Biofilm formation enables free-living nitrogen-fixing rhizobacteria to fix nitrogen under aerobic conditions

Abstract

The multicellular communities of microorganisms known as biofilms are of high significance in agricultural setting, yet it is largely unknown about the biofilm formed by nitrogen-fixing bacteria. Here we report the biofilm formation by Pseudomonas stutzeri A1501, a free-living rhizospheric bacterium, capable of fixing nitrogen under microaerobic and nitrogen-limiting conditions. P. stutzeri A1501 tended to form biofilm in minimal media, especially under nitrogen depletion condition. Under such growth condition, the biofilms formed at the air-liquid interface (termed as pellicles) and the colony biofilms on agar plates exhibited nitrogenase activity in air. The two kinds of biofilms both contained large ovoid shape 'cells' that were multiple living bacteria embedded in a sac of extracellular polymeric substances (EPSs). We proposed to name such large 'cells' as A1501 cyst. Our results suggest that the EPS, especially exopolysaccharides enabled the encased bacteria to fix nitrogen while grown under aerobic condition. The formation of A1501 cysts was reversible in response to the changes of carbon or nitrogen source status. A1501 cyst formation depended on nitrogen-limiting signaling and the presence of sufficient carbon sources, yet was independent of an active nitrogenase. The pellicles formed by Azospirillum brasilense, another free-living nitrogen-fixing rhizobacterium, which also exhibited nitrogenase activity and contained the large EPS-encapsuled A1501 cyst-like 'cells'. Our data imply that free-living nitrogen-fixing bacteria could convert the easy-used carbon sources to exopolysaccharides in order to enable nitrogen fixation in a natural aerobic environment.

Valorization of Lignin as an Immobilizing Agent for Bioinoculant Production using Azospirillum brasilense as a Model Bacteria

“…In this study, we took advantage of the ability of A. brasilense to form biofilms and its consequent adherence to prepare a bioinoculant with organosolv-recovered lignin. We tracked the biofilm formation on lignin particles in different culture media [19,20]. We observed a good adhesion of bacterial particles to lignin as shown before; this could be explained in many ways, but we can attribute the facility of adherence to the fact that A. brasilense forms more stable biofilms in hydrophobic surfaces [20], which is the case of lignin.…”

“…We observed a good adhesion of bacterial particles to lignin as shown before; this could be explained in many ways, but we can attribute the facility of adherence to the fact that A. brasilense forms more stable biofilms in hydrophobic surfaces [20], which is the case of lignin. Also, it seems that lack of nitrogen negatively affected the biofilm formation after 3 days, after which they started to diminish in relative biomass; different from Pseudomonas stutzeri A1501, which forms more biofilms under nitrogen-depleted conditions [19].…”

“…Biofilm biomass quantification was carried out, as previously described, with slight modifications [19,27]. Following the specifications mentioned above, A. brasilense was grown in 125 mL Erlenmeyer flasks containing 50 mL of one of three different media: LB, OAB with nitrogen source (OAB+), or OAB without nitrogen (OAB−) [28] and 50 mg of lignin.…”

Community assemblage of free-living diazotrophs along the elevational gradient of Mount Gongga

“…Diazotrophs are either free-living or symbiotically associated with plants. Intensive studies have been conducted on symbiotic diazotrophs, while more and more studies have revealed the importance of free-living diazotrophs to N inputs in natural ecosystems (Reed et al, 2011;Reed et al, 2013;Wang et al, 2017a).…”

On the Contribution of Cell Aggregation and Extracellular DNA to Biofilm Formation and Stabilization in Azospirillum brasilense Bacteria

“…Оценка содержания нуклеиновых кислот в матриксе биопленок. Выделение компонентов матрикса и определение содержания в них нуклеиновых кислот проводили согласно рекомендациям, приведенным в работе [18]. Биомассу 6-суточных биопленок, сформированных на поверхности стекла, после удаления планктонных бактерий промывали 50 мМ ФБ (pH 7.0) и смывали пипетированием этим буфером.…”