Laminitis, a condition of the hoof, is one of the most common and devastating conditions that affects the horse (Equus caballus). The most common factors responsible for triggering laminitis are intake of excess grain or exposure to lush pastures where high starch rich diets are consumed, although the connection between the gut and the hoof is not well understood. When horses are fed starch rich diets, like grain commonly found in the diets of race and performance horses, the balance of microbial species present in the digestive tract of the horse become disturbed, leading to lactic acidosis in the horse. The objective of this study was to identify and quantify the fecal microbial populations in horses maintained on various diets. Microbial populations present in the equine hindgut were assayed by using bacterial ribosomal DNA fragments present in fecal samples. Identification and quantification of specific bacterial species, using bacterial primers and florescent probes, can be detected by quantitative real-time PCR (qPCR). This study focused on equine hindgut streptococcal species (EHSS), including Streptococcus lutetiensis, which accounts for approximately 70% of the microbiota present in the hindgut prior to the onset of laminitis. Our results suggest that avoidance of pasture for all laminitic prone horses may need to be reevaluated as the hindgut microbe sensitivity to diet is unique for each individual horse. The use of these assays will be valuable in future work exploring the changing microbial populations present in the equine hindgut.