Guanyang Zhang

The Use of Mean Instead of Smallest Interspecific Distances Exaggerates the Size of the “Barcoding Gap” and Leads to Misidentification

DNA barcoding is one of the best funded and most visible large-scale initiatives in systematic biology and has generated both much interest and controversy. DNA barcoding has also attracted significant support from foundations that had previously shown little interest in systematics. Yet, the project is controversial because many systematists feel that currently the conceptual foundation of DNA barcoding remains weak. This problem can only be alleviated through additional research that can lead to improved tools and concepts. Here, we scrutinize a key concept of DNA barcoding, the so-called barcoding gap (Meyer and Paulay, 2005), and use empirical data to document that it needs to be computed based on the smallest instead of the mean interspecific distances. In the literature on DNA barcoding, the “barcoding gap” (Meyer and Paulay, 2005) refers to the separation between mean intraand interspecific sequence variability for congeneric COI sequences. The barcoding gap is so essential to barcoding that a widely cited publication was dedicated to documenting these gaps across major metazoan taxa (Hebert et al., 2003b). It is also regularly mentioned in articles promoting barcoding to a broader audience (Check, 2005; Cognato and Caesar, 2006; Dasmahapatra and Mallet, 2006) and is one of the few metrics included in the Web-based identification system BOLD, “The Barcode of Life Data System,” which is a major identification tool for the DNA barcoding community (http://www.barcodinglife.org; Ratnasingham and Hebert, 2007). Large barcoding gaps are routinely used to predict DNA-barcoding success for the taxon under study (Hebert et al., 2003a, 2003b, 2004a, 2004b; Hogg and Hebert, 2004; Powers, 2004; Zehner et al., 2004; Armstrong and Ball, 2005; Ball et al., 2005; Barrett and Hebert, 2005; Lorenz et al., 2005; Saunders, 2005; Smith et al., 2005, 2006; Ward et al., 2005; Cywinska et al., 2006; Hajibabaei et al., 2006a, 2006b; Lefebure et al., 2006; Clare et al., 2007; Seifert et al., 2007). However, here we argue and document that barcoding gaps are currently incorrectly computed and that the values reported in the barcoding literature are misleading. The main problem is that the barcoding gap is generally quantified as the difference between intraspecific and mean interspecific, congeneric distances, whereas we will argue here that for species identification only the smallest interspecific distance should be used. Other authors have also pointed out that the use of smallest interspecific distances would be more appropriate (see Sperling, 2003; Moritz and Cicero, 2004; Vences et al., 2005a, 2005b; Cognato, 2006; Meier et al., 2006; Meyer and Paulay, 2005; Roe and Sperling, 2007), but currently we lack a comparative study that documents that the two measures yield different results. Here we provide evidence based on 43,137 COI sequences from 12,459 Metazoan species that barcoding gaps based on mean interspecific distances are artificially inflated and that only smallest interspecific distances correctly reflect that species identification gets more difficult as more species are sampled. Using DNA barcodes for species identification is analogous to identifying an unidentified specimen by comparing it to a reference collection of identified specimens. Initially one may compare an unidentified specimen to all identified material in the same genus, but ultimately the identification problem pares down to deciding whether a specimen belongs to one of a few, very similar, congeneric species. Determining an unidentified specimen to species is straightforward if the intraspecific variability is small—i.e., the unidentified specimen is a good match to a referenced species—and the differences between the best-matching species and the next best match is large—i.e., the specimen is a good match to only one of the referenced species. Analogously, the ease with which a query sequence can be identified to species is only dependent on how different it is from the most similar allospecific sequence, whereas its distinctness from a hypothetical “average” congeneric species does not matter (see Sperling, 2003; Moritz and Cicero, 2004; Vences et al., 2005a, 2005b; Cognato, 2006; Meier et al., 2006; Meyer and Paulay, 2005; Roe and Sperling, 2007). Yet, DNA barcoding publications and BOLD continue to report the mean instead of the smallest interspecific distances for congeneric species.

Molecular phylogeny of Harpactorini (Insecta: Reduviidae): correlation of novel predation strategy with accelerated evolution of predatory leg morphology

Much research and discussion have focused on the effects of key innovations on lineage diversification, whereas little has been done to investigate their role in morphological evolution using phylogenetic approaches. Here we present the first comprehensive molecular phylogeny of the Harpactorini (Insecta: Reduviidae), the largest assassin bug tribe, sampling 229 terminal taxa and using five gene segments (28S D2, D3–D5, 16S, COI, and Deformed). Employing comparative phylogenetic methods, we demonstrate the correlation of a putative key innovation, the sticky trap predation strategy, with accelerated rates of morphological evolution of the predatory fore leg in assassin bugs. We show that bugs exhibiting sticky trap predation have evolved more slender and longer fore femora than non‐sticky bugs. Using phylogenetically independent contrast analyses, we document correlated evolution between femoral thickness and length. We argue that the novel sticky trap predation strategy may allow sticky bugs to alleviate functional constraints on the fore femur and thus to attain a higher rate of evolution than other Harpactorini or Reduviidae. We discuss the possibility that sticky bugs represent a case of adaptive radiation. We also test historical supra‐generic groups within the Harpactorini, and show that most of them are not monophyletic. We confirm the paraphyly of Harpactorini with respect to Rhaphidosomini.

Model selection in statistical historical biogeography of Neotropical insects—The Exophthalmus genus complex (Curculionidae: Entiminae)

Statistical historical biogeographic methods rely on models that represent various biogeographic processes. Until recently model selection in this domain was not widely used, and the impact of differential model selection on inferring biogeographic scenarios was not well understood. Focusing on Neotropical weevils in the Exophthalmus genus complex (EGC) (Insecta: Curculionidae: Entiminae), we compare three commonly used biogeographic models - DIVA (Dispersal-Vicariance Analysis), DEC (Dispersal-Extinction-Cladogenesis) and BayArea (Bayesian Analysis of Biogeography), and examine the impact of modeling founder-event jump dispersal on historical biogeographic reconstructions. We also investigate the biogeographic events that have shaped patterns of distribution, diversification, and endemism in this weevil lineage. We sample representatives of 65 species of the EGC and 26 outgroup terminals from the Neotropics, including Caribbean islands and the mainland. We reconstruct a molecular phylogeny based on six genes and apply molecular dating using a relaxed clock with three fossil calibration points. Historical biogeographic estimations and alternative biogeographic models are computed and compared with the R package BioGeoBEARS. Model selection strongly favors biogeographic models that include founder-event jump dispersal. Without modeling jump dispersal, estimations based on the three biogeographic models are dramatically different, especially for early-diverging nodes. When jump dispersal is included, the three biogeographic models perform similarly. Accordingly, we show that the Neotropical mainland was colonized by Caribbean species in the early Miocene, and that in situ diversification accounts for a majority (∼75%) of the biogeographic events in the EGC. Our study highlights the need to assess wide-ranging historical biogeographic processes - including founder-event jump dispersal - for best-fitting statistical Caribbean biogeographic reconstructions. Moreover, colonization of the Neotropical mainland from the Caribbean reinforces the notion that islands can be an important source of continental diversity.

Matching dimorphic sexes and immature stages with adults: resolving the systematics of the Bekilya group of Malagasy assassin bugs (Hemiptera: Reduviidae: Peiratinae)

The Madagascar endemic assassin bugs Bekilya Villiers and Hovacoris Villiers were described from macropterous male specimens with striking colour patterns, and are currently monotypic. Mutillocoris Villiers, with two species from Madagascar, was based on brachypterous female specimens that resemble female mutillid wasps. To investigate the validity of the three genera, recently collected specimens from Madagascar were studied with both morphological and molecular techniques. Morphology alone appeared to be of limited value for associating males with females, and immature stages with adults, because of drastic differences between the sexes and the life stages. However, the use of morphology in conjunction with molecular data resolved these associations and showed that species of Mutillocoris represent females of Bekilya and Hovacoris, which we transfer accordingly to these two genera and refer to them as the Bekilya group. The type species of Mutillocoris belongs in Bekilya, resulting in the synonymy of these two genera (Mutillocorissyn.n.). The Bekilya group is diagnosed and several new species are described: Bekilya mahafalyasp.n., Bekilya tenebrasp.n., Bekilya tulearasp.n., Hovacoris bicolornotumsp.n., Hovacoris melanocepssp.n. and Hovacoris rufiventrissp.n. A total of ten species are recognized within the Bekilya group. The monophyly of Bekilya, Hovacoris and the Bekilya group is confirmed by morphological and molecular phylogenetic analyses.

Infection Rates of Triatoma protracta (Uhler) with Trypanosoma cruzi in Southern California and Molecular Identification of Trypanosomes

We report Trypanosoma cruzi infection rates of the native kissing bug Triatoma protracta in southern California. The rates are within the historically reported range, but differ significantly between the two sites (19% in Escondido and 36% in Glendora). Identification of T. cruzi in T. protracta was conducted for the first time by using partial 18S ribosomal RNA and 24Sα ribosomal RNA sequences. Incongruence of 24Sα ribosomal RNA phylogeny with current T. cruzi genotype classification supports non-clonality of some T. cruzi genotypes.

Emerging semantics to link phenotype and environment

Understanding the interplay between environmental conditions and phenotypes is a fundamental goal of biology. Unfortunately, data that include observations on phenotype and environment are highly heterogeneous and thus difficult to find and integrate. One approach that is likely to improve the status quo involves the use of ontologies to standardize and link data about phenotypes and environments. Specifying and linking data through ontologies will allow researchers to increase the scope and flexibility of large-scale analyses aided by modern computing methods. Investments in this area would advance diverse fields such as ecology, phylogenetics, and conservation biology. While several biological ontologies are well-developed, using them to link phenotypes and environments is rare because of gaps in ontological coverage and limits to interoperability among ontologies and disciplines. In this manuscript, we present (1) use cases from diverse disciplines to illustrate questions that could be answered more efficiently using a robust linkage between phenotypes and environments, (2) two proof-of-concept analyses that show the value of linking phenotypes to environments in fishes and amphibians, and (3) two proposed example data models for linking phenotypes and environments using the extensible observation ontology (OBOE) and the Biological Collections Ontology (BCO); these provide a starting point for the development of a data model linking phenotypes and environments.

Molecular phylogeny of Harpactorinae and Bactrodinae uncovers complex evolution of sticky trap predation in assassin bugs (Heteroptera: Reduviidae)

Sticky trap predation, the use of adhesive substances to trap and capture prey, is an intriguing yet poorly studied predatory strategy. Unique among known sticky trap predators, assassin bugs (Reduviidae) have evolved both exogenous and endogenous sticky trap predatory mechanisms: some trap their prey with sticky plant resins, some scavenge insects entrapped by sticky plant trichomes and others self‐produce sticky secretions. The evolution of these different strategies in assassin bugs is poorly understood due to the lack of comprehensive phylogenies. We reconstruct a phylogeny of Reduviidae (141 taxa; > 5000 bp) focusing on the Harpactorinae and Bactrodinae that engage in sticky trap predation. Ancestral state reconstruction, and temporal and geographical divergence analyses show that sticky trap predation techniques in assassin bugs evolved at least seven times independently since the late Cretaceous: use of sticky plant trichomes evolved as many as four times, resin‐use twice independently and once as a transition from trichome use, and ‘self‐stickiness’ once. Exogenous and endogenous sticky traps first appeared in the Neotropics, with the two exogenous mechanisms (resin and trichome use) subsequently evolving independently in the Old World. This study illustrates, for the first time, the complex evolutionary pattern of sticky trap predation within assassin bugs.

Specimens versus sequences.

In his Perspective “GenBank—natural history in the 21st century?” (24 October 2008, [p. 537][1]), B. J. Strasser claims that GenBank follows the tradition of natural history studies. I argue that GenBank is inconsistent with some important aspects of the tradition of natural history and it

ZELUS RENARDII AND Z. TETRACANTHUS (HEMIPTERA: REDUVIIDAE): BIOLOGICAL ATTRIBUTES AND THE POTENTIAL FOR DISPERSAL IN TWO ASSASSIN BUG SPECIES

ABSTRACT The leafhopper assassin bug, Zelus renardii Kolenati, is a natural enemy and stands out among species in the large New World genus Zelus Fabricius (∼60 spp.) by its introduction to and establishment in 3 biogeographic regions. We here present documentation of the distribution and habitat of Z. renardii in its native range in North and Central America and compare it with Z. tetracanthus Stål, a wide-ranging New World congener that apparently has not dispersed outside of its native range. In addition, we document and compare predatory and reproductive behaviors in the 2 species. Zelus renardii is widely distributed in the Western USA and shows a continuous geographic range south to Guatemala; Z. tetracanthus is broadly distributed across North and Central America and also occurs in Brazil. In Riverside County, California, Z. renardii is common in suburban and disturbed habitats in addition to certain natural areas, whereas Z. tetracanthus is usually restricted to natural areas. The behavioral comparison under laboratory conditions indicated that Z. renardii caught prey faster and that feeding duration in this species was shorter than in Z. tetracanthus. The duration of pre-copulatory behaviors in Z. renardii was shorter than in Z. tetracanthus, resulting in a shorter overall mating duration. Based on the higher percentage of egg batches that produced first instars in Z. renardii, this species may establish large populations under adverse conditions faster than Z. tetracanthus. Our observations on distribution and biology contribute toward an understanding of the differences in invasiveness between the 2 species.

Endosymbiont diversity and evolution across the weevil tree of life

As early as the time of Paul Buchner, a pioneer of endosymbionts research, it was shown that weevils host diverse bacterial endosymbionts, probably only second to the hemipteran insects. To date, there is no taxonomically broad survey of endosymbionts in weevils, which preclude any systematic understanding of the diversity and evolution of endosymbionts in this large group of insects, which comprise nearly 7% of described diversity of all insects. We gathered the largest known taxonomic sample of weevils representing four families and 17 subfamilies to perform a study of weevil endosymbionts. We found that the diversity of endosymbionts is exceedingly high, with as many as 44 distinct kinds of endosymbionts detected. We recovered an ancient origin of association of Nardonella with weevils, dating back to 124 MYA. We found repeated losses of this endosymbionts, but also cophylogeny with weevils. We also investigated patterns of coexistence and coexclusion.