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Unit II - Microbiology

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Unit II - Microbiology Sources: Pelczar M.J. and Chan E.C.S., Elements of Microbiology, McGraw-Hill, Inc. 1981 Paustian, T. , http://www.bact.wisc.edu/MicrotextBook ... – PowerPoint PPT presentation

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Title: Unit II - Microbiology


1
Unit II - Microbiology
  • Sources
  • Pelczar M.J. and Chan E.C.S., Elements of
    Microbiology, McGraw-Hill, Inc. 1981
  • Paustian, T. , http//www.bact.wisc.edu/MicrotextB
    ook/University of Wisconsin-Madison 2002
    (Textbook on the web)

2
Student Objectives
  • 1. To become familiar with the ubiquitous nature
    of microorganisms.
  • 2. To learn some basic bacteriological lab
    techniques.

3
  • 3. To understand how bacteria grows.
  • 4. To become familiar with major groups of
    microorganisms.

4
.
  • 5. To know how microorganisms can be controlled.
  • 6. To be familiar with diseases caused by
    microorganisms.

5
Section One
  • History of Microbiology

6
The First Microscope.
  • Invented in late 1500s
  • A simple microscope like a magnifying lens

7
The discovery of microorganisms.
  • Leeuwenhoek Dutch naturalist in late 1600s

8
  • Ground lenses and made microscopes.
  • First to record observations of microscopic
    organisms seen in rainwater.
  • Called the little organisms animalcules.

9
  • Basic kinds of bacteria as observed by
    Leeuwenhoek
  • spirilla spiral-shape

10
  • cocci spherical

11
  • bacilli rod-shaped

12
Spontaneous Generation
  • Belief that organisms arose from non life
  • Disproved by use of beef broth experiments
  • Louis Pasteur the gooseneck flask

13
Pasteur
14
Schwanns flame
Schulze acid base
Schroeders cotton plug
Pasteurs
15
Fermentation
  • Used for many years.
  • Pasteur discovered microbes were responsible

16
Disease
  • 1540s first theories that small organisms
    caused disease.

17
Key Discoveries
  • Pasteur isolated anthrax causing microbes
  • Koch discovered anthrax was caused by bacilli
    bacteria
  • Koch developed culture media

18
Kochs postulates
  • A specific microbe is associated with a given
    disease.
  • That microbe can be isolated and grown.
  • The pure culture can be used to infect a host
    organism.
  • Microbes can be recovered from infected organism

19
Koch
20
Immunization
  • Any process that develops resistance in a host to
    a specific disease.

21
Pasteur discovered immunization accidentally.
  • By using old cultures, injected chickens remained
    healthy.
  • He injected a new culture in those chickens and
    again they remained healthy.

22
Source http//www.tutorvista.com/content/biolog
y/biology-iv/immune-system/vaccination-and-immunis
ation.php
23
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24
  • Source http//www.bbc.co.uk/schools/gcsebitesize
    /history/shp/modern/indrevmethodsrev2.shtml -
    This source says Charles Chamberland (France
    1880) discovered by chance (when he left bacteria
    exposed to air) that injecting chickens with an
    attenuated (weakened) form of chicken cholera
    gave them immunity to the disease (ie he
    discovered the principle of inoculation).
    Pelczar and Chan credit this discovery to Pasteur.

25
Other breakthroughs
  • Jenner and smallpox immunization
  • Vaccinations (from vacca cow)
  • Rabies vaccines.
  • Antisepsis
  • Many more !!

26
Todays Microscope.
  • Bright field dark objects in bright area
  • Compound multiple lenses. Up to 2000 x
  • Electron maximum magnification because of short
    wavelengths. Up to 400,000 x

27
The oil immersion lens.
  • is 100 X
  • Requires oil to prevent light refraction
  • Gives maximum magnification for compound light
    microscope.

28
Quiz History of Microbiology
  • COMING SOON
  • GET READY !!!!

29
Section 2
  • Laboratory Techniques

30
Microscopes
  • Light microscopes max 2000 X
  • Electron microscopes max 400,000 X

31
Bright-field Microscopy
  • Light background and dark objects.

32
The Compound Light Microscope
  • Main components for directing light.
  • Eyepiece, objectives, stage, condenser, iris
    diaphragm, fine and course adjustment knobs,
    base, mirror, etc.

33
Resolving Power
  • Resolving Power wavelength
  • NA objective NA
    condenser
  • NA numerical aperature
  • Wavelength rane is 400 to 700 nm
  • NA for high dry is about 0.85
  • NA for oil-immersion objective 1.2 to 1.4

34
  • Whatever the resolving power, if two objects are
    closer together than the numerical resolving
    power, they appear as one.

35
  • The resolving power of a microscope is the
    ability of the devise to distinguish between two
    objects. A light microscope can only resolve two
    objects that are bigger than 250 nanometers
    because objects smaller than this fall well below
    the size of a wavelength of visible light (500
    650 nanometers). Non lens system can ever resolve
    two dots that are closer together than half the
    wavelength of the light that is used to view
    them. Source http//www.blurtit.com/q571512.h
    tml

36
  • No doubt you have seen a sunrise and a sunset.
    The sky turns red, orange, yellow and even purple
    because the colored wavelengths of light travel
    farther through the air (atmosphere). When the
    sun is low in the sky, this long journey through
    the atmosphere means the colors with shorter
    wavelengths, like blue, have already scattered or
    bounced off in numerous directions.
  • Orange sunsets (yellow and red light waves)
    appear when the air is clean. Sunsets that are
    the most spectacular occur when red wavelengths
    reflect off of overhead clouds.
  • Spectators continue to see light in the sky long
    after it has turned dark on the ground. Why is
    this? Because night doesn't "fall". It actually
    rises from the ground as the sun goes farther
    below the horizon.
  • Civil twilight occurs when the sun is 6 below
    the horizon. This is from the time that the sun
    drops below the horizon until artificial lights
    (street or home) are needed.
  • Astronomical twilight occurs when the sun is 18
    below the horizon. This is when there is no
    sunlight on the western horizon and stars can be
    seen.
  • Twilight is shorter in the tropics because the
    sun's path is more perpendicular to the Earth's
    plane and it takes less time to go from 6 to 18
    below the horizon at this angle.
  • White nights occur in extreme northern latitudes
    where the evening twilight merges with the
    morning twilight.
  • http//sci.odu.edu/sci/Scire/05Edition/sunset.html

37
Preparation of Specimens
  • Wet mount - short term mount high dry
  • Hanging Drop high dry
  • Stained smears best for bacterial form - oil
    immersion

38
Wet mount source http//www.greatscopes.com/a
ct005.htm
39
Hanging Drop Slide Source http//homepage.smc.
edu/wissmann_paul/microbiology/motility.html
40
Negative Staining
  • Stains background and makes bacteria stand out
    more clearly.
  • Nigrosine stain

41
Staining the Bacteria (positive)
  • Stains bacteria and not the background.
  • Main stains are methylene blue, gentian violet
    (crystal violet), and carbol fuchsin.

42
The Staining Process
  • Three basic steps.
  • Smearing
  • Fixing
  • Staining

43
Step One the smear
  • A very clean slide
  • Draw circle on back of slide and label back of
    slide.
  • Sterilize your loop.
  • Place very small drop of sterile water on slide

44
  • Sterilize loop.
  • Obtain sample on loop
  • Remember Bacteria are very small !!
  • Smear bacteria in water

45
Step two Fixing the smear
  • Air dry the smear
  • Pass slide through burner flame up to 3 times
  • Do not flame the slide until all water has
    disappeared !!!

46
Step three - staining
  • Cover the smear with stain for a few seconds.
  • Wash all the stain off the slide.
  • Dry using filter paper

47
Special Staining
  • Gram Stain procedure divides bacteria into two
    groups.
  • Gram positive organisms that are stained
    purple.
  • Gram negative organisms that retain
    counterstain color.

48
  • Acid-fast staining to classify those bacteria
    which retain stain when washed with acids or
    alcohol.

49
Growing Bacteria in the Lab
  • Culture and Media

50
Growth Requirements
  • energy source
  • a source of carbon
  • required nutrients,
  • proper O2, pH, temp, etc.

51
Culture
  • A population of microorganisms cultivated in a
    medium

52
Culture media .
  • are designed to provide all the essential
    nutrients in solution for bacterial growth

53
Pure Culture
  • Culture containing only one species of organism.

54
Types of Media
  • Liquid for pure batch cultures
  • Solid isolation of pure cultures. Agar.

55
Assignment
  • Read online textbook section Culture Media for
    the Growth of Bacteria Page 16 of Word document
    on CD

56
Section 3
  • Survey of the Microbial World

57
Terminology
  • Microbiology study of small living organisms of
    microscopic size.
  • Protist a microorganism in the Kingdom
    Protista. (includes most microorganisms)

58
  • Prokaryote - a cell in which the nuclear
    substance is not enclosed in a membrane.
    (includes the bacteria)

59
  • Eukaryote a cell that has a definitive or true
    nucleus (these cells make up the bodies of all
    non protist organisms)

60
  • Virus obligate intracellular parasitic organism
    that is smaller than bacteria. They can only
    reproduce in cells of host organisms.

61
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62
The Cell A review
  • General Structure
  • Membrane -
  • Nucleus contains nucleoproteins
  • Cytoplasm (cyto cell, plasm formed substance)

63
  • Characteristics of all living things
  • Reproduce
  • Energy needs and growth
  • Elimination of waste
  • Response to stimuli
  • Susceptible to mutations

64
  • Look for the 3 general structures in each of the
    following three organisms.

65
Paramecium a representative cell
66
A Typical Bacterial Cell
67
Euglena another typical cellular organism
68
Prokaryotic cells
  • No internal membranes
  • Division by fission not by mitosis
  • Cell wall with mucopeptide material.

69
Some Prokaryotic cells
  • have capsule or slime layer covering cell wall
  • have flagella (whip) for motility

70
Prokaryotic cytoplasm
  • May contain
  • Ribosomes proteins and RNA
  • Granules - chemicals
  • Nuclear material - DNA
  • Mesosomes - folds of Cytoplasmic membrane

71
Eukaryotic cells
  • Contain internal membranes - called Endoplasmic
    Reticulum (ER) distinguishing it from the
    prokaryotic cells.

72
Eucaryotic cell structures
  • ER
  • Nucleus
  • Golgi apparatus
  • Mitochondria
  • Chloroplasts
  • Vacuole

73
  • Microtubules
  • Microfilaments
  • Flagella
  • Cilia
  • Cell walls

74
Groups of Microorganisms
  • Prokaryotic protists
  • Eucaryotic protists
  • Viruses

75
Please note
  • Classification schemes often change.
  • Some literature lists Bacteria as a member of
    Kingdom Monera and not Protista.
  • Others split bacteria into two kingdoms called
    Archaebacteria and Eubacteria.

76
Prokaryotic Protists
  • Bacteria
  • 3 basic shapes
  • Size range from 0.5 mm to 2.5 mm
  • Cyanobacteria
  • Contain chlorophyll and photosynthesize

77
Eucaryotic Protists
  • Fungi
  • Rigid cell walls, no chlorophyll
  • Protozoa
  • Include flagellates, amebas, ciliates, and
    sporozoa more on that later !
  • Algae
  • Contain chlorophyll

78
Paramecium a protozoan
79
Viruses
  • Not cellular
  • Obligate parasites
  • 20 nm to 300 nm in size
  • Host specific

80
Section 4
  • - Protozoans

81
Protozoa introduced
  • Classified by locomotion
  • Flagellates
  • Amebas
  • Ciliates
  • Sporozoa

Many plankton are protozoans !
82
Flagellates
  • Cell is surrounded by pellicle gives the
    organism shape.

83
Disease-causing Flagellates
  • Giardia lablia that nasty intestinal parasite
  • Trypanosoma (African sleeping sickness spread
    by tsetse flies)
  • Trichomonas - vaginitis, spread as venereal
    disease

84
Amebas
  • Name comes from Greek word meaning change
  • Pseudopodia false feet are protoplasmic
    extensions

85
Disease-causing Amebas
  • Entamoeba histolytica cause of amebic
    dysentery.
  • (yes there are non disease causing Amebas like
    Ent. Gingivalis and Ent. Coli)

86
Ciliates
  • Contain at least one macronucleus and one
    micronuclei per cell.
  • May have partial covering of cilia or complete
    covering like Paramecium

87
Disease-causing Ciliates
  • Only one !!
  • Balantidium coli causes bloody diarrhea which
    is transmitted by infected food or water from
    swine feces.

88
Sporozoans
  • All are parasitic.
  • Feed on hosts cells or body fluids.
  • All produce spores a resistant or dormant cell.

89
Disease-causing Sporozoans
  • Plasmodium - Malaria-causing sporozoans
  • Toxoplasma gondii - mild infection usually
    requiring no treatment but can cause birth
    defects if mother is infected.

90
The changing face of Protozoans
  • Cyst formation -
  • Trophozoites - organism in stage of active
    growth, also called vegetative stage.

91
Section 5
Many phytoplankton are algae !
  • Algae

92
Algae Shapes
  • Single cells may be
  • Spherical
  • Rod-shaped
  • Club-shaped
  • Spindle-shaped

93
Algae Growth
  • May grow as single cells.
  • May grow in multicellular colonies.
  • Colonies may be made of identical cells.
  • Colonies may be made of different cells that
    specialize.

94
Algae Cellular Structure
  • Algae is Eukaryotic
  • Contain chloroplasts.
  • If motile, they have flagella.
  • May have stalks or spines for anchoring.

95
Classification of Algae
  • Based on
  • Pigment
  • Chemistry
  • Flagella
  • Cell wall features
  • Cell organization (single or colony)
  • Life cycles and reproduction.

96
Some major Divisions
  • Chlorophycophyta green algae
  • Rhodophycophyta red algae
  • Chrysophycophyta golden algae
  • Bacillariophycophyta - diatoms

97
Diatoms
98
Blue-green Bacteria/algae
99
Reproduction
  • Assexual
  • By binary fission (cell division producing two
    daughter cells)
  • Spore formation.

100
  • Sexual
  • Gametes
  • Zygote

101
Growth Requirements
  • Algae are aerobic need oxygen
  • Algae are photosynthetic need light
  • Algae require moisture need water
  • Algae require nutrients
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