Principles and methods of geometric morphometrics

The basic concepts, notions and methods of geometric morphometrics (GM) are considered. This approach implies multivariate analysis of landmark coordinates located following certain rules on the surface of a morphological object. The aim of GM is to reveal differences between morphological objects by their shapes as such, the "size factor" being excluded. The GM is based on the concept of Kendall's space (KS) defined as a hypersphere with points distributed on its surface. These points are the shapes defined as aligned landmark configurations. KS is a non-Euclidian space, its metrics called Procrustes is defined by landmark configuration of a reference shape relative to which other shapes are aligned and compared. The differences among shapes are measured as Procrustes distances between respective points. For the linear methods of multivariate statistics to be applied to comparison of shapes, the respective points are projected onto the tangent plane (tangent space), the tangent point being defined by the reference. There are two principal methods of shape comparisons in GM: the Procrustes superimposition (a version of the least squares analysis) and thin-plate spline analysis. In the first case, Procrustes residuals are the outcome shape variables which remain after isometric alignment of the shapes being compared. Their summation over all landmarks yields Procrustes distances among these shapes. The Procrustes distances can be used in multivariate analyses just as the Euclidian distances. In the second case, the shapes are fitted to the references by stretching/compressing and shearing until complete identity of their landmark configurations. Eigenvectors of resulting bending energy matrix are defined as new shape variables, principal warps which yield another shape space with the origin defined by the reference. Projections of the shapes being compared onto principal warps yield partial warps, and their covariance matrix decomposition into eigenvectors yields relative warps which are similar to principal components (in particular, they are mutually orthogonal). Both partial and relative warps can be used in many multivariate statistic analyses as quantitative shape variables. Results of thin-plate spline analysis can be represented graphically by transformation grid which displays type, amount and localization of the shape differences. Basis rules of sample composition and landmark positioning to be used in GM are considered. At present, rigid (with minimal degrees of freedom) 2D morphological objects are most suitable for GM applications. It is important to recognize three type of real landmarks, and additionally semi-landmarks and "virtual" landmarks. Some procedures of thin-plate spline analysis are considered exemplified by some study cases, as well as applications of some standard multivariate methods to GM results. They make it possible to evaluate correlation between different shapes, as well as between a shape and some non-shape variables (linear measurements etc); to evaluate the differences among organisms by shape of a morphological structure; to identify landmarks which most accounted for both correlation and differences between the shapes. An annotated list of most popular softwares for GM is provided.     

GEOMETRIC MORPHOMETRICS OF THE SKULL ROOF OF STEREOSPONDYLS (AMPHIBIA: TEMNOSPONDYLI)

Abstract:  Geometric morphometric analysis using relative warps is applied to the skull roof of 62 species of stereospondyls and their closest outgroups (i.e. basal archegosauriforms) from among temnospondyl amphibians. Twenty-one landmarks and five taxonomic groups are used for comparisons. Their skull evolution is quantified in a morphospace defined by two relative warps axes. The majority of groups show poor concordance between morphological and phylogenetic distances. The only exception is represented by Yates and Warren's study of stereospondyl relationships, in which concordance is high. Only basal archegosauriforms and rhinesuchids show significant overlap in morphospace, although this might be due to low sample sizes. Regression of estimated mean disparity against taxon sample size shows that species within both the trematosauroid and the rhytidostean groups are more widely dispersed in morphospace than species belonging to any of the remaining stereospondyl groups. Stereospondyl skull evolution was characterized by divergence between major clades and convergence within those clades. Changes in patterns of morphospace occupation through time agree with the hypothesis of an 'explosive' radiation in the early Early Triassic, after the extinction of basal archegosauriforms at the end of the Permian.     

Evolution of shape in oligocene and mioceneNotocarinovalva (Ostracoda, Crustacea): A multivariate statistical study

Largely due to L. F. Bookstein, geometric morphometrics has been developed as an amalgamation of techniques drawn from mathematical statistics, non-Euclidean geometry and computer graphics, applied to labelled points (landmarks) and the biological images upon which they are registered. A tool of fundamental importance is the method of interpolation known as the thin-plate spline. Bookstein's sample-oriented procedure of relative warps, applied to data composed of coordinate pairs observed on eight landmarks on three samples of two species of fossil marine ostracods (bivalved microcrustacean),Notocarinovalva airella andN. yulecartensis, separated in time, is used to obtain weight matrices, which, when appropriately partitioned, constitute the familiar data matrices of multivariate statistical analysis. Standard multivariate statistical analysis of samples may be carried out in the tangent space to shape space at the Procrustes average shape. Linear discriminant function scores were used for assessing, approximately, the evolutionary relevance of shape change and change in size (based on standard distance measures) in the species from the upper and lower Oligocene and lower middle Miocene of Victoria, Australia. The findings indicate that a small, though consistent, change in non-uniform shape of the carapace has taken place. The analysis indicates that the evolution in shape of the carapace could have been caused by random genetic drift in moderately large populations, tempered by weak selection.     

Computing the uniform component of shape variation

Any change in shape of a configuration of landmark points in two or three dimensions includes a uniform component, a component that is a wholly linear (affine) transformation. The formulas for estimating this component have been standardized for two-dimensional data but not for three-dimensional data. We suggest estimating the component by way of the complementarity between the uniform component and the space of partial warps. The component can be estimated by regression in either one space or the other: regression on the partial warps, followed by their removal, or regression on a basis for the uniform component itself. Either of the new methods can be used for both two- and three-dimensional landmark data and thus generalize Bookstein's (1996, pages 153-168 in Advances in morphometrics [L. F. Marcus et al., eds.], Plenum, New York) linearized Procrustes formula for estimating the uniform component in two dimensions.     

黄斑蜂族的系统学研究：几何形态测量方法（膜翅目： 切叶蜂科）

利用采集于土耳其安纳托利亚中部(Middle Anatolia) 的切叶蜂科6个种92头标本,选取虫体上的30个特征点(landmarks)(其中前翅16个,后翅6个,头部8个),利用UPGMA等数值分类方法对不同属和种的亲缘关系进行了探讨。据此对一些种的归属和一些属的划分提出了一些与传统分类观点相同或不同的见解,但各种分析方法的结果都支持切叶蜂族Megachilini起源的单系性。     

A taxonomic study of phlebioid fungi (Basidiomycota)

The genus Phlebia has long been regarded as a polyphyletic or paraphyletic taxon, including distinct groups of more closely related species. Consequently, the delimitation of the genus has been given different interpretations and several rearrangements have been proposed by various authors. In the present study, DNA sequences (25S, rDNA) were obtained for twenty species of the genus Phlebia and a phylogenetic analysis was performed. Because of the presumably paraphyletic nature of the genus, different outgroups were used for different sets of taxa. A core group of species, including the type Ph. radiata , is well distinguished. For the delimitation of the genus, however, the wider scope of a more weakly supported clade is proposed. This clade also includes Phlebiopsis gigantea and its combination in Phlebia is reaffirmed. Two species, Ph. griseoflavescens and Ph. tristis , are distinctly separated from this clade and should be removed from the genus. Morphological characters were used in a separate phylogenetic analysis but the result did not conform with analysis from sequence data.     

DETECTING SHAPE VARIATION IN OAK LEAF MORPHOLOGY: A COMPARISON OF ROTATIONAL-FIT METHODS

Rotational-fit methods were used to examine shape variation in oak leaf morphology by reference to a set of 14 landmarks for each leaf. Within-tree, between-tree, and between-species variations were examined. Generalized least-squares and generalized resistant-fit analyses revealed patterns of landmark variation that could be related to leaf architecture. The two species (Quercus palustris and Q. velutina) did not illustrate the same patterns of within-tree variability nor did they reveal similar degrees of between-tree variability. Incorporating an affine transformation algorithm resulted in little insight in some comparisons but suggested strong uniform shape change in other comparisons (especially between species). Resistant-fit mean square statistics were equally variable 1) in upper vs. lower crown samples within trees, 2) between trees within each species, and 3) between species. In addition, resistant-fit mean square statistics were found to be a poor measure of similarity, whether derived by comparison to a single reference object or by way of pairwise comparisons. Fundamental species differences in leaf shape are suggested by relationships among particular sets of landmarks, although overall shape differences cannot be explained fully by these analyses.     

Geometric morphometric analysis of Eysarcoris guttiger,E.annamita and E.ventralis (Hemiptera: Pentatomidae)

The genus Eysarcoris can be easily distinguished from other genera through the two spots in the basal angle of the scutellum.Nevertheless,Eysarcoris species show complex variances.Geometric morphometric methods have been increasingly applied to distinguish species and to define the boundary of genera among insects.In the present study,geometric morphometric approach was firstly employed to evaluate the shape variation of three characters (fore wing,hind wing and pygophore) of E.guttiger,E.annamita and E.ventralis using E.aeneus as outgroup to ascertain whether this approach is a reliable method for the taxonomy of Eysarcoris.Analysis was conducted on the landmarks of the three characters of these species.Multivariate regression of procrustes coordinates against centroid size was conducted to test the presence of allometry.Principal component analysis (PCA),canonical variate analysis (CVA)and cluster analysis were utilized to describe variations in shapes among the studied species.For all of the three characters,though PCA analysis showed some overlap among species,p-values for procrustes distance and mahalanobis distance were all less than 0.0001.The distribution of the three studied species corresponds with their species status.This study demonstrates that the geometric morphometrics of both the fore wing and the hind wing might represent a possible tool for the identification of species within this genus.     

Classification and typing methods of black-pigmented Gram-negative anaerobes

Abstract Until recently, black-pigmented Gram-negative anaerobes were classified as ‘black-pigmented Bacteroides ’. At present, 11 distinct species are recognized in this group. Because of major differences with Bacteroides fragilis , the type species of the genus Bacteroides , new genera have been proposed: Porphyromonas for three asaccharolytic species, and Prevotella for the saccharolytic species. Typing methods have been developed for some species of black-pigmented Gram-negative anaerobes. These include biotyping and serotyping, but relatively few types can be distinguished with these methods. Recently, DNA restriction endonuclease analysis has been used for typing of P. gingivalis, Pr. intermedia and P. endodontalis strains. Great heterogeneity was observed within all three species. This typing method can be useful for epidemiological studies.     

Phylogenic and phenotypic characterization of some Eubacterium-like isolates from human feces: description of Solobacterium moorei Gen. Nov., Sp. Nov

Three isolated strains from human feces were characterized by biochemical tests and 16S rDNA analysis. Phylogenetic analysis revealed that these isolated strains were members of the Clostridium subphylum of gram-positive bacteria. The phenotypic characters resembled those of the genus Eubacterium, but these strains were shown to be phylogenetically distant from the type species of the genus, Eubacterium limosum. The strains showed a specific phylogenetic association with Holdemania filiformis and Erysipelothrix rhusiopathiae. Based on a 16S rDNA sequence divergence of greater than 12% with H. filiformis and E. rhusiopathiae, a new genus, Solobacterium, is proposed for three strains, with one species, Solobacterium moorei. The type strain of Solobacterium moorei is JCM 10645T.     

Details of Retropositional Genome Dynamics That Provide a Rationale for a Generic Division: The Distinct Branching of All the Pacific Salmon and Trout (Oncorhynchus) from the Atlantic Salmon and Trout (Salmo)

Salmonid species contain numerous short interspersed repetitive elements (SINEs), known collectively as the HpaI family, in their genomes. Amplification and successive integration of individual SINEs into the genomes have occurred during the evolution of salmonids. We reported previously a strategy for determining the phylogenetic relationships among the Pacific salmonids in which these SINEs were used as temporal landmarks of evolution. Here, we provide evidence for extensive genomic rearrangements that involved retropositions and deletions in a common ancestor of all the Pacific salmon and trout. Our results provide genetic support for the recent phylogenetic reassignment of steelhead and related species from the genus Salmo to the genus Oncorhynchus. Several other informative loci identified by insertions of HpaI SINEs have been isolated, and previously proposed branching orders of the Oncorhynchus species have been confirmed. The authenticity of our phylogenetic tree is supported both by the isolation of more than two informative loci per branching point and by the congruence of all our data, which suggest that the period between succesive speciations was sufficiently long for each SINE that had been amplified in the original species to become fixed in all individuals of that species.     

链霉菌属菌株AS4.693和AS4.702的分类学研究

链霉菌属“Setae”种群原为北里孢菌属Kitasatosporia (Omura,1982)。1992年,Wellington根据16S rRNA序列分析结果将其并入链霉菌属,并建立“Setae”种群。通过对保藏的链霉菌AS 4.693、AS 4.702进行的形态学、细胞化学、分子遗传分类研究结果表明,它们与链霉菌属“Setae”种群中的典型种——西唐链霉菌Streptomyces setae(JCM3304’)具有相似性。它们的rDNA相似性高达100％,证明它们应归属于同一种群。AS.4.693定名为西唐链霉菌不规则新亚种Streptomyces setae subsp.irregularis nov.,AS 4.702定名为西唐链霉菌波曲弗氏新亚种Streptomyces setae subsp.flexuofradiae nov.。     

链孢囊菌属3种分子分类方法的对比

链孢囊菌属（Streptosporangium）是链孢囊菌科（Streptosporangiaceae）的模式属,包含13个种．种的鉴别通常是多相分类方法,其中尤以DNA同源性分析为国际公认的定种标准;全基因组杂交同源性在70%以下的为不同种．但在进行大量菌株的比对时操作比较复杂．本实验以链孢囊菌属15株标准菌株为实验菌株,选择适宜引物,对其基因组DNA的16S-23S rDNA 间隔区序列（ITS）和REP序列进行了扩增,分别获得了两种基因指纹图谱,并通过UPGMA聚类法构建了相应的进化距离树图．结果表明,对于链孢囊菌属中不同种的区分,两种基因图谱技术的分辨力相当,且两种方法呈现的菌株间同源性与DNA-DNA杂交的结果吻合,有望为链孢囊菌属分类学的研究提供简单、准确、快速的标准程序．     

THE CYANOBACTERIAL GENUS BRASILONEMA,GEN. NOV., A MOLECULAR AND PHENOTYPIC EVALUATION1

Twelve populations of filamentous, heterocytous scytonematoid cyanobacteria from subaerophytic (mainly epiphytic) habitats in subtropical and tropical Brazil (São Paulo) were studied. The populations form a uniform cluster, which differs from the traditional scytonematoid genera genetically and by several indistinct, but typical morphological characters (fasciculation of filaments, rare false branching). Two strains were isolated in monospecific cultures. Molecular analyses were performed on these strains from natural populations taken from bromeliad leaves and iron water pipes. Because the results from 16S rRNA gene sequencing indicated the separate position of this cyanobacterium at the generic level, a new genus, Brasilonema, is proposed with the type species Brasilonema bromeliae. The new genus is described using combined molecular and cytomorphological criteria, in accordance with the nomenclatorial recommendations of both the Bacteriological Code and the Botanical Code of Nomenclature (cf. Oren 2004 ). The ultrastructure of the type species was studied, and vacuole‐like structures in the cells were explained. The genus Brasilonema is commonly distributed, particularly in subaerophytic habitats from southeastern Brazil. The type species (B. bromeliae) has a rather curious ecology, living epiphytically (in phytothelmes) inside the vase‐shaped leaf rosettes of bromeliads.     

Characterization of <Emphasis Type="Italic">Penicillium</Emphasis> Species by Ribosomal DNA Sequencing and BOX,ERIC and REP-PCR Analysis

The genus Penicillium is one of the largest and widely distributed fungal genera described to date. As a result, its taxonomic classification and species discrimination within this genus has become complicated. In this study, 52 isolates that belonged to the Penicillum genus and other related genera were characterized using two DNA-based methods: (i) analysis of the nucleotide sequences of internal transcribed spacers in ribosomal DNA and (ii) analysis of DNA fingerprints that were generated by polymerase chain reactions with specific primers for enterobacterial repetitive intergenic consensus (ERIC) and repetitive extragenic palindromic (REP) sequences, and BOX elements. Using both methods, Penicillium species were discriminated from other fungal genera. Furthermore, Penicillium species that include strains which are used as biocontrol agents, such as P. glabrum, P. purpurogenum, and P. oxalicum, could be distinguished from other Penicillium species using these techniques. Based on our findings, we propose that a polyphasic approach that includes analysis of the nucleotide sequences of ribosomal DNA and detecting the presence of highly conserved, repeated nucleotide sequences can be used to determine the genetic relationships between different Penicillium species. Furthermore, we propose that our results can be used as a start point to develop a strategy to monitor the environmental presence of particular strains of Penicillium species when they are used as biocontrol agents.     

Geometric morphometries and the analysis of higher taxa: a case study based on the metendosternite of the Scarabaeoidea (Coleoptera)

A geometric morphometric analysis of families belonging to the Scarabaeoidea, using landmarks from three two-dimensional views (frontal, dorsal and lateral) of an internally situated rigid structure, the metendosternite, was carried out. Procrustes distances matrices were obtained to produce phenograms from the three analyses. Relative warp analyses were also performed and the first two relative warps for the three data sets plotted against each other. The metendosternite is an internally situated anatomical structure that has largely been neglected in past studies. However, it proved to be useful in geometric morphometric studies, because it is rigid and landmarks can be easily identified. Geometric morphometries is shown to be a powerful tool to identify phenetic relationships between higher level taxa, and the metendosternite is a new structure available for beetle systematisists.