The adaptation of enzymes to temperature: catalytic characterization of glucosephosphate isomerase homologues isolated from Mytilus edulis and Isognomon alatus, bivalve molluscs inhabiting different thermal environments

Homologues of glucosephosphate isomerase (GPI, EC 5.3.1.9) were purified to homogeneity and kinetically characterized from Mytilus edulis and Isognomon alatus, two bivalve molluscs experiencing contrasting thermal environments. The enzyme isolated from I. alatus functions at warmer temperatures (25-35 C) than GPI from M. edulis, a species that inhabits colder marine littoral habitats (5-20 C). The former exhibits apparent first-order (with respect to substrate) catalytic rate constants (Vmax/KM) in vitro that become progressively greater than the mussel enzyme as the assay temperature is raised. Apparent zero-order catalytic rate constants (Vmax) are relatively less differentiated. Catalytic efficiency, defined as the rate at which a catalytic event occurs in either reaction direction for reference standard states (substrate concentrations), is greater for the enzyme from the tropical species (I. alatus) at all realistic combinations of temperature and substrate concentration except for the lowest temperatures and highest substrate concentrations, where the GPI from the boreal/temperate M. edulis is more efficient. This pattern of catalytic divergence appears to be due primarily to differentiation in Vmax/KM. These results and other published data are reviewed and shown to be inconsistent with claims that adaptation of enzymes to higher cell temperatures requires a loss in catalytic efficiency.      

Comparative cytotoxicity of fumonisin B1 in two cell lines derived from normal human bronchial epithelial cells using four distinct bioassay techniques

This study focuses on the cytotoxic effects of fumonisin B₁ (FB₁) on both immortalised and immortalised and subsequently transfected normal human bronchial epithelial (NHBE) cells of human origin using four bioassays. While the MTT, Neutral Red and hexosaminidase colorimetric assays showed little difference between the toxic effects on the two related cell lines, the clonogenic assay, measuring cell survival and proliferation, indicated that FB₁ had a more toxic effect on the nontransfected cells. This kind ofin vitro approach using cells which retain many characteristics of normal cell growth and differentiation can go some way to developing evaluation models for food safety in the case of mycotoxin contamination without resorting totally to whole animal testing. Nevertheless, one or two cytotoxicity tests may be inadequate for a complete appraisal of toxic potential: rather, as wide a range of methodologies as feasible should be employed initially before meaningful conclusions may be drawn.     

Phylogeographic differentiation in the mitochondrial control region in the koala, Phascolarctos cinereus (Goldfuss 1817)

The koala, Phascolarctos cinereus, is a geographically widespread species endemic to Australia, with three currently recognized subspecies: P.c. adustus, P.c. cinereus, and P.c. victor. Intraspecific variation in the mitochondrial DNA (mtDNA) control region was examined in over 200 animals from 16 representative populations throughout the species’ range. Eighteen different haplotypes were defined in the ≈ 860 bp mtDNA control region, as determined by heteroduplex analysis/temperature gradient gel electrophoresis (HDA/TGGE). Any single population typically possessed only one or two haplotypes yielding an average within-population haplotypic diversity of 0.180 ± 0.003, and nucleotide diversity of 0.16%. Overall, mtDNA control region sequence diversity between populations averaged 0.67%, and ranged from 0% to 1.56%. Nucleotide divergence between populations averaged 0.51%, and ranged from 0% to 1.53%. Neighbour-joining methods revealed limited phylogenetic distinction between geographically distant populations of koalas, and tentative support for a single evolutionarily significant unit (ESU). This is consistent with previous suggestions that the morphological differences formalized by subspecific taxonomy may be interpreted as clinal variation. Significant differentiation in mtDNA-haplotype frequencies between localities suggested that little gene flow currently exists among populations. When combined with microsatellite analysis, which has revealed substantial differentiation among koala populations, we conclude that the appropriate short-term management unit (MU) for koalas is the local population.     

Gray matter imaging in multiple sclerosis: what have we learned?

At the early onset of the 20^th century, several studies already reported that the gray matter was implicated in the histopathology of multiple sclerosis (MS). However, as white matter pathology long received predominant attention in this disease, and histological staining techniques for detecting myelin in the gray matter were suboptimal, it was not until the beginning of the 21^st century that the true extent and importance of gray matter pathology in MS was finally recognized. Gray matter damage was shown to be frequent and extensive, and more pronounced in the progressive disease phases. Several studies subsequently demonstrated that the histopathology of gray matter lesions differs from that of white matter lesions. Unfortunately, imaging of pathology in gray matter structures proved to be difficult, especially when using conventional magnetic resonance imaging (MRI) techniques. However, with the recent introduction of several more advanced MRI techniques, the detection of cortical and subcortical damage in MS has considerably improved. This has important consequences for studying the clinical correlates of gray matter damage. In this review, we provide an overview of what has been learned about imaging of gray matter damage in MS, and offer a brief perspective with regards to future developments in this field.     

Detection of Xanthomonas fragariae and presumptive detection of Xanthomonas arboricola pv. fragariae, from strawberry leaves, by real-time PCR

Real-time (TaqMan) PCR assays were developed to detect the strawberry angular leaf spot pathogen Xanthomonas fragariae (Xf) and the strawberry bacterial blight pathogen Xanthomonas arboricola pv. fragariae (Xaf). The Xf PCR (Xf gyrB) was designed within regions of the gyraseB gene, unique to Xf, after generating gyraseB DNA sequence data from Xf and other closely related strains. The Xaf PCR (Xaf pep) was designed within regions of the pep prolyl endopeptidase gene that were unique to Xaf, after generating pep DNA sequence data from Xf and Xaf strains. The Xf gyrB PCR detected only Xf strains amongst a panel of 20 Xanthomonas-related spp. and pathovars. The Xaf pep PCR assay detected all Xaf strains tested plus two other (of three tested) X. arboricola pathovars. An existing genomic DNA extraction protocol was modified to facilitate detection of both pathogens to 10(3) cells per strawberry leaf disc.     

Lifestyle modifications to prevent and control hypertension. 1. Methods and an overview of the Canadian recommendations. Canadian Hypertension Society,Canadian Coalition for High Blood Pressure Prevention and Control,Laboratory Centre for Disease Control at Health Canada,Heart and Stroke Foundation of Canada

OBJECTIVE: To provide updated, evidence-based recommendations for health care professionals on lifestyle changes to prevent and control hypertension in otherwise healthy adults (except pregnant women). OPTIONS: For people at risk for hypertension, there are a number of lifestyle options that may avert the condition--maintaining a healthy body weight, moderating consumption of alcohol, exercising, reducing sodium intake, altering intake of calcium, magnesium and potassium, and reducing stress. Following these options will maintain or reduce the risk of hypertension. For people who already have hypertension, the options for controlling the condition are lifestyle modification, antihypertensive medications or a combination of these options; with no treatment, these people remain at risk for the complications of hypertension. OUTCOMES: The health outcomes considered were changes in blood pressure and in morbidity and mortality rates. Because of insufficient evidence, no economic outcomes were considered. EVIDENCE: A MEDLINE search was conducted for the period January 1996 to September 1996 for each of the interventions studied. Reference lists were scanned, experts were polled, and the personal files of the authors were used to identify other studies. All relevant articles were reviewed, classified according to study design and graded according to level of evidence. VALUES: A high value was placed on the avoidance of cardiovascular morbidity and premature death caused by untreated hypertension. BENEFITS, HARMS AND COSTS: Lifestyle modification by means of weight loss (or maintenance of healthy body weight), regular exercise and low alcohol consumption will reduce the blood pressure of appropriately selected normotensive and hypertensive people. Sodium restriction and stress management will reduce the blood pressure of appropriately selected hypertensive patients. The side effects of these therapies are few, and the indirect benefits are well known. There are certainly costs associated with lifestyle modification, but they were not measured in the studies reviewed. Supplementing the diet with potassium, calcium and magnesium has not been associated with a clinically important reduction in blood pressure in people consuming a healthy diet. RECOMMENDATIONS: (1) It is recommended that health care professionals determine the body mass index (weight in kilograms/[height in metres]2) and alcohol consumption of all adult patients and assess sodium consumption and stress levels in all hypertensive patients. (2) To reduce blood pressure in the population at large, it is recommended that Canadians attain and maintain a healthy body mass index. For those who choose to drink alcohol intake should be limited to 2 or fewer standard drinks per day (maximum of 14/week for men and 9/week for women). Adults should exercise regularly. (3) To reduce blood pressure in hypertensive patients, individualized therapy is recommended. This therapy should emphasize weight loss for overweight patients, abstinence from or moderation in alcohol intake, regular exercise, restriction of sodium intake and, in appropriate circumstances, individualized cognitive behaviour modification to reduce the negative effects of stress. VALIDATION: The recommendations were reviewed by all of the sponsoring organizations and by participants in a satellite symposium of the fourth international Conference on Preventive Cardiology. They are similar to those of the World Hypertension League and the Joint National committee, with the exception of the recommendations on stress management, which are based on new information. They have not been clinically tested. SPONSORS: The Canadian Hypertension Society, the Canadian Coalition for High Blood Pressure Prevention and Control, the Laboratory Centre for Disease Control at health Canada, and the Heart and Stroke Foundation of Canada.     

Lifestyle modifications to prevent and control hypertension. 5. Recommendations on dietary salt. Canadian Hypertension Society,Canadian Coalition for High Blood Pressure Prevention and Control,Laboratory Centre for Disease Control at Health Canada,Heart and Stroke Foundation of Canada

OBJECTIVE: To provide updated, evidence-based recommendations concerning the effects of dietary salt intake on the prevention and control of hypertension in adults (except pregnant women). The guidelines are intended for use in clinical practice and public education campaigns. OPTIONS: Restriction of dietary salt intake may be an alternative to antihypertensive medications or may supplement such medications. Other options include other nonpharmacologic treatments for hypertension and no treatment. OUTCOMES: The health outcomes considered were changes in blood pressure and in morbidity and mortality rates. Because of insufficient evidence, no economic outcomes were considered. EVIDENCE: A MEDLINE search was conducted for the period 1966-1996 using the terms hypertension, blood pressure, vascular resistance, sodium chloride, sodium, diet, sodium or sodium chloride dietary, sodium restricted/reducing diet, clinical trials, controlled clinical trial, randomized controlled trial and random allocation. Both trials and review articles were obtained, and other relevant evidence was obtained from the reference lists of the articles identified, from the personal files of the authors and through contacts with experts. The articles were reviewed, classified according to study design and graded according to level of evidence. In addition, a systematic review of all published randomized controlled trials relating to dietary salt intake and hypertension was conducted. VALUES: A high value was placed on the avoidance of cardiovascular morbidity and premature death caused by untreated hypertension. BENEFITS, HARMS AND COSTS: For normotensive people, a marked change in sodium intake is required to achieve a modest reduction in blood pressure (there is a decrease of 1 mm Hg in systolic blood pressure for every 100 mmol decrease in daily sodium intake). For hypertensive patients, the effects of dietary salt restriction are most pronounced if age is greater than 44 years. A decrease of 6.3 mm Hg in systolic blood pressure and 2.2 mm Hg in diastolic blood pressure per 100 mmol decrease in daily sodium intake was observed in people of this age group. For hypertensive patients 44 years of age and younger, the decreases were 2.4 mm Hg for systolic blood pressure and negligible for diastolic blood pressure. A diet in which salt is moderately restricted appears not to be associated with health risks. RECOMMENDATIONS: (1) Restriction of salt intake for the normotensive population is not recommended at present, because of insufficient evidence demonstrating that this would lead to a reduced incidence of hypertension. (2) To avoid excessive intake of salt, people should be counselled to choose foods low in salt (e.g., fresh fruits and vegetables), to avoid foods high in salt (e.g., pre-prepared foods), to refrain from adding salt at the table and minimize the amount of salt used in cooking, and to increase awareness of the salt content of food choices in restaurants. (3) For hypertensive patients, particularly those over the age of 44 years, it is recommended that the intake of dietary sodium be moderately restricted, to a target range of 90-130 mmol per day (which corresponds to 3-7 g of salt per day). (4) The salt consumption of hypertensive patients should be determined by interview. VALIDATION: These recommendations were reviewed by all of the sponsoring organizations and by participants in a satellite symposium of the fourth International Conference on Preventive Cardiology. They have not been clinically tested. SPONSORS: The Canadian Hypertension Society, the Canadian Coalition for High Blood Pressure Prevention and Control, the Laboratory Centre for Disease Control at Health Canada, and the Heart and Stroke Foundation of Canada.