Benefits and costs of a hypercapsule and the mechanism of its loss in a clinical isolate of Acinetobacter baumannii

Abstract

Acinetobacter baumannii is an opportunistic pathogen in which capsule production is closely linked to immune evasion and environmental persistence. Recent studies have described two seemingly contradictory phenomena--increasing prevalence of capsule-overproducing clinical isolates and frequent isolation of capsule-deficient variants. The biological significance of these phenomena remains unclear. In this study, we analyzed a clinical isolate, OCU_Ac16b, which spontaneously gives rise to two phenotypically distinct variants: the L type forming large colonies with an extremely thick capsule, and the S type forming small colonies with a substantially reduced or absent capsule. When L-type cells were cultured in test tubes under low-shaking conditions, S-type variants reproducibly emerged, constituting 45-80% of the population within 24 hours. Whole-genome sequencing revealed that this conversion is driven by distinct mutations in the capsular polysaccharide synthesis cluster, including insertion sequence (IS) insertions and a single-nucleotide deletion. PCR analysis confirmed that IS insertions in wzy (polysaccharide polymerase) occur within individual L-type colonies prior to liquid culture, providing a likely explanation for the rapid and reproducible emergence of S-type variants. Phenotypic characterization demonstrated a biological trade-off, with L-type cells exhibiting enhanced resistance to serum killing, desiccation, and certain {beta}-lactam antibiotics, whereas S-type cells showed superior surface attachment, increased biofilm formation, and a growth advantage under oxygen-limited conditions. Our findings uncover a highly reproducible, mutation-driven capsule switching mechanism that enables rapid phenotypic adaptation to changing environments. This phenotypic heterogeneity has significant implications for pathogenesis, persistence, diagnostic evaluation, and clinical management. ImportanceAcinetobacter baumannii is a clinically important opportunistic pathogen that exhibits striking phenotypic diversity. In particular, some clinical isolates produce unusually thick capsules, which are thought to contribute to immune evasion and persistence, while others lack capsule altogether. However, the biological significance of these contrasting phenotypes has remained unclear. In this study, we analyzed a clinical isolate that spontaneously gives rise to capsule-deficient variants from a hypercapsulated form. We found that the conversion is driven by spontaneous mutations in capsule biosynthesis genes that arise naturally within colonies, while the expansion of capsule-deficient cells is promoted under oxygen-limited conditions. The two variants differed in serum resistance, desiccation tolerance, growth characteristics, and antibiotic responses, revealing a trade-off between protective barriers and environmental adaptability. These findings provide new insights into how A. baumannii balances survival strategies through genetic and phenotypic heterogeneity, with important implications for diagnosis, treatment, and bacterial persistence in clinical settings.