Alexandro S. Nunes

Purification and identification of metabolites produced by Bacillus cereus and B. subtilis active against Meloidogyne exigua, and their in silico interaction with a putative phosphoribosyltransferase from M. incognita

To contribute to the development of products to control Meloidogyne exigua, the bacteria Bacillus cereus and B. subtilis were cultivated in liquid medium to produce metabolites active against this plant-parasitic nematode. Fractionation of the crude dichloromethane extracts obtained from the cultures afforded uracil, 9H-purine and dihydrouracil. All compounds were active against M. exigua, the latter being the most efficient. This substance presented a LC50 of 204 µg/mL against the nematode, while a LC50 of 260 µg/mL was observed for the commercial nematicide carbofuran. A search for protein-ligand complexes in which the ligands were structurally similar to dihydrouracil resulted in the selection of phosphoribosyltransferases, the sequences of which were used in an in silico search in the genome of M. incognita for a similar sequence of amino acids. The resulting sequence was modelled and dihydrouracil and 9H-purine were inserted in the active site of this putative phosphoribosyltransferase resulting in protein-ligand complexes that underwent molecular dynamics simulations. Calculation of the binding free-energies of these complexes revealed that the dissociation constant of dihydrouracil and 9H-purine to this protein is around 8.3 x 10-7 and 1.6 x 10-6 M, respectively. Consequently, these substances and the putative phosphoribosyltransferase are promising for the development of new products to control M. exigua.

Changes in metabolites in plant roots after inoculation with Meloidogyne incognita

The study of the metabolic variations in plant roots up to 96 h after their inoculation with second-stage juveniles (J2) of Meloidogyne incognita revealed that soybean cultivars resistant to this nematode produced more soluble phenols and alkaloids. In tomato plants the resistance to M. incognita correlates with the production of soluble phenols, the concentrations of which were always higher in the resistant cultivar. For common bean plants the production of soluble carbohydrates, especially sucrose, increased after their inoculation. However, the extracts of roots from plants resistant to M. incognita did not increase the in vitro mortality of J2.

Effect of plant and fungous metabolites on Meloidogyne exigua

As nematodes cause great damage to Brazilian coffee production, effective methods to control these parasites are necessary. In a previous work Allium cepa L., Cajanus cajan (L.) Mill., Crotalaria juncea L., Ficus elastica Roxb., Ruta graveolens L., Stylosanthes guianensis Aubl., Leucaena leucocephala (Lam.) Dewit., Brachiaria decumbens Stapf., Catharanthus roseus G. Don, Tagetes minuta L., Ricinus communis L. and Coffea arabica L. produced active substances against Meloidogyne exigua Goeldi, a nematode widely disseminated through Brazilian coffee fields. Thus, aqueous extracts of such plants, collected in a different season from that of the previous work, as well as crude metabolites produced in liquid medium by Fusarium moniliforme Shelden and Cylindrocarpon magnusianum (Sacc.) Woll., were submitted to in vitro assays with M. exigua second-stage juveniles (J2). All plants and fungi produced active substances against J2. Therefore, their metabolites were applied to six-month-old coffee plants inoculated with M. exigua. After 90 days in a greenhouse, those samples obtained from A. cepa, L. leucocephala, R. graveolens and F. moniliforme inhibited the production of galls and eggs by M. exigua, demonstrating potential to control such parasite.