Document Type

Thesis - University Access Only

Award Date

1995

Degree Name

Master of Science (MS)

Department / School

Biology and Microbiology

First Advisor

Michael B. Hildreth

Abstract

Echinococcus multilocularis is the causative agent for Alveolar Hydatid Disease (ARD), a zoonotic cestode that can be lethal in humans if not promptly diagnosed or if left untreated. Infected dogs and cats can serve as a source of human exposure to E. multilocularis eggs. The ability to safely diagnose E. multilocularis infections in dogs and cats is an extremely important component in controlling the spread of this parasite to human population. This thesis investigated two problems associated with the use of molecular techniques for diagnosing E. multilocularis infections in pet dogs and cats. The first investigation dealt with developing a method to kill (or inactivate) E. multilocularis eggs by utilizing ultracold temperatures. Results from this investigation demonstrated the lethal effect of -70°C on E. multilocularis eggs frozen either within the intestinal contents of its host or in commonly used cryoprotectants. Plunging the E. multilocularis eggs into liquid nitrogen for even 15 minutes also prevented the eggs from developing into hydatid cysts when inoculated into rodents. The second investigation dealt with developing a method for disrupting the embryophore from E. multilocularis eggs and efficiently isolating genomic DNA from the oncosphere. Several physical and chemical methods were explored in an attempt to isolate the maximum quantity of DNA present in the eggs following standard phenol-chloroform extraction and ethanol precipitation of the DNA. A combination of exposure of the eggs to microwaves for two minutes, sonication for 10 minutes, and to proteinase K digestion for one hour yielded 50% of the theoretical quantity of DNA present in 1,000 eggs. The polymerase chain reaction method for identifying E. multilocularis DNA was found to be more sensitive, specific and reliable than the dot blot hybridization method.

Library of Congress Subject Headings

Echinococcus
Dogs -- Parasites
Cats -- Parasites
Dogs
Cats
Animals as carriers of disease

Format

application/pdf

Publisher

South Dakota State University

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Rights Statement

In Copyright