Brief report: Bereaved parents informing research design: The place of a pilot study.

BACKGROUND: Risk minimization in research with bereaved parents is important. However, little is known about which research methods balance the sensitivity required for bereaved research participants and the need for generalizable results. AIM: To explore parental experiences of participating in mixed method bereavement research via a pilot study. DESIGN: A convergent parallel mixed method design assessing bereaved parents' experience of research participation. SETTING/PARTICIPANTS: Eleven parents whose child was treated for cancer at The Royal Children's Hospital, Brisbane completed the questionnaire/interview being piloted (n = 8 mothers; n = 3 fathers; >6 months and <6 years bereaved). Of these, eight parents completed the pilot study evaluation questionnaire, providing feedback on their experience of participation. RESULTS: Participants acknowledged the importance of bereaved parents being central to research design and the development of bereavement programs. Sixty-three per cent (n = 5/8) of parents described completion of the questionnaire as 'not at all/a little bit' of a burden. Seventy-five per cent (n = 6/8) of parents opting into the telephone interview described participation as 'not at all/a little bit' of a burden. When considering the latest timeframes for participation in bereavement research 63% (n = 5/8) of parents indicated 'no endpoint.' Findings from the pilot study enabled important adjustments to be made to a large-scale future study. CONCLUSIONS: As a research method, pilot studies may be utilized to minimize harm and maximize the potential benefits for vulnerable research participants. A mixed method approach allows researchers to generalize findings to a broader population while also drawing on the depth of the lived experience.

Are hybrid species more fit than ancestral parent species in the current hybrid species habitats?

Hybrid speciation is thought to be facilitated by escape of early generation hybrids into new habitats, subsequent environmental selection and adaptation. Here, we ask whether two homoploid hybrid plant species (Helianthus anomalus, H. deserticola) diverged sufficiently from their ancestral parent species (H. annuus, H. petiolaris) during hybrid speciation so that they are more fit than the parent species in hybrid species habitats. Hybrid and parental species were reciprocally transplanted into hybrid and parental habitats. Helianthus anomalus was more fit than parental species in the H. anomalus actively moving desert dune habitat. The abilities to tolerate burial and excavation and to obtain nutrients appear to be important for success in the H. anomalus habitat. In contrast, H. deserticola failed to outperform the parental species in the H. deserticola stabilized desert dune habitat, and several possible explanations are discussed. The home site advantage of H. anomalus is consistent with environmental selection having been a mechanism for adaptive divergence and hybrid speciation and supports the use of H. anomalus as a valuable system for further assessment of environmental selection and adaptive traits.

Phenotypic selection on leaf ecophysiological traits in Helianthus.

Habitats that differ in soil resource availability are expected to differ for selection on resource-related plant traits. Here, we examined spatial and temporal variation in phenotypic selection on leaf ecophysiological traits for 10 Helianthus populations, including two species of hybrid origin, Helianthus anomalus and Helianthus deserticola, and artificial hybrids of their ancestral parents. Leaf traits assessed were leaf size, succulence, nitrogen (N) concentration and water-use efficiency (WUE). Biomass and leaf traits of artificial hybrids indicate that the actively moving dune habitat of H. anomalus was more growth limiting, with lower N availability but higher relative water availability than the stabilized dune habitat of H. deserticola. Habitats differed for direct selection on leaf N and WUE, but not size or succulence, for the artificial hybrids. However, within the H. anomalus habitat, direct selection on WUE also differed among populations. Across years, direct selection on leaf traits did not differ. Leaf N was the only trait for which direct selection differed between habitats but not within the H. anomalus habitat, suggesting that nutrient limitation is an important selective force driving adaptation of H. anomalus to the active dune habitat.

UV-induced changes in the skin: can they be repaired?

The damaging effects of UVB light have been described previously and include a number of changes to multiple cell types. At previous meetings of this society, we have shown that Langerhans' cells are the most susceptible to UVB induced damage which can be shown as ultrastructural changes in dendrites, nucleus and cytoplasm by transmission electron microscopy. We have also shown that their patterns of migration from skin to regional lymph node and their ability to present antigens to autologous T cells have been profoundly altered by UVB irradiation. The aim of this work was to establish if it was possible to reverse any of the damage done to Langerhans' cells by UVB exposure by topical application of a DNA repair enzyme such as T4N5 endonuclease. These experiments were undertaken in a sheep model that allowed collection of cells as they migrate from the skin. This allowed for a direct examination of the migration characteristics and ultrastructural features of all Langerhans' cells before, during, and for 2 weeks after exposure to a single dose of UVB. Results obtained from this project indicate that treatment by topical application of DNA repair enzyme immediately after UVB irradiation may restore a number of normal immune parameters associated with the structure and function of migrating Langerhans' cells. It appears that there is a dose related correction of the increased tempo of cell migration and some improvements in the number of ultrastructurally damaged Langerhans' cells have also been associated with application of higher doses of DNA repair enzyme. These preliminary findings indicate that some potential therapeutic benefits are associated with the use of such agents in reversing the immunological damage caused by exposure to erythemal doses of UVB light.

Predawn disequilibrium between plant and soil water potentials in two cold-desert shrubs.

Classical water relations theory predicts that predawn plant water potential should be in equilibrium with soil water potential (soil Ψw) around roots, and many interpretations of plant water status in natural populations are based on this expectation. We examined this expectation for two salt-tolerant, cold-desert shrub species in glasshouse experiments where frequent watering assured homogeneity in soil Ψw and soil-root hydraulic continuity and where NaCl controlled soil Ψw. Plant water potentials were measured with a pressure chamber (xylem Ψp) and thermocouple psychrometers (leaf Ψw). Soil Ψw was measured with in situ thermocouple psychrometers. Predawn leaf Ψw and xylem Ψp were significantly more negative than soil Ψw, for many treatments, indicating large predawn soil-plant Ψw disequilibria: up to 1.2 MPa for Chrysothamnus nauseosus (0 and 100 mM NaCl) and 1.8 MPa for Sarcobatus vermiculatus (0, 100, 300, and 600 mM NaCl). Significant nighttime canopy water loss was one mechanism contributing to predawn disequilibrium, assessed by comparison of xylem Ψp for bagged (to minimize transpiration) and unbagged canopies, and by gas exchange measurements. However, nighttime transpiration accounted for only part of the predawn disequilibrium. Other mechanisms that could act with nighttime transpiration to generate large predawn disequilibria are described and include a model of how leaf apoplastic solutes could contribute to the phenomenon. This study is among the first to conclusively document such large departures from the expectation of predawn soil-plant equilibrium for C3 shrubs, and provides a general framework for considering relative contributions of nighttime transpiration and other plant-related mechanisms to predawn disequilibrium.

Quercus species differ in water and nutrient characteristics in a resource-limited fall-line sandhill habitat.

We compared co-occurring mature Quercus laevis Walt. (turkey oak), Q. margaretta Ashe (sand post oak) and Q. incana Bartr. (bluejack oak) trees growing in resource-limited sandhill habitats of the southeastern United States for water and nutrient characteristics. The Quercus spp. differed in their distribution along soil water and nutrient gradients, and in their access to and use of water, even though the study year was wetter than average with no mid-season drought. Quercus laevis had the greatest access to soil water (least negative pre-dawn water potential, psi(pd)) and the most conservative water-use strategy based on its relatively low stomatal conductance (g(s)), high instantaneous water-use efficiency (WUE), least negative midday water potential (psy(md)) and high leaf specific hydraulic conductance (K(L)). Quercus margaretta had the least conservative water-use characteristics, exhibiting relatively high g(s), low instantaneous WUE, most negative psi(md), and low K(L). Quercus margaretta also had a low photosynthetic nitrogen-use efficiency (PNUE), but a high leaf phosphorus concentration. Quercus incana had the poorest access to soil water, but intermediate water-use characteristics and leaf nutrient characteristics more similar to those of Q. laevis. There were no species differences for photosynthesis (A), leaf nitrogen on an area basis, or seasonally integrated WUE (delta13C). Both A and g(s) were positively correlated for each species, but A and g(s) were generally not correlated with psi(pd), psi(md) or delta psi(pd-md). Although we found differences in resource use and resource status among these sandhill Quercus spp., the results are consistent with the interpretation that they are generally drought avoiders. Quercus laevis may have an advantage on xeric ridges because of its greater ability to access soil water and use it more conservatively compared with the other Quercus spp.

Malignant hyperthermia.

Malignant hyperthermia is a pharmacogenetic disease of skeletal muscle characterized by hypermetabolism that occurs on exposure to a triggering agent or agents. The most common agents are halogenated inhalational anesthetics and succinylcholine, a depolarizing muscle relaxant. Patients who experience malignant hyperthermia are generally transferred to the ICU for ongoing treatment and monitoring for secondary complications of the disorder. Critical care practitioners must be both knowledgeable and competent to prevent and treat perioperative episodes of malignant hyperthermia. A thorough preoperative interview should be done to determine risk factors and susceptible patients. This article provides critical care nurses with sound information on the pathophysiology of malignant hyperthermia, the ability to assess the disease properly and treat the patient both before and after the crisis, and the ability to provide support and teaching to patients and patients' families to prevent the recurrence of malignant hyperthermia.

High apoplastic solute concentrations in leaves alter water relations of the halophytic shrub, Sarcobatus vermiculatus.

Predawn plant water potential (Psi(w)) is used to estimate soil moisture available to plants because plants are expected to equilibrate with the root-zone Psi(w). Although this equilibrium assumption provides the basis for interpreting many physiological and ecological parameters, much work suggests predawn plant Psi(w) is often more negative than root-zone soil Psi(w). For many halophytes even when soils are well-watered and night-time shoot and root water loss eliminated, predawn disequilibrium (PDD) between leaf and soil Psi(w) can exceed 0.5 MPa. A model halophyte, Sarcobatus vermiculatus, was used to test the predictions that low predawn solute potential (Psi(s)) in the leaf apoplast is a major mechanism driving PDD and that low Psi(s) is due to high Na+ and K+ concentrations in the leaf apoplast. Measurements of leaf cell turgor (Psi(p)) and solute potential (Psi(s)) of plants grown under a range of soil salinities demonstrated that predawn symplast Psi(w) was 1.7 to 2.1 MPa more negative than predawn xylem Psi(w), indicating a significant negative apoplastic Psi(s). Measurements on isolated apoplastic fluid indicated that Na+ concentrations in the leaf apoplast ranged from 80 to 230 mM, depending on salinity, while apoplastic K+ remained around 50 mM. The water relations measurements suggest that without a low apoplastic Psi(s), predawn Psi(p) may reach pressures that could cause cell damage. It is proposed that low predawn apoplastic Psi(s) may be an efficient way to regulate Psi(p) in plants that accumulate high concentrations of osmotica or when plants are subject to fluctuating patterns of soil water availability.

Adjustments in hydraulic architecture of Pinus palustris maintain similar stomatal conductance in xeric and mesic habitats.

We investigated relationships between whole-tree hydraulic architecture and stomatal conductance in Pinus palustris Mill. (longleaf pine) across habitats that differed in soil properties and habitat structure. Trees occupying a xeric habitat (characterized by sandy, well-drained soils, higher nitrogen availability and lower overstory tree density) were shorter in stature and had lower sapwood-to-leaf area ratio (A(S):A(L)) than trees in a mesic habitat. The soil-leaf water potential gradient (psiS - psiL) and leaf-specific hydraulic conductance (kL) were similar between sites, as was tissue-specific hydraulic conductivity (Ks) of roots. Leaf and canopy stomatal conductance (gs and Gs, respectively) were also similar between sites, and they tended to be somewhat higher at the xeric site during morning hours when vapour pressure deficit (D) was low. A hydraulic model incorporating tree height, A(S):A(L) and psiS-psiL accurately described the observed variation in individual tree G(Sref) (G(S) at D = 1 kPa) across sites and indicated that tree height was an important determinant of G(Sref) across sites. This, combined with a 42% higher root-to-leaf area ratio (A(R):A(L)) at the xeric site, suggests that xeric site trees are hydraulically well equipped to realize equal--and sometimes higher potential for conductance compared with trees on mesic sites. However, a slightly more sensitive stomatal closure response to increasing D observed in xeric site trees suggests that this potential for higher conductance may only be reached when D is low and when the capacity of the hydraulic system to supply water to foliage is not greatly challenged.

Water potential and ionic effects on germination and seedling growth of two cold desert shrubs.

We tested expectations that two desert shrubs would differ in germination and seedling relative growth rate (RGR) responses to Na and Ψ(s) stress. The study species, Chrysothamnus nauseosus ssp. consimilis and Sarcobatus vermiculatus (hereafter referred to by genus), differ in their distribution along salinity gradients, with Chrysothamnus inhabiting only less saline areas. In growth chamber studies, declining Ψ(s) (-0.82 to -2.71 MPa) inhibited germination of both species, and Chrysothamnus was less tolerant of Ψ(s) stress than Sarcobatus. Germination fell below 10% for Chrysothamnus at -1.64 MPa (NaCl and PEG), and for Sarcobatus at -2.4 MPa PEG. Neither species exhibited ion toxicity. There was substantial ion enhancement for Sarcobatus in lower Ψ(s), allowing for 40% germination in -2.71 MPa NaCl. For seedling RGR, species were not different at -0.29 or -0.82 MPa (0 and 100 mmol/L NaCl, respectively), but Chrysothamnus RGR declined substantially at -1.3 MPa (200 mmol/L NaCl). The greater stress tolerance of Sarcobatus was not associated with a lower RGR under nonsaline conditions. Species differences in seed and seedling Ψ(s) stress tolerance probably contribute to the restricted distribution of Chrysothamnus to less saline areas. The Na uptake of Sarcobatus seedlings enhances its ability to deal with declining Ψ(s) and establish in more saline areas.

Differential desensitization of A1 adenosine receptor-mediated inhibition of cardiac myocyte contractility and adenylate cyclase activity. Relation to the regulation of receptor affinity and density.

Effects of chronic exposure of cultured atrial myocytes to R-N6-(2-phenylisopropyl)-adenosine (R-PIA) on the A1 adenosine receptor-mediated inhibition of adenylate cyclase activity and myocyte contractility were examined. Chronic exposure of atrial myocytes cultured from 14-day-old chick embryos to R-PIA desensitized the myocyte to the inhibitory effects of R-PIA on contractility and adenylate cyclase activity in a time- and dose-dependent manner. Desensitization of the negative inotropic response was only partial, whereas the adenosine receptor-mediated inhibition of adenylate cyclase activity was almost completely absent after 24 hours of R-PIA (1 microM) exposure. Furthermore, the contractile response to R-PIA desensitized more slowly than the desensitization of A1 adenosine receptor-mediated inhibition of adenylate cyclase (t1/2 = 11.4 +/- 0.7 hours versus 7.5 +/- 1 hours, mean +/- SEM, n = 12 and 6, respectively). Thus, the two A1 adenosine receptor-linked functional responses desensitized differently in response to chronic exposure of the myocyte to R-PIA. Binding of the antagonist radioligand [3H]-8-cyclopentyl-1,3-dipropylxanthine [( 3H]CPX) in membranes from myocytes preexposed to R-PIA demonstrated a time-dependent decrease in receptor density without any change in the affinity for the antagonist radioligand. Computer analyses of agonist competition with [3H]CPX binding in membranes from control and R-PIA-treated myocytes revealed a conversion of the high-affinity A1 adenosine receptor to a low-affinity form such that after 24 hours of 1 microM R-PIA exposure, all of the receptors were in a low-affinity form.(ABSTRACT TRUNCATED AT 250 WORDS)

Genetic variation in Pueraria lobata (Fabaceae), an introduced, clonal, invasive plant of the southeastern United States.

Pueraria lobata (kudzu), a clonal, leguminous vine, is invading the southeastern United States at a rate of 50 000 ha per year. Genetic variability and clonal diversity were measured in 20 southeastern U.S. populations using 14 allozyme loci. Within its U.S. range, 92.9% of the loci were polymorphic and overall genetic diversity was 0.290. Such high levels of genetic diversity are consistent with its history of multiple introductions over an extended period of time. The average proportions of polymorphic loci and genetic diversity within populations were 55.7% (range = 28.6-85.7%) and 0.213 (range = 0.114-0.317), respectively. The proportion of total genetic diversity found among populations was similar to species with equivalent life history characters (G(ST) = 0.199). No regional patterns of variation were seen. The number of putative genotypes in each population ranged from 2 to 26. Mean genotypic diversity was 0.694, ranging from 0.223 to 0.955. Such high levels of genotypic diversity indicate that local sites are often colonized by several propagules (most likely seeds) and/or that sexual reproduction occurs within populations after establishment. An excess of heterozygosity was observed in populations with few unique genets, implying that selection for highly heterozygous individuals may occur in populations of P. lobata.

Night-time conductance in C3 and C4 species: do plants lose water at night?

Significant night-time stomatal conductance and transpiration were found for 11 out of 17 species with a range of life histories (herbaceous annual, perennial grass, shrub, tree), photosynthetic pathways (C(3), C(4)), and habitats in the western United States. Across species and habitats, higher night-time conductance and transpiration were associated with higher daytime values. The prevalence, mechanisms and ecological implications of substantial night-time water loss deserve further investigation.

Species-specific patterns of hydraulic lift in co-occurring adult trees and grasses in a sandhill community.

Plants can significantly affect ecosystem water balance by hydraulic redistribution (HR) from dry to wet soil layers via roots (also called hydraulic lift, HL, when the redistribution is from deep to shallow soil). However, the information on how co-occurring species in natural habitats differ in HL ability is insufficient. In a field study, we compared HL ability of four tree species (including three congeneric oak species) and two C4 bunch grass species that co-occur in subxeric habitats of fall-line sandhills in southeastern USA. Soil water potentials (psi(s)) were recorded hourly for 3 years both in large chambers that isolated roots for each species and outside the chambers. Outside of root chambers, soil drying occurred periodically in the top 25 cm and corresponded with lack of precipitation during the summer growing season. Soil moisture was continuously available at a 1 m depth. HL activity was observed in three of the tree species, with greater frequency for Pinus palustris than for Quercus laevis and Q. incana. The fourth tree species Q. margaretta did not exhibit HL activity even though it experienced a similar psi(s) gradient. For the C4 bunch grasses, Aristida stricta exhibited a small amount of HL activity, but Schizachyrium scoparium did not. The capacity for HL activity may be linked to the species ecological distribution. The four species that exhibited HL activity in this subxeric habitat are also dominant in adjacent xeric sandhill habitats, whereas the species that did not exhibit HL are scarcely found in the xeric areas. This is consistent with other studies that found greater fine root survival in dry soil for the four xeric species exhibiting HL activity. The differential ability of these species to redistribute water from the deep soil to the rapidly drying shallow soil likely has a strong effect on the water balance of sandhill plant communities, and is likely linked to their differential distribution across edaphic gradients.

Transgressive character expression in a hybrid sunflower species.

Diploid hybrid lineages often are ecologically distinct from their parental species. However, it is unclear whether this niche divergence is typically achieved via hybrid intermediacy, a mixture of parental traits, and/or the evolution of extreme (transgressive) morphological and ecophysiological features. Here we compare an extensively studied hybrid sunflower species, Helianthus anomalus, with its putative parents, H. annuus and H. petiolaris, for 41 morphological and 12 ecophysiological traits. Helianthus anomalus was morphologically intermediate for one trait (2.4%), parental-like for 23 traits (56.1%), and transgressive for 17 traits (41.5%). For ecophysiological traits, H. anomalus was not significantly different from one or both parents for nine traits (75%), and was transgressive for the remaining three (25%). Thus, H. anomalus appears to be a mosaic of parental-like and transgressive phenotypes. Although the fitness effects of the transgressive characters are not yet known, many of these characters are consistent with adaptations reported for other sand dune plants. Genetic studies are currently underway to ascertain whether these extreme characters arose as a direct byproduct of hybridization or whether they evolved via mutational divergence.

Environment-dependent performance and fitness of Iris brevicaulis, I. fulva (Iridaceae), and hybrids.

We tested the relative fitness of two Louisiana Iris species (Iris brevicaulis and I. fulva) and their first-generation backcross hybrids in three experimental watering treatments: dry, field capacity, and flooded. Leaf area expansion rate, gas exchange (A(max), g(s), c(i)), and biomass at final harvest were measured for each species and hybrid class in all three environmental treatments. Fitness (based on total biomass) of the four genotypic classes differed significantly with environment. All genotypic classes performed most poorly in the dry treatment. The fitness ranking of genotypic class also changed across environments (significant genotypic class by treatment interaction) with hybrid genotype fitness shifting relative to parental genotypes. Integrating over all treatments, backcrosses to I. fulva showed the lowest fitness, whereas backcrosses to I. brevicaulis outperformed I. fulva. The differences in fitness were apparently achieved by a combination of differences in photosynthesis and allocation. In this system, hybrids are not necessarily less fit than their parents, and the relationship between hybrid and parental fitness is influenced by environmental conditions, lending support to the Hybrid Novelty model of hybrid zone evolution.

Functional characterization of type IV pili expressed on diarrhea-associated isolates of Aeromonas species.

Our past work has shown that long, flexible type IV pili (single or in bundles) are the predominant pili expressed on fecal isolates of diarrhea-associated species of Aeromonas (Aeromonas veronii biovar sobria and A. caviae). They represent a family of type IV pili which we have designated Bfp (for bundle-forming pili). Reports from Japan suggest that Bfp are intestinal colonization factors. This study presents compelling evidence to support this conclusion. Aeromonas bacteria and/or Bfp purified from a strain of A. veronii biovar sobria were shown to adhere to epithelial and intestinal cell lines, freshly isolated human enterocytes, and fresh and fixed human and rabbit intestinal tissues, as determined by light and electron microscopy and immunohistochemical detection. Removal of Bfp by mechanical means decreased adhesion to cell lines by up to 80%. Purified Bfp blocked adhesion of the test strain to intestinal cells in a dose-dependent manner. Adhesion was also blocked by the Fab fraction of anti-Bfp immunoglobulin G. Moreover, ultrastructural studies (ruthenium red staining and transmission and scanning electron microscopy) demonstrated for the first time that Aeromonas adhesion to human enterocytes is pilus mediated and suggested that Bfp may also promote colonization by forming bacterium-to-bacterium linkages. Bfp-positive isolates examined for type IV pilus-mediated twitching motility in agar and slide culture assays developed for Pseudomonas aeruginosa did not, however, exhibit this function.