Developing Liquid Rhizobium Inoculants with Enhanced Long-Term Survival, Storage Stability, and Plant Growth Promotion Using Ectoine Additive.

Liquid microbial inoculants have recently received great attention due to their vital roles for sustainable agricultural practices. However, long-term conservation under ambient temperature conditions and deleterious environmental factors might negatively impact microbial cell survival and limit their efficacy in the field. Thus, developing efficient liquid formulation providing prolonged survival of rhizobia in the final product and after an application is crucial. Therefore, this study investigates the effect of various additives on the long-term survival of rhizobia stored in liquid cultures at room temperature (25 °C) for 12 months. Various yeast sucrose media amended with polyvinylpyrrolidone (PVP) or gum arabic as colloidal agents in combination with ectoine (as a compatible solute) and/or glycerol were evaluated. A dramatic decline in viable cell count was obtained in formulas amended only with PVP from Log 8.5 to Log 5 in the first six months and then to Log 1.5 after 12 months. In contrast, rhizobia stored at PVP-based formulas amended with 10 mg L ectoine exhibited almost constant survival level till the end of the storage period. The same trend was obtained using formulas based on gum arabic as a colloidal dispersing agent; however, less decline in cell count using a formula containing gum arabic alone as compared to using PVP. On the other hand, PVP based formulas exhibited higher viscosity compared with another formula. Increased viscosity till the 8th month of storage was achieved in the presence of ectoine indicating the increase of exopolymeric substances production. Electrophoretic protein pattern of rhizobial cells (stored for 12 months) exhibited several low molecular weight protein bands in cells stored in PVP based formula with ectoine as compared to the other treatments. Thus, the amendment of the liquid formulation of rhizobia bioinoculant with PVP plus ectoine not only improved cell survival but also enhanced the culture viscosity and consequently ameliorate the colonization and performance of rhizobial inoculants.