PREVALENCE OF BACILLUS CEREUS AND ITS ENTEROTOXIN IN SOME COOKED AND HALF COOKED CHICKEN PRODUCTS

Assiut Vet. Med. J. Vol. 52 No. 109 April 2006

Department of Food Hygiene, Animal Health Research Institute, Dokki, Giza.

PREVALENCE OF BACILLUS CEREUS AND ITS ENTEROTOXIN IN SOME COOKED AND HALF COOKED CHICKEN PRODUCTS

(With 4 Tables)

By ENSHRAH K.). MIRA and SUSAN M.A. ABUZIED

(Received at 5/3/2006)

مدى تواجد ميکروب الباسلس سيرس وسمومه في بعض منتجات

الدواجن المطهية والنصف مطهية انشراح خليل ميرة ، سوزان احمد ابو زيد

أجريت هذه الدراسة على خمسة أنواع من منتجات الدواجن (ناجتس، فيليه صدور، فاهيتا، دبوس وأجنحة) وکانت هذه الأنواع عبارة عن منتجات دواجن جاهزة للأکل تم تجميعها من محلات الوجبات السريعة ومنتجات دواجن نصف مطهية مجمدة تم تجميعها من السوبر مارکت وکذلک نفس العينات المجمدة النصف مطهية التي تم إعدادها طبقا لطريقة الطهي المدونة على الغلاف الخارجي لکل منتج. وتم فحص جميع العينات المجمعة للکشف عن تواجد میکروب الباستس سيرس وقدرة العترات المعزولة على إفراز السموم المعوية. وأثبتت نتائج الدراسة تواجد میکروب الباسکس سيرس في جمي

في جميع انواع العينات بنسب مختلفة وکانت اعلى نسبة عزل للميکروب من عينات الوجبات الجاهزة يليها المنتجات النصف مطهية المجمدة في حين أثبتت الدراسة أن إتباع طرق الطهي العينات النصف مطهية المدونة على العبوات أنقصت من نسبة تواجد الميکروب بالعينات. بالنسبة لقدرة العترات المعزولة من ميکروب الباسلس سيرس على إنتاج السموم المعوية باستخدام

B. cereus enterotoxin reversed passive latex agglutination kit أظهرت نتائج البحث أن ۱۳ عترة من

۳۳ عترة (۳۹٫4%) و

عترات من ۳۰ (۲۰%) أيجابية لإفراز السموم المعوية بعينات منتجات الدواجن المطهية الجاهزة والعينات المجمدة النصف مطهية على التوالي بينما وجدت عترتان فقط من ، عترات معزولة من العينات المطهية طبقا للطرق المدونة لها القدرة على إفراز السموم.

SUMMARY

Five types of chicken products (Nuggets, Tenders, Fajieta, Drum sticks and Wings) were used in this study. These types included ready-to-eat chicken products which were collected from fast food shops and frozen

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half cooked chicken products collected from supermarkets. Some samples of frozen half cooked products were cooked according to the cooking instruction printed on the package label. All samples were examined for the detection of B. cereus and the ability of the isolated strains to produce diarrheal enterotoxin. B. cereus was isolated from all the types of the examined samples with different percentages. Ready-to eat chicken product samples recorded the highest incidence of isolation followed by frozen half cooked chicken products. While applying cooking instruction reduced the incidence of B. cereus.

By using B. cereus enterotoxin reversed passive latex agglutination kit, Bacillus diarrheal enterotoxin was detected from 13 out of 33 strains (39.4%) isolated from ready-to-eat samples, and 6 out of 30 isolates (20%) from frozen half cooked samples. Whereas, only 2 out of 6 isolates were diarrheal enterotoxigenic producing strains isolated from cooked chicken product samples.

Key words: Cereus, chicken products, B. cereus enterotoxin, passive

latex agglutination technique.

INTRODUCTION

Bacillus cereus is an aerobic spore-forming bacterium commonly found in the environment. It is likely to be found in food because of its widespread occurrence and because of certain factors that favor its survival and presence in food production environments.

B. cereus is an important foodborne pathogen, and its strain can be characterized as mesophilic or psychrotrophic. Mesophilic strains have a growth range of 15-55 °C and their spores tends to be more heat resistant. Whereas, psychrotrophic strains have a growth range of 4 to 35 °C and their spores tend to be less heat resistant (Choma et al. 2000 and Granum et al. 2000).

Human gastroenteritis attributed to B. cereus is a food intoxication after ingestion of food containing preformed enterotoxins rather than a result of colonization or infection of host.

B. cereus food poisoning was attributed to two enterotoxins, diarrhoeal which has an onset period of 10-15 hours after ingestion and mainly associated with proteinaceous food such as meat products and milk, and emetic which has a short onset period of 1-5 hours after ingestion and it is mainly associated with farinaceous foods, particularly rice, and cereal derivatives such as flour (Kramer and Gilbert, 1989).

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A variety of different chicken product types, breaded, half or fully cooked are widely spread in Egyptian markets and most preferable especially by children and youth. Poultry is likely to be contaminated with B. cereus during grow out, from dusty housing condition, from contaminated chicks, or from feed. Hatchery environments, facilities and equipment have been shown to be contaminated with cells, which could be contaminating chicks (Willinghan et al. 1996).

Spores survive feed manufacture and readily colonize the gut of the chicken (Jadamus et al. 2001). As would be expected B. cereus has been found in meat and poultry products (Hatakka, 1998, Fang et al. 2002; Tessi et al. 2002).

Ingredients typically added to meat products, such as spices, seasoning and protein supplements, have been found to contain B. cereus (Konuma et al. 1998 and Te Giffel et al. 1996). Similarly, ingredients typical of those used in breaded coatings for chickens, such as wheat products and flour, have been shown to contain B. cereus (Te Giffel et al. 1996). Food packaging proper has been reported to harbor B. cereus (Vaisanen et al. 1991), which could lead to post cook exposure.

Therefore the aim of this study was to determine whether B. cereus or its enterotoxin could be detected in several types of retail chicken products.

MATERIALS and METHODS

Samples:

This study was performed on five types of chicken products (Nuggets, Tenders, Fajieta, Drum-sticks and Wings) where ten samples of each item were obtained. First: A survey was conducted on the ready-to-eat aforementioned products served in Giza Governorate restaurants, a total of 50 samples (10 samples of each product were obtained). Second: 50 samples of frozen half cooked chicken products (the same types of the products) were collected from supermarkets and divided into two groups. The first group was examined without cooking whereas the second group of the samples was cooked according to the cooking instructions written on the package labels.

All samples were subjected to bacterial analysis for isolation and identification of B. cereus. - Isolation of B. cereus was performed as described in the

Bacteriological Analytical Manual (US FDA, 1995) by using 25 gm of

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the sample enriched in 225 ml of trypticase soy-polymixin broth which stomachered for 1 min and incubated for 18-24 h at 30°C. then 0.1 ml was plated on mannitol-egg yolk-polymixin and incubated at 30°C for

18-24 h - Suspected B. cereus colonies were identified morphologically and

biochemically according to Cowan and Steel, (1974). - - Detection of B. cereus enterotoxin was done by reversed passive latex agglutination technique (BCET-RPLA SEIKEN, Japan, Tokyo, 103)

RESULTS

Table 1: Incidence of B. cereus isolated from ready-to-eat chicken

product samples Samples No. of samples

Positive samples

No. Nuggets

10 Tender Fajita Drumsticks Wings

60

10

10

50

aoua

10

80

10

Table 2: Incidence of B. cereus isolated from

chicken product samples

frozen half cooked

Samples

No. of samples

10

Positive samples No.

80 40 40

Nuggets Tender Fajita Drumsticks Wings

10

10 10 10

80

60

Table 3: Incidence of B. cereus isolated from chicken product samples

after application of cooking instructions. Samples No. of samples No of +ve samples Reduction %

0

1

0

Nuggets

10 Tender

10 Fajita T10 T Drumsticks

10 Wings

10

100 100 100 60

O

4

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Table 4: Incidence of enterotoxigenic strains of B. cereus isolated from

ready-to-eat, frozen half cooked and cooked chicken product

samples: Samples Ready-to-eat

Frozen half cooked

Cooked No. of Toxin 1 % No. of Toxin % No. of Toxin

+ve production

tve production

+ve production Nuggets

258 ! 2 25 0

0 Tender

6

1

16 4 1 -

- 0

0 Fajita

3 6014

-

- 0

0 Drumsticks 8

5 62.5 8

2 25 4 12 Wings

6 2 33.3 6 2

33.3 2 Total I 33 | 13 39.4 30 6 20 6 2

0

0

10

on 10

NONOO

50

0

33.3

DISCUSSION

A survey was conducted on ready to eat five chicken products (Table 1) indicated that B. cereus was isolated from all the examined samples. The highest incidence of B. cereus was found in nuggets and drumsticks (80%) followed by tenders and wings (60%) then fajieta! (50%).

Nearly similar results were obtained by Harmon and Kautter (1991): Tessi et al. (2002) and Murindamombe et al. (2005), while lo percentages of occurrence were reported in other studies (Mosupye and Von Holy, 1999, 2000 and Umoh and Odoba, 1999).

The high incidence of B. cereus isolation may be due to the temperature of holding the chicken product, environmental contamination, surviving spores or chicken under cooking. Monitoring of cooking temperature on site was impossible, since shops served ready to eat fast food didn't consent to have their preparation examined.

Smith et al. (2004) reported that some types of cooked products are possible to mishandling and temperature abuse, which could potentially lead to the growth of B. cereus and toxin production, sometimes ready to eat chicken products are held at room or outdoor temperatures before served. During which time large population of bacteria, including pathogens, can proliferate, (Bryan et al., 1982a,b). This practice potentially hazardous for products that support the growth of pathogenic bacteria and should be prevented.

Results in Table (2) shows that B. cereus was isolated from all types of frozen half cooked chicken products samples, the highest incidence was from nuggets and drum stick (80%) followed by wings (60%) then, tenders and fajieta (40%).

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The presence of B. cereus in frozen half cooked products may be attributed to spore surviving from raw poultry, post processing contamination with either spices, seasoning or protein supplements that may contain B. cereus (Te Giffel, 1996 Konuma et al., 1998 and Smith et al., 2004).Wheat products and flour used in breaded coatings have been also shown to contain B. cereus (Te Giffel et al. 1996). Similarly, food packaging has been reported to harbor B. cereus (Vaisanen et al.,

1991).

The effect of preparing chicken products samples (cooking) according to the instruction printed on labels- on the incidence of B. cereus isolation is illustrated in Table (3). The results indicated that B. cereus failed to be isolated from nuggets, tenders and fajieta samples. This result indicated that cooking process was efficient to achieve 100% reduction of B. cereus isolated from samples before the cooking process and lead to conclude that the proper time-temperature exposure can be effective in killing B. cereus (Fruin and Guthertz, 1982 and ICMSF, 1996). Although, the cooking instruction for drum stick and wings samples were applied, B. cereus could survive and be isolated after cooking with reduction rate 60% and 80% respectively.

Patterson and Gibbs (1973) illustrated that cooking can kill heat sensitive microbes while allow heat resistant forms including B. cereus to survive. In addition, ICMSF (1996) reported that vegetative forms of pathogens in portions that are still frozen, in thick masses and in cavities or layers insulated by stuffing survive cooking temperature. Smith et al. (2004) found that chicken skin harbor B. cereus which may explain their presence in drumsticks and wings samples after cooking.

From the results achieved in Table (4), the enterotoxin analysis of the isolates in which the production of the diarrheal toxin was confirmed by the reversed passive latex agglutination test demonstrated that, 25, 16, 60, 62.53 and 33.3% of the isolates recovered from the five ready to eat samples elaborated the diarrheogenic toxin, respectively. While frozen half cooked nuggets, drum sticks and wing samples contained 2 (25%) of 8,2 (25%) of 8 and 2 (33.3%) of 6, isolates that were shown to be enterotoxin producers, respectively.

Strains of B. cereus isolated from drumsticks and wings after applying the cooking instructions revealed that only 2 out of 4 isolates recovered from drumsticks samples had the ability to produce the diarrheogenic enterotoxin.

Several studies proved that B. cereus isolated from frozen, cooked and ready to eat chicken products had the ability to produce

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enterotoxin. The results achieved by Smith et al. (2004) and Murindamombe et al. (2005) were approximately near to that obtained in the present study. While Choma et al. (2000) recorded that B. cereus isolates were enterotoxigenic in slightly higher percentage.

Therefore, finding indicated that at least for the types of products represented by these samples, control measures should be directed at preventing the germination and out growth of spores in cooked food. Heat-resistant bacterial spores especially B. cereus can be reduced by

able time-temperature to ensure safety. Also, by storing the cooked food at high temperature or keeping it at refrigeration temperature until use since B. cereus don't have the ability to grow and produce toxin at temperature below 4 °C (Van Netten et al., 1990).

Further researches are required to determine the scope of the problem and the source of contamination.

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