Pangenome Analysis and Genome-Guided Probiotic Evaluation of Cyclic Dipeptides Producing Levilactobacillus brevis DY55bre Strain from a Lactic Acid Fermented Shalgam to Assess Its Metabolic, Probiotic Potentials, and Cytotoxic Effects on Colorectal Cancer Cells

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Yetiman A. E., Horzum M., Kanbur E., Çadir M., Bahar D., Gürbüz Ş., ...Daha Fazla

Probiotics and Antimicrobial Proteins, 2025 (SCI-Expanded, Scopus) identifier identifier identifier identifier

  • Yayın Türü: Makale / Tam Makale
  • Basım Tarihi: 2025
  • Doi Numarası: 10.1007/s12602-025-10760-7
  • Dergi Adı: Probiotics and Antimicrobial Proteins
  • Derginin Tarandığı İndeksler: Science Citation Index Expanded (SCI-EXPANDED), Scopus, BIOSIS, CAB Abstracts, Food Science & Technology Abstracts, MEDLINE, Veterinary Science Database
  • Anahtar Kelimeler: Cholesterol, Antimicrobial activity, Pangenome, Probiotic, Anticancer, <italic>Levilactobacillus</italic>
  • Açık Arşiv Koleksiyonu: AVESİS Açık Erişim Koleksiyonu
  • Erciyes Üniversitesi Adresli: Evet

Özet

This study investigates the genetic, metabolic, and probiotic characteristics of Levilactobacillus brevis DY55bre, a strain isolated from the traditional Turkish fermented beverage, shalgam. Whole-genome sequencing revealed a circular genome of 2.485 Mb with a GC content of 45.72%, predicted 2791 genes, and multiple CRISPR-Cas systems. Pangenome analysis demonstrated an open structure, with 18.9% core genes and 103 strain-specific genes, highlighting its genetic diversity. The DY55bre exhibits heterofermentative carbohydrate metabolism due to the presence of the araBAD operon and the lack of 1-phosphofructokinase (pfK) and fructose-1,6-bisphosphate aldolase enzymes. Probiotic evaluation revealed firm survival under simulated gastrointestinal conditions, including resistance to acidic pH (as low as 3.0) and bile salts (up to 1%), along with significant adhesion to intestinal epithelial cell lines (HT29;59.3%, Caco-2;87%, and DLD-1;60.8%). The strain exhibited high auto-aggregation (84.55%) and cell surface hydrophobicity (56.69%), essential for gut colonization. Safety assessments confirmed its non-hemolytic nature and absence of horizontally acquired antibiotic resistance genes. Notably, GC-MS analysis identified bioactive cyclic dipeptides, Cyclo(D-Phe-L-Pro) and Cyclo(L-Leu-L-Pro), which demonstrated cytotoxic effects against colorectal cancer cell lines, with IC50 values of 7.71 mg/mL for HT29 and 3.19 mg/mL for DLD-1. The cell-free supernatant exhibited antimicrobial activity against pathogens, likely due to the synergistic effects of cyclic dipeptides, organic acids, and other metabolites. Antioxidant assays revealed significant ABTS+ (76.63%) and DPPH (34.25%) radical scavenging activities, while cholesterol assimilation tests showed a 27.29% reduction. These findings position the DY55bre as a promising candidate for functional foods, nutraceuticals, and therapeutic applications, warranting further in vivo validation.