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 O157:H7 outbreak, Shigella, Listeria
Antibiotic sensitivity
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
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
G+ G- AF WL IC
Bacteria
Bacterial classification
Cell morphology Shapes
Rod Cocci Spiral
Associations Individual Diplo- Staphylo- Strepto-
G+ G- AF WL IC
Bacteria
Rod Cocci Rod Cocci Spiral
Bacterial classification
Growth characteristics Oxygen requirement
Aerobic Anaerobic
Microaerophilic, aerotolerant
Facultative Spore formation Intracellular/extracellular Fastidious/non-fastidious
G+ G- AF WL IC
Bacteria
Rod Cocci Rod Cocci Spiral
+ spore - + +/- -O2
Classification & Diagnosis
Type of colonies Appearance
Color, shape, size and smoothness On differential media
Blood, MacConkey, EMB On selective media
MacConkey, Thayer-Martin
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
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
Immunological detection
Conventional diagnosis methods
Conventional method
Depend on ability to culture Treponema pallidum
Slow, esp. for fastidious species Mycobacterium spp.
Not always definitive
Molecular diagnosis
Ribotyping Restriction fragment length polymorphism
(RFLP) DNA hybridization PCR, RT-PCR and RAPD Nucleic acid sequence analysis Phage-GFP (TB)
RFLP
GGATCCCCTAGG
DNA hybridization
In situHybridization
Rate of increase2n
PCR RT-PCR
RAPD of P. aeruginosa
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
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 ( or )
Staphylococci
S. aureus: coagulase + S. epidermidis: coagulase -
Streptococci
Streptococci On blood agar Growth inhibition disc
S. pyogenes (group A) -hemolytic Sensitive to bacitracin
S. agalactiae (group B) -hemolytic Resistant to bacitracin
S. pneumoniae (pneumococcus) -hemolytic Sensitive to optochin
Viridans -hemolytic Resistant to optochin
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
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
BacteriaGram+ Gram- Acid Fast
IntraCellular
WallLess
Cocci Rod CocciRodSpiral
Staph. Strep. Non-spore Spore
StraightCurve
+O2 -O2+/-O2 Other
S. a.S. e.S. s.
ABPnVir
Fil Rod
A.i. C.d.L. m.
M.t.M.l.N.c.
+O2 -O2
B.a.B.c.
C.b.C.t.C.p.C.d.
TreponemaBorreliaLeptospira
NeisseriaMoraxella
P.a. Enteric Bact.
Resp. Zoo GU
Bordetella.H. influenzaeLegionella
YersiniaPasteurellaBrucellaFrancisellaStreptobacillus
H. ducreyiGardnerellaCalymmatobacterium
RickettsiaCoxiellaErlichiaChlamydia
Mycoplasma
VibrioCampylobacterHelicobacter