Spatial distribution and stability of the eight microbial species of the altered schaedler flora in the mouse gastrointestinal tract

Abstract

The overall complexity of the microbial communities in the gastrointestinal (GI) tracts of mammals has hindered observations of dynamics and interactions of individual bacterial populations. However, such information is crucial for understanding the diverse disease-causing and protective roles that gut microbiota play in their hosts. Here, we determine the spatial distribution, interanimal variation, and persistence of bacteria in the most complex defined-flora (gnotobiotic) model system to date, viz., mice colonized with the eight strains of the altered Schaedler flora (ASF). Quantitative PCR protocols based on the 16S rRNA sequence of each ASF strain were developed and optimized to specifically detect as few as 10 copies of each target. Total numbers of the ASF strains were determined in the different regions of the GI tracts of three C.B-17 SCID mice. Individual strain abundance was dependent on oxygen sensitivity, with microaerotolerant Lactobacillus murinus ASF361 present at 10(5) to 10(7) cells/g of tissue in the upper GI tract and obligate anaerobic ASF strains being predominant in the cecal and colonic flora at 10(8) to 10(10) cells/g of tissue. The variation between the three mice was small for most ASF strains, except for Clostridium sp. strain ASF502 and Bacteroides sp. strain ASF519 in the cecum. A comparison of the relative distribution of the ASF strains in feces and the colon indicated large differences, suggesting that fecal bacterial levels may provide a poor approximation of colonic bacterial levels. All ASF strains were detected by PCR in the feces of C57BL/6 restricted flora mice, which had been maintained in an isolator without sterile food, water, or bedding for several generations, providing evidence for the stability of these strains in the face of potential competition by bacteria introduced into the gut.

FIG. 1.

Distribution of ASF strains in different sections of the GI tracts of three defined-flora C.B-17 mice. The number of total bacterial cells of ASF strains (A), ASF356 (B), ASF457 (C), ASF492 (D), ASF500 (E), ASF361 (F), ASF502 (G), and ASF519 (H) in mouse 1 (solid circles), mouse 2 (open squares), and mouse 3 (open diamonds) are shown. The sections are taken from the esophagus (Esop.) (section E1), stomach (sections S1 and S2), small intestine (sections I1 to I6), ileocecal junction and apical cecum (sections C1 and C2, respectively), and colon (sections L1 to L3).

FIG. 1.

Distribution of ASF strains in different sections of the GI tracts of three defined-flora C.B-17 mice. The number of total bacterial cells of ASF strains (A), ASF356 (B), ASF457 (C), ASF492 (D), ASF500 (E), ASF361 (F), ASF502 (G), and ASF519 (H) in mouse 1 (solid circles), mouse 2 (open squares), and mouse 3 (open diamonds) are shown. The sections are taken from the esophagus (Esop.) (section E1), stomach (sections S1 and S2), small intestine (sections I1 to I6), ileocecal junction and apical cecum (sections C1 and C2, respectively), and colon (sections L1 to L3).

FIG. 2.

Fecal levels of the ASF strains in three defined-flora C.B-17 mice were determined by QPCR. The numbers of ASF bacteria/g (wet weight) in mouse 1 (white columns), mouse 2 (shaded columns), and mouse 3 (black columns) are shown.

FIG. 3.

Specific PCR products obtained by amplification of fecal DNA extracted from C57BL/6 RF mice using the ASF-specific primers. These mice, which originally harbored the defined flora, were bred and maintained under nonsterile conditions for 3 years. Lanes 1 and 10, 100-bp ladder; lanes 2 to 9, RF fecal pellet DNA amplified with primers specific for ASF356 (417 bp), ASF360 (131 bp), ASF361 (182 bp), ASF457 (95 bp), ASF492 (167 bp), ASF500 (285 bp), ASF502 (427 bp), and ASF519 (429 bp) (expected sizes shown in parentheses), respectively.