Non-antibiotic disinfectant synchronously interferes methane production and antibiotic resistance genes propagation during sludge anaerobic digestion: Activation of microbial adaptation and reconfiguration of bacteria-archaea synergies

Abstract

Waste activated sludge (WAS) presents both resource recovery potential and pollution risks, making its efficient treatment challenging. Anaerobic digestion is broadly recognized as a green and sustainable approach to WAS treatment, whose efficiency is easily impacted by the exogeneous pollutants in WAS. However, the impact of polyhexamethylene guanidine (PHMG), as a widely-used non-antibiotic disinfectant, on WAS digestion under semi-continuous flow conditions remains unclear. In this study, CH₄ production decreased from 16.1 mL/g volatile suspended solids (VSS) in the control to 13.2 mL/g VSS and 0.3 mL/g VSS under low and high PHMG exposure, respectively, while PHMG increased the number of antibiotic resistance gene (ARG) copies per bacterium by 4.6–12.7 %. Molecular docking analysis revealed that PHMG could spontaneously bind to and disintegrate WAS (binding energy:2.35 and -9.62 kcal/mol), increasing the likelihood of microbial exposure to PHMG. This led to an increase in bacterial abundance and a reduction in archaeal populations, resulting in bacterial dominance in ecological niches. The network topology index in PHMG-treated reactors was consistently lower than in the control, with a higher proportion of negatively correlated links, indicating a more antagonistic relationship between bacteria and archaea. Consequently, PHMG significantly interfered with key genes involved in CH₄ biosynthesis (e.g., mch and mtd). Interestingly, methanogenic activity and archaeal chemotaxis (e.g., rfk and cheA) partially recovered under low PHMG exposure due to archaeal adaptation through quorum sensing and two-component systems. However, this adaptation process also contributed to the propagation of ARGs through horizontal gene transfer, facilitated by the enhancement of mobile genetic elements and ARGs hosts. These findings confirm the ecological risks of PHMG and highlight the need for effective WAS disposal strategies.