Metabolic Signatures of Triatomine Vectors of <i>Trypanosoma cruzi</i> Unveiled by Metabolomics

<div><p>Chagas disease is a trypanosomiasis whose causative agent is the protozoan parasite <i>Trypanosoma cruzi</i>, which is transmitted to humans by hematophagous insects known as triatomines and affects a large proportion of South America. The digestive tract of the insect vectors in which <i>T. cruzi</i> develops constitutes a dynamic environment that affects the development of the parasite. Thus, we set out to investigate the chemical composition of the triatomine intestinal tract through a metabolomics approach. We performed Direct Infusion Fourier Transform Ion Cyclotron Resonance Mass Spectrometry on fecal samples of three triatomine species (<i>Rhodnius prolixus</i>, <i>Triatoma infestans</i>, <i>Panstrongylus megistus</i>) fed with rabbit blood. We then identified groups of metabolites whose frequencies were either uniform in all species or enriched in each of them. By querying the Human Metabolome Database, we obtained putative identities of the metabolites of interest. We found that a core group of metabolites with uniform frequencies in all species represented approximately 80% of the molecules detected, whereas the other 20% varied among triatomine species. The uniform core was composed of metabolites of various categories, including fatty acids, steroids, glycerolipids, nucleotides, sugars, and others. Nevertheless, the metabolic fingerprint of triatomine feces differs depending on the species considered. The variable core was mainly composed of prenol lipids, amino acids, glycerolipids, steroids, phenols, fatty acids and derivatives, benzoic acid and derivatives, flavonoids, glycerophospholipids, benzopyrans, and quinolines. Triatomine feces constitute a rich and varied chemical medium whose constituents are likely to affect <i>T. cruzi</i> development and infectivity. The complexity of the fecal metabolome of triatomines suggests that it may affect triatomine vector competence for specific <i>T. cruzi</i> strains. Knowledge of the chemical environment of <i>T. cruzi</i> in its invertebrate host is likely to generate new ways to understand the factors influencing parasite proliferation as well as methods to control Chagas disease.</p></div

Metabolic classes in the uniform core.

<p>Frequency is given in number of hits per metabolic category in the uniform core. Only metabolic classes with 10 or more hits are displayed.</p

Metabolic classes in the variable core.

<p>Frequency is given in number of hits per metabolic category in the following comparisons: <i>P. megistus</i> vs. <i>T. infestans</i> and <i>R. prolixus</i> (white bars), <i>R. prolixus</i> vs. <i>T. infestans</i> and <i>P. megistus</i> (gray bars), and <i>T. infestans</i> vs. <i>R. prolixus</i> and <i>P. megistus</i> (black bars). Only metabolic classes with 5 or more hits are displayed.</p

Representation of relative metabolite distribution.

<p>Most metabolites were found at very low rates (below 0.025%). The distribution extends above 0.05, up to 14%, but was not shown for clarity.</p

Boolean operations on metabolite rate differences.

<p>Venn diagrams are given for all replicate combinations considering the following triatomine comparisons: <i>R. prolixus</i> vs. <i>T. infestans</i> AND <i>P. megistus</i> (A), <i>T. infestans</i> vs. <i>R. prolixus</i> AND <i>P. megistus</i> (B), <i>P. megistus</i> vs. <i>T. infestans</i> AND <i>R. prolixus</i> (C), and all comparisons above (D).</p

Distributions of metabolite rate differences.

<p>Three examples are given for the distributions of these differences among feces of different triatomine species, i.e., <i>T. infestans</i> vs. <i>R. prolixus</i> (A), <i>T. infestans</i> vs. <i>P. megistus</i> (B) and <i>R. prolixus</i> vs. <i>P. megistus</i> (C). The histograms focus on the significant part of the samples in terms of representativeness, but the values were found in a larger interval. In all panels, ∼95% of pair differences are found between −0.15 and +0.15 (n = 2,086).</p

Differences in ion frequencies among triatomine replicates.

<p>Plots are given for all replicate combinations considering the following triatomine pairs: <i>T. infestans</i> vs. <i>R. prolixus</i> (A), <i>T. infestans</i> vs. <i>P. megistus</i> (B) and <i>R. prolixus</i> vs. <i>P. megistus</i> (C). Dots between dashed lines are for the metabolites with small differences among pairs of triatomine species. Dots outside the dashed lines are for the metabolites displaying large differences among pairs of triatomine species (at <i>p</i>≤0.05). For plotting convenience, the scale of the <i>y</i> axis has been limited to the interval −1 to +1. Some pairs exist outside this range (data not shown).</p

Metabonomics Reveals Drastic Changes in Anti-Inflammatory/Pro-Resolving Polyunsaturated Fatty Acids-Derived Lipid Mediators in Leprosy Disease

<div><p>Despite considerable efforts over the last decades, our understanding of leprosy pathogenesis remains limited. The complex interplay between pathogens and hosts has profound effects on host metabolism. To explore the metabolic perturbations associated with leprosy, we analyzed the serum metabolome of leprosy patients. Samples collected from lepromatous and tuberculoid patients before and immediately after the conclusion of multidrug therapy (MDT) were subjected to high-throughput metabolic profiling. Our results show marked metabolic alterations during leprosy that subside at the conclusion of MDT. Pathways showing the highest modulation were related to polyunsaturated fatty acid (PUFA) metabolism, with emphasis on anti-inflammatory, pro-resolving omega-3 fatty acids. These results were confirmed by eicosanoid measurements through enzyme-linked immunoassays. Corroborating the repertoire of metabolites altered in sera, metabonomic analysis of skin specimens revealed alterations in the levels of lipids derived from lipase activity, including PUFAs, suggesting a high lipid turnover in highly-infected lesions. Our data suggest that omega-6 and omega-3, PUFA-derived, pro-resolving lipid mediators contribute to reduced tissue damage irrespectively of pathogen burden during leprosy disease. Our results demonstrate the utility of a comprehensive metabonomic approach for identifying potential contributors to disease pathology that may facilitate the development of more targeted treatments for leprosy and other inflammatory diseases.</p></div

Comparison of the relative levels^a of metabolites of the arachidonic acid pathway in sera from BT and LL patients before and after antibiotic treatment.

a<p>Absent values were substituted with the limit of detection, represented by the lowest intensity value of any given sample. Then, averaged values from the untreated BT serum samples (n = 4) were normalized to 100% and other samples were normalized accordingly. SD are shown in parentheses. PG, prostaglandin; LT, leukotriene; TX, thromboxane; EET, epoxyeicosatrienoic acid; oxo-ETE, oxoicosatetraenoic acid; HETE, hydroxyeicosatetraenoic acid; HPETE, hydroperoxyeicosatetraenoic acid; DHET, dihydroxyeicosatrienoic acid.</p

Serum levels of eicosanoids in borderline tuberculoid and polar lepromatous patients determined by EIAs.

<p>Box-plots represent serum levels of PGD₂ (a), PGE₂ (b), LTB₄ (c) and LXA₄ (d) assessed in healthy controls, BT and LL patients, as indicated. Median values are indicated by lines. Outliers were detected using the Grubbs' test and removed. Group comparisons were evaluated with Kruskall–Wallis non-parametric analysis of variance (ANOVA) and Dunn's multiple-range post hoc test. PGD₂, prostaglandin D₂; PGE₂, prostaglandin E₂; LTB₄, leukotriene B₄; LXA₄, lipoxin A₄. <i>P</i>-values higher than 0.05 are not shown.</p