Anaerobic digestion (AD) is a biological process occurring in the absence of oxygen, where microorganisms degrade organic matter to produce biogas, a mixture of methane (CH4), carbon dioxide (CO2), and trace gases like hydrogen sulfide (H2S) and ammonia (NH3). In assessing AD of RAS sludge, understanding how aquacultural sludge characteristics affect the digestion process and the microbial community inside the digester will determine the feasibility of treatment. The breakdown of abundant nitrogen-rich organic compounds, such as proteins, in fish sludge leads to excess ammonia formation inside an AD system. Ammonia can exist in two forms (ammonium ion, or NH4+, and free ammonia, or NH3) inside the digester, depending on the pH of the sludge. As the pH increases, ammonium ions are rapidly converted to toxic free ammonia, which might destabilize the AD process and inhibit desirable CH4 production.
In this study, RAS sludge from a Steelhead trout grow-out tank was mixed with deionized water varying the initial concentration of total nitrogen (five treatments). Subsequently, the sludge was mixed with an inoculum source in lab-scale batch reactors. The ratio of microorganisms to organic matter (inoculum-to-substrate ratio or ISR) was kept constant in each treatment. The results showed an increase in the lag phase period for CH4 production as the concentration of total N increased in the treatments. However, at the end of the study, the cumulative CH4 production values were similar for most treatments, signifying that the microbial consortia may have acclimated to the high ammonia conditions. While not significantly different, it was noted that the mixture with 40% sludge and 60% deionized water resulted in the highest normalized cumulative CH4 production (397 ± 3 mL CH4/g VS).
Microbial analysis of the final digested samples was also conducted using high-throughput sequencing (HTS) to understand the acclimation process of the microbial community. The results showed that the increase in the relative abundance of the ammonia-tolerant genus Methanosarcina and a simultaneous decrease in the ammonia-sensitive genus Methanosaeta corresponded with the increase in the initial total nitrogen concentration of the treatments. Due to the increase in the ammonia concentrations, the methanogenic archaeal community gradually shifted towards genera that were more resistant to high ammonia concentrations, which may have resulted in the extended lag phase observed for the treatments with high total nitrogen concentrations.