A polymerase chain reaction protocol for the detection of Xanthomonas albilineans, the causal agent of sugarcane leaf scald disease

A polymerase chain reaction (PCR) protocol was developed that amplified a 360-bp DNA product unique to Xanthomonas albilineans (Xa), the causal agent of sugarcane leaf scald disease. The assay utilizes previously described PCR primers that target the intergenic transcribed spacer (ITS) region between the 16S and 23S rRNA genes. Primer pair Ala4/L1 allowed amplification of a 360-bp DNA fragment from 71 Xa strains including representatives of serovars I, II, and III. Fragments of different sizes were also amplified from three unidentified saprophytic bacteria from sugarcane. Xa could be detected at a lower bacterial concentration with the PCR protocol than with a serological dot blot assay. With PCR, as little as 1.25 pg of Xa genomic DNA (125 fg if followed by Southern blot hybridization), or as few as 0 to 5 CFU of Xa per reaction were detected from infected sugarcane sap and leaf diffusate. Five CFU of Xa per reaction were detected from suspension culture. The PCR protocol provides a rapid, reliable, and economical tool for routine detection and identification of Xa.