The fungus Magnaporthe oryzae presents high variability in virulence which can vary depending on the wheat genotypes. There is also a intense demand for sources of resistance to wheat blast. Since 2016, the Aegilops ventricosa 2NS chromosomal segment in wheat genotypes has been used as the sole source of blast resistance. The objectives of this study were to: a) characterize and compare the virulence of M. oryzae isolates in Brazil based on the reaction of a set of wheat genotypes; b) compare the wheat genotypes in relation to the reaction to M. oryzae infection on seedlings and detached heads; and c) relate the presence of the chromosomal segment 2NS of A. ventricosa in wheat genotypes and resistance to blast. Seedlings and detached heads of wheat genotypes were used in experiments conducted under controlled conditions in which individualized suspensions of conidia of 91 wheat M. oryzae isolates were used in inoculation procedures. Presence of the 2NS chromosomal segment in wheat genotypes was determined via PCR. It was verified that M. oryzae virulence groups are widely distributed in the country and that, due to the high resistance to blast demonstrated, Santa Fe wheat cultivar, carrying 2NS chromosomal segment, plays a prominent role as a source of resistance to blast. However, the presence of the 2NS chromosomal segment in the genome does not necessarily associated with resistance to blast. The evaluation of wheat M. oryzae isolates for virulence generates a very distinct classification patterns depending on whether the object of infection analysis is leaf or head.