Origins of social parasitism: the importance of divergence ages in phylogenetic studies

Phylogenetic studies on insect social parasites have found very close host-parasite relationships, and these have often been interpreted as providing evidence for sympatric speciation. However, such phylogenetic inferences are problematic because events occurring after the origin of parasitism, such as extinction, host switching and subsequent speciation, or an incomplete sampling of taxa, could all confound the interpretation of phylogenetic relationships. Using a tribe of bees where social parasitism has repeatedly evolved over a wide time-scale, we show the problems associated with phylogenetic inference of sympatric speciation. Host-parasite relationships of more ancient species appear to support sympatric speciation, whereas in a case where parasitism has evolved very recently, sympatric speciation can be ruled out. However, in this latter case, a single extinction event would have lead to relationships that support sympatric speciation, indicating the importance of considering divergence ages when analysing the modes of social parasite evolution.     

Impact of duplicate gene copies on phylogenetic analysis and divergence time estimates in butterflies

Background

Dentin sialophosphoprotein (DSPP) is the largest member of the SIBLING family and is the most abundant noncollagenous protein in dentin. DSPP is also expressed in non-mineralized tissues including metabolically active ductal epithelia and some cancers. Its function, however, is poorly defined. The carboxy-terminal fragment, dentin phosphoprotein (DPP) is encoded predominantly by a large repetitive domain that requires separate cloning/sequencing reactions and is, therefore, often incomplete in genomic databases. Comparison of DPP sequences from at least one member of each major branch in the mammalian evolutionary tree (including some "toothless" mammals) as well as one reptile and bird may help delineate its possible functions in both dentin and ductal epithelia.

Results

The BMP1-cleavage and translation-termination domains were sufficiently conserved to permit amplification/cloning/sequencing of most species' DPP. While the integrin-binding domain, RGD, was present in about half of species, only vestigial remnants of this tripeptide were identified in the others. The number of tandem repeats of the nominal SerSerAsp phosphorylation motif in toothed mammals (including baleen whale and platypus which lack teeth as adults), ranged from ~75 (elephant) to >230 (human). These repeats were not perfect, however, and patterns of intervening sequences highlight the rapidity of changes among even closely related species. Two toothless anteater species have evolved different sets of nonsense mutations shortly after their BMP1 motifs suggesting that while cleavage may be important for DSPP processing in other tissues, the DPP domain itself may be required only in dentin. The lizard DSPP had an intact BMP1 site, a remnant RGD motif, as well as a distinctly different Ser/Asp-rich domain compared to mammals.

Conclusions

The DPP domain of DSPP was found to change dramatically within mammals and was lost in two truly toothless animals. The defining aspect of DPP, the long repeating phosphorylation domain, apparently undergoes frequent slip replication and recombination events that rapidly change specific patterns but not its overall biochemical character in toothed animals. Species may have to co-evolve protein processing mechanisms, however, to handle increased lengths of DSP repeats. While the RGD domain is lost in many species, some evolutionary pressure to maintain integrin binding can be observed.     

Impact of duplicate gene copies on phylogenetic analysis and divergence time estimates in butterflies

Background  

The increase in availability of genomic sequences for a wide range of organisms has revealed gene duplication to be a relatively common event. Encounters with duplicate gene copies have consequently become almost inevitable in the context of collecting gene sequences for inferring species trees. Here we examine the effect of incorporating duplicate gene copies evolving at different rates on tree reconstruction and time estimation of recent and deep divergences in butterflies.     

Estimation of DNA sequence divergence from comparison of restriction endonuclease digests.

An estimation of the DNA sequence divergence between defined DNA secquences of individuals or species may be made from comparison by gel electrophoresis of restriction endonuclease digests. This analysis is applicable to purified DNA sequence of moderate complexity (1-100 X 10(6) daltons) which have diverged by base substitution of 0.5 to 25% of nucleotides.     

Making a robust biomolecular time scale for phylogenetic studies

The further evolution of informational molecular sequences should depend on the number of viable alternatives possible for the sequences as set by selection, the unrepaired mutation rate, and time. Most biomolecular clocks are based on Kimura's nearly neutral mutation random-drift hypothesis. This clock assumes that informational sequences are in equilibrium, i.e., the nucleotides mutate at a uniform rate and the number of nucleotides unconstrained by selection remains constant. Correcting for deviations from these assumptions should produce a more accurate clock. Informational molecules probably formed from polynucleotides having some other function such as nitrogen or nucleotide storage, thus being initially functionally unselected. At any time the rate of development of functionality in a protein may be expected to be proportional to the number of viable alternatives of sequence in its potentially interacting regions. Assuming the rate of unrepaired mutations is constant, these clocks should exponentially slow as they evolve, each with a different rate toward individual equilibria. Also if the degree of selection changes, its clock rate should change. For a more precise clock two approaches are suggested to estimate these time dependent changes in evolutionary rate. An improved clock could improve estimation of phylogeny and put a time scale on that phylogeny.     

Perspective: gene divergence, population divergence, and the variance in coalescence time in phylogeographic studies

Molecular methods as applied to the biogeography of single species (phylogeography) or multiple codistributed species (comparative phylogeography) have been productively and extensively used to elucidate common historical features in the diversification of the Earth's biota. However, only recently have methods for estimating population divergence times or their confidence limits while taking into account the critical effects of genetic polymorphism in ancestral species become available, and earlier methods for doing so are underutilized. We review models that address the crucial distinction between the gene divergence, the parameter that is typically recovered in molecular phylogeographic studies, and the population divergence, which is in most cases the parameter of interest and will almost always postdate the gene divergence. Assuming that population sizes of ancestral species are distributed similarly to those of extant species, we show that phylogeographic studies in vertebrates suggest that divergence of alleles in ancestral species can comprise from less than 10% to over 50% of the total divergence between sister species, suggesting that the problem of ancestral polymorphism in dating population divergence can be substantial. The variance in the number of substitutions (among loci for a given species or among species for a given gene) resulting from the stochastic nature of DNA change is generally smaller than the variance due to substitutions along allelic lines whose coalescence times vary due to genetic drift in the ancestral population. Whereas the former variance can be reduced by further DNA sequencing at a single locus, the latter cannot. Contrary to phylogeographic intuition, dating population divergence times when allelic lines have achieved reciprocal monophyly is in some ways more challenging than when allelic lines have not achieved monophyly, because in the former case critical data on ancestral population size provided by residual ancestral polymorphism is lost. In the former case differences in coalescence time between species pairs can in principle be explained entirely by differences in ancestral population size without resorting to explanations involving differences in divergence time. Furthermore, the confidence limits on population divergence times are severely underestimated when those for number of substitutions per site in the DNA sequences examined are used as a proxy. This uncertainty highlights the importance of multilocus data in estimating population divergence times; multilocus data can in principle distinguish differences in coalescence time (T) resulting from differences in population divergence time and differences in T due to differences in ancestral population sizes and will reduce the confidence limits on the estimates. We analyze the contribution of ancestral population size (theta) to T and the effect of uncertainty in theta on estimates of population divergence (tau) for single loci under reciprocal monophyly using a simple Bayesian extension of Takahata and Satta's and Yang's recent coalescent methods. The confidence limits on tau decrease when the range over which ancestral population size theta is assumed to be distributed decreases and when tau increases; they generally exclude zero when tau/(4Ne) > 1. We also apply a maximum-likelihood method to several single and multilocus data sets. With multilocus data, the criterion for excluding tau = 0 is roughly that l tau/(4Ne) > 1, where l is the number of loci. Our analyses corroborate recent suggestions that increasing the number of loci is critical to decreasing the uncertainty in estimates of population divergence time.     

Estimating divergence times in large phylogenetic trees

A new method, PATHd8, for estimating ultrametric trees from trees with edge (branch) lengths proportional to the number of substitutions is proposed. The method allows for an arbitrary number of reference nodes for time calibration, each defined either as absolute age, minimum age, or maximum age, and the tree need not be fully resolved. The method is based on estimating node ages by mean path lengths from the node to the leaves but correcting for deviations from a molecular clock suggested by reference nodes. As opposed to most existing methods allowing substitution rate variation, the new method smoothes substitution rates locally, rather than simultaneously over the whole tree, thus allowing for analysis of very large trees. The performance of PATHd8 is compared with other frequently used methods for estimating divergence times. In analyses of three separate data sets, PATHd8 gives similar divergence times to other methods, the largest difference being between crown group ages, where unconstrained nodes get younger ages when analyzed with PATHd8. Overall, chronograms obtained from other methods appear smoother, whereas PATHd8 preserves more of the heterogeneity seen in the original edge lengths. Divergence times are most evenly spread over the chronograms obtained from the Bayesian implementation and the clock-based Langley-Fitch method, and these two methods produce very similar ages for most nodes. Evaluations of PATHd8 using simulated data suggest that PATHd8 is slightly less precise compared with penalized likelihood, but it gives more sensible answers for extreme data sets. A clear advantage with PATHd8 is that it is more or less instantaneous even with trees having several thousand leaves, whereas other programs often run into problems when analyzing trees with hundreds of leaves. PATHd8 is implemented in freely available software.     

Molecular phylogenetic and divergence time estimation analyses of the sawfly subfamily Selandriinae (Hymenoptera: Tenthredinidae)

Selandriinae, a subfamily of family Tenthredinidae (Hymenoptera: Symphyta), comprises multiple tribes, each of which has a relationship with specific plant group. The host specificity of the Selandriinae taxa provides a good model to examine the coevolution between hosts and insects. However, few phylogenetic studies for the Selandriinae obscure the evolutionary scenario with their host‐plants. The present study is a molecular phylogenetic analysis of 19 selandriine species based on mitochondrial genes (12S: 461 sites, 16S: 262sites and COI: 495 sites) and nuclear genes (18S: 773 sites and 28S: 495 sites). The results suggested three of six studied tribes are genetically isolated. Moreover, estimation of the time of molecular divergence showed that the Selandriinae clearly diverged at the same time as their host‐plants (monocots and ferns). These results suggested that the Selandriinae species might have codiversified with their hosts.     

Estimation of coalescence probabilities and population divergence times from SNP data

We present a method called the G(A|B) method for estimating coalescence probabilities within population lineages from genome sequences when one individual is sampled from each population. Population divergence times can be estimated from these coalescence probabilities if additional assumptions about the history of population sizes are made. Our method is based on a method presented by Rasmussen et al. (2014) to test whether an archaic genome is from a population directly ancestral to a present-day population. The G(A|B) method does not require distinguishing ancestral from derived alleles or assumptions about demographic history before population divergence. We discuss the relationship of our method to two similar methods, one introduced by Green et al. (2010) and called the F(A|B) method and the other introduced by Schlebusch et al. (2017) and called the TT method. When our method is applied to individuals from three or more populations, it provides a test of whether the population history is treelike because coalescence probabilities are additive on a tree. We illustrate the use of our method by applying it to three high-coverage archaic genomes, two Neanderthals (Vindija and Altai) and a Denisovan.Subject terms: Rare variants, Evolutionary genetics

One of the goals of population genetics is to estimate the divergence time of isolated populations. We will review several methods that have been proposed and present a new method that is closely related to two existing methods. We will emphasize the assumptions made when using different methods. It will be useful to make the distinction between estimating coalescence probabilities within populations and estimating population divergence times. We will also introduce a test for a treelike population history based on our method.For distantly related populations, the numbers of mutational differences between sequences indicate relative times of divergence. Relative times are converted to absolute times by assuming a mutation rate. This method traces to Zuckerkandl and Pauling (1962, 1965) and has been used and refined extensively. This class of methods estimates genomic divergence times. Using it to estimate population or species divergence times assumes that those times are so large that the difference between them can be ignored.For recently diverged populations, the numbers of mutational differences probably do not provide a reliable estimate of population divergence times both because there may be too few mutations that differentiate populations and because the difference between the genomic and population divergence times may be substantial. To overcome this problem, Green et al. (2010) (in Supplement 14) introduced a method that accounts for the difference between genomic and population divergence. This method was used in later papers from the same group (Meyer et al. 2012; Prüfer et al. 2014, 2017).The Green et al. (2010) method is applicable when one genome is sampled from each of two populations. It depends on the statistic F(A|B), which is the fraction of sites in population A that carry the derived allele when that site is heterozygous in population B. Green et al. (2010) showed by simulation that the expectation of F(A|B) decreases roughly exponentially with the separation time of A and B. The rate of decrease depends on the history of population sizes both in B and in the population ancestral to A and B. Green et al. (2010) estimated population divergence times by interpolating their simulation results.More recently, Schlebusch et al. (2017), in Section 9.1 of their supplementary materials, introduced a similar method, called the TT method. Their method is based on analytic expressions for the configuration probabilities of SNPs that are polymorphic in the two populations. The TT method assumes that ancestral and derived alleles can be distinguished and the population before divergence was of constant size. The TT method is developed and elaborated on by Sjödin et al. (2020).In the present paper, we present a new method that is closely related to the F(A|B) and TT methods. We call it the G(A|B) method to emphasize its similarity to F(A|B). Our method is based on a method presented by Rasmussen et al. (2014) to test whether an ancient DNA sequence is from a population directly ancestral to a present-day population. We will show that our method provides a way to test whether the history of three or more populations is accurately represented by a population tree even if the demographic histories of those populations are not known.     

Sequence length bounds for resolving a deep phylogenetic divergence

In evolutionary biology, genetic sequences carry with them a trace of the underlying tree that describes their evolution from a common ancestral sequence. The question of how many sequence sites are required to recover this evolutionary relationship accurately depends on the model of sequence evolution, the substitution rate, divergence times and the method used to infer phylogenetic history. A particularly challenging problem for phylogenetic methods arises when a rapid divergence event occurred in the distant past. We analyse an idealised form of this problem in which the terminal edges of a symmetric four-taxon tree are some factor (λ) times the length of the interior edge. We determine an order λ² lower bound on the growth rate for the sequence length required to resolve the tree (independent of any particular branch length). We also show that this rate of sequence length growth can be achieved by existing methods (including the simple ‘maximum parsimony’ method), and compare these order λ² bounds with an order λ growth rate for a model that describes low-homoplasy evolution. In the final section, we provide a generic bound on the sequence length requirement for a more general class of Markov processes.     

Molecular evidence for deep phylogenetic divergence in Mandrillus sphinx

Mandrills (Mandrillus sphinx) are forest primates indigenous to western central Africa. Phylogenetic analysis of 267 base pairs (bp) of the cytochrome b gene from 53 mandrills of known and 17 of unknown provenance revealed two phylogeographical groups, with haplotypes differentiated by 2.6% comprising seven synonymous transitions. The distribution of the haplotypes suggests that the Ogooué River, Gabon, which bisects their range, separates mandrill populations in Cameroon and northern Gabon from those in southern Gabon. The haplotype distribution is also concordant with that of two known mandrill simian immunodeficiency viruses, suggesting that these two mandrill phylogroups have followed different evolutionary trajectories since separation.     

Extreme morphological divergence: phylogenetic position of a termite ectoparasite

Species of Termitaria are lesion-forming ectoparasites occurring worldwide on a diverse group of termites. The reduced thallus consists of a basal cell layer from which haustorial cells penetrate the termite and a darkly pigmented sporodochium. One species, Termitaria snyderi, has been the subject of several morphological studies, but its phylogenetic position has remained enigmatic. Here we provide evidence of a close relationship between T. snyderi and the morphologically distinct ascomycetes, Kathistes analemmoides and K. calyculata, based on phylogenetic analysis of molecular characters derived from portions of the nuclear-encoded small-subunit ribosomal RNA gene (ssu rDNA) and supplemental evidence from the ?-tubulin gene. Trees were derived using parsimony and maximum-likelihood criteria. Bayesian analysis and parsimony bootstrap methods were used to assess support for the tree nodes.     

Ultrastructural and phylogenetic studies on Blastocystis isolates from cockroaches

Four Blastocystis isolates from cockroaches were established and these isolates were morphologically confirmed as Blastocystis organisms by light and/or electron microscopy. As these isolates were morphologically indistinguishable from Blastocystis isolated from other animals, phylogenetic analyses were conducted using their small subunit ribosomal RNA genes. A analyses of these sequences with previously reported ones that had been classified into nine Blastocystis clades indicated the presence of a new clade that comprised only Blastocystis organisms from cockroaches (clade X). A clade comprised of amphibian and reptilian Blastocystis organisms (clade IX) was located at the basal position of the Blastocystis tree together with the common ancestor of Proteromonas and Protoopalina, clade X emerged after the divergences of these two basal clades and its branching position was clearly supported by bootstrap analysis.     

Estimation of population divergence times from non-overlapping genomic sequences: examples from dogs and wolves

Despite recent technological advances in DNA sequencing, incomplete coverage remains to be an issue in population genomics, in particular for studies that include ancient samples. Here, we describe an approach to estimate population divergence times for non-overlapping sequence data that is based on probabilities of different genealogical topologies under a structured coalescent model. We show that the approach can be adapted to accommodate common problems such as sequencing errors and postmortem nucleotide misincorporations, and we use simulations to investigate biases involved with estimating genealogical topologies from empirical data. The approach relies on three reference genomes and should be particularly useful for future analysis of genomic data that comprise of nonoverlapping sets of sequences, potentially from different points in time. We applied the method to shotgun sequence data from an ancient wolf together with extant dogs and wolves and found striking resemblance to previously described fine-scale population structure among dog breeds. When comparing modern dogs to four geographically distinct wolves, we find that the divergence time between dogs and an Indian wolf is smallest, followed by the divergence times to a Chinese wolf and a Spanish wolf, and a relatively long divergence time to an Alaskan wolf, suggesting that the origin of modern dogs is somewhere in Eurasia, potentially southern Asia. We find that less than two-thirds of all loci in the boxer and poodle genomes are more similar to each other than to a modern gray wolf and that--assuming complete isolation without gene flow--the divergence time between gray wolves and modern European dogs extends to 3,500 generations before the present, corresponding to approximately 10,000 years ago (95% confidence interval [CI]: 9,000-13,000). We explicitly study the effect of gene flow between dogs and wolves on our estimates and show that a low rate of gene flow is compatible with an even earlier domestication date ～30,000 years ago (95% CI: 15,000-90,000). This observation is in agreement with recent archaeological findings and indicates that human behavior necessary for domestication of wild animals could have appeared much earlier than the development of agriculture.     

Early diverging Ascomycota: phylogenetic divergence and related evolutionary enigmas

The early diverging Ascomycota lineage, detected primarily from nSSU rDNA sequence-based phylogenetic analyses, includes enigmatic key taxa important to an understanding of the phylogeny and evolution of higher fungi. At the moment six representative genera of early diverging ascomycetes (i.e. Taphrina, Protomyces, Saitoella, Schizosaccharomyces, Pneumocystis and Neolecta) have been assigned to "Archiascomycetes" sensu Nishida and Sugi ama (1994) or the subphylum "Taphrinomycotina" sensu Eriksson and Winka (1997). The group includes fungi that are ecologically and morphologically diverse, and it is difficult therefore to define the group based on common phenotypic characters. Bayesian analyses of nSSU rDNA or combined nSSU and nLSU rDNA sequences supported previously published Ascomycota frameworks that consist of three major lineages (i.e. a group of early diverging Ascomycota. [Taphrinomycotina], Saccharomycotina and Pezizomycotina); Taphrinomycotina is the sister group of Saccharomycotina and Pezizomycotina. The 50% majority rule consensus of 18000 Bayesian MCMCMC-generated trees from multilocus gene sequences of nSSU rDNA, nLSU rDNA (D1/D2), RPB2 and beta-tubulin also showed the monophyly of the three subphyla and the basal position of Taphrinomycotina in Ascomycota with significantly higher statistical support. However to answer controversial questions on the origin, monophyly and evolution of the Taphrinomycotina, additional integrated phylogenetic analyses might be necessary using sequences of more genes with broader taxon sampling from the early diverging Ascomycota.     

Precision and accuracy of divergence time estimates from STR and SNPSTR variation

Inference of intraspecific population divergence patterns typically requires genetic data for molecular markers with relatively high mutation rates. Microsatellites, or short tandem repeat (STR) polymorphisms, have proven informative in many such investigations. These markers are characterized, however, by high levels of homoplasy and varying mutational properties, often leading to inaccurate inference of population divergence. A SNPSTR is a genetic system that consists of an STR polymorphism closely linked (typically < 500 bp) to one or more single-nucleotide polymorphisms (SNPs). SNPSTR systems are characterized by lower levels of homoplasy than are STR loci. Divergence time estimates based on STR variation (on the derived SNP allele background) should, therefore, be more accurate and precise. We use coalescent-based simulations in the context of several models of demographic history to compare divergence time estimates based on SNPSTR haplotype frequencies and STR allele frequencies. We demonstrate that estimates of divergence time based on STR variation on the background of a derived SNP allele are more accurate (3% to 7% bias for SNPSTR versus 11% to 20% bias for STR) and more precise than STR-based estimates, conditional on a recent SNP mutation. These results hold even for models involving complex demographic scenarios with gene flow, population expansion, and population bottlenecks. Varying the timing of the mutation event generating the SNP revealed that estimates of divergence time are sensitive to SNP age, with more recent SNPs giving more accurate and precise estimates of divergence time. However, varying both mutational properties of STR loci and SNP age demonstrated that multiple independent SNPSTR systems provide less biased estimates of divergence time. Furthermore, the combination of estimates based separately on STR and SNPSTR variation provides insight into the age of the derived SNP alleles. In light of our simulations, we interpret estimates from data for human populations.