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PubMed2005

The regulator of G-protein signaling proteins involved in sugar and abscisic acid signaling in Arabidopsis seed germination.

The regulator of G-protein signaling (RGS) proteins, recently identified in Arabidopsis (Arabidopsis thaliana; named as AtRGS1), has a predicted seven-transmembrane structure as well as an RGS box with GTPase-accelerating activity and thus desensitizes the G-protein-mediated signaling. The roles of AtRGS1 proteins in Arabidopsis seed germination and their possible interactions with sugars and abscisic acid (ABA) were investigated in this study. Using seeds that carry a null mutation in the genes encoding RGS protein (AtRGS1) and the alpha-subunit (AtGPA1) of the G protein in Arabidopsis (named rgs1-2 and gpa1-3, respectively), our genetic evidence proved the involvement of the AtRGS1 protein in the modulation of seed germination. In contrast to wild-type Columbia-0 and gpa1-3, stratification was found not to be required and the after-ripening process had no effect on the rgs1-2 seed germination. In addition, rgs1-2 seed germination was insensitive to glucose (Glc) and sucrose. The insensitivities of rgs1-2 to Glc and sucrose were not due to a possible osmotic stress because the germination of rgs1-2 mutant seeds showed the same response as those of gpa1-3 mutants and wild type when treated with the same concentrations of mannitol and sorbitol. The gpa1-3 seed germination was hypersensitive while rgs1-2 was less sensitive to exogenous ABA. The different responses to ABA largely diminished and the inhibitory effects on seed germination by exogenous ABA and Glc were markedly alleviated when endogenous ABA biosynthesis was inhibited. Hypersensitive responses of seed germination to both Glc and ABA were also observed in the overexpressor of AtRGS1. Analysis of the active endogenous ABA levels and the expression of NCED3 and ABA2 genes showed that Glc significantly stimulated the ABA biosynthesis and increased the expression of NCED3 and ABA2 genes in germinating Columbia seeds, but not in rgs1-2 mutant seeds. These data suggest that AtRGS1 proteins are involved in the regulation of seed germination. The hyposensitivity of rgs1-2 mutant seed germination to Glc might be the result of the impairment of ABA biosynthesis during seed germination.

Abscisic Acid↗
PubMed2003

Genetics of drought tolerance during seed germination in tomato: inheritance and QTL mapping.

A BC1 population (N = 1000) of an F1 hybrid between a stress-sensitive Lycopersicon esculentum breeding line (NC84173; maternal and recurrent parent) and a germination stress-tolerant Lycopersicon pimpinellifolium accession (LA722) was evaluated for seed germination rate under drought stress (DS) (14% w/v polyethyleneglycol-8000, water potential approximately -680 kPa), and the most rapidly germinating seeds (first 3% to germinate) were selected. The 30 selected BC1 seedlings were grown to maturity and self pollinated to produce BC1S1 progeny seeds. Twenty of the 30 selected BC1S1 progeny families were evaluated for germination rate under DS and their average performance was compared with that of a "nonselected" BC1S1 population of the same cross. Results indicated that selection for rapid germination under DS significantly improved progeny germination rate under DS (selection gain = 19.6%), suggesting a realized heritability of 0.47 for rate of germination under DS in this population. The 30 selected BC1 plants were subjected to restriction fragment length polymorphism (RFLP) analysis, and marker allele frequencies for 119 RFLP markers which spanned 1153 cM of the 12 tomato chromosomes were determined. A distributional extreme marker analysis, which measures statistical differences in marker allele frequencies between a selected and a nonselected population, detected four quantitative trait loci (QTLs) for rate of germination under DS in this population. Of these, two QTLs, located on chromosomes 1 and 9, were contributed by the L. pimpinellifolium donor parent and had larger effects than the other two QTLs, located on chromosomes 8 and 12, which were contributed by the L. esculentum recurrent parent. A few BC1S1 families were identified with all or most of the identified QTLs and with germination rates comparable with that of LA722. These families should be useful for the development of germination drought-tolerant tomato lines using marker-assisted selection (MAS). The overall results indicate that drought tolerance during seed germination in tomato is genetically controlled and potentially could be improved by directional phenotypic selection or MAS.

Chromosome Mapping↗
PubMed2003

[A comparative study on seed germination of 15 grass species in Keeqin Sandyland].

A laboratory study was made on the germination characteristics of freshly-collected seeds of grass species at the Wulanaodu area of Keeqin Sandyland in Eastern Inner-Mongolia. Of the 15 species examined, 8 species including Clinelymus dahuricus, Cleistogenes squarrosa, Pappophorum boreale, Spodiopogon sibiricus, Phragmites communis, Chloris virgata, Arundinella hirta, Pennisetum alopecuroides had a germination rate of over 80%, but 4 species including Echinochloa hispidula, Hemarthria compressa, Tragus berteronianus and Setaria viridis had a value of less than 10%. Spodiopogon sibiricus, Eragrostis pilosa, Phragmites communis, Chloris virgata, Clinelymus dahuricus, Pappophorum boreale, Digitaria cilliaris and Cleistogenes squrrosa began to germinate within 1-3 days after the test began, while Setaria viridis, Tragus berteronianus and Hemarthria compressa failed to germinate in a period of more than 10 days. For the species such as Digitaria cilliaris, Echinochloa hispidula, Phragmites communis, Eragrostis pilosa and Spodiopogon sibiricus, their germination period was less than 10 days, while Clinelymus dahuricus and Pappophorum boreale had a germination period of more than 20 days. The days required for half the final germination rate to be reached were: 2 days for Chloris virgata, 3 days for Phragmites communis, 4 days for Spodiopogon sibiricus, 5 days for Clinelymus dahuricus and Cleistogenes squarrosa, 7 days for Arundinella hirta and Pappophorum boreale, and 10 days for Pennisetum alopecuriodes. Compared with the Sheffield region in Britain, the Wulanaodu area of Kerqin Sandyland had a higher proportion of annul grasses with a low germination rate and a longer germination period, and the perennial grasses at the Wulanaodu area had an approximately same germination rate, but a longer germination period. During germination, ruderals showed the potential for risk-sharring, and thus, they had a relatively higher disturbance-resistance capacity.

Germination↗
PubMed2005

Environmental and genetic influences on the germination of Arabidopsis thaliana in the field.

Seasonal germination timing strongly influences lifetime fitness and can affect the ability of plant populations to colonize and persist in new environments. To quantify the influence of seasonal environmental factors on germination and to test whether pleiotropy or close linkage are significant constraints on the evolution of germination in different seasonal conditions, we dispersed novel recombinant genotypes of Arabidopsis thaliana into two geographic locations. To decouple the photoperiod during seed maturation from the postdispersal season that maternal photoperiod predicts, replicates of recombinant inbred lines were grown under short days and long days under controlled conditions, and their seeds were dispersed during June in Kentucky (KY) and during June and November in Rhode Island (RI). We found that postdispersal seasonal conditions influenced germination more strongly than did the photoperiod during seed maturation. Genetic variation was detected for germination responses to all environmental factors. Transgressive segregation created novel germination phenotypes, revealing a potential contribution of hybridization of ecotypes to the evolution of germination. A genetic trade-off in germination percentage across sites indicated that determinants of fitness at or before the germination stage may constrain the geographic range that a given genotype can inhabit. However, germination timing exhibited only weak pleiotropy across treatments, suggesting that different sets of genes contribute to variation in germination behavior in different seasonal conditions and geographic locations. Thus, the genetic potential exists for rapid evolution of appropriate germination responses in novel environments, facilitating colonization across a broad geographic range.

Arabidopsis↗
PubMed1991

Kinetics of the tingible body macrophage response in mouse germinal center development and its depression with age.

Although tingible body macrophages (TBM) have been recognized in germinal centers for over 100 years, their role in the germinal center response is not clear. In this study, the kinetics of the TBM response was quantitatively assessed and correlated with the kinetics of germinal center development in young mice. The TBM response in old mice (which have an age-related depression of germinal center development; Szakal et al., 1990) was analyzed for comparison. Young and old immune mice were challenged with human serum albumin and 0, 1, 3, 5, 7, 10, and 14 days later the popliteal and axillary lymph nodes were evaluated. Germinal centers were localized histochemically in alternate serial sections using horseradish peroxidase conjugated peanut agglutinin. TBM numbers were determined per germinal center on adjacent sections by the presence of tingible bodies or histochemically by using the monoclonal antibody Mac-2. Analysis of lymph nodes from young mice showed that TBM numbers decreased with the dissociation of preexisting germinal centers. TBM reappeared 5 days after challenge and the TBM kinetics paralleled the increase in size of de novo germinal centers. In fact, a constant ratio of one TBM to every 350-450 B cells was maintained from day 5 to day 10. In old lymph nodes, TBM were generally absent throughout germinal center development. The lack of TBM prior to germinal center development and their absence in aged mice are inconsistent with the concept that TBM are required for the induction of the germinal center reaction. However, the data are consistent with a role for TBM in regulating the magnitude of the germinal center reaction.

Aging↗
PubMed1996

Germination and Viability of Capilliconidia of Neozygites floridana (Zygomycetes: Entomophthorales) under Constant Temperature, Humidity, and Light Conditions

The germination of capilliconidia of Neozygites floridana on the cassava green mite, Mononychellus tanajoa, and the viability of these conidia collected on glass coverslips were studied at different temperatures, humidities (expressed as saturation deficit (SD), measured in mg Hg), and light conditions. Germination began within 2 hr of their attachment to the mites, at which time more than 66.0% of the conidia maintained at between 18 and 28°C, at SDs 0 and 0.2, and in the dark produced germ tubes. Percent germination pooled across all factors significantly increased from 15.7% after 2 hr to 28.2% after 6 hr and 45.0% after 14 hr. Germination at the extreme temperatures tested, 24.5% at 13°C and 14.8% at 33°C, was significantly lower than 40.0, 47.2, and 36.5% recorded at 18, 23, and 28°C, respectively. Percent germination increased with increasing humidity from 2.8% at SD 1.2 to 60.9% at SD 0. Although germination was observed when both light and dark conditions were tested, the 44.9% germination recorded under continuous darkness after 6 hr was significantly higher than 20.3% under continuous light. Rate of germination of capilliconidia declined with increasing storage time from 93.0% before storage to a pooled mean of 81.0% after 1 day and 3.5% after 10 days. Higher germination rates were maintained at lower temperatures. Whereas germination after 7 days did not exceed 8.0% at 28 and 33°C, it was 85.9 at SD 10 in the dark at 18°C. Germination at SD 10 was significantly higher (43.3%) than at SDs 6, 2, and 0 (between 31.34 and 34.95%). Continuous light reduced viability as an average of 47.1% of the conidia maintained in the dark germinated, compared to 23.5% of those in the light.

Journal Article↗
PubMed1978

Seed germination as a thermobiological problem.

Thermal effects on seed germination are considered through the changes brought about by temperature in the germination capacity, in the germination rate, and in the distribution of the relative frequency of germination along the incubation times. A number of questions of general thermobiological interest are thus raised, entailing the need of an analysis of the temperature dependence of the seed germination rate. A treatment of these rates by the activation-energy approach cannot be general, for their Arrhenius plots are not always linear. Moreover, it is shown that any process displaying a temperature optimum (as happens in the germination of most seed species) cannot follow one of the fundamental tenets of the collision rate theory. The need of a theoretical treatment stressing the essential role of the partition of energy within the seed system has led to an anlysis using the absolute reaction rate theory. New experimental prospects for the physiology of seed germination are thus raised, concerning the meaning of the temperature cardinal points, the growth pattern of the embryo in germinating seeds, the dual effect of protein thermodenaturation, the effects of high hydrostatic pressures, and a whole pharmacological line of work. The cybernetic counterpart of the thermodynamic view of seed germination appears in the study of the distribution of the relative frequency of germination along the isothermal incubation time. In some species of seeds the thermal communication between the environment and the seed growth effector can be shown to proceed by molecular collisions at all germination isotherms. In the seeds of Dolichos biflorus this communication through random thermal noise prevails only at temperatures close to both extreme limits of germination. Both in this species and in Calotropis procera there is a temperature range (encompassing the optimum) within which a temperature signal is superimposed upon the gaussian noise. An interpretation is proposed according to which the temperature signal is transduced in a protein-conformation code.

Protein Denaturation↗
PubMed1998

Influence of several environmental factors on the initiation of germination and inactivation of Bacillus cereus by high hydrostatic pressure.

The influence of pH, aw, L-alanine, and fat concentration of milk on the initiation of germination and inactivation by high hydrostatic pressure (HHP) (250 mPa at 25 degrees C for 15 min and 690 mPa at 40 degrees C for 2 min) of Bacillus cereus sporulated at 20, 30 and 37 degrees C was investigated. B. cereus sporulated at the lowest temperature was found to be the most resistant to the initiation of germination and inactivation by HHP. At ambient pressure, the rate and extension of germination induced by L-alanine were also lower in B. cereus sporulated at 20 than 30 or 37 degrees C. The optimum pH for the germination and inactivation of B. cereus depended on the sporulation temperature. At 250 mPa the extent of germination for the three suspensions increased with higher pH. At 690 mPa, the pH barely affected the germination of B. cereus sporulated at 20 degrees C (3 log cycles), but the inactivation increased as the pH of the medium was lowered. After the same treatment, pressure optimally initiated the germination of B. cereus sporulated at 30 and 37 degrees C (6-7 log cycles) around neutral pH. Higher inactivation was obtained at pH 6. High concentrations of sucrose protected the spores from the germinating and inactivating effect of HHP. At aw 0.92, no germination was detected when the spores were pressurized at 250 mPa, and only 1 log cycle of B. cereus sporulated at 20 and 30 degrees C and 2 log cycles of B. cereus sporulated at 37 degrees C were germinated at 690 mPa. In addition, no inactivation was observed at aW 0.92 even at 690 mPa. The presence of L-alanine in the medium of pressurization increased the germination initiated by HHP at 250 mPa, but not at 690 mPa. A combination of 250 mPa at 25 degrees C with L-alanine (100 mM) was found to give an additive response. The initiation of germination and inactivation by HHP were not affected by the fat concentration of the milk.

Alanine↗
PubMed2002

Use of a novel method to characterize the response of spores of non-proteolytic Clostridium botulinum types B, E and F to a wide range of germinants and conditions.

AIMS: Limited information is available on the germination triggers for spores of non-proteolytic Clostridium botulinum. An automated system was used to study the effect of a large number of potential germinants, of temperature and pH, and aerobic and anaerobic conditions, on germination of spores of non-proteolytic Cl. botulinum types B, E and F. METHODS AND RESULTS: A Bioscreen analyser was used to measure germination by decrease in optical density. Results were confirmed by phase-contrast light microscopy. Spores of strains producing type B, E and F toxin gave similar results. Optimum germination occurred in L-alanine/L-lactate, L-cysteine/L-lactate and L-serine/L-lactate (50 mmol l(-1) of each). A further 12 combinations of factors induced germination. Sodium bicarbonate, sodium thioglycollate and heat shock each enhanced germination, but were not essential. Germination was similar in aerobic and anaerobic conditions. The optimum pH range was 5.5-8.0, germination occurred at 1-40 degrees C, but not at 50 degrees C, and was optimal at 20-25 degrees C. CONCLUSIONS: The automated system enabled a systematic study of germination requirements, and provided an insight into germination in spores of non-proteolytic Cl. botulinum. SIGNIFICANCE AND IMPACT OF THE STUDY: The results extend understanding of germination of non-proteolytic Cl. botulinum spores, and provide a basis for improving detection of viable spores.

Aerobiosis↗
PubMed2002

Analysis of spore cortex lytic enzymes and related proteins in Bacillus subtilis endospore germination.

The location and function of recognized cortex-lytic enzymes of Bacillus subtilis have been explored, and the involvement in germination of a number of related proteins tested. The SleB and CwlJ proteins are cortex-lytic enzymes, partially redundant in function, that are required together for effective cortex hydrolysis during B. subtilis spore germination. Spores were fractionated, and Western blotting of individual fractions suggests that the CwlJ protein is localized exclusively to the outer layers, or integument. The second spore-lytic enzyme, SleB, is localized both in the inner membrane of the spore and in the integument fraction. Neither protein changes location or size as the spore germinates. The ypeB gene is the second gene in a bicistronic operon with sleB. The SleB protein is absent from ypeB mutant spores, suggesting that YpeB is required for its localization or stabilization. In fractions of wild-type spores, the YpeB protein is found in the same locations as SleB - in both the inner membrane and the integument. As the absence of CwlJ protein does not affect the overall RP-HPLC profile of peptidoglycan fragments in germinating spores, this enzyme's hydrolytic specificity could not be defined. The effects of inactivation of several homologues of cortex-lytic enzymes of as yet undefined function were examined, by testing null mutants for their germination behaviour by OD(600) fall and by RP-HPLC of peptidoglycan fragments from dormant and germinating spores. The YaaH enzyme is responsible for a likely epimerase modification of peptidoglycan during spore germination, but the loss of this activity does not appear to affect the spore's ability to complete germination. Unlike the other cortex-lytic enzymes, the YaaH protein is present in large amounts in the spore germination exudate of B. subtilis. Mutants lacking either YdhD or YvbX, both homologues of YaaH, had no detectable alteration in either dormant or germinating spore peptidoglycan, and germinated normally. The ykvT gene, which encodes a protein of the SleB/CwlJ family, has no apparent association with germination: the gene is expressed in vegetative cells, and mutants lacking YkvT have no detectable phenotype.

Amidohydrolases↗
PubMed1992

Appearance of Endoproteolytic Enzymes during the Germination of Barley.

Barley endoproteolytic enzymes are important to germination because they hydrolyze endosperm storage proteins to provide precursors for new protein synthesis. We recently developed an electrophoretic method utilizing gel-incorporated protein substrates to study the endoproteinases of 4-d-germinated barley (Hordeum vulgare L. cv Morex) grain. This work extends those findings to determine the temporal pattern of the appearance of the endoproteinases during germination, the sensitivities of the proteinases to class-specific proteinase inhibitors, and where, in germinating caryopses, the proteinases reside. Six endoproteinase activity bands (representing a minimum of seven enzymes) were present in 5-d-germinated barley grain extracts subjected to electrophoresis in nondenaturing gels at pH 8.8. The activities of two of the enzyme bands ("neutral" proteinases) increased as the pH was increased from 3.8 to 6.5. The activities of the remaining four ("acidic") bands diminished abruptly as the pH increased above 4.7. Two proteinase bands hydrolyzed gelatin but not edestin, four of the proteinases hydrolyzed both gelatin and edestin at nearly the same rates, and one enzyme degraded only edestin. One neutral endoproteinase was sensitive to diisopropyl fluorophosphate inhibition, and the other was not inhibited by any of inhibitors tested. Four of acidic enzymes were cysteine proteinases [inhibited by trans-epoxysuccinyl-l-leucylamido(4-guanidino)butane and N-ethylmaleimide]; the other was an aspartic acid endoproteinase (sensitive to pepstatin). Only the aspartic proteinase was detected in either ungerminated or steeped barley grain. During the germination (malting) process, the aspartic endoproteinase activity decreased until the second day of germination and then increased until germination day 5. The first endoproteinase(s) induced during germination was a neutral enzyme that showed activity on the 1st day of the germination phase after steeping. Most of the endoproteinases became active on the 2nd or 3rd germination day, but one cysteine proteinase was not detected until the 5th day. Acid cysteine proteinases were present in the aleurone, scutellum, and endosperm tissues but not in shoots and roots. The aleurone layer and endosperm contained almost exclusively band B1 neutral proteinases, whereas the scutellum, shoots, and roots contained both B1 and B2 bands. This work shows that germinating barley contains a complex set of proteinases whose expression is temporally and spatially controlled. But, at the same time, it also shows that this electrophoretic method for separating and studying individual enzymes of this complex will allow us to more readily characterize and purify them.

Journal Article↗
PubMed1986

Requirement for Ethylene Synthesis and Action during Relief of Thermoinhibition of Lettuce Seed Germination by Combinations of Gibberellic Acid, Kinetin, and Carbon Dioxide.

Application of exogenous ethylene in combination with gibberellic acid (GA(3)), kinetin (KIN), and/or CO(2) has been reported to induce germination of lettuce seeds at supraoptimal temperatures. However, it is not clear whether endogenous ethylene also plays a mediatory role when germination under these conditions is induced by treatment regimes that do not include ethylene. Therefore, possible involvement of endogenous ethylene during the relief of thermoinhibition of lettuce (Lactuca sativa L. cv Grand Rapids) seed germination at 32 degrees C was investigated. Combinations of GA(3) (0.5 millimolar), KIN (0.05 millimolar), and CO(2) (10%) were used to induce germination. Little germination occurred in controls or upon treatment with ethylene, KIN, or CO(2). Neither KIN nor CO(2) affected the rate of ethylene production by seeds. Both germination and ethylene production were slightly promoted by GA(3). Treatments with GA(3)+CO(2), GA(3)+KIN, or GA(3)+CO(2)+KIN resulted in approximately 10-to 40-fold increases in ethylene production and 50 to 100% promotion of germination as compared to controls. Initial ethylene evolution from the treated seeds was greater than from the controls and a major surge in ethylene evolution occurred at the time of visible germination. Application of 1 millimolar 2-aminoethoxyvinyl glycine (AVG), an inhibitor of ethylene synthesis, in combination with any of above three treatments inhibited the ethylene production to below control levels. This was accompanied by a marked decline in germination percentage. Germination was also inhibited by 2,5-norbornadiene (0.25-2 milliliters per liter), a competitive inhibitor of ethylene action. Application of exogenous ethylene (1-100 microliters per liter) overcame the inhibitory effects of AVG and 2,5-norbornadiene on germination. The results demonstrate that endogenous ethylene synthesis and action are essential for the alleviation of thermoinhibition of lettuce seeds by combinations of GA(3), KIN, and CO(2). It also appears that these treatment combinations do not act exclusively via promotion of ethylene evolution as the application of exogenous ethylene alone did not promote germination.

Journal Article↗
PubMed2006

Development of a population-based threshold model of conidial germination for analysing the effects of physiological manipulation on the stress tolerance and infectivity of insect pathogenic fungi.

Entomopathogenic fungi are being used as biocontrol agents of insect pests, but their efficacy can be poor in environments where water availability is reduced. In this study, the potential to improve biocontrol by physiologically manipulating fungal inoculum was investigated. Cultures of Beauveria bassiana, Lecanicillium muscarium, Lecanicillium longisporum, Metarhizium anisopliae and Paecilomyces fumosoroseus were manipulated by growing them under conditions of water stress, which produced conidia with increased concentrations of erythritol. The time-course of germination of conidia at different water activities (water activity, aw) was described using a generalized linear model, and in most cases reducing the water activity of the germination medium delayed the onset of germination without affecting the distribution of germination times. The germination of M. anisopliae, L. muscarium, L. longisporum and P. fumosoroseus was accelerated over a range of aw levels as a result of physiological manipulation. However, the relationship between the effect of physiological manipulation on germination and the osmolyte content of conidia varied according to fungal species. There was a linear relationship between germination rate, expressed as the reciprocal of germination time, and aw of the germination medium, but there was no significant effect of fungal species or physiological manipulation on the aw threshold for germination. In bioassays with M. anisopliae, physiologically manipulated conidia germinated more rapidly on the surface of an insect host, the melon cotton aphid Aphis gossypii, and fungal virulence was increased even when relative humidity was reduced after an initial high period. It is concluded that physiological manipulation may lead to improvements in biocontrol in the field, but choice of fungal species/isolate will be critical. In addition, the population-based threshold model used in this study, which considered germination in terms of physiological time, also called hydrotime, could have general application in mycology and environmental microbiology.

Animals↗
PubMed1985

Germination of spores from Clostridium botulinum B-aphis and Ba410.

The germination of spores from Clostridium botulinum B-aphis and Ba410 was examined. In a complex medium, heat activation of spores from both strains doubled the germination rates and was required for germination in the presence of 2% NaCl. In a defined medium (CTB [D. B. Rowley and F. Feeherry, J. Bacteriol. 104:1151-1157, 1970]), the parent strain B-aphis germinated at a rate of 0.77% min-1 in the absence of NaCl and was not affected by 2% NaCl. A salt-tolerant derivative, strain Ba410, germinated at rates of 0.16% min-1 in CTB and 0.04% min-1 in CTB containing 2% NaCl. L-Alanine-triggered spores germinated faster than did L-cysteine-triggered spores from both strains. When both amino acids were present, B-aphis germinated rapidly in the absence of NaCl and had biphasic kinetics in the presence of NaCl. Strain Ba410 had biphasic kinetics in the absence of NaCl and germinated slowly with single-phase kinetics in the presence of NaCl. L-Alanine- and L-cysteine-triggered germinations were each inhibited by both D-alanine and D-cysteine, indicating a common germinant-binding site for both alanine and cysteine. Attempts to select for variants with amino acid-specific germinant-binding sites were unsuccessful. Differences in the germination kinetics of both strains could not be explained by ultrastructural differences. Transmission electron micrographs revealed striking similarities between the strains.

Clostridium botulinum↗
PubMed2000

Comparative study of pressure- and nutrient-induced germination of Bacillus subtilis spores.

Germination experiments with specific germination mutants of Bacillus subtilis, including a newly isolated mutant affected in pressure-induced germination, suggest that a pressure of 100 MPa triggers the germination cascades that are induced by the nutrient germinant alanine (Ala) and by a mixture of asparagine, glucose, fructose, and potassium ions (AGFK), by activating the receptors for alanine and asparagine, GerA and GerB, respectively. As opposed to germination at 100 MPa, germination at 600 MPa apparently short-cuts at least part of the Ala- and AGFK-induced germination pathways. Inhibitors of nutrient-induced germination (HgCl(2) and Nalpha-P-tosyl-L-arginine methyl ester) also inhibit pressure-induced germination at 600 MPa, suggesting that germination at 600 MPa involves activation of a true physiological germination pathway and is therefore not merely a physico-chemical process in which water is forced into the spore protoplast.

Alanine↗
PubMed1975

Isolation, characterization, and mapping of Bacillus subtilis 168 germination mutants.

After mutagenesis with nitrosoguanidine, germination mutants of Bacillus subtilis 168 were selected by killing, with heat, spores that germinated at 42 C and collecting survivors at 30 C. The germination properties of nine mutants variously affected in amino acid biosynthesis and sugar utilization were studied in detail. They were divided into two groups: (i) Ger-ALA mutants, failed to germinate in 10 mM L-alanine but germinated in complex media (some of these mutants were temperature sensitive); (ii) Ger-PAB mutants, germinated poorly, even in complex media, suggesting that they were blocked in important germination functions. All the mutants failed to germinate in L-alpha-amino-n-butyrate or L-valine (including temperature-sensitive mutants only at the restrictive temperature) showing that there is a step necessary for germination affected by all three acids. The mutants had normal growth rates, indicating that the defective gene products were specific for germination functions. These defects were not identified. Eight of the mutants were mapped by transduction with phage PBS-1. The recombinants were scored either by observations, by microscopy of phase darkening of the spores, or by a plate test involving the reduction of tetrazolium by heated colonies of spores. Five of the mutations, of at least three phenotypes, were between thr-5 and cysB3 away from all the sporulation markers that have been previously mapped. A linked ald (alanine dehydrogenase) locus was on the other side of thr-5. The other Ger markers were located in at least two additional positions. Auxotrophic strains that were used for mapping germinated normally, but germination of the Ger mutants differed slightly in different genetic backgrounds.

Bacillus subtilis↗
PubMed1980

Germination properties as marker events characterizing later stages of Bacillus subtilis spore formation.

At various stages during spore formation sporangia were shocked by cold treatment or with toluene, and the germination requirements of the prespores were examined. Up to 5 h after induction of sporulation (t5) germination was spontaneous; i.e., it occurred without any added germinants. After t5, during stages V and VI, the capacity for spontaneous germination diminished progressively, and the spores acquired a need for externally added germinants. At t6 this need was satisfied by either L-alanine or a mixture of KCl, glucose, and fructose. By t8, the latter response had disappeared. The spores germinated only with L-alanine, and the response was much slower. Experiments with chloramphenicol showed that the germination properties of the spores appearing between t6 and t8 were the expression of events in protein synthesis that had occurred before t5. Although the germination requirements developed at about the same time as heat resistance, they could be dissociated from heat resistance in wild-type and mutant cells. The germination properties of the developing spores are additional marker events characterizing the later stages of sporulation, as follows: (i) spontaneous germination (up to the end of stage IV); (ii) germination requirements that are satisfied by KCl-glucose-fructose or L-alanine (stage V); and (iii) slow germination response with L-alanine only (stage VI).

Alanine↗
PubMed1964

RESPONSES OF BACILLUS SUBTILIS SPORES TO IONIC ENVIRONMENTS DURING SPORULATION AND GERMINATION.

Fleming, H. P. (University of Illinois, Urbana), and Z. John Ordal. Responses of Bacillus subtilis spores to ionic environments during sporulation and germination. J. Bacteriol. 88:1529-1537. 1964.-The ionic environments of germination and sporulation menstrua had a prominent influence on characteristics of Bacillus subtilis spores. There was a synergistic effect for l-alanine and inorganic ions on spore germination. The maximal rate of germination in solutions of l-alanine was dependent on ionic concentration and species. Germination was negligible in l-alanine at low ionic strength but increased as the ionic strength was increased up to about 10(-1)m with a variety of salts. Phosphate was the most active ion tested, and divalent cations were the least active in supporting germination in l-alanine. Germination progressed slowly at 45 C in sodium chloride or sodium phosphate alone but not in CaCl(2) alone. Germination rates in l-alanine were retarded at high ionic strengths (mu in the range of 0.1 to 1.0). Inhibitory effects of high concentrations of certain divalent cations on germination were related to the binding abilities of these metals. High concentrations of NaCl (10(-1) to 1.0 m) in the sporulation medium resulted in lowered heat resistance and germination rate of the resulting spores. The addition of calcium (5 x 10(-2)m CaCl(2)) to the sporulation medium relieved the repression of NaCl on germination and caused the spores to have a greater heat resistance. Calcium and dipicolinic acid (DPA) contents of the spores were unaffected by NaCl in the sporulation medium. The calcium, but not the DPA, content of spores increased as a result of supplementing the sporulation medium with calcium. Possible roles of ions in the germination of spores are discussed.

Alanine↗