ارزيابي اثر پرتو فرابنفش بر رشد اشريشيا كلي و باسيلوس سريوس جدا شده از شيرخام و برنج خام

Study the effects of ultraviolet radiation on the growth of Escherichia coli and Bacillus cereus isolated from raw milk and raw rice

اشريشياكلي و باسيلوس سريوس علاوه بر ايجاد فساد در مواد غذايي مي توانند در مصرف‌كنندگان مواد غذايي حاوي مقدار كافي از اين باكتري ها مسموميت ايجاد كنند. به همين دليل حذف و يا كنترل اين باكتري ها هميت پيدا مي كند. در اين مطالعه اثر ضد باكتريايي پرتو فرا بنفش در مدت زمان‌هاي مختلف، بر رشد اشريشيا كلي و باسيلوس سريوس مورد ارزيابي قرار گرفت. بدين منظور ابتدا اشريشيا كلي و باسيلوس سريوس را به ترتيب از شير خام و برنج جداسازي و سپس به ترتيب در محيط كشت 4EMB و5 MYP به مدت 24 تا 48 ساعت كشت داده شدند؛ در مرحله ي بعد پس از انجام آزمايشات تاييدي مناسب، كشت خالص مربوط به هر يك از باكتري ها تهيه شد. بررسي‌ها نشان داد كه بيشترين اثر پرتو فرابنفش بر باكتري ها در طول موج حدود 260 نانومتر است. مشاهده شد كه باسيلوس سريوس نسبت به اشريشيا كلي مقاومت بيشتري به پرتو فرابنفش نشان مي دهد به شكلي كه تعداد كلني هاي اشريشيا كلي پس از 80 ثانيه قرار گرفتن در مجاورت با طول موج 254 نانومتر، به صفر نزديك مي شود اما در مورد باسيلوس سريوس پس از اين مدت به 1500 مي‌رسد؛ و نيز با افزايش مدت زمان پرتو دهي و در غياب نور مريي تعداد ميكروارگانيسم ها بيشتر كاهش مي يابد.

Introduction: One of the most important aspects of food preservation is controlling the growth of microorganisms, which if overlooked it leads to uncontrolled growth of microorganisms associated with food spoilage and food poisoning. Microbial contamination of foods is important because of pathogens are capable to transfer to foods during the processing, distribution, and storage. Escherichia coli and Bacillus cereus can cause spoilage in food; intake food contains plenty of bacteria and toxic. Therefore it is important to eliminate or control these bacteria safely. Ultraviolet (UV) radiation is considered as non-ionizing radiation and the first time in 1940 was used as a method for infection elimination in air. This approach nowadays is widely used for controlling microbial growth and as disinfection in food industry. Wavelength range of ultraviolet radiation is approximately 328-210 nm. The beam is naturally present in the sunlight. The bactericidal effect of UV irradiation depends on the type of bacteria, the distance, and dose of radiation. The most cytotoxic effect of UV irradiation is obtained at the wavelength of 260 nm, which corresponds to the intense absorption of energy by organic bases in the nucleic acid. UV irradiation causes radicals generation, which subsequently attack the nucleic acid and develop mutations in their genomes and gene transcription and translation processes. In this study, the antibacterial effect of different exposure times of ultraviolet radiation on the growth of E. coli and B. cereus was evaluated. Materials and methods: All media used in this study was procured from Merck Company. Ultraviolet device (Camag, USA) was used at wavelength of 254 nm, Nr= 29000, Amp= 0.25. B. cereus isolation: 1 mL of different dilutions of rice (0.1, 0.01, and 0.001) was transferred to Brain-heart infusion (BHI) and it was incubated at 32 °C for 24 h. A loop containing the bacteria was then transferred to Mannitol Egg Yolk Polymyxin (MYP) agar and it was incubated at 35 °C for 24 h. B. cereus produce big and round colonies, with a halo around the colonies. Starch test was carried out as confirming test for B. cereus colonies. Briefly, some colonies of B. cereus were added to test tube with sterile distilled water containing starch and a few drops of lugol. Development of blue color indicates the presence of B. cereus due to starch hydrolysis. E. coli isolation: E. coli was isolated from raw milk following the method described by Kargar et al. (2005). Briefly, raw milk was first homogenized; 0.1 ml of each dilution of homogenized raw milk was inoculated on Escherichia coli broth medium containing 20 mg novobiocin. E. coli was then isolated after transferring the former media on EMB specific culture and incubation at 36 °C for 24 h. After confirming colonies by Durham tube and complementary tests, pure cultures were obtained from them by streak-plate method. UV irradiation: A loop of E. coli colonies was transferred to nutrient broth and it was treated with UV beam (254 nm) at three times (40, 60, and 80 s). After preparing dilution of 0.0001 for each of the treatments and incubating for 24 h, survival curve was plotted. These operations were also carried out on B. ceruse colonies. A control sample also was considered for each examined bacterium. Results and Discussion: Rate of Bacillus cereus growth was reduced under UV radiation. As it is shown, Death curve of E. coli, E. coli count was decreased by increasing the time of UV radiation, so that count of this bacteria reached to about zero after UV radiation for 80 s. However, reduction of B. cereus count was less than E. coli count at same wavelength (254 nm) and time of irradiation. This revealed that B. cereus have more resistance to UV radiation compared to E. coli. These results were consistent to observation of Sharp (1940) who evaluate the effects of UV light on bacteria suspended in air and reported that required energy for air sterilization containing B. cereus is more than twice the energy is needed to eliminate E. coli. UV light more penetrates to cell wall of Gram-negative bacteria compared to Gram-positive bacteria due to having a small amount of peptidoglycan in the cell wall and caused mutations in regulating genes of transcription and translation. Conclusion: The efficiency of the two main processes of cell is reduced in the presence of UV irradiation and leads to growth reduction and death. The more resistance of B. cereus can be for several reasons, such as having a thicker cell wall compared to E. coli, and the capability to produce spore, and the capability to proofing mutations. Keywords: Escherichia coli, Bacillus cereus, Raw milk, Rice, Ultraviolet.