Staphylococcus epidermidis bacteriocin A37 kills natural competitors with a unique mechanism of action.

Many bacteria produce antimicrobial compounds such as lantibiotics to gain advantage in the competitive natural environments of microbiomes. Epilancins constitute an until now underexplored family of lantibiotics with an unknown ecological role and unresolved mode of action. We discovered production of an epilancin in the nasal isolate Staphylococcus epidermidis A37. Using bioinformatic tools, we found that epilancins are frequently encoded within staphylococcal genomes, highlighting their ecological relevance. We demonstrate that production of epilancin A37 contributes to Staphylococcus epidermidis competition specifically against natural corynebacterial competitors. Combining microbiological approaches with quantitative in vivo and in vitro fluorescence microscopy and cryo-electron tomography, we show that A37 enters the corynebacterial cytoplasm through a partially transmembrane-potential-driven uptake without impairing the cell membrane function. Upon intracellular aggregation, A37 induces the formation of intracellular membrane vesicles, which are heavily loaded with the compound and are essential for the antibacterial activity of the epilancin. Our work sheds light on the ecological role of epilancins for staphylococci mediated by a mode of action previously unknown for lantibiotics.

Genome-scale metabolic models consistently predict in vitro characteristics of Corynebacterium striatum.

Introduction: Genome-scale metabolic models (GEMs) are organism-specific knowledge bases which can be used to unravel pathogenicity or improve production of specific metabolites in biotechnology applications. However, the validity of predictions for bacterial proliferation in in vitro settings is hardly investigated. Methods: The present work combines in silico and in vitro approaches to create and curate strain-specific genome-scale metabolic models of Corynebacterium striatum. Results: We introduce five newly created strain-specific genome-scale metabolic models (GEMs) of high quality, satisfying all contemporary standards and requirements. All these models have been benchmarked using the community standard test suite Metabolic Model Testing (MEMOTE) and were validated by laboratory experiments. For the curation of those models, the software infrastructure refineGEMs was developed to work on these models in parallel and to comply with the quality standards for GEMs. The model predictions were confirmed by experimental data and a new comparison metric based on the doubling time was developed to quantify bacterial growth. Discussion: Future modeling projects can rely on the proposed software, which is independent of specific environmental conditions. The validation approach based on the growth rate calculation is now accessible and closely aligned with biological questions. The curated models are freely available via BioModels and a GitHub repository and can be used. The open-source software refineGEMs is available from https://github.com/draeger-lab/refinegems.

Sampling of Human Microbiomes to Screen for Antibiotic-Producing Commensals.

Soil-derived microorganisms have been sampled intensively throughout the last decades in order to discover bacterial strains that produce new antibiotics. The increasing emergence of multidrug-resistant bacteria and the constant high demand for new antibiotic classes are leading to the sampling and investigation of new microbiomes that contain antimicrobial producers. Human-associated microbiomes are therefore gaining more and more attention. This chapter presents a detailed description of how human microbiomes can be sampled and how microbiota members from skin and nasal samples can be isolated. Different methods for antimicrobial compound screening are presented.

Mixed Populations and Co-Infection: Pseudomonas aeruginosa and Staphylococcus aureus.

The human pathogens Pseudomonas aeruginosa and Staphylococcus aureus are frequently co-isolated from chronic wounds or cystic fibrosis patient airways. Clinical studies analysing the impact of co-infection on patient clinical outcomes lead to contradictory results. However, laboratory approaches suggest that the two pathogens co-colonize the same infection niches and form a mixed-species biofilm, therefore favouring their resistance to antibiotics and immune response. In parallel, many recent studies have focused on the different interactions between the two bacterial species. It has long been recognized that P. aeruginosa usually outcompetes S. aureus, and the molecular mechanisms involved in this state of bacterial competition are now well understood. However, several recent studies show that interactions between P. aeruginosa and S. aureus can be diverse and evolve over time. Thus, many CF isolates of P. aeruginosa and S. aureus can coexist and develop cooperative behaviours. In this chapter, we will provide an overview of the current knowledge on the mixed populations of P. aeruginosa and S. aureus, from their mechanisms of establishment to their impacts on bacterial physiology and clinical outcomes.

How Bacterial Adaptation to Cystic Fibrosis Environment Shapes Interactions Between Pseudomonas aeruginosa and Staphylococcus aureus.

Pseudomonas aeruginosa and Staphylococcus aureus are the two most prevalent bacteria species in the lungs of cystic fibrosis (CF) patients and are associated with poor clinical outcomes. Co-infection by the two species is a frequent situation that promotes their interaction. The ability of P. aeruginosa to outperform S. aureus has been widely described, and this competitive interaction was, for a long time, the only one considered. More recently, several studies have described that the two species are able to coexist. This change in relationship is linked to the evolution of bacterial strains in the lungs. This review attempts to decipher how bacterial adaptation to the CF environment can induce a change in the type of interaction and promote coexisting interaction between P. aeruginosa and S. aureus. The impact of coexistence on the establishment and maintenance of a chronic infection will also be presented, by considering the latest research on the subject.

From genotype to phenotype: adaptations of Pseudomonas aeruginosa to the cystic fibrosis environment.

Pseudomonas aeruginosa is one of the main microbial species colonizing the lungs of cystic fibrosis patients and is responsible for the decline in respiratory function. Despite the hostile pulmonary environment, P. aeruginosa is able to establish chronic infections thanks to its strong adaptive capacity. Various longitudinal studies have attempted to compare the strains of early infection with the adapted strains of chronic infection. Thanks to new '-omics' techniques, convergent genetic mutations, as well as transcriptomic and proteomic dysregulations have been identified. As a consequence of this evolution, the adapted strains of P. aeruginosa have particular phenotypes that promote persistent infection.

Trophic cooperation promotes bacterial survival of Staphylococcus aureus and Pseudomonas aeruginosa.

In the context of infection, Pseudomonas aeruginosa and Staphylococcus aureus are frequently co-isolated, particularly in cystic fibrosis (CF) patients. Within lungs, the two pathogens exhibit a range of competitive and coexisting interactions. In the present study, we explored the impact of S. aureus on the physiology of P. aeruginosa in the context of coexistence. Transcriptomic analyses showed that S. aureus significantly and specifically affects the expression of numerous genes involved in P. aeruginosa carbon and amino acid metabolism. In particular, 65% of the strains presented considerable overexpression of the genes involved in the acetoin catabolic (aco) pathway. We demonstrated that acetoin is (i) produced by clinical S. aureus strains, (ii) detected in sputa from CF patients and (iii) involved in P. aeruginosa's aco system induction. Furthermore, acetoin is catabolized by P. aeruginosa, a metabolic process that improves the survival of both pathogens by providing a new carbon source for P. aeruginosa and avoiding the toxic accumulation of acetoin on S. aureus. Due to its beneficial effects on both bacteria, acetoin catabolism could testify to the establishment of trophic cooperation between S. aureus and P. aeruginosa in the CF lung environment, thus promoting their persistence.

Impact of Coexistence Phenotype Between Staphylococcus aureus and Pseudomonas aeruginosa Isolates on Clinical Outcomes Among Cystic Fibrosis Patients.

Staphylococcus aureus (SA) is the major colonizer of the lungs of cystic fibrosis (CF) patients during childhood and adolescence. As patients age, the prevalence of SA decreases and Pseudomonas aeruginosa (PA) becomes the major pathogen infecting adult lungs. Nonetheless, SA remains significant and patients harboring both SA and PA are frequently found in the worldwide cohort. The overall impact of co-infection remains controversial. Furthermore, co-infecting isolates may compete or coexist. The aim of this study was to analyse if co-infection and the coexistence of SA and PA could lead to worse clinical outcomes. The clinical and bacteriological data of 212 Lyon CF patients were collected retrospectively, and patients were ranked into three groups, SA only (n = 112), PA only (n = 48) or SA plus PA (n = 52). In addition, SA and PA isolates from co-infected patients were tested in vitro to define their interaction profile. Sixty five percent (n = 34) of SA/PA pairs coexist. Using univariate and multivariate analysis, we confirm that SA patients have a less severe clinical condition than others, and PA induces a poor outcome independently of the presence of SA. Regarding co-infection, no significant difference in clinical outcomes was observed between patients with coexisting pairs and patients with competitive pairs. However, when compared to SA mono-infected patients, patients with coexisting pair presented higher frequency and length of hospitalizations and more exacerbations. We suggest that coexistence between SA and PA may be an important step in the natural history of lung bacterial colonization within CF patients.

Coexistence with Pseudomonas aeruginosa alters Staphylococcus aureus transcriptome, antibiotic resistance and internalization into epithelial cells.

Cystic fibrosis (CF) is the most common life-threatening genetic disease among Caucasians. CF patients suffer from chronic lung infections due to the presence of thick mucus, caused by cftr gene dysfunction. The two most commonly found bacteria in the mucus of CF patients are Staphylococcus aureus and Pseudomonas aeruginosa. It is well known that early-infecting P. aeruginosa strains produce anti-staphylococcal compounds and inhibit S. aureus growth. More recently, it has been shown that late-infecting P. aeruginosa strains develop commensal-like/coexistence interaction with S. aureus. The aim of this study was to decipher the impact of P. aeruginosa strains on S. aureus. RNA sequencing analysis showed 77 genes were specifically dysregulated in the context of competition and 140 genes in the context of coexistence in the presence of P. aeruginosa. In coexistence, genes encoding virulence factors and proteins involved in carbohydrates, lipids, nucleotides and amino acids metabolism were downregulated. On the contrary, several transporter family encoding genes were upregulated. In particular, several antibiotic pumps belonging to the Nor family were upregulated: tet38, norA and norC, leading to an increase in antibiotic resistance of S. aureus when exposed to tetracycline and ciprofloxacin and an enhanced internalization rate within epithelial pulmonary cells. This study shows that coexistence with P. aeruginosa affects the S. aureus transcriptome and virulence.

Molecular evolution of hemojuvelin and the repulsive guidance molecule family.

Repulsive guidance molecules (RGMs) are found in vertebrates and chordates and are involved in embryonic development and iron homeostasis. Members of this family are GPI-linked membrane proteins that contain an N-terminal signal peptide, a C-terminal propeptide, and a conserved RGD motif. Vertebrates are known to possess three paralogues; RGMA and RGMB (sometimes called Dragon) are expressed in the nervous system and are thought to play various roles in neural development. Hemojuvelin (HJV; also called repulsive guidance molecule c, RGMC) is the third member of this family, and mutations in this gene result in a form of juvenile hemochromatosis (type 2A). Phylogenetic analyses of 55 different RGM family sequences from 21 different species support the existence of a novel gene, found only in fish, which we have labeled RGMD. The pattern of conserved residues in each family identifies new candidates for important functional roles, including ligand binding.