Journal of Research in Microbes
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Original Research
Occurrence of Listeria monocytogenes in Ready-To-Eat foodstuffs in Windhoek, Namibia Authors: Thuli K. Mogomotsi1 and Kazhila C. Chinsembu. Institution: University of Namibia, Faculty of Science, Department of Biological Sciences, P/Bag 13301, Windhoek, Namibia. 1. P. O. Box 97, Manyana, Botswana. Corresponding author: Kazhila C. Chinsembu.
ABSTRACT: Listeria monocytogenes (Family Corynebacteriaceae; Order Eubacteriales) is a Gram-positive and facultative intracellular pathogen that causes listeriosis. Most cases of human listeriosis are caused by consumption of contaminated ready-to-eat (RTE) foodstuffs purchased from supermarkets and cafeterias. Although L. monocytogenes poses risks for food-borne listeriosis, data on levels of L. monocytogenes in RTE foods in Namibia are non-existent. This scenario makes it difficult to devise preventive regulatory guidelines against possible incidences of human listeriosis. In order to provide baseline data, we microbiologically surveyed for the prevalence of L. monocytogenes in several RTE foods sold in selected retail outlets and university cafeterias of Windhoek, Namibia. The occurrence of L. monocytogenes was statistically higher on hotdogs and polony than other RTE foods such as apples, meat salads, egg salads, and coleslaw. But the loads of L. monocytogenes were too low to warrant fears of possible outbreaks of human listeriosis. However, to help build consumer confidence, we recommend that workers who sell RTE foods should be given training in food hygiene and safety management procedures based on Hazard Analysis and Critical Control Point (HACCP) principles.
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Article Citation: Thuli K. Mogomotsi and Kazhila C. Chinsembu. Occurrence of Listeria monocytogenes in Ready-To-Eat foodstuffs in Windhoek, Namibia. Journal of Research in Microbes (2012) 1(2): 065-070
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Listeria monocytogenes, contamination, ready-to-eat foods, Windhoek, Namibia.
Dates: Received: 23 Jul 2012
Accepted: 10 Aug 2012
Published: 22 Sep 2012
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Mogomotsi and Chinsembu,2012 In Namibia, people enjoy a diet consisting of
INTRODUCTION Listeria Corynebacteriaceae;
monocytogenes Order
(Family
Eubacteriales)
is
large quantities of beef, pork, lamb, chicken, and sea
a
foods such as fish. Most of these ‘meats’ are
Gram-positive, motile, non-spore forming, facultative
incorporated in RTE foods that are sold by retail outlets
intracellular bacterial pathogen that causes disease in
such as official supermarkets, open markets, and
mammals and birds (Gilmour et al., 2010). It was
cafeterias. Although L. monocytogenes poses risks for
recognized as a human pathogen in 1929 (Gray and
food-borne
Killinger, 1966). L. monocytogenes causes an infection
L. monocytogenes in RTE foods in Namibia are
called listeriosis which is characterized by clinical
non-existent. This scenario makes it difficult to devise
symptoms such as gastrointestinal disorders (non-bloody
preventive
diarrhea, nausea, and vomiting), influenza-like sickness
incidences of human listeriosis. In order to provide
(high fever, headache, and myalgia), and serious
baseline data, we microbiologically surveyed for the
septicemia and meningitis (Indrawattana et al., 2011).
prevalence of L. monocytogenes in several RTE foods
Persons susceptible to symptomatic listeriosis include the
sold in selected retail outlets of Windhoek, Namibia.
list eriosis,
regulatory
data
guidelines
on
levels
against
of
possible
elderly, infants, pregnant women, and individuals with impaired immune systems due to HIV/AIDS, major
MATERIALS AND METHODS
surgery, malnutrition, gastric acidity, and lack of
Sampling
physical fitness (Swaminathan and Gerner-Smidt, 2007).
Purposive sampling was carried out between
L. monocytogenes is ubiquitous in nature and has
March and July 2010. This sampling method was chosen
been isolated from soil, dust, water and sewage (Weis
because
and Seeliger, 1975). The vast majority of human
contaminated with L. monocytogenes were chosen so as
listeriosis cases are foodborne and the most commonly
to save time and money. Sampling was done on monthly
implicated sources of infection are ready-to-eat (RTE)
basis at supermarkets in Windhoek’s city centre and
food products such as meat, dairy, seafood, and fresh
Eros, and cafeterias at the University of Namibia
produce that are contaminated with L. monocytogenes
(UNAM). The sampled RTE foodstuffs included apples,
during
hotdogs, coleslaw, egg salads, meat salads, and polony.
processing
(Gilmour
et
al.,
2010).
only
sites
that
were
suspected
to
be
L. monocytogenes may be introduced into RTE foods and food processing facilities due to cross-contamination with environmental sources or from the faeces of food production animals such as poultry, sheep, and cattle (Esteban et al., 2009). L. monocytogenes persists in food processing facilities for a much longer period of time (up to >10 years) because it has the ability to form biofilms that promote attachment to food processing surfaces, and can grow at wide-ranging temperatures (0.4 to 45°C) and pH values ranging 4 to 9.6 (Gilmour et al., 2010). Thirteen serotypes of L. monocytogenes are known but most human infections (>90%) are caused by the three serotypes 1/2a, 1/2b, and 4b (Miya et al., 2010). 066
Figure 1: Listeria monocytogenes colonies on selective media Journal of Research in Microbes (2012) 1(2): 065-070
Mogomotsi and Chinsembu,2012 Mean CFUs of Listeria monocytogenes (± SD) Mean CFUs of Listeria monocytogenes (±SD)
Mean CFUs monocytogenes Mean CFUsofofListeria Listeria monocytogenes ( (±SD) SD) 14
14
12
12
10
10
8
C F U s
CFUs
CFUs
C F U s
6
8 6
4
4
2
2
0
0 Apples
Hotdog
Cafeteria 1
Cafeteria 2
Cafeteria 3
Figure 2: Prevalence of Listeria monocytogenes on apples and hotdogs at UNAM
Figure 3: Prevalence of Listeria monocytogenes on food at different cafeterias at UNAM
Names of RTE food outlets are concealed for ethical
allowed to cool to about 50oC and 2 vials of supplement
reasons.
1 and two vials of supplement 2 were added. The agar
Preparation of L. monocytogenes enrichment broth
was then poured into plates.
An amount of 500 ml of distilled water was
Microbiological analysis of the food samples
placed in a conical flask and 27.2 g of Oxoid
An amount of 9 ml of the L. monocytogenes
L. monocytogenes enrichment broth was added. Then the
enrichment broth was poured into test tubes. For
contents were autoclaved for 15 minutes under a
enrichment, the food samples were crushed into smaller
o
temperature of 121 C and a pressure of 121 Pascals. The o
pieces and 1 g of each of the samples were added into the
broth was allowed to cool around 50 C and 1 vial of
different test tubes containing 9 ml enrichment broth.
supplement 143 was added. The broth contents included
These test tubes were then incubated for 48 hours.
pancreatic digest of casein, proteose peptone, beef
A spreading technique was used to plate 1 ml from each
extract, yeast
extract, sodium chloride, disodium
of the enrichment broth test tubes onto RAPID Listeria
phosphate, monopotassium phosphate, esculin, nalidixic
agar plates. The plates were incubated for 48 hours. After
acid and acriflavine. Some of these components are
the incubation period, the appearance of blue colonies
selective only for L. monocytogenes. For example,
was indicative of L. monocytogenes (Figure 1).
pancreatic digest of casein is a source of amino acids and
white colony observed was regarded as a negative result.
supports growth of L. monocytogenes because the genus
Colony forming units (CFU) of L. monocytogenes were
is composed of fastidious microorganisms. Acriflavine
determined for each of the sampled foodstuffs.
Any
prevents the growth of fungi while esculin is a differential component for organisms that hydrolyze
RESULTS
esculin to react with ferric citrate. L. monocytogenes
The prevalence of L. monocytogenes on apples
does hydrolyze esculin.
and hotdogs at UNAM is shown in Figure 2. The results
Preparation of L. monocytogenes selective agar
showed that apples, with a mean CFU of 6.27 ± 0.46
An
a mo u nt
of
71. 5g
of
R AP I D
(SD)
were
less
significantly
contaminated
by
L. monocytogenes selective agar was placed in 1 litre
L. monocytogenes than hotdogs, with a mean CFU of
conical flask. Then 950 ml of distilled water was added.
10.87 ± 0.79 (SD) (ANOVA F-value 25.15, p < 0.000;
The contents were allowed to dissolve completely and
df = 1, 28).
later sterilized by autoclaving. The contents were then Journal of Research in Microbes (2012) 1(2): 065-070
067
Mogomotsi and Chinsembu,2012 Mean CFUsofofListeria Listeria monocytogenes (± (±SD) SD) Mean CFUs monocytogenes
Mean CFUs Listeriamonocytogenes monocytogenes(±(±SD) Mean CFUs of of Listeria SD)
18
25
16 14
20
12
C F 10 U 8 s 6
CFUs
CFUs
C F U s
4
15
10
5
2 0
0
Cafeteria 1/Hotdog
Cafeteria 1/Apples
Cafeteria 2/Hotdog
Cafeteria 2/Apples
Cafeteria 3/Hotdog
Figure 4 presents the prevalence of Listeria monocytogenes on food types at the three cafeterias at UNAM
analysis
of
variance
and
Scheff’s
post-hoc tests showed diminishing significant differences in
Coleslaw
Egg salad
Meat salad
Polony
Figure 5: Prevalence of Listeria monocytogenes on RTE foods at a retail shop in central Windhoek
In terms of the three cafeterias at UNAM, univariate
Apples
Cafeteria 3/Apples
egg salads (10.40 ± 1.75), apples (9.80 ± 1.50) and coleslaw (9.20 ± 1.15) (ANOVA F-value = 17.80, p < 0.000; df= 4, 20).
the mean CFUs (± SD) of L. monocytogenes:
cafeteria 1(10.20 ± 1.28), cafeteria 2(8.10 ± 0.96), and
DISCUSSION
cafeteria 3(7.40 ± 0.83) (ANOVA F-value = 4.486,
Although the global incidence of foodborne
p < 0.022; df = 2, 24). Figure 3 shows the occurrence of
illnesses is increasing (Nguz, 2007), the loads of
L. monocytogenes at the three cafeterias at UNAM.
L. monocytogenes present in the sampled RTE foods in
Figure
of
Windhoek, Namibia, did not pose any public health risks
L. monocytogenes on food types at the three cafeterias at
for listeriosis outbreaks because they were below the
UNAM. Univariate analysis of variance showed that the
most probable number of 107/g (Miya et al., 2010).
interaction between the two factors (food type*cafeteria)
Notwithstanding, hotdogs had more L. monocytogenes
was
contamination than apples (Figure 2). This is possibly
not
4
presents
significant ly
the
prevalence
different
(ANOVA
F-value = 2.176, p > 0.135; df = 2, 24).
because hotdogs contain meat that may have been
In some RTE foods sold at a retail shop in central
cross-contaminated by L. monocytogenes through contact
Windhoek, ANOVA and Scheffe’s post-hoc tests showed
with animal faeces or during meat processing. In this
that
light,
the
prevalence
of
L.
monocytogenes
was
it
may
be
plausible
to
suggest
that
significantly higher in polony with a mean of 22.20
L. monocytogenes is less frequent on fruits but more
CFUs (± 1.16 SD) than in egg salads (13.80 ± 0.58),
frequent on meat products.
apples (13.60 ± 0.81), meat salads (10.40 ± 0.81), and
At UNAM, the presence of L. monocytogenes
coleslaw (9.60 ± 1.86) (F-value = 19.30, p < 0.000;
differed from cafeteria to cafeteria (Figure 3). This
df = 4, 20). These results are plotted in Figure 5.
variation can be explained by differences in cleaning
The pattern of prevalence of L. monocytogenes
regimes, food sources, handling, processing, or storage
on RTE foods in a retail shop in Eros (Figure 6), an
protocols. However, it is worth noting that because
eastern suburb of Windhoek, was similar to that found in
L. monocytogenes is ubiquitous in the environment, its
central Windhoek. The mean number of CFUs (± SD)
presence in the cafeterias does not automatically translate
was significantly higher for polony (22.00 ± 1.00) than
into a risk for disease. Hence, the cafeterias should not
the other RTE foods such as meat salads (12.40 ± 0.51),
be stigmatized as being unhygienic.
068
Journal of Research in Microbes (2012) 1(2): 065-070
Mogomotsi and Chinsembu, 2012 In order to allay fears about L. monocytogenes
Mean CFUs monocytogenes(±(±SD) Mean CFUsof of Listeria Listeria monocytogenes SD)
contamination of RTE foods and help build consumer
25
confidence, we recommend that workers who sell RTE
20
foods should be given training in food hygiene and safety C 15 F U s 10
CFUs
management procedures based on Hazard Analysis and Critical Control Point (HACCP) principles in order to ensure satisfactory microbiological quality of RTE foods.
5
Personnel in RTE food outlets should also be routinely
0 Apples
Coleslaw
Egg salad
Meat salad
Polony
Figure 6: Prevalence of Listeria monocytogenes on RTE foods at a retail shop in Eros, Windhoek Figure 4 showed that the overall prevalence of
tested
for
various
microbial
agents
including
L. monocytogenes. ACKNOWLEDGEMENTS
L. monocytogenes was not simultaneously dependent on
We are thankful to the Department of Biological
the type of food and cafeteria. Thus, the effect of food
Sciences at UNAM for financial and technical support.
type is completely separate from the effect of cafeteria;
We also thank the UNAM Research and Publications
there were additive effects and no statistical interaction.
Office for paying the publication fees.
On corollary to this, we suggest that the results of similar experiments in the future will still be valid even if they
REFERENCES
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Journal of Research in Microbes (2012) 1(2): 065-070
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Mogomotsi and Chinsembu,2012 Miya S, Takahashi H, Ishikawa T, Fujii T and Kimura B. 2010. Risk of Listeria monocytogenes contamination of raw ready-to-eat seafood products available at retail outlets in Japan. Appl. Environ Microbiol. 76(10):3383-3386. Nguz K. 2007. Assessing food safety systems in sub-Saharan countries: an overview of key issues. Food Control 18:131-134. Swaminathan B and Gerner-Smidt P. 2007. The epidemiology of human listeriosis. Microbes Infect 9:1236-1243. Weis J and Seeliger HP.
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