Graduate Studies

 

First Advisor

Samodha C. Fernando

Degree Name

Doctor of Philosophy (Ph.D.)

Department

Complex Biosystems

Date of this Version

8-2024

Document Type

Dissertation

Citation

A dissertation presented to the faculty of the Graduate College of the University of Nebraska in partial fulfillment of requirements for the degree of Doctor of Philosophy

Major: Complex Biosystems

Under the supervision of Professor Samodha C. Fernando

Lincoln, Nebraska, August 2024

Comments

Copyright 2024, Alison Clare Neujahr. Used by permission

Abstract

With the ever-increasing demand for animal protein and the continual need for the assurance of proper animal welfare, investigations of preventative strategies for diseases is greatly needed. Specifically in the cattle industry, infectious bovine keratoconjunctivitis (IBK), commonly known as pinkeye, causes a large economic loss to the producer. However, effective mitigation strategies are lacking. Partly, this is due to the lack of studies investigating the ocular microbiome. Therefore, studies investigating the bovine ocular microbiome are greatly needed to better understand IBK disease etiology. To this end, we investigated the changes in the ocular microbiome in animals before, during and after IBK infection using shotgun metagenomic sequencing and compared microbiome changes associated with infection.

Over the last several decades, an increasing emergence of novel bacterial and viral species and variants has been observed. Among the pathogens emerging, viruses have been identified as one of the major concerns and therefore rapid and accurate diagnostic tests are greatly needed. Currently, the APHIS surveillance program routinely tests for five high consequence pathogens within the swine industry, which include Foot and Mouth Disease, Highly Pathogenic Avian Influenza, Classical Swine Fever, and African Swine Fever. The major detection methods for such diseases include real-time PCR assays and pathogen-specific antibodies. However, due to genetic drift or -shift in virus genomes, failure to detect such pathogens is a risk with devastating consequences. Additionally, the emergence of novel pathogens requires non-biased detection methods for discovery with no prior information. Here we developed a novel sequence-based surveillance/diagnostic tool to rapidly and accurately detect viral and bacterial pathogens simultaneously using Oxford Nanopore Technologies MinION™ sequencing platform.

Prophylactic use of antibiotics has been used in the livestock industry to prevent disease. However, this has resulted in an increase of antibiotic resistant bacteria. Thus, there is a need for new methods and technologies to reduce the antibiotic burden in the livestock industry. Within the beef cattle industry, tylosin is used as a prophylactic to reduce the occurrence of liver abscesses. In addition to lost revenue associated with the liver and surrounding muscles, animal productivity and wellbeing are substantially decreased. Investigations have attempted to isolate and utilize a direct fed microbial as an alternative to antibiotics. However, success has been limited. Interestingly, previous studies from the Fernando lab have identified biosynthetic gene clusters (BGCs) from organisms within the rumen to harbor genes that inhibit pathogens which cause disease. Here we isolate microbes from the rumen and screen these microbes for the synthesis of inhibitory agents against pathogens. Such organisms were isolated, the genes of interest were identified through genome sequencing, and genes were introduced to GRAS approved strains of Bacillus pumilus to develop genetically engineered probiotics with increased functions. Such engineered probiotics can be used to increase animal health and performance in the livestock industry.

Advisor: Samodha C. Fernando

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