(176ap) Characterization of Microbiome Dynamics Using Flow Cytometry for Napier Grass Anaerobic Digestion

Pennisetum purpureum, commonly known as Napier grass, has great potential as feedstock for the production of biogas. In anaerobic digestion of Napier grass, the microbiome plays a key role, which is crucial for effective biomethane production. Conventional methods used previously for microbiome dynamics studies were labor-intensive and time-consuming. In this study, flow cytometry was used as a robust, high-throughput approach for the rapid characterization of microbiome dynamics in anaerobic digestion of Napier grass. Flow cytometry allows for the effective monitoring of variations in microbial community structure in response to environmental perturbations such as pH changes, soluble chemical oxygen demand, and volatile fatty acid production. This approach provides immediate insight into the dynamics of microbial communities by utilizing the comprehensive multidimensional data acquired through flow cytometry. This multiparametric technique allows for real-time monitoring and optimization of anaerobic digestion processes, ultimately increasing biogas generation efficiency from Napier grass. This study aims to analyze microbiome dynamics using flow cytometry and correlate the flow cytometric data with the 16S rRNA gene sequencing data for Napier grass anaerobic digestion. In this study, Napier grass was given with chemical and biological pretreatment that might help in better hydrolysis and biomethane generation. 1 and 2 % sodium hydroxide and calcium carbonate solutions were used for chemical pretreatment. Two fungal strains, Trichoderma longibrachiatum and Pycnoporous sanguineus, were used for biological pretreatment. After 60 days of digestion, Napier grass digested with 2% sodium hydroxide showed the highest methane production, i.e., 266.91 ± 10.71 mL/g VS added. The metagenomics analysis showed relatively high archaeal abundance of Methanothrix, Methanobacterium, and Methanosarcina in 2% NaOH pretreated Napier grass compared to the other samples. The statistical analysis of different flow cytometric populations showed a positive correlation with biogas and methane production, improving the understanding of the microbial dynamics in anaerobic digestion systems. By leveraging the abundant data obtained from flow cytometric analysis, this technique helps to increase biogas production from renewable biomass sources like Napier grass.