Metabolomics Insights into Chemical Convergence in Xanthomonas perforans and Metabolic Changes Following Treatment with the Small Molecule Carvacrol

Abstract

Microbes are natural chemical factories and their metabolome comprise diverse arrays of chemicals. The genus Xanthomonas comprises some of the most important plant pathogens causing devastating yield losses globally and previous studies suggested that species in the genus are untapped chemical minefields. In this study, we applied an untargeted metabolomics approach to study the metabolome of a globally spread important xanthomonad, X. perforans. The pathogen is difficult to manage, but recent studies suggest that the small molecule carvacrol was efficient in disease control. Bacterial strains were treated with carvacrol, and samples were taken at time intervals (1 and 6 h). An untreated control was also included. There were five replicates for each sample and samples were prepared for metabolomics profiling using the standard procedure. Metabolomics profiling was carried out using a thermo Q-Exactive orbitrap mass spectrometer with Dionex ultra high-performance liquid chromatography (UHPLC) and an autosampler. Annotation of significant metabolites using the Metabolomics Standards Initiative level 2 identified an array of novel metabolites that were previously not reported in Xanthomonas perforans. These metabolites include methoxybrassinin and cyclobrassinone, which are known metabolites of brassicas; sarmentosin, a metabolite of the Passiflora-heliconiine butterfly system; and monatin, a naturally occurring sweetener found in Sclerochiton ilicifolius. To our knowledge, this is the first report of these metabolites in a microbial system. Other significant metabolites previously identified in non-Xanthomonas systems but reported in this study include maculosin; piperidine; β-carboline alkaloids, such as harman and derivatives; and several important medically relevant metabolites, such as valsartan, metharbital, pirbuterol, and ozagrel. This finding is consistent with convergent evolution found in reported biological systems. Analyses of the effect of carvacrol in time-series and associated pathways suggest that carvacrol has a global effect on the metabolome of X. perforans, showing marked changes in metabolites that are critical in energy biosynthesis and degradation pathways, amino acid pathways, nucleic acid pathways, as well as the newly identified metabolites whose pathways are unknown. This study provides the first insight into the X. perforans metabolome and additionally lays a metabolomics-guided foundation for characterization of novel metabolites and pathways in xanthomonad systems.

Impact of Weissella cibaria BYL4.2 and its supernatants on Penicillium chrysogenum metabolism

“…Macrocarpal I compounds have no bacteriostatic activity against gram-negative bacteria and fungi, but have bacteriostatic activity against gram-positive bacteria ( Wei, 2015 ); PS[20:4(5Z,8Z,11Z,14Z)/14:0] is a glycerophospholipid that promotes the activation of glycerophospholipid metabolism and GPC pathway cycling ( Wu et al, 2021 ). Sarmentosin is an unsaturated γ-hydroxynitrile glucoside produced by plants and is often chelated by lepidopterans, sarmentosin was previously reported only in plant systems and reported this metabolite for the first time in microbial systems ( Jibrin et al, 2021 ). 2-isopropyl malic acid is an intermediate product of leucine biosynthesis ( Suzuki et al, 2007 ).…”

Potential and Metabolic Pathways of Eugenol in the Management of Xanthomonas perforans, a Pathogen of Bacterial Spot of Tomato

“…Metabolite annotation was performed according to Level 2 of the Metabolomics Standards Initiative, as described previously [ 45 ]. The preliminary results of the one-way ANOVA after data normalization are shown in the Supplementary file (Figures S1a,b and S2a,b) .…”

“…The pathway analyses suggested that eugenol targets several pathways in X. perforans . The pathways that were significantly regulated by the putatively annotated metabolites in both untargeted search and when targeted against the Pseudomonas putida KT2440 are widely present in Gram-negative bacteria, including X. perforans [ 45 , 46 ]. At least 10 pathways and 12 pathways were significantly affected by the putatively annotated metabolites in the positive and negative ion phases, respectively.…”

“…Carvacrol, 3-indolylacetonitrile (IAN), and N-acetylcysteine (NAC) were shown to enhance the control of copper-resistant bacterial spot strains in south Florida [ 7 , 8 , 10 , 11 ]. Carvacrol improved disease management in greenhouse and field trials against X. perforans compared to the untreated control, with multiple pathways in the bacterium being altered [ 7 , 10 , 45 ]. IAN significantly reduced disease severity compared to the untreated control when applied three days prior to inoculation [ 8 ].…”

Bacterial Spot of Tomato and Pepper in Africa: Diversity, Emergence of T5 Race, and Management

“…Transcriptomics can improve understanding of the interaction of bacterial spot pathogens and their hosts ( Strauss et al, 2012 ; Du et al, 2015 ; Shi and Panthee, 2020 ). Recently, known and novel metabolites from nonbacterial spot systems were reported in X. perforans ( Jibrin et al, 2021b ). Combining metabolomics and transcriptomics in studying isolates of the bacterial spot pathogens from Africa could provide additional insights into the diversity of molecules that are important in host–pathogen or other ecological interactions and their pathways and expressed genes.…”