Development of a new semiselective medium for isolating Xanthomonas campestris pv. manihotis from plant material and soil

An effective control for bacterial blight of cassava (Manihot esculenta), c aused by Xanthomonas campestris pv, manihotis, requires the use of non cont aminated cuttings and seeds. Using classical agar plating techniques for sc reening planting material for contamination has not been very successful be cause of the lack of a reliable semiselective agar medium. The pathogen gro ws slowly on general plating media and is easily overgrown by saprophytic b acteria during isolation from diseased plants. In an effort to develop a se miselective medium, the utilization of several carbon and nitrogen sources was studied. Results of these tests provided information used to design a b asal medium allowing good growth of the target organism while suppressing g rowth of several common saprophytes. Additional selectivity was achieved by incorporating three antibiotics into the basal medium. The new semiselecti ve agar medium, designated cefazolin trehalose agar (CTA) medium, contained (per liter) 3.0 g of K2HPO4, 1.0 g of NaH2PO4, 0.3 g of MgSO4. 7H(2)O, 1.0 g of NH4Cl, 9.0 g of D(+)-trehalose, 1.0 D(+)-glucose, 1.0 g of yeast extr act, 0.025 g of cefazolin, 0.0012 g of lincomycin, 0.0025 g of phosphomycin , 0.25 g of cycloheximide, and 14.0 g of agar. In comparison to a starch-ba sed semiselective medium (SXM), plating efficiencies using pure cultures of 10 strains of X. campestris pv. manihotis were significantly higher on CTA , with an average of 85 and 50%, respectively. Likewise, isolation and reco very of X. campestris pv. manihotis from infected cassava leaves and contam inated soil were much higher on CTA than on SXM agar. When X, campestris pv . manihotis occurs in high concentrations in diseased tissue, the standard yeast trehalose glucose agar medium supplemented with 250 mu g of cyclohexi mide per mi appears to be satisfactory. The newly developed CTA medium shou ld prove useful for control strategies to identify and remove infected plan ting material of cassava, as well as for basic ecological studies of the pa thogen.