The purpose of this study was to identify fast and effectively the Nosema disease of bumble bees from Gangwon province in Korea. Bumble bees are crucial pollinators of various crops and microsporidia are the critical infections of these hosts. The origin of the bumble bees is probably being known from the Britain. Also, the following species of bumble bees have been used around the world: Bombus terrestris, Bombus lucorum, Bombus occidentalis, Bombus ignites and Bombus impatiens. When bumble bees are infected with N. bombi, their abdomens can become distended, paralyzed and infected workers and they often become sluggish and die early. We have identified the morphology of the microsporidium by light and electron microscopy and have found that the morphology of the microsporidium is rounded spore morph, with fairly small spores as described before in many other articles. For the specific and sensitive diagnosis of the microsporidian parasite N. bombi in bumble bees, we have developed the improved method of polymerase chain reaction (PCR) conditions for expeditious diagnosis. Two pairs of primers were tested on N. bombi and the related microsporidia Nosema apis and Nosema sp., both of which infect bumble bees and honey bees and further we have verified and analyzed DNA sequence data of N. bombi in bumble bees by using the BLAST server at the National Center for Biotechnology Information.