Putative Nitrogen-Fixing Bacteria Associated With the Rhizosphere and Root Endosphere of Wheat Plants Grown in an Andisol From Southern Chile

Abstract

Acidic ash derived volcanic soils (Andisols) support 50% of cereal production in Chile. Nitrogen (N) is essential for cereal crops and commonly added as urea with consequent environmental concerns due to leaching. Despite the relevance of N to plant growth, few studies have focused on understanding the application, management and ecological role of N2-fixing bacterial populations as tool for improve the N nutrition of cereal crops in Chile. It is known that N2-fixing bacteria commonly inhabits diverse plant compartments (e.g., rhizosphere and root endosphere) where they can supply N for plant growth. Here, we used culture-independent and dependent approaches to characterize and compare the putative N2-fixing bacteria associated with the rhizosphere and root endosphere of wheat plants grown in an Andisol from southern Chile. Our results showed significantly greater bacterial loads in the rhizosphere than the root endosphere. Quantitative PCR results indicated that the copy number of the 16S rRNA gene ranged from 1012~1013 and 107~108 g−1 sample in rhizosphere and root endosphere, respectively. The nifH gene copy number ranged from 105~106 and 105 g−1 sample in rhizosphere and root endosphere, respectively. The total culturable bacteria number ranged from 109~1010 and 107~108 CFU g−1 sample in rhizosphere and 104~105 and 104 CFU g−1 sample in root endosphere using LB and NM-1 media, respectively. Indirect counts of putative N2-fixing bacteria were 103 and 102~103 CFU g−1 sample in rhizosphere and root endosphere using NFb medium, respectively. Sequencing of 16S rRNA genes from randomly selected putative N2-fixing bacteria revealed the presence of members of Proteobacteria (Bosea and Roseomonas), Actinobacteria (Georgenia, Mycobacterium, Microbacterium, Leifsonia, and Arthrobacter), Bacteroidetes (Chitinophaga) and Firmicutes (Bacillus and Psychrobacillus) taxa. Differences in 16S rRNA and putative nifH-containing bacterial communities between rhizosphere and root endosphere were shown by denaturing gradient gel electrophoresis (DGGE). This study shows a compartmentalization between rhizosphere and root endosphere for both the abundance and diversity of total (16S rRNA) and putative N2-fixing bacterial communities on wheat plants grown in Chilean Andisols. This information can be relevant for the design and application of agronomic strategies to enhance sustainable N-utilization in cereal crops in Chile.

In Situ Cultivation Approach to Increase the Culturable Bacterial Diversity in the Rhizobiome of Plants

“…In this study, the bacterial diversity obtained was higher than those reported in wheat rhizobiome studies based on conventional cultivation, which typically report isolates from 14 to 20 bacterial genera (da Moreira et al 2016;Rascovan et al 2016;Verma et al 2015). A recent study focused on screening of putative nitrogen-fixing bacteria associated to wheat plants grown in an Andisols has reported isolation of nine strains belonged to different genera; therefore, the use of MWCs to in situ cultivation methods seems to be an effective alternative to achieve broad of bacterial diversity from wheat microbiome (Rilling et al 2018). Phylogenetically, the three most abundant genera identified were Pseudomonas, Bacillus, and Stenotrophomonas.…”

Bacterial Community Selection of Russula griseocarnosa Mycosphere Soil

“…The members of the genus Burkholderia occur simultaneously with fungal taxa (Stopnisek et al, 2015), and the co-occurring might be due to Burkholderia's ability to migrate with the growing hyphae (Nazir et al, 2012). Mycobacterium has nitrogen fixation functions (Rilling et al, 2018) and can provide nitrogen for the growth of R. griseocarnosa. Sorangium has rich xylan-degrading enzymes that can degrade biological macromolecules, cellulose, hemicellulose, and xylan (Tamaru et al, 2010), which is beneficial for increased mushroom productivity (Zhou et al, 2017).…”

Improved Isolation of Uncultured Anaerobic Bacteria using Medium Prepared with Separate Sterilization of Agar and Phosphate

“…The PS protocol has been shown to improve the culturability of a wide range of microorganisms, including slow-growing heterotrophs (Kato et al, 2018), hard-to-culture Actinobacteria (Adam et al, 2018), and alkane-degrading bacteria (Zheng et al, 2018). Furthermore, many research groups reported the isolation of novel bacterial taxa using the PS protocol (Nishioka et al, 2016;Huang et al, 2017;Kitzinger et al, 2018;Rilling et al, 2018). However, these studies only targeted aerobic microorganisms.…”