BACTERIAL CLASSIFICATION AND DIAGNOSIS OF BACTERIAL DISEASE

1
BACTERIAL CLASSIFICATION AND DIAGNOSIS OF
BACTERIAL DISEASE

• Disease causing agent
• Bacterial, fungal, viral or other?
• Treatment
• Antibiotic sensitivity
• Source of infection
• Food, air or contact?
• Epidemiology
• Cholera, Diphtheria
• Prevention
• E. coli O157H7 outbreak, Shigella, Listeria

2
Antibiotic sensitivity
3
Site of sampling

• Sterile sites
• Blood
• Cerebrospinal fluid (CSF)
• Body fluids (Peritoneal and pleural)
• Non-sterile (normal flora)
• Respiratory tract
• Ear, eye and mouth
• Skin (wound and abscess)
• Urine (mid-stream)
• Feces

4
Bacterial classification

• Wall structure
• Gram
• Staphylococcus, Streptococcus, Clostridium,
  Bacillus
• Gram -
• Enteric, respiratory and others
• Acid-fast
• Mycobacterium
• Wall-less
• Mycoplasma
• Unusual
• Obligate intracellular
• Rickettsia, Chlamydia

Bacteria
G G- AF WL IC
5
Bacterial classification

• Cell morphology
• Shapes
• Rod
• Cocci
• Spiral
• Associations
• Individual
• Diplo-
• Staphylo-
• Strepto-

Bacteria
G G- AF WL IC
Rod Cocci
Rod Cocci Spiral
6
Bacterial classification

• Growth characteristics
• Oxygen requirement
• Aerobic
• Anaerobic
• Microaerophilic, aerotolerant
• Facultative
• Spore formation
• Intracellular/extracellular
• Fastidious/non-fastidious

Bacteria
G G- AF WL IC
Rod Cocci
Rod Cocci Spiral
/- -O2
spore -
7
Classification Diagnosis

• Type of colonies
• Appearance
• Color, shape, size and smoothness
• On differential media
• Blood, MacConkey, EMB
• On selective media
• MacConkey, Thayer-Martin

8
Classification Diagnosis

• Metabolism
• Utilization of specific substrates
• Lactose (Sal/Shi/Yer/)-
• Citrate (E. coli-/Klebsiella)
• Production of certain end products
• Fermentation end products
• Acid (acetate, propionic acid, butyric acid etc.)
• Acetoin
• Alcohol
• Amine
• H2S

9
Classification Diagnosis

• Specialized tests
• Immunological
• O-, H- K-Ag (serotype)
• Precipitation, agglutination
• Specialized enzymes
• Catalase--- Staph. vs. Strep-.
• Coagulase---S. aureus vs. S. epidermidis-
• Oxidase---Neisseria gonorrhoea
• Urease---Proteus, Helicobacter
• Antibiogram pattern
• Phage typing
• Fatty acid profile

10
Immunological detection
11
Conventional diagnosis methods
12
Conventional method

• Depend on ability to culture
• Treponema pallidum
• Slow, esp. for fastidious species
• Mycobacterium spp.
• Not always definitive

13
Molecular diagnosis

• Ribotyping
• Restriction fragment length polymorphism (RFLP)
• DNA hybridization
• PCR, RT-PCR and RAPD
• Nucleic acid sequence analysis
• Phage-GFP (TB)

14
RFLP
GGATCC CCTAGG
15
DNA hybridization
16
In situHybridization
17
PCR
RT-PCR
Rate of increase 2n
18
RAPD of P. aeruginosa
19
Molecular diagnosis

• Reduce reliance on culture
• Faster
• More sensitive
• More definitive
• More discriminating
• Techniques adaptable to all pathogens

• Technically demanding
• Relatively expensive
• Can be too sensitive
• Provides no information if results are negative

20
Bioterrorism

• Pathogen detection
• Fast and accurate
• Mobile
• Inexpensive
• Source investigation

21
Differentiating Staphylococci from Streptococci

• Gram stain and morphology
• Both Gram
• Staphylococci bunched cocci
• Streptococci chained cocci (S. pneumoniae form
  diplococcus)
• Enzyme tests
• Staphylococci catalase
• Streptococci catalase -
• Growth
• Staph. large colonies (non-fastidious), some
  hemolytic
• Strep. small colonies (fastidious), many
  hemolytic (a or b)

22
Staphylococci

• S. aureus coagulase
• S. epidermidis coagulase -

23
Streptococci
24
Differentiating the Gram- bacteria

• Cocci
• Neisseria
• Rods
• Type of disease they cause
• Enteric Gram- rods
• API test
• Curved
• Vibrio, Campylobacter, Helicobacter
• Spiral Gram- organisms
• Spirochetes

25
Gram negative
Curved rods
Straight rods
Lactose Lactose-
TCBS agar Yellow Oxidase Vibrio
Campy blood agar 42oC 25oC- Campylobacter
Citrate Citrate-
H2S H2S-
Klebsiella E. coli Salmonella
Shigella
26
Bacteria
Intra Cellular
Wall Less
Gram
Gram-
Acid Fast
Cocci
Rod
Cocci
Rod
Spiral
M.t. M.l. N.c.
Rickettsia Coxiella Erlichia Chlamydia
Mycoplasma
Staph.
Strep.
Non-spore
Spore
Treponema Borrelia Leptospira
Neisseria Moraxella
Fil
Rod
O2
-O2
Curve
B.a. B.c.
C.b. C.t. C.p. C.d.
S. a. S. e. S. s.
A B Pn Vir
A.i.
C.d. L. m.
Straight
Vibrio Campylobacter Helicobacter
O2
-O2
/-O2
Other
P.a.
Enteric
Bact.
Resp.
Zoo
GU
H. ducreyi Gardnerella Calymmatobacterium
Yersinia Pasteurella Brucella Francisella Streptob
acillus
Bordetella. H. influenzae Legionella