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Journal
of
Appl ied
Bacteriology 1995,
79
360 367
Eff iciency of d i f ferent enr ich m ent and iso lat ion pro ced ures for
the detectio n of Salmonella seroty pes in edib le of fa l
G. A r r o yo and J.A. Arroyo
Departamento de Microbiologia 11 Facultad de Farmacia and 'Departamento de Microbiologia 111 Facultad de
Biologia Universidad Complutense Madrid Spain
5209/01/95: rece ived 24 January 1995, rev ised 5 Apr i l 1995 and accepted 10 Apr i l 1995
G . A R R O Y O
A N D
J . A . A R R O Y O . 1995. Rapid detection systems for Salmonella in foodstuffs are
currently being developed. However, existing standards still call for application of traditional
methods employing pre-enrichment followed by selective enrichment and isolation. The
efficacy of various methods was tested using
264
chicken and lamb organ meats.
Pre-enrichment was carried out in Tryptone Soy Broth (TSB) and enrichment in
Tetrathionate Brilliant Green Broth (TT B) at 37 C, Selenite Broth with Brilliant Green and
Sulphapyridine at 37C and 43 C, and Rappaport-Vassiliadis Broth (RV 10) at 42C. The
isolation media were Brilliant Green Agar (BGA), Deoxycholate Citrate Agar, Hektoen
Enteric Agar (HEA) and Salmonella-Shigella Agar.
no.
6579
and enrichment in
TTB/37 C
followed by isolation in HEA, no longer
recommended
by
that standard, produced the best results. Low percentages of positive
samples and difficulties in detecting
Salmonella
are the result of interference by competing
organisms (Enterobacteriaceae) and the number of salmonellas present after enrichment.
A total of 528 samples (TSB, eggs, lamb liver and chicken liver) were inoculated with
Sal m. enteritidis Sal m. kapemba
and
Salm . virchow
and the preceding experiment was
repeated. All the TSB and egg samples tested positive, but the percentage of positive samples
from the lamb and chicken liver was only 81-92%. Recovery of the salmonellas did not
depend upon the method employed or the serotype inoculated but instead on interference by
competing flora and the numbers of
Salmonella
present in the samples.
Enrichment in RV/42 C followed by isolation on BGA as recommended by I S 0 standard
I N T R O D U C T I O N
Salmonellas are responsible for most cases of food poison-
ing in
the developed world. Foods such as meat, eggs,
poultry and organ meats are common vehicles of salmonel-
losis. For that reason, rapid detection methods based pri-
marily on immunological and genetic characteristics are
under development. However, the standards presently in
force in many countries recommend traditional methods,
which are slower, requiring
a t
least
5
d, though they can be
quite reliable when applied by experienced laboratory tech-
nicians.
A variety of methods are in use, and their success rates
depend upon a number of different factors. They employ
pre-enrichment in buffered peptone water, followed by
Correspondence
t o
:
Dr G.
r royo Jul ian
Romea
9
28003-Madrid
Spain.
selective enrichment in more than one medium, usually
Muller-Kauffmann Tetrathionate Broth and Selenite
Cystine Broth incubated a t 37C or
a t
43 C, and subse-
quently selective isolation on various solid media (Anon.
1981). Van Schotthorst
et a l .
(1987) recommended enrich-
ment
in
Rappaport-Vassiliadis (RV) broth
a t
43 C, and
Beckers
e t
a l . (1987b) proposed replacing the Tetrathionate
method with the RV method
in
the I S 0 standard (Anon.
1990).
The selective effects of the different media are mainly
based on the addition of substances that inhibit the growth
of contaminating microflora and prevent the proliferation of
competing microflora, on the incubation temperature and
the inoculum size employed. Studies on some of these
inhibitors (Arroyo and Arroyo 1995) have often yielded dis-
appointing results. The numbers of competing Entcrobac-
teriaceae and Pseudomonadaceae
in
many natural foodstuffs
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FOR SALMONELLA D E T E C T I O N 301
must be assumed to be higher than the numbers of Salmon-
ella
and those competing organisms are also capable of
withstanding the same inhibitor concentrations as salmon-
ellas during pre-enrichment and isolation, thu s masking th e
presence of salmonellas and giving rise to false positives
(Rhodes and Quesnel 1986).
The addition of stains, e.g. brilliant green, to the pre-
enrichment media combined with raising the incubation
temperature to 43C has produced varying results for
Tetrathionate Broth (Van Schothorst et al. 1977). Arroyo
(1990) reported that temperature affected the growth of
certain Salmonella serotypes, making detection more diffi-
cult.
T he use of antibiotics and chemical therap eutic agents in
the media has also been proposed. Osborne and Stokes
(1955) recommended adding sulphapyridine to enhance the
effectiveness of S elenite Br oth.
The success of the isolation media depends basically
upon the enrichment step employed and the number of
competing organisms that survive that step.
Besides the international standards, the recommen-
dations of the National Reference Center at the Carlos I11
Health Institute in Madrid are usually followed when ana-
lysing food and drink in Spain (Pascual Anderson 1993).
The object of the present study was therefore to evaluate
the efficacy of the recommended enrichment media and the
influence of incubation temperature using Selenite Broth
with added brilliant green and sulphapyridine. T h e efficacy
of four commonly used solid isolation media was also
tested.
T he foodstuffs employed in this study were typical natu-
rally contaminated sources of salmonellas, namely, lamb
organ meats and chicken liver, purchased at markets in
Madrid (Experiment 1). Tryptone Soy Broth (TSB), eggs
and lamb and chicken liver were also artificially contami-
nated with three Salmonella serotypes (Experiment 2).
MATERIAL S AND METHODS
Experiment 1: Naturally contaminated samples
A
total of 264 organ meats purchased retail at 10 markets in
Madrid were examined. The organs used were chicken
livers (78) and lamb organs (186), including liver (46), lung
(46), heart (46), oesophagus (46), and spleen
(2).
The organ
meats were transported to the laboratory in
a
portable
cooler at 4C. Examinations were performed on the day of
purchase.
Twenty-five grams of each sample were weighed out
using sterile instruments under aseptic conditions, added to
225 ml of Trypto ne Soy Broth (T SB), and homogenized in
sterile bags in a Stomacher model 400 Lab Blender for 2
min.
Microbiological examinations
The microbiological examinations employed differed some-
what from standard m ethods.
(a) Pre-enrichme nt was performed in T S B incubated a t
37C for 18 h (Arroyo 1990).
(b) Selective enrichment was performed by transferring
10
ml of the pre-enrichment culture to a flask containing
100 ml of Muller-Kauffmann Tetrath ionate Broth
(T TB ) containing
1
ml
of
a 0.1% (w/v) solution of bril-
liant green, followed by incubation at 37C for 24 h.
Two flasks, each containing 100 ml of Selenite Broth
(SC ) containing 1 ml of a 0.1 (w/v) solution of brilliant
green and sulphapyridine were likewise inoculated with
10
ml of the pre-enrichment hom ogena te; one of the
flasks was incubated at 37 C, the other at 43 C, for 24 h
(Arroyo 1990). Finally,
0.1
ml of pre-enrichment
medium was transferred to 10 ml Rappaport-Vassiliadis
Broth (RV 10) (Vassiliadis 1983) and incuba ted at 42 C
for 24 h.
(c) Selective isolation was performed by streaking the sur-
faces of plates containing Brilliant Green Agar (BGA),
Hektoen Enteric Agar (HEA), Salmonella-Shigella Agar
(SSA) and Deoxycholate Citrate Agar (DCA) with the
T T B and S C selective enrichment media. T he plates
were all incubated at 37C for 24-48 h. T h e RV selective
enrichment medium was inoculated only onto BGA
(Vassiliadis
et al.
1978) and incubated a t 37C for 24-48
h. Colonies suspected of belonging to the genus Salmon-
ella were isolated in test tubes containing Tryptone
Soy
Agar (TS A) incubated at 37C for 24-48 h.
(d) Identification of salmonellas was carried out using mor-
phological, biochemical (H olt 1993) and serological tests.
Strains identified as Salmonella were serotyped at the
National Reference Center at the Carlos I11 Health Insti-
tute in Madrid.
Experiment
2:
Arti fic ially contaminated samples
Pure cultures.
Pure cultures of
Salm . enteritidis Salm .
kapemba
and
Salm. virchow
isolated from lamb and chicken
organ meats were grown on TSA slants for 24 h and then
washed from the slant with sterile physiological saline
solu-
tion. T he suspension was diluted with sterile saline solution
to an optical density of 0.2 at 540 nm measured with a
Bausch and Lomb model Spectronic 20 spectrophotometer.
Serial dilutions were prepared from this stock suspension to
995 The Society for Appl ied Bacter io logy, Journal of Applied Bacteriology 79, 360 367
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362
G . A R R O Y O A N D
J . A .
A R R O Y O
yield the appropriate number of cells. That number was
determined by plating 0.1 ml of the diluted suspension on
TSA plates and incubating at 37C for 24 h.
Table 2 Efticiency of the four enrichment procedures in terms of
the number of isolated serotypes
Enrichment Isolationt
Y
Serotype
No.
I n o c u l a t i o n . Inoculation took place as follows :
(a) Three sets of 44 samples of 225 ml of TSB were each
inoculated with cultures containing 1000 cells of S a l m .
enteritidis Salm.
kapemba
and S a l m .
virchow
respec-
tively. These
132
samples were incubated at
37C
for
18
h. Inoculation of the enrichm ent and isolation media and
detection of salmonellas were then as already described
in Experiment
1
(b)
A
total
of 132
pasteurized eggs purchased at
10
markets
in Madrid were washed in sterile water under aseptic
conditions, cracked and the contents of each egg homog-
enized in 225 ml of TSB. Thre e groups
of
44
of
these
samples were then inoculated with cultures containing
1000 cells of each of the three aforementioned serotypes,
respectively, and incubated at 37C for 18 h. Inoculation
of the enrichment and isolation media and detection of
salmonellas were then as already described in Experi-
ment
1 .
(c) A total of 132 samples of each lamb liver and chicken
liver, purchased at 10 markets in Madrid, were homoge-
nized in a Stomacher model 400 Lab Blender (each
sample consisting of 25 g of organ meat in 225 ml of
TSB). Three groups
of 44
samples
of
each
of
these two
organ meats were inoculated with cultures containing
1000
cells of each
of
the aforementioned serotypes,
respectively, and incubated at
37C for 18
h. Inoculation
Table
1
Number and percentage of the different Salmonella
serotypes detected in the 260 processed and confirmed strains
TTB/37 C*
SC137 C
SC/43 C
RV142 C
Total
47 18.07 Salm. enteritidis
t yphimurium
virchow
worthington
kapemba
give
n antis
cubana
Salm. autoagglut.
96 36.92 enteritidis
typhimurium
virchow
worthington
kapemba
give
havana
arizona
58 22.30 enteritidis
virchow
worthington
kapemba
Salm.
autoagglut.
59 22.69 enteritidis
typhimurium
virchow
morthington
kapemba
infantis
brandenburg
anatum
agona
260 99.98
10
7
14
4
3
3
3
1
2
4
7
40
25
5
4
4
7
6
37
12
1
2
5
8
19
4
3
12
3
2
3
260
Serotype No.
Salm . enteritidis
typhimurium
virchow
worthington
infantis
kapemba
give
brandenburg
havana
anatum
arizona
agona
cubana
Salm.
autoagglutinable
14 serotypes
25
22
110
45
15
12
7
3
4
2
7
3
1
4
260
9.61
8.46
42.30
17.30
5.76
4.61
2.69
1.15
1.53
0.76
2.69
1.15
0.38
1.53
99.92
* Enrichment broths and incubation temperature : TTB, Tetra-
thionate Brilliant Green Broth, 37C; SC, Selenite Broth (with
brilliant green and sulphapyridine), 37C and 43C; RV,
Rappaport-Vassiliadis Broth (RV lo), 42C.
t
Numb er and percentage of positive isolation.
of the enrichment and isolation media and detection of
salmonellas were as already described in Exp erim ent 1 .
R E S U L T S
Exper iment1
Of the 264 lamb and chicken organ meats examined, 83
(31.43%) tested positive for Salmonel la . A total of 260
strains isolated were confirmed to be on e of
14
serotypes of
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S A L M O N E L L A
D E T E C T I O N
363
Table
3
Efficiency of enrichment and
isolation procedures in terms of positive
strains of Salmonella detected
Isolation procedu re7
Enrichment procedure BGA DCA HEA
SSA
No. ( )
TTB/37 C* 7
(2.69)
SC/37 C 14
(5.38)
SC/43 C 13
(5)
RV/42 C 59
(22.69)
No. 93
( /.I (35.76)
15
(5.76)
27
(10.38)
15
(5.76)
57
(21.9)
15 10
(5.76) (3.84)
28 27
(10.76) (10.38)
12 18
(4.61) (6.92)
55 55
(21.13) (21.13)
47
(18.05)
96
(36.9)
58
(22.29)
59
(22.69)
260
(99.93)
Enrichment broths and incubation temperature : TTB, Tetrathionate Brilliant Green
Broth, 37C; SC, Selenite Broth (with brilliant green and sulphapyridine), 37C and 43C;
RV, Rappaport-Vassiliadis Broth (RV lo), 42C.
t
Isolation media: BGA, Brilliant Green Agar; DCA, Deoxycholate Citrate Agar; HEA,
Hektoen Enteric Agar; SSA Salmonella-Shigella Agar.
No.,
Number of strains detected.
Percentage
of
positive isolations given in paren theses.
Table
4 Serotypes detected according to the enrichment and isolation procedures used
Isolation procedure
Enrichment
procedure BGA DCA HEA
SSA
TTB/37 C
Sal m typhimurium Sal m enteritidis
virchow typhimurium
virchow
worthington
infantis
give
Salm
autoagglut.
SC/37 C
SC/43 C
RV/42 C
enteritidis
virchow
worthington
give
arizona
virchow
worthington
Salm autoagglut.
enteritidis
typhimurium
virchow
worthington
kapemba
infantis
brandenburg
anatum
agona
virchow
worthington
arizona
enteritidis
virchow
worthington
Salm enteritidis Sa lm typhimurium
typhimurium virchow
virchow kapemba
worthington infantis
in antis give
give
cubana
Salm
autoagglut.
virchow
worthington
kapemba
typhimurium
virchow
worthington
kapemba
enteritidis
virchow
worthington
kapemba
give
havana
arizona
enteritidis
virchow
worthington
For abbreviations see Table 3.
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Bacteriology,
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364
G .
A R R O Y O A N D J . A . A R R O Y O
Salmonella. The distribution of serotypes, including
autoagglutinable strains, appears in T able 1 .
Table
2
shows that
96 Salmonella
strains
(36.92%)
were
detected using the
SC
enrichment medium incubated at
37C. Incubation of this same medium at 43C resulted in
only
58 Salmonella
strains
(22.30%).
Enrichment in TTB
incubated at
37C
yielded
47
positive strains
(18.07%),
while enrichment in RV 10 incubated at 42C yielded 59
(22.69%) positive isolations. Table 2 also lists the different
serotypes detected using each of the media. TTB/37 C and
RV/42 C
produced nine
of
the
14
serotypes detected and
SC/37 C
produced eight.
SC/43 C
yielded the fewest
serotypes (5).
Salmonella virchow
was the most abundant serotype and
was detected with
all
four of the enrichment methods used.
On the other hand, not all the minor serotypes present were
detected by all the different methods.
Table
3
gives the number and percentage of positive iso-
lations on the solid media for each of the selective enrich-
ment media employed. BGA yielded
93
positive isolations
(35.76%). Detection rates differed substantially, with TTB/
37C
yielding only seven positive isolations
(2.69%)
and
RV/42 C
yielding
59 (22.69%).
The results obtained using
enrichment in SC/37 C and SC/43 C were quite similar, 14
(538%)
and
13 ( 5 )
positive isolations, respectively. The
results obtained using
DCA,
HEA and SSA were more
uniform. T h e best results, 27 (10.38%) and 28 (10.76%)
positive isolations, were achieved following enrichment in
SC/37 C; results were not as good when enrichment was
carried out in
SC/43 C.
In terms of the variety
of
serotypes detected (Table
4),
the best results were obtained by BGA after enrichment in
RV/42 C (9
serotypes) and
HEA
after enrichment in TT B /
37C (8
serotypes). On the whole, interference by com-
peting organisms on the isolation media was lower
following enrichment in T T B and RV.
Experlment
2
Table
5
presents the recovery of
Sal m. enteritidis Salm.
kapemba
and
Salm. virchow
in the different samples. The
results show that direct inoculation with
1000
cells in T S B
yielded a recovery rate of 100% positive samples, irrespec-
tive
of
the enrichment and isolation method used and the
serotype inoculated.
Inoculation of a foodstuff devoid
or
with negligible levels
of contamination, namely, pasteurized eggs, with 1000 cells
and homogenization with T S B also produced a
100%
recovery rate.
As
in the preceding case, the recovery rate
was the sam e for all the serotypes inoculated.
Table 5 Recovery of Salmonella
Sample Enrichment procedure enteritidis Salm. kapemba and
Salm.
No. TTB/37OC
sc-37 ~ S C / ~ ~ O C
V/42OC virchow using different procedures
of
Inocula Nature
enrichment
Salm. enteritidis TSB
Eggs
Lamb liver
Chicken liver
Salm.
kapemba
TSB
Eggs
Lamb liver
Chicken liver
Salm. virchow TSB
Eggs
Lamb liver
Chicken liver
4 4 4 4
(100)
4 4 4 4
(loo)
44 38
44 36
4 4 4 4
(loo)
4 4 4 4
(100)
44 38
44 38
4 4 4 4
(100)
4 4 4 4
(100)
44 39
44 39
(86.4)
(81.8)
(86.4)
(86.4)
(88.6)
(88.6)
44
44
39
(88.6)
38
(86.4)
44
(100)
44
(100)
40
(90.9)
39
(88.6)
44
(loo)
44
(100)
40
(90.9)
41
(92.3)
(100)
(100)
44
(100)
44
(loo)
37
(84.1)
36
(81.8)
44
(100)
44
(100)
37
(84.1)
38
(86.4)
44
(loo)
44
(loo)
38
(86.4)
36
(81.8)
44
(100)
44
(1W
40
(90.9)
37
(84.1)
44
( 1 0 )
44
( loo )
39
(88.6)
41
(92.3)
44
W
44
41
(92.3)
39
(88.6)
Percentage
of
positive samples in parentheses.
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D E T E C T I O N 365
In contrast, inoculation of the lamb and chicken livers,
foodstuffs with normally high levels of contamination
(Enterobacteriaceae counts of up to lo7), with 1000 cells in
TSB homogenate yielded more uneven results, ranging
from
a
low of
81.81%
positive isolations to a high of
92.27% positive isolations and a slight difference in favour of
RV/42 C. SC/43 C
produced the lowest recovery rates.
Again, the results were independent of the salmonella
serotype inoculated.
The results obtained for the selective isolation media
tested in turn depended upon the enrichment broth
employed. On the whole, interference by competing
organisms was high, particularly on the BGA, making
detection of
Salmonella
difficult even though the samples
had been inoculated with that bacterium.
D I S C U S S I O N
The success of enrichment and isolation media is based on
the presence of inhibitors intended to act against Gram-
positive contaminating microflora and Gram-negative com-
petitive flora, normally Enterobacteriaceae.
A
study of 12
inhibitors
a t
similar concentrations (Arroyo and Arroyo
1995) demonstrated that the level of inhibition on salmon-
ellas was the same as on the othe r Enterobacteriaceae exam-
ined and accordingly that inhibition of the multiplication of
such organisms during enrichment was difficult. Direct
counts of total Enterobacteriaceae in lamb and chicken
organ meats on Violet-Red-Bile-Glucose-Agar (VRB G)
reached up to lo7 cfu
g- '
(Arroyo and Arroyo 1995). T h e
high initial levels of Enterobacteriaceae and their further
growth in the enrichment media masked the presence of
Salmonella
giving rise to false positives (Watson and
Walker 1978; Rhodes and Quesnel 1986). Salmonellas
cannot begin to multiply during enrichment until the
number of competitors has fallen (Van Schothorst and
Renaud 1983; Rhodes and Quesnel 1986). This decrease is
brought about by a fall in pH and depends upon the food-
stuff in question, bacterial metabolism, and the presence of
large numbers of Gram-positive bacteria, including lactic
acid bacteria (Van Schothorst and Renaud
1985).
In the
samples examined by the authors, there were rather high
numbers of Enterobacteriaceae with low numbers of Gram-
positive organisms, and t he p H did not fall below 6.2
(unpublished data).
The results for the enrichment media in Experiment
1
varied. The greatest variety of different
Salmonella
serotypes were detected using RV 10 (Vassiliadis et al.
1981
;
Vassiliadis 1983) and the largest number of positive
isolations using SC/37 C, though it should be noted that
most of these belonged to the two most ab undan t serotypes,
Salm. virchow
and
Salm. worthington
and that detection of
the minor serotypes using this latter medium was more dif-
ficult. Fagerberg and Avens (1976) reported that certain
serotypes were easier to detect in certain media than in
others.
TTB, which Beckers
et al. (1987a,
b) recommended
replacing with RV in the I S 0 standard (Anon. 1990), pro-
duced the fewest positive isolations, but the number of dif-
ferent serotypes detected was higher than using SC/37 C.
This suggests that certain inhibitors may affect the growth
and multiplication of some of the minor serotypes present.
However, according to Van Schothorst et al. (1977), inhibi-
tion depends less upon the serotype than upon the sensi-
tivity of certain cells to the inhibitor. Patil and Parhad
(1986) reported that T T B was the best enrichment medium
when E.
coli
and other lactose-positive Enterobacteriaceae
were the predominant competing organisms, whereas
SC
yielded better results when organisms of the genus Proteus
predominated.
Raising the incubation temperature may increase the
number of positive isolations (Carlson and Snoeyenbos
1972). However, tetrathionate with brilliant green at 43C
is toxic to many salmonellas (Harvey and Price
1979;
Arroyo
1990).
Both the number of positive isolations and
the variety of serotypes detected decreased using SC/43 C.
Studies with pure cultures have indicated that certain Sal-
monella serotypes cannot multiply at high incubation tem-
peratures (Carlson and Snoeyenbos 1974; Van Schothorst
et al. 1977). On the other hand, survival of most of the
Enterobacteriaceae at 43C and at 44C (e.g. E.
coli
whose
presence as a faecal coliform is detected in lactose broth
incubated at
44 C),
made isolation of salmonellas extremely
difficult. Van Schothorst and Renaud (1983) reported that
isolation of salmonellas was virtually impossible when the
number of lactose-positive Enterobacteriaceae was
10'
g '
or m l-' , hence raising the temperature would appear to be
unnecessary when using th e SC medium.
Isolation on BGA following enrichment in SC/37 C and
SC/43 C
was quite difficult because of the large numbers
of competing bacteria present.
For
the salmonellas
to
be
detectable on that medium, their concentration in the
enrichment m edium after incubation m ust reach a level of
lo3
m l -
'
(Van Leusden
et al.
1982);
at lower levels they
remain undetectable, even if the level of the competing
flora drops to below lo 3 ml
.
Detection on BGA is facilitated by greater inhibition of
competing organisms in RV
10
(Vassiliadis
1983).
Com-
peting bacteria belong mainly to the genera
Proteus Morga-
nella and Shigella. The presence of Yersinia is important,
because the Rappaport-Wauters medium is used for enrich-
ment of that genus. T ha t medium is similar to RV, and the
colonies of that genus on BGA do not differ in appearance
from Salmonella colonies.
Isolation o n HEA and DCA after enrichment in T T B
yielded the most positive isolations, and anaerobic culture is
R ? 1995 The Society for Applied Bacteriology.
Journal
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79,
360-367
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366 G . A R R O Y O A N D J . A . A R R O Y O
not required when using DCA (Edgar and Soar 1979).
However, contradicting the findings of Moriiiigo
e t
al.
(1989),
these media did not inhibit the growth of
Pseudo-
monas aeruginosa
or other Pseudomonadaceae and Vibriona-
ceae. Based on these results, RV would appear to be the
most appropriate enrichment medium, but complete rejec-
tion of T T B would appear to be premature, since that
medium yielded the greatest variety in the number of
serotypes and lower interference by competing organisms
than SC.
Th e results of Experiment 2 corroborated the results of
Experiment 1 . In the TSB and commercial pasteurized egg
samples, in which there was no interference by competing
organisms,
100%
of the samples tested positive, irrespective
of the serotype inoculated and the method employed, even
when lower concentrations of inocula were used (results not
presented here). Previously, inoculation with small
numbers of
Salm. montevideo
and
Salm. heidelberg
yielded
excellent recoveries using Selenite Broth, whereas the con-
centration of the inoculum had to be increased to 500 cells
when using Tetrathio nate B roth (Bailey
e t
al.
1981).
In the
present experiment the pre-enrichment broth was inocu-
lated to facilitate the recovery of salmonellas in the T T B
(Taylor and Silliker
1962).
The level of competing organisms was high in the lamb
and chicken liver samples, and the percentage of positive
isolations was lower, though the results did not vary much
with the method used and the serotype inoculated. Accord-
ingly, successful detection of salmonellas in those organ
meats was dependent upon interference by competitors.
The similar behaviour exhibited by
Salm. k a p emb a
a minor
serotype in Experiment
1,
indicates that it is able to tolerate
the same inhibitor concentrations as
S a l m. en t er i t i d i s
and
S a l m . virchow
and provides further confirmation that sal-
monella concentrations must be
l o3
ml - after enrichment.
Difficulties in detecting salmonellas are heightened when
there are cross-reactions with other Enterobacteriaceae,
such as
Proteus Morganel la
and
Citrobacter freundii.
Increasing interest in developing fast methods of analysis
and future application of those methods may help to miti-
gate these difficulties.
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