Sheep-Derived Lactobacillus johnsonii M5 Enhances Immunity and Antioxidant Capacity, Alleviates Diarrhea, and Improves Intestinal Health in Early-Weaned Lambs

Abstract

The early weaning of lambs frequently leads to weakened immunity, impaired intestinal function, and increased susceptibility to intestinal disease. Lactobacillus plays a role in regulating immunity, enhancing antioxidant capacity, and maintaining intestinal health. This study aims to isolate a strain of Lactobacillus with favorable probiotic properties from sheep feces and investigate its effects on the intestinal health of early-weaned lambs. In this study, the growth characteristics, acid production capacity, bacteriostatic capacity, bile salt tolerance, gastrointestinal fluid tolerance, self-coagulation capacity, and surface hydrophobicity of Lactobacillus isolated from sheep feces were analyzed for in vitro probiotic properties. Lactobacilli with strong probiotic properties were used for in vivo validation. A total of 72 Hu lambs were allocated into four groups: a ewe-reared group (ER), early-weaning group (EW), low-dose Lactobacillus group (LL), and high-dose Lactobacillus group (HL). Early weaning was performed in the EW, LL, and HL groups at the age of 28 days. Lactobacillus johnsonii M5 (L. johnsonii M5), isolated from sheep feces, exhibited strong probiotic properties in vitro. Feeding EW lambs with a low dose of L. johnsonii M5 significantly reduced their diarrhea rate (p < 0.05). Its supplementation increased the levels of superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GSH-Px) and total antioxidant capacity (T-AOC) in serum and jejunal mucosa and decreased levels of malondialdehyde (MDA) (p < 0.05). Compared to the EW group, serum immunoglobulin G (IgG) levels were significantly increased in the LL group (p < 0.05). Compared to the EW group, feeding with L. johnsonii M5 increased the content of anti-inflammatory cytokines, while reducing the content of pro-inflammatory cytokines in serum and jejunal mucosa (p < 0.05). Feeding early-weaned lambs with L. johnsonii M5 also decreased jejunal crypt depth and increased occludin and claudin-1 in jejunal mucosa (p < 0.05). These findings indicate that feeding early-weaned lambs with L. johnsonii M5 enhances their immunity and antioxidant capacity, improving intestinal health, and mitigates diarrhea in early-weaned lambs.

Keywords: Lactobacillus; antioxidant; early weaning; immunity; lambs.

Figure 1

(A) Colony morphology of Lactobacillus strains isolated from sheep feces on MRS agar medium and the microscopy results of Gram staining. (a) Strain M5; (b) strain M13; (c) strain M15; (d) strain M19. (B) Phylogenetic tree derived from 16S rRNA gene sequence of Lactobacillus isolated from sheep feces.

Figure 2

(A) Growth curves of the four Lactobacillus strains isolated from sheep feces after 24 h. (B) Acid production curves of the four Lactobacillus strains isolated from sheep feces after 24 h.

Figure 3

(A) Inhibitory zone diameters of four strains of Lactobacillus isolated from sheep feces against E. coli, S. aureus, and E. enteritidis. Survival of the four strains of Lactobacillus in (B) 0.3% bile salts, (C) artificial gastric juice at pH 3.0, and (D) artificial intestinal juice at pH 6.8. (E) Hydrophobicity and (F) auto-agglutination rate of the four Lactobacillus strains. The same superscript letters indicate no significant difference (p > 0.05), while different letters indicate significant differences (p < 0.05).

Figure 4

Hemolytic results of the four Lactobacillus strains isolated from sheep feces.

Figure 5

Effects of different doses of L. johnsonii M5 on antioxidant indices in weaned lambs (n = 6). MDA, malondialdehyde; SOD, superoxide dismutase; CAT, catalase; GSH-Px, peroxidase, glutathione; T-AOC, total antioxidant capacity. In a box plot, the same letters above the box indicate no significant difference (p > 0.05), while different letters indicate significant differences (p < 0.05). The levels of (A) MDA, (B) SOD, (C) CAT, (D) T-AOC and (E) GSH-Px in serum of lambs in each group. The levels of (F) MDA, (G) SOD, (H) CAT, (I) T-AOC and (J) GSH-Px in jejunal mucosa in each group.

Figure 6

Effects of different doses of L. johnsonii M5 on serum immune parameters of weaned lambs (n = 6). IgA, immunoglobulin A; IgG, immunoglobulin G; IgM, immunoglobulin M; TNF-α, tumor necrosis factor-α; IL-1β, interleukin-1β; IL-6, interleukin-6; IL-10, interleukin-10. In a box plot, the same letters above the box indicate no significant difference (p > 0.05), while different letters indicate significant differences (p < 0.05). The levels of (A) IgG, (B) IgA, (C) IgM, (D) TNF-α, (E) IL-1β, (F) IL-6 and (G) IL-10 in serum of lambs in each group. The levels of (H) TNF-α, (I) IL-1β, (J) IL-4, (K) IL-6, (L) IL-10, (M) IL-17 in jejunal mucasa of lambs in each group.

Figure 7

Effects of different doses of L. johnsonii M5 on jejunal histomorphology of weaned lambs (n = 6). VH:CD, villus height/crypt depth. In a box plot, the same letters above the box indicate no significant difference (p > 0.05), while different letters indicate significant differences (p < 0.05). The (A) villus height, (B) crypt depth and (C) VH:CH of the jejunum in each group of lambs.

Figure 8

Effects of different doses of L. johnsonii M5 on tight junction protein of weaned lambs (n = 6). ZO-1, Zonula Occludens-1. In a box plot, the same letters above the box indicate no significant difference (p > 0.05), while different letters indicate significant differences (p < 0.05). The levels of (A) ZO-1, (B) claudin-1, (C) occludin in jejunal mucasa of lambs in each group.