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Infect. Immun., 09 1997, 3768-3773, Vol 65, No. 9
Copyright © 1997, American Society for Microbiology

Exposure to low oxygen tension and increased osmolarity enhance the ability of Mycobacterium avium to enter intestinal epithelial (HT-29) cells

LE Bermudez, M Petrofsky and J Goodman
Kuzell Institute for Arthritis and Infectious Diseases, California Pacific Medical Center Research Institute, San Francisco 94115, USA.

Current evidence indicates that Mycobacterium avium infection in patients with AIDS is acquired mostly through the gastrointestinal (GI) tract and that M. avium binds to and invades GI mucosal cells in vitro. Since M. avium is exposed to specific environmental conditions in the GI tract such as changes in pH, low oxygen (O2) tension, increased osmolarity, and low concentration of iron, we investigated the effects of these conditions on the bacterium's ability to enter HT-29 intestinal cells. M. avium 101 (serovar 1) was cultured in 7H9 broth and then exposed to pH 4.5 to 8.0, low O2 tension, 0.1 to 0.3 M dextrose, and absence of iron for 2 h. After washing, bacteria (10(7)/ml) were used in the invasion assay. Confluent HT-29 cells were exposed to 10(6) bacteria for 1 h and then treated with amikacin (200 microg/ml) for 2 h to selectively kill extracellular but not intracellular M. avium. The supernatant was then removed, the monolayer was lysed, and the lysate was plated onto 7H10 agar plates. While exposure to acidic and basic pHs, as well as iron-free medium, had no significant effect on M. avium invasion of intestinal epithelial cells, low O2 tension and increased osmolarity enhanced invasion 11- and 9- fold, respectively, compared with the control. Exposure of M. avium to the combination of low O2 concentration and hyperosmolarity resulted in an approximate 10- to 15-fold increase in penetration of HT-29 cells. Hyperosmolarity and low O2 tension induced the invasive M. avium phenotype and can be useful for the identification of genes associated with M. avium invasion of intestinal mucosa.

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