- Volume 131, Issue 8, 1985

Summary: The colonization of the intestinal tract of suckling mice by Campylobacter jejuni was examined by orally challenging the mice with a wild-type strain and several nonmotile mutant strains which were isolated after treating the wild-type strain with mutagens. The wild-type strain had colonized the lower portion of the small intestine, the caecum and the colon 2 d after inoculation. Two nonmotile strains, one of which (M8) had lost all the flagellar structure including the filament, the hook and the basal structure, and the other (M1) which had lost only the filament region, were both cleared from the intestinal tract 2 d after challenge. Another nonmotile strain (M14), which had a complete flagellar structure like that of the wild-type strain, did not colonize and was cleared from the intestinal tract like the other nonmotile and nonflagellated strains. One atypically motile strain (M5), which had a shorter flagellar filament than that of the wild-type strain, colonized the intestinal tract only when mice were challenged with a large inoculum. None of the mice challenged with either the wild-type or any of the mutant strains showed signs of illness. We concluded that motility is an important factor in the colonization of the intestinal tract of suckling mice by C. jejuni.

Summary: A new type of amphipathic antigen was extracted from whole cells of Streptococcus sanguis ATCC 10557 (biotype B, serotype II) by the phenol/water method. The extract was treated with nuclease P1, and was applied to a column of Sepharose 6B. Each fraction was checked by passive haemagglutination (PHA) and immunodiffusion tests against anti-10557 serum which was obtained by immunizing rabbits with whole cells of strain ATCC 10557. Strong PHA activity was demonstrated in the first hexose-containing peak (peak 1) eluted near the void volume, while the second hexose-containing peak (peak 2) produced a heavy band against anti-10557 serum in an immunodiffusion test. The third peak (peak 3) which partially overlapped with peak 2 reacted with concanavalin A, but not with the antiserum, in agar gel. Peaks 2 and 3 had no PHA activity. Peak 1 contained only 1% phosphorus, indicating that cells of strain ATCC 10557 possess an amphipathic antigen which differs from the lipoteichoic acids that are common in many Gram-positive bacteria. Peak 1 was a fatty acid-substituted heteropolysac-charide composed of glucose, galactose, mannose, glycerol and fatty acids in a molar ratio of approximately 1·0 :1·3 : 2·7 : 0·3 : 1·0. PHA activity was inhibited in the presence of polymerized mannose. Peak 2 was composed of glucose, galactose, rhamnose and N-acetylgalactosamine in a molar ratio of approximately 1·0:1·4:0·8:0·8, which was essentially identical to the serotype II carbohydrate antigen reported previously.

Summary: Vibrio strains isolated from diseased turbot in an experimental fish farm on the Atlantic coast of northwest Spain were identified as Vibrio anguillarum. The isolates shared many biochemical characteristics with V. anguillarum strains obtained from other sources, and harboured a plasmid species that showed extensive homology with plasmid pJM1, carried by V. anguillarum strain 775 isolated from an epizootic in North America. Restriction endonuclease analysis showed that the two plasmids were very similar albeit not identical. The presence of the plasmid in the turbot isolates was associated with their ability to cause disease in fish. Plasmid-carrying bacteria could also grow under conditions of iron limitation. Two outer membrane proteins, of 86 and 79 kDal, were induced, and a similar siderophore activity to that produced by V. anguillarum 775 was also detected under these conditions. The 86 kDal outer membrane protein cross-reacted immunologically with antiserum raised against the outer membrane protein OM2 produced by strain 775. Nonvirulent plasmidless derivatives were unable to grow under iron-limiting conditions, and were also unable to produce either siderophore activity or the 86 kDal outer membrane protein, suggesting the plasmid-mediated nature of these components.

Summary: Tunicamycin is an antimicrobial agent which inhibits the first reaction of the dolichol pathway leading to N-glycosylation of proteins. The effect of tunicamycin on the growth of the dimorphic fungus Candida albicans differed depending on the growth phase of the organism. Addition of tunicamycin to stationary phase yeast cells inhibited the resumption of growth of those cells in either morphology, as cultures failed to initiate either yeast bud or germ tube formation. When tunicamycin was added to growing cells, growth was inhibited but not immediately. When it was added to germ tube cultures, nuclear division and septum formation continued for some time before ceasing. Addition of the drug to exponential phase yeast cultures resulted in an approximately 45% increase in cell number before cell division ceased and yeast accumulated in both budded and unbudded stages of the cell cycle. Accumulation of trichloroacetic acid precipitable radiolabelled protein and nucleic acid continued unchanged for some time following addition of tunicamycin; however, after a while a reduced rate of accumulation was noted.

Summary: The isolation of Nectria galligena mutants resistant to benzoic acid (BA), the phytoalexin of immature apples, is reported. Those in which pathogenicity on mature apples was similar to the wild-type were inoculated in immature fruits. One mutant resistant to BA was more aggressive than the wild-type in young apples at the end of their maturation cycle. Aggressiveness of this mutant and the wild-type could be increased by growing the fungus before inoculation into the fruit on a medium supplemented with BA. Amounts of BA in mutant-infected and wild-type-infected tissues were similar. We conclude that BA is involved in the resistance of immature apples infected by N. galligena.

Summary: Ammonium-grown cultures of Sporobolomyces roseus developed the capacity to assimilate nitrate when they were incubated in nitrate or nitrogen-free medium. Cycloheximide, 6-methyl purine and antimycin A inhibited this process. Nitrate assimilation required aerobic energy metabolism and was inhibited completely by ammonium. The rapid inhibition of nitrate assimilation by ammonium was not the result of an inhibition of nitrate reductase (NR) activity. Nitrite also inhibited nitrate assimilation. NR in cell-free extracts of S. roseus was NADPH-specific and its activity was repressed in cultures containing ammonium and derepressed during nitrogen starvation. Nitrate stimulated the appearance of NR in these cultures. Nitrate assimilation was not limited by apparent (potential) NR activity.

Summary: The existence of energy-dependent copper influx was demonstrated in protoplasts of Penicillium ochro-chloron. Protoplasts and mycelium were tolerant of copper over the pH range 3·0 to 5·5 but sensitive at pH 6·0. Uptake of copper was approximately ten times greater at pH 6·0 than in the lower range. At pH 3·0, influx showed saturation kinetics with a half-maximal influx at an external Cu2+ concentration of 390 μ;M and a maximum influx rate of 22 nmol h−1 (107 cells)−1. Saturation kinetics were not observed at pH 6·0.

Summary: Puromycin is a potent inhibitor of bacterial protein synthesis, but puromycin-producing Streptomyces alboniger KCC S-0309 is tolerant to the antibiotic in vivo. Puromycin bound to both 30S and 50S ribosomal subunits from S. alboniger and inhibited polyuridylate-directed polyphenylalanine synthesis by the ribosomes. However, the organism possessed a novel puromycin-inactivating enzyme which acetylated the antibiotic at the 2″-NH2 group of the O-methyltyrosine moiety.

Summary: Mutants of Rhizobium leguminosarum 300 which were unable to utilize one or more organic acids as growth substrates were obtained by Tn5 mutagenesis. Mutant strain MNF3080 was defective in dicarboxylate transport and was unable to grow on succinate. Strain MNF3085 was defective in phosphoenolpyruvate carboxykinase and hence could not carry out gluconeogenesis. This strain did not grow on pyruvate, succinate, glutamate or arabinose but grew on glucose and on glycerol. Strain MNF3075 was unable to utilize pyruvate; the biochemical lesion in this mutant was not identified. MNF3085 and MNF3075 were symbiotically effective. MNF3080 nodulated peas, but the nodules were ineffective in N2 fixation and displayed morphological abnormalities. These data support previous findings which suggest that utilization of exogenous dicarboxylates is essential for effective nodule development by R. leguminosarum.

Summary: Gluconeogenesis in Rhizobium leguminosarum MNF3841 takes place via the derepressible enzymes phosphoenolpyruvate carboxykinase (PEPCK) and fructose bisphosphate aldolase. A Tn5-induced PEPCK deficient mutant (MNF3085) fails to grow on a wide range of simple carbon sources. PEPCK and fructose bisphosphate aldolase are rapidly derepressed when cells are transferred from a sugar to an organic acid as the sole carbon source. The addition of 0·1 mM-sucrose causes an 80% inhibition of PEPCK and fructose bisphosphate aldolase synthesis, and 0·4 mM-sucrose causes a complete inhibition of PEPCK synthesis. Pea bacteroids of MNF3841 purified on a Percoll gradient contain low levels of PEPCK and fructose bisphosphate aldolase. This bacteroid-associated PEPCK activity is clearly of bacterial not plant origin because of its nucleotide requirement, and the fact that bacteroids of MNF3085 contain no PEPCK activity at all. Although the mutant lacks PEPCK it is still able to nodulate and fix N2 as effectively as the parent. The capacity to synthesize sugars via gluconeogenesis is not required for an effective symbiosis. Furthermore, these data suggest that pea bacteroids receive sufficient sugar to compensate for the gluconeogenic defect in strain MNF3085, but insufficient to completely repress the synthesis of PEPCK in the wild-type.

Summary: The free lipids of a sample of Mycobacterium leprae were extracted by a procedure designed to produce separate non-polar and polar fractions. The composition of these lipids was analysed semi-quantitatively by five special thin-layer chromatographic systems covering the total range of mycobacterial lipid polarities. In order of increasing polarity, the major lipids were dimycocerosates of phthiocerol A, phthiocerol B and phthiodiolone A, glycosyl phenolphthiocerol dimycocerosates and phospholipids, including monoacylphosphatidylinositol di- and pentamannosides. The diacylated forms of these latter lipids, found in most mycobacteria, were not present. The composition of the free lipids of the leprosy bacillus, surveyed over the total polarity range for the first time, showed that the patterns were particularly related to those of Mycobacterium bovis, Mycobacterium kansasii and Mycobacterium marinum.

Summary: The mycolic and fatty acids of three samples each of Mycobacterium leprae and Mycobacterium gordonae were compared. Acids released by whole-organism alkaline hydrolysis were converted to 4-nitrobenzyl esters and mycolic acids were further derivatized to t-butyldimethylsilyl ethers. Thin-layer chromatography of the derivatized long-chain extracts showed that all three M. leprae preparations contained so-called α-mycolates and ketomycolates but that the M. gordonae samples had a methoxymycolate in addition to the above types. Silica gel normal-phase high-performance liquid chromatography of the total mycolic acid derivatives confirmed the lack of detectable amounts of methoxymycolates in M. leprae and reverse-phase chromatography of the individual mycolate types demonstrated the homogeneity of the chain lengths of the mycolic acids in each species. Non-hydroxylated fatty acid 4-nitrobenzyl esters were transformed to methyl esters and examined by gas chromatography. Tuberculostearic (10-methyloctadecanoic) acid was a major component of the lipids of all three M. leprae preparations but it was absent in one M. gordonae strain and a very minor component in the other representatives of this latter species. On the basis of fatty and mycolic acid compositions, therefore, a previously suggested close relationship between M. leprae and M. gordonae was not supported.

Summary: Representative strains of coagulase-positive and coagulase-negative staphylococci were degraded by acid methanolysis and the resultant fatty acid methyl esters analysed by gas chromatography. The quantitative data obtained were examined by cluster analysis. The coagulase-positive strains formed six major and one single-member cluster at the 90% S-level. The Staphylococcus intermedius aggregate cluster included the single-member cluster and major clusters 1 and 2. The four remaining clusters contained S. aureus strains and were homogeneous and distinct. The coagulase-negative strains were recovered in ten major and three single-member clusters at the 90% S-level. Five of the ten major clusters were reasonably homogeneous with respect to the existing classification. Thus, three S. capitis strains and five of the six S. epidermidis strains, two of the three S. hominis strains and five of the six S. simulans strains were recovered in separate clusters. Cluster 7 was divided into two subclusters; one contained five of the six S. hyicus strains and the other contained the two representatives of S. lentus. The remaining clusters were heterogeneous with regard to the named strains they contained.

Summary: Use of a pyruvate-based medium in which the supply of combined N limited growth enabled detection of diazotrophy (as C2H2 reduction) in batch cultures of 12 out of 15 strains of Desulfovibrio representing 5 species and including 2 strains hitherto believed to be non-diazotrophic.