جداسازی و شناسایی مولکولی باکتری لاکتوباسیلوس پلانتاروم از محصولات لبنی سنتی
محورهای موضوعی : میکروب شناسی صنعتی مواد غذائی
مریم کاظمی میرکی
1
,
الهام معظمیان
2
,
محمد جواد مختاری
3
,
mehrdad gholamzad
4
1 - دانشجو
2 - 2. گروه میکروبیولوژی، دانشکده علوم، کشاورزی و فناوری های نوین، واحد شیراز، دانشگاه آزد اسلامی، شیراز، ایران
3 - استادیار، دانشکده علوم، دانشگاه آزاد اسلامی واحد زرقان، زرقان. ایران
4 - Tehran Islamic Azad university of medical science
کلید واژه: باکتری اسیدلاکتیک, لاکتوباسیلوس پلانتاروم, مواد لبنی, شناسایی مولکولی, تست بیوشیمیایی ,
چکیده مقاله :
لاکتوباسیلوس پلانتاروم یک باکتری مهم در صنایع غذایی است که به عنوان یک سویه مفید و کاربردی شناخته می‌شود. پتانسیل بالقوه ی پروبیوتیکی و توانایی ایجاد ایمنی از جمله مزیت‌های آنها می‌باشد. در این مقاله، جداسازی و شناسایی باکتری لاکتوباسیلوس پلانتاروم از مواد لبنی مورد بررسی قرار گرفته است. 40 نمونه از مواد لبنی سنتی از مناطق مختلف استان فارس جمع‌آوری شد. نمونه ها در محیط کشت MRSتلقیح و مورفولوژی کلنی و شکل آنها بررسی شد. سپس رنگ آمیزی گرم و شناسایی ویژگی‌های بیوشیمیایی مانند تست کاتالاز، اکسیداز، تخمیر قندها و سیترات انجام شدند. در نهایت، با استفاده از روش مولکولی واکنش زنجیره‌ای پلی مراز، تکثیر و توالی‌یابی ژن16S rRNA انجام شد. با استفاده از روش‌های تشخیصی و بیوشیمیایی، 11 جدایه‌ی باکتری لاکتوباسیلوس با موفقیت جداسازی شد و پس از واکنش زنجیره‌ای پلی‌مراز و توالی‌یابی، 4 جدایه لاکتوباسیلوس پلانتاروم تایید شدند. هر 4 جدایه گرم مثبت، کاتالاز ، اکسیداز و سیترات منفی بودند و گلوکز، مانوز، ساکاروز و سوربیتول را تخمیر می‌کردند. تشخیص نهایی لاکتوباسیلوس پلانتاروم از طریق شناسایی مولکولی و تعیین توالی ژن16S rRNA تأیید شد. نتایج نشان می‌دهد که باکتری لاکتوباسیلوس پلانتاروم در مواد لبنی وجود دارد و می‌توان با استفاده از بررسی شکل، ساختار، روش‌های بیوشیمیایی و مولکولی، این گونه‌ی مهم را از مواد لبنی جداسازی کرد. همچنین، استفاده از روش‌های مورد استفاده در این تحقیق امکان جداسازی لاکتوباسیلوس پلانتاروم را فراهم می‌کند.
Lactobacillus plantarum is an important bacterium in the food industry recognized as a useful and practical strain. The advantages of lactic acid bacteria, including Lactobacillus species, are their potential probiotic properties and their ability to induce immune responses. This study investigated the isolation and identification of Lactobacillus plantarum from dairy products. Forty samples of traditional dairy products were collected from different regions of the Fars province. The samples were cultured on MRS agar media and their colony morphology and shape were investigated. Then, Gram staining and biochemical tests such as catalase, oxidase, sugar fermentation, and citrate utilization were performed to identify the biochemical characteristics of Lactobacillus species. Finally, using molecular methods such as polymerase chain reaction (PCR) targeting the 16S rRNA gene and sequencing the desired sequence, the genetic identification of Lactobacillus species was performed. Using diagnostic and biochemical methods, 11 Lactobacillus isolates were successfully isolated, and after polymerase chain reaction and sequencing, 4 isolates were confirmed as Lactobacillus plantarum. All 4 isolates were Gram-positive, catalase-negative, oxidase-negative, and citrate-negative, and they fermented glucose, mannose, sucrose, and sorbitol. The final diagnosis of this bacterium was confirmed by molecular identification and sequencing of the 16S rRNA gene. The results indicate the presence of Lactobacillus plantarum in dairy products and it is possible to isolate this important species from dairy products by examining the shape, biochemical and molecular methods. Furthermore, the use of the methods employed in this study as diagnostic indicators enables isolation of Lactobacillus plantarum.
Abadi, M. E. G. M., Hosseini-Safa, A., Habibi, S., Dehghan, M., Forouzani-Moghaddam, M. J., & Oshaghi, M. (2023). Isolation and characterization of the lactobacillus strain from honey and its probiotic properties. Iranian journal of microbiology, 15(3), 439.
Asaadi, H., Eshaghi Milasi, Y., & Yazdansetad, S. (2019). Isolation and molecular identification of carotenoid-producing bacteria. Nova Biologica Reperta, 6(1), 61-69.
Behera, S. S., Ray, R. C., & Zdolec, N. (2018). Lactobacillus plantarum with functional properties: an approach to increase safety and shelf‐life of fermented foods. BioMed research international, 2018(1), 9361614.
Cretin, B., Philippi, N., Bousiges, O., Dibitonto, L., Sellal, F., Martin-Hunyadi, C., & Blanc, F. (2017). Do we know how to diagnose epilepsy early in Alzheimer's disease? Revue neurologique, 173(6), 374-380.
da Silva Sabo, S., Vitolo, M., González, J. M. D., & de Souza Oliveira, R. P. (2014). Overview of Lactobacillus plantarum as a promising bacteriocin producer among lactic acid bacteria. Food Research International, 64, 527-536.
Davati, N. (2018). Isolation and Identification of Indigenous Lactic Acid Bacteria from Traditional Yogurt Produced from Ewe's Milk from Alvand Nomads Region and Evaluation of Their Acidifying Potential. Journal of food science and technology(Iran), 15(74), 222-213.
Dehghani Champiri, I., Bamzade, Z., Rahimi, E., & Rouhi, L. (2023). Isolation of Lactic Acid Bacteria from Local Yogurt Samples in Chaharmahal and Bakhtiari Province: The Assessment of Probiotic Characteristics. Iran J Med Microbiol 17(6), 669-679.
Dorri, K., Namdar, N., & HemayatkhahJahromi, V. (2013). Isolation of lactobacilli from dairy products and their effects on the main pathogenic bacteria in stomach and intestine. Medical Laboratory Journal, 7(1), 22-28.
Dubernet, S., Desmasures, N., & Guéguen, M. (2002). A PCR-based method for identification of lactobacilli at the genus level. FEMS microbiology letters, 214(2), 271-275.
Elagöz, A., Abdi, A., Hubert, J.-C., & Kammerer, B. (1996). Structure and organisation of the pyrimidine biosynthesis pathway genes in Lactobacillus plantarum: a PCR strategy for sequencing without cloning. Gene, 182(1-2), 37-43.
Emami, A., & Noeiaghdam, R. (2008). Cholesterol Assimilation with Isolated lactobacilli Strains of Fars’ Local Dairy Products. Armaghane Danesh, 13(3), 45-56.
Guidone, A., Zotta, T., Ross, R. P., Stanton, C., Rea, M. C., Parente, E., & Ricciardi, A. (2014). Functional properties of Lactobacillus plantarum strains: A multivariate screening study. LWT-Food Science and Technology, 56(1), 69-76.
Haghshenas, B., Nami, Y., Almasi, A., Abdullah, N., Radiah, D., Rosli, R., Barzegari, A., & Khosroushahi, A. Y. (2017). Isolation and characterization of probiotics from dairies. Iranian journal of microbiology, 9(4), 234.
Hassan, A. A. M., Sakr, S. S., Ali, A. A., Mohamed Ahmed, I. A., & Elkashef, H. (2023). Isolation, identification, and biochemical characterization of five Lacticaseibacillus strains from Oggtt: A traditional fermented and dried buttermilk. Food Science & Nutrition, 11(2), 1040-1050.
Hazards, E. P. o. B., Ricci, A., Allende, A., Bolton, D., Chemaly, M., Davies, R., Girones, R., Koutsoumanis, K., Lindqvist, R., & Nørrung, B. (2018). Update of the list of QPS‐recommended biological agents intentionally added to food or feed as notified to EFSA 7: suitability of taxonomic units notified to EFSA until September 2017. Efsa Journal, 16(1), e05131.
Hou, Q., Bai, X., Li, W., Gao, X., Zhang, F., Sun, Z., & Zhang, H. (2018). Design of primers for evaluation of lactic acid bacteria populations in complex biological samples. Frontiers in Microbiology, 9, 2045.
Khushboo, Karnwal, A., & Malik, T. (2023). Characterization and selection of probiotic lactic acid bacteria from different dietary sources for development of functional foods [Original Research]. Frontiers in Microbiology, 14.
Koohsari, H., Rashti, Z., & Arab, S. (2019). The Isolation of lactic acid bacteria from local dairy products of Gorgan township with the ability to inhibit the growth of some gastrointestinal pathogens. Journal of Food Microbiology, 6(3), 22-36.
Li, P., Gu, Q., Yang, L., Yu, Y., & Wang, Y. (2017). Characterization of extracellular vitamin B12 producing Lactobacillus plantarum strains and assessment of the probiotic potentials. Food chemistry, 234, 494-501.
Liu, C., Xue, W.-j., Ding, H., An, C., Ma, S.-j., & Liu, Y. (2022). Probiotic potential of Lactobacillus strains isolated from fermented vegetables in Shaanxi, China. Frontiers in Microbiology, 12, 774903.
Naji, M., Norouzi, J., & Saffarian, P. (2021). Antimicrobial properties of lactobacilli isolated from soya on E. coli and Staphylococcus aureus. Research in Medicine: Journal of Research in Medical Sciences, 45(2).
Narimani, T., Tarinejad, A., & Hejazi, M. A. (2013). Isolation and identification of lactic acid bacteria from traditional dairy products of Kleibar, Heris and Varzaghan. Food Hygiene, 3(Autumn 3(11)), 23-37.
Nouri, S., Nazari, S., & Hosseyni, P. (2018). Isolation and Biochemical and Molecular Identification of Lactobacillus Plantarum Bacteria from Rhizosphere of Lenjan Rice. Biological Journal of Microorganism, 7(27), 61-71.
Padasht, N., Issazadeh, K., & Shah Illi, M. (2023). Cytotoxic Effects of Lactobacillus Cytoplasmic Extract isolated from Guilan province dairy products on Colon Cancer Cell Line (HT-29). Biological Journal of Microorganism, 12(46), 13-25.
Panda, S. K., Ray, R. C., Mishra, S. S., & Kayitesi, E. (2018). Microbial processing of fruit and vegetable wastes into potential biocommodities: a review. Critical Reviews in Biotechnology, 38(1), 1-16.
Rahi, A., & Marhamatizadeh, M. H. (2022). Investigating the Inhibitory Effect of Probiotic Lactobacillus Species Isolated From Dairy Products. Avicenna Journal of Clinical Microbiology and Infection, 9(3), 103-108.
Ray, R. C., & Joshi, V. (2014). Fermented foods: past, present and future. Microorganisms and fermentation of traditional foods, 1-36.
Shuhadha, M., Panagoda, G., Madhujith, T., & Jayawardana, N. (2017). Evaluation of probiotic attributes of Lactobacillus sp. isolated from cow and buffalo curd samples collected from Kandy. The Ceylon medical journal, 62(3), 159-166.
Silva, L. F., Casella, T., Gomes, E. S., Nogueira, M. C. L., De Dea Lindner, J., & Penna, A. L. B. (2015). Diversity of lactic acid bacteria isolated from Brazilian water buffalo mozzarella cheese. Journal of food science, 80(2), M411-M417.
Sun, J., Silander, O., Rutherfurd-Markwick, K., Wen, D., Davy, T. P.-p., & Mutukumira, A. N. (2022). Phenotypic and genotypic characterisation of Lactobacillus and yeast isolates from a traditional New Zealand Māori potato starter culture. Current Research in Food Science, 5, 1287-1294.
Swain, M. R., & Ray, R. C. (2016). Nutritional values and bioactive compounds in lactic acid fermented vegetables and fruits. Lactic acid fermentation of fruits and vegetables, 1, 37-52.
Tadesse, B. T., Tesfaye, A., Muleta, D., Bahiru, A., Terefework, Z., & Wessel, G. (2018). Isolation and molecular identification of lactic acid bacteria using 16s rRNA genes from fermented Teff (Eragrostis tef (Zucc.)) dough. International journal of food science, 2018(1), 8510620.
Taye, Y., Degu, T., Fesseha, H., & Mathewos, M. (2021). Isolation and identification of lactic acid bacteria from cow milk and milk products. The Scientific World Journal, 2021(1), 4697445.
Uppada, S. R., Akula, M., Bhattacharya, A., & Dutta, J. R. (2017). Immobilized lipase from Lactobacillus plantarum in meat degradation and synthesis of flavor esters. Journal of Genetic Engineering and Biotechnology, 15(2), 331-334.
Urshev, Z., Doynova, D., Prasev, I., Denkova-Kostova, R., Koleva, A., Denkova, Z., Goranov, B., & Kostov, G. (2024). Identification of Lactic Acid Bacteria Strains Isolated from Sourdoughs Prepared with Different Flour Types. Applied Sciences, 14(5), 2093.
Vantsawa, P. A., Maryah, U. T., & Bulus, T. (2017). Isolation and identification of lactic acid bacteria with probiotic potential from fermented cow milk (nono) in Unguwar Rimi Kaduna State Nigeria. American Journal of Molecular Biology, 7(2), 99-106.
Xue, Z., Brooks, J. T., Quart, Z., Stevens, E. T., Kable, M. E., Heidenreich, J., McLeod, J., & Marco, M. L. (2021). Microbiota assessments for the identification and confirmation of slit defect-causing bacteria in milk and Cheddar cheese. Msystems, 6(1), 10.1128/msystems. 01114-01120.
Yi, R., Tan, F., Liao, W., Wang, Q., Mu, J., Zhou, X., Yang, Z., & Zhao, X. (2019). Isolation and identification of Lactobacillus plantarum HFY05 from natural fermented yak yogurt and its effect on alcoholic liver injury in mice. Microorganisms, 7(11), 530.
Zafar mokhtarian, E., Rezazadeh Bari, M., & Amiri, S. (2018). Isolation and molecular identification of exopolysaccharide producing lactic acid bacteria from sheep milk and yogurt. Journal of food science and technology(Iran), 15(79), 243-252.