# Reducing the sensitivity of Halomonas sp. to oxygen availability through adaptive laboratory evolution

## TL;DR

Scientists evolved a salt-loving bacterium to grow better in low-oxygen conditions, improving its usefulness for industrial processes.

## Contribution

A new framework for reducing microbial sensitivity to oxygen through adaptive evolution is proposed.

## Key findings

- Adaptive evolution improved Halomonas sp. growth robustness by 62% in micro-aerobic environments.
- PHB production yield increased by 21%, while ectoine production decreased by 79%.
- Genetic mutations in transport and enzymatic pathways were linked to improved oxygen tolerance.

## Abstract

Halophiles attract increasing attention to serve as sustainable industrial hosts for prolonged continuous processes or in open-vat fermentations owing to the reduced risk of contamination enabled by high salt concentration in the medium. Despite growing interest, their application in large-scale manufacturing remains limited partly due to bioprocessing challenges. A robust host that performs consistently well across scales should withstand variations in oxygen availability since local hypoxic regions often manifest in large-scale tanks even under strict operational control. In this work, we modified Halomonas sp. to achieve robust growth profiles in micro-aerobic environments by gradually exposing a continuous culture to stress induced by reduced oxygen availability using adaptive laboratory evolution. Nominal contamination was observed at 8.5% salt concentration even during non-aseptic operation, offering a suitable environment for prolonged continuous cultivation. Following adaptive laboratory evolution, Halomonas sp. achieved comparable growth at low-to-moderate (0.5%-3.0%) and moderate-to-high (3.0–5.0%) dissolved oxygen availability (OD600 = 4.46 ± 0.29 & µ = 0.40 h− 1 and 4.96 ± 0.81 & µ = 0.35 h− 1, respectively). Adaptive evolution improved the growth robustness of Halomonas sp. by 62% and its PHB production yield by 21%, while reducing ectoine production by 79% indicting improved metabolic perception of oxygen availability and reduced stress response. Mutations in cellular transport mechanisms and enzymatic pathways enabled this response as indicated by whole genome sequencing. The proposed framework is an effective strategy to reduce the oxygen sensitivity of microbial strains in general to improve their physiological properties, and to genetically design populations suitable for large scale bioprocess operations.

The online version contains supplementary material available at 10.1186/s12934-026-02951-w.