David C. Marshall

Cryptic Failure of Partitioned Bayesian Phylogenetic Analyses: Lost in the Land of Long Trees

Partitioned Bayesian phylogenetic analyses of routine genetic data sets, constructed using MrBayes (Ronquist and Huelsenbeck 2003), can become trapped in regions of parameter space characterized by unrealistically long trees and distorted partition rate multipliers. Such analyses commonly fail to reach stationarity during hundreds of millions of generations of sampling-many times longer than most published analyses. Some data sets are so prone to this problem that paired MrBayes runs begun from different starting trees repeatedly find the same incorrect long-tree solutions and consequently pass the most commonly employed tests of stationarity, including the average standard deviation of split frequencies (ASDSF) and the potential scale reduction factor (PSRF) statistics offered by MrBayes (Gelman and Rubin 1992). In these situations, failure to reach stationarity is recognizable only in light of prior knowledge of model parameters, such as the expectation that third-codon-position sites usually evolve fastest in protein-coding genes. The conditions that lead to the long-tree problem are frequently encountered in phylogenetic studies today, and I present 6 demonstration examples from the literature. Although the effects on tree length (TL) are often dramatic, effects on topology appear to be subtle. Susceptibility to the problem is sometimes predicted by the difference between the true TL and the starting TL. In some cases, the problems described here can be avoided or reduced by manipulation of the starting TL and/or by adjustments to the prior on branch lengths. In more difficult situations, accurate branch length estimation may not be possible with Bayesian methods because of dependence of the solution on the branch length prior.

Differentiating between Hypotheses of Lineage Sorting and Introgression in New Zealand Alpine Cicadas (Maoricicada Dugdale)

Lineage sorting and introgression can lead to incongruence among gene phylogenies, complicating the inference of species trees for large groups of taxa that have recently and rapidly radiated. In addition, it can be difficult to determine which of these processes is responsible for this incongruence. We explore these issues with the radiation of New Zealand alpine cicadas of the genus Maoricicada Dugdale. Gene trees were estimated from four putative independent loci: mitochondrial DNA (2274 nucleotides), elongation factor 1-alpha (1275 nucleotides), period (1709 nucleotides), and calmodulin (678 nucleotides). We reconstructed phylogenies using maximum likelihood and Bayesian methods from 44 individuals representing the 19 species and subspecies of Maoricicada and two outgroups. Species-level relationships were reconstructed using a novel extension of gene tree parsimony, whereby gene trees were weighted by their Bayesian posterior probabilities. The inferred gene trees show marked incongruence in the placement of some taxa, especially the enigmatic forest and scrub dwelling species, M. iolanthe. Using the species tree estimated by gene tree parsimony, we simulated coalescent gene trees in order to test the null hypothesis that the nonrandom placement of M. iolanthe among gene trees has arisen by chance. Under the assumptions of constant population size, known generation time, and panmixia, we were able to reject this null hypothesis. Furthermore, because the two alternative placements of M. iolanthe are in each case with species that share a similar song structure, we conclude that it is more likely that an ancient introgression event rather than lineage sorting has caused this incongruence.

Allochronic speciation, secondary contact, and reproductive character displacement in periodical cicadas (Hemiptera: Magicicada spp.): genetic, morphological, and behavioural evidence

Periodical cicadas have proven useful in testing a variety of ecological and evolutionary hypotheses because of their unusual life history, extraordinary abundance, and wide geographical range. Periodical cicadas provide the best examples of synchronous periodicity and predator satiation in the animal kingdom, and are excellent illustrations of habitat partitioning (by the three morphologically distinct species groups), incipient species (the year classes or broods), and cryptic species (a newly discovered 13‐year species, Magicicada neotredecim). They are particularly useful for exploring questions regarding speciation via temporal isolation, or allochronic speciation. Recently, data were presented that provided strong support for an instance of allochronic speciation by life‐cycle switching. This speciation event resulted in the formation of a new 13‐year species from a 17‐year species and led to secondary contact between two formerly separated lineages, one represented by the new 13‐year cicadas (and their 17‐year ancestors), and the other represented by the pre‐existing 13‐year cicadas. Allozyme frequency data, mitochondrial DNA (mtDNA), and abdominal colour were shown to be correlated genetic markers supporting the life‐cycle switching/allochronic speciation hypothesis. In addition, a striking pattern of reproductive character displacement in male call pitch and female pitch preference between the two 13‐year species was discovered. In this paper we report a strong association between calling song pitch and mtDNA haplotype for 101 individuals from a single locality within the M. tredecim/M. neotredecim contact zone and a strong association between abdomen colour and mtDNA haplotype. We conclude by reviewing proposed mechanisms for allochronic speciation and reproductive character displacement.

Steady Plio-Pleistocene diversification and a 2-million-year sympatry threshold in a New Zealand cicada radiation.

Estimation of diversification rates in evolutionary radiations requires a complete accounting of cryptic species diversity. The rapidly evolving songs of acoustically signaling insects make them good model organisms for such studies. This paper examines the timing of diversification of a large (30 taxon) group of New Zealand cicadas (genus Kikihia Dugdale). We use Bayesian relaxed-clock methods and phylogenetic trees based on nuclear and mitochondrial DNA data, and we apply alternative combinations of evolutionary rate priors and geological calibrations. The extant Kikihia taxa began to diversify near the Miocene/Pliocene boundary around the time of increased mountain-building, and both the mitochondrial and nuclear-gene trees confirm early splits of lineages currently represented by lowland forest-dwelling taxa. Most lineages originated in the Pleistocene, and sustained diversification occurred rapidly at over 0.5 lineages/my, a rate comparable to that of the Hawaiian silverswords. Diversification rate tests suggest an increase in the early to mid-Pliocene, followed by constant diversification from the Late Pliocene onward. No descendants of the many Pleistocene-age splits have evolved the ability to coexist in sympatry, and, where they do come into contact, hybrid zones have been documented based on acoustic and DNA evidence. In contrast, lineages separated in time by approximately 2Myr often overlap in distribution with no evidence of hybridization. This suggests that at least 2Myr has been required to achieve the level of divergence required for reproductive isolation.

Glacial refugia in a maritime temperate climate: Cicada (Kikihia subalpina) mtDNA phylogeography in New Zealand

Understanding the biological significance of Pleistocene glaciations requires knowledge of the nature and extent of habitat refugia during glacial maxima. An opportunity to examine evidence of glacial forest refugia in a maritime, Southern Hemisphere setting is found in New Zealand, where the extent of Pleistocene forests remains controversial. We used the mitochondrial phylogeography of a forest‐edge cicada (Kikihia subalpina) to test the hypothesis that populations of this species survived throughout South Island during the Last Glacial Maximum. We also compared mitochondrial DNA phylogeographic patterns with male song patterns that suggest allopatric divergence across Cook Strait. Cytochrome oxidase I and II sequences were analyzed using network analysis, maximum‐likelihood phylogenetic estimation, Bayesian dating and Bayesian skyline plots. K. subalpina haplotypes from North Island and South Island form monophyletic clades that are concordant with song patterns. Song divergence corresponds to approximately 2% genetic divergence, and Bayesian dating suggests that the North Island and South Island population‐lineages became isolated around 761 000 years bp. Almost all South Island genetic variation is found in the north of the island, consistent with refugia in Marlborough Sounds, central Nelson and northwest Nelson. All central and southern South Island and Stewart Island haplotypes are extremely similar to northern South Island haplotypes, a ‘northern richness/southern purity’ pattern that mirrors genetic patterns observed in many Northern Hemisphere taxa. Proposed southern South Island forest habitat fragments may have been too small to sustain populations of K. subalpina, and/or they may have harboured ecological communities with no modern‐day analogues.

Temporal Separation and Speciation in Periodical Cicadas

Abstract Speciation, the set of processes by which two populations of one species become distinct species, is an important topic in evolutionary biology. It is usually impractical to conduct experiments on how new species form, but occasionally the natural history of a species places it in a context that may be thought of as a “natural experiment” with regard to speciation. One such natural experiment involves the periodical cicadas of eastern North America, a group in which populations have become isolated in time and space. Some of these isolated populations appear to have evolved into distinct genetic lineages. A rare life-cycle switching event brought two such lineages into contact in the relatively recent past, and the two lineages are now behaving as distinct species. This natural experiment provides important insights into species differences and the processes that underlie species formation.

Versatile aggressive mimicry of cicadas by an Australian predatory katydid.

Background In aggressive mimicry, a predator or parasite imitates a signal of another species in order to exploit the recipient of the signal. Some of the most remarkable examples of aggressive mimicry involve exploitation of a complex signal-response system by an unrelated predator species. Methodology/Principal Findings We have found that predatory Chlorobalius leucoviridis katydids (Orthoptera: Tettigoniidae) can attract male cicadas (Hemiptera: Cicadidae) by imitating the species-specific wing-flick replies of sexually receptive female cicadas. This aggressive mimicry is accomplished both acoustically, with tegminal clicks, and visually, with synchronized body jerks. Remarkably, the katydids respond effectively to a variety of complex, species-specific Cicadettini songs, including songs of many cicada species that the predator has never encountered. Conclusions/Significance We propose that the versatility of aggressive mimicry in C. leucoviridis is accomplished by exploiting general design elements common to the songs of many acoustically signaling insects that use duets in pair-formation. Consideration of the mechanism of versatile mimicry in C. leucoviridis may illuminate processes driving the evolution of insect acoustic signals, which play a central role in reproductive isolation of populations and the formation of species.

Reconstructing asymmetrical reproductive character displacement in a periodical cicada contact zone.

Selection against costly reproductive interactions can lead to reproductive character displacement (RCD). We use information from patterns of displacement and inferences about predisplacement character states to investigate causes of RCD in periodical cicadas. The 13‐year periodical cicada Magicicada neotredecim exhibits RCD and strong reproductive isolation in sympatry with a closely related 13‐year species, Magicicada tredecim. Displacement is asymmetrical, because no corresponding pattern of character displacement exists within M. tredecim. Results from playback and hybridization experiments strongly suggest that sexual interactions between members of these species were possible at initial contact. Given these patterns, we evaluate potential sources of selection for displacement. One possible source is ‘acoustical interference’, or mate‐location inefficiencies caused by the presence of heterospecifics. Acoustical interference combined with the species‐specificity of song pitch and preference appears to predict the observed asymmetrical pattern of RCD in Magicicada. However, acoustical interference does not appear to be a complete explanation for displacement in Magicicada, because our experiments suggest a significant potential for direct sexual interactions between these species before displacement. Another possible source of selection for displacement is hybrid failure. We evaluate the attractiveness of inferred hybrid mating signals, and we examine the viability of hybrid eggs. Neither of these shows strong evidence of hybrid inferiority. We conclude by presenting a model of hybrid failure related to life cycle differences in Magicicada.

Developmental Plasticity of Life-Cycle Length in Thirteen-Year Periodical Cicadas (Hemiptera: Cicadidae)

ABSTRACT Speciation in periodical cicadas (Magicicada Davis) is closely tied to changes in life-cycle length, which presents a paradox because these organisms depend on emergence synchrony for survival. Recently proposed speciation models invoke developmental plasticity as a possible solution: Environmentally triggered “4-yr accelerations” occur in 17-yr cicadas, suggesting that canalization of induced plasticity could change 17-yr populations into temporally isolated 13-yr populations. However, the reverse shift, 13-yr cicadas emerging in 17 yr, has never been documented. We searched 4 yr after the normal emergence of a 13-yr brood (and in a year with no expected periodical cicada emergences anywhere) and found periodical cicadas active at 26 of 92 sites, with examples of all four 13-yr species. At one location, we found evidence of at least 1,724 cicadas per ha emerging. Few males were heard singing at most sites, so these off-schedule cicadas apparently did not survive long in the face of predation. We also found one 13-yr species singing 8 yr late within the range of a different 13-yr brood, suggesting an 8-yr delayed emergence or consecutive generations of 4-yr delayed cicadas. Developmental plasticity in life-cycle length seems to be similar in 13- and 17-yr cicadas—both types possess the ability to switch to the opposite life cycle and to emerge 1 yr early and/or late. The confirmation of a reverse life-cycle switch in 13- cicadas suggests improvements to theories of life-cycle evolution in Magicicada and strengthens the case for developmental plasticity in speciation.

Inflation of Molecular Clock Rates and Dates: Molecular Phylogenetics, Biogeography, and Diversification of a Global Cicada Radiation from Australasia (Hemiptera: Cicadidae: Cicadettini)

Dated phylogenetic trees are important for studying mechanisms of diversification, and molecular clocks are important tools for studies of organisms lacking good fossil records. However, studies have begun to identify problems in molecular clock dates caused by uncertainty of the modeled molecular substitution process. Here we explore Bayesian relaxed-clock molecular dating while studying the biogeography of ca. 200 species from the global cicada tribe Cicadettini. Because the available fossils are few and uninformative, we calibrate our trees in part with a cytochrome oxidase I (COI) clock prior encompassing a range of literature estimates for arthropods. We show that tribe-level analyses calibrated solely with the COI clock recover extremely old dates that conflict with published estimates for two well-studied New Zealand subclades within Cicadettini. Additional subclade analyses suggest that COI relaxed-clock rates and maximum-likelihood branch lengths become inflated relative to EF-1[Formula: see text] intron and exon rates and branch lengths as clade age increases. We present corrected estimates derived from: (i) an extrapolated EF-1[Formula: see text] exon clock derived from COI-calibrated analysis within the largest New Zealand subclade; (ii) post hoc scaling of the tribe-level chronogram using results from subclade analyses; and (iii) exploitation of a geological calibration point associated with New Caledonia. We caution that considerable uncertainty is generated due to dependence of substitution estimates on both the taxon sample and the choice of model, including gamma category number and the choice of empirical versus estimated base frequencies. Our results suggest that diversification of the tribe Cicadettini commenced in the early- to mid-Cenozoic and continued with the development of open, arid habitats in Australia and worldwide. We find that Cicadettini is a rare example of a global terrestrial animal group with an Australasian origin, with all non-Australasian genera belonging to two distal clades. Within Australia, we show that Cicadettini is more widely distributed than any other cicada tribe, diverse in temperate, arid and monsoonal habitats, and nearly absent from rainforests. We comment on the taxonomic implications of our findings for thirteen cicada genera.