Substantial Extracellular Metabolic Differences Found Between Phylogenetically Closely Related Probiotic and Pathogenic Strains of Escherichia coli

Abstract

Since its first isolation a century ago, the gut inhabitant Escherichia coli strain Nissle 1917 has been shown to have probiotic activities; however, it is yet not fully elucidated which differential factors play key roles in its beneficial interactions with the host. To date, no metabolomics studies have been reported investigating the potential role of small molecules in functional strain differentiation of Nissle from its genetically close neighbors. Here, we present results of liquid chromatography coupled to high-resolution mass spectrometry characterization of extracellular metabolomes of E. coli strains as a proxy of their bioactivity potential. We found that phylogroup B2 strains exported a more diverse arsenal of metabolites than strains of other phylogroups. Zooming into the phylogroup B2 metabolome identified consistent substantial differences between metabolic output of E. coli Nissle and other strains, particularly in metabolites associated to the Argimine biosynthesis pathway. Nissle was found to release higher levels of Ornithine and Citrulline whilst depleting greater amounts of Arginine from the medium. Moreover, a novel Nissle-specific metabolite not reported before in bacteria, 5-(Carbamoylamino)-2-hydroxypentanoic acid (Citrulline/Arginic Acid related) was observed. Finally, Nissle, CFT073 and NCTC12241/ATCC25922 shared the excretion of N5-Acetylornithine, whereas other strains released N2-Acetylornithine or no N-Acetylornithine at all. Thus, we found substantial metabolic differences in phylogenetically very similar E. coli strains, an observation which suggests that it is justified to further investigate roles of small molecules as potential modulators of the gut environment by probiotic, commensal, and pathogenic strains, including E. coli Nissle 1917.

Integrating in vitro metabolomics with a 96-well high-throughput screening platform

“…Here, a monophasic solvent system consisting of 1:3:1 (v/v/v) water:methanol:chloroform was used (above), which was a modification of the classic monophasic solvent system 0.8:2:1 (v/v/v) water:methanol:chloroform (Bligh & Dyer, 1959 ) because a separation of polar and non-polar solvent phases was occasionally observed for this classic method (likely due to intracellular water, salts or residual wash solvent). The ratio of 1:3:1 (v/v/v) water:methanol:chloroform has been reported previously in metabolomics studies (Baptista et al, 2018 ; van der Hooft et al, 2019 ) and, as shown above, was effective at extracting polar metabolites for positive and negative ionisation modes by nESI-DIMS. However, attempts to analyse lipids in these extracts resulted in unacceptable electrospray instability (data not shown) that was likely due to the presence of more polar metabolites, prompting the evaluation of separate monophasic extractions of polar metabolites and lipids in a manner that would allow for automation.…”

“…In principle, omics technologies align well with the described shift in toxicity testing due to their ability to generate molecular mechanistic information to support decision making in regulatory applications, including chemical grouping and mode of action (MoA) prediction (Sperber et al, 2019;van Ravenzwaay et al, 2016;Viant et al, 2019). The value of metabolomics to risk assessment, defined as the profiling of metabolites (low-molecular-weight biochemicals) in cells, tissues or biofluids, has been acknowledged by several regulatory bodies (European Chemicals Agency, 2016;European Food Safety Authority et al, 2018).…”

Role of Carbon Monoxide in Host–Gut Microbiome Communication

“…Probiotic E. coli strains have also been shown to enhance the synthesis of antimicrobial β-defensin-2, engage in host signaling for immune-modulation, induce anti-inflammatory effects, and inhibit colonization of pathogens . Specifically, the gut resident and probiotic E. coli Nissle 1917 has been shown to reduce S. typhimurium colonization by causing iron deprivation via competition for heme …”

Metabolomics Study on Pathogenic and Non-pathogenic E. coli with Closely Related Genomes with a Focus on Yersiniabactin and Its Known and Novel Derivatives

“…However, so far there are hardly any metabolomic studies available on the differences of the small extracellular metabolites of these E. coli strains. A recently published study compared the metabolome of EcN and some other E. coli strains from the phylogroup B2, which are closely related to EcN and included CFT073 and 83972, and found differences in the metabolite profile of arginine biosynthesis-related metabolites [45].…”