Aims: The present study aimed at gaining an insight into the abundance and genetic diversity of culturable N-fixing epiphyte bacteria on the phyllosphere of maize in arid and semi-arid regions of Iran. Methods and Results: Leaf samples of the maize variety, 'single cross 704' (Zea mays L.) were collected from different locations in Iran. The community of culturable N-fixing epiphyte bacteria present was examined by 16S rRNA sequencing, BOXAIR-polymerase chain reaction (PCR) and restricted fragment length polymorphisms analysis of 16S rRNA gene (16S-RFLP). Approximately, 31Á82% of the 242 isolates were identified as N-fixers by cultivation of bacteria in Rennie medium and detection of their nifH gene. The N-fixers were affiliated with four bacterial phyla: Firmicutes, Proteobacteria, Actinobacteria and Bacteroidetes. 16S rRNA sequencing detected 16 genera and 24 different species in the identified phyla. The most dominant genus was Bacillus and the species identified were B. pumilus, B. amyloliquefaciens, B. subtilis, B. paralicheniformis, B. licheniformis, B. niabensis and B. megaterium. In total, 22 RFLP groups were present among the isolates originally identified as N-fixing bacteria. BOXAIR-PCR showed that there was a low similarity level among the N-fixing bacteria isolates, and genetic differentiation of individual strains was relatively great. Conclusions: Our findings suggest that nitrogen-fixing epiphyte bacteria on the phyllosphere of maize may provide significant nitrogen input into arid and semi-arid ecosystem. Significance and Impact of the Study: This research implies that phyllosphere epiphyte diazotrophs have much to offer in sustainable agriculture and can be an alternative to chemical N-fertilizers for providing nitrogen to crops arid and semi-arid regions.

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In order to develop nanotechnology application in the agricultural systems achieving more sustainability in the environment, we have used different nano-carriers for phosphate solubilizing bacteria. The viability and efficacy of two bacterial species; Pseudomonas putida (PP20) and Pseudomonas kilonensis (PK11) in solubilizing phosphate sources (i.e., tricalcium phosphate and hydroxyapatite) with different nano-carriers including nanoclay, natural char micro-particles (NCMPs), nanoclay + alginate, NCMPs + alginate, and natural char nano-particles (NCNPs)+alginate were investigated. Clay, talc powder, and natural char (NC) were included for comparison. The synthesized NCNPs and NCMPs were characterized using FTIR, SEM and Boehm titration analyses. The results confirmed that the chemical oxidation of pristine char made many oxygenated functional groups on the surface of tiny and spherical NCNPs (14.8 nm) which caused their effective incorporation in the matrix of alginate beads. Results of phosphate solubilizing study showed that P. kilonensis was the superior species for viability and stability of its performance on solubilizing phosphorus. The six months evaluation showed that NCNPs + alginate and nanoclay + alginate carriers at both temperatures (4 °C and 28 °C), were the proficient carriers for preserving both bacteria. The results of solubilizing phosphorus sources revealed that both bacteria solubilized tricalcium phosphate more than hydroxyapatite and PK11 showed more privilege in this regard. In addition, the solubilizing index determined after storage for 6 months at 4 °C was higher for all the carriers. Analysis of variance for phosphatase activity revealed that embedding both bacteria in nanoclay + alginate carrier guaranteed the highest phosphatase activity, even though differences between this carrier and NCNPs + alginate and NCMPs + alginate were not significant for the PK11.