Use of confocal microscopy to detect Salmonella typhimurium within host cells associated with Spv-mediated intracellular proliferation

The major limitation in histological examination of orally inoculated mice of Salmonella typhimurium has been the difficulty of attaining high enough levels for immunochemical detection. This problem has been solved by the use of confocal laser scanning microscopy (CLSM) analysis, which allows detection of bacteria in the immunostained sections of mouse spleens at a minimum rate of approximately 1000 colony-forming units (cfu)/spleen. Here, we demonstrate that over 80% of salmonellae of the wild type of S. typhimurium were detected intracellularly within Mac-1 positive cells by the CLSM analysis of immunostained sections from spleens in orally or subcutaneously inoculated mice. Only 40% of salmonellae of the spy-deleted strain were detected inside Mac-1 positive cells. These data suggest that the spy genes play a key role in intracellular proliferation within phagocytes in the mouse spleen. (C) 2000 Academic Press.