New semiselective medium for isolation of Xanthomonas campestris pv. vesicatoria
A new semiselective medium (CMC-E) for isolation of Xanthomonas campestris pv. vesicatoria (EPPO A2 organism) has been developed in the USA. In plating efficiency tests the CMC-E medium was more efficient than Tween medium B agar or nutrient agar. With the CMC-E medium X. c. pv. vesicatoria was successfully isolated from pepper seed, irrigation water and air samples.
The new medium is composed and prepared (for 1 liter of CMC-E) as follows:
1,5 g KH2PO4,
6,0 g K2HPO4,
0,2 g KCL,
1,0 g MgSO4.H2O,
1,0 g yeast extract,
2,0 g eosin Y,
0,4 g methylene blue,
5,0 ml stock solution of minor elements
1 drop of anti-bubble agent
all dissolved in deionized water in a Waring blender at low speed.
3,0 g agar and 26,0 g high-viscosity carboxymethyl cellulose sodium salt (CMC) are added to the solution in the blender which has to be set to high speed to prevent lumping.
The preparation must be sterilized at 121ø C for 15 min..
2,0 g l-1 gelatine are sterilized in seperate 100 ml of the basal medium (without CMC and agar) and added to the medium immediately after the sterilization.
ÿ64 mg l-1 cephalexin dissolved in 24 ml sterile distilled water at 80-90ø C,
ÿ12 mg l-1 5-fluorouracil dissolved in 24 ml sterile distilled water,
100 mg l-1 bacitracin dissolved in 20 ml sterile distilled water,
ÿ10 mg l-1 neomycin sulfate dissolved in 10 ml sterile distilled water,
100 mg l-1 cycloheximide dissolved in 2 ml ethanol and
ÿÿ1 ml tobramycin (concentration 0,4 mg ml-1) are filter sterilized and aseptically
added to the basal medium immediately after sterilization.
The resulting 1 l medium must be poured into the plates immediately to prevent clumping.
Sources
Gitaitis, R.D.; Chang, C.J.; Sijam, K.; Dowler, C.C. (1991) A differential medium for semiselective isolation of Xanthomonas campestris pv. vesicatoria and other cellulolytic Xanthomonads from various natural sources.
Plant Disease 75, 1274-1278.