1. Bacteriology & the Archaea
Lab #2
Bacteriology & the Archaea

2. Bacterial Groups
PROTEOBACTERIA
Subgroup: Alpha Proteobacteria
Rhizobium (arrows)
2.5 µm
Subgroup: Beta Proteobacteria
Nitrosomonas
1 µm
Subgroup: Gamma Proteobacteria
Chromatium
0.5 µm
Subgroup: Delta Proteobacteria
Chrondromyces
crocatus
10 µm
2 µm
Subgroup: Epsilon Proteobacteria
Heliocobacter pylori
5 µm
Bdellovibrio
bacteriophorus
1. Proteobacteria: diverse group of gram negative bacteria
5 well-established groups
alpha
beta
gamma
delta
epsilon
newer groups being discovered today – e.g. zeta

3. Bacterial Groups 1. Proteobacteria:
a. alpha: live in close association with eukaryotes
Rhizobium – lives in nodules within the roots of legumes – convert atmospheric N2 into compounds that the plants can use (nitrogen fixation)
some strains can cause tumors in plants – Agrobacterium – used to genetically modify plants
root nodules

4. Bacterial Groups 1. Proteobacteria: b. beta: nutritionally diverse
Nitrosomonas – soil bacteria that plays a role in N2 recycling by oxidizing ammonium (NH4) into nitrites (NO2-)

5. Bacterial Groups 1. Proteobacteria:
c. delta: slime secreting myxobacteria
when the soil dries out – they form into aggregations called fruiting bodies & release spores into the environment for the establishment of new colonies in better environments
Bdellovibrio bacteria “charge” at other bacteria at speeds equivalent to 240km/hr
drills into its prey using its flagella and digestive enzymes

6. Bacterial Groups 1. Proteobacteria:
d. gamma: includes many well-known strains of gram-negative bacteria
autotrophic & heterotrophic species
include the older classification known as purple-sulfur bacteria (e.g. Thiomargarita namibiensis)
obtain energy by oxidizing H2S  producing sulfur as a waste
many heterotrophic strains are pathenogenic (e.g. Legionella, Salmonella and Vibrio cholerae)
non-pathenogenic strain = E. coli
Vibrio cholerae

7. Bacterial Groups 1. Proteobacteria:
e. epsilon: many are pathenogenic to humans and other animals
includes Campylobacter = blood poisoning
Helicobacter pylori = stomach ulcers
f. zeta: relatively new classification
Helicobacter pylori

8. Bacterial Groups
2. Gram positive bacteria: rival proteobacteria in diversity
2 major subgroups based on cell shape: cocci and bacillus
most decompose organic matter in soil
two strains of Actinomycetes cause leprosy and tuberculosis
Streptomyces used by pharmaceutical companies to produce antibiotics
numerous strains are very pathogenic: Bacillus anthracis, Clostridium botulinum, Staphylococcus and Streptococcus
GRAM-POSITIVE BACTERIA
5 µm
Streptomyces
Mycoplasmas covering
a human fibroblast cell

9. Bacterial Groups 3. Chlamydias 4. Spirochetes
can only survive in animal cells – depend on their hosts for ATP
Chlamydia trachomatis – cause of non-gonococcal urethritis (most common STD)
4. Spirochetes
move through rotation provided by internal flagella-like filaments
Treponema pallidum – causes syphilis
Borrelia burgdorferi – causes Lyme disease
Leptospira – causes leptospirosis (field fever)
5 µm
SPIROCHETES
Leptospira
2.5 µm
CHLAMYDIAS
Chlamydia (arrows)

10. Bacterial Groups 5. Cyanobacteria photoautotrophs
only prokaryotes with plant-like, oxygen-generating photosynthesis
abundant components of fresh water and marine phytoplankton
CYANOBACTERIA
Oscillatoria
50 µm

11. Bacterial classification
colony morphology
bacterial colonies grow from single cells
colony is composed of millions of bacteria
each colony has a characteristic size, shape, consistency, texture and color
common colony shapes:
punctiform = each colony is less than 1mm
round
filamentous – often confused with fungus (which is more “fuzzy”)
irregular

12. Bacterial classification
cell morphology
bacilli (rod)
cocci (spherical)
spirilla or spiral
many bacterial cells adhere to each other and form clusters or chains
under some environments – many different species may associate with each other – creating a community called a biofilm
biofilms are usually found where nutrients are plentiful
soils, water pipes, surface of your teeth

13. Gram staining
both Gram-positive and Gram-negative bacteria take up the same amounts of crystal violet (CV) and iodine (I).
CV-I complex is trapped inside the Gram-positive cell by the washing of the bacteria with 95% ethanol
results in dehydration and limits the loss of CV-I complex = PURPLE STAIN
thin peptidoglycan layer of the gram negative bacteria does not impede extraction of the CV-I complex
plus the outer membrane limits the amount of CV-I complex that can reach the PG layer = CLEAR STAIN

14. Gram Staining
1. Place a slide with a bacterial smear on a staining rack.
2. STAIN the slide with crystal violet for 1-2 min.
3. Pour off the stain and rinse with water thoroughly. 4. Flood slide with Gram's iodine for 1-2 min.
5. Pour off the iodine and rinse with water thoroughly.
6. Decolorize by washing the slide briefly with acetone (2-3 seconds) or 95% ethanol
7. Wash slide thoroughly with water to remove the acetone/ethanol
8. Flood slide with safranin counterstain for 2 min.
9. Wash with water.
10. Blot excess water and dry

15. Neisseria gonnorhea – coccus, gram -ve

16. Neisseria gonnorhea

17. Mycobacterium tuberculosis – bacillus, gram –ve

18. Mycobacterium tuberculosis

19. Mycobacterium tuberculosis

20. Treponema pallidum – gram -ve

21. Treponema pallidum

22. gram +ve and -ve

24. gram +ve