Culture, Microscopic, and Sampling Methods
227
Swab/Swab-Rinse Methods
Swabbing is the oldest and most widely used method for the microbiological examination of surfaces
not only in the food and dairy industries but also in hospitals and restaurants. The swab-rinse method
was developed by W.A. Manheimer and T. Ybanez. Either cotton or calcium alginate swabs are
used. If one wishes to examine given areas of a surface, templates may be prepared with openings
corresponding to the size of the area to be swabbed, for example, 1 in
2 or 1 cm
2 . The sterile template
is placed over the surface, and the exposed area is rubbed thoroughly with a moistened swab. The
exposed swab is returned to its holder (test tube) containing a suitable diluent and stored at refrigerator
temperatures until plated. The diluent should contain a neutralizer, if necessary. When cotton swabs
are used, the organisms must be dislodged from the fibers. When calcium alginate swabs are used, the
organisms are released into the diluent upon dissolution of the alginate by sodium hexametaphosphate.
The organisms in the diluent are enumerated by a suitable method such as SPC, but any of the culture
media may be used to test specifically for given groups of organisms. In an innovation in the swab-rinse
method presented by Koller,
68 1.5 ml of fluid is added to a flat surface, swabbed for 15 seconds over
a 3-cm
2 area, and volumes of 0.1 and 0.5 ml collected in microliter pipettes. The fluid may be surface
or pour plated using plate count agar or selective media.
Concerning the relative efficacy of cotton and calcium alginate swabs, most investigators agree that
higher numbers of organisms are obtained by use of the latter. Using swabs, some researchers recovered
as few as 10% of organisms from bovine carcasses,
87 47% of Bacillus subtilis spores from stainlesssteel surfaces,
7 and up to 79% from meat surfaces.
22,93 Swab results from bovine carcasses were on the
average 100 times higher than by contact plate method, and the deviation was considerably lower.
87
The latter investigators found the swab method to be best suited for flexible, uneven, and heavily
contaminated surfaces. The ease of removal of organisms depends on the texture of the surface and the
nature and types of biota. Even with its limitations, the swab-rinse method remains a rapid, simple,
and inexpensive way to assess the microbiological biota of food surfaces and utensils.
The use of the ATP assay system to detect the presence of cells within 2–5 minutes after swabbing
allows it to be used on-line. Although the ATP assay as used in this regard is not specific for bacteria, it
provides valuable information on the level of cell contamination of a surface and can be used to make
quick assessments of the relative efficacy of surface cleaning methods. The basis of the ATP assay is
described in Chapter 11.
Contact Plate
The replicate organism direct agar contact (RODAC) method employs special Petri plates, which
are poured with 15.5–16.5 ml of an appropriate plating medium, resulting in a raised agar surface.
When the plate is inverted, the hardened agar makes direct contact with the surface. Originated by
Gunderson and Gunderson in 1945, it was further developed in 1964 by Hall and Hartnett. When
surfaces are examined that have been cleaned with certain detergents, it is necessary to include a
neutralizer (lecithin, Tween 80, and so on) in the medium. Once exposed, plates are covered and
incubated, and the colonies enumerated.
Perhaps the most serious drawbacks to this method are the covering of the agar surface by spreading
colonies, and its ineffectiveness for heavily contaminated surfaces. These can be minimized by using
plates with dried agar surfaces and by using selective media.
28 The RODAC plate has been shown
to be the method of choice when the surfaces to be examined are smooth, firm, and nonporous.
7,87
Although it is not suitable for heavily contaminated surfaces, it has been estimated that a solution that
227
Swab/Swab-Rinse Methods
Swabbing is the oldest and most widely used method for the microbiological examination of surfaces
not only in the food and dairy industries but also in hospitals and restaurants. The swab-rinse method
was developed by W.A. Manheimer and T. Ybanez. Either cotton or calcium alginate swabs are
used. If one wishes to examine given areas of a surface, templates may be prepared with openings
corresponding to the size of the area to be swabbed, for example, 1 in
2 or 1 cm
2 . The sterile template
is placed over the surface, and the exposed area is rubbed thoroughly with a moistened swab. The
exposed swab is returned to its holder (test tube) containing a suitable diluent and stored at refrigerator
temperatures until plated. The diluent should contain a neutralizer, if necessary. When cotton swabs
are used, the organisms must be dislodged from the fibers. When calcium alginate swabs are used, the
organisms are released into the diluent upon dissolution of the alginate by sodium hexametaphosphate.
The organisms in the diluent are enumerated by a suitable method such as SPC, but any of the culture
media may be used to test specifically for given groups of organisms. In an innovation in the swab-rinse
method presented by Koller,
68 1.5 ml of fluid is added to a flat surface, swabbed for 15 seconds over
a 3-cm
2 area, and volumes of 0.1 and 0.5 ml collected in microliter pipettes. The fluid may be surface
or pour plated using plate count agar or selective media.
Concerning the relative efficacy of cotton and calcium alginate swabs, most investigators agree that
higher numbers of organisms are obtained by use of the latter. Using swabs, some researchers recovered
as few as 10% of organisms from bovine carcasses,
87 47% of Bacillus subtilis spores from stainlesssteel surfaces,
7 and up to 79% from meat surfaces.
22,93 Swab results from bovine carcasses were on the
average 100 times higher than by contact plate method, and the deviation was considerably lower.
87
The latter investigators found the swab method to be best suited for flexible, uneven, and heavily
contaminated surfaces. The ease of removal of organisms depends on the texture of the surface and the
nature and types of biota. Even with its limitations, the swab-rinse method remains a rapid, simple,
and inexpensive way to assess the microbiological biota of food surfaces and utensils.
The use of the ATP assay system to detect the presence of cells within 2–5 minutes after swabbing
allows it to be used on-line. Although the ATP assay as used in this regard is not specific for bacteria, it
provides valuable information on the level of cell contamination of a surface and can be used to make
quick assessments of the relative efficacy of surface cleaning methods. The basis of the ATP assay is
described in Chapter 11.
Contact Plate
The replicate organism direct agar contact (RODAC) method employs special Petri plates, which
are poured with 15.5–16.5 ml of an appropriate plating medium, resulting in a raised agar surface.
When the plate is inverted, the hardened agar makes direct contact with the surface. Originated by
Gunderson and Gunderson in 1945, it was further developed in 1964 by Hall and Hartnett. When
surfaces are examined that have been cleaned with certain detergents, it is necessary to include a
neutralizer (lecithin, Tween 80, and so on) in the medium. Once exposed, plates are covered and
incubated, and the colonies enumerated.
Perhaps the most serious drawbacks to this method are the covering of the agar surface by spreading
colonies, and its ineffectiveness for heavily contaminated surfaces. These can be minimized by using
plates with dried agar surfaces and by using selective media.
28 The RODAC plate has been shown
to be the method of choice when the surfaces to be examined are smooth, firm, and nonporous.
7,87
Although it is not suitable for heavily contaminated surfaces, it has been estimated that a solution that