In recent years, the number of wheat, Triticum aestivum L., fields heavily infested by Hessian fly, Mayetiola destructor (Say), has increased in the Great Plains of the United States. Historically, resistance genes in wheat have been the most efficient means of controlling this insect pest. To determine which resistance genes are still effective in this area, virulence of six Hessian fly populations from Texas, Oklahoma, and Kansas was determined, using the resistance genes H3, H4, H5, H6, H7H8, H9, H10, H11, H12, H13, H16, H17, H18, H21, H22, H23, H24, H25, H26, H31, and Hdic. Five of the tested genes, H13, H21, H25, H26, and Hdic, conferred high levels of resistance (> 80% of plants scored resistant) to all tested populations. Resistance levels for other genes varied depending on which Hessian fly population they were tested against. Biotype composition analysis of insects collected directly from wheat fields in Grayson County, TX, revealed that the proportion of individuals within this population virulent to the major resistance genes was highly variable (89% for H6, 58% for H9, 28% for H5, 22% for H26, 15% for H3, 9% for H18, 4% for H21, and 0% for H13). Results also revealed that the percentages of biotypes virulent to specific resistance genes in a given population are highly correlated (r2 = 0.97) with the percentages of susceptible plants in a virulence test. This suggests that virulence assays, which require less time and effort, can be used to approximate biotype composition.