Keywords

1.1 Introduction

The production of crops must be increased to fulfill the global need, but environmental conditions and limited mineral and water sources are making this goal harder. These difficulties will be worsened by the predicted rise in human population over the coming decades, and crops are badly impacted by climate change due to global warming. Soil and climate control the spread of various plant species, and the production of crops is heavily influenced by biotic and abiotic pressures. The anticipated crop losses for grains such as wheat and maize as a result of climate change are around 3.8–5.5% (Lobell et al. 2011; Singh et al. 2016a, 2021a, b). Among the biotic stresses, fungal diseases are major responsible factors for crop loss. More than 19,000 fungal species have been identified that infect crop plants, worldwide. Approximately 30% of crop diseases are caused by pathogenic fungus (Jain et al. 2019). Agricultural techniques must concentrate on managing soil health and crop protection alongside their natural partners, beneficial microorganisms, in order to meet this challenge. Environmental pressures are frequently applied to plants in both natural and cultivated systems. The issue of the short supply of water for agriculture is getting worse in many places of the world. Excessive use of groundwater jeopardizes future irrigation capacity. Urbanization and industrialization are disturbing important wetlands and aquatic ecosystems. Water quality problems such salinization, nutrient overloads, and pesticide pollution are very common, worldwide (Brodt et al. 2011).

The sustainable crop production is accomplished by applying biotechnology techniques, traditional plant breeding, and agronomic practices. The conventional breeding involves choosing genotypes with the highest productive crop; genetic modification, such as gene insertions or induced mutations; and agronomic practices, such as the use of fertilizer. Because of the genetic makeup of plants and diverse environmental variables, the standard plant breeding and agronomic methods are not always effective. GM crops are commercially not so successful because of lesser acceptance in public. Numerous symbiotic or free-living soil bacteria contribute significantly to the viability, development, and productivity of plants under different environmental and biotic challenges. Numerous studies have established the plant and microbe interaction that are promoting plant development and shield plants from abiotic and microbial threats. The productivity of crops heavily depends on microbial communities present in rhizospheric soil; within the last few decades, PGPR has emerged as significant and promising tools for the sustainable agriculture practices (Yadav et al. 2022a, 2023). According to the Kloepper and Schroth (1981), the term “PGPR” refers to bacteria that may colonize plant roots and stimulate plant growth. When they reintroduced microbial consortium into the soil, about 2% to 5% of rhizobacteria were showing positive impact plant development, called PGPR (Antoun and Prévost 2006). The majority of PGPR bacteria belongs to the Bacillus, Acetobacter, Azospirillum, Azotobacter, Burkholderia, Klebsiella, Pseudomonas, and Serratia genera (Glick 1995, Singh et al. 2016a, b). Many bacterial species from different genera have been characterized as PGPR, but Bacillus and Pseudomonas spp. are major contributors and have been extensively explored (Podile and Kishore 2006; Singh et al. 2016a, b). Bacillus spp. can produce endospores, which enable them to remain viable for a long time in unfavorable environmental circumstances. Mature endospores of Bacillus are resistant to heat, UV, γ-radiation, and various toxic and hydrolytic enzyme treatments (Nicholson et al. 2000). The pellets of Bacillus subtilis endospores can survive in free space exposed to the solar radiation (Horneck et al. 1984). Most of Bacillus species are rod shaped, Gram-positive, endosporic, aerobic or facultative anaerobic, and ubiquitous in nature. Members of this genus have an extensive range of physiological adaptations that allow them to survive in various types of natural habitats. They are present in all type of habitats and water bodies and can be frequently isolated from the clinical samples (Carter 1990). They also survive in extreme condition habitats such as hot springs (Panda et al. 2013), hydrothermal vents (Marteinsson et al. 1996), tidal flats (Jung et al. 2011), shallow marine water (Maugeri et al. 2001), high-salt environments (Rivadeneyra et al. 1993), acidic (Mahdavi et al. 2010) and alkaline environments (Gessesse and Gashe 1997), heavy metal-contaminated sites (Egidi et al. 2016), dumping sites (Yadav et al. 2022b), and radioactive sites (Asker et al. 2007). Because of their versatile physiological ability, Bacillus spp. has been exploited for the production of enzymes, antibiotics, metabolites, drugs, farming, and industrial processes. Bacillus species are the source of two well-known antibiotics, bacitracin and polymyxin (Wu et al. 2022; Choi et al. 2009). In comparison to many other genera, the genus Bacillus is enormous and exhibits a high level of genetic variation. The second edition of Bergey’s manual (2009) listed 141 species of Bacillus (Vos et al. 2009). Bacillus spp. can thrive in a variety of conditions, which illustrates their vast metabolic ranges and ubiquitous distribution. More than 200 Bacillus species have been distinguished and categorized using genomic and conventional methods combined, and their role in the environment and pathological aspect have been identified (Singh et al. 2016a, Mageshwaran et al. 2022, Shafi et al. 2023, Shahid et al. 2022a, b). B. subtilis, as a model organism of prokaryotes, is well explored in molecular and cell biology research area and has been frequently used as a model organism in numerous experiments (Borriss et al. 2018; Mageshwaran et al. 2022). Many Bacillus species, e.g., B. megaterium, B. circulans, B. coagulans, B. subtilis, B. azotofixans, B. macerans, B. velezensis, etc. are extensively researched for their PGPR actions (Hashem et al. 2019; Goswami et al. 2016; Basu et al. 2021; Yadav et al. 2022a, 2023; Gupta et al. 2022). Enhancement of nutrient uptake (N, P, K, and other vital minerals) and stimulation of plant hormones are direct actions of PGPR, while promoting plant growth by inhibiting plant pathogen and induction of ISR are indirect actions of PGPR (Ahemad and Kibret 2014, Yadav et al. 2022a, 2023). The genus Bacillus holds largest share in microbe-based agricultural commercial products. The first Bacillus species-based commercialized biofertilizer was Alinit, which enhanced crop production up to 40% (Kilian et al. 2000). Kodiak (B. subtilis GB03), Quantum-400 (B. subtilis GB03), RhizoVital (B. amyloliquefaciens FZB42), Serenade (B. subtilis QST713), and YIB (Bacillus spp.) are some examples of Bacillus-based products that have been successfully commercialized (Brannen and Kenney 1997; Ngugi et al. 2005; Cawoy et al. 2011). Because of the more efficient metabolite synthesis and spore-forming nature of Bacillus spp., which increases the viability of cells in commercially prepared products, Bacillus-based biofertilizers are more efficient than Pseudomonas-based fertilizers (Haas and Défago 2005). The Bacillus species are frequently regarded as an ideal candidate for bioformulations because of their rapid growth, ease of handling, and better colonizing abilities (Dimkić et al. 2022; Malviya et al. 2020b, 2022a, b).

This chapter deals with the many PGPR candidates from Bacillus genera, their abilities and mechanisms to enhance plant growth, their contribution to crop protection against both abiotic and biotic stressors, and their significance in ongoing strategy for increasing yields from agriculture. The Bacillus-based bioformulations for broad-spectrum application against several biotic and abiotic issues are also addressed. In this chapter we will discuss about the mechanism of Bacillus-mediated crop protection and their wide application (Singh et al. 2021a, b, Sahu et al. 2021.

1.2 Bacillus spp. and Their Role as PGPR

The rhizosphere is the small region of soil where a plant’s roots have a significant influence. Amino acids and sugars are excreted by the roots’ system as exudates, which are a major source of energy and nutrition for microorganisms in the rhizosphere. The bacterial communities are more prevalent in the rhizosphere than any other microbial communities (Sahu et al. 2019). The research evidence indicates that Gram-positive bacteria are dominating among the microbial community in rhizospheric soil. The recent findings lend credence to this idea that Bacillus species are dominating populations in the chrysanthemum of barley and grass (Smalla et al. 2001). Arabidopsis thaliana root system can secrete up to 30% of their photosynthates via exudates that are utilized by the associated B. subtilis and in return provide the plant with many growth-promoting traits (Allard-Massicotte et al. 2016). The overall actions of PGPR are depicted in Fig. 1.1, and it can be divided into two categories: (1) direct modes of action, such as stimulation of VOC and phytohormone, dropping down the level of ethylene in plant, assisting in resource acquisition (NPK and other essential minerals), N2-fixation, and stimulation of induced systemic resistance (ISR); (2) indirect modes of actions, such as when PGPR act like biocontrol agents reducing diseases, beneficial symbioses, or degradation of xenobiotic compounds present in contaminated soils (Jacobsen 1997; Sahu et al. 2020a, b; Singh et al. 2021a, b).

Fig. 1.1
A schematic diagram depicts the direct actions of P G P R, including nitrogen fixation, siderophore production, uptake of P, K, and other vital micronutrients, and volatile organic compounds. Indirect actions of P G P R include abiotic and biotic stress alleviation and induced systemic resistance.

An overview of direct and indirect PGPR actions of Bacillus spp.

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PGPR are categorized into four types on the basis of their functions by Somers et al. (2004): biofertilizers (enhance the accessibility of nutrients to the plants), phytostimulants (promote growth of plant, typically by the stimulation of phytohormones), rhizoremediators (degradation of organic pollutants), and biopesticides (produce antimicrobial metabolites to control microbial diseases). In various agricultural regimes, such as permanent grassland, grassland that has been converted into cultivable land, soil DNA from these fields was analyzed by PCR and PCR-DGGE techniques; the abundance of Bacillus-related genera including Paenibacillus, Alicyclobacillus, Aneurinibacillus, Virgibacillus, Salibacillus, and Gracilibacillus was observed (Garbeva et al. 2003). Up to 95% Gram-positive bacteria with low G + C% from various agricultural area were inferred as Bacillus species: B. mycoides, B. pumilus, B. megaterium, B. thuringiensis, B. firmus, and Paenibacillus. Out of all the soil DNA samples, less than 6% of the clones associated with Arthrobacter species and Frankia were obtained (Antoun and Prévost 2006). The monitoring of temporal and geographical variety of B. benzoevorans and related bacilli and their abundance in the bacterial population, by applying specific PCR-primer with DGGE techniques, this species was found as cosmopolitan. Such cosmopolitan bacteria play an important role in soil ecosystems ((Tzeneva et al. 2004). In 1997, whole genome of B. subtilis 168 was published with 4100 protein-coding genes making up the 4.2 Mbp-long genome (Kunst et al. 1997). Many PGPR strains of P. polymyxa species have been identified as having the ability to boost plant development and offer significant resistance to biotic and abiotic stress in plants (Timmusk and Wagner 1999; Singh and Wesemael 2022). However, this bacterium colonized only in root tips and form biofilm, and it was not detected in aerial tissues (Timmusk et al. 2005). Only culture-based methods of PGPR isolation may give false results because of resistant nature of endospores (Bent and Chanway 2002). The plant growth of many plants has been observed to be boosted by Bacillus species, and they are also highly successful as biocontrol agents in many plant microbial ailments (De Freitas et al. 1997; Kokalis–Burelle et al. 2002). The Bacillus consortium mediated induction of ISR against specific pathogens: Sclerotium rolfsii which caused southern blight of the tomato, Colletotrichum gloeosporioides which caused anthracnose of the long cayenne pepper, and Cucumber mosaic virus which caused cucumber mosaic disease were studied under greenhouse conditions. According to the findings, various PGPR mixes consistently reduced disease in all host plants. One PGPR combination, B. amyloliquefaciens strain IN937a + B. pumilus strain IN937b, substantially (P = 0.05) shielded plants from every pathogen evaluated in experiment (Jetiyanon et al. 2003). The antibiotic-producing strain B. megaterium KL39 has a broad antifungal spectrum against many pathogenic agents such as Rhizoctonia solani, Pyricularia oryzae, Monilinia fructicola, Botrytis cinerea, Alternaria kikuchiana, Fusarium oxysporum, and F. solani. For Phytophthora capsici, a MIC value of 10 g/mL was evaluated while using radioactive [3H-adenine] as the precursor; macromolecular incorporation tests with P. capsici revealed that the antibiotic KL39 drastically inhibits the DNA manufacture of the fungal cell (Jung and Kim 2005). B. subtilis isolate K18 (BS-K18) increased wilt tolerance in susceptible chickpea variety JG-62, and it also enhanced plant root and shoot development in both resistant and susceptible varieties. This putative antagonist with PGPR action may be utilized to control chickpea wilt (Suthar et al. 2017). B. subtilis RB14-C, a lpa-14-gene dependent antibiotics iturinA and surfactin producer, was found effective in reducing mortality rate of tomato seedlings caused by Rhizoctonia solani (Asaka and Shoda 1996). This strain is also resistant to chemical pesticide, flutolanil (Kondoh et al. 2001). Eighty-three isolates of the Bacillus genus from salty soils in Tunisia were evaluated for the biocontrol of dry rot of potato tubers caused by Fusarium; the most effective isolates were identified as B. cereus, B. lentimorbus, and B. licheniformis. These isolates were able to reduce the disease up to 66–89% (Sadfi et al. 2001). Their antifungal properties were linked to the ability to produce volatile compounds and a variety of complex lytic and chitinase enzymes. Increased bioavailability of nutrients, excretion of enzymes or chemical compounds to biocontrol the disease-causing agents, induction of systemic responses, enhanced secondary metabolism, and improved plant defensive responses against abiotic and biotic stresses are some main advantages of PGPR application. The PGPR traits of some Bacillus spp. and their effects on different host plants and mechanism are summarized in Table 1.1.

Table 1.1 Examples of Bacillus spp. and their PGPR action mechanisms and effects on host plant
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1.3 Bacillus spp. and N2-Fixation

Nitrogen is a major vital element required for overall plant growth. Urea and other inorganic fertilizers have been used as a source of nitrogen on a variety of crops, worldwide. Although these nitrogen fertilizers significantly increase yields, overuse of such fertilizers poses considerable risks to the environment and public health (Ahmed et al. 2017). Reducing reliance on synthetic fertilizers is indispensable for environmentally friendly agricultural practices. The capacity to fix atmospheric nitrogen and make it accessible to host plant has been explored in microorganisms. Many Bacillus and Paenibacillus spp. such as B. cereus, B. circulans, B. firmus, B. pumilus, B. licheniformis, B. megaterium, B. subterraneous, B. aquimaris, B. vietnamensis, B. aerophilus, Paenibacillus sp. WLY78, and P. mucilaginosus are known to fix atmospheric N2 (Ding et al. 2005; Shi et al. 2016; Ma et al. 2018; Liu et al. 2019). Enzyme nitrogenase is essential for nitrogen fixation; it is made of two protein parts: reductase and iron (Fe) protein and catalytic molybdenum iron (MoFe) protein (Owens and Tezcan 2018). For quantification of N2 fixation, acetylene reduction assay (ARA) (Soper et al. 2021) and for the diversity of N2-fixers and identification, nifH gene as genetic marker is widely accepted. The Paenibacillus nif operon is 11 kbp in size, and it is made up of a cluster of nif genes (Dasgupta et al. 2021). Fe-protein and Mo-fe-protein are encoded by the nifH gene and nifDK genes, respectively. The conserved region within the nifH gene has importance in the study of phylogeny and detection of molecular potential for N2-fixation in any environment (Mehta et al. 2003). P. azotofixans, a diazotrophic strain, is highly effective in fixing atmospheric nitrogen, and the ability of these bacteria to fix nitrogen is unaffected by the presence of nitrate (Choo et al. 2003). Based on acetylene reduction assay (ARA), B. circulans and B. polymyxa were identified as non-symbiotic N2-fixing in tussock-grassland soils in New Zealand (Line and Loutit 1971). Recently, P. odorifer, P. graminis, P. peoriae, and P. brasiliensis have been described as nitrogen fixers (Beneduzi et al. 2010; Li et al. 2019; Von der Weid et al. 2002). During a comprehensive study of the diversity of N2-fixing PGPR procured from sugarcane fields, an assessment of the N2-fixation ability of 22 isolates was done using the nifH gene. The most potential N2-fixing isolates, CY5 and CA1, were identified as B. megaterium and B. mycoides, respectively. Along with the nitrogen fixation ability, these two isolates were also able to biocontrol Sporisorium scitamineum and Ceratocystis paradoxa and enhance the expression of gene related to various defense and tolerance mechanisms in Saccharum spp. (Singh et al. 2020). An attempt was made to investigate the capability of heterotrophic Bacillus to fix nitrogen based on the presence of the nifH-gene; B. megaterium, B. flexus, and B. circulans were isolated from the soil samples of estuarine and coastlines in India. B. megaterium was the most dominant and strong N2-fixer bacteria among all the isolates (Yousuf et al. 2017). Non-symbiotic N2-fixing bacteria from soil of many lakes in East Java, Indonesia, were explored; isolate B2 has the most activity and was identified as B. paramycoides, a B. cereus group member (Nafisah et al. 2022). The PGPR-mediated fixation of atmospheric nitrogen and symbiotically supplying to the host plants is an important mechanism for the maintenance of nitrogen levels in agricultural lands. The biofertilizer based on B. subtilis successfully replaced 50% urea by lowering the nitrogen loss by 54% when applied to the fields. Additionally, this approach led to a 5.0% increase in crop output and an 11.2% improvement in nitrogen usage efficiency. The use of biofertilizer also led to an increase in bacterial communities of Bacteroidetes and Chloroflexi, which are crucial in the breakdown of soil organic matters (Sun et al. 2020).

1.4 Bacillus and Phosphate Solubilization

The absorption of phosphate by plant roots, which is considered to be the second most important nutrient for plants after nitrogen, is needed for optimum plant development (Bechtaoui et al. 2021). According to reports, the average amount of phosphorus (P) in soil is almost 1200 mg/kg (Tiessen 2008), but P is the least accessible to the plants in comparison to N and K, especially in low pH soils with lower organic matter and poor water-holding capacity (Yan et al. 2020). P plays an important role in the sustainability of the agricultural production system and the maintenance of soil fertility. P-based fertilizers have long been utilized successfully to replenish depleted soil phosphorus so that it is readily accessible to plants. However, the use of commercial fertilizers is a costly strategy, and since the mineral may easily be lost from the soil and subsequently mix with local streams and can pollute both terrestrial and aquatic habitats, phosphorus is frequently rendered inaccessible to plants (Adesemoye and Kloepper 2009). There are many bacteria that can mobilize insoluble forms of phosphorus, known as P-solubilizing bacteria (PSB). Many beneficial bacteria from Bacillus genus living in the soil as free or in association of plant roots are capable of solubilizing insoluble soil P and make it bioavailable to the host plants. Many species of Bacillus like B. circulans, B. cereus, B. fusiformis, B. pumilus, B. megaterium, B. mycoides, B. coagulans, B. chitinolyticus, B. subtilis, etc. have been reported as phosphorus solubilizers (Sharma et al. 2013). Due to their capacity to harness the precipitated phosphorus (P) in the soil, PSB has established themselves as significant biofertilizer that improve crop production and help in the nurture the soil health (Prakash and Arora 2019). According to Sundara et al. (2002), PSB can lower the dependency on synthetic P by 25%. Better results can be achieved when PSB are inoculated as a consortium with other PGPR or arbuscular micorrhizal fungi (AMF); its effect can rise up to 50% (Khan et al. 2009). The P solubilization mechanism of PSB is mediated by their capacity to release organic acids, i.e., gluconic and keto-gluconic acids, through their hydroxyl and carboxyl groups; their ability to chelate the cations bound to phosphate, siderophore, protons, hydroxyl ions, and CO2; as well as their ability to release extracellular enzymes and degrade substrate (Alori et al. 2017; Richardson and Simpson 2011; Heidari et al. 2020). The phosphate availability was found highest at an acidic pH (5.5) and poorest at a near-neutral pH in the crops Brassica campestris, Medicago sativa, and Oryza sativa (Barrow et al. 2020). Using chicken bones, fish bones, and ash as substrates, three PSB strains, B. megaterium PCM 1855, B. cereus PCM 1948, and B. subtilis PCM 1938, were tested to examine their ability to liberate phosphorus from unavailable structures. All the isolates were producing organic acid such as gluconic, lactic, acetic, succinic, and propionic, and maximum concentration of P was recovered from the fish bone as substrate inoculated with B. megaterium; however, this strain was not producing propionic acid (Saeid et al. 2018). PSB solubilize the mineral P by lowering the pH of micro-environments and make it available for plant uptake (Riaz et al. 2021). Isolate GQYP101, procured from the rhizospheric soil of Lycium barbarum L., was showing potential as a PSB. A pot trial with maize was conducted to assess the PGPR capabilities of the PSB, and whole genome sequencing was done to investigate the possible mechanism. This strain, identified as B. altitudinis, GQYP101 proved capable to boost the fresh weight and maximum leaf area and increased stem diameter and nitrogen (100%) and phosphorus (47.9%) levels of aerial portion in maize seedlings (Zhao et al. 2022). Alkaliphilic bacterial strain, isolated from mangrove ecosystem, B. marisflavi, was capable of tri-calcium phosphate solubilization by producing alkaline phosphatase (Prabhu et al. 2018). In addition to having a strong capacity to dissolve insoluble inorganic phosphates, a PGPR strain of B. firmus NCIM-2636 was inoculated in the fields of the Jaya and IR-8 varieties of rice (Oryza sativa L.) in Nagaland, India. For 2 years, the rice fields were supplied with single super phosphate (SSP) and Mussoorie rock phosphate (RP) as fertilizers. The findings from experiments indeed show that the bacteria generated the desired effect more strongly when implemented in combination with RP than with SSP (Datta et al. 1982). PGPR strain CB7, isolated from apple root soil, was identified as B. circulans in Himachal Pradesh, India. This isolate showed traits of P-solubilization, siderophore and auxin production, ACC-deaminase and nitrogenase activity, and antagonistic effects against Dematophora necatrix (Mehta et al. 2015). P-solubilizing Bacillus represents an economic and ecological and sustainable strategy to improve crop production. Therefore, more research is necessary to study potent biofertilizers and plant growth-stimulating properties at the field trial.

1.5 Bacillus and Potassium Uptake and Translocation

Potassium (K) is important for plant physiology, cellular development, water movement, transfer of nutrients and metabolites, and responses to stresses (Sardans and Peñuelas 2021). Although K is the most prevalent inorganic cation, only 1–2% of it can be utilized by plants; the rest of it is bonded with different minerals and is therefore inaccessible to plants. K supplied as fertilizers change the physiochemical and biological characteristics of the lands in addition to increasing crop yield. Consequently, consistent application of artificial fertilizers results in a decrease of organic matter in the soil and an overall decrease in agricultural soil health. Chemical fertilizers used in excess make the soil more hard, reduce soil fertility, pollute ecosystems, and deplete essential nutrients of soils (Pahalvi et al. 2021). Bacteria that can increase bioavailability of K by releasing the K from complex minerals and convert it to soluble forms for plant uptake are called K-solubilizing bacteria (KSB). Many Bacillus spp. such B. aryabhattai, B. mucilaginosus, B. edaphicus, B. circulans, B. cereus, B. subtilis, B. coagulans, B. amyloliquefaciens, B. megaterium, and Paenibacillus spp. have the capacity to solubilize K minerals (Yadav and Sidhu 2016; Etesami et al. 2017). KSBs solubilize K minerals in their surrounding micro-environment by altering the pH and chelation of the cations bound to K. KSB change pH by releasing the organic acids including citric, tartaric, and oxalic acids and proton pump mechanism. KSBs release K-rich minerals and other nutrients from nonsoluble mineral deposits in the soil and help in K-uptake and improve plant health (Sharma et al. 2016). B. mucilaginosus has the ability to dissolve silicate mineral in culture solutions, and the rates of dissolution considerably depend on the silicate mineral type. B. mucilaginosus produced extracellular polysaccharides that allowed it to form bacterial-mineral complexes. These bacterial-mineral complexes show different micro-environmental physicochemical characteristics in comparison to their surroundings in terms of pH value, viscosity, and organic acid concentrations. These bacterial-mineral complexes were dissolved and later transformed into secondary silicate minerals by the combined action of H+ ion exchange and acetate (Mo and Lian 2011). The potassium content was raised in cotton and rape plants growing in soils treated with insoluble potassium and inoculated with strain NBT by 30% and 26%, respectively. Additionally, higher N and P levels of above-ground plant components were a result of bacterial inoculation. This bacterial strain was able to colonize and grow in the cotton and rape rhizospheric soil (Sheng 2005). Bacillus subtilis has KtrAB, KtrCD, and KimA transporter for K uptake. KtrAB and KimA are high-affinity transporters that permit fast development at micromolar K concentrations, but KtrCD is a low-affinity transporter that is crucial for potassium absorption in a broad range of environmental circumstances (Gundlach et al. 2017). In China, B. aryabhattai SK1-7 strain was tested for K-solubilizing ability and applied it to Populus alba L. By converting insoluble potassium in the soil into usable potassium, strain SK1-7 can boost plant K concentration and plant growth (Chen et al. 2020). The tomato crop was inoculated with B. megaterium and P. mucilaginosus with biochar for 2 years in a greenhouse. The findings showed that adding 75 L/ha of microbial inoculants to greenhouse, tomato yield increased by 23.41% and concentrations of soluble sugar and vitamin C by 13.62% and 14.41%, respectively (Yang et al. 2023b). To grow sudangrass, mica waste was added to two different types of alfisols, and B. mucilaginosus was utilized as a biofertilizer. Dry biomass yield, absorption, and percent K-recoveries were found higher in bacteria-treated trial in comparison to the untreated control. B. mucilaginosus mediated a higher degree of mica solubilization in soils which was confirmed by X-ray diffraction analysis (Basak and Biswas 2009). When tea plants were cultivated on mica waste and a KSB B. pseudomycoides was inoculated, similar findings were obtained. Soil treated with mica waste and bacteria had increased potassium availability, which led to improved potassium absorption in tea plants (Pramanik et al. 2019). The K-solubilizing bacteria as biofertilizer are gaining popularity as a key component of global food security and integrated nutrient management solutions (INM) (Shrivastava et al. 2016).

1.6 Bacillus and Zinc Uptake

Zinc (Zn) is a crucial co-factor for many enzymes and DNA-binding Zn-finger proteins in animal and microorganisms. Additionally, Zn is essential for metabolism, mitosis, growth, and mitochondrial function (Frassinetti et al. 2006). Zn can be found in all kinds of enzymes and is the second most prevalent transition metal for living things (Keith et al. 2000). As per World Health Organization (WHO) reports, 4–73% of global population is facing scarcity of zinc, which impacts 31% of people globally. The top ten main risk factors for illness in developing nations are associated with zinc deficiency. The lack of micronutrients like zinc poses a serious threat to sustainability of global health. Almost 50% of soils is deficit of zinc that affects majority of agricultural production. Up to 95% of the population of Gram-positive bacteria in plant rhizosphere is composed by Bacillus spp. One of the most extensively researched bacterial species, Bacillus, has the capacity to dissolve zinc minerals (Masood et al. 2022). Numerous processes, including chelation, pH alteration, and proton extrusion, have been described in bacteria that aid in the solubilization of zinc (Yadav et al. 2022a, 2023). The sequestration of cations in bacterial cell and release of organic acids such as gluconic acid, butyric acid, lactic acid, and oxalic acid and decrease the pH in rhizospheric region enhance the bioavailability of Zn to the plant roots (Mumtaz et al. 2019). Agricultural productivity and environmental sustainability goals might benefit from the possible contribution of Bacillus genera in Zn solubilization. In a study with isolates B. aryabhattai and B. subtilis, maximum solubilization of zinc was obtained with B. aryabhattai. These isolates were suggested as prospective bio-inoculants for the biofortification of maize to help solve the issues of Zn malnutrition because of their capacity to stimulate growth (Mumtaz et al. 2017). Isolate CDK25, isolated from cow dung, was identified as B. megaterium through the 16S rRNA gene sequencing. The pot trial of Capsicum annuum L., inoculated with CDK25, enhanced growth parameters and increased zinc content in fruit (0.25 mg/100 g) (Bhatt and Maheshwari 2020). Bacterial isolates BT3 and CT8, isolated from the rhizospheric soil of chickpea from different parts of Indo-Gangetic Plains (IGP), India, were identified as B. altitudinis. Chickpea growth measures were enhanced by the inoculation BT3 and CT8, and the plant’s intake of zinc increased by 3.9–6.0%. For increasing zinc intake and chickpea development, BT3 and CT8 have the outstanding ability to solubilization of insoluble Zn compounds such oxides, phosphates, and carbonates of Zn (Kushwaha et al. 2021; Yadav et al. 2022a, 2023).

1.7 Bacillus and Siderophore Production

Small iron-chelating molecules known as siderophores have a strong affinity for metal ions. The structure of siderophores exhibits a broad range of variation. A short polypeptide chain with different coordinating iron-ligating groups makes up the majority of several siderophores (Ahmed and Holmström 2014). The Bacillus species generate a vast range of siderophores that are important to their survival, including bacillibactin, pyoverdine, pyochelin, schizokinen, and petrobactin (Crosa and Walsh 2002; Khan et al. 2016). Bacterial siderophores are divided into four main classes based on their structural characteristics, kinds of ligands, and iron-coordinating functional groups: carboxylate, hydroxamates, phenol catecholates, and pyoverdines (Chimiak et al. 1984). Many species of Bacillus, i.e., B. anthracis, B. thuringiensis, B. cereus, B. velezensis, B. atrophaeus, B. mojavensis, B. licheniformis, B. pumilus, B. halodenitrificans, and B. subtilis, are known as prolific producer of siderophores (Ramadoss et al. 2013; Goswami et al. 2014; Khan et al. 2016). In agriculture, siderophore and its derivatives are widely used to improve soil fertility, biofortification of nutrients, and biocontrol fungi pathogens (Crowley 2006). Siderophore is also used for mitigating environmental metal pollution, particularly from soil and water (Ali and Vidhale 2013). B. subtilis LSBS2, procured from sesame plant rhizosphere in Tamil Nadu, India, was producing catecholate siderophore bacillibactin (296 mg/L). This isolate promoted plant growth and increased oil content in sesame plants (Nithyapriya et al. 2021). B. megaterium and B. subtilis were found capable of siderophore production under alkaline condition. A higher iron-chelating siderophore was obtained by B. megaterium (pH 9.0,) followed by B. subtilis and A. vinelandii (Ferreira et al. 2019). Siderophores bind iron and make it less available to competing microbes and help in biocontrol of plant pathogens (Kesaulya et al. 2018). Hydroxamate siderophores producing strain Bacillus sp. PZ-1 potential enhanced the phytoextraction of Pb from soil when inoculated with Brassica juncea (Yu et al. 2017). Siderophore also forms complexes with many other vital metal ions, i.e., Mo, Mn, Co, and Ni, and make them available to the host.

1.8 Stimulation/Production of Phytohormone

PGPR found in the rhizosphere secrete the hormones auxin (IAA), gibberellin (GA), cytokinin (CK), ethylene (ET), and abscisic acid (ABA) for manipulating the balance of plant hormones, which in turn controls root/shoot growth and development as well as the plant response to stress (Sgroy et al. 2009; Antar et al. 2021). Many members of Bacillus and Paenibacillus such as B. velezensis, B. subtilis, B. megaterium, B. aquimaris, B. licheniformis, B. amyloliquefaciens, and P. polymyxa are known as potent phytohormone producer (Lim and Kim 2009). Indole-3-acetic acid (IAA) is a powerful signaling necessary for interactions between plants and microbes and has direct effects on plant development (Matsuda et al. 2018). Bacterial auxins alter the auxin pool to either a suboptimal or optimal level which enhances plant root development, particularly the formation of secondary roots, and increases root surface area, thus enhancing plant nutrition and leading to improved plant growth and productivity. B. amyloliquefaciens FZB42 can synthesize auxin through a trp-dependent pathway, and plant growth promotion was found functionally correlated in Lemna (duckweed). IAA secretion increased five times when 5 mM tryptophan was added in the medium (Idris et al. 2007). Higher auxin-producing B. cereus (So3II) (35.8 mg/mL) and B. subtilis (Mt3b) (36.6 mg/mL) were isolated from the rhizospheric soils of S. nigrum and M. tricuspidatum. Both strains were able to produce auxin without adding any substrate as a precursor. Both isolates were also producing siderophore and ACC deaminase (Wagi and Ahmed 2019). Through the transcriptome analysis of B. amyloliquefaciens SQR9 and B. subtilis 168, six genes were identified for IAA biosynthesis. IPyA pathway for IAA biosynthesis, consisting of patB, yclC, and dhaS genes, was confirmed through homologous and heterologous expression in both strains (Shao et al. 2015). B. licheniformis MML2501 can produce IAA (23 μg/mL) under optimized conditions such as pH 7.0, temperature 35 °C, and tryptophan conc. 16 mM. However, this strain failed to solubilize phosphate, but it was having a positive effect on seed germination and growth parameters when inoculated in groundnut pot trials (Prashanth and Mathivanan 2010). The mass production of IAA with cost-effective medium was done with B. circulans E9 (Sarmiento-López et al. 2022). A significant amount of ABA, IAA, CKs, and GAs was observed in a culture growth medium of B. aryabhattai strain SRB02. Plants treated with SRB02 were substantially less prone to heat stress than untreated plants. High levels of IAA, JA, GA12, GA4, and GA7, were detected in SRB02-treated plants (Park et al. 2017). Phytohormone producer B. megaterium was used in study to confirm the role of cytokinin-receptor CRE1, AHK2 and AHK3, and RPN12 in growth of A. thaliana and P. vulgaris. Plants with triple knockouts for ctk receptor CRE1-12/AHK2-2/AHK3-3 were found insensitive to bacterial inoculation in terms of growth promotion and root developmental responses, while B. megaterium was exhibited in AHK2 single and double mutation in combinations with RPN12. These findings indicate that ctk receptors function as a complementary in B. megaterium-mediated plant growth (Ortíz-Castro et al. 2008).

1.9 Bacillus and Antimicrobial Metabolites

Bacteria have biosynthetic gene clusters (BGCs) to produce different types of secondary metabolites, i.e., polyketides, terpenes, siderophores, and ribosomal and non-ribosomal synthesized peptides (Harwood et al. 2018). B. thuringiensis, B. cereus, B. velezensis, and B. licheniformis belonging to the genus Bacillus, especially B. subtilis and B. amyloliquefaciens, are extensively studied for their antimicrobial properties and their use as biocontrol agents against multiple soil-borne or postharvest plant pathogens. Many commercial biocontrol products based on Bacillus spp. have been commercialized successfully (Table 1.2). About 4–5% genome of B. subtilis is devoted to antibiotic production (Stein 2005). Many PGPR candidates of Bacillus genus have been exploited as microbial biopesticides (Kiesewalter et al. 2021). Approximately, 50% of commercially available bacterial biocontrol agents are Bacillus-based products (Arguelles-Arias et al. 2009). Antimicrobial compounds erucamide, behenic acid, palmitic acid, phenylacetic acid, and β-sitosterol were recovered, extracted, and purified from B. megaterium and tested against A. tumefaciens (T-37), E. carotovora (EC-1), and R. solanacearum (RS-2). Among all five compounds, phenylacetic acid was found most effective against all tested plant pathogens (Xie et al. 2021). Macrolactin A, 7-O-malonyl macrolactin AB, and surfactin B-producing B. amyloliquefaciens HR62 was applied on tomato with a biofertilizer, BIO62, against R. solanacearum. This combination successfully inhibits the growth of R. solanacearum, thereby decreasing disease incidence to 65% (Huang et al. 2014). B. amyloliquefaciens strain FZB42 can inhibit fire blight in orchard trees caused by Erwinia amylovora, by producing antibacterial polyketide difficidin and dipeptide bacilysin (Chen et al. 2009). Organic solvent-based crude extract of B. velezensis DTU001 strain can inhibit the growth of plant pathogenic fungi, A. uvarum, P. ulaiense, and P. expansum. The main bioactive lipopeptides were characterized as iturins, fengycins, and surfactins. Secondary metabolites, non-ribosomal production of antimicrobial chemicals, and siderophores account for 15.4% of the whole genome of DTU001 (Devi et al. 2019, Malviya et al. 2020a, b, 2022a, b). B. subtilis 22 and Trichoderma atroviride SG3403 used together shown improved antifungal activity against Fusarium graminearum. The lack of antimicrobial activity of the fungus SG3403 as a single culture shows that interactions between the two microorganisms in a coculture setting may result in the development of certain antifungal compounds. LC–MS analysis revealed eight specific compounds including mevastatin and koninginin which were present only in co-culture (Li et al. 2020).

Table 1.2 Some examples of commercial biocontrol products based on Bacillus spp. and their applications
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1.10 Bacillus and Volatile Organic Compounds (VOCs)

Microbial volatile organic compounds (MVOCs) are a group of chemicals produced by fungal and bacterial metabolic reactions. Bacterial VOCs occur in a variety of chemical forms, such as ketones, alcohols, terpenoids, sulfur compounds, alkenes, etc. Some chemicals are shared by the entire group of microorganisms, while others are exclusive to distinct strains. A single bacterium can produce up to 100 distinct VOCs (Sidorova et al. 2021). VOCs are important for intra- and interspecific communication between bacteria and hosts. Plants synthesize VOCs in order to protect themselves from herbivores and diseases, communicate with other plants, or feed microbes. Microorganisms release VOCs to communicate, attack, or defend themselves. Due to their capacity to either trigger the plant defenses mechanisms, limit the expansion of plant diseases, or encourage plant growth and development, microbial VOCs (MVOCs) play significant role in to plant’s health status (Poveda 2021). Bacillus spp. are considered as factories of plant protective VOCs and have various roles in plant protection against bacterial and fungal diseases, induction of ISR, and plant growths (Poulaki and Tjamos 2023). Several Bacillus spp. particularly B. amyloliquefaciens, B. velezensis, B. subtilis, and B. altitudinis are most explored for their VOC-producing abilities (Grahovac et al. 2023). A number of VOC compounds such as 2-decanone, benzothiazole, phenol (4-chloro-3-methyl) 2,3,5-trimethylpyrazine, 2-nonanone, 2-dodecanone, styrene, 1-tetradecene (Yuan et al. 2012; Guevara-Avendaño et al. 2019; Gao et al. 2017; Insam and Seewald 2010) have been isolated from different Bacillus spp. and found effective against many plant pathogens. VOCs produced by B. subtilis AP-3 have positive effects on plant biomass of shoot and roots of soyabean by 88% and 18%, respectively, compared to the untreated control. The root architecture of soybeans exposed to VOCs was found altered with increased length, diameter, surface area, and numbers (Bavaresco et al. 2020). VOCs produced by B. amyloliquefaciens CPA-8 strain were showing promising results in sweet cherry fruit against postharvest pathogens Monilinia laxa, M. fructicola, and Botrytis cinerea. The VOCs were identified as 1,3-pentadiene, acetoin (3-hydroxy-2-butanone), and thiophene by solid-phase micro-extraction (SPME)-gas chromatography. The mycelial development of all target pathogens could be suppressed by all identified VOCs; however, thiophene was the most potent VOC, displaying more than 82% reduction of mycelial growth (Gotor-Vila et al. 2017). VOCs can modify the plant hormone levels by directly acting on related genes. In response to B. subtilis SYST2 VOCs albuterol and 1,3-propanediol, tomato plants displayed differential expression of genes involved in the production or metabolism of IAA (SlIAA1 and SlIAA3), GAs (GA20ox-1), Cks (SlCKX1), expansin (Exp2, Exp9, Exp18), and ET (ACO1) in plants. VOCs produced by B. methylotrophicus BCN2 and B. thuringiensis BCN10 can suppress the growth of postharvest pathogens F. oxysporum, Botryosphaeria sp., T. atroviride, C. gloeosporioides, and P. expansum (He et al. 2020).

1.11 Bacillus Thuringiensis (Bt) and CRY Proteins

B. thuringiensis (Bt), B. popilliae, and B. sphaericus are well known for their insecticidal properties. However, Bt is most studied and commercialized as biopesticide because it is safe for humans and has a broad-spectrum insecticidal nature (Ibrahim et al. 2010). Bt-based biopesticides are considered as the safest and most environment-friendly insecticides (Caballero et al. 2020). Bt produces crystal (Cry) and cytolytic (Cyt) proteins in the sporulation phase and Vip and Sip proteins in the vegetative growth phase (Gonzalez-Vazquez et al. 2021). The cry protein consists of proteins called δ-endotoxin. Cry proteins create pores and disrupt the gut epithelial membranes of juvenile insects (Latham et al. 2017). These proteins are toxic against a wide range of insect orders such as Coleoptera, Lepidoptera, Diptera, Hemiptera, and Hymenoptera, nematodes, and human cancer cells (Palma et al. 2014). The Cry protein family is categorized into at least 50 subgroups with more than 200 members (Bravo et al. 2007). Biopesticide manufactured from B. thuringiensis (Bt) contributes to over three quarters of the 3 billion dollar global market. Cry proteins were obtained by transgenic expression of the Bt gene in E. coli BL21, and their toxicity was evaluated against H. armigera larvae. On behalf of LD50 and ID50, Cry1Ab, Cry1Ac, and Cry2Aa, Cry proteins were found most toxic (Li and Bouwer 2012). A patent (No. 336230) was granted on “a process for the mass production of Bacillus thuringiensis (Bt) biocide using millet grain based agro-medium” invented by ICAR-VPKAS, Almora, India. This cold-tolerant strain Bacillus thuringiensis subsp. galleriae/colmeri (MTCC 8997) was isolated from North-Western Himalaya, India, and exhibited tendency of producing larger quantity of highly insecticidal cry proteins against many plant-harming insect (Kant and Shahid 2022).

1.12 Bacillus-Mediated Abiotic Stress Tolerance in Plants

Drought or excessive water condition, low or high temperature, unusual pH, salinity, metal pollution, and UV radiation all work against the growth and development of plants, limiting their ability to flourish and lowering agricultural productivity (He et al. 2018). PGPR-mediated physiochemical changes in plants that help in the alleviation of abiotic stress are called “induced systemic tolerance” (IST) (Yang et al. 2009). Low crop yield due to drought conditions is a cumulative result of diminished stomatal function, decreased transpiration and photosynthesis rates, and early crop maturity. PGPR can alter the root structure, phytohormone pool, osmolyte deposits, antioxidant enzymatic activity, and expression of defense-associated genes (Seleiman et al. 2021). Alleviation of drought stress in A. thaliana with the inoculation of PGPR P. polymyxa was confirmed by Timmusk and Wagner (1999). This strain successfully stimulates drought tolerance responsive gene, ERD15. In response to abiotic stressors, PGPR can modify the levels of the phytohormones JA, ABA, BR, IAA, GAs, SA, CKs, and ET through different pathways. PGPR has ACC deaminase enzymes that convert ACC into α-ketobutyrate and ammonia, deplete ET ethylene level, and enhance abiotic stress tolerance in plants (Gupta and Pandey 2019; Misra et al. 2017). Potential traits of the mitigation of drought and heavy metal toxicity were shown by B. cereus and B. haynesii, isolated from Vigna mungo and Phaseolus vulgaris. Both were showing ACC-deaminase activity, EPS, and IAA production and tolerant to As, Ba, and Ni metals (Andy et al. 2023). The dual culture inoculation of arbuscular mycorrhizal fungus R. irregularis and B. megaterium in maize enhanced tolerance against drought and high temperature stress (Romero-Munar et al. 2023). B. endophyticus PB3, B. altitudinis PB46, and B. megaterium PB50 were compared to effectiveness in eliciting of drought tolerance in rice. B. megaterium PB50 was most effective as it improved the contents of water, total sugar, proteins, proline, phenolics, K, Ca, ABA, and IAA and upregulated the expression of stress-related genes (LEA, RAB16B, HSP70, SNAC1, and bZIP23) (Arun et al. 2020). The enhancement of antioxidant enzymes glutathione peroxidase and peroxidase as well as upregulation of genes associated with stress suppression, B. butanolivorans KJ40, can reduce the drought stress in pepper (Kim et al. 2022). When four varieties of wheat crop (V1: Akbar 2019; V2: Dilkash 2021; V3: Faisalabad 2008; and V4: Subhani 2020) seed were treated with Bacillus subtilis NA2, plants were found more tolerant to salinity stress in comparison to control (Gul et al. 2023). Bacillus altitudinis WR10 can improve the health of rice plant under high-salinity and low-phosphorus conditions. There are numerous examples of Bacillus-mediated tolerance against abiotic stresses in plants (Table 1.3).

Table 1.3 Bacillus-mediated abiotic stress tolerance in different hosts and their effects on plants
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1.13 Bacillus-Mediated ISR in Plants and Biotic Stress Tolerance

PGPR can mitigate biotic stress through the hyperparasitism, production of antimicrobial compounds (De Vrieze et al. 2018), ISR induction (Alizadeh et al. 2013; Martínez-Medina et al. 2017; Romera et al. 2019), and competition for nutrients and niche space (Recep et al. 2009; Vanitha and Ramjegathesh 2014; Lastochkina et al. 2019). The resistance induced against pathogens in plants by nonpathogenic antagonistic rhizobacteria is known as induced systemic resistance (ISR) in general (Ryu et al. 2004; Walters et al. 2005). B. amyloliquefaciens, B. subtilis, B. pasteurii, B. cereus, B. pumilus, B. mycoides, and B. sphaericus can significantly reduce the occurrence or impact of several diseases on a variety of hosts, through the ISR induction (Choudhary et al. 2007) (Table 1.4). Systemic acquired resistance (SAR) is mediated by SA and accretion of PR proteins, while ISR is mostly dependent on JA and ET signaling (Pieterse et al. 2001). The signaling pathway may be different and PGPR host specific. The pathogenesis-related (PR) gene may not be necessary for ISR to function because it is largely a JA- and ET-dependent mechanism (Romera et al. 2019; Alizadeh et al. 2013). ISR enables the whole plant to develop more resistance to pathogens. PGPR can induce transcription factor MYB72, hormones, and signal molecules such as auxins and nitric oxide (Zamioudis et al. 2015). The process of PGPR-induced ISR is not fully understood, although research findings suggested that VOCs and microbe-associated molecular patterns (MAMPS) are some of the significant elicitors. Phenylacetic acid (PAA), extracted from B. fortis IAGS162, was found potential ISR elicitor that significantly reduced Fusarium wilt disease and altered metabolite profile of tomato (Akram et al. 2016). B. pumilus INR7 induced ISR against bacterial spot caused by X. axonopodis pv. vesicatoria in pepper. The combination of INR7 and benzothiadiazole (BTH) leads to the enhanced expression of defense-related marker genes CaPR1, CaTin1, and CaPR4 (Yi et al. 2013). PGPR strain B. cereus AR156 can provide resistance to a variety of diseases including P. syringae pv. tomato DC3000. NPR1-dependent activation of SA, JA, and ET signaling pathways and simultaneous expression of defense-related genes PR1, PR2, PR5, and PDF1.2 were observed when Arabidopsis was treated with AR156 (Niu et al. 2011). B. subtilis and P. fluorescens share same mechanism of ISR induction through the JA/ET and NPR1-dependent against B. cinerea but in case of Pst DC3000, B. subtilis elicit ISR induction relies on SA, JA/ET, and NPR1 while P. fluorescens was found depend on SA pathway only. ABA signaling is also very crucial along with JA/ET signaling in primed systemic immunity by beneficial bacteria against Pst DC3000, but not against B. cinerea (Nguyen et al. 2020). B. proteolyticus strain OSUB18 elicited the ISR against P. syringae and Botrytis cinerea in Arabidopsis plants. Plant treated with OSUB18 showed higher expression gene related to plant hormone SA (PR1, PR2, PR5, EDS5, and SID2) and JA (PDF1.2, LOX3, JAR1, and COI1) in comparison to the control (Yang et al. 2023a).

Table 1.4 Bacillus-mediated and ISR responses and biotic stress tolerance in plants
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1.14 Future Aspects

Putting together the information found in the literature, in conclusion, we saw that many Bacillus spp. are able to regulate plant growth, development, and tolerance against abiotic and biotic stresses, which certainly increases the importance of their practical application in an effort to boost agricultural productivity and sustainability. The findings from the literature show Bacillus genera have huge potential use as biofertilizers, biostimulants, biopesticides, and ISR elicitors against biotic stresses and defense against abiotic stress. Understanding their underlying mechanisms of physiochemical activity on host plants under various environmental and biotic stresses is crucial to maximizing the potential and application of PGPR in sustainable agricultural output.