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At least 523 records · Page 29Linked to original sources
PubMed2005

Bacteria associated with spores of the arbuscular mycorrhizal fungi Glomus geosporum and Glomus constrictum.

Spores of the arbuscular mycorrhizal fungi (AMF) Glomus geosporum and Glomus constrictum were harvested from single-spore-derived pot cultures with either Plantago lanceolata or Hieracium pilosella as host plants. PCR-denaturing gradient gel electrophoresis analysis revealed that the bacterial communities associated with the spores depended more on AMF than host plant identity. The composition of the bacterial populations linked to the spores could be predominantly influenced by a specific spore wall composition or AMF exudate rather than by specific root exudates. The majority of the bacterial sequences that were common to both G. geosporum and G. constrictum spores were affiliated with taxonomic groups known to degrade biopolymers (Cellvibrio, Chondromyces, Flexibacter, Lysobacter, and Pseudomonas). Scanning electron microscopy of G. geosporum spores revealed that these bacteria are possibly feeding on the outer hyaline spore layer. The process of maturation and eventual germination of AMF spores might then benefit from the activity of the surface microorganisms degrading the outer hyaline wall layer.

Asteraceae↗
PubMed2005

Photodynamic inactivation of Bacillus spores, mediated by phenothiazinium dyes.

Spore formation is a sophisticated mechanism by which some bacteria survive conditions of stress and starvation by producing a multilayered protective capsule enclosing their condensed DNA. Spores are highly resistant to damage by heat, radiation, and commonly employed antibacterial agents. Previously, spores have also been shown to be resistant to photodynamic inactivation using dyes and light that easily destroy the corresponding vegetative bacteria. We have discovered that Bacillus spores are susceptible to photoinactivation by phenothiazinium dyes and low doses of red light. Dimethylmethylene blue, methylene blue, new methylene blue, and toluidine blue O are all effective, while alternative photosensitizers such as Rose Bengal, polylysine chlorin(e6) conjugate, a tricationic porphyrin, and a benzoporphyrin derivative, which easily kill vegetative cells, are ineffective. Spores of Bacillus cereus and B. thuringiensis are most susceptible, B. subtilis and B. atrophaeus are also killed, and B. megaterium is resistant. Photoinactivation is most effective when excess dye is washed from the spores, showing that the dye binds to the spores and that excess dye in solution can quench light delivery. The relatively mild conditions needed for spore killing could have applications for treating wounds contaminated by anthrax spores, for which conventional sporicides would have unacceptable tissue toxicity.

Bacillus↗
PubMed2005

Incidence and diversity of potentially highly heat-resistant spores isolated at dairy farms.

The presence of highly heat-resistant spores of Bacillus sporothermodurans in ultrahigh-temperature or sterilized consumer milk has emerged as an important item in the dairy industry. Their presence is considered undesirable since they hamper the achievement of commercial sterility requirements. By using a selective 30-min heat treatment at 100 degrees C, 17 Belgian dairy farms were screened to evaluate the presence, sources, and nature of potentially highly heat-resistant spores in raw milk. High numbers of these spores were detected in the filter cloth of the milking equipment and in green crop and fodder samples. About 700 strains were isolated after the selective heating, of which 635 could be screened by fatty acid methyl ester analysis. Representative strains were subjected to amplified ribosomal DNA restriction analysis, 16S rRNA gene sequencing, percent G+C content, and DNA-DNA reassociations for further identification. The strain collection showed a remarkable diversity, with representatives of seven aerobic spore-forming genera. Bacillus licheniformis and Bacillus pallidus were the most predominant species overall. Twenty-three percent of the 603 spore-forming isolates proved to belong to 18 separate novel species. These findings suggest that the selective heating revealed a pool of unknown organisms with a higher heat-resistant character. This study showed that high spore counts can occur at the dairy farm and that feed and milking equipment can act as reservoirs or entry points for potentially highly heat-resistant spores into raw milk. Lowering this spore load by good hygienic measures could probably further reduce the contamination level of raw milk, in this way minimizing the aerobic spore-forming bacteria that could lead to spoilage of milk and dairy products. Assessment and characterization of this particular flora are of great importance to allow the dairy or food industry to adequately deal with newly arising microbiological problems.

Animals↗
PubMed1962

Gamma-ray sterilization and residual toxicity studies of ground beef inoculated with spores of Clostridium botulinum.

Inoculated packs of cooked and raw ground beef were sterilized with gamma radiation from cobalt-60. With inocula of 5,000,000 Clostridium botulinum 213B spores per g of cooked ground beef, 3.8 megarad were required for sterilization; in raw ground beef, 3.72 megarad sterilized the meat when inocula of 1,700,000 C. botulinum 213B spores were used per g. Using C. botulinum 62A spores, cooked ground beef inoculated with 5,200,000 spores per g was sterilized with 3.85 megarad; raw ground beef, inoculated with 2,670,000 spores per g, was sterilized with 3.6 megarad. Cans of meat that were considered sterile by lack of culture growth after incubation for at least 6 months and, in some instances, as long as 5 years, were tested for the presence of botulinus toxin. No toxin was found in any meat taken from inoculated packs prepared from C. botulinum 213B spores; however, all cans of meat that had been inoculated with more than 2,670,000 C. botulinum 62A spores per g of meat, contained type A toxin. It was shown that these latter inocula of heat-shocked spores, by themselves, contained sufficient toxin to kill mice. However, more toxin appeared to be present than could be ascribed to the unirradiated spores alone. This finding is discussed.

Animals↗
PubMed1962

Heat activation and heat-induced dormancy of Bacillus stearothermophilus spores.

Heat-induced dormancy was observed when spores of two strains of Bacillus stearothermophilus were heated in distilled water at 80, 90, and 100 C. At temperatures above 100 C, true activation occurred; however, maximal activation was not achieved until temperatures of 110 to 115 C were employed. A heat treatment of 115 C for 3 min was required to induce maximal activation in one suspension of strain 1518 spores, whereas a heat treatment of 110 C for 7 to 10 min was adequate for the other suspension of strain 1518 spores. Spores from both strain M suspensions required heat treatments of 110 C for 9 to 15 min for maximal activation. The degree to which the spores could be activated was strain dependent and variable among spore suspensions of the same strain. The germination and outgrowth of all spores, regardless of strain and suspensions source, were significantly reduced when the spores were heated in m/120 phosphate buffer at maximal or near maximal activating temperatures. It was suspected that phosphate lowered the heat resistance of the spores to the extent that the heat treatments were lethal to a portion of the populations.

Bacillus↗
PubMed1970

Chemical states of bacterial spores: heat resistance and its kinetics at intermediate water activity.

Bacterial spore heat resistance at intermediate water activity, like aqueous and strictly dry heat resistance, is a property manipulatable by chemical pretreatments of the dormant mature spore. Heat resistances differ widely, and survival is prominently nonlogarithmic for both chemical forms of the spore. Log survival varies approximately as the cube of time for the resistant state of Bacillus stearothermophilus spores and as the square of time for the sensitive state. A method for measuring heat resistance at intermediate humidity was designed to provide direct and unequivocal control of water vapor concentration with quick equilibration, maintenance of known spore state, and dispersion of spores singly for valid survivor counting. Temperature characteristics such as z, E(a), and Q(10) cannot be determined in the usual sense (as a spore property) for spores encapsulated with a constant weight of water. Effect on spore survival of temperature induced changes of water activity in such systems is discussed.

Bacillus↗
PubMed1970

Resistance of Bacillus subtilis spores to inactivation by gamma irradiation and heating in the presence of a bactericide. I. Suitability of viable count procedures.

A statistical evaluation of viable count procedures utilized for obtaining treatment survival curve data for Bacillus subtilis NCTC 8236 spores is described. Within the various recovery conditions tested, incubation on nutrient agar containing 1% dextrose for 48 hr at 37 C was found to promote the highest count of viable spores surviving a variety of bactericidal treatments involving gamma irradiation, heat, and chlorocresol. The count of viable spores on the medium was not significantly altered when the dextrose was added to the nutrient agar either before autoclaving or aseptically at 50 to 55 C from a solution sterilized by filtration. The volume of medium which promoted the highest count of viable spores was 20 ml per 85 mm of diameter in disposable plastic plates. Counts of viable spores were reproducible on successive batches of media. The carry-over of variable concentrations of chlorocresol into the medium from serial dilutions affected the count of viable spores. Spores in the aqueous stock suspension used for all experiments were uniformly distributed after shaking and did not diminish significantly in viability after 16 months of storage at 5 C. Grouping of indexes of dispersion, calculated from quintuplicate plate colony counts, indicated that the suitability of the viable count procedures, employed for the enumeration of spores surviving the various bactericidal treatments, tended to diminish as the level of spore inactivation exceeded 95%.

Agar↗
PubMed1970

Dry-heat destruction of Bacillus subtilis spores on surfaces: effect of humidity in an open system.

Bacillus subtilis var. niger spores were tested for dry-heat resistance on stainless-steel strips hung in an oven. Heat resistance was dependent on the relative humidity before and during treatment, which in turn affected the water content of the spores. Higher humidities increased the heat resistance of the spores. D-values ranged from 16.1 min for spores conditioned at <2% relative humidity (RH) and treated at 0.34% RH to 37.6 min for spores conditioned at 89% RH and treated at 1.1% RH. The y-intercept of the regression line ranged from 6.94 x 10(4) for spores conditioned and treated at the low humidities to 2.00 x 10(5) for spores conditioned at 89% RH and treated at 0.34% RH. For a constant value of N(0), the y-intercept appears to be lowered by low-humidity conditions. The statistic log y(0)/log N(0) is used to measure the downward displacement of the regression line. Values obtained in this experiment range from 0.90 for spores conditioned at <2% RH and treated at 0.34% RH to 1.04 for spores conditioned at <2% RH and treated at 1.1% RH. A combination of linear regression and analysis of variance methods was used for data analysis. The former estimates D-values and y-intercepts, whereas the latter is sensitive to differences between treatments.

Analysis of Variance↗
PubMed2002

Dtrlp, a multidrug resistance transporter of the major facilitator superfamily, plays an essential role in spore wall maturation in Saccharomyces cerevisiae.

The de novo formation of multilayered spore walls inside a diploid mother cell is a major landmark of sporulation in the yeast Saccharomyces cerevisiae. Synthesis of the dityrosine-rich outer spore wall takes place toward the end of this process. Bisformyl dityrosine, the major building block of the spore surface, is synthesized in a multistep process in the cytoplasm of the prospores, transported to the maturing wall, and polymerized into a highly cross-linked macromolecule on the spore surface. Here we present evidence that the sporulation-specific protein Dtrlp (encoded by YBR180w) plays an important role in spore wall synthesis by facilitating the translocation of bisformyl dityrosine through the prospore membrane. DTR1 was identified in a genome-wide screen for spore wall mutants. The null mutant accumulates unusually large amounts of bisformyl dityrosine in the cytoplasm and fails to efficiently incorporate this precursor into the spore surface. As a result, many mutant spores have aberrant surface structures. Dtrlp, a member of the poorly characterized DHA12 (drug:H+ antiporter with 12 predicted membrane spans) family, is localized in the prospore membrane throughout spore maturation. Transport by Dtrlp may not be restricted to its natural substrate, bisformyl dityrosine. When expressed in vegetative cells, Dtrlp renders these cells slightly more resistant against unrelated toxic compounds, such as antimalarial drugs and food-grade organic acid preservatives. Dtrlp is the first multidrug resistance protein of the major facilitator superfamily with an assigned physiological role in the yeast cell.

Cell Wall↗
PubMed2004

Morphogenetic pathway of spore wall assembly in Saccharomyces cerevisiae.

The Saccharomyces cerevisiae spore is protected from environmental damage by a multilaminar extracellular matrix, the spore wall, which is assembled de novo during spore formation. A set of mutants defective in spore wall assembly were identified in a screen for mutations causing sensitivity of spores to ether vapor. The spore wall defects in 10 of these mutants have been characterized in a variety of cytological and biochemical assays. Many of the individual mutants are defective in the assembly of specific layers within the spore wall, leading to arrests at discrete stages of assembly. The localization of several of these gene products has been determined and distinguishes between proteins that likely are involved directly in spore wall assembly and probable regulatory proteins. The results demonstrate that spore wall construction involves a series of dependent steps and provide the outline of a morphogenetic pathway for assembly of a complex extracellular structure.

Cell Membrane↗
PubMed2006

ExsY and CotY are required for the correct assembly of the exosporium and spore coat of Bacillus cereus.

The exosporium-defective phenotype of a transposon insertion mutant of Bacillus cereus implicated ExsY, a homologue of B. subtilis cysteine-rich spore coat proteins CotY and CotZ, in assembly of an intact exosporium. Single and double mutants of B. cereus lacking ExsY and its paralogue, CotY, were constructed. The exsY mutant spores are not surrounded by an intact exosporium, though they often carry attached exosporium fragments. In contrast, the cotY mutant spores have an intact exosporium, although its overall shape is altered. The single mutants show altered, but different, spore coat properties. The exsY mutant spore coat is permeable to lysozyme, whereas the cotY mutant spores are less resistant to several organic solvents than is the case for the wild type. The exsY cotY double-mutant spores lack exosporium and have very thin coats that are permeable to lysozyme and are sensitive to chloroform, toluene, and phenol. These spore coat as well as exosporium defects suggest that ExsY and CotY are important to correct formation of both the exosporium and the spore coat in B. cereus. Both ExsY and CotY proteins were detected in Western blots of purified wild-type exosporium, in complexes of high molecular weight, and as monomers. Both exsY and cotY genes are expressed at late stages of sporulation.

Bacillus cereus↗
PubMed2006

Role of SpoVA proteins in release of dipicolinic acid during germination of Bacillus subtilis spores triggered by dodecylamine or lysozyme.

The release of dipicolinic acid (DPA) during the germination of Bacillus subtilis spores by the cationic surfactant dodecylamine exhibited a pH optimum of approximately 9 and a temperature optimum of 60 degrees C. DPA release during dodecylamine germination of B. subtilis spores with fourfold-elevated levels of the SpoVA proteins that have been suggested to be involved in the release of DPA during nutrient germination was about fourfold faster than DPA release during dodecylamine germination of wild-type spores and was inhibited by HgCl(2). Spores carrying temperature-sensitive mutants in the spoVA operon were also temperature sensitive in DPA release during dodecylamine germination as well as in lysozyme germination of decoated spores. In addition to DPA, dodecylamine triggered the release of amounts of Ca(2+) almost equivalent to those of DPA, and at least one other abundant spore small molecule, glutamic acid, was released in parallel with Ca(2+) and DPA. These data indicate that (i) dodecylamine triggers spore germination by opening a channel in the inner membrane for Ca(2+)-DPA and other small molecules, (ii) this channel is composed at least in part of proteins, and (iii) SpoVA proteins are involved in the release of Ca(2+)-DPA and other small molecules during spore germination, perhaps by being a part of a channel in the spore's inner membrane.

Amines↗
PubMed1974

Ultraviolet sensitivity of Bacillus subtilis spores upon germination and outgrowth.

A strain of Bacillus subtilis, UVSSP-42-1, which produces ultraviolet (UV)-sensitive spores and vegetative cells, was found to possess germinated spores 25 times more UV resistant than the resting spores. This relative resistance achieved upon germination was associated with the transition of the heat-resistant refractile spores to the heat-sensitive phase-dark forms. Several generations of outgrowth were required before the cells attained the level of UV sensitivity characteristic of the vegetative cell. The UV sensitivity of germinated spores was compared with other strains with various combinations of mutations affecting deoxyribonucleic acid repair capabilities. The presence of hcr and ssp mutations which are known to abolish the removal of photoproducts from deoxyribonucleic acid did not alter significantly the sensitivity of the germinated forms. However, the addition of the recA mutation and, to some extent, the pol mutation increased the UV sensitivity of the germinated spores. These results indicate that deoxyribonucleic acid repair mechanisms dependent on the recA gene are active in the germinated spores. The chemical nature of the damage repaired by the recA gene product is not known. This study indicates that the life cycle of sporulating bacilli consists of at least three photobiologically distinct forms: spore, germinated spore, and vegetative cell.

Bacillus subtilis↗
PubMed1976

Ultrastructure and elemental composition of dormant and germinating Diplodia maydis spores.

The ultrastructure of Diplodia maydis spores was studied in thin sections with a transmission electron microscope. Storage vacuoles were evenly distributed in the two cells. Some of the vacuoles that contained a dense osmiophilic sphere(s) were surrounded by a membrane, and had membranous aggregates around their periphery. The sport wall was composed of an electron-dense layer and an electron-translucent layer. An inner cytoplasmic membrane was present. Dormant and germinating spores were studied with scanning electron microscopy and also with a Si (Li) energy-dispersive X-ray analyzer. The dormant spore was ovate and usually two-celled with a central septum. Germination proceeded via a germ tube from the side of one end of the cell. Of several methods for preparation of specimens for X-ray analysis studied, freeze-dried spores mounted on carbon stubs and then further carbon coated gave the best results. X-ray analyses revealed that spore populations contained large amounts of Si, P, Cl, and K, smaller amounts of S and Ca, and trace amounts of Mg and Al. Analyses of single spores revealed high K and Cl and low P and Mg at one end of the cell with concomitant low K and Cl and high P and Mg in the central portion and other end of the cell. In two-celled germinating spores, high K and Cl occurred in the end of the nongerminating spore cell, whereas the germinating cell contained high P and Mg and low K and Cl. X-ray image maps revealed that K and Cl were located together at one end of the spore.

Calcium↗
PubMed1978

In vitro translation of polyadenylate-containing RNAs from dormant and germinating spores of the fungus Botryodiplodia theobromae.

Polyadenylated RNA isolated by oligodeoxythymidylate-cellulose chromatography from spores of the fungus Botryodiplodia theobromae was translated in a cell-free protein-synthesizing system derived from wheat embryo. Reaction conditions which would yield efficient and accurate in vitro translation of the spore RNA were established. Dual isotopically labeled mixtures of in vitro translation products from germinated and dormant spore polyadenylated RNA, as well as polyadenylated RNAs from intermediate stages of germination, produced qualitatively similar gel electrophoresis patterns, with polypeptides of 10,000 to 55,000 molecular weight. Proteins synthesized in vivo and extracted from germinating spores at three different stages possessed a greater size range, with molecular weights up to 85,000, although the in vitro synthesis apparently did yield the lower-molecular-weight proteins which were synthesized in vivo. Tryptic digest patterns of proteins translated in vitro from polyadenylated RNA of dormant and germinated spores were found to be identical in positions in only 40% of the spots. Furthermore, a dual-label comparison by isoelectric focusing of proteins translated from polyadenylated RNA of germinated and dormant spores also showed qualitative and quantitative differences among the in vitro translation products. We conclude that there are differences between the in vitro translation products of mRNA from dormant and germinated spores and that the mRNA preserved in the dormant spores contains genetic information which is qualitatively different from that of the germinated spores.

Cell-Free System↗
PubMed1978

Purification and characterization of additional low-molecular-weight basic proteins degraded during germination of Bacillus megaterium spores.

Dormant spores Bacillus megaterium contained a group of low-molecular-weight (5,000 to 11,000) basic (pI greater than 9.4) proteins (termed D, E, F, and G proteins) which could be extracted from disrupted spores with strong acids. These proteins were distinct from the previously described A, B, and C proteins which are degraded during spore germination. However, the D, E, F, and G proteins were also rapidly degraded during spore germination, accounting for 10 to 15% of the protein degraded. Proteins similar to the D, E, F, and G species were also present in spores of other bacterial species. In B. megaterium, the D, E, F, and G proteins were low or absent (less than 15% of the spore level) in vegetative and young sporulating cells and appeared only late in sporulation. The D, E, F, and G proteins were purified to homogeneity, and all contained a high percentage of hydrophilic amino acids; one protein (G) contained 31% basic amino acids and also contained tryptophan. All four proteins were rapidly degraded in vitro by dormant spore extracts. Two proteins (D and F) were degraded in vitro by the previously described spore protease which initiates degradation of the A, B, and C proteins in vivo; the spore enzyme (s) degrading proteins E and G have not been identified.

Amino Acids↗
PubMed1981

Germination properties of a spore coat-defective mutant of Bacillus subtilis.

The presence of the gerE36 mutation in strains of Bacillus subtilis 168 resulted in poor germination of their spores in a range of germinants, as measured by the fall in absorbance of spore suspensions. Although resistant to heat and organic solvents, spores were sensitive to lysozyme; electron microscopy revealed that their coat structure was incomplete. These spores responded to germinants by losing heat resistance and changing from phase bright to phase gray. The release of dipicolinic acid and the fall in absorbance of spore suspensions reached only 75 and 50% of wild-type levels, respectively, but followed the same time course as the loss of heat resistance. Although the germination response was incomplete, the concentration of L-alanine required to elicit it was the same for the mutant as for the wild type. The properties of mutant spores suggest that an intact spore coat is not required for the initial interaction between germinant and spore, but that the coat layers may contain molecules important in later stages of germination. In transduction with phage SPP1, the gerE36 mutation mapped between citF and ilvB and was 90% cotransduced with citF2. The gerE mutation identifies the location of a gene important for the progress of late stages of spore formation.

Bacillus subtilis↗
PubMed1982

Noninvolvement of the spore cortex in acquisition of low-molecular-weight basic proteins and UV light resistance during Bacillus sphaericus sporulation.

Two major low-molecular weight, acid-soluble proteins (termed A and B proteins) were purified from Bacillus sphaericus spores and had properties similar to those of the analogous proteins from spores of other Bacillus species. These proteins were accumulated late in sporulation, when the developing spores became resistant to UV light, and were degraded during spore germination by a spore protease. A mutant of B. sphaericus unable to make spore cortex because of a block in diaminopimelic acid (DAP) biosynthesis accumulated and maintained levels of the A and B proteins similar to those in the DAP+ parent or the DAP- strain in which cortex formation was restored by growth with DAP. In addition, the DAP- strain grown without DAP acquired a level of UV light resistance identical to that of wild-type spores and at the time of appearance of the A and B proteins. These findings indicate that formation of little, if any, spore cortex is required for acquisition of UV light resistance or maintenance of high levels of A and B proteins. The data provide further support for a role of the A and B proteins in the spore's UV light resistance.

Bacillus↗