Search

Among wild relatives of cultivated almond and plum, a desert almond species, but not a montane plum species, showed higher drought resistance than the common rootstocks routinely used in orchards. Water shortage is a severe environmental factor causing growth disruption and yield-\loss in many agricultural plant species. As fruit trees are likely to suffer from the effects of severe drought in the future, wild relatives of cultivated crops can provide plant breeders a unique material to improve the drought resistance of modern crop varieties. We conducted a drought and rewatering greenhouse experiment along 51 days with young trees of almond (the desert wild species Prunus ramonensis vs. the commonly used rootstock hybrid Prunus dulcis × Prunus persica) and plum (the montane wild species Prunus ursina vs. the rootstock Prunus cerasifera × Prunus persica). To decipher the drought resistance mechanisms in these trees we monitored physiological responses. Expression dynamics of cellular water channels from the plasma intrinsic protein (PIP) aquaporin family were measured in the almond species. Our results indicate a higher drought resistance in wild almond compared to the rootstock, but not in the wild plum species. Under drought, P. ramonensis had ~ ninefold higher photosynthesis activity, ~ 50-fold higher water-use efficiency and lower vulnerability to embolism than the rootstock. In the almond species, PIP downregulation was linked with maintenance of hydraulic conductivity, and vice versa for upregulation. This study implies that there is a link between drought resistance in wild tree species and their native habitat conditions, with an advantage for the desert, but not the montane, species. Finally, our study highlights the need to protect and conserve wild relatives of fruit tree species, partly as potential plant materials to be used by breeders to improve the resilience of orchard tree species to drought.

The lychee fruit is in high demand worldwide. However, the yields of many cultivars are low, including the high-quality cultivars “Nuomici” (NMC) and ”Fei Zi Xiao” (FZX), which are very tasty and produce large fruit with a small seed, but tend to shed their fruitlets. In a previous work, we found that cross-hand pollination of “Mauritius” (MA) with pollen of another cultivar increased fruit set and reduced fruit-drop in comparison to self-hand pollination. In the current research, we aimed to identify the optimal pollen donor for three of the main cultivars grown in Israel: MA, FZX, and “Tamuz” (TA). We compared the effect of different pollinizers and found that the Vietnamese cultivar “Hong Long” (HL), which is becoming an important cultivar in Israel, was the optimal pollinizer for the three cultivars. In addition, we found that FZX and TA were not self-fertile under the Israeli environmental conditions since they tend to shed fruitlets that originated from self-fertilization. In contrast, MA is able to fertilize itself, although cross-pollination greatly increased its fruit number and size. We also identified a new PCR marker for lychee, M3, that enabled us to distinguish between self- and cross-fertilized FZX fruits pollinated by HL. Our results indicate that cross-pollination, particularly by HL, has beneficial effects on the production of lychee and it is especially important for cultivars that generate small seeds and tend to shed their fruitlets.

The lychee fruit is in high demand worldwide. However, the yields of many cultivars are low, including the high-quality cultivars "Nuomici" (NMC) and "Fei Zi Xiao" (FZX), which are very tasty and produce large fruit with a small seed, but tend to shed their fruitlets. In a previous work, we found that cross-hand pollination of "Mauritius" (MA) with pollen of another cultivar increased fruit set and reduced fruit-drop in comparison to self-hand pollination. In the current research, we aimed to identify the optimal pollen donor for three of the main cultivars grown in Israel: MA, FZX, and "Tamuz" (TA). We compared the effect of different pollinizers and found that the Vietnamese cultivar "Hong Long" (HL), which is becoming an important cultivar in Israel, was the optimal pollinizer for the three cultivars. In addition, we found that FZX and TA were not self-fertile under the Israeli environmental conditions since they tend to shed fruitlets that originated from self-fertilization. In contrast, MA is able to fertilize itself, although cross-pollination greatly increased its fruit number and size. We also identified a new PCR marker for lychee, M3, that enabled us to distinguish between self- and cross-fertilized FZX fruits pollinated by HL. Our results indicate that cross-pollination, particularly by HL, has beneficial effects on the production of lychee and it is especially important for cultivars that generate small seeds and tend to shed their fruitlets.

Erwinia amylovora causes fire blight, a bacterial disease of pome fruits. For 15 years, fire blight management in Israel was based solely on the antibiotic oxolinic acid but, in response to decreased disease-control efficacy, copper agents were added in 2014. However, concern was raised as to whether repeated applications of copper might have impaired pollen germination and thereby impaired yields. In Israel, apple cultivars, except for the early-blooming cultivar ‘Cripps pink’ (trademark Pink Lady), appeared to be markedly less sensitive to the infection than pear cultivars (‘Spadona’ and ‘Costia’). The aims of the present study were to monitor the effects of treatments with copper-based agents during blossoming in reducing fire blight damage, while recording their effects on pollen-tube germination and fruit-yield parameters. Copper tribasic sulfate and copper hydroxide reduced disease levels by more than 70% compared with uninfected controls. Pollen-tube germination rates in both apples and pears were significantly reduced after two successive daily applications of copper tribasic sulfate, but usually they recovered to control levels after introduction of a single-day interval. Four to seven seasonal applications of copper agents on apple and pear blossoms did not significantly reduce fruit weight and size, seed number, seed-set or fruit-set. The efficacy of oxolinic acid, applied either alone or tank-mixed with copper, resembled that of copper alone. In summary, copper application against fire blight in pears and apples during their blooming in Israel is both effective and harmless to yields.

A hyperpolarised-NMR acquisition approach that is sensitive to the process of glucose-6-phosphate anomerization is presented. Using selective depolarisation of one of the anomer's signals, it is possible to observe the replenishing of this signal due to the fast anomeric exchange of this compound. The forward to reverse reaction rate constants ratio was ca. 1.6.

Trees of the genus Prunus produce some of the most widely consumed fruits globally. The combination of climate change-related warming and increased drought stress, scarcity of freshwater resources for irrigation, and increasing demands due to population growth creates a need for increased drought tolerance in these tree species. Recently, we have shown in the field that a native wild pear species performs better under drought than two cultivated pear species. Here, a comparative field study was conducted in Israel to investigate traits associated with drought tolerance in almond (cultivated Prunus dulcis (Mill.) D. A. Webb vs wild Prunus ramonensis Danin) and plum (cultivated Prunus domestica L. vs wild Prunus ursina Kotschy). Measurements of xylem embolism and shoot and root carbon reserves were done along a year, including seasonal drought in the wild and a 35-day drought experiment in the orchards. Synchronous measurements of native xylem embolism and shoot water potential showed that cultivated and wild almond trees lost ~50% of hydraulic conductivity at −2.3 and −3.2 MPa, respectively. Micro-CT images confirmed the higher embolism ratio in cultivated versus wild almond, whereas the two plum species were similar. Dynamics of tissue concentrations of nonstructural carbohydrates were mostly similar across species, with higher levels in cultivated versus wild plum. Our results indicate an advantage for the wild almond over its cultivated relative in terms of xylem resistance to embolism, a major risk factor for trees under drought stress. This result is in line with our previous experiment on pear species. However, the opposite trends observed among the studied plum species mean that these trends cannot be generalized. It is possible that the potential for superior drought tolerance in wild tree species, relative to their cultivated relatives, is limited to wild species from dry and hot habitats.

A non-radioactive 2-deoxyglucose (2DG) analog has been developed here for hyperpolarized magnetic resonance investigations. The analog, [13C6,D8]2DG, showed 13% polarization in solution (27,000-fold signal enhancement at the C1 site), following a dissolution-DNP hyperpolarization process. The phosphorylation of this analog by yeast hexokinase (yHK) was monitored in real-time with a temporal resolution of 1 s. We show that yHK selectively utilizes the β anomer of the 2DG analog, thus revealing a surprising anomeric specificity of this reaction. Such anomeric selectivity was not observed for the reaction of yHK or bacterial glucokinase with a hyperpolarized glucose analog. yHK is highly similar to the human HK-2, which is overexpressed in malignancy. Thus, the current finding may shed a new light on a fundamental enzyme activity which is utilized in the most widespread molecular imaging technology for cancer detection – positron-emission tomography with 18F-2DG.

Honeybees (HB) are strongly attracted to lychee flowers and serve as the main pollen vector in most lychee orchards throughout the world. Despite precise protocols for managing HB in Israeli lychee orchards, we suspected that the full pollination potential of the orchards was not being achieved and that yields could be increased. Since bumblebees (BB) forage in harsh weather and at lower temperatures than HB, in previous studies we questioned if adding BB to HB in apple and pear orchards would increase fruit set and yield. Encouraged by the important results, we examined this approach in lychee in the present work. The experiments were conducted for three years, resulting in a significant increase in lychee fruit set and yield. Our observations revealed that BB contribute to lychee pollination not only because they are active in conditions that HB are not, but also because HB collect mainly nectar from the lychee flowers and barely contact the reproductive organs, while BB collect mainly pollen and hence pollination increases. As a result of this work, since 2018 10 BB hives per hectare have been introduced into many lychee orchards in Israel in addition to the standard 2.5 HB hives per hectare.

Reports on gadolinium deposits in the body and brains of adults and children who underwent contrast-enhanced MRI examinations warrant development of new, metal free, contrast agents for MRI. Nitrate is an abundant ion in mammalian biochemistry and sodium nitrate can be safely injected intravenously. We show that hyperpolarized [15N]nitrate can potentially be used as an MR tracer. The 15N site of hyperpolarized [15N]nitrate showed a T1 of more than 100 s in aqueous solutions, which was prolonged to more than 170 s below 20 °C. Capitalizing on this effect for polarization storage we obtained a visibility window of 9 min in blood. Conversion to [15N]nitrite, the bioactive reduced form of nitrate, was not observed in human blood and human saliva in this time frame. Thus, [15N]nitrate may serve as a long-lived hyperpolarized tracer for MR. Due to its ionic nature, the immediate applications appear to be perfusion and tissue retention imaging.

The ischemic penumbra in stroke is not clearly defined by today's available imaging tools. This study aimed to develop a model system and noninvasive biomarkers of ischemic brain tissue for an examination that might potentially be performed in humans, very quickly, in the course of stroke triage. Perfused rat brain slices were used as a model system and 31 P spectroscopy verified that the slices were able to recover from an ischemic insult of about 3.5 min of perfusion arrest. This was indicated as a return to physiological pH and adenosine triphosphate levels. Instantaneous changes in lactate dehydrogenase (LDH) and pyruvate dehydrogenase (PDH) activities were monitored and quantified by the metabolic conversions of hyperpolarized [1-13 C]pyruvate to [1-13 C]lactate and [13 C]bicarbonate, respectively, using 13 C spectroscopy. In a control group (n = 8), hyperpolarized [1-13 C]pyruvate was administered during continuous perfusion of the slices. In the ischemia group (n = 5), the perfusion was arrested 30 s prior to administration of hyperpolarized [1-13 C]pyruvate and perfusion was not resumed throughout the measurement time (approximately 3.5 min). Following about 110 s of the ischemic insult, LDH activity increased by 80.4 ± 13.5% and PDH activity decreased by 47.8 ± 25.3%. In the control group, the mean LDH/PDH ratio was 16.6 ± 3.3, and in the ischemia group, the LDH/PDH ratio reached an average value of 38.7 ± 16.9. The results suggest that monitoring the activity of LDH and PDH, and their relative activities, using hyperpolarized [1-13 C]pyruvate, could serve as an imaging biomarker to characterize the changes in the ischemic penumbra.

The ischemic penumbra in stroke is not clearly defined by today's available imaging tools. This study aimed to develop a model system and noninvasive biomarkers of ischemic brain tissue for an examination that might potentially be performed in humans, very quickly, in the course of stroke triage. Perfused rat brain slices were used as a model system and

The clinical characterization of small hepatocellular carcinoma (HCC) lesions in the liver and differentiation from heterogeneous inflammatory or fibrotic background is important for early detection and treatment. Metabolic monitoring of hyperpolarized 13C-labeled substrates has been suggested as a new avenue for diagnostic magnetic resonance. The metabolism of hyperpolarized [1-13C]pyruvate was monitored in mouse precision-cut liver slices (PCLS) of aged MDR2-KO mice, which served as a model for heterogeneous liver and HCC that develops similarly to the human disease. The relative in-cell activities of lactate dehydrogenase (LDH) to alanine transaminase (ALT) were found to be 0.40 &plusmn, 0.06 (n = 3) in healthy livers (from healthy mice), 0.90 &plusmn, 0.27 (n = 3) in heterogeneously inflamed liver, and 1.84 &plusmn, 0.46 (n = 3) in HCC. Thus, the in-cell LDH/ALT activities ratio was found to correlate with the progression of the disease. The results suggest that the LDH/ALT activities ratio may be useful in the assessment of liver disease. Because the technology used here is translational to both small liver samples that may be obtained from image-guided biopsy (i.e., ex vivo investigation) and to the intact liver (i.e., in a non-invasive MRI scan), these results may provide a path for differentiating heterogeneous liver from HCC in human subjects.

Adaptations of fruit trees to future climate are a current research priority due to the rapid increase in air temperature and changes in precipitation patterns. This is aimed at securing sustainable food production for our growing populations. Key physiological traits in trees conferring drought tolerance are resistance to embolism and stomatal control over water loss. Recently, we have shown in the field that a native wild pear species performs better under drought than two cultivated pear species. A comparative greenhouse study was conducted to investigate traits associated with drought tolerance in four ecotypes of a wild pear species (Pyrus syriaca Boiss), compared with a wild pear species (Pyrus betulifolia Bunge) commonly used as a pear rootstock. Seed sources were collected from semi-arid, sub-humid and humid sites across northern Israel. Measurements of water relations, leaf physiology, hydraulic conductivity and percent loss of hydraulic conductivity (PLC) were conducted under standard irrigation, drought and recovery from drought. The four P. syriaca ecotypes maintained significantly higher leaf gas exchange values and water-use efficiency and had lower PLC than the rootstock species under prolonged drought as well as during recovery. Across the four ecotypes, stomatal closure occurred at stem water potential (Ψ) around −3.5 MPa; however, Ψ at 50% PLC ranged from −4.1 MPa in the humid ecotype to −5.2 MPa in one of the semi-arid ecotypes, rendering the latter with a higher hydraulic safety margin (the Ψ difference between stomatal closure and 50% PLC). Divergence of the ecotypes in xylem vulnerability to embolism closely matched the mean annual precipitation at their seed sources. Thus, selection of pear ecotypes from populations in semi-arid sites may be better than the currently used plant material for preparing our cultivated species for hotter and drier future climate.

Cardiovascular diseases account for more than 30% of all deaths worldwide and many could be ameliorated with early diagnosis. Current cardiac imaging modalities can assess blood flow, heart anatomy and mechanical function. However, for early diagnosis and improved treatment, further functional biomarkers are needed. One such functional biomarker could be the myocardium pH. Although tissue pH is already determinable via MR techniques, and has been since the early 1990s, it remains elusive to use practically. The objective of this study was to explore the possibility to evaluate cardiac pH noninvasively, using in-cell enzymatic rates of hyperpolarized [1-13 C]pyruvate metabolism (ie, moles of product produced per unit time) determined directly in real time using magnetic resonance spectroscopy in a perfused mouse heart model. As a gold standard for tissue pH we used 31 P spectroscopy and the chemical shift of the inorganic phosphate (Pi) signal. The nonhomogenous pH distribution of the perfused heart was analyzed using a multi-parametric analysis of this signal, thus taking into account the heterogeneous nature of this characteristic. As opposed to the signal ratio of hyperpolarized [13 C]bicarbonate to [13 CO2 ], which has shown correlation to pH in other studies, we investigated here the ratio of two intracellular enzymatic rates: lactate dehydrogenase (LDH) and pyruvate dehydrogenase (PDH), by way of determining the production rates of [1-13 C]lactate and [13 C]bicarbonate, respectively. The enzyme activities determined here are intracellular, while the pH determined using the Pi signal may contain an extracellular component, which could not be ruled out. Nevertheless, we report a strong correlation between the tissue pH and the LDH/PDH activities ratio. This work may pave the way for using the LDH/PDH activities ratio as an indicator of cardiac intracellular pH in vivo, in an MRI examination.

The ability to directly monitor in vivo brain metabolism in real time in a matter of seconds using the dissolution dynamic nuclear polarization technology holds promise to aid the understanding of brain physiology in health and disease. However, translating the hyperpolarized signal observed in the brain to cerebral metabolic rates is not straightforward, as the observed in vivo signals reflect also the influx of metabolites produced in the body, the cerebral blood volume, and the rate of transport across the blood brain barrier. We introduce a method to study rapid metabolism of hyperpolarized substrates in the viable rat brain slices preparation, an established ex vivo model of the brain. By retrospective evaluation of tissue motion and settling from analysis of the signal of the hyperpolarized [1-13C]pyruvate precursor, the T1s of the metabolites and their rates of production can be determined. The enzymatic rates determined here are in the range of those determined previously with classical biochemical assays and are in agreement with hyperpolarized metabolite relative signal intensities observed in the rodent brain in vivo.

In a previous work, it was found that ‘Spadona’ the main European pear cultivar grown in Israel, is semi-compatible with each of its three pollinators ‘Gentile’,‘Coscia’ and ‘Spadochina’. Thus, the full yield potential of these cultivars is not being realized. The Japanese pear (Pyrus pyrifolia) can fertilize European pears. In Israel, the environmental conditions for growing pears are sub-optimal for most pear cultivars. However, the Japanese pear ‘Hosui’ is well adapted to these conditions; moreover, it is fully compatible and flowers simultaneously with the cultivated European pears. In this work, we report on a 10-year evaluation of pollination of ‘Spadona’ and ‘Coscia’ pollinated by ‘Hosui’. The experiments were conducted in three stages: hand pollination; ‘bouquet insertion’; and a combined European-Asian pear orchard. The results indicate that this is a beneficial means for significantly improving fruit quality and quantity.

Organoids are a powerful tool in the quest to understand human diseases. As the developing brain is extremely inaccessible in mammals, cerebral organoids (COs) provide a unique way to investigate neural development and related disorders. The aim of this study was to utilize hyperpolarized 13C NMR to investigate the metabolism of COs in real-time, in a non-destructive manner. The enzymatic activity of lactate dehydrogenase (LDH) was determined by quantifying the rate of [1-13C]lactate production from hyperpolarized [1-13C]pyruvate. Organoid development was assessed by immunofluorescence imaging. Organoid viability was confirmed using 31P NMR spectroscopy. A total of 15 organoids collated into 3 groups with a group total weight of 20–77 mg were used in this study. Two groups were at the age of 10 weeks and one was at the age of 33 weeks. The feasibility of this approach was demonstrated in both age groups, and the LDH activity rate was found to be 1.32 ± 0.75 nmol/s (n = 3 organoid batches). These results suggest that hyperpolarized NMR can be used to characterize the metabolism of brain organoids with a total tissue wet weight of as low as 20 mg (3) and a diameter ranging from 3 to 6 mm.

Pollination is limiting for avocado production. We examined whether adding bumblebees (BBs, ca. 10 hives/ha) to conventional honeybees (HB, 5 hives/ha) would improve ‘Hass’ avocado pollination and yields. A preliminary trial (2017/18) in an avocado orchard with four consecutive rows of ‘Hass’ followed by one row of ‘Ettinger’ serving as a pollenizer (20% ‘Ettinger’) showed a considerable increase in ‘Hass’ yield in rows adjacent to (up to 80 m from) the BB hives vs. distant rows (=controls). In 2018/19, the trials were extended to three additional orchards. A significant yield increase was obtained in the BB hive-adjacent trees compared to BB hive-distant ones. Similar results were obtained in 2019/20, in experiments conducted throughout the country. The SNP analysis, to determine the parents of ‘Hass’ fruit at varying distances from the BB hives, showed no differences in the cross-pollination rate (‘Hass’ × ‘Ettinger’). However, pollination rates and the number of germinating pollen grains per stigma decreased with distance from the hives, and correlated to the negative gradient in yield. Taken together, our data suggest that adding BB hives to ‘Hass’ avocado orchards, at ca. 10 hives/ha resulting in 0.5–1.0 BB visits/tree per min, increases pollination and, accordingly, total yield.

The dissolution-dynamic nuclear polarization technology had previously enabled nuclear magnetic resonance detection of various nuclei in a hyperpolarized state. Here, we show the hyperpolarization of 31P nuclei in important biological phosphates (inorganic phosphate and phosphocreatine) in aqueous solutions. The hyperpolarized inorganic phosphate showed an enhancement factor >11,000 (at 5.8 T, 9.3% polarization) in D2O (T1 29.4 s). Deuteration and the solution composition and pH all affected the lifetime of the hyperpolarized state. This capability opens up avenues for real-time monitoring of phosphate metabolism, distribution, and pH sensing in the live body without ionizing radiation. Immediate changes in the microenvironment pH have been detected here in a cell-free system via the chemical shift of hyperpolarized inorganic phosphate. Because the 31P nucleus is 100% naturally abundant, future studies on hyperpolarized phosphates will not require expensive isotope labeling as is usually required for hyperpolarization of other substrates., Real-time monitoring of phosphate metabolism and distribution in the live body without ionizing radiation is highly desirable. Here, the authors show dissolution-dynamic nuclear polarization technology can enable nuclear magnetic resonance detection of hyperpolarized 31P of important biological phosphates in aqueous solutions.

The ecto-nucleoside triphosphate diphosphohydrolase-1 (E-NTPDase-1, CD39) enzyme is responsible for the breakdown of extracellular ATP to ADP and then to AMP by a two-step process. Defective CD39 activity has been described in a variety of medical conditions including malignancy and rheumatic diseases and has been proved to be of major diagnostic and clinical importance. Here we show for the first time that a 31P NMR spectroscopy methodology enables the quantification of these two steps in a single blood sample. We have applied this assay to determine the E-NTPDase activity on human mononuclear cells taken from two siblings affected by a stop-codon mutation in the ENTPD1 gene, their obligatory heterozygous parents, and healthy volunteers. The affected subjects presented low ATP breakdown activity, mainly expressed as low AMP production.

[1-13C]pyruvate, the most widely used compound in dissolution-dynamic nuclear polarization (dDNP) magnetic resonance (MR), enables the visualization of lactate dehydrogenase (LDH) activity. This activity had been demonstrated in a wide variety of cancer models, ranging from cultured cells, to xenograft models, to human tumors in situ. Here we quantified the LDH activity in precision cut tumor slices (PCTS) of breast cancer xenografts. The Michigan Cancer Foundation-7 (MCF7) cell-line was chosen as a model for the luminal breast cancer type which is hormone responsive and is highly prevalent. The LDH activity, which was manifested as [1-13C]lactate production in the tumor slices, ranged between 3.8 and 6.1 nmole/nmole adenosine tri-phosphate (ATP) in 1 min (average 4.6 &plusmn, 1.0) on three different experimental set-ups consisting of arrested vs. continuous perfusion and non-selective and selective RF pulsation schemes and combinations thereof. This rate was converted to an expected LDH activity in a mass ranging between 3.3 and 5.2 &micro, mole/g in 1 min, using the ATP level of these tumors. This indicated the likely utility of this approach in clinical dDNP of the human breast and may be useful as guidance for treatment response assessment in a large number of tumor types and therapies ex vivo.

Investigation of hyperpolarized substrate metabolism has been showing utility in real-time determination of in-cell and in vivo enzymatic activities. Intracellular reaction rates may vary during the course of a measurement, even on the very short time scales of visibility on hyperpolarized MR, due to many factors such as the availability of the substrate and co-factors in the intracellular space. Despite this potential variation, the kinetic analysis of hyperpolarized signals typically assumes that the same rate constant (and in many cases, the same rate) applies throughout the course of the reaction as observed via the build-up and decay of the hyperpolarized signals. We demonstrate here an acquisition approach that can null the need for such an assumption and enable the detection of instantaneous changes in the rate of the reaction during an ex vivo hyperpolarized investigation, (i.e. in the course of the decay of one hyperpolarized substrate dose administered to a viable tissue sample ex vivo). This approach utilizes hyperpolarized product selective saturating-excitation pulses. Similar pulses have been previously utilized in vivo for spectroscopic imaging. However, we show here favorable consequences to kinetic rate determinations in the preparations used. We implement this acquisition strategy for studies on perfused tissue slices and develop a theory that explains why this particular approach enables the determination of changes in enzymatic rates that are monitored via the chemical conversions of hyperpolarized substrates. Real-time changes in intracellular reaction rates are demonstrated in perfused brain, liver, and xenograft breast cancer tissue slices and provide another potential differentiation parameter for tissue characterization.

Reports on gadolinium deposits in the body and brains of adults and children who underwent contrast-enhanced MRI examinations warrant development of new, metal free, contrast agents for MRI. Nitrate is an abundant ion in mammalian biochemistry and sodium nitrate can be safely injected intravenously. We show that hyperpolarized [

Precision-cut liver slices (PCLS) are widely used in liver research as they provide a liver model with all liver cell types in their natural architecture. The purpose of this study was to demonstrate the use of PCLS for hyperpolarized metabolic investigation in a mouse model, for potential future application in liver biopsy cores. Fresh normal liver was harvested from six mice. 500 μm PCLS were prepared and placed in a 10 mm NMR tube in an NMR spectrometer and perfused continuously.

The ability to directly monitor in vivo brain metabolism in real time in a matter of seconds using the dissolution dynamic nuclear polarization technology holds promise to aid the understanding of brain physiology in health and disease. However, translating the hyperpolarized signal observed in the brain to cerebral metabolic rates is not straightforward, as the observed in vivo signals reflect also the influx of metabolites produced in the body, the cerebral blood volume, and the rate of transport across the blood brain barrier. We introduce a method to study rapid metabolism of hyperpolarized substrates in the viable rat brain slices preparation, an established ex vivo model of the brain. By retrospective evaluation of tissue motion and settling from analysis of the signal of the hyperpolarized [1-13C]pyruvate precursor, the T1s of the metabolites and their rates of production can be determined. The enzymatic rates determined here are in the range of those determined previously with classical biochemical assays and are in agreement with hyperpolarized metabolite relative signal intensities observed in the rodent brain in vivo.

The ecto-nucleoside triphosphate diphosphohydrolase-1 (E-NTPDase-1, CD39) enzyme is responsible for the breakdown of extracellular ATP to ADP and then to AMP by a two-step process. Defective CD39 activity has been described in a variety of medical conditions including malignancy and rheumatic diseases and has been proved to be of major diagnostic and clinical importance. Here we show for the first time that a

Water availability is becoming a limiting factor with increasing world population that challenges global food security. Thus, we need to enhance cultivation in increasingly drier and hotter climate and prepare fruit trees for the ongoing climate change. Wild tree species might offer vital information and plant material in face of these challenges. A year-long comparative field study was conducted to investigate the mechanisms underlying drought tolerance in pear species (cultivated Pyrus communis and Pyrus pyrifolia vs. the wild Pyrus syriaca). We confirmed the hypothesis of higher drought tolerance in wild pear compared to its cultivated relative. P. syriaca xylem had fewer, narrower vessels, and lower vulnerability to embolism. It showed higher intrinsic water-sue efficiency and more robust seasonal patterns of photosynthesis, hydraulic conductivity, and PIP (plasma intrinsic protein) aquaporin expression. Across species, we identified a ubiquitous gene (PIP1:5/1:6), nine drought-inhibited genes, and two drought-induced genes (PIP1:4 and 2:6/2:7, confirming previous studies). Our study highlights the potential of using wild relatives of fruit tree species to prepare key crops to a drier and hotter future. The study of PIPs leads the way to a more focused research of the role of these cellular water channels in minimizing tree water loss under drought, while ensuring hydration of specific tissues.

Precision-cut liver slices (PCLS) are widely used in liver research as they provide a liver model with all liver cell types in their natural architecture. The purpose of this study was to demonstrate the use of PCLS for hyperpolarized metabolic investigation in a mouse model, for potential future application in liver biopsy cores. Fresh normal liver was harvested from six mice. 500 μm PCLS were prepared and placed in a 10 mm NMR tube in an NMR spectrometer and perfused continuously. 31 P spectra were acquired to evaluate the presence of adenosine triphosphate (ATP) and validate viability in all samples. Hyperpolarized [1-13 C]pyruvate was flushed into the NMR tube in the spectrometer. Consecutive 13 C NMR spectra were acquired immediately after the injection using both non-selective (five injections, two livers) and selective RF excitation (six injections, three livers). The 31 P spectra showed the characteristic signals of ATP, confirming the viability of the PCLS for more than 2.5 h in the spectrometer. After each of the [1-13 C]pyruvate injections, both [1-13 C]lactate and [1-13 C]alanine signals were detected. Selective RF excitation aimed at both [1-13 C]lactate and [1-13 C]alanine enabled better visualization and quantification of the metabolic activity. Using this acquisition approach only the newly formed metabolites are observed upon excitation, and their intensities relative to those of hyperpolarized pyruvate enable quantification of metabolite production rates. This rate of lactate and alanine production appeared to be constant throughout the measurement time, with alanine production about 2.3 times higher than lactate. In summary, the viability of PCLS in an NMR spectrometer was demonstrated and hyperpolarized [1-13 C]pyruvate metabolism was recorded. This study opens up the possibility of evaluating alanine aminotransferase (ALT) and lactate dehydrogenase (LDH) activities in human liver biopsies, while preserving the tissue architecture and viability. In healthy, well-perfused liver slices the ratio of ALT to LDH activity is about 2.3.

The original version of the Supplementary Information associated with this Article contained an error in Supplementary Figure 2 and Supplementary Figure 5 in which the 31P NMR spectral lines were missing. The HTML has been updated to include a corrected version of the Supplementary Information.

Determination of the genetic compatibility between self-incompatible cultivars is crucial in agriculture. The Rosaceae family carries the S -RNase-mediated gametophytic self-incompatibility (GSI) system. Each haplotype is conferred by an S -locus. The S -locus contains two highly polymorphic genes, S-RNase and SFB , which are characteristic of each haplotype and therefore these genes are ideal markers for molecular S -genotyping. In this study 43 Japanese plum cultivars grown in Israel were S -genotyped based on their S-RNase gene sequences. Four alleles, S b , S c , S e and S h are widespread and together are responsible for 87% of the S -haplotypes therefore many of the cultivar combinations are semi-compatible. In Israel semi-compatibility was shown to correlate with low yield. However, two cultivars, ‘Wickson’ S f S k and ‘Shiro’ S f S g carry rare S -haplotypes and, therefore, are fully compatible with most of the analyzed cultivars.

Japanese plum belongs to the Rosaceae, which carry the S-RNase-based gametophytic self-incompatibility system. Therefore, these plum cultivars depend on cross-pollination for setting fruit. However, cultivar combinations may be genetically semi- or fully compatible. In apples and pears cultivated in Israel, where conditions for pollination are usually unfavorable, it was demonstrated that yields may be correlated with compatibility levels of pollinators. In the present study fruit-set of semi- and fully compatible plum couples was determined for a wide range of cultivars, in unfavorable and favorable conditions. Full compatibility was found superior to semi-compatibility, in both unfavorable and favorable conditions for pollination. These findings should be taken into consideration in determining the orchard designs of cultivars.

Japanese plum (Prunus salicina Lindl.), a species of the Rosaceae family, carries the S-RNase-mediated gametophytic self-incompatibility system. Self-incompatibility is manifested if the S-haplotype of the pollen is carried also by the pollinated flower. Thus, for fertilization to occur, the cultivars have to be genetically compatible. The haplotype is conferred by an S-locus, which contains the style-specific expressed S-RNase and the pollen-specific expressed F-box genes (SFB). Since both the S-RNase and the SFB genes are multiallelic and are characteristic of each of the S-haplotypes, they are ideal markers for molecular S-typing. In this work, seven SFBs, from eight japanese plum cultivars, were cloned and sequenced. Five of the alleles were published recently and two SFBg and SFBk are new. The physical linkage of SFBb and SFBc to their adjacent S-RNase was determined; it is 544 base pairs (bp) and 404 bp for the Sb and Sc loci, respectively.

SummaryJapanese plum (Prunus salicina Lindl.) belongs to the Rosaceae family, which carries the S-RNase-mediated gametophytic self-incompatibility system, which prevents self-fertilisation, and thus promotes out-crossing. The plum cultivar ‘Black Diamond’® has become, one of the most important cultivars in Israel in the last decade, yet its yield is low in comparison with its known potential. Honeybees (Apis mellifera) are the most important pollinators for plums and several studies have demonstrated an apparent relationship between the number of honeybees and the size of the fruit crop. Therefore, in this study, we focussed on improving bee management in a ‘Black Diamond’ orchard. In four consecutive years of experiments, we examined the effects of increasing the density of bee colonies, and of making multiple introductions of colonies, on honeybee activity and on their effectiveness as pollinators of ‘Black Diamond’. We showed that four separate introductions, each of 0.625 colonies ha–1, every 2 – 3 d ...

SummaryThe Japanese plum (Prunus salicina Lindl.) belongs to the Rosaceae, which carries the gametophytic self-incompatibility system. Therefore, commercial plum orchards must contain at least two cultivars, which are able to fertilize each other. However, the S-allele content, which determines the fertilization compatibility, has not yet been identi®ed in most of the Japanese plum varieties. This lack of knowledge may lead to cases of reduced yields due to semi, instead of full, compatibility. In the present study, ®ve S-alleles were cloned from three Japanese plum cultivars. The genes were isolated, from genomic DNA, by applying the PCR (Polymerase Chain Reaction), resulting in fragments containing exons, as well as introns. Four of the ®ve clones turned out to be new alleles: from `Wickson' S3 and S6, and from `Royal-Zee', S4 and S5. The ®fth allele, S1, which was isolated from `Red-Beaut', has been cloned previously as a cDNA sequence from `Sordum' (named Sa, GenBank accession no. AB026981). Sequence ...

In the gametophytic self-incompatibility system, growth of self-pollen tubes in the style is inhibited in a haplotype-specific manner by S-RNase. The mechanism by which S-RNase confers its specificity is unknown. However, a hypervariable region (RHV in Rosaceae and HVa-HVb in Solanaceae) that differs among the many cloned S-RNase alleles has been proposed to be involved in conferring the S-haplotype specificity of the S-RNase. Region swapping experiments between S-RNases and crystallography of the enzyme support this assumption. However, the deduced amino acid sequences of Sn-RNase and Si-RNase alleles from the European pear (Pyrus communis) were recently found to have an identical RHV. In the present study it is shown that Sn-RNase does not prevent fertilization by Si-pollen haplotype, thus presenting a case in which RHV is not required for the determination of specific pollen rejection by S-RNase, and implying that other regions in the enzyme may be sufficient for this specificity.