- Volume 141, Issue 8, 1995

Cryptococcus neoformans is an opportunistic fungal pathogen which preferentially localizes to the inositol-rich environment of the central nervous system. One of its distinguishing traits is its capacity to catabolize inositol. Inositol is a precursor for the synthesis of phosphatidylinositol (PI). This study demonstrated that C. neoformans synthesizes inositol. Three inositolcontaining sphingolipids were identified in C. neoformans: ceramide-(P-inositol)2mannose, ceramide-P-inositol-mannose, and ceramide-P-inositol. These inositol-containing sphingolipids are typical of fungi but not higher eukaryotes. The effect of inositol on the membrane lipid composition of C. neoformans was also examined. In contrast to the nonpathogenic yeast Saccharomyces cerevisiae, neither the PI composition nor the synthesis of methylated phospholipids was altered by exogenous inositol. Hence, C. neoformans appears to have a metabolic mechanism for maintaining a steady lipid composition regardless of the inositol in its environment.

A Rhizobium meliloti strain that forms effective nodules on soybean (Glycine max) and alfalfa (Medicago sativa) is described. The strain, 042B, was isolated from root nodules of alfalfa in Xinjiang Autonomous Region of China. Experiments showed that strain 042B was able to nodulate soybean as effectively as Bradyrhizobium japonicum USDA110, a widely used inoculant strain. Under hydroponic conditions, both strains performed similarly in many respects such as time required for the appearance of nodules, total nitrogenase activity, plant top dry weight and total plant nitrogen. In soil pot experiments, total nitrogen, seed weight and seed nitrogen of soybean plants inoculated with strains 042B or USDA110 were much higher than those of the uninoculated control, but there were no differences between plants with strain 042B relative to strain USDA110. However, when strain 042B was the inoculant, the number of nodules was greater, but the nodules were smaller than those of strain USDA110. Results from ELISA showed that nodule occupancy of strain 042B in soybean ranged from 82-90% and that of strain USDA110 ranged from 78-86%. Strain 042B can grow in the presence of 5% (w/v) NaCl, at 42 °C, and at pH 10·7. Strain 042B was shown to have two large plasmids (molecular sizes 200 kb and > 1000 kb, respectively). The DNA G + C mol% of strain 042B was 63·6. The DNA level of homology between strain 042B and type-strain R. meliloti USDA1002 was 76%, while DNA homologies of strain 042B were 10% and 12% with R. fredii USDA205 and B. japonicum USDA6, respectively.

Roseobacter denitrificans has rough (R)-type lipopolysaccharide, containing 2-keto-3-deoxyoctonate but no heptoses. Its lipid A has a glucosamine-containing, phosphorylated backbone. It contains the rare 3-oxotetradecanoic (3-oxomyristic) acid as the only amide-bound fatty acid and ester-bound 3-hydroxydecanoic acid, this pattern being characteristic for the a 3 subgroup of Proteobacteria. Treatment of the major outer-membrane protein (porin, apparent molecular mass 88 kDa) of Roseobacter denitrificans with EDTA (2 mM, 30°C, 20 min) resulted in the dissociation of the oligomers into monomers (apparent molecular mass 35 kDa). EDTA-sensitive dissociation has so far been observed only within the α-3 subgroup of Proteobacteria. The 12 N-terminal amino acids of the monomers exhibit sequence homology with the porins of Rhodobacter capsulatus, Rhodobacter sphaeroides and Rhodopseudomonas blastica. Renaming of Roseobacter denitrificans as Rhodobacter denitrificans is suggested.

The Bacillus subtilis 168 circular chromosome yielded ten fragments on I-Ceui endonuclease digestion. I-Ceul recognizes a 26 bp sequence that is located within the gene encoding the 235 subunit of the rRNA in Chlamydomonas eugametos, Escherichia coli and Salmonella typhimurium. The precise locations of the I-Ceul sites of the B. subtilis chromosome were determined on a Notl-Sfil physical map by (i) double digestion analyses with I-Ceul and Sfil, (ii)comparison of mutant strains lacking a specific rrn operon, (iii)using an I-Ceul linking clone and (iv) analysis of nucleotide sequence data of some rrn operons. In conclusion, all the I-Ceul sites were located within the B. subtilis rrn operons and the I-Ceul sites were conserved in all the B. subtilis 168 derivatives tested. Thus, variations in size of the I-Ceul fragments must be due to genome alterations. A B. subtilis 168 strain was investigated with I-Ceul. We demonstrated that the aberrant structure was the outcome of the inversion of an ~ 1700 kb DNA segment.