Osmotic adaptation by gram-negative bacteria: possible role for periplasmic oligosaccharides.

The cyclic (1----2)-beta-D-glucans produced by species of Agrobacterium and Rhizobium resemble the membrane-derived oligosaccharides of Escherichia coli in their periplasmic localization, intermediate size, and (1----2)-beta-D-glucan backbones. The regulation of the biosynthesis of cyclic (1----2)-beta-D-glucan by Agrobacterium tumefaciens is now shown to parallel the osmotic regulation of membrane-derived oligosaccharide biosynthesis in Escherichia coli. This result suggests a general role for periplasmic oligosaccharides in the osmotic adaptation of Gram-negative bacteria as ecologically diverse as enteric and soil bacteria.

Human monoclonal antibodies that recognize conserved epitopes in the core-lipid A region of lipopolysaccharides.

Epstein-Barr virus (EBV)-transformed human B lymphocytes were fused with a murine-human heteromyeloma to produce stable hybrid cell lines that secreted human monoclonal antibodies (mAbs) of the IgM class that recognized conserved epitopes in the core-lipid A region of lipopolysaccharides (LPS). Three of the mAbs reacted with epitopes on the lipid A moiety, while a fourth recognized a determinant in the core oligosaccharide. The lipid A-specific mAbs cross-reacted with heterologous rough LPS and with lipid As released by acid hydrolysis of different intact (smooth) LPS. Carbohydrate groups in the O-side chain and core oligosaccharide of isolated, smooth LPS restricted antibody access to antigenic sites on lipid A. Yet, one lipid A-reactive mAb recognized its epitope on the surfaces of a variety of intact bacteria. These findings confirm the presence of highly conserved epitopes in the core-lipid A complex and prove the existence of human B cell clones with the potential for secreting high avidity IgM antibodies that react with these widely shared determinants. Such human mAbs might provide protective activity against disease caused by diverse gram-negative bacteria.

Crystallization and preliminary X-ray analysis of porin from Rhodobacter capsulatus.

Porin monomers of the phototrophic bacterium Rhodobacter capsulatus were purified. Crystals were obtained from a solution of porin solubilized with the detergent octyltetraoxyethylene within 5 days using the vapor phase equilibration technique. The crystals were rhombohedral with an edge length of 0.4 mm. The space group was trigonal R3 with unit cell constants of a = b = 95 A, c = 147 A. Reflexions were observed to 3.2 A.

Cat scratch disease. Identification of bacteria in seven cases of lymphadenitis.

A retrospective study of lymph node biopsy specimens from nine patients with the clinical findings and histologic features of cat scratch disease was undertaken to determine whether the recent report by Wear et al. that pleomorphic bacteria are present in the lymph nodes of cat scratch disease could be confirmed. In seven of our nine cases, pleomorphic bacteria were demonstrated with the Warthin-Starry (WS) silver stain. These were gram-negative with the Brown-Hopps tissue Gram stain and were almost at the limit of microscopic resolution. Lymph node specimens from 13 additional patients with nonspecific lymphadenitis who had neither clinical nor histologic findings of cat scratch disease were studied similarly; in none of these were bacteria demonstrated with the WS silver stain. After examining the distribution of the organisms and the related morphologic features in cat scratch disease, we conclude that demonstration of pleomorphic, gram-negative, WS-positive bacteria in the appropriate clinical and histologic setting can firmly establish the diagnosis of cat scratch disease.

Rapid diagnosis of bacterial meningitis by the detection of a fatty acid marker in CSF with gas chromatography-mass spectrometry and selected ion monitoring.

A chemical marker of bacterial meningitis was sought by comparing derivatives of sterile cerebrospinal fluid (CSF) with cultures of organisms in spinal fluid and artificial media. The technique of gas chromatography-mass spectrometry with selected ion monitoring (GC-MS-SIM) was used, optimised for the analysis of fatty acids. Twenty candidate ions were screened, and an ion of mass: charge ratio (m/e) 268 was chosen for detection in clinical specimens. The origin of this marker is unknown, but it is probably the molecular ion of a C16:1 fatty acid. In 135 clinical specimens of CSF examined, the m/e 268 ion was found to be a useful marker for the common organisms that cause bacterial meningitis, giving a sensitivity of 88% and a specificity of 98%. The method was more rapid and more sensitive than conventional microscopy and culture, but CSF containing coagulase-negative staphylococci, Mycobacterium tuberculosis, Cryptococcus neoformans and some other uncommon pathogens gave inconsistent results. Many organisms produced characteristic ion profiles with multiple-ion monitoring, and this method of chemical analysis holds promise for the rapid diagnosis of bacterial infections to genus or species level.

Are ECG Welsh cup electrodes effectively cleaned?

Re-usable Welsh electrocardiograph (ECG)-electrodes are potential vehicles for cross infection. This study confirmed that in-use ECG-electrodes are frequently contaminated with organisms such as coagulase-negative staphylococci and Gram-negative bacilli. The efficacy of five cleaning procedures was evaluated. Immersing the electrodes in water at 60 degrees C for one hour was the most effective method of decontamination tested, following challenge with a standardised suspension of Staphylococcus saprophyticus. Significant contamination persisted following simple cleaning measures. It is suggested that this was promoted by the inadequate removal of electrode gel which provided a protective environment for microorganisms.

Responses of equine neutrophils to contagious equine metritis organism and its lipopolysaccharides.

Morphology and function of equine neutrophils were evaluated after combination with contagious equine metritis organism (CEMO) or 1 of 2 CEMO lipopolysaccharides (LPS). The 2 LPS (LPS-a; LPS-p) isolated from the CEMO contained 14- and 16-carbon fatty acids, ketodeoxyoctanate, hexose, and heptose, but were morphologically distinct. Neutrophils exposed to LPS had fewer granules, whereas those exposed to CEMO had more granules than did the controls (phosphate-buffered saline solution). Neutrophil iodination was significantly increased with 10 and 25 micrograms of LPS-a, but not significantly altered by LPS-p or CEMO. Staphylococcus aureus ingestion was not influenced by CEMO and was mildly decreased by LPS-a. These results indicate that CEMO may have at least 2 functionally and morphologically distinct, but chemically similar, LPS and that 1 of these LPS (LPS-a) may enhance neutrophil killing by stimulating neutrophil iodinating mechanisms.

Characterization of eugonic fermenters group EF-4 by polyacrylamide gel electrophoresis and protein immunoblot analysis.

Whole-cell lysates and proteinase K-extracted lipopolysaccharide (LPS) of 19 strains of the group eugonic fermenter-4 (EF-4) were analyzed by electrophoresis and protein immunoblotting. These strains were isolated from dog- and cat-bite abscesses in human beings, ferret and human gastric lesions, and cat-lung infections. These strains represent 2 biovar groupings; EF-4a biovars ferment glucose and possess arginine dihydrolase activity, whereas EF-4b biovars do not. Electrophoresis of whole-cell lysates could distinguish between these biovars groups. Electrophoresis of LPS extracts revealed that all strains of EF-4 possess smooth chemotypes. Two strains of EF-4a reacted weekly in protein immunoblots and revealed distinct LPS profiles. These studies suggests that subgroups of EF-4 biovars may exist.

[Lipopolysaccharide-protein complexes of the outer membrane of gram-negative bacteria]. / Lipopolisakharid-belkovye kompleksy vneshnei membrany gramotritsatel'nykh bakterii.

The evidence for occurring lipopolysaccharide-protein complexes in the outer membrane of gram-negative bacteria has been summarized. The composition and supramolecular structure of these complexes as well as their functions in microbial envelope and substantial role in membrane organization have been discussed. The biological properties of the complexes as endotoxins and O-specific antigens have been considered.

[Placental smears in a maternity ward. Importance of early diagnosis of neonatal bacterial infections caused by materno-fetal contamination]. / Frottis placentaires en maternité. Intérêt pour le diagnostic précoce des infections bactériennes néonatales par contamination materno-foetale.

Direct cytobacteriological investigation of the amniochorionic plate of the placenta has been used systematically in the Maternity Hospital as a test to detect amniotic infection. This test was practised on fresh placentae in the laboratory of Histology, preceding the examination of the placenta. The test gave, within 30 minutes of delivery, information about the presence of bacteria of fungi on the amniotic surface, their abundance and their morphology. The results of the placental smears were compared to the results of bacteriological cultures obtained from the placenta, the infant and occasionally the mother. The diagnosis of neonatal infection was established from clinical, biological and bacteriological criteria which were defined in advance. Two series are presented: the first concerns 2,514 cases selected in the delivery room over a period of 4 years for high risks infection, i.e. 1 in 6 of the 15,377 deliveries registered during this period. The rate of positive smears was 9%: 63% Gram positive, 17% Gram negative, and 20% mixed. The most common bacteriologically proven neonatal infections were streptococcus B (40%) and Escherichia coli (33%). The second is a prospective survey of 400 unselected consecutive cases observed during a period of 7 weeks: 1% of the smears were positive - and the test would have been justified in 1 in 5 cases. From these two surveys, it appears that placental smear is a good test to detect maternofetal contamination. The positive predictive value of the test is 80%. The negative predictive value is 98% and approaches 100% if cases where the mother received antibiotics before delivery are excluded.

[Improved method of lipopolysaccharide isolation from gram-negative bacteria]. / Uluchshennyi metod vydeleniia lipopolisakharidov iz gramotritsatel'nykh bakterii.

A modification of the traditional method for lipoplysaccharide isolation from the cells of grammnegative bacteria was elaborated on the basis of extraction by the hot water solution of phenol (the method of Westfahl). To make the method simpler and to raise the yield of the product it was proposed to use the water-phenol extract without its division for plases. The nucleic acids are eliminated by precipitation from dialyzed extract at pH 3,2-3,4 achieved by addition of acetic acid. The comparative isolation of lipopolysaccharides by the classic and modified methods has confirmed the advantages of a new technique.

Simple, rapid, and quantitative release of periplasmic proteins by chloroform.

We introduce a method by which periplasmic proteins can be released rapidly, simply, and quantitatively by treating cells with chloroform. All the amino acid-binding proteins tested maintained their activity during chloroform treatment. This method makes practical the analysis of the periplasmic protein complement of a large number of strains.

Bacterial cell walls and membranes. Discovery of the teichoic acids.

Teichoic acids are major wall components of most Gram-positive bacteria. Their discovery followed that of their nucleotide precursors. Lipoteichoic acids associated with the cell membrane were discovered at the same time. Events leading to these discoveries and the probable function of teichoic acids in cation control are described.

Peptidoglycan-polysaccharide complex in the cell wall of the filamentous prochlorophyte Prochlorothrix hollandica.

A peptidoglycan-polysaccharide complex composed of N-acetylglucosamine, N-acetylmuramic acid, muramic acid 6-phosphate, L-alanine, D-alanine, D-glutamic acid, meso-diaminopimelic acid, N-acetylmannosamine, mannose, galactose, glucose, and phosphate was isolated from cell walls of the filamentous prochlorophyte Prochlorothrix hollandica; this complex was similar in chemical composition and structure to that found in cyanobacteria. Peptide patterns of partial acid hydrolysates of the isolated peptidoglycan revealed an A1 gamma structure with direct cross-linkage (m-diaminopimelic acid-D-alanine) of the peptide side chains. The degree of cross-linkage (63%) was found to be in the range of values obtained for gram-positive bacteria and cyanobacteria.

The structure of a novel polysaccharide isolated from Zymomonas mobilis determined by nuclear magnetic resonance spectroscopy.

A novel polysaccharide has been observed in vivo by 13C nuclear magnetic resonance (NMR) spectroscopy of the gram-negative bacterium, Zymomonas mobilis. The polysaccharide, which was not removed from the Z. mobilis cell by washing with 0.86% saline, was extracted by mild acid treatment. The structure was elucidated by a combination of NMR techniques, including proton and proton-carbon two-dimensional methods. The structure was determined to be -alpha-fructofuranosyl-(2-1)-beta-fructofuranosyl-(2-6)-. The polysaccharide is unlike levan, a beta-(2-6)-fructose polymer which is produced by many bacteria, including Z.mobilis, when grown on sucrose. The polysaccharide isolated here is produced by cells cultured on glucose, fructose or sucrose.