CARACTERIZAÇÃO GÉNETICA DE POPULAÇÕES DE Heterodera glycines DE MUNICÍPIOS DA REGIÃO DA ESTRADA DE FERRO NO ESTADO DE GOIÁS

Abstract

Heterodera glycines, also known as Soybean Cyst Nematode (SCN), since it is detected in Brazil in the 1991/1992 crop, has been confusing among the phytosanitary problems that make it difficult to obtain high yields in soybean (Glycine max) due to their capacity of dissemination and the high variability of physiological races. To correctly identify the physiological race of this nematode present in the cultivation area is the first step for the correct and safe use of the resistant cultivars in order to avoid the selection pressure of new breeds. In view of this problem, the objective of the present study was the genetic characterization of populations of Heterodera glycines from soybean producing municipalities of the region of the Estrada de Ferro (state of Goiás) in races and also the molecular characterization employing RAPD markers. The studies were conducted at the Goiano IF - Urutaí campus, in Urutaí - GO. Soil samples (in soybean commercial areas) were collected in the municipalities of the region of the Estrada de Ferro and later, a nematological analysis was performed to confirm the presence of NCS in the inoculum to be multiplied in a greenhouse. For the race test, the experiment was arranged in DIC with 30 (NCS) populations x 6 (soybean differentiators) with ten replicates. The seeds of the soybean differentiators (Pickett, Peking, PI88788, PI90763, Hartwig and Lee) were pre-germinated in cell trays. At the moment of the transplanting of these differentiators for tubes (containing only autoclaved sand), inoculation (4000 eggs and juveniles of second stage) of each SCN population was carried out. Thirty days after inoculation, the number of females on each differentiator was calculated to calculate the female index according to the methodology of Riggs & Schmitt (1988). For the molecular characterization, 28 RAPD primers were used and DNA extraction was performed by the 2% CTAB method. It then quantified the DNA by visual comparisons of bands generated on agarose gel (0.8%) stained with ethidium bromide (0.5 mgL-1). The RAPD reactions were assembled to a final volume of 25 μl, composed of 10 mM Tris-HCL (pH 8.3), 50 mM KCl, 2.4 mM MgCl 2, 100 μM of each deoxyribonucleotide, 0.4 μM of the " primer ", 1 U of Taq DNA polymerase enzyme, and 30 ng of DNA. PCR reactions were done in a thermal cycler and the amplified products were separated by horizontal electrophoresis, 1.2% agarose gel containing ethidium bromide, and submerged in TBE buffer (0.09 M Tris-Borate, 0.002 M EDTA). The RAPD data obtained were evaluated binary, assigning values of (1) for presence and (0) absence of the amplified band for each primer. The binary matrix was used to estimate the genetic distance between populations based on Jaccard dissimilarity. The clustering analysis based on the calculated genetic dissimilarity matrix was obtained by the Ward method using the statistical program R version 3.0.3. There were found 9 distinct physiological races of Heterodera glycines in municipalities of the region of the Estrada de Ferro (State of Goiás). In the municipality of Orizona, races 3, 6, 9 and 10 were found. In Vianópolis, races 3 and 10 were found. In Silvânia and Leopoldo de Bulhões, races 3 and 1 were detected in both municipalities. Races 3, 4+, 6, 10 and 14+ were detected in Ipameri. In Campo Alegre de Goiás, he identified races 6, 13 and 14. In Gameleira de Goiás and Catalão, he detected only race 3. Of the 28 RAPD primers, only 10 were polymorphic among the NCS populations and the rest did not amplify or have had low amplification of the DNA fragment. The use of RAPD markers allowed to verify differences in the gene pool that compose different breeds of Heterodera glycines. Five genetically distinct groups were obtained among the population accesses of Heterodera glycines from municipalities of the region of the Railroad, whose genetic variability among them was high.