Extensive use of antimicrobial agents such as cotrimoxazole has been associated with raising of antimicrobial resistance. Current study is focused on assessing the prevalence of cotrimoxazole resistance  in klebsiella pneumoniae and the frequency of related genes. 155 isolates of klebsiella pneumoniae were collected during Mar.2007 to Apr.2012 from Ilam hospitals and Milad hospital of Tehran. Antibiotic susceptibility test done to screening resistance isolates according to Kirby-Bauer method. sul1, sul2, sul3, dfrA1, dfrA5, and Int1 genes were detected by PCR. Among 155 species, forty isolates (26%) were resistance to cotrimoxazole. Frequency of sul1 gene was 32 isolates (80%) and 24 isolates of dfrA1(60%), none isolates of dfrA5 (0%), 28 isolates of int (70%), 25 isolates of sul2 (62.5%), and no isolates of sul3 (0%) has been detected. 17 (42.5%)  isolates have sul1 and sul2 simultaneously, and 18 (45%) isolates have int1 and dfrA1. 11 isolates have  sul1, sul2,int1 and dfrA1 genes concurrence by 27.5% frequency. Our study shown resistance to  cotromoxazole in klebsiella isolated from Ilam hospitals and Milad hospital of Tehran is moderate and Sul genes have the highest frequency in  resistance isolates.