Characterization of the fungicidal activity of Calothrix elenkinii using chemical methods and microscopy.

An investigation was directed towards biochemical characterization of cyanobacterium Calothrix elenkinii and analysis of the chemical nature and mode of action of its fungicidal metabolite(s) against oomycete Pythium debaryanum. Biochemical characterization of the culture in terms of carbohydrate utilization revealed the facultative nature of C. elenkinii. Unique antibiotic markers were also found for this strain. 16S rDNA sequencing of the strain revealed 98% similarity with Calothrix sp. PCC7101. The fungicidal activity was tested by disc diffusion assay of different fractions of the culture filtrate. A minimum inhibitory concentration of 10 microl was recorded for ethyl acetate fraction of the 7-weeks old culture filtrates. HPLC, followed by NMR spectral analysis demonstrated the presence of a substituted benzoic acid in the ethyl acetate fraction. Microscopic examination revealed distinct granulation, followed by disintegration of the hyphae of Pythium sp., indicating the presence of an active metabolite in the culture filtrates of Calothrix sp. The fungicidal activity of C. elenkinii can be attributed to the presence of 3-acetyl-2-hydroxy-6-methoxy-4-methyl benzoic acid. This is the first report of a benzoic acid derivative having fungicidal activity in cyanobacteria.

Development of cyanobacterium-based biofilms and their in vitro evaluation for agriculturally useful traits.

The ability of cyanobacteria to be useful as matrices for agriculturally important bacteria was evaluated. Biofilms were generated with the selected strain Anabaena torulosa after co-culturing with Azotobacter chroococcum, Pseudomonas striata, Serratia marcescens, and Mesorhizobium ciceri. The biochemical attributes were compared with individual bacterial and cyanobacterial cultures. The biofilms were characterized in terms of proteins, chlorophyll, IAA production, acetylene-reducing activity, phosphate solubilization, and antagonism towards selected phytopathogenic fungi. An enhancement in the population counts was recorded in A. torulosa-S. marcescens and A. torulosa-P. striata biofilms. The A. torulosa-A. chroococcum and A. torulosa-M. ciceri biofilms were also able to utilize new saccharides as compared to the individual cultures. Such novel biofilms with agriculturally useful traits can provide additional advantages including the broader spectrum of activity and the presence or formation of biologically active compounds; they also suggest the way to effective inoculants for sustainable and environment friendly agriculture.

Developing biochemical and molecular markers for cyanobacterial inoculants.

Markers for evaluating the establishment of cyanobacteria based on their sensitivity or resistance to antibiotics, saccharide utilization patterns and PCR generated fingerprints were developed. Four selected strains (isolates from rhizosphere soils of diverse agro-ecosystems) have shown potential as diazotrophs and exhibited plant growth promoting abilities. Different responses were obtained on screening against 40 antibiotics, which aided in developing selectable antibiotic markers for each strain. Biochemical profiles generated using standardized chromogenic identification system (including saccharide utilization tests) revealed that 53 % of the saccharides tested were not utilized by any strain, while some strains exhibited unique ability for utilization of saccharides such as melibiose, cellobiose, maltose and glucosamine. PCR based amplification profiles developed using a number of primers based on repeat sequences revealed the utility of 3 primers in providing unique fingerprints for the strains.