New insights of inoculating thermo-tolerant CIP-degrading microbiota for promoting livestock manure compost humification under CIP stress: from biological and abiotic perspectives

Antibiotics containing primary amino groups are known to form humic acid-like substances (HALs) during composting. However, for antibiotics that generate humic acid (HA) precursors via bio-degradation, it remains unclear whether they can directly contribute to HA formation in compost and how inoculating microorganisms capable of degrading such antibiotics influences humus formation. This study focused on ciprofloxacin (CIP), a representative quinolone antibiotic. Here, we isolated thermotolerant CIP-degrading microbiota (TCM), primarily composed of Enterococcus and Aspergillus , and combined them with psychrotrophic cellulose-degrading fungi (PCDFs) to enhance humification under CIP stress in cold environments. Using maize straw and pig manure as substrates, TCM inoculation achieved 70.6 % CIP removal and significantly accelerated humification through dual biotic-abiotic pathways. Key findings include the synthesis of HALs via the polyphenol-protein pathway, driven by CIP-derived primary amine compounds (PACs) and quinones. Spectroscopic results demonstrated that TCM enhanced the aromaticity and structural complexity of humic substances (HAs), as evidenced by a 7.56 % reduction in the E4/E6 ratio and increased FTIR absorption at 1641 cm −1 . Metagenomic analysis revealed that TCM enriched functional genera ( Jeotgalicoccus , Halopseudomonas , and Caldibacillus ) and upregulated pathways associated with HA precursor synthesis. This study introduces a dual strategy integrating biodegradation and abiotic humification to transform CIP into stable humus, providing a novel “waste-to-resource” approach for managing antibiotic-contaminated manure in cold regions. The results offer critical microbial resources and mechanistic insights for advancing sustainable agricultural waste management.