Eggshell membrane structure and penetration by Salmonella Typhimurium

Eggshell membrane was removed from 10 broiler hatching eggs at approximately monthly intervals through the productive life of a commercial flock. A piece of membrane (2 by 2 cm) was used to cover an opening in an apparatus designed to test penetration by Salmonella Typhimurium. The membrane was placed between two chambers as the only means of liquid transfer. The chamber above the membrane was filled with a suspension of Salmonella Typhimurium cells. Sampling of the bottom chamber provided cultural evidence of the penetration by a marker strain of Salmonella Typhimurium. Samples were drawn at 15 min and at 1, 3, 5, 7, 12, 24, and 48 h. Following the cultural penetration experiment, the same pieces of membrane were removed and stained for microscopic examination. Membrane structure was examined with the use of a confocal laser scanning microscope. Each image consisted of a composite of eight 1-mu m optical slices showing the fibers making up the outer surface of the membrane. These images were transferred to an image analysis software package that allowed the measurement of the interfiber area. No clear correlation could be made between the average size of all interfiber areas or the total interfiber area measured in the outer 8 mu m of the outer membrane and the ability of Salmonella Typhimurium to penetrate the same piece of eggshell membrane.