Improving the extraction and purification of leaf and phloem sugars for oxygen isotope analyses.

RATIONALE: The oxygen isotopic composition (here shown as the δ18 O value) of soluble sugars in leaves and phloem tissue holds valuable information about plant functions in response to climatic changes. However, δ18 O analysis of sugars is prone to error, and thoroughly tested methods are lacking. METHODS: We performed three experiments to test if sample preparation modifies the δ18 O values of sugars. In experiment 1, we tested the effects of oven-drying versus freeze-drying, whereas in experiment 2 we focused on the extraction and purification of leaf sugars. In experiment 3, we investigated the exudation and purification of twig phloem sugars as a function of exudation time and different ethylenediaminetetraacetic acid (EDTA) exudation media. RESULTS: Freeze-drying produced more consistent δ18 O values than oven-drying for sucrose but not for phloem sugars. The extraction and purification of leaf sugars can be performed without a significant modification of their δ18 O values; yet the purified leaf and phloem sugars possessed higher δ18 O values than the fraction of water-soluble compounds. Moreover, the exudation time significantly modulated the δ18 O values of phloem sugars, which is probably related to changes in the sugar composition. The addition of EDTA did not improve the determination of the δ18 O values of phloem sugars. CONCLUSIONS: We show that the sample preparation of plant sugars for the reliable determination of δ18 O values requires a strict protocol, which is described in this paper. For phloem sugar, we recommend a maximum exudation time of 1 h to reduce the degradation of sucrose and minimise oxygen isotope exchange reactions between the resulting hexoses and water.

Employing stable isotopes to determine the residence times of soil water and the temporal origin of water taken up by Fagus sylvatica and Picea abies in a temperate forest.

We assessed how the seasonal variability of precipitation δ2 H and δ18 O is propagated into soil and xylem waters of temperate trees, applied a hydrological model to estimate the residence time distribution of precipitation in the soil, and identified the temporal origin of water taken up by Picea abies and Fagus sylvatica over 4 yr. Residence times of precipitation in the soil varied between a few days and several months and increased with soil depth. On average, 50% of water consumed by trees throughout a year had precipitated during the growing season, while 40% had precipitated in the preceding winter or even earlier. Importantly, we detected subtle differences with respect to the temporal origin of water used by the two species. We conclude that both current precipitation and winter precipitation are important for the water supply of temperate trees and that winter precipitation could buffer negative impacts of spring or summer droughts. Our study additionally provides the means to obtain realistic estimates of source water δ2 H and δ18 O values for trees from precipitation isotope data, which is essential for improving model-based interpretations of δ18 O and δ2 H values in plants.

Temperate tree species show identical response in tree water deficit but different sensitivities in sap flow to summer soil drying.

Temperate forests are expected to be particularly vulnerable to drought and soil drying because they are not adapted to such conditions and perform best in mesic environments. Here we ask (i) how sensitively four common temperate tree species (Fagus sylvatica, Picea abies, Acer pseudoplatanus and Fraxinus excelsior) respond in their water relations to summer soil drying and seek to determine (ii) if species-specific responses to summer soil drying are related to the onset of declining water status across the four species. Throughout 2012 and 2013 we determined tree water deficit (TWD) as a proxy for tree water status from recorded stem radius changes and monitored sap flow rates with sensors on 16 mature trees studied in the field at Lägeren, Switzerland. All tree species responded equally in their relative maximum TWD to the onset of declining soil moisture. This implies that the water supply of all tree species was affected by declining soil moisture and that none of the four species was able to fully maintain its water status, e.g., by access to alternative water sources in the soil. In contrast we found strong and highly species-specific responses of sap flow to declining soil moisture with the strongest decline in P. abies (92%), followed by F. sylvatica (53%) and A. pseudoplatanus (48%). F. excelsior did not significantly reduce sap flow. We hypothesize the species-specific responses in sap flow to declining soil moisture that occur despite a simultaneous increase in relative TWD in all species reflect how fast these species approach critical levels of their water status, which is most likely influenced by species-specific traits determining the hydraulic properties of the species tree.

Functional and motor outcome 5 years after stroke is equivalent to outcome at 2 months: follow-up of the collaborative evaluation of rehabilitation in stroke across Europe.

BACKGROUND AND PURPOSE: Recovery of patients within the first 6 months after stroke is well documented, but there has been little research on long-term recovery. The aim of this study was to analyze functional and motor recovery between admission to rehabilitation centres and 5 years after stroke. METHODS: This follow-up of the Collaborative Evaluation of Rehabilitation in Stroke Across Europe study, included patients from 4 European rehabilitation centres. Patients were assessed on admission, at 2 and 6 months, and 5 years after stroke, using the Barthel Index, Rivermead Motor Assessment Gross Function, Leg and Trunk function, and Arm function. Linear mixed models were used, corrected for baseline characteristics. To account for the drop-out during follow-up, the analysis is likelihood-based (assumption of missingness at random). RESULTS: A total of 532 patients were included in this study, of which 238 were followed up at 5 years post stroke. Mean age at stroke onset was 69 (±10 SD) years, 53% were men, 84% had ischemic strokes, and 53% had left-sided motor impairment. Linear mixed model analysis revealed a significant deterioration for all 4 outcomes between 6 months and 5 years (P<0.0001). Scores at 2 months were not statistically significant different from scores at 5 years after stroke. Higher age (P<0.0001) and increasing stroke severity on admission (P<0.0001) negatively affected long-term functional and motor recovery. CONCLUSIONS: Five-year follow-up revealed deterioration in functional and motor outcome, with a return to the level measured at 2 months. Increasing age and increasing stroke severity negatively affected recovery up to 5 years after stroke.

Long-term prediction of functional outcome after stroke using single items of the Barthel Index at discharge from rehabilitation centre.

PURPOSE: To determine the prognostic value of single items of the Barthel Index (BI) at discharge from rehabilitation, in predicting independence in personal activities of daily living (ADL) (BI score ≥ 95/100) at five years after stroke. METHOD: People with stroke were recruited consecutively from four European rehabilitation centres. BI was assessed on discharge and at five years after stroke. Stepwise multivariate logistic regression analysis was used to determine independent predictors of BI score ≥ 95/100 at five years after stroke. Thereupon, percentage chance of reaching BI ≥ 95/100 at five years after stroke was calculated. RESULTS: Data were available for 153 patients. Independence in dressing (odds ratio (OR)=5.22, 95% confidence interval (CI)=1.85-14.76, p=0.002) and bathing (OR=8.10, 95% CI=3.40-19.32, p<0.0001) were independent predictors. Independence in both items resulted in 74.1% (57.6-85.8) chance of reaching BI ≥ 95/100 at five years after stroke. Dependence in both items resulted in 6.3% (5.1-7.9) chance. Independence in bathing, but dependence in dressing resulted in 35.4% (30.7-40.4) chance whereas the opposite resulted in 26.1% (20.7-32.3) chance. CONCLUSION: Simple assessment of dressing and bathing on discharge from rehabilitation enables therapeutic staff to predict prognosis for long-term independence in personal ADL. This method can be used for early identification of persons with stroke who need intensive follow-up. Implications for Rehabilitation (In)dependence for dressing and bathing at discharge from a rehabilitation centre are significant factors in the prediction of (in)dependence in personal ADL at five years after stroke. This predictive tool can be used for targeting inpatient stroke rehabilitation and early identification of those patients who need intensive follow-up.

Five-year mortality and related prognostic factors after inpatient stroke rehabilitation: a European multi-centre study.

OBJECTIVE: To determine 5-year mortality and its association with baseline characteristics and functional status 6 months post-stroke for patients who received inpatient rehabilitation. DESIGN: A prospective rehabilitation-based cohort study. SUBJECTS: A total of 532 consecutive stroke patients from 4 European rehabilitation centres. METHODS: Predictors were recorded on admission. Barthel Index was assessed at 6 months (BI6mths) and patients were followed for 5 years post-stroke. Survival probability was computed using Kaplan-Meier analysis and compared across 3 BI6mths-classes (0-60, 65-90, 95-100) (log-rank test). Significant independent predictors were determined using multivariate Cox regression analysis (hazard ratio (HR)). RESULTS: Five-year cumulative risk of death was 29.12% (95% confidence interval (CI): 22.86-35.38). Age (HR=1.06, 95% CI: 1.04-1.09), cognitive impairment (HR=1.77, 95% CI: 1.21-2.57), diabetes mellitus (HR=1.68, 95% CI: 1.16-2.41) and atrial fibrillation (HR=1.52, 95% CI: 1.08-2.14) were independent predictors of increased mortality. Hyperlipidaemia (HR=0.66, 95% CI: 0.46-0.94), and higher BI6mths (HR=0.98, 95% CI: 0.97-0.99) were independent predictors of decreased mortality. Five-year survival probability was 0.85 (95% CI: 0.80-0.89) for patients in BI6mths-class: 95-100, 0.72 (95% CI: 0.63-0.79) in BI6mths-class: 65-90 and 0.50 (95% CI: 0.40-0.60) in BI6mths-class: 0-60 (p<0.0001). CONCLUSION: Nearly one-third of rehabilitation patients died during the first 5 years following stroke. Functional status at 6 months was a powerful predictor of long-term mortality. Maximum functional independence at 6 months post-stroke should be promoted through medical interventions and rehabilitation. Future studies are recommended to evaluate the direct effect of rehabilitation on long-term survival.

Variations in follow-up services after inpatient stroke rehabilitation: a multicentre study.

BACKGROUND: Care after discharge from inpatient stroke rehabilitation units varies across Europe. The aim of this study was to compare service delivery after discharge. METHODS: A total of 532 consecutive patients after stroke were recruited from 4 European rehabilitation centres in Germany, Switzerland, Belgium and the UK. At 2-month intervals, clinical assessments and structured interviews were carried out to document functional status and delivery of services after discharge. Significant factors for receiving follow-up services were analysed using a logistic generalized estimating equation model. RESULTS: After controlling for case-mix, the results showed that Belgian patients were most likely to receive physical therapy but least likely to receive occupational therapy. German patients were least likely to receive nursing care. UK patients were less likely to receive medical care from their general practitioner compared with the other patient groups. CONCLUSION: Clinical characteristics did not explain the variations in service delivery after discharge from in-patient stroke rehabilitation. The decision-making processes involved in the provision of follow-up services need to be better documented. To improve our understanding of events post-discharge, the influence of non-clinical factors, such as healthcare regulations, should be explored further.

Anxiety and depression in the first six months after stroke. A longitudinal multicentre study.

PURPOSE: To document the prevalence, severity and time course of anxiety and depression in stroke rehabilitation patients in four European countries. METHOD: At two, four and six months post-stroke, the prevalence and severity of anxiety and depression were determined in 532 consecutively recruited patients, using the Hospital Anxiety and Depression Scale. Time course of prevalence and severity was examined, using Cochran-Q and Friedman-tests, respectively. We identified whether the numbers of anxious/depressed patients at each time point comprised the same individuals. RESULTS: Prevalence of anxiety ranged between 22% and 25%; depression between 24% and 30%. Median severity ranged between 4 and 5. No significant differences between centres occurred (p > 0.05). Prevalence of both disorders was not significantly different over time. Severity of anxiety decreased between four and six months; severity of depression remained stable. About 40% of the patients with initial anxiety remained anxious at six months. Some 11% and 7% of those initially not anxious became anxious at four or six months after stroke, respectively. Depression showed a similar pattern. CONCLUSIONS: Despite differences in patient profiles and intensity of rehabilitation, no significant differences occurred between centres in prevalence and severity of both disorders. Anxiety was almost as common as depression and additional patients became anxious/depressed at each time point.

Motor and functional recovery after stroke: a comparison of 4 European rehabilitation centers.

BACKGROUND AND PURPOSE: Outcome after first stroke varies significantly across Europe. This study was designed to compare motor and functional recovery after stroke between four European rehabilitation centers. METHODS: Consecutive stroke patients (532 patients) were recruited. They were assessed on admission and at 2, 4, and 6 months after stroke with the Barthel Index, Rivermead Motor Assessment of Gross Function, Rivermead Motor Assessment of Leg/Trunk, Rivermead Motor Assessment of Arm, and Nottingham Extended Activities of Daily Living (except on admission). Data were analyzed using random effects ordinal logistic models adjusting for case-mix and multiple testing. RESULTS: Patients in the UK center were more likely to stay in lower Rivermead Motor Assessment of Gross Function classes compared with patients in the German center (DeltaOR, 2.4; 95% CI, 1.3 to 4.3). In the Swiss center, patients were less likely to stay in lower Nottingham Extended Activities of Daily Living classes compared with patients in the UK center (DeltaOR, 0.7; 95% CI, 0.5 to 0.9). The latter were less likely to stay in lower Barthel Index classes compared with the patients in the German center (DeltaOR, 0.6; 95%CI, 0.4 to 0.8). Recovery patterns of Rivermead Motor Assessment of Leg/Trunk and Rivermead Motor Assessment of Arm were not significantly different between centers. CONCLUSIONS: Gross motor and functional recovery were better in the German and Swiss centers compared with the UK center, respectively. Personal self-care recovery was better in the UK compared with the German center. Previous studies in the same centers indicated that German and Swiss patients received more therapy per day. This was not the result of more staff but of a more efficient use of human resources. This study indicates potential for improving rehabilitation outcomes in the UK and Belgian centers.

Effect of socioeconomic status on functional and motor recovery after stroke: a European multicentre study.

BACKGROUND: Previous studies have shown an inverse gradient in socioeconomic status for disability after stroke. However, no distinction has been made between the period in the stroke rehabilitation unit (SRU) and the period after discharge. The purpose of this study was to examine the impact of education and equivalent income on motor and functional recovery for both periods. METHODS: 419 consecutive patients were recruited from six SRUs across Europe. The Barthel Index (BI) and Rivermead Motor Assessment (RMA) were measured on admission, at discharge and 6 months after stroke. Ordinal logistic regression models were used, adjusting for case mix. Cumulative odds ratios (OR) were calculated to measure differences in recovery between educational levels and income groups with adjustments for case mix. RESULTS: Patients with a low educational level were less likely to improve on the BI (OR 0.53; 95% CI 0.32 to 0.87) and the RMA arm during inpatient stay (OR 0.54; 95% CI 0.31 to 0.94). For this period, no differences in recovery were found between income groups. After discharge, patients with a low equivalent income were less likely to improve on all three sections of the RMA: gross function (OR 0.20; 95% CI 0.06 to 0.66), leg and trunk (OR 0.22; 95% CI 0.09 to 0.55) and arm (OR 0.30; 95% CI 0.10 to 0.87). No differences were found for education. CONCLUSIONS: During inpatient rehabilitation, educational level was a determinant of recovery, while after discharge, equivalent income played an important role. This study suggests that it is important to develop a better understanding of how socioeconomic factors affect the recovery of stroke patients.

Stroke rehabilitation in Europe: what do physiotherapists and occupational therapists actually do?

BACKGROUND AND PURPOSE: Physiotherapy (PT) and occupational therapy (OT) are key components of stroke rehabilitation. Little is known about their content. This study aimed to define and compare the content of PT and OT for stroke patients between 4 European rehabilitation centers. METHODS: In each center, 15 individual PT and 15 OT sessions of patients fitting predetermined criteria were videotaped. The content was recorded using a list comprising 12 therapeutic categories. A generalized estimating equation model was fitted to the relative frequency of each category resulting in odds ratios. RESULTS: Comparison of PT and OT between centers revealed significant differences for only 2 of the 12 categories: ambulatory exercises and selective movements. Comparison of the 2 therapeutic disciplines on the pooled data of the 4 centers revealed that ambulatory exercises, transfers, exercises, and balance in standing and lying occurred significantly more often in PT sessions. Activities of daily living, domestic activities, leisure activities, and sensory, perceptual training, and cognition occurred significantly more often in OT sessions. CONCLUSIONS: This study revealed that the content of each therapeutic discipline was consistent between the 4 centers. PT and OT proved to be distinct professions with clear demarcation of roles.

Use of time by stroke patients: a comparison of four European rehabilitation centers.

BACKGROUND AND PURPOSE: Differences exist between European countries in the proportion of patients who die or become dependent after stroke. The aim of the present study was to identify differences in the use of time by stroke patients in 4 rehabilitation centers in 4 European countries. METHODS: In each of the 4 centers, 60 randomly selected stroke patients were observed at 10-minute intervals using behavioral mapping. Observations took place on 30 weekdays selected at random, on equal numbers of morning, afternoon, and evening sessions. A logistic generalized estimating equation model with correction for differences in case mix and multiple testing was used for the analysis. RESULTS: Overall time available from different professions was the highest in the United Kingdom, but patients in the United Kingdom spent on average only 1 hour per day in therapy. This was significantly less than patients in Belgium and Germany, who spent approximately 2 hours, and patients in Switzerland who spent approximately 3 hours per day in therapy. In all centers, patients spent less than half their time in interactions and >72% of the time in nontherapeutic activities. CONCLUSIONS: Important differences in the use of time were established, which appeared dependent on management decisions rather than the number of staff available. Patients in the Swiss and German centers spent more time in therapy, possibly because of the structured organization of rehabilitation. Further studies will verify whether this has an effect on outcome.