Relative sensitivity of photosynthesis and respiration to freeze-thaw stress in herbaceous species : importance of realistic freeze-thaw protocols

The relative effect of a freeze-thaw cycle on photosynthesis, respiration, and ion leakage of potato leaf tissue was examined in two potato species, Solanum acaule Bitt. and Solanum commersonii Dun. Photosynthesis was found to be much more sensitive to freezing stress than was respiration, and demonstrated more than a 60% inhibition before any impairment of respiratory function was observed. Photosynthesis showed a slight to moderate inhibition when only 5 to 10% of the total electrolytes had leaked from the tissue (reversible injury). This was in contrast to respiration which showed no impairment until temperatures at which about 50% ion leakage (irreversible injury) had occurred. The influence of freeze-thaw protocol was further examined in S. acaule and S. commersonii, in order to explore discrepancies in the literature as to the relative sensitivities of photosynthesis and respiration. As bath cooling rates increased from 1°C/hour to about 3 or 6°C/hour, there was a dramatic increase in the level of damage to all measured cellular functions. The initiation of ice formation in deeply supercooled tissue caused even greater damage. As the cooling rates used in stress treatments increased, the differential sensitivity between photosynthesis and respiration nearly disappeared. Examination of agriculturally relevant, climatological data from an 11 year period confirmed that air cooling rates in the freezing range do not exceed 2°C/hour. It was demonstrated, in the studies presented here, that simply increasing the actual cooling rate from 1.0 to 2.9°C/hour, in frozen tissue from paired leaflet halves, meant the difference between cell survival and cell death.     

Effect of gestation on GnRH-induced LH and FSH release in buffalo (Bubalus bubalis)

This study examined the effect of gestation on the hypophyseal responsiveness of buffalo to GnRH-induced LH and FSH release. Peripheral plasma LH and FSH concentrations were measured at 1 h before and upto 6 h after administration of GnRH (1 ug/kg body weight) or saline at Days 60, 150 and 240 of gestation in 2 groups of buffalo (n = 4 each). Basal LH concentrations did not vary at the 3 stages of gestation, while basal FSH concentrations exhibited a significant reduction (P < 0.05) from Day 60 to Day 150 of gestation. There was a significant reduction in the total LH (P < 0.05) and FSH (P < 0.01) released in response to GnRH from Day 60 to Day 240 of gestation. The duration of LH and FSH peaks and the time to attain peak concentration was not affected by the stage of gestation. The results of the present study point to a progressive decline in LH and FSH release responses to GnRH during the advancement of gestation in the buffalo.     

Use of lysophosphatidylethanolamine, a natural lipid, to retard tomato leaf and fruit senescence

We studied the influence of lysophosphatidylethanolamine (LPE) on the pattern and rate of ethylene production and respiration of tomato ( Lycopersicon esculentum cv. H7155) leaflets and fruit. Leaflets that had been senescing on the plant showed a climacteric-like rise in ethylene production but not in respiration rate which decreased continuously with leaf age. Detached leaflets had a climacteric-like pattern in respiration whether they were incubated in complete darkness or in light. Detached leaflets incubated in the dark had higher rates of ethylene production and CO₂ evolution than did light-incubated leaves. There was no change in the pattern of ethylene production or CO₂ evolution as a result of LPE treatment. However, LPE-treated attached and detached leaflets had consistently lower rates of CO₂ evolution. The reduction in CO₂ evolution by LPE was most pronounced at the climacteric-like peak of the detached leaves. LPE-treated leaflets had a higher chlorophyll content and fresh weight and lower electrolyte leakage than the control. LPE-treated fruits had lower rates of ethylene and CO₂ production than did the control. LPE-treated fruits also had higher pericarp firmness and lower electrolyte leakage than the control. The results of the present study provide evidence that LPE is able to retard senescence of attached leaves and detached leaves and tomato fruits. Several recent studies suggest that lysolipids can act in a specific manner as metabolic regulators. Our results suggest a specific role of lysolipid LPE in aging and senescence     

Effect of turgor pressure on water permeability ofAllium cepa epidermis cell membranes

Summary Using onion epidermis layer a very accurate method for measuring the permeability of epidermis cells to water was standardized. In this method a 1.4 cm diameter epidermis disc was soaked in tritiated water (500 Ci/ml) for about 1 hr. Next the disc was mounted in a specially designed elution chamber where it was held flat and washed on the noncuticular side with ordinary water. A constant flow rate, high enough to minimize unstirred layer effect, was used. Permeability was calculated in the usual way after separating different exponentials from the efflux curve of tritiated water. Turgor pressure of the cell was regulated by soaking the disc in mannitol solutions containing tritiated water and washing it in the chamber with same concentration mannitol solution containing no radioactivity. Water permeability values were found to decrease less than 8% when the turgor pressure was decreased from 8 atm (full turgor) to zero. Turgor pressure had no significant effect on the water permeability of onion epidermal cells. Our results are contradictory to the findings of Zimmerman and Steudle (1974,J. Membrane Biol. 16:331) but are similar to the findings of Tazawa and Kamiya (1966,Aust. J. Biol. Sci. 19:339) and Kiyosawa and Tazawa (1972,Protoplasma 74:257).     

Comparative mapping of loci controlling winter survival and related traits in oilseed Brassica rapa and B. napus

Winter survival is an important characteristic of oilseedBrassica that is seeded in the fall in northern climates,and it may be affected by genetic variation for other cold-regulated traits,such as freezing tolerance and vernalization responsive flowering time. Weanalyzed immortalized populations of oilseed Brassica rapa(recombinant inbred lines) and B. napus (double haploidlines) derived from crosses of annual and biennial types in order to comparethe map positions and effects of quantitative trait loci controlling wintersurvival, nonacclimated and acclimated freezing tolerances, and flowering time.The B. napus population was evaluated in multiple winters,and six of the 16 total significant QTL for winter survival were detected inmore than one winter. Correspondence in the map positions of QTL controllingdifferent traits within species provided evidence that some alleles causinggreater acclimated freezing tolerance and later flowering time also contributedto increased winter survival. Correspondence in the map positions of QTLbetween species provided evidence for allelic variation at homologous loci inB. rapa and B. napus. The potentialrole of some candidate genes in regulating these traits is discussed.     

Plasma Membrane ATPase Activity following Reversible and Irreversible Freezing Injury

Plasma membrane ATPase has been proposed as a site of functional alteration during early stages of freezing injury. To test this, plasma membrane was purified from Solanum leaflets by a single step partitioning of microsomes in a dextran-polyethylene glycol two phase system. Addition of lysolecithin in the ATPase assay produced up to 10-fold increase in ATPase activity. ATPase activity was specific for ATP with a K_m around 0.4 millimolar. Presence of the ATPase enzyme was identified by immunoblotting with oat ATPase antibodies. Using the phase partitioning method, plasma membrane was isolated from Solanum commersonii leaflets which had four different degrees of freezing damage, namely, slight (reversible), partial (partially reversible), substantial and total (irreversible). With slight (reversible) damage the plasma membrane ATPase specific activity increased 1.5- to 2-fold and its K_m was decreased by about 3-fold, whereas the specific activity of cytochrome c reductase and cytochrome c oxidase in the microsomes were not different from the control. However, with substantial (lethal, irreversible) damage, there was a loss of membrane protein, decrease in plasma membrane ATPase specific activity and decrease in K_m, while cytochrome c oxidase and cytochrome c reductase were unaffected. These results support the hypothesis that plasma membrane ATPase is altered by slight freeze-thaw stress.     

Global change‐driven effects on dissolved organic matter composition: Implications for food webs of northern lakes

Northern ecosystems are experiencing some of the most dramatic impacts of global change on Earth. Rising temperatures, hydrological intensification, changes in atmospheric acid deposition and associated acidification recovery, and changes in vegetative cover are resulting in fundamental changes in terrestrial–aquatic biogeochemical linkages. The effects of global change are readily observed in alterations in the supply of dissolved organic matter (DOM)—the messenger between terrestrial and lake ecosystems—with potentially profound effects on the structure and function of lakes. Northern terrestrial ecosystems contain substantial stores of organic matter and filter or funnel DOM, affecting the timing and magnitude of DOM delivery to surface waters. This terrestrial DOM is processed in streams, rivers, and lakes, ultimately shifting its composition, stoichiometry, and bioavailability. Here, we explore the potential consequences of these global change‐driven effects for lake food webs at northern latitudes. Notably, we provide evidence that increased allochthonous DOM supply to lakes is overwhelming increased autochthonous DOM supply that potentially results from earlier ice‐out and a longer growing season. Furthermore, we assess the potential implications of this shift for the nutritional quality of autotrophs in terms of their stoichiometry, fatty acid composition, toxin production, and methylmercury concentration, and therefore, contaminant transfer through the food web. We conclude that global change in northern regions leads not only to reduced primary productivity but also to nutritionally poorer lake food webs, with discernible consequences for the trophic web to fish and humans.     

Response of wheat growth,grain yield and water use to elevated CO2 under a Free‐Air CO2 Enrichment (FACE) experiment and modelling in a semi‐arid environment

The response of wheat crops to elevated CO₂ (eCO₂) was measured and modelled with the Australian Grains Free‐Air CO₂ Enrichment experiment, located at Horsham, Australia. Treatments included CO₂ by water, N and temperature. The location represents a semi‐arid environment with a seasonal VPD of around 0.5 kPa. Over 3 years, the observed mean biomass at anthesis and grain yield ranged from 4200 to 10 200 kg ha^?1 and 1600 to 3900 kg ha^?1, respectively, over various sowing times and irrigation regimes. The mean observed response to daytime eCO₂ (from 365 to 550 μmol mol^?1 CO₂) was relatively consistent for biomass at stem elongation and at anthesis and LAI at anthesis and grain yield with 21%, 23%, 21% and 26%, respectively. Seasonal water use was decreased from 320 to 301 mm (P = 0.10) by eCO₂, increasing water use efficiency for biomass and yield, 36% and 31%, respectively. The performance of six models (APSIM‐Wheat, APSIM‐Nwheat, CAT‐Wheat, CROPSYST, OLEARY‐CONNOR and SALUS) in simulating crop responses to eCO₂ was similar and within or close to the experimental error for accumulated biomass, yield and water use response, despite some variations in early growth and LAI. The primary mechanism of biomass accumulation via radiation use efficiency (RUE) or transpiration efficiency (TE) was not critical to define the overall response to eCO₂. However, under irrigation, the effect of late sowing on response to eCO₂ to biomass accumulation at DC65 was substantial in the observed data (~40%), but the simulated response was smaller, ranging from 17% to 28%. Simulated response from all six models under no water or nitrogen stress showed similar response to eCO₂ under irrigation, but the differences compared to the dryland treatment were small. Further experimental work on the interactive effects of eCO₂, water and temperature is required to resolve these model discrepancies.     

Isolation and characterization of embryonic stem cell-like cells from in vitro-produced buffalo (Bubalus bubalis) embryos

This study was carried out to isolate and characterize buffalo embryonic stem (ES) cell-like cells from in vitro-produced embryos. Inner cell mass (ICM) cells were isolated either mechanically or by enzymatic digestion from 120 blastocysts whereas 28 morulae were used for the isolation of blastomeres mechanically. The ICM cells/ blastomeres were cultured on mitomycin-C-treated feeder layer. Primary cell colony formation was higher (P < 0.05) for hatched blastocysts (73.1%, 30/41) than that for early/expanded blastocysts (25.3%, 20/79). However, no primary cell colonies were formed when blastomeres obtained from morulae were cultured. Primary colonies were formed in 14.1% (12/85) of intact blastocyst culture, which was significantly lower (P < 0.05) than that of 41.6% for ICM culture. These colonies were separated by enzymatic or mechanical disaggregation. Using mechanical disaggregation method, the cells remained undifferentiated and two buffalo ES cell-like cell lines (bES1, bES2) continued to grow in culture up to eight passages. However, disassociation through enzymatic method resulted in differentiation. Undifferentiated cells exhibited stem cell morphological features, normal chromosomal morphology, and expressed specific markers such as alkaline phosphatase (AP) and Oct-4. Cells formed embryoid bodies (EBs) in suspension culture; extended culture of EBs resulted in formation of cystic EBs. Following prolonged in vitro culture, these cells differentiated into several types of cells including neuron-like and epithelium-like cells. Furthermore, the vitrified-thawed ES cell-like cells also exhibited typical stem cell characteristics. In conclusion, buffalo ES cell-like cells could be isolated from in vitro-produced blastocysts and maintained in vitro for prolonged periods of time.