Levels of insect infestation, insect spatial distribution, and the relationship between the number of insect-damaged kernels (IDK) and the number of insects present in grain samples in three-hopper railcars transporting wheat from country elevators to a mill were studied. Six of eight sampled railcars were infested with more than two species of insects. The most abundant species collected were the lesser grain borer, Rhyzopertha dominica (F.), and rusty grain beetle, Cryptolestes ferrugineus (Stephens), with the larval stage of the two species being the most prevalent (>90%). The spatial distributions of these two species within the grain mass were typically clumped in railcar compartments containing >0.4 insect/2.75-kg sample of wheat, and these foci of high-infestation levels varied in compartments within the railcars and among the sampled railcars. There were no significant correlations between IDK and insect density for any of the different stage-specific insect populations that were collected in the grain samples. Mean numbers of immatures and IDK differed among railcars and compartments within railcars, but not among grain depths. Number of insects in the first discharge sample was not correlated with mean numbers of insects in the entire compartment. This indicates that each compartment of a railcar should be sampled to determine level of insect infestation but that sampling at different depths within a compartment is less important.