# Estuaries of the Tinto, Odiel and Piedras rivers as source of new species of Pseudomonas with biofertilizer potential under stress conditions

## TL;DR

This study identifies new Pseudomonas species from polluted estuaries that can boost plant growth under stress, offering potential for sustainable agriculture.

## Contribution

The discovery and characterization of three new Pseudomonas species with biofertilizer potential from metal-contaminated estuaries.

## Key findings

- Three new Pseudomonas species were identified using phylogenomic and genomic analyses.
- The new species showed plant growth-promoting traits and enhanced legume biomass and nitrogen content in greenhouse tests.
- The N8T strain improved shoot, root biomass, and nodule numbers when co-inoculated with rhizobia.

## Abstract

The estuaries of the Tinto, Odiel, and Piedras rivers have been extensively studied by several groups due to ecological aspects such as heavy metal contamination, nutrient deficiency and degradation influenced by climate change. Aimed to recover these ecosystems, a large collection of plant growth-promoting bacteria (PGPB), several of them previously identified as new species, was isolated from the rhizosphere and tissues of native plants. In this study, we analyse the strains N4, N8T, L1T, and SDT3T as potential new species. A phylogenomic analysis revealed that four strains belong to the genus Pseudomonas, forming distinct clades with full support. Digital DNA-DNA hybridization and average nucleotide identity values were below the species delineation thresholds (70% and 95–96%), confirming the novelty of strains N8T, L1T, and SDT3T, while N4 and N8T were identified as the same species. As a result, we propose the names Pseudomonas medicaginis sp. nov., Pseudomonas onubensis sp. nov., and Pseudomonas spartinae sp. nov., with P. medicaginis comprising two strains. These strains, previously identified as PGPB, harboured genes related to PGP traits and were further characterized phenotypically and biochemically. To evaluate the agroecological potential of N8T strain, it was tested in co-inoculation with the corresponding rhizobia on lentil, pea, alfalfa, and bean under greenhouse conditions. In all legumes, N8T enhanced shoot and root biomass, increased nodule numbers, and improved nitrogen content compared with single rhizobia inoculation. In summary, these estuarine ecosystems are not only ecologically significant for their microbial diversity but also serve as reservoirs of beneficial bacteria with promising agroecological applications.

The online version contains supplementary material available at 10.1038/s41598-025-31256-y.