Cyanobacteria in natural environments exist in close interaction, forming associations with other microorganisms. These interactions are based on principles of mutualism, where each member of the consortium complements the functions of the others, thereby enhancing the efficiency of nutrient exchange and metabolic processes. Such cooperation provides benefits for all participants, increasing their growth, productivity, and survival.
Under laboratory conditions, algal-bacterial consortia can be artificially constructed to improve biotechnological processes, for example, to accelerate microalgal biomass production or to obtain value-added products for biotechnological applications. Investigating these interactions opens new opportunities for the development of environmentally friendly and efficient technologies. In particular, algal–bacterial consortia are widely applied in the production of biofertilizers and bioactive compounds, as well as in wastewater treatment and the bioremediation of contaminated environments.
For this study, the cyanobacterial species Nostoc commune was selected as a model organism due to its ability to perform photosynthesis in vegetative cells and to fix atmospheric nitrogen in specialized cells known as heterocysts. It forms filamentous structures surrounded by a mucilaginous sheath, which facilitates attachment and interactions with other microorganisms. Owing to its physiological characteristics, Nostoc commune represents a convenient and promising model for studying interactions within algal-bacterial consortia.
The effective application of algal-bacterial associations is not feasible without a comprehensive characterization of each constituent member. Of critical importance is not only the taxonomic composition of the association, but also a detailed examination of the physiological and biochemical properties, as well as the productivity parameters of the interacting organisms. Therefore, the objective of this study was to analyze the bacterial component of the algal-bacterial association formed by Nostoc commune.
Algal-bacterial consortia represent stable coexisting systems composed of cyanobacteria and diverse heterotrophic bacteria, in which mutual benefits are established. These interactions are most commonly driven by the exchange of essential metabolites. Microalgae and cyanobacteria receive nitrogen, vitamins, iron, amino acids, and other growth-promoting compounds from associated bacteria. In return, bacterial partners utilize photosynthetically fixed carbon released by cyanobacteria in the form of dissolved organic compounds.
The material used in this study was a culture of Nostoc commune maintained in the collection of the Department of Biochemistry and Biotechnology at Yuriy Fedkovych Chernivtsi National University.
To identify the associated microflora, the following culture media were employed: meat-peptone agar, Sabouraud agar, thioglycollate medium, Lactobacillus agar, and Bifidobacterium-selective medium. These media were selected based on their selective properties for microorganisms with different physiological and ecological requirements. After five days of cultivation, the growth of associated microorganisms was observed on four out of the five media tested.
The highest number of colonies (4.3 × 10² CFU/mL) was recorded on thioglycollate medium, indicating a close association between the cyanobacteria and anaerobic or facultatively anaerobic microorganisms. A lower, yet still considerable, number of colonies (3.1 × 10² CFU/mL) was observed on Sabouraud agar. These results suggest the probable presence of yeasts or micromycetes within the associated microflora of the Nostoc commune culture.
On meat-peptone agar (MPA), a colony count of 1.9 × 10² CFU/mL was recorded, representing a moderate level of growth compared with the other culture media used. This result indicates the probable presence of both Gram-positive and Gram-negative bacteria capable of growth under aerobic conditions.
On Bifidobacterium-selective medium, a moderate growth level of 1.05 × 10² CFU/mL was observed. In contrast, no growth was detected on Lactobacillus agar, which is designed for the cultivation of lactic acid bacteria (e.g., members of the genera Lactobacillus and Lactococcus). This finding suggests the absence of such microorganisms in the associated microflora of Nostoc commune.
Subsequently, colony morphology was analyzed based on key characteristics, including color, shape, edge structure, size, elevation, consistency, and transparency. In addition, fixed microscopic preparations stained using the Gram staining method were prepared for further analysis.
During cultivation on meat-peptone agar, three morphologically distinct colony types were detected, indicating the presence of at least three different microorganisms associated with the Nostoc commune culture. An analysis of macroscopic colony characteristics combined with microscopic examination suggests that the first colony type may correspond to the bacterial species Microbacterium laevaniformans.
Based on the observed phenotypic traits, the second colony morphotype is most likely represented by microorganisms belonging to the genus Pseudomonas. The third colony type exhibited characteristics most consistent with Bacillus licheniformis.
Cultivation on Sabouraud agar also revealed three morphologically distinct colony types. Two of these displayed staining properties and microscopic features identical to those observed on meat-peptone agar. Consequently, these isolates were considered to represent the same microorganisms, namely Bacillus licheniformis and members of the genus Pseudomonas. This finding provides additional evidence for their stable presence within the associated microflora of the Nostoc commune culture.
In addition, white, matte, circular colonies were observed on Sabouraud agar. Microscopic analysis revealed the presence of hyphae or hyphae-like structures, which are characteristic of fungal organisms. Taking these features into account, the isolate was tentatively classified as a yeast-like fungus, with Saprochaete suaveolens considered the most probable representative.
Two morphologically distinct colony types were identified on thioglycollate medium. One of them exhibited staining characteristics and microscopic morphology identical to those of isolates detected on both meat-peptone agar and Sabouraud agar and was therefore attributed to Bacillus licheniformis. The second colony type observed on this medium is likely represented by microorganisms belonging to the genus Acinetobacter.
Cultivation on Bifidobacterium-selective medium revealed two morphologically distinct colony types, which, according to our assumptions, belong to the group of actinomycetes. The first colony type is presumably represented by microorganisms of the genus Streptomyces, whereas the second is likely affiliated with the genus Actinomyces.
Overall, the investigation of the bacterial component of the algal–bacterial culture Nostoc commune demonstrated a high level of diversity within its associated microflora. Representatives of the genus Bacillus were identified, including the probable species Bacillus cereus, Bacillus subtilis, and Bacillus licheniformis. The possible presence of bacteria belonging to the genera Pseudomonas spp. or Sphingomonas spp., which are involved in the degradation of organic polymers, was also noted, as well as Acinetobacter spp., capable of utilizing cellular metabolites and thereby reducing environmental toxicity. Furthermore, Microbacterium laevaniformans, a Gram-positive bacterium known for its ability to synthesize levan - a polymer involved in biofilm formation - was detected.
The associated microflora of Nostoc commune is taxonomically diverse, comprising microorganisms that produce antibiotics and enzymes, species that degrade cellular metabolites, as well as symbionts involved in biofilm formation and maintenance. Among these, filamentous bacteria such as Acinetobacter spp. and Streptomyces spp. are likely present and are capable of forming mycelium-like structures that contribute to biofilm stabilization.
Particular attention should also be given to the yeast-like fungus Saprochaete suaveolens, whose occurrence is likely related to elevated levels of organic compounds in the medium. This fungus presumably utilizes the exopolysaccharides of Nostoc commune as a nutrient source, metabolizing its products without exerting any negative effects on the cyanobacterium.
The results of this study demonstrate that the associated microflora of Nostoc commune is highly diverse, encompassing bacteria with distinct physiological and ecological traits, as well as yeast-like fungi. The culture harbors Gram-positive and Gram-negative bacteria, including representatives of Bacillus, Microbacterium, Pseudomonas, Acinetobacter, Streptomyces, and Actinomyces, which participate in nutrient cycling, degradation of cellular metabolites, and biofilm formation. Additionally, the presence of the yeast-like fungus Saprochaete suaveolens suggests interactions with cyanobacterial exopolysaccharides, reflecting a complex and mutually supportive algal–bacterial association. Overall, these findings highlight the functional diversity of the Nostoc commune microflora and its potential ecological and biotechnological significance.
References
1. León-Miranda E., Tejada-Jiménez M. Microalgal and nitrogen-fixing bacterial consortia: From interaction to biotechnological potential. Plants. 2023. Vol. 12(13), P. 2476.
2. Cooper M. B., Smith A. G. Exploring mutualistic interactions between microalgae and bacteria in the omics age. Current Opinion in Plant Biology. 2015. Vol. 26. P. 147–153.
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