RESUMO
A multiblocked mutant strain (ΔAHBA and Δasm12, asm21) of Actinosynnema pretiosum, the producer of the highly toxic maytansinoid ansamitocin, has been used for the mutasynthetic production of new proansamitocin derivatives. The use of mutant strains that are blocked in the biosynthesis of an early building block as well as in the expression of two tailoring enzymes broadens the scope of chemo-biosynthetic access to new maytansinoids. Remarkably, a ring-expanded macrolactone derived from ansamitocin was created for the first time.
Assuntos
Actinobacteria/química , Maitansina/biossíntese , Actinobacteria/genética , Actinobacteria/metabolismo , Maitansina/análogos & derivados , Maitansina/química , Estrutura Molecular , MutaçãoRESUMO
In the biosynthesis of the tripyrrolic pigment prodigiosin, PigB is a predicted flavin-dependent oxidase responsible for the formation of 2-methyl-3-amylpyrrole (MAP) from a dihydropyrrole. To prove which dihydropyrrole is the true intermediate, both possibilities, 5-methyl-4-pentyl-3,4-dihydro-2H-pyrrole (5 a, resulting from transamination of the aldehyde of 3-acetyloctanal) and 2-methyl-3-pentyl-3,4-dihydro-2H-pyrrole (6, resulting from transamination of the ketone), were synthesised. Only 5 a restored pigment production in a strain of Serratia sp. ATCC 39006 blocked earlier in MAP biosynthesis. PigB is membrane-associated and inactive when its transmembrane domain was deleted, but HapB, its homologue in Hahella chejuensis, lacks the transmembrane domain and is active in solution. Two colourimetric assays for PigB and HapB were developed, and the HapB-catalysed reaction was kinetically characterised. Ten analogues of 5 a were synthesised, varying in the C2 and C3 side chains, and tested as substrates of HapB inâ vitro and for restoration of pigment production in Serratia ΔpigD inâ vivo. All lengths of side chain tested at C3 were accepted, but only short side chains at C2 were accepted. The knowledge that 5 a is an intermediate in prodigiosin biosynthesis and the ease of synthesis of analogues of 5 a makes a range of prodigiosin analogues readily available by mutasynthesis.
Assuntos
Antibacterianos/biossíntese , Proteínas de Bactérias/química , Gammaproteobacteria/enzimologia , Monoaminoxidase/química , Prodigiosina/biossíntese , Serratia/enzimologia , Especificidade por SubstratoRESUMO
Mutasynthesis of pyrichalasinâ H from Magnaportheâ griseaâ NI980 yielded a series of unprecedented 4'-substituted cytochalasin analogues in titres as high as the wild-type system (≈60â mg L-1 ). Halogenated, O-alkyl, O-allyl and O-propargyl examples were formed, as well as a 4'-azido analogue. 4'-O-Propargyl and 4'-azido analogues reacted smoothly in Huisgen cycloaddition reactions, whereas p-Br and p-I compounds reacted in Pd-catalysed cross-coupling reactions. A series of examples of biotin-linked, dye-linked and dimeric cytochalasins was rapidly created. Inâ vitro and inâ vivo bioassays of these compounds showed that the 4'-halogenated and azido derivatives retained their cytotoxicity and antifungal activities; but a unique 4'-amino analogue was inactive. Attachment of larger substituents attenuated the bioactivities. Inâ vivo actin-binding studies with adherent mammalian cells showed that actin remains the likely intracellular target. Dye-linked compounds revealed visualisation of intracellular actin structures even in the absence of phalloidin, thus constituting a potential new class of actin-visualisation tools with filament-barbed end-binding specificity.
Assuntos
Citocalasinas , Actinas , Animais , Citocalasinas/síntese química , Citocalasinas/química , Citocalasinas/farmacologia , Citoesqueleto , FaloidinaRESUMO
The argyrins are secondary metabolites from myxobacteria with antibiotic activity against Pseudomonas aeruginosa. Studying their structure-activity relationship is hampered by the complexity of the chemical total synthesis. Mutasynthesis is a promising approach where simpler and fully synthetic intermediates of the natural product's biosynthesis can be biotechnologically incorporated. Here, we report the synthesis of a series of tripeptide thioesters as mutasynthons containing the native sequence with a dehydroalanine (Dha) Michael acceptor attached to a sarcosine (Sar) and derivatives. Chemical synthesis of the native sequence á´ -Ala-Dha-Sar thioester required revision of the sequential peptide synthesis into a convergent strategy where the thioester with sarcosine was formed before coupling to the Dha-containing dipeptide.
RESUMO
Prodiginines are a group of naturally occurring pyrrole alkaloids produced by various microorganisms and known for their broad biological activities. The production of nature-inspired cyclic prodiginines was enabled by combining organic synthesis with a mutasynthesis approach based on the GRAS (generally recognized as safe) certified host strain Pseudomonas putida KT2440. The newly prepared prodiginines exerted antimicrobial effects against relevant alternative biotechnological microbial hosts whereas P.â putida itself exhibited remarkable tolerance against all tested prodiginines, thus corroborating the bacterium's exceptional suitability as a mutasynthesis host for the production of these cytotoxic secondary metabolites. Moreover, the produced cyclic prodiginines proved to be autophagy modulators in human breast cancer cells. One promising cyclic prodiginine derivative stood out, being twice as potent as prodigiosin, the most prominent member of the prodiginine family, and its synthetic derivative obatoclax mesylate.
RESUMO
Natural compounds are often structurally complex and their synthesis is still highly challenging. The review intends to give an overview on developments in biotechnology and their role for the production of natural products and active agents. In vitro and in vivo methods are presented side by side beginning with rather simple but smart single step conversions, followed by cascade reactions, and finishing with complex bio-, semi- and mutasynthesis endeavours. All the enzymatic approaches do obviously complement traditional synthetic methods; with their particular strengths, the combined repertoire will lead to an increased efficiency in natural product synthesis as well as in providing analogues.
Assuntos
Produtos Biológicos/metabolismo , Enzimas/metabolismo , Produtos Biológicos/química , Cinética , Estrutura Molecular , SoluçõesRESUMO
Thiomarinol and mupirocin are assembled on similar polyketide/fatty acid backbones and exhibit potent antibiotic activity against methicillin-resistant Staphylococcus aureus (MRSA). They both contain a tetrasubstituted tetrahydropyran (THP) ring that is essential for biological activity. Mupirocin is a mixture of pseudomonic acids (PAs). Isolation of the novel compound mupirocinâ P, which contains a 7-hydroxy-6-keto-substituted THP, from a ΔmupP strain and chemical complementation experiments confirm that the first step in the conversion of PA-B into the major product PA-A is oxidation at the C6â position. In addition, nine novel thiomarinol (TM) derivatives with different oxidation patterns decorating the central THP core were isolated after gene deletion (tmlF). These metabolites are in accord with the THP ring formation and elaboration in thiomarinol following a similar order to that found in mupirocin biosynthesis, despite the lack of some of the equivalent genes. Novel mupirocin-thiomarinol hybrids were also synthesized by mutasynthesis.
Assuntos
Antibacterianos/farmacologia , Staphylococcus aureus Resistente à Meticilina/efeitos dos fármacos , Mupirocina/análogos & derivados , Mupirocina/farmacologia , Policetídeo Sintases/genética , Antibacterianos/química , Antibacterianos/metabolismo , Testes de Sensibilidade Microbiana , Conformação Molecular , Mupirocina/biossíntese , Mupirocina/química , Mutação , Policetídeo Sintases/metabolismoRESUMO
Filamentous fungi have the ability to produce a wide range of secondary metabolites some of which are potent toxins whereas others are exploited as food additives or drugs. Fungal natural products still play an important role in the discovery of new chemical entities for potential use as pharmaceuticals. However, in most cases they cannot be directly used as drugs due to toxic side effects or suboptimal pharmacokinetics. To improve drug-like properties, including bioactivity and stability or to produce better precursors for semi-synthetic routes, one needs to generate non-natural derivatives from known fungal secondary metabolites. In this minireview, we describe past and recent biosynthetic approaches for the diversification of fungal natural products, covering examples from precursor-directed biosynthesis, mutasynthesis, metabolic engineering and biocombinatorial synthesis. To illustrate the current state-of-the-art, challenges and pitfalls, we lay particular emphasis on the class of fungal cyclodepsipeptides which have been studied longtime for product diversification and which are of pharmaceutical relevance as drugs.
Assuntos
Aspergillus/metabolismo , Produtos Biológicos , Fungos/metabolismo , Engenharia Metabólica/métodos , Aspergillus/genética , Depsipeptídeos/isolamento & purificação , Descoberta de Drogas/métodos , Fungos/genéticaRESUMO
Enzymes have been used in organic chemistry and biotechnology for 100 years, but their widespread application has been prevented by a number of limitations, including the often-observed limited thermostability, narrow substrate scope, and low or wrong stereo- and/or regioselectivity. Directed evolution provides a means to address and generally solve these problems, especially since recent methodology development has made this protein engineering method faster, more efficient, and more reliable than in the past. This Darwinian approach to asymmetric catalysis has led to a number of industrial applications. Metabolic-pathway engineering, mutasynthesis, and fermentation are likewise enzyme-based techniques that enrich chemistry. This account outlines the scope, and particularly, the limitations, of biocatalysis. The complementary nature of enzymes and man-made catalysts is emphasized.
Assuntos
Técnicas de Química Sintética/métodos , Química Orgânica/métodos , Enzimas/metabolismo , Engenharia de Proteínas/métodos , Biocatálise , Enzimas/genética , Química Verde , EstereoisomerismoRESUMO
BACKGROUND: Sansanmycins are uridyl peptide antibiotics (UPAs), which are inhibitors of translocase I (MraY) and block the bacterial cell wall biosynthesis. They have good antibacterial activity against Pseudomonas aeruginosa and Mycobacterium tuberculosis strains. The biosynthetic gene cluster of sansanmycins has been characterized and the main biosynthetic pathway elucidated according to that of pacidamycins which were catalyzed by nonribosomal peptide synthetases (NRPSs). Sananmycin A is the major compound of Streptomyces sp. SS (wild type strain) and it bears a non-proteinogenic amino acid, meta-tyrosine (m-Tyr), at the N-terminus of tetrapeptide chain. RESULTS: ssaX deletion mutant SS/XKO was constructed by the λ-RED mediated PCR targeting method and confirmed by PCR and southern blot. The disruption of ssaX completely abolished the production of sansanmycin A. Complementation in vivo and in vitro could both recover the production of sansanmycin A, and the overexpression of SsaX apparently increased the production of sansanmycin A by 20%. Six new compounds were identified in the fermentation culture of ssaX deletion mutant. Some more novel sansanmycin analogues were obtained by mutasynthesis, and totally ten sansanmycin analogues, MX-1 to MX-10, were purified and identified by electrospray ionization mass spectrometry (ESI-MS) and nuclear magnetic resonance (NMR). The bioassay of these sansanmycin analogues showed that sansanmycin MX-1, MX-2, MX-4, MX-6 and MX-7 exhibited comparable potency to sansanmycin A against M. tuberculosis H37Rv, as well as multi-drug-resistant (MDR) and extensive-drug-resistant (XDR) strains. Moreover, sansanmycin MX-2 and MX-4 displayed much better stability than sansanmycin A. CONCLUSIONS: We demonstrated that SsaX is responsible for the biosynthesis of m-Tyr in vivo by gene deletion and complementation. About twenty novel sansanmycin analogues were obtained by mutasynthesis in ssaX deletion mutant SS/XKO and ten of them were purified and structurally identified. Among them, MX-2 and MX-4 showed promising anti-MDR and anti-XDR tuberculosis activity and greater stability than sansanmycin A. These results indicated that ssaX deletion mutant SS/XKO was a suitable host to expand the diversity of the N-terminus of UPAs, with potential to yield more novel compounds with improved activity and/or other properties.
Assuntos
Antibacterianos/química , Proteínas de Bactérias/genética , Mutação , Oligopeptídeos/química , Oligopeptídeos/genética , Peptídeo Sintases/genética , Streptomyces/metabolismo , Uridina/análogos & derivados , Antibacterianos/biossíntese , Antibacterianos/farmacologia , Proteínas de Bactérias/metabolismo , Cromatografia Líquida de Alta Pressão , Farmacorresistência Bacteriana Múltipla/efeitos dos fármacos , Espectroscopia de Ressonância Magnética , Testes de Sensibilidade Microbiana , Conformação Molecular , Família Multigênica , Mycobacterium tuberculosis/efeitos dos fármacos , Oligopeptídeos/biossíntese , Oligopeptídeos/farmacologia , Peptídeo Sintases/metabolismo , Plasmídeos/metabolismo , Pseudomonas aeruginosa/efeitos dos fármacos , Espectrometria de Massas por Ionização por Electrospray , Streptomyces/genética , Uridina/biossíntese , Uridina/química , Uridina/genética , Uridina/farmacologiaRESUMO
Penicillium chrysogenum, which lacks the roqA gene, processes synthetic, exogenously added histidyltryptophanyldiketopiperazine (HTD) to yield a set of roquefortine-based secondary metabolites also produced by the wild-type strain. Feeding a number of synthetic HTD analogues to the ΔroqA strain gives rise to the biosynthesis of a number of new roquefortine D derivatives, depending on the nature of the synthetic HTD added. Besides delivering semisynthetic roquefortine analogues, the mutasynthesis studies presented here also shed light on the substrate preferences and molecular mechanisms employed by the roquefortine C/D biosynthesis gene cluster, knowledge that may be tapped for the future development of more complex semisynthetic roquefortine-based secondary metabolites.
Assuntos
Proteínas Fúngicas/genética , Indóis/química , Indóis/metabolismo , Mutação , Penicillium chrysogenum/genética , Penicillium chrysogenum/metabolismo , Compostos Heterocíclicos de 4 ou mais Anéis/química , Compostos Heterocíclicos de 4 ou mais Anéis/metabolismo , Família Multigênica/genética , Piperazinas/química , Piperazinas/metabolismoRESUMO
Myxopyronin is a natural α-pyrone antibiotic from the soil bacterium Myxococcus fulvus Mx f50. Myxopyronin inhibits bacterial RNA polymerase (RNAP) by binding to a part of the enzyme not targeted by the clinically used rifamycins. This mode of action makes myxopyronins promising molecules for the development of novel broad-spectrum antibacterials. We describe the derivatization of myxopyronins by an advanced mutasynthesis approach as a first step towards this goal. Site-directed mutagenesis of the biosynthetic machinery was used to block myxopyronin biosynthesis at different stages. The resulting mutants were fed with diverse precursors that mimic the biosynthetic intermediates to restore production. Mutasynthon incorporation and production of novel myxopyronin derivatives were analyzed by HPLC-MS/MS. This work sets the stage for accessing numerous myxopyronin derivatives, thus significantly expanding the chemical space of f α-pyrone antibiotics.
Assuntos
Antibacterianos/biossíntese , Produtos Biológicos/metabolismo , Mutação , Myxococcus/genética , Myxococcus/metabolismo , Pironas/metabolismo , Antibacterianos/química , Produtos Biológicos/química , RNA Polimerases Dirigidas por DNA/química , RNA Polimerases Dirigidas por DNA/genética , RNA Polimerases Dirigidas por DNA/metabolismo , Modelos Moleculares , Conformação Proteica , Pironas/química , Thermus thermophilus/enzimologiaRESUMO
Streptomyces hygroscopicus is a natural producer of geldanamycin. Mutasynthetic supplementation of an AHBA-blocked mutant with all possible monofluoro 3-aminobenzoic acids provided new fluorogeldanamycins. These showed strong antiproliferative activity and inhibitory effects on human heat shock protein Hsp90. Binding to Hsp90 in the low nanomolar range was determined from molecular modelling, AFM analysis and by calorimetric studies.
Assuntos
Antineoplásicos/química , Antineoplásicos/farmacologia , Benzoquinonas/química , Proteínas de Choque Térmico HSP90/antagonistas & inibidores , Lactamas Macrocíclicas/química , Streptomyces/metabolismo , Antineoplásicos/metabolismo , Calorimetria/métodos , Linhagem Celular Tumoral/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Fluorbenzenos/metabolismo , Fluorbenzenos/farmacologia , Proteínas de Choque Térmico HSP90/metabolismo , Humanos , Espectroscopia de Ressonância Magnética , Microscopia de Força Atômica , Modelos Moleculares , Quinonas/química , Streptomyces/genética , meta-Aminobenzoatos/metabolismo , meta-Aminobenzoatos/farmacologiaRESUMO
Supplementing an AHBA(-) mutant strain of Amycolatopsis mediterranei, the rifamycin producer, with a series of benzoic acid derivatives yielded new tetraketides containing different phenyl groups. These mutasynthetic studies revealed unique reductive properties of A. mediterranei towards nitro- and azidoarenes, leading to the corresponding anilines. In selected cases, the yields of mutaproducts (fermentation products isolated after feeding bacteria with chemically prepared analogs of natural building blocks) obtained are in a range (up to 118 mg L(-1)) that renders them useful as chiral building blocks for further synthetic endeavors. The configuration of the stereogenic centers at C6 and C7 was determined to be 6R,7S for one representative tetraketide. Importantly, processing beyond the tetraketide stage is not always blocked when the formation of the bicyclic naphthalene precursor cannot occur. This was proven by formation of a bromo undecaketide, an observation that has implications regarding the evolutionary development of rifamycin biosynthesis.
Assuntos
Actinomycetales/química , Antibacterianos/biossíntese , Azidas/química , Produtos Biológicos/química , Compostos Bicíclicos com Pontes/síntese química , Hidroliases/síntese química , Complexos Multienzimáticos/biossíntese , Complexos Multienzimáticos/química , Naftalenos/síntese química , Policetídeo Sintases/química , Rifamicinas/química , Rifamicinas/síntese química , Sequência de Aminoácidos , Antibacterianos/química , Compostos Bicíclicos com Pontes/química , Hidroliases/química , Hidroliases/metabolismo , Espectroscopia de Ressonância Magnética , Estrutura Molecular , Naftalenos/química , Policetídeo Sintases/metabolismo , Rifamicinas/biossínteseRESUMO
Isopropylstilbene is a natural product from Photorhabdus luminescens TT01, with multiple biological activities. A mutant deficient in the production of both anthraquinones and cinnamic acid was constructed, thus giving a clean background according to UV detection. This anthraquinone and stilbene deficient (ASD) mutant was used in mutasynthesis experiments to obtain new stilbene derivatives, which were detected by GC-MS. The structures of the new derivatives were confirmed by detailed MS analysis and then chemically synthesised; all of the natural and synthetic compounds were tested against protozoa that cause tropical diseases. Two compounds obtained by mutasynthesis showed the highest activity against Trypanosoma cruzi, the causative agent of Chagas disease, and Leishmania donovani, which causes leishmaniasis.
Assuntos
Photorhabdus/metabolismo , Estilbenos/química , Estilbenos/farmacologia , Tripanossomicidas/química , Tripanossomicidas/farmacologia , Trypanosoma cruzi/efeitos dos fármacos , Antraquinonas/metabolismo , Doença de Chagas/tratamento farmacológico , Cinamatos/metabolismo , Humanos , Mutação , Photorhabdus/química , Photorhabdus/genética , Estilbenos/síntese química , Estilbenos/metabolismo , Tripanossomicidas/síntese química , Tripanossomicidas/metabolismoRESUMO
A combination of mutasynthesis, precursor-directed biosynthesis and semisynthesis provides access to new ansamitocin derivatives including new nanostructured particle-drug conjugates. These conjugates are based on the toxin ansamitocin and superparamagnetic iron oxide-silica core shell particles. New ansamitocin derivatives that are functionalized either with alkynyl- or azido groups in the ester side chain at C-3 are attached to nanostructured iron oxide core-silica shell particles. Upon exposure to an oscillating electromagnetic field these conjugates heat up and the ansamitocin derivatives are released by a retro-Diels-Alder reaction. For example, one ansamitocin derivative exerts strong antiproliferative activity against various cancer cell lines in the lower nanomolar range while the corresponding nanostructured particle-drug conjugate is not toxic. Therefore, these new conjugates can serve as dormant toxins that can be employed simultaneously in hyperthermia and chemotherapy when external inductive heating is applied.
Assuntos
Compostos Férricos/química , Maitansina/análogos & derivados , Nanoestruturas/química , Dióxido de Silício/química , Linhagem Celular Tumoral , Proliferação de Células , Reação de Cicloadição , Febre/induzido quimicamente , Humanos , Magnetismo , Maitansina/biossíntese , Maitansina/química , Estrutura MolecularRESUMO
The recent development of thiopeptide analogues of antibiotics has allowed some of the limitations inherent to these naturally occurring substances to be overcome. Chemical synthesis, semisynthetic derivatization, and engineering of the biosynthetic pathway have independently led to complementary modifications of various thiopeptides. Some of the new substances have displayed improved profiles, not only as antibiotics, but also as antiplasmodial and anticancer drugs. The design of novel molecules based on the thiopeptide scaffold appears to be the only strategy to exploit the high potential they have shown inâ vitro. Herein we present the most relevant achievements in the production of thiopeptide analogues and also discuss the way the different approaches might be combined in a multidisciplinary strategy to produce more sophisticated structures.
Assuntos
Antibacterianos/química , Peptídeos/química , Compostos de Sulfidrila/química , Antibacterianos/síntese química , Antibacterianos/farmacologia , Bactérias Gram-Negativas/efeitos dos fármacos , Bactérias Gram-Positivas/efeitos dos fármacos , Peptídeos/síntese química , Peptídeos/farmacologia , Relação Estrutura-AtividadeRESUMO
The preparation of alkyne-modified ansamitocins by mutasynthetic supplementation of Actinosynnema pretiosum mutants with alkyne-substituted aminobenzoic acids is described. This modification paved the way to introduce a thiol linker by Huisgen-type cycloaddition which can principally be utilized to create tumor targeting conjugates. In bioactivity tests, only those new ansamitocin derivatives showed strong antiproliferative activity that bear an ester side chain at C-3.
RESUMO
Polyketides from (hydroxy)benzoates are an interesting group of plant polyphenolic compounds, whose biotechnological production is so far underrepresented due to their challenging heterologous biosynthesis. Efficient heterologous production of 2,4,6-tri- and 2,3',4,6-tetrahydroxybenzophenone, 3,5-dihydroxybiphenyl, and 4-hydroxycoumarin by whole-cell biocatalysis in combination with in situ product extraction with an organic solvent was demonstrated. Production was highly dependent on the used CoA ligase and polyketide synthase type III. Therefore, different combinations of polyketide synthases and benzoate-CoA ligases were evaluated for their biosynthesis performance in the solvent-tolerant Pseudomonas taiwanensis VLB120. A solvent screening yielded 2-undecanone as biocompatible, extraction-efficient solvent with good phase separation. In aqueous-organic two-phase cultivations, this solvent extraction circumvents product instability in the aqueous cultivation medium, and it increases yields by reducing inhibitory effects. Complete de novo synthesis from glucose of all (hydroxy)benzoate-derived polyketides was achieved in two-phase cultivations with metabolically engineered strains. Additionally, mutasynthesis was applied to obtain fluorinated benzophenone derivatives.
Assuntos
Policetídeos , Benzoatos , Plantas , Pseudomonas , SolventesRESUMO
Bacterial secondary metabolites exhibit diverse remarkable bioactivities and are thus the subject of study for different applications. Recently, the individual effectiveness of tripyrrolic prodiginines and rhamnolipids against the plant-parasitic nematode Heterodera schachtii, which causes tremendous losses in crop plants, was described. Notably, rhamnolipid production in engineered Pseudomonas putida strains has already reached industrial implementation. However, the non-natural hydroxyl-decorated prodiginines, which are of particular interest in this study due to a previously described particularly good plant compatibility and low toxicity, are not as readily accessible. In the present study, a new effective hybrid synthetic route was established. This included the engineering of a novel P. putida strain to provide enhanced levels of a bipyrrole precursor and an optimization of mutasynthesis, i.e., the conversion of chemically synthesized and supplemented monopyrroles to tripyrrolic compounds. Subsequent semisynthesis provided the hydroxylated prodiginine. The prodiginines caused reduced infectiousness of H. schachtii for Arabidopsis thaliana plants resulting from impaired motility and stylet thrusting, providing the first insights on the mode of action in this context. Furthermore, the combined application with rhamnolipids was assessed for the first time and found to be more effective against nematode parasitism than the individual compounds. To obtain, for instance, 50% nematode control, it was sufficient to apply 7.8 µM hydroxylated prodiginine together with 0.7 µg/ml (~ 1.1 µM) di-rhamnolipids, which corresponded to ca. » of the individual EC50 values. In summary, a hybrid synthetic route toward a hydroxylated prodiginine was established and its effects and combinatorial activity with rhamnolipids on plant-parasitic nematode H. schachtii are presented, demonstrating potential application as antinematodal agents. Graphical Abstract.