Razi Journal of Medical Sciences

fa اثر سویه‌های پروبیوتیکی معده زنبور عسل در کاهش گلوکز سرم نمونه های دیابتی در شرایط آزمایشگاهی Effect of Honey Bee Stomach Probiotic Strains on in vitro Glucose lowering in Diabetic Serum Samples میکروبیولوژی Microbiology پژوهشي Research <div style="text-align: justify;">زمینه و هدف: دیابت نوع 2 یک بیماری چند عاملی است که عوامل ژنتیکی و محیطی می‌توانند در ایجاد آن نقش داشته باشند. با شیوع دیابت در سراسر جهان لزوم کاربرد درمان‌های نوین مورد توجه قرار گرفته است. هدف این مطالعه، بررسی توانایی سویه‌های پروبیوتیک جداشده از معده زنبور عسل در کاهش غلظت گلوکز در سرم بیماران دیابتی نوع 2 در شرایط آزمایشگاهی  بود. روش کار: باکتری‌های پروبیوتیک از معده 20 زنبور عسل جداسازی شدند و با استفاده از آزمون‌های بیوشیمیایی، تست مقاومت آنتی بیوتیکی، ارزیابی توانایی تخمیر دکستروز بعنوان منبع کربن، مورد بررسی قرار گرفتند. جهت شناسایی و تایید سویه های جداسازی شده، استخراج DNA و بررسی مولکولی بر اساس 16srRNA انجام شد. اثر پروبیوتیک های مورد مطالعه، بر کاهش گلوکز سرم جدا شده از 50 بیمار دیابت نوع 2، در شرایط ازمایشگاهی بررسی شد. داده‌ها آنالیز آماری شدند. یافته‌ها: از معده 20 زنبور عسل، 30 ایزوله پروبیوتیک جداسازی شد. بر اساس تست‌های بیوشیمیایی و روش مولکولی، ایزوله‌ها شامل لاکتوباسیلوس اسیدوفیلوس، لاکتوباسیلوس پلانتاروم، لوکونوستوک مزنتروئیدس و باسیلوس سوبتیلیس بودند که اثر معنی‌داری را درکاهش گلوکز در بازه زمانی 6 ساعت پس از تلقیح باکتری به سرم خون بیماران دیابتی نوع 2، در شرایط آزمایشگاهی نشان دادند (05/0>p). نتیجه‌گیری: نتایج نشان داد که پروبیوتیک‌های معده زنبور عسل، می توانند در شرایط آزمایشگاهی پتانسل کاهش گلوکز سرم بیماران دیابتی را داشته باشند. برای تایید اثرات درمانی در بیماران دیابتی، مطالعات بالینی گسترده با حجم نمونه بیشتر و تنوع سویه‌های فراوان‌تر، توصیه می‌گردد.</div> <div style="text-align: justify;">Background & Aims: Type 2 diabetes mellitus is a multifactorial disease with different causes, consist of genetic damage, environmental disorders and lifestyle risk factors (1-6). Other funders to the pathogenesis of this disease contain entrance to healthy food and changes in the gut microbiota. Alterations in gut microbiota in diabetic patients can damage to glycemic control and show significantly roles in glucose metabolism. Therefore, could have arisen new treatment programs (11-21). Conversely, current researches demonstrated that probiotics, mainly the lactobacillus species, hold antioxidant effects and may be control and treatment diabetes by controlling the gut microbiota in patients (15-30). General studies have presented that probiotics as, Lactobacillus acidophilus, Lactobacillus casei, and plantarum incorporate major antidiabetic properties (33-40). The use of probiotics represents a promising intervention. Specific probiotic strains, particularly from the Lactobacillus genus have demonstrated significant potential in improving blood glucose regulation and reducing oxidative stress through their interaction with the gut microbiome. Honey, a natural substance, has been identified as a potential vector for these beneficial microbes, as it can contain viable probiotic bacteria, including those originating from the bee gut. This research aims were isolation and identification probiotic bacteria from the stomach of honey bees that exhibit antibacterial properties. So, Investigation and validation the in vitro glucose-lowering efficacy of these isolated probiotic strains when introduced to diabetic serum samples. This work focuses on antidiabetic potential of probiotics. Methods: This study employed a comprehensive multi-step process, contain of: 1. Isolation and Preliminary Identification: Source: Stomachs of 20 honey bees. Culture Medium: De Man, Rogosa and Sharpe (MRS) agar, selective for Lactic Acid Bacteria. Biochemical and Morphological Tests:  Gram staining: Isolates were likely Gram-positive rods. Catalase test: Isolates were likely catalase-negative. Oxidase test: Isolates were likely oxidase-negative. Carbohydrate fermentation: Used to create a metabolic profile for identification. Growth at different temperatures, motility, acid and bile salt resistance. 2. Assessment of Probiotic Properties: Antibacterial Activity: The isolated probiotics showed a statistically significant (P < 0.05) ability to inhibit the growth of three major pathogens (P. mirabilis, E. coli, S. aureus). The triple-repeat measurements ensure the results are reliable and reproducible. Acid and Bile Tolerance: Isolates were confirmed to survive the harsh conditions of the human gastrointestinal tract. Antibiotic Resistance: The profile was determined using the CLSI standard method. 3. Specialized Functional Assessment (Diabetes Focus): Glucose Tolerance: The ability of the bacteria to consume glucose at different concentrations was tested in vitro. Clinical Application: This property was directly linked to a clinical study involving 50 diabetic patients. The results, presented as Mean ± SEM, showed that the probiotic application led to a statistically significant (p < 0.05) reduction in blood sugar levels compared to a control group. 4. Molecular Identification: DNA Extraction and PCR: The 16S rRNA gene, the gold standard for bacterial identification, was amplified. Sequencing and Phylogenetics: By sequencing the PCR product and comparing it to the NCBI database via BLAST, the researchers conclusively identified the isolated strain(s) to the species level. 5. Statistical Analysis: Software: SPSS. Test: One-way ANOVA followed by Tukey's post-hoc test. This is appropriate for comparing the means of more than two groups. Significance: A P-value < 0.05 confirms that the observed differences were very unlikely to be due to random chance. Results: In the current research, 30 insulated of probiotic bacteria which had lactobacillus species characters were identified from Stomachs of 20 honey bees’ samples. Findings include: 1.Isolation and Identification:30 bacterial isolates with characteristics of Lactobacillus species were obtained from 20 honey bee samples. They were initially identified using standard morphology and biochemical tests. 2.Probiotic Properties: Antimicrobial Activity: All 30 isolates demonstrated the ability to inhibit the growth of the pathogenic bacteria Proteus mirabilis. Antibiotic Sensitivity: The isolates showed high sensitivity (50%) to penicillin and ampicillin but complete resistance (100%) to ceftazidime. Tolerance to Gut Conditions: All isolates survived simulated gut conditions, specifically acidic environments and the presence of bile salts. 3.Glucose-Lowering Effect (In Vitro): Out of the 30 isolates, 9 showed a significant ability to reduce glucose levels in a solution (dextrose powder) at various concentrations over a 6-hour period. Crucially, these 9 strains also significantly lowered blood glucose levels in vitro in serum samples taken from 50 diabetic patients. This effect was observed over 6 hours and was statistically significant (p < 0.05). 4.Molecular Confirmation: Molecular analysis (16S rRNA gene sequencing) was used to precisely identify the 9 most effective strains. They were found to be: Lactobacillus acidophilus, Lactobacillus plantarum, Leuconostoc mesenteroides, Bacillus subtilis. This molecular confirmation validated the initial biochemical identification methods. Conclusion: This study's in vitro results demonstrate that the isolated and identified several strains of bacteria from honey bees that exhibit strong probiotic potential. These probiotic strains effectively reduced blood sugar levels in samples from diabetic patients. They pronounced ability to lower glucose levels in vitro, including in diabetic serum samples. Thus, these bee-derived probiotics could be promising candidates for further research into managing blood sugar levels, potentially leading to applications for diabetes management.  Current research is focused on the ability of various probiotic strains, to regulate blood glucose levels in diabetic patients (15-20).  However, presents several challenges that must be addressed (29-36). Key limitations include: Strain-Specific Effects: Different probiotic strains possess unique biological and metabolic properties and interact with the host's immune system in distinct ways. Consequently, their effects can vary significantly across different populations. Delivery Method: The vehicle of administration can influence the efficacy and integration into standardized treatment regimens. Dosage and Duration: Determining the optimal dosage, treatment duration, and the ideal timing for intervention remains unclear and appears to vary between strains. In conclusion, while this study indicates that probiotics may assist in diabetes treatment, their application requires rigorous validation. Future research must include long-term human studies to confirm efficacy, establish safety profiles, and rule out any potential toxicity associated with prolonged use. There is a critical need for large-scale, placebo-controlled clinical trials on diabetic patients from diverse populations, utilizing various  probiotic strains under standardized protocols to realize their therapeutic potential.</div> باکتری پروبیوتیک, زنبورعسل, دیابت نوع 2, کاهش گلوکز Probiotic Bacteria, Honey Bee, Type 2 Diabetes, Glucose Lowering 1 17 http://rjms.iums.ac.ir/browse.php?a_code=A-10-8659-1&slc_lang=fa&sid=1 Elham Siasi الهام سیاسی emi_biotech2006@yahoo.ca 3900319475328460091337 3900319475328460091337 Yes Effect of Honey Bee Stomach Probiotic Strains on in vitro Glucose lowering in Diabetic Serum Samples دانشیار، گروه میکروبیولوژی، واحد تهران شمال، دانشگاه آزاد اسلامی، تهران، ایران Mohaddeseh Karimi Nori محدثه کریمی نوری 3900319475328460091338 3900319475328460091338 No MSc degree, Department of Microbiology, NT.C., Islamic Azad University, Tehran, Iran کارشناسی ارشد، گروه میکروبیولوژی، واحد تهران شمال، دانشگاه آزاد اسلامی، تهران، ایران Laya Takbiri لعیا تکبیری 3900319475328460091339 3900319475328460091339 No Assistant Professor, Department of Microbiology, NT.C., Islamic Azad University, Tehran, Iran استادیار، گروه میکروبیولوژی، واحد تهران شمال، دانشگاه آزاد اسلامی، تهران، ایران