The research question focuses on determining the effectiveness of different PGPB species and application methods (foliar vs. soil) on enhancing maize growth, physiology, yield, and seed quality under Mediterranean conditions. The context is the global push for more sustainable agricultural practices, with PGPB emerging as a potential biostimulant. The study's purpose is to assess the impact of various PGPB strains and application techniques on maize, a major cereal crop. The importance lies in the potential to improve maize productivity and quality using environmentally friendly methods, reducing reliance on chemical fertilizers. This study builds upon existing research demonstrating the positive effects of PGPB on various plant species, but seeks to refine understanding by examining different application methods and their interactions with specific PGPB strains. The lack of comprehensive data on the effects of PGPB on maize yield and seed quality under Mediterranean conditions specifically motivated this investigation.

Previous research has extensively explored the use of various beneficial bacteria as biostimulants in agriculture, highlighting the positive impacts of species such as *Arthrobacter*, *Acinetobacter*, *Enterobacter*, *Ochrobactrum*, *Pseudomonas*, *Rhodococcus*, and *Bacillus*. While the mechanisms are not fully elucidated, studies suggest PGPB enhance plant growth through growth regulator production, nitrogen fixation, antagonism against phytopathogens, and phosphate solubilization. Some hypotheses propose PGPB interact with plant signaling processes and mitigate stress responses. The use of indigenous strains adapted to local environmental conditions is often emphasized due to their superior competitiveness and effectiveness. Both foliar and soil application methods have been explored, with foliar application requiring further investigation into the leaf surface-microbe interaction. Previous studies have shown positive impacts of foliar application on various crops, including increased yield and improved quality characteristics. Soil inoculation, whether on seeds, seedlings, or directly into soil, has also demonstrated efficacy in improving plant physiology and disease resistance. However, knowledge on PGPB's effects on nutritional value and harvested product quality remains limited, emphasizing the need for further investigation, particularly with significant crops like maize.

A field experiment was conducted in Oropos, Greece, from April to August 2019 using a corn hybrid (Zea mays, GW 8002). The experiment employed a completely randomized design (CRD) with three replications and 12 treatments (two application methods × six PGPB treatments). The PGPB treatments included *Azotobacter chroococcum*, *Bacillus subtilis*, *Bacillus megatherium*, and two 1:1 mixes (A. chroococcum + B. subtilis and A. chroococcum + B. megatherium), along with a control. Each replication covered 12 m². PGPB solution (7 L/ha, diluted 1:100) was applied once at 40 days after sowing (DAS), either folia (spraying) or to the soil near sowing rows. Soil analysis was performed before sowing to determine nutrient levels (Table 1). The bacterial strains used were isolated from maize-cultivated soil and identified via 16s rDNA sequencing. Growth parameters (dry weight, chlorophyll content), physiological parameters (photosynthetic rate, transpiration rate, stomatal conductance), and yield were measured at 52, 72, and 88 DAS. Seed quality parameters (size, sphericity, texture, total solids, protein, and crude fiber content) were determined after harvest and drying. Two-way ANOVA and Duncan's test were used for statistical analysis.

The study revealed statistically significant effects of PGPB and application methods on various parameters. *A. chroococcum* treatment increased chlorophyll content by up to 6.1%, photosynthetic rate by up to 18.4%, and transpiration rate by up to 34.3%. Soil application yielded significantly higher chlorophyll content and dry weight compared to foliar application at all measurement points. Soil application of *B. megatherium* and the *A. chroococcum*/ *B. subtilis* mix resulted in the highest yields (244.67 g and 243.67 g per plant, respectively), exceeding the control by 5.5-13.4%. *B. subtilis* treatment significantly increased total solids content (92%) and crude fiber content (46%) in harvested seeds compared to the control. Soil application generally yielded better results than foliar application. A significant interaction between PGPB type and application method was observed in photosynthetic rate (at 72 DAS), yield, and total solids.

The findings support the hypothesis that PGPB application can positively impact maize growth, physiology, yield, and seed quality under Mediterranean conditions. Soil application consistently outperformed foliar application, likely due to better bacterial establishment and longevity in the soil. The variation in the effectiveness of different PGPB strains highlights the importance of selecting appropriate strains based on specific environmental conditions and the crop. The significant increase in yield and improved seed quality parameters (total solids and crude fiber) demonstrate the potential of PGPB as a sustainable alternative to chemical fertilizers. This study's results align with previous research showing PGPB's beneficial effects on various crops and contribute to the broader understanding of PGPB application optimization for enhanced agricultural sustainability. The significant interaction between PGPB type and application method underscores the need for more research to elucidate the complex interactions between different PGPB strains and application methods.

This study demonstrates that both the type of PGPB and the application method significantly impact maize growth, physiology, yield, and seed quality. Soil application of PGPB, particularly *B. megatherium* and the *A. chroococcum*/ *B. subtilis* mix, resulted in the highest yield. *B. subtilis* significantly improved seed quality. Future research should focus on optimizing PGPB application strategies, investigating underlying mechanisms, and expanding studies to other cereal crops and diverse environmental conditions. Further research could explore the long-term effects of PGPB application, and the development of commercially viable PGPB-based biostimulants.

The study was conducted at a single location under specific Mediterranean conditions, limiting the generalizability of the findings to other geographical regions or soil types. The experiment spanned only one growing season, preventing assessment of long-term effects. Further research should investigate the optimal timing and frequency of PGPB application, and examine a wider range of PGPB strains and their combinations. The precise mechanisms by which the different PGPB treatments influence maize growth and quality characteristics requires further detailed investigation.