Deletion of <i>hmsP</i> Enables Excessive Biofilm Production in the <i>csrA</i>-deficient Mutant.

<p>Relative biofilm production of CO92, CO92Δ<i>csrA</i>, CO92Δ<i>hmsP</i>, and CO92Δ<i>csrA</i>Δ<i>hmsP</i> following 24 hours post-inoculation in HIB medium. Error bars reflect standard deviation from the mean of 2 independent experiments, each consisting of 6 technical replicates. * P-value < 0.005 as determined by Tukey’s HSD post-hoc analysis.</p

<i>Y</i>. <i>pestis crp</i>-deficient Mutant 26°C Growth Kinetics.

<p>(A) Growth curves of CO92, CO92Δ<i>crp</i>, and plasmid complemented CO92Δ<i>crp</i>: pBluescript/<i>crp</i> when grown in BCS medium supplemented with 0.2% glucose or 0.2% K-gluconate. Growth of the CO92 <i>crp</i>-deficient mutant when cultured in medium supplemented with 0.2% K-gluconate was significantly impaired relative to all other bacterial strain and media conditions (Tukey’s HSD P < 0.05). (B) Growth curves of KIM6+ and KIM6+Δ<i>crp</i> when cultured in BCS medium supplemented with either 0.2% glucose, 0.2% glycerol, or a combination of both 0.2% glucose and 0.2% glycerol. Growth of the KIM6+ <i>crp</i>-deficient mutant when cultured in medium solely supplemented with 0.2% glycerol was significantly impaired relative to all other bacterial strain and media conditions (Tukey’s HSD P < 0.05). (C) Growth kinetics of the CO92 and KIM6+ <i>crp</i>-deficient mutants and respective isogenic controls when grown in HIB medium at 26°C.</p

Bacterial Strains and Plasmids.

<p>* All <i>Y</i>. <i>pestis</i> strains contained the pMT1 and pPCP plasmids.</p><p>Bacterial Strains and Plasmids.</p

<i>Y</i>. <i>pestis csrA</i>-deficient Mutant 26°C Growth Kinetics.

<p>(A) Growth of CO92, CO92Δ<i>csrA</i>, and the chromosomal restoration mutant CO92Δ<i>csrA csrA</i>’ in BCS medium supplemented with either 0.2% glucose or 0.2% K-gluconate during incubation at 26°C. (B) Growth kinetics of the CO92 and KIM6+ <i>csrA</i>-deficient mutants and respective controls when cultured in HIB medium at 26°C. No significant alteration in growth kinetics as determined by repeated measures ANOVA was calculated among the <i>csrA</i>-deficient mutants and the respective controls, regardless of media type or available carbon source.</p

<i>Y</i>. <i>pestis</i> Biofilm Production is not Influenced by Exogenous cAMP.

<p>(A) Relative biofilm production of KIM6+ and KIM6+Δ<i>crp</i> when grown in BCS medium supplemented with 0.2% glucose containing either 3 mM cAMP prepared in K-phosphate buffer or an equal volume of plain K-phosphate buffer. (B) Relative biofilm production of CO92 and CO92Δ<i>crp</i> when grown in BCS medium supplemented with 0.2% glucose containing either 3 mM cAMP prepared in K-phosphate buffer or an equal volume of plain K-phosphate buffer. Identical source inoculum per strain was utilized for each media type. Error bars reflect standard deviation from the mean derived from two independent experiments, each consisting of 6 technical replicates. No statistically significant change in crystal violet absorption as determined by one-way ANOVA was calculated for both the <i>crp</i>-deficient mutants and the respective parental controls when cultured in media containing 3mM cAMP or an equal volume of K-phosphate buffer.</p

Comparison of different routes of infection of mice with the same isogenic strains, C-267 derivatives.

<p>*, <i>P</i><0.05; **, <i>P</i><0.01. Mann-Whitney test. 10 sc/10 id, 10 cfu via subcutaneous/intradermal route. The actual injection doses were equal to 5 (C-267), 6 (C-267pKP3455), 7 (C-267pKPEV) cfu per mouse.</p

Kinetics of survival in subcutaneously inoculated mice.

<p>(A and B) The dose-dependent survival assays and (C, D) mean time to death post infection of mice (<i>n</i> = 8 in a group) inoculated with three different doses of the strains (A, C) C-267 and (B, D) 231 isogenic derivatives (Pla (-)) deficient in Pla or producing its I259 (pKP3455) or T259 (pKPEV) isoform. *, <i>P</i><0.05, **, <i>P</i><0.01, ***, <i>P</i><0.001. The planned injection dose of 100 cfu was actually equal to 98 (C-267), 115 (C-267pKP3455), 123 (C-267pKPEV), 103 (231pPst^-), 95 (231pPst^-pKP3455) or 75 (231pPst^-pKPEV) cfu.</p