Food Biochemistry and Food Processing

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Table 31.4.

Comparison of Pathogen Subtyping and Verification Methods

Amount of

Method

Advantages

Disadvantages

DNA required

RFLP

• Simplicity


• Requires pure cultures of pathogen

3–5



g

• Can be done with restriction


• Many bands to compare; confusing,

endonucleases for serotype

may be hard to see differences

level discrimination or combined

• Cannot compare organisms at the genus or

with PCR for lineage comparisons

family level

• Partial digestion and faint bands may be a

problem

AFLP

• PCR to amplify a few genes and simplify


• Requires pure cultures of pathogen

10–100 ng

result analysis

• Cannot compare organisms at the genus or
  
  
  • Can compare differences between bands on a gel
  
  
  
  

family level

• Fewer bands to compare than with RFLP• Can compare among labs

RAPD

• Universal primers that are not gene specific


• Requires pure cultures of pathogen

10 ng

• Is not very reproducible between gels or labs• Cannot compare organisms at the genus or

family level

Ribotyping

• Similar to RFLP, but visualizes only bands


• Requires pure cultures of pathogen

1



g

correlating to the

rrn

portion of ribosomes

so fewer bands to compare

• Can compare organisms at the genus and

family level

Pulsed-field

• PulseNet database available to


• Time consuming—3 days for results

Measured by

gel

coordinate and compare

• Requires pure cultures of pathogen

turbidity of

Electrophoresis

results—can track outbreaks

• Patterns may change after intestinal

culture–lysis

• Can compare differences between

passage; differences in patterns may

and DNA

bands on a gel

not indicate actual strain

digestion

• Results are comparable among labs

differences

performed in

• Cannot compare organisms at the

agarose plugs

genus or family level

FISH

• May detect one pathogen in a mixed


• Cells must be fixed and

Single cell

community of microbes

permeabilized before visualization

level

Source:

Adapted from Gurtler and Mayall 2001, Savelkuoh et al. 1999.