BACTERIAL HAZARDS OF CHICKEN SHAWERMA SANDWICHES

Assiut University web-site: www.aun.edu.eg

BACTERIAL HAZARDS OF CHICKEN SHAWERMA SANDWICHES

EL-DOSOKY, H.F.A. ¹; SHEREEN, S. MOSTAFA ² and RAMADAN, A.H.²

¹ Food Hygiene Dept. Mansoura Provential Lab, Animal Health Research Institute

² Microbiololgy Dept. Mansoura Provential Lab, Animal Health Research Institute

Received: 31 March 2018;     Accepted: 31 April 2018

INTRODUCTION

Chicken and chicken products provide animal protein of high biological value for consumers at all ages, where they contain all the essential amino acids required for growth with high proportion of unsaturated fatty acids and low cholesterol value moreover, poultry meat is a good source of different types of vitamins and minerals (Zaki and Shehata, 2008).

Meanwhile, Braun et al. (2005) mentioned that these foods are manipulated extensively during processing and therefore have a potential for high bacterial contamination levels on the meat surface, as well as the inside. As a result, there is an increased risk of pathogens surviving and transferring not only by cross-contamination, but also by undercooking as observed in this kind of fast-food industry.

Ghafir et al. (2008) mentioned that several bacterial indicators   are   used   to  evaluate the hygiene during

Corresponding author: Dr. EL-DOSOKY, H.F.A

E-mail address: rafat552008@yahoo.com

Present address: Food Hygiene Dept. Mansoura Provential lab, Animal Health Research Institute

meat slaughtering process including the counts of E.coli and other Enterobacteriaceae and aerobic bacteria while, (Ahmad et al., 2013) analyzed fresh shawerma for E.coli, Salmonella, aerobes and coliforms which were found in order of aerobes >E. coli> Salmonella, there is variation among the number of bacteria in every part aerobes were greater in number as compared to E. coli and Salmonella. (Logue and Nde 2007) mentioned that symptoms of food poisoning such as vomiting, nausea, cramps, diarrhea with or without blood, abdominal pain, or fever might appear after hours or even few days after consuming contaminated food. They are often mild and a person can recover alone at home but some people need to refer to the hospital. Risks of getting an infection are higher in infants or children since they don’t have a well-developed immune system and in old people as the response of their immune system becomes weaker.

Bennett et al. (2008) reported that 1229 food borne outbreaks caused by Bacillus cereus and Staph. aureus from 1998 to 2008 in U.S., 39% were with a confirmed etiology. Vomiting was commonly reported in Bacillus cereus (75%) of cases and (87%) of Staph. aureus outbreaks in addition to Berdgoll (1989) concluded that a very small amount of Staph. aureus enterotoxins ranging from 20 ng to < 1 μg is needed to cause a typical symptoms of staphylococcal food poisoning. Furthermore, Asao et al. (2003) mentioned that an outbreak in Japan caused by low-fat milk contaminated with SEA showed that the total intake of SEA per individual was estimated to be 20–100 ng. More recently, Ostyn et al. (2010) in French found an outbreak caused by contaminated cheese, doses of SEE ingested by symptomatic persons were estimated to be about 90 ng. Presence of Salmonella in frozen meat products cause serious infection if not cooked properly (Dominguez and Schaffner, 2009).

Therefore, this work was planned to evaluate some of the bacteriological dangers in chicken shawerma sandwiches sold in Mansoura city.

MATERIALS AND METHODS

A- Collection of samples: A total of 100 random samples of chicken shawerma sandwiches were collected separately from different restaurants with variant localities at Mansoura city and aseptically transferred without delay in an insulated ice box to the laboratory, then subjected to the following.

B - Bacteriological examination:

1- Preparation of food homogenate: according to the technique recommended by ISO, 6887-2 (2003) 25 g of each sample were taken by a sterile knife and stomached using Seward stomacher 80 biomaster England with 225 ml sterile buffered peptone water (0.1%) to give a homogenate of 1/10 dilution from which six fold serial dilutions were prepared and subjected to the following bacteriological examination.

2- Aerobic Plate Count (APC): It was carried out according to APHA (2001)

3- Staph. aureus count: FDA (2002) using Baird-Parker agar plates, incubated at 35^oC for 48 hr and the suspected Staph. aureus colonies were  isolated, and confirmed bycatalase, coagulase, thermos table nuclease  and Voges-Proskauer tests.

4- Bacillus cereus count: according to the technique recommended by ISO, (2004) using Mannitol egg yolk polymexinphenol red agar and confirmed biochemically by catalase, nitrate reduction test, Voges-Proskauer test and detection of haemolysis.

5- Counting and Identification of Beta-glucuronidase-positive E. coli according to the technique recommended by ISO, 16649/2(2001)      E. coli was detected by using sorbitol Mac Conkey agar medium (Oxoid, England). Pure suspected colonies were biochemicaly identified by using urease, Indol, methyl red, H₂S production test and citrate utilization test according to (Koneman et al., 1997). And Salmonella according to the technique recommended by ISO, 6579 (2002) on enrichment Rappaport vassiliades broth at 42⁰C for 24-48h, platting on XLD agar at 35⁰C for 24h. The presumptive colonies were confirmed biochemically and by multiplex PCR.

C- Detection of virulence genes: The isolated Staph. aureus, E. coli and Salmonella  were examined  by using PCR for detection of Staph. aureus enterotoxins; E. coli (stx1 and stx2) and Salmonella typhimurium (invA, and stn). Reference Lab. for Quality Control on Poultry Production, Animal Health Research Institute, Dokki -Egypt.

1- DNA extraction for isolated Staph. aureus:

a- DNA extraction from samples was performed using the QIA amp DNA Mini kit (Qiagen, Germany, GmbH) with modifications from the manufacturer’s recommendations. Briefly, 200 µl of the sample suspension was incubated with 10 µl of proteinase K and 200 µl of lysis buffer at 56^OC for 10 min. After incubation, 200 µl of 100% ethanol was added to the lysate. The sample was then washed and centrifuged following the manufacturer’s recommendations. Nucleic acid was eluted with 100 µl of elution buffer provided in the kit.

b- Oligonucleotide Primers used were supplied from Metabion (Germany) are listed in Table 1.

c- For multiplex PCR of enterotoxins, Primers were utilized in a 50- µl reaction containing 25 µl of Emerald Amp Max PCR Master Mix (Takara, Japan), 1 µl of each primer of 20 pmolconcentration, 8 µl of water, and 7 µl of DNA template. The reaction was performed in an Appliedbiosystem 2720 thermal cycler.

d- Analysis of the PCR Products: The products of PCR were separated by electrophoresis on 1.5% agarose gel (Applichem, Germany, GmbH) in 1x TBE buffer at room temperature using gradients of 5V/cm. For gel analysis, 30 µl of the multiplex PCR products were loaded in each gel slot. Gelpilot 100 bp DNA ladder (Qiagen, Germany, GmbH) was used to determine the fragment sizes. The gel was photographed by a gel documentation system (Alpha Innotech, Biometra) and the data was analyzed through computer software.

2- DNA extraction for isolated E. coli:

a- DNA extraction from samples was performed using the QIA amp DNA Mini kit (Qiagen, Germany, GmbH) with modifications from the manufacturer’s recommendations. Briefly, 200 µl of the sample suspension was incubated with 10 µl of proteinase K and 200 µl of lysis buffer at 56^OC for 10 min. After incubation, 200 µl of 100% ethanol was added to the lysate. The sample was then washed and centrifuged following the manufacturer’s recommendations. Nucleic acid was eluted with 100 µl of elution buffer provided in the kit.

b- Oligonucleotide Primers used were supplied from Metabion (Germany) are listed in Table 1.

c- PCR amplification For stx1, stx2 duplex PCR, primers were utilized in a 50- µl reaction containing 25 µl of Emerald Amp Max PCR Master Mix (Takara, Japan), 1 µl of each primer of 20 pmol concentration, 13 µl of water, and 8 µl of DNA template. The reaction was performed in an Applied biosystem 2720 thermal cycler.

d- Analysis of the PCR Products: The products of PCR were separated by electrophoresis on 1.5% agarose gel (Applichem, Germany, GmbH) in 1x TBE buffer at room temperature using gradients of 5V/cm. For gel analysis, 20 µl of the uniplex PCR products and 30 µl of the duplex PCR products were loaded in each gel slot. Generuler 100 bp DNA ladder (Fermentas, sigma) was used to determine the fragment sizes. The gel was photographed by a gel documentation system (Alpha Innotech, Biometra) and the data was analyzed through computer software.

3- DNA extraction for isolated Salmonella typhimurium:

a- DNA extraction from samples was performed using the QIA amp DNA Mini kit (Qiagen, Germany, GmbH) with modifications from the manufacturer’s recommendations. Briefly, 200 µl of the sample suspension was incubated with 10 µl of proteinase K and 200 µl of lysis buffer at 56^OC for 10 min. After incubation, 200 µl of 100% ethanol was added to the lysate. The sample was then washed and centrifuged following the manufacturer’s recommendations. Nucleic acid was eluted with 100 µl of elution buffer.

b- Oligonucleotide Primers used were supplied from Metabion (Germany) are listed in Table 1.

c- PCR amplification Primers were utilized in a 25- µl reaction containing 12.5 µl of Emerald Amp Max PCR Master Mix (Takara, Japan), 1 µl of each primer of 20 pmol concentration, 4.5 µl of water, and 6 µl of DNA template. The reaction was performed in an Applied biosystem 2720 thermal cycler.

d- Analysis of the PCR Product: The products of PCR were separated by electrophoresis on 1.5% agarose gel (Applichem, Germany, GmbH) in 1x TBE buffer at room temperature using gradients of 5V/cm. For gel analysis, 20 µl of the products was loaded in each gel slot. Gelpilot 100 bp DNA Ladder (Qiagen, Germany, GmbH) and generuler 100 bp ladder (Fermentas, Germany) were used to determine the fragment sizes. The gel was photographed by a gel documentation system (Alpha Innotech, Biometra) and the data was analyzed through computer software.

Table 1: Primers sequences, target genes, amplicon sizes and cycling conditions for Staph.aureus, E. coli and Salmonella typhimurium used in multiplex PCR.

Statistical analysis:

The results are expressed as log mean ± Standard Error (SE). Data were statistically analyzed using statistical analysis systems.

Table 2: Statistical Analytical Results of Bacterial Incidence inthe examined Chicken Shawerma Sandwich esexpressed as log mean±SE. (n=100).

Fig (1): Agarose gel electrophoresis of Staph. aureus PCR products using enterotoxins Staphylococcus primer.

Pos=positive control, Neg=negative control, L=100 bp DNA ladder

Lane "1": Positive amplification of 278bp for enterotoxin D

Lane "2" and Lane "4": positive amplification of 209 bp for enterotoxin E

Lane "3" Lane "5" were negative

Fig (2): Agarose gel electrophoresis of E. coli PCR products using stx1 and stx2 primers.

Lane "1"; Lane "3 "; Lane "4 "and Lane "5" were negative

Lane "2": Positive amplification of 614 bp for stx1 gene

DISCUSSION

Poultry are known to be a reservoir of large number of bacteria which may be pathogenic to human being. Typically, these occur in low sanitation levels and may pose a threat to the consumer if the product is not handled in a safe manner. Therefore, the production, transportation and sale of meat products must be performed with the almost care and preferably be subjected to HACCP evaluation, to prevent the presentation of any undue hazard (Madden, 1994). Hence, the importance of food as a vehicle in transmission of several diseases has been documented especially in developing countries where hygienic standards are not strictly followed or enforced (Harakeh et al., 2005). Foodborne diseases do not always result in acute gastroenteritis but also, food represents an important vehicle for pathogens (Flint et al., 2005).

The obtained results of aerobic platecount in Table 2 of chicken shawerma sandwiches were 5.5±3.2 log₁₀cfu/g these results were nearly in accordance with  those achieved by (Sharaf and Sabra, 2012) in cooked chicken shawerma which were 1.2x10⁵cfu/g and lower than the results recorded by (Salem et al., 2016) in chicken shawerma sandwiches which were 2.8x10⁶±1.0x10⁶cfu/g. this difference may be due to contamination of raw food, food handlers, utensils, unefficient cooking process and post cooking contamination.

Obviously, for the safety of shawerma sandwiches, it is necessary to cut a not more than 2-cm piece of fillet every time to be sure that these pieces are efficiently cooked and the latter is ready-made. Microbiological quality problems of Shawerma  sandwiches depends greatly on the quality of raw meat and other ingredients, efficiency of cooking process, proper sanitary practices for personnel and for cooking/processing utensils (Kayaardi et al., 2006). The achieved resultsof Staph. aureus in Table 2 declared  that the mean count of Staph. aureus in chicken shawerma sandwiches were 2.2± 1.6log₁₀cfu/g with incidence percent of 18% and the results of Fig (1) showed  that 3 out of 18 isolated strains were enterotoxigenic produced sed and see virulent genes.

These results were nearly similar to Hatakka (1998) who found that hot foods have been the source of outbreaks of Staph. aureus food poisoning and Nimri et al. (2014) examined chicken shawerma sandwiches for Staph. aureus which were 8.3% and some strains are capable of producing a heat-stable enterotoxin that causes food poisoning in humans also, Salem et al. (2016) found Staph. aureus in 26% of the examined samples. (Todd 2017) isolate Staph. aureus from shawerma and Chen et al. (2015) mentioned that Staph. aureus in cooked meat products were 4.84% in addition to (Hu ShouKui et al., 2013) Surveyed that the positive rates of Staph. aureus in cooked meat were 10.0% and 43.7% of the isolated strains produced enterotoxins while, Abou Hussein (2007) and Sharaf and Sabra (2012) can^,t detect Staph. aureus in cooked chicken shawerma. 

The main sources of pathogenic bacteria in food are contaminated raw food, food handlers, dust, water, utensils & insects (Ray, 1996). The results of examined chicken shawerma sandwiches in Table2 indicated that the mean count of E. coli were 3.3±1.8 log₁₀cfu/g with incidence percent of 22% and the results of Fig (2)  showed  that presence of stx1 in one isolate and the other isolates were not toxigenic. these results were higher than that obtained by Abou Hussein (2007) who mentioned that the incidence of E. coli in cooked chicken shawerma was 13.33% while nearly similar results obtained by Sharaf and Sabra (2012) who examined cooked chicken shawerma samples for E. coli count which were 3.9x10²cfu/g with an incidence percent 20%, while higher percentage were recently reported by (Nimri et al., 2014) examined shawerma (donair kebab) where the predominant species was E. coli (28.3%), with six isolates of serotype O157:H7 and Banna and Nawas (2016) tested chicken shawerma sandwiches for incidence of E. coli which were 20%.

Shinagawa (1990) and Sousa and Stamford (2013) stated that  Bacillus cereus is able to form spores, founded in ready meals and it is responsible for causing two distinct types of food poisoning: an emetic toxin pre-formed in food where cooked rice is the most common vehicle, the diarrhoeal type is caused by an enterotoxin. Its virulence factors include the production of haemolysins and phospholipases toxins and the symptoms are similar to those of Staph. aureus intoxication. The heat resistance of Bacillus cereus spores and the non fastidious nature of the organism facilitate its survival and/or growth in a wide variety of foods.

The achieved results in Table 2 declared that the mean count of Bacillus cereus were 2.4±1.9 log₁₀cfu/g with incidence percent of 32%, these results were higher than Chen et al. (2015) who detected Bacillus cereus in 12.66% of cooked meat products and lower than those recorded by Salem et al. (2016) who examined chicken shawerma sandwiches for the presence of Bacillus cereus incidence which were identified in 40% of examined samples.

Cooking grilled chicken fillet dish shawerma has been found to ensure its freedom from Salmonella only in a piece less than 2 cm thickness where Deeper layers of chicken and its juice that accumulates in the grill tray may remain be Salmonella-contaminated throughout the heat treatment (Sergevnin et al., 2012). The incidence results of Salmonellain the examined chicken shawerma sandwiches were 10% meanwhile, Salmonella typhimurium could not be detected. These results were lower than Nimri et al. (2014) who found Salmonella spp. In 26.3% from the examined Shawarma sandwiches and Banna and Nawas (2016) who tested chicken shawerma sandwiches for incidence of Salmonella which was 30%.

CONCLUSION

This study confirms that chicken shawerma sandwiches may serve as a source of foodborne pathogens and accordingly a potential public health hazard. Corrective action needs to be employed to minimize the risk of consuming this type of fast food, knowing that even small doses of the organism may lead to food poisoning Such action must aim at minimizing the bacterial contamination during the production of chicken shawarma (cleaning, cutting, seasoning and stacking), its cooking and serving. More attention should be given to the cleanliness of utensils used in preparing the sandwiches (rod, knives, etc.) in addition to the personal hygiene of the workers preparing and stuffing the sandwiches. New regulations that will ensure the safety of the consumer.

REFERENCES

Abou Hussein, R.A. (2007): Detection of food mediated pathogens in some meat and chicken products by using recent techniques. Ph. D. Thesis (Meat Hygiene), Faculty of Veterinary Medicine, Banha University, Egypt.

Ahmad, F.; Abbas, S.; Butt, Z.A.; Ali, A.S.; Mahmood, R.; Ahmad, M.; Farhan, M.; Nawaz, M.A. and Iftikhar, A. (2013): Bacterial contamination in processed chicken Shawarma (meat) sold in various parts of Lahore, Pakistan. Pakistan J. of Nutrition. 12 (2): 130-134.

APHA (American Public Health Association) (2001): Compendium Methods for the Microbiological Examination of Food. Washington DC.

Asao T.; Kumeda, Y. and Kawai, T. (2003): An extensive outbreak of staphylococcal food poisoning due to low-fat milk in Japan estimation of enterotoxin A in the incriminated milk and powdered skim milk. Epidemiol Infect. 130: 33-40.

Banna, Hanin and Nawas, T. (2016): Salmonella: A Common Contaminant of Chicken Shawarma in Ras Beirut Restaurants.  J. of Environmental Science, Toxicology and Food Technology.10, 8; 19-22.

Bennett, S.D.; Walsh, K.A. and Gould, L.H. (2008): Foodborne disease outbreaks caused by Bacillus cereus, Clostridium perfringens, and Staph. aureus-United States 1998-2008. J. Clin. Infect. Dis. 57(3): 425-433.

Berdgoll, M.S. (1989): Staph. aureus In Doyle MP. editor. Food-Borne Bacterial Pathogens. New York: Marcel Dekker. 464-523.

Braun, P.; Kirk, M.; Scallan, E.; Fitzgerald, M.;  Adak, GK.; Sockett, P.; Ellis, A.; Hall, G.; Gargouri, N.; Walke, H. and Braam, P. (2005): Estimating the burden of acute gastroenteritis, foodborne disease, and pathogens commonly transmitted by food: An international review. J. Clin. Infect. Dis. 41: 698-704.

Chen, Y.; Shen, L.; Pan, HongYu and Chen, X.Y. (2015): Monitoring of food borne pathogens of food in Panzhihua, 2011-2013. Modern Preventive Medicine. 42(21): 3871-3873.

Dipineto, L.; Santaniello, A.; Fontanella, M.; Lagos, K.; Fioretti, A. and Menna, L.F. (2006): Presence of Shiga toxin-producing E. coli O157:H7 in living layer hens. Letters in Applied Microbiology 43; 293–295.

Dominguez, S.A. and Schaffner, D.W. (2009): Survival of Salmonella in processed chicken products during frozen storage. J. Food Prot. 72: 2088-2092.

FDA (Food and Drug Administration) (2002): Enumeration of coliform bacteria and  E. coli. Bacteriological Analytical Manual. Chapter 4.

Flint, J.A.; Van Duynhoven, Y.T.; Angulo, F.J.; DeLong, S.M.; Okafo, C.N.; Umoh, V.J. and Galadima, M. (2010): Occurrence of pathogens on vegetables harvested from soils irrigated with contaminated streams. Sci. Total Environ. 311: 49-56.

Flint, J.A.; Van Duynhoven, Y.T.; Angulo, F.J.; DeLong, S.M.; Braun, P.; Kirk, M.; Scallan, E.; Fitzgerald, M.; Adak, G.K.; Sockett, P.; Ellis, A.; Hall, G.; Gargouri, N.; Walke, H. and Braam, P. (2005): Estimating the burden of acute gastroenteritis, foodborne disease, and pathogens commonly transmitted by food: An international review. J. Clin. Infect. Dis. 41: 698-704.

Ghafir, Y.; China, B.; Dierick, K.; De Zutter, L. and Daube, G. (2008): Hygiene indicator microorganisms for selected pathogens on beef, pork, and poultry meats in Belgium. J. Food Prot. 71: 35-45.

Harakeh, S.; Yassinea, H.; Ghariosb, M.; Barbourc, E.; Hajjara, S.; El-Fadeld, M.; Toufeilib, I. and Tannous, R. (2005): Isolation molecular characterization and antimicrobial resistance patterns of Salmonella and E. coli isolated from meat-based fast food in Lebanon. Sci. Total Environ 341: 33-44.

Hatakka, M. (1998): Microbiological quality of hot meals served by airlines. J. Food Protec. 61(8): 1052-1056. 

Hu ShouKui; Liu ShiYun; HuWan Fu; Zheng TianLi and XuJianGuo (2013): Molecular biological characteristics of Staph. aureus isolated from food. European Food Research and Technology. 236(2): 285-291.

ISO (2004): The International Organization for Standardization 2004a Horizontal method for the enumeration of presumptive Bacillus cereus-colony count technique at 30c. ISO 7932: 2004, Geneva, Switzerland.

ISO, 16649/2 (2001): Microbiology of Food and Animal Feeding Stuffs-Horizontal Method for the enumeration of beta - glucuronidase - positive E. coli: part 2: colony-count technique at 44^OC using 5-bromo -4-chloro-3-indoly beta-glucuronide.

ISO, 6579 (2002): Microbiology of Food and Animal Feeding Stuffs-Horizontal Method for the Detection of Salmonella spp.

ISO, 6887/2 (2003): Microbiology of Food and Animal Feeding Stuffs -Preparation of Test Samples, Initial Suspension and Decimal Dilutions for Microb. Exam Egypt. J. Chem. Environ. Health, 1 (1): 686-69.

Kayaardi, S.; Kayacier, Q.A. and Gok, V. (2006): Sensory and Chemical Analysis of Doner Kebab Made from Turkey Meat. J. Muscle Food. 17: 165-173.

Koneman, E.W.; Allen, S.D.; Dowell, V.R. and Sommers, H.M. (1997): Colour Atlas and Textbook of Diagnostic Microbiology. 2nd Ed, Lippincott, Philadelphia, New York, London.

Logue, C.M. and Nde, C.W. (2007): Salmonella contamination of turkey from processing to final product – A process to product perspective. Foodborne Pathogens and Disease, 4 (4), 491–504.    

Madden, N.K. (1994): Microbial hazards in animal Products. Proceeding of the Nutrition Society, 53: 309-312.

Mehrotra, M.; Wang, G. and Johnson, M.W. (2000): Multiplex PCR for detection of genes for Staph. aureus enterotoxin, exfoliative toxins, toxic shock syndrome toxin 1, and Methicillin resistance. J. of Clinical Microbiology, 38: 1032-1035.

Murugkar, H.V.; Rahman, H. and Dutta, P.K. (2003): Distribution of virulence genes in Salmonella serovars isolated from man & animals. Indian J. Med. Res., 117: 66-70.

Nimri, L.; Al-Dahab, F.A. and Batchoun, R. (2014): Foodborne bacterial pathogens recovered from contaminated shawarma meat in northern Jordan. J. of Infection in Developing Countries. 8(11): 1407-1414. 

Olivera, S.D.; Rodenbusch, C.R.; Ce, M.C.; Rocha, S.L.S. and Canal, C.W. (2003): Evaluation of selective and non selective en-richment PCR procedures for Salmonella de-tection. Lett. Appl. Microbiol., 36: 217-221.

Ostyn, A.; De Buyser, M.L.; Guillier, F.; Groult, J.; Felix, B; Salah, S.; Delmas, G. and Hennekinne, J.A. (2010): First evidence of a food poisoning outbreak due to staphylococcal enterotoxin type E, France, 2009. Euro Surveill 15.

Ray, B. (1996): Fundamental Food Microbiology. CRC Press, Inc. Tokyo, New York

Salem, Nehad I.E.; Adel M. El Gamel; Khalifa, Amal M.A.; Abd-El-Hady, Hala A.M. and Abou Zeid, Mayada A.M. (2016): Microbial Status of Shawerma Sandwiches in Kafr El-Sheikh Governorate. Alex. J. of Vet. Sc. 51(2): 303-309.

Sergevnin, V.I.; Udavikhina, L.S.; Gorokhova, S.V.; Istomina, L.F.; Khasanov, R.Kh.; Sarmometov, E.V.; Novoselov, V.G. (2012): Microbiological evaluation of chicken shawerma as a factor for Salmonella transmission. Gigiena i Sanitariya (1): 29-32. 

Sharaf, Eman M. and Sabra, Sherifa M. (2012): Microbiological Loads for Some Types of Cooked Chicken. J. of World Appl. Sc. 17 (5): 593-597.

Shinagawa, K. (1990): Analytical methods for Bacillus cereus and other Bacillus species. Int. J. Food Microbiol. 10 (2): 125-141.

Sousa, J.P.de and Stamford, T.L.M. (2013): Bacillus cereus: an important pathogen in food. Higiene Alimentar. 27(220/221): 88-93.

Todd, E.C.D. (2017): Foodborne disease and food control in the Gulf States. Food Control; 73(Part B): 341-366.

Zaki, Eman, M.S. and Shehata, Amal, A. (2008): Incidence of some enterotoxigenic food poisoning microorganisms in chicken meat products. Giza, Vet. Med. J.; 56(3): 255-266.