Several microorganisms display beneficial effects on plants in nature and may be used to enhance growth and crop yields as an alternative to chemical fertilizers. Bacillus species constitute the largest class of growth-promoting bacteria. Bacillus-plant interactions induce plant growth through growth-responsive genes, proteins, phytohormones, and metabolites [11]. Plant growth promotion analyses were performed to evaluate whether B. aryabhattai could promote plant growth in different plant species. Arabidopsis (Fig. 1) and N. tabacum (Fig. 2) plants treated with the bacterium showed increased growth, and significant increases were obtained in plant size and fresh weight between the treated and nontreated plants. Notably, although this endophytic bacterium was isolated from wild plant species, it exhibited a robust impact on the growth of nonhost plants. The results show that B. aryabhattai can enhance the growth of different plant species, thus allowing it to be used to enhance the growth of plants under field conditions.
Diverse bacteria from the Bacillus genus were shown to be good plant growth-promoting bacteria [11, 13, 19, 21, 26]. The first commercial biofertilizer was obtained from Bacillus spp. and enhanced crop yields by 40% [16]. Additionally, biofertilizers using Bacillus species are more effective at producing diverse metabolite, forming spores, and maintaining cell viability. These characteristics allow the generation of formulated products suited for commercial use [27]. Biofertilizers are good candidates as alternatives to chemical fertilizers to promote plant growth and yields [12]. Additionally, Bacillus species related to roots or rhizospheres could be used to develop biofilms to enhance plant growth [28].
The size and weight of shoots, leaves, and roots from various plant species were enhanced after the application of B. insolitus, B. subtilis, and B. methylotrophicus, respectively [13, 15]. The production of phytohormones such as indole-3-acetic acid (IAA), cytokinins, gibberellic acid (GA), and spermidines was increased in plants treated with B. subtilis and B. methylotrophicus and induced plant growth [15, 19, 26]. Additionally, the induction of endogenous proteins, amino acids, and minerals by B. megaterium and B. methylotrophicus promoted plant growth [15, 29]. Interestingly, B. aryabhattai displayed some plant growth-promoting features, resulting in improved growth of Arabidopsis and N. tabacum plants. However, these effects need to be investigated under natural conditions and with crop species. Considering the effect of B. aryabhattai on plants, this bacterium could be used to promote plant growth as a biofertilizer under field conditions.
Plant-beneficial bacterial interactions have been extensively analyzed. However, it is not clear which specific molecular pathways are associated with these interactions. This information is important to enhance the potential of these classes of bacteria under field conditions. In the current study, we used RNA sequencing to analyze the genes expressed during Arabidopsis-B. aryabhattai interactions. A high number of novel genes involved in metabolite biosynthesis were differentially expressed in our dataset. The results reveal new insights into plant and bacterial gene expression and assist in our understanding of the molecular events implicated during Arabidopsis-B. aryabhattai interactions. Notably, GO and KEGG analyses showed significant changes between treated and nontreated plants. B. aryabhattai has a remarkable impact on plants. Our data indicated that B. aryabhattai triggered important molecular pathways related to plant growth.
Curiously, cinnamyl alcohol dehydrogenase, apyrase, thioredoxin H8, benzaldehyde dehydrogenase, indoleacetaldoxime dehydratase, berberine bridge enzyme-like, gibberellin-regulated protein, maturase K, tetratricopeptide repeat (TPR)-like superfamily protein, BTB/POZ and TAZ domain-containing protein and auxin-responsive GH3 family protein genes were highly induced during the Arabidopsis-B. aryabhattai interaction.
Cinnamyl alcohol dehydrogenase is a key enzyme during plant secondary metabolism, especially lignin synthesis, and it is closely related to plant growth and development. Lignin constitutes one of the major components of plant cell walls and has the function of connecting cells. Previously, this enzyme was expressed in lateral roots and in root tips in sweet potato, and its activity was induced by abscisic acid [30]. Additionally, cinnamyl alcohol dehydrogenase genes are related to lignin biosynthesis during the final developmental phases of soybean seeds [31]. Most likely, B. aryabhattai promotes the synthesis of lignin during the growing phase of Arabidopsis and N. tabacum plants, resulting in the robust phenotype observed for the plants treated with the bacterium.
Likewise, the extracellular nucleotides can be regulated by apyrases. Apyrases are involved in the control of plant growth and development. Specifically, apyrases have influence in the auxin transport and stomatal aperture. Removal of the apyrases activity can leads to growth inhibition [32]. Potato plants with apyrase gene silenced showed phenotypic changes, retardation of growth, increasing of tuber number per plant, and effect on tuber morphology [33]. Meanwhile, the expression of apyrases gene in Arabidopsis plants had a marked effect on the growth of plant tissues and accumulation of auxin levels [34, 35]. Thus, there was significant evidence that apyrases developed a crucial role in regulating the growth of Arabidopsis and N. tabacum plants.
Besides, benzaldehyde dehydrogenase is an important enzyme involves in the processing of benzaldehyde to benzoic acid. The growth, mineral composition, and chlorophyll content of soybean plants were influenced by benzoic acid [36]. Benzoic acid had a remarked effect on the growth and yield of tomato plants. Additionally, there was a positive effect of benzoic on fruit yield [37]. Interestingly, B. aryabhattai could be indirectly inducing the plant growth through this enzyme, which displays a key role in the benzoic acid pathway.
Gibberellins are involved in plant growth, development processes, stem elongation, flowering, and seed germination [38]. Vegetative and reproductive growth were severely affected in rice plants expressing a gibberellin-regulated gene in antisense orientation [39]. In addition, maturase K gene was highly expressed in Anoectochilus roxburghii plants treated with endophytic fungi [40]. Recently, the maturase K gene was induced during the Arabidopsis - Bacillus altitudinis interaction [41].
Further, proteins with tetratricopeptide repeat motifs are basic components for gibberellin and ethylene responses. A silencing of an Arabidopsis chloroplast-localized tetratricopeptide repeat protein gene affected the plant growth, leaf greening, chloroplast, and genes involved in photosynthesis [42]. Recently, an endophytic B. altitudinis induced Arabidopsis tetratricopeptide repeat-like superfamily proteins genes with a marked effect on plant growth [41]. Moreover, broad-complex, tram track, and bric-a-brac family proteins (BTB) genes had a high influence on transcription, protein modification, chromatin, cytoskeletal, and hormone pathways in tomato [43]. These kind of genes could be implicated indirectly in the activation of phytohormone related with plant growth.
The bacterium B. aryabhattai induced the plant growth by the trigger of key molecular pathways, involved in the production of phytohormones and transcription factors. The root development, shoot growth, and fruit ripening were regulated by Aux/IAA family genes [44]. Auxin influences numerous stages of plant development and growth by directing the expression of auxin-activate genes [45]. Auxin controls plant development and growth by changing the induction of different genes [46].
While much of what happens in the plant during interactions with endophytic bacteria is known, it is also important to understand what happens in the bacteria during its interaction with the plant, such as which genes are expressed in the bacterium that may contribute to the growth phenotype of treated plants. Interestingly, arginine decarboxylase, D-hydantoinase, ATP synthase gamma chain and 2-hydroxyhexa-2,4-dienoate hydratase genes were highly induced in B. aryabhattai during the interaction with the plant, which constitutes the first evidence of the expression of these kinds of genes in this species. We speculate that the overexpression of these genes in B. aryabhattai might enhance plant growth.
The activity of arginine decarboxylase was implicated in the effect of hormones on plant growth [47]. Arginine decarboxylase is an important enzyme responsible for putrescine biosynthesis. Arginine decarboxylase expression correlates with cell growth and stress responses in apple plants [48]. This enzyme is involved in efficient ROS elimination and its influence on root growth, which is conducive to drought tolerance [49].
In addition, an active D-hydantoinase from Pseudomonas fluorescens was related to the synthesis of D-amino acids [50]. Although plants are constantly exposed to D-amino acids (D-AAs) in the rhizosphere, these compounds have inhibitory effects on plant growth. A recent characterization of D-AA-stimulated ethylene production in Arabidopsis showed the physiological function of a specific D-AA and its metabolizing enzyme in plants [51]. Most likely, the regulation of plant D-AA content could influence the composition of the rhizosphere [52].