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. 2002 Apr;70(4):1984-90.
doi: 10.1128/iai.70.4.1984-1990.2002.

Helicobacter Pylori Uses Motility for Initial Colonization and to Attain Robust Infection

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Free PMC article

Helicobacter Pylori Uses Motility for Initial Colonization and to Attain Robust Infection

Karen M Ottemann et al. Infect Immun. .
Free PMC article

Abstract

Helicobacter pylori has been shown to require flagella for infection of the stomach. To analyze whether flagella themselves or motility is needed by these pathogens, we constructed flagellated nonmotile mutants. This was accomplished by using both an insertion mutant and an in-frame deletion of the motB gene. In vitro, these mutants retain flagella (Fla(+)) but are nonmotile (Mot(-)). By using FVB/N mice, we found that these mutants had reduced ability to infect mice in comparison to that of their isogenic wild-type counterparts. When these mutants were coinfected with wild type, we were unable to detect any motB mutant. Finally, by analyzing the 50% infectious dose, we found that motility is needed for initial colonization of the stomach mucosa. These results support a model in which motility is used for the initial colonization of the stomach and also to attain full infection levels.

Figures

FIG. 1.
FIG. 1.
PCR analysis of motB mutants.(A) Schematic representations of wild-type motB (i), motB::cat (ii), and ΔmotB (iii). Arrows represent primers used for PCR. Primer 1 is motB1, primer 2 is motB3, and primer 3 is colicat1 (sequences are in the text). Numbers at the top of panel i indicate nucleotide numbers for the motB open reading frame, while numbers at the bottom represent the genomic sequence numbers from the TIGR sequence (42). The location of the BclI site used for inserting the cat/aphA3-sacB is marked. (B) Agarose gel analysis of PCR amplification products from genomic DNA of SS1, SS1 ΔmotB and SS1 motB::cat, and G27 and G27 motB::cat. The source of genomic DNA is shown at the top of the figure; the primer set (as in panel A) is shown at the bottom. The arrows indicate full-length motB, and the diamonds indicate altered motB (either motB::cat or ΔmotB). Kilobase markers are shown on the left side. Although in some lanes multiple bands can be seen, in all cases the dominant band is the band of interest. Similar results were obtained with G27ΔmotB.
FIG. 2.
FIG. 2.
H. pylori strains lacking motB do not spread in soft agar. Photograph of a brucella broth-5% FBS soft agar plate after 4 days of incubation. The strains were stabbed into the middle of each growth area on day 0. The numerals 1, 2, and 3 indicate wild-type SS1, SS1 ΔmotB, and SS1 motB::cat, respectively. Similar results were obtained with G27.
FIG. 3.
FIG. 3.
H. pylori SS1 lacking motB is flagellated. Electron micrographs of SS1, stained with phosphotungstate. (A) Wild-type SS1; (B) SS1 ΔmotB; (C) SS1 motB::cat. Flagella are visible in all samples; one filament in each panel is marked with an arrowhead.
FIG. 4.
FIG. 4.
motB mutants do not colonize mouse stomachs as well as the wild-type strain. Symbols representing numbers of CFU per gram of stomach in mice infected for 2 weeks with wild-type SS1 (filled circles) or SS1 ΔmotB (filled triangles) are shown. Mice infected by wild-type SS1 were dosed with 9 × 107 CFU, while mice receiving SS1 ΔmotB received 1.9 × 108 CFU. Each point represents one mouse. Three of the ΔmotB mice had no detectable H. pylori; because the limit of detection is estimated to be 250 CFU/gram of stomach, we placed these mice at this level. This detection limit is marked by a dotted line.

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