Research Article - (2024) Volume 7, Issue 3
Received: 08-Aug-2024, Manuscript No. pollution-24-144864;
Editor assigned: 10-Aug-2024, Pre QC No. P-144864;
Reviewed: 24-Aug-2024, QC No. Q-144864;
Revised: 30-Aug-2024, Manuscript No. R-144864;
Published:
06-Sep-2024
, DOI: 10.37421/2684-4958.2024.7.341
Citation: Bandi, Lasya, Chandra Deepthi, Shubhangi Kanjilal and Kasala Akshitha Reddy, et al. “Cyanobacteria-Fungi: Friends to Waste Water Treatment.” Pollution 7 (2024): 341.
Copyright: © 2024 Bandi L, et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
In nature, many relationships are found amongst various species, which inhabit and find a way of living together in this ecosystem. The aquatic cyanobacterium species have been known to have certain mutualistic interactions with the non-aquatic fungal species, but not all such interactions have been ventured in depth. The cyanobacterium species Scytonema was extracted from a polluted lake while the fungal species Aspergillus flavus was isolated from rotten apples and pears. These have originated in different environments and were exposed to different media. They were co-cultured for 45 days, as well as cultured separately in BG11 medium, cyanobacteria spent medium, fungal spent medium and domestic primary effluent collected from a local open drainage. The observation portrayed their mutualistic relationship to one another and their efficiency in treating wastewater. Such a relationship has led to a decrease in the percentage of biomass and thus in-turn, helping us analyzes its effect in treating wastewater systems.
Scytonema-Aspergillus relationship • Cyanobacterium • Filamentous fungi • Wastewater treatment
Microorganisms have significantly influenced our environment through their life cycles, which impact the dynamics of surrounding organisms. The interactions among these microbes have resulted in an array of advantages, such as the synthesis of vital enzymes and the breakdown of non-biodegradable waste materials. Cyanobacteria (Scytonema, Nostoc, Anabaena) and fungus (Aspergillus species) are found in different habitats, demonstrating the ecological diversity of microorganisms.
The latest studies [1] emphasize the intricate nature of interactions between algae and fungi, showcasing their promise in the advancement of eco-friendly strategies for treating wastewater. These interactions play a vital role in the biological treatment of wastewater, as mutualistic connections and biomass generation make important contributions. Furthermore, research [2] highlights the diverse capacities of microorganisms, namely their ability to break down chitin and to prevent oxidation. These abilities are particularly important in the interactions between bacteria and fungi.
Although the importance of algal-fungal interactions is widely acknowledged, a thorough examination of several sub-species is still necessary. An unknown interac- tion exists between Scytonema and Aspergillus [3-5]. Although there is separate research available on these species, the main objective here is to comprehend their ecological interactions and their contribution to biomass production. The existence of cytotoxins [6-10], anti-fungal enzymes [11,12], etc. introduces additional levels of complexity and uncertainty to this interrelationship, emphasizing the necessity for further investigation[13].
To enhance our understanding of environmentally friendly wastewater treatment utilizing living organisms, it is crucial to conduct a thorough investigation into the relationship between Scytonema and Aspergillus [14].
The objective is to examine the dynamics of their interactions by subjecting them to precisely controlled conditions, including parameters such as pH, temperature, and light exposure, whether these microbes demonstrate mutualistic interactions or engage in competitive connections [15,16].
Gaining a thorough comprehension of this complex subject not only enhances our understanding of interactions among microorganisms but also offers vital insights into their influence on the growth of living organisms[17,18]. These insights have the potential to completely transform the process of treating wastewater, leading to the development of more advanced and environmentally sustainable solutions to protect the environment [19].
Methods
The cyanobacterium chosen for this investigation was acquired from stagnant wastewater of HMT Lake, HMT nagar, Nacharam, Hyderabad, Telangana, India(Figure 1). All the media was purchased from Himedia, India. The collection consisted of many species, including Nostoc, Anabaena, and colonies of Scytonema found among the isolated algae. Scytonema was sub-cultured utilizing Bristol agar medium to obtain pure colonies. Meanwhile, fungal species were extracted from decomposed sections of apples and pears, spread onto PDA plates, and placed in an incubator for 2-3 days. Different fungal species were detected. Amongst them, Aspergillus flavus was selected and sub-cultured on PDA plates to get pure cultures. The A. flavus and Scytonema were cultured separately in BG11 medium using particular conditions: static growth, with a 10:14 hr light/dark cycle, and an average temperature of 25.6 ℃. Additionally, the experiment involved using filter-sterilized air that was enriched with 1% (v/v) CO2 to supply inorganic carbon. Under these conditions, all cultures were consistently preserved, with glasswares and media being sterilized using autoclave.
Figure 1. Mixed culture of cyanobacteria obtained from HMT Lake, HMT nagar, Nacharam, Hyderabad, Telangana, India 500076.
Cyanobacteria isolation
Cyanobacterial cultures were obtained from a stagnant lake containing mixed cultures of microorganisms. The algal flocks were mostly separated and cultivated on Bristol medium, which consisted of 25 g/L NaNO3, 2.5 g/L CaCl2·2H2O, 7.5 g/L MgSO4·7H2O, 7.5 g/L K2HPO4, 17.5 g/L KH2PO4, and 2.5 g/L NaCl solution. Deionized water was introduced to obtain a volume of 1 liter. This medium was used to carry out the initial serial dilution process to decrease the concentration of algal colonies. The cyanobacterial culture concentration thus achieved through dilution [20-23] was sub-cultured to obtain pure strains of Scytonema. This was subsequently utilized for further experimentation in the study. [6,13] After culturing cyanobacteria for 19 days in the medium, the cyanobacterial colonies were filtered out and the remaining medium was used as a spent medium.
Aspergillus flavus isolation
To separate Aspergillus strains, decomposed sections of apples and pears were spread onto PDA plates and placed in an incubator at a temperature of 25.6 ℃, while keeping them in darkness. The observed colonies consisted of black mycelium, white-green mycelium, and white mycelium (Figure 2). The white cottony growth colonies were transferred on new PDA plates and kept in an incubator for 19 days (Figures 3 and 4). On the fourth day, we confirmed the successful sub-culturing of pure cultures. On the 19th day, fungal spores were isolated by rinsing with deionized water, and passing through a sterile glass wool filter. The spore cultures were subsequently placed in deionized water and incubated under comparable circumstances. [24] After culturing fungal colonies for 19 days in the medium, the colonies were filtered out and the remaining medium was used as a spent medium.
Figure 2. Culturing fungal species isolated from rotten apple and rotten pear. a) and d) have successfully been observed to have pure colonies, while b) and c) have shown to have mixed cultures. a) and d) have been used for further sub-culturing.
Figure 3. Subculturing pure strains of Aspergillus culture on PDA medium. The petri dish b) with green colonies, i.e, Aspergillus flavus, has been used for the experiment.
Figure 4. Cyanobacteria and fungi co-culture and individual cultures grown in a) BG11 medium, b) Fungal spent medium, c) Cyanobacteria spent medium and d) domestic waste water.
Co-culturing Cyanobacteria and fungi
The Scytonema strain was sub-cultured in BG11 medium to produce inoculum. The culturing was done in static growth, with an illumination intensity of 100 µmol/m2/s and a light/dark cycle of 10:14 hours. The temperature was maintained at 28 ± 1°C. Inorganic carbon was added through the addition of filter-sterilized air that was enhanced with 1% (v/v) CO2. The BG11 medium composition consisted of 1.5 g/L NaNO3, 40 mg/L K2HPO4, 75 mg/L MgSO4 7H2O, 36 mg/L CaCl2·2H2O, 6 mg/L citric acid, 6 mg/L ferric ammonium citrate, 1 mg/L EDTA-Na2, 20 mg/L Na2CO3, and 1 mg/L A5. Where A5, in turn, was a trace metal solution containing 2.86 g/L H3BO3, 1.86 g/L MnCl24H2O, 0.22 g/L ZnSO47H2O, 0.39 g/L Na2MO42H2O, 0.08 g/L CuSO4 5H2O, and 0.05 g/L Co (NO3)26H2O [25,26].
Co-culture of Cyanobacteria (Scytonema) and fungus (Aspergillus flavus)
The interaction among cyanobacteria (Scytonema) and fungus (A. flavus) were cultured in BG-11 medium. [20-27] When these species were cultured separately in BG-11, no significant growth was detected for Aspergillus, while Scytonema had significant growth. When Scytonema was cultured with the A. flavus, the green clumps became prominent during incubation (Figure 5). Hyphae of A. flavus with Scytonema cells beneath them were observed under microscope [20]. This observation indicates that A. flavus grows in the presence of Scytonema but cannot propagate alone in the BG-11 medium. Further, their coexistence points to a potential mutualistic relationship contributing to forming a synthetic lichen (Figure 6) [28,29].
Figure 5. Observed growth in BG-11 medium for a) Cyanobacteria-fungi co-culture, b) Fungi individual culture and c) Cyanobacteria individual culture.
Figure 6. Visible associations of clumps of a Aspergillus spores with Scytonema.
Fungal growth in spent medium of Scytonema
The nutrition source for fungal growth was assessed in the Scytonema spent medium. There were two conditions implied: BG-11 medium and Scytonema spent medium (Figure 7). In the Scytonema spent medium, a noticeable growth of fungi was observed while there was no growth in the BG-11 medium. These results imply that Scytonema spent medium impart vital nutrients sufficient for fungal growth [30,31].
Figure 7. Observations made on cyanobacteria BG-11 spent medium for a) Cyanobacteria individual culture, b) Fungi individual culture and c) Cyanobacteria-fungi co-culture.
Scytonema growth in fungal spent medium
Scytonema growth was evaluated in the Fungal spent medium for the nutritional requirements. The two conditions were implied: BG-11 medium and Fungal spent medium (Figure 8) [32]. Scytonema demonstrated comparatively more growth in the Fungal spent medium than in the BG-11 medium. This results in the presence of adequate nutrients in the Fungal spent medium [33,34].
Figure 8. Observations made on fungal spent medium (PDA) for a) Cyanobacteria-fungi co-culture, b) Fungi individual culture and c) Cyanobacteria individual culture.
Co-culture in domestic primary effluent for wastewater treatment
Cyanobacteria-fungus co-culture for wastewater treatment was applied by culturing the microorganisms in domestic primary effluent [1,25]. A decrease in the Optical Density (O.D) values at 620 nm was initially observed and was stable later. This occurrence indicates possible wastewater treatment with microorganisms utilizing the organic content present in the primary effluent for their growth. The stabilization of O.D values infers an equilibrium point in the treatment process (Figures 9-11) [35].
Figure 9. Observations made on domestic waste water collected from an open drainage in Mallapur, Medchal, Hyderabad, Telangana, India. 500076; a) Cyanobacteria-Fungi coculture, b) Fungi individual culture and c) Cyanobacteria individual culture.
Figure 10. Graphical representation showcasing the increase in O.D values for cyanobacteria and fungi co-culture in BG-11 medium (blue) and decrease in the O.D value of domestic waste water that had the co-culture of cyanobacteria and fungi.
Figure 11. Percentage of transmittance in the case of BG11 medium and domestic wastewater as calculated by colorimeter.
The co-culturing of Cyanobacteria and Fungi disclosed a symbiotic relationship among each other. Their collaborative presence in different mediums showed green clump formation suggesting a mutualistic association with a potential synthetic lichen formation. The Cyanobacteria-Fungi cultures grown in BG-11 media had an accelerated rate of Optical Density (O.D) values which was evident with the overall increase in the size of their colonies and biomass. In the case of primary effluent collected from a local drain, a decrease in optical density values was observed over a period of time in the co-culture. This was due to the fact that the nutrients present in the water were utilized by the cyanobacteria-fungi co-culture and thus, their mutualistic relationship proved to be efficient in treating domestic primary effluent. Relative interactions between Cyanobacteria and Fungi coincidences with previous studies [24]. The spore density and light conditions have an effect on nitrogen fixation and algal growth. This allineates with our study, drawing attention to the significance of factors influencing such interactions for possible benefits in a wide range of applications. The capability of Cyanobacteria to transform carbon dioxide into useful chemicals and fuels would be a latent environmental benefit. The Cyanobacteria-Fungi relationship ensures for sustainable practices in agriculture, industry and wastewater treatment.
In conclusion, these results provide insights into the mutualistic relationship between Cyanobacteria (Scytonema) and Fungi (Aspergillus flavus) along with probable applications in nutrient recycling and wastewater treatment. The divergent assemblage of secondary metabolites induced by cyanobacteria includes complexity to the interactions and unfolds new approaches for future research. Comprehending the specific compounds involved and their mechanisms can pave the way for sustainable applications in agriculture, medicine, industry and environmental management.
We sincerely thank Rev. Sr. Alphonsa.p, correspondent, Rev. Sr. Velangini Kumari, principal, St. Pious X Degree & PG College for Women for giving her kind consent and providing all the facilities to carry out our project work in this esteemed institution.
We owe our gratitude to Poosa Padmalatha, lecturer, Head of Department of Biotechnology. We are extremely thankful and indebted to her for sharing expertise, conscientious guidance and encouragement to accomplish this project.
We take this opportunity to thank all the people who aided directly or indirectly in the completion of this project.
Ethical approval
Not applicable.
Data availability
The data is publicly available at https://doi.org/10.6084/m9.figshare.25329913.v1
The funding for this project has been aided by St. Pious x Degree and PG College for Women.
Bandi Lasya, Shubhangi Kanjilal carried out literature reviews. Kasala Akshitha Reddy, Chandra Deepthi drafted the manuscript and performed the statistical analysis. Poosa Padmalatha offered guidance and planned the procedure. All contributing authors performed the experiment equally. All authors read and finalized the final manuscript.
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