Molecular Clock Rate Research Articles

Phylogeny and biogeography of harmochirine jumping spiders (Araneae: Salticidae)

We use ultraconserved elements (UCE) and Sanger data to study the phylogeny, age, and biogeographical history of harmochirine jumping spiders, a group that includes the species-rich genus Habronattus, whose remarkable courtship has made it the focus of studies of behaviour, sexual selection, and diversification. We recovered 1947 UCE loci from 43 harmochirine taxa and 4 outgroups, yielding a core dataset of 193 UCEs with at least 50 % occupancy. Concatenated likelihood and ASTRAL analyses confirmed the separation of harmochirines into two major clades, here designated the infratribes Harmochirita and Pellenita. Most are African or Eurasian with the notable exception of a clade of pellenites containing Habronattus and Pellenattus of the Americas and Havaika and Hivanua of the Pacific Islands. Biogeographical analysis using the DEC model favours a dispersal of the clade’s ancestor from Eurasia to the Americas, from which Havaika’s ancestor dispersed to Hawaii and Hivanua’s ancestor to the Marquesas Islands. Divergence time analysis on 32 loci with 85 % occupancy, calibrated by fossils and island age, dates the dispersal to the Americas at approximately 4 to 6 million years ago. The explosive radiation of Habronattus perhaps began only about 4 mya. The phylogeny clarifies both the evolution of sexual traits (e.g., the terminal apophyses was enlarged in Pellenes and not subsequently lost) and the taxonomy. Habronattus is confirmed as monophyletic. Pellenattus is raised to the status of genus, and 13 species moved into it as new combinations. Bianor stepposus Logunov, 1991 is transferred to Sibianor, and Pellenes bulawayoensis Wesołowska, 1999 is transferred to Neaetha. A molecular clock rate estimate for spider UCEs is presented and its utility to inform prior distributions is discussed.

Unravelling the complex interplay between antibiotic consumption and adaptive changes in methicillin-resistant Staphylococcus aureus.

This study aims to elucidate the genomic dynamics driving the emergence of antimicrobial resistance (AMR), with a specific focus on the interplay between AMR and antimicrobial usage. We conducted a comprehensive analysis using a ST239 methicillin-resistant Staphylococcus aureus (MRSA) dataset over a continuous 12-year period from a single hospital. Genomic analyses were performed tracking the changes in MRSA populations, particularly the emergence of reduced vancomycin susceptibility, and assessing the impact of glycopeptide use on these emergence events. Our findings reveal a significant correlation between hospital glycopeptide usage and the selection of MRSA strains with reduced vancomycin susceptibility. Genomic analyses provided insights into the molecular mechanisms driving resistance emergence, including the slowing of the molecular clock rate in response to heightened antimicrobial consumption. In conclusion, this study the highlights the complex dynamics between AMR and antimicrobial use at the hospital level. The observed correlation between antimicrobial consumption and the development of less susceptible MRSA strains underscores the importance of antimicrobial stewardship programmes and the establishment of optimal consumption thresholds for mitigating AMR effectively.

Mutation rates and adaptive variation among the clinically dominant clusters of Mycobacterium abscessus

Mycobacterium abscessus (Mab) is a multidrug-resistant pathogen increasingly responsible for severe pulmonary infections. Analysis of whole-genome sequences (WGS) of Mab demonstrates dense genetic clustering of clinical isolates collected from disparate geographic locations. This has been interpreted as supporting patient-to-patient transmission, but epidemiological studies have contradicted this interpretation. Here, we present evidence for a slowing of the Mab molecular clock rate coincident with the emergence of phylogenetic clusters. We performed phylogenetic inference using publicly available WGS from 483 Mab patient isolates. We implement a subsampling approach in combination with coalescent analysis to estimate the molecular clock rate along the long internal branches of the tree, indicating a faster long-term molecular clock rate compared to branches within phylogenetic clusters. We used ancestry simulation to predict the effects of clock rate variation on phylogenetic clustering and found that the degree of clustering in the observed phylogeny is more easily explained by a clock rate slowdown than by transmission. We also find that phylogenetic clusters are enriched in mutations affecting DNA repair machinery and report that clustered isolates have lower spontaneous mutation rates in vitro. We propose that Mab adaptation to the host environment through variation in DNA repair genes affects the organism's mutation rate and that this manifests as phylogenetic clustering. These results challenge the model that phylogenetic clustering in Mab is explained by person-to-person transmission and inform our understanding of transmission inference in emerging, facultative pathogens.

Phylogenetic analysis of Brachymystax and Hucho genera—Summary on evolutionary status within the Salmoninae subfamily

In this study comparative phylogenetic analyses of huchonid fishes (Brachymystax and Hucho genera) were carried out by application of various computational methods based on the supermatrix data retrieved from NCBI Gene Bank. For this purpose, the available mitochondrial (3575 bp) and nuclear (2037 bp) DNA data originated from all extant huchonid species were used. Additionally, molecular dating was carried out to estimate time background of huchonids divergence. Phylogenetic analyses were conducted using the Neighbor-joining (NJ), Maximum likelihood (ML), Maximum parsimony (MP) and Bayesian approach (BA) methods. The ancestral position of Brachymystax/Hucho cluster within Salmoninae subfamily was resolved and dated to emerge c.a. 24.7–25.6 million years ago (Mya). The evolutionary age of the most recent ancestor of contemporary huchonids was estimated to 10.6–13.6 Mya, while the oldest lineage within huchonid fishes was the genus Brachymystax (5.5–6.1 Mya). All three extant Hucho species were dated to split about 2.5 Mya, suggesting their disjunctive distribution during speciation. The highest heterogeneity was observed within Hucho taimen and Hucho bleekeri species, which clustered into two distinct phylogeographic groups, implying their vicariant origin from different glaciation refugees. The lowest level of genetic differentiation was recorded in Hucho hucho species, which may reflect past low population size, bottlenecks, founder effect and/or low rate of molecular clock. The carried out comparative analyses evidenced the discordance between nuclear and mitochondrial DNA data, indicating probable introgression between H. bleekeri and Brachymystax lenok that was estimated to take place around 5.5 Mya. The monotypic Parahucho genus was resolved as evolutionary younger and separate from huchonid lineage, that emerged around 15.8–13.2 Mya.

Quantifying bacterial evolution in the wild: A birthday problem for Campylobacter lineages.

Measuring molecular evolution in bacteria typically requires estimation of the rate at which nucleotide changes accumulate in strains sampled at different times that share a common ancestor. This approach has been useful for dating ecological and evolutionary events that coincide with the emergence of important lineages, such as outbreak strains and obligate human pathogens. However, in multi-host (niche) transmission scenarios, where the pathogen is essentially an opportunistic environmental organism, sampling is often sporadic and rarely reflects the overall population, particularly when concentrated on clinical isolates. This means that approaches that assume recent common ancestry are not applicable. Here we present a new approach to estimate the molecular clock rate in Campylobacter that draws on the popular probability conundrum known as the ‘birthday problem’. Using large genomic datasets and comparative genomic approaches, we use isolate pairs that share recent common ancestry to estimate the rate of nucleotide change for the population. Identifying synonymous and non-synonymous nucleotide changes, both within and outside of recombined regions of the genome, we quantify clock-like diversification to estimate synonymous rates of nucleotide change for the common pathogenic bacteria Campylobacter coli (2.4 x 10−6 s/s/y) and Campylobacter jejuni (3.4 x 10−6 s/s/y). Finally, using estimated total rates of nucleotide change, we infer the number of effective lineages within the sample time frame–analogous to a shared birthday–and assess the rate of turnover of lineages in our sample set over short evolutionary timescales. This provides a generalizable approach to calibrating rates in populations of environmental bacteria and shows that multiple lineages are maintained, implying that large-scale clonal sweeps may take hundreds of years or more in these species.

Regional effect on the molecular clock rate of protein evolution in Eutherian and Metatherian genomes

BackgroundDifferent types of proteins diverge at vastly different rates. Moreover, the same type of protein has been observed to evolve with different rates in different phylogenetic lineages. In the present study we measured the rates of protein evolution in Eutheria (placental mammals) and Metatheria (marsupials) on a genome-wide basis and we propose that the gene position in the genome landscape has an important influence on the rate of protein divergence.ResultsWe analyzed a protein-encoding gene set (n = 15,727) common to 16 mammals (12 Eutheria and 4 Metatheria). Using sliding windows that averaged regional effects of protein divergence we constructed landscapes in which strong and lineage-specific regional effects were seen on the molecular clock rate of protein divergence. Within each lineage, the relatively high rates were preferentially found in subtelomeric chromosomal regions. Such regions were observed to contain important and well-studied loci for fetal growth, uterine function and the generation of diversity in the adaptive repertoire of immunoglobulins.ConclusionsA genome landscape approach visualizes lineage-specific regional differences between Eutherian and Metatherian rates of protein evolution. This phenomenon of chromosomal position is a new element that explains at least part of the lineage-specific effects and differences between proteins on the molecular clock rates.

Meta-analysis of Genetic Diversity of the VP1 Gene Among the Circulating O, A, and SAT2 Serotypes and Vaccine Strains of FMD Virus in Egypt.

IntroductionThree strains of the FMD virus (A, O, and SAT 2) were recognised as causes of the FMD circulating in Egypt. The aims of this study were to trace the FMDV isolates from outbreaks in Egypt to understand their epidemiology and evolution and to understand the situation of the vaccine strains compared with the circulating serotypes.Material and MethodsA meta-analysis was carried out by using the data available for FMD outbreaks in Egypt from GenBank and the World Reference Laboratory for Foot-and-Mouth Disease (WRLFMD); a comparison was done with both data sets for the three serotypes. MEGA-X was used for the evolution analysis, through constructions of phylogenetic trees for all sequences recorded in GenBank for each serotype in different Egyptian outbreaks in different years and also within the same year. Additionally, nucleotide substitution rate, molecular clock, and mean evolutionary rates were estimated for the three serotypes to understand and compare their evolution.ResultsAbsence of some records of certain serotype outbreaks from the WRLFMD database was noted as were subsequent missing appropriate vaccine programmes. Genetic variation was recorded among the virus isolates within the same years and also the vaccine strain was associated with up to 26 amino acid substitutions. The evolution rate of the SAT2 strain was the highest of the circulating strains. SAT2 had high amino acid substitution per year at an important immunogenic site (130–170), serotype A had less, and serotype O the least.ConclusionThe need for different strategies for vaccine serotype selection is indicated.

Population genomics of louping ill virus provide new insights into the evolution of tick-borne flaviviruses.

The emergence and spread of tick-borne arboviruses pose an increased challenge to human and animal health. In Europe this is demonstrated by the increasingly wide distribution of tick-borne encephalitis virus (TBEV, Flavivirus, Flaviviridae), which has recently been found in the United Kingdom (UK). However, much less is known about other tick-borne flaviviruses (TBFV), such as the closely related louping ill virus (LIV), an animal pathogen which is endemic to the UK and Ireland, but which has been detected in other parts of Europe including Scandinavia and Russia. The emergence and potential spatial overlap of these viruses necessitates improved understanding of LIV genomic diversity, geographic spread and evolutionary history. We sequenced a virus archive composed of 22 LIV isolates which had been sampled throughout the UK over a period of over 80 years. Combining this dataset with published virus sequences, we detected no sign of recombination and found low diversity and limited evidence for positive selection in the LIV genome. Phylogenetic analysis provided evidence of geographic clustering as well as long-distance movement, including movement events that appear recent. However, despite genomic data and an 80-year time span, we found that the data contained insufficient temporal signal to reliably estimate a molecular clock rate for LIV. Additional analyses revealed that this also applied to TBEV, albeit to a lesser extent, pointing to a general problem with phylogenetic dating for TBFV. The 22 LIV genomes generated during this study provide a more reliable LIV phylogeny, improving our knowledge of the evolution of tick-borne flaviviruses. Our inability to estimate a molecular clock rate for both LIV and TBEV suggests that temporal calibration of tick-borne flavivirus evolution should be interpreted with caution and highlight a unique aspect of these viruses which may be explained by their reliance on tick vectors.

Phylogenetic relationships within Pseudamnicola Paulucci, 1878 (Caenogastropoda: Truncatelloidea) indicate two independent dispersal events from different continents to the Balearic Islands

Until recently, the evolutionary origin of aquatic organisms in the Balearic archipelago (western Mediterranean) had been scarcely addressed. The freshwater gastropod genus Pseudamnicola Paulucci, 1878 (Hydrobiidae), consisting of ∼60 species in the Mediterranean area, is widely distributed across the major islands of Majorca and Minorca. From phylogenetic datasets of limited geographic coverage, a recent well-supported split has been inferred between the Minorca and Iberian Pseudamnicola species, but the phylogenetic position of the Majorca clade composed of three species remained uncertain. In this study, we extend published sequence data for the COI and 16S rRNA mitochondrial genes and the 28S rRNA nuclear gene in western Mediterranean populations of Pseudamnicola with homologous sequences from individuals of the non-surveyed region of Morocco to assess the origin of the Balearic species from a wider geographic perspective. Our maximum likelihood and Bayesian inference analyses recovered a strongly supported clade comprising the Majorcan species and four phylogenetic lineages in Morocco, two of which are new to science (described here as P. ramosae sp. nov. and P. ouarzazatensis sp. nov.). Dating the nodes using an external molecular clock rate, younger ages were estimated for the Majorcan species (∼1 Mya) than for the Moroccan ones (∼5 Mya), which were still younger than the formation of the Balearic Islands (∼30 Mya). An independent well-supported clade grouped the Minorca and Iberian species diverging at ∼4 Mya. Thus, we found two independent evolutionary origins within the Balearic archipelago likely attributable to long-distance dispersal events from the African and Eurasian continents.http://zoobank.org:pub:24D17C1D-4A42-4722-9867-D8D84465F55Ahttp://zoobank.org:act:85EC4DF1-9BFA-4B28-B3F2-C091212AFCF1http://zoobank.org:act:C790EAC6-F22B-489A-93EB-CABF9EEAF282

Sequencing a Juglans regia\u2009\xd7\u2009J. microcarpa hybrid yields high-quality genome assemblies of parental species

Members of the genus Juglans are monecious wind-pollinated trees in the family Juglandaceae with highly heterozygous genomes, which greatly complicates genome sequence assembly. The genomes of interspecific hybrids are usually comprised of haploid genomes of parental species. We exploited this attribute of interspecific hybrids to avoid heterozygosity and sequenced an interspecific hybrid Juglans microcarpa × J. regia using a novel combination of single-molecule sequencing and optical genome mapping technologies. The resulting assemblies of both genomes were remarkably complete including chromosome termini and centromere regions. Chromosome termini consisted of arrays of telomeric repeats about 8 kb long and heterochromatic subtelomeric regions about 10 kb long. The centromeres consisted of arrays of a centromere-specific Gypsy retrotransposon and most contained genes, many of them transcribed. Juglans genomes evolved by a whole-genome-duplication dating back to the Cretaceous-Paleogene boundary and consist of two subgenomes, which were fractionated by numerous short gene deletions evenly distributed along the length of the chromosomes. Fractionation was shown to be asymmetric with one subgenome exhibiting greater gene loss than the other. The asymmetry of the process is ongoing and mirrors an asymmetry in gene expression between the subgenomes. Given the importance of J. microcarpa × J. regia hybrids as potential walnut rootstocks, we catalogued disease resistance genes in the parental genomes and studied their chromosomal distribution. We also estimated the molecular clock rates for woody perennials and deployed them in estimating divergence times of Juglans genomes and those of other woody perennials.

Reconstructing foot-and-mouth disease outbreaks: a methods comparison of transmission network models

A number of transmission network models are available that combine genomic and epidemiological data to reconstruct networks of who infected whom during infectious disease outbreaks. For such models to reliably inform decision-making they must be transparently validated, robust, and capable of producing accurate predictions within the short data collection and inference timeframes typical of outbreak responses. A lack of transparent multi-model comparisons reduces confidence in the accuracy of transmission network model outputs, negatively impacting on their more widespread use as decision-support tools. We undertook a formal comparison of the performance of nine published transmission network models based on a set of foot-and-mouth disease outbreaks simulated in a previously free country, with corresponding simulated phylogenies and genomic samples from animals on infected premises. Of the transmission network models tested, Lau’s systematic Bayesian integration framework was found to be the most accurate for inferring the transmission network and timing of exposures, correctly identifying the source of 73% of the infected premises (with 91% accuracy for sources with model support >0.80). The Structured COalescent Transmission Tree Inference provided the most accurate inference of molecular clock rates. This validation study points to which models might be reliably used to reconstruct similar future outbreaks and how to interpret the outputs to inform control. Further research could involve extending the best-performing models to explicitly represent within-host diversity so they can handle next-generation sequencing data, incorporating additional animal and farm-level covariates and combining predictions using Ensemble methods and other approaches.

Concordant patterns of mtDNA and nuclear phylogeographic structure reveal Pleistocene vicariant event in the green crab Carcinus aestuarii across the Siculo-Tunisian Strait

This study focuses on the population genetic structure of the green crab Carcinus aestuarii along part of the African Mediterranean coast, with the main target to confirm genetic subdivision across the well documented genetic boundary of the Siculo-Tunisian Strait. For this purpose, the mitochondrial COI (cytochrome oxidase I) gene and five polymorphic microsatellite loci were analysed in 144 and 120 specimens, respectively. Our results show the existence of two distinct haplogroups, separated by 16 mutational steps and revealed a non random distribution of the genetic variation along the African Mediterranean coast. Dating analyses, based on the use of different molecular clock models and rates, placed the divergence among both haplogroups at 1.91 Myr (95% HPD: 1.11–2.68 Myr) to 0.69 Myr (95% HPD: 0.44–0.98 Myr). This range of divergence time estimation corresponds to the Early Pleistocene. The particular pattern of genetic divergence among Eastern and Western African Mediterranean populations of C. aestuarii, detected by 2-level AMOVA at the mitochondrial level, was consistent with that inferred from microsatellite analysis and suggests a vicariant event in C. aestuarii. Demographic reconstruction, inferred from mismatch distribution and BSP analyses, yielded different patterns of demographic history between both African Mediterranean groups. The distribution pattern of the two haplogroups across the African Mediterranean coast, along with results of Bayesian analysis of genetic structure revealing an intermediate geographic group between the two divergent groups of the African coast, support the hypothesis of a secondary contact between two historically isolated groups. Although this hypothetical contact zone, thought to be located near the Siculo-Tunisian Strait, still needs to be verified, the asymmetric gene flow from Western to Eastern African Mediterranean, as inferred by the results of a MIGRATE analysis, reinforces the previously mentioned results.

Spatial and temporal evolution of Lassa virus in the natural host population in Upper Guinea.

This study aimed at reconstructing the spatial and temporal evolution of Lassa virus (LASV) in the natural host population. To this end, we generated 132 partial nucleoprotein sequences of LASV from M. natalensis trapped in 12 villages around Faranah, Upper Guinea, over a period of 12 years. This study reveals two main features of LASV evolution in M. natalensis. First, the virus evolves in the reservoir with a molecular clock rate of 9 (7–11) × 10–4 position–1 year–1 implying that contemporary LASV lineages circulate in the Faranah area since less than 100 years. Second, viruses circulating in a specific village are diverse and polyphyletic. We observed, however, there are monophyletic clusters at village and sub-village level at specific points in time. In conclusion, our data indicate that the temporal and spatial pattern of LASV evolution in the natural reservoir is characterized by a combination of stationary circulation within a village and virus movement between villages. The latter feature is relevant for rodent control strategies, as it implies that recurrence of the virus from neighbouring villages may occur in villages where the virus has previously been eradicated.

Epidemic history of major genotypes of hepatitis C virus in Uruguay

Worldwide, more than 170 million people are chronically infected with the hepatitis C virus (HCV) and every year die more than 350,000 people from HCV-related liver diseases. Recently, HCV was reclassified into seven major genotypes and 67 subtypes. Some subtypes as 1a, 1b and 3a, have become epidemic as a result of the new parenteral transmission routes and are responsible for most HCV infections in Western countries. HCV 1a subtype have been sub-categorized into two separate sub clades. Recent studies based on the analysis of NS5B genome region, reveal that HCV epidemics in Argentina and Brazil are characterized by multiple introductions events of subtypes 1a, 1b and 3a, followed by subsequent local dispersion. There is no data about HCV genotypes circulating in Uruguay and their evolutionary and demographic history. To this end, a total of 153 HCV NS5B gene sequences were obtained from Uruguayan patients between 2005 and 2011. 86 (56%) sequences grouped with subtype 1a, 40 (26%) with subtype 3a and 27 (18%) with subtype 1b. Furthermore, subtype 1a sequences were distributed among both clades, 1 (n=62, 72%) and 2 (n=24, 28%). Four local HCV clades were found: UY-1a(I), UY-1a(II), UY-1a(III) and UY-3a; comprising a 39% of all HCV viruses analyzed in this study. HCV epidemic in Uruguay has been driving by multiple introductions of subtypes 1a, 1b and 3a and by local dissemination of a few country-specific strains. The evolutionary and demographic history of the major Uruguayan HCV clade UY-1a(I) was reconstructed under two different molecular clock rate models and displayed an epidemic history characterized by an initial phase of rapid expansion followed by a more recent reduction of growth rate since 2000–2005. This is the first comprehensive study about the molecular epidemiology and epidemic history of HCV in Uruguay.

The molecular clock of neutral evolution can be accelerated or slowed by asymmetric spatial structure.

Over time, a population acquires neutral genetic substitutions as a consequence of random drift. A famous result in population genetics asserts that the rate, K, at which these substitutions accumulate in the population coincides with the mutation rate, u, at which they arise in individuals: K = u. This identity enables genetic sequence data to be used as a “molecular clock” to estimate the timing of evolutionary events. While the molecular clock is known to be perturbed by selection, it is thought that K = u holds very generally for neutral evolution. Here we show that asymmetric spatial population structure can alter the molecular clock rate for neutral mutations, leading to either K<u or K>u. Our results apply to a general class of haploid, asexually reproducing, spatially structured populations. Deviations from K = u occur because mutations arise unequally at different sites and have different probabilities of fixation depending on where they arise. If birth rates are uniform across sites, then K ≤ u. In general, K can take any value between 0 and Nu. Our model can be applied to a variety of population structures. In one example, we investigate the accumulation of genetic mutations in the small intestine. In another application, we analyze over 900 Twitter networks to study the effect of network topology on the fixation of neutral innovations in social evolution.

A molecular phylogeny of the harriers (Circus, Accipitridae) indicate the role of long distance dispersal and migration in diversification

The monophyly of the raptorial Circus genus (harriers) has never been in question, but the specific status of many, often vulnerable island endemic, taxa remains uncertain. Here we utilise one mitochondrial and three nuclear loci from all currently recognised Circus taxa (species and subspecies) to infer a robust phylogeny, to estimate the divergence date and to reconstruct the biogeographic origins of the Circus group. Our phylogeny supports both the monophyly of Circus and polyphyly of the genus Accipiter. Depending on the rate of molecular clock used, the emergence of the harrier clade took place between 4.9 and 12.2mya which coincides with the worldwide formation of open habitats which extant harriers now exploit. The sister relationship of the Northern Harrier C. cyaneus hudsonius and the Cinereous Harrier C. cinereus contradicts previous classifications that treated the former as conspecific with the Hen Harrier C. cyaneus cyaneus. Thus both should be elevated to species status: C. hudsonius and C. cyaneus. Further, the African Marsh C. ranivorus and the European Marsh C. aeruginosus Harriers emerge as sister species. The remaining marsh harriers exhibit very little genetic diversity, and are all recently diverged taxa that exhibit allopatric distributions. Considering their sister relationship and geographic proximity, we recommend treating C. approximans and C. spilonotus spilothorax as subspecies of C. approximans. For C. spilonotus spilonotus C. maillardi maillardi and C. maillardi macrosceles, their plumage and morphometric differences, phylogenetic relationship and geographic distributions make lumping of these taxa as a single species complicated. We thus propose to recognise as separate, recently evolved species: C. spilonotus, C. maillardi and C. macrosceles. Biogeographic inferences on the ancestral origin of harriers are uncertain, indicating that the harriers emerged in either the Neotropics, Palearctic or Australasia. We are, however, able to show that speciation within the harriers was driven by long range dispersal and migration events.

Time dependence of evolutionary metrics during the 2009 pandemic influenza virus outbreak.

With the expansion of DNA sequencing technology, quantifying evolution during emerging viral outbreaks has become an important tool for scientists and public health officials. Although it is known that the degree of sequence divergence significantly affects the calculation of evolutionary metrics in viral outbreaks, the extent and duration of this effect during an actual outbreak remains unclear. We have analyzed how limited divergence time during an early viral outbreak affects the accuracy of molecular evolutionary metrics. Using sequence data from the first 25 months of the 2009 pandemic H1N1 (pH1N1) outbreak, we calculated each of three different standard evolutionary metrics—molecular clock rate (i.e., evolutionary rate), whole-gene dN/dS, and site-wise dN/dS—for hemagglutinin and neuraminidase, using increasingly longer time windows, from 1 month to 25 months. For the molecular clock rate, we found that at least 3–4 months of temporal divergence from the start of sampling was required to make precise estimates that also agreed with long-term values. For whole-gene dN/dS, we found that at least 2 months of data were required to generate precise estimates, but 6–9 months were required for estimates to approach their long term values. For site-wise dN/dS estimates, we found that at least 6 months of sampling divergence was required before the majority of sites had at least one mutation and were thus evolutionarily informative. Furthermore, 8 months of sampling divergence was required before the site-wise estimates appropriately reflected the distribution of values expected from known protein-structure-based evolutionary pressure in influenza. In summary, we found that evolutionary metrics calculated from gene sequence data in early outbreaks should be expected to deviate from their long-term estimates for at least several months after the initial emergence and sequencing of the virus.

New insight into HCV E1/E2 region of genotype 4a.

IntroductionHepatitis C virus (HCV) genome contains two envelope proteins (E1 and E2) responsible for the virus entry into the cell. There is a substantial lack of sequences covering the full length of E1/E2 region for genotype 4. Our study aims at providing new sequences as well as characterizing the genetic divergence of the E1/E2 region of HCV 4a using our new sequences along with all publicly available datasets.MethodsThe genomic segments covering the whole E1/E2 region were isolated from Egyptian HCV patients and sequenced. The resulting 36 sequences 36 were analyzed using sequence analysis techniques to study variability within and among hosts in the same time point. Furthermore, previously published HCV E1/E2 sequence datasets for genotype 4a were retrieved and categorized according to the geographical location and date of isolation and were used for further analysis of variability among Egyptian over a period of 15 years, also compared with non-Egyptian sequences to figure out region-specific variability.ResultsPhylogenetic analysis of the new sequences has shown variability within the host and among different individuals in the same time point. Analysis of the 36 sequences along with the Egyptian sequences (254 sequences in E1 in the period from 1997 to 2010 and 8 E2 sequences in the period from 2006 to 2010) has shown temporal change over time. Analysis of the new HCV sequences with the non-Egyptian sequences (182 sequences in E1 and 155 sequences in the E2) has shown region specific variability. The molecular clock rate of E1 was estimated to be 5E-3 per site per year for Egyptian and 5.38E-3 for non-Egyptian. The clock rate of E2 was estimated to be 8.48E per site per year for Egyptian and 6.3E-3 for non-Egyptian.ConclusionThe results of this study support the high rate of evolution of the Egyptian HCV genotype 4a. It has also revealed significant level of genetic variability among sequences from different regions in the world.Electronic supplementary materialThe online version of this article (doi:10.1186/s12985-014-0231-y) contains supplementary material, which is available to authorized users.

Land-bridge calibration of molecular clocks and the post-glacial Colonization of Scandinavia by the Eurasian field vole Microtus agrestis.

Phylogeography interprets molecular genetic variation in a spatial and temporal context. Molecular clocks are frequently used to calibrate phylogeographic analyses, however there is mounting evidence that molecular rates decay over the relevant timescales. It is therefore essential that an appropriate rate is determined, consistent with the temporal scale of the specific analysis. This can be achieved by using temporally spaced data such as ancient DNA or by relating the divergence of lineages directly to contemporaneous external events of known time. Here we calibrate a Eurasian field vole (Microtus agrestis) mitochondrial genealogy from the well-established series of post-glacial geophysical changes that led to the formation of the Baltic Sea and the separation of the Scandinavian peninsula from the central European mainland. The field vole exhibits the common phylogeographic pattern of Scandinavian colonization from both the north and the south, however the southernmost of the two relevant lineages appears to have originated in situ on the Scandinavian peninsula, or possibly in the adjacent island of Zealand, around the close of the Younger Dryas. The mitochondrial substitution rate and the timescale for the genealogy are closely consistent with those obtained with a previous calibration, based on the separation of the British Isles from mainland Europe. However the result here is arguably more certain, given the level of confidence that can be placed in one of the central assumptions of the calibration, that field voles could not survive the last glaciation of the southern part of the Scandinavian peninsula. Furthermore, the similarity between the molecular clock rate estimated here and those obtained by sampling heterochronous (ancient) DNA (including that of a congeneric species) suggest that there is little disparity between the measured genetic divergence and the population divergence that is implicit in our land-bridge calibration.

A review of phylogeographic analyses of animal taxa from the Aegean and surrounding regions

We provide a review and synthesis of key findings in phylogeographic research on terrestrial animals in the Aegean archipelago (Greece) and surrounding regions (Greek mainland, southern Balkans and Asia Minor). A critical review of more than 100 phylogeographic articles on 76 animal genera (30 invertebrates and 46 vertebrates) that have been published so far for the region leads to the recognition of three types of distribution patterns: old ‘colonizers’ diversified in the Aegean before the formation of the mid-Aegean Trench (MAT – i.e. before 9 Mya), post-MAT colonizers that arrived to the Aegean in late Miocene and Pliocene and new colonizers that inhabited the region in the Pleistocene or even the Holocene. Several problems, mainly regarding the use of calibration points and/or molecular clock rates for clade chronology, have been identified in many analyses. Despite the large amount of phylogeographic work concerning the Aegean and surrounding regions, many groups remain unstudied (especially invertebrates, micro-organisms, fungi and plants), and many issues still remain unresolved. The relative roles of extinction, speciation, dispersal and vicariance, as well as the effects of adaptive and non-adaptive components of diversification, need further analyses with modern tools that can provide deeper insights. A more detailed reconstruction of the palaeogeography of the region is also of prime importance. The critical views presented herein may prove useful also for the evaluation of similar work in other regions.