Pooja Vaswani, Author at Everflow Global

Unveiling the Hidden World of Algal Biofilms: Nature’s Microscopic…

October 31, 2023October 31, 2023
by Pooja Vaswani

Biofilms are clusters of microorganisms that adhere to each other and a solid surface where water is present for extended periods. In nature, people often refer to photosynthetic biofilms as periphytic or algal biofilms, which consist mostly of algae, cyanobacteria, and heterotrophic bacteria living in symbiosis. In addition to these primary members, biofilms often contain additional microorganisms like flagellates and desmids and non-biological elements like silt, sand, and minerals.

Algae, which are chlorophyll-containing organisms, are widespread and lack traditional roots, stems, and leaves (Kesaano M, Sims RC et al., 2014). Biofilms are intricate communities of microorganisms that form on any surface, enclosed within a matrix of extracellular polymeric substances (EPS). These biofilms typically consist of unicellular or simple filamentous organisms, often known as microfouling or slimes. In specific conditions, such as high water flow or exposure to toxins, macroalgae like Enteromorpha and Ectocarpus can exhibit stunted growth on anti-fouling surfaces. In such cases, these algae may adapt by adopting a more compact growth form or integrating into the microfouling slime layer. The term “algal biofilms” refers to communities primarily dominated by microalgae that colonize well-lit surfaces when moisture and nutrients are present (Leadbeater BS et al., 1992, Jarvie HP, Neal C, et al., 2002).

Adherence and Attachment

Two methods can achieve microalgae immobilization: passive immobilization, accomplished through natural or induced biofilm formation, and active immobilization, achieved by trapping the cells within hydrogel polymer matrices. Researchers have utilized various immobilization media, including alginates, carrageenans, and polyacrylamide gel, for various purposes (Muñoz et al., 2009; de-Bashan and Bashan, 2010; He and Xue, 2010; He et al., 2014; Gagliano et al., 2017). Immobilization based on these polymeric matrices provides low mechanical strength and can result in restrictive diffusion of the substrate (Qureshi et al., 2005; Moreno-Garrido, 2008). This approach capitalizes on microalgae’s natural tendency to form biofilms.

Inorganic and organic compounds adhere to the surface of a substrate, creating a conducive environment for microbial growth (Qureshi et al., 2005; Stephens et al., 2015). Once microalgae and bacteria establish themselves on the surface, they secrete extracellular substances comprising nucleic acids, proteins, polysaccharides, and phospholipids. These substances serve a dual purpose: enhancing adherence to the substrate and simultaneously capturing and concentrating essential nutrients required for cell growth (Mohsenpour et al., 2021; Qureshi et al., 2005).

Algal Biofilm Habitats: Nature’s Adaptive Colonizers

In natural ecosystems such as rainforests, deserts, and ocean floors, algal biofilms are commonly present on surfaces like river stones (see Fig. 1), seashores, garden walls, tree bark, reed stems, and bamboo pipes. These microalgal biofilms thrive when they have sufficient moisture and light, and people typically collect them by scraping. They can grow on various materials, including plastics such as polyvinylchloride, polyethylene, polyurethane polymethyl methacrylate, polystyrene, polycarbonate, and polyamide; and other materials such as glass, cardboard, and ceramic tiles.

Notably, algal biofilms exhibit adaptability to environmental changes, maintaining colonies on surfaces and detaching either as single colonies or in clumps.

Composition of algal biofilms

The polysaccharide matrix called EPS embeds microorganisms in biofilms, providing cohesion and enabling component interactions (Flemming & Wingender et al., 2010).

Nutrients and dissolved gases diffuse through the boundary layer above the biofilm, then through the EPS matrix to reach biofilm cells, while waste gases and products either recycle within or diffuse outward. Surfaces concentrate charged particles and molecules, potentially serving as nutritional sources due to mineral and organic content. Various inorganic and organic molecules accumulate on surfaces, and particle-associated bacteria exhibit higher nutrient uptake rates, as shown by (Paerl and Merkel et al.,1982).

Diatoms are common early colonizers in algal biofilms, often making up the majority of associated bacteria and other unicellular organisms (Jackson & Jones, 1988). Single-celled diatoms are known for their silica frustules and their golden brown appearance, which is attributed to the fucoxanthin pigment present in their chloroplasts. They vary in size and attachment mechanisms (Daniel et al., 1987). Just a few diatom cells can start surface attachment, multiplying to form a compact biofilm.

In freshwater environments, green algae and cyanobacteria, particularly in aerial conditions, can dominate algal biofilms (Grant et al., 1982). For instance, Pleurococcus, a green unicellular alga, thrives in damp conditions on various surfaces. Trentepohlia, a filamentous green alga, can disfigure exterior surfaces in humid regions (Wee & Lee et al.,1980). Cyanobacteria, though usually minor, can become dominant due to their resilience and nitrogen-fixing abilities.

The biofilm is a dynamic community in equilibrium with its environment, undergoing growth, death, sloughing, and regeneration processes similar to bacterial biofilms but with less organization (see fig 2).

mature algal biofilms — Fig 2 Mature Algal Biofilm

Factor affecting algal biofilm.

The development and adhesion of algal biofilms are influenced by various factors, including the physico-chemical characteristics of the substrate, operational parameters (see fig 3), the specific microalgal strain employed, and the interactions between microalgal cells, the substrate, and the liquid medium.

factors affecting algal biofilms — Fig 3 Factors Affecting algal biofilm

Light

Light is essential for algal photosynthesis and microalgal growth. Green algae dominate in high-light conditions during early colonization, while heterotrophic bacteria thrive in low-light environments. Diatoms often prevail in light-limited algal biofilms.

Increasing photon flux density (PFD) enhances growth rates in planktonic algae, but excessive PFD can result in photoinhibition and photooxidation, leading to culture stagnation or demise. Microalgae have the capacity to adjust to changing light intensities to optimize efficiency and protect against photodamage. The specific threshold for photoinhibition or light limitation varies among biofilm communities.

Under high irradiance, thick biofilms dominated by green algae resembling Scenedesmus were observed. Biofilms cultivated in a similar light range exhibited higher cellular nutrient content compared to those grown under higher light regimes. Algal Biofilms cultured under low irradiance using the same inoculum were thinner, more compact, and featured a wider variety of species, including cyanobacteria.

Hultberg et al. demonstrated that light quality has a direct impact on biofilm formation when using monochromatic illumination, suggesting the potential for enhanced cellular growth and lipid content through light quality optimization.

Temperature

Temperature plays a pivotal role in microalgal growth, biomass production, and biochemical processes. The optimal temperature range for microalgal cultivation typically spans from 20°C to 25°C. Below 16°C, growth slows down, while exceeding 35°C can be fatal for certain species. High temperatures, depending on the species, can stimulate cell growth, adhesion, and EPS production, promoting the formation of biofilms (Qureshi et al., 2005).

Claquin et al. (Claquin et al.; 2008) conducted research highlighting temperature’s significant impact on marine microalgae, influencing growth and the secretion of transparent exopolymeric particles (TEP) due to shifts in microbial and grazing activities (Honda Y, Matsumoto J, et al., 1983). Temperature also affects algal growth rates, species composition, and grazing activity within biofilm communities(Rao TS et al.; 2010). Tuchman and Blinn (1979) observed a rise in algal densities as temperature increased.

The Arrhenius relationship elucidates how temperature influences algal growth rates under consistent light and optimal nutrient conditions(Goldman JC et al.; 1974, Raven JA et al.; 1988). Biofilms grown in thin water layers are more sensitive to temperature fluctuations compared to suspended cultures (Posadas et al., 2013).

Furthermore, temperature stress and the accumulation of calcium can have adverse effects on EPS production in algal cells (Domozych, et al., 2007). Fica and Sims (2016) determined that elevating wastewater temperature (7–27°C) and increasing organic carbon levels (300–1,200 mg L−1) significantly enhance biomass growth in algae-based biofilm systems. Temperature serves as the precise conductor orchestrating the dynamics of microalgae.

Nutrients

The abundance of algal species within biofilms is strongly influenced by nutrient concentration and light intensity, with a preference for algal dominance under conditions of high inorganic and low organic carbon levels(Unnithan VV et al.; 2014).

Nutrient availability plays a pivotal role in shaping algal growth, biofilm characteristics, succession patterns, and species composition (Nils RP et al.; 2003, Sekar R et al.; 2002). Environments enriched with biodegradable organic matter tend to favor the development of heterotrophic biofilms, whereas phototrophic biofilms thrive in response to light and the presence of inorganic nutrients (Hillebrand H et al.; 2002, Olapade OA et al.; 2006). The transport of nutrients to algal cells occurs through a concentration boundary layer via diffusion mechanisms.

Nitrogen and phosphorus availability exert distinct influences on microalgal metabolism. In instances of nitrogen starvation, microalgae pivot towards the production of lipids and/or carbohydrates as opposed to proteins (Gojkovic et al., 2020). There exists a mutual reliance between nitrogen and phosphorus uptake, with green algae initially dominating biofilms in nitrogen-deficient conditions, followed by diatoms, and ultimately cyanobacteria. The presence of phosphorus enhances nitrogen uptake, and in the presence of nitrogen, excess phosphorus can be absorbed by algae, a phenomenon known as luxury uptake (Bougaran et al., 2010)t.

Monod’s Kinetic Expression in Algal Biofilm Research

Monod’s kinetic expression finds application in algal biofilm research to elucidate growth patterns concerning nutrient concentrations and to predict rates of substrate utilization. For instance, Hill et al. 2009 established a growth saturation threshold of 25 μg/L soluble reactive phosphorus (SRP) for algal growth in stream biofilms.

Microalgae harness dissolve for in-organic carbon sources such as CO2 (aq) and HCO3 − found in wastewater, along with atmospheric CO2, and organic carbon derived from bacterial degradation (E. Posadas et al.; 2013, L.B. Christenson et al.; 2011).

Carbon, nitrogen, phosphorus, and silicon (for diatoms) are pivotal elements governing microalgae growth. The ratios of C:N:P serve as valuable indicators of nutrient limitation within algal communities, with the Redfield ratio (106:16:1 molar basis) representing a characteristic benchmark for optimally growing phytoplankton (R.S. Stelzer et al.; 2001, H. Hillebrand et al; 1999)

Elevated levels of nitrogen and phosphorus, coupled with increased inorganic carbon concentrations and heightened light intensity, augment the accumulation of photosynthetic biomass and its relative proportions in comparison to non-photosynthetic biomass. Additionally, advanced cultures and elevated temperature and nutrient loading rates contribute significantly to an upsurge in the proportions of cyanobacteria within photosynthetic biofilms.

pH

The pH level plays a crucial role in nutrient availability, including factors such as the solubility of ammonium and phosphate ions, as well as the formation of precipitates. Elevated pH values, typically at 9 or above, can trigger the formation of calcium phosphate, rendering it inaccessible to microalgae (Laliberté et al., 1997). Conversely, a decrease in pH levels can influence enzymatic activities, albeit depending on the specific microalgal species, thereby decelerating cell growth and product production.

Furthermore, pH values determine the charge of functional groups present on the surface of microalgae, consequently affecting their capacity to bind substances such as heavy metal ions.

CO2

Carbon stands as an indispensable element vital for the growth and productivity of microorganisms. Photoautotrophic organisms like microalgae possess the capacity to employ light energy to photosynthetically convert carbon dioxide (CO2) into biomass (S.B. Patwardhan et al.; 2022). Additionally, microalgae can utilize soluble carbonates as a carbon source for cellular growth, either via direct uptake or by catalyzing the conversion of carbonate into free carbon dioxide through enzymatic carboanhydrase activity.

Hence, the availability of an appropriate level of CO2 is imperative for the proper growth and metabolic activities of microalgae. Deviations from this optimum CO2 level, whether below or above, can exert adverse impacts on growth and productivity (W. Blanken et al 2014, B. Clement-Larosiere et al.; 2014). For instance, CO2 concentrations falling below the optimal level may result in carbon limitation, thereby retarding growth and productivity. Conversely, upon the introduction of an elevated concentration of CO2, microalgae assimilate a portion of the carbon through photosynthesis. Simultaneously, the excess carbon is converted into carbonic acid (H2CO3), which subsequently induces acidification of the medium.

Substratum

Algal adhesion studies primarily explore surface characteristics and material composition’s influence on biofilm formation to enhance cell attachment and biofilm growth. Ozkan and Berberoglu (2013) found differences in cell attachment between green algae and diatoms related to surface hydrophobicity, a concept emphasizing hydrophobic entities’ preference for each other to minimize water contact (Palmer et al., 2007).

Surface texture is crucial; rough or porous surfaces, with increased surface area and shear force protection, promote higher cell attachment (Gross et al., 2016; Huang et al., 2018). Increasing surface roughness, as suggested by Cao et al. (2009), creates slower flow zones that aid algal settlement. Studies by Huang et al. (2018) and Kardel et al. (2018) demonstrated weaker shear stress on grooved surfaces, influencing cell attachment based on groove shapes.

Material properties matter. Christenson and Sims (2012) found superior algal growth on cellulose-based natural polymer surfaces over synthetic polymers on various substrates. Sekar et al. (2004) observed greater attachment of certain algal species to hydrophobic surfaces (e.g., titanium, perspex, stainless steel), with exceptions for copper and its alloys, which hindered attachment due to toxicity (aluminum and admiralty brass).

Hydrophobicity

Hydrophobicity is a factor that affects the surface of the algal cell and substrate, for this reason considered an interface and a cellular factor. In diverse multi-species microbial biofilm, the presence of fimbriae, proteinaceous bacterial appendages rich in hydrophobic amino acids, can increase cell surface hydrophobicity (Barros et al., 2018). Flagellated cells show an increased ability to attach to surfaces. Flagellar motility may serve to overcome initial electrostatic surface repulsion (Bullitt and Makowski, 1995; Qureshi et al., 2005; Krasowska and Sigler, 2014). However, the microalgae biofilm on a hydrophilic surface might decrease and then increase with the increasing concentration of DOMs(dissolvable organic matter) and inorganic salts. Furthermore, many studies have shown that the DOMs can adsorb onto the substratum surface, leading to the development of surface conditioning films (Hwang et al., 2012)

Flow Velocity

The flow velocity of the liquid medium housing the algal biofilm plays a pivotal role in governing both cellular growth and attachment dynamics. This is primarily attributed to the liquid medium’s essential function as a source of vital nutrients for sustaining microalgal cells. However, it is imperative to acknowledge that elevated flow velocities can impose shear stress upon the biofilm (P. Choudhary et al.; 2017).

Furthermore, it is well-documented that turbulent flows occurring within the liquid medium have the potential to induce the detachment of cells from the biofilm, leading to a subsequent reduction in the overall thickness of the biofilm (L. Katarzyna et al.; 2015). It is noteworthy that the specific arrangement of microalgal biofilms may involve their placement within a rotational environment, resulting in periodic exposures to gaseous and liquid phases. This unique configuration introduces an added layer of complexity to the intricate interplay between flow velocity, shear stress, and the dynamics of microalgal cell growth and attachment within the biofilm milieu.

Extracellular polymeric substances (EPS)

Extracellular Polymeric Substances (EPS) in algal biofilm communities consist of polysaccharides, proteins, nucleic acids, lipids, and humic acids (A.M. Romani et al.; 2008, F. Di Pippo et al .; 2009) influencing their physical and chemical properties. EPS serves as nutrient reservoirs, with embedded enzymes breaking down EPS and inert solids (Sutherland IW et al.; 1999), and they act as ion exchange resins, trapping nutrients through sorption (Flemming HC et al.; 2007, Wolfaardt GM et al; 1999). EPS roles include facilitating cell movement (D.J. Smith et al.; 1998), preventing cell desiccation, protecting against toxins (J.V. Garcia-Meza et al.; 2005), and providing structural stability (I.W. Sutherland et al.; 2001).

Environmental Factors Influencing EPS Production

Microalgae adjust EPS production in response to environmental factors during biofilm formation. Light intensity significantly affects EPS accumulation (H. Ge et al.; 2014), e.g., Nostoc sp. produces more EPS at 206.20 mg/g DW under 80 μE.m^−2.s^−1 compared to 155.49 mg/g DW at 40 μE.m^−2.s^−1 .

Adhesion materials enhance EPS production in diatom Amphora coffaeformis (Becker K et al .; 1996). Growth materials influence EPS, as shown by Shen et al. (H. Ge et al.; 2014), and increased nutrient levels, especially nitrogen, boost EPS in diatom and green algae. Operational factors like light, temperature, nutrients, and culture density affect both biomass and EPS secretion. Li et al. noted EPS concentrations of 1.25 and 1.75 g.L^−1 with C/N ratios of 0.96 and 12.82, respectively (.H. Li, L. Ji et al.; 2020).

Young, grazed algal biofilms exhibit high EPS-to-biomass ratios for survival (C. Barranguet et al.; 2005). Cyanobacteria and diatom biomass correlate positively with EPS in wastewater-based biofilms. Light is linked to EPS production, and even in darkness, some species like Cylindrotheca closterium, Navicula perminuta, and Nitzschia sigma secrete EPS using stored glucan as a carbon source (D.J. Smith et al.; 1998).

Wastewater algal biofilms face grazing and changing conditions, necessitating species selection for EPS production and carbon management.

Species interaction

Natural biofilms encompass a diverse array of microbial constituents, including fungi, algae, protozoa, flagellates, and bacteria ( B.S.C. Leadbeater et al.; 1992, F. Di Pippo et al.; 2009). Within photosynthetic biofilms, one finds a myriad of algae, bacteria, cyanobacteria, protozoa, and multicellular microorganisms. Notably, diatoms, green algae, and filamentous algae play substantial roles in contributing to biofilm biomass, displaying both autotrophic and heterotrophic capacities (Liang Y,Sarkany N et al; 2009). The bacterial component encompasses cyanobacteria and heterotrophic and autotrophic bacterial species.

Within algal biofilm, organisms engage in symbiotic interactions where heterotrophic bacteria serve as a source of carbon dioxide for photosynthetic organisms, thereby enabling the production of biomass and oxygen during respiration. Furthermore, excreted carbohydrates, vitamins, and organic compounds serve as vital nutrients for both algae and bacteria. This nutrient exchange is facilitated by the initial colonization of bacteria, which expedites the formation of algal biofilms (G. Roeselers et al.; 2007, É. Ács et a.; 2007).

Developmental Stages and Species Succession

The developmental stage of a biofilm exerts a discernible influence on species succession, resulting in alterations in the relative abundance and proportions of algae, bacteria, and extracellular polymeric substances (EPS) . In the early stages of photosynthetic biofilm formation, a higher proportion of EPS and bacteria is observed in comparison to algae and cyanobacteria, a phenomenon colloquially referred to as “conditioning”. Subsequently, following the establishment of the EPS matrix, algae exhibit rapid growth in the upper layers of the biofilm, thereby prompting bacteria to form aerial colonies in competition for essential nutrients.

Succession Patterns in Algal Groups

Biofilm maturity also exerts discernible effects on the dominance of specific algal groups. Diatoms are found to predominate in the early stages of biofilm development (typically within the first 15–20 days), while filamentous chlorophytes become more prevalent in later stages (Besemer K, Singer et al.; 2007, Johnson Reet al.; 1997). Some investigations have suggested a succession pattern characterized by an initial dominance of green algae during the early phase (1–4 days), followed by diatoms during the subsequent phase (5–9 days), and ultimately giving way to cyanobacteria during the third phase (10–15 days) (Sekar R et al.; 2004). Cyanobacteria are identified as late successional microorganisms (Zippel B et al.; 2005) with their distribution and prevalence being influenced by a multitude of factors (Rao TS et al.; 2010, Barranguet C et al.;2005, Sekar R et al.; 2004 ).

The presence of grazers, including Chironomids, Gastropods, Trichopteran larvae, Ephemeropteran larvae, and crustaceans, has a pronounced impact on reducing algal biomass throughout the seasons, with summer exhibiting the most significant effects (Hillebrand H et al.; 2001). Notably, filamentous species and chain-forming diatoms are more susceptible to grazing than single-celled algae species (Hillebrand H et al.;2002).

The succession of algal species within photosynthetic biofilms is governed by a complex interplay of factors, including light intensity, temperature, nutrient concentrations, and shear rates, with seasonal variations playing a pivotal role in shaping these dynamics see fig 4.

algal biofilms applivation — Fig 4 Different factors for algal biofilm and its application

Metabolic pathway

Metabolic pathways in microalgae cultivation are influenced by the availability and form of nutrients, primarily driven by the assimilation of available carbon (Markou G et al., 2014). These pathways play a crucial role in enhancing biomass production and shaping the composition of intracellular metabolites like proteins, lipids, and pigments. They rely on inputs such as light and carbon sources (Wang J et al.,2014).

Heterotrophic cultivation of microalgae is considered the most promising approach for achieving higher biomass yield and optimizing biochemical composition (Chen F. et al.,1996, Yang C et al.,2000). Generally, there are three distinct metabolic pathways in microalgae, determined by their nutritional requirements for growth and valuable biochemical production.

The main cultivation conditions are photoautotrophic cultivation, heterotrophic cultivation, and mixotrophic cultivation.

Photoautotrophic cultivation

The photosynthetic pathway involves converting sunlight and carbon dioxide into energy and cellular carbon dioxide, directing biochemical production in microalgae (Burkholder JM et al., 2008).

In photoautotrophic cultivation, microalgae exhibit varying lipid contents, ranging from 5% to 68%, depending on the species (Chen CY et al.; 2011). To boost lipid content during growth, nitrogen or nutrient limitations can be applied (Mata TM et al.,2010). The highest reported lipid productivity in microalgae under photoautotrophic conditions reached 179 mg/L/d using Chlorella sp. with 2% carbon dioxide and 0.25 vvm aeration (Chiu SY et al.; 2008).

Heterotrophic cultivation

Heterotrophic cultivation involves microalgae using organic compounds as both carbon and energy sources (Chojnacka K et al.; 2004). Some microalgae species can thrive in both photoautotrophic and heterotrophic conditions.

Only a few microalgae species have succeeded on a large scale in the heterotrophic mode (Lee DU et al.; 2001), but it is more efficient in terms of cell production per unit energy compared to the autotrophic mode (Yang C et al.; 2000). For instance, Chlorella protothecoides experienced a 40% increase in lipid content when shifting from photoautotrophic to heterotrophic cultivation ( Xu H et al.; 2006).

Microalgae can utilize various organic carbon sources during growth, including sucrose, glucose, lactose, galactose, glycerol, and fructose (Liang Y et al.;2009). Some microalgal species are obligate heterotrophs and are cultivated in heterotrophic mode, especially for lipid and pigment production from wastewater. This approach allows for achieving high cell densities, up to 100 g/L, facilitating microalgae biomass harvesting (Morales-Sánchez D et al.; 2015).

Mixotrophic cultivation

Mixotrophic cultivation combines autotrophic and heterotrophic modes, enhancing microalgae growth and resource utilization (Burkholder JM et al.; 2008). Microalgae use organic carbon compounds and CO2 as carbon sources, recycling CO2 during phototrophic conditions when light is available (Mata TM et al.; 2010). This approach aids global carbon dioxide reduction by utilizing carbon dioxide for microalgae growth. Industrial wastewater serves as a carbon source when combined with a light source, offering an efficient and cost-effective microalgae cultivation method (see fig 5).

Conclusion

Algal biofilms, a captivating realm within microbiology and environmental science, showcase intricate communities of microorganisms residing within a matrix of extracellular polymeric substances (EPS). Dominated by microalgae, cyanobacteria, and heterotrophic bacteria, these biofilms are versatile colonizers, populating a range of surfaces where moisture and light are abundant, from river stones to plastic surfaces and tree bark. Their ability to thrive and adapt in diverse conditions, using three primary metabolic pathways (photoautotrophic, heterotrophic, and mixotrophic cultivation), underscores their resourcefulness.

Beyond their aesthetic appeal, algal biofilms play an active ecological role. They contribute to water purification, nutrient cycling, and the production of valuable biomass and bioactive compounds, reflecting the dynamic nature of natural ecosystems. In a world grappling with environmental challenges, algal biofilms offer hope and insight, serving as a model of resilience. These microorganisms, without traditional plant structures, harness the power of photosynthesis, adapting to changing conditions, and exemplifying the ingenuity and tenacity of life.

Algal biofilms are not just subjects of scientific inquiry; they are living testaments to the intricate interplay of microorganisms in nature, demonstrating the remarkable potential for adaptation and survival. In a changing world, their adaptability and resourcefulness inspire us to explore and understand the complex relationships within ecosystems.

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Bio Remediation/ Phycoremediation

Biofilms: A Dynamic Community in Equilibrium with the Environment

September 11, 2023September 12, 2023
by Pooja Vaswani

Biofilm, a complex assembly of microbial cells anchored to surfaces within a polysaccharide matrix, forms a harmonious and organized microbial community. This strategy enables microorganisms to thrive in favorable conditions, avoiding displacement by currents. Comprising cells and extracellular polymeric substances, biofilms exhibit dynamic equilibrium with their surroundings, cycling through growth, death, and regeneration.

Microorganisms gain remarkable advantages from biofilm cultivation, including protection from harsh conditions, enhanced resilience to stress, and cooperative metabolic and gene expression adjustments. Microorganisms thriving in biofilms exhibit an astonishing ability to adapt, demonstrating a captivating collective and synchronized behavior that intrigues both scientists and enthusiasts. (Donlan et al., 2002; Chmielewski and Frank et al., 2004; Van Houdt and Michiels et al., 2010). Multispecies biofilms are prevalent in nature, significantly impacting ecosystems (Donlan et al., 2002; Hall-Stoodley et al., 2004).

1. Microbial Biofilm Formation: A Complex Developmental Process in Bacteria

Biofilm formation is a captivating process wherein bacteria transition from single cells to structured multicellular communities. These parallels other bacterial developmental processes like sporulation (Dunny GM, Leonard BA. Et al.,1997), fruiting body formation (Plamann L et al.,1995, Shimkets et al.;1999, Wall D et al.,1999), and stalked-cell formation (Fukuda AK et al.,1977, Hecht GB et al., 1995, Trun NJ et al.,1990, Quon KC et al., 1996, Wu J et al.,1997). In nature, biofilms exist as diverse microbial communities, with bacteria joining, departing, exchanging genetic material, and occupying specific niches, resembling complex cities rather than developed organisms.

1.1 Biofilm Formation in Non-Motile Species

In non-motile species, during favorable conditions for biofilm formation, bacteria increase adhesin expression, enhancing stickiness for cell-to-cell and cell-to-surface adherence (Gotz, 2002). Surface proteins like Bap(biofilm-associated protein) in staphylococci aid cell interactions and matrix creation (Lasa and Penades et al., 2006). Similar proteins exist in other species, often with repeated domains undergoing recombination within the bap gene, yielding variable-length proteins (Latasa et al., 2006). Additionally, species that lack motility also generate exopolysaccharides (EPS), which subsequently become crucial constituents of the extracellular matrix. An illustrative instance of this is the production of PIA(Polysaccharide intercellular adhesion )/PNAGS (Poly-N-acetylglucosamine )EPS by the gene products originating from the ica operon in staphylococcal species. Thus, altered surface proteins and EPS production are key in initiating biofilm formation in non-motile bacteria.

1.2 Biofilm Formation in Motile Species

In motile species, conducive conditions trigger bacteria to adhere to a surface, lose motility, and form biofilms. An extracellular matrix binds them together. Flagella are vital for biofilm initiation; flagella-minus mutants show reduced biofilm formation (Pratt and Kolter et al., 1998; Watnick and Kolter 1999; Lemon et al., 2007). Listeria monocytogenes recover initial adhesion with directed movement, indicating motility’s role in overcoming repulsion for biofilm formation (Lemon et al., 2007). The initial surface encounter leads to transient adherence, determining stable biofilm development or return to planktonic state.

2. Genetically Distinct Stages of Biofilm Formation

Biofilm formation can be divided into five genetically distinct stages:

1. Initial surface attachment

2. Monolayer formation

3. Migration to form multilayered microcolonies

4. Production of extracellular matrix

5. Biofilm maturation with characteristic three-dimensional architecture (O’Toole et al., 2000)(fig 1).

Fig 1 Decoding Bacterial Biofilms: Approaches to Regulate and Control

3. Regulatory Variations in Biofilm Formation

In numerous motile bacteria, initial surface attachment relies on flagella-mediated motility. However, specific Gram-negative bacteria require type IV pili-associated surface motility for microcolony and three-dimensional architecture formation. Notably, this motility is absent in Gram-positive bacteria, except for Clostridia ssp. (Varga et al., 2006; O’Toole et al., 2000)

4. Extensive Cellular Differentiation in Biofilm Environment

Once the bacteria have successfully adhered to the surface, they begin producing an extracellular matrix. This matrix serves as a crucial organizational element, enabling the formation of structured communities within the biofilm. As a result, extensive cellular differentiation can occur within the biofilm environment.

5. Biofilm Composition and Architecture

Its composition hinges on the inoculum’s characteristics, while external factors like substrate, nutrients, competition, and grazing shape colonization and growth (Baier et al., 1980; Characklis & Cooksey et al., 1983; Marshall et al., 1985).

The architecture of biofilm is shaped by hydrodynamics, nutrients, bacterial motility, communication, exopolysaccharides, and proteins. Altered biofilm morphology in mutants lacking components of extracellular polymeric substances (EPS) illustrates their impact. Exopolysaccharides in Vibrio cholerae and colanic acid in Escherichia coli influence three-dimensional biofilm formation.

5.1 Biofilm Architecture in Bacillus subtilis

In the case of Bacillus subtilis biofilm, the matrix consists of an exopolysaccharide and the secreted protein Tas A, both of which are essential for maintaining the structural integrity of the matrix and facilitating the development of biofilm architecture resembling fruiting body-like structures (Fig 2).

6. Factors affecting the formation of biofilm.

Biofilms form at interfaces of aqueous or gaseous phases and substratum surfaces in diverse systems. Metabolic substrates required for growth must be accessible in the aqueous phase (e.g., medical catheters) or between aqueous and solid phases (e.g., minerals). Nutrient quantity and ratios in the liquid phase impact growth-limiting factors (van Loosdrecht et al., 2002).

Biofilm development is influenced by factors like temperature, pH, Oxygen levels, hydrodynamics, osmolarity, ions, nutrients, and biotic elements, collectively shaping bacterial behavior as mentioned in Fig 3 (van Loosdrecht et al., 2002).

6.1 The Relevence of EPS in Biofilm Composition

In biofilms, microorganisms make up less than 10% of the dry mass, with over 90% being the EPS matrix. EPS facilitates cell adhesion, cohesion, and interactions, including cell-cell communication, promoting the formation of micro consortia. It acts as an external digestive system, retaining enzymes to sequester nutrients from the water phase. Biofilm morphology varies, from smooth and flat to rough, fluffy, or filamentous, even forming mushroom-shaped colonies surrounded by water-filled voids. The structure adapts in response to nutritional changes, supporting diverse habitats and the coexistence of mixed-species consortia (Klausen et al., 2003).

Table 1 Function of EPS for Biofilm Formation ( Flemming HC, Wingender J et al., 2010)

6.2 Surface factor

Interactions between bacterial cell walls and surfaces are influenced by interfacial electrostatic and van der Waals forces (McClaine JW et al.,2002, Vigeant MA et al.,2002). Other factors, such as hydration forces, hydrophobic interactions, and steric forces, also impact cell attachment (Findenegg GH. JN Israelachvili et al., 1985). Hydrophobic interactions (low surface energy) and electrostatic interactions charge have been extensively studied.

6.2.1 Interactions Driving Bacterial Cell Attachment to Surfaces: Electrostatic Force

Electrostatic forces drive bacterial cell attachment to surfaces, given bacteria’s net negative charge (Soni KA et al., 2008, Katsikogianni MG et al 2010). Positively charged surfaces facilitate rapid attachment, while negative surfaces hinder it. Extracellular organelles aid in overcoming repulsion (Bullitt E et al., 1995). High ionic strength reduces charge effects. Bacterial cell walls expose functional groups interacting with substrates (Hong Y et al.,2008). Adsorbed molecules influence surface chemistry and charge, aiding adsorption and biofilm growth. Hydrophobic groups and organelles stabilize interactions after repulsion.

Biofilms form on various materials in contact with bacteria-containing fluids, influenced by surface roughness, chemistry, and conditioning films (Donlan et al., 2002). Hydrophobic surfaces promote faster attachment through interactions with flagella, fimbriae, and pili (Donlan et al., 2002; Donlan and Costerton et al., 2002). However, exceptions like Listeria monocytogenes prefer hydrophilic surfaces (Chavant et al., 200). Clinical isolates of Staphylococcus epidermidis favor biofilm formation on hydrophobic substrates (Cerca et al., 2005). Hydrophilic surfaces generally exhibit higher bacterial attachment than hydrophobic surfaces (Donlan, 2002).

6.2.2 High energy surface

Thermodynamic analysis of surface energies reveals insights into bacterial adhesion (Absolom DR 1983). Hydrophilic surfaces enhance bacterial adhesion when cell wall surface tension exceeds the surrounding liquid (Absolom DR et al.,1983). Fluorinated materials with large contact angles indicate low-energy surfaces (Absolom DR et al., 1983). Oxidation of fluorinated surfaces reduces initial bacterial attachment due to altered hydrophilic properties (Davidson CA et al., 2004). Bacterial attachment depends on surface free energy; high surface free energy surfaces like stainless steel and glass are more hydrophilic (Davidson CA et al., 2004). Biofilm growth depends on various factors, including nutrient concentrations and light availability, once bacterial cells adhere and achieve confluence.

6.3 Role of Biosurfactants in Bacterial Attachment and Biofilm Formation

Biosurfactants with antibacterial and antifungal properties are crucial for bacterial attachment to and detachment from oil droplets (Kim HJ et al., 2011 ). Research interest in environmentally friendly chemicals has grown, including biosurfactants (Kim HJ, Boedicker JQ, Choi JW, Ismagilov RF et al., 2008, Eun YJ et al.,2009 ). Microorganisms produce biosurfactants at the air-water interface, influencing surface tension and gas exchange in surface waters (Flickinger ST et al.,2011). Rhamnolipids, found in the EPS matrix of P. aeruginosa, act as surfactants, contributing to microcolony formation, bacterial migration, and biofilm dispersion (Boedicker JQ et al.,2009) (Vincent ME et al.,2010, Renner LD, Weibel DB et al.,2011) (Harmsen M et al., 2010).

6.4 Influence of Quorum Sensing (QS) on Biofilm Formation

Quorum sensing (QS) is a crucial regulatory mechanism for biofilm formation (Parsek and Greenberg 2005). Microorganisms release auto-inducers (AIs) that induce or repress QS-controlled genes, impacting biofilm structure and 3-dimensional organization (Steinmoen et al. 2002). QS also influences population size and dispersion in biofilms (Lewis et al., 2001; Davies et al., 2003; Jesaitis et al. 2003). Additionally, QS can induce behaviors and control group activities, affecting biofilm development and resistance to stress (Camilli and Bassler et al.,2006). Cell properties like hydrophilicity, fimbriae, and flagella also influence microbial attachment in biofilms.

6.5 Chemotaxis triggers biofilm formation

Chemotaxis is vital for biofilm formation in bacteria like Pseudomonas aeruginosa, E. coli, and Vibrio cholerae (O’Toole GA et al., 1998, Pratt LA et al.,1998, Watnick PI et al.,1999). Bacteria sense chemical stimuli, use flagella to swim toward nutrients, and attach to the substrate. Flagella and/or type IV pili are essential for initial cell adhesion, leading to microcolony formation (Stelmack PL et al.,1999). E. coli also utilizes type I pili for initial attachment. Bacterial chemotaxis aids biofilm growth and spread as it develops.

6.6 Importance of Horizontal Gene Transfer (HGT) in Microbial Communities

Horizontal gene transfer (HGT) is crucial for microbial evolution, facilitated by mobile genetic elements like plasmids, transposons, and bacteriophages (Koonin EV et al., 200) Bacterial biofilms show distinct gene expression compared to planktonic counterparts, influenced by the surface they settle on (Keyhani NO et al.,1996) HGT occurs within biofilms, allowing gene transfer between bacteria (Roberts AP et al.,1999).HGT also plays a significant role in animal evolution, diseases, and intercellular processes, like Agrobacterium transferring genetic material to plants (Guo M et al.,2019). Chemotaxis influences HGT, seen in Agrobacterium’s DNA transfer to plant cells (Guo M et al.,2019)., Niehus R et al.,2015). Other pathogenic bacteria also employ chemotaxis-derived HGT within host cells (Dougherty K et al.,2014 , Desmond E et al.,2007).

6.7 Interspecies Interactions in Bacterial Adhesion: Implications for Biofilm Stabilization and Commensal Relationships

McEldowney and Fletcher (McEldowney S, Fletcher M et al.,1987) showed that bacterial adhesion to a surface can affect other species differently, with negative, positive, or neutral outcomes. Inhibitory interactions may involve cell blockage or secretion of inhibitory macromolecules (Belas MR, Colwell RR et al.,1982).

6.7.1 Positive Interactions during Adhesion

Positive interactions during adhesion can result from bacterial products modifying the conditioning film or direct cell-to-cell contact. Dental plaque exemplifies direct interactions, where ligand-receptor interactions cause interspecies coaggregation (Kolenbrander PE et al., 1989). Hydroxyapatite attachment enhances the adhesion of co-aggregating pairs (Ciardi JE, McCray GF et al.,1987, Schwarz SU, Ellen RP 1987 ). Other interactions, like mutans streptococci adhering to immobilized oral bacteria, play a role in biofilm formation (Lamont RJ et al.,1990). Interspecies interactions are crucial during the initial stages of biofilm formation.

6.7.2 Neutral Adhesion Interactions

The attachment of one species to a substrate may not always be influenced by another species’ attachment (Cowan MM et al.,1991, McEldowney S et al.,1987). Neutral interactions could result from separate binding sites on the substrate for each species, including multiple high- and low-affinity sites within species (Gibbons RJ et al.,1983, Korber DR et al.,1994). For example, a nonmotile mutant strain of Pseudomonas fluorescens attached to glass independently of the parent strain, suggesting different binding sites (, Korber DR et al.,1994). Adhesion interactions depend on the surface and species involved (McEldowney S, Fletcher M et al.,1987), leading to variations in adhesion mechanisms for different substrates. Vibrio proteolytica’s adhesion to hydrophobic substrates was inhibited by proteases, while hydrophilic substrates were unaffected (Paul JH et al.,1985).

6.8 Commensal Interactions in Biofilm Stabilization

One intriguing aspect of biofilm stabilization is that it can be seen as a commensal interaction, where one species benefits from another’s ability to form a stable film.

6.8.1 Types of Commensal Interactions in Biofilms

Commensal interactions in biofilms occur when one population benefits without affecting the other. Oxygen consumption by aerobic/facultative microorganisms creates oxygen gradients, enabling the growth of obligate anaerobes (Costerton JW et al.,1994,De Beer D et al.,1994, Lewandowski Z et al.,1994). This interaction is crucial for sulfate-reducing bacteria growth in anaerobic microniches, contributing to microbially-induced corrosion (Hamilton WA. et al.,1985, Lee W, Lewandowski Z et al.,1993).

6.8.2 Commensalism in Dental Plaque Development

In dental plaque development, commensalism involving oxygen consumption occurs, with aerobic bacteria declining, and anaerobic Veillonella spp. predominating in deeper layers, while aerobic species dominate upper layers (Ritz HL et al.,1967 1969). Also in waste-water treatment biofilms, layering of aerobic and anaerobic bacterial species illustrates commensal interactions (Alleman JE, Veil JA et al.,1982). Other commensal interactions like substrate provision may be common in biofilms but require further research (Ritz HL et al.,1967).

7. Symbiotic Relationships and Enhanced Growth in Bacterial-Associated Algal Biofilms

Bacteria initiate EPS matrix formation in algal biofilms, leading to symbiotic relationships and competition with algae (Davis LS, Hoffmann JP et al.,1990, Mack WN et al., 1975). Studies show significant benefits of bacterial presence for algal recruitment and growth (Irving TE et al.,2011, Hodoki Y et al.,2005, Sekar R et al., 2004, Holmes PE et al., 1986). Algal biomass increases with rising pre-conditioned bacterial biofilm concentrations (Hodoki Y et al.,2005). Mixed community biofilms exhibit higher algal growth compared to pure algal cultures (Holmes PE et al., 1986).

Algae biofilms in non-sterile wastewater show increased thickness, switch to a sessile state, and resist shear stress better due to bacteria and EPS presence (Irving TE et al.,2011).Irving and Allen (Irving TE et al.,2011) demonstrated a nine-fold increase in algae biofilm thickness when grown on non-sterile secondary wastewater effluent compared to sterile wastewater. They also found that in the presence of unsterile wastewater, algae cells had a greater tendency to switch from a planktonic (free-floating) to a sessile (attached) state and exhibited enhanced resistance to shear stress.

Fig 8 Bacterial association with microalgae

8. Impactful Roles of Biofilm in Chemistry and Environmental Restoration

Biofilms are valuable in chemical synthesis processes, including ethanol, poly-3-hydroxybutyrate, and benzaldehyde production (Kunduru and Pometto, 1996; Li et al., 2006; Zhang et al., 2004). They also play essential roles in wastewater treatment, phenol bioremediation, and the degradation of dinitrophenols and toxic metals in environmental remediation (Lendenmann et al., 1998; Luke and Burton, 2001; Nicolella et al., 2000; Singh and Cameotra, 2004). Biofilms are highly resilient and serve as valuable tools for beneficial purposes in various chemical processes and environmental applications.

9. Conclusion

Biofilms represent dynamic and well-organized microbial communities that play a vital role in various ecological and industrial processes. They form through a complex developmental process, transitioning bacteria from a free-floating state to sedentary, multi-layered communities. The formation of biofilms involves various factors, such as surface characteristics, chemotaxis, and horizontal gene transfer, which contribute to the unique three-dimensional architecture and stability of these communities.
Interspecies interactions within biofilms further enhance their complexity, leading to commensal relationships and symbiotic associations that benefit different microbial species within the community. Bacteria initiate biofilm formation, and their presence fosters the recruitment and growth of algae, creating symbiotic relationships within mixed community biofilms.
The significance of biofilms extends beyond microbial interactions, as they also find applications in chemical synthesis and environmental restoration. Biofilms play crucial roles in wastewater treatment, bioremediation of contaminated sites, and the production of various chemicals, showcasing their potential as valuable tools for beneficial purposes.
Overall, the study of biofilms provides valuable insights into the intricacies of microbial communities, their interactions, and their adaptability to diverse environments. Understanding biofilm formation and behaviour can lead to the development of innovative strategies for improving industrial processes, environmental management, and human health. As researchers delve deeper into the world of biofilms, they uncover the fascinating intricacies of these dynamic microbial communities and their impact on the ecosystem and human activities.

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Algal Biotechnology

Microalgae: A Powerful Tool for Climate Change and Water…

July 4, 2023July 7, 2023
by Pooja Vaswani

Global climate change has emerged as a pressing global issue in contemporary times. The primary contributors to this phenomenon are the rising levels of greenhouse gases, predominantly originating from human activities and the operations of power plants (Watanabe Y, et., al; 2017).

Human activities, such as fossil fuel usage and industrial production, as well as natural processes involving the Earth’s oceans, soil, plants, animals, and volcanoes, release greenhouse gases, including CO_2, CH₄, and N₂O, into the atmosphere. However, since the Industrial Revolution, human activities have emerged as the primary contributors to greenhouse gas emissions (Baba Mohammad et al., 2015; Mirzaei et al., 2015).

Specific Sources of CO₂ Emissions

According to Le Quere (Le Quere et al. 2013, 2015), the combustion of fossil fuels, namely coal, natural gas, and oil, is responsible for about 87% of human-produced CO₂emissions. Among these fuels, coal combustion contributes to 43% of CO₂ emissions from fuel burning, followed by oil at 36% and natural gas at 20%. Power plants, vehicles, aircraft, and industrial facilities primarily use these fossil fuels to generate heat, electricity, and power. The largest share of man-made CO₂emissions, approximately 41%, originates from power generation and heating activities. Transportation ranks as the second largest source, accounting for around 22% of CO₂ emissions resulting from fossil fuel burning. Industries contribute approximately 20% of CO₂ emissions from fossil fuel combustion. Other human-related sources include deforestation and land-use changes, contributing to roughly 9%, and industrial processes like cement manufacturing, which account for around 4%.

The emission of CO₂ into the atmosphere involves not only human activities but also natural processes. According to a study conducted by Denman et al. In 2007, researchers found that natural sources significantly contribute to CO2 emissions. The Earth’s oceans, through the exchange between the ocean and the atmosphere, account for a substantial 42.84% of these natural emissions. Additionally, sources like plant and animal respiration, soil respiration, and decomposition contribute to 28.56% of the total CO₂ emissions generated naturally.

CO₂: A major Green House Gas

Research efforts are ongoing and focused on tackling global warming by actively mitigating greenhouse gas emissions. While carbon dioxide naturally exists as a greenhouse gas, the concentration of CO₂in the atmosphere has undergone a substantial increase because of industrialization and human activities (National Research Council et al.,2011; Siegenthaler U et al., 2005; Chang EH et al, 2003).

According to a report by the French National Center for Scientific Research, atmospheric CO₂levels have soared to 380 parts per million (ppm), marking a twofold increase compared to the previous century (Morais and Costa, et al.,2007). The notable surge in CO₂ concentration directly attributes to the combination of population growth and industrialization. Future projections indicate that by 2100, CO₂ emissions in the atmosphere could reach a staggering 26 billion tons, surpassing the emissions recorded in the past century by 18.5 billion tons (Chiu et al., 2008).

Power plants play a significant role in releasing CO₂ into the atmosphere through the combustion of fossil fuels (Benemann JR et al., 1993). To tackle the challenge of CO₂ emissions from this source, it is imperative to prioritize efforts in enhancing the efficiency of power plant generation and transitioning towards cleaner and more sustainable alternatives.

Biological CO₂ Sequestration Technology/Method

In 1990, the Research Institute of Innovative Technology for the Earth (RITE) launched the Biological CO₂ Fixation and Utilization Project under the sponsorship of the Ministry of International Trade and Industry (MITI). The New Energy and Industrial Technology Development Organization (NEDO) commissioned this pioneering initiative (Murakami, N, et al., 1996).

Among the array of available CO₂ capture methods, the biological approach stands out as an attractive alternative. This approach harnesses the power of photosynthesis, which enables the conversion of carbon dioxide into organic matter, fueled by sunlight as an energy source. In this regard, photosynthetic microorganisms have emerged as a preferred choice, owing to their remarkable characteristics. These microorganisms possess the unique ability to assimilate CO₂ into carbohydrates, lipids, and proteins using solar energy. They exhibit higher rates of CO₂ fixation compared to land plants and offer better compatibility for integrating CO₂ removal systems into industrial processes when compared to other photosynthetic systems involving higher plants.

Algae potential players for CO₂ Sequestration.

Within the realm of photosynthesis, there are two fundamental processes at play: the light-dependent reaction, which entails a series of intricate steps reliant on the availability of light, and the light-independent reaction, also recognized as carbon fixation. In the context of microalgae, they have developed a specialized mechanism known as the CO₂concentrating mechanism (CCM) to secure an ample reservoir of inorganic carbon required to support cellular growth and proliferation during the process of photosynthesis (Brueggeman et al., 2012).

The CO₂concentrating mechanism (CCM) plays a crucial role in microalgae by effectively increasing the concentration of CO₂ at the active site of the enzyme Rubisco (ribulose-1,5-bisphosphate carboxylase oxygenase). These microorganisms possess specialized inorganic carbon transporters located in the plasma membrane and thylakoid membrane, each with varying affinities and flux rates for HCO₃ (bicarbonate) and CO₂ (Beardall and Raven, 2017). Carbonic anhydrase (CA) is responsible for converting HCO₃ into CO₂, and the proper functioning of both CA and the CCM is highly dependent on the availability of CO₂ (Zou et al., 2004; Zhou et al., 2016) See Fig 1.

Carbon-capture-mechanism-CCM-by-algae for climate change

As a result, the availability of CO2 intricately ties to the activity of these enzymes. While elevated CO₂ levels can promote the growth of algae, they may also impede the activity of carbonic anhydrase (CA) and the functioning of the CO₂concentrating mechanism (CCM) (Xia and Gao, 2005).

Microalgae surpass terrestrial plants in their efficiency in converting CO₂ into organic compounds. They exhibit approximately 10 times greater efficacy in CO₂ fixation compared to land plants.

Algae cultivation in Wastewater an alternative approach for wastewater treatment & CO₂Sequestration.

The combination of biological CO₂ fixation using microalgae and wastewater treatment presents significant advantages in terms of both economic feasibility and environmental sustainability. By cultivating microalgae in nutrient-rich wastewater, these microorganisms have ample access to the nutrients needed for their growth. As they flourish, microalgae absorb CO₂ and convert it into biomass, which can be utilized as a valuable feedstock for biofuel production (Kannan DC et., 2022, Magar et al., 2022).

The integration of CO₂ capture, wastewater treatment, and biofuel production creates a synergistic and compelling approach. It not only contributes to reducing CO₂ emissions by capturing and utilizing CO₂ but also addresses the crucial requirement for effective wastewater treatment. This combined strategy demonstrates a promising solution that not only helps mitigate climate change but also promotes the efficient utilization of resources while generating a valuable end product (Wang B et al., 2008).

The integration of microalgae in wastewater treatment presents a sustainable and eco-conscious solution that minimizes or eliminates the requirement for chemical treatments in wastewater plants. This groundbreaking technology utilizes the inherent abilities of microalgae to effectively extract nutrients, specifically nitrogen and phosphorus compounds, from wastewater. In addition to facilitating CO₂ capture through photosynthesis, this approach successfully tackles the pressing challenge of nutrient removal in wastewater.

Mechanism of Microalgae-based Wastewater Treatment

Microalgae offer an invaluable solution to address the challenge of microcontaminants found in wastewater effluents, which can pose significant risks when they enter drinking water treatment processes (Beelen ES et al., 2007). By efficiently eliminating these microcontaminants, microalgae greatly improve the overall quality of water. Additionally, microalgae play a vital role in reducing nutrient levels, such as carbon, nitrogen, and phosphorus, while simultaneously releasing significant amounts of oxygen. This dual action not only aids in the decomposition of organic matter by bacteria but also helps mitigate odor-related issues commonly associated with wastewater (De Pauw N et al., 1983).

The abundant availability of nutrients in municipal wastewater makes it an ideal resource for harnessing the potential of microalgae. Furthermore, the close proximity of wastewater treatment plants to power plants, which generate significant amounts of CO₂ in flue gases through the combustion of fossil fuels, further emphasizes the benefits of this approach. This strategic positioning enables the efficient utilization of readily available CO₂ resources.

Microalgae cultivation in wastewater presents versatile opportunities, allowing for the use of either open or closed systems. These approaches enable precise control over the growth media, optimizing the removal of nutrients from wastewater and maximizing the production of valuable microalgae biomass. This customized approach ensures effective and sustainable wastewater treatment while generating valuable resources in the form of microalgae biomass.

Open ponds

High-rate algal ponds, commonly known as HRAPs or raceway ponds, are the prevailing large-scale production systems in practice. These open, shallow ponds have been employed since the 1950s and utilize a paddle wheel to ensure the circulation of algae and nutrients. While raceway ponds are relatively affordable to construct and operate, their productivity is often hindered by challenges such as contamination, inadequate mixing, the presence of dark zones, and inefficient utilization of CO₂ (Chisti, 2007; Mata et al., 2010).

In theory, raceway ponds should achieve production levels of 50–60 g m−2 day−1, and there have been instances of reaching such levels in a single day (Sheehan et al., 1998). However, in practical applications, it proves difficult to attain even 10–20 g m−2 day−1 productivity (Shen et al., 2009). While the high evaporation rate of open ponds is often perceived as a limitation, it does provide some benefits by aiding in temperature regulation through evaporative cooling (U.S. DOE, 2010).

Moreover, the HRP system’s simplicity and symbiotic nature make it a perfect candidate for emulating natural ecosystems in laboratory and field settings, aligning harmoniously with the principles of ecological engineering (Cunningham M et al., 2010).

An important conclusion drawn from cost analysis studies conducted by the U.S. Department of Energy’s Aquatic Species Program was that there are limited prospects for alternative systems to replace the open pond approach, particularly when considering the need for low fuel costs (Sheehan et al., 1998).

Closed reactors

According to Chisti (2007), tubular photobioreactors are the exclusive type of closed systems used on a large scale. These reactors come in vertical, horizontal, and helical designs, with helical ones being regarded as the easiest to expand (Carvalho et al., 2006). In comparison to open ponds, tubular photobioreactors offer several advantages such as improved control over pH and temperature, better protection against contamination, enhanced mixing, reduced evaporation, and higher cell densities (Mata et al., 2010).

Reported productivities typically range from 20 to 40 grams per square meter per day (Shen et al., 2009). However, despite these benefits, tubular reactors have not gained significant usage due to various challenges. Issues such as the accumulation of toxic oxygen, unfavorable pH and CO₂ gradients, overheating, bio-fouling, and high material and maintenance costs have limited their widespread adoption (Mata et al., 2010; Molina Grima et al., 1999). Among these challenges, the removal of oxygen is considered one of the most difficult problems to overcome, particularly when scaling up the system. It effectively imposes limitations on the length of tubes or panels and necessitates a more complex or modular design (Carvalho et al., 2006

How Algae CO₂Sequestration Potential is Calculated?

To tackle the challenge of reducing CO₂ emissions, researchers have explored various avenues, which can be broadly divided into two groups: (i) methods based on chemical reactions and (ii) biological methods (Benemann JR et al., 1993, Wang B et al.,2008). Among these, biological processes have emerged as compelling solutions to combat global warming, attracting significant attention from the scientific community (Yun YS et al., 1997).

Chisti emphasizes that microalgal biomass typically consists of around 50% carbon, allowing for the capture of approximately 1.83 kg of CO₂ per kilogram of biomass produced (Chisti et al., 2007). In the field of CO₂ capture using microalgae, many studies rely on measuring CO₂ concentrations at the inlet and outlet of cultivation reactors. However, this approach may pose challenges as it does not guarantee that all consumed CO₂ is solely attributed to the growth of microalgae. Alternatively, estimating the carbon content can provide a more precise assessment of the CO₂ consumption by microalgae cells, assuming that the culture medium does not contain carbon sources other than CO₂ (Yun YS et al., 1997; Tang D et al., 2011).

Based on this premise, researchers can estimate the rate of CO₂ capture (RCO₂) using the following formula.

RCO₂ = P·CCO₂·MCO₂/MC,

where P represents biomass productivity (g L-1 day-1), CCO₂ denotes the carbon content of microalgae biomass obtained from CO₂, and MCO₂and MC correspond to the molecular weights of carbon dioxide and carbon, respectively.

CO₂ concentration and CO₂ uptake efficiency of Microalgae

The cultivation of photoautotrophic microalgae relies on carbon dioxide (CO₂) as the primary source of carbon. Despite the physiological significance of CO₂ concentration on microalgae cell growth, researchers have conducted limited research to analyze CO₂ uptake under relevant process conditions, indicating the need for further investigation (Cheng L et al., 2006). Atmospheric air contains minimal amounts of CO₂, which cannot sufficiently support microalgae cell growth due to limited mass transfer driving forces. To overcome this challenge, researchers can obtain CO₂ from pure sources or harness it from flue gases, thereby addressing the environmental concern associated with CO₂ capture (Acién Fernández FG et al., 2012).

Microalgae and cyanobacteria have the ability to adapt their photosynthetic properties, including the carbon concentrating mechanism (CCM), to high concentrations of CO₂ (Baba et al., 2011). Certain microalgae and cyanobacteria species, such as Chlamydomonas reinhardtii, can quickly acclimate to a CO₂ concentration of 20% within a few days (Hanawa et al., 2007).

When developed at air-equilibrated CO₂ levels, microalgae and cyanobacteria can effectively utilize low levels of dissolved inorganic carbon (DIC). Cyanobacteria actively increase calcite deposition at higher CO₂ concentrations, which plays a crucial role in CO₂ removal. This increase leads to a 1.5- to 2.5-fold enhancement in CO₂ fixation rates (Ramanan et al., 2009, 2010; Badger and Price, 1994).

Enhancing CO₂Mass Transfer with Microbubbles

Some of the important physical factors that change the mass transfer rate of CO₂ from the air to liquid are air bubble size, the pore size of the sparger, flow rate, air holding time of the liquid system, etc. Microbubble size is a very essential aspect to enhance the CO₂ mass transfer rate (McGinn PJ et al., 2011). However, high CO₂ concentrations hinder the growth of microalgae and cyanobacteria (Ramanan et al., 2010).

Previous research has reported that optimal CO₂ fixation occurs at concentrations ranging from 2% to 5% CO₂, with 3% CO₂ being approximately 489 times higher than the atmospheric CO₂concentration (Douskova et al., 2009; Ibn-Mohammed et al., 2013). These studies indicate that many algal strains may not require undermined haphazard high CO₂ feeding. Excess-supplied CO₂ mostly goes to waste, and it is necessary to determine the critical CO₂ concentration required by each strain.(Chaitanya Magar et al. 2019, Rambhiya, S. J. et al 2021).

Conclusion

In summary, the utilization of microalgae cultures presents a promising solution for both CO₂capture and biomass production. By cultivating microalgae in wastewater media, it becomes possible to remove significant pollutants from the wastewater while efficiently capturing CO₂ and converting it into valuable biomass feedstock. The selection of suitable algae strains and optimization of key parameters such as pH, temperature, and CO₂concentration is crucial in maximizing CO₂ fixation efficiency in biological systems. Certain algae species, such as Scenedesmus dimorphus, Scenedesmus incrassatulus, Chlorella sp., and Scenedesmus obliquus, demonstrate remarkable CO₂ fixation rates.

Moreover, integrating CO₂ capture from flue gases with microalgae-based wastewater treatment holds immense potential for sustainable biofuel production. This integrated approach offers the opportunity to address environmental concerns by capturing CO₂ emissions, treating wastewater, and producing valuable biofuel resources concurrently. To enhance the efficiency and effectiveness of microalgae cultivation, the development of comprehensive dynamic models becomes essential. These models should accurately depict the growth rates of microalgae under various limiting factors, enabling the design of optimized control systems and the identification of optimal operational conditions.

By advancing our understanding of the complex interactions and dynamics involved in microalgae-based systems, we can unlock the full potential of this technology. Ultimately, this will contribute to reducing reliance on fossil fuels, mitigating CO₂ emissions, and promoting a more sustainable and environmentally friendly approach to energy and wastewater management.

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Unveiling the Hidden World of Algal Biofilms: Nature’s Microscopic…

Adherence and Attachment

Algal Biofilm Habitats: Nature’s Adaptive Colonizers

Composition of algal biofilms

Factor affecting algal biofilm.

Light

Temperature

Nutrients

Monod’s Kinetic Expression in Algal Biofilm Research

pH

CO2

Substratum

Hydrophobicity

Flow Velocity

Extracellular polymeric substances (EPS)

Environmental Factors Influencing EPS Production

Species interaction

Developmental Stages and Species Succession

Succession Patterns in Algal Groups

Metabolic pathway

Photoautotrophic cultivation

Heterotrophic cultivation

Mixotrophic cultivation

Conclusion

References

Reference

Reference

References

References

References

References

Reference

Reference

Biofilms: A Dynamic Community in Equilibrium with the Environment

1. Microbial Biofilm Formation: A Complex Developmental Process in Bacteria

1.1 Biofilm Formation in Non-Motile Species

1.2 Biofilm Formation in Motile Species

2. Genetically Distinct Stages of Biofilm Formation

3. Regulatory Variations in Biofilm Formation

4. Extensive Cellular Differentiation in Biofilm Environment

5. Biofilm Composition and Architecture

5.1 Biofilm Architecture in Bacillus subtilis

6. Factors affecting the formation of biofilm.

6.1 The Relevence of EPS in Biofilm Composition

Table 1 Function of EPS for Biofilm Formation ( Flemming HC, Wingender J et al., 2010)

6.2 Surface factor

6.2.1 Interactions Driving Bacterial Cell Attachment to Surfaces: Electrostatic Force

6.2.2 High energy surface

6.3 Role of Biosurfactants in Bacterial Attachment and Biofilm Formation

6.4 Influence of Quorum Sensing (QS) on Biofilm Formation

6.5 Chemotaxis triggers biofilm formation

6.6 Importance of Horizontal Gene Transfer (HGT) in Microbial Communities

6.7 Interspecies Interactions in Bacterial Adhesion: Implications for Biofilm Stabilization and Commensal Relationships

6.7.1 Positive Interactions during Adhesion

6.7.2 Neutral Adhesion Interactions

6.8 Commensal Interactions in Biofilm Stabilization

6.8.1 Types of Commensal Interactions in Biofilms

6.8.2 Commensalism in Dental Plaque Development

7. Symbiotic Relationships and Enhanced Growth in Bacterial-Associated Algal Biofilms

8. Impactful Roles of Biofilm in Chemistry and Environmental Restoration

9. Conclusion

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Microalgae: A Powerful Tool for Climate Change and Water…

Specific Sources of CO2 Emissions

CO2: A major Green House Gas

Biological CO2 Sequestration Technology/Method

Algae potential players for CO2 Sequestration.

Algae cultivation in Wastewater an alternative approach for wastewater treatment & CO2 Sequestration.

Mechanism of Microalgae-based Wastewater Treatment

Open ponds

Closed reactors

How Algae CO2 Sequestration Potential is Calculated?

CO2 concentration and CO2 uptake efficiency of Microalgae

Enhancing CO2 Mass Transfer with Microbubbles

Specific Sources of CO₂ Emissions

CO₂: A major Green House Gas

Biological CO₂ Sequestration Technology/Method

Algae potential players for CO₂ Sequestration.

Algae cultivation in Wastewater an alternative approach for wastewater treatment & CO₂Sequestration.

How Algae CO₂Sequestration Potential is Calculated?

CO₂ concentration and CO₂ uptake efficiency of Microalgae

Enhancing CO₂Mass Transfer with Microbubbles