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Carbon catabolism and synthesis of macromolecules during spore germination of Microsporum gypseum.

Either d- or l-leucine (10(-3)m) and unsaturated long-chain fatty acids such as oleic, linoleic, and arachidonic (10(-4)m) significantly stimulated macroconidia germination of Microsporum gypseum. Saturated long-chain fatty acids did not affect germination, whereas saturated short-chain fatty acids such as caprylic, hexanoic, and butyric were completely inhibitory. Germination was followed by an increase in endogenous respiration and a decrease in dry weight of approximately 5% at 4 hr. Endogenous fatty acids and soluble carbohydrates were utilized significantly during germination. Tritiated leucine, uridine, and thymidine were incorporated respectively into protein, ribonucleic acid (RNA), and deoxyribonucleic acid (DNA) fractions within the first 5 min of germination. Incorporation of oleic-1-C(14) into RNA and protein was significantly increased after germ tube development. Net synthesis of RNA and protein started prior to germ tube protrusion. Increase in DNA could be detected only later. A significant increase in RNA and protein during the 4th hr of germination was correlated with vegetative development. Inhibition of respiration and incorporation of leucine-H(3) and uridine-H(3) into corresponding macromolecules by dl-fluorophenylalanine and phenethyl alcohol started before germ tube appearance. Griseofulvin significantly inhibited incorporation of uridine-H(3) and thymidine-H(3), but not of leucine-H(3). This inhibition occurred only after initial vegetative development. In contrast to the two other inhibitors, which substantially inhibited germination, griseofulvin only slightly retarded the period of germination and did not affect respiration.

Alcohols↗

Comparative study of susceptibilities of germinated and ungerminated conidia of Aspergillus fumigatus to various antifungal agents.

Conidia are used as inocula for the in vitro susceptibility testing of Aspergillus fumigatus. Since the MIC is defined on the basis of visible mycelial growth, conidia should germinate and produce sporelings (germinated conidia) for monitoring of the growth inhibition and fungicidal activity of a drug. If a compound is capable of inhibiting germination of conidia while affecting or not affecting the growth of the organism, the MIC obtained will be the concentration of the drug required for the inhibition of conidial germination but not necessarily that required for inhibition of the growth of the organism. We investigated the susceptibility of germinated and ungerminated conidia to amphotericin B, itraconazole, voriconazole, and SCH56592. The MICs of various antifungal agents for germinated conidia were almost identical to those obtained for ungerminated conidia. In addition, both the germinated and ungerminated conidia were killed with almost equal efficiency by all of the compounds tested when exposed to the drugs for 24 h. These results suggest that either germinated or ungerminated conidia could be used as inocula for in vitro susceptibility studies of A. fumigatus with identical results.

Amphotericin B↗

Characterization of cDNA clones specific for sequences developmentally regulated during Dictyostelium discoideum spore germination.

Spore germination in the slime mold Dictyostelium discoideum was used as a model to study the developmental regulation of protein and mRNA synthesis. Changes in the synthesis of these macromolecules occur during the transition from dormant spore to amoebae. The study of the mechanisms which regulate the quantity and quality of protein synthesis can best be accomplished with cloned genes. cDNA clones which hybridized primarily with mRNAs from only spores or germinating spores and not with growing amoebae were collected. Three such clones, denoted pLK109, pLK229, and pRK270, were isolated and had inserts of approximately 500, 1,200, and 690 base pairs, respectively. Southern blot hybridization experiments suggested that each of the genes is present in multiple copies in the D. discoideum genome. RNA blot hybridizations were performed to determine the sizes of the respective mRNAs and their developmental regulation. The mRNA that hybridized to pLK109 DNA was present predominantly in spores and at 1 h after germination but was absent in growing amoebae. Its concentration dramatically dropped at 3 h. The mRNA present in spores is apparently larger (approximately 0.5 kilobase) than in the later stages of germination (0.4 kilobase), indicating processing of the RNA during germination. The mRNA that hybridized to pLK229 DNA was approximately 1.0 kilobase and was present in very low amounts during growth. Its concentration rose until 1 h after spore germination and decreased thereafter. pRK270-specific RNA was approximately 2.7 kilobases and was found predominantly at 1 h after germination. It was present in lower concentrations at 2 and 3 h after germination and was absent in spores and amoebae. In vitro translation of mRNA selected from 1-h polyadenylated RNA which was hybridized to pLK109 or pLK229 DNA gave proteins of molecular weights consistent with the sizes of the mRNAs as determined by the RNA blot analysis.

Cloning, Molecular↗

Ultrastructure and lipid identification during conidium germination of Stemphylium sarcinaeforme.

Multicelled conidia of Stemphylium sarcinaeforme germinate in water forming several germ tubes. Individual cells within conidia are connected by pores which are plugged in ungerminated conidia and open in germinated ones. During germination, vacuoles enlarge, endoplasmic reticulum profiles increase in number, and mitochondria change from spherical to elongate. The germ tube wall is laid down at the site of emergence from the conidium. Shortly after germination, a septum with a central pore forms where ther germ tube emerged. The germ tube wall is surrounded by a fibrillar sheath. Lipid bodies are closely associated with vacuoles during germination. The ultrastructural location of lipid was found by extraction of conidia with lipid solvents. Total lipid decreases from 14.4% of the dry weight of ungerminated conidia to 13.4% of the dry weight of conidia germinated for 10 h. No qualitative changes occurred in the major lipid classes of conidia during germination. The activities of lipase and acid phosphatase were detected in ungerminated and germinated conidia.

Acid Phosphatase↗

The role of Ca2+ during spore germination in Dictyostelium: autoactivation is mediated by the mobilization of Ca2+ while amoebal emergence requires entry of external Ca2+.

One of the developmental pathways used by the social amoeba Dictyostelium discoideum produces dormant spores. As with any temporary resistant stage, these spores must be able to germinate rapidly in response to positive environmental stimuli. One such stimulus is the autoactivator, an endogenous, diffusible molecule that is secreted by spores. Previous work has shown that three phases of germination, autoactivation, spore swelling and amoebal emergence, require the activity of the Ca(2+)-dependent, regulatory protein calmodulin, implicating Ca2+ as an essential cation during germination. In this study we used a pharmacological approach coupled with the direct measurement of Ca2+ levels in germinating spore populations by atomic adsorption to examine Ca(2+)-dependent signal transduction during spore activation and germination in D. discoideum. Inhibitors of both phospholipase C and internal Ca2+ release inhibited autoactivation while exogenously added Ins(1,4,5)P3, acted synergistically with the autoactivator. The antagonists specifically affected spore activation as mediated by the autoactivator, since neither had any effect on heat-activated spores. In contrast, La3+, an inhibitor of Ca2+ uptake, had little or no effect on either autoactivation or the swelling of autoactivated spores. However, an inhibition of Ca2+ influx by La3+ inhibited both the swelling of heat-activated spores and amoebal emergence following each period of autoactivation or heat activation. Ca2+ levels change in the spore population during germination. During activation and swelling, Ca2+ efflux occurs from the spores. Both of the activating stimuli used here, the autoactivator and heat, caused this Ca2+ efflux. The efflux is reversed during emergence when there is a net Ca2+ uptake by the spores and cells from the medium. Together these data provide the first evidence that autoactivation is mediated by Ca(2+)-dependent signal transduction, leading to Ca2+ efflux, and that the late event of germination, amoebal emergence, requires Ca2+ uptake to proceed. The data also suggest that the responses of the spore to the each of autoactivator and heat, i.e. Ca2+ movements and germination, are mediated by different mechanisms.

Animals↗

Nutritional and physiological factors affecting germination of heterothallic Saccharomyces cerevisiae ascospores.

Saccharomyces cerevisiae strain AP-3 was examined with respect to those nutritional requirements and physiological conditions which influence its germination rate. It was found that glucose as a carbon source supported the most rapid rate of germination for this heterothallic strain. In contrast, strain AP-3 spore germination was supported the least by the carbon sources potassium acetate and lactose. Of the nitrogen sources tested in culture medium containing glucose, the complex nitrogen sources peptone and casein hydrolysate appeared to be capable of stimulating germination better than a control culture containing ammonium sulphate. None of the amino acids screened were found to stimulate strain AP-3 germination compared with ammonium sulphate. The optimal culture medium pH for ascospore germination was 4.5 although spore germination could still be initiated by glucose between pH 3.0 and pH 7.5. Germination initiation by glucose was observed over a temperature range from 25 degrees C to 50 degrees C, but the optimal temperature appeared to be 40 degrees C.

Hydrogen-Ion Concentration↗

Effects of basic oxygen furnace slag and inorganic nutrients on the germination of resting cysts of two toxic dinoflagellates.

Effects of basic oxygen furnace (BOF) slag, inorganic nutrients and H2S on the germination of resting cysts of two toxic dinoflagellates Alexandrium catenella/tamarense and Gymnodinium catenatum were studied in batch cultures. The germination rate of the test species has increased by 23-25%, when the concentration of NO3--N or H2S in culture medium has increased to 2.0 ppm. At the treatment of enriched NH4+-N and PO43--p, the germination of resting cyst was increased. Nevertheless, the increased range in germination rates was less than those of NO3--N and H2S. When BOF slag in culture medium increased to 50 mg/ml (or 500 g/m2), the cyst germination rate fell to less than 5%. At higher level of concentrations germination was completely inhibited. Adding BOF slag to the culture medium reduced the concentration of inorganic salts and H2S in seawater and sediments, resulting in the inhibition of cyst germination. These findings demonstrate the potential use of BOF slag on the sediments seed bank of red tide organism because it has an ability to inhibit resting cysts germination.

Animals↗

Evolving abundance and clonal pattern of human germinal center B cells during childhood.

Childhood is a critical period for the development of the memory B-lymphocyte repertoire necessary in protective humoral immunity. This study addressed the natural history of memory B cells based on the previous identification of germinal center and mantle zone cells as the probable precursor and mature memory cell populations, respectively. Using flow cytometric quantitation of these B-cell subpopulations in human tonsil, we found that germinal center cells were abundant (70% of tonsil B cells) during early childhood (2 to 3 years), but decline by early adolescence (8 to 14 years) to a low level (33%, P = .0003). To study the clonal evolution of these B-cell subpopulations, germinal center and mantle zone B cells were isolated using a preparative magnetic immunobead method, and analyzed using a novel polymerase chain reaction-based quantitative assay to measure the abundance of B-cell clones bearing certain rearranged VH subfamilies. Two VH subfamilies were informative: VH1N clones were uniquely deficient in germinal center B cells at the early age period, but became abundant in later childhood; and VH3L clones were absent among germinal center cells regardless of age. In contrast, B-cell clones bearing each VH subfamily were abundant in the mantle zone subpopulation throughout childhood. These findings suggest that the abundance and clonal pattern of germinal center B cells evolves during childhood, presumably due to changing antigenic or ontogenic processes. Moreover, the distinct clonal pattern of germinal center versus mantle zone B cells suggests that a major phase of clonal selection occurs after germinal center emigration.

Adolescent↗

Temperature and seed germination.

Temperature can affect the percentage and rate of germination through at least three separate physiological processes. 1. Seeds continuously deteriorate and, unless in the meanwhile they are germinated, they will ultimately die. The rate of deterioration depends mainly on moisture content and temperature. The Q10 for rate of loss of viability in orthodox seeds consistently increases from about 2 at -10 degrees C to about 10 at 70 degrees C. 2. Most seeds are initially dormant. Relatively dry seeds continuously lose dormancy at a rate which is temperature-dependent. Unlike enzyme reactions, the Q10 remains constant over a wide range of temperature at least up to 55 degrees C, and typically has a value in the region of 2.5-3.8. Hydrated seeds respond quite differently: high temperatures generally reinforce dormancy or may even induce it. Low temperatures may also induce dormancy in some circumstances, but in many species they are stimulatory (stratification response), especially within the range -1 degree C to 15 degrees C. Small, dormant, hydrated seeds are usually also stimulated to germinate by alternating temperatures which typically interact strongly and positively with light (and often also with other factors including nitrate ions). The most important attributes of alternating temperatures are amplitude, mean temperature, the relative periods spent above and below the median temperature of the cycle (thermoperiod) and the number of cycles. 3. Once seeds have lost dormancy their rate of germination (reciprocal of the time taken to germinate) shows a positive linear relation between the base temperature (at and below which the rate is zero) and the optimum temperature (at which the rate is maximal); and a negative linear relation between the optimal temperature and the ceiling temperature (at and above which the rate is again zero). The optimum temperature for germination rate is typically higher than that required to achieve maximum percentage germination in partially dormant or partially deteriorated seed populations. None of the sub-cellular mechanisms which underlie any of these temperature relations are understood. Nevertheless, the temperature responses can all be quantified and are fundamental to designing seed stores (especially long term for genetic conservation), prescribing germination test conditions, and understanding seed ecology (especially that required for the control of weeds).

Plant Development↗

Identification and biological activity of germination-inhibiting long-chain fatty acids in animal-waste composts.

Long-chain fatty acids in germination-inhibiting animal-waste composts were identified by gas chromatography-mass spectrometry as myristic, palmitic, stearic, oleic, linoleic, and linolenic acids. These acids were found at concentrations greater than 0.25 mg (g dry compost)-1. The identified acids, together with lauric acid, and five kinds of short- and medium-chain fatty acid, were tested for their effects on the germination process of sorghum seeds. The authentic long-chain fatty acids, which were dissolved in a 1:9 (v/v) mixture of methanol and distilled water at 40 mg liter-1, significantly reduced the alpha-amylase activity, physiological water uptake, and ATP content of the germinating seeds during the first 24 h of inhibition, as well as the rate of germination of seeds. Among the tested fatty acids, myristic and palmitic acids were the most potent inhibitors of germination. The inhibitory effects of long-chain fatty acids were stronger than those of the phenolic acids. The short- and medium-chain fatty acids did not have any significant germination-inhibitory effects at 40 mg liter-1. The results indicate that the long-chain fatty acids are the dominant inhibitors of germination in animal-waste composts, and that the inhibition of the alpha-amylase activity in germinating sorghum seeds is one aspect of the mode of action of these long-chain fatty acids.

Animals↗

Effect of soaking, germination, autoclaving and cooking on chemical and biological value of guar compared with faba bean.

Raw, soaked, germinated, autoclaved and cooked both of guar and faba bean were chemically and biologically evaluated. Crude protein, crude fiber and ash contents of guar and faba bean decreased as a result of all treatments with the exception of protein content in both germinated guar and faba bean. A slight decrease in lipid content was noticed during germination, while there were no changes in lipids after soaking, autoclaving and cooking. Protein content of guar was higher than of faba bean, and showed higher values for all amino acids than faba bean protein except for lysine and tryptophan. Germination caused a slight increase in total essential amino acids, while soaking caused a slight decrease in both of guar and faba bean. There was a slight decrease in the mineral contents by the heat processes with the exception of Mg and Mn for guar and Na, Mn for faba bean due to the cooking process. Germinated guar and faba bean showed noticeable decrease in Na, K, Cu, Mn and Mg and a slight increase in Fe and Zn than the raw materials. Germination was more effective in reducing phytic acid than heat treatment, and therefore it improves the nutritional quality of both guar and faba bean. Protein efficiency ratio (PER), true digestibility (DT) and biological value (BV) were estimated in all treatments under study using rat bioassay procedures. Data revealed pronounced improvements of the nutritive value as a result of all processes under study especially germination being the most effective.

Amino Acids↗

Limited period of graviresponsiveness in germinating spores of Ceratopteris richardii.

Rhizoids of the fern Ceratopteris richardii Brogn. usually emerge 40 h after germination is initiated by light, and more than 90% of them emerge growing in a downward direction. However, when the spores are germinated on a clinostat, the emerging rhizoids show no preferential orientation. This indicates that under normal 1 g conditions the initial growth direction of rhizoids can be oriented by gravity. If the orientation of the spores is changed 3 h or less after the start of germination, the growth direction of most emerging rhizoids becomes downward relative to the new orientation. However, if the orientation of the spores is changed by 180 degrees 8 h or more after germination is initiated by light, most rhizoids emerge growing upward; i.e., the same direction as if there had been no orientation change. Emerged rhizoids also do not change their direction of growth if their orientation is changed. These results indicate that the growth direction of emerging rhizoids is set by gravity prior to actual emergence, and that the time of full orientation responsiveness is limited to a period ranging from the initiation of germination to about 3-4 h after the start of germination. There is a gravity-oriented nuclear movement beginning at about 13 h after germination, and this movement appears to predict the initial growth direction of rhizoids.

Cell Nucleus↗

Estimating germinability of Plasmopara viticola oospores by means of neural networks.

Neural networks are trained to estimate the germination percentages of Plasmopara viticola oospores, overwintered in natural conditions in two viticultural areas in northern Italy, by using climatic (temperature and rainfall) data, as well as the previous germination measurement, as input variables. The 288 available patterns consist of a set of selected independent variables associated with the corresponding germination percentage. All 12 networks investigated converge to a non-linear relationship between the selected independent variables and oospore germination. The highest correlation coefficient (equal to 0.83) between the real and estimated germination percentages is obtained by considering, as input to the network, the climatic data (both temperature and rainfall) recorded during the 40 days before sampling and the germination percentage assessed in the germination assay carried out immediately before the present sampling.

Germination↗

Enhanced post-germinative growth of encapsulated somatic embryos of Siberian ginseng by carbohydrate addition to the encapsulation matrix.

This experiment was carried out to enhance conversion and ex vitro survival of encapsulated somatic embryos of Siberian ginseng (Eleutherococcus senticosus). Cotyledonary somatic embryos were encapsulated with 3.0% sodium alginate; 96% of the encapsulated embryos converted to plantlets with well-elongated epicotyls in Perlite containing sucrose as a carbon source. However, although they germinated, post-germinative growth of encapsulated embryos was suppressed on Perlite that did not contain sucrose. Instead of sucrose addition to Perlite, addition of carbon sources to the encapsulation matrix enhanced post-germinative growth of encapsulated embryos. In the encapsulation matrix with 2% sucrose, post-germinative growth of encapsulated embryos was more than twice (23.5%) that of the control capsules without sucrose (10.0%). Embryos encapsulated with both 2% sucrose and 1% starch powder showed the highest post-germinative growth percentage (42.1%). Iodine staining and analysis of starch content in the encapsulation matrix revealed that starch in the encapsulation matrix decomposed during embryo germination. This result indicates that carbohydrate treatment in the encapsulation matrix enhanced post-germinative growth of encapsulated embryos of Siberian ginseng.

Cell Culture Techniques↗

Seed size and germination response: a relationship for fire-following plant species exposed to thermal shock.

Thermal shock is well known to be an important stimulus for the germination of soil-stored seeds in fire-prone plant communities. Nevertheless, while the overall germination response of different species is known to vary, the interaction between seed size and germination to a range of thermal-shock temperatures is poorly understood. This interaction may be important in regulating post-fire plant community establishment, since larger seeds are able to emerge from deeper within the soil profile than smaller seeds, and are therefore likely to be insulated against high above-ground temperatures by a deeper soil covering. In this experiment we examined how germination of eight co-occurring Western Australian fire-followers was influenced by thermal shock, and whether germination was significantly correlated with seed size. We found that small-seeded species not only showed enhanced germination at higher temperatures, but that their ability to germinate at higher temperatures was also greater than that displayed by larger-seeded species. These findings suggest that while seed size may be a useful general predictor of post-fire recruitment success, under different fire regimes the interaction between seed size, maximum seedling emergence depth, and the ability to withstand different thermal-shock temperatures is complex and may confound recent predictive models.

Adaptation, Physiological↗

Effects of germinated barley foodstuff on dextran sulfate sodium-induced colitis in rats.

Germinated barley foodstuff (GBF), derived from the aleurone and scutellum fractions of germinated barley, is rich in glutamine and low-lignified hemicellulose, and increases mucosal protein, RNA, and DNA content in the intestine when fed to normal rats. The aim of this study was to evaluate the effects of feeding GBF or germinated gramineous seeds on experimental ulcerative colitis. Sprague-Dawley rats that received 3% dextran sulfate sodium in their diets were used as an experimental colitis model. The effects of sulfasalazine, a drug used to treat inflammatory bowel disease, were compared with those of GBF. After rats had consumed diets containing GBF or various aleurone and scutellum fractions, mucosal damage; the content of mucosal protein, RNA, and DNA in the colo-rectum; and serum interleukin-8 and alpha1-acid glycoprotein levels were assessed. GBF and germinated seeds more effectively prevented bloody diarrhea and mucosal damage in colitis compared with controls and rats receiving sulfasalazine, but non-germinated samples did not have a protective effect. GBF increased mucosal protein and RNA content in the colitis model. The consumption of GBF appears to prevent inflammation in a colitis model, and its effect seems to be related to the germination process. GBF and germinated seeds have the potential to serve as nutritional therapy for ulcerative colitis.

Animals↗

Alteration of the embryo transcriptome of hexaploid winter wheat (Triticum aestivum cv. Mercia) during maturation and germination.

Grain dormancy and germination are areas of biology that are of considerable interest to the cereal community. We have used a 9,155-feature wheat unigene cDNA microarray resource to investigate changes in the wheat embryo transcriptome during late grain development and maturation and during the first 48 h of postimbibition germination. In the embryo 392 mRNAs accumulated by twofold or greater over the time course from 21 days postanthesis (dpa) to 40 dpa and on through 1 and 2 days postgermination. These included mRNAs encoding proteins involved in amino acid biosynthesis and metabolism, cell division and subsequent cell development, signal transduction, lipid metabolism, energy production, protein turnover, respiration, initiation of transcription, initiation of translation and ribosomal composition. A number of mRNAs encoding proteins of unknown function also accumulated over the time course. Conversely 163 sequences showed decreases of twofold or greater over the time course. A small number of mRNAs also showed rapid accumulation specifically during the first 48 h of germination. We also examined alterations in the accumulation of transcripts encoding proteins involved in abscisic acid signalling. Thus, we describe changes in the level of transcripts encoding wheat Viviparous 1 (Vp1) and other interacting proteins. Interestingly, the transcript encoding wheat Viviparous-interacting protein 1 showed a pattern of accumulation that correlates inversely with germination. Our data suggests that the majority of the transcripts required for germination accumulate in the embryo prior to germination and we discuss the implications of these findings with regard to manipulation of germination in wheat.

Computational Biology↗

Germination characteristics of native Japanese dandelion autopolyploids and their putative diploid parent species.

Germination characteristics of native Japanese Taraxacum lineages of Taraxacum platycarpum (diploid), T. venustum (triploid and tetraploid), and T. albidum (pentaploid) have been studied at different temperatures. Taraxacum platycarpum ssp. hondoense is the putative diploid parent of T. venustum. Diploid T. platycarpum ssp. hondoense and the polyploids T. venustum and T. albidum are found in different areas of Japan, and distribution differences may reflect divergent ecological and physiological traits among ploidy levels. In this study, to prevent mixing of seeds of different polyploidy we used flow cytometry to examine the ploidy level of the plants from which seeds were collected. Results from seed-germination experiments showed that dependence on temperature of final percentage germination was qualitatively similar for both autopolyploids T. venustum and diploids T. platycarpum--germination was suppressed at high and low temperatures. It was also shown that seed germination of autopolyploids was suppressed more than that of the ancestral diploid at low temperatures and that seed germination for polyploids was higher than for the diploid. Threshold variations at low temperatures might affect the distribution of native dandelions. Taraxacum venustum, which occurs in cool climates, might have developed a distinctly lower germination threshold at low temperatures whereas T. albidum, which is native to warm climates, might have developed an adaptive threshold at high temperatures.

Adaptation, Physiological↗