poultry strains to aid in a better understanding the IBV status inCalifornia and establishing preventive measures.
Animal Health Component
100%
Research Effort Categories
Basic
100%
Applied
(N/A)
Developmental
(N/A)
Goals / Objectives
Infectious Bronchitis Virus (IBV) can be detrimental to the health of chickens and the economy. Information is needed on IBV in backyard chickens to determine the effects of the virus to causes of morbidity or mortality in backyard chickens, and to establish relationships between backyard flocks (BYF) and the commercial industry. The goal of this proposal is surveillance of IBV status in backyard chickens, including the pathological and diagnostic features of IBV infection and the molecular characterization of the IBV isolates from backyard flocks.We will collate and organize data on: the prevalence of IBV detection and contribution to disease in BYFs, the pathological lesions, diagnostic methodologies, and locations. On positive samples, we will perform the molecular characterization of IBV isolates: virus isolation, sequencing and phylogenetic analysis, and compare with known vaccine or field strains. Finally, we will compare the histopathological, immunohistochemical and molecular diagnostic methods for IBV detection.
Project Methods
-Case data: Backyard chickens submitted to the Northern (Davis) and Southern (San Bernardino) laboratories of the California AnimalHealth and Food Safety Laboratory System (CAHFS) for necropsies during a period of three months will be included in the study.Case reports will be analyzed for the locations of the flocks at the county level, and information on age, sex, breed of chicken, as well as the cause(s) of death and secondary and/or underlying pathological findings. The IBV positive chickens will be mapped using a web service mapping tool GeoNames®.-Histology and IBV testing: Two sets of trachea, kidney and cecal tonsil tissues will be collected during the time of necropsy fromeach chicken. One set will be fixed in 10% buffered formalin and cut at 4μm sections for hematoxylin & eosin (HE) staining and IBVimmunohistochemistry (IHC), and another set collected in whirl pack bags and frozen at -80°C for PCR.Histopathology will be performed on HE sections of trachea and kidneys when tissues are available and the histological lesionsscored based on mild (+1), moderate (+2) and severe (+3) lesions. Lesions to be scored in the trachea are a) single-cell necrosis inthe epithelium, and b) leukocytic infiltrations in the epithelium and lamina propria. The criteria in the kidney are a) tubular necrosischaracterized by distention of convoluted tubules, degeneration/coagulative necrosis of the tubules, and sloughed epithelial cellsand heterophils in the lumen, b) lymphoplasmacytic interstitial infiltrations, and c) gout.IHC will be performed on all three tissues using monoclonal antibodies per CAHFS SOPs. Interpretation and scoring of theimmunostaining will be conducted dependent on specific immunostaining of cytoplasm of epithelial cells and of mononuclear cells,ranging between focal (+1), multifocal (+2) and extensive (+3) staining.-Real-time reverse transcriptase polymerase chain reaction (RT-qPCR) targeting the S1 gene hypervariable region will be carried outinitially on the pool of frozen trachea, kidney, and cecal tonsil. When positive, the tissues each will be tested separately by RT-PCR,and the Ct values recorded. Virus isolation will be attempted on RT-PCR positive tissues. The homogenate from tissue swabs will beinoculated into SPF chicken eggs (Charles River) and incubated for up to 5 days.