Characterization and Evaluation of Lactobacillus plantarum LC5.2 Isolated from Thai Native Pigs for its Probiotic Potential in Gut Microbiota Modulation and Immune Enhancement

Abstract

Probiotic supplementation, particularly with Lactobacillus species, enhances growth performance, maintains gastrointestinal microbial balance, and prevents infections in livestock. This study isolated Lactobacillus strains from the feces of healthy native pigs in southern Thailand and assessed their probiotic properties and safety through both in vitro and in vivo evaluations. Nine Lactobacillus strains showed probiotic potential, with Lactobacillus plantarum LC5.2 demonstrating the best characteristics. This strain tolerated both acid and bile (100% tolerance) and exhibited strong adhesion properties, including high auto-aggregation (69.74%), cell surface hydrophobicity (77.14%), adhesion to Caco-2 cells (9.31%), and biofilm formation. It also exhibited antibacterial activity, inhibiting EHEC, EPEC, and Salmonella Typhimurium through organic acid production. Co-aggregation with these pathogens ranged from 60.83% to 74.09%. Safety evaluations showed no hemolytic activity, susceptibility to antibiotics, and co-existence with other probiotics. In mice, L. plantarum LC5.2 showed no toxicity, with normal food intake, behavior, and weight gain. No abnormalities were found in the small intestine, colon, liver, or spleen. Mice administered the probiotic had significantly higher intestinal IgA levels. Gut microbiome analysis revealed no notable structural alterations but indicated an increase in beneficial bacteria, including Lactobacillus. These results suggest that L. plantarum LC5.2 demonstrates strong probiotic potential, safety, and benefits for gut health, making it a promising candidate for livestock applications.

Keywords: Lactobacillus plantarum; Thai native pigs; gut microbiota modulation; probiotic properties; safety assessment.

Fig. 1. Total secretory IgA levels in the small intestine of mice in the control and Lactobacillus plantarum LC5.2 groups.

Asterisk (*) indicates a significant difference (p < 0.05).

Fig. 2. Histological examinations of small intestine in mice. (A) Control and (B) Lactobacillus plantarum LC5.2 treatment group.

Paraffin-embedded tissues were sectioned and stained with hematoxylin and eosin (H&E) to assess histological changes. Hematoxylin stains cell nuclei blue, and eosin staining cytoplasm and extracellular matrix pink. Sections were viewed at 20× magnification.

Fig. 3. Histological examinations of colon in mice. (A) Control and (B) Lactobacillus plantarum LC5.2 treatment group.

Paraffin-embedded tissues were sectioned and stained with hematoxylin and eosin (H&E) to assess histological changes. Hematoxylin stains cell nuclei blue, and eosin staining cytoplasm and extracellular matrix pink. Sections were viewed at 20× magnification.

Fig. 4. Histological examinations of liver in mice. (A) Control and (B) Lactobacillus plantarum LC5.2 treatment group.

Paraffin-embedded tissues were sectioned and stained with hematoxylin and eosin (H&E) to assess histological changes. Hematoxylin stains cell nuclei blue, and eosin staining cytoplasm and extracellular matrix pink. Sections were viewed at 20× magnification.

Fig. 5. Histological examinations of spleen in mice. (A) Control and (B) Lactobacillus plantarum LC5.2 treatment group.

Paraffin-embedded tissues were sectioned and stained with hematoxylin and eosin (H&E) to assess histological changes. Hematoxylin stains cell nuclei blue, and eosin staining cytoplasm and extracellular matrix pink. Sections were viewed at 20× magnification.

Fig. 6. Relative abundance of bacteria at all taxonomic levels in the fecal microbiota of mice administered Lactobacillus plantarum LC5.2 compared to the control (PBS) group.

Stacked bar chart showing the relative abundance of bacterial taxa at all classification levels (phylum to species) in the fecal microbiota of mice treated with Lactobacillus plantarum LC5.2 compared to the PBS control group. Each color represents different bacterial taxa identified in the samples. Taxonomic profiles were generated from 16S rRNA gene sequencing data and the figure was created using MicrobiomeAnalyst.

Fig. 7. LEfSe comparison of differentially abundant bacterial taxa between mice treated with Lactobacillus plantarum LC5.2 and the control group.

Blue bars indicate taxa enriched in the control group, red bars in the LC5.2 group, with bar length reflecting the LDA score.

Fig. 8. Alpha diversity of the fecal microbiota in Lactobacillus plantarum LC5.2-treated and control (PBS) mice, based on Shannon, Simpson, and Chao1 indices.

The median is shown by the line inside each box, with whiskers extending to the min and max values. Outliers and individual samples are represented by dots.

Fig. 9. Beta diversity of the microbiome between Lactobacillus plantarum LC5.2-treated and control (PBS) mice, based on Bray–Curtis distance.

Red dots represent LC5.2, and blue dots represent control.

Fig. 10. Beta diversity of the microbiome between Lactobacillus plantarum LC5.2-treated and control (PBS) mice, based on Jaccard distance.

Red dots represent LC5.2, and blue dots represent control.