ABSTRACT
Background: Fusobacterium nucleatum (Fn) is a common resident of the human GI tract and has been recognized as an opportunistic pathogen implicated in inflammatory bowel disease (IBD). Few studies had focused on the quantitative analysis of Fn in IBD patients. Aims: To establish a novel absolute quantitative real-time PCR method for detection of Fn in fecal samples of Crohn's disease (CD) patients and to study the correlation between fecal Fn and the common inflammatory indicators of CD. Methods: Fecal samples of 57 CD patients and 41 healthy subjects from Suzhou Municipal Hospital were collected and the genomic DNA was extracted. NusG (transcription antitermination protein) of Fusobacterium nucleatum subsp. nucleatum ATCC 25586 was constructed into pUC57 to form a standard plasmid. An absolute quantitative standard curve was built by detecting the gradient diluted standard plasmid via SYBR Green real-time PCR method. The sensitivity, specificity and stability of this novel method were assessed, and then the novel method was used to detect the abundance of Fn in fecal sample of CD patients and healthy controls. Correlations between fecal Fn and fecal calprotectin (FC), C-reactive protein (CRP) and erythrocyte sedimentation rate (ESR) were analyzed. Results: The absolute quantitative real-time PCR method for the detection of fecal Fn established in this study was sensitive, specific and stable. The detection rate and mean abundance of fecal Fn were significantly higher in CD patients than in healthy controls (P=0.034; P=0.039). Fecal Fn abundance in CD patients was positively correlated with FC level (r=0.459, P=0.011). Conclusions: The absolute quantitative real-time PCR established in this study is a promising method for human fecal Fn detection. In Suzhou, Jiangsu Province, the detection rate and abundance of fecal Fn are significantly increased in CD patients. Fecal Fn abundance in CD patients is correlated with FC level, and may reflect the disease activity.