Subject index

Barley seeds were treated with ethyl methanesulphonate (EMS) for 5 h, washed for 18 h, exposed for 3 h to caffeine, re-dried to 30% water content and either immediately sown or stored for 10 days at 25°. The caffeine post-treatment considerably potentiated the EMS-induced M₁ germination and the M₁ seedling height reduction, but had no influence on the EMS-induced M₁ sterility or percentage of M₂ chlorophyll mutants. The caffeine post-treatment also had no significant influence on the “storage” recovery from the EMS-induced genetic effects.     

Storage of ethyl methanesulphonate-treated barley seeds with 15 per cent moisture. Influence of treatment and washing conditions

Barley seeds were treated with ethyl methanesulphonate (EMS), washed for 24 h, redried to 15 per cent moisture and stored at 25°C. The criteria used for expressing the effect of storage were chromosomal aberrations in root tips, M₁ germination, M₁ seedling height, M₁ survival and the frequency of M₂ chlorophyll mutants. The increase of the M₁ biological injury due to storage was not influenced:

1. by applying EMS solutions at pH 2, pH 7 and pH 10,
2. by lowering the EMS concentration and increasing the treatment time,
3. by different variations of washing with water and by washings with 0.005 N NaOH, 200 mM cysteine or 200 mM thiourea. The rate of the increase of the M₁ injury due to storage depends on the EMS dose. With a decrease in the EMS dose the storage effect is more delayed.

     

Viability and virulence of blastospores of Metarhizium anisopliae (Metch.) Sorokin after storage in various liquids at different temperatures

Blastospores of three strains of Metarhizium anisopliae were stored in 18 liquids at 4°C, 20°C and 35°C for 18 weeks, 12 weeks or 9 days respectively. Viability was quantified by determination of their germination. In bioassays the virulence of stored blastospores was studied using adults and third instars of Locusta migratoria migratorioides (R. & F.) and compared to those of freshly produced blastospores and conidia. Generally, there was great variability in the viability of blastospores, depending on the fungal strain and the liquids used. Blastospores survived best at 4°C in 10% hydroxyethyl starch; for example, germination of M. anisopliae strain 97 still amounted to more than 80% after storage for 18 weeks. Other suitable liquids were deionized water, 25% Ringer's solution and 1% sodium alginate. The viability of blastospores stored at 20°C was considerably shorter than at 4°C. During storage for 12 weeks at 20°C the best protective liquids for M. anisopliae strain 97 were 25% Ringer's solution (43% germination), deionized water (23%) and 10% hydroxyethyl starch (23%). At 35°C, 45% of M. anisopliae strain 97 blastospores still germinated after storage for 7 days in 25% glycerol. The bioassays revealed that the virulence of blastospores after storage was comparable to that of fresh ones and even better than that of fresh conidia. In general, the LT₅₀ was about 4–6 days at an alternating day/night temperature of 28/20°C.     

Storage of ethyl methanesulphonate-treated barley seeds with low moisture contents

Barley seeds were treated with ethyl methanesulphonate (EMS) for 3 h at 25° C, washed with tap water for 24 h at 25° C, redried at 40° C to different moisture contents below 15% and stored at 25° C in desiccators or in sealed plastic bags. The criteria used for expressing the effect of storage were the M₁ seedling height and the frequency of chromosomal aberrations. With 14·9% seed moisture a strong increase of biological injury occurred in the course of a 2-week storage, while storage of seeds having an initial moisture content of 11·7% led to a significant increase of injury only after 6 weeks. Superdry EMS-treated seeds with 5% or less moisture can be stored at 25° C without any changes in the biological effects. A method is recommended to avoid the EMS-storage effects.     

Effect of gibberellic acid- and water-based pre-soaking treatments under different temperatures and photoperiods on the seed germination of Allium stracheyi Baker: An endangered alpine species of Central Himalaya,India

Allium stracheyi Baker (Alliaceae, 2600–3000 m asl), an endangered species of Central Himalaya, India, has low seed germination in its natural habitat. This study is an attempt to improve seed germination by determining the seed viability with a low mean germination time (MGT) and germination index (GI) under optimum temperature, light, and pre-soaking treatments. The seeds were pre-soaked in hot water (80°C), cold water (10°C), and gibberellic acid (GA₃ at 50 and 100 mg/l) for 24 h and subjected to light (12 h light and 12 h dark) and continuous dark (24 h) conditions with different temperature regimes (10, 15, 20, 25, and 30°C). The viability varied between 66.0% and 69.67% and declined rapidly after 12 months of storage. Our studies suggest that the 100 mg/l GA₃ treatment was beneficial for seed germination and seedling growth. Pre-soaking in a 100 mg/l GA₃ solution and incubation at 20°C under light conditions enhanced the germination significantly (p < 0.05) and resulted in the highest (97.3%) germination with the lowest MGT = 5.7 days, with GI = 8.11. The recommendations of this study support the conservation of alpine A. stracheyi via simple and cost-effective techniques for optimal seed germination.     

The ecology of germination and reproduction of less frequent and vanishing species of the Czechoslovak flora II.Pulsatilla slavica Reuss

The conditions of embryo development, of germination and sprouting in the speciesPulsatilla slavica (north Carpathian endemite) were ascertained. By studies of germination ecology ofPulsatilla slavica it was found that the development of the underdeveloped embryo and germination set in at about 10 °C to about 30 °C. Of the temperatures examined, the temperature of about 25 °C, and alternating ones, the temperature of 10/30 °C appeared to be the optima. At the temperature of 25 °C, the rate of loss of germination capacity was ascertained partly in fully ripe achenes, partly in achenes collected in the stage of wax-ripeness. After 39 weeks of room storage, 62.6% of fully ripe achenes and 32.6% of achenes collected in the stage of waxripeness germinated. The achenes sown after full ripeness in the middle of June sprouted in the middle of July, those sown at the beginning of October sprouted during the first decade of May, and those sown at the beginning of April sprouted during the, third decade of May-Pulsatilla slavica reproduces spontaneously only in the generative way, but it can also be propagated vegetatively by root cuttings.     

Germination responses of four native terrestrial orchids from south-west Western Australia to temperature and light treatments

We report an investigation into the impact of temperature and illumination on in vitro symbiotic and asymbiotic germination of the threatened taxon Caladenia huegelii, and three other orchid spp. namely—Caladenia latifolia, Microtis media and Pterostylis sanguinea, all species from south-west Western Australia, a recognized biodiversity hotspot. High symbiotic germination on oatmeal agar (OMA + fungal symbionts specific to each species) was recorded in three species in continuous dark incubation i.e. C. huegelii seeds (98 % germination at 25 °C), and M. media and P. sanguinea (93 and 98 % respectively at 20 °C). Highest symbiotic germination for C. latifolia (100 %) was observed at 15 and 20 °C under light treatment (12/12 h light/dark). Low temperature incubation (10 °C) significantly suppressed symbiotic germination/development of seedlings across all species. Asymbiotic media treatments assessed (OMA minus fungal symbionts, Pa₅ and ½ MS), failed to stimulate any germination with C. latifolia seeds at 20 °C in either light or dark treatments after an 8 week incubation period. Seeds of M. media sown onto ½ MS medium resulted in higher germination in all developmental stages (3–5) in dark treatment than OMA and Pa₅. Seeds of P. sanguinea sown onto ½ MS medium resulted in higher overall germination in all developmental stages (3–5) in light and dark incubation compared to OMA and Pa₅. OMA supported the highest asymbiotic germination (100 %) in both light and dark incubation with M. media (only to stage 3) but did not support germination and development with other spp. tested. Caladenia huegelii seeds reached developmental Stage 3 (i.e. germinated), but only on Pa₅ medium and only at a relatively low rate in either light (2.6 %) or dark (2.1 %). Germination was higher and development of seedlings faster overall in all test species in symbiotic compared with asymbiotic media treatments. P. sanguinea seeds demonstrated the best response (among species tested) to asymbiotic germination on ½ MS with 40–53 % of germinated seeds spread over developmental stages 3–5 in light or dark incubation (at 20 °C) respectively. Illumination had no effect on fungal symbiont growth across all species, however incubation temperature treatments (10, 15, 20 and 25 °C) affected fungal growth rate. Growth of the fungal symbionts of C. huegelii, M. media and C. latifolia demonstrated significantly lower activity at 10 °C, but the cumulative radial growth rate of the P. sanguinea fungal symbiont reached 64 cm² after only 2 weeks at all temperatures tested, including 10 °C. The study highlights differences in symbiotic and aysmbiotic germination and early protocorm development in vitro between co-occurring herbaceous terrestrial Australian orchid taxa in response to variations in basal media, temperature and light.     

Effect of temperature and cold stratification on seed germination of the Mediterranean wild aromatic Clinopodium sandalioticum (Lamiaceae)

A germination study was carried out on seeds of Clinopodium sandalioticum (Bacch. & Brullo) Bacch. & Brullo ex Peruzzi & Conti (Lamiaceae), a wild aromatic plant endemic to Sardinia. Seeds were incubated at a range of constant (5–25°C) and an alternating temperatures regime (25/10°C), with 12 hours of irradiance per day. The results achieved at 10°C were also compared with those obtained after a period of cold stratification at 5°C for three months. Final seed germination ranged from ca. 28% (5°C) to ca. 72% (25/10°C). A base temperature for germination (T_b) of ca. 5°C and a thermal constant for 50% germination (S) of 89.3°Cd were identified and an optimal temperature for germination (T_o) was estimated to be comprised between 20 and 25°C. Cold stratification negatively affected seed viability and germination at 10°C. Although a typical “Mediterranean germination syndrome”, could not be detected for C. sandalioticum seeds, these results were coherent with those previously reported for other Mediterranean Lamiaceae species.     

Effect of Temperature Regimes on Germination of Dimorphic Seeds of Atriplex prostrata

Dimorphic seeds of Atriplex prostrata were removed from cold dry storage monthly over a one year period to test for fluctuations in seed dormancy and germination rate. For each seed type, four replicates of 25 seeds were exposed to four alternating night/day temperature regimes mimicking seasonal fluctuations in Ohio: 5/15 °C; 5/25 °C; 15/25 °C and 20/35 °C with a corresponding 12-h photoperiod (20 μmol m⁻² s⁻¹; 400 – 700 nm). We found a significant three-way interaction of seed size, temperature and month for both percent germination and the rate of germination. Large seeds showed the greatest germination at the 20/35 °C and 5/25 °C temperature regimes and small seeds at the 5/25 °C regime. Large seeds had greater germination at all temperatures as compared to small seeds. Large seeds had the fastest germination rates at 20/35 °C followed by 5/25 °C whereas small seeds had the fastest rates at 5/25 °C followed by 20/35 °C. This revised version was published online in July 2006 with corrections to the Cover Date.     

Persistence of <Emphasis Type="Italic">Isaria fumosorosea</Emphasis> (Hypocreales: Cordycipitaceae) SFP-198 Conidia in Corn Oil-Based Suspension

Long-term persistence of entomopathogenic fungi as biopesticides is a major requirement for successful industrialization. Corn oil carrier was superior in maintaining germination rates of Isaria fumosorosea SFP-198 conidia during exposure to 50°C for 2 h, when compared with other oils, such as soybean oil, cottonseed oil, paraffin oil, and methyl oleate. The corn oil-based conidial suspension (91.6% germination) was also better in this regard than conidial powder (28.4% germination) after 50°C for 8 h. Long-term storage stabilities of corn oil-based conidial suspension and conidial powder at 4 and 25°C for 24 months were investigated, based on the correlation of germination rate with insecticidal activity against greenhouse whiteflies, Trialeurodes vaporariorum. Viability of conidia in corn oil was more than 98.4% for up to 9 months of storage at 25°C, and followed by 23% at 21 months. However, conidial powder had only 34% viability after 3 months of storage at 25°C, after which its viability rapidly decreased. The two conidial preparations stored at 4°C had better viabilities than those at 25°C, showing the same pattern as above. These results indicate that corn oil-based conidial suspension can be used to improve conidial persistence in long-term storage and be further applied to the formulation of other thermo-susceptible biological control agents.     

Effects of pH,NH<Subscript>4</Subscript>-N concentration,temperature, and storage period on basidiospore germination in an ectomycorrhizal ammonia fungus <Emphasis Type="Italic">Hebeloma vinosophyllum</Emphasis>

Basidiospore germination in an ectomycorrhizal ammonia fungus Hebeloma vinosophyllum was stimulated by 10–500 mM NH₄Cl aqueous solution at pH 4.5–9.0, but not by pure water. The basidiospores germinated at 10°–35°C with an optimum at 25°–30°C. The highest germination percentage (83.0%) was observed in 100 mM NH₄Cl aqueous solution adjusted to pH 8.0 by KOH, when the basidiospores were incubated at a density of 10⁶ spores/ml at 30°C for 14 days. The percent germination value decreased with the increased duration of storage under both dry and wet conditions. Humidity and temperature affected the longevity of H. vinosophyllum basidiospores. The basidiospores maintained their germination ability longer under a dry condition than under a wet condition. The greatest longevity was accomplished by storage at 15°C under a dry condition.     

Effect of storage time and pretreatment on seed germination of the threatened coniferous species Fokienia hodginsii

We report the effects of storage time and pretreatment on seed germination of Fokienia hodginsii. Lower mean germination was observed in seeds stored for 2 years (6.41 ± 1.23 seeds/replicate) compared with those stored for 1 year (8.52 ± 1.06 seeds/replicate). Seeds collected from a southern location had statistically higher mean germination (9.67 ± 1.28 seeds/replicate) than those collected from a northern location (7.99 ± 1.36 seeds/replicate). Higher mean T₅₀ was observed in seeds stored for 2 years (37.02 ± 4.43 days) compared with those stored for 1 year (30.69 ± 5.06 days). Mean germination of untreated fresh seeds was 9.97 ± 1.34 seeds/replicate and that of treated fresh seeds in 60°C water was 12.95 ± 1.24 seeds/replicate. Fresh seeds treated with 50°C and 70°C water had a significantly lower mean germination compared with untreated seeds and seeds treated in 60°C water. Mean T₅₀ was lowest in seeds treated with 60°C water.     

Staining Mitochondria With Hematoxylin After Formalin-Sublimate Fixation; A Rapid Method

Mitochondria were stained in liver, kidney, pancreas, adrenal and intestinal mucosa of rat and mouse. Tissues 1 mm thick, were fixed in a mixture of saturated aqueous HgCl₂, 90 ml; formalin (37-38% HCHO), 10 ml, at room temperature (25°C) for 1 hr. Deparaffinized sections 3-4μ thick were treated with Lugol's iodine (U.S.P.) followed by Na₂S₂O₃ (5%), rinsed in water and the ribonucleic acid removed by any of the following procedures: 0.2 M McIlavaine's buffer, pH 7.0, 2 hr, or 0.2 M phosphate buffer, pH 7.0, 2 hr at 37°C; 0.1% aqueous ribonuclease, 2 hr at 37°C; 5% aqueous trichloracetic acid overnight at 37°C; or 1% KOH at room temperature for 1 hr. After washing in water, sections were treated with a saturated solution of ferric ammonium alum at 37°C for 8-12 hr and colored by Regaud's ripened hematoxylin for 18 hr. They were then differentiated in 1% ferric ammonium alum solution while under microscopic observation.     

Effects of temperature, light, storage conditions, sowing time, and burial depth on the seed germination of Cardiocrinum cordatum var. glehnii (Liliaceae)

In this study, we conducted experiments to accumulate practical information on the propagation and establishment of a population of Cardiocrinum cordatum var. glehnii by seed sowing. C. cordatum var. glehnii seeds require approximately 19 months from seed dispersal to cotyledon emergence in the field. However, the period from seed dispersal to radicle emergence was shortened to approximately 7–8 months by the temperature transition of 25/15°C (60 days) → 15/5°C (30 days) → 0°C (120 days) → 15/5°C (i.e., 15/5°C represents alternating temperature treatment wherein the seeds were placed at 15°C for 12 h during the day and then at 5°C for 12 h during the night). More than 90% of the seeds, which were stored dry at 5°C for 12 months and sown in pots in the field, showed cotyledon emergence, whereas in seeds stored dry at 25°C, dry at room temperature, and non-dry at room temperature, cotyledon emergence was decreased by less than 1%. More than 88% of the seeds that were stored dry at 5°C and sown in the field in October 2002 immediately after collecting, November, and from April to July 2003 showed cotyledon emergence in spring 2004. However, seeds sown in August, September, and October 2003 showed cotyledon emergences of 57.6%, 0%, and 0% in spring 2004, respectively. Seeds collected in October 2002 and sown until July 2003 in the field received adequate high temperature in summer, moderate temperature in autumn, and cold temperature in winter; therefore, the percentage of cotyledon emergence was high in spring 2004. On the other hand, seeds sown in August 2003 or later could not receive enough high temperature; thus, cotyledons emerged from only a few seeds.     

Effect of temperature on germination,growth, and infectivity of the mosquito pathogen Tolypocladium cylindrosporum (deuteromycotina; hyphomycetes)

The mosquito pathogen Tolypocladium cylindrosporum was examined with regard to its response to temperature. Similar temperature ranges were found for growth, germination, and infectivity of blastospores and conidia. Germination occurred at 8° and 33°C but not at 6° and 35°C. Optimal germination and growth was noted between 24° and 27°C for both spore types. Infectivity of blastospores and conidia at different temperatures was examined by exposing L₂Aedes sierrensis larvae to concentrations of 5 × 10⁵ blastospores/ml or 5 × 10⁶ conidia/ml. Larvae were incubated at 12°, 15°, 25°, and 30°C. Infection occurred at all temperatures tested with LT₅₀ values ranging from 22.7 days (12°C) to 5.6 (25°C) days for conidia and 4.7 days (12°C) to 0.6 day (25°C) for blastospores. These results confirmed earlier findings that blastospores infected and killed host larvae more rapidly than conidia and suggested that this difference is largely due to the more rapid germination rate of blastospores. These experiments demonstrated that T. cylindrosporum can be active against mosquito larvae over a broad range of temperatures encompassing both the cold-water habitat of certain temperate mosquito species as well as the habitat of tropical vector species.     

The germination of grass seeds after storage at different temperatures in aluminium foil and manilla paper packets

The germination of seeds of three species of forage grasses, Lolium perenne, Festuca pratensis and Dactylis glomerata, was studied after storage for 3–5 years under five different storage conditions: in aluminium foil packets at —25°C, 0°C and laboratory temperature (c. 18°C), and in manilla paper packets at 0°C and laboratory temperature. With Lolium perenne and Festuca pratensis high germination values at 3 and 7 days were obtained from seed stored at — 25 °C and 0°C in foil packets (5% moisture), but at laboratory temperatures, seed from foil packets gave lower germination values than those from manilla paper packets. At all three temperatures Dactylis glomerata germination after 7 and 14 days was higher in seed stored in foil than in manilla packages. With all three species stored in manilla packets, germination was higher after laboratory than cold storage.     

Propagation of three orchid genera using encapsulated protocorm-like bodies

Summary Protocorm-like bodies (plbs) of the orchid Dendrobium ‘Sonia’ were obtained from fractionated plb explants on Murashiga and Skoog (1962) medium (MS) supplemented with 4.44 μM 6-benzylaminopurine (BA). The developmental stage of plbs to be encapsulated, concentrations of sodium alginate (2–5%) and calcium chloride (25–100 mM), and concentration of MS salts in the matrix were assessed. Fractionated plbs 13–15 d after culture were at the leaf primordia stage and found suitable for encapsulation studies. The best encapsulation response was observed with 3% sodium alginate upon complexation with 75 mM CaCl₂.2H₂O. An encapsulation matrix prepared with MS medium (three-quarter strength) supplemented with 0.44 μM BA and 0.54 μM α-naphthaleneacetic acid gave 100% conversion of encapsulated plbs to plants after storage. A viability of >85% of encapsulated plbs of Dendrobium ‘Sonia’ was observed for 75 d at 4°C. Encapsulated plbs of orchids Dendrobium, Oncidium, and Cattleya were stored at 4°C for 75, 60 and 30 d, respectively, with >88% germination. All plants survived potting in media having either wood charcoal pieces alone or in combination with brick pieces.     

Germination behaviour of seeds of Withania somnifera (L.) Dunal: a high value medicinal plant

In order to evolve a quick method for smooth and optimum germination for Withania somnifera- a medicinally efficacious multipurpose plant, present investigation was carried to study the effect of physico-chemical treatments, storage, temperature, photoperiod and growth regulators (GA₃, IAA, IBA, 2–4 D and BA) on germinability. The most effective treatment is GA₃ at 150 μg/ml concentration at 25 °C. The optimal temperature for germination is 25 °C and continuous light favored germination showing that photoperiod has a significant role. The seedlings derived from seeds performed well when grown in a glasshouse. The data have implications for conservation and cultivation of the species studied.     

Post-harvest age-induced seed dormancy of Acer ginnala and its alleviation by growth regulator and low temperature treatments

One-month-old fruits of Acer ginnala with winged pericarp attached gave 44% germination and this was not increased by cold treatment at 4°C for 0, 10, 20, or 30 days, gibberellic acid treatment at 0, 1, 10, 100 or 1000 mg litre^-1, or ethephon treatment at 0, 2, 20, 200 or 2000 mg litre^-1. After 6 months of storage at 20–25 °C, germination of untreated fruits fell to 5% but could be restored to that of 1-month-old fruits by incubation at 4 °C for 30 days. After 9 months storage, no germination occurred in untreated fruits. Cold treatment (30 days at 4 °C partially restored germination (26%). Treatment with either gibberellic acid (1000 mg litre^-1) and 30 days at 4 °C (40%) or ethephon (100 mg litre^-] and 30 days at 4 °C improved germination (69%). The combination of all three treatments, i.e. 100 mg litre^-1 gibberellic acid, 100 mg litre^-1 ethephon and 30 days at 4 °C, optimised germination (86%). Thus, dormancy of A. ginnala developed during storage but could be reversed by a combination of treatment with low temperature and growth regulators. The highest germination (86%) was achieved after low temperature and growth regulator treatment of stored fruit.     

Thermodynamic characterization of ethidium bromide binding to a unique site on yeast tRNAphe.

A self-consistent thermodynamic characterization of the binding of ethidium to yeast phenylalanine-specific tRNA at 25°C, pH 7.0, in 11 nM MgCl₂, 375 nM NaCl, and 25 mM sodium phosphate has been obtained. Two ethidium molecules bind per tRNA under these conditions. The stronger site has a dissociation constant equal to 1.9 ± 0.5 μM and ΔH_dis°′ = 12 ± 1 Kcal/mol, and the weaker sites has a dissociation constant equal to 24 ± 9 μM and ΔH_dis°′ = 8.9 ± 1.5 Kcal/mol. The average calorimetric ΔH_dis°′ for the to sites 10.6 ± 0.4 kcal/mol. The thermodynamics of binding to the stranger sites are most probably the thermodynamics of interaction between A·U (6) and A·U (7), the unique site identified by Jones and Kearns. The binding is enthalpically driven and classical hydrophobic interactions do not appear to be important in the binding reaction.