Table 1 describes various Bordetella strains and plasmids used in the study, including wild type strains and mutant derivatives lacking genes like bvgS, fla, and frlAB. Figures 1-5 show results of experiments examining the role of these genes in biofilm formation, flagella expression, and other phenotypes over time or under different conditions like shaking or static growth.

1. Table 1 Strain or Plasmid Description Reference Bordetella RB50 Wild type New Zealand White Rabbit isolate RB54 ΔbvgS derivative of RB50, Bvg - Rev1 fha driven frlAB synthesis, RB50 frl r RBA5 Δfla derivative of Rev1 RB54Δfla Δfla derivative of RB54 RBA2 ΔfrlAB derivative of RB50 RB50T GFP expressing derivative of RB50 This Study Rev1T GFP expressing derivative of Rev1 This Study RBA2T GFP expressing derivative of RBA2 This Study RBA5T GFP expressing derivative of RBA5 This Study plasmids pTac-GFP Tac driven GFP, CM r derivative of pBBR1-GFP pKK233-2 hybrid trp/lac (tac) promotor pBBR1-GFP promotorless GFP, CAT vector

2. Fig.1 BvgA frlAB fliA ( σ F,28 ) flaA Other frl regulated loci BvgR

3. 0 0.1 0.2 0.3 0.4 0.5 1 8 32 24 48 72 Time (h) OD 540 Fig.2A WT Bvg - Rev1 Rev1 Δ fla

4. 0 0.1 0.2 0.3 0.4 1 2 3 6 24 Time (h) OD 540 Fig.2B WT Rev1 WT Δ frlAB WT Δ fla Rev1 Δ fla Bvg - Bvg - Δ fla

5. 0 10 20 30 40 50 60 70 80 90 100 Percent biofilm cells Fig.2C WT Bvg - Rev1

6. WT Bvg - Rev1 Δ fla Rev1 Fig.3 2h 72h

7. 6h 48h 72h WT Rev1 WT Δ frlAB Fig.4

8. Real Time 2hrs 8hrs 48hrs Expression of flaA relative to RecA Attached Attached Planktonic Planktonic WT Rev1 0 5 10 15 20 25 30 35 Fig.5

9. Real Time expression of Flagella 0 5 10 15 20 25 30 35 40 45 50 Fla Standardized to RpoD Attached Attached Plank. Plank. WT Rev1 Shaking Shaking Tubing Tubing 2hrs 8hrs 48hrs 15hrs