bioRxiv Science⌕ Search
Home/Archive/bioRxiv

bioRxiv · 10.1101/2025.05.13.653867

Co-infection with Streptococcus and Rothia spp. drives prophage dynamics in Pseudomonas aeruginosa in an artificial sputum model

Wright, L. L.·Cazares, A.·Figueroa, W.·Chatterley, L. C.·Willmott, T. O.·O'Brien, S.·Fothergill, J.
View original source ↗Export metadata ↓

Abstract

ObjectivesChronic bacterial lung infections with Pseudomonas aeruginosa are common in people with cystic fibrosis (CF) but interactions with other commonly seen members of the CF lung community, such as Streptococcus and Rothia spp., are not well understood. The aim of this study was to determine the impact of other species on P. aeruginosa using an artificial sputum medium (ASM) model designed to mimic the conditions in the CF lung. MethodsP. aeruginosa LESB58, representing a CF epidemic strain, was inoculated into ASM either alone or in co-culture with Streptococcus anginosus, Streptococcus vestibularis, Rothia mucilaginosa or Rothia dentocariosa. Gene expression was assessed using RNAseq at 72 and 96 h timepoints. Free phage numbers were assessed by plaque assay and qPCR. ResultsDifferential expression of P. aeruginosa genes, including those related to virulence factors such as prophage, type III secretion, exopolysaccharides and pyochelin production, was seen in co-culture conditions compared with single-species culture. In particular, at 72 h, there was lower expression of most LES prophage genes under co-culture conditions, and this corresponded with lower overall free phage at the same timepoint. Specifically, during co culture, there was a reduction in the levels of a D3112-like transposable phage, previously shown to have the ability to drive CF-like adaptations in an experimental evolution model, and an increase in levels of Pf1-like filamentous phage, associated with altered biofilm formation and immune responses. ConclusionGene expression changes during co-culture with Streptococcus and Rothia spp. were identified affecting key P. aeruginosa virulence factors including phage. Further work indicated changes to free phage levels of two important P. aeruginosa LESB58 prophage with the potential to affect P. aeruginosa adaptation, antibiotic resistance and host immune response in the CF lung. This study emphasises the importance of understanding the influence of lung commensals on focal pathogens in a complex microbiome.

Source connections

Explore related subjects

microbiologyPreprint

Keep this discovery

Explore connections, maps & timelines

BibTeXRIS

Wright, L. L., Cazares, A., Figueroa, W., Chatterley, L. C., Willmott, T. O., O'Brien, S., Fothergill, J.. 2025-05-13. Co-infection with Streptococcus and Rothia spp. drives prophage dynamics in Pseudomonas aeruginosa in an artificial sputum model. https://doi.org/10.1101/2025.05.13.653867

Cite the original work for its findings. Save a collection to share your selection of sources.

KEEP EXPLORING

Related preprints

bioRxiv2026

Reverse Transcription Loop-Mediated Isothermal Amplification Combined with Lateral Flow Assay (RT-LAMP-LFA) for Rapid Detection of the 2026 Bundibugyo Virus (BDBV) Outbreak Strain

Making accurate diagnosis is a critical first step in patient management and control for emerging and re-emerging disease outbreaks. Bundibugyo virus (BDBV) is a filovirus that causes Ebola disease, and the 2026 outbreak highlights the need for rapid diagnostic approaches that can be deployed in settings with limited laboratory infrastructure. Here, we developed a reverse transcription loop-mediated isothermal amplification-lateral flow assay (RT-LAMP-LFA) for rapid detection of the 2026 BDBV outbreak strain. LAMP primers were designed based on the genome sequence of the 2026 outbreak strain, and the conservation of the primer binding sites among all available 2026 outbreak genomes was confirmed in silico. LAMP primers were evaluated for analytical specificity and sensitivity using in vitro-transcribed RNA (ivtRNA) from BDBV and other filoviruses, as well as RNA derived from arboviruses and mammalian cells. Amplification products were detected using a Chromatography-Printed Array Strip (C-PAS) lateral flow platform. The newly designed primers specifically detected the presence of BDBV RNA without nonspecific amplification of the other viral or cellular RNA samples tested. Consistent amplification was observed down to 103 copies per reaction. In addition, BDBV viral RNA was successfully amplified, and the amplification products were detected using C-PAS, with the total workflow being completed within 25 min. These findings demonstrate the analytical specificity and sensitivity of the developed RT-LAMP-LFA assay and its compatibility with rapid visual lateral flow detection, supporting its potential application as a point-of-care approach for BDBV detection.

microbiology↗
bioRxiv2026

Divergent methane production and microbial community responses to carbon enrichment in freshwater wetland sediments

Methane (CH4) accumulation in wetland sediments reflects interactions among organic matter, sediment chemistry, and microbial communities. We combined depth-resolved field characterization across four wetland habitats in northern Israel with complementary incubations to examine methane accumulation and responses to labile-carbon enrichment. Sediment cores were characterized using Rock-Eval analysis, porewater chemistry, oxygen profiling at three sites, and mcrA amplicon sequencing. High CH4 concentrations occurred under contrasting conditions: Ein Nimfit (EN) contained approximately 12% total organic carbon and reached 2.1 mM CH4, whereas Ein Afek site EA2 contained approximately 5% carbon and accumulated 0.7-1.7 mM CH4. Both showed sulfate depletion, but EN exhibited pronounced sulfide accumulation, whereas EA2 showed greater dissolved iron and nutrient enrichment. Their classified methanogenic assemblages also differed. EA1 remained methane poor despite increasing carbon content with depth, while EA3 combined low carbon content and CH4 accumulation with detectable methanogens and lower methanogenic diversity. In a 36-day bottle experiments using sediments from two of the four wetland habitats, sucrose-amended EA3 exhibited a transient maximum of 18.8 mmol collected CH4 kg-1 wet sediment during Days 22-29, whereas EN reached its highest collected amount during Days 29-36. Methane proportions increased before the large output increase in EA3 but remained comparatively stable and high in EN. Soluble chemical oxygen demand trajectories and microbial community changes also differed. These findings identify distinct biogeochemical settings supporting methane accumulation and demonstrate methanogenic potential in low-CH4 sediment, emphasizing the complementary information provided by field profiles, microbial characterization, and exploratory enrichment experiments.

microbiology↗
bioRxiv2026

The absence of microbiome in germ-free mice exacerbates the pathogenicity of mouse-adapted SARS-CoV-2 MA10

Germ-free mice, which are devoid of microbiota, provide a powerful system to define the contribution of the microbiome to host responses during infection. Severe COVID-19 outcomes are associated with multiple comorbidities that are themselves characterized by microbiome dysbiosis; however, the causal role of the microbiota in SARS-CoV-2 pathogenesis remains poorly understood. Here, we use germ-free mouse models to evaluate the impact of the microbiome on disease severity following infection with mouse-adapted SARS-CoV-2 MA10. Comparison with conventionally housed mice revealed that the absence of the microbiome markedly exacerbates SARS-CoV-2-induced disease in both C57BL/6 and Swiss Webster backgrounds. Germ-free mice exhibited greater weight loss, impaired lung function, worsened pulmonary pathology, and pronounced alterations in circulating cytokine and chemokine profiles compared with conventional controls. These findings show that resident microbiota limit SARS-CoV-2 pathogenicity and establish germ-free COVID-19 models as a powerful platform to identify protective microbial taxa and microbiota-based interventions to reduce severe disease.

microbiology↗