Fundamental Food Microbiology, Third Edition - Fuad Fathir

DETECTION OF MICROORGANISMS IN FOOD AND FOOD ENVIRONMENT 561
C. Enumeration of Injured Microbial Groups by Selective Media
Sublethally injured coliforms and pathogenic bacterial cells, when enumerated by
selective agar media, may die because of their developed sensitivity to selective
agents in the media and thus cannot be detected (see Chapter 9). They are first
allowed to repair the injury in nonselective media (broth or agar) for a short period,
which enables the cells to regain resistance to the selective agents. Following repair,
they can be exposed to selective media. For enumerating coliforms in food that may
contain injured coliforms, the diluted sample can first be pour plated in nonselective
plate count agar and incubated for 1–2 h at 25–35�C, thus enabling the repair of the
cells. Double-strength violet red bile agar can then be overlaid on the plates and the
plates incubated at 35�C for 24 h for selective growth of coliforms. 1–3
D. Dilution Scheme, Plating, Incubation, Selection of Plates
for Counting CFUs, and Reporting Results
The standard and recommended method books listed above have detailed descriptions
on these steps. They should be followed as accurately as possible. The average
of CFUs from duplicate or triplicate plates from a selected dilution should be used
to report the counts of the specific microbial group per ml or g, or cm 2 of the food
sample. This data should be used in the proper interpretation of microbiological
quality of the food.
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VI. QUALITATIVE METHODS TO ISOLATE MICROORGANISMS
IN FOODS
A. Isolation of Pathogens
The main objective of this method is to determine whether a sample contains viable
cells or spores of a specific pathogen. Foods are tested for several pathogens, such
as Salmonella, Esc. coli O157:H7, Lis. monocytogenes, Vibrio cholerae, and Shigella
spp., by the specific isolation procedure when necessary. For other pathogens, such
as enteropathogenic Esc. coli, Yer. enterocolitica, and Cam. jejuni, isolation procedures
are not generally used; instead, enumeration procedures are used.
An isolation procedure generally contains several steps, such as nonselective
preenrichment, followed by selective enrichment, and then testing on an agar medium
containing selective and differential agents. It is assumed that a food normally
contains a low population of a pathogen as compared with associative microorganisms,
and the pathogens could be in the injured state. The food sample (e.g., 25 g)
is first preenriched in a nonselective broth and incubated for the injured cells to
repair and then multiply in order to reach moderately high numbers (along with
many other associated microorganisms). An aliquot is then transferred from the
preenrichment broth to a selective enrichment broth and incubated. It is expected
that during incubation, the specific pathogen and closely related microorganisms
will selectively grow to a high number, whereas many of the associated microor-