Robin M. Bush

Ecological and immunological determinants of influenza evolution

In pandemic and epidemic forms, influenza causes substantial, sometimes catastrophic, morbidity and mortality. Intense selection from the host immune system drives antigenic change in influenza A and B, resulting in continuous replacement of circulating strains with new variants able to re-infect hosts immune to earlier types. This ‘antigenic drift’ often requires a new vaccine to be formulated before each annual epidemic. However, given the high transmissibility and mutation rate of influenza, the constancy of genetic diversity within lineages over time is paradoxical. Another enigma is the replacement of existing strains during a global pandemic caused by ‘antigenic shift’—the introduction of a new avian influenza A subtype into the human population. Here we explore ecological and immunological factors underlying these patterns using a mathematical model capturing both realistic epidemiological dynamics and viral evolution at the sequence level. By matching model output to phylogenetic patterns seen in sequence data collected through global surveillance, we find that short-lived strain-transcending immunity is essential to restrict viral diversity in the host population and thus to explain key aspects of drift and shift dynamics.

Predicting adaptive evolution.

Phylogenetic trees reconstruct past evolution and can provide evidence of past evolutionary pressure on genes and on individual codons. In addition to tracing past evolutionary events, molecular phylogenetics might also be used to predict future evolution. Our ability to verify adaptive hypotheses using phylogenetics has broad implications for vaccine design, genomics and structural biology.

Molecular evolution of the Chlamydiaceae

Phylogenetic analyses of surface antigens and other chlamydial proteins were used to reconstruct the evolution of the Chlamydiaceae. Trees for all five coding genes [the major outer-membrane protein (MOMP), GroEL chaperonin, KDO-transferase, small cysteine-rich lipoprotein and 60 kDa cysteine-rich protein] supported the current organization of the family Chlamydiaceae, which is based on ribosomal, biochemical, serological, ecological and DNA-DNA hybridization data. Genetic distances between some species were quite large, so phylogenies were evaluated for robustness by comparing analyses of both nucleotide and protein sequences using a variety of algorithms (neighbour-joining, maximum-likelihood, maximum-parsimony with bootstrapping, and quartet puzzling). Saturation plots identified areas of the trees in which factors other than relatedness may have determined branch attachments. All nine species were clearly differentiated by distinctness ratios calculated for each gene. The distribution of virulence traits such as host and tissue tropism were mapped onto the consensus phylogeny. Closely related species were no more likely to share virulence characters than were more distantly related species. This phylogenetically disjunct distribution of virulence traits could not be explained by lateral transfer of the genes we studied, since we found no evidence for lateral gene transfer above the species level. One interpretation of this observation is that when chlamydiae gain access to a new niche, such as a new host or tissue, significant adaptation ensues and the virulence phenotype of the new species reflects adaptation to its environment more strongly than it reflects its ancestry.

Mechanism of 150-cavity formation in influenza neuraminidase

The recently discovered 150-cavity in the active site of group-1 influenza A neuraminidase (NA) proteins provides a target for rational structure-based drug development to counter the increasing frequency of antiviral resistance in influenza. Surprisingly, the 2009 H1N1 pandemic virus (09N1) neuramidase was crystalized without the 150-cavity characteristic of group-1 NAs. Here we demonstrate, through a total sum of 1.6 μs of biophysical simulations, that 09N1 NA exists in solution preferentially with an open 150-cavity. Comparison with simulations using avian N1, human N2 and 09N1 with a I149V mutation and an extensive bioinformatics analysis suggests that the conservation of a key salt bridge is crucial in the stabilization of the 150-cavity across both subtypes. This result provides an atomic-level structural understanding of the recent finding that antiviral compounds designed to take advantage of contacts in the 150-cavity can inactivate both 2009 H1N1 pandemic and avian H5N1 viruses.

Evidence for the adaptive significance of allozymes in forest trees

Positive correlations between allozyme heterozygosity and fitness measures, primarily growth and fecundity, have been reported for a number of forest tree species. Because the amount of variation in growth explained by allozyme genotype is usually on the order of only a few percent, there has been little effort made towards using electrophoretic screening of allozymes as a tool in early selection on seedlings in production nurseries. We review the progress made in studies of heterozygosity in forest trees, focusing on how recent studies have utilized careful experimental design to allow testing of hypotheses as to the causative nature of the heterozygosity-fitness phenomena. We discuss evidence suggesting a deleterious nature for rare allozyme alleles, and present a case of apparent balancing selection across life history stages acting to maintain rare alleles in Pinus taeda. We also review the apparently common trend in natural stands toward increasing heterozygosity over time, and suggest how gains might be made through artificial selection based on allozyme survey data.

The Fitness Consequences Of Multiple‐Locus Heterozygosity: The Relationship Between Heterozygosity And Growth Rate In Pitch Pine (Pinus Rigida Mill.)

Positive correlations between measures of “fitness” and the number of electrophoretic loci for which an individual is heterozygous have been observed in many species. Two major hypotheses have been proposed to explain this phenomenon: inbreeding depression and overdominance. Until recently, there has been no way to distinguish between these hypotheses. The overdominance model devised by Smouse (1986) is used here in a reanalysis of Ledig et al.‘s (1983) study of heterozygosity and growth rate in eight populations of pitch pine and is contrasted with an inbreeding‐depression analysis. Ledig et al. (1983) regressed mean growth rate per heterozygosity class on the number of heterozygous loci, a method of analysis which, although it points to general trends in the data, does not differentiate between hypotheses. The correlations they obtained in four populations were significant only because regressing on the means eliminates most of the sum of squares for error and does not weight the unequally sized heterozygosity classes. Reanalysis of Ledig et al.‘s data using individuals, not means, showed no significant correlations between heterozygosity and fitness.

Prediction of the membrane‐spanning β‐strands of the major outer membrane protein of Chlamydia

There is preliminary experimental evidence indicating that the major outer‐membrane protein (MOMP) of Chlamydia is a porin. We tested this hypothesis for the MOMP of the mouse pneumonitis serovar of Chlamydia trachomatis using two secondary structure prediction methods. First, an algorithm that calculates the mean hydrophobicity of one side of putative β‐strands predicted the positions of 16 transmembrane segments, a structure common to known porins. Second, outer loops typical of porins were assigned using an artificial neural network trained to predict the topology of bacterial outer‐membrane proteins with a predominance of β‐strands. A topology model based on these results locates the four variable domains (VDs) of the MOMP on the outer loops and the five constant domains on β‐strands and the periplasmic turns. This model is consistent with genetic analysis and immunological and biochemical data that indicate the VDs are surface exposed. Furthermore, it shows significant homology with the consensus porin model of the program FORESST, which contrasts a proposed secondary structure against a data set of 349 proteins of known structure. Analysis of the MOMP of other chlamydial species corroborated our predicted model.

THE IMPACT OF ELECTROPHORETIC GENOTYPE ON LIFE HISTORY TRAITS IN PINUS TAEDA

Reports of positive associations between allozymic heterozygosity and measures of fitness are routine, but it has not been possible to distinguish between the two preeminent explanations of the phenomenon, dominance and overdominance. We tested several of the assumptions of these hypotheses in our study of the relationship between electrophoretic genotype and three life history traits in loblolly pines (Pinus taeda L.). Traits examined included the survival and growth of selfed and outcrossed progeny of 45 maternal trees, and maternal fecundity, measured as the number of surviving progeny per mother tree. Inbreeding depression was severe; the relative fitness of the selfed progeny was only 8% that of the outcrossed progeny. We found a heterozygote fecundity advantage, which should have resulted in an excess of rare alleles in the progeny. Instead, there was evidence of severe survival selection against rare alleles in both heterozygous and homozygous forms. The deficit of rare alleles averaged 69 and 50% in the selfed and outcrossed progeny, respectively.

Barriers to antigenic escape by pathogens: trade-off between reproductive rate and antigenic mutability.

BackgroundA single measles vaccination provides lifelong protection. No antigenic variants that escape immunity have been observed. By contrast, influenza continually evolves new antigenic variants, and the vaccine has to be updated frequently with new strains. Both measles and influenza are RNA viruses with high mutation rates, so the mutation rate alone cannot explain the differences in antigenic variability.ResultsWe develop a new hypothesis to explain antigenic stasis versus change. We first note that the antigenically static viruses tend to have high reproductive rates and to concentrate infection in children, whereas antigenically variable viruses such as influenza tend to spread more widely across age classes. We argue that, for pathogens in a naive host population that spread more rapidly in younger individuals than in older individuals, natural selection weights more heavily a rise in reproductive rate. By contrast, pathogens that spread more readily among older individuals gain more by antigenic escape, so natural selection weights more heavily antigenic mutability.ConclusionThese divergent selective pressures on reproductive rate and antigenic mutability may explain some of the observed differences between pathogens in age-class bias, reproductive rate, and antigenic variation.

Evolution of β-glucuronidase regulation in the genus mus

Despite the central role suggested for regulatory mutations in many evolutionary scenarios, there is relatively little information available about the type and extent of regulatory differences between species, or to what extent differences between species are independent of variation within species. To address this issue we have studied the regulatory system of β‐glucuronidase, a gene implicated in a murine androgen‐inducible pheromone‐signalling system. We examined the changes in β‐glucuronidase hormonal regulation which have occurred during the radiation of a group of 12 closely related species of mice by assaying β‐glucuronidase activity in six different tissues after treatment with estrogen, and with androgen alone and in combination with either estrogen or growth hormone. We also examined in some detail the extent of variation in regulatory responses within species. We found extensive variation in regulatory phenotypes both within and among the species surveyed, suggesting that many of the species examined are currently polymorphic for various regulatory factors that affect inducibility of β‐glucuronidase. The variation we observed reflects changes in the ability of the β‐glucuronidase gene to respond to hormonal influences, rather than changes in aspects of the hormonal signalling system exterior to the gene. The marked differences among species in the renal and uterine responses to hormonal induction of β‐glucuronidase are not easily related to the phylogeny of the genus Mus. If hormonal induction of the gene for β‐glucuronidase is subject to natural selection, it appears to be subject to widely fluctuating selective forces. We review evidence that the apparently disorderly evolution of the hormonal responsiveness of β‐glucuronidase does not appear to be a unique property of this regulatory system. In contrast to the evolution of many protein sequences, which are tightly correlated with phylogeny and proceed at a relatively constant rate, some, perhaps many, regulatory phenotypes are in rapid evolutionary flux, providing an extensive range of phenotypes upon which selection can act.