Sugar-beet seed steep treatments to improve germination under cold,wet conditions

Modifying the present commercial sugar-beet steep treatment (12h in 0.2% thiram suspension, 25°C) to include an initial 2h steep in 0.3M hydrochloric acid, decreased fruit weight and cortex density and improved the performance of the inherently slower part of the population under cold, wet conditions. Adding gibberellins (GA_4/7) or an N-substituted phthalimide (AC 94377) to the thiram steep was also beneficial whereas kinetin of N⁶-benzyladenine gave no improvement. Germination was even more rapid and better synchronised following a 4-day seed advancement sequence, particularly when this started with the acid steep. Overall, it was possible to increase the proportion of seeds which gave a root or produced hypocotyls at least 2cm tall by 9% and 14% respectively; the thermal time needed for the 90th seed to germinate was reduced from 73 to 30 day degrees and synchrony could be improved at least two-fold.     

Changes in composition of phenolic compounds and antioxidant properties of <Emphasis Type="Italic">Vitis amurensis</Emphasis> seeds germinated under osmotic stress

The research focused on the changes of phenolic compounds as well as their antiradical activity and reducing power isolated from Amur grape (Vitis amurensis) seeds during germination under optimal conditions and under osmotic stress. The seeds were found to contain tannins, (+) catechin, (−) epicatechin, and gallic acid (in free, ester- and glycoside-bound forms). Extracts from the seeds were also shown to contain two other phenolic acids: caffeic and p-coumaric acids, in very low levels. During a 3-day seed germination test under osmotic stress (−0.5 MPa), the content of total phenolics, tannins and phenolic acids declined as compared to the control. However, seed germination under stress conditions led to a significant increase in the amount of catechins. Because catechin is the one of the units in condensed tannins, its dynamic increase during seed germination may be involved in metabolism of tannins under osmotic stress. It is also likely that the synthesis of catechins is greater under stress conditions and these compounds may be engaged in the process of acclimatization of grapevines to stress conditions. The content of total phenolic compounds in seed extracts is positively correlated with their antioxidant properties. The extracts from seeds germinated under optimal conditions exhibited strong antiradical properties against the DPPH (2,2-diphenyl-1-picrylhydrazyl) radical as well as reducing power. As regards the extracts from grape seeds germinated under osmotic stress, this capability was much weaker. The research demonstrated that antioxidants could interfere with the oxidation process induced by various stresses by acting as oxygen scavengers, therefore the tolerance to drought stress might be correlated with an increase in the antioxidant potential.     

Assembly of cortical microtubules during cold treatment of the coenocytic green alga,Chaetomorpha moniligera

The effects of cold treatment on the cortical microtubules (MTs) of Chaetomorpha moniligera Kjellman were investigated by immunofluorescence microscopy. Cortical MTs in Chaetomorpha thallus are arranged longitudinally. In this study, 70–75% of MTs disassembled within 4 h on ice while the others remained stable under these conditions. Reticulate background immunofluorescence, assumed to indicate the presence of a tubulin monomer, was distributed about the stable MTs. Immunofluorescence was prominent in only 50% of the cells. Tubulin polymerization was noted where the background and MT immunofluorescence was strong. New MTs grew transversely as single strings or clusters from the sides of MTs after cold treatment for 4 h and elongated with time to take on a reticulate form at 24 h. The significance of this tubulin polymerization under cold treatment is discussed.Abbreviations MT microtubule - MTOC microtubule-organizing center     

Life cycle and adaptive strategies of three non-annual,ruderalCardueae: Onopordum nervosum,Carthamus arborescens,andCirsium scabrum

The life cycle and autoecology of three Mediterranean thorny species,Onopordum nervosum (Iberian endemic),Cirsium scabrum (Western Mediterranean) andCarthamus arborescens (Iberian-North African), of potential use in agriculture as bioenergetic, forage or oil producing plants were studied. These threeCardueae spp. are good examples of ways in which Mediterranean plants adapt their typical annual cycle to avoid summer droughts.O. nervosum is a perennial, monocarpic, heliophilous species which flowers in early summer and grows even under the extreme xeric conditions of the Mediterranean summers. It is a basiphilous plant usually found in sandy-loamy and sandy-clayey-loamy soils with a marked geographical variability in its seed germination.C. scabrum is a perennial, monocarpic species which has a long life cycle, flowering in summer and growing only during the wet months. It is not well-adapted to droughts and is usually restricted to acid soils.C. arborescens is a perennial, polycarpic species which flowers in spring and dries up in summer. It is usually found in basic loamy-sandy or sandy-loamy soils poor in organic matter.     

Cold shock response inLactococcus lactis ssp.diacetylactis

The acquired freeze-thaw tolerance was investigated forLactococcus lactis ssp.diacetylactis. Pretreatment of microorganisms at less severe temperatures to initiate cold tolerance gaveL. lactis ssp.diacetylactis improved cell viability after successive freezings and thawings. The ability of cells to survive freeze-thaw was dependent on factors experienced prior to freezing. Factors affecting lactic acid bacteria survival during freeze-thaw cycles were found to be different diluents, growth phase, and different cold temperatures. Viability experiments showed that this strain displaying cold shock cryotolerance had an improved survival capacity in stationary phase. The plasmid contents of lactic acid bacteria isolated from different types, DRC-2 and DRC-2C, were examined and compared with the plasmid contents of culture collection strains both before and after cold shock treatment. Using agarose gel electrophoresis, no obvious correlation between the cold shock response and the number of plasmids in the cell could be observed.     

Abscisic acid relationships in the chill-related dormancy mechanism in apple seeds

Abscisic acid (ABA) levels in seeds from three cultivars of apple (Malus domestica Borkh.) which have substantially different chilling requirements were investigated by gas chromatography mass-spectrometry selected ion monitoring (GCMS-SIM) during stratification. The ABA content of dormant unchilled seeds was similar in the three cultivars, suggesting no relationship between the chilling requirement of those seeds and their ABA status. That chilling is not related to ABA changes during stratification was confirmed by warm (20°C) and cold (5°C) stratification experiments. ABA content dropped rapidly and nearly identically under both temperature regimes, but only cold stratification promoted germination. The decline in ABA during stratification was due in large part to leaching from the seed coat and nucellar membrane; the ABA content of the embryo remained nearly constant. The radicle in intact seeds stratified at 5°C began growing 20–30 days after the ABA in the seed coat and nucellar membrane had nearly disappeared. Radicle growth did not occur in unchilled seeds, even though ABA had leached from them as well. It is possible that the leaching of ABA from the seed allows certain promotive forces to develop, but if so, these can develop only at chilling temperatures. Studies were also conducted on 2-trans ABA relationships to apple seed dormancy, but no association was evident.Report No. 12, Department of Fruit and Vegetable Science, Cornell University.     

<Emphasis Type="Italic">In vitro</Emphasis> propagation of brown oak (<Emphasis Type="Italic">Quercus semecarpifolia</Emphasis> Sm.) from seedling explants

An efficient and reproducible method for the regeneration of multiple shoots of brown oak (Quercus semecarpifolia Sm.) has been developed in which a part of the petiolar tube containing a primary shoot is used as the explant. Explants derived from in vitro grown seedlings were cultured either on Murashige and Skoog or Woody Plant medium (WPM) containing different concentrations of benzyladenine (BAP) throughout the range of 1–20 μM. WPM supplemented with 20 μM BAP was found to be best for adventitious shoot induction and for the multiplication of individual shoots. In-vitro-produced shoots were rooted using a two-step method. Firstly, shoots were cultured on WPM containing indolebutyric acid (IBA) at either 50 or 100 μM for 24 or 48 h. Secondly, the shoots were transferred to plant-growth-regulator-free half-strength WPM. The second step not only considerably improved the rooting percentage but also minimized the formation of basal callus. The most effective first-step treatment was found to be 100 μM IBA for 24 h, which initiated rooting at a frequency of 100%. Well-rooted plants were transferred to plastic cups containing nonsterile, sieved soil and farmyard manure, hardened under greenhouse conditions, and then successfully established in pots. This procedure is suitable for use in large-scale production of plants and may have potential application in additional oak species.     

<Emphasis Type="Italic">Ds</Emphasis> insertion mutagenesis as an efficient tool to produce diverse variations for rice breeding

The availability of diversified germplasm resources is the most important for developing improved rice varieties with higher seed yield or tolerance to various biotic or abiotic stresses. Here we report an efficient tool to create increased variations in rice by maize Ac/Ds transposon (a gene trap system) insertion mutagenesis. We have generated around 20,000 Ds insertion rice lines of which majority are homozygous for Ds element. We subjected these lines to phenotypic and abiotic stress screens and evaluated these lines with respect to their seed yields and other agronomic traits as well as their tolerance to drought, salinity and cold. Based on this evaluation, we observed that random Ds insertions into rice genome have led to diverse variations including a range of morphological and conditional phenotypes. Such differences in phenotype among these lines were accompanied by differential gene expression revealed by GUS histochemical staining of gene trapped lines. Among the various phenotypes identified, some Ds lines showed significantly higher grain yield compared to wild-type plants under normal growth conditions indicating that rice could be improved in grain yield by disrupting certain endogenous genes. In addition, several 1,000s of Ds lines were subjected to abiotic stresses to identify conditional mutants. Subsequent to these screens, over 800 lines responsive to drought, salinity or cold stress were obtained, suggesting that rice has the genetic potential to survive under abiotic stresses when appropriate endogenous genes were suppressed. The mutant lines that have higher seed yielding potential or display higher tolerance to abiotic stresses may be used for rice breeding by conventional backcrossing combining with molecular marker-assisted selection. In addition, by exploiting the behavior of Ds to leave footprints upon remobilization, we have shown an alternative strategy to develop new rice varieties without foreign DNA sequences in their genome. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Three zinc‐finger RNA‐binding proteins in cabbage (Brassica rapa) play diverse roles in seed germination and plant growth under normal and abiotic stress conditions

Despite the increasing understanding of the stress‐responsive roles of zinc‐finger RNA‐binding proteins (RZs) in several plant species, such as Arabidopsis thaliana, wheat (Triticum aestivum) and rice (Oryza sativa), the functions of RZs in cabbage (Brassica rapa) have not yet been elucidated. In this study, the functional roles of the three RZ family members present in the cabbage genome, designated as BrRZ1, BrRZ2 and BrRZ3, were investigated in transgenic Arabidopsis under normal and environmental stress conditions. Subcellular localization analysis revealed that all BrRZ proteins were exclusively localized in the nucleus. The expression levels of each BrRZ were markedly increased by cold, drought or salt stress and by abscisic acid (ABA) treatment. Expression of BrRZ3 in Arabidopsis retarded seed germination and stem growth and reduced seed yield of Arabidopsis plants under normal growth conditions. Germination of BrRZ2‐ or BrRZ3‐expressing Arabidopsis seeds was delayed compared with that of wild‐type seeds under dehydration or salt stress conditions and cold stress conditions, respectively. Seedling growth of BrRZ3‐expressing transgenic Arabidopsis plants was significantly inhibited under salt, dehydration or cold stress conditions. Notably, seedling growth of all three BrRZ‐expressing transgenic Arabidopsis plants was inhibited upon ABA treatment. Importantly, all BrRZs possessed RNA chaperone activity. Taken together, these results indicate that the three cabbage BrRZs harboring RNA chaperone activity play diverse roles in seed germination and seedling growth of plants under abiotic stress conditions as well as in the presence of ABA.     

Cold and water stresses produce changes in endogenous jasmonates in two populations of <Emphasis Type="Italic">Pinus pinaster</Emphasis> Ait

There is considerable evidence suggesting that jasmonates (JAs) play a role in plant resistance against abiotic stress. It is well known that in Angiosperms JAs are involved in the defense response, however there is little information about their role in Gymnosperms. Our proposal was to study the involvement of JAs in Pinus pinaster Ait. reaction to cold and water stress, and to compare the response of two populations of different provenances (Gredos and Bajo Tiétar) to these stresses. We detected 12-oxo-phytodienoic acid (OPDA), jasmonic acid (JA), and the hydroxylates 11-hydroxyjasmonate and 12-hydroxyjasmonate in foliage and shoots of P. pinaster plants. The response of the Gredos population to cold stress differed from that of Bajo Tiétar. Gredos plants showed a lower JA-basal level than Bajo Tiétar; under cold stress JA increased twofold at 72 h, while it decreased in Bajo Tiétar plants. The hydroxylates slightly increased in both populations due to cold stress treatment. Under water stress, plants from Gredos showed a remarkable JA-increase; thus the JA-response was much more prominent under water stress than under cold stress. In contrast, no change was found in JA-level in Bajo Tiétar plants under water stress. The level of JA-precursor, OPDA, was very low in control plants from Gredos and Bajo Tiétar. Under water stress OPDA increased only in plants from Bajo Tiétar. Therefore, we inform here of a different JAs-accumulation pattern after the stress treatment in P. pinaster from two provenances, and suggest a possible correlation with adaptations to diverse ecological conditions.     

Field assessment of <Emphasis Type="Italic">Fusarium oxysporum</Emphasis> based mycoherbicide for control of <Emphasis Type="Italic">Striga hermonthica</Emphasis> in Nigeria

Fusarium oxysporum (isolate PSM 197) based mycoherbicide was evaluated for its efficacy under field conditions in trials conducted during 1999--2001 cropping seasons in the Nigerian savanna. In the 1999 cropping season, spot application of 5--10 g of mycoherbicide was found to give effective control of Striga hermonthica. Results of on-farm trials at Barhim and Dutsen-Ma areas showed the application of the mycoherbicide to significantly (p= 0.05) increase stand count at both 3 weeks and at harvest, reduced Striga shoot count and increased crop yield in both improved and local sorghum varieties, as compared with the same varieties not treated with the mycoherbicide. Results establish the efficacy of F. oxysporum as a mycoherbicide and the need for further development of the mycoherbicide into formulated granules or seed treatment for use in control of S. hermonthica under field conditions.     

水引发对油松种子和幼苗的生理效应及其作用机理

为了解决油松种子在直播造林中发芽率低、幼苗抗旱性差的问题,研究了油松种子水引发的方法及引发对其种子和幼苗的生理效应,并对水引发后幼苗的抗旱机理进行了探索。结果表明:油松种子水引发的最佳条件为10℃,闭光条件下分批给种子加水,加水总量为种子初始质量的30%,引发10 d。水引发可显著(P0.05)降低种子的半数发芽天数和硬实率,显著提高种子的发芽指数、最终发芽率和活力指数;水引发种子的可溶性糖及可溶性蛋白含量相比对照也得到提高。水引发可增加油松幼苗的茎粗、鲜重及根长,降低其相对苗高;并极显著(P0.01)提高了幼苗过氧化氢酶、过氧化物酶和根内脱氢酶活性,降低了游离氨基酸含量。干旱胁迫下,引发种子的幼苗脯氨酸和可溶性糖含量分别高于对照39.6%、118.9%,丙二醛含量低于对照93.4%,均达极显著水平。综合分析认为,水引发提高了种子可溶性糖及可溶性蛋白含量为种子萌发提供了保障,从而提高了种子发芽率,并通过增强幼苗抗氧化酶活性、降低丙二醛含量,进而增强幼苗抗氧化系统的功能,最终明显提高其抗旱性。     

Elicitation of volatile compounds in photomixotrophic cell culture of Petroselinum crispum

Photomixotrophic cells of Petroselinum crispum accumulated >500 mg chlorophyll per kg wet weight and grew well in a broad range of phytoeffector conditions. Autoclaved fungal cells were lethal for photoheterotrophic cells, but induced in photomixotrophic cells the formation of volatile n-alkanes, phthalides, coumarins, and elemicine. Most of the compounds elicited reached a concentration maximum between 20 and 30 h after addition of the mycelium, whereas the group of n-alkanes increased steadily during the 90 h monitored. Maximum concentrations were: 12 mg of graveolone, 1 mg of bergapten, 0.5 mg of sedanenolide, and 0.5 mg of n-tetradecane per 1 nutrient medium. A dose/effect relationship was found; 10 to 25 g of fungal wet weight per 1 culture medium resulted in maximum accumulation of volatiles. The formation of volatiles by photomixotrophic in vitro cells is discussed as an integral part of plant responses to ecological stress.     

Increased Germination Rates of Baldcypress and Pondcypress Seed Following Treatments Affecting the Seed Coat

The effects of seed-coat removal, warm-water soaking, cold stratification, dry-cold storage and acid treatment on the germination of baldcypress (Taxodium distichum (L.) Rich.) and pondcypress (Taxodium distichum var. nutans (Ait.) Sweet) seed were studied. Removal of the hard seed coat resulted in a prompt and high percent germination, indicating that there was no physiological embryo dormancy. Cold stratification, warm-water soaking and acid treatment did not increase percent germination over the control after a 60-day germination period. They did promote the rate of germination however, having a significantly higher germination at 30 days. Dry-cold storage promoted the initial rate of germination, but resulted in a decreased percent germination below the control after 60 days. The above results suggest that moisture, and not cold, was the crucial component of stratification. The improved germination rate following acid treatment further suggests that any treatment which rendered the seed coat more permeable or softened it was more effective in increasing the rate of germination of baldcypress and pondcypress seed.     

Drop off rhythm and survival periods of <Emphasis Type="Italic">Amblyomma lepidum</Emphasis> (Acari: Ixodidae) under field conditions

In this study, engorged Amblyomma lepidum ticks were found to drop off in two peaks, one in the morning and one in the evening. Most larvae and females engorged during the morning hours between 06.00 h and 10.00 h with a peak around 08.00 h, whereas the majority of the nymphs dropped in the evening between 18.00 h and 24.00 h with the peak around 22.00 h. Although the effect of time on drop-off patterns of the ticks was statistically significant (p≤ 0.001), there were no significant seasonal influences. Survival of unfed stages of A. lepidum was also studied and was found to increase from larvae to adult ticks. The longest survival periods of 10, 11 and 14 weeks were recorded during the wet season for larvae, nymphs and adults, respectively. It is concluded that environmental conditions required for survival of A. lepidum are optimal only during the wet season and that during other seasons the tick depends primarily on prevailing micro-climatic conditions for its survival.     

Effect of wet and dry cycles in calcareous soil on mineral nutrient uptake of two grasses,Agrostis stolonifera L. and Festuca ovina L.

Shoot uptake of mineral nutrients (Ca, Cu, Fe, K, mg, Mn, P, S, Zn) by Agrostis stolonifera L. was compared with Festuca ovina L. under wet and dry cycles. Such conditions are typical for A. stolonifera sites, whereas F. ovina is growing mostly on consistently drier and better-drained soils. Plants were grown in a glasshouse, at controlled temperature and light conditions, using two moisture regimes, one constant at 60% WHC (water holding capacity), one wet/dry fluctuating between 35 and 100% WHC. Above ground and total biomass production was lower under wet/dry treatment than at constant water regime in F. ovina, but did not differ between regimes in A. stolonifera. Shoot uptake of most elements was severely reduced in F. ovina at the wet/dry regime. Shoot uptake and concentrations of most elements studied (Cu, K, Mn, P, S, Zn) were lower (p<0.05) under wet/dry treatment than at constant regime in A. stolonifera and tended to be lower also of Fe and Mg. Differences in biomass production observed are consistent with field evidence that A. stolonifera grows in sites which are periodically flooded but may become quite dry during other periods, and that F. ovina is limited to sites which are consistently drier and better drained. Evidence from the present study, however, does not support any view that alternating wet and dry cycles, as typical of A. stolonifera field sites, would be beneficial to nutrient acquisition of this species but that biomass production may develop normally at the lower uptake of most mineral nutrients measured under the wet/dry regime. Such regimes are decidedly unfavourable to both growth and nutrient acquisition of F. ovina.     

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.     

Wind tunnel assays of olfactory responses of femaleRhagoletis pomonella flies to apple volatiles: effect of wind speed and odour release rate

Little is known about how adults of the corn leafhopper,Dalbulus maidis (DeLong & Wolcott), and its congeners survive subfreezing temperatures at high elevations during the dry winter in Mexico. In the laboratory, duration of survival at −5°C was measured for four MexicanDalbulus species:D. maidis, D. elimatus (Ball),D. gelbus DeLong andD. quinquenotatus DeLong & Nault; and a closely related North American species,Baldulus tripsaci Kramer & Whitcomb. Adult leafhoppers reared under environmental conditions that simulated the beginning of the dry winter season during October in Mexico (‘October-reared’) were at least twice as tolerant of −5°C than adults reared under environmental conditions that simulated the beginning of the wet summer season during June (‘June-reared’).Dalbulus species found primarily at high elevations, such asD. elimatus, were seven times more tolerant of −5°C thanD. quinquenotatus, a species which overwinters at low to mid elevations on itsTripsacum hosts. October-rearedD. maidis adults survived relatively short periods at −5°C (LT₅₀=8.9h) compared to October-rearedD. elimatus adults (LT₅₀=42.3h). This suggests that in Mexico,D. maidis either overwinters in protected habitats at higher elevations or it migrates to lower, frost-free regions. October-rearedB. tripsaci adults, which overwinter in the egg stage, were intolerant of −5°C (LT₅₀=2.6h). A conditioning period for 1 h at +5°C before and after exposure to −5°C significantly improved survival forD. maidis. Supercooling points (SCPs) were between −23 and −20°C, indicating that mortality of these leafhoppers at −5°C was due to cold shock injury rather than internal ice formation.