Small reactive carbonyl compounds as tissue lipid oxidation products; and the mechanisms of their formation thereby

Small reactive carbonyl compounds (RCCs) such as formaldehyde, acetaldehyde, acrolein, crotonaldehyde, glyoxal, methylglyoxal, glycolaldehyde, glycidaldehyde, and 2-butene-1,4-dial are involved in carbonyl and oxidative stress-related physiological disorders. While some evidence indicates that lipid oxidation could be an important source of these compounds in vivo, this has sometimes been doubted because the mechanisms of their formation thereby are poorly understood. Here, representative literature supporting the significant formation of these compounds during lipid oxidation under physiologically relevant conditions are highlighted, and the strengths and weaknesses of previously proposed mechanisms of their formation thereby are considered. In addition, based on the current understanding of lipid oxidation chemistry, some new pathways of their formation are suggested. The suggested pathways also generate 4-hydroxy-2-butenal, a precursor of the carcinogen furan, whose endogenous formation in tissues has hitherto not been seriously considered.     

mtDNA diversity in rhesus monkeys reveals overestimates of divergence time and paraphyly with neighboring species

Reconstructions of the human-African great ape phylogeny by using mitochondrial DNA (mtDNA) have been subject to considerable debate. One confounding factor may be the lack of data on intraspecific variation. To test this hypothesis, we examined the effect of intraspecific mtDNA diversity on the phylogenetic reconstruction of another Plio- Pleistocene radiation of higher primates, the fascicularis group of macaque (Macaca) monkey species. Fifteen endonucleases were used to identify 10 haplotypes of 40-47 restriction sites in M. mulatta, which were compared with similar data for the other members of this species group. Interpopulational, intraspecific mtDNA diversity was large (0.5%- 4.5%), and estimates of divergence time and branching order incorporating this variation were substantially different from those based on single representatives of each species. We conclude that intraspecific mtDNA diversity is substantial in at least some primate species. Consequently, without prior information on the extent of genetic diversity within a particular species, intraspecific variation must be assessed and accounted for when reconstructing primate phylogenies. Further, we question the reliability of hominoid mtDNA phylogenies, based as they are on one or a few representatives of each species, in an already depauperate superfamily of primates.      

Phenotypic Variants of Staphylococci and Their Underlying Population Distributions Following Exposure to Stress

This study investigated whether alterations in environmental conditions would induce the formation of small colony variant phenotypes (SCV) with associated changes in cell morphology and ultra-structure in S. aureus, s. epidermidis, and S. lugdunensis. Wild-type clinical isolates were exposed to low temperature (4°C), antibiotic stress (penicillin G and vancomycin; 0-10,000 µg mL^-1), pH stress (pH 3-9) and osmotic challenge (NaCl concentrations of 0-20%). Changes in cell diameter, cell-wall thickness, and population distribution changes (n ≥ 300) were assessed via scanning and transmission electron microscopy (SEM and TEM), and compared to control populations. Our analyses found that prolonged exposure to all treatments resulted in the subsequent formation of SCV phenotypes. Observed SCVs manifested as minute colonies with reduced haemolysis and pigmentation (NaCl, pH and 4°C treatments), or complete lack thereof (antibiotic treatments). SEM comparison analyses revealed significantly smaller cell sizes for SCV populations except in S. aureus and S. epidermidis 10% NaCl, and S. epidermidis 4°C (p<0.05). Shifts in population distribution patterns were also observed with distinct sub-populations of smaller cells appearing for S. epidermidis, and S. lugdunensis. TEM analyses revealed significantly thicker cell-walls in all treatments and species except S. lugdunensis exposed to 4°C. These findings suggest that staphylococci adapted to environmental stresses by altering their cell size and wall thickness which could represent the formation of altered phenotypes which facilitate survival under harsh conditions. The phenotypic response was governed by the type of prevailing environmental stress regime leading to appropriate alterations in ultra-structure and size, suggesting downstream changes in gene expression, the proteome, and metabolome.     

Demography of a population with a long history of poaching and the utility of the individual identification technique as a tool for monitoring intermittently studied elephant populations

In many elephant populations in Africa, adverse effects of poaching including altered age and sex structure and long calving intervals continue to negatively impact the rate of recovery. The study reported here characterized the demographic status of the elephant population in Meru National Park, Kenya, so as to determine the size, age and sex structure, and calving interval of the population and to compare its demographic parameters against those from the relatively stable elephant population at Amboseli. Additionally, a demographic study conducted 7 years earlier enabled us to determine the demographic performance of the population and to explicitly test the utility of the individual identification technique in monitoring intermittently studied populations. We found 392 elephants in the park. The proportion of older individuals was lower, and the calving interval was longer than estimates from Amboseli. We recognized 16% of elephants indexed in the previous study. Our results suggest that the individual identification technique may be useful in monitoring demographic changes including poaching in intermittently studied elephant populations. Overall, our results suggest that although the Meru elephant population showed signs of recovery, poaching continues to imperil its recovery.     

An analysis of haematological parameters in patients and individual residents of a Plasmodium falciparum malaria holoendemic area of western Kenya

Many clinical symptoms of malaria are associated with alterations in certain haematological parameters during acute and subclinical infections. Total leucocyte and erythrocyte counts, haemoglobin concentration, haematocrit and other minor indices, were investigated in five cohort groups of individuals resident in a malaria hyperendemic area of western Kenya. The groups included age- and sex-matched adults with acute Plasmodium falciparum malaria, aparasitaemic adults, children with acute malaria, aparasitaemic children and asymptomatic-parasitaemic school children. The study aimed at defining what constitutes immunity to malaria which may be important in the critical evaluation of malaria vaccine antigens. Anaemia was more severe in adults and children with acute malaria than in their age- and sex-matched aparasitaemic and asymptomatic-aparasitaemic school children. Lymphocyte counts were significantly higher in asymptomatic-aparasitaemic school children than in aparasitaemic adults, suggesting a possible functional role for lymphocytes in the anti-disease immunity in the former group.     

Serosurvey of Brucella spp. infection in the Kafue lechwe (Kobus leche kafuensis) of the Kafue flats in Zambia

One of the diseases of veterinary and public health importance affecting the Kafue lechwe (Kobus leche kafuensis) on the Kafue flats is brucellosis, for which only scant information is available. During the 2003 (October), 2004 (December), and 2008 (July-December) hunting seasons in the Kafue flats, we conducted a study to determine the seroprevalence of Brucella spp. in the Kafue lechwe and to evaluate serologic tests for detection of Brucella spp. antibodies in lechwe. The Rose Bengal Test (RBT), competitive enzyme-linked immunosorbent assay (cELISA), and fluorescence polarization assay (FPA) were used. A total of 121 Kafue lechwe were hunted for disease investigations in 2003, 2004, and 2008 in the Kafue Flat Game Management Area. Of these, 21.6%, (95% confidence interval [CI]: 14.2-29.1%) had detectable antibodies to Brucella spp. The Kafue lechwe in Lochnivar National Park had higher antibody results than those in Blue Lagoon National Park (odds ratio=3.0; 95% CI: 0.94-9.4). Infection levels were similar in females (21.6%) and males (21.7%). Results were similar among RBT, FPA, cELISA tests, suggesting that these could effectively be used in diagnosing brucellosis in the Kafue lechwe. Our study demonstrates the presence of Brucella infections in the Kafue lechwe in two national parks located in the Kafue flats and further highlights the suitability of serologic assays for testing the Kafue lechwe. Because the Kafue lechwe is the most hunted wildlife species in Zambia, hunters need to be informed of the public health risk of Brucella spp. infection.     

Dynamic moisture sorption characteristics of enzyme-resistant recrystallized cassava starch

The interaction of moisture with enzyme-resistant recrystallized starch, prepared by heat-moisture treatment of debranched acid-modified or debranched non-acid-modified cassava starch, was investigated in comparison with the native granules. Crystallinities of the powdered products were estimated by X-ray diffraction. Moisture sorption was determined using dynamic vapor sorption analyzer and data fitted to various models. Percent crystallinities of native starch (NS), non-acid-modified recrystallized starch (NAMRS), and acid-modified recrystallized starch (AMRS) were 39.7, 51.9, and 56.1%, respectively. In a(w) below 0.8, sorption decreased in the order NS > NAMRS > AMRS in line with increasing sample crystallinities but did not follow this crystallinity dependence at higher a(w) because of condensation and polymer dissolution effects. Adsorbed moisture became internally absorbed in NS but not in NAMRS and AMRS, which might explain the high resistance of the recrystallized starches to digestion because enzyme and starch cannot approach each other over fairly sufficient surface at the molecular level.     

Oleanane-type triterpenes of Embelia schimperi leaves

An investigation of an ethyl acetate extract of Embelia schimperi leaves has led to the isolation of 10 oleanane-type triterpenes characterized as 3beta,16alpha-di-O-acetyl-13beta, 28-epoxyoleanane (1), 3beta-acetyl-16-oxo-13beta, 28-epoxyoleanane (2), 3beta-acetyl-16alpha-hydroxy-13beta, 28-epoxyoleanane (3), 3beta-acetyl-16alpha-hydroxyoleanane-13beta, 28-olide (4), 3beta-acetyl-28-hydroxy-16-oxo-12-oleanene (5), 3beta, 28-di-O-acetyl-16alpha-hydroxy-12-oleanene (6), 3beta-acetyl-11alpha, 28-dihydroxy-16-oxo-12-oleanene (7), 3beta, 11alpha, 16alpha, 28-tetrahydroxy-12-oleanene (8), 3beta-acetyl-16alpha, 28alpha-dihydroxy-13beta, 28-oxydooleanane (9) and 3beta, 28alpha-dihydroxy-16-oxo-13beta, 28-oxydooleanane (10). The known compounds isolated from the same extract included 3beta, 16alpha-dihydroxy-13beta, 28-epoxyoleanane (protoprimulagenin A) (11), 3beta-hydroxy-16-oxo-13beta, 28-epoxyoxyoleanane (aegicerin) (12), 3, 16-dioxo-13beta, 28-epoxyoleanane (embilionone) (13), 3beta, 28-dihydroxy-16-oxo-12-oleanene (schimperinone) (14), taraxerone (15), taraxerol (16) and stigmasterol (17). Structure elucidations were carried out spectroscopically.     

Multiplexed expression and screening for recombinant protein production in mammalian cells

Background  

A variety of approaches to understanding protein structure and function require production of recombinant protein. Mammalian based expression systems have advantages over bacterial systems for certain classes of protein but can be slower and more laborious. Thus the availability of a simple system for production and rapid screening of constructs or conditions for mammalian expression would be of great benefit. To this end we have coupled an efficient recombinant protein production system based on transient transfection in HEK-293 EBNA1 (HEK-293E) suspension cells with a dot blot method allowing pre-screening of proteins expressed in cells in a high throughput manner.