Applications of machine learning in phylogenetics.

Machine learning has increasingly been applied to a wide range of questions in phylogenetic inference. Supervised machine learning approaches that rely on simulated training data have been used to infer tree topologies and branch lengths, to select substitution models, and to perform downstream inferences of introgression and diversification. Here, we review how researchers have used several promising machine learning approaches to make phylogenetic inferences. Despite the promise of these methods, several barriers prevent supervised machine learning from reaching its full potential in phylogenetics. We discuss these barriers and potential paths forward. In the future, we expect that the application of careful network designs and data encodings will allow supervised machine learning to accommodate the complex processes that continue to confound traditional phylogenetic methods.

The use of Lentilactobacillus buchneri PJB1 and Lactiplantibacillus plantarum MTD1 on the ensiling of whole-plant corn silage, snaplage, and high-moisture corn.

Experiments were conducted over a 3-yr period to evaluate the effects of bacterial inoculants on the fermentation profile and aerobic stability of whole-plant corn silage (WPC), snaplage (SNA), and high-moisture corn (HMC). Whole-plant corn was inoculated with Lentilactobacillus buchneri PJB1 in combination with Lactiplantibacillus plantarum MTD1 or with Lpb. plantarum alone (experiments 1 and 2). Snaplage (experiment 3) and HMC (experiments 4 and 5) were inoculated with Len. buchneri in combination with Lpb. plantarum or with Len. buchneri alone. After inoculation, the feedstuffs were ensiled in 7.57-L silos and stored at 21 ± 2°C for 30 or 90 d. In experiment 5, silage was subjected to air stress for 2 h every 2 wk through 42 d and then for 2 h/wk until 90 d and had samples analyzed for their bacterial community composition by metagenomics. Overall, in all experiments, silages inoculated with Len. buchneri alone or in combination with Lpb. plantarum had more acetic acid and 1,2-propanediol and fewer yeasts than uninoculated silages. After 30 d of ensiling, inoculation with Len. buchneri alone or in combination with Lpb. plantarum did not affect the aerobic stability of SNA, but it slightly increased the stability of WPC and markedly improved the stability of HMC. After 90 d of ensiling, inoculation with Len. buchneri alone or in combination with Lpb. plantarum markedly improved the aerobic stability of WPC, SNA, and HMC. In experiment 5, inoculation increased the relative abundance (RA) of Lactobacillaceae and reduced the RA of Enterobacteriaceae and Leuconostocaceae in HMC at 30 and 90 d and the RA of Clostridiaceae in non-air-stressed HMC at 90 d. Air-stressed HMC inoculated with Len. buchneri had less lactic acid, more acetic acid and 1,2-propanediol, and markedly greater aerobic stability than uninoculated air-stressed HMC at 90 d. In conclusion, inoculation with Len. buchneri PJB1 alone or in combination with Lpb. plantarum MTD1 increased the production of acetic acid and 1,2-propanediol, inhibited yeasts development, and improved the aerobic stability of WPC, SNA, and HMC. In HMC, inoculation markedly improved aerobic stability as soon as after 30 d of ensiling, and after 90 d, inoculation improved stability even under air stress conditions.

Phylogenetic inference using generative adversarial networks.

MOTIVATION: The application of machine learning approaches in phylogenetics has been impeded by the vast model space associated with inference. Supervised machine learning approaches require data from across this space to train models. Because of this, previous approaches have typically been limited to inferring relationships among unrooted quartets of taxa, where there are only three possible topologies. Here, we explore the potential of generative adversarial networks (GANs) to address this limitation. GANs consist of a generator and a discriminator: at each step, the generator aims to create data that is similar to real data, while the discriminator attempts to distinguish generated and real data. By using an evolutionary model as the generator, we use GANs to make evolutionary inferences. Since a new model can be considered at each iteration, heuristic searches of complex model spaces are possible. Thus, GANs offer a potential solution to the challenges of applying machine learning in phylogenetics. RESULTS: We developed phyloGAN, a GAN that infers phylogenetic relationships among species. phyloGAN takes as input a concatenated alignment, or a set of gene alignments, and infers a phylogenetic tree either considering or ignoring gene tree heterogeneity. We explored the performance of phyloGAN for up to 15 taxa in the concatenation case and 6 taxa when considering gene tree heterogeneity. Error rates are relatively low in these simple cases. However, run times are slow and performance metrics suggest issues during training. Future work should explore novel architectures that may result in more stable and efficient GANs for phylogenetics. AVAILABILITY AND IMPLEMENTATION: phyloGAN is available on github: https://github.com/meganlsmith/phyloGAN/.

Social Support for Rural Gender Diverse Youth Compared to Cisgender Peers.

PURPOSE: Gender diverse youth (GDY) experience higher rates of mental health concerns than their cisgender peers, but these can be ameliorated by feeling support from family, school, and community. Little is known about how youth perceptions of support vary by gender identity, especially for younger adolescents and those living in rural areas. METHODS: Youth ages 12-19 years completed anonymous surveys including measures of perceived support and a two-step gender identity question. GDY (n = 206) were further categorized into binary and nonbinary gender identities. An additional 500 randomly selected cisgender youth were included for comparison. Multivariate analyses of variance with Tukey post hoc tests were employed to test GDY group differences while accounting for the interaction between scaled measures. RESULTS: Cisgender youth had the highest perceived support across all support measures while youth who shared both binary and nonbinary aspects of their gender identity had the lowest rates of perceived support. The F tests for between-subject effects were statistically significant (p <.001) for all six support measures, and multivariate group testing was statistically significant with Wilks' λ 6.38(18,1621.17) = 0.82; p <.001. DISCUSSION: Despite research demonstrating a strong association between perceived support and improved mental health outcomes, GDY in our sample had lower rates of perceived support at the family, school, and community levels. GDY with both binary and nonbinary gender identities had the lowest levels of perceived support. Further research is needed to see if this finding is consistent in other populations and to develop targeted interventions to improve perceived support for this population.

Species limits and diversification of the Dendropsophus rubicundulus subgroup (Anura, Hylidae) in Neotropical savannas.

Understanding the processes that generate and maintain biodiversity at and below the species level is a central goal of evolutionary biology. Here we explore the spatial and temporal drivers of diversification of the treefrog subgroup Dendropsophus rubicundulus, a subgroup of the D. microcephalus species group, over periods of pronounced geological and climatic changes in the Neotropical savannas that they inhabit. This subgroup currently comprises 11 recognized species distributed across the Brazilian and Bolivian savannas, but the taxonomy has been in a state of flux, necessitating reexamination. Using newly generated single nucleotide polymorphism (SNP) data from restriction-site associated DNA sequencing (RADseq) and mitochondrial 16S sequence data for ∼150 specimens, we inferred phylogenetic relationships, tested species limits using a model-based approach, and estimated divergence times to gain insights into the geographic and climatic events that affected the diversification of this subgroup. Our results recognized at least nine species: D. anataliasiasi, D. araguaya, D. cerradensis, D. elianeae, D. jimi, D. rubicundulus, D. tritaeniatus, D. rozenmani, and D. sanborni. Although we did not collect SNP data for the latter two species, they are likely distinct based on mitochondrial data. In addition, we found genetic structure within the widespread species D. rubicundulus, which comprises three allopatric lineages connected by gene flow upon secondary contact. We also found evidence of population structure and perhaps undescribed diversity in D. elianeae, which warrants further study. The D. rubicundulus subgroup is estimated to have originated in the Late Miocene (∼5.45 million years ago), with diversification continuing through the Pliocene and Early Pleistocene, followed by the most recent divergence of D. rubicundulus lineages in the Middle Pleistocene. The epeirogenic uplift followed by erosion and denudation of the central Brazilian plateau throughout the Pliocene and Pleistocene, in combination with the increasing frequency and amplitude of climatic fluctuations during the Pleistocene, was important for generating and structuring diversity at or below the species level in the D. rubicundulus subgroup.

Possible role of caffeine in nicotine use onset among early adolescents: Evidence from the Young Mountaineer Health Study Cohort.

BACKGROUND: Preventing nicotine use onset among children and youth is an important public health goal. One possible contributor that has received little empirical investigation is caffeine use. The goal of this study was to examine the possible contribution of caffeine to nicotine onset during early adolescence. METHODS: We used data from the Young Mountaineer Health Study Cohort. Survey data were collected from 1,349 (response rate: 80.7%) 6th grade students (mean age at baseline 11.5 years) in 20 middle schools in West Virginia during the fall of 2020 and spring of 2021. We limited our analyses to students reporting never having used any form of nicotine at baseline. Logistic regression was employed in analyses. RESULTS: Approximately 8% of participants reported having used nicotine at least once between baseline and the follow-up, and 4.7% reported solely using electronic nicotine delivery systems (ENDS) and no other forms of nicotine. In multivariable analyses, we controlled for many environmental, social, and behavioral variables known to influence nicotine use such as alcohol use, peer substance use, and perceived access to nicotine. We formulated our main independent variable, caffeine consumption, as continuous deciles. Any nicotine use, as well as ENDS use only at follow-up, were modeled as dependent variables. Caffeine was significantly associated with nicotine use in both models with ORs of 1.15 (1.04-1.27) and 1.13 (1.00-1.28). CONCLUSIONS: Caffeine consumption among 6th grade non-nicotine users was associated with nicotine use at approximately 6-months follow-up.

There and Back Again: The Unexpected Journeys of Metridium de Blainville, 1824 between the Old Oceans and throughout the Modern World.

AbstractMembers of the sea anemone genus Metridium are abundant in temperate rocky habitats and fouling communities. Their biogeographic history is expected to reflect changes in currents and habitats that have influenced benthic communities, such as the climate-influenced changes that occurred during the Last Glacial Maximum. More recently, however, anthropogenic influences such as shipping transportation and the creation of artificial habitat have altered and affected the composition of modern-day marine communities. Here we use sequence-capture data to examine the genetic structure of Metridium across its shallow-water distribution to (1) evaluate species boundaries within Metridium, (2) elucidate the dispersal history of Metridium between and among oceans, and (3) assess the influence of anthropogenic movement on modern-day populations. We find strong evidence for two species within Metridium: M. farcimen and M. senile. Dispersal from the Pacific to the Atlantic included a subsequent isolation of a small population in or above the Bering Sea, which has presumably moved southward. Within the native range of M. senile, admixture is prevalent even between oceans as a result of anthropogenic activities. The nonnative populations in Chile and the Falkland Islands came from at least two distinct introduction events originating from both coasts of the United States in the North Pacific and North Atlantic Oceans. Hybridization between M. senile and M. farcimen is documented as occurring in anthropogenically influenced habitats. The heavy influence from anthropogenic activities will continue to impact our understanding of marine organisms, particularly within the native range and for those that are easily transported across long distances.

Social Support and Perceptions of COVID-19-related Emotional Impact on Mental Health Among Early Adolescents in Appalachia.

BACKGROUND: Young people who experience higher levels of social support from their schools and families have been shown to be less likely to develop symptoms of negative mental health outcomes such as depression and anxiety.1-4 This raises questions concerning how young people's stress and psychological changes due to the COVID-19 pandemic as well as social support during this time have affected their overall mental health. The aim of this study was to assess the association between sources of parental- and school-level social support and youth perceptions of COVID-19-related emotional impact on mental health among early adolescent girls and boys in Appalachia. METHODS: Using linear regression, we analyzed the first and third wave of survey data from the larger parent study (Young Mountaineer Health Study) cohort, collected in 20 middle schools throughout West Virginia in the fall of 2020 and fall of 2021 (N = 1349, mean age: 11.5, response rate: 80.7%). RESULTS: Approximately half of participants reported knowing someone that had been sick with COVID-19. Those experiencing higher levels of perceived COVID-19-related emotional impact reported greater levels of depression, anxiety, and anger. Both parental and school-level social support were associated with better mental health outcomes. CONCLUSIONS: Early adolescent perceptions of COVID-19-related emotional impact were associated with depression, anxiety, and anger and moderated by social support at home and in school among 11-12-year-old youth in Appalachia.

The Prevalence of Gender-Diverse Youth in a Rural Appalachian Region.

This study discusses a survey that explores youth gender identity in the Appalachian region.

Caffeine consumption and onset of alcohol use among early adolescents.

Preventing or delaying the onset of alcohol use among children and youth is an important public health goal. One possible factor in alcohol use onset among early adolescents is caffeine. The aim of this study was to assess the possible contribution of caffeine to the onset of alcohol use during early adolescence. We used data from the Young Mountaineer Health Study Cohort. Survey data were collected from 1349 (response rate: 80.7%) 6th grade students (mean age at baseline 11.5 years) in 20 middle schools in West Virginia during the fall of 2020, and again approximately 6 months later in spring of 2021. We limited our analyses to students reporting never having used any form of alcohol at baseline. Logistic regression was employed in multivariable analyses and both Odds Ratios and Relative Risks reported. At follow-up, almost 14% of participants reported having consumed alcohol at least once and 57% used caffeine of 100 mg + daily. In multivariable analyses we controlled for social and behavioral variables known to impact tobacco use. Caffeine use was operationalized as a three-level factor: no use, <100 mg per day, and 100 + mg per day, with the latter being the approximate equivalent of the minimum of a typical cup of coffee or can of energy drink. Caffeine use of 100 mg + per day was significantly related to alcohol use at 6-months follow-up (OR: 1.79, RR: 1.56, p = .037). We conclude that caffeine consumption among 11-12-year-old adolescents may be a factor in early onset of alcohol use.

Risk and Resilience Pathways, Community Adversity, Decision-making, and Alcohol Use Among Appalachian Adolescents: Protocol for the Longitudinal Young Mountaineer Health Study Cohort.

BACKGROUND: Alcohol use impairs psychosocial and neurocognitive development and increases the vulnerability of youth to academic failure, substance use disorders, and other mental health problems. The early onset of alcohol use in adolescents is of particular concern, forecasting substance abuse in later adolescence and adulthood. To date, evidence suggests that youth in rural areas are especially vulnerable to contextual and community factors that contribute to the early onset of alcohol use. OBJECTIVE: The objective of the Young Mountaineer Health Study is to investigate the influence of contextual and health behavior variables on the early onset of alcohol use among middle school-aged youth in resource-poor Appalachian rural communities. METHODS: This is a program of prospective cohort studies of approximately 2200 middle school youth from a range of 20 rural, small town, and small city (population <30,000) public schools in West Virginia. Students are participating in 6 waves of data collection (2 per year) over the course of middle school (sixth to eighth grades; fall and spring) from 2020 to 2023. On the basis of an organizational arrangement, which includes a team of local data collection leaders, supervising contact agents in schools, and an honest broker system to deidentify data linked via school IDs, we are able to collect novel forms of data (self-reported data, teacher-reported data, census-linked area data, and archival school records) while ensuring high rates of participation by a large majority of youth in each participating school. RESULTS: In the spring of 2021, 3 waves of student survey data, 2 waves of data from teachers, and a selection of archival school records were collected. Student survey wave 1 comprised 1349 (response rate 80.7%) participants, wave 2 comprised 1649 (response rate 87%) participants, and wave 3 comprised 1909 (response rate 83.1%) participants. The COVID-19 pandemic has had a negative impact on the sampling frame size, resulting in a reduced number of eligible students, particularly during the fall of 2020. Nevertheless, our team structure and incentive system have proven vitally important in mitigating the potentially far greater negative impact of the pandemic on our data collection processes. CONCLUSIONS: The Young Mountaineer Health Study will use a large data set to test pathways linking rural community disadvantage to alcohol misuse among early adolescents. Furthermore, the program will test hypotheses regarding contextual factors (eg, parenting practices and neighborhood collective efficacy) that protect youth from community disadvantage and explore alcohol antecedents in the onset of nicotine, marijuana, and other drug use. Data collection efforts have been successful despite interruptions caused by the COVID-19 pandemic in 2020 and 2021. INTERNATIONAL REGISTERED REPORT IDENTIFIER (IRRID): DERR1-10.2196/40451.

The role of multiple Pleistocene refugia in promoting diversification in the Pacific Northwest.

Pleistocene glacial cycles drastically changed the distributions of taxa endemic to temperate rainforests in the Pacific Northwest, with many experiencing reduced habitat suitability during glacial periods. In this study, we investigate whether glacial cycles promoted intraspecific divergence and whether subsequent range changes led to secondary contact and gene flow. For seven invertebrate species endemic to the PNW, we estimated species distribution models (SDMs) and projected them onto current and historical climate conditions to assess how habitat suitability changed during glacial cycles. Using single nucleotide polymorphism (SNP) data from these species, we assessed population genetic structure and used a machine-learning approach to compare models with and without gene flow between populations upon secondary contact after the last glacial maximum (LGM). Finally, we estimated divergence times and rates of gene flow between populations. SDMs suggest that there was less suitable habitat in the North Cascades and Northern Rocky Mountains during glacial compared to interglacial periods, resulting in reduced habitat suitability and increased habitat fragmentation during the LGM. Our genomic data identify population structure in all taxa, and support gene flow upon secondary contact in five of the seven taxa. Parameter estimates suggest that population divergences date to the later Pleistocene for most populations. Our results support a role of refugial dynamics in driving intraspecific divergence in the Cascades Range. In these invertebrates, population structure often does not correspond to current biogeographic or environmental barriers. Rather, population structure may reflect refugial lineages that have since expanded their ranges, often leading to secondary contact between once isolated lineages.

Assessing model adequacy for Bayesian Skyline plots using posterior predictive simulation.

Bayesian skyline plots (BSPs) are a useful tool for making inferences about demographic history. For example, researchers typically apply BSPs to test hypotheses regarding how climate changes have influenced intraspecific genetic diversity over time. Like any method, BSP has assumptions that may be violated in some empirical systems (e.g., the absence of population genetic structure), and the naïve analysis of data collected from these systems may lead to spurious results. To address these issues, we introduce P2C2M.Skyline, an R package designed to assess model adequacy for BSPs using posterior predictive simulation. P2C2M.Skyline uses a phylogenetic tree and the log file output from Bayesian Skyline analyses to simulate posterior predictive datasets and then compares this null distribution to statistics calculated from the empirical data to check for model violations. P2C2M.Skyline was able to correctly identify model violations when simulated datasets were generated assuming genetic structure, which is a clear violation of BSP model assumptions. Conversely, P2C2M.Skyline showed low rates of false positives when models were simulated under the BSP model. We also evaluate the P2C2M.Skyline performance in empirical systems, where we detected model violations when DNA sequences from multiple populations were lumped together. P2C2M.Skyline represents a user-friendly and computationally efficient resource for researchers aiming to make inferences from BSP.

Application of 3-nitrooxypropanol and canola oil to mitigate enteric methane emissions of beef cattle results in distinctly different effects on the rumen microbial community.

BACKGROUND: The major greenhouse gas from ruminants is enteric methane (CH4) which in 2010, was estimated at 2.1 Gt of CO2 equivalent, accounting for 4.3% of global anthropogenic greenhouse gas emissions. There are extensive efforts being made around the world to develop CH4 mitigating inhibitors that specifically target rumen methanogens with the ultimate goal of reducing the environmental footprint of ruminant livestock production. This study examined the individual and combined effects of supplementing a high-forage diet (90% barley silage) fed to beef cattle with the investigational CH4 inhibitor 3-nitrooxypropanol (3-NOP) and canola oil (OIL) on the rumen microbial community in relation to enteric CH4 emissions and ruminal fermentation. RESULTS: 3-NOP and OIL individually reduced enteric CH4 yield (g/kg dry matter intake) by 28.2% and 24.0%, respectively, and the effects were additive when used in combination (51.3% reduction). 3-NOP increased H2 emissions 37-fold, while co-administering 3-NOP and OIL increased H2 in the rumen 20-fold relative to the control diet. The inclusion of 3-NOP or OIL significantly reduced the diversity of the rumen microbiome. 3-NOP resulted in targeted changes in the microbiome decreasing the relative abundance of Methanobrevibacter and increasing the relative abundance of Bacteroidetes. The inclusion of OIL resulted in substantial changes to the microbial community that were associated with changes in ruminal volatile fatty acid concentration and gas production. OIL significantly reduced the abundance of protozoa and fiber-degrading microbes in the rumen but it did not selectively alter the abundance of rumen methanogens. CONCLUSIONS: Our data provide a mechanistic understanding of CH4 inhibition by 3-NOP and OIL when offered alone and in combination to cattle fed a high forage diet. 3-NOP specifically targeted rumen methanogens and partly inhibited the hydrogenotrophic methanogenesis pathway, which increased H2 emissions and propionate molar proportion in rumen fluid. In contrast, OIL caused substantial changes in the rumen microbial community by indiscriminately altering the abundance of a range of rumen microbes, reducing the abundance of fibrolytic bacteria and protozoa, resulting in altered rumen fermentation. Importantly, our data suggest that co-administering CH4 inhibitors with distinct mechanisms of action can both enhance CH4 inhibition and provide alternative sinks to prevent excessive accumulation of ruminal H2.

Using all Gene Families Vastly Expands Data Available for Phylogenomic Inference.

Traditionally, single-copy orthologs have been the gold standard in phylogenomics. Most phylogenomic studies identify putative single-copy orthologs using clustering approaches and retain families with a single sequence per species. This limits the amount of data available by excluding larger families. Recent advances have suggested several ways to include data from larger families. For instance, tree-based decomposition methods facilitate the extraction of orthologs from large families. Additionally, several methods for species tree inference are robust to the inclusion of paralogs and could use all of the data from larger families. Here, we explore the effects of using all families for phylogenetic inference by examining relationships among 26 primate species in detail and by analyzing five additional data sets. We compare single-copy families, orthologs extracted using tree-based decomposition approaches, and all families with all data. We explore several species tree inference methods, finding that identical trees are returned across nearly all subsets of the data and methods for primates. The relationships among Platyrrhini remain contentious; however, the species tree inference method matters more than the subset of data used. Using data from larger gene families drastically increases the number of genes available and leads to consistent estimates of branch lengths, nodal certainty and concordance, and inferences of introgression in primates. For the other data sets, topological inferences are consistent whether single-copy families or orthologs extracted using decomposition approaches are analyzed. Using larger gene families is a promising approach to include more data in phylogenomics without sacrificing accuracy, at least when high-quality genomes are available.

Genomic evidence of an ancient inland temperate rainforest in the Pacific Northwest of North America.

The disjunct temperate rainforests of the Pacific Northwest of North America (PNW) are characterized by late-successional dominant tree species Thuja plicata (western redcedar) and Tsuga heterophylla (western hemlock). The demographic histories of these species, along with the PNW rainforest ecosystem in its entirety, have been heavily impacted by geological and climatic changes the PNW has experienced over the last 5 million years, including mountain orogeny and repeated Pleistocene glaciations. These environmental events have ultimately shaped the history of these species, with inland populations potentially being extirpated during the Pleistocene glaciations. Here, we collect genomic data for both species across their ranges to test multiple demographic models, each reflecting a different phylogeographical hypothesis on how the ecosystem-dominating species may have responded to dramatic climatic change. Our results indicate that inland and coastal populations in both species diverged ~2.5 million years ago in the early Pleistocene and experienced decreases in population size during glacial cycles, with subsequent population expansion. Importantly, we found evidence for gene flow between coastal and inland populations during the mid-Holocene. It is likely that intermittent migration in these species during this time has prevented allopatric speciation via genetic drift alone. In conclusion, our results from combining genomic data and demographic inference procedures establish that populations of the ecosystem dominants Thuja plicata and Tsuga heterophylla persisted in refugia located in both the coastal and inland regions of the PNW throughout the Pleistocene, with populations expanding and contracting in response to glacial cycles with occasional gene flow.

Assessing model adequacy leads to more robust phylogeographic inference.

Phylogeographic studies base inferences on large data sets and complex demographic models, but these models are applied in ways that could mislead researchers and compromise their inference. Researchers face three challenges associated with the use of models: (i) 'model selection', or the identification of an appropriate model for analysis; (ii) 'evaluation of analytical results', or the interpretation of the biological significance of the resulting parameter estimates, delimitations, and topologies; and (iii) 'model evaluation', or the use of statistical approaches to assess the fit of the model to the data. The field collectively invests most of its energy in point (ii) without considering the other points; we argue that attention to points (i) and (iii) is essential to phylogeographic inference.

Species Tree Inference Methods Intended to Deal with Incomplete Lineage Sorting Are Robust to the Presence of Paralogs.

Many recent phylogenetic methods have focused on accurately inferring species trees when there is gene tree discordance due to incomplete lineage sorting (ILS). For almost all of these methods, and for phylogenetic methods in general, the data for each locus are assumed to consist of orthologous, single-copy sequences. Loci that are present in more than a single copy in any of the studied genomes are excluded from the data. These steps greatly reduce the number of loci available for analysis. The question we seek to answer in this study is: what happens if one runs such species tree inference methods on data where paralogy is present, in addition to or without ILS being present? Through simulation studies and analyses of two large biological data sets, we show that running such methods on data with paralogs can still provide accurate results. We use multiple different methods, some of which are based directly on the multispecies coalescent model, and some of which have been proven to be statistically consistent under it. We also treat the paralogous loci in multiple ways: from explicitly denoting them as paralogs, to randomly selecting one copy per species. In all cases, the inferred species trees are as accurate as equivalent analyses using single-copy orthologs. Our results have significant implications for the use of ILS-aware phylogenomic analyses, demonstrating that they do not have to be restricted to single-copy loci. This will greatly increase the amount of data that can be used for phylogenetic inference.[Gene duplication and loss; incomplete lineage sorting; multispecies coalescent; orthology; paralogy.].

The Frequency and Topology of Pseudoorthologs.

Phylogenetics has long relied on the use of orthologs, or genes related through speciation events, to infer species relationships. However, identifying orthologs is difficult because gene duplication can obscure relationships among genes. Researchers have been particularly concerned with the insidious effects of pseudoorthologs-duplicated genes that are mistaken for orthologs because they are present in a single copy in each sampled species. Because gene tree topologies of pseudoorthologs may differ from the species tree topology, they have often been invoked as the cause of counterintuitive results in phylogenetics. Despite these perceived problems, no previous work has calculated the probabilities of pseudoortholog topologies or has been able to circumscribe the regions of parameter space in which pseudoorthologs are most likely to occur. Here, we introduce a model for calculating the probabilities and branch lengths of orthologs and pseudoorthologs, including concordant and discordant pseudoortholog topologies, on a rooted three-taxon species tree. We show that the probability of orthologs is high relative to the probability of pseudoorthologs across reasonable regions of parameter space. Furthermore, the probabilities of the two discordant topologies are equal and never exceed that of the concordant topology, generally being much lower. We describe the species tree topologies most prone to generating pseudoorthologs, finding that they are likely to present problems to phylogenetic inference irrespective of the presence of pseudoorthologs. Overall, our results suggest that pseudoorthologs are unlikely to mislead inferences of species relationships under the biological scenarios considered here.[Birth-death model; orthologs; paralogs; phylogenetics.].