In restriction fragment length polymorphism (RFLP), a restriction
enzyme is used to cut DNA into a number of fragments depending on the
number and location of restriction sites present. Separation of these
fragments by electrophoresis will reveal a fingerprint of that particular
organism. Ribotyping, a particular type of RFLP where the probes used
are specific for rRNA genes, has been automated and equipment is
available commercially that can ribotype an organism in 8 hours.
A commonly encountered problem is that RFLP patterns are exces-
sively complex with many poorly resolved bands. Use of restriction
enzymes with fewer sites on the bacterial chromosome overcomes this
problem but specialist electrophoretic techniques are needed to separate
the large DNA fragments produced. This is pulsed field gel elect-
rophoresis (PFGE) which uses an alternating electric field to tease apart
the large DNA molecules.
10.7 Laboratory Accreditation
From what has already been said it should be clear that there can be a
number of different ways of detecting the same organism in a food
matrix. The choice of method used can be governed by several factors
and the relative merits of different methods is a topic of constant
investigation and debate. This can however lead to the situation where
differences in a result reported by two laboratories simply reflect the
different method used.
In addition to problems arising from intrinsic differences in the
performance of different methods, the same method in different labora-
tories can be subject to variation introduced by factors such as differences
in procedures, equipment and its calibration. Some possible examples
would include autoclave temperature profile when sterilizing media, time
and temperature of incubation, sources of medium components and, of
course, competence and experience of laboratory personnel.
A number of approaches are adopted to avoid such potential prob-
lems. Several national and international bodies approve standard meth-
ods for conducting certain analyses and one of these should be adopted
for routine work and strictly adhered to wherever possible. Testing
laboratories also often participate in proficiency testing schemes where
a central body distributes standard samples for analysis, often specifying
the precise time this should be conducted and the method to be used.
Results are reported back, collated and a report circulated to participat-
ing laboratories which can then judge their performance against that of
others. Finally, laboratories can seek some form of third party, inde-
pendent recognition. There are quality systems such as the Good Labo-
ratory Practice scheme and standards such as (ISO 9000 series) which are
concerned with the quality of management within the organization but
394 Methods for the Microbiological Examination of Foods