Principles of
Bacteriology
Prepared by
Hamad ALAssaf
Alassaf_h@yahoo.com
2015
1
Bacterial Structure
 Peptidoglycan : gives rigid support, protect
against osmotic pressure.
Capsule: protect against phagocytosis.
Polysaccharides except Bacillus anthrax, which
contains D-glutamate.
Spore: provides resistance to dehydration, heat,
and chemicals. Keratin-like coat; dipicolinic
acid.
2
Gram Stain Limitations
• These organism do not Gram stain well:
1- Treponema (too thin to be visualized).
2- Rickettsia (intracellular parasite).
3- Mycobacteria (No cell wall).
4- Legionella pnumophila ( primarily intracellular).
5- Chlamydia (intracellular parasite; lacks muramic acid in cell wall).
3
Stains
• Giemsa : Borrelia, Plasmodium, Trypanosomes, Chlamydia.
• PAS (periodic acid-Schiff): Stains glycogen, mucopolysaccharides; used to
diagnose Whipple’s disease (Tropheryma whippelii).
• Ziehl-Neelsen: Acid-fast organism.
• Indian ink: Cryptococcus neoformans, and used to stain thick polysaccharide capsule
red.
• Silver stain: Fungi (e.g. Pneumocystis), Legionella.
4
Silver Stain
Indian ink
Giemsa Stain
ZN stain
5
Special culture requirements
Microrganism Media used for isolation
H. influenzae
N. gonorrhoeae
B. pertussis
C. diphtheriae
M. tuberculosis
M. pneumoniae
Legionella
Fungi
Enteric pathogens
Vibrio cholerae
Chocolate agar with factors V (NAD) and X (hematin).
Thayer-Martin Media (VPN)
Bordet-Gengou agar
Tellurite plate, Loffler’s media
Lowenstein-Jensen agar
Eaton’s agar
Charcoal yeast extract agar buffered with cysteine
Sabouraud’s agar
Hektone enteric agar or Xylose-Lysine-Deoxycholate agar
TCBS (alkaline growth medium)
6
•Encapsulated Bacteria: Positive Quellung reaction e.g.
Streptoccoccus pneumonia, Klebsiella pneumonia, Haemophilus
influnesae type B, Neisseria meningitides, Salmonella, Group B
streptococcus.
•Urease-positive bugs: Proteus, Klebsiella, H.pylori, Ureaplasma.
•Pigment-producing bacteria: Actinomyces israelii (yellow –
sulfur), S.aureus (yellow pigment), Pseudomonas aeruginosa (bluegreen pigment), Serratia marcescens (red pigment).
7
Bacterial virulence factors
• These promote evasion of host immune response.
1- Protein A (S. aureus): Binds Fc region of Ig. Prevent opsonization and
phagocytosis.
2- IgA protease: Enzyme that cleaves IgA. Secreted by S.pneumoniae, H.
influenza type B, and Neisseria in order to colonize respiratory mucosa.
3- M protein (group A streptococcus): helps prevent phagocytosis.
8
Normal Flora
• Is found on body surfaces contiguous with the outside environment.
• Is semi-permanent, varying with major life changes.
• Can cause infection:
- if misplaced , e.g. fecal flora to urinary tract or abdominal cavity, or skin flora to catheter.
- if person become compromised, normal flora may overgrow (oral thrush).
• contribute to health:
- Protective host defense by maintaining conditions such as pH so other organism may not
grow.
- Serves nutritional function by synthesizing: K and B12 vitamins.
 Normal Flora e.g. Nose (S.aureus), cutaneous (Staphylococcus epidermidis), Oropharynx
(Viridans streptococci), Vagina (Lactobacillus), colon (E.coli).
Blood and Stomach: No Normal Flora present.
9
Colonization
• The first stage of microbial infection is colonization:
• Pathogens usually colonize host tissues that are in contact with the external
environment.
• Sites of entry in human hosts include the urogenital tract, the digestive tract,
the respiratory tract and the conjunctiva.
• Adherence to cell surfaces involves:
- Pili/fimbriae: primary mechanism in most gram negative cells.
- Teichoic acids: primary mechanism of gram positive cells.
- Adhesion: colonizing factor adhesions, pertussis toxin, and hemagglutinins.
- IgA proteses: cleaved Fc portion may coat bacteria and bind them to cellular
Fc receptor.
10
Definitions
• Carrier: person colonized by a potential pathogen without overt
disease.
• Bacteremia: bacteria in bloodstream without overt clinical signs.
• Septicemia: bacteria in bloodstream (multiplying) with clinical
symptoms.
 Spores of fungi have a reproductive role.
11