Date of Award

5-2007

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Legacy Department

Biological Sciences

Advisor

Rice, Charles D

Abstract

Little is known about the immunobiology of fish responses to parasitic infections. The aim of this study was to describe a model for the examination of fish-nematode interactions on the immunological front, and to develop new reagents for the study of this model. Gulf killifish, Fundulus grandis, were collected over two seasons from Simmon's Bayou, Jackson County, MS, USA, pre Hurricane Katrina, and examined for endoparasites, particularly the larval nematode Eustrongylides ignotus. A baseline study was preformed to determine the prevalence, intensity, and histologic features of E. ignotus infection in F. grandis, and this study serves as the first description of its kind for this host-parasite model. In addition, two monoclonal antibodies, (mAb) M24-2 and 2C11, were characterized for their use in this project, as well as for general applications in fish immunobiology. Monoclonal antibody M24-2 was previously developed in this lab and found to recognize a ~ 14.5 kD protein in serum from all fish tested to date, and this antibody also recognized denatured hen egg lysozyme (HEL). Using mAb M24-2, cellular lysozyme protein was also found in lymphoid cell lysates, formalin-fixed and permeabilized lymphoid cells adhered to glass cover slips, and in plastic embedded head kidney and spleen tissue. In addition, mAb M24-2 was found to recognize most cells separated off percoll density gradients in a fraction known to contain fish phagocytes, primarily macrophages and neutrophils. Monoclonal antibody 2C11 was developed and determined to recognize a ~17 kD protein found in fish cell lysates, but in not plasma. Upon immune staining of glass adherent cells, it was determined that mAb 2C11 recognizes granule content of a small population of highly granulated cells, most likely eosinophilic granular cells (EGCs). Like mAb M24-2, mAb 2C11 was found to be cross reactive in all species of fish tested; however, it is unable to bind its antigen in plastic embedded tissues.
Upon dissection of F. grandis, the larval nematode E. ignotus was found, as well as the apicomplexan Calyptospora funduli and metacercaria of a digenetic trematode. Histological descriptions of normal and infected head kidneys, spleen and livers from F. grandis, along with infected tissues probed with mAbs M24-2 and 2C11, were preformed. There were no observable differences of cell types surrounding parasites in tissues, nor of immune staining, between infected and un-infected fish. There was no correlation between fish size, sex, parasite load and other parameters such as number of melanocyte macrophage centers in head kidney and spleen tissue, fatty liver and 2C11 positive cells.

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Biology Commons

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