Molecular identification of Oxalobacter formigenes with the polymerase chain reaction in fresh or frozen fecal samples

Objective To develop a simple and rapid polymerase chain reaction (PCR) met hod for detecting Oxalobacter formigenes (which degrades oxalate in the gut ) in fecal specimens from healthy volunteers and patients with urolithiasis , and to determine whether O. formigenes can be detected in frozen or fresh fecal samples. Materials and methods Whole bacterial DNA was isolated directly from fresh and frozen fecal samples obtained from 30 healthy volunteers free from urol ithiasis and from fresh fecal samples obtained from 38 patients with urolit hiasis. Genus-specific oligonucleotide sequences were designed, correspondi ng to homologous regions residing in the oxc gene that encodes for oxalyl-c oenzyme A decarboxylase. A PCR-based assay was used on both fresh and froze n fecal samples, and the nucleotide sequences analysed to confirm oxc. Results A PCR. product of 416 bp encoding the oxc gene was detected in 23 ( 77%) of 30 healthy volunteers free from urolithiasis and in 14 (37%) of 38 patients with urolithiasis. In healthy volunteers, the results of PCR for t he fresh and the frozen samples were identical in each subject. The nucleot ide sequence analysis showed that the sequence of the amplified product was compatible with that of oxc. Conclusion O. formigenes can be identified easily and efficiently using thi s PCR-based detection system. The colonization rate of O. formigenes in pat ients with urolithiasis was significantly lower than that in healthy volunt eers known to be free from urolithiasis. Furthermore, as the PCR-based assa y results in the frozen fecal samples were identical to those from fresh sa mples in each subject, immediate processing of fecal samples may not be nec essary to detect O. formigenes in the clinical setting.