BACKGROUND: Fall armyworm (Spodoptera frugiperda) is a highly destructive maize pest that significantly threatens agricultural productivity. Existing control methods, such as chemical insecticides and entomopathogens, lack effectiveness, necessitating alternative approaches.
METHODS: Gut-associated bacteria were isolated from the gut samples of fall armyworm and screened based on their chitinase and protease-producing ability before characterization through 16S rRNA gene sequence analysis. The efficient chitinase-producing Bacillus licheniformis FGE4 and Enterobacter cloacae FGE18 were chosen to test the biocontrol efficacy. As their respective cell suspensions and extracted crude chitinase enzyme, these two isolates were applied topically on the larvae, supplemented with their feed, and analyzed for their quantitative food use efficiency and survivability.
RESULTS: Twenty-one high chitinase and protease-producing bacterial isolates were chosen. Five genera were identified by 16S rRNA gene sequencing: Enterobacter, Enterococcus, Bacillus, Pantoea, and Kocuria. In the biocontrol efficacy test, the consumption index and relative growth rate were lowered in larvae treated with Enterobacter cloacae FGE18 by topical application and feed supplementation. Similarly, topical treatment of Bacillus licheniformis FGE4 to larvae decreased consumption index, relative growth rate, conversion efficiency of ingested food, and digested food values.
CONCLUSIONS: The presence of gut bacteria with high chitinase activity negatively affects insect health. Utilizing gut-derived bacterial isolates with specific insecticidal traits offers a promising avenue to control fall armyworms. This research suggests a potential strategy for future pest management.

Introduction: Bacillus licheniformis (B.licheniformis) was widely used in poultry feeds. However, it is still unclear about how B.licheniformis regulates the growth and development of Pekin ducks. Methods: The experiment was designed to clarify the effect and molecular mechanism of B. licheniformis on the lipid metabolism and developmental growth of Pekin ducks through multiomics analysis, including transcriptomic and metabolomic analyses. Results: The results showed that compared with the control group, the addition of 400 mg/kg B. licheniformis could significantly increase the body weight of Pekin ducks and the content of triglyceride (p < 0.05), at the same time, the addition of B. licheniformis could affect the lipid metabolism of liver in Pekin ducks, and the addition of 400 mg/kg B. licheniformis could significantly increase the content of lipoprotein lipase in liver of Pekin ducks. Transcriptomic analysis revealed that the addition of B. licheniformis primarily impacted fatty acid and glutathione, amino acid metabolism, fatty acid degradation, as well as biosynthesis and elongation of unsaturated fatty acids. Metabolomic analysis indicated that B. licheniformis primarily affected the regulation of glycerol phospholipids, fatty acids, and glycerol metabolites. Multiomics analysis demonstrated that the addition of B. licheniformis to the diet of Pekin ducks enhanced the regulation of enzymes involved in fat synthesis via the PPAR signaling pathway, actively participating in fat synthesis and fatty acid transport. Discussion: We found that B. licheniformis effectively influences fat content and lipid metabolism by modulating lipid metabolism-associated enzymes in the liver. Ultimately, this study contributes to our understanding of how B. licheniformis can improve the growth performance of Pekin ducks, particularly in terms of fat deposition, thereby providing a theoretical foundation for its practical application. Conclusion: B. licheniformis can increase the regulation of enzymes related to fat synthesis through PPAR signal pathway, and actively participate in liver fat synthesis and fatty acid transport, thus changing the lipid metabolism of Pekin ducks, mainly in the regulation of glycerol phospholipids, fatty acids and glycerol lipid metabolites.

Tibetan tea changes during microorganism fermentation. Research on microorganisms in Tibetan tea has focused on their identification, while studies on the influence of specific microorganisms on the components and health functions of Tibetan tea are lacking. Bacillus licheniformis was inoculated into Tibetan tea for intensive fermentation, and the components of B. licheniformis-fermented tea (BLT) were detected by liquid chromatography with tandem mass spectrometry (UHPLC-TOF-MS), and then the effects of BLT on intestinal probiotic functions were investigated by experiments on mice. The results revealed the metabolites of BLT include polyphenols, alkaloids, terpenoids, amino acids, and lipids. Intensified fermentation also improved the antioxidant capacity in vivo and the protective effect on the intestinal barrier of Tibetan tea. In addition, the enhanced fermentation of Tibetan tea exerted intestinal probiotic effects by modulating the relative abundance of short-chain fatty acid-producing bacteria in the intestinal flora. Therefore, intensive fermentation with B. licheniformis can improve the health benefits of Tibetan tea.

Aflatoxin B1 (AFB1) contamination is a serious threat to nutritional safety and public health. The CotA-laccase from Bacillus licheniformis ANSB821 previously reported by our laboratory showed great potential to degrade AFB1 without redox mediators. However, the use of this CotA-laccase to remove AFB1 in animal feed is limited because of its low catalytic efficiency and low expression level. In order to make better use of this excellent enzyme to effectively degrade AFB1, twelve mutants of CotA-laccase were constructed by site-directed mutagenesis. Among these mutants, E186A and E186R showed the best degradation ability of AFB1, with degradation ratios of 82.2% and 91.8% within 12 h, which were 1.6- and 1.8-times higher than those of the wild-type CotA-laccase, respectively. The catalytic efficiencies (kcat/Km) of E186A and E186R were found to be 1.8- and 3.2-times higher, respectively, than those of the wild-type CotA-laccase. Then the expression vectors pPICZαA-N-E186A and pPICZαA-N-E186R with an optimized signal peptide were constructed and transformed into Pichia pastoris GS115. The optimized signal peptide improved the secretory expressions of E186A and E186R in P. pastoris GS115. Collectively, the current study provided ideal candidate CotA-laccase mutants for AFB1 detoxification in food and animal feed and a feasible protocol, which was desperately needed for the industrial production of CotA-laccases.

Enhancing the aerobic stability of whole-plant corn silage is essential for producing high-quality silage. Our research assessed the effect of inoculation with Lactobacillus buchneri or Bacillus licheniformis and its modulation of the bacterial and fungal microbial community structure in an aerobic stage of whole-plant corn silage. Following treatment with a distilled sterile water control, Lactobacillus buchneri, and Bacillus licheniformis (2 × 105 cfu/g), whole-plant corn was ensiled for 60 days. Samples were taken on days 0, 3, and 7 of aerobic exposure, and the results showed that inoculation with Lactobacillus buchneri or Bacillus licheniformis improved the aerobic stability of silage when compared to the effect of the control (p < 0.05). Inoculation with Bacillus licheniformis attenuated the increase in pH value and the decrease in lactic acid in the aerobic stage (p < 0.05), reducing the filamentous fungal counts. On the other hand, inoculation with Lactobacillus buchneri or Bacillus licheniformis increased the diversity of the fungal communities (p < 0.05), complicating the correlation between bacteria or fungi, reducing the relative abundance of Acetobacter and Paenibacillus in bacterial communities, and inhibiting the tendency of Monascus to replace Issatchenkia in fungal communities, thus delaying the aerobic spoilage process. Due to the prevention of the development of aerobic spoilage microorganisms, the silage injected with Lactobacillus buchneri or Bacillus licheniformis exhibited improved aerobic stability.

This study investigates the co-precipitation of calcium and barium ions in hypersaline wastewater under the action of Bacillus licheniformis using microbially induced carbonate precipitation (MICP) technology, as well as the bactericidal properties of the biomineralized product vaterite. The changes in carbonic anhydrase activity, pH, carbonate and bicarbonate concentrations in different biomineralization systems were negatively correlated with variations in metal ion concentrations, while the changes in polysaccharides and protein contents in bacterial extracellular polymers were positively correlated with variations in barium concentrations. In the mixed calcium and barium systems, the harvested minerals were vaterite containing barium. The increasing concentrations of calcium promoted the incorporation and adsorption of barium onto vaterite. The presence of barium significantly increased the contents of O-CO, N-CO, and Ba-O in vaterite. Calcium promoted barium precipitation, but barium inhibited calcium precipitation. After being treated by immobilized bacteria, the concentrations of calcium and barium ions decreased from 400 and 274 to 1.72 and 0 mg/L (GB/T15454-2009 and GB8978-1996). Intracellular minerals were also vaterite containing barium. Extracellular vaterite exhibited bactericidal properties. This research presents a promising technique for simultaneously removing and recycling hazardous heavy metals and calcium in hypersaline wastewater.

Glycoside hydrolase family 5 (GH5) encompasses enzymes with several different activities, including endo-1,4-β-mannosidases. These enzymes are involved in mannan degradation, and have a number of biotechnological applications, such as mannooligosaccharide prebiotics production, stain removal and dyes decolorization, to name a few. Despite the importance of GH5 enzymes, only a few members of subfamily 7 were structurally characterized. In the present work, biochemical and structural characterization of Bacillus licheniformis GH5 mannanase, BlMan5_7 were performed and the enzyme cleavage pattern was analyzed, showing that BlMan5_7 requires at least 5 occupied subsites to perform efficient hydrolysis. Additionally, crystallographic structure at 1.3 Å resolution was determined and mannoheptaose (M7) was docked into the active site to investigate the interactions between substrate and enzyme through molecular dynamic (MD) simulations, revealing the existence of a - 4 subsite, which might explain the generation of mannotetraose (M4) as an enzyme product. Biotechnological application of the enzyme in stain removal was investigated, demonstrating that BlMan5_7 addition to washing solution greatly improves mannan-based stain elimination.

Bacillus licheniformis and bacitracin have a huge application market and value in the fields of medicine, chemistry, aquaculture, agricultural, and sideline products. Therefore, the selection of B. licheniformis with high production of bacitracin is of great importance. In this experimental protocol, Bacillus with a high yield of bacitracin was isolated, purified, and identified from the fresh feces of healthy pigs. The inhibitory effect of secondary metabolite bacitracin on Micrococcus luteus was also tested. Thin-layer chromatography and high-performance liquid chromatography were used for the qualitative and quantitative detection of bacitracin. The physiological and biochemical characteristics of B. licheniformis were determined by relevant kits. The phylogenetic relationships of B. licheniformis were determined and constructed using gene sequence detection. This protocol describes and introduces the standard isolation, purification, and identification process of B. licheniformis from animal fresh feces from multiple perspectives, providing a method for the large-scale utilization of B. licheniformis and bacitracin in factories.

Candida albicans, a significant human pathogenic fungus, employs hydrolytic proteases for host invasion. Conventional antifungal agents are reported with resistance issues from around the world. This study investigates the role of Bacillus licheniformis extracellular proteins (ECP) as effective antifungal peptides (AFPs). The aim was to identify and characterize the ECP of B. licheniformis through LC-MS/MS and bioinformatics analysis. LC-MS/MS analysis identified 326 proteins with 69 putative ECP, further analyzed in silico. Of these, 21 peptides exhibited antifungal properties revealed by classAMP tool and are predominantly anionic. Peptide-protein docking revealed interactions between AFPs like Peptide chain release factor 1 (Q65DV1_Seq1: SASEQLSDAK) and Putative carboxy peptidase (Q65IF0_Seq7: SDSSLEDQDFILESK) with C. albicans virulent SAP5 proteins (PDB ID 2QZX), forming hydrogen bonds and significant Pi-Pi interactions. The identification of B. licheniformis ECP is the novelty of the study that sheds light on their antifungal potential. The identified AFPs, particularly those interacting with bonafide pharmaceutical targets SAP5 of C. albicans represent promising avenues for the development of antifungal treatments with AFPs that could be the pursuit of a novel therapeutic strategy against C. albicans. SIGNIFICANCE OF STUDY: The purpose of this work was to carry out proteomic profiling of the secretome of B. licheniformis. Previously, the efficacy of Bacillus licheniformis extracellular proteins against Candida albicans was investigated and documented in a recently communicated manuscript, showcasing the antifungal activity of these proteins. In order to achieve high-throughput identification of ES (Excretory-secretory) proteins, the utilization of liquid chromatography tandem mass spectrometry (LC-MS) was utilized. There was a lack of comprehensive research on AFPs in B. licheniformis, nevertheless. The proteins secreted by B. licheniformis in liquid medium were initially discovered using liquid chromatography-tandem mass spectrometry (LC-MS) analysis and identification in order to immediately characterize the unidentified active metabolites in fermentation broth.

Bacillus licheniformis is a gram-positive, endospore-forming, saprophytic and facultative anaerobe that is resistant to heat and environmental conditions. This study was the first to isolate and confirm B. licheniformis as a cause of bovine mastitis in Iran. In the summer of 2020, 105 samples of mastitic milk were collected from dairy farms around Tehran and sent to the microbiology laboratory of the Faculty of Veterinary Medicine at the University of Tehran. The bacterial pathogens were identified using selective and differential culture media and confirmed by PCR to contain the toxin synthetase genes licA, licB and licC in mastitic isolates of B. licheniformis. Resistance patterns to 19 antibiotics were determined for two isolates of B. licheniformis. Staphylococcus aureus and Escherichia coli were identified as the most important organisms in the samples. B. licheniformis was isolated from the two samples containing all three genes. Both isolates were resistant to streptomycin, trimethoprim-sulfamethoxazole, cefixime, ampicillin, bacitracin, clindamycin, and gentamicin. B. licheniformis was reported for the first time in Iran as a cause of bovine mastitis with clinical symptoms. The first isolation of toxin-producing strains of B. licheniformis from mastitic cows in Iran raises concerns about the safety of dairy products. In principle, selected strains with toxigenic potential should not be used as feed additives and animal feed. However, whole genome sequencing is proposed to search for genes coding for toxins.