A constitutively mannose-sensitive agglutinating Salmonella enterica subsp. enterica serovar typhimurium strain, carrying a transposon in the fimbrial usher gene stbC, exhibits multidrug resistance and flagellated phenotypes

Abstract

Static broth culture favors Salmonella enterica subsp. enterica serovar Typhimurium to produce type 1 fimbriae, while solid agar inhibits its expression. A transposon inserted in stbC, which would encode an usher for Stb fimbriae of a non-flagellar Salmonella enterica subsp. enterica serovar Typhimurium LB5010 strain, conferred it to agglutinate yeast cells on both cultures. RT-PCR revealed that the expression of the fimbrial subunit gene fimA, and fimZ, a regulatory gene of fimA, were both increased in the stbC mutant when grown on LB agar; fimW, a repressor gene of fimA, exhibited lower expression. Flagella were observed in the stbC mutant and this phenotype was correlated with the motile phenotype. Microarray data and RT-PCR indicated that the expression of three genes, motA, motB, and cheM, was enhanced in the stbC mutant. The stbC mutant was resistant to several antibiotics, consistent with the finding that expression of yhcQ and ramA was enhanced. A complementation test revealed that transforming a recombinant plasmid possessing the stbC restored the mannose-sensitive agglutination phenotype to the stbC mutant much as that in the parental Salmonella enterica subsp. enterica serovar Typhimurium LB5010 strain, indicating the possibility of an interplay of different fimbrial systems in coordinating their expression.

Figure 1

Observation of Salmonella enterica subsp. enterica serovar Typhimurium LB5010 and the stbC mutant strain grown in static broth culture. (a) Salmonella enterica subsp. enterica serovar Typhimurium LB5010 strain grown in static LB broth condition at 37°C for 48 h produced fimbrial appendages (arrow). No flagella structures were observed. (b) Salmonella enterica subsp. enterica serovar Typhimurium stbC mutant strain grown in static LB broth condition at 37°C for 48 h produced fimbrial appendages (arrow) and flagella structures (arrowhead). Bacterial cells were negatively stained with 2% of phosphotungstic acid (20,000x).

Figure 2

Observation of Salmonella enterica subsp. enterica serovar Typhimurium LB5010 and the stbC mutant strain grown on solid agar. (a) Salmonella enterica subsp. enterica serovar Typhimurium LB5010 grown on LB agar at 37°C for 16 h did not produce fimbrial appendages. (b) The Salmonella enterica subsp. enterica serovar Typhimurium stbC mutant grown on LB agar at 37°C for 16 h exhibited flagella structures (arrowhead) but no fimbrial appendages were observed (3,000x). Bacterial cells were negatively stained with 2% of phosphotungstic acid (20,000x).

Figure 3

Salmonella enterica subsp. enterica serovar Typhimurium LB 5010 and the stbC mutant grown on MSRV agar medium. (a) Salmonella enterica subsp. enterica serovar Typhimurium LB5010 did not exhibit any “halo” appearance on the agar surface. (b) a gray-white zone was observed extending from the inoculated drop of the stbC mutant.

Figure 4

Effect of a transposon inserted in stbC on transcription within the fim gene cluster. RT-PCR assays were used to monitor fim gene transcription in Salmonella enterica subsp. enterica serovar Typhimurium LB5010, the stbC mutant, stbC (pStbC), and stbC (vector) cultured on static LB broth and solid LB agar. The intensities of the bands on the gel were determined by densitometry and are expressed relative to the value for Salmonella enterica subsp. enterica serovar Typhimurium LB5010 grown on LB agar.