Resumen de Biological control approaches of meloidogyne spp. In vegetable crops: from application of selected antagonists to suppressive soils
Root-knot nematodes (RKN), Meloidogyne spp., are the most economically important genus of plant parasitic nematodes that cause considerable damage and yield losses of horticultural crops worldwide. RKN management strategies tend to reduce chemical nematicides by encouraging alternative control methods like the use of plants bearing resistance genes (R-genes) and/or by microbe-inducing plant resistance, and the antagonistic potential of soils. In the thesis, two biological control approaches of Meloidogyne spp. were evaluated: 1) the application of selected nematode antagonists, the fungus Pochonia chlamydosporia and the bacteria Bacillus firmus I-1582 (Bf I-1582), to know its ability to induce plant resistance, and 2) the level of soil suppressiveness of vegetable production sites conducted under organic or integrated standards.
Regarding the ability of P. chlamydosporia and Bf I-1582 to induce plant resistance, the results of this thesis provide evidence that two out of five P. chlamydosporia isolates (M10.43.21 and M10.55.6) and Bf I-1582 are able to induce systemic resistance against M. incognita in the susceptible tomato (Solanum lycopersicum) cv. Durinta but not in the cucumber (Cucumis sativus) cv. Dasher II in split-root experiments. In addition, the cardinal temperatures for the Bf I-1582 growth and biofilm formation were determined in order to improve its use in field conditions. Moreover, Bf I-1582 was transformed with GFP to study its effect on nematode eggs and on tomato and cucumber root colonization. In tomato, the number of egg masses and the number of eggs per plant were reduced by M10.43.21 and M10.55.6 P. chlamydosporia isolates and by Bf I-1582. P. chlamydosporia isolates colonized both tomato and cucumber roots, being the M10.43.21 and the M10.55.6 isolates the best root colonizers in tomato and cucumber, respectively. In the case of Bf I-1582, the bacteria colonized endophytically roots of both plants, but highest values were recorded in tomato. The dynamic regulation of genes related to jasmonic acid (JA) and salicylic acid (SA) was determined by RT-qPCR at three different times after nematode inoculation (dani). Bf I-1582 primed tomato plants by both SA and JA at all the times in tomato, but only SA at 7 dani in cucumber. Regarding P. chlamydosporia, isolate M10.43.21, induced the expression of the SA pathway in tomato at 0, 7 and 42 dani. The JA pathway was also up regulated at 7 dani. These results show the similar model of dynamic regulation of these plant hormone pathways related to plant defense mechanisms against the nematode. Also, Bf I-1582 grew and formed biofilm between 15 and 45 ºC, being 35 ºC the optimal temperature. Bf I-1582GFP was adhered to the egg shell and inside the eggs. Also, Bf I-1582GFP colonized root hairs and epidermal cells and some bacteria were found inside the tomato root. In cucumber, few bacteria were observed on epidermal cells and the bacteria were no found inside the root.
In relation to the level of soil suppressiveness of vegetable production it was carried out a study in four organic and two integrated vegetables production standards sites located in north-eastern Spain. The fluctuation both of Meloidogyne population density and fungal egg parasitism were determined during the rotation sequences in two years (2015-2016). Five out of six of these sites were suppressive soils to Meloidogyne spp. The percentage of fungal egg parasitism ranged from 11.2 to 55 % and P. chlamydosporia was the only fungal species isolated from the eggs. In parallel, two tomato pots experiments were carried out using sterilized and non-sterilized soils from each site and inoculated with second-stage juveniles (J2) to achieve a rate of 1 J2 cm-3 of soil. In both, in five of them the number of nematode eggs per plant was reduced in all nonsterilized soils compared to the sterilizes ones. Also, P. chlamydosporia was the only fungal species isolated from parasitized nematode eggs.
