Bacillus Species Research Articles

Bioengineering Bacillus spp. for Sustainable Crop Production: Recent Advances and Resources for Biotechnological Applications

AbstractThe goal of sustainable agriculture is to meet the rising need for food, while minimizing adverse impacts on the environment, protecting natural resources, and ensuring agricultural output over the long term. The pressing need to increase agricultural yield through sustainable agriculture is being emphasized. Several Bacillus species have been used as commercial biopesticides since they can act against plant pathogens by potentially suppressing them. At the same time, they can act as plant growth-promoting rhizobacteria and are known for their diverse characteristics and beneficial properties, making them potential candidates for use sustainable crop production programs. Knowledge of genetic information opens the door of possibility for understanding the way these microorganisms behave. By applying biotechnological tools to Bacillus, strategies can be adopted for the purpose of increasing the yield of crops and managing pests and pathogens that infect them. In this review, we identify the genes in the most significant Bacillus spp. that contribute to plant improvement. The most important biotechnological tools and advance computational approaches are described to provide an extended vision on this topic. However, increasing the crop production through application of beneficial microbial strains requires a multifaceted approach that considers ecological, economic, and social aspects. By implementing these strategies and practices, we can work towards a sustainable and resilient agricultural system that meets the growing food demand, while preserving the environment for future generations.

Long-term garlic‒maize rotation maintains the stable garlic rhizosphere microecology.

Crop rotation is a sophisticated agricultural practice that can modify the demographic structure and abundance of microorganisms in the soil, stimulate the growth and proliferation of beneficial microorganisms, and inhibit the development of harmful microorganisms. The stability of the rhizosphere microbiome is crucial for maintaining both soil ecosystem vitality and crop prosperity. However, the effects of extended garlic‒maize rotation on the physicochemical characteristics of garlic rhizosphere soil and the stability of its microbiome remain unclear. To investigate this phenomenon, soil samples from the garlic rhizosphere were collected across four different lengths of rotation in a garlic-maize rotation. There were notable positive associations between the total nitrogen and total phosphorus contents in the soil and the duration of rotation. Prolonged rotation could increase the maintenance of microbiome α diversity. The number of years of rotation and the soil organic carbon (SOC) content emerged as principal determinants impacting the evolution of the bacterial community structure, with the SOC content playing a pivotal role in sculpting the species diversity within the garlic rhizosphere bacterial community. Additionally, SOC remains predominant in shaping the root-associated bacterial community's β-nearest taxon index. However, these factors do not have a notable effect on the fungal community inhabiting the garlic rhizosphere. In comparison with monoculture, rotation can amplify the interconnectivity and intricacy of microbial ecological networks. Long-term rotation can further maintain the stability of both microbial ecological networks and interactions between bacterial and fungal communities. It can enlist a plethora of beneficial Bacillus species microorganisms within the garlic rhizosphere to form a biological barricade that aids in safeguarding garlic against encroachment by the pathogenic fungus Fusarium oxysporum, consequently diminishing disease incidence. This study provides a theoretical foundation for the sustainable development of garlic through long-term crop rotation with maize. Our research results indicate that long-term garlic‒maize rotation maintains stable garlic rhizosphere microecology. Our study provides compelling evidence for the role of long-term crop rotation in maintaining microbiota and community stability, emphasizing the importance of cultivating specific beneficial microorganisms to enhance rotation strategies for garlic farming, thereby promoting sustainability in agriculture.

Biocontrol Efficacy of Trichoderma and Bacillus Isolates against Sclerotium rolfsii under in vitro Conditions

Background: Sclerotium rolfsii is a polyphagous pathogen which is uneconomical and difficult to control due to its broad host range and persistence due to its tough resting structures, i.e., sclerotia. Chemical control has been supplanted by biological phytopathogen control. Antibiotic microorganisms are regarded as one of the viable management techniques in an integrated approach for ecofriendly management. Methods: Twenty three Trichoderma spp. and twenty one Bacillus species were isolated at Groundnut Pathology Laboratory, ICRISAT, Patancheru from rhizospheric soils of Telangana during rabi 2022-23 and 2023-24 and examined for antagonistic activity against Sclerotium rolfsii in an effort to design an effective biocontrol system for the management of Sclerotium rolfsii in groundnut. Result: The highest antagonistic activity against Sclerotium rolfsii has been observed in Trichoderma viride and Bacillus cereus. In the dual culture assay, Trichoderma viride and Bacillus cereus inhibited Sclerotium rolfsii by up to 65.33 per cent and 58.22 per cent, respectively. In the metabolites assay, Trichoderma viride and Bacillus cereus inhibited Sclerotium rolfsii by 63.33 per cent and 55.40 per cent, respectively. Thus, Trichoderma viride and Bacillus cereus displayed promising biofungicidal capabilities against the stem rot pathogen. ​

The Effectiveness of Biofloc Technology and Its Application Prospects in Sea Cucumber (Apostichopus japonicus) Aquaculture: A Review

This review aims to advance the development of biofloc technology (BFT), providing more sustainable and efficient practices for the farming of the Japanese sea cucumber (Apostichopus japonicus). BFT is a sustainable aquaculture method that promotes nutrient recycling and effective carbon source management, offering significant advantages such as improving water quality, enhancing growth performance, and boosting the physiological activity and disease resistance of cultured animals. In A. japonicus farming, the optimal carbon source is glucose, and the ideal carbon-to-nitrogen (C/N) ratio ranges between 15 and 20. Microbial additives, such as the Bacillus species, have been shown to enhance biofloc formation and growth, as well as the immune responses in A. japonicus. However, the technology also faces limitations, including finding suitable biofloc culture protocols that match the physiological habits of A. japonicus and potential challenges with biofloc stability under varying environmental conditions. Based on existing research, this review discusses these limitations in the farming of A. japonicus. Additionally, it compares biofloc farming models for other economically important aquatic species. By addressing these key aspects, this review offers insights to enhance BFT performance, ultimately contributing to more efficient and sustainable A. japonicus aquaculture practices.

Prevalence and identification of Bacillus species in cellophane, foil, and disposable plastic containers used for food packaging

Background: Foil, cellophane, and disposable plastic containers are commonly employed in food packaging, namely as coatings on food items. Nevertheless, the existence of microbes on the surface and inside the internal composition of these materials gives rise to concerns, as they have the capacity to multiply under ideal circumstances and spread to food items, thereby jeopardizing the safety and quality of the food. Objective: This study aimed at examining the number of microorganisms and identifying specific bacterial species within frequently used food packaging materials like foil, cellophane, and disposable plastic containers. Methods: samples of foil, cellophane, and disposable plastic containers were collected from various popular brands in Iran. The overall bacterial count wضas determined using defibering, flooding, and smear techniques followed by culturing on Tryptone Glucose Extract Agar (TGEA) medium. To identify the bacterial species, various biochemical tests were performed, including fermentation, motility testing, catalase test, oxidase test, and methyl red-Voges Proskauer (MR-VP) test. Results: Bacterial quantity ranged from 0 CFU/g to 8.3 × 103 CFU/g in analyzed samples. All the samples had primary contaminants mainly Bacillaceae family bacteria that could produce spores. Disposable plastic containers had the lowest bacterial count whereas cellophane showed the highest bacterial contamination level. Among Bacillaceae family members Bacillus licheniformis was dominant. Conclusion: The study's findings emphasize the possibility of microbial contamination in food packaging materials, including cellophane. These results indicate that the food packaging industry should adopt a rigorous approach to hazard evaluation and Important Control Points (HACCP) in order to guarantee the microbiological safety and efficacy of packaging materials.

Impact of Bacillus Inoculation on Rhizosphere Bacterial Community Structure: A Review

The rhizosphere is a specialized zone where plant roots interact with the soil microbiome. Among various beneficial microbes, the genus Bacillus stands out due to its diverse functionalities and potential to boost plant growth and resilience. Bacillus is a key genus of plant growth-promoting rhizobacteria (PGPR) known for enhancing nutrient availability, producing phytohormones, and inducing plant resistance to pathogens. Inoculating Bacillus species into the rhizosphere can significantly alter the bacterial community's structure and composition. These alterations are driven by the competitive and cooperative interactions between Bacillus and the native rhizosphere microorganisms. Incorporating soil microorganisms into the host plant's beneficial bacterial community improves soil nutrient cycling and nutrient use efficiency. Using microbial inoculants is an effective strategy to address crop succession challenges, enhance microbial community structure, and improve soil fertility, thereby promoting crop growth. This review thoroughly examines the current understanding of how Bacillus inoculation impacts rhizosphere bacterial community structure.

Prospects of iron solubilizing Bacillus species for improving growth and iron in maize (Zea mays L.) under axenic conditions

Iron (Fe) deficiency in calcareous soils is a significant agricultural challenge, affecting crop productivity and nutritional quality. This study aimed to isolate, characterize, and evaluate Fe solubilizing rhizobacterial isolates from maize rhizosphere in calcareous soils as potential biofertilizers. Forty bacterial isolates coded as SG1, SG2, …, SG40 were isolated and screened for siderophore production, with ten showing significant Fe solubilizing capabilities. These isolates were further assessed for phosphate solubilization and exopolysaccharides production. The selected bacterial isolates were also screened under axenic conditions for their ability to improve maize growth. The isolates SG8, SG13, SG24, SG30 and SG33 significantly enhanced growth parameters of maize. Notably, SG30 showed highest increment in shoot length (58%), root length (54%), root fresh and dry biomass (67% and 76%), SPAD value (67%), relative water contents (69%), root surface area (61%), and Fe concentration in shoots (79%) as compared to control. The biochemical characterization of these strains showed that all these strains have capability to solubilize insoluble phosphorus, produce indole-3-acetic acid (IAA), and ammonia with catalase, urease and protease activity. Molecular identification through 16s rRNA gene sequencing confirmed high similarity (99.7–100%) of the selected isolates to various Bacillus species, including B. pyramidoids, B. firmicutes, and B. cereus. The study provides a strong base for developing eco-friendly, cost-effective biofertilizers to address Fe deficiency in crops and promote sustainable agriculture.

Plant growth promoting Rhizobacteria for sustainable tomato production

Numerous phosphate solubilizing bacteria (PSB) are found in the rhizosphere, benefiting the host plant in different mechanisms and promoting sustainable agriculture. They improve soil fertility with minimum cost requirements using an eco-friendly approach, which is an essential attribute for small-scale farm production. Thus, isolation, characterization, and evaluation of symbiotic effectiveness are hierarchical procedures to develop bioinoculants. Using a dual (plate and liquid medium) screening technique and a greenhouse trial, four potential PSB strains (Mk-1–25, Mk-13–16, Mk-20–7, and Mk-20–20) were selected from tomato rhizosphere soil. All isolates exhibited the following traits: produced colony clear-zone, lowered liquid medium pH, gram-positive, produced urease and IAA, dissolved substantial P from various substrates, symbiotically effective, and improved tomato growth and yield. Taxonomically, they belong to Bacillus and Priestia species (i.e., Mk-1–25 Bacillus halotolerance, Mk-13–16 Bacillus amyloliquefaciens, Mk-20–7 Bacillus megaterium, and Mk-20–20 Priestia megaterium). Among the current strains, Bacillus halotolerance recorded higher SI (3.11), was able to grow in extreme conditions (10% salt, up to 45 °C), produced HCN and siderophore, and improved tomatoes’ shoot length and weight, in contrast, Mk-20–20 produced HCN, siderophore, was able to fix nitrogen, improved tomato germination, shoot dry weight, and fruit weight. Various P-supplements (Ca3(PO4)2, bonemeal, FePO4, AlPO4, compost, and DAP fertilizer) were added to induce tomato-strain symbiotic interaction; hence, compost substantially promoted the interaction and resulted in higher symbiotic effectiveness. Ca3(PO4)2 was a good media composite preferred by most PSB strains; consequently, a higher concentration (219 µg/ml) of dissolved P was recorded from the liquid medium after 10 days of incubation than other substrates. Based on the data obtained from laboratory and greenhouse trials, the current strains were found to be potential candidate. Future work should confirm their efficacy at the field level using model host crops at different sites to recommend them as farm inputs and biofertilizer.

Outside-in enhancement of phosphate solubilizing bacteria by sludge biochar for phenol remediation in soil: Pollution stress reduction, electron transfer gain and secretion regulation

The high toxicity of phenol poses a significant barrier to bacterial bio-removal capacity. This study innovatively proposes the utilization of sludge biochar (SB) to embed phosphate-solubilizing bacteria (CZB1), creating a composite system (SB-CZB1) that enhances the bio-removal efficiency of phenol. The research findings indicate that after being embedded with biochar, Phosphorus solubilizing bacteria (PSB) achieved a significant enhancement (49.7 %-67.0 %) in phenol removal efficiency across different concentration gradients (1500, 2000, 2500 mg/L). SB component in the SB-CZB1 composite system provides a stable and favorable growth environment for CZB1, significantly enhancing microbial metabolic activity. Notably, it stimulates CZB1 to secrete succinic acid and malic acid (with increases of 3.8 % and 23.9 %, respectively), effectively mitigating the detrimental effects of phenol toxicity on microorganisms. Three-dimensional fluorescence and electrochemical analysis demonstrate that SB not only exhibits exceptional electrochemical performance but also stimulates CZB1 to generate redox-active substances (quinone components in humic acid). The electron transfer rates between microorganisms and phenol in the SB-CZB1 system are 9.75–21.71 times and 23.94–63.95 times higher than those in SB and CZB1 alone, respectively. Meanwhile, the abundant oxygen-containing functional groups (COH/COC, COOH, CO) on the surface of SB-CZB1 play a pivotal role in the adsorption and removal of phenol. Furthermore, pot experiments further reveal that the SB-CZB1 composite system significantly enhances the removal efficiency of phenol in soil (98.1 %). Additionally, the application of SB-CZB1 effectively modulated soil properties, notably increasing available potassium and available phosphorus content by 40.86 % and 136.69 %. This application not only enriched soil microbial diversity but also promoted the proliferation of native organic pollutant remediation bacteria in the soil. Notably, the proliferation of Bacillus species affiliated with CZB1 was the most significant, increasing by 82.1 %. This result confirms that biochar embedding technology effectively enhanced the colonization of CZB1 in the soil, thereby significantly accelerating the bio-removal process of phenol.

In vivo biocontrol potential of Bacillus plant growth-promoting rhizobacteria against pectinolytic plant pathogens.

Bacillus is well known for producing a wide range of compounds that inhibit microbial phytopathogens. From this perspective, we were interested in evaluating the biocontrol potential of 5 plant growth-promoting rhizobacteria Bacillus species (PGPR-Bacillus) on 21 microbial pectinolytic plant pathogens isolated from previous studies. Phytopathogenicity and in vivo biocontrol potential of PGPR curative and preventive treatments were investigated from this angle. Overall, the pathogenicity test on healthy tomato, zucchini, and mandarin showed low rot to no symptoms for all PGPR strain culture treatments. Conversely, zucchini pre-treated with PGPR strains B. circulans and B. cereus for 72h showed no signs of soft rot and remained healthy when in vitro contaminated with phytopathogens (Neisseria cinerea and Pichia anomala). Additionally, the PGPR-Bacillus strains were shown to be effective in mitigating the symptoms of soft rot in tomatoes, zucchini, and oranges using in vivo curative treatment. It is true that the majority of pectinolytic phytopathogenic strains exhibited antibiotic resistance. In vivo tests revealed that PGPR-Bacillus cell culture was effective against plant pathogens. Thus, PGPR-Bacillus can be considered a potential biocontrol agent for pectinolytic plant pathogens.

A phosphate starvation induced small RNA promotes Bacillus biofilm formation

Currently, almost all known regulators involved in bacterial phosphorus metabolism are proteins. In this study, we identified a conserved new small regulatory RNA (sRNA), named PhoS, encoded in the 3’ untranslated region (UTR) of the phoPR genes in Bacillus velezensis and B. subtilis. Expression of phoS is strongly induced upon phosphorus scarcity and stimulated by the transcription factor PhoP. Conversely, PhoS positively regulates PhoP translation by binding to the ribosome binding site (RBS) of phoP mRNA. PhoS can promote Bacillus biofilm formation through, at least in part, enhancing the expression of the matrix-related genes, such as the eps genes and the tapA-sipW-tasA operon. The positive regulation of phoP expression by PhoS contributes to the promoting effect of PhoS on biofilm formation. sRNAs regulating biofilm formation have rarely been reported in gram-positive Bacillus species. Here we highlight the significance of sRNAs involved in two important biological processes: phosphate metabolism and biofilm formation.

A The Effects of Bio-Priming on Seed Germination and Seedling Growth of Italian Ryegrass (Lolium multiflorum Lam.)

Seed bio-priming applications with plant growth promoting rhizobacteria (PGPR) have been widely used recently to improve germination and seedling growth. Therefore, the aim of this study was to investigate the effects of bio-priming with different bacterial strains on germination and seedling development of Italian ryegrass seeds. The sterilized seeds of the Elif variety (Lolium multiflorum Lam) were inoculated with nine different bacterial strains belonging to Bacillus species (108 cfu/mL bacterial suspension) for 15 min at 120 rpm and then dried at room temperature. The treated seeds were germinated in petri dishes with 25 seeds between 3 filter papers at 22 ±2 ˚C. The study was carried out in a completely randomized design with three replications. As a result of the study, no significant difference was obtained between the treatments in germination percentage and root length, but it was determined that SY2 and SY5 (Bacillus isolates) showed superior performance compared to the control in terms of shoot length and seedling fresh and dry weights.

Bacillus-inoculated organomineral fertilizer in soybean crop during short-term: comparing sources and doses with mineral fertilizer

Bacillus species and organomineral fertilizers can improve various crop responses. The objective of this study was to evaluate the efficiency of organomineral fertilizer with B. subtilis and B. licheniformis, in comparison to mineral fertilizer, on soybean growth, leaf nutrient content, grain yield, and soil characteristics. Two soybean field experiments (305 km apart) were set up in southeastern Brazil. Two sources of fertilizers (inoculated granulated organomineral, 5-26-0 + 0-0-32 N-P2O5-K2O, and coated mineral fertilizer, 10-52-0 + 0-0-60 N-P2O5-K2O), and four doses of P plus K (no fertilizer, 50, 75, 100 kg ha−1 of each P2O5 and K2O) were applied. At 60 days after sowing, plant height and leaf analysis were assessed. Weight of 1000 seeds, grain yield, and soil analysis were evaluated (120 days after sowing). In both fields, there was interaction between source and P plus K dose factors for soybean grain yield. In Site 1, the highest grain yield (4,543.69 kg ha−1) was observed for 100 kg ha−1 of P2O5 and K2O dose of coated mineral fertilizer; in Site 2, the highest grain yield (4,638.55 kg ha−1) was observed for 75 kg ha−1 of P2O5 and K2O dose of organomineral fertilizer. These factors also affected N, P, and K leaf contents. Less effects were observed on plant height, weight of 1000 seeds, and soil characteristics. Inoculated organomineral fertilizer can be used for total or partial supply of nutrients without a clear difference from mineral fertilization. Smaller doses of inoculated organomineral fertilizer could be used without compromising soybean grain yield.

Isolation and Characterization of Arsenic-Tolerable Bacteria from Groundwater and Their Implementation on Rice Seedling's Shoot and Root Enhancement.

Arsenic exerts detrimental impacts on primary metabolism in plants, leading to reduced crop yield. Some arsenic-resistant plant growth-promoting bacteria (PGPB) help plants by providing some plant hormones to sustain their growth and development under arsenic stress. Here, seven different species of Bacillus were isolated from arsenic-contaminated groundwater of West Bengal, India. Those species were capable of growing in the presence of > 3.12g/L arsenate (AsV) and > 0.65g/L arsenite (AsIII) salts and also resist different heavy metals like Cu2+, Fe2+, Co2+, Zn2+, Pb2+, etc. They were susceptible to multiple groups of antibiotics like beta-lactam, aminoglycosides, etc. All species were capable of detoxifying arsenite and influenced rice seedlings' growth in the presence of arsenic salts by their capabilities likenitrogen-fixing ability, phosphate solubilization, indole 3-acetic acid (IAA), gibberellic acid (GA), proline production, etc. Most species helped enhance root and shoot lengths under arsenic stress. These primary findings suggest that those Bacillus spp. could be used as potential bio-fertilizers in arsenic-contaminated agricultural fields.

Enhancing Broccoli growth by bacillus rhizosphere bacteria

This study examines the enhancement of broccoli growth by Bacillus Rhizosphere. Bacteria Broccoli is a nutrient-rich vegetable known for its high content of polyphenols, flavonoids, vitamins, minerals, fiber, and low calorie count, making it a recommended daily food. Tropical regions grow broccoli in mountainous areas. Sustainable agriculture emphasizes minimizing chemical fertilizers, and biofertilizers like Plant Growth Promoting Rhizobacteria (PGPR), such as Bacillus, can reduce their use. Bacillus improves plant growth by fixing nitrogen, solubilizing phosphate and potassium, and producing phytohormones. Understanding Bacillus growth dynamics is vital for optimizing its agricultural application. This study aimed to analyze growth curves of different Bacillus species and investigate their potential role in promoting broccoli growth. This study was carried out in January–April 2024 at the Soil Biology Laboratory, Faculty of Agriculture, Universitas Padjadjaran, and Bumi Agro Technology Company, Lembang. The research was divided into 3 stages: growth curve determination, Bacillus liquid inoculant preparation, and application of treatment to plants. The findings showed that isolates of B. safensis strain MDL5, B. altitudinis strain RPW2, B. subtilis strain YPS4, and Bacillus sp. strain SZ057 reached the highest point in their growth curves after 4 and 5 days of incubation. After that, the curves slowly went down. Inoculating Bacillus into broccoli plants resulted in an increase in plant height by 43.28–58.75% compared to the control.

Immune protective, stress indicators, antioxidant, histopathological status, and heat shock protein gene expression impacts of dietary Bacillus spp. against heat shock in Nile tilapia, Oreochromis niloticus

This research evaluated the efficacy of mixed Bacillus strains probiotic supplements in mitigating acute thermal-induced stress in Nile tilapia (Oreochromis niloticus). Three experimental fish groups involved 135 Nile tilapia (49 ± 2 g); one control (no added probiotics), 0.5, and 1% of selected Bacillus strains (B. subtilis, B. licheniformis, and B. pumilus) for 58 days. After the feeding period, growth parameters, immunological parameters, stress biochemical markers, and antioxidant parameters in addition to genes related to stress and histopathological changes in fish, were assessed; subsequently subjected to heat shock at 36 ± 0.5 ◦C for 2 h. Before the heat challenge, our results exhibited a marked increase in the growth efficacy (P < 0.05), lower marked serum levels of triglycerides and cholesterol, and tissue malondialdehyde (MDA) levels along with significantly increased superoxide dismutase (SOD) and catalase (CAT) enzymes activity in fish-fed Bacillus probiotic at 0.5% concerning the control group (P < 0.05). There were no significant changes in the serum levels of glucose, cortisol, lactate, phagocytic activity, respiratory burst (ROS), total immunoglobulin Ig, alanine aminotransferase (ALT), aspartate aminotransferase (AST), lactate dehydrogenase (LDH), total protein, albumin, globulin, uric acid, urea, creatinine, as well as HSP70, GST, and GPx mRNA expression in most of the probiotic groups compared to the control group (P > 0.05). When Nile tilapia was exposed to heat stress, supplementation with Bacillus probiotic in the diet significantly decreased most of the indices related to serum biochemical (ALT (P < 0.01; P < 0.001), AST (P < 0.01), LDH (P < 0.01), urea (P < 0.05), and creatinine (P < 0.01)), triglycerides (P < 0.001; (P < 0.01)), cholesterol (P < 0.01; (P < 0.05)), glucose (P < 0.001), and cortisol (P < 0.01; (P < 0.05)), with tissue oxidative stress MDA levels (P < 0.05), and HSP70 mRNA expression (P < 0.01; P < 0.001), aligned with the stressed control group. In addition, a notable upsurge in the total protein, albumin, globulin, phagocytic and ROS activities, and total Ig, as well as the enzymatic antioxidant ability (SOD, CAT) (P < 0.01), with GST and GPx mRNA expression (P < 0.05; P < 0.01), were shown in fish-fed Bacillus spp. post-exposure compared with the stressed control group. Besides, no histopathological alterations were revealed in the spleen and brain of fish pre- and post-heat exposure. According to our findings, diet supplementation of Bacillus species has the potential to combat the suppressive effects of heat shock in Nile tilapia.

Thiophanate-methyl and its major metabolite carbendazim weaken rhizobacteria-mediated defense responses in cucumbers against Fusarium wilt

AbstractThe effect of fungicides on the plant-rhizosphere microbiome is a subject of ongoing debate, but whether any alteration in the rhizosphere microbiome could affect plant health is an issue that has not been thoroughly investigated. To address this deficiency, we analyzed the rhizosphere microbiome of wilt disease—resistant and disease-susceptible cucumber cultivars to determine whether (and which) plant-associated microorganisms have a role in disease resistance. We further assessed whether the fungicides thiophanate-methyl and carbendazim affect the rhizosphere microbiome, which may contribute to the plant’s immune response. Based on results acquired with both radicle-inoculation and soil-inoculation methods, cultivars Longyuanxiuchun (LYXC) and Shuyan2 (SY2) were identified as being disease resistant, whereas Zhongnong6 (ZN6) and Zhongnong38 (ZN38) were susceptible. The microbiome structure differed substantially between the resistant and susceptible plants, with LYXC and SY2 each having a significantly greater Shannon index than Zhongnong38. These results revealed that the disease-resistant cucumber cultivars recruited more beneficial bacteria, i.e., Bacillus, in their rhizosphere soil; as such, Bacillus was identified as a keystone genus in the microbial co-occurrence network. Thus, the presence of Bacillus may help cucumbers defend against fungal pathogens within the rhizosphere. Bacillus subtilis strain LD15, which was isolated from LYXC rhizosphere soil, could suppress pathogen growth, in vitro, and reduce disease severity in pot assays. Moreover, evidence also confirmed the accumulation of LD1 in the rhizosphere soil of resistant cucumber cultivars. For LYXC, application of thiophanate-methyl or carbendazim altered the microbiome structure, decreased bacterial diversity, and reduced the abundance of Bacillus species. Finally, pot assays verified that fungicide application decreased the proportion of LD15 in rhizosphere soil. From a microbial perspective, thiophanate-methyl and carbendazim may weaken the rhizobacteria-mediated defense response of cucumbers against cucumber Fusarium wilt disease. Our findings reveal a role for the rhizosphere microbiome in protecting plants from pathogens and constitute a reference for assessing the ecotoxicological risk of pesticides to non-target soil microorganisms.

Evaluation of the probiotic ability of Cellulolytic Bacillus subtilis SBMP4 (FSP20) isolated from waste-dump site in FIIRO, Lagos Nigeria

BACKGROUND: Current invention in the field of biotechnology includes research into the probiotic potentials of enzyme-producing bacteria species. OBJECTIVE: This study evaluated the probiotic ability of cellulolytic Bacillus subtilis SBMP4 isolated from waste dump site. METHODS: Isolated Bacillus species was screened for cellulolytic ability on carboxyl methylcellulose agar (CMCA), and hydrolysis efficiency was calculated. Probiotic ability of cellulolytic Bacillus species was carried out as follows: pathogenic potentials, bacteriocin producing ability, antibiotic susceptibility testing, gastric acid, and bile tolerance. Molecular identification was done by means of 16S rRNA gene sequencing, blast search analysis and phylogenetic analysis. RESULTS: Zone of clearance of 30 ± 2.08 mm on CMCA with hydrolysis efficiency of 114 % depicted the isolate’s cellulolytic potential. Bacillus subtilis SBMP4 showed γ-hemolytic phenotype with negative lecithinase activity. Antibacterial activity of bacteriocin observed against Staphylococcus aureus ATCC 25923 reference culture, had maximum inhibition zone of 14±1.0 mm at 72 hour of incubation. Substantial susceptibility towards 9 out of 10 commonly used antibiotics further depicted the isolate’s probiotic ability. Bacillus subtilis SBMP4 exhibited good gastric acid tolerance (pH 2.5) of 5.857 Log cycle/ml and good tolerance to 5 % and 10 % bile salt concentrations with 6.134 and 6.301 Log cycle/ml of viable bacteria cells. Molecular identification of the selected bacteria species revealed it to be Bacillus subtilis SBMP4 with RefSeq accession number NR_118383, showing maximum sequence similarity of 98%. CONCLUSION: Cellulolytic Bacillus subtilis SBMP4 possesses good probiotic ability and has the potentials of being applied as additive in livestock feeding to enhance overall productivity. KEYWORDS: Cellulolytic, Probiotic, CelA gene, Bacteriocin, Bacillus subtilis

Citric Acid by-Product Fermentation by Bacillus subtilis I9: A Promising Path to Sustainable Animal Feed

Citric acid by-products in animal feed pose a sustainability challenge. Bacillus species are commonly used for fermenting and improving the nutritional quality of feedstuffs or by-products. An experiment was conducted to enhance the nutritional value of citric acid by-products through fermentation with Bacillus subtilis I9 for animal feed. The experiment was carried out in 500 mL Erlenmeyer flasks with 50 g of substrate and 200 mL of sterile water. Groups were either uninoculated or inoculated with B. subtilis I9 at 107 CFU/mL. Incubation occurred at 37 °C with automatic shaking at 150 rpm under aerobic conditions for 0, 24, 48, 72, and 96 h. Inoculation with B. subtilis I9 significantly increased Bacillus density to 9.3 log CFU/mL at 24 h (p &lt; 0.05). CMCase activity gradually increased, reaching a maximum of 9.77 U/mL at 72 h. After 96 h of fermentation with inoculated B. subtilis I9, the citric acid by-product exhibited a significant decrease (p &lt; 0.05) in crude fiber by 10.86%, hemicellulose by 20.23%, and cellulose by 5.98%, but an increase in crude protein by 21.89%. Gross energy decreased by 4% after inoculation with B. subtilis in comparison to the uninoculated control (p &lt; 0.05). Additionally, the non-starch polysaccharide (NSP) degradation due to inoculation with B. subtilis I9 significantly reduced (p &lt; 0.05) NSP by 24.37%, while galactose, glucose, and uronic acid decreased by 22.53%, 32.21%, and 18.11%, respectively. Amino acid profile content increased significantly by more than 12% (p &lt; 0.05), including indispensable amino acids such as histidine, isoleucine, lysine, methionine, phenylalanine, tryptophan, and valine and dispensable amino acids like alanine, aspartic acid, glutamic acid, glutamine, glycine, proline, and tyrosine. Furthermore, citric acid by-products inoculated with B. subtilis I9 exhibited changes in the cell wall structure under scanning electron microscopy, including fragmentation and cracking. These results suggest that fermenting citric acid by-products with B. subtilis I9 effectively reduces dietary fiber content and improves the nutritional characteristics of citric acid by-products for use in animal feed.

Bioactive compounds from food-grade Bacillus.

Bacillus species have attracted significant attention in recent years due to their potential for producing various bioactive compounds with diverse functional properties. This review highlights bioactive substances from food-grade Bacillus strains and their applications in functional foods and nutraceuticals. The metabolic capacities of Bacillus species have allowed them to generate a wide range of bioactive substances, including vitamins, enzymes, anti-microbial peptides, and other non-ribosomal peptides. These substances have a variety of positive effects, including potential cholesterol-lowering and immune-modulatory qualities in addition to anti-oxidant and anti-bacterial actions. The uses and mechanisms of action of these bioactive chemicals can be used to improve the functional qualities and nutritional profile of food products. Examples include the use of anti-microbial peptides to increase safety and shelf life, as well as the use of Bacillus-derived enzymes in food processing to improve digestibility and sensory qualities. The exploitation of bioactive compounds from food-grade Bacillus strains presents a promising frontier in the development of functional foods and nutraceuticals with enhanced health benefits. Due to their wide range of activity and applications, they are considered as important resources for the development of novel medications, agricultural biocontrol agents, and industrial enzymes. Ongoing research into the biosynthetic pathways, functional properties, and applications of these compounds is essential to fully realize their potential in the food industry. This review underscores the significance of various bioactive compounds generated from Bacillus in tackling global issues like environmental sustainability, sustainable agriculture, and antibiotic resistance. Future developments in microbiology and biotechnology will enable us to fully utilize the potential of these amazing microbes, resulting in novel approaches to industry, agriculture, and health. © 2024 Society of Chemical Industry.