Polymerase chain reaction (PCR) and newly designed primers, XAF1/XAR1, were tested for selective detection of the causal agent of leaf scald of sugarcane, Xanthomonas albilineans. The efficiency and reliability of PCR were compared with dot immunobinding assay (DIA), ELISA and classical isolation techniques for detecting X. albilineans in suspensions of pure cells and extracts of field-collected stalk and leaf samples of sugarcane. In addition, classical PCR and BIO-PCR (biological amplification followed by PCR) were compared with isolation on a semiselective agar medium. Classical PCR and BIO-PCR techniques had the advantage of not requiring pathogenicity tests to confirm the identity of colonies tentatively identified as X. albilineans on modified semiselective XAM agar medium. The m-XAM medium and BIO-PCR techniques were the most sensitive; however, the former required seven days whereas the latter required only four days. The BIO-PCR technique was as sensitive as the semiselective medium technique and eliminated the need to conduct any additional tests to confirm the identification.