Search PubMed⌕ Search

Biomedical subjects

E C Holmes

Publications and source records attributed to E C Holmes.

At least 19 recordsLinked to original sources
PubMed1999

Widespread intra-serotype recombination in natural populations of dengue virus.

Diversity analysis of 71 published dengue virus gene sequences revealed several strains that appeared to be mosaics comprising gene regions with conflicting evolutionary histories. Subsequent maximum likelihood breakpoint estimation identified seven recombinants, including members of three of the four dengue virus serotypes, with breakpoints in the premembrane/membrane gene, the envelope gene, and at the junction of the envelope and first nonstructural genes. Many of the individual recombinants contain sequence representing separate genetic subtypes. The results were highly statistically significant and were confirmed by phylogenetic analysis of the regions of interest. These findings indicate that recombination may play a very significant role in shaping genetic diversity in dengue virus and, as such, have important implications for its biology and its control.

Dengue Virus↗
PubMed1999

Reconstructing the complex evolutionary history of hepatitis B virus.

A detailed analysis of the evolutionary history of hepatitis B virus (HBV) was undertaken using 39 mammalian hepadnaviruses for which complete genome sequences were available, including representatives of all six human genotypes, as well as a large sample of small S gene sequences. Phylogenetic trees of these data were ambiguous, supporting no single place of origin for HBV, and depended heavily on the underlying model of DNA substitution. In some instances genotype F, predominant in the Americas, was the first to diverge, suggesting that the virus arose in the New World. In other trees, however, sequences from genotype B, prevalent in East Asia, were the most divergent. An attempt was also made to determine the rate of nucleotide substitution in the C open reading frame and then to date the origin of HBV. However, no relationship between time and number of substitutions was found in two independent data sets, indicating that a reliable molecular clock does not exist for these data. Both the pattern and the rate of nucleotide substitution are therefore complex phenomena in HBV and hinder any attempt to reconstruct the past spread of this virus.

Americas↗
PubMed1999

Evolutionary history of MHC class I genes in the mammalian order Perissodactyla.

We carried out an analysis of partial sequences from expressed major histocompatibility complex (MHC) class I genes isolated from a range of equid species and more distantly related members of the mammalian order Perissodactyla. Phylogenetic analysis revealed a minimum of six groups, five of which contained genes and alleles that are found in equid species and one group specific to the rhinoceros. Four of the groups contained only one, or very few sequences, indicating the presence of relatively nonpolymorphic loci, while another group contained the majority of the equid sequences identified. These data suggest that a diversification of MHC genes took place after the split between the Equidae and the Rhinocerotidae yet before the speciation events within the genus Equus.

Animals↗
PubMed1999

Amplification and overexpression of the cyclin D1 and epidermal growth factor receptor genes in non-small-cell lung cancer. Lung Cancer Study Group.

PURPOSE: To study the structure and expression of the cyclin D1 and the epidermal growth factor receptor (EGFR) genes in a cohort of 298 non-small-cell lung cancer (NSCLC) specimens. METHODS: Gene structure was studied by Southern analysis, and gene expression was studied by Northern analysis and immunohistochemical analysis. RESULTS: Amplification of the cyclin D1 locus was found in 14/298 (5%) specimens. All 12/12 specimens with amplification of the cyclin D1 gene for which RNA was available were found to express the cyclin D1 transcript, and 11/12 overexpressed the transcript to levels higher than that of uninvolved lung. The EGFR gene was amplified in 17/286 samples of NSCLC tested, and was overexpressed in 22/169 (13%) cases tested, including 12/13 cases with amplification of the gene for which RNA was available. Cyclin D1 gene amplification was associated with advanced lymph node involvement (P = 0.043), but not with larger tumor size or adverse outcome. Cyclin D1 gene amplification and overexpression occurred independently of retinoblastoma tumor-suppressor gene (RB) inactivation, but tumors with amplification of the cyclin D1 gene were more likely to have EGFR gene amplification (P < 0.005). No correlation of EGFR gene amplification or overexpression with tumor size, lymph node involvement, patient demographic data, or survival was identified. CONCLUSIONS: These data indicate that the cyclin D1 and EGFR genes are amplified and overexpressed in NSCLC, and amplification of the cyclin D1 gene occurs frequently in conjunction with amplification of the EGFR gene.

Adenocarcinoma↗
PubMed1999

Biological consequences of deletions within the 3'-untranslated region of flaviviruses may be due to rearrangements of RNA secondary structure.

It was previously reported that deletions introduced into the 3'-untranslated region (3'-UTR) of dengue type 4 (DEN 4) virus (Men, R., Bray, M., Clark, D., Chanock, R.M., Lai, C.J., 1996. DEN 4 virus mutants containing deletions in the 3'-noncoding region of the RNA genome: analysis of growth restriction in cell culture and altered viremia pattern and immunogenicity in Rhesus monkeys. J. Virol. 70, 3930-3937), tick-borne encephalitis (TBE) virus (Mandl, C.W., Holzmann, H., Meixner, T., Rauscher, S., Stadler, P.F., Allison, S.L. , Heinz, F.X., 1998. Spontaneous and engineered deletions in the 3'-noncoding region of TBE virus: construction of highly attenuated mutants of a flavivirus. J. Virol. 72, 2132-2140) and subgenomic replicons of Kunjin virus (Khromykh, A.A., Westaway, E.G., 1997. Subgenomic replicons of the flavivirus Kunjin: construction and applications. J. Virol. 71, 1497-1505) altered the infectivity of the mutants and reduced the efficiency of RNA replication. Here, these deletions were superimposed onto the models of secondary structure we constructed previously and the folding of the modified 3'-UTR sequences was simulated. The analysis showed that most of the deletions disrupted or reshaped conserved elements of secondary structure and that the biological effects of these deletions are likely to represent structural rearrangements in the 3'-UTR, rather than the loss of sequence motifs. The analysis also suggested that the overall structural integrity of the flaviviral 3'-UTR is essential for optimal performance of its promotor function, although two distinct parts can be defined: the most 3'-terminal structures and sequences which may be critical for the initiation of minus-strand RNA synthesis, and more proximal structures and sequences that possibly function as enhancers of viral RNA replication. The functional significance of certain structural elements and their possible effect on the efficiency of viral replication in different cells are also discussed.

3' Untranslated Regions↗
PubMed1999

Population dynamics of HIV-1 inferred from gene sequences.

A method for the estimation of population dynamic history from sequence data is described and used to investigate the past population dynamics of HIV-1 subtypes A and B. Using both gag and env gene alignments the effective population size of each subtype is estimated and found to be surprisingly small. This may be a result of the selective sweep of mutations through the population, or may indicate an important role of genetic drift in the fixation of mutations. The implications of these results for the spread of drug-resistant mutations and transmission dynamics, and also the roles of selection and recombination in shaping HIV-1 genetic diversity, are discussed. A larger estimated effective population size for subtype A may be the result of differences in time of origin, transmission dynamics, and/or population structure. To investigate the importance of population structure a model of population subdivision was fitted to each subtype, although the improvement in likelihood was found to be nonsignificant.

Genes, Viral↗
PubMed1999

Genealogical evidence for positive selection in the nef gene of HIV-1.

The pattern and process of evolution in the nef gene of HIV-1 was analyzed within and among patients. Using a maximum likelihood method that allows for variable intensity of selection pressure among codons, strong positive selection was detected in a hemophiliac patient over 30 mo of infection. By reconstructing the process of allele substitution in this patient using parsimony, the synapomorphic amino acid changes separating each time point were found to have high probabilities of being under positive selection, with selective coefficients of at least 3.6%. Positive selection was also detected among 39 nef sequences from HIV-1 subtype B. In contrast, multiple pairwise comparisons of nonsynonymous and synonymous substitution rates provided no good evidence for positive selection and sliding window analyses failed to detect most positively selected sites. These findings demonstrate that positive selection is an important determinant of nef gene evolution and that genealogy-based methods outperform pairwise methods in the detection of adaptive evolution. Mapping the locations of positively selected sites may also be of use in identifying targets of the immune response and hence aid vaccine design.

Acquired Immunodeficiency Syndrome↗
PubMed1999

Phylogenetic evidence for recombination in dengue virus.

A split decomposition analysis of dengue (DEN) virus gene sequences revealed extensive networked evolution, indicative of recombination, among DEN-1 strains but not within serotypes DEN-2, DEN-3, or DEN-4. Within DEN-1, two viruses sampled from South America in the last 10 years were identified as recombinants. To map the breakpoints and test their statistical support, we developed a novel maximum likelihood method. In both recombinants, the breakpoints were found to be in similar positions, within the fusion peptide of the envelope protein, demonstrating that a single recombination event occurred prior to the divergence of these two strains. This is the first report of recombination in natural populations of dengue virus.

Dengue↗
PubMed1999

The influence of recombination on the population structure and evolution of the human pathogen Neisseria meningitidis.

The extent to which recombination disrupts the bifurcating treelike phylogeny and clonal structure imposed by binary fission on bacterial populations remains contentious. Here, we address this question with a study of nucleotide sequence data from 107 isolates of the human pathogen Neisseria meningitidis. Gene fragments from 12 house-keeping loci distributed around the meningococcal chromosome were analyzed, showing that (1) identical alleles are disseminated among genetically diverse isolates, with no evidence for linkage disequilibrium; (2) different loci give distinct and incongruent phylogenetic trees; and (3) allele sequences are incompatible with a bifurcating treelike phylogeny at all loci. These observations are consistent with the hypothesis that meningococcal populations comprise organisms assembled from a common gene pool, with alleles and allele fragments spreading independently, together with the occasional importation of genetic material from other species. Further, they support the view that recombination is an important genetic mechanism in the generation new meningococcal clones and alleles. Consequently, for anything other than the short-term evolution of this species, a bifurcating treelike phylogeny is not an appropriate model.

Alleles↗
PubMed1999

Networks and groups within the genus Neisseria: analysis of argF, recA, rho, and 16S rRNA sequences from human Neisseria species.

To understand the pattern of nucleotide sequence variation among bacteria that frequently exchange chromosomal genes, we analyzed sequences of the recA, argF, and rho genes, as well as part of the small-subunit (16S) rRNA gene, from about 50 isolates of human commensal Neisseria species and the pathogenic N. meningitidis and N. gonorrhoeae. Almost all isolates of these species could be assigned to five phylogenetic groups that are found for all genes examined and generally are supported by high bootstrap values. In contrast, the phylogenetic relationships among groups varied according to the gene analyzed with notable incongruences involving N. cinerea and N. lactamica. Further analysis using split decomposition showed that for each gene, including 16S rRNA, the patterns of sequence divergence within N. meningitidis and closely related species were inconsistent with a bifurcating treelike phylogeny and better represented by an interconnected network. These data indicate that the human commensal Neisseria species can be separated into discrete groups of related species but that the relationships both within and among these groups, including those reconstructed using 16S rRNA, have been distorted by interspecies recombination events.

Base Sequence↗
PubMed1999

The mid-depth method and HIV-1: a practical approach for testing hypotheses of viral epidemic history.

We introduce the mid-depth method, a practical approach for testing hypotheses of demographic history using genealogies reconstructed from sequence data. The relative positions of internal nodes within a genealogy contain information about past population dynamics. We explain how this information can be used to (1) test the null hypothesis of constant population size and (2) estimate the growth rate and current population size of an exponentially growing population. Simulation tests indicate that, as expected, estimates of exponential growth rates are sometimes biased. The mid-depth method is computationally rapid and does not require knowledge of the sample's mutation rate. However, it does assume that the reconstructed genealogy is correct and is therefore best suited to the analysis of variation-rich viral data sets. When applied to HIV-1 sequence data, the mid-depth method provides phylogenetic evidence of different exponential growth rates for subtypes A and B. We posit that this difference in growth rate reflects the different transmission routes and epidemiological histories of the two subtypes.

HIV-1↗
PubMed1998

Molecular epidemiology and evolution of emerging infectious diseases.

Molecular epidemiology is an emerging science. The development of new and rapid protocols to isolate and identify pathogens, coupled with the sophisticated phylogenetic analysis of their gene sequences, is providing a new and fascinating insight into the biology, origin and spread of infectious diseases. In this essay, I describe some of the ways in which the techniques of modern molecular biology and evolution have equipped us to face the challenge of these new infections.

Dengue↗
PubMed1998

Tracing the origins of louping ill virus by molecular phylogenetic analysis.

The nucleotide and deduced amino acid sequences of louping ill (LI) virus isolates, collected from representative regions of the British Isles and Norway, were determined for either the entire envelope gene (20 isolates) or for a portion of the envelope gene that spans a hypervariable region and includes an LI virus specific marker sequence (53 isolates). Phylogenetic analysis reveals the presence of three major geographical populations of LI virus in the British Isles, viz. Irish, Welsh and British LI viruses, which all cause encephalomyelitis in animals, predominantly sheep, and co-habit the same tick population. British LI virus occurs throughout Scotland, England, Ireland and Norway. Irish and Welsh LI viruses occur only in Ireland and Wales, respectively. Phylogenetic analysis also predicts that LI virus initially emerged in Ireland and that a descendant was introduced into Great Britain via Wales and was subsequently transported to the borders of Scotland, from where it was dispersed throughout Scotland, northern England and Norway. More recently, the British LI virus was reintroduced into Ireland and also into south-west England. Dates of lineage divergence, calculated from the synonymous substitution rate, indicate that LI virus emerged in the British Isles less than 800 years ago and most LI virus dispersal occurred during the last 300 years. By combining these data with historical records it appears that livestock movement can be implicated in the dispersal of LI virus.

Animals↗
PubMed1997

Secondary structure of the 3' untranslated region of flaviviruses: similarities and differences.

Genetic algorithm-based RNA secondary structure prediction was used in combination with comparative sequence analysis to construct models of folding for the distal part of the 3'-untranslated region of flaviviruses belonging to four serological groups. Elements of RNA secondary structure that are preserved among all the flaviviruses studied were revealed, despite the high degree of sequence divergence between them. At the same time, structural elements were observed that distinguish members of different serological groups and, in particular, a region of remarkable structural divergence between the tick-borne and mosquito-borne flaviviruses was found. Application of the genetic algorithm also revealed that the 3'-terminus of flaviviral genomic RNA may take on alternative conformations, which are not observed in the 3'-terminus of complementary minus strand RNA. These alternative folding patterns may have roles in the regulation of transcription and translation initiation and in the switch between them.

Animals↗
PubMed1997

Complete sequence of two tick-borne flaviviruses isolated from Siberia and the UK: analysis and significance of the 5' and 3'-UTRs.

The complete nucleotide sequence of two tick-transmitted flaviviruses, Vasilchenko (Vs) from Siberia and louping ill (LI) from the UK, have been determined. The genomes were respectively, 10928 and 10871 nucleotides (nt) in length. The coding strategy and functional protein sequence motifs of tick-borne flaviviruses are presented in both Vs and LI viruses. The phylogenies based on maximum likelihood, maximum parsimony and distance analysis of the polyproteins, identified Vs virus as a member of the tick-borne encephalitis virus subgroup within the tick-borne serocomplex, genus Flavivirus, family Flaviviridae. Comparative alignment of the 3'-untranslated regions revealed deletions of different lengths essentially at the same position downstream of the stop codon for all tick-borne viruses. Two direct 27 nucleotide repeats at the 3'-end were found only for Vs and LI virus. Immediately following the deletions a region of 332-334 nt with relatively conserved primary structure (67-94% identity) was observed at the 3'-non-coding end of the virus genome. Pairwise comparisons of the nucleotide sequence data revealed similar levels of variation between the coding region, and the 5' and 3'-termini of the genome, implying an equivalent strong selective control for translated and untranslated regions. Indeed the predicted folding of the 5' and 3'-untranslated regions revealed patterns of stem and loop structures conserved for all tick-borne flaviviruses suggesting a purifying selection for preservation of essential RNA secondary structures which could be involved in translational control and replication. The possible implications of these findings are discussed.

Animals↗
PubMed1997

Uroscopy in the 21st century: high-field NMR spectroscopy.

From the experiments described, it can be seen that there are different research approaches that can be taken and these are summarized in Table 1. Whereas much scientific research is principally hypothesis led, there remains, nevertheless, an important place for exploratory research. High resolution NMR can measure, directly and simultaneously, a wide range of endogenous metabolites in biological fluids and has the unique capability of providing structural information on the metabolites detected. It has proved to be a powerful research tool with which to study inherited metabolic diseases, renal disease, drug metabolism, and toxicity, and can be used to monitor the effects of drug therapy. For instance, by using a library of experimental toxins one can map the metabolic profile of site-specific nephron injury. With this approach in man one could eventually take an unknown disease such as Balkan nephropathy and predict the initial site of tubular injury, the mode of injury and therefore the kind of toxin capable of producing that injury. NMR spectroscopic techniques are still advancing rapidly, with ever increasing sensitivity and sophistication of NMR pulse sequences to enhance structural elucidation in complex mixtures. Given the advances in directly coupled HPLC-NMR and even HPLC-NMR-mass spectroscopy it is likely that these technologies in conjunction with pattern recognition will make major contribution to our understanding of renal processes and provide new diagnostic insights in the 21st century.

Forecasting↗