Tissue culture propagation of Banana (Musa spp.) proved to be the alternative commercial tool for traditional vegetative propagation through suckers. The aim of this study was to explore the application of bacterial culture supernatant as a substitution for commercial indole acetic acid (IAA) for improving plant tissue culture production.

Up to 200 bacterial isolates were obtained from different banana plant spheres (rhizosphere, endorhiza, and stem-endosphere) and screened for their in vitro ability to produce IAA. About 50% of isolates were able to produce IAA with different amounts; the most promising IAA producing isolates were further identified according to the 16S rRNA gene sequencing. The chemical composition of the culture supernatants of most promising IAA producing isolate; Brevibacillus brevis-R39 and Klebsiella variicola-R19 were determined using GC-MS analysis, then the culture supernatants of Brevibacillus brevis-R39 was selected as Murashige and Skoog (MS) medium supplement at different doses (5, 10, 20, and 40 ml/l), corresponding to 0.21, 0.42, 0.4, and 1.68 mg IAA/l, respectively) in comparison to commercial IAA (0.5, 1, 2, and 4 mg/l). The results demonstrated that the culture supernatant of Brevibacillus brevis-R39 at 10 ml/l significantly showed the highest average length of banana shoots (11.9% over the control), followed by its 5ml/l (7.8% over the control). The greatest width of leaves (24% over the control) was recorded with the bacterial extract at concentration of 5 ml/l, followed by 13.2% over the control with bacterial extract at 10 ml/l. The greatest length of roots (49% over the control) was recorded with the bacterial extract at 10 mg/l, followed by 22% over the control with IAA at 2 mg/l. Our results recommend the use of Brevibacillus brevis-R39 culture supernatant as a medium supplement as a substitution for commercial IAA to improve banana plantlet properties.