Anaerobic Culture Methods

1. Anaerobic Culture Methods • Reducing media • Contain chemicals (thioglycollate or oxyrase) that combine O2 • Heated to drive off O2

2. Anaerobic Culture Methods • Anaerobic jar Figure 6.5

3. Anaerobic Culture Methods • Anaerobic chamber Figure 6.6

4. Capnophiles Require High CO2 • Candle jar • CO2-packet Figure 6.7

5. Selective Media • Suppress unwanted microbes and encourage desired microbes. Figure 6.9b–c

6. Differential Media • Make it easy to distinguish colonies of different microbes. Figure 6.9a

7. Enrichment Media • Encourages growth of desired microbe • Assume a soil sample contains a few phenol-degrading bacteria and thousands of other bacteria • Inoculate phenol-containing culture medium with the soil and incubate • Transfer 1 ml to another flask of the phenol medium and incubate • Transfer 1 ml to another flask of the phenol medium and incubate • Only phenol-metabolizing bacteria will be growing

8. A pure culture contains only one species or strain. • A colony is a population of cells arising from a single cell or spore or from a group of attached cells. • A colony is often called a colony-forming unit (CFU).

9. Streak Plate Figure 6.10a–b

10. Preserving Bacteria Cultures • Deep-freezing: –50°to –95°C • Lyophilization (freeze-drying): Frozen (–54° to –72°C) and dehydrated in a vacuum

11. Reproduction in Prokaryotes • Binary fission • Budding • Conidiospores (actinomycetes) • Fragmentation of filaments

12. Binary Fission Figure 6.11

13. Figure 6.12b

14. If 100 cells growing for 5 hours produced 1,720,320 cells:

15. Figure 6.13

16. Figure 6.14

17. Direct Measurements of Microbial Growth • Plate counts: Perform serial dilutions of a sample Figure 6.15, step 1

18. Plate Count • Inoculate Petri plates from serial dilutions Figure 6.16

19. Plate Count • After incubation, count colonies on plates that have 25-250 colonies (CFUs) Figure 6.15

20. Direct Measurements of Microbial Growth • Filtration Figure 6.17

21. Direct Measurements of Microbial Growth • Multiple tube MPN test. • Count positive tubes and compare to statistical MPN table. Figure 6.18b

22. Direct Measurements of Microbial Growth • Direct microscopic count

23. Direct Measurements of Microbial Growth Figure 6.19, steps 1, 3

24. Estimating Bacterial Numbers by Indirect Methods • Turbidity Figure 6.20

25. Direct methods Plate counts Filtration MPN Direct microscopic count Dry weight Indirect methods Turbidity Metabolic activity Dry weight Measuring Microbial Growth