The microbe SSAU-2 displayed the efficient capability for indole-3-acetic acid (IAA) synthesis. Numerous studies have previously highlighted the active involvement of Bacillus species, such as Bacillus subtilis, Bacillus megaterium, and Bacillus amyloliquefaciens SQR9, in the phyllosphere and their significant contributions to soil micro-engineering [13, 14, 15, 16].
The SSAU-2 has been observed to exhibit a unique behavior of producing indole-3-acetic acid (IAA), even under highly stressful conditions. This microbe has the ability to thrive even in environments with high concentrations of PEG-6000, which could be advantageous for arid or semi-arid regions across the globe. Additionally, the bacteria can resist artificial drought conditions and effectively produce IAA even in concentrations of 15% of the aforementioned compound, which could be a significant development for regions with low precipitation levels. The abilities to release multiple plant growth promoting compounds can enhance the barren land around the globe. The EPS secretion by these microbes is one of the crucial mechanisms to improve the soil fertility by enhancing the soil aggregation and water capture. Ashry et al. (2022) and Rashid et al. (2021) have produced similar results, highlighting the potential of Bacillus sp. in producing plant growth hormones under artificial drought conditions [17, 18]. These microbes have proven to be essential to the ecosystem, particularly in alleviating drought stress [19].
Another unique property of the SSAU-2 is its ability to produce IAA under the presence of Cr (VI). The bacteria can remove Cr (VI) up to 50 ppm and tolerate it at high concentrations, making it a versatile microbe for diverse ecosystems. These characteristics can be used to alleviate the Cr (VI) contaminated land and make it re-fertile. Srinivas Ravi has also demonstrated that Bacillus spp. can enhance soil fertility by releasing IAA under Cr (VI) stress [20]. Furthermore, another study has shown that these microbes play an active role in enhancing the productivity of various crops under the abiotic stress of Cr (VI) [21].
Bacillus spp. SSAU-2's efficient IAA biosynthesis potential, particularly at basic pH levels, suggests its applicability in industrial sites contaminated with high-pH conditions. Interestingly, these findings contrast with a study by Kumar et al. (2023), which reported optimal IAA biosynthesis at acidic pH [22]. However, our results align with research conducted by Lebrazi et al. (2020) and Suliasih et al. (2020), demonstrating that certain rhizobacterial strains can optimally synthesize IAA under basic pH conditions [23, 24]. This suggests that Bacillus spp. SSAU-2 can exhibit high efficiency in soils with excessively alkaline pH levels, presenting it as a valuable candidate for sustainable plant growth promotion.
Salinity-related studies indicate that a higher saline environment is necessary for increased auxin synthesis, but salinity levels greater than 30 g/l reduce auxin synthesis efficiency, as the microbe can only thrive up to a certain salinity level. In contrast to the findings of Lebrazi et al. (2020), who reported 2% salinity as the optimum for IAA biosynthesis, this study identified 3% salinity as the optimal condition for IAA synthesis by Bacillus spp. SSAU-2 [23]. The demonstrated ability of SSAU-2 to synthesize IAA under alkaline and saline conditions underscores its potential to play a comprehensive role in revitalizing semi-arid or salinity-affected soils.
The dependence on an initial tryptophan concentration of up to 1% indicates that the microbe possesses a pathway induced by tryptophan presence in the media. However, the optimal tryptophan concentration for auxin synthesis is 1%, beyond which the microbe does not exhibit enhanced synthesis characteristics. In the absence of tryptophan, IAA synthesis was not observed in strain SSAU2, indicating that the strain requires the presence of tryptophan as an inducer for auxin synthesis. Co-culturing with tryptophan-producing microbes can enhance IAA synthesis, as demonstrated by Raut et al. (2017) using Azolla and IAA producer microbes [25]. Similar findings were reported by Kumar et al. (2023), showing that increasing tryptophan concentration proportionally improves IAA biosynthesis by the microbe [22]. Additionally, Lebrazi et al. (2020) and Suliasih et al. (2020) reported that after reaching a certain concentration of tryptophan, IAA biosynthesis reaches equilibrium, consistent with the results of this study [23, 24]. This research provides insights into the impact of different carbon and nitrogen sources on auxin synthesis by Bacillus spp. SSAU-2, contributing to our understanding of its growth and auxin production dynamics.
The nitrogen source findings highlight the positive impact of nitrogen sources containing tryptophan on auxin synthesis by Bacillus spp. SSAU-2. Consistent with our results, Kumar et al. (2023) showed that peptone and yeast extract supported high IAA synthesis, while ammonium chloride negatively impacted it [22]. However, in this study, ammonium chloride only slightly affected IAA biosynthesis. Suliasih et al. (2020) also reported similar results [24]. These insights into the influence of nitrogen sources on auxin synthesis provide valuable information for optimizing microbial auxin production and understanding the microbe's nutritional requirements.
TLC (thin-layer chromatography) studies revealed the formation of various auxins when different carbon and nitrogen sources were used. This finding, not reported elsewhere, is a key phenomenon in this study. Kumar et al. (2023) used propanol: water as a mobile phase in the ratio of 8:2 and reported an Rf value of 0.91 for IAA [22]. Panigrahi et al. (2020) reported an Rf value of 0.77 for synthesized IAA, which differs from our findings [29]. Parvin et al. (2020) explored the effect of different mobile phases on the Rf value of IAA, indicating that variations in mobile phases can impact the Rf value [30]. The different colors observed after spraying with Salkowski's reagent also indicate the synthesis of different types of auxins by Bacillus Sp. SSAU-2. This phenomenon of different colors of auxins with Salkowski's reagent has been reported by Sumera et al. (1981) [31]. The significant differences in Rf values depending on carbon and nitrogen sources highlight the complexity of IAA synthesis and its potential applications as a bio-fertilizer.
The microbe's diverse capabilities for synthesizing various phytohormones and other plant growth-promoting compounds suggest its unique potential in fertility restoration. Similar findings by Desoky et al. (2020) highlight the capabilities of Bacillus cereus in secreting IAA and other plant growth-promoting compounds, confirming the effectiveness of Bacillus species in enhancing soil fertility and plant growth through diverse mechanisms [32]. Bacillus spp. SSAU-2 presents a promising bio-fertilizer candidate, capable of comprehensively improving plant health and soil quality [33, 34]. In the previous reports by our lab it is clear that SSAU-2 not only exhibits plant growth-promoting activities but also demonstrates its potential in enhanced seed development and controlling soil parameters [35]. The consortia approach with Nostoc commune algae further enhances the beneficial effects, offering a promising strategy for sustainable agriculture and soil restoration [36].