Alignment-free Sequence Comparison Research Articles

SHARK enables sensitive detection of evolutionary homologs and functional analogs in unalignable and disordered sequences

Intrinsically disordered regions (IDRs) are structurally flexible protein segments with regulatory functions in multiple contexts, such as in the assembly of biomolecular condensates. Since IDRs undergo more rapid evolution than ordered regions, identifying homology of such poorly conserved regions remains challenging for state-of-the-art alignment-based methods that rely on position-specific conservation of residues. Thus, systematic functional annotation and evolutionary analysis of IDRs have been limited, despite them comprising ~21% of proteins. To accurately assess homology between unalignable sequences, we developed an alignment-free sequence comparison algorithm, SHARK (Similarity/Homology Assessment by Relating K-mers). We trained SHARK-dive, a machine learning homology classifier, which achieved superior performance to standard alignment-based approaches in assessing evolutionary homology in unalignable sequences. Furthermore, it correctly identified dissimilar but functionally analogous IDRs in IDR-replacement experiments reported in the literature, whereas alignment-based tools were incapable of detecting such functional relationships. SHARK-dive not only predicts functionally similar IDRs at a proteome-wide scale but also identifies cryptic sequence properties and motifs that drive remote homology and analogy, thereby providing interpretable and experimentally verifiable hypotheses of the sequence determinants that underlie such relationships. SHARK-dive acts as an alternative to alignment to facilitate systematic analysis and functional annotation of the unalignable protein universe.

Alignment-Free Sequence Comparison: A Systematic Survey From a Machine Learning Perspective.

The encounter of large amounts of biological sequence data generated during the last decades and the algorithmic and hardware improvements have offered the possibility to apply machine learning techniques in bioinformatics. While the machine learning community is aware of the necessity to rigorously distinguish data transformation from data comparison and adopt reasonable combinations thereof, this awareness is often lacking in the field of comparative sequence analysis. With realization of the disadvantages of alignments for sequence comparison, some typical applications use more and more so-called alignment-free approaches. In light of this development, we present a conceptual framework for alignment-free sequence comparison, which highlights the delineation of: 1) the sequence data transformation comprising of adequate mathematical sequence coding and feature generation, from 2) the subsequent (dis-)similarity evaluation of the transformed data by means of problem-specific but mathematically consistent proximity measures. We consider coding to be an information-loss free data transformation in order to get an appropriate representation, whereas feature generation is inevitably information-lossy with the intention to extract just the task-relevant information. This distinction sheds light on the plethora of methods available and assists in identifying suitable methods in machine learning and data analysis to compare the sequences under these premises.

Biochemical Property Based Positional Matrix: A New Approach Towards Genome Sequence Comparison.

The growth of the genome sequence has become one of the emerging areas in the study of bioinformatics. It has led to an excessive demand for researchers to develop advanced methodologies for evolutionary relationships among species. The alignment-free methods have been proved to be more efficient and appropriate related to time and space than existing alignment-based methods for sequence analysis. In this study, a new alignment-free genome sequence comparison technique is proposed based on the biochemical properties of nucleotides. Each genome sequence can be distributed in four parameters to represent a 21-dimensional numerical descriptor using the Positional Matrix. To substantiate the proposed method, phylogenetic trees are constructed on the viral and mammalian datasets by applying the UPGMA/NJ clustering method. Further, the results of this method are compared with the results of the Feature Frequency Profiles method, the Positional Correlation Natural Vector method, the Graph-theoretic method, the Multiple Encoding Vector method, and the Fuzzy Integral Similarity method. In most cases, it is found that the present method produces more accurate results than the prior methods. Also, in the present method, the execution time for computation is comparatively small.

Interpreting alignment-free sequence comparison: what makes a score a good score?

Alignment-free methods are alternatives to alignment-based methods when searching sequence data sets. The output from an alignment-free sequence comparison is a similarity score, the interpretation of which is not straightforward. We propose objective functions to interpret and calibrate outputs from alignment-free searches, noting that different objective functions are necessary for different biological contexts. This leads to advantages: visualising and comparing score distributions, including those from true positives, may be a relatively simple method to gain insight into the performance of different metrics. Using an empirical approach with both DNA and protein sequences, we characterise different similarity score distributions generated under different parameters. In particular, we demonstrate how sequence length can affect the scores. We show that scores of true positive sequence pairs may correlate significantly with their mean length; and even if the correlation is weak, the relative difference in length of the sequence pair may significantly reduce the effectiveness of alignment-free metrics. Importantly, we show how objective functions can be used with test data to accurately estimate the probability of true positives. This can significantly increase the utility of alignment-free approaches. Finally, we have developed a general-purpose software tool called KAST for use in high-throughput workflows on Linux clusters.

Michael Waterman's Contributions to Computational Biology and Bioinformatics.

On the occasion of Dr. Michael Waterman's 80th birthday, we review his major contributions to the field of computational biology and bioinformatics including the famous Smith-Waterman algorithm for sequence alignment, the probability and statistics theory related to sequence alignment, algorithms for sequence assembly, the Lander-Waterman model for genome physical mapping, combinatorics and predictions of ribonucleic acid structures, word counting statistics in molecular sequences, alignment-free sequence comparison, and algorithms for haplotype block partition and tagSNP selection related to the International HapMap Project. His books Introduction to Computational Biology: Maps, Sequences and Genomes for graduate students and Computational Genome Analysis: An Introduction geared toward undergraduate students played key roles in computational biology and bioinformatics education. We also highlight his efforts of building the computational biology and bioinformatics community as the founding editor of the Journal of Computational Biology and a founding member of the International Conference on Research in Computational Molecular Biology (RECOMB).

Genome skimming approach reveals the gene arrangements in the chloroplast genomes of the highly endangered Crocus L. species: Crocus istanbulensis (B.Mathew) Rukšāns.

Crocus istanbulensis (B.Mathew) Rukšāns is one of the most endangered Crocus species in the world and has an extremely limited distribution range in Istanbul. Our recent field work indicates that no more than one hundred individuals remain in the wild. In the present study, we used genome skimming to determine the complete chloroplast (cp) genome sequences of six C. istanbulensis individuals collected from the locus classicus. The cp genome of C. istanbulensis has 151,199 base pairs (bp), with a large single-copy (LSC) (81,197 bp), small single copy (SSC) (17,524 bp) and two inverted repeat (IR) regions of 26,236 bp each. The cp genome contains 132 genes, of which 86 are protein-coding (PCGs), 8 are rRNA and 38 are tRNA genes. Most of the repeats are found in intergenic spacers of Crocus species. Mononucleotide repeats were most abundant, accounting for over 80% of total repeats. The cp genome contained four palindrome repeats and one forward repeat. Comparative analyses among other Iridaceae species identified one inversion in the terminal positions of LSC region and three different gene (psbA, rps3 and rpl22) arrangements in C. istanbulensis that were not reported previously. To measure selective pressure in the exons of chloroplast coding sequences, we performed a sequence analysis of plastome-encoded genes. A total of seven genes (accD, rpoC2, psbK, rps12, ccsA, clpP and ycf2) were detected under positive selection in the cp genome. Alignment-free sequence comparison showed an extremely low sequence diversity across naturally occurring C. istanbulensis specimens. All six sequenced individuals shared the same cp haplotype. In summary, this study will aid further research on the molecular evolution and development of ex situ conservation strategies of C. istanbulensis.

Numerical Characterization of DNA Sequences for Alignment-free Sequence Comparison - A Review.

Biological macromolecules, namely, DNA, RNA, and protein, have their building blocks organized in a particular sequence and the sequential arrangement encodes the evolutionary history of the organism (species). Hence, biological sequences have been used for studying evolutionary relationships among the species. This is usually carried out by Multiple Sequence Algorithms (MSA). Due to certain limitations of MSA, alignment-free sequence comparison methods were developed. The present review is on alignment-free sequence comparison methods carried out using the numerical characterization of DNA sequences. The graphical representation of DNA sequences by chaos game representation and other 2-dimensional and 3-dimensional methods are discussed. The evolution of numerical characterization from the various graphical representations and the application of the DNA invariants thus computed in phylogenetic analysis are presented. The extension of computing molecular descriptors in chemometrics to the calculation of a new set of DNA invariants and their use in alignment-free sequence comparison in an N-dimensional space and construction of phylogenetic trees are also reviewed. The phylogenetic tress constructed by the alignment-free sequence comparison methods using DNA invariants were found to be better than those constructed using alignment-based tools such as PHLYIP and ClustalW. One of the graphical representation methods is now extended to study viral sequences of infectious diseases for the identification of conserved regions to design peptidebased vaccines by combining numerical characterization and graphical representation.

Standardized Phylogenetic Evolutionary Analysis based on an Alignment Free Sequence Comparison Technique

Standardized Phylogenetic Evolutionary Analysis based on an Alignment Free Sequence Comparison Technique

Alignment-free sequence comparison for virus genomes based on location correlation coefficient.

Coronaviruses (especially SARS-CoV-2) are characterized by rapid mutation and wide spread. As these characteristics easily lead to global pandemics, studying the evolutionary relationship between viruses is essential for clinical diagnosis. DNA sequencing has played an important role in evolutionary analysis. Recent alignment-free methods can overcome the problems of traditional alignment-based methods, which consume both time and space. This paper proposes a novel alignment-free method called the correlation coefficient feature vector (CCFV), which defines a correlation measure of the L-step delay of a nucleotide location from its location in the original DNA sequence. The numerical feature is a 16×L-dimensional numerical vector describing the distribution characteristics of the nucleotide positions in a DNA sequence. The proposed L-step delay correlation measure is interestingly related to some types of L+1 spaced mers. Unlike traditional gene comparison, our method avoids the computational complexity of multiple sequence alignment, and hence improves the speed of sequence comparison. Our method is applied to evolutionary analysis of the common human viruses including SARS-CoV-2, Dengue virus, Hepatitis B virus, and human rhinovirus and achieves the same or even better results than alignment-based methods. Especially for SARS-CoV-2, our method also confirms that bats are potential intermediate hosts of SARS-CoV-2.

Alignment free sequence comparison methods and reservoir host prediction.

MotivationThe emergence and subsequent pandemic of the SARS-CoV-2 virus raised urgent questions about its origin and, particularly, its reservoir host. These types of questions are long-standing problems in the management of emerging infectious diseases and are linked to virus discovery programs and the prediction of viruses that are likely to become zoonotic. Conventional means to identify reservoir hosts have relied on surveillance, experimental studies and phylogenetics. More recently, machine learning approaches have been applied to generate tools to swiftly predict reservoir hosts from sequence data.ResultsHere, we extend a recent work that combined sequence alignment and a mixture of alignment-free approaches using a gradient boosting machines (GBMs) machine learning model, which integrates genomic traits (GT) and phylogenetic neighbourhood (PN) signatures to predict reservoir hosts. We add a more uniform approach by applying Machine Learning with Digital Signal Processing (MLDSP)-based structural patterns (M-SP). The extended model was applied to an existing virus/reservoir host dataset and to the SARS-CoV-2 and related viruses and generated an improvement in prediction accuracy.Availability and implementationThe source code used in this work is freely available at https://github.com/bill1167/hostgbms.Supplementary informationSupplementary data are available at Bioinformatics online.

A new graph-theoretic approach to determine the similarity of genome sequences based on nucleotide triplets

Methods of finding sequence similarity play a significant role in computational biology. Owing to the rapid increase of genome sequences in public databases, the evolutionary relationship of species becomes more challenging. But traditional alignment-based methods are found inappropriate due to their time-consuming nature. Therefore, it is necessary to find a faster method, which applies to species phylogeny. In this paper, a new graph-theory based alignment-free sequence comparison method is proposed. A complete-bipartite graph is used to represent each genome sequence based on its nucleotide triplets. Subsequently, with the help of the weights of edges of the graph, a vector descriptor is formed. Finally, the phylogenetic tree is drawn using the UPGMA algorithm. In the present case, the datasets for comparison are related to mammals, viruses, and bacteria. In most of the cases, the phylogeny in the present case is found to be more satisfactory as compared to earlier methods.

CRAFT: Compact genome Representation toward large-scale Alignment-Free daTabase.

Rapid developments in sequencing technologies have boosted generating high volumes of sequence data. To archive and analyze those data, one primary step is sequence comparison. Alignment-free sequence comparison based on k-mer frequencies offers a computationally efficient solution, yet in practice, the k-mer frequency vectors for large k of practical interest lead to excessive memory and storage consumption. We report CRAFT, a general genomic/metagenomic search engine to learn compact representations of sequences and perform fast comparison between DNA sequences. Specifically, given genome or high throughput sequencing data as input, CRAFT maps the data into a much smaller embedding space and locates the best matching genome in the archived massive sequence repositories. With 102-104-fold reduction of storage space, CRAFT performs fast query for gigabytes of data within seconds or minutes, achieving comparable performance as six state-of-the-art alignment-free measures. CRAFT offers a user-friendly graphical user interface with one-click installation on Windows and Linux operating systems, freely available at https://github.com/jiaxingbai/CRAFT. Supplementary data are available at Bioinformatics online.

Information Theory in Computational Biology: Where We Stand Today.

“A Mathematical Theory of Communication” was published in 1948 by Claude Shannon to address the problems in the field of data compression and communication over (noisy) communication channels. Since then, the concepts and ideas developed in Shannon’s work have formed the basis of information theory, a cornerstone of statistical learning and inference, and has been playing a key role in disciplines such as physics and thermodynamics, probability and statistics, computational sciences and biological sciences. In this article we review the basic information theory based concepts and describe their key applications in multiple major areas of research in computational biology—gene expression and transcriptomics, alignment-free sequence comparison, sequencing and error correction, genome-wide disease-gene association mapping, metabolic networks and metabolomics, and protein sequence, structure and interaction analysis.

The number of k-mer matches between two DNA sequences as a function of k and applications to estimate phylogenetic distances.

We study the number Nk of length-k word matches between pairs of evolutionarily related DNA sequences, as a function of k. We show that the Jukes-Cantor distance between two genome sequences-i.e. the number of substitutions per site that occurred since they evolved from their last common ancestor-can be estimated from the slope of a function F that depends on Nk and that is affine-linear within a certain range of k. Integers kmin and kmax can be calculated depending on the length of the input sequences, such that the slope of F in the relevant range can be estimated from the values F(kmin) and F(kmax). This approach can be generalized to so-called Spaced-word Matches (SpaM), where mismatches are allowed at positions specified by a user-defined binary pattern. Based on these theoretical results, we implemented a prototype software program for alignment-free sequence comparison called Slope-SpaM. Test runs on real and simulated sequence data show that Slope-SpaM can accurately estimate phylogenetic distances for distances up to around 0.5 substitutions per position. The statistical stability of our results is improved if spaced words are used instead of contiguous words. Unlike previous alignment-free methods that are based on the number of (spaced) word matches, Slope-SpaM produces accurate results, even if sequences share only local homologies.

Alignment-free genomic sequence comparison using FCGR and signal processing

BackgroundAlignment-free methods of genomic comparison offer the possibility of scaling to large data sets of nucleotide sequences comprised of several thousand or more base pairs. Such methods can be used for purposes of deducing “nearby” species in a reference data set, or for constructing phylogenetic trees.ResultsWe describe one such method that gives quite strong results. We use the Frequency Chaos Game Representation (FCGR) to create images from such sequences, We then reduce dimension, first using a Fourier trig transform, followed by a Singular Values Decomposition (SVD). This gives vectors of modest length. These in turn are used for fast sequence lookup, construction of phylogenetic trees, and classification of virus genomic data. We illustrate the accuracy and scalability of this approach on several benchmark test sets.ConclusionsThe tandem of FCGR and dimension reductions using Fourier-type transforms and SVD provides a powerful approach for alignment-free genomic comparison. Results compare favorably and often surpass best results reported in prior literature. Good scalability is also observed.

Afann: bias adjustment for alignment-free sequence comparison based on sequencing data using neural network regression

Alignment-free methods, more time and memory efficient than alignment-based methods, have been widely used for comparing genome sequences or raw sequencing samples without assembly. However, in this study, we show that alignment-free dissimilarity calculated based on sequencing samples can be overestimated compared with the dissimilarity calculated based on their genomes, and this bias can significantly decrease the performance of the alignment-free analysis. Here, we introduce a new alignment-free tool, Alignment-Free methods Adjusted by Neural Network (Afann) that successfully adjusts this bias and achieves excellent performance on various independent datasets. Afann is freely available at https://github.com/GeniusTang/Afann.

Alignment-Free Sequence Comparison With Multiple k Values.

Alignment-free sequence comparison approaches have become increasingly popular in computational biology, because alignment-based approaches are inefficient to process large-scale datasets. Still, there is no way to determine the optimal value of the critical parameter k for alignment-free approaches in general. In this article, we tried to solve the problem by involving multiple k values simultaneously. The method counts the occurrence of each k-mer with different k values in a sequence. Two weighting schemes, based on maximizing deviation method and genetic algorithm, are then used on these counts. We applied the method to enhance the three common alignment-free approaches D2, D2S, and D2*, and evaluated its performance on similarity search and functionally related regulatory sequences recognition. The enhanced approaches achieve better performance than the original approaches in all cases, and much better performance than some other common measures, such as Pcc, Eu, Ma, Ch, Kld, and Cos.

Benchmarking of alignment-free sequence comparison methods

BackgroundAlignment-free (AF) sequence comparison is attracting persistent interest driven by data-intensive applications. Hence, many AF procedures have been proposed in recent years, but a lack of a clearly defined benchmarking consensus hampers their performance assessment.ResultsHere, we present a community resource (http://afproject.org) to establish standards for comparing alignment-free approaches across different areas of sequence-based research. We characterize 74 AF methods available in 24 software tools for five research applications, namely, protein sequence classification, gene tree inference, regulatory element detection, genome-based phylogenetic inference, and reconstruction of species trees under horizontal gene transfer and recombination events.ConclusionThe interactive web service allows researchers to explore the performance of alignment-free tools relevant to their data types and analytical goals. It also allows method developers to assess their own algorithms and compare them with current state-of-the-art tools, accelerating the development of new, more accurate AF solutions.

Stein’s method for functions of multivariate normal random variables

Par le theoreme de l’application continue, si une suite $(\mathbf{W}_{n})_{n\geq1}$ de vecteurs aleatoires de dimension $d$ converge en loi vers une loi normale multivariee $\Sigma^{1/2}\mathbf{Z}$, alors la suite des variables aleatoires $(g(\mathbf{W}_{n}))_{n\geq1}$ converge en loi vers $g(\Sigma^{1/2}\mathbf{Z})$ si $g:\mathbb{R}^{d}\rightarrow\mathbb{R}$ est continue. Dans cet article, nous developpons la methode de Stein pour obtenir des bornes explicites sur la distance entre $g(\mathbf{W}_{n})$ et $g(\Sigma^{1/2}\mathbf{Z})$, pour des metriques lisses sur l’espaces des probabilites. Nous obtenons plusieurs bornes dans le cas ou la $j$-eme coordonnee de $\mathbf{W}_{n}$ est donnee par $W_{n,j}=\frac{1}{\sqrt{n}} \sum_{i=1}^{n}X_{ij}$, ou les $X_{ij}$ sont independants. En particulier, si $g$ verifie certains conditions de derivabilite et de croissance, nous obtenons une borne d’ordre $n^{-(p-1)/2}$, pour des fonctions-test lisses, si les $p$ premiers moments des $X_{ij}$ coincident avec ceux de la loi normale. Si $p$ est un entier pair et $g$ est une fonction paire, ce taux de convergence peut etre encore ameliore en $n^{-p/2}$. Nous montrons que ces taux de convergence sont d’ordre optimal. Nous appliquons nos bornes generales a quelques exemples, incluant l’approximation en loi de statistiques suivant asymptotiquement une loi du chi-deux; l’approximation d’esperances de fonctions lisses de variables aleatoires de loi binomiales ou de Poisson; des taux de convergence pour la methode $\delta$; et une approximation variance-gamma quantitative de la statistique $D_{2}^{*}$ pour la comparaison sans alignement, dans le cas de suites binaires.

Genome classification improvements based on k-mer intervals in sequences

Given the vast amount of genomic data, alignment-free sequence comparison methods are required due to their low computational complexity. k-mer based methods can improve comparison accuracy by extracting an effective feature of the genome sequences. The aim of this paper is to extract k-mer intervals of a sequence as a feature of a genome for high comparison accuracy. In the proposed method, we calculated the distance between genome sequences by comparing the distribution of k-mer intervals. Then, we identified the classification results using phylogenetic trees. We used viral, mitochondrial (MT), microbial and mammalian genome sequences to perform classification for various genome sets. We confirmed that the proposed method provides a better classification result than other k-mer based methods. Furthermore, the proposed method could efficiently be applied to long sequences such as human and mouse genomes.