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Plant growth-promoting bacteria (PGPB) can mitigate the effect of abiotic stresses on plant growth and development; however, the degree of plant response is host-specific. The present study aimed to assess the growth-promoting effect of Herbaspirillum (AP21, AP02), Azospirillum (D7), and Pseudomonas (N7) strains (single and co-inoculated) in perennial ryegrass plants subjected to drought. The plants were grown under controlled conditions and subjected to water deficit for 10 days. A significant increase of approximately 30% in dry biomass production was observed using three co-inoculation combinations (p < 0.01). Genomic analysis enabled the detection of representative genes associated with plant colonization and growth promotion. In vitro tests revealed that all the strains could produce indolic compounds and exopolysaccharides and suggested that they could promote plant growth via volatile organic compounds. Co-inoculations mostly decreased the in vitro-tested growth-promoting traits; however, the co-inoculation of Herbaspirillum sp. AP21 and Azospirillum brasilense D7 resulted in the highest indolic compound production (p < 0.05). Although the Azospirillum strain showed the highest potential in the in vitro and in silico tests, the plants responded better when PGPB were co-inoculated, demonstrating the importance of integrating in silico, in vitro, and in vivo assessment results when selecting PGPB to mitigate drought stress.

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Perennial ryegrass is a forage commonly used in temperate regions for livestock feeding; however, its yield is affected by reduced biomass production under water deficit. In a previous study, three co-inoculations of beneficial bacteria were selected based on their ability to promote plant growth under reduced water availability. The aim of this work was to elucidate some mechanisms by which the selected bacteria can help improve the response of perennial ryegrass to water deficit. Ryegrass plants were inoculated with each of the co-inoculations (Herbaspirillum sp. AP02–Herbaspirillum sp. AP21; Herbaspirillum sp. AP02–Pseudomonas sp. N7; Herbaspirillum sp. AP21–Azospirillum brasilense D7) and subjected to water deficit for 10 days. Physiological and biochemical measurements were taken 10 days after stress and shortly after rehydration. The results showed that bacteria had a positive effect on shoot biomass production, dissipation of excess energy, and proline and chlorophyll pigments during the days of water deficit (p < 0.05). The leaf water status of the inoculated plants was 12% higher than that of the uninoculated control after rehydration. Two Herbaspirillum strains showed greater potential for use as biofertilizers that help ameliorate the effects of water deficit.