Pierre Baudin

Evaluation of selective media and immunoassays for detection of Xanthomonas albilineans, causal agent of sugarcane leaf scald disease

A selective medium (XAS medium) was developed for the isolation of Xanthamonas albilineans (Ashby) Dowson, which causes leaf scald disease of sugarcane. XAS medium supported high plating efficiencies of the pathogen. The growth rate, morphology, and pigmentation of colonies on the medium were useful differential characteristics to identify the pathogen. XAS medium consisted of a modification of Wilbrinks medium that was supplemented with 5 g of KBr, 100 mg of cycloheximide, 2 mg of benomyl, 25 mg of cephalexin, 30 mg of novobiocin, and 50 mg of kasugamycin per liter. The pathogen was isolated on XAS medium with greater than 98% frequency from symptomatic sugarcane in Florida, Guadeloupe, and the Dominican Republic, and was isolated less frequently from asymptomatic sugacane

Intraspecific genomic variation within Xanthomonas albilineans, the sugarcane leaf scald pathogen.

ABSTRACT To better understand the nature of recent outbreaks of leaf scald disease of sugarcane in a number of sugarcane production regions of the world including Florida, Guadeloupe, Louisiana, Mauritius, Taiwan, and Texas, a study of the worldwide genetic variation of the pathogen was undertaken. A total of 218 strains from 31 geographic locations were examined. Genomic DNA of each strain was digested with the rare cutting restriction enzyme SpeI, and the fragments were separated by pulsed-field gel electrophoresis (PFGE). A total of 102 bands were identified, and 54 different DNA banding patterns (haplotypes) were observed. Eight groups of banding patterns, designated PFGE groups A through H, were consistently detected by visual, principal component, and cluster analyses. Five groups were comprised of multiple haplotypes representing numerous strains, and three were comprised of single haplotypes representing one strain each. The leaf scald outbreaks in Florida, Louisiana, Texas, and possibly Guadeloupe and Taiwan could be attributed to the introduction of strains belonging to PFGE group B. When infection by two strains each of the newly introduced strains (PFGE group B) and those previously present in Florida (PFGE group A) was analyzed in 22 sugarcane cultivars by reisolation 24 weeks after inoculation, a significantly greater mean frequency was detected for PFGE group B strains and no cultivar by PFGE group interaction was observed. Inadvertent dispersal of the pathogen among plants, possibly by means of aerosols or splashing water, was detected in a subsequent experiment. Strains of PFGE group B were recovered from the internal tissues of some plants inoculated with PFGE group A strains and were also found to be epiphytic colonizers of nonsymptomatic, noninoculated plants adjacent to the inoculated plants; whereas strains of PFGE group A were recovered only from plants that had been inoculated with them. Thus, the possibility became more apparent that strain variation might be associated, at least in part, with factors governing plant-to-plant spread of the pathogen in nature.

Intraspecific Variability of Xanthomonas albilineans

Summary Sixty five Xanthomonas albilineans strains originating from 22 countries were analysed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) of whole-cell proteins, indirect-immunofluorescent antibody typing and gas-liquid chromatographic analysis of fatty acid methyl esters (FAMEs). The strains were separated into two major SDS-PAGE protein clusters, three serovars and three FAME groups. SDS-PAGE protein cluster 1 contained 48 strains which represented a world-wide geographic distribution (with the exception of Fiji and tropical Africa), the majority of which (47) belonged to serovar 1. SDS-PAGE protein cluster 2 comprised 17 strains mainly from tropical Africa and Fiji, which belonged either to serovars 2 or 3 (except strain LMG 478). There was less correlation between the grouping based on FAME composition and those based on SDS-PAGE protein profiles or serological properties. The genomic relationship between X. albilineans and other Xanthomonas species was investigated by DNA: DNA hybridization. A DNA relatedness of 97% was found within X. albilineans, whereas low DNA hybridization values (6–37%) were observed between X. albilineans type strain LMG 494 and representative strains from the other 20 Xanthomonas DNA hybridization groups so far determined. Within the genus Xanthomonas, X. albilineans clearly constitutes a well-delineated species.