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Minimizing pathogenic bacteria, including spores, in indoor air.

Five experiments were conducted to assess whether aerosolized bacteria, including spores, respond like particulate contaminants to the primary (electrical) forces that control the distribution of small particulate contaminants in indoor air. Such response would suggest an approach to minimizing infection in offices, hospitals, nursing homes, and other facilities. It also would have implications for protection against intentionally introduced pathogenic bacteria, including spores. The experiments used two different genera and five different strains of bacteria, including spores. Micrococcus luteus was used as a surrogate for Gram-positive cocci, because M. luteus is similar in size, shape, and cell wall composition to staphylococci, streptococci, and enterococci. Similarly, spore-forming and vegetative Bacillus subtilis were used as surrogates for Gram-positive bacilli such as Bacillus anthracis. The experiments were conducted in a dedicated aerosol physics test facility with culture-based measurements made at timed intervals. The results indicate that the organisms do respond like particulate contaminants to typical electrical forces in a room.

Aerosols↗

Bioactive actinomycetes from Krishna River sediments of Andhra Pradesh.

Sediment samples from Krishna River at Nagayalanka of Andhra Pradesh, India were investigated as a source of actinomycetes to screen for the production of novel bioactive compounds. During our investigation on fresh water actinomycetes from 5 different river sediment samples, a total of 80 actinomycetes were isolated. Out of these 80 isolates, 30 isolates which showed distinct macromorphological characteristics were selected. The antimicrobial and enzymatic activities were studied for all the 30 isolates. The preliminary study for antimicrobial activity by cross streak method indicated that 16 isolates (53.3%) have excellent antagonistic properties. All these 16 isolates were subjected to detailed submerged fermentation studies. It was observed that 12 isolates (40.0%) exhibited antibacterial activity, 9 isolates (30.0%) showed antifungal activity while 5 isolates (16.6%) showed both antibacterial and antifungal activities. All the 30 isolates were also subjected for the determination of enzymatic activities 25 isolates (83.3%) exhibited amylolytic activity while 27 isolates (90.0%) showed proteolytic activity. Among these isolates, six promising isolates were selected for detailed morphological, cultural, physiological and biochemical studies. It was established that these isolates belong to the Streptomyces genus by virtue of their cell wall composition pattern and were identified as strains of different Streptomyces species like S. rochei, S. alanosinicus, S. erumpens, S. griseoplanus, S. gancidicus and S. nigrogriseolus.

Actinobacteria↗

[Studies on the classification of thermophilic actinomycetes. IV. Determination of thermophilic Streptomyces hygroscopicus group].

The thermophilic streptomyces hygroscopicus is one of the pathogens of farmer's lung disease. The identified 60 strains of thermophilic streptomyces hygroscopicus coming from the isolates from the haystack, moldy hay and sputa collected from Jiangsu, Hubei province and Shanghai in China. These strains are grown around 50 degrees C and have moist patches on the surface of colonies. It is proved that protease can be extracted from cultured H9-4 strain. This protease is provided with antigen, the farmer's lung disease of rabbit can be induced in animals experiments. On clinical diagnosis the farmer's lung disease can be detected by sera test. From the identification of the 60 thermophilic streptomyces hygroscopicus strains, we found the morphological cultural, physiological characteristics and cell wall composition of H9-4 and T562 well different from description hitherto. So two of them are identified as new species. H9-4 is named as Streptomyces thermoendus sp. nov. and T562 is named as Streptomyces thermobicorno-hygroscopicus sp. nov.

Animals↗

Luciferin-luciferase assay of adenosine triphosphate from bacteria: a comparison of dimethylsulphoxide (DMSO) and acetone with other solvents.

The extraction of adenosine triphosphate (ATP) from six strains of bacteria, chosen for differences in cell-wall composition and habitat, was performed. The solvents were two in common use, Tris-ethylenediaminetetraacetate (Tris-EDTA) and trichloroacetic acid (TCA), and two promising, though less utilized solvents, dimethylsulphoxide (DMSO) and acetone. ATP was determined by the luciferin-luciferase reaction. Of the solvents used, DMSO and acetone were the most effective considering the different kinds of bacteria tested and, of these two, DMSO was the most convenient to use. Tris-EDTA was not as effective as the other solvents tested and TCA, which was effective with most strains, gave low yields when used with cultures grown in artificial seawater broth. Internal standards were used to determine if there were substances present that could inhibit the reaction of released ATP with the luciferin-luciferase reagent. Extracts of ATP, stored at -20 degrees C, were stable for up to 3 weeks.

Acetone↗

The faecal actinomycete flora of Protracheoniscus amoenus (woodlice; Isopoda).

Following a marked order of preference, adults of Protracheoniscus amoenus consumed in preference tests the fallen, weathered leave species of forest litter much more selectively than those of the millipede Chromatoiulus projectus. Under natural circumstances, the individuals (prejuveniles, juveniles, young adults and adults) of these two species live frequently together in the forest litter-layer, feeding on weathered leaves of the very same tree species and placing their faecal pellets in the vicinity of each other. Accordingly, there is a high probability of mutual contamination with their faecal bacteria. In contrast, promicromonospora-type intestinal actinomycetes characterized by cell-wall composition type VI colonize only the faeces of Ch. projectus, and are completely lacking of the dropped pellets of woodlice. Diverse species of streptomycetes were detected in the fresh faecal matter of both litter-dwelling animal species, but presumably they were only survivors of gut passage and passive travellers through the digestive canal.

Actinomycetales↗

[New concept of the taxonomic position of anaerobic corynebacteria].

The new phenotypic and genotypic characteristics of the anaerobic coryneforms allow us to individualize them in a new sub-genus: Coryneformis nv. sub-gen so defined: Corynebacteriaceae with morphology of true Corynebacterium from which they differ by pathogenicity, the ability to synthesize reticulo-stimulin, their cell-wall composition, thier G + C%, their anaerobic growth and their salmon-pink colonies. They differ also from the true Propionibacterium which are not pathogenic, do not synthesize the reticulo-stimulin, have cream-grey, yellor or red-brown colonies, and do synthesize cobalamine (vitamin B12).

Actinomycetales↗

Cell-wall structure and composition of Penicillium janczewskii as affected by inulin.

Penicillium janczewskii, a filamentous fungus isolated from the rhizosphere of Vernonia herbacea (Asteraceae), grows rapidly on media containing either sucrose or inulin as carbon sources. Maintenance of P. janczewskii on inulin medium induces secretion of proteins with high inulinase activity but results in a mycelium that easily collapses and breaks. We evaluated the influence of inulin on fungal growth and colony morphology and on cell-wall structure and composition in comparison with growth and wall characteristics on sucrose-containing medium. P. janczewskii grown on Czapek medium with agar containing 1% (w/v) sucrose or inulin showed differences in the color and morphology of the colonies, although growth rates were similar on both carbon sources. Scanning-electron microscopy revealed that the hyphae from fungus grown on inulin-containing medium are much thinner than those from fungus cultivated on sucrose. Ultrastructural analysis of 5 d old cultures using transmission-electron microscopy indicated significant differences in the cell-wall thickness between hyphae grown on inulin or sucrose media. No differences were detected in the overall carbohydrate and protein contents of cell walls isolated from cultures grown on the two carbon sources. Glycosyl composition analyses showed glucose and galactose as the predominant neutral monosaccharides in the walls but showed no differences attributable to the carbon source. Glycosyl linkage composition analyses indicated a predominance of 3-linked glucopyranosyl in the hyphal walls when P. janczewskii was grown on inulin-containing medium. Our results suggest that growth on inulin as the sole carbon source results in structural changes in the mycelia of P. janczewskii that lead to mycelial walls with altered physical and biological properties.

Carbohydrates↗

Taxonomic significance of fucose in the class Urediniomycetes: distribution of fucose in cell wall and phylogeny of urediniomycetous yeasts.

The carbohydrate compositions of cell wall were determined in the strains of class Urediniomycetes, mainly ballistoconidium-forming yeasts and related taxa. The major component of cell wall was mannose, and glucose was included as the second component, but xylose was not detected in any strain. Out of 41 strains examined, 39 contained galactose, 14 contained arabinose and 12 contained rhamnose. As a minor component, fucose was detected in 30 strains but not in 11 strains. A phylogenetic tree based on 18S rDNA sequences indicated that the fucose-lacking strains, Erythrobasidium hasegawianum, Rhodotolura aurantiaca, R. lactosa, R. minuta, Sakaguchia dacryoidea, Sporobolomyces coprosmae, S. elongatus, S. folicola, S. gracilis, S. kluyverinielii and S. oryzicola, constituted a distinct cluster from those strains which contained fucose. This cluster corresponded to one of the five subclusters, the Erythrobasidium cluster, in the phylogenetic tree of class Urediniomycetes. The carbohydrate composition of cell wall is believed to reflect the phylogenetic relationships among basidiomycetous fungi. The presence or absence of fucose in cell wall should be regarded as an important phenotypic characteristic in the taxonomy of basidiomycetes.

Basidiomycota↗

Cell wall and lipid composition of Isosphaera pallida, a budding eubacterium from hot springs.

Isosphaera pallida is an unusual gliding, budding eubacterium recently isolated from North American hot springs. Electron micrographs of ultrathin sections revealed a cell wall atypical of eubacteria: two electrondense layers separated by an electron-transparent layer, with no evident peptidoglycan layer. Growth was not inhibited by penicillin. Cell walls were isolated from sheared cells by velocity sedimentation. The rigid-layer fraction, prepared from cell walls by treatment with boiling 10% sodium dodecyl sulfate, was hydrolyzed and chemically analyzed for muramic acid. This essential component of peptidoglycan was absent. Amino acid analysis demonstrated a proteinaceous wall structure. Pitlike surface structures seen in negatively stained whole cells and thin sections were correlated with periodically spaced perforations of the rigid sacculus. An analysis of the lipid composition of I. pallida revealed typical ester-linked lipids with unbranched fatty acids, in contrast to the isoprenyl ether-linked lipids of archaebacteria, which also have proteinaceous cell walls. Capnoids, unusual sulfonolipids which are present in gliding bacteria of the Cytophaga-Flexibacter group, were absent.

Amino Acids↗

Altered pectin composition in primary cell walls of korrigan, a dwarf mutant of Arabidopsis deficient in a membrane-bound endo-1,4-beta-glucanase.

Korrigan (kor) is a dwarf mutant of Arabidopsis thaliana (L.) Heynh. that is deficient in a membrane-bound endo-1,4-beta-glucanase. The effect of the mutation on the pectin network has been studied in kor by microscopical techniques associated with various probes specific for different classes of pectic polysaccharides. The localisation of native crystalline cellulose was also examined using the cellobiohydrolase I-gold probe. The investigations were focused on the external cell walls of the epidermis, a cell layer that, in a number of plant species, has been shown to be growth limiting. Anionic sites associated with pectic polymers were quantified using the cationic gold probe. Homogalacturonans were quantified using polyclonal anti-polygalacturonic acid/rhamnogalacturonan I antibodies recognising polygalacturonic acid, and monoclonal JIM7 and JIM5 antibodies recognising homogalacturonans with a high or low degree of methyl-esterification, respectively. Rhamnogalacturonans were quantified with two monoclonal antibodies, LM5, recognising beta-1,4 galactan side chains of rhamnogalacturonan I, and CCRCM2. Our results show a marked increase in homogalacturonan epitopes and a decrease in rhamnogalacturonan epitopes in kor compared to the wild type. A substantial decrease in cellobiohydrolase I-gold labelling was also observed in the mutant cell walls. These findings demonstrate that a deficiency in an endo-1,4-beta-glucanase, which is in principle not directly implicated in pectin metabolism, can induce important changes in pectin composition in the primary cell wall. The changes indicate the existence of feedback mechanisms controlling the synthesis and/or deposition of pectic polysaccharides in primary cell walls.

Antibodies, Monoclonal↗

Analysis of the amino acid and sugar composition of streptococcal cell walls by gas chromatography-mass spectrometry.

A procedure for determining the amino acid and sugar composition of streptococcal peptidoglycan-polysaccharide complexes by capillary gas chromatography-mass spectrometry (GC-MS) was established. Amino acids are analysed as butyl heptafluorobutyl derivatives and sugars as alditol acetates. These two different groups of compounds are derivatized independently but chromatography in both cases utilizes the same OV-1701 fused-silica capillary column which simplifies GC-MS analysis. The butyl heptafluorobutyl procedure incorporates new pre- and post-derivatization clean-up steps. Additionally, selected-ion monitoring MS allows amino acids to be readily analysed without interference from background noise.

Amino Acids↗

Chemical composition of Streptococcus mutans cell walls and their susceptibility to Flavobacterium L-11 enzyme.

The susceptibility to a cell wall lytic L-11 enzyme from Flavobacterium sp. and the quantitative and/or qualitative composition of the cell walls of some strains of cariogenic Streptococcus mutans and a non-cariogenic strain of Streptococcus mitis were determined. The purified cell walls of S. mutans strains HS-1 (serotype a), BHT (b), NCTC10449 (c), C67-1 (c), C67-25 (c), OMZ 176 (d), MT703 (e), MT557 (f), OMZ65 (g), and AHT (g), and S. mitis CHT contained glutamic acid, alanine, and lysine as well as muramic acid and glucosamine as a peptidoglycan component. Besides these amino acids, significant amounts of threonine were detected in strains HS-1, OMZ65, and AHT cell walls, and considerable amounts of aspartic acid and/or threonine as well as several other amino acids in OMZ176, OMZ65, and CHT cell walls. Rhamnose was a common special component of the cell walls of S. mutans strains BHT, NCTC10449, MT703, B2 (e), MT557, and AHT, and S. mitis CHT. An additional sugar component, glucose, was detected in the cell walls of all of these strains except BHT, and galactose was found in BHT, AHT, and CHT cell walls. Galactosamine was present in S. mitis CHT cell walls. Varying amounts of phosphorus were detected in the cell walls of all the strains examined. The cell walls of all these streptococcal strains except MT703, 6715, and AHT were susceptible to the lytic action of the L-11 enzyme to various extents. No consistent relationship was observed between the amino acid and sugar composition of these cell walls and their susceptibility to the L-11 enzyme. The chemical composition of these cell walls is discussed in terms of the serological classification of S. mutans.

Amino Acids↗

Cell wall and phospholipid composition and their contribution to the salt tolerance of Halomonas elongata.

The salt-tolerant bacterium Halomonas elongata makes a variety of physiological adaptations in response to increases in the salt concentration of its growth medium. The cell walls become more compact and internally coherent. The overall lipid pattern shows an increased amount of negatively charged lipids. In addition, the peptidoglycan composition of H. elongata, although not changing in response to increased NaCl, contains the hydrophobic amino acid leucine which is unique among bacterial species. The results suggest that H. elongata is able to live in a wide variety of salt concentrations because it alters its cell physiology in ways which increase both structural integrity and the amount of less-mobile, "structured" cell water, making the cells less susceptible to NaCl-induced dehydration.

Acclimatization↗

Carbohydrate composition of the isolated cell walls of dermatophytes.

In an attempt to evaluate taxonomic character of sugar composition of dermatophytes, the purified cell walls from 13 species are analyzed on neutral sugar composition by gas liquid chromatography. The results were principally compatible with those obtained by conventional morphological examination. Neutral sugar components of dermatophytes cell walls were mannose and glucose in the ratio of 1:2.7 for Epidermophyton and 1:1.4 for Microsporum. There were two types in Trichophyton, in which the ratios of mannose to glucose were 1:1.6 and 1:3.8. The cases of Trichophyton ferrugineum and Trichophyton mentagrophytes were exceptional. The ratio of the former was 1:1.4, which implied the relation to Microsporum group, and the ratio of the latter was 1:2.3, which was supposed to be the intermediate of two types of Trichophyton group. Albino type cell wall of Epidermophyton floccosum was more rich in glucose than pigmented type one.

Acetylglucosamine↗

Differences in virulence of Sporothrix schenckii conidia related to culture conditions and cell-wall components.

A murine model was used to evaluate the virulence of Sporothrix schenckii conidia cultured for 4, 7, 10 or 12 days in Sabouraud's dextrose broth (SDB). A correlation was observed between length of culture and virulence. Mice infected intravenously with S. schenckii conidia cultured for 4 or 7 days showed 40-100% cumulative mortality. In contrast, mice infected with conidia from cultures grown for 10 or 12 days in SDB showed no mortality (100% survival). A much greater accumulation of fungal colony forming units (cfu) was observed in the lungs, livers and spleens of mice inoculated with conidia of S. schenckii cultured for 7 days than in mice infected with conidia cultured for 12 days. The livers of mice from the former group showed a widespread granulomatous reaction whereas mice inoculated with S. schenckii cultured for 12 days showed a more limited response with fewer granulomas. No difference in viability or replicative capacity was discerned for these S. schenckii cultured cells. However, the more virulent forms of the fungus showed differences in cell-wall sugar composition with rhamnose:mannose molar ratios of 1.7:1.0 for cells cultured for 4 days and 1.0:1.7 for conidia cultured for 12 days. These results suggest that the virulence of S. schenckii conidia may be determined by their cell-wall composition.

Agglutination Tests↗

[Polysaccharide composition of the cell wall of Streptomyces antibioticus RIA-594(39), a producer of the antibiotic oleandomycin].

The main polysaccharide components of the cell wall in S. antibioticus RIA-594 (39) i.e. peptidoglycan, teichoic acid and polysaccharide were studied. Peptidoglycan consists of the polysaccharide fraction containing equimolar quantities of N-acetylglucosamine and muramic acid and the peptide subunits including alanine, glutamic and L,L-diaminopimelic acids and glycine at a ratio of 1.4:0.9:1:0.9. It is characteristic that certain peptide subunits of the streptomycete contain no alanine. A polysaccharide differing from glycerol teichoic acid and containing galactose and N-acetylglucosamine was isolated from the cell wall. During the streptomycete development the quantity of peptidoglycan remained constant, the quantity of teichoic acid lowered and the quantity of polysaccharide increased. Correlation between the presence of aminosugars in the composition of teichoic acid and polysaccharide specific of the streptomycete cell wall and the presence of aminosugars in the structure of oleandomycin was shown. This is probably connected with characteristic features of the organism physiology.

Amino Acids↗

Effects of phosphate limitation of growth on the cell-wall and lipid composition of Saccharomyces cerevisiae.

The phosphorus content of phosphate-limited Saccharomyces cerevisiae was only 71% of that of non-limited yeast. Walls prepared from phosphate-limited cells contained slightly less phosphorus than control walls. No evidence was obtained for the presence in these walls of uronic acid or succinyl residues. The carbohydrate content of walls of phosphate-limited yeast was less than that of non-limited walls, and this was reflected in a decreased glucan content. There was only a slight decrease in glucosamine content while the protein content increased. The major change in the lipid composition of phosphate-limited yeast was a decrease in both sterol esters and triacylglycerols. There was a decrease in total lipid content, but increased production of phosphatidylethanolamine and phosphatidylcholine. The phosphatidylserine content was decreased. These results suggest that there are fewer intracellular low-density vesicles in phosphate-limited yeast.

Carbohydrates↗