The vasorelaxation of cerebral arteries by carbon monoxide

Carbon monoxide (CO) is known to increase cerebral blood flow, but the effect of CO on the vascular tone of large cerebral arteries is uncertain. We tested whether CO affects cerebral artery tone by measuring tension generated by ex vivo segments of dog basilar artery upon exposure to CO. In cerebral artery segments contracted with either KCl or prostaglandin F(2alpha), CO caused a concentration-related relaxation beginning with a concentration of 57 microM. Relaxation did not occur if CO was administered in the presence of bubbling carboxygen (95% O(2):5% CO(2)), which reduces greater than 99% of CO from the solution. Furthermore, the CO-induced relaxation of cerebral artery segments was reduced in the presence of the guanylyl cyclase inhibitor 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ, 10 microM)or the potassium channel blocker tetraethylammonium (TEA, 1 mM). Neither ODQ nor TEA completely eliminated the relaxation caused by CO and there was no additive effect if ODQ and TEA were administered together. These results suggest that cerebral arteries are directly relaxed by CO and that this relaxation depends upon the activation of guanylyl cyclase and the opening of potassium channels.     

Scaled quantum mechanical force field for alanyl-alanine peptide in solution

We have obtained ab initio scale factors and assigned frequencies for the alanine-alanine peptide in water. Calculations were performed on the isolated acidic and basic Ala-Ala structures, two one-water basic Ala-Ala supermolecules, and one two-water acidic and one two-water basic Ala-Ala supermolecules. Force constants were scaled using the experimentally determined Raman and Fourier transform infrared vibrational frequencies of four isotopic species of Ala-Ala in water at pH 13 and pH 1. Most of the 4-31G scale factors were transferable from smaller molecules. All but one scale factor were directly transferable between the pH 1 and pH 13 species for coordinates unchanged by protonation in both the isolated and two-water supermolecule structures. Scale factors for nonpolar coordinates were transferable between all Ala-Ala species with only a few small changes. Good agreement was obtained between the calculated and experimental frequencies for all isotopic species and structures.     

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.     

DO FRESHWATER FISHES DIVERSIFY FASTER THAN MARINE FISHES? A TEST USING STATE‐DEPENDENT DIVERSIFICATION ANALYSES AND MOLECULAR PHYLOGENETICS OF NEW WORLD SILVERSIDES (ATHERINOPSIDAE)

Freshwater habitats make up only ～0.01% of available aquatic habitat and yet harbor 40% of all fish species, whereas marine habitats comprise >99% of available aquatic habitat and have only 60% of fish species. One possible explanation for this pattern is that diversification rates are higher in freshwater habitats than in marine habitats. We investigated diversification in marine and freshwater lineages in the New World silverside fish clade Menidiinae (Teleostei, Atherinopsidae). Using a time‐calibrated phylogeny and a state‐dependent speciation–extinction framework, we determined the frequency and timing of habitat transitions in Menidiinae and tested for differences in diversification parameters between marine and freshwater lineages. We found that Menidiinae is an ancestrally marine lineage that independently colonized freshwater habitats four times followed by three reversals to the marine environment. Our state‐dependent diversification analyses showed that freshwater lineages have higher speciation and extinction rates than marine lineages. Net diversification rates were higher (but not significant) in freshwater than marine environments. The marine lineage‐through time (LTT) plot shows constant accumulation, suggesting that ecological limits to clade growth have not slowed diversification in marine lineages. Freshwater lineages exhibited an upturn near the recent in their LTT plot, which is consistent with our estimates of high background extinction rates. All sequence data are currently being archived on Genbank and phylogenetic trees archived on Treebase.     

DIVERSITY‐DEPENDENT CLADOGENESIS AND TRAIT EVOLUTION IN THE ADAPTIVE RADIATION OF THE AUKS (AVES: ALCIDAE)

Through the course of an adaptive radiation, the evolutionary speed of cladogenesis and ecologically relevant trait evolution are expected to slow as species diversity increases, niches become occupied, and ecological opportunity declines. We develop new likelihood‐based models to test diversity‐dependent evolution in the auks, one of only a few families of seabirds adapted to underwater “flight,” and which exhibit a large variety of bill sizes and shapes. Consistent with the expectations of adaptive radiation, we find both a decline in rates of cladogenesis (a sixfold decline) and bill shape (a 64‐fold decline) evolution as diversity increased. Bill shape diverged into two clades at the basal cladogenesis event with one clade possessing mostly long, narrow bills used to forage primarily on fish, and the other with short thick bills used to forage primarily on plankton. Following this initial divergence in bill shape, size, a known correlate of both prey size and maximum diving depth, diverged rapidly within each of these clades. These results suggest that adaptive radiation in foraging traits underwent initial divergence in bill shape to occupy different food resources, followed by size differentiation to subdivide each niche along the depth axis of the water column.     

Development of a multiplex PCR assay for simultaneous detection of Theileria annulata,Babesia bovis and Anaplasma marginale in cattle

Tropical theileriosis, bovine babesiosis and anaplasmosis are tick-borne protozoan diseases that impose serious constraints on the health and productivity of domestic cattle in tropical and sub-tropical regions of the world. A common feature of these diseases is that, following recovery from primary infection, animals become persistent carriers of the pathogen and continue to play a critical role in disease epidemiology, acting as reservoirs of infection. This study describes development and evaluation of multiplex and single PCR assays for simultaneous detection of Theileria annulata, Babesia bovis and Anaplasma marginale in cattle. Following in silico screening for candidate target genes representing each of the pathogens, an optimised multiplex PCR assay was established using three primer sets, cytob1, MAR1bB2 and bovar2A, for amplification of genomic DNA of T. annulata, A. marginale and B. bovis respectively. The designed primer sets were found to be species-specific, generating amplicons of 312, 265 and 166 base pairs, respectively and were deemed suitable for the development of a multiplex assay. The sensitivity of each primer pair was evaluated using serial dilutions of parasite DNA, while specificity was confirmed by testing for amplification from DNA of different stocks of each pathogen and other Theileria, Babesia and Anaplasma species. Additionally, DNA preparations derived from field samples were used to evaluate the utility of the single and multiplex PCRs for determination of infection status. The multiplex PCR was found to detect each pathogen species with the same level of sensitivity, irrespective of whether its DNA was amplified in isolation or together with DNA representing the other pathogens. Moreover, single and multiplex PCRs were able to detect each species with equal sensitivity in serially diluted DNA representing mixtures of T. annulata, B. bovis and A. marginale, and no evidence of non-specific amplification from non-target species was observed. Validation that the multiplex PCR efficiently detects single and mixed infections from field samples was demonstrated. The developed assay represents a simple and efficient diagnostic for co-detection of tropical theileriosis, bovine babesiosis and anaplasmosis, and may be a valuable tool for epidemiological studies aimed at assessing the burden of multiple infection with tick-borne pathogens and improving control of the associated diseases in endemic regions.     

Whole-Genome Sequencing of Theileria parva Strains Provides Insight into Parasite Migration and Diversification in the African Continent

The disease caused by the apicomplexan protozoan parasite Theileria parva, known as East Coast fever or Corridor disease, is one of the most serious cattle diseases in Eastern, Central, and Southern Africa. We performed whole-genome sequencing of nine T. parva strains, including one of the vaccine strains (Kiambu 5), field isolates from Zambia, Uganda, Tanzania, or Rwanda, and two buffalo-derived strains. Comparison with the reference Muguga genome sequence revealed 34 814–121 545 single nucleotide polymorphisms (SNPs) that were more abundant in buffalo-derived strains. High-resolution phylogenetic trees were constructed with selected informative SNPs that allowed the investigation of possible complex recombination events among ancestors of the extant strains. We further analysed the dN/dS ratio (non-synonymous substitutions per non-synonymous site divided by synonymous substitutions per synonymous site) for 4011 coding genes to estimate potential selective pressure. Genes under possible positive selection were identified that may, in turn, assist in the identification of immunogenic proteins or vaccine candidates. This study elucidated the phylogeny of T. parva strains based on genome-wide SNPs analysis with prediction of possible past recombination events, providing insight into the migration, diversification, and evolution of this parasite species in the African continent.     

Postprandial metabolism of docosapentaenoic acid (DPA, 22:5n−3) and eicosapentaenoic acid (EPA, 20:5n−3) in humans

The study of the metabolism of docosapentaenoic acid (DPA, 22:5n?3) in humans has been limited by the unavailability of pure DPA and the fact that DPA is found in combination with eicosapentaenoic acid (EPA, 20:5n?3) and docosahexaenoic acid (DHA, 22:6n?3) in natural products. In this double blind cross over study, pure DPA and EPA were incorporated in meals served to healthy female volunteers. Mass spectrometric methods were used to study the chylomicron lipidomics. Plasma chylomicronemia was significantly reduced after the meal containing DPA compared with the meal containing EPA or olive oil only. Both EPA and DPA were incorporated into chylomicron TAGs, while there was less incorporation into chylomicron phospholipids. Lipidomic analysis of the chylomicron TAGs revealed the dynamic nature of chylomicron TAGs. The main TAG species that EPA and DPA were incorporated into were EPA/18:1/18:1, DPA/18:1/16:0 and DPA/18:1/18:1. There was very limited conversion of DPA and EPA to DHA and there were no increases in EPA levels during the 5 h postprandial period after the DPA meal. In conclusion, EPA and DPA showed different metabolic fates, and DPA hindered the digestion, ingestion or incorporation into chylomicrons of the olive oil present in the meal.     

Inclusion of Plasma Lipid Species Improves Classification of Individuals at Risk of Type 2 Diabetes

Background

A significant proportion of individuals with diabetes or impaired glucose tolerance have fasting plasma glucose less than 6.1 mmol/L and so are not identified with fasting plasma glucose measurements. In this study, we sought to evaluate the utility of plasma lipids to improve on fasting plasma glucose and other standard risk factors for the identification of type 2 diabetes or those at increased risk (impaired glucose tolerance).

Methods and Findings

Our diabetes risk classification model was trained and cross-validated on a cohort 76 individuals with undiagnosed diabetes or impaired glucose tolerance and 170 gender and body mass index matched individuals with normal glucose tolerance, all with fasting plasma glucose less than 6.1 mmol/L. The inclusion of 21 individual plasma lipid species to triglycerides and HbA1c as predictors in the diabetes risk classification model resulted in a statistically significant gain in area under the receiver operator characteristic curve of 0.049 (p<0.001) and a net reclassification improvement of 10.5% (p<0.001). The gain in area under the curve and net reclassification improvement were subsequently validated on a separate cohort of 485 subjects.

Conclusions

Plasma lipid species can improve the performance of classification models based on standard lipid and non-lipid risk factors.