Definition of common carotid wall thickness affects risk classification in relation to degree of internal carotid artery stenosis: the Plaque At RISK (PARISK) study

Background

Mean or maximal intima-media thickness (IMT) is commonly used as surrogate endpoint in intervention studies. However, the effect of normalization by surrounding or median IMT or by diameter is unknown. In addition, it is unclear whether IMT inhomogeneity is a useful predictor beyond common wall parameters like maximal wall thickness, either absolute or normalized to IMT or lumen size. We investigated the interrelationship of common carotid artery (CCA) thickness parameters and their association with the ipsilateral internal carotid artery (ICA) stenosis degree.

Methods

CCA thickness parameters were extracted by edge detection applied to ultrasound B-mode recordings of 240 patients. Degree of ICA stenosis was determined from CT angiography.

Results

Normalization of maximal CCA wall thickness to median IMT leads to large variations. Higher CCA thickness parameter values are associated with a higher degree of ipsilateral ICA stenosis (p?<?0.001), though IMT inhomogeneity does not provide extra information. When the ratio of wall thickness and diameter instead of absolute maximal wall thickness is used as risk marker for having moderate ipsilateral ICA stenosis (>50%), 55 arteries (15%) are reclassified to another risk category.

Conclusions

It is more reasonable to normalize maximal wall thickness to end-diastolic diameter rather than to IMT, affecting risk classification and suggesting modification of the Mannheim criteria.

Trial registration

Clinical trials.gov NCT01208025.

     

Biological Control of Sheep Parasites using Duddingtonia flagrans: Trials on Commercial Farms in Sweden

   

Trials were conducted on 3 commercial sheep farms in Sweden to assess the effect of administering spores of the nematode trapping fungus, Duddingtonia flagrans, together with supplementary feed to lactating ewes for the first 6 weeks from turn-out on pastures in spring. Also control groups of ewes, receiving only feed supplement, were established on all 3 farms. Groups were monitored by intensive parasitological investigation. The ewes and their lambs were moved in late June to saved pastures for summer grazing, the lambs receiving an anthelmintic treatment at this time. After approximately 6 weeks on summer pasture the lambs were weaned, treated a second time with anthelmintic, and returned to their original lambing pastures for finishing. Decisions as to when lambs were to be marketed were entirely at the discretion of the farmer co-operators. No difference in lamb performance was found between the two treatments on all three farms. This was attributed to the high levels of nutrition initially of the ewes limiting their post-partum rise in nematode faecal egg counts in spring, which in turn resulted in low levels of nematode infection on pastures throughout the autumn period. Additionally, pastures were of good quality for the lambs during the finishing period, so they grew at optimal rates as far as the farmers were concerned.     

Exchanging the yellow fever virus envelope proteins with Modoc virus prM and E proteins results in a chimeric virus that is neuroinvasive in SCID mice

A chimeric flavivirus infectious cDNA was constructed by exchanging the premembrane (prM) and envelope (E) genes of the yellow fever virus vaccine strain 17D (YF17D) with the corresponding genes of Modoc virus (MOD). This latter virus belongs to the cluster of the "not-known vector" flaviviruses and is, unlike YF17D, neuroinvasive in SCID mice. Replication of in vitro-transcribed RNA from this chimeric flavivirus was shown by [(3)H]uridine labeling and RNA analysis. Expression of the MOD prM and E proteins was monitored by radioimmunoprecipitation and revealed that the MOD proteins were correctly and efficiently produced from the chimeric precursor protein. The MOD E protein was shown to be N-linked glycosylated, whereas prM, as predicted from the genome sequence, did not contain N-linked carbohydrates. In Vero cells, the chimeric virus replicated with a similar efficiency as the parental viruses, although it formed smaller plaques than YF17D and MOD. In SCID mice that had been infected intraperitoneally with the chimeric virus, the viral load increased steadily until death. The MOD/YF virus, like MOD from which it had acquired the prM and E structural proteins, but unlike YF, proved neuroinvasive in SCID mice. Animals developed neurological symptoms about 15 days after inoculation and died shortly thereafter. The distribution of MOD/YF RNA in the brain of infected mice was similar to that observed in MOD-infected mice. The observations provide compelling evidence that the determinants of neuroinvasiveness of flaviviruses are entirely located in the envelope proteins prM and E.     

All subgenomic mRNAs of equine arteritis virus contain a common leader sequence.

During the replication of equine arteritis virus (EAV) six subgenomic mRNAs are synthesized. We present evidence that the viral mRNAs form a 3'-coterminal nested set and contain a common leader sequence of 208 nucleotides which is encoded by the 5'-end of the genome. The leader is joined to the bodies of mRNA 5 and 6 at positions defined by the sequence 5' UCAAC 3'. The part of the leader sequence flanking the UCAAC motif is very similar to the 5'-splice site of the Tetrahymena pre-rRNA. A possible internal guide sequence has been identified 43 nucleotides downstream of the leader sequence on the genome. Hybridization analysis shows that all EAV intracellular RNAs contain the leader sequence. These data imply that the viral subgenomic mRNAs are composed of leader and body sequences which are non-contiguous on the genome.     

Pathway analysis in metabolomics: Recommendations for the use of over-representation analysis

Over-representation analysis (ORA) is one of the commonest pathway analysis approaches used for the functional interpretation of metabolomics datasets. Despite the widespread use of ORA in metabolomics, the community lacks guidelines detailing its best-practice use. Many factors have a pronounced impact on the results, but to date their effects have received little systematic attention. Using five publicly available datasets, we demonstrated that changes in parameters such as the background set, differential metabolite selection methods, and pathway database used can result in profoundly different ORA results. The use of a non-assay-specific background set, for example, resulted in large numbers of false-positive pathways. Pathway database choice, evaluated using three of the most popular metabolic pathway databases (KEGG, Reactome, and BioCyc), led to vastly different results in both the number and function of significantly enriched pathways. Factors that are specific to metabolomics data, such as the reliability of compound identification and the chemical bias of different analytical platforms also impacted ORA results. Simulated metabolite misidentification rates as low as 4% resulted in both gain of false-positive pathways and loss of truly significant pathways across all datasets. Our results have several practical implications for ORA users, as well as those using alternative pathway analysis methods. We offer a set of recommendations for the use of ORA in metabolomics, alongside a set of minimal reporting guidelines, as a first step towards the standardisation of pathway analysis in metabolomics.     

Improved analyses of human mtDNA sequences support a recent African origin for Homo sapiens

New quantitative methods are applied to the 135 human mitochondrial sequences from the Vigilant et al. data set. General problems in analyzing large numbers of short sequences are discussed, and an improved strategy is suggested. A key feature is to focus not on individual trees but on the general "landscape" of trees. Over 1,000 searches were made from random starting trees with only one tree (a local optimum) being retained each time, thereby ensuring optima were found independently. A new tree comparison metric was developed that is unaffected by rearrangements of trees around many very short internal edges. Use of this metric showed that downweighting hypervariable sites revealed more evolutionary structure than studies that weighted all sites equally. Our results are consistent with convergence toward a global optimum. Crucial features are that the best optima show very strong regional differentiation, a common group of 49 African sequences is found in all the best optima, and the best optima contain the 16 !Kung sequences in a separate group of San people. The other 86 sequences form a heterogeneous mixture of Africans, Europeans, Australopapuans, and Asians. Thus all major human lineages occur in Africa, but only a subset occurs in the rest of the world. The existence of these African-only groups strongly contradicts multiregional theories for the origin of Homo sapiens that require widespread migration and interbreeding over the entire range of H. erectus. Only when the multiregional model is rejected is it appropriate to consider the root, based on a single locus, to be the center of origin of a population (otherwise different loci could give alternative geographic positions for the root). For this data, several methods locate the root within the group of 49 African sequences and are thus consistent with the recent African origin of H. sapiens. We demonstrate that the time of the last common ancestor cannot be the time of major expansion in human numbers, and our results are thus also consistent with recent models that differentiate between the last common ancestor, expansion out of Africa, and the major expansion in human populations. Such a two-phase model is consistent with a wide range of molecular and archeological evidence.      

A covariotide model explains apparent phylogenetic structure of oxygenic photosynthetic lineages

The aims of the work were (1) to develop statistical tests to identify whether substitution takes place under a covariotide model in sequences used for phylogenetic inference and (2) to determine the influence of covariotide substitution on phylogenetic trees inferred for photosynthetic and other organisms. (Covariotide and covarion models are ones in which sites that are variable in some parts of the underlying tree are invariable in others and vice versa.) Two tests were developed. The first was a contingency test, and the second was an inequality test comparing the expected number of variable sites in two groups with the observed number. Application of these tests to 16S rDNA and tufA sequences from a range of nonphotosynthetic prokaryotes and oxygenic photosynthetic prokaryotes and eukaryotes suggests the occurrence of a covariotide mechanism. The degree of support for partitioning of taxa in reconstructed trees involving these organisms was determined in the presence or absence of sites showing particular substitution patterns. This analysis showed that the support for splits between (1) photosynthetic eukaryotes and prokaryotes and (2) photosynthetic and nonphotosynthetic organisms could be accounted for by patterns arising from covariotide substitution. We show that the additional problem of compositional bias in sequence data needs to be considered in the context of patterns of covariotide/covarion substitution. We argue that while covariotide or covarion substitution may give rise to phylogenetically informative patterns in sequence data, this may not always be so.