Immune response to lipopolysaccharide in the american

Immune Response to Lipopolysaccharide in the American Alligator (Alligator Mississippiensis)
Mandi P. Sims

Department of Chemistry, McNeese State University, Lake Charles, LA, 70609 June 13,2005

Abstract:
The immune response of the American alligator to LPS from different bacterial species was studied. Alligators were injected with different doses of a mixture of LPS from Escherichiacoli, Klebsiella pneumaniae and Pseudomonas aeruginosa. The doses used were lOmg/kg body weight, 1.0mg/kg body weight, O.lmg/kg body weight and O.Olmg/kgbody weight. From this a dose response was seen. This was determined by determining the change in the number of leukocytes over eleven days. Also a differential leukocyte count was done. This involved looking at the change in the number ofheterophils, lymphocytes, basophiles, monocytes and eosinophils over time. In the dose response, a sharp increase in the number of leukocytes on day one has been determined to be due to a sharp increase in the number of heterophils. After analyzing the immune response to the mixture of LPS from all three bacterial species, a kinetic study was performed looking at the difference in the response between thespecies. Three alligators were injected with a dose of O.5mg/kgbody weight of LPS from Escherichia coli, three with the same dose ofLPS from Klebsiella pneumaniae and three more with the LPS from Pseudomonas aeruginosa. In this study, the dose was held constant, due to the fact that dose response was not being studied. In this experiment, the response to the LPS from the different bacterial specieswas the same. The data obtained showed no difference in the responses. Like the dose response study, the number of leukocytes increases at day one and decreased over time, due to the increase in the number of heterophils at day one and declining over time.

Introduction:
The immune system of the American alligator is one that is very fascinating. These animals are exposed to an environmentthat has the potential to be very infectious. Despite the microbial laden environment in which they live, alligators heal rapidly and do so with little or no sign of infection. This led to the interest in studying their immune system. Previously in this Biochemistry laboratory it has been found that alligator serum possesses potent antibacterial a ctivity.l T his activity w as determined by assessingthe activity alligator serum had on different species of cultured infectious bacteria and fungi. It has also been found the serum from the American alligator possesses antiviral activity as well.2 The findings in these experiments led to the curiosity of how the immune system responds to infectious bacteria when injected directly into the body. Two studies were performed this summer; one was akinetic experiment as well as a dose response experiment. This was looking at the immune response to different doses of a solution of a mixture of lipopolysaccharide, LPS, taken from three different bacterial species, over the course of eleven days. The other study was just a kinetic experiment, which was looking at the immune response to LPS from individual bacterial species over the course ofseven days. The dose ofLPS injected was the same for each species.

Experimental:
In the first experiment the LPS mixture that contained equal parts of LPS from three different bacterial species, Escherichia coli, Klebsiella pneumaniae and Pseudomonas aeruginosa. A total of twelve alligators were used in this experiment. Three different doses of the LPS mixture were used. Three alligators wereinjected with 10mg/kg body weight of the LPS mixture, three were injected with Img/kg body weight of the LPS solution, three with O.lmg/kg body weight and three with O.Olmg/kg body weight. The bleeding of these alligators was done at day 0,1,2,3,4,5,7,9 and 11. This was done using a 20-gauge I-inch needle equipped with a 3mL syringe. The bleeding method used was the cardiac bleed. Going directly...