Abstract: As a major cash crop, cotton has been widely cultivated for several decades in Xinjiang Uygur Autonomous Region (referred to as Xinjiang). Due to continuous cotton cropping over a long period, soil-borne diseases (e.g., Fusarium sp. and Verticillium dahlia) have worsened in cotton fields in recent years, resulting in substantial yield declines. The objective of this study was to determine the effects of long-term cultivation of cotton on soil biological activity, soil microbial community structure, microbial genetic diversity (e.g., total bacterial and total fungi) and specific soil microbe (e.g., fusarium and bacillus). Three cotton treatments continuous mono-cropping for 5 consecutive years (CtN5), for 10 consecutive years (CtN10) and for 15 consecutive years (CtN15) were set up in an oasis farmland in northern Xinjiang. Then soil peroxidase, invertase, arylsulfatase, dehydrogenase, and protease activities were measured, while soil microbial community structure diversities of total bacteria, total fungi, fusarium soil-borne pathogenic bacteria and bacillus soil antagonistic bacteria were determined using PCR-DGGE fingerprint approach. There was a significantly decreasing tendency in soil catalase, invertase and dehydrogenase activities with increasing years of continuous cropping. For instance, compared with CtN10, CtN15 treatment decreased activities by 15.0% for catalase and by 6.4% for invertase. However, arylsulfatase and protease activities decreased from continuous cotton cropping of 5 years (CtN5) to 10 years (CtN10) followed by an increasing tendency from continuous cotton cropping of 10 years (CtN10) to 15 years (CtN15). The Shannon-Wiener and Simpson indices of soil total bacteria and total fungi declined markedly with increasing years of continuous cotton cropping. Compared with CtN10, the quantity of soil total bacteria gene band (DGGE fingerprint) decreased by 7.41% under CtN15. It decreased by 1.72% from CtN5 treatment to CtN10 treatment. The values of soil total fungal gene band and Shannon-Wiener diversity index were 78 and 3.22 under CtN15 treatment, suggesting respectively 17.02% and 5.29% decreases under CtN15 compared with CtN5. Compared with CtN10 treatment, the Shannon-Wiener and Simpson indices of soil bacillus community increased respectively by 54.8% and 14.5% under CtN15 treatment. Moreover, the amount of fusarium gene and bacillus gene bands as well as the related diversity indices decreased from CtN5 to CtN10 treatment, but then increased from CtN10 to CtN15. In conclusion therefore, soil enzyme activity and microbial community structure and diversity decreased with increasing years of continuous cotton cropping. Long-term, continuous cultivation of cotton had an adverse effect on soil biological characteristics in northern Xinjiang.