Growth of Candidate Phyla Radiation bacteria in groundwater incubations reveals widespread adaptations to oxic conditions

Background : The Candidate Phyla Radiation (CPR) comprises a widespread but poorly understood group of bacteria with limited cultured representatives, largely due to their metabolic dependencies on microbial hosts. In laboratory incubations, CPR often decline sharply in relative abundance, even when samples originate from natural environments where they dominate, such as groundwater, where they can represent over 50% of the microbiome. Suitable enrichment conditions and host interactions remain poorly defined. Results : Here, we analyzed 16S rRNA gene amplicon data from 397 groundwater incubation samples across 31 treatments, including 22 under oxic conditions, to identify factors that promote CPR survival and growth. Despite an initial decline, CPR abundances recovered over longer incubation times, reaching up to 11-30% of the microbial community. In total, we detected 1,410 CPR amplicon sequence variants (ASVs), spanning six major CPR classes commonly found in groundwater. Enrichment success was treatment-specific: Cand. Saccharimonadia dominated in incubations with polysaccharides (up to 31.4%), while Cand. Parcubacteria were enriched (>23%) in treatments stimulating methylotrophs and autotrophs. ASV-specific growth rates based on quantitative PCR showed that some CPR doubled within 1-2 days, comparable to faster-growing non-CPR groundwater bacteria, while most CPR had doubling times around 15 days. Strikingly, CPR reached higher absolute abundances under oxic conditions than under anoxic conditions. Metabolic network analysis based on metagenome-assembled genomes revealed that up to 62% of annotated genes were associated with functions linked to oxic conditions. In fact, 25 CPR genomes encoded enzymes that directly utilize oxygen, challenging the long-standing view of CPR as strictly anaerobic, fermentative organisms. Conclusions : Our findings demonstrate that diverse CPR lineages not only survive but actively grow in groundwater incubations, even under oxic conditions. The discovery of genes for oxygen-dependent reactions and substantial CPR enrichment in oxic treatments reveals unexpected metabolic flexibility, helping to explain their persistence and ecological success across a wide range of environments.