Novel methods of disease surveillance in wildlife

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Novel methods of disease surveillance in wildlife

Hansen, Cristina M.

Date: 2015-05

Type: Thesisphd

Table of Contents: 1. Tularemia in Alaska, 1938-2010 -- 1.1. Introduction -- 1.2. Tularemia in wildlife in Alaska -- 1.3. History of human tularemia in Alaska -- 1.3. History of human tularemia in Alaska -- 1.4. Epidemiology of reported cases in Alaska 1946-2010 -- 1.5. Molecular subtyping of recent F. tularensis isolates -- 1.6. Conclusions -- 1.7. Competing interests -- 1.8. Authors contributions -- 1.9. Acknowledgments -- 1.10. References -- 2. Development and implementation of a broad-based polymerase chain reaction surveillance method for bacterial DNA in Alaskan wildlife tissues -- 2.1. Introduction -- 2.2. Materials and methods -- 2.2.1. Sample collection -- 2.2.2. DNA extraction -- 2.2.3. Primers -- 2.2.4. End point PCR reactions -- 2.2.5. qPCR reactions -- 2.2.6. qPCR detection limit and dilution factor determination -- 2.2.7. Sequence analysis -- 2.3. Results -- 2.4. Discussion -- 2.5. Acknowledgments -- 2.6. Sources and manufacturers -- 2.7. Declaration of conflicting interests -- 2.8. Funding -- 2.9. References -- 3. Microbial infection as a source of embryo mortality in wild geese on the Arctic Coastal of Alaska -- 3.1. Introduction -- 3.2. Materials and methods -- 3.2.1. Sample collection -- 3.2.2. Bacterial culture and identification -- 3.2.3. Microscopy -- 3.2.4. DNA extraction -- 3.2.5. PCR and sequencing -- 3.2.6. Embryonated egg infections -- 3.2.7. Histopathology -- 3.3. Results -- 3.3.1 Samples collected -- 3.3.2. Microbiology, PCR, and sequencing -- 3.3.3. Cloacal and nest swabs -- 3.3.4. Morphology -- 3.3.5. Neisseria phylogenetics -- 3.3.6. Embryonated egg infections -- 3.3.7. Sequence accession numbers -- 3.4. Discussion -- 3.5. Acknowledgments -- 3.6. References -- 4. Use of cellulose filter paper to quantify whole blood mercury in two marine mammals: validation study -- 4.1. Introduction -- 4.2. Materials and methods -- 4.2.1. Filter paper and samples -- 4.2.2. Sample preparation -- 4.2.3. Mercury analysis -- 4.2.4. Calculations and statistics -- 4.3. Results -- 4.4. Discussion -- 4.5. Conclusion -- 4.6. Acknowledgments -- 4.7. Literature cited -- General conclusions -- References.

Abstract:

Both infectious and noninfectious disease agents in wildlife impact human health and accurate research, monitoring, and diagnostic methods are necessary. The objectives of the research reported here were to develop and implement novel methods for bacterial and toxicological disease agent surveillance in wildlife. This dissertation begins with a review of tularemia, an important zoonotic disease to the state of Alaska and the Northern hemisphere. In chapter two, I show the development and implementation of broad-based PCR and quantitative PCR (qPCR) surveillance methods for bacterial DNA in tissue samples; 1298 tissue samples were assayed, numerous potential bacterial pathogens were identified and qPCR detection limits were quantified for various tissue matrices. Chapter three describes an investigation into microbial infection as a source of embryo mortality in greater white-fronted geese (Anser albifrons) in Arctic Alaska. This chapter builds upon our previously developed PCR surveillance techniques by which I demonstrated that bacterial infection is responsible for some greater white-fronted goose embryo mortality in Arctic Alaska. Chapter four describes the development and validation of a cellulose filter paper method for quantifying total mercury in whole blood. I determined that filter paper technology is useful for monitoring total mercury in whole blood, with excellent recoveries (82 - 95% of expected values) and R2 values (0.95 - 0.97) when regressed against the concentration of total mercury in whole blood, the technique generally considered as the "gold standard" for mercury detection. These methods will aid in the accurate detection of disease agents in wildlife as demonstrated by our white-fronted goose work.