Coastal microbial biofilms constitute an important biogenic interface controlling trace metal retention, transformation and biological utilization in marine environments, yet their response to anthropogenic metal inputs coupled with increasing oceanic acidification (OA) remain poorly understood. As declining seawater pH alters metal speciation and bioavailability, understanding the mechanisms regulating biological metal sequestration is essential for predicting future trace metal cycling.

In this study, we investigated the combined effects of reduced pH (7.4) and environmentally relevant Pb exposure (20 ppb) on Pb partitioning in intertidal microphytobenthic biofilms grown in filtered, autoclaved seawater in a continuously stirred tank reactor (50rpm, 14:10 light: dark photoperiod). Time-series Pb partitioning between seawater and biofilms was coupled with elemental concentration analysis, physicochemical monitoring, biochemical and spectroscopic characterisation of biofilm and extracellular polysaccharide (EPS) and genome-resolved metagenomics to resolve the mechanisms governing Pb sequestration under OA. On exposure to reduced pH, dissolved Pb progressively declined (67% removal by Day 14) as Pb accumulated within biofilm, where the EPS matrix constituted the dominant Pb sink. Pb-acidified biofilms exhibited reduced chlorophyll content, photosynthetic oxygen production and cell viability accompanied by elevated oxidative stress. Substantial increase in EPS production was observed, particularly carbohydrate and uronic acid-rich fractions with intensified carboxylate and glycosidic FTIR signatures indicating enrichment of high-affinity Pb-binding ligands. Simultaneous depletion of phosphate in the seawater indicated its enhanced utilization in Pb sequestration by the biofilm. Genome-resolved metagenomics revealed pronounced community restructuring, with Cyanobacteria becoming the dominant phylum under combined OA and pH stress along with increased abundance of genes involved in EPS biosynthesis, heavy-metal transport, oxidative stress defence and metallothionein production.
Integrating these biogeochemical observations, a hierarchical four-tier detoxification mechanism was proposed: (i) rapid Pb immobilization through EPS ion-exchange, (ii) long-term stabilization via uronic-acid rich carboxylate complexation in the EPS, (iii) intracellular sequestration through polyphosphate binding granule formation and extracellular Pb-phosphate-mineralization, (iv) terminal cytosolic detoxification by metallothionein. These findings demonstrate that OA fundamentally reshapes biogenic Pb sequestration by promoting coordinated extracellular and intracellular detoxification processes in Cyanobacteria-enriched coastal biofilms. This study helps to establish a mechanistic link between climate-driven acidification, microbial adaptation and Pb biogeochemistry providing process-level insights into trace-metal cycling under future ocean conditions.