Implications for assessing sludge hygienization: Differential responses of the bacterial community, human pathogenic bacteria, and fecal indicator bacteria to sludge pretreatment-anaerobic digestion

Sewage sludge is the byproduct of wastewater treatment plants, which host enormous diversity of microbes including potential pathogens. However, there are still challenges in assessing hygienization during sludge stabilization due to the complex relationships between dominant microbes and human pathogenic bacteria (HPB), and the accuracy of fecal indicator bacteria (FIB) is also disputed. Here, the responses of the bacterial community, HPB, and FIB to sludge pretreatment-anaerobic digestion (AD) were comprehensively compared using culture-based and 16S rRNA gene molecular analysis methodologies. Bacterial and HPB communities differed in response to sludge pretreatment-AD. AD drove the variation of bacterial community, but led to the convergence of HPB communities in pretreated sludge, indicating the existence of ecological niches that favors HPB dissemination in digesters. The correlation analysis indicated that FIB was suitable for characterizing general pathogen removal instead of showing the real pattern of HPB (i.e., each HPB), implying the need for comprehensive assessment approaches. Moreover, AD-related parameters including pH, total solids destruction, and methane yield were found to play important role in assessing pathogen inactivation given their correlation. This work provides theoretical basis for the selection of appropriate sludge stabilization approaches and future supervision of biosolids biosafety, which finally benefits human health.