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REVISED 121213
Identification of presumptive
Salmonella
colonies from selective/differential agar media as
Salmonella
1
spp. has historically been achieved using a variety of biochemical and serological procedures. In the
2
case of food and environmental sample analysis, these procedures are specified in reference methods
3
such as those in the U.S. Food and Drug Administration’s
Bacteriological Analytical Manual
(BAM) [1]
4
and the U.S. Department of Agriculture’s
Microbiology Laboratory Guidebook
(MLG) [2]. These methods
5
include conventional biochemical tests, miniaturized biochemical test devices, automated biochemical
6
identification platforms, and serological agglutination tests using
Salmonella
-specific antisera. The
7
biochemical identification procedures, although accurate and reliable, generally require 6-24 h to obtain
8
results. The serological procedures may be rapid, but often require subculture to enhance antigen
9
expression, especially in the case of flagellar (H) antigen typing.
10
As an adjunct or alternative to biochemical and serological procedures, nucleic acid-based identification
11
methods hold promise for providing timely and accurate results. This has been acknowledged, for
12
example, by reference in both BAM and MLG to use of nucleic acid-based methods for identification of
13
Listeria monocytogenes
[3, 4].
14
The ANSR®
Salmonella
assay was originally developed for rapid screening of enriched food and
15
environmental samples. The assay is an isothermal nucleic acid amplification procedure, based on the
16
nicking enzyme amplification reaction (NEAR) technology [5]. The ANSR method has been evaluated in
17
three AOAC Performance Tested Method
SM
validation studies, leading to certification as PTM method
18
061203, with claims for a wide variety of food and environmental sample types [6-7]. In these studies,
19
the ANSR method exhibited sensitivity comparable to that of the FDA/BAM and USDA/MLG reference
20
culture methods by probability of detection statistical analysis, as well as > 99% inclusivity and 100%
21
exclusivity in testing of target and non-target bacteria.
22
This method performance, coupled with the simplicity and rapidity of the assay (less than 40 min.),
23
suggested that the method could also serve as a useful tool for identification of presumptive
Salmonella
24
spp. isolates from selective/differential agar plating media. A pre-collaborative study has been
25
completed in which colonies of 113
Salmonella
spp. strains and 37 non-
Salmonella
strains were picked
26
from tryptic soy agar (TSA) and 6 selective/differential agar media [Hektoen enteric agar (HE), xylose
27
lysine deoxycholate agar (XLD), bismuth sulfite agar (BS), brilliant green sulfa agar (BGS), xylose lysine
28
tergitol agar (XLT-4), and double-modified lysine iron agar (DMLIA)] and tested in the ANSR assay. The
29
former 3 media are specified for use in the BAM reference method, while the latter 3 are specified in the
30
MLG method. One hundred twelve
Salmonella
spp. strains produced positive results from all 7 media,
31
2