Spatialized Metabolomic Annotation Combining MALDI Imaging and Molecular Networks.

MALDI mass spectrometry imaging has gained major interest in the field of chemical imaging. This technique makes it possible to locate tens to hundreds of ionic signals on the sample surface without any a priori. One of the current challenges is still the limited ability to annotate signals in order to convert m/z values into probable chemical structures. At the same time, data obtained by LC-MS/MS have benefited from the development of numerous chemoinformatics tools, in particular molecular networks, for their efficient annotation. For the first time, we present here the combination of MALDI-FT-ICR imaging with molecular networks from MALDI-MS/MS data directly acquired on plant tissue sections. Annotation improvements are demonstrated, paving the way for new annotation pipelines for MALDI imaging.

Nitrogen Enriched Solid-State Cultivation for the Overproduction of Azaphilone Red Pigments by Penicillium sclerotiorum SNB-CN111.

Azaphilones are microbial specialized metabolites employed as yellow, orange, red or purple pigments. In particular, yellow azaphilones react spontaneously with functionalized nitrogen groups, leading to red azaphilones. In this study, a new two-step solid-state cultivation process to produce specific red azaphilones pigments was implemented, and their chemical diversity was explored based on liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) and a molecular network. This two-step procedure first implies a cellophane membrane allowing accumulating yellow and orange azaphilones from a Penicillium sclerotiorum SNB-CN111 strain, and second involves the incorporation of the desired functionalized nitrogen by shifting the culture medium. The potential of this solid-state cultivation method was finally demonstrated by overproducing an azaphilone with a propargylamine side chain, representing 16% of the metabolic crude extract mass.

Comparative metabolomic study of fungal foliar endophytes and their long-lived host Astrocaryum sciophilum: a model for exploring the chemodiversity of host-microbe interactions.

Introduction: In contrast to the dynamics observed in plant/pathogen interactions, endophytic fungi have the capacity to establish enduring associations within their hosts, leading to the development of a mutually beneficial relationship that relies on specialized chemical interactions. Research indicates that the presence of endophytic fungi has the ability to significantly modify the chemical makeup of the host organism. Our hypothesis proposes the existence of a reciprocal exchange of chemical signals between plants and fungi, facilitated by specialized chemical processes that could potentially manifest within the tissues of the host. This research aimed to precisely quantify the portion of the cumulative fungal endophytic community's metabolome detectable within host leaves, and tentatively evaluate its relevance to the host-endophyte interplay. The understory palm Astrocaryum sciophilum (Miq.) Pulle was used as a interesting host plant because of its notable resilience and prolonged life cycle, in a tropical ecosystem. Method: Using advanced metabolome characterization, including UHPLC-HRMS/MS and molecular networking, the study explored enriched metabolomes of both host leaves and 15 endophytic fungi. The intention was to capture a metabolomic "snapshot" of both host and endophytic community, to achieve a thorough and detailed analysis. Results and discussion: This approach yielded an extended MS-based molecular network, integrating diverse metadata for identifying host- and endophyte-derived metabolites. The exploration of such data (>24000 features in positive ionization mode) enabled effective metabolome comparison, yielding insights into cultivable endophyte chemodiversity and occurrence of common metabolites between the holobiont and its fungal communities. Surprisingly, a minor subset of features overlapped between host leaf and fungal samples despite significant plant metabolome enrichment. This indicated that fungal metabolic signatures produced in vitro remain sparingly detectable in the leaf. Several classes of primary metabolites were possibly shared. Specific fungal metabolites and/or compounds of their chemical classes were only occasionally discernible in the leaf, highlighting endophytes partial contribution to the overall holobiont metabolome. To our knowledge, the metabolomic study of a plant host and its microbiome has rarely been performed in such a comprehensive manner. The general analytical strategy proposed in this paper seems well-adapted for any study in the field of microbial- or microbiome-related MS and can be applied to most host-microbe interactions.

Combining OSMAC, metabolomic and genomic methods for the production and annotation of halogenated azaphilones and ilicicolins in termite symbiotic fungi.

We gathered a collection of termite mutualistic strains from French Guiana to explore the metabolites of symbiotic microorganisms. Molecular networks reconstructed from a metabolomic analysis using LC-ESI-MS/MS methodology led us to identify two families of chlorinated polyketides, i.e., azaphilones from Penicillium sclerotiorum and ilicicolins from Neonectria discophora. To define the biosynthetic pathways related to these two types of scaffolds, we used a whole genome sequencing approach followed by hybrid assembly from short and long reads. We found two biosynthetic gene clusters, including two FAD-dependent halogenases. To exploit the enzymatic promiscuity of the two identified FAD halogenases, we sought to biosynthesize novel halogenated metabolites. An OSMAC strategy was used and resulted in the production of brominated analogs of ilicicolins and azaphilones as well as iodinated analogs of azaphilones.

Classification of Environmental Strains from Order to Genus Levels Using Lipid and Protein MALDI-ToF Fingerprintings and Chemotaxonomic Network Analysis.

During the last two decades, MALDI-ToF mass spectrometry has become an efficient and widely-used tool for identifying clinical isolates. However, its use for classification and identification of environmental microorganisms remains limited by the lack of reference spectra in current databases. In addition, the interpretation of the classical dendrogram-based data representation is more difficult when the quantity of taxa or chemotaxa is larger, which implies problems of reproducibility between users. Here, we propose a workflow including a concurrent standardized protein and lipid extraction protocol as well as an analysis methodology using the reliable spectra comparison algorithm available in MetGem software. We first validated our method by comparing protein fingerprints of highly pathogenic bacteria from the Robert Koch Institute (RKI) open database and then implemented protein fingerprints of environmental isolates from French Guiana. We then applied our workflow for the classification of a set of protein and lipid fingerprints from environmental microorganisms and compared our results to classical genetic identifications using 16S and ITS region sequencing for bacteria and fungi, respectively. We demonstrated that our protocol allowed general classification at the order and genus level for bacteria whereas only the Botryosphaeriales order can be finely classified for fungi.

Antiparasitic Ovalicin Derivatives from Pseudallescheria boydii, a Mutualistic Fungus of French Guiana Termites.

Social insects are in mutualism with microorganisms, contributing to their resistance against infectious diseases. The fungus Pseudallescheria boydii SNB-CN85 isolated from termites produces ovalicin derivatives resulting from the esterification of the less hindered site of the ovalicin epoxide by long-chain fatty acids. Their structures were elucidated using spectroscopic analysis and semisynthesis from ovalicin. For ovalicin, these compounds displayed antiprotozoal activities against Plasmodium falciparum and Trypanosoma brucei, with IC50 values of 19.8 and 1.1 µM, respectively, for the most active compound, i.e., ovalicin linoleate. In parallel, metabolomic profiling of a collection of P. boydii strains associated with termites made it possible to highlight this class of compounds together with tyroscherin derivatives in all strains. Finally, the complete genome of P. boydii strains was obtained by sequencing, and the cluster of potential ovalicin and ovalicin biosynthesis genes was annotated. Through these metabolomic and genomic analyses, a new ovalicin derivative named boyden C, in which the 6-membered ring of ovalicin was opened by oxidative cleavage, was isolated and structurally characterized.

Treating leishmaniasis in Amazonia, part 2: Multi-target evaluation of widely used plants to understand medicinal practices.

ETHNOPHARMACOLOGICAL RELEVANCE: Leishmaniasis are widely distributed among tropical and subtropical countries, and remains a crucial health issue in Amazonia. Indigenous groups across Amazonia have developed abundant knowledge about medicinal plants related to this pathology. AIM OF THE STUDY: We intent to explore the weight of different pharmacological activities driving taxa selection for medicinal use in Amazonian communities. Our hypothesis is that specific activity against Leishmania parasites is only one factor along other (anti-inflammatory, wound healing, immunomodulating, antimicrobial) activities. MATERIALS AND METHODS: The twelve most widespread plant species used against leishmaniasis in Amazonia, according to their cultural and biogeographical importance determined through a wide bibliographical survey (475 use reports), were selected for this study. Plant extracts were prepared to mimic their traditional preparations. Antiparasitic activity was evaluated against promastigotes of reference and clinical New-World strains of Leishmania (L. guyanensis, L. braziliensis and L. amazonensis) and L. amazonensis intracellular amastigotes. We concurrently assessed the extracts immunomodulatory properties on PHA-stimulated human PBMCs and RAW264.7 cells, and on L. guyanensis antigens-stimulated PBMCs obtained from Leishmania-infected patients, as well as antifungal activity and wound healing properties (human keratinocyte migration assay) of the selected extracts. The cytotoxicity of the extracts against various cell lines (HFF1, THP-1, HepG2, PBMCs, RAW264.7 and HaCaT cells) was also considered. The biological activity pattern of the extracts was represented through PCA analysis, and a correlation matrix was calculated. RESULTS: Spondias mombin L. bark and Anacardium occidentale L. stem and leaves extracts displayed high anti-promatigotes activity, with IC50 ≤ 32 µg/mL against L. guyanensis promastigotes for S. mombin and IC50 of 67 and 47 µg/mL against L. braziliensis and L. guyanensis promastigotes, respectively, for A. occidentale. In addition to the antiparasitic effect, antifungal activity measured against C. albicans and T. rubrum (MIC in the 16-64 µg/mL range) was observed. However, in the case of Leishmania amastigotes, the most active species were Bixa orellana L. (seeds), Chelonantus alatus (Aubl.) Pulle (leaves), Jacaranda copaia (Aubl.) D. Don. (leaves) and Plantago major L. (leaves) with IC50 < 20 µg/mL and infection rates of 14-25% compared to the control. Concerning immunomodulatory activity, P. major and B. orellana were highlighted as the most potent species for the wider range of cytokines in all tested conditions despite overall contrasting results depending on the model. Most of the species led to moderate to low cytotoxic extracts except for C. alatus, which exhibited strong cytotoxic activity in almost all models. None of the tested extracts displayed wound healing properties. CONCLUSIONS: We highlighted pharmacologically active extracts either on the parasite or on associated pathophysiological aspects, thus supporting the hypothesis that antiparasitic activities are not the only biological factor useful for antileishmanial evaluation. This result should however be supplemented by in vivo studies, and attracts once again the attention on the importance of the choice of biological models for an ethnophamacologically consistent study. Moreover, plant cultural importance, ecological status and availability were discussed in relation with biological results, thus contributing to link ethnobotany, medical anthropology and biology.

Antimicrobial Activity and Synergy Investigation of Hypericum scabrum Essential Oil with Antifungal Drugs.

The chemical composition of Lebanese Hypericum scabrum essential oil (EO) was analyzed by gas chromatography (GC) and gas chromatography-mass spectrometry (GG-MS). Its antimicrobial activity was evaluated by determining its minimal inhibitory concentrations (MICs) against a Gram-negative and a Gram-positive bacterium, one yeast, and five dermatophytes. H. scabrum EO was most active on filamentous fungi (MIC values of 32-64 µg/mL). Synergy within the oil was investigated by testing each of the following major components on Trichophyton rubrum: α-pinene, limonene, myrcene, ß-pinene and nonane, as well as a reconstructed EO. The antifungal activity of the natural oil could not be reached, meaning that its activity might be due, in part, to minor constituent(s). The interactions between H. scabrum EO and commercially available antifungals were assessed by the checkerboard test. A synergistic effect was revealed in the combination of the EO with amphotericin B.

Identification of Antagonistic Compounds between the Palm Tree Xylariale Endophytic Fungi and the Phytopathogen Fusarium oxysporum.

To discover microorganisms that naturally possess chemical weapons against the phytopathogen Fusarium oxysporum, the biological and chemical diversity of plant leaf endophytes was investigated. Endophytes were isolated from the palm tree Astrocaryum sciophyllum collected in pristine forests of French Guiana. Several Xylariaceae inhibited the growth of F. oxysporum and were further explored. Antifungal specialized metabolites were isolated from the Xylariaceae BSNB-0294 strain in confrontation with the phytopathogen and led to the identification of undescribed compounds, i.e., two depsipeptides named xylariaceins, two metabolites containing a 3-imidazolinone moiety, and four new compounds including a nitro-phenylpropanamide and three phenylalanine analogues named xylariains A-D. In parallel, the chemical investigation of the phytopathogen during the coculture led to the identification of an unknown compound, which we named focicin. The production of focicin was exacerbated during the competition. Matrix-assisted laser desorption/ionization coupled to time-of-flight mass spectometry (MALDI-TOF MS) imaging of the competition between BSNB-0294 (endophytic strain) and F. oxysporum f.sp. ciceris (phytopathogen) highlighted time-dependent chemical interactions between the two microorganisms.

Characterization of Pseudomonas aeruginosa Quorum Sensing Inhibitors from the Endophyte Lasiodiplodia venezuelensis and Evaluation of Their Antivirulence Effects by Metabolomics.

The opportunistic pathogen Pseudomonas aeruginosa is one of the "critical priority pathogens" due to its multidrug resistance to a wide range of antibiotics. Its ability to invade and damage host tissues is due to the use of quorum sensing (QS) to collectively produce a plethora of virulence factors. Inhibition of QS is an attractive strategy for new antimicrobial agents because it disrupts the initial events of infection without killing the pathogen. Highly diverse microorganisms as endophytes represent an under-explored source of bioactive natural products, offering opportunities for the discovery of novel QS inhibitors (QSI). In the present work, the objective was to explore selective QSIs within a unique collection of fungal endophytes isolated from the tropical palm Astrocaryum sciophilum. The fungi were cultured, extracted, and screened for their antibacterial and specific anti-QS activities against P. aeruginosa. The endophytic strain Lasiodiplodia venezuelensis was prioritized for scaled-up fractionation for its selective activity, leading to the isolation of eight compounds in a single step. Among them, two pyran-derivatives were found to be responsible for the QSI activity, with an effect on some QS-regulated virulence factors. Additional non-targeted metabolomic studies on P. aeruginosa documented their effects on the production of various virulence-related metabolites.

Isolation and Identification of Isocoumarin Derivatives With Specific Inhibitory Activity Against Wnt Pathway and Metabolome Characterization of Lasiodiplodia venezuelensis.

The Wnt signaling pathway controls multiple events during embryonic development of multicellular animals and is carcinogenic when aberrantly activated in adults. Breast cancers are dependent on Wnt pathway overactivation mostly through dysregulation of pathway component protein expression, which necessitates the search for therapeutically relevant compounds targeting them. Highly diverse microorganisms as endophytes represent an underexplored field in the therapeutic natural products research. In the present work, the objective was to explore the chemical diversity and presence of selective Wnt inhibitors within a unique collection of fungi isolated as foliar endophytes from the long-lived tropical palm Astrocaryum sciophilum. The fungi were cultured, extracted with ethyl acetate, and screened for their effects on the Wnt pathway and cell proliferation. The endophytic strain Lasiodiplodia venezuelensis was prioritized for scaled-up fractionation based on its selective activity. Application of geometric transfer from analytical HPLC conditions to semi-preparative scale and use of dry load sample introduction enabled the isolation of 15 pure compounds in a single step. Among the molecules identified, five are original natural products described for the first time, and six are new to this species. An active fraction obtained by semi-preparative HPLC was re-purified by UHPLC-PDA using a 1.7 µm phenyl column. 75 injections of 8 µg were necessary to obtain sufficient amounts of each compound for structure elucidation and bioassays. Using this original approach, in addition to the two major compounds, a third minor compound identified as (R)-(-)-5-hydroxymellein (18) was obtained, which was found to be responsible for the significant Wnt inhibition activity recorded. Further studies of this compound and its structural analogs showed that only 18 acts in a highly specific manner, with no acute cytotoxicity. This compound is notably selective for upstream components of the Wnt pathway and is able to inhibit the proliferation of three triple negative breast cancer cell lines. In addition to the discovery of Wnt inhibitors of interest, this study contributes to better characterize the biosynthetic potential of L. venezuelensis.

Dereplication, Annotation, and Characterization of 74 Potential Antimicrobial Metabolites from Penicillium Sclerotiorum Using t-SNE Molecular Networks.

Microorganisms associated with termites are an original resource for identifying new chemical scaffolds or active metabolites. A molecular network was generated from a collection of strain extracts analyzed by liquid chromatography coupled to tandem high-resolution mass spectrometry, a molecular network was generated, and activities against the human pathogens methicillin-resistant Staphylococcus aureus, Candida albicans and Trichophyton rubrum were mapped, leading to the selection of a single active extract of Penicillium sclerotiorum SNB-CN111. This fungal species is known to produce azaphilones, a colorful family of polyketides with a wide range of biological activities and economic interests in the food industry. By exploring the molecular network data, it was shown that the chemical diversity related to the P. sclerotiorum metabolome largely exceeded the data already reported in the literature. According to the described fragmentation pathways of protonated azaphilones, the annotation of 74 azaphilones was proposed, including 49 never isolated or synthesized thus far. Our hypothesis was validated by the isolation and characterization of eight azaphilones, among which three new azaphilones were chlorogeumasnol (63), peniazaphilone E (74) and 7-deacetylisochromophilone VI (80).

Identification and dereplication of endophytic Colletotrichum strains by MALDI TOF mass spectrometry and molecular networking.

The chemical diversity of biologically active fungal strains from 42 Colletotrichum, isolated from leaves of the tropical palm species Astrocaryum sciophilum collected in pristine forests of French Guiana, was investigated. The collection was first classified based on protein fingerprints acquired by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) correlated with cytotoxicity. Liquid chromatography coupled to high-resolution tandem mass spectrometry (LC-HRMS/MS) data from ethyl acetate extracts were acquired and processed to generate a massive molecular network (MN) using the MetGem software. From five Colletotrichum strains producing cytotoxic specialized metabolites, we predicted the occurrence of peptide and cytochalasin analogues in four of them by MN, including a similar ion clusters in the MN algorithm provided by MetGem software. Chemoinformatics predictions were fully confirmed after isolation of three pentacyclopeptides (cyclo(Phe-Leu-Leu-Leu-Val), cyclo(Phe-Leu-Leu-Leu-Leu) and cyclo(Phe-Leu-Leu-Leu-Ile)) and two cytochalasins (cytochalasin C and cytochalasin D) exhibiting cytotoxicity at the micromolar concentration. Finally, the chemical study of the last active cytotoxic strain BSNB-0583 led to the isolation of four colletamides bearing an identical decadienamide chain.

Paecilosetin Derivatives as Potent Antimicrobial Agents from Isaria farinosa.

Fifty-seven entomopathogenic microorganisms were screened against human pathogens and subjected to mass spectrometry molecular networking based dereplication. Isaria farinosa BSNB-1250, shown to produce potentially novel biologically active metabolites, was grown on a large scale on potato dextrose agar, and paecilosetin (1) and five new analogues (2-6) were subsequently isolated. The structures of the new compounds were elucidated using 1D and 2D NMR. The absolute configurations of compounds 1-6 were determined using Mosher ester derivatives (1, 3, 4), comparison of experimental and calculated ECD spectra (2-4 and 6), and single-crystal X-ray diffraction analysis (5). Compounds 1 and 5 exhibited strong antibacterial activity against MSSA and MRSA with MIC values of 1-2 µg/mL.

Atmospheric pressure photoionization versus electrospray for the dereplication of highly conjugated natural products using molecular networks.

Natural products are sources of inspiration and reservoir of high valuable molecules. Recently, analytical tools based on liquid chromatography coupled to tandem mass spectrometry to generate molecular network became widely employed for dereplication. This strategy greatly accelerates the identification of known and structural hypothesis of unknown. Despite the availability of different ionization sources, alternatives to classical electrospray ionization (ESI), such as atmospheric pressure chemical ionization (APCI) or photoionization (APPI), have been neglected. In particular, APPI has been described for its ionization efficiency on non-polar molecules bearing no acid or basic groups. For that reason, we investigated APPI potential to generate molecular network and compare it to ESI on several criteria that are generation of ion species, sensitivity and signal-to-noise ratio (SNR) for different extracts rich in highly conjugated natural products. We first optimized APPI experimental conditions on crude extract from a fungus, Penicillium sclerotiorum, producing polyketones belonging to the azaphilone family. Then we compared APPI and ESI on different fractions of the fungus and on two plant extracts, French Guyanese Swartzia panacoco (Aubl.) R.S. Cowan (arial parts) and Indian Cassia auriculata L. (leaves) containing phenolic compounds, such as flavonoids. While ESI generated more ion species and displayed a better sensitivity, APPI generated only protonated adduct and better SNR. Comparing ESI and APPI generated species on molecular network reveal that both strategies overlap for the majority of protonated ions.

Potent and Non-Cytotoxic Antibacterial Compounds Against Methicillin-Resistant Staphylococcus aureus Isolated from Psiloxylon mauritianum, A Medicinal Plant from Reunion Island.

With the occurrence of antibiotic-resistant Staphylococcus aureus strains, identification of new anti-staphylococcal drugs has become a necessity. It has long been demonstrated that plants are a large and diverse source of antibacterial compounds. Psiloxylon mauritianum, an endemic medicinal plant from Reunion Island, was chemically investigated for its reported biological activity against S. aureus. Aspidin VB, a phloroglucinol derivative never before described, together with Aspidin BB, were first isolated from the ethyl acetate extract of P. mauritianum leaves. Their structures were elucidated from spectroscopic data. Aspidin VB exhibited strong antibacterial activity against standard and methicillin-resistant S. aureus strains, with a minimal inhibition concentration (MIC) of 0.25 µg/mL, and no cytotoxicity was observed at 10-5 M in MRC5 cells. Due to its biological activities, Aspidin VB appears to be a good natural lead in the fight against S. aureus.

Alsinol, an arylamino alcohol derivative active against Plasmodium, Babesia, Trypanosoma, and Leishmania: past and new outcomes.

Malaria, babesiosis, trypanosomosis, and leishmaniasis are some of the most life-threatening parasites, but the range of drugs to treat them is limited. An effective, safe, and low-cost drug with a large activity spectrum is urgently needed. For this purpose, an aryl amino alcohol derivative called Alsinol was resynthesized, screened in silico, and tested against Plasmodium, Babesia, Trypanosoma, and Leishmania. In silico Alsinol follows the Lipinski and Ghose rules. In vitro it had schizontocidal activity against Plasmodium falciparum and was able to inhibit gametocytogenesis; it was particularly active against late gametocytes. In malaria-infected mice, it showed a dose-dependent activity similar to chloroquine. It demonstrated a similar level of activity to reference compounds against Babesia divergens, and against promastigotes, and amastigotes stages of Leishmania in vitro. It inhibited the in vitro growth of two African animal strains of Trypanosoma but was ineffective in vivo in our experimental conditions. It showed moderate toxicity in J774A1 and Vero cell models. The study demonstrated that Alsinol has a large spectrum of activity and is potentially affordable to produce. Nevertheless, challenges remain in the process of scaling up synthesis, creating a suitable clinical formulation, and determining the safety margin in preclinical models.

Carneic Acids from an Endophytic Phomopsis sp. as Dengue Virus Polymerase Inhibitors.

Thirteen carneic acids were isolated from the fungal endophyte Phomopsis sp. SNB-LAP1-7-32. Their structures were identified by mass spectrometry and extensive one- and two-dimensional NMR spectroscopy and through comparison with data reported in the literature. Compounds 1-13 were investigated for their antipolymerase activities against DENV polymerase and Zika NS5. Five of them exhibited significant inhibition of dengue polymerase with IC50 values in the 10 to 20 µM range without cytotoxicity. None inhibited Zika virus NS5 protein.

Annotation and quantification of N-acyl homoserine lactones implied in bacterial quorum sensing by supercritical-fluid chromatography coupled with high-resolution mass spectrometry.

In recent years, use of supercritical-fluid chromatography (SFC) with CO2 as the mobile phase has been expanding in the research laboratory and industry since it is considered to be a green analytical method. This technique offers numerous advantages, such as good separation and sensitive detection, short analysis times, and stability of analytes. In this study, a method for quantification of N-acyl homoserine lactones (AHLs), signaling molecules responsible for cell-to-cell communication initially discovered in bacteria, by SFC coupled with high-resolution mass spectrometry (HRMS) was developed. The SFC conditions and MS ionization settings were optimized to obtain the best separation and greatest sensitivity. The optimal analysis conditions allowed quantification of up to 30 AHLs in a single run within 16 min with excellent linearity (R2 > 0.998) and sensitivity (picogram level). This method was then applied to study AHL production by one Gram-negative endophytic bacterium, Paraburkholderia sp. BSNB-0670. Nineteen known AHLs were detected, and nine abundant HSLs were quantified. To further investigate the production of uncommon AHLs, a molecular networking approach was applied on the basis of the SFC-HRMS/MS data. This led to additional identification of four unknown AHLs annotated as N-3-hydroxydodecanoylol homoserine lactone, N-3-hydroxydodecadienoyl homoserine lactone, and N-3-oxododecenoyl homoserine lactones (two isomers).

Towards the optimization of botanical insecticides research: Aedes aegypti larvicidal natural products in French Guiana.

Natural products have proven to be an immeasurable source of bioactive compounds. The exceptional biodiversity encountered in Amazonia, alongside a rich entomofauna and frequent interactions with various herbivores is the crucible of a promising chemodiversity. This prompted us to search for novel botanical insecticides in French Guiana. As this French overseas department faces severe issues linked to insects, notably the strong incidence of vector-borne infectious diseases, we decided to focus our research on products able to control the mosquito Aedes aegypti. We tested 452 extracts obtained from 85 species originating from 36 botanical families and collected in contrasted environments against an Ae. aegypti laboratory strain susceptible to all insecticides, and a natural population resistant to both pyrethroid and organophosphate insecticides collected in Cayenne for the most active of them. Eight species (Maytenus oblongata Reissek, Celastraceae; Costus erythrothyrsus Loes., Costaceae; Humiria balsamifera Aubl., Humiriaceae; Sextonia rubra (Mez) van der Werff, Lauraceae; Piper hispidum Sw., Piperaceae; Laetia procera (Poepp.) Eichl., Salicaceae; Matayba arborescens (Aubl.) Radlk., Sapindaceae; and Cupania scrobitulata Rich., Sapindaceae) led to extracts exhibiting more than 50% larval mortality after 48 h of exposition at 100 µg/mL against the natural population and were considered active. Selectivity and phytochemistry of these extracts were therefore investigated and discussed, and some active compounds highlighted. Multivariate analysis highlighted that solvents, plant tissues, plant family and location had a significant effect on mortality while light, available resources and vegetation type did not. Through this case study we highlighted that plant defensive chemistry mechanisms are crucial while searching for novel insecticidal products.