Search Results (192)

Jackfruit seeds, a by-product of the jackfruit processing industry, comprise a substantial proportion of starch. As a result, jackfruit seeds are emerging as a viable source of fermentable sugars for fermentation processes. In this study, α-amylase from Bacillus licheniformis TKU004 was employed to hydrolyze gelatinized jackfruit seed starch slurry, and the hydrolysis conditions were systematically optimized using the Box–Behnken design (BBD) coupled with response surface methodology (RSM). Three independent variables, including incubation temperature (40–60 °C), enzyme-to-substrate ([E]/[S]) ratio (5–10 U/g), and reaction time (2–6 h), were evaluated, with dextrose equivalent (DE, %) as the response. The optimal hydrolysis parameters were determined to be 47 °C, an [E]/[S] ratio of 10 U/g, and a reaction time of 5.1 h, yielding a predicted DE of 31.72%. Experimental validation confirmed a DE of 32.85 ± 1.12%, in close agreement with the model prediction. HPLC (high-performance liquid chromatography) analysis of the hydrolysate revealed a composition of 14.20% glucose, 56.51% maltose, and 29.29% maltooligosaccharides, indicating that this process is well-suited for producing high-maltose syrup. In short, this study demonstrates the feasibility of valorizing jackfruit seed waste into value-added carbohydrate products through enzymatic hydrolysis with B. licheniformis α-amylase. Full article

Bacteria in concrete has been studied as an additive to repair microcracks and reduce permeability, as well as increase compressive strength. Within the broad spectrum of bacteria, two types promise to be effective agents against external sulfate attack: (i) Bacillus subtilis, which could indirectly prevent the entry of sulfates through the mechanism of sealing by calcium precipitation, and (ii) Bacillus licheniformis, which could encapsulate the sulfates that enter by diffusion and prevent the consequences of the pathology, such as expansion and loss of strength. This research evaluates the impact of B. subtilis and B. licheniformis on the performance of cementitious mixes against external sulfate attack, measuring compressive strength, expansion, permeability, and effects on the microstructure. Results show that both bacteria can produce compressive strength improvements of up to 20% at 28 days and 50% at 180 days. Moreover, in the presence of sulfates, improvements of up to 90% can be observed over control mixes. However, this result should be carefully evaluated because although B. licheniformis produces better results in the long term, it results in lower strength in the presence of sulfates in the short term. At the same time, B. licheniformis significantly reduces expansion against external sulfate attack, decreasing it by up to 80%, because it generates less ettringite and gypsum. Thus, B. licheniformis is an effective agent against external sulfate attack. Based on the results, it is estimated that both bacteria can be used to improve performance; however, care must be taken with concentration, which affects homogeneity or generates negative effects. In particular, it is noteworthy that calcium carbonate loss was observed from the mixes due to continuous curing and that calcium precipitation can generate negative effects against sulfates in the long term. Full article

In recent years, bacteria-based self-healing has emerged as a promising bioengineering strategy to address the self-repair of cracks in cement-based materials, which represent one of the persistent durability challenges. This approach relies on microbiologically induced calcium carbonate (CaCO₃) precipitation (MICP), in which metabolically active bacteria promote CaCO₃ formation of crystals that can heal cracks and restore material integrity. This study compares the self-healing potential of a natural (N-) alkaline soil Bacillus licheniformis strain with a UV-strain (phenotypic mutant) generated through controlled UV exposure followed by adaptive evolution. Both strains were evaluated under conditions relevant to cementitious environments. The UV-strain exhibited enhanced ureolytic performance, reaching urease activity of 0.32 U/mg compared to 0.24 U/mg in the N-strain. This translated into improved biomineralization, with CaCO₃ precipitation reaching 2.37 mg versus 2.23 mg/100 mL in the N-strain. Additionally, the UV-strain showed increased cell hydrophobicity and aggregation, indicating improved nucleation potential and surface-mediated mineral deposition. Multivariate analysis confirmed strong correlations between ureolytic metabolism, alkalization, and mineral formation, while artificial neural network (ANN) modeling (MLP 6-10-14) successfully predicted biomineralization-related parameters with high accuracy (R² > 0.90 for urease activity, NH₄⁺, ΔpH, and CaCO₃). The results demonstrate that UV-induced phenotypic adaptation can enhance biomineralization efficiency with minor trade-offs in physiological robustness. For the first time, that controlled UV-induced phenotypic adaptation can be used as a targeted strategy to enhance biomineralization efficiency in B. licheniformis, while maintaining functional stability under cement-relevant conditions. These findings provide a novel framework for tailoring bacterial performance in self-healing systems for construction biotechnology. Full article

Bacteria adapted to elevated temperatures are commonly associated with geothermal environments and are recognized for their functional diversity. In this study, cultivable bacteria were isolated from a geothermal spring in northern Sinaloa, Mexico, and characterized through physicochemical analysis, molecular identification, growth kinetics, and functional screening. The isolates were identified as Bacillus licheniformis (strains J1, J3, and J8) and Brevibacillus borstelensis (strains J6 and J9). Growth analyses showed that, in nutrient broth at 45 °C, the evaluated strains exhibited specific growth rates ranging from 1.25 to 1.78 h⁻¹ and short doubling times between 23 and 33 min, with B. borstelensis J6 displaying the highest rate. At 50 °C, μmax values ranged from 0.77 to 1.08 h⁻¹, indicating sustained growth at elevated temperatures. Functional assays demonstrated extracellular proteolytic, amylolytic, and cellulolytic activities, mainly associated with B. licheniformis strains, in addition to tolerance to the pesticides fluazinam and benomyl. Antagonistic tests showed that B. licheniformis J8 inhibited the phytopathogenic fungi Sclerotinia sclerotiorum and Sclerotium rolfsii, while qualitative mineral solubilization assays indicated the ability of selected isolates to mobilize phosphate and potassium. These findings highlight geothermal ecosystems as valuable reservoirs of thermotolerant bacteria with enzymatic versatility and environmental relevance, supporting further molecular and process-optimization studies. Full article

Sporulation represents a complex metabolic reprogramming process in bacteria. In this study, we used CRISPR-Cpf1 to knock out spoIIE and rsfA in Bacillus licheniformis. The ΔspoIIE strain completely lost sporulataion capacity, while ΔrsfA showed a 25% reduction. Although viable cell counts decreased by 80.7% and 45.7%, respectively, glucose consumption and 2,3-butanediol synthesis remained unchanged, and acetoin synthesis increased by 19% in ΔspoIIE. Per-cell metabolic rates were significantly enhanced: glucose uptake increased 2.7–3.4-fold, acetoin synthesis 2.3–4.2-fold, 2,3-butanediol synthesis 1.7-fold, and heterologous protein expression 10–15-fold. These findings demonstrate that blocking sporulation liberates metabolic resources and enhances the specific productivity of vegetative cells, providing a strategy for engineering high-performance B. licheniformis cell factories. Full article

Heat stress (HS) is one of the most challenging environmental conditions, responsible for impaired growth and reproduction in living systems. It also leads to altering the release of different biochemicals responsible for controlling the metabolic pathway. Five White Wistar rats were exposed at 42 ± 1 °C inside a closed chamber for the induction of hyperthermia. Their rectal temperature was recorded before and after heat exposure. The semi-digested food from the gut (colon) of sacrificed rats was collected under sterilized conditions for the isolation of gut bacteria on a nutrient agar plate at 50 °C, 60 °C, and 70 °C. The sample was incubated for 24 h and isolates were further purified. The proteolytic, amylolytic, cellulolytic, and xylanolytic activities were measured via plate assay and the enzymatic index was calculated. Total protein and estimation of heat shock protein 70 (HSP70) were also quantified. Initially, the rectal temperature of the animal was 37.1 ± 0.2 °C, but after exposure to heat, the temperature was 40.8 ± 0.2 °C. The number of purified isolates was recorded, i.e., at 50 °C (04), at 60 °C (01), and at 70 °C (03). Among eight isolates, Bacillus licheniformis (50 °C) showed all four enzymatic activities with a higher enzymatic index. Further, this novel isolate also exhibited a maximum concentration of HSP70. This preliminary study reveals the survival of a bacterium (B. licheniformis) capable of producing key metabolites, highlighting its significance in supporting host physiology and other pathophysiological conditions. As a probiotic, it may serve as a potential therapeutic bridge connecting HSP70, host physiological function, and gut health. Full article

This study investigated the physiological benefits of probiotic supplementation during refeeding after short-term fasting in Nile tilapia (Oreochromis niloticus). A total of 180 fish were assigned to three groups: continuously fed control or subjected to 5 days of fasting followed by 15 days of refeeding with either a basal or probiotic-enriched diet containing Bacillus subtilis, B. licheniformis, and B. pumilus. Growth performance indices (body weight, length, weight gain, specific growth rate, condition factor, relative feed intake, and feed conversion ratio) were measured. Muscle samples were collected for histomorphological evaluation and quantitative real-time PCR analysis of antioxidant genes catalase (cat) and superoxide dismutase 2 (sod-2), growth-related genes insulin-like growth factor 1 (igf-1) and suppressor of cytokine signaling 2 (soc-2), anti-inflammatory gene transforming growth factor beta (tgf-β), and myostatin genes. Fasting significantly reduced (p < 0.05) body weight compared to control, confirming the impact of nutrient deprivation. Upon refeeding, fish on the basal diet showed partial growth recovery but remained below control levels, whereas probiotic-fed fish exhibited superior recovery, surpassing both control and basal groups in body weight, length and weight gain. Condition factor exhibited insignificant changes among all groups after fasting and upon refeeding. Specific growth rate of the entire experiment was highest in the probiotic group, while insignificant. Relative feed intake decreased in both refed groups, yet feed conversion ratio improved, particularly with probiotics. Gene expression analysis revealed fasting-induced upregulation of antioxidant (cat and sod-2) and myostatin (p < 0.05), alongside downregulation of growth-related (igf-1 and soc-2) and anti-inflammatory (tgf-β) genes (p < 0.05). Basal refeeding restored most expressions, whereas probiotics enhanced antioxidant, growth, and anti-inflammatory genes while normalizing myostatin (p > 0.05 vs. control). Histological evaluation showed fasting-induced muscle atrophy, which was most effectively reversed by probiotics. Overall, probiotics accelerated recovery, highlighting their potential to optimize post-fasting growth in aquaculture. Full article

Bacteria represent promising tools for reducing the use of synthetic inputs in crop production. In this study, we evaluated the effects of two bacterial strains isolated from chicken compost tea—Bacillus licheniformis and Pseudomonas mendocina—on the yield and quality of strawberry. Experimental assays were conducted in two seasons (2023 and 2024) under macro-tunnel conditions, with the following treatments: control without applications (Con); commercial NPK fertilizer (FerC); application of B. licheniformis (BL) and P. mendocina (PM) solution in soil once a month. Both bacterial treatments enhanced soil properties. Fruit individual weight significantly increased in BL treatment compared to the control. Similar trends were observed for anthocyanin and ascorbic acid content (increases > 25%), as well as for antioxidant activity (increases of more than 20% and 13% for BL and PM, respectively). The differences were more significant in 2023. In addition, both strains showed positive in vitro results for phytase, siderophore, and IAA production (5.8–8.8 and 9.3–13 µg IAA/mL for BL and PM after 15 days). Although further field validation is required, these results indicate that bacteria (particularly B. licheniformis) show strong potential as bioinoculants to enhance the productivity and quality of strawberry. Full article

Bacillus species are among the most widely used microbial agents in agricultural biocontrol, reflecting their ecological resilience, functional diversity, and long history of practical application. The antagonistic activity of Bacillus spp. against plant pathogens and their plant growth–promoting effects are well established. However, these biological functions are frequently considered in isolation from safety evaluations and regulatory decision-making, resulting in a fragmented evidence base. This review addresses this gap by providing an integrated synthesis of agriculturally relevant Bacillus taxa, explicitly linking biocontrol performance with strain-level safety considerations and regulatory assessment. This review focuses on the principal groups currently applied in agriculture, including the Bacillus subtilis lineage, notably B. amyloliquefaciens, B. velezensis, B. pumilus, and B. licheniformis, as well as B. thuringiensis and Cytobacillus firmus. Key mechanisms underlying biocontrol efficacy are examined alongside evidence from greenhouse and field applications. These mechanisms include the production of secondary metabolites and volatile compounds, biofilm formation, rhizosphere colonisation, and the induction of plant defence responses. Attention is given to environmental and operational factors that influence the consistency of performance. A central contribution of this review is the integration of functional evidence with safety-relevant considerations, such as realistic metabolite exposure, antimicrobial resistance potential, and ecological effects. Regulatory approaches in the European Union, the United States, and selected Organisation for Economic Co-operation and Development countries are compared to illustrate how such evidence informs risk assessment and supports the sustainable use of Bacillus-based biocontrol agents in modern agriculture. Full article

As synthetic biology advances, prokaryotic microorganisms have become critical platforms for heterologous biosynthesis in cell factory applications. However, conventional free enzyme systems encounter substantial challenges, including inefficient intermediate transfer, toxic intermediate accumulation, and vulnerability to temperature and pH fluctuations. Enzyme complex catalytic systems offer promising solutions to these limitations. Bacillus licheniformis, a Generally Recognized as Safe (GRAS) host with exceptional protein secretion capacity, represents an ideal chassis for enzyme complex construction. This study developed a peptide-mediated platform in B. licheniformis to enable enzyme complex self-assembly and evaluated its effects on metabolic pathway performance. Five peptide elements were screened through fusion with enhanced orange/green fluorescent proteins (eOFP/eGFP) and transglutaminase (TGase). Effective peptide pairs were identified by measuring fluorescence intensity, visualizing complex formation via laser confocal microscopy, and assessing TGase activity. Subsequently, recombinant strains expressing peptide-fused key metabolic enzymes (gadTt and KdgA) were constructed for whole-cell biotransformation using gluconate as substrate to investigate the impact of peptide-mediated enzyme complexes on pyruvate synthesis. In the fluorescent protein system, P18/D18—amphipathic peptides that drive enzyme self-assembly via intermolecular hydrophobic interactions—increased extracellular fluorescence intensity of eOFP and eGFP by 31.11% and 25.21%, respectively. The D18 peptide significantly elevated TGase activity by enhancing structural stability to over 1.3-fold that of the control. For pyruvate synthesis, the peptide-mediated enzyme complex exhibited remarkable advantages in substrate conversion rate (up to 53.08%) and thermostability, confirming the platform’s ability to enhance substrate channeling despite no optimization for absolute yield. This study established a novel peptide-mediated multienzyme self-assembly platform in B. licheniformis, providing a valuable strategy for artificial metabolic channel design in synthetic biology. Full article

Rice bran, a nutrient-rich by-product of rice milling, is an underutilized resource in sustainable crop utilization. This study aimed to investigate the characteristics, total phenolic content, and antioxidant activities of rice bran protein hydrolysates (RBPHs) produced using proteases from Bacillus licheniformis (RBPH-B) and α-chymotrypsin (RBPH-C), along with their protein fractions (F1; >100 kDa, F2; 10–100 kDa, F3; 1–10 kDa, F4; <1 kDa). Molecular weight, color, surface hydrophobicity, secondary structure, total phenolic content, and antioxidant activities of the hydrolysates were assessed. Both enzymatic hydrolysis and ultrafiltration reduced molecular weight and surface hydrophobicity, enhanced lightness, and increased α-helix content. Among all samples, the <1 kDa peptide fraction derived from α-chymotrypsin hydrolysis (RBPH-C-F4) exhibited the strongest antioxidant activity, with the lowest EC₅₀ values for ABTS (0.94 mg/mL) and DPPH (210 µg/mL), as well as the highest inhibition of metal chelating activity (1.35 mmol EDTA/g sample) and linoleic peroxidation (90.62%). Enzymatic hydrolysis enhanced total phenolic content compared with native rice bran protein. These findings highlight the potential of rice bran-derived peptides as antioxidant candidates and indicate that further validation in food systems is required. Full article

This study systematically characterized the L-cysteine biosynthetic pathway in Bacillus licheniformis and demonstrated that exogenous serine supplementation significantly upregulated the expression of pathway-associated genes, confirming serine as the primary precursor driving L-cysteine synthesis. Through targeted gene deletions, we generated knockout strains BL2ΔglyA, BL2ΔsdaAA, BL2ΔmetC, BL2Δ2, and BL2Δ3 to minimize precursor diversion and product degradation. Combinatorial overexpression of the feedback-resistant mutant cysE^f and the transporter eamA yielded an engineered strain achieving 1.075 g/L L-cysteine in shake-flask fermentation with an 18.69% molar conversion yield. These findings highlight the potential of B. licheniformis as a platform for sulfur metabolic engineering and provide a sustainable fermentation strategy to replace traditional high-pollution hydrolysis-based L-cysteine production. Additionally, this work reveals fundamental differences in sulfur metabolism networks between Gram-positive and Gram-negative bacteria, elucidating microbial metabolic diversity and the cross-regulatory mechanisms linking sulfur, carbon, and nitrogen metabolism. Full article

Bacillus licheniformis Mb1, a thermophilic strain isolated from the Yungas rainforest in northwestern Argentina, was analyzed through genomic and experimental approaches to explore its biotechnological potential. Phylogenomic analysis confirmed its close relationship with B. licheniformis reference strains. The genome revealed multiple genes associated with hydrolytic, oxidative, carbohydrate-active, and polyester-degrading activities, indicating a wide enzymatic capacity. Experimental assays demonstrated strong extracellular hydrolytic activities and efficient degradation of bis(2-hydroxyethyl) terephthalate (BHET), a key polyethylene terephthalate (PET) intermediate. In liquid cultures with 3 mg/mL BHET, B. licheniformis Mb1 achieved 99.9% depletion within four days, with transient BHET dimer accumulation and progressive terephthalic acid (TPA) production, reaching 1.17 mg/mL after 15 days. Mono (2-hydroxyethyl) terephthalate (MHET) and vanillic acid were not detected. Complete BHET and dimer degradation suggests the presence of versatile hydrolases acting on short-chain polyester intermediates. Sequence and molecular docking analyses identified a BHETase-like carboxylesterase as the main enzyme candidate, featuring a truncated lidC region that generates a more open catalytic cleft. This structural trait, not previously reported in bacterial BHETases, enables the accommodation of bulkier substrates such as BHET dimer. These findings highlight B. licheniformis Mb1 as a promising biocatalyst for polyester depolymerization and a valuable microbial resource for future enzyme discovery and plastic bioremediation strategies. Full article

Despite increasing interest in probiotics as antibiotic alternatives in swine production, few studies have directly compared the functional efficacy of different commercial probiotic formulations under controlled conditions. We conducted an in vitro study using porcine intestinal epithelial (IPEC-J2) and macrophage-like (3D4/21) cell models to compare the efficacy of three commercial probiotic consortia (C1: three strains of Bacillus velezensis; C2: B. licheniformis + B. subtilis; C3: Clostridium butyricum). Treatments were evaluated for their ability to inhibit pathogenic Escherichia coli, Clostridium perfringens, and Salmonella spp., enhance epithelial barrier integrity, and modulate immune responses. Experimental endpoints included pathogen inhibition assays, adhesion to IPEC-J2 cells, transepithelial electrical resistance (TEER), tight junction protein expression, and cytokine profiling via RT-qPCR and proteomics. Data were analyzed using the Kruskal–Wallis test with false discovery rate (FDR) control at 5%. C1 cell-free supernatant (CFS) strongly inhibited pathogen growth (84.8 ± 5.3% inhibition of ETEC F4⁺F18⁻ vs. medium control; p < 0.05), whereas C2 had no effect, and C3 inhibited only one isolate. The coculture of IPEC-J2 cells with C1 CFS increased the expression of TJ proteins ZO-1, MUC13, and MUC20 (+12.9–46.6% vs. control; p < 0.001) and anti-inflammatory TGF-β; reduced pro-inflammatory IL-6 in LPS-stimulated 3D4/21 cells. In comparison, C2 and C3 showed minimal impact on epithelial barrier integrity and immune modulation, as indicated by negligible changes in TEER values, tight junction protein expression (ZO-1, MUC13, MUC20), and cytokine profiles relative to the control. In conclusion, C1 demonstrated greater in vitro efficacy than C2 (B. licheniformis + B. subtilis) and C3 (Clostridium butyricum), including pathogen inhibition assays, epithelial adhesion, TEER measurements, and cytokine modulation, suggesting its potential as a leading candidate for functional probiotic applications. Full article

Recycled bio-wastes such as compost and vermicompost, and bioenergy byproducts such as digestate and biochar are widely acknowledged for their role as soil conditioners capable of preserving soil fertility, maintaining soil health, and acting as a bio-adsorbent of organic soil pollutants (BIOSORs). Moreover, they are attracting increasing attention for use as effective carriers of microbial consortia into arable soils. This study aims to combine selection of bacteria tolerating contaminants of emerging concern (CECs) and their use to fortify BIOSORs. Seventeen bacterial strains isolated from commercial bio-stimulant formulations were studied together with three strains previously isolated and identified as Bacillus subtilis, Bacillus licheniformis, and Serratia plymuthica. All the strains were tested in vitro for their ability to grow under increasing concentrations (0, 0.2, 0.5 and 1 mg L⁻¹) of CECs: bisphenol A, 4-nonylphenol, penconazole, and S-metolachlor. Results highlighted a variability in the tolerance of the bacteria to the tested CECs. The B. subtilis, B. licheniformis, and S. plymuthica were the most promising strains, individually or as consortium, to tolerate individual CECs and their mix. Moreover, they exhibited metabolic activity when inoculated in the BIOSORs. Nevertheless, additional investigations such as quantitative assessment of CECs are needed to validate the methodology. This work contributes to investigate the feasibility of stable and functionally active microbially enriched bio-sorbents (Me-BIOSORs) and provides preliminary evidence supporting the potential to be used in soil–plant systems at the field scale. Full article