The molecular epidemiology of the virus and mapping helps understand the epidemics' evolution and apply quick control measures. This study provides genomic evidence of multiple severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) introductions into Sri Lanka and virus evolution during circulation. Whole-genome sequences of four SARS-CoV-2 strains obtained from coronavirus disease 2019 (COVID-19) positive patients reported in Sri Lanka during March 2020 were compared with sequences from Europe, Asia, Africa, Australia and North America. The phylogenetic analysis revealed that the sequence of the sample of the first local patient collected on 10 March, who contacted tourists from Italy, was clustered with SARS-CoV-2 strains collected from Italy, Germany, France and Mexico. Subsequently, the sequence of the isolate obtained on 19 March also clustered in the same group with the samples collected in March and April from Belgium, France, India and South Africa. The other two strains of SARS-CoV-2 were segregated from the main cluster, and the sample collected from 16 March clustered with England and the sample collected on 30 March showed the highest genetic divergence to the isolate of Wuhan, China. Here we report the first molecular epidemiological study conducted on circulating SARS-CoV-2 in Sri Lanka. The finding provides the robustness of molecular epidemiological tools and their application in tracing possible exposure in disease transmission during the pandemic.

When the iconic DNA genetic code is expressed in terms of energy differentials, one observes that information embedded in chemical sequences, including some biological outcomes, correlate with distinctive free energy profiles. Specifically, we find correlations between codon usage and codon free energy, suggestive of a thermodynamic selection for codon usage. We also find correlations between what are considered ancient amino acids and high codon free energy values. Such correlations may be reflective of the sequence-based genetic code fundamentally mapping as an energy code. In such a perspective, one can envision the genetic code as composed of interlocking thermodynamic cycles that allow codons to ‘evolve’ from each other through a series of sequential transitions and transversions, which are influenced by an energy landscape modulated by both thermodynamic and kinetic factors. As such, early evolution of the genetic code may have been driven, in part, by differential energetics, as opposed exclusively by the functionality of any gene product. In such a scenario, evolutionary pressures can, in part, derive from the optimization of biophysical properties (e.g. relative stabilities and relative rates), in addition to the classic perspective of being driven by a phenotypical adaptive advantage (natural selection). Such differential energy mapping of the genetic code, as well as larger genomic domains, may reflect an energetically resolved and evolved genomic landscape, consistent with a type of differential, energy-driven ‘molecular Darwinism’. It should not be surprising that evolution of the code was influenced by differential energetics, as thermodynamics is the most general and universal branch of science that operates over all time and length scales.

All organisms encounter pathogens, and birds are especially susceptible to infection by malaria parasites and other haemosporidians. It is important to understand how immune genes, primarily innate immune genes which are the first line of host defense, have evolved across birds, a highly diverse group of tetrapods. Here, we find that innate immune genes are highly conserved across the avian tree of life and that although most show evidence of positive or diversifying selection within specific lineages or clades, the number of sites is often proportionally low in this broader context of putative constraint. Rather, evidence shows a much higher level of negative or purifying selection in these innate immune genes – rather than adaptive immune genes – which is consistent with birds' long coevolutionary history with pathogens and the need to maintain a rapid response to infection. We further explored avian responses to haemosporidians by comparing differential gene expression in wild birds (1) uninfected with haemosporidians, (2) infected with Plasmodium and (3) infected with Haemoproteus (Parahaemoproteus). We found patterns of significant differential expression with some genes unique to infection with each genus and a few shared between ‘treatment’ groups, but none that overlapped with the genes included in the phylogenetic study.

Geographically isolated environments such as the conifer forests atop the Madrean “sky islands” in southeastern Arizona provide natural laboratories for studying factors involved in speciation and origins of biodiversity. Using molecular and geospatial analyses, we examine beetle population phylogeny, regional climate records, and the Quaternary paleobiogeography of forests to evaluate four hypothetical scenarios regarding the current geographic and population genetic patterns of Scaphinotus petersi. Scaphinotus petersi is a large, flightless beetle that resides in the Madrean conifer forests above ~ 1900 m asl. Our results do not support the current hypothesis that S. petersi populations found on seven separate mountain ranges are genetically distinct and separated as temperatures warmed after the Last Glacial Maximum (LGM). Rather, we show that only some of the ranges hold genetically distinct populations, and the timing of separation among the populations does not appear to coincide with specific climatic events such as warming trends. In addition, we show that predicted changes to the climate of the Madrean sky islands may result in the disappearance of S. petersi from some of the lower ranges by the end of this century.

Retinitis pigmentosa (RP) comprises several heritable diseases that involve photoreceptor, and ultimately retinal, degeneration. Currently, mutations in over 50 genes have known links to RP. Despite advances in clinical characterization, molecular characterization of RP remains challenging due to the heterogeneous nature of causal genes, mutations, and clinical phenotypes. In this study, we compiled large datasets of two important visual genes associated with RP: rhodopsin, which initiates the phototransduction cascade, and the retinoid isomerase RPE65, which regenerates the visual cycle. We used a comparative evolutionary approach to investigate the relationship between interspecific sequence variation and pathogenic mutations that lead to degenerative retinal disease. Using codon-based likelihood methods, we estimated evolutionary rates (d N/d S) across both genes in a phylogenetic context to investigate differences between pathogenic and nonpathogenic amino acid sites. In both genes, disease-associated sites showed significantly lower evolutionary rates compared to nondisease sites, and were more likely to occur in functionally critical areas of the proteins. The nature of the dataset (e.g., vertebrate or mammalian sequences), as well as selection of pathogenic sites, affected the differences observed between pathogenic and nonpathogenic sites. Our results illustrate that these methods can serve as an intermediate step in understanding protein structure and function in a clinical context, particularly in predicting the relative pathogenicity (i.e., functional impact) of point mutations and their downstream phenotypic effects. Extensions of this approach may also contribute to current methods for predicting the deleterious effects of candidate mutations and to the identification of protein regions under strong constraint where we expect pathogenic mutations to occur.

The rate of new emerging infectious diseases entering the human population has increased over the past century, with pathogens originating from animals or from products of animal origin accounting for the vast majority. Primary risk factors for the emergence and spread of emerging zoonoses include expansion and intensification of animal agriculture and long-distance live animal transport, live animal markets, bushmeat consumption and habitat destruction. Developing effective control strategies is contingent upon the ability to test causative hypotheses of disease transmission within a statistical framework. Broadly speaking, molecular phylogeography offers a framework in which specific hypotheses regarding pathogen gene flow and dispersal within an ecological context can be compared. A number of different methods has been developed for this application. Here, our intent is firstly to discuss the application of a wide variety of statistically based methods (including Bayesian reconstruction, network parsimony analysis and regression) to specific viruses (influenza, salmon anaemia virus, foot and mouth disease and Rift Valley Fever) that have been associated with animal farming/movements; and secondly to place them in the larger framework of the threat of potential zoonotic events as well as the economic and biosecurity implications of pathogen outbreaks among our animal food sources.

Trypanosomatids represent the causative agents of major diseases in humans, livestock and plants, with inevitable suffering and economic hardship as a result. They are also evolutionarily highly divergent organisms, and the many unique aspects of trypanosome biology provide opportunities in terms of identification of drug targets, the challenge of exploiting these putative targets and, at the same time, significant scope for exploration of novel and divergent cell biology. We can estimate from genome sequences that the degree of divergence of trypanosomes from animals and fungi is extreme, with perhaps one third to one half of predicted trypanosome proteins having no known function based on homology or recognizable protein domains/architecture. Two highly important aspects of trypanosome biology are the flagellar pocket and the nuclear envelope, where in silico analysis clearly suggests great potential divergence in the proteome. The flagellar pocket is the sole site of endo- and exocytosis in trypanosomes and plays important roles in immune evasion via variant surface glycoprotein (VSG) trafficking and providing a location for sequestration of various invariant receptors. The trypanosome nuclear envelope has been largely unexplored but, by analogy with higher eukaryotes, roles in the regulation of chromatin and most significantly, in controlling VSG gene expression are expected. Here we discuss recent successful proteomics-based approaches towards characterization of the nuclear envelope and the endocytic apparatus, the identification of conserved and novel trypanosomatid-specific features, and the implications of these findings.

Stomatopod crustaceans possess apposition compound eyes that contain more photoreceptor types than any other animal described. While the anatomy and physiology of this complexity have been studied for more than two decades, few studies have investigated the molecular aspects underlying the stomatopod visual complexity. Based on previous studies of the structure and function of the different types of photoreceptors, stomatopod retinas are hypothesized to contain up to 16 different visual pigments, with 6 of these having sensitivity to middle or long wavelengths of light. We investigated stomatopod middle- and long-wavelength-sensitive opsin genes from five species with the hypothesis that each species investigated would express up to six different opsin genes. In order to understand the evolution of this class of stomatopod opsins, we examined the complement of expressed transcripts in the retinas of species representing a broad taxonomic range (four families and three superfamilies). A total of 54 unique retinal opsins were isolated, resulting in 6–15 different expressed transcripts in each species. Phylogenetically, these transcripts form six distinct clades, grouping with other crustacean opsins and sister to insect long-wavelength visual pigments. Within these stomatopod opsin groups, intra- and interspecific clusters of highly similar transcripts suggest that there has been rampant recent gene duplication. Some of the observed molecular diversity is also due to ancient gene duplication events within the stem crustacean lineage. Using evolutionary trace analysis, 10 amino acid sites were identified as functionally divergent among the six stomatopod opsin clades. These sites form tight clusters in two regions of the opsin protein known to be functionally important: six in the chromophore-binding pocket and four at the cytoplasmic surface in loops II and III. These two clusters of sites indicate that stomatopod opsins have diverged with respect to both spectral tuning and signal transduction.

Neuropeptides are ubiquitous signalling molecules in all metazoans possessing nervous systems, from the simple nerve nets of the cnidarians to the immensely complex systems of mammals. While the discipline of peptide neuroendocrinology was born through the study of higher vertebrates, there now exists a plethora of information regarding neuropeptides and peptidic regulatory factors in invertebrates. Such phylogenetic studies have revealed that peptidic neurotransmission is of early evolutionary origin and that, while invertebrates have nervous systems which are simpler in terms of nerve cell number and organisation when compared with vertebrates, the complexity of the peptidic ‘vocabulary’ of invertebrate neurones is of a similar order of magnitude. Most research on invertebrate neuropeptides has been directed towards representative members of groups such as the insects and molluscs and it is only in recent years that efforts have been focused on the helminths (platyhelminths and nematodes). Here, the putative origins of peptidic transmitters is discussed and the current state of knowledge on helminth neuropeptides is reviewed. In order to place the study of helminth neuropeptides in an historical and conceptual perspective, methodological development and conceptual modifications in the disciplines of vertebrate and higher invertebrate peptide neuroendocrinology have been summarised.

Partial 28S ribosomal RNA (rRNA) gene sequences, including the variable domains D1, D2 and D3, were determined for representative species from the 4 Schistosoma species groups. On an alignment of 1345 bp from S. mansoni, S. haematobium, S. spindale and S. japonicum (with Heterobilharzia americana chosen as an outgroup), both maximum likelihood and maximum parsimony analyses provide a robust molecular phytogeny for the genus; ((((S. haematobium, S. spindale), S. mansoni), S. japonicum), H. americana). When analysed separately, both domain D1 and domain D2 yielded similarly informative data whereas D3 failed to resolve the phylogeny. These results confirm a phylogeny previously suggested by 18S rRNA gene sequences, corroborating the status of S. spindale as a sister taxon to S. haematobium, and demonstrate the utility of 28S rRNA gene sequence data for resolving phylogenies within the Schistosomatidae.

Note: nucleotide sequence data reported in this paper are available in the EMBL database under the accession numbers Z46503/04/05/06/21.

A parasite can be considered to be the device of a nucleic acid which allows it to exploit the gene products of other nucleic acid–the host organisms. In this view, all parasites are ‘molecular parasites’. But it is interesting to restrict our attention to nucleic acids which do not encode organisms, as these live in a purely molecular world which lacks emergent features such as fangs and ovipositors. Viruses and transposons are molecular parasites in this sense. Most viral nucleic acids do code for some proteins, such as replicases and the protein shell in which they travel between their cellular oases. Some, however, do not even have a shell and code for nothing at all–these are the ‘viroids’ (Reisner & Gross, 1985), the smallest parasites in the world.

We hypothesize that forms of antagonistic coevolution have forged strong links between positive selection at the molecular level and increased cancer risk. By this hypothesis, evolutionary conflict between males and females, mothers and foetuses, hosts and parasites, and other parties with divergent fitness interests has led to rapid evolution of genetic systems involved in control over fertilization and cellular resources. The genes involved in such systems promote cancer risk as a secondary effect of their roles in antagonistic coevolution, which generates evolutionary disequilibrium and maladaptation. Evidence from two sources: (1) studies on specific genes, including SPANX cancer/testis antigen genes, several Y-linked genes, the pem homebox gene, centromeric histone genes, the breast cancer gene BRCA1, the angiogenesis gene ANG, cadherin genes, cytochrome P450 genes, and viral oncogenes; and (2) large-scale database studies of selection on different functional categories of genes, supports our hypothesis. These results have important implications for understanding the evolutionary underpinnings of cancer and the dynamics of antagonistically-coevolving molecular systems.

It is widely believed that the current DNA–RNA–protein-based life forms have evolved from preceding RNA–protein-based life forms, and these again, from mere RNA replicons. By rationale, it can be assumed that the early RNA replicons were fully heterotrophic in terms of obtaining all their building blocks from their environment. In the absence of protein catalysts, their essential life functions had to be mediated by simple functional structures and mechanisms, such as RNA secondary structures, RNA–RNA interactions and RNA-mediated catalysis, and possibly by catalytic minerals or clays. The central role of RNA catalysts in early life forms is supported by the fact that several catalytic RNAs still perform central biological functions in current life forms, and at least some of these may be derived as molecular relicts from the early RNA-based life. The RNA-catalysed metabolic reactions and molecular fossils are more conserved in the eukaryotic life forms than in the prokaryotes, suggesting that the linear eukaryote genomes may more closely resemble the structure and function of the early RNA replicons, than what do the circular prokaryote genomes. Present-day RNA viruses and viroids utilize ultimately simple life strategies, which may be similar to those used by the early RNA replicons. Thus, molecular and functional properties of viruses and viroids may be considered as examples or models of the structures and replication mechanisms, which might have been used for the replication of the early biopolymers.

The stability of a trans-acting delta ribozyme was studied under various conditions. Although in vitro (i.e., in the presence of protein extracts) this delta ribozyme appears to be only slightly more stable than a hammerhead ribozyme, in vivo (i.e., after cell transfection) it exhibits an outstanding stability that manifests itself in the calculated half-life of over 100 h regardless of the means of transfection. The P2 stem, which includes both the 5′ and 3′ ends, is shown to play a critical role in this stability. Direct mutagenesis of the most nuclease susceptible nucleotides failed to generate a more stable ribozyme that retained the same catalytic potential. Clearly, delta ribozyme appears to be well adapted to the human cell environment, and is therefore ideal for the development of a gene-inactivation system.

Catalytic RNAs are often regarded as molecular fossils from the RNA World, yet it is usually difficult to get more specific information about their evolution. Here we have investigated the coevolution of group II intron RNA structures with their intron-encoded reverse transcriptases (RTs). Unlike group I introns, there has been no obvious reshuffling between intron RNA structures and ORFs. Of the six classes of intron structures that encode ORFs, three are conventional forms of group II A1, B1, and B2 secondary structures, whereas the remaining classes are bacterial, are possibly associated with the most primitive ORFs, and have unusual features and hybrid features of group IIA and group IIB intron structures. Based on these data, we propose a new model for the evolution of group II introns, designated the retroelement ancestor hypothesis, which predicts that the major RNA structural forms of group II introns developed through coevolution with the intron-encoded protein rather than as independent catalytic RNAs, and that most ORF-less introns are derivatives of ORF-containing introns. The model is supported by the distribution of ORF-containing and ORF-less introns, and by numerous examples of ORF-less introns that contain ORF remnants.

As models for molecular evolution, immune response, and local search algorithms, various authors have used a stochastic process called the evolutionary walk, which goes as follows. Assign a random number to each vertex of the infinite N-ary tree, and start with a particle on the root. A step of the process consists of searching for a child with a higher number and moving the particle there; if no such child exists, the process stops. The average number of steps in this process is asymptotic, as N → ∞, to log N, a result first proved by Macken and Perelson. This paper relates the evolutionary walk to a process called random bisection, familiar from combinatorics and number theory, which can be thought of as a transformed Poisson process. We first give a thorough treatment of the exact walk length, computing its distribution, moments and moment generating function. Next we show that the walk length is asymptotically normally distributed. We also treat it as a mixture of Poisson random variables and compute the asymptotic distribution of the Poisson parameter. Finally, we show that in an evolutionary walk with uniform vertex numbers, the ‘jumps’, ordered by size, have the same asymptotic distribution as the normalized cycle lengths in a random permutation.

Internal transcribed spacer (ITS) nrDNA sequences of 42 accessions (representing 28 species) of Aframomum revealed an unusually low level of sequence variation, suggesting a recent radiation of the genus in Africa. The sample of species analysed includes all the main morphological variation and is based on wide geographical sampling. The Aframomum sequences varied from 187bp to 190bp (ITS 1) and 215bp to 216bp (ITS 2). Pair-wise sequence difference between accessions varied from 0% (e.g. A. luteoalbum and A. thonneri) to 2.74% (e.g. A. sp. nov. B to A. pseudostipulare). This contrasts with a comparable data set for the SE Asian genus Alpinia in the same tribe (Alpineae) in which maximum pair-wise difference is six times greater (range 0.5–15.6%). A parsimony analysis of the in-group and out-group taxa supports the monophyly of the genus Aframomum, but does not resolve the relationships between the in-group species. Four putative multi-species groups, however, have some jackknife support. The species sampled vary greatly in vegetative, floral and fruit characters. This morphological variation is not reflected in the ITS sequence data. This may be a result of rapid radiation under conditions of Pleistocene climatic change and effective dispersal of seeds by primates.

Dimeric proteins can arise by the swapping of structural domains between monomers. The prevalence of this occurrence is unknown. Ribonuclease A (RNase A) is assumed to be a monomer near physiological conditions. Here, this hypothesis is tested and found to be imprecise. The two histidine residues (His12 and His119) in the active site of RNase A arise from two domains (S-peptide and S-protein) of the protein. The H12A and H119A variants have 105-fold less ribonucleolytic activity than does the wild-type enzyme. Incubating a 1:1 mixture of the H12A and H119A variants at pH 6.5 and 65 °C results in a 103-fold increase in ribonucleolytic activity. A large quantity of active dimer can be produced by lyophilizing a 1:1 mixture of the H12A and H119A variants from acetic acid. At pH 6.5 and 65 °C, the ribonucleolytic activity of this dimer converges to that of the dimer formed by simply incubating the monomers, as expected for a monomer–dimer equilibrium. The equilibrium dissociation constant for the dimer is near 2 mM at both 65 and 37 °C. This value of Kd is only 20-fold greater than the concentration of RNase A in the cow pancreas, suggesting that RNase A dimers exist in vivo. The intrinsic ability of RNase A to form dimers under physiological conditions is consistent with a detailed model for the evolution of homodimeric proteins. Dimers of “monomeric” proteins could be more prevalent than is usually appreciated.

The structure–functional convergence between two Zn-dependent proteases, namely thermolysin and mitochondrial processing peptidase (MPP), is described. These two families of nonhomologous enzymes show not only functional convergence of several active site residues as in chymotrypsin and subtilisin, but also structural convergence of overall molecular architectures including the β-sheet arrangement and packing of the surrounding α-helices. The major functionally important structural elements are present in both enzymes with different topological connections and often in reverse main-chain orientation, but display similar packing. The structural comparison helps to rationalize sequence “inversion” of the HEXXH thermolysin consensus present as HXXEH in MPP. The described structural convergence may be due to a limited number of alternatives to build a Zn-protease that utilizes hydrogen bonding between a substrate main chain and the enzyme β-sheet for substrate binding.

Resemblance between animal taxa may be due to convergence rather than to recent common ancestry. Constraints on biological materials and adaptation to particular habits or habitats will produce widespread convergence. How may we distinguish the two causes of resemblance? The relationship between convergence and taxonomy is discussed, demonstrating that the choice of taxonomic method will itself determine the extent to which convergence is perceived. In particular, cladistic analysis based on parsimony will tend to minimise and thus conceal convergence: neither the resulting cladogram nor a consistency index derived from it can be used to assess the prevalence of convergence. With any taxonomic system, there can be no substitute for evaluation of the morphological characters used. Complementary use of molecular characters shows promise: we wait further understanding of constraints in genetic evolution and of the possibilities of convergence at this level also.

These general principles are illustrated with a range of examples from within and between invertebrate phyla: the phylogeny of Cnidaria and Platyhelminthes cannot be traced with certainty, but where the fossil record allows clear rooting, as for the echinoderms and in particular the echinoids, combination of morphological and molecular methods has made much progress. Sub-groups within a phylum, for example opisthobranch molluscs and the dipteran Phoridae, may show an uncontested phylogeny, and here studies have precisely identified convergence and shown that it may be the commoner cause of resemblance. Adaptation to exacting environments shown by terrestial and freshwater nemertines may also result in a predominance of convergent resemblance.

Traditional grouping of phyla breaks down on re-examination of supposedly key characters, such as segmentation, body cavities, germ layers and symmetry, each of which must have had multiple origins: nor are developmental stages (especially not larvae) a reliable guide to relationships. Demarcation of phyla may be difficult, as with arthropods, and location of phyla is even more difficult, due to their early and rapid radiation. Over-simplified definition of characters has bedevilled invertebrate classification and the use of molecular data has not yet resolved the major controversies.

The question ‘How common is convergence?’ remains unanswered and may be unanswerable. Our examples indicate that even the minimum detectable levels of convergence are often high, and we conclude that at all levels convergence has been greatly underestimated.