Comparative bioacoustics: a roadmap for quantifying and comparing animal sounds across diverse taxa

Animals produce a wide array of sounds with highly variable acoustic structures. It is possible to understand the causes and consequences of this variation across taxa with phylogenetic comparative analyses. Acoustic and evolutionary analyses are rapidly increasing in sophistication such that choosing appropriate acoustic and evolutionary approaches is increasingly difficult. However, the correct choice of analysis can have profound effects on output and evolutionary inferences. Here, we identify and address some of the challenges for this growing field by providing a roadmap for quantifying and comparing sound in a phylogenetic context for researchers with a broad range of scientific backgrounds. Sound, as a continuous, multidimensional trait can be particularly challenging to measure because it can be hard to identify variables that can be compared across taxa and it is also no small feat to process and analyse the resulting high-dimensional acoustic data using approaches that are appropriate for subsequent evolutionary analysis. Additionally, terminological inconsistencies and the role of learning in the development of acoustic traits need to be considered. Phylogenetic comparative analyses also have their own sets of caveats to consider. We provide a set of recommendations for delimiting acoustic signals into discrete, comparable acoustic units. We also present a three-stage workflow for extracting relevant acoustic data, including options for multivariate analyses and dimensionality reduction that is compatible with phylogenetic comparative analysis. We then summarize available phylogenetic comparative approaches and how they have been used in comparative bioacoustics, and address the limitations of comparative analyses with behavioural data. Lastly, we recommend how to apply these methods to acoustic data across a range of study systems. In this way, we provide an integrated framework to aid in quantitative analysis of cross-taxa variation in animal sounds for comparative phylogenetic analysis. In addition, we advocate the standardization of acoustic terminology across disciplines and taxa, adoption of automated methods for acoustic feature extraction, and establishment of strong data archival practices for acoustic recordings and data analyses. Combining such practices with our proposed workflow will greatly advance the reproducibility, biological interpretation, and longevity of comparative bioacoustic studies.     

STBase: One Million Species Trees for Comparative Biology

Comprehensively sampled phylogenetic trees provide the most compelling foundations for strong inferences in comparative evolutionary biology. Mismatches are common, however, between the taxa for which comparative data are available and the taxa sampled by published phylogenetic analyses. Moreover, many published phylogenies are gene trees, which cannot always be adapted immediately for species level comparisons because of discordance, gene duplication, and other confounding biological processes. A new database, STBase, lets comparative biologists quickly retrieve species level phylogenetic hypotheses in response to a query list of species names. The database consists of 1 million single- and multi-locus data sets, each with a confidence set of 1000 putative species trees, computed from GenBank sequence data for 413,000 eukaryotic taxa. Two bodies of theoretical work are leveraged to aid in the assembly of multi-locus concatenated data sets for species tree construction. First, multiply labeled gene trees are pruned to conflict-free singly-labeled species-level trees that can be combined between loci. Second, impacts of missing data in multi-locus data sets are ameliorated by assembling only decisive data sets. Data sets overlapping with the user’s query are ranked using a scheme that depends on user-provided weights for tree quality and for taxonomic overlap of the tree with the query. Retrieval times are independent of the size of the database, typically a few seconds. Tree quality is assessed by a real-time evaluation of bootstrap support on just the overlapping subtree. Associated sequence alignments, tree files and metadata can be downloaded for subsequent analysis. STBase provides a tool for comparative biologists interested in exploiting the most relevant sequence data available for the taxa of interest. It may also serve as a prototype for future species tree oriented databases and as a resource for assembly of larger species phylogenies from precomputed trees.     

Genomic regulatory regions: insights from comparative sequence analysis

Comparative sequence analysis is contributing to the identification and characterization of genomic regulatory regions with functional roles. It is effective because functionally important regions tend to evolve at a slower rate than do less important regions. The choice of species for comparative analysis is crucial: shared ancestry of a clade of species facilitates the discovery of genomic features important to that clade, whereas increased sequence divergence improves the resolution at which features can be discovered. Recent studies suggest that comparative analyses are useful for all branches of life and that, in the near future, large-scale mammalian comparative sequence analysis will provide the best approach for the comprehensive discovery of human regulatory elements.     

ProfileGrids as a new visual representation of large multiple sequence alignments: a case study of the RecA protein family

Background  

Multiple sequence alignments are a fundamental tool for the comparative analysis of proteins and nucleic acids. However, large data sets are no longer manageable for visualization and investigation using the traditional stacked sequence alignment representation.     

Estimating evolution of temporal sequence changes: a practical approach to inferring ancestral developmental sequences and sequence heterochrony

Developmental biology often yields data in a temporal context. Temporal data in phylogenetic systematics has important uses in the field of evolutionary developmental biology and, in general, comparative biology. The evolution of temporal sequences, specifically developmental sequences, has proven difficult to examine due to the highly variable temporal progression of development. Issues concerning the analysis of temporal sequences and problems with current methods of analysis are discussed. We present here an algorithm to infer ancestral temporal sequences, quantify sequence heterochronies, and estimate pseudoreplicate consensus support for sequence changes using Parsimov-based genetic inference [PGi]. Real temporal developmental sequence data sets are used to compare PGi with currently used approaches, and PGi is shown to be the most efficient, accurate, and practical method to examine biological data and infer ancestral states on a phylogeny. The method is also expandable to address further issues in developmental evolution, namely modularity.     

Facial morphology as determined by anthropometry: keeping it simple

Anthropometry remains an efficient, noninvasive method for describing craniofacial morphology in spite of the appearance of more sophisticated technologies. The major advantage afforded by anthropometry is its technical simplicity, a fact which makes it a readily available tool for evaluating patients, planning facial surgery, or delineating basic features of craniofacial syndromes. Anthropometry lacks the detail of more powerful technologies, but it is better suited for populational studies because of the availability of comparative, normal databases. The standard z-scores produced by such comparisons lend themselves to multivariate analysis. This type of comparative analysis is not yet possible for computerized tomography, three-dimensional imaging, or photogrammetry. To illustrate the utility of this technique an example is cited from an ongoing study of hypohidrotic ectodermal dysplasia (HED) in which anthropometry reveals details of facial morphology overlooked in previous studies. These include the presence of reduced facial height and a striking reduction in the size of the facial features in spite of the fact that facial widths are comparatively normal. Gene carriers show a similar though nonidentical pattern of defects. Like all morphometric approaches, anthropometry has its limitations. Well-designed protocols minimize these limitations by incorporating multiple facial dimensions in the analysis and by emphasizing careful collection of data with standard instruments and methodology.     

An evaluation of primers for detecting denitrifiers via their functional genes

Microbial populations provide nitrogen cycling ecosystem services at the nexus of agriculture, environmental quality and climate change. Denitrification, in particular, impacts socio-environmental systems in both positive and negative ways, through reduction of aquatic and atmospheric nitrogen pollution, but also reduction of soil fertility and production of greenhouse gases. However, denitrification rates are quite variable in time and space, and therefore difficult to model. Microbial ecology is working to improve the predictive ecology of denitrifiers by quantifying and describing the diversity of microbial functional groups. However, metagenomic sequencing has revealed previously undescribed diversity within these functional groups, and highlighted a need to reevaluate coverage of existing DNA primers for denitrification functional genes. We provide here a comprehensive in silico evaluation of primer sets that target diagnostic genes in the denitrification pathway. This analysis makes use of current DNA sequence data available for each functional gene. It contributes a comparative analysis of the strengths and limitations of each primer set for describing denitrifier functional groups. This analysis identifies genes for which development of new tools is needed, and aids in interpretation of existing datasets, both of which will facilitate application of molecular methods to further develop the predictive ecology of denitrifiers.     

Serial analysis of gene expression (SAGE): unraveling the bioinformatics tools

Serial analysis of gene expression (SAGE) is a powerful technique that can be used for global analysis of gene expression. Its chief advantage over other methods is that it does not require prior knowledge of the genes of interest and provides qualitative and quantitative data of potentially every transcribed sequence in a particular cell or tissue type. This is a technique of expression profiling, which permits simultaneous, comparative and quantitative analysis of gene-specific, 9- to 13-basepair sequences. These short sequences, called SAGE tags, are linked together for efficient sequencing. The sequencing data are then analyzed to identify each gene expressed in the cell and the levels at which each gene is expressed. The main benefit of SAGE includes the digital output and the identification of novel genes. In this review, we present an outline of the method, various bioinformatics methods for data analysis and general applications of this important technology.     

Tackling the plant proteome: practical approaches, hurdles and experimental tools

The study of complex biological questions through comparative proteomics is becoming increasingly attractive to plant biologists as the rapidly expanding plant genomic and expressed sequence tag databases provide improved opportunities for protein identification. This review focuses on practical issues associated with comparative proteomic analysis, including the challenges of effective protein extraction and separation from plant tissues, the pros and cons of two-dimensional gel-based analysis and the problems of identifying proteins from species that are not recognized models for functional genomic studies. Specific points are illustrated using data from an ongoing study of the tomato and pepper fruit proteomes.     

Theoretical and Applied Aspects of Comparative Genomics of Vertebrates

The Human Genome Project stimulated the development of efficient strategies and relevant hardware for complete genome sequencing. The comparative genomic approach extends the possibilities of using the sequencing data to identify new genes or conserved regulatory regions by means of nucleotide sequence alignment of the particular regions of the mouse and human genomes, or to trace the evolutionary events resulting in the genome structure of modern mammals. The review focuses on the use of new molecular cytogenetic methods along with computer-aided analysis of the genomes in vertebrates. Several factors hindering data analysis are considered. The currently available information on gene evolution rate inferred from comparative genomic data is presented. The origin and evolution of the genomes of several species are discussed.     

Fundamental and applied aspects of comparative genomics of vertebrates

The Human Genome Project stimulated the development of efficient strategies and relevant hardware for complete genome sequencing. The comparative genomic approach extends the possibilities of using the sequencing data to identify new genes or conserved regulatory regions by means of nucleotide sequence alignment of the particular regions of the mouse and human genomes, or to trace the evolutionary events resulting in the genome structure of modern mammals. The review focuses on the use of new molecular cytogenetic methods along with computer-aided analysis of the genomes in vertebrates. Several factors hindering data analysis are considered. The currently available information on gene evolution rate inferred from comparative genomic data is presented. The origin and evolution of the genomes of several species are discussed.     

Insights from the protein sequence and structure analysis of PgHsc70 and OsHsp70 genes

Heat shock proteins are induced in a wide range of abiotic and biotic stresses. They are well known for cellular chaperone activities and play an important role in protecting plants through regulation of homeostasis and survival. A comprehensive characterization and comparative analysis of the Hsp70 family members within the closely related plant species helps in better interpretation of these proteins'' contribution to cell function and response to specific environmental stresses. Therefore, it is of interest to glean insights from the protein sequence analysis of PgHsc 70 and OsHsp70 genes. Thus, we document data from the sequence and structure analysis of PgHsc 70 and OsHsp 70 gene a.     

Assignment of position-specific error probability to primary DNA sequence data.

DNA sequence predicted from polyacrylamide gel-based technologies is inaccurate because of variations in the quality of the primary data due to limitations of the technology, and to sequence-specific variations due to nucleotide interactions within the DNA molecule and with the gel. The ability to recognize the probability of error in the primary data will be useful in reconstructing the target sequence of a DNA sequencing project, and in estimating the accuracy of the final sequence. This paper describes the use of linear discriminant analysis to assign position-specific probabilities of incorrect, over- and under-prediction of nucleotides for each predicted nucleotide position in primary sequence data generated by a gel-based DNA sequencing technology. Using this method, most of the error potential in primary sequence data can be assigned to a limited number of discrete positions. The use of probability values in the sequence reconstruction process, and in estimating the accuracy of consensus sequence determination is described.     

Identification of Soybean Genes Involved in Circadian Clock Mechanism and Photoperiodic Control of Flowering Time by In Silico Analyses

Glycine max is a photoperiodic short-day plant and the practical consequence of the response is latitude and sowing period limitations to commercial crops. Genetic and physiological studies using the model plants Arabidopsis thaliana and rice （Oryza sativa） have uncovered several genes and genetic pathways controlling the process, however information about the corresponding pathways in legumes is scarce. Data mining prediction methodologies, including multiple sequence alignment, phylogeneUc analysis, bioinformaUcs expression and sequence motif pattern identification, were used to identify soybean genes involved in day length perception and photoperiodic flowering induction. We have investigated approximately 330 000 sequences from open-access databases and have identified all bona fide central oscillator genes and circadian photoreceptors from A. thaliana in soybean sequence databases. We propose a working model for the photoperiodic control of flowering time in G. max, based on the identified key components. These results demonstrate the power of comparative genomics between model systems and crop species to elucidate the several aspects of plant physiology and metabolism.     

DraGnET: Software for storing,managing and analyzing annotated draft genome sequence data

Background  

New "next generation" DNA sequencing technologies offer individual researchers the ability to rapidly generate large amounts of genome sequence data at dramatically reduced costs. As a result, a need has arisen for new software tools for storage, management and analysis of genome sequence data. Although bioinformatic tools are available for the analysis and management of genome sequences, limitations still remain. For example, restrictions on the submission of data and use of these tools may be imposed, thereby making them unsuitable for sequencing projects that need to remain in-house or proprietary during their initial stages. Furthermore, the availability and use of next generation sequencing in industrial, governmental and academic environments requires biologist to have access to computational support for the curation and analysis of the data generated; however, this type of support is not always immediately available.     

Multiple sequence alignments of partially coding nucleic acid sequences

Background  

High quality sequence alignments of RNA and DNA sequences are an important prerequisite for the comparative analysis of genomic sequence data. Nucleic acid sequences, however, exhibit a much larger sequence heterogeneity compared to their encoded protein sequences due to the redundancy of the genetic code. It is desirable, therefore, to make use of the amino acid sequence when aligning coding nucleic acid sequences. In many cases, however, only a part of the sequence of interest is translated. On the other hand, overlapping reading frames may encode multiple alternative proteins, possibly with intermittent non-coding parts. Examples are, in particular, RNA virus genomes.     

PLAN: a web platform for automating high-throughput BLAST searches and for managing and mining results

Background  

BLAST searches are widely used for sequence alignment. The search results are commonly adopted for various functional and comparative genomics tasks such as annotating unknown sequences, investigating gene models and comparing two sequence sets. Advances in sequencing technologies pose challenges for high-throughput analysis of large-scale sequence data. A number of programs and hardware solutions exist for efficient BLAST searching, but there is a lack of generic software solutions for mining and personalized management of the results. Systematically reviewing the results and identifying information of interest remains tedious and time-consuming.     

The phylogeography of dusky dolphins (Lagenorhynchus obscurus): a critical examination of network methods and rooting procedures

We investigated the phylogeography and evolutionary history of dusky dolphins (Lagenorhynchus obscurus) using DNA sequences of the full mitochondrial cytochrome b gene in 124 individuals from the putative stocks off Peru, Argentina and Southwest Africa. While genetic differentiation within oceans is surprisingly low, there is no evidence for recent female gene flow between Atlantic and Pacific waters. Highest genetic variability in terms of sequence divergence and number of haplotypes is found in the Atlantic. Our analyses also indicate that the eastern South Pacific dusky dolphins stock should be considered a separate management unit. Given the high level of mortality experienced by the Peruvian dusky dolphin in local fishery activities, these findings have important implications for an objective management of the species. Furthermore, we analysed our mitochondrial sequence data with several widely used network estimation and rooting methods. The resulting intraspecific gene genealogies and rooting inferences exhibited substantial differences, underlying the limitations of some algorithms. Given that scientific hypotheses and management decisions depend strongly on inferred tree or network topologies, there is a clear need for a systematic comparative analysis of available methods. Finally, the present study indicates that (i) the dusky and the Pacific white-sided dolphins are sister species and (ii) not only the Westwind Drift hypothesis but also other models of dispersion are compatible with the current geographical distribution of dusky dolphins.