Phylogenetic Relationships of the Mutualistic Fungi Associated with Macrotermes subhyalinus in Oman.

The symbiotic association between fungus-gardening termites Macrotermes and its fungal symbiont has a moderate degree of specificity-although the symbiotic fungi (Termitomyces) form a monophyletic clade, there is not a one-to-one association between termite species and their fungus-garden associates. Here, we aim to determine the origin and phylogenetic relationships of Termitomyces in Oman. We used sequences of the internal transcribed spacer region (ITS) and the nuclear large subunit ribosomal RNA (LSU rRNA, 25S) gene and analyzed these with sequences of Termitomyces from other geographic areas. We find no evidence for more than a single colonization of Oman by Termitomyces. Unexpectedly, we find Termitomyces in Oman is most closely related to the symbiont of M. subhyalinus in West Africa rather than to those of geographically closer populations in East Africa.

Are asymmetric inheritance systems an evolutionary trap? Transitions in the mechanism of paternal genome loss in the scale insect family Eriococcidae.

Haplodiploidy and paternal genome elimination (PGE) are examples of asymmetric inheritance, where males transmit only maternally inherited chromosomes to their offspring. Under haplodiploidy, this results from males being haploid, whereas under PGE, males inherit but subsequently exclude paternally inherited chromosomes from sperm. Their evolution involves changes in the mechanisms of meiosis and sex determination and sometimes also dosage compensation. As a result, these systems are thought to be an evolutionary trap, meaning that once asymmetric chromosome transmission evolves, it is difficult to transition back to typical Mendelian transmission. We assess whether there is evidence for this idea in the scale insect family Eriococcidae, a lineage with PGE and the only clade with a suggestion that asymmetric inheritance has transitioned back to Mendelian inheritance. We conduct a cytological survey of 13 eriococcid species, and a cytological, genetic, and gene expression analysis of species in the genus Cystococcus, to investigate whether there is evidence for species in this family evolving Mendelian chromosome transmission. Although we find that all species we examined exhibit PGE, the mechanism is extremely variable within Eriococcidae. Within Cystococcus, in fact, we uncover a previously undiscovered type of PGE in scale insects that acts exclusively in meiosis, where paternally inherited chromosomes in males are present, uncondensed, and expressed in somatic cells but eliminated prior to meiosis. Broadly, we fail to find evidence for a reversion from PGE to Mendelian inheritance in Eriococcidae, supporting the idea that asymmetric inheritance systems such as PGE may be an evolutionary trap.

Host phylogeny and ecological associations best explain Wolbachia host shifts in scale insects.

Wolbachia are among the most prevalent and widespread endosymbiotic bacteria on Earth. Wolbachia's success in infecting an enormous number of arthropod species is attributed to two features: the range of phenotypes they induce in their hosts, and their ability to switch between host species. Whilst much progress has been made in elucidating their induced phenotypes, our understanding of Wolbachia host-shifting is still very limited: we lack answers to even fundamental questions concerning Wolbachia's routes of transfer and the importance of factors influencing host shifts. Here, we investigate the diversity and host-shift patterns of Wolbachia in scale insects, a group of arthropods with intimate associations with other insects that make them well suited to studying host shifts. Using Illumina multitarget amplicon sequencing of Wolbachia-infected scale insects and their direct associates we determined the identity of all Wolbachia strains. We then fitted a generalized additive mixed model to our data to estimate the influence of host phylogeny and the geographical distribution on Wolbachia strain sharing among scale insect species. The model predicts no significant contribution of host geography but strong effects of host phylogeny, with high rates of Wolbachia sharing among closely related species and a sudden drop-off in sharing with increasing phylogenetic distance. We also detected the same Wolbachia strain in scale insects and several intimately associated species (ants, wasps and flies). This indicates putative host shifts and potential routes of transfers via these associates and highlights the importance of ecological connectivity in Wolbachia host-shifting.

Wolbachia in scale insects: a distinct pattern of infection frequencies and potential transfer routes via ant associates.

Wolbachia is one of the most successful endosymbiotic bacteria of arthropods. Known as the 'master of manipulation', Wolbachia can induce a wide range of phenotypes in its host that can have far-reaching ecological and evolutionary consequences and may be exploited for disease and pest control. However, our knowledge of Wolbachia's distribution and the infection rate is unevenly distributed across arthropod groups such as scale insects. We fitted a distribution of within-species prevalence of Wolbachia to our data and compared it to distributions fitted to an up-to-date dataset compiled from surveys across all arthropods. The estimated distribution parameters indicate a Wolbachia infection frequency of 43.6% (at a 10% prevalence threshold) in scale insects. Prevalence of Wolbachia in scale insects follows a distribution similar to exponential decline (most species are predicted to have low prevalence infections), in contrast to the U-shaped distribution estimated for other taxa (most species have a very low or very high prevalence). We observed no significant associations between Wolbachia infection and scale insect traits. Finally, we screened for Wolbachia in scale insect's ecological associates. We found a positive correlation between Wolbachia infection in scale insects and their ant associates, pointing to a possible route of horizontal transfer of Wolbachia.

Identifying genetic markers for a range of phylogenetic utility-From species to family level.

Resolving the phylogenetic relationships of closely related species using a small set of loci is challenging as sufficient information may not be captured from a limited sample of the genome. Relying on few loci can also be problematic when conflict between gene-trees arises from incomplete lineage sorting and/or ongoing hybridization, problems especially likely in recently diverged lineages. Here, we developed a method using limited genomic resources that allows identification of many low copy candidate loci from across the nuclear and chloroplast genomes, design probes for target capture and sequence the captured loci. To validate our method we present data from Eucalyptus and Melaleuca, two large and phylogenetically problematic genera within the Myrtaceae family. With one annotated genome, one transcriptome and two whole-genome shotgun sequences of one Eucalyptus and four Melaleuca species, respectively, we identified 212 loci representing 263 kbp for targeted sequence capture and sequencing. Of these, 209 were successfully tested from 47 samples across five related genera of Myrtaceae. The average percentage of reads mapped back to the reference was 57.6% with coverage of more than 20 reads per position across 83.5% of the data. The methods developed here should be applicable across a large range of taxa across all kingdoms. The core methods are very flexible, providing a platform for various genomic resource availabilities and are useful from shallow to deep phylogenies.

Turnover of southern cypresses in the post-Gondwanan world: extinction, transoceanic dispersal, adaptation and rediversification.

Cupressaceae subfamily Callitroideae has been an important exemplar for vicariance biogeography, but its history is more than just disjunctions resulting from continental drift. We combine fossil and molecular data to better assess its extinction and, sometimes, rediversification after past global change. Key fossils were reassessed and their phylogenetic placement for calibration was determined using trait mapping and Bayes Factors. Five vicariance hypotheses were tested by comparing molecular divergence times with the timing of tectonic rifting. The role of adaptation to fire (serotiny) in its spread across a drying Australia was tested for Callitris. Our findings suggest that three transoceanic disjunctions within the Callitroideae probably arose from long-distance dispersal. A signature of extinction, centred on the end-Eocene global climatic chilling and drying, is evident in lineages-through-time plots and in the fossil record. Callitris, the most diverse extant callitroid genus, suffered extinctions but surviving lineages adapted and re-radiated into dry, fire-prone biomes that expanded in the Neogene. Serotiny, a key adaptation to fire, likely evolved in Callitris coincident with the biome shift. Both extinction and adaptive shifts have probably played major roles in this chronicle of turnover and renewal, but better understanding of biogeographical history requires improved taxonomy of fossils.

A newly recognised species of Cryptes Maskell 1892 (Hemiptera: Coccidae) from Western Australia.

Cryptes utzoni Lin, Kondo Cook sp. n. (Hemiptera: Coccidae) is described based on adult female morphology and DNA sequences from mitochondrial and nuclear loci. This Australian endemic species was found on the stem of Acacia aneura (Fabaceae) in Western Australia. All phylogenetic analyses of three independent DNA loci show that C. utzoni is closely related to C. baccatus (Maskell), the type and only species of Cryptes Maskell, 1892. The adult female of C. utzoni is described and illustrated and a table is provided of the characters that differ among adult females of the two species of Cryptes now recognised (C. baccatus and C. utzoni) and a morphologically similar Western Australian species, Austrolichtensia hakearum (Fuller). There is deep genetic divergence in COI among samples of C. baccatus, suggesting the possibility of a species complex in this taxon.

Nomenclatural changes in the Australasian gall-inducing genus Apiomorpha Rübsaamen (Hemiptera: Coccomorpha: Eriococcidae).

Apiomorpha Rübsaamen, 1894 was erected as a replacement name for Brachyscelis Schrader, 1863 that was preoccupied in the Coleoptera (Chrysomelidae: Brachyscelis Germar, 1834). Apiomorpha is a genus of eriococcid scale insects that induce galls on Eucalyptus (Myrtaceae) in Australia and New Guinea (Szent-Ivany & Womersley 1962; Gullan 1984; Gullan et al. 2005). In his original description of the genus, Schrader (1863a) included six species, of which B. citricola Schrader was subsequently recognised as a nomen nudum (Froggatt 1921). Among the other five, B. pileata Schrader was later designated as the type species of Apiomorpha by Lindinger (1937). Also amongst these five was B. ovicola Schrader, for which Schrader described and illustrated galls of males and females on twigs and leaves of Eucalyptus haemastoma (Schrader 1863a, plate II, figs a, e) in or near Sydney, New South Wales (NSW). He stated that his species names reflected the shape of the galls of adult females; hence those of B. ovicola can be interpreted as being egg-shaped and were illustrated as such by Schrader (1863a). Galls of males of B. ovicola he described and illustrated as trumpet-shaped.

A newly recognised Australian endemic species of Austrolecanium Gullan & Hodgson 1998 (Hemiptera: Coccidae) from Queensland.

Austrolecanium cryptocaryae Lin & Cook sp. n. is described based on adult female morphology and DNA sequences from mitochondrial and nuclear loci. This Australian endemic species was found on the underside of leaves of Cryptocarya microneura (Lauraceae) in Queensland. All phylogenetic analyses of four independent DNA loci and a concatenated dataset show that A. cryptocaryae is monophyletic and closely related to A. sassafras Gullan & Hodgson, the type species of Austrolecanium Gullan & Hodgson. The adult female of A. cryptocaryae is described and illustrated and a table is provided of the characters that differ among adult females of the three species of Austrolecanium currently recognised (A. cappari (Froggatt), A. cryptocaryae sp. n. and A. sassafras).

Species delimitation in asexual insects of economic importance: The case of black scale (Parasaissetia nigra), a cosmopolitan parthenogenetic pest scale insect.

Asexual lineages provide a challenge to species delimitation because species concepts either have little biological meaning for them or are arbitrary, since every individual is monophyletic and reproductively isolated from all other individuals. However, recognition and naming of asexual species is important to conservation and economic applications. Some scale insects are widespread and polyphagous pests of plants, and several species have been found to comprise cryptic species complexes. Parasaissetia nigra (Nietner, 1861) (Hemiptera: Coccidae) is a parthenogenetic, cosmopolitan and polyphagous pest that feeds on plant species from more than 80 families. Here, we implement multiple approaches to assess the species status of P. nigra, including coalescence-based analyses of mitochondrial and nuclear genes, and ecological niche modelling. Our results indicate that the sampled specimens of P. nigra should be considered to comprise at least two ecotypes (or "species") that are ecologically differentiated, particularly in relation to temperature and moisture. The presence of more than one ecotype under the current concept of P. nigra has implications for biosecurity because the geographic extent of each type is not fully known: some countries may currently have only one of the biotypes. Introduction of additional lineages could expand the geographic extent of damage by the pest in some countries.

Congruent biogeographical disjunctions at a continent-wide scale: Quantifying and clarifying the role of biogeographic barriers in the Australian tropics.

AIM: To test whether novel and previously hypothesized biogeogaphic barriers in the Australian Tropics represent significant disjunction points or hard barriers, or both, to the distribution of plants. LOCATION: Australian tropics: Australian Monsoon Tropics and Australian Wet Tropics. METHODS: The presence or absence of 6,861 plant species was scored across 13 putative biogeographic barriers in the Australian Tropics, including two that have not previously been recognised. Randomizations of these data were used to test whether more species showed disjunctions (gaps in distribution) or likely barriers (range limits) at these points than expected by chance. RESULTS: Two novel disjunctions in the Australian Tropics flora are identified in addition to eleven putative barriers previously recognized for animals. Of these, eleven disjunction points (all within the Australian Monsoon Tropics) were found to correspond to range-ending barriers to a significant number of species, while neither of the two disjunctions found within the Australian Wet Tropics limited a significant number of species' ranges. MAIN CONCLUSIONS: Biogeographic barriers present significant distributional limits to native plant species in the Australian Monsoon Tropics but not in the Australian Wet Tropics.

Three explanations for biodiversity hotspots: small range size, geographical overlap and time for species accumulation. An Australian case study.

To understand the generation and maintenance of biodiversity hotspots, we tested three major hypotheses: rates of diversification, ecological limits to diversity, and time for species accumulation. Using dated molecular phylogenies, measures of species' range size and geographical clade overlap, niche modelling, and lineages-through-time plots of Australian Fabaceae, we compared the southwest Australia Floristic Region (SWAFR; a global biodiversity hotspot) with a latitudinally equivalent non-hotspot, southeast Australia (SEA). Ranges of species (real and simulated) were smaller in the SWAFR than in SEA. Geographical overlap of clades was significantly greater for Daviesia in the SWAFR than in SEA, but the inverse for Bossiaea. Lineage diversification rates over the past 10 Myr did not differ between the SWAFR and SEA in either genus. Interaction of multiple factors probably explains the differences in measured diversity between the two regions. Steeper climatic gradients in the SWAFR probably explain the smaller geographical ranges of both genera there. Greater geographical overlap of clades in the SWAFR, combined with a longer time in the region, can explain why Daviesia is far more species-rich there than in SEA. Our results indicate that the time for speciation and ecological limits hypotheses, in concert, can explain the differences in biodiversity.

Clock model makes a large difference to age estimates of long-stemmed clades with no internal calibration: a test using Australian grasstrees.

BACKGROUND: Estimating divergence times in phylogenies using a molecular clock depends on accurate modeling of nucleotide substitution rates in DNA sequences. Rate heterogeneity among lineages is likely to affect estimates, especially in lineages with long stems and short crowns ("broom" clades) and no internal calibration. We evaluate the performance of the random local clocks model (RLC) and the more routinely employed uncorrelated lognormal relaxed clock model (UCLN) in situations in which a significant rate shift occurs on the stem branch of a broom clade. We compare the results of simulations to empirical results from analyses of a real rate-heterogeneous taxon - Australian grass trees (Xanthorrhoea) - whose substitution rate is slower than in its sister groups, as determined by relative rate tests. RESULTS: In the simulated datasets, the RLC model performed much better than UCLN: RLC correctly estimated the age of the crown node of slow-rate broom clades, whereas UCLN estimates were consistently too young. Similarly, in the Xanthorrhoea dataset, UCLN returned significantly younger crown ages than RLC (mean estimates respectively 3-6 Ma versus 25-35 Ma). In both real and simulated datasets, Bayes Factor tests strongly favored the RLC model over the UCLN model. CONCLUSIONS: The choice of an unsuitable molecular clock model can strongly bias divergence time estimates. In particular, for data predicted to have more rate variation among than within clades, dating with RLC is much more likely to be accurate than with UCLN. The choice of clocks should be informed by the biology of the study group (e.g., life-form) or assessed with relative rate tests and post-hoc model comparisons.

Australia's arid-adapted butcherbirds experienced range expansions during Pleistocene glacial maxima.

A model of range expansions during glacial maxima (GM) for cold-adapted species is generally accepted for the Northern Hemisphere. Given that GM in Australia largely resulted in the expansion of arid zones, rather than glaciation, it could be expected that arid-adapted species might have had expanded ranges at GM, as cold-adapted species did in the Northern Hemisphere. For Australian biota, however, it remains paradigmatic that arid-adapted species contracted to refugia at GM. Here we use multilocus data and ecological niche models (ENMs) to test alternative GM models for butcherbirds. ENMs, mtDNA and estimates of nuclear introgression and past population sizes support a model of GM expansion in the arid-tolerant Grey Butcherbird that resulted in secondary contact with its close relative--the savanna-inhabiting Silver-backed Butcherbird--whose contemporary distribution is widely separated. Together, these data reject the universal use of a GM contraction model for Australia's dry woodland and arid biota.

Rapid chromosomal evolution in a morphologically cryptic radiation.

Cryptic species occur within most of the major taxonomic divisions, and a current challenge is to determine why some lineages have more cryptic species than others. It is expected that cryptic species are more common in groups where there are life histories or genetic architectures that promote speciation in the absence of apparent morphological differentiation. Chromosomal rearrangements have the potential to lead to post-zygotic isolation and might be an important factor leading to cryptic species. Here we investigate the potential role of chromosomal change in driving speciation in the karyotypically diverse scale insect genus Apiomorpha, focussing on four species placed in the same species group (the A. minor species group Gullan, 1984). Using mitochondrial and nuclear DNA sequence data, we find that Apiomorpha minor is not monophyletic and consists of at least nine cryptic species. Diploid chromosome counts range from 2n=4 to 2n=84 across the four currently recognized species, and some of the chromosomal variation exists in the absence of other genetic or host use differences, consistent with karyotypic changes being involved in lineage divergence and the generation of cryptic species.

Evolutionary consequences of shifts to bird-pollination in the Australian pea-flowered legumes (Mirbelieae and Bossiaeeae).

BACKGROUND: Interactions with pollinators are proposed to be one of the major drivers of diversity in angiosperms. Specialised interactions with pollinators can lead to specialised floral traits, which collectively are known as a pollination syndrome. While it is thought that specialisation to a pollinator can lead to either an increase in diversity or in some cases a dead end, it is not well understood how transitions among specialised pollinators contribute to changes in diversity. Here, we use evolutionary trait reconstruction of bee-pollination and bird-pollination syndromes in Australian egg-and-bacon peas (Mirbelieae and Bossiaeeae) to test whether transitions between pollination syndromes is correlated with changes in species diversity. We also test for directionality in transitions that might be caused by selection by pollinators or by an evolutionary ratchet in which reversals to the original pollination syndrome are not possible. RESULTS: Trait reconstructions of Australian egg-and-bacon peas suggest that bee-pollination syndrome is the ancestral form and that there has been replicated evolution of bird-pollination syndromes. Reconstructions indicate potential reversals from bird- to bee-pollination syndromes but this is not consistent with morphology. Species diversity of bird-pollination syndrome clades is lower than that of their bee-pollination syndrome sisters.We estimated the earliest transitions from bee- to bird-pollination syndrome occurred between 30.8 Ma and 10.4 Ma. Geographical structuring of pollination syndromes was found; there were fewer bird-pollination species in the Australian southeast temperate region compared to other regions of Australia. CONCLUSIONS: A consistent decrease in diversification rate coincident with switches to bird pollination might be explained if greater dispersal by bird pollinators results in higher levels of connectivity among populations and reduced chances of allopatric speciation.The earliest transitions overlap with the early diversification of Australian honeyeaters - the major lineage of pollinating birds in Australia. Our findings are consistent with the idea that environment and availability of pollinators are important in the evolution of pollination syndromes. Changes in flower traits as a result of transitions to bird-pollination syndrome might also limit reversals to a bee-pollination syndrome.

Matsucoccus macrocicatrices (Hemiptera: Matsucoccidae): first report, distribution, and association with symptomatic eastern white pine in the southeastern United States.

We provide the first report of Matsucoccus macrocicatrices Richards (Hemiptera: Matsucoccidae) feeding and reproducing on eastern white pine, Pinus strobus L., in the southeastern United States. Until now, M. macrocicatrices had been reported only from the Canadian Atlantic Maritimes, New Hampshire, and Massachusetts. Entomological holdings of 27 major museums in eastern North America have no historical records for M. macrocicatrices from the southeastern region. However, our field surveys and molecular analyses (DNA barcoding) have resulted in the collection and positive identification of M. macrocicatrices in Georgia, North Carolina, South Carolina, Tennessee, Virginia, and West Virginia In addition to the new geographic range, M. macrocicatrices is also being associated with dieback and mortality of all diameter classes of P. strobus leading to concern about a potential shift from its historically nonpestiferous presence on the host tree. On P. strobus, M. macrocicatrices was found embedded in cankers or present on top of the bark with necrotic tissue under their feeding area, indicating that they may be creating wounds for opportunistic pathogenic fungi to infest. Further, we found M. macrocicatrices living outside of the epiphytic mats of its symbiotic fungus, Septobasidium pinicola Snell. This study shows that M. macrocicatrices is now widespread in the southeastern United States, with implications for the future survival and regeneration of P. strobus in eastern North America.

A multilocus coalescent analysis of the speciational history of the Australo-Papuan butcherbirds and their allies.

Changes in geology, sea-level and climate are hypothesised to have been major driving processes of evolutionary diversification (speciation and extinction) in the Australo-Papuan region. Here we use complete species-level sampling and multilocus (one mitochondrial gene, five nuclear loci) coalescent analyses to estimate evolutionary relationships and test hypotheses about the role of changes in climate and landscape in the diversification of the Australo-Papuan butcherbirds and allies (Cracticinae: Cracticus, Strepera, Peltops). Multilocus species trees supported the current classification of the morphologically, ecologically and behaviourally divergent Australian Magpie (Cracticus tibicen (previously Gymnorhina tibicen)) as a member of an expanded genus Cracticus, which includes seven other species with 'butcherbird' morphology and behaviour. Non-monophyly of currently recognised species within Peltops and the white-throated butcherbird species-group (C.argenteus, C.mentalis, C. torquatus) at both mtDNA and nuclear loci suggest that a comprehensive taxonomic revision is warranted for both of these groups. The time-calibrated multilocus species tree revealed an early divergence between the New Guinean rainforest-restricted Peltops lineage and the largely open-habitat inhabiting Cracticus (butcherbirds and magpies) plus Strepera (currawongs) lineage around 17-28Ma, as well as a relatively recent radiation of lineages within Cracticus over the past 8Ma. Overall, patterns and timings of speciation were consistent with the hypothesis that both the expansion of open sclerophyllous woodlands 25-30Ma and the formation of extensive grassland-dominated woodlands 6-8Ma allowed the radiation of lineages adapted to open woodland habitats.

Phylogenetic niche conservatism: what are the underlying evolutionary and ecological causes?

Phylogenetic niche conservatism (PNC) is the tendency of lineages to retain their niche-related traits through speciation events. A recent surge in the availability of well-sampled molecular phylogenies has stimulated phylogenetic approaches to understanding ecological processes at large geographical scales and through macroevolutionary time. We stress that PNC is a pattern, not a process, and is found only in some traits and some lineages. At the simplest level, a pattern of PNC is an inevitable consequence of evolution - descent with modification and divergence of lineages - but several intrinsic causes, including physicochemical, developmental and genetic constraints, can lead directly to a marked pattern of PNC. A pattern of PNC can also be caused indirectly, as a by-product of other causes, such as extinction, dispersal limitation, competition and predation. Recognition of patterns of PNC can contribute to understanding macroevolutionary processes: for example, release from constraint in traits has been hypothesized to trigger adaptive radiations such as that of the angiosperms. Given the multiple causes of patterns of PNC, tests should address explicit questions about hypothesized processes. We conclude that PNC is a scientifically useful concept with applications to the practice of ecological research.

Testing for ecological limitation of diversification: a case study using parasitic plants.

Imbalances in phylogenetic diversity could be the result of variable diversification rates, differing limits on diversity, or a combination of the two. We propose an approach to distinguish between rates and limits as the primary cause of phylogenetic imbalance, using parasitic plants as a model. With sister-taxon comparisons, we show that parasitic plant lineages are typically much less diverse than their autotrophic sisters. We then use age estimates for taxa used in the sister-taxon comparisons to test for correlations between clade age and clade diversity. We find that parasitic plant diversity is not significantly correlated with the age of the lineage, whereas there is a strong positive correlation between the age and diversity of nonparasitic sister lineages. The Ericaceae sister pair Monotropoideae (parasitic) and Arbutoideae (autotrophic) is sufficiently well sampled at the species level to allow more parametric comparisons of diversification patterns. Model fitting for this group supports ecological limitation in Monotropoideae and unconstrained diversification in Arbutoideae. Thus, differences in diversity between parasitic plants and their autotrophic sisters might be caused by a combination of ecological limitation and exponential diversification. A combination of sister-taxon comparisons of diversity and age, coupled with model fitting of well-sampled phylogenies of focal taxa, provides a powerful test of likely causes of asymmetry in the diversity of lineages.