classificationof bacteria
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Classificationof BacteriaTRANSCRIPT
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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, Treponema
G+ G- AF WL IC
Bacteria
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Bacterial classification
Cell morphology Shapes
Rod Cocci Spiral
Associations Individual Diplo- Staphylo- Strepto- Filaments
G+ G- AF WL IC
Bacteria
Rod Cocci Rod Cocci Spiral
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Gram stains
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Gram Stains of Mixtures
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Cell shapes
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Bacterial classification
Growth characteristics Oxygen requirement
AerobicAnaerobic
Microaerophilic, aerotolerantFacultative
Spore formation Intracelular/extracellular Fastidious/non-fastidious
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Endospore
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Classification & Diagnosis
Type of colonies Appearance
Color, shape, size and smoothness On differential media
Blood, MacConkey, EMB, Manitol salt agar
On selective mediaMacConkey, EMB, Manitol salt agar
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Colony morphology/color
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Bacterial colony morphology
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Capsule
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MacConkey Agar Plate
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Blood Agar Plates and Hemolysis
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Mannitol salt agar
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Classification & Diagnosis Metabolism
Utilization of specific substrates Lactose (Sal/Shi/Yer/)-
Citrate (E.coli-/Klebsiela+)
Production of certain end products Fermentation end products
Acid (acetate, propionic acid, butyric acid etc.)
Aceton Alcohol Amine H2S
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Triple Sugar Iron Agar Slants
TSI•Fermentation of glucose, lactose, and/or sucrose•Reduction of sulfur to hydrogen sulfide•Gas formation
Used for Enterobacteriaceae
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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
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Urease Test
Urea CO2 + NH3 NH4+ + OH-
H2O
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Indole Test
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API test strip
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Bacteriophage Plaques
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Conventional method
Depend on ability to culture Treponema pallidum
Slow, esp. for fastidious species Mycobacterium spp.
Not always definitive
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Immunological detection
ELISA
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Instant View tests
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Molecular diagnosis
Ribotyping Restriction fragment length
polymorphism (RFLP) DNA hybridization PCR, RT-PCR and RAPD Nucleic acid sequence
analysis Phage-GFP (TB)
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RFLP
GGATCCCCTAGG
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DNA hybridization
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In situHybridization
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Rate of increase2n
PCR
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Real time PCRQR
5'3' 5'
3'
5' 3' 3' 5'
QR
5' 3' 3' 5'
QR
Q
5' 3' 3' 5'
R
PrimPrimerer
PrimePrimerr
ProbeProbe
Taq DNA Taq DNA PolPol
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RAPD of P. aeruginosa
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DNA and protein chips
A B C D
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Microarray array mediated pathogen detection
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Ab chip mediated pathogen detection
-O157:H7
-Salmonella
-O157:H7
-Salmonella
E.coli O157:H7
Salmonellatyphimurium
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Molecular diagnosis
Reduce reliance on culture
Faster High 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
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Bioterrorism
Pathogen detection Fast and accurateMobile Inexpensive
Source investigation
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Biosensor Mediated Pathogen Detection
Step 1: Attach Capture Antibody to Fiber Optic Waveguide
Step 2: Apply Sample. Target Organism Binds to WaveguideStep 3: Wash Away Other Cells and Debris
Y Y Y Y YY YY Y YY YY
YY YYYY
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Step 1: Attach Capture Antibody to Fiber Optic WaveguideStep 2: Apply Sample. Target Organism Binds to WaveguideStep 3: Wash Away Other Cells and DebrisStep 4: Add Specific Detection Antibody Step 5: Turn on Laser. Detection Antibody Fluoresces,
Indicating a Positive Sample.
Y Y Y Y YY YY Y YYY
Y YYYY
Y Y Y YY YY YY
Y Y YY Y YY Y
Y Y
Biosensor Mediated Pathogen Detection
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Biosensors
Analyte 2000TM
for laboratory research and development
RAPTORTM automated version for field
analysis
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Differentiating Staphylococci from Streptococci
Gram stain and morphology Both Gram + Staphylococci: bunched cocci Streptococci: chained cocci
S. pneumoniae: diplococcus
Enzyme tests Staphylococci: catalase + Streptococci: catalase -
Growth Staph.: large colonies (non-fastidious), some hemolytic Strep.: small colonies (fastidious), many hemolytic (
or )
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Staphylococci
S. aureus : coagulase + S. epidermidis : coagulase -
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Streptococci
Streptococci On blood agar Growth inhibition discS. pyogenes (group A) -hemolytic Sensitive to bacitracinS. agalactiae (group B) -hemolytic Resistant to bacitracin
S. pneumoniae (pneumococcus) -hemolytic Sensitive to optochinViridans -hemolytic Resistant to optochin
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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
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Gram negative
Straight rods Curved rods
Lactose+ Lactose-
Citrate+ Citrate- H2S+ H2S-
Klebsiella E.coli Salmonella Shigella
Campy blood agar42oC+ 25oC-
Campylobacter
TCBS agarYellowOxidase+
Vibrio
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BacteriaGram+
Cocci Rod
Staph. Strep. Non-spore Spore
S. a.S. e.S. s.
ABPnVir
Fil Rod
A.i. C.d.L. m.
+O2 -O2
B.a.B.c.
C.b.C.t.C.p.C.d.
Acid Fast
M.t.M.l.N.c.
IntraCellular
RickettsiaCoxiellaErlichiaChlamydia
WallLess
Mycoplasma
Gram-
CocciRodSpiral
StraightCurve
+O2 -O2+/-O2 Other
TreponemaBorreliaLeptospira
NeisseriaMoraxella
P.a. Enteric Bact.
Resp. Zoo GU
Bordetella.H. influenzaeLegionella
YersiniaPasteurellaBrucellaFrancisllaStreptobacillus
H. ducreyiGardnerellaCalymmatobacterium
VibrioCampylobacterHelicobacter