Marine antifouling biocide 4,5-dichloro-2-n-octyl-4-isothiazolin-3-one disrupts sediment microbiome structure and function: insights from absolute quantification and enzyme activity dynamics

ABSTRACT The organic booster biocide DCOIT (4,5-dichloro-2-n-octyl-4-isothiazolin-3-one) is widely used in marine antifouling systems, yet its ecological impacts on sediment microbiomes remain poorly understood. Here, we integrated absolute quantitative 16S rRNA gene sequencing, metagenomics, and enzyme activity assays to examine microbial responses to DCOIT exposure (0–50 μg/g sediment) over 30 days. DCOIT induced oxidative stress and bioenergetic impairment, accompanied by reduced microbial activity and inhibition of key enzyme-mediated processes involved in organic matter turnover and nitrogen transformation. Absolute quantification revealed a compensatory increase in total microbial abundance by Day 30, despite persistent diversity loss and community restructuring. Metagenomic analysis showed that DCOIT disturbed functional potentials related to carbon and nitrogen cycling. Kordiimonas , Aliikangiella , and Neptuniibacter emerged as potential contributors to nitrogen transformation, whereas Marinobacter was more closely associated with potential DCOIT transformation. DCOIT exposure also enriched adaptive traits, including chemotaxis, motility, quorum sensing, and biofilm regulation, and was accompanied by increased multidrug efflux systems and heavy metal resistance determinants. Our findings provide novel insights into the ecotoxicological risks of isothiazolinone biocides and highlight the potential for DCOIT to undermine sediment ecosystem functions and microbial habitat health. Given its extensive application, this study emphasizes the need to consider the microbial ecological consequences of DCOIT accumulation in seafloor environments. IMPORTANCE DCOIT is widely used in marine antifouling coatings and can accumulate in benthic sediments, yet its effects on sediment microbiomes remain poorly defined. This study shows that DCOIT disrupts microbial energy status, enzyme activities, community structure, and nitrogen-cycling functions while selecting for adaptive traits and resistance-related determinants. By integrating absolute quantification, metagenomics, and enzyme assays, our work demonstrates that DCOIT poses microbial ecological risks beyond toxicity to macroorganisms and should be considered in assessments of antifouling biocides.