Bacteriology & the Archaea Lab #2 Bacteriology & the Archaea

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

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

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-)

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

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

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

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

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)

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

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

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

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

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 http://www.youtube.com/watch?v=OQ6C-gj_UHM

Neisseria gonnorhea – coccus, gram -ve

Neisseria gonnorhea

Mycobacterium tuberculosis – bacillus, gram –ve

Mycobacterium tuberculosis

Mycobacterium tuberculosis

Treponema pallidum – gram -ve

Treponema pallidum

gram +ve and -ve

gram +ve