Isolation, improvement and characterization of an ammonium excreting mutant strain of the heterocytous cyanobacterium, Anabaena variabilis PCC 7937

• Stable mutant strains were obtained after double random mutagenesis treatments.
• Ammonium excreting activities after partial inhibition of glutamine synthetase.
• Evolutionnary engineering was used to isolate a more efficient strain.
• Ammonium excretion induced partial inhibition of the culture growth.
• The main phenotypic trait linked to ammonium release in this multicellular cyanobacteria is a decrease of the filaments lengths.

Besides potential applications in the agriculture field as natural nitrogen fertilizer, N2-fixing cyanobacteria have recently gained some attentions for new applications linked to the potential production of biologically active molecules or biohydrogen. Ammonium bioproduction is also gaining attention with the potential use of microalgae in biofuels production and the concerns about the increasing needs for nitrogen substrates. This study has investigated some phenotypic traits linked to biomass production and ammonium release in multicellular cyanobacteria, Anabaena variabilis PCC 7937. It confirms that this wild-type strain has no natural ability for ammonium excretion under diazotrophic conditions. A mutant strain, A. variabilis PCC 7937-C9, was obtained after double random mutagenesis treatments with ethyl methane–sulfonate and screening in batch cultures for resistance to the effect of a glutamine synthetase inhibitor, l-methionine-d,l-sulfoximine (MSX). Although significantly characterized by shorter cell filaments, the growth parameters in photobioreactors of the mutant strain cultures were in the same range of values than those of the wild type. In the presence of MSX this strain was shown to produce extracellular ammonium, with specific rates up to 4.9 μmol NH4+ mg Chl a−1 h−1. The efficiency of this strain, estimated by its specific rate of ammonium excretion, was shown to be improved after consecutive batch cultures with increasing concentrations of MSX. Such mutant strains are of potential use for investigating ways to improve extracellular ammonium bioproduction.

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