Bioflocculant-Producing Bacteria from Complex Waste Streams: A Sustainable Approach to Microbial Flocculation, EPS Characterization, and Sludge Minimization for Industrial Effluent Remediation

The global pursuit of sustainable and eco-friendly alternatives to chemical coagulants has intensified interest in microbial bioflocculants for wastewater treatment applications. In this study, 63 indigenous bacterial isolates were obtained from diverse environmental matrices in Gujarat, India, including municipal solid waste sites, pharmaceutical effluent treatment plants (ETPs), aeration tanks, sedimentation tanks, and activated sludge systems. Detailed morphological, biochemical, and gram-staining analysis revealed significant taxonomic and phenotypic diversity among the isolates, representing both gram-positive and gram-negative bacteria with distinct colony traits, pigmentation, and cellular morphologies. Isolation of bioflocculant-producing strains was carried out through serial dilution, plating on nutrient agar and luria bertani agar, and subsequent morphological differentiation based on colony texture, EPS production, and mucoid appearance. Preliminary screening, based on ropy colony formation, viscous loop tests, and visual EPS assessment, shortlisted promising bioflocculant producers. Optimized cultivation protocols were implemented, including incubation for 24 to 48 hours followed by sedimentation periods of 5 to 20 minutes using kaolin clay suspensions to evaluate flocculation activity. Isolates ASW5, ASW9, SED11, A13, and S14 exhibited exceptional flocculating activity exceeding 90%, highlighting their potential for efficient suspended solids removal, turbidity reduction, and sludge minimization in wastewater treatment processes. Further characterization included viable biomass quantification (CFU/mL), EPS production assessment, and sludge volume index (SVI) determination. Comparative evaluations using conventional chemical coagulants such as ferric chloride (FeCl₃), ferrous sulfate (FeSO₄), and commercial oxyfloc formulations demonstrated competitive or superior performance of select bacterial isolates, particularly in enhancing sludge settling efficiency, reducing sludge retention time, and lowering SVI values. Advanced statistical correlation matrix analyses identified significant associations between flocculating efficiency, colony morphology, eps production, and sedimentation behavior. Source-wise isolate distribution, taxonomic affiliations, and morphological diversity were visualized using Sankey diagrams, providing a comprehensive ecological overview of microbial communities associated with engineered wastewater treatment systems. This study demonstrates the potential of native bacterial strains as efficient, eco-friendly bioflocculant producers, offering sustainable, cost-effective alternatives to chemical coagulants. The work advances microbial biotechnology for enhanced wastewater clarification, sludge management, and effluent quality improvement, supporting global efforts toward circular bioeconomy strategies and green wastewater treatment technologies.