IMPROVED METHODOLOGY FOR ISOLATION OF ACETOBACTER-DIAZOTROPHICUS AND CONFIRMATION OF ITS ENDOPHYTIC HABITAT

Nitrogen-free, semi-solid defined medium with crystallized cane sugar (100 g/l) supplemented with cane juice (5 ml/l) was the most selective for isolating Acetobacter diazotrophicus. Surveys of A. diazotrophicu s using this medium showed that > 10(3) cells/g fresh wt were present at all sites in all parts of the sugar cane plant and in all trash sam ples examined, reaching up to 10(7)/g. Additional samples, from forage grasses and cereals and from weed species collected within the sugar cane fields, were all negative. Heat treatment (50-degrees-C for 30 mi n) of the sugar cane setts did not affect A. diazotrophicus numbers wi thin the plant. Nitrogenase activity of intact soil-plant systems in p ots planted with heat-treated setts did not respond to inoculation wit h A. diazotrophic us. The endophytic habitat of this diazotroph and it s propagation within the stem cuttings was confirmed.