Cryptic diversity and discordance in single‐locus species delimitation methods within horned lizards (Phrynosomatidae: Phrynosoma)

Biodiversity reduction and loss continues to progress at an alarming rate, and thus, there is widespread interest in utilizing rapid and efficient methods for quantifying and delimiting taxonomic diversity. Single‐locus species delimitation methods have become popular, in part due to the adoption of the DNA barcoding paradigm. These techniques can be broadly classified into tree‐based and distance‐based methods depending on whether species are delimited based on a constructed genealogy. Although the relative performance of these methods has been tested repeatedly with simulations, additional studies are needed to assess congruence with empirical data. We compiled a large data set of mitochondrial ND4 sequences from horned lizards (Phrynosoma) to elucidate congruence using four tree‐based (single‐threshold GMYC, multiple‐threshold GMYC, bPTP, mPTP) and one distance‐based (ABGD) species delimitation models. We were particularly interested in cases with highly uneven sampling and/or large differences in intraspecific diversity. Results showed a high degree of discordance among methods, with multiple‐threshold GMYC and bPTP suggesting an unrealistically high number of species (29 and 26 species within the P. douglasii complex alone). The single‐threshold GMYC model was the most conservative, likely a result of difficulty in locating the inflection point in the genealogies. mPTP and ABGD appeared to be the most stable across sampling regimes and suggested the presence of additional cryptic species that warrant further investigation. These results suggest that the mPTP model may be preferable in empirical data sets with highly uneven sampling or large differences in effective population sizes of species.     

A rosette by any other name: species diversity in the Bangiales (Rhodophyta) along the South African coast

The Bangiales is an order of Rhodophyta, widely distributed around the globe and best known for its economic value in the nori industry. The morphological simplicity of the group offers limited distinguishing characters for species identification. We therefore delimited species of the Bangiales along the South African coast based on two unlinked loci, the mitochondrial cox1 gene and the plastid rbcL gene, supplemented with additional sequence data from a third gene, the nuclear nSSU. Application of DNA-based species delimitation methods including the Automatic Barcode Gap Discovery (ABGD), General Mixed Yule Coalescent (GYMC) and Poisson Tree Processes (PTP), resulted in the recognition of 10 Porphyra and three Pyropia species in South Africa, only three of which had been previously described. Additional species of Bangiales previously recorded along the South African coast were added to our final species list despite not being found in the present study, resulting in an estimate of 14–16 Bangiales species occurring along this shoreline. Most of this extensive genetic diversity has been misidentified as the commonly rosette-forming species P. capensis. The name P. capensis currently refers to a species complex and cannot be attached to any one species with certainty. All species in this complex, confirmed using genetic data, are endemic to South Africa. Our results compare well with other Southern Hemisphere countries, such as Chile and New Zealand, where high genetic diversity, species richness and endemicity have also been found.     

Molecular data reveal cryptic lineages within the northeastern Atlantic and Mediterranean small mussel drills of the Ocinebrina edwardsii complex (Mollusca: Gastropoda: Muricidae)

We used a molecular phylogenetic approach to investigate species delimitations and diversification in the mussel drills of the Ocinebrina edwardsii complex by means of a combination of nuclear (internal transcribed spacer 2, ITS2) and mitochondrial [cytochrome oxidase subunit I (COI) and 16S] sequences. Our sample included 243 specimens ascribed to seven currently accepted species from 51 sites. Five of the samples were from either the type locality of a nominal species or a close nearby locality (O. edwardsii from Corsica, O. carmelae and O. piantonii from the Kerkennah Islands, O. hispidula from the Gulf of Gabès and O. leukos from the Canary Islands), one from the inferred original locality (O. ingloria from Venice Lagoon), and specimens assigned in the recent literature to O. nicolai. We used a combination of distance‐ and tree‐based species delimitation methods to identify Molecular Operational Taxonomic Units (MOTUs) to compare with the a priori species identifications. The consensus tree obtained by BEAST on the COI alignment allows the recognition of several distinct clades supported by the three species delimitation methods employed. The eight‐MOTUs scenario, shared by the Automatic Barcode Gap Discovery (ABGD) and Generalized Mixed Yule‐Coalescent (GMYC) methods, comprises the following major clades: clade A contains the south Tunisian species Ocinebrina piantonii Cecalupo, Buzzurro & Mariani from which the sympatric taxon O. carmelae Cecalupo, Buzzurro & Mariani (new synonym) cannot be separated; clades B and C bring together all populations from the Aegean Sea and some from the Ionian Sea, respectively; clade D groups, on the one hand, the south Tunisian samples morphologically assigned to O. hispidula Pallary and, on the other, Atlantic and Alboran Sea samples (including the Canarian taxon O. leukos Houart); clade E includes a sample from the type locality of O. edwardsii and several samples from the Tyrrhenian Sea; clades F and G correspond to a few samples from the Venice Lagoon and the Tyrrhenian Sea, respectively; clade H groups the bulk of samples from the Adriatic Sea, including samples from the Venice Lagoon morphologically identified as Ocinebrina ingloria (Crosse), and some from the Ionian Sea. No final conclusions could be reached to reconcile the currently recognized morphological taxa with the clades suggested by the COI data. The geographical structure proposed by the mitochondrial markers is similar to that found in other marine invertebrates and partially corresponds to the species defined by shell characters. We propose here a framework for the revision of the Ocinebrina edwardsii species complex, suggesting a geographical pattern for the diversification of this group in the studied area. © 2013 The Linnean Society of London     

A new integrative framework for large‐scale assessments of biodiversity and community dynamics,using littoral gastropods and crabs of British Columbia,Canada

Improving our understanding of species responses to environmental changes is an important contribution ecologists can make to facilitate effective management decisions. Novel synthetic approaches to assessing biodiversity and ecosystem integrity are needed, ideally including all species living in a community and the dynamics defining their ecological relationships. Here, we present and apply an integrative approach that links high‐throughput, multicharacter taxonomy with community ecology. The overall purpose is to enable the coupling of biodiversity assessments with investigations into the nature of ecological interactions in a community‐level data set. We collected 1195 gastropods and crabs in British Columbia. First, the General mixed Yule‐coalescent (GMYC) and the Poisson Tree Processes (PTP) methods for proposing primary species‐hypotheses based on cox1 sequences were evaluated against an integrative taxonomic framework. We then used data on the geographic distribution of delineated species to test species co‐occurrence patterns for nonrandomness using community‐wide and pairwise approaches. Results showed that PTP generally outperformed GMYC and thus constitutes a more effective option for producing species‐hypotheses in community‐level data sets. Nonrandom species co‐occurrence patterns indicative of ecological relationships or habitat preferences were observed for grazer gastropods, whereas assemblages of carnivorous gastropods and crabs appeared influenced by random processes. Species‐pair associations were consistent with current ecological knowledge, thus suggesting that applying community assembly within a large taxonomical framework constitutes a valuable tool for assessing ecological interactions. Combining phylogenetic, morphological and co‐occurrence data enabled an integrated view of communities, providing both a conceptual and pragmatic framework for biodiversity assessments and investigations into community dynamics.     

生物物种资源监测原则与指标及抽样设计方法

生物物种资源监测是了解生物物种资源现状、开展生物物种资源保护与管理的基础工作和重要手段.阐述了生物物种资源监测的科学性原则、可操作性原则和持续性原则.提出了监测计划的制定程序;监测计划应充分考虑所具有的人力、资金和后勤保障等条件,并进行定期评估.分析了指示物种在物种资源监测中的作用与不足;认为应选择具有不同生态需求和生活史的生物类群作为监测对象.讨论了监测指标的选取方法;监测指标应可测量、有科学基础、易被公众接受、低成本和高效益;监测方法应具有科学性,能检测到相应的变化,应采用高效率、低成本的标准化监测方法.分析了现有监测计划在抽样设计方面存在的问题,探讨了空间变异性和可检测率对监测数据误差的影响及其处理方式,讨论了样本量确定和监测样地的大小、形状及位置设计.监测样地要有较好的代表性,能在有限的监测面积中较好地反映监测区域内群落种类组成与数量特征.最后,讨论了生物物种资源监测的尺度和标准化问题.     

Integrative taxonomy of Peniculus,Metapeniculus, and Trifur (Siphonostomatoida: Pennellidae), copepod parasites of marine fishes from Chile: species delimitation analyses using DNA barcoding and morphological evidence

Pennellidae is a family of copepod parasites of widely distributed marine fishes. The pennellid species are usually morphologically differentiated by cephalothorax, neck, trunk, and abdomen shape. These characters, however, show high polymorphism and therefore using only this type of data, delimitation at species level of this genus is difficult. In this study, we explored the genetic distances calculated from sequences of a DNA barcoding marker (COI mt) (678 base pairs). We also explored the genetic distances of 25 Peniculus specimens associated within nine marine fish species, four Metapeniculus specimens associated within one marine fish species, and four Trifur specimens associated within one marine fish species. All specimens were collected in Antofagasta Bay, Chile and were calculated from sequences of a DNA barcoding marker (COI mt) (678 base pairs). The genetic distance among the Peniculus specimens was 0.95% from the different host species, the Metapeniculus specimens distance was 0.44% and the Trifur specimens was 2.25%. Genetic difference between Peniculus and Metapeniculus was 17.86% and Peniculus differ from T. tortuosus by 18.16%. We analysed the barcoding gene fragment using Bayesian Inference (BI) for phylogenetic reconstruction using three outgroups. Based on the phylogenetic analysis an ultrametric tree was built and a general mixed Yule-coalescent (bGMYC) model was conducted for species delimitation. Morphometrics analyses were made with Bayesian statistics. Mean and credibility limit (95%) for each parameter was calculated. Results show that based on morphology the individuals collected can be assigned to P. cf. fistula von Nordmann, 1832, Metapeniculus antofagastensis Castro-Romero & Baeza-Kuroki, 1985, and Trifur cf. tortuosus Wilson, 1917. High morphological polymorphism was observed for the lineage of Peniculus associated to several host species of marine fishes. Similar results were obtained for Trifur cf. tortuosus parasites on Chilean marine fishes.     

A molecular assessment of species diversity in Tympanopleura and Ageneiosus catfishes (Auchenipteridae: Siluriformes)

In order to test the congruence of genetic data to the morphologically defined Neotropical catfish genera Tympanopleura and Ageneiosus and explore species diversity, we generated 17 DNA barcodes from five of six species of Tympanopleura and 12 of 13 species of Ageneiosus. To discriminate limits between species, an automatic barcode gap discovery (ABGD), a generalised mixed yule-coalescent model (GYMC) and fixed distance thresholds Kimura two-parameter (K2P; 3%) were used to discriminate putative species limits from the DNA barcodes. The ABGD, GMYC and K2P methods agreed by each generating 13 clusters: six in Tympanopleura (five nominal plus one undescribed species) and seven in Ageneiosus. These clusters corresponded broadly to the described species, except in the case of the Ageneiosus ucayalensis group (A. akamai, A. dentatus, A. intrusus, A. ucayalensis, A. uranophthalmus and A. vittatus). Haplotype sharing and low divergences may have prevented molecular methods from distinguishing these species. We hypothesise that this is the result of a recent radiation of a sympatric species group distributed throughout the Amazon Basin. One putative new species of Tympanopleura was also supported by the molecular data. These results taken together highlight the utility of molecular methods such as DNA barcoding in understanding patterns of diversification across large geographic areas and in recognising overlooked diversity.     

Genetic and morphological studies of species status for poorly known endemic Trochulus phorochaetius (Bourguignat, 1864) (Gastropoda: Pulmonata: Hygromiidae), and its comparison with closely related taxa

The taxonomic status of Trochulus phorochaetius and its phylogenetic relationships to Trochulus plebeius and Trochulus hispidus were evaluated based on molecular, morphological, and genital anatomy data. The canonical discriminant analysis (CDA) of shell morphology allowed the clear differentiation between these three nominal species, whereas the genitalia revealed their high similarity. Analyses of cytochrome c oxidase subunit I (COI) sequences were not always congruent with the differentiation between these three species by shell characters. None of them formed a monophyletic group covering all its sequences. Instead, many sequences obtained from individuals classified to the same morphospecies, and/or usually collected from the same region or country, created highly supported separate clades and delimited clusters. Three distinct clades containing sequences of two morphospecies originating from the same country were identified in molecular phylogenetic and species delimitation studies: (1) T. plebeius + T. hispidus from Great Britain; (2) T. plebeius + T. hispidus from Poland; and (3) T. phorochaetius + T. hispidus from France. In the latter case some of the sequences were even identical. Their genetic similarity could indicate the ability to hybridize, which may be evidenced by the lack of major differences in their reproductive system. The assignment of distinctive morphospecies, and thus existing taxonomic names, to genetically defined evolutionary lineages is premature and arbitrary to some extent at this stage. © 2013 The Linnean Society of London     

SpedeSTEM: a rapid and accurate method for species delimitation

We describe a software package (SpedeSTEM) that allows researchers to conduct a species delimitation analysis using intraspecific genetic data. Our method operates under the assumption that a priori information regarding group membership is available, for example that samples are drawn from some number of described subspecies, races or distinct morphotypes. SpedeSTEM proceeds by calculating the maximum likelihood species tree from all hierarchical arrangements of the sampled alleles and uses information theory to quantify the model probability of each permutation. SpedeSTEM is tested here against empirical and simulated data; results indicate that evolutionary lineages that diverged as few as 0.5N generations in the past can be validated as distinct using sequence data from little as five loci. This work enables speciation investigations to identify lineages that are evolutionarily distinct and thus have the potential to form new species before these lineages acquire secondary characteristics such as reproductive isolation or morphological differentiation that are commonly used to define species.     

A general framework to quantify and compare ecological impacts under temporal dynamics

Biodiversity is diminishing at alarming rates due to multiple anthropogenic drivers. To mitigate these drivers, their impacts must be quantified accurately and comparably across drivers. To enable that, we present a generally applicable framework introducing fundamental principles of ecological impact quantification, including the quantification of interactions between multiple drivers. The framework contrasts biodiversity variables in impacted against those in unimpacted or other reference situations while accounting for their temporal dynamics through modelling. Properly accounting for temporal dynamics reduces biases in impact quantification and comparison. The framework addresses key questions around ecological impacts in global change science, namely, how to compare impacts under temporal dynamics across stressors, how to account for stressor interactions in such comparisons, and how to compare the success of management actions over time.     

Plant species delimitation: A comparison of morphological and molecular markers

Abstract

Species delimitation is fundamental in many areas of biology. Despite its importance, there is no agreement on criteria for species delimitation mostly due to divergence on the point of view adopted by the different biological disciplines. Two main groups of diagnostic characters are commonly used to distinguish species: the traditional morphological ones and the molecular ones. Field species recognition and sampling are generally based on morphological characters, but they can either fail to discriminate species and mask the presence of cryptic species or discriminate different species while in reality there is only one. To overcome this problem it is common to compare clusters obtained on the basis of the observed polymorphism of both characters, and to analyse their agreement. Here we compile a set of studies that have examined species delimitation with both markers. This provides a review of the different morphological and molecular markers, and of the sampling strategy and clustering methodology generally employed to delimitate species. Some conclusions are drawn with regard to species delimitation, when comparing diagnostic morphological and molecular markers.     

Single-locus species delimitation: a test of the mixed Yule-coalescent model, with an empirical application to Philippine round-leaf bats

Prospects for a comprehensive inventory of global biodiversity would be greatly improved by automating methods of species delimitation. The general mixed Yule-coalescent (GMYC) was recently proposed as a potential means of increasing the rate of biodiversity exploration. We tested this method with simulated data and applied it to a group of poorly known bats (Hipposideros) from the Philippines. We then used echolocation call characteristics to evaluate the plausibility of species boundaries suggested by GMYC. In our simulations, GMYC performed relatively well (errors in estimated species diversity less than 25%) when the product of the haploid effective population size (N(e)) and speciation rate (SR; per lineage per million years) was less than or equal to 10(5), while interspecific variation in N(e) was twofold or less. However, at higher but also biologically relevant values of N(e) × SR and when N(e) varied tenfold among species, performance was very poor. GMYC analyses of mitochondrial DNA sequences from Philippine Hipposideros suggest actual diversity may be approximately twice the current estimate, and available echolocation call data are mostly consistent with GMYC delimitations. In conclusion, we consider the GMYC model useful under some conditions, but additional information on N(e), SR and/or corroboration from independent character data are needed to allow meaningful interpretation of results.     

Using metabarcoding to compare the suitability of two blood‐feeding leech species for sampling mammalian diversity in North Borneo

The application of high‐throughput sequencing (HTS) for metabarcoding of mixed samples offers new opportunities in conservation biology. Recently, the successful detection of prey DNA from the guts of leeches has raised the possibility that these, and other blood‐feeding invertebrates, might serve as useful samplers of mammals. Yet little is known about whether sympatric leech species differ in their feeding preferences, and whether this has a bearing on their relative suitability for monitoring local mammalian diversity. To address these questions, we collected spatially matched samples of two congeneric leech species Haemadipsa picta and Haemadipsa sumatrana from lowland rainforest in Borneo. For each species, we pooled ~500 leeches into batches of 10 individuals, performed PCR to target a section of the mammalian 16S rRNA locus and undertook sequencing of amplicon libraries using an Illumina MiSeq. In total, we identified sequences from 14 mammalian genera, spanning nine families and five orders. We found greater numbers of detections, and higher diversity of OTUs, in H. picta compared with H. sumatrana, with rodents only present in the former leech species. However, comparison of samples from across the landscape revealed no significant difference in mammal community composition between the leech species. We therefore suggest that H. picta is the more suitable iDNA sampler in this degraded Bornean forest. We conclude that the choice of invertebrate sampler can influence the detectability of different mammal groups and that this should be accounted for when designing iDNA studies.     

A pre-processing pipeline to quantify,visualize, and reduce technical variation in protein microarray studies

Technical variation, or variation from non-biological sources, is present in most laboratory assays. Correcting for this variation enables analysts to extract a biological signal that informs questions of interest. However, each assay has different sources and levels of technical variation, and the choice of correction methods can impact downstream analyses. Compared to similar assays such as DNA microarrays, relatively few methods have been developed and evaluated for protein microarrays, a versatile tool for measuring levels of various proteins in serum samples. Here, we propose a pre-processing pipeline to correct for some common sources of technical variation in protein microarrays. The pipeline builds upon an existing normalization method by using controls to reduce technical variation. We evaluate our method using data from two protein microarray studies and by simulation. We demonstrate that pre-processing choices impact the fluorescent-intensity based ranks of proteins, which in turn, impact downstream analysis.     

Biogeography and species delimitation of the rheophilic suckermouth catfish genus Pseudolithoxus (Siluriformes: Loricariidae), with the description of a new species from the Brazilian Amazon

The rapids-dwelling suckermouth catfish genus Pseudolithoxus was previously only known from the Guiana-Shield-draining Orinoco and Casiquiare river systems of Colombia and Venezuela, but new records have expanded this range considerably further into the Amazon basin of Brazil, and include occurrences from rivers draining the northern Brazilian Shield. These highly disjunct records are now placed in an evolutionary and phylogeographic context using a dated species tree constructed from mitochondrial (Cytb) and nuclear (RAG1) gene sequence data. Due to mito-nuclear discordance, we also delimit the putative species using statistical coalescent models and a range of additional metrics. We infer that at least two species of Pseudolithoxus are present in the Amazon basin: P. nicoi, previously only recorded from the río Casiquiare, but now also reported from the upper rio Negro, and a new species, which we describe herein from south-draining Guiana Shield and north-draining Brazilian Shield. Our data reject a simple model of Miocene vicariance in the group following uplift of the Uaupés Arch separating the Orinoco and Amazon systems, and instead suggest more complex dispersal scenarios through palaeo-connections in the Pliocene and also via the contemporary rio Negro and rio Madeira in the late Pleistocene.

www.zoobank.org/urn:lsid:zoobank.org:pub:46A6BEC1-1A1B-4244-80-A6-FD9CF8DA0384.