1. Bacillus

2. Bacillus sp. • GP rod • Spore former • Examples • B. anthracis • B. cereus • B. subtilis • B. megatarium • Exoenzymes • Lipase • Amylase • Casease • Gelatinase

3. Spores

4. Agar Characteristics

5. Starch Hydrolysis • Enzymes • Amylase • Maltase • Substrate • Starch • Reaction • Clear zone

6. Protein Hydrolysis • Enzymes • Proteases • Caseases • Substance • Casein • Reaction • Clear zone

7. Lipid Hydrolysis • Enzymes • Lipase • Substrate • Tributyrin • Reaction • Clear zone

8. Bacillus Tri-Plate

9. Collagen Hydrolysis • Enzyme • Gelatinase • Use loop (vs. needle) • Inoculate several times • Stab deep • Substrate • Gelatin • Reaction • Clear Zone on Agar • Liquefy Gelatin in tube

10. MSA • Selective • Differential • Substrate • Mannitol • Salt (7.5%) • pH = phenol red • Reaction • Growth • Fermentation

11. Nitrate Reduction • Inoculate with loop • Incubate • Additional solutions • Sulfanilic acid (A) • Naphthylamine (B) • (+) = Red • Nitrate to Nitrite • No color, add Zinc • (+) = not red • (-) = red

12. Citrate • Enzyme • Citrase • Substrate • Citrate • pH • Bromthymol blue • Reaction • Keto acids • Oxaloacetic • Pyruvate • Acetic acid • CO2 • CO2 + NaH2CO3 • Prussian blue = +

13. Bile Esculin Agar (BEA) • Selective • Bile Salts • Differential • Esculin (CH2O) • Fe-NH3-citrate • Reaction • Growth in bile • Hydrolyze esculin • Esculetin-Iron Rxn • Black precipitate

14. Results • Exoenzymes • Starch (amylase, maltase) • Lipid (lipase) • Proteins (Casease, protease) • Collagen (Gelatinase) • Reaction • (+) = clearing • (-) = no clearing

15. Starch Hydrolysis

16. Lipid Hydrolysis

17. Protein Hydrolysis

18. Gelatin (Collagen) Hydrolysis

19. BEA and Citrate Results + + Bile Esculin Citrate

21. Hydrolysis Results Summary casein lipid Gelatin Starch