Sebastian Kvist | University of Toronto (original) (raw)

Papers by Sebastian Kvist

Zoologica Scripta, 2018

At this meeting, the community reached consensus to revise nemertean taxonomy at the class level,... more At this meeting, the community reached consensus to revise nemertean taxonomy at the class level, based on the compiled evidence from studies on nemertean systematics published in the last 15 years (Andrade et al., 2014, 2012 ; Thollesson & Norenburg, 2003). Previous classifications (e.g., Stiasny-Wijnhoff, 1936) are not based on phylogenetic grounds, and the use of these names is therefore nowadays not wholly informative. With the purpose of facilitating the practical use of the nemertean taxonomy and also making nemertean taxonomy reflect a wealth of more recent information, we conclude that the ranks Anopla and Enopla should be eliminated with the following argumentation: "Enopla" has for long held no more information than the name "Hoplonemertea". "Anopla" is paraphyletic and the name usually corresponds to the following traits: (a) not bearing stylet; and (b) mouth and proboscis having separate openings. This information is equally well presented in a system that also bears information on natural groups. The names "Anopla" and "Enopla" are hence dismissed and the following system of the Nemertea phylum is implemented:

Hydrobiologia, 2022

The bloodfeeding leech genus Placobdella is dominated by North American diversity, with only a si... more The bloodfeeding leech genus Placobdella is dominated by North American diversity, with only a single nominal species known from Central America and one from the Palearctic region. This is likely due to considerable underestimation of Palearctic biodiversity, but investigations into potential hidden diversity are lacking. To shed light on this, the present study introduces new data for specimens initially identified as Placobdella costata from Ukraine (close to the type locality),

Leeches play important roles in food webs due to their abundance, diversity and feeding habits. S... more Leeches play important roles in food webs due to their abundance, diversity and feeding habits. Studies using invertebrate-derived DNA (iDNA) extracted from leech gut contents to target vertebrate DNA have focused on the Indo-Pacific region and mainly leveraged the leech family Haemadipsidae, composed of haematophagous terrestrial leeches, while the aquatic haematophagous, liquidosomatophagous and macrophagous counterparts have largely been disregarded. While there is general knowledge regarding the taxonomic groups that leeches prefer to feed on, detailed taxonomic resolution is still missing and therefore, their potential use for monitoring animals is not known. In this study, 116 non-haemadipsid leeches belonging to 12 species and spanning the three feeding habits were collected in Mexico and Canada. We used DNA metabarcoding to investigate their diet and assess their potential use for vertebrate monitoring. We detected vertebrate taxa from five orders including fish, turtles and...

BioInvasions Records, 2018

We report two new, curious records of freshwater worms collected in the Panama Canal, Panama: the... more We report two new, curious records of freshwater worms collected in the Panama Canal, Panama: the ribbon worm, Apatronemertes albimaculosa Wilfert and Gibson, 1974, and the leech, Placobdella parasitica (Say, 1824). Identification of these specimens relied on morphological examinations and, more heavily, on molecular barcoding using the mitochondrial COI locus. Both phylogenetic and genetic distance analyses were employed to robustly establish the identities of the specimens. Whereas the native range of A. albimaculosa is currently unknown, P. parasitica was first described from Minnesota, USA and is possibly native to southern Canada and most of the USA. Neither species has previously been recorded from Central America. We briefly discuss possible scenarios for the dispersal of both species, and conclude that the introduction of P. parasitica into Panamanian waters was likely human-mediated in association with the introduction of its turtle host. Insofar as ours is the first record of wild-caught A. albimaculosa worldwide, we do not exclude the possibility of Panama being within the native range of the species. However, it is also possible that the species has been transported with ballast water into the Panama Canal; a relatively common phenomenon.

The Journal of parasitology, Jan 16, 2018

Leeches (Annelida: Hirudinea) possess powerful salivary anticoagulants and, accordingly, are freq... more Leeches (Annelida: Hirudinea) possess powerful salivary anticoagulants and, accordingly, are frequently employed in modern, authoritative medicine. Members of the almost exclusively marine family Piscicolidae account for 20% of leech species diversity, and feed on host groups (e.g., sharks) not encountered by their freshwater and terrestrial counterparts. Moreover, some species of Ozobranchidae feed on endangered marine turtles and have been implicated as potential vectors for the tumor-associated turtle herpesvirus. In spite of their ecological importance and unique host associations, there is a distinct paucity of data regarding the salivary transcriptomes of either of these families. Using next generation sequencing, we profiled transcribed, putative anticoagulants and other salivary bioactive compounds that have previously been linked to bloodfeeding from 7 piscicolid species (3 elasmobranch-feeders; 4 non-cartilaginous fish-feeders) and 1 ozobranchid species (2 samples). In tot...

Memoirs of Museum Victoria, 2014

Magesh, M., Kvist S. and Glasby, C.J. 2014. Incipient speciation within the Namalycastis abiuma (... more Magesh, M., Kvist S. and Glasby, C.J. 2014. Incipient speciation within the Namalycastis abiuma (Annelida: Nereididae) species group from southern India revealed by combined morphological and molecular data. Memoirs of Museum Victoria 71: 169-176. Namalycastis abiuma (Grube, 1872), originally described from Brazil, comprises a species complex of morphologically similar forms occurring circumtropically, including India. Apart from the Namalycastis abiuma species group, four other Namalycastis species are presently known from India: N.

PLOS ONE, 2016

Whereas many nemerteans (ribbon worms; phylum Nemertea) can be identified from external character... more Whereas many nemerteans (ribbon worms; phylum Nemertea) can be identified from external characters if observed alive, many are still problematic. When it comes to preserved specimens (as in e.g. marine inventories), there is a particular need for specimen identifier alternatives. Here, we evaluate the utility of COI (cytochrome c oxidase subunit I) as a single-locus barcoding gene. We sequenced, data mined, and compared gene fragments of COI for 915 individuals representing 161 unique taxonomic labels for 71 genera, and subjected different constellations of these to both distance-based and character-based DNA barcoding approaches, as well as species delimitation analyses. We searched for the presence or absence of a barcoding gap at different taxonomic levels (phylum, subclass, family and genus) in an attempt to understand at what level a putative barcoding gap presents itself. This was performed both using the taxonomic labels as species predictors and using objectively inferred species boundaries recovered from our species delimitation analyses. Our data suggest that COI works as a species identifier for most groups within the phylum, but also that COI data are obscured by misidentifications in sequence databases. Further, our results suggest that the number of predicted species within the dataset is (in some cases substantially) higher than the number of unique taxonomic labels-this highlights the presence of several cryptic lineages within well-established taxa and underscores the urgency of an updated taxonomic backbone for the phylum.

The Journal of parasitology, Jan 4, 2015

One of the recalcitrant questions regarding the evolutionary history of clitellate annelids invol... more One of the recalcitrant questions regarding the evolutionary history of clitellate annelids involves the feeding preference of the common ancestor of extant rhynchobdellid (proboscis bearing) and arhynchobdellid (jaw bearing) leeches. Whereas early evidence, based on morphological data, pointed towards independent acquisitions of blood feeding in the 2 orders, molecular-based phylogenetic data suggest that the ancestor of modern leeches was a sanguivore. Here, we use a comparative transcriptomic approach in order to increase our understanding of the diversity of anticoagulation factors for 3 species of the genus Placobdella, for which comparative data have been lacking, and inspect these in light of archetypal anticoagulant data for both arhynchobdellid and other rhynchobdellid species. Notwithstanding the varying levels of host-specificity displayed by the 3 different species of Placobdella, transcriptomic profiles with respect to anticoagulation factors were largely similar - this...

Proceedings of the Biological Society of Washington, 2014

Namalycastis glasbyi Fernando & Rajasekaran, 2007 was originally described from the creek-lined c... more Namalycastis glasbyi Fernando & Rajasekaran, 2007 was originally described from the creek-lined coast north of Mumbai in western India. However, the original description includes some oddities that make species recognition problematic and may confuse future taxonomic evaluations of the genus. Moreover, the type material of the species has been lost and, therefore, this species is herein redescribed, a neotype is assigned, and the distinguishing morphological features are clearly illustrated. The species shares some features with two other Indian species: Namalycastis indica Southern, 1921 and the N. abiuma Müller in Grube, 1871 species group. However, N. glasbyi differs from N. indica in having comparatively smaller antennae, shorter posterior dorsal cirri, a higher count of notochaetae (maximum of three) in the mid and anterior body, and finely serrated blades in sub-and supra-neuroacicular falcigers in the anterior and mid-body. Further, sub-neuroacicular spinigers have finely serrated blades in all parapodia in N. glasbyi, as opposed to the coarsely serrated spinigers from antero-mid body parapodia of N. indica. Moreover, N. glasbyi differs from the N. abiuma species group in lacking coarsely serrated sub-neuroacicular spinigers in the parapodia of mid and posterior parts of the body and in possessing a tripartite pygidium. In addition to these morphological differences, the mitochondrial COI sequence (DNA barcode) of the neotype of Namalycastis glasbyi clearly distinguishes it from other Indian species.

Ecology and Evolution, 2013

Although medicinal leeches have long been used as treatment for various ailments because of their... more Although medicinal leeches have long been used as treatment for various ailments because of their potent anticoagulation factors, neither the full diversity of salivary components that inhibit coagulation, nor the evolutionary selection acting on them has been thoroughly investigated. Here, we constructed expressed sequence tag libraries from salivary glands of two species of medicinal hirudinoid leeches, Hirudo verbana and Aliolimnatis fenestrata, and identified anticoagulant-orthologs through BLASTx searches. The data set then was augmented by the addition of a previously constructed EST library from the macrobdelloid leech Macrobdella decora. The identified orthologs then were compared and contrasted with well-characterized anticoagulants from a variety of leeches with different feeding habits, including non-sanguivorous species. Moreover, four different statistical methods for predicting signatures of positive and negative evolutionary pressures were used for 10 rounds each to assess the level and type of selection acting on the molecules as a whole and on specific sites. In total, sequences showing putative BLASTx-orthology with five and three anticoagulant-families were recovered in the A. fenestrata and H. verbana EST libraries respectively. Selection pressure analyses predicted high levels of purifying selection across the anticoagulant diversity, although a few isolated sites showed signatures of positive selection. This study represents a first attempt at mapping the anticoagulant repertoires in a comparative fashion across several leech families.

Journal of the Marine Biological Association of the United Kingdom, 2015

The heteronemertean species Micrura dellechiajei is thus far only known from its type locality in... more The heteronemertean species Micrura dellechiajei is thus far only known from its type locality in the Gulf of Naples (Italy) and has not been recorded in 120 years. During two oceanographic surveys conducted in Spanish Mediterranean waters, several nemertean specimens were collected, and thorough morphological examination indicated that some of these pertained to the species M. dellechiajei, suggesting that populations may be more widespread than previously thought. Because of the rarity of this species coupled with the fact that its last morphological narrative was given 120 years ago, we here provide a redescription of the species based on the new specimens, complete with illustrations and new data concerning its morphology, and we also place some of the collected specimens in a molecular phylogenetic framework.

ZooKeys, 2012

Namalycastis jaya sp. n. (Polychaeta: Nereididae: Namanereidinae) is described from the southern ... more Namalycastis jaya sp. n. (Polychaeta: Nereididae: Namanereidinae) is described from the southern coast of Kerala in southwest India. One important characteristic feature of the species is the lack of notochaetae in all parapodia, a characteristic that it shares with at least two other species, Namalycastis elobeyensis Glasby, 1999 and Namalycastis hawaiiensis Johnson, 1903. It differs from N. elobeyensis by virtue of its smaller antennae, unequal eye size, bilobed acicular neuropodial ligule and multi-incised pygidium rim. Moreover, it differs from N. hawaiiensis by having fewer teeth on the serrated blades of the sub-neuroacicular falciger in chaetiger 10, and by possessing finely serrated falcigers in posterior segments. Beyond morphological analyses, molecular phylogenetics was used for the first time for Namalycastis to support population monophyly and recognition of the new species. The analysis, using both mitochondrial and nuclear data, corroborated the morphological analysis in suggesting that our specimens represent an as yet undescribed species, Namalycastis jaya sp. n., which forms a monophyletic group among the sampled nereidid taxa. Finally, a taxonomic key for Namalycastis species recorded from the Indian region is provided.

PLoS ONE, 2014

In spite of the high relevance of lumbricid earthworms ('Oligochaeta': Lumbricidae) for soil stru... more In spite of the high relevance of lumbricid earthworms ('Oligochaeta': Lumbricidae) for soil structure and functioning, the taxonomy of this group of terrestrial invertebrates remains in a quasi-chaotic state. Earthworm taxonomy traditionally relies on the interpretation of external and internal morphological characters, but the acquisition of these data is often hampered by tedious dissections or restricted access to valuable and rare museum specimens. The present state of affairs, in conjunction with the difficulty of establishing primary homologies for multiple morphological features, has led to an almost unrivaled instability in the taxonomy and systematics of certain earthworm groups, including Lumbricidae. As a potential remedy, we apply for the first time a non-destructive imaging technique to lumbricids and explore the future application of this approach to earthworm taxonomy. High-resolution micro-computed tomography (mCT) scanning of freshly fixed and museum specimens was carried out using two cosmopolitan species, Aporrectodea caliginosa and A. trapezoides. By combining two-dimensional and three-dimensional dataset visualization techniques, we demonstrate that the morphological features commonly used in earthworm taxonomy can now be analyzed without the need for dissection, whether freshly fixed or museum specimens collected more than 60 years ago are studied. Our analyses show that mCT in combination with soft tissue staining can be successfully applied to lumbricid earthworms. An extension of the approach to other families is poised to strengthen earthworm taxonomy by providing a versatile tool to resolve the taxonomic chaos currently present in this ecologically important, but taxonomically neglected group of terrestrial invertebrates.

Journal of Parasitology, 2012

Invertebrate Biology, 2013

Freshwater, marine, and terrestrial leeches that depend on a diet of fresh blood have evolved sal... more Freshwater, marine, and terrestrial leeches that depend on a diet of fresh blood have evolved salivary peptide components that inhibit normal thrombus formation by prey. Although bloodfeeding in leeches has long been of interest to biologists and medical practitioners alike, only a few studies have comprehensively examined the anticoagulant repertoires of hematophagous leeches, and these have largely been confined to representatives of Glossiphoniidae, Hirudinidae, and Macrobdellidae. Here, we present 454 pyrosequencing data from the salivary transcriptome of the hematophagous terrestrial leech Haemadipsa interrupta (Haemadipsidae). Assembled transcripts were annotated using both similarity scores (BLAST) and gene ontology (Blast2GO). Subsequently, transcripts were examined within alignments containing well-characterized anticoagulants and other select bioactive (i.e., affecting the living cells of the prey) salivary proteins and phylogenies were reconstructed for each protein data set to verify orthology predictions. In total, transcripts significantly matching 20 salivary proteins of interest were found in the transcriptome, representing several different antagonistic pathways. After reviewing gene ontologies, alignments, and phylogenetic trees, sequences for 15 out of the 20 hits were deemed correctly annotated. Additionally, we recovered matches against several proteins that have previously been linked to anticoagulation (e.g., cathepsin and disintegrins), but the specific function of which in leeches needs further investigation. Finally, in light of these data as well as those previously published, we discuss our current understanding of the distribution and evolution of anticoagulants in leeches.

Cladistics, 2013

We present phylogenomic analyses of the most comprehensive molecular character set compiled for A... more We present phylogenomic analyses of the most comprehensive molecular character set compiled for Annelida and its constituent taxa, including over 347 000 aligned nucleotide sites for 39 taxa. The nucleotide data set was recovered using a pre-existing amino acid data set of almost 48 000 aligned sites as a backbone for tBLASTn searches against NCBI. In addition, orthology determinations of the loci in the original amino acid data set were scrutinized using an All vs All Reciprocal Best Hit approach, employing BLASTp, and examining for statistical interdependency among the loci. This approach revealed considerable sequence redundancy among the loci in the original data set and a new data set was compiled, with the redundancy removed. The newly compiled nucleotide data set, the original amino acid data set, and the new reduced amino acid data set were subjected to parsimony analyses and two forms of bootstrap resampling. The last-named data set also was analysed using a maximum-likelihood approach. There were two main objectives to these analyses: (i) to examine the general topology, including support, resulting from the analyses of the new data sets and (ii) to assess the consistency of the branching patterns across optimality criteria by comparison with previous probabilistic approaches. The phylogenetic hypotheses resulting from analyses of the three data sets are largely unsupported, reflecting the continued difficulty of finding numerous, reliable, and suitable loci for a group as ancient as Annelida. Resulting parsimonious hypotheses disagree, in some respects, with the previous probabilistic approaches; Sedentaria and, in most cases, Errantia are not supported as monophyletic groups but Pleistoannelida is recovered as a (unsupported) monophyletic group in one of the three parsimony analyses as well as the likelihood analysis. In addition, we performed missing data titration studies to estimate the impact of missing data on overall support and support for specific clades.

Mitochondrial DNA, 2010

Species identifications based on DNA barcoding rely on the correct identity of previously barcode... more Species identifications based on DNA barcoding rely on the correct identity of previously barcoded specimens, but little attention has been given to whether deposited barcodes include correspondence to the species' name-bearing type. The information content associated with COX1 sequences in the two most commonly used repositories of barcodes, GenBank and the Barcode of Life Data System (BOLD), is often insufficient for subsequent evaluation of the robustness of the identification procedure. We argue that DNA barcoding and taxonomy alike will benefit from more information content in the annotations of barcoded specimens as this will allow for validation and re-evaluation of the initial specimen identification. The aim should be to closely connect specimens from which reference barcodes are generated with the holotype through straightforward taxonomy, and geographical and genetic correlations. Annotated information should also include voucher specimens and collector/identifier information. We examine two case studies based on empirical data, in which barcoding and taxonomy benefit from increased information content. On the basis of data from the first case study, we designate a barcoded neotype of the European medicinal leech, Hirudo medicinalis, on morphological and geographical grounds.

The description of Helobdella stagnalis (Linnaeus, 1758) has emphasized the presence of a nuchal,... more The description of Helobdella stagnalis (Linnaeus, 1758) has emphasized the presence of a nuchal, chitinous scute located on the dorsal surface in the first third of the body as the diagnostic character for the species. Historically, identifications of species of Helobdella have relied heavily on this character and, as a result, Helobdella stagnalis has been reported from an inordinately broad geographic range, including Europe, Asia, Africa, North America, and South America. In addition to a few earlier investigations, a recent analysis showed that great genetic distances (orders of magnitude greater than previous estimations of intraspecific divergence in leeches) are present between scute-bearing specimens identified as H. stagnalis from Europe and North America, implying that H. stagnalis does not occur in North America. The present study expands the geographic boundaries of taxon sampling for both European and North American taxa, and reexamines the phylogenetic relationships and cytochrome c oxidase subunit I (COI) variation within scute-bearing species of the genus Helobdella. Our analyses include specimens putatively identified as "Helobdella stagnalis" from Sweden, and Iran, as well as numerous localities covering Canada and the USA. Our results corroborate previous studies in that European and west Asian specimens form a clade, including the neotype, which is separate from North American taxa. To alleviate future taxonomic confusion, we redescribe H. stagnalis and designate a neotype from the inferred type locality. The designation of a neotype stabilizes the taxonomy of scute-bearing leeches of the genus Helobdella and enables us to definitively correct erroneous identifications reported in previous studies. We also note that at least four lineages of scute-bearing, North American species of Helobdella lack formal descriptions.

Leeches of the family Erpobdellidae are important members of benthic freshwater environments, whe... more Leeches of the family Erpobdellidae are important members of benthic freshwater environments, where they are voracious predators of other invertebrates and an important source of nutrition for several species of vertebrates. Beset by a lack of reliable diagnostic morphological characters and destructive identification processes, molecular approaches have, in recent years, been employed to illuminate the relationships within this family, and DNA barcoding has been employed for identification purposes. However, an understanding of the levels of genetic variation across the geographic distributions of members of the genus is still lacking. Herein, we sequence the mitochondrial COI locus for 249 newly collected North American individuals, representing 5 species, as well as mitochondrial 12S rDNA, nuclear 18S rDNA, and nuclear 28S rDNA for a select subset of these. Our COI dataset was leveraged to detect potential cryptic species, and to calculate genetic distances as a proxy for the degree of gene flow between populations. Augmented by numerous sequences from GenBank, the multilocus dataset was used to reconstruct a phylogenetic hypothesis for worldwide members of the genus. Beyond corroborating previous overarching phylogenetic frameworks, our results show that an undescribed species that is morphologically and genetically similar to Erpobdella punctata exists in sympatry with this species-the new species has likely been overlooked in previous studies due to its morphological similarity with Erpobdella punctata. Erpobdella bucera is reported from Canada for the first time; and Erpobdella microstoma is newly reported from Saskatchewan and placed in a phylogeny for the first time. Finally, we find evidence for genetic structure in both E. cf. punctata and Erpobdella obscura that is correlated with major river drainage basin boundaries in North America.

Blood-feeding leeches of the genus Placobdella have acquired intracellular alphaproteobacterial e... more Blood-feeding leeches of the genus Placobdella have acquired intracellular alphaproteobacterial endosymbionts of the genus Reichenowia that potentially aid in the production of B vitamins, thereby ameliorating the lack of these essential nutrients in the diet of the hosts. For Placobdella associates, little is still known about the diversity, genetic makeup, and the mode of transmission of bacteria between leeches. We aimed to (i) place the bacterial symbionts in a phylogenetic context, (ii) compare patterns of cladogenesis between the bacteria and hosts to search for evidence of co-speciation, and (iii) explore the mode of bacterial transmission between leech parent and offspring. DNA sequencing of the bacterial 16S rDNA and 23S rDNA loci suggests that, whereas Reichenowia forms a monophyletic group within the alphaproteobacterial family Rhizobiaceae, no evidence for co-speciation between hosts and bacteria can be traced. Attempts at DNA amplification for ovarial tissues were negative for a range of species, but two 16S rDNA sequences retrieved from the testisacs of P. rugosa showed very high similarity with Reichenowia. Although we cannot rule out that this may be a contamination, or a different, potentially free-living species of bacteria, our results may indicate that Reichenowia is transferred from leech parent to offspring via the testisacs.

Zoologica Scripta, 2018

At this meeting, the community reached consensus to revise nemertean taxonomy at the class level,... more At this meeting, the community reached consensus to revise nemertean taxonomy at the class level, based on the compiled evidence from studies on nemertean systematics published in the last 15 years (Andrade et al., 2014, 2012 ; Thollesson & Norenburg, 2003). Previous classifications (e.g., Stiasny-Wijnhoff, 1936) are not based on phylogenetic grounds, and the use of these names is therefore nowadays not wholly informative. With the purpose of facilitating the practical use of the nemertean taxonomy and also making nemertean taxonomy reflect a wealth of more recent information, we conclude that the ranks Anopla and Enopla should be eliminated with the following argumentation: "Enopla" has for long held no more information than the name "Hoplonemertea". "Anopla" is paraphyletic and the name usually corresponds to the following traits: (a) not bearing stylet; and (b) mouth and proboscis having separate openings. This information is equally well presented in a system that also bears information on natural groups. The names "Anopla" and "Enopla" are hence dismissed and the following system of the Nemertea phylum is implemented:

Hydrobiologia, 2022

The bloodfeeding leech genus Placobdella is dominated by North American diversity, with only a si... more The bloodfeeding leech genus Placobdella is dominated by North American diversity, with only a single nominal species known from Central America and one from the Palearctic region. This is likely due to considerable underestimation of Palearctic biodiversity, but investigations into potential hidden diversity are lacking. To shed light on this, the present study introduces new data for specimens initially identified as Placobdella costata from Ukraine (close to the type locality),

Leeches play important roles in food webs due to their abundance, diversity and feeding habits. S... more Leeches play important roles in food webs due to their abundance, diversity and feeding habits. Studies using invertebrate-derived DNA (iDNA) extracted from leech gut contents to target vertebrate DNA have focused on the Indo-Pacific region and mainly leveraged the leech family Haemadipsidae, composed of haematophagous terrestrial leeches, while the aquatic haematophagous, liquidosomatophagous and macrophagous counterparts have largely been disregarded. While there is general knowledge regarding the taxonomic groups that leeches prefer to feed on, detailed taxonomic resolution is still missing and therefore, their potential use for monitoring animals is not known. In this study, 116 non-haemadipsid leeches belonging to 12 species and spanning the three feeding habits were collected in Mexico and Canada. We used DNA metabarcoding to investigate their diet and assess their potential use for vertebrate monitoring. We detected vertebrate taxa from five orders including fish, turtles and...

BioInvasions Records, 2018

We report two new, curious records of freshwater worms collected in the Panama Canal, Panama: the... more We report two new, curious records of freshwater worms collected in the Panama Canal, Panama: the ribbon worm, Apatronemertes albimaculosa Wilfert and Gibson, 1974, and the leech, Placobdella parasitica (Say, 1824). Identification of these specimens relied on morphological examinations and, more heavily, on molecular barcoding using the mitochondrial COI locus. Both phylogenetic and genetic distance analyses were employed to robustly establish the identities of the specimens. Whereas the native range of A. albimaculosa is currently unknown, P. parasitica was first described from Minnesota, USA and is possibly native to southern Canada and most of the USA. Neither species has previously been recorded from Central America. We briefly discuss possible scenarios for the dispersal of both species, and conclude that the introduction of P. parasitica into Panamanian waters was likely human-mediated in association with the introduction of its turtle host. Insofar as ours is the first record of wild-caught A. albimaculosa worldwide, we do not exclude the possibility of Panama being within the native range of the species. However, it is also possible that the species has been transported with ballast water into the Panama Canal; a relatively common phenomenon.

The Journal of parasitology, Jan 16, 2018

Leeches (Annelida: Hirudinea) possess powerful salivary anticoagulants and, accordingly, are freq... more Leeches (Annelida: Hirudinea) possess powerful salivary anticoagulants and, accordingly, are frequently employed in modern, authoritative medicine. Members of the almost exclusively marine family Piscicolidae account for 20% of leech species diversity, and feed on host groups (e.g., sharks) not encountered by their freshwater and terrestrial counterparts. Moreover, some species of Ozobranchidae feed on endangered marine turtles and have been implicated as potential vectors for the tumor-associated turtle herpesvirus. In spite of their ecological importance and unique host associations, there is a distinct paucity of data regarding the salivary transcriptomes of either of these families. Using next generation sequencing, we profiled transcribed, putative anticoagulants and other salivary bioactive compounds that have previously been linked to bloodfeeding from 7 piscicolid species (3 elasmobranch-feeders; 4 non-cartilaginous fish-feeders) and 1 ozobranchid species (2 samples). In tot...

Memoirs of Museum Victoria, 2014

Magesh, M., Kvist S. and Glasby, C.J. 2014. Incipient speciation within the Namalycastis abiuma (... more Magesh, M., Kvist S. and Glasby, C.J. 2014. Incipient speciation within the Namalycastis abiuma (Annelida: Nereididae) species group from southern India revealed by combined morphological and molecular data. Memoirs of Museum Victoria 71: 169-176. Namalycastis abiuma (Grube, 1872), originally described from Brazil, comprises a species complex of morphologically similar forms occurring circumtropically, including India. Apart from the Namalycastis abiuma species group, four other Namalycastis species are presently known from India: N.

PLOS ONE, 2016

Whereas many nemerteans (ribbon worms; phylum Nemertea) can be identified from external character... more Whereas many nemerteans (ribbon worms; phylum Nemertea) can be identified from external characters if observed alive, many are still problematic. When it comes to preserved specimens (as in e.g. marine inventories), there is a particular need for specimen identifier alternatives. Here, we evaluate the utility of COI (cytochrome c oxidase subunit I) as a single-locus barcoding gene. We sequenced, data mined, and compared gene fragments of COI for 915 individuals representing 161 unique taxonomic labels for 71 genera, and subjected different constellations of these to both distance-based and character-based DNA barcoding approaches, as well as species delimitation analyses. We searched for the presence or absence of a barcoding gap at different taxonomic levels (phylum, subclass, family and genus) in an attempt to understand at what level a putative barcoding gap presents itself. This was performed both using the taxonomic labels as species predictors and using objectively inferred species boundaries recovered from our species delimitation analyses. Our data suggest that COI works as a species identifier for most groups within the phylum, but also that COI data are obscured by misidentifications in sequence databases. Further, our results suggest that the number of predicted species within the dataset is (in some cases substantially) higher than the number of unique taxonomic labels-this highlights the presence of several cryptic lineages within well-established taxa and underscores the urgency of an updated taxonomic backbone for the phylum.

The Journal of parasitology, Jan 4, 2015

One of the recalcitrant questions regarding the evolutionary history of clitellate annelids invol... more One of the recalcitrant questions regarding the evolutionary history of clitellate annelids involves the feeding preference of the common ancestor of extant rhynchobdellid (proboscis bearing) and arhynchobdellid (jaw bearing) leeches. Whereas early evidence, based on morphological data, pointed towards independent acquisitions of blood feeding in the 2 orders, molecular-based phylogenetic data suggest that the ancestor of modern leeches was a sanguivore. Here, we use a comparative transcriptomic approach in order to increase our understanding of the diversity of anticoagulation factors for 3 species of the genus Placobdella, for which comparative data have been lacking, and inspect these in light of archetypal anticoagulant data for both arhynchobdellid and other rhynchobdellid species. Notwithstanding the varying levels of host-specificity displayed by the 3 different species of Placobdella, transcriptomic profiles with respect to anticoagulation factors were largely similar - this...

Proceedings of the Biological Society of Washington, 2014

Namalycastis glasbyi Fernando & Rajasekaran, 2007 was originally described from the creek-lined c... more Namalycastis glasbyi Fernando & Rajasekaran, 2007 was originally described from the creek-lined coast north of Mumbai in western India. However, the original description includes some oddities that make species recognition problematic and may confuse future taxonomic evaluations of the genus. Moreover, the type material of the species has been lost and, therefore, this species is herein redescribed, a neotype is assigned, and the distinguishing morphological features are clearly illustrated. The species shares some features with two other Indian species: Namalycastis indica Southern, 1921 and the N. abiuma Müller in Grube, 1871 species group. However, N. glasbyi differs from N. indica in having comparatively smaller antennae, shorter posterior dorsal cirri, a higher count of notochaetae (maximum of three) in the mid and anterior body, and finely serrated blades in sub-and supra-neuroacicular falcigers in the anterior and mid-body. Further, sub-neuroacicular spinigers have finely serrated blades in all parapodia in N. glasbyi, as opposed to the coarsely serrated spinigers from antero-mid body parapodia of N. indica. Moreover, N. glasbyi differs from the N. abiuma species group in lacking coarsely serrated sub-neuroacicular spinigers in the parapodia of mid and posterior parts of the body and in possessing a tripartite pygidium. In addition to these morphological differences, the mitochondrial COI sequence (DNA barcode) of the neotype of Namalycastis glasbyi clearly distinguishes it from other Indian species.

Ecology and Evolution, 2013

Although medicinal leeches have long been used as treatment for various ailments because of their... more Although medicinal leeches have long been used as treatment for various ailments because of their potent anticoagulation factors, neither the full diversity of salivary components that inhibit coagulation, nor the evolutionary selection acting on them has been thoroughly investigated. Here, we constructed expressed sequence tag libraries from salivary glands of two species of medicinal hirudinoid leeches, Hirudo verbana and Aliolimnatis fenestrata, and identified anticoagulant-orthologs through BLASTx searches. The data set then was augmented by the addition of a previously constructed EST library from the macrobdelloid leech Macrobdella decora. The identified orthologs then were compared and contrasted with well-characterized anticoagulants from a variety of leeches with different feeding habits, including non-sanguivorous species. Moreover, four different statistical methods for predicting signatures of positive and negative evolutionary pressures were used for 10 rounds each to assess the level and type of selection acting on the molecules as a whole and on specific sites. In total, sequences showing putative BLASTx-orthology with five and three anticoagulant-families were recovered in the A. fenestrata and H. verbana EST libraries respectively. Selection pressure analyses predicted high levels of purifying selection across the anticoagulant diversity, although a few isolated sites showed signatures of positive selection. This study represents a first attempt at mapping the anticoagulant repertoires in a comparative fashion across several leech families.

Journal of the Marine Biological Association of the United Kingdom, 2015

The heteronemertean species Micrura dellechiajei is thus far only known from its type locality in... more The heteronemertean species Micrura dellechiajei is thus far only known from its type locality in the Gulf of Naples (Italy) and has not been recorded in 120 years. During two oceanographic surveys conducted in Spanish Mediterranean waters, several nemertean specimens were collected, and thorough morphological examination indicated that some of these pertained to the species M. dellechiajei, suggesting that populations may be more widespread than previously thought. Because of the rarity of this species coupled with the fact that its last morphological narrative was given 120 years ago, we here provide a redescription of the species based on the new specimens, complete with illustrations and new data concerning its morphology, and we also place some of the collected specimens in a molecular phylogenetic framework.

ZooKeys, 2012

Namalycastis jaya sp. n. (Polychaeta: Nereididae: Namanereidinae) is described from the southern ... more Namalycastis jaya sp. n. (Polychaeta: Nereididae: Namanereidinae) is described from the southern coast of Kerala in southwest India. One important characteristic feature of the species is the lack of notochaetae in all parapodia, a characteristic that it shares with at least two other species, Namalycastis elobeyensis Glasby, 1999 and Namalycastis hawaiiensis Johnson, 1903. It differs from N. elobeyensis by virtue of its smaller antennae, unequal eye size, bilobed acicular neuropodial ligule and multi-incised pygidium rim. Moreover, it differs from N. hawaiiensis by having fewer teeth on the serrated blades of the sub-neuroacicular falciger in chaetiger 10, and by possessing finely serrated falcigers in posterior segments. Beyond morphological analyses, molecular phylogenetics was used for the first time for Namalycastis to support population monophyly and recognition of the new species. The analysis, using both mitochondrial and nuclear data, corroborated the morphological analysis in suggesting that our specimens represent an as yet undescribed species, Namalycastis jaya sp. n., which forms a monophyletic group among the sampled nereidid taxa. Finally, a taxonomic key for Namalycastis species recorded from the Indian region is provided.

PLoS ONE, 2014

In spite of the high relevance of lumbricid earthworms ('Oligochaeta': Lumbricidae) for soil stru... more In spite of the high relevance of lumbricid earthworms ('Oligochaeta': Lumbricidae) for soil structure and functioning, the taxonomy of this group of terrestrial invertebrates remains in a quasi-chaotic state. Earthworm taxonomy traditionally relies on the interpretation of external and internal morphological characters, but the acquisition of these data is often hampered by tedious dissections or restricted access to valuable and rare museum specimens. The present state of affairs, in conjunction with the difficulty of establishing primary homologies for multiple morphological features, has led to an almost unrivaled instability in the taxonomy and systematics of certain earthworm groups, including Lumbricidae. As a potential remedy, we apply for the first time a non-destructive imaging technique to lumbricids and explore the future application of this approach to earthworm taxonomy. High-resolution micro-computed tomography (mCT) scanning of freshly fixed and museum specimens was carried out using two cosmopolitan species, Aporrectodea caliginosa and A. trapezoides. By combining two-dimensional and three-dimensional dataset visualization techniques, we demonstrate that the morphological features commonly used in earthworm taxonomy can now be analyzed without the need for dissection, whether freshly fixed or museum specimens collected more than 60 years ago are studied. Our analyses show that mCT in combination with soft tissue staining can be successfully applied to lumbricid earthworms. An extension of the approach to other families is poised to strengthen earthworm taxonomy by providing a versatile tool to resolve the taxonomic chaos currently present in this ecologically important, but taxonomically neglected group of terrestrial invertebrates.

Journal of Parasitology, 2012

Invertebrate Biology, 2013

Freshwater, marine, and terrestrial leeches that depend on a diet of fresh blood have evolved sal... more Freshwater, marine, and terrestrial leeches that depend on a diet of fresh blood have evolved salivary peptide components that inhibit normal thrombus formation by prey. Although bloodfeeding in leeches has long been of interest to biologists and medical practitioners alike, only a few studies have comprehensively examined the anticoagulant repertoires of hematophagous leeches, and these have largely been confined to representatives of Glossiphoniidae, Hirudinidae, and Macrobdellidae. Here, we present 454 pyrosequencing data from the salivary transcriptome of the hematophagous terrestrial leech Haemadipsa interrupta (Haemadipsidae). Assembled transcripts were annotated using both similarity scores (BLAST) and gene ontology (Blast2GO). Subsequently, transcripts were examined within alignments containing well-characterized anticoagulants and other select bioactive (i.e., affecting the living cells of the prey) salivary proteins and phylogenies were reconstructed for each protein data set to verify orthology predictions. In total, transcripts significantly matching 20 salivary proteins of interest were found in the transcriptome, representing several different antagonistic pathways. After reviewing gene ontologies, alignments, and phylogenetic trees, sequences for 15 out of the 20 hits were deemed correctly annotated. Additionally, we recovered matches against several proteins that have previously been linked to anticoagulation (e.g., cathepsin and disintegrins), but the specific function of which in leeches needs further investigation. Finally, in light of these data as well as those previously published, we discuss our current understanding of the distribution and evolution of anticoagulants in leeches.

Cladistics, 2013

We present phylogenomic analyses of the most comprehensive molecular character set compiled for A... more We present phylogenomic analyses of the most comprehensive molecular character set compiled for Annelida and its constituent taxa, including over 347 000 aligned nucleotide sites for 39 taxa. The nucleotide data set was recovered using a pre-existing amino acid data set of almost 48 000 aligned sites as a backbone for tBLASTn searches against NCBI. In addition, orthology determinations of the loci in the original amino acid data set were scrutinized using an All vs All Reciprocal Best Hit approach, employing BLASTp, and examining for statistical interdependency among the loci. This approach revealed considerable sequence redundancy among the loci in the original data set and a new data set was compiled, with the redundancy removed. The newly compiled nucleotide data set, the original amino acid data set, and the new reduced amino acid data set were subjected to parsimony analyses and two forms of bootstrap resampling. The last-named data set also was analysed using a maximum-likelihood approach. There were two main objectives to these analyses: (i) to examine the general topology, including support, resulting from the analyses of the new data sets and (ii) to assess the consistency of the branching patterns across optimality criteria by comparison with previous probabilistic approaches. The phylogenetic hypotheses resulting from analyses of the three data sets are largely unsupported, reflecting the continued difficulty of finding numerous, reliable, and suitable loci for a group as ancient as Annelida. Resulting parsimonious hypotheses disagree, in some respects, with the previous probabilistic approaches; Sedentaria and, in most cases, Errantia are not supported as monophyletic groups but Pleistoannelida is recovered as a (unsupported) monophyletic group in one of the three parsimony analyses as well as the likelihood analysis. In addition, we performed missing data titration studies to estimate the impact of missing data on overall support and support for specific clades.

Mitochondrial DNA, 2010

Species identifications based on DNA barcoding rely on the correct identity of previously barcode... more Species identifications based on DNA barcoding rely on the correct identity of previously barcoded specimens, but little attention has been given to whether deposited barcodes include correspondence to the species' name-bearing type. The information content associated with COX1 sequences in the two most commonly used repositories of barcodes, GenBank and the Barcode of Life Data System (BOLD), is often insufficient for subsequent evaluation of the robustness of the identification procedure. We argue that DNA barcoding and taxonomy alike will benefit from more information content in the annotations of barcoded specimens as this will allow for validation and re-evaluation of the initial specimen identification. The aim should be to closely connect specimens from which reference barcodes are generated with the holotype through straightforward taxonomy, and geographical and genetic correlations. Annotated information should also include voucher specimens and collector/identifier information. We examine two case studies based on empirical data, in which barcoding and taxonomy benefit from increased information content. On the basis of data from the first case study, we designate a barcoded neotype of the European medicinal leech, Hirudo medicinalis, on morphological and geographical grounds.

The description of Helobdella stagnalis (Linnaeus, 1758) has emphasized the presence of a nuchal,... more The description of Helobdella stagnalis (Linnaeus, 1758) has emphasized the presence of a nuchal, chitinous scute located on the dorsal surface in the first third of the body as the diagnostic character for the species. Historically, identifications of species of Helobdella have relied heavily on this character and, as a result, Helobdella stagnalis has been reported from an inordinately broad geographic range, including Europe, Asia, Africa, North America, and South America. In addition to a few earlier investigations, a recent analysis showed that great genetic distances (orders of magnitude greater than previous estimations of intraspecific divergence in leeches) are present between scute-bearing specimens identified as H. stagnalis from Europe and North America, implying that H. stagnalis does not occur in North America. The present study expands the geographic boundaries of taxon sampling for both European and North American taxa, and reexamines the phylogenetic relationships and cytochrome c oxidase subunit I (COI) variation within scute-bearing species of the genus Helobdella. Our analyses include specimens putatively identified as "Helobdella stagnalis" from Sweden, and Iran, as well as numerous localities covering Canada and the USA. Our results corroborate previous studies in that European and west Asian specimens form a clade, including the neotype, which is separate from North American taxa. To alleviate future taxonomic confusion, we redescribe H. stagnalis and designate a neotype from the inferred type locality. The designation of a neotype stabilizes the taxonomy of scute-bearing leeches of the genus Helobdella and enables us to definitively correct erroneous identifications reported in previous studies. We also note that at least four lineages of scute-bearing, North American species of Helobdella lack formal descriptions.

Leeches of the family Erpobdellidae are important members of benthic freshwater environments, whe... more Leeches of the family Erpobdellidae are important members of benthic freshwater environments, where they are voracious predators of other invertebrates and an important source of nutrition for several species of vertebrates. Beset by a lack of reliable diagnostic morphological characters and destructive identification processes, molecular approaches have, in recent years, been employed to illuminate the relationships within this family, and DNA barcoding has been employed for identification purposes. However, an understanding of the levels of genetic variation across the geographic distributions of members of the genus is still lacking. Herein, we sequence the mitochondrial COI locus for 249 newly collected North American individuals, representing 5 species, as well as mitochondrial 12S rDNA, nuclear 18S rDNA, and nuclear 28S rDNA for a select subset of these. Our COI dataset was leveraged to detect potential cryptic species, and to calculate genetic distances as a proxy for the degree of gene flow between populations. Augmented by numerous sequences from GenBank, the multilocus dataset was used to reconstruct a phylogenetic hypothesis for worldwide members of the genus. Beyond corroborating previous overarching phylogenetic frameworks, our results show that an undescribed species that is morphologically and genetically similar to Erpobdella punctata exists in sympatry with this species-the new species has likely been overlooked in previous studies due to its morphological similarity with Erpobdella punctata. Erpobdella bucera is reported from Canada for the first time; and Erpobdella microstoma is newly reported from Saskatchewan and placed in a phylogeny for the first time. Finally, we find evidence for genetic structure in both E. cf. punctata and Erpobdella obscura that is correlated with major river drainage basin boundaries in North America.

Blood-feeding leeches of the genus Placobdella have acquired intracellular alphaproteobacterial e... more Blood-feeding leeches of the genus Placobdella have acquired intracellular alphaproteobacterial endosymbionts of the genus Reichenowia that potentially aid in the production of B vitamins, thereby ameliorating the lack of these essential nutrients in the diet of the hosts. For Placobdella associates, little is still known about the diversity, genetic makeup, and the mode of transmission of bacteria between leeches. We aimed to (i) place the bacterial symbionts in a phylogenetic context, (ii) compare patterns of cladogenesis between the bacteria and hosts to search for evidence of co-speciation, and (iii) explore the mode of bacterial transmission between leech parent and offspring. DNA sequencing of the bacterial 16S rDNA and 23S rDNA loci suggests that, whereas Reichenowia forms a monophyletic group within the alphaproteobacterial family Rhizobiaceae, no evidence for co-speciation between hosts and bacteria can be traced. Attempts at DNA amplification for ovarial tissues were negative for a range of species, but two 16S rDNA sequences retrieved from the testisacs of P. rugosa showed very high similarity with Reichenowia. Although we cannot rule out that this may be a contamination, or a different, potentially free-living species of bacteria, our results may indicate that Reichenowia is transferred from leech parent to offspring via the testisacs.

In all but one case, commensal alphaproteobacteria form close relationships with plant hosts, aid... more In all but one case, commensal alphaproteobacteria form close relationships with plant hosts, aiding in, among other things, nitrogen fixation and nodulation. The odd one out, Reichenowia parasitica, is an obligate endosymbiont of the freshwater glossiphoniid leech Placobdella parasitica. Housed in the leech mycetomes (esophageal organs, the only known function of which is to house bacteria), the bacteria have been hypothesized to provide their host with essential vitamins otherwise lacking from its strict hematophagous diet. A previous phylogenetic analysis, based solely on the ribosomal DNA gene 16S, placed R. parasitica within the family Rhizobiaceae, including known plant pathogens and symbionts. However, because the bacteria proved refractory to a series of attempts at in vitro culturing, isolation of a mixed pool of DNA from the leech mycetomes and subsequent sequencing quickly becomes awkward. Therefore, we performed 454 pyrosequencing on a DNA sample isolated from 39 pairs of mycetomes and used subtractive scaffolding, employing a scoring scheme, to separate leech and bacterial DNA in silico. Using MUMmer 3, we then identified housekeeping regions with valuable phylogenetic information and identified orthologs across a series of alphaproteobacteria. In total, 20 contigs were identified and jointly subjected to phylogenetic analysis to provide a more rigorous hypothesis of the position of R. parasitica within the alphaproteobacteria. Results suggest that the bacterium is in fact part of the Rhizobiaceae family, its closest relative being the pathogenic plant-parasite Agrobacterium. Our technique of in silico a posteriori subtractive scaffolding is apparently novel and will likely prove effective for other non-cultivable, obligate endosymbionts.