Lactobacillus delbrueckii subsp. bulgaricus KLDS 1.0207 Exerts Antimicrobial and Cytotoxic Effects in vitro and Improves Blood Biochemical Parameters in vivo Against Notable Foodborne Pathogens

Abstract

Globally, foodborne diseases (FBDs) result in millions of sicknesses and deaths annually. Cumulative evidence suggests that the use of probiotic lactic acid bacteria (LAB) strains could be a viable alternative in inhibiting the activities of foodborne pathogens. This study aims to evaluate the in vitro antimicrobial, cytotoxic, and tolerance levels of Lactobacillus bulgaricus KLDS 1.0207 against two notable foodborne pathogens - Escherichia coli ATCC25922 and Staphylococcus aureus ATCC25923. Afterward, a 48 BALB/c mice-trial was used to assess its ameliorative effects on weight and serum biochemical parameters. Results showed that the cell-free supernatant (CFS) of this strain significantly inhibited both pathogens, but these effects were abolished at pH 6.5 and 7.0 (P < 0.05). Also, 6.96 ± 0.02 log CFU mL^-1 of L. bulgaricus KLDS 1.0207 was still viable after three hours in simulated gastric juice and at pH 3.0, indicating that this strain was a potential probiotic candidate. Also, inflammatory activities in RAW264.7 cells were significantly inhibited using 10⁹ CFU mL^-1 of L. bulgaricus KLDS 1.0207 cells (P < 0.05). Significant weight losses were also prevented in the T _LBSA (from 19.42 ± 1.04 to 19.55 ± 0.55 g) and T _LBEC (from 22.86 ± 0.90 to 14.77 ± 9.86 g) groups compared to their respective model groups (T _SA - from 21.65 ± 1.80 to 20.14 ± 1.84, and T _EC - from 21.45 ± 0.82 to 14.45 ± 9.70 g). Besides, there was a slight weight gain in the S. aureus prevention group (T _LBSA ) compared to the model group (T _SA ). Serum biochemical analyses revealed that the total cholesterol (TC), triglycerides (TG), low-density lipoprotein (LDL), and some mineral levels were markedly increased by S. aureus and E. coli administrations but were reversed to normalcy in both prevention groups (T _LBSA and T _LBEC ). Interestingly, high-density lipoprotein (HDL) levels, which were initially disrupted in the model groups, were restored in the prevention groups (T _LBSA and T _LBEC ). This study presents L. bulgaricus KLDS 1.0207 as a promising probiotic candidate with antimicrobial, anti-inflammatory, acid, and bile tolerant and lipid-regulating applications. It also gives valuable insights for targeted future in vivo treatment and prevention studies involving other probiotic LAB candidates. Future in vivo studies elucidating specific mechanisms behind the in vitro antimicrobial, cytotoxic, and in vivo ameliorative effects are warranted.

Keywords: HDL; L. bulgaricus; LDL; anti-inflammation; biochemical; probiotics; tolerance; weight.

FIGURE 1

In vitro cytotoxic effects of different concentrations (CFU mL^–1) of L. bulgaricus KLDS 1.0207 culture in RAW264.7 cells, Control - RAW264.7 cells; T1 - RAW264.7 cells + 1 × 10⁵ L. bulgaricus KLDS 1.0207; T2 - RAW264.7 cells + 1 × 10⁶ L. bulgaricus KLDS 1.0207; T3 - RAW264.7 cells + 1 × 10⁷ L. bulgaricus KLDS 1.0207; T4 - RAW264.7 cells + 1 × 10⁸ L. bulgaricus KLDS 1.0207; T5 - RAW264.7 cells + 1 × 10⁹ L. bulgaricus KLDS 1.0207, All values were obtained in triplicate and expressed as Mean ± SD. **Significant difference compared to the control (P < 0.05).

FIGURE 2

Weekly weights (g) of study animals before and after S. aureus ATCC25923 infection. The final weights of study animals after two weeks suggest that L. bulgaricus KLDS 1.0207 effectively prevented weight loss caused by S. aureus administration (P > 0.05).

FIGURE 3

Weekly weights (g) of study animals before and after E. coli ATCC25922 infection. After two weeks of study, slight improvements were observed in the prevention group (T_LBEC) compared to the model group (T_EC) (P > 0.05). **Significant difference compared to the control (P < 0.05).

FIGURE 4

Aspartate aminotransferase (AST), alanine aminotransferase (ALT), total protein (TP), and albumin (ALB) levels (U/L) of BALB/c mice in the control, and all trial groups. All values were obtained in triplicate and expressed as Mean ± SD.

FIGURE 5

Total glucose (TG), total cholesterol (TC), high-density lipoprotein (HDL), and low-density lipoprotein (LDL) levels (mmol/L) of BALB/c mice in the control and all trial groups. All values were obtained in triplicate and expressed as Mean ± SD.

FIGURE 6

Mineral levels of BALB/c mice in the control and all trial groups. All values were obtained in triplicate and expressed as Mean ± SD.