BIOL 260-General Microbiology. Instructor: Seana Davidson

Transcription

1 BIOL 260-General Microbiology Instructor: Seana Davidson

2 Welcome to BIOL 260: Microbiology! First day: Review of Syllabus Sign-in Introduce the course, review course expectations Begin with first lab Exercise 3: Microscope Lab

3 What is microbiology? The scientific discipline which studies microbes or microorganisms Biology of microbes The interaction of microbes with other microbes, the environment, and humans

4 What are examples of microbes? Algae Fungi Protozoa Bacteria Viruses

5 1. Algae 2. Fungi 3. Protozoa 4. Bacteria 5. Viruses Which microbes are eukaryotes? Which are prokaryotes? Which can perform photosynthesis? Which are classified based on locomotion? Which have cell walls? Which have some type of nucleic acid?

6 Types of Microbes fungi protozoa algae viruses

7 Types of Microbes: Algae Microscopic Diatoms and Volvox Macroscopic

8 Volvox, microscopic aglae that form colonies By Tomachi from

9 Types of Microbes: Protozoa

10 Types of Microbes: Fungi mold yeast mushrooms Can form macroscopic structures

11 Types of Microbes: Bacteria

12 Viruses, Viroids, Prions

13 Microorganisms are associated with Disease Cause of many epidemics in history Bubonic plague ( )-bacteria-still around-yersinia pestis transmitted by fleas Small pox--virus HIV--virus Malaria--protozoa

14 Bacteria are also associated with healthy conditions Normal microbiota (normal flora) The bacteria that are present on our bodies

15 Bacteria are everywhere and provide many services 3.5 billion years of diversification and evolution the environment Soils, Rhizobium associated with plants Food products much of our food is fermented with bacteria in some way Recombinant DNA products Enzymes for many uses Fading of blue jeans, degradation of cellulose, synthesis of nylon

16 History of Microbiology It all started with the microscope! Zacharis Janssen (1600 or so) Dutch spectacle maker. Possibly first telescope and microscope Antoni van Leewenhoek ( )-crafted lense and saw animalcules Robert Hooke (1665) microscopic mushrooms (penicillum mold)

17 Zacharis Janssen s microscope Modeled after the telescope Consisted of two lenses Magnified images 3-10X

18 Leewenhoek s microscope

19 Where do organisms come from? Debunking Spontaneous generation Francesco Redi (1668) flies come from eggs of other flies, not generation from meat years more to prove the same for microbes. John Needham (1745)-boiled broth, microbes still grew Lazzaro Spallanzani (1765)-boiled longer, melted necks of flasks, remained sterile until neck broken Louis Pasteur (1861) showed microbes were in the air, cotton ball experiment, and swan necked flask experiment. What were these? Biogenesis Rudolf Virchow (1858) father of modern medicine-discredit humorism, bringing more science to medicine. Diseases are caused by organisms.

20 Pasteur s flasks Tyndall had trouble repeating this. Why? What was in the lab? What did he discover?

21 John Tyndall questions Pasteur s experiments Could not reproduce Pasteur s results Found that there were heat resistant forms of microbes Same year (1876) Ferdinand Cohn discovers heat resistant forms of bacteria called endospores 1877 Robert Koch demonstrates that anthrax caused by Bacillus anthracis His method of proof became known at Koch s Postulates.

22 New cells need to be placed in categories Aristotle-plant or animal kingdom Kingdom Protista (1866) Electron microscope (1940 s) Kingdom Procaryotae (1968) Carl Woese proposed 3 Domains (1978)

23 Three Domain System

24 Prokaryotes (Bacteria) Bacteria Domain (Eubacteria) Gram negative (thinner cell wall) Gram positive (cell wall) No cell walls Archaea Domain (Archaebacteria)No cell walls Methanogens Halophiles Thermophiles

25 Binomial system of nomenclature Genus and species Escherichia coli, E. coli Both names are in italics or underlined Note that there are higher levels of divisions. Order, Family, then Genus and Species.

26 Relationship of size and resolution

27 Types of microscopes Brightfield Darkfield Phase Contrast Fluorescent Electron

28 Microscopy-Brightfield

29 Some Principles of Light Microscopy Resolution: the ability to distinguish two adjacent objects as separate and distinct Resolution is determined by the wavelength of light used and numerical aperture of lens Limit of resolution for light microscope is about 0.2 µ m Shorter wavelengths give higher resolution

30 Electron Microscopy Transmission Electron Microscopy (TEM) Electromagnets function as lenses System operates in a vacuum High magnification and resolution (0.2 nm) (traveling electrons have very short wavelength) Enables visualization of structures at the molecular level (Figure 2.10a and b) Specimen must be very thin (20 60 nm) and be stained

31 Microscopy Electron microscopes - maximum magnification 100,000X Scanning EM: picture of surfaces Transmission EM: picture of a slice through the cell: interior

32 Oil has same refractive index as glass High magnification lenses use oil to improve image (oil immersion lens)

33 Ways to improve contrast and visualization DIFFERENT METHODS OF LIGHTING Phase contrast / differential interference contrast(dic) Darkfield STAINING Simple: cells all stained the same Differential: stains different types of cells so that they can be distinguished (Gram stain)

34 Microscopy Stained specimen Wet mount

35 2012 Pearson Education, Inc. Examples of: Bright field Phase contrast Dark field

36 Microscopic Techniques: Dyes and Staining Simple stains Basic dyes with positive charge stick to cells Acid dyes provide background stain Differential stains Gram stain - separates bacteria into two categories based on type of cell wall Gram-positive Gram-negative Acid Fast Stain

37 Figure 2.4a Step 1 Result: All cells purple Flood the heat-fixed smear with crystal violet for 1 min Step 2 Result: All cells remain purple Step 3 Result: Gram-positive cells are purple; gram-negative cells are colorless Step 4 G - Result: Gram-positive (G + ) cells are purple; gram-negative (G - ) cells are pink to red G + Add iodine solution for 1 min Decolorize with alcohol briefly about 20 sec Counterstain with safranin for 1 2 min 2012 Pearson Education, Inc.

38 Gram-positive Cell Wall Thick layer of peptidoglycan Teichoic acids

39 Gram-negative Cell Wall Thin layer of peptidoglycan Outer membrane and cytoplasmic membrane lipopolysaccharide (LPS)

40 Differential Stain: Acid Fast Stains particular group of bacteria based on their cell membrane composition (Mycobacterium) Mycolic acid (waxy) makes it difficult to get dye in, so stain with carbol fuschin, heat, then wash with acid alcohol. Dye is retained only in the mycolic acidcontaining cells. (pink)

41 Special stain: Capsule Stain India ink excluded by the gel-like exopolysaccharide capsule

42 Special stain: Endospore Stain Malachite green stains the spores, Safranin is used as counter stain for cells (pink). Gram stain does not stain spores.

43 Special stain: Flagella Stain Stain with basic stain that will stick to the cells and the flagella, making them visible.

44 Morphology of Prokaryotic Cells: Cell Shapes Coccus Rod Vibrio Spirillum Spirochaete

45 Morphology of Prokaryotic Cells: Cell Groupings