Orientations of the Bacteroides fragilis capsular polysaccharide biosynthesis locus promoters during symbiosis and infection

Abstract

Orientations of the seven invertible polysaccharide biosynthesis locus promoters of Bacteroides fragilis were determined from bacteria grown in vitro, from feces of monoassociated and complex colonized mice, and from B. fragilis-induced murine abscesses. Bacteria grown in vivo have greater variability in orientation of polysaccharide locus promoters than culture-grown organisms.

FIG. 1.

PS locus promoter orientations from B. fragilis grown in vitro. (A) Schematic diagram of the PCR digestion method used to determine psa promoter orientation. All PCR programs had an initial incubation of 2 min at 94°C, followed by 30 cycles of 94°C for 30 s, 59°C for 30 s, and 72°C for 90 s and a final 5-min incubation at 72°C. The DNA fragments were separated by electrophoresis on 2% agarose gels and visualized by ethidium bromide staining. (B) Ethidium bromide-stained agarose gels demonstrating the fragments resulting from PCR digestion analysis of bacteria from three independent broth cultures. Each lane represents the data from one independent broth culture.

FIG. 2.

B. fragilis PS locus promoter orientations from bacteria grown in broth cultures (triangles), from fecal samples taken from monocolonized mice at 9 weeks of age (wk 9 mono, diamonds), from fecal samples taken 2 weeks later from the same mice that remained in the gnotobiotic isolator (wk 11 mono, circles) or that were removed from the isolator at 9 weeks and acquired a complex flora (wk 11 complex, squares), and from murine intraperitoneal abscesses (X's). Each data point represents one independent broth culture, mouse, or abscess. Pertaining to the fecal samples, open symbols indicate that the mice were from litter A and filled symbols indicate that the mice were from litter B. Fecal samples from the mice of litter B that remained in the isolator for the duration of the experiment were not analyzed at the first time point. The horizontal bar represents the median.