Effects of Bacillus amyloliquefaciens LFB112 on Growth Performance, Carcass Traits, Immune, and Serum Biochemical Response in Broiler Chickens

This study aimed to investigate the effects of Bacillus amyloliquefaciens LFB112 on the growth performance, carcass traits, immune response, and serum biochemical parameters of broiler chickens. A total of 396 1 day old, mixed-sex commercial Ross 308 broilers with similar body weights were allotted into six treatment groups. The assigned groups were the CON group (basal diet with no supplement), AB (antibiotics) group (basal diet + 150 mg of aureomycin/kg), C+M group (basal diet + 5 × 108 CFU/kg B. amyloliquefaciens LFB112 powder with vegetative cells + metabolites), C group (basal diet + 5 × 108 CFU/kg B. amyloliquefaciens LFB112 vegetative cell powder with removed metabolites), M group (basal diet + 5 × 108 CFU/kg B. amyloliquefaciens LFB112 metabolite powder with removed vegetative cells), and CICC group (basal diet + 5 × 108 CFU/kg Bacillus subtilis CICC 20179). Results indicated that chickens in the C+M, C, and M groups had higher body weight (BW) and average daily gain (ADG) (p < 0.05) and lower feed conversion ratio (FCR) (p = 0.02) compared to the CON group. The C+M group showed the lowest abdominal fat rate compared to those in the CON, AB, and CICC groups (p < 0.05). Compared to the CON group, serum IgA and IgG levels in the C+M, C, and M groups significantly increased while declining in the AB group (p < 0.05). B. amyloliquefaciens LFB112 supplementation significantly reduced the serum triglyceride, cholesterol, urea, and creatinine levels, while increasing the serum glucose and total protein (p < 0.05). In conclusion, B. amyloliquefaciens LFB112 significantly improved the growth performance, carcass traits, immunity, and blood chemical indices of broiler chickens and may be used as an efficient broiler feed supplement

Expression, Purification and Characterization of a Novel Hybrid Peptide CLP with Excellent Antibacterial Activity

CLP is a novel hybrid peptide derived from CM4, LL37 and TP5, with significantly reduced hemolytic activity and increased antibacterial activity than parental antimicrobial peptides. To avoid host toxicity and obtain high-level bio-production of CLP, we established a His-tagged SUMO fusion expression system in Escherichia coli. The fusion protein can be purified using a Nickel column, cleaved by TEV protease, and further purified in flow-through of the Nickel column. As a result, the recombinant CLP with a yield of 27.56 mg/L and a purity of 93.6% was obtained. The purified CLP exhibits potent antimicrobial activity against gram+ and gram- bacteria. Furthermore, the result of propidium iodide staining and scanning electron microscopy (SEM) showed that CLP can induce the membrane permeabilization and cell death of Enterotoxigenic Escherichia coli (ETEC) K88. The analysis of thermal stability results showed that the antibacterial activity of CLP decreases slightly below 70 °C for 30 min. However, when the temperature was above 70 °C, the antibacterial activity was significantly decreased. In addition, the antibacterial activity of CLP was stable in the pH range from 4.0 to 9.0; however, when pH was below 4.0 and over 9.0, the activity of CLP decreased significantly. In the presence of various proteases, such as pepsin, papain, trypsin and proteinase K, the antibacterial activity of CLP remained above 46.2%. In summary, this study not only provides an effective strategy for high-level production of antimicrobial peptides and evaluates the interference factors that affect the biological activity of hybrid peptide CLP, but also paves the way for further exploration of the treatment of multidrug-resistant bacterial infections

Yeast Expressed Hybrid Peptide CLP Abridged Pro-Inflammatory Cytokine Levels by Endotoxin Neutralization

The aim of this study was to apply a strategy to express a recombinant CLP peptide and explore its application as a product derived from natural compounds. The amphiphilic CLP peptide was hybridized from three parent peptides (CM4, LL37, and TP5) and was considered to have potent endotoxin-neutralizing activity with minimal cytotoxic and hemolytic activity. To achieve high secretion expression, an expression vector of pPICZαA-HSA-CLP was constructed by the golden gate cloning strategy before being transformed into Pichia pastoris and integrated into the genome. The recombinant CLP was purified through the Ni-NTA affinity chromatography and analyzed by SDS-PAGE and mass spectrometry. The Limulus amebocyte lysate (LAL) test exhibited that the hybrid peptide CLP inhibited lipopolysaccharides (LPS) in a dose-dependent manner and was significantly (p p p < 0.05) with the treatment of CLP. Consequently, we concluded that the hybrid peptide CLP might be used as a therapeutic agent