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Prevalence of Bacillus cereus and Associated Risk Factors in Chinese-Style Fried

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DOI: 10.1089/fpd.2011.0969 · Source: PubMed

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FOODBORNE PATHOGENS AND DISEASE
Volume 9, Number 2, 2012
ª Mary Ann Liebert, Inc.
DOI: 10.1089/fpd.2011.0969

Prevalence of Bacillus cereus and Associated

Risk Factors in Chinese-Style Fried Rice
Available in the City of Colombo, Sri Lanka

Manosha Lakmali Perera1 and Gerard Ranjan Ranasinghe 2

Abstract
The present study aimed to assess the prevalence of Bacillus cereus and its associated risk factors in Chinese-style
fried rice available in Colombo city. In 200 samples of fried rice the prevalence of B. cereus was 56%. The
prevalence by variety of fried rice was chicken (20.0%), vegetable (18.0%), seafood (10.0%), egg (5.0%), mixed
(2.0%), and beef (1.0%). Of analyzed samples, 28 (14%) had colony counts > 106 colony forming units per gram
(cfu/g), the infectious dose for B. cereus food borne outbreaks. Occurrence of > 106 cfu/g of B. cereus were
associated with storage of boiled rice at room temperature ( p = 0.030), > 4 hours of storage at room temperature
( p = 0.042) and cooking frequency of more than once per dining session ( p = 0.017). The type of rice and the
quantity boiled per day were not independent risk factors for high B. cereus counts. Majority of B. cereus isolates
(53.7%) in this study were not typable. The serotypes observed included H15 (14.3%), H19 (14.3%), and H20
(10.7%). These serotypes are known to be associated with both emetic and diarrheal syndromes. All isolates were
sensitive to ciprofloxacin (100%), erythromycin (100%), gentamicin (100%), chloramphenicol (100%), and ami-
kacin (100%) whereas 100% resistance was observed for penicillin with minimal inhibitory concentration range
of 32–256 lg/mL.

Introduction food poisoning, this organism is implicated in wound infec-

tions, bacteremia, endocarditis, and severe eye infections such

B acillus cereus is ubiquitous in the environment and is

able to survive and proliferate in soil, water, and vege-
tation. The highly resistant spores can survive heating, dry-
as keratitis, endophthalmitis, and panophthalmitis. Other less
frequent infections include postoperative meningitis, gingi-
vitis, and periodontitis (Kotiranta et al., 2000).
ing, radiation, freezing, and pasteurization (Kotiranta et al., Cooked rice is usually associated with the emetic syndrome
2000). It is able to contaminate wide variety of raw and pro- of food poisoning. B. cereus has been documented worldwide
cessed food items such as cereals, dairy products, meat, fish, in outbreaks associated with cooked rice (Little et al., 2002;
vegetables, herbs, spices, cooked rice, soups, and desserts Lake et al., 2004). Since 1971, contaminated Chinese-style fried
(Granum, 2001). rice has been responsible for more than 40 incidents of emetic
B. cereus is responsible for two distinct types of food poi- type food poisoning in the United Kingdom (Taylor and
soning. The diarrheal syndrome is caused by enterotoxins Gilbert, 1975). In a risk assessment for B. cereus rice associated
released in the small intestine after ingestion of contaminated food poisoning outbreaks by Notermans and Batt (1998),
food. The emetic syndrome is caused by a heat stable peptide ‘‘Chinese-style food’’ was the commonest food vehicle in
preformed in food (Vilas-Boas et al., 2007). The emetic syn- Canada (1985–1986, 17/39 outbreaks), the United States
drome is rapid in onset, occurring within 1–5 hours of in- (1988–1992, 12/21 outbreaks), and the Netherlands (1992–
gestion and causes nausea, vomiting, and abdominal cramps. 1994, 17/40 outbreaks). In addition, in England and Wales 12
In contrast, the diarrheal syndrome is delayed in onset, oc- outbreaks occurred after consumption of Chinese or Indian
curring after 8–16 hours and is characterized by watery di- style takeaways during 1992 to 2000 (Little et al., 2002). Also,
arrhea, cramps, and abdominal pain. Usually for both cooked rice associated outbreaks have been reported from
syndromes symptoms do not persist beyond 24 hours and the Japan (Shinagawa et al., 1985; Shiota et al., 2010), Singapore
illness is mild and self limiting (Lake et al., 2004). In addition to (Tay et al., 1982), Scotland (Dickson and Morgan, 1995),

1
Molecular Biology Section, Medical Research Institute, Colombo, Sri Lanka.
2
Microbiology Division, Department of Pathology, Provincial General Hospital, Badulla, Sri Lanka.

125
126 PERERA AND RANASINGHE

Finland (Raevuori et al., 1976), India (Granum and Baird- (Oxoid Ltd) using a Stomacher 400 Circulator (Seward Ltd).
Parker, 2000), and New Zealand (Lake et al., 2004). Con- With this initial 1:10 (10 - 1) dilution, serial dilutions from 10 - 2
sequently, B. cereus has gained prominence as a priority to 10 - 7 were prepared in 0.1% peptone water (Bridson, 2006).
pathogen for enteric diseases task force work plan 2009–2010
drawn up by the WHO (WHO, 2009). Presumptive B. cereus count
From 2005 to 2010, 7620 food poisoning cases have been
The medium used for isolation of B. cereus was prepared
reported in Sri Lanka out of which 600 have occurred in Co-
with B. cereus Selective Agar Base (CM617), B. cereus Selective
lombo district (Ministry of Healthcare and Nutrition Sri
Supplement/Polymyxin B (SR99), and Egg Yolk Emulsion
Lanka, 2005). Clinical symptoms of many of these cases were
(SR47) (Oxoid Ltd). Each batch of prepared medium was
similar to B. cereus emetic type food poisoning. However,
quality controlled using B. cereus ATCC 10876 as the positive
most were not investigated due to the mild and self limiting
control and Bacillus subtilis ATCC 6633 as the negative control.
nature of the illness. Also, the majority of cases were sporadic
Duplicate plates of B. cereus selective agar were inoculated
and not associated with outbreaks. In the recent past, B. cereus
with 0.1 mL of 10 - 1 to 10 - 7 dilutions. Each inoculum was
counts ranging from 1.0 · 103 to 8.4 · 108 have been reported
evenly spread on the surface of the medium with a sterile
in cooked rice and fried rice (Unpublished data from Food
glass spreading rod. Plates were incubated at 35C for 24
and Water Laboratory, Medical Research Institute, Sri Lanka).
hours and cursorily examined for Bacillus colonies. Thereafter,
Sri Lanka is a popular tourist destination and local cuisine
the plates were left at room temperature for 24 hours to allow
which includes fried rice is very much in demand. Therefore,
colonies to develop typical morphology of B. cereus (Bridson,
ensuring food safety is a prime concern.
2006). The characteristic colonies were crenated, *5 mm in
The predominance of B. cereus outbreaks linked to Chinese-
diameter, peacock blue, and surrounded by a conspicuous
style fried rice is associated with the common practice of
egg yolk precipitate of the same color. All plates from the
boiling large quantities of rice and storing at room tempera-
dilution series was examined with the naked eye and one set
ture before frying. This is done to avoid refrigeration which
containing an estimated 15–150 typical colonies was selected
makes the grains to become sticky and clump together. Raw
for manual counting (Sri Lanka Standard: 516: Part 8, 1983;
rice frequently contains heat resistant spores that survive
USFDA, 2001). This number expressed as cfu/g fried rice was
boiling. When boiled rice is stored at room temperature for
the presumptive colony count.
prolonged periods, these spores germinate, proliferate, and
produce heat stable emetic toxin (Little et al., 2002). Vegetative
Confirmed B. cereus count
cell growth is stimulated by the nutrient content of eggs and
meat added to the rice (Morita and Woodburn, 1977). The From each positive sample, five peacock blue, lecithinase-
organism is thus able to achieve the infectious dose of counts positive colonies were picked and transferred to nutrient agar
exceeding 106 colony forming units per gram (cfu/g) of food slants. These were incubated for 24 hours at 30C and used for
(USFDA, 2001). performing morphological and biochemical confirmatory
This study was aimed at assessing the prevalence of B. cereus tests. With Gram staining (212539, BD Difco BBL Stains),
in Chinese-style fried rice available in Colombo city, evaluating B. cereus appears as large Gram-positive bacilli in chains with
selected risk factors for significant counts ( > 106 cfu/g), ser- ellipsoidal, central, or paracentral spores which do not swell
otyping, and antibiotic sensitivity testing of isolates. the sporangium. The rapid confirmatory staining procedure
(Holbrook and Anderson, 1980) using malachite green (90903;
Materials and Methods Fluka), Sudan Black (199664; Sigma-Aldrich), and aqueous
0.5% w/v safranin (212532; BD Difco BBL Stains) showed the
Sampling method characteristic morphology of pale green spores and black lipid
During a period of 1 year, 200 samples of Chinese-style globules in red stained cytoplasm.
fried rice were collected by systematic random cluster sampling The principle biochemical tests used for identification of B.
of eateries within Colombo city. The samples were categorized cereus group were positive catalase (H3410; Sigma-Aldrich),
according to the main ingredient added to fried rice during positive Voges-Proskauer test (39484; Fluka), reduction of
preparation. These were vegetables, chicken, seafood, eggs, nitrate (72548; Fluka), hydrolysis of tyrosine (T3754; Sigma-
beef or a mixture of all. The distribution of 200 samples included Aldrich), growth in 0.001% w/v lysozyme broth (62970;
70 (35%) vegetable fried rice, 59 (29.5%) chicken fried rice, 36 Fluka), liquefaction of gelatin (48723; Fluka), and production
(18%) seafood fried rice, 18 (9%) egg fried rice, 10 (5%) beef fried of acid from glucose (49159; Fluka), xylose (X1500; Sigma-
rice, and 7 (3.5%) mixed fried rice. Samples were transported Aldrich), and arabinose (10850; Fluka). These biochemical
without freezing and analysis commenced within 3 hours. tests were chosen from standard references for isolation and
A structured interviewer administered questionnaire was enumeration of B. cereus from food (Kramer et al., 1982; Sri
designed to elicit information on practices related to prepa- Lanka Standard: 516: Part 8, 1983; USFDA, 2001). All bio-
ration and storage of fried rice. The questionnaire was pre- chemical media were quality controlled for performance with
tested on an appropriate sample population. Data processing recommended positive and negative controls. For each batch
and analysis was conducted using SPSS 13 statistical software of testing B. cereus ATCC 10876 and B. subtilis ATCC 6633
package. were used as additional controls.
However, these reactions are identical for all members of
the B. cereus group namely, B. cereus, Bacillus mycoides, Bacillus
Preparation of sample dilutions
thuringiensis, and Bacillus anthracis. Therefore, additional
Ten grams from each fried rice sample was randomly taken tests were performed by inoculating the isolates on Sheep
and homogenized in 90 mL of 0.1% Peptone Water (CM0009) Blood Agar (CM0854; Oxoid Ltd), Nutrient Agar (CM0003;
PREVALENCE OF BACILLUS CEREUS IN FRIED RICE 127

Oxoid Ltd) and staining 3–4 day old cultures with warm 0.5% by comparative disk diffusion method. The antibiotics tested
basic fuchsin (212545; BD Difco BBL Stains) for toxin crystals. were ciprofloxacin (5 lg), erythromycin (15 lg), gentamicin
B. cereus isolates were identified as those showing active (10 lg), penicillin (10 lg), chloramphenicol (30 lg), and ami-
motility, strong hemolysis, no rhizoid growth, and no protein kacin (30 lg) (Oxoid Ltd). The resistance profile to penicillin
toxin crystals (USFDA, 2001). The number of B. cereus was was further investigated with Penicillin Etest (Biomerieux)
calculated by using the proportion of selected colonies con- minimal inhibitory concentration (MIC) method which was
firmed as B. cereus and the dilution factor (Presumptive col- performed and interpreted per manufacturer’s instructions
ony count · proportion confirmed as B. cereus · dilution and CLSI, M45-A, Guideline (CLSI, 2006).
factor · 10). The confirmed colony counts were expressed as
cfu/g of fried rice. Results
Of the 200 fried rice samples analyzed, B. cereus was de-
Assessment of risk factors tected in 112 (56.0%) whereas 44% (88) of the samples were
Occurrence of > 106 cfu/g of B. cereus were assessed with negative for B. cereus. Among these positive samples, 28 (14%)
regard to five presumed risk factors. These included the type had colony counts > 106 which corresponds to the infectious
of rice (Basmathi, Samba, others), amount of rice boiled per dose. Simultaneously 84 (42%) samples contained B. cereus
day ( > 10, £ 10 kg), method of storage of boiled rice (steaming colony counts £ 106. For counts exceeding 106 the highest
hot, room temperature, refrigerated), duration of storage of prevalence was detected in chicken fried rice (23.7%). None of
boiled rice before frying ( £ 4, > 4 hours), and frequency of the beef fried rice contained B. cereus in excess of 106. With
preparation of boiled rice per dining session (1, > 1). Bivariate regard to counts less than 106, the highest prevalence of 44.4%
associations were assessed and those which were statistically was individually observed in both seafood and egg fried rice
significant ( p-value < 0.05) were included for binary logistic whereas beef fried rice recorded the lowest prevalence of 20%
regression analysis. (Table 1).
In the assessment of risk factors for the occurrence of B.
Serotyping of B. cereus cereus in counts > 106 cfu/g three parameters emerged as in-
dependent risk factors (Table 2). These were storage at room
Serotyping of B. cereus isolates detected at > 106 cfu/g was temperature before frying ( p = 0.030), > 4 hours of storage
carried out at the Food Safety Microbiology Laboratory of prior to frying ( p = 0.042), and frequency of cooking more than
PHLS Central Public Health Laboratory, United Kingdom. once ( p = 0.017).
Actively motile strains obtained by several passages through The majority of B. cereus isolates present in counts exceed-
semisolid nutrient agar were cultured in nutrient broth for 5 ing 106 were not typable. Amongst the typable isolates, the
hours at 35C with constant agitation. After confirming the most prominent serotypes were H15 (14.3%) and H19 (14.3%).
presence of active motility by phase contrast microscopy, Serotype H20 was observed at 10.7% (Table 3). Antibiotic
formalin 1% (v/v) was added to obtain antigen suspensions. sensitivity demonstrated an identical pattern. All isolates
Flagellar ‘‘H’’ antisera prepared by intravenous inoculation of were sensitive to ciprofloxacin (100%), erythromycin (100%),
rabbits was used for typing the isolates. Doubling dilutions of gentamicin (100%), chloramphenicol (100%), and amikacin
0.25 mL portions of antirsera were made in agglutination (100%). Conversely, 100% resistance was observed for peni-
trays and equal volumes of antigen suspension added. Final cillin. This was confirmed by E-test which showed MIC to be
dilutions were 1:40 to 1:640. These were incubated at 50C for within the range of 32–256 lg/mL for all isolates.
2 hours. Macroscopic agglutination was observed with dark
field illumination (Taylor and Gilbert, 1975). Discussion
In many food poisoning outbreaks due to B. cereus, cooked
Antibiotic sensitivity testing of B. cereus isolates
rice has been implicated as the source. Table 4 indicates some
For B. cereus isolates occurring at counts > 106 cfu/g, sus- of the notable outbreaks reported since 1971. Almost all out-
ceptibility to a standard panel of antibiotics was determined breaks were associated with symptoms of emetic syndrome.

Table 1. Prevalence of Bacillus cereus Classified as Counts of £ 106 cfu/g, > 106 cfu/g,
and Nondetectable by Variety of Chinese-Style Fried Rice

Not detected £ 106 cfu/g > 106 cfu/g Total number analyzed from each variety

Variety No: % No: % No: % No: %

Vegetable 34 48.57 29 41.43 07 10.00 70 35.00

Chicken 19 32.20 26 44.07 14 23.73 59 29.60
Seafood 16 44.44 16 44.44 04 11.11 36 18.00
Egg 08 44.44 08 44.44 02 11.11 18 9.00
Beef 08 80.00 02 20.00 – – 10 5.00
Mixed 03 42.86 03 42.86 01 14.29 07 3.50
Total 88 44.00a 84 42.00 28 14.00 200 100.0
a
Total number of samples with Bacillus cereus detected = (200–88) 112 (56.00%).
cfu/g, colony forming units per gram.
128 PERERA AND RANASINGHE

Table 2. Risk Factors for Bacillus cereus Counts > 106 cfu/g in Fried Rice
with Adjusted Odds Ratios and 95% Confidence Intervals

Factor Frequency % p-value Adjusted OR 95% CI

Step1
Frequency of cooking per dining session 0.067 2.80 0.93–8.41
Only once 07
More than once 21
Temperature of storage before frying 0.034 5.41 1.14–25.71
Room temperature 26
Storage in fridge/hot steaming 2
Time of storage before frying 0.064 4.39 0.91–21.05
£ 4 hours 2
> 4 hours 26
Step 2
Frequency of cooking per dining session 0.017 3.16 1.23–8.14
Only once 07
More than once 21
Temperature of storage before frying 0.030 5.54 1.18–25.97
Room temperature 26
Storage in fridge/hot steaming 2
Time of storage before frying 0.042 4.55 1.2–21.43
£ 4 hours 16
> 4 hours 12

This may be attributed to the stimulatory effect of starch on Studies reveal that the degree of B. cereus contamination of
growth of B. cereus and production of emetic toxin (Kramer boiled rice was directly related to the temperature of storage
and Gilbert, 1989). and length of time the rice was kept before serving (Gilbert
This study revealed 56% of fried rice samples to be con- et al., 1974; Little et al., 2002). It is recommended that boiled
taminated with B. cereus. This compares favorably with rice not be stored at ambient temperature for more than four
prevalence for fried rice reported by other investigators, hours as this would permit resistant spores to germinate and
Notermans and Batt (1998) 12%–86% in the Netherlands, proliferate. This findings were corroborated by our study
Bryan et al. (1981) 85.7% in the United States, and Schiemann which showed statistically significant association between
(1978) 33% in the United Kingdom. Similar high prevalence of B. cereus in counts > 106 cfu/g and storage of boiled rice at
B. cereus has been observed for boiled rice; 100% in India room temperature for > 4 hours before preparation. Two
(Kamat et al., 1989), 92.9% (Bryan et al., 1981) and 91.7% outbreaks described by Khodr et al. (1994) and Shiota et al.
(Harmon and Kautter, 1991) in the United States, 10%–93% in (2010), where rice was stored at room temperature for a pro-
the Netherlands (Notermans and Batt, 1998), and 93.9% in the longed period and reheated are classic examples for time
United Kingdom (Nichols et al., 1999). The colony counts and temperature abuse mentioned above.
observed in these studies ranged from < 102 to ‡ 107. The Bulk cooking of rice once per day and cooling at room
colony counts observed in the current study were 103 (13.0%), temperature were shown to be associated with high B. cereus
104 (9.0%), 105 (11.5%), 106 (8.5%), 107 (3.5%), 108 (2.5%), 109 counts (WHO, 2000). This has been attributed to the presence
(4.5%), and 1010 (3.5%). B. cereus becomes a hazard when of favorable growth conditions. However, the findings of
present in numbers exceeding 106 cfu/g. Our study revealed present study contradicted this notion as it showed frequency
14% of samples to have this count which is considerably of cooking more than once to be an independent risk factor for
higher than 0.4% reported by Nichols et al. (1999) and 3.1% by
Little et al. (2002).
The high counts of B. cereus associated with boiled rice
could result from heat activation and germination of spores Table 3. Serotypes of Bacillus cereus Isolates
present in raw rice. Prevalence of B. cereus in raw rice has been Obtained from Fried Rice Samples
reported as 38%–68% by coworkers in India (Vijayalakshmi Containing > 106 cfu/g
et al., 1981), the United States (Ankolekar et al., 2009), Le- Total number % of
bannon (Hassan and Nabbut, 1996), the United Kingdom Serovar ‘‘H’’ of each serovar each serovar
(Blakey and Priest, 1980), and Korea (Park et al., 2009). In
addition, B. cereus vegetative cells may be present in meat Nontypable 15 53.57
and seafood whereas dried foods and spices contain spores 15 04 14.29
(Bryan et al., 1981; Little et al., 2002). There were clearly evi- 19 04 14.29
dent variations in prevalence of B. cereus by the variety of 20 03 10.71
Polyaglutination 01 3.57
fried rice in our study. Therefore, it could be contended that
Autoagglutination 01 3.57
addition of meat, sea food, herbs, and spices may have con- Total 28 100
tributed to the observed variations which merits further
investigation. Serotyping was performed by using B. cereus H-specific antisera.
PREVALENCE OF BACILLUS CEREUS IN FRIED RICE 129

Table 4. Bacillus cereus Outbreaks Associated with Cooked Rice

Bacillus cereus
Year Country Food item Symptoms/syndrome count (cfu/g) Reference

1971 United Kingdom Chinese-style V, N, (D) 106–109 Taylor and

fried rice Emetic Type Gilbert (1975)
1973–1982 Japan Rice and chicken, V, N 104–108 Shinagawa et al. (1985)
Fried rice Emetic Type
1974–1999 Japan Boiled rice, V, N 106–108 Agata et al. (2002)
fried rice, Emetic Type
sushi rice,
rice gruel
1976 Finland Boiled rice V, C, (D) 1.7 · 108 Raevuori et al. (1976)
Emetic type
1981 Singapore Fried rice V, C, (D), Not documented Tay et al. (1982)
headache, fever
Emetic type
1985 United States Cooked rice V, N, (D) Not documented CDC (1986)
Emetic Type
1985 United States Rice and chicken D, C, N Not documented Baddour et al. (1986)
Diarrheal type
1991 India Cooked rice V, N Not documented Granum and
Emetic Type Baird-Parker (2000)
1993 United States Fried rice N, V, C, (D) > 106 Khodr et al. (1994)
with chicken Emetic Type
1994 United States Cooked rice V, N Not documented Granum and
and macaroni Emetic Type Baird-Parker (2000)
1994 Scotland Fried rice V, N, C, 2.0 · 107 Dickson and
Emetic Type Morgan (1995)
1996 India Cooked rice V, N Not documented Granum and
Emetic Type Baird-Parker (2000)
2000 United Kingdom Cooked rice V > 106 Ripabelli et al. (2000)
Emetic type
2000 The Netherland Vegetarian rice V, C 105–108 The EFSA
Emetic type Journal (2005)
2008 Japan Fried rice V, N Not documented Shiota et al. (2010)
Emetic Type

V, vomiting; N, nausea; C, cramps; D, diarrhea; (D), mild/occasional diarrhea.

significant B. cereus counts. Possible explanations for this re- MIC ranging from 32 to 256 lg/mL was demonstrably higher
sult could be postcooking contamination from inadequately than 95% reported for the US isolates (Luna et al., 2007).
cleaned utensils and addition of left over rice to new batches
(Bryan et al., 1981). But the present study did not include ex- Conclusions
amination of postcooking contamination. The type of rice and
With B. cereus prevalence of 56%, Chinese-style fried rice in
the quantity boiled per day was not a contributing factor for
Colombo city could be a hazard for foodborne outbreaks.
high B. cereus counts. These two parameters have not been
Common practices in preparation such as storage of boiled
previously investigated. Small sample size with predictably
rice at room temperature for prolonged periods and a greater
wide 95% confidence intervals was a limitation.
frequency of cooking emerged as significant risk factors for B.
Majority of our B. cereus isolates were not typable. Untyp-
cereus contamination. Therefore, food-handlers should be in-
able strains of B. cereus have been responsible for both emetic
structed on safe practices which include keeping boiled rice ei-
and diarrheal syndrome (Kramer and Gilbert, 1989). The
ther at > 60C (hot steaming) or cooling rapidly and transferring
present study revealed serotypes H15, H19, and H20. H19 has
to a refrigerator within 4 hours (WHO, 2000). Local serotypes
been associated with both emetic and diarrheal syndrome
could be associated with both emetic and diarrheal syndromes.
(Kramer et al., 1982). H15 and H20 have been reported from
Aminoglycosides, ciprofloxacin, and chloramphenicol are suit-
diarrheal and emetic syndromes respectively. Further, H15
able therapeutic options for indigenous infections.
has been isolated from raw rice, H19 from boiled rice, and H20
from both (Gilbert and Kramer, 1986).
Acknowledgments
The 100% sensitivity for ciprofloxacin, erythromycin, gen-
tamicin, chloramphenicol, and amikacin in our study is sim- This project was supported by a grant from the Medical
ilar to that observed by Stec (1990) and Meena et al. (2000), Research Institute, Colombo, Sri Lanka. For professional ad-
differing from that of Luna et al. (2007) where erythromycin vice and assistance we gratefully acknowledge Professor
sensitivity was 84%. The uniform resistance to penicillin with Chandra Kodikara and Dr. Karven Cooray.
130 PERERA AND RANASINGHE

Disclosure Statement Kramer JM and Gilbert RJ. Bacillus cereus and other Bacillus
species. In: Food Borne Bacterial Pathogens. Doyle MP (ed.).
No competing financial interests exist. New York: Marcel Dekker, 1989, pp. 21–70.
Kramer JM, Turnball PCB, Munshi G, and Gilbert RJ. Identifi-
References
cation and characterization of B. cereus and other Bacillus
Agata N, Ohta M, and Yokoyama K. Production of Bacillus cereus species associated with foods and food poisoning. In: Isolation
emetic toxin (cereulide) in various foods. Int J Food Microbiol and Identification Methods for Food Poisoning Organisms. Society
2002;73:23–27. for Applied Bacteriology Technical Series. Corry J, Roberts D,
Ankolekar C, Rahmati T, and Labbé RG. Detection of toxigenic and Skinner FA (eds.). London: Academic Press, 1982, pp.
Bacillus cereus and Bacillus thuringiensis spores in U.S. rice. Int J 261–286.
Food Microbiol 2009;128:460–466. Lake R, Hudson A, and Cressey P. Risk profile: Bacillus spp. in
Baddour LM, Gaia SM, Griffin R, and Hudson R. A hospital Rice [Client report FW0319]. New Zealand: Institute of En-
cafeteria-related food-borne outbreak due to Bacillus cereus: vironmental Science and Research, 2004.
unique features. Infect Control 1986;7:462–465. Little CL, Barnes J, and Mitchel RT. Microbiological quality of
Blakey LJ and Priest FG. The occurrence of Bacillus cereus in take away cooked rice and chicken sandwiches; effectiveness
some dried foods including pulses and cereals. J Appl Bac- of food hygiene training of the management. Commun Dis
teriol 1980;48:297–302. Public Health 2002;5:289–298.
Bridson EY (ed.). Oxoid Manual, ninth edition. UK: Oxoid Lim- Luna V, King D, Gulledge J, Cannons A, Amuso P, and Cattani J.
ited, 2006. Susceptibility of Bacillus anthracis, Bacillus cereus, Bacillus my-
Bryan FL, Bartleson CA, and Christopherson N. Hazard analy- coides, Bacillus pseudomycoides and Bacillus thuringiensis to 24
ses, in reference to Bacillus cereus, of boiled and fried rice in antimicrobials using Sensititre automated microbroth dilution
Cantonese-style restaurants. J Food Prot 1981;44:500–512. and Etest agar gradient diffusion methods. J Antimicrob
[CDC] Centers for Disease Control and Prevention. Bacillus ce- Chemother 2007;60:555–567.
reus-maine. Morb Mortal Wkly Rep 1986;35:408–410. Meena BS, Kapoor KN, and Agarwal RK. Occurrence of multi-
[CLSI] Clinical and Laboratory Standards Institute. Methods for drug resistant Bacillus cereus in foods. J Food Sci Technol
Antimicrobial Dilution and Disk Susceptibility Testing of Infre- 2000;37:289–291.
quently Isolated or Fastidious Bacteria: Approved Guideline M45- Ministry of Healthcare and Nutrition Sri Lanka, Epidemiology
A. Wayne, PA: CLSI, 2006. Unit. Quarterly Epidemiological Bulletins. Volumes 45–51. 2005.
Dickson A and Morgan M. Scottish Centre for Infection and Available at www.epid.gov.lk/Epidemiological%20Bulletin
Environmental Health Weekly Report. Glasgow, UK: Scottish .htm, accessed June 14, 2011. (Online.)
Centre for Infection and Environmental Health. 1995;29:7. Morita TN and Woodburn MJ. Stimulation of Bacillus cereus
Gilbert RJ and Kramer JM. Bacillus cereus food poisoning. In: growth by protein in cooked rice combinations. J Food Sci
Progress in Food Safety. Oliver DO and Cochrane BA (eds.). 1977;42:1232–1235.
University of Wisconsin-Madison: Food Research Institute, Nichols GL, Little CL, Mithani V, and de Louvois J. The mi-
1986, pp. 85–93. crobiological quality of cooked rice from restaurants and take-
Gilbert RJ, Stringer MF, and Peace TC. Survival and growth of away premises in the United Kingdom. J Food Prot
Bacillus cereus in boiled and fried rice in relation to outbreaks 1999;62:877–882.
of food poisoning. J Hyg 1974;73:433–444. Notermans S and Batt CA. A risk assessment approach for food-
Granum PE. Bacillus cereus. In: Food Microbiology Fundamentals borne Bacillus cereus and its toxins. J Appl Microbiol Symp
and Frontiers. Doyle MP (ed.). Washington, DC: ASM Press, Suppl 1998;84:51S–61S.
2001, pp. 373–381. Park YB, Kim JB, Shin SW, Kim JC, Cho SH, et al. Prevalence,
Granum PE and Baird-Parker TC. Bacillus species. In: Micro- genetic diversity, and antibiotic susceptibility of Bacillus cereus
biological Safety and Quality of Food, Volume 2. Lund BM, Baird- strains isolated from rice and cereals collected in Korea. J Food
Parker TC, and Gould GW (eds.). Gaithersburg: Aspen Pub- Prot 2009;72:612–617.
lishers, 2000, pp. 1029–1039. Raevuori M, Kiutamo T, Niskanen A, and Salminen K. An
Harmon SM and Kautter DA. Incidence and growth potential of outbreak of Bacillus cereus food poisoning in Finland associ-
Bacillus cereus in ready to serve foods. J Food Prot 1991;54:372– ated with boiled rice. J Hyg 1976;76:319–327.
374. Ripabelli G, McLauchlin J, Mithani V, and Threlfall EJ. Epi-
Hassan G and Nabbut N. Prevalence and characterization of demiological typing of Bacillus cereus by amplified frag-
Bacillus cereus isolates from clinical and natural sources. J Food ment length polymorphism. Lett Appl Microbiol 2000;
Prot 1996;59:193–196. 30:358–363.
Holbrook R and Anderson JM. An improved selective and di- Schiemann DA. Occurrence of Bacillus cereus and the bacterio-
agnostic medium for the isolation and enumeration of Bacillus logical quality of Chinese ‘‘take-out’’ foods. J Food Prot
cereus in foods. Can J Microbiol 1980;26:753–759. 1978;41:450–454.
Kamat AS, Nerkar DP, and Nair PM. Bacillus cereus in some Shinagawa K, Matsusaka N, Konuma H, and Kurata H. The
Indian foods, incidence and antibiotic, heat and radiation re- relation between the diarrheal and other biological activities of
sistance. J Food Saf 1989;10:31–41. Bacillus cereus involved in food poisoning outbreaks. Nippon
Khodr M, Hills S, Stiefel S, Comer-Morrison C, and Lee S. Ba- Juigaku Zasshi 1985;47:557–565.
cillus cereus food poisoning associated with fried rice at two Shiota M, Saitou K, Mizumoto H, Matsusaka M, Agata N, et al.
child day care centers in Virginia, 1993. Morb Mortal Wkly Rapid detoxification of cereulide in Bacillus cereus food poi-
Rep 1994;43:177–178. soning. Pediatrics 2010;125:E951–E955.
Kotiranta A, Lounatmaa K, and Haapasalo M. Epidemiology Sri Lanka Standard: 516: Part 8. Microbiological Test Methods—
and pathogenesis of B. cereus infections. Microbes Infect Detection and Enumeration of Bacillus cereus. UDC 664:543.9.078.
2000;2:189–198. Colombo: Sri Lanka Standards Institution, 1983.
 PREVALENCE OF BACILLUS CEREUS IN FRIED RICE 131

Stec E. Sensitivity to antiobiotics and selected compounds Vilas-Boas GT, Peruca AP, and Arantes OM. Biology and tax-
of Bacillus cereus isolated from food products and onomy of Bacillus cereus, Bacillus anthracis and Bacillus thir-
cases of food poisoning. Recz Panstw Zakl Hig 1990;41: ungiensis. Can J Microbiol 2007;53:673–687.
269–272. [WHO] World Health Organization. Foodborne Disease: A Focus
Tay L, Goh KT, and Tan SE. An outbreak of Bacillus cereus food for Health Education. Geneva, 2000.
poisoning. Singapore Med J 1982;23:214–217. [WHO] World Health Organization. WHO Initiative to Estimate the
Taylor AJ and Gilbert RJ. An outbreak of B. cereus poisoning: a Global Burden of Foodborne Diseases. Second formal meeting of
provisional serotyping scheme. J Med Microbiol 1975;8:543– the Foodborne Diseases Burden Epidemiology Reference Group
550. (FERG). Geneva, 2009, p. 18. Available at www.who.int/entity/
The EFSA Journal. Opinion of the Scientific Panel on Biological foodsafety/foodborne_disease/FERG2_report.pdf, accessed Jan-
Hazards on Bacillus cereus and other Bacillus spp in foodstuffs. uary 14, 2010. (Online.)
European Food Safety Auth J 2005;175:1–48.
[USFDA] US Food and Drug Administration, Centre for Food Address correspondence to:
Safety and Applied Nutrition. Bacteriological analytical manual Manosha Lakmali Perera, M.Sc.
online. 2001. Available at www.fda.gov/Food/ScienceResearch/ Molecular Biology Section
LaboratoryMethods/BacteriologicalAnalyticalManualBAM/ Medical Research Institute
ucm070875.htm, accessed January 14, 2010. (Online.) Colombo 8
Vijayalakshmi G, Dwarakanath CT, and Murthy VS. Studies on Sri Lanka
Bacillus cereus contamination of rice and rice preparation. J
Food Sci Technol 1981;18:231–234. E-mail: [email protected]

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