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Phylogenetic analysis of Placobdella (Hirudinea: Rhynchobdellida: Glossiphoniidae) with consideration of COI variation
Abstract
Placobdella is a genus of blood-feeding leeches in the family Glossiphoniidae. Historically, species of Placobdella have posed difficulty for systematists owing to a lack of informative morphological characters and the preponderance of inadequate or incomplete species descriptions. Here, we conduct a phylogenetic analysis of 55 individuals representing 20 of the 24 currently recognized nominal taxa using COI, ND1, 12S rDNA and ITS sequences under parsimony, maximum likelihood and Bayesian inference. We also examine the isolated COI phylogeny for the genus using an expanded dataset encompassing three additional species not included in the concatenated dataset. Finally, we assess genetic variation at the COI locus to validate initial specimen identifications and estimate how COI variation may reflect species boundaries. We conclude that Placobdella is a monophyletic group that places as the sister group to a clade formed by the genera Haementeria and Helobdella. We discuss the evolutionary implications of several internal relationships that are robustly resolved by all three optimality criteria, paying particular attention to the apparent fluidity of morphological characters exhibited by members of Placobdella. We also find preliminary evidence for the presence of cryptic and undescribed diversity within the genus.
... The partial sequence of mitochondrial cytochrome c oxidase subunit I (COI), nuclear 18S rRNA (18S) and Internal Transcribed Spacer 1 and 2 flanking the 5.8S rRNA (ITS) of representative specimens were PCR-amplified using the primers LCO1490-HCO2198 and LCO-inerpo2-HCO-out (Folmer et al. 1994;Nakano 2016), A-B (Medlin et al. 1988;Apakupakul et al. 1999), and ITS5-ITS4 (White et al. 1990) respectively. The PCR and cycle sequencing reactions were performed according to Nakano (2012) for COI and 18S, and de Carle et al. (2017) for ITS, except for the use of the PCR reaction kit, EmeraldAmp PCR Master Mix (Takara Bio). The newly determined sequences were deposited in the International Nucleotide Sequence Databases (INSD) through the DNA Data Bank of Japan (DDBJ). ...
... Our COI alignment matrix comprised 1267 bp representing 31 haplotypes of Japanese Torix. The maximum uncorrected pairwise genetic distance among Torix collected in this study was 5.2% ( Table 2) that is smaller than the range of interspecific variation observed in other glossiphoniid species (de Carle et al. 2017;Kaygorodova et al. 2020;Jovanović et al. 2021). ...
... The ITS data revealed low genetic diversity within Japanese Torix individuals (uncorrected p-distance <1.1%) that is within the range of intraspecific variation in Placobdella (de Carle et al. 2017). Network analysis indicated that Torix in Japan can be broadly divided into the eastern and western lineages (Fig. 3), as indicated in the phylogenetic analysis of COI (Fig. 2). ...
Although most members of the freshwater leech family Glossiphoniidae have mid-body somites divided into three annuli, the genus Torix Blanchard, 1893 is distinguished by two-annuli somites. Torix has high species richness in Far East Asia, and three nominal species have been recognised in the Japanese Archipelago and adjacent regions that can be distinguished by a combination of both internal and external morphological characteristics. However, recent studies have shown that these diagnostic features are ontogenetically variable and this has resulted in taxonomic confusion among Torix species endemic to the Japanese Archipelago. In this study, we revisit the taxonomic accounts of T. orientalis (Oka, 1925) and T. tagoi (Oka, 1925), in addition to that of the recently redescribed T. tukubana (Oka, 1935) to clarify the diagnostic characteristics for each of the three species. Our morphological and molecular phylogenetic analyses demonstrated that the three Torix species in Japan are indistinguishable. We therefore conclude that these species should be synonymised and treated as a single species. The specific names orientalis and tagoi were simultaneously established under the genus Oligobdella Moore, 1918, therefore we acted as First Reviser and gave precedence to the name O. tagoi, thus the valid name for the Far East Asian Torix species is T. tagoi unless T. orientalis and T. tagoi are treated as distinct species. There are several type localities for T. tagoi and the name-bearing types have been lost, therefore we designate a neotype for this species to obviate zoological and nomenclatural issues.
ZooBank: urn:lsid:zoobank.org:pub:E2DBF999-3B51-456F-AB04-A7D138E0AF2D
... In addition, the SH-aLRT branch test was used to evaluate the tree branch supports. The tree was rooted at Haementeria ghilianii Filippi, 1849 following the previous phylogenetic hypotheses (e.g., [18]) and visualized with iTOL [19]. ...
... The combination mentioned above should now be regarded as a potential synapomorphy shared by these three Placobdella species. Additionally, de Carle and coworkers [18] suggested that compact salivary tissue is plesiomorphic for Placobdella, and that diffuse salivary glands have evolved independently several times within the genus. Therefore, the organization of the salivary glands showed great variety among studied Placobdella species. ...
... Moreover, several other species with only diffuse salivary glands have been reported (e.g. P. pediculata, P. cryptobranchii, P. phalera, P. michiganensis, and P. picta) [18,45]. These species were included by Barta and Sawyer [51] in the genus Desserobdella, in light of their possession of diffuse salivary tissue versus the two pairs of compact salivary glands distinguishing species of the genus Placobdella. ...
Background
Herein, we describe a new species of turtle blood-feeding leech, Placobdella nabeulensis sp. nov. from Palearctic North Africa (Tunisia and Algeria). The new species is described based on detailed morphological analyses using light and scanning electron microscopes.
Results
Apart from the detailed morphology of the atrium, morphological features alone do not sufficiently separate the species from congeners due to the absence of distinct diagnostic characters. Therefore, we turned to molecular data to better distinguish this new species from other members of the genus and establish a basis for its genetic separation. Four DNA fragments were successfully amplified, including mitochondrial COI and 12S rDNA, as well as nuclear 28S rDNA and histone H3. We then provided the molecular descriptor of the taxon, based on redundant diagnostic nucleotide combinations in DNA sequence alignment within the Folmer region. Results of the phylogenetic analysis and species delimitation methods (ABGD, ASAP, and bPTP) based on the COI locus support the species rank of the Tunisian-Algerian Placobdella.
Conclusions
The new species is most closely related to the European species Placobdella costata (Fr. Müller, 1846) and the present study indicates that Placobdella nabeulensis sp. nov. has likely been confused with the European counterpart in several previous studies. This article is registered at www.zoobank.org under urn:lsid:zoobank.org:pub:4A4B9C1D-2556-430F-8E4B-0CE99F2012F5.
... DNA from Alaskan specimens was amplified according to our prior work (Tessler et al., 2018a) for 28S, 18S, 16S and COI; 12S amplification followed protocols from our other prior work (de Carle et al., 2017). Throughout the course of our research, we noticed that acanthobdellidan DNA is difficult to amplify and sequence. ...
... Paracanthobdella livanowi is sister to A. peledina ( Fig. 4; Supporting Information, Files S2-S4), as was expected based on morphology. The genetic distance between the two species at the COI locus (13.20%) is higher than values for other hirudinean species pairs, which has been reported at ~8% (Oceguera-Figueroa et al., 2010;de Carle et al., 2017;Iwama et al., 2019). Interestingly, although the divergence between A. peledina and P. livanowi is substantial, the split is relatively recent, especially in comparison to the long branch length for Acanthobdellida and the level of variability found in the closely related branchiobdellidans and leeches. ...
... RADSeq) would be useful for determining whether gene flow exists between the Alaskan and Nordic localities. Unfortunately, COI, the most common marker for determining differences between leech species and populations (de Carle et al., 2017;Tessler et al., 2018c;Mack et al., 2019), and additional nuclear loci did not amplify for these samples, potentially leading to some issues with missing data. Furthermore, given that no external morphological differences were noted between samples of Nordic and Alaskan A. peledina, we refrain from formal species or population delimitation analyses at this time. ...
Acanthobdellida gnaw into the sides of salmonid fishes in frigid Arctic lakes and rivers, latching on with fearsome facial hooks. Sister to leeches, they are an ancient lineage with two described species. Unfortunately, Acanthobdellida are rarely collected, leading to a paucity of literature despite their unique morphology. Populations range from Eurasia to Alaska (USA), but few specimens of Acanthobdella peledina are represented in molecular studies, and no molecular data exist for Paracanthobdella livanowi, making their taxonomic position difficult to assess. We use phylogenetics and morphology to determine whether allopatric populations of A. peledina are distinct species and assess the current classification scheme used for Acanthobdellida. We produce a new suborder, Acanthobdelliformes, to match the *Corresponding authors.
... DNA from Alaskan specimens was amplified according to our prior work (Tessler et al., 2018a) for 28S, 18S, 16S and COI; 12S amplification followed protocols from our other prior work (de Carle et al., 2017). Throughout the course of our research, we noticed that acanthobdellidan DNA is difficult to amplify and sequence. ...
... Paracanthobdella livanowi is sister to A. peledina ( Fig. 4; Supporting Information, Files S2-S4), as was expected based on morphology. The genetic distance between the two species at the COI locus (13.20%) is higher than values for other hirudinean species pairs, which has been reported at ~8% (Oceguera-Figueroa et al., 2010;de Carle et al., 2017;Iwama et al., 2019). Interestingly, although the divergence between A. peledina and P. livanowi is substantial, the split is relatively recent, especially in comparison to the long branch length for Acanthobdellida and the level of variability found in the closely related branchiobdellidans and leeches. ...
... RADSeq) would be useful for determining whether gene flow exists between the Alaskan and Nordic localities. Unfortunately, COI, the most common marker for determining differences between leech species and populations (de Carle et al., 2017;Tessler et al., 2018c;Mack et al., 2019), and additional nuclear loci did not amplify for these samples, potentially leading to some issues with missing data. Furthermore, given that no external morphological differences were noted between samples of Nordic and Alaskan A. peledina, we refrain from formal species or population delimitation analyses at this time. ...
Acanthobdellida gnaw into the sides of salmonid fishes in frigid Arctic lakes and rivers, latching on with fearsome
facial hooks. Sister to leeches, they are an ancient lineage with two described species. Unfortunately, Acanthobdellida
are rarely collected, leading to a paucity of literature despite their unique morphology. Populations range from Eurasia
to Alaska (USA), but few specimens of Acanthobdella peledina are represented in molecular studies, and no molecular
data exist for Paracanthobdella livanowi, making their taxonomic position difficult to assess. We use phylogenetics
and morphology to determine whether allopatric populations of A. peledina are distinct species and assess the current
classification scheme used for Acanthobdellida. We produce a new suborder, Acanthobdelliformes, to match the taxonomy within Hirudinea. Scanning electron micrographs indicate species-level differences in the anterior sucker and facial hooks; molecular phylogenetics mirrors this divergence between species. We assign both species to the family Acanthobdellidae and abandon the family Paracanthobdellidae. Alaskan and European A. peledina populations are morphologically similar, but appear phylogenetically divergent. Our data strongly suggest that members of the order Acanthobdellida diverged relatively recently in their ancient history, but based on genetic distance, this divergence appears to pre-date the most recent cycles of glaciation.
... The species is externally characterized by a dark, dorsomedial stripe interrupted by areas of lighter pigmentation; five dorsal rows of papillae, with heavy secondary papillation, and a distinctive pattern of two rows of three papillae, followed by two pairs of paramedial papillae adjacent to the anus; a repeated marginal alternation of two light-coloured annuli, followed by one dark annulus in mid-body segments; and an unpigmented 'mask' near the two pairs of coalesced eyespots (Moser et al. 2012). Previous observations of P. rugosa, however, demonstrate that these characteristics are not universal: instead, papillation and coloration are highly variable among individuals, more so than in other species of Placobdella (Klemm 1985;Moore 1905;Moser et al. 2012;de Carle et al. 2017;Langer et al. 2018) (see also Figure 1). Furthermore, there has been over a century of taxonomic confusion between P. rugosa and Placobdella ornata (Verrill 1872). ...
... A recent investigation into phylogenetic relationships and species delimitation within the genus Placobdella revealed a lack of phylogeographic structure among the sampled specimens of P. rugosa (de Carle et al. 2017). The nine specimens included in the study displayed significant variation in patterns of dorsal papillation and pigmentation, characters which are commonlyand reliablyused to diagnose congeneric species (Klemm 1985). ...
... In spite of this, phylogenetic analyses revealed that all nine specimens nested together with short branch lengths, and their relationships did not reflect geographic structure. Observed genetic distances in these specimens were also low relative to other leech species: average intraspecific distance at the cytochrome c oxidase subunit I (COI) locus was 0.9% (minimum ¼ 0.000%; maximum ¼ 2.2%) (de Carle et al. 2017). These findings were surprising for a number of reasons. ...
Placobdella rugosa has long presented challenges to leech biologists. Its extreme morphological variability and similarity to some congeneric species has confounded classification for over a century. Recent molecular analyses revealed a surprising lack of genetic variation among morphologically disparate, geographically widespread specimens of P. rugosa. Given the lack of any obvious mechanism by which this species could disperse between distant habitats, it was expected that widespread populations would be genetically isolated from each other. In the present study, we investigate the relationship between geographic distance and genetic diversity in P. rugosa using COI sequences from specimens collected across Canada and the United States. Although we find preliminary evidence for a barrier to gene flow between eastern and western collecting localities, our vastly expanded dataset largely corroborates prior studies, showing minimal phylogeographic signal among the sequences and negligible levels of genetic isolation by distance. A recent range expansion following the last ice age and/or host-mediated dispersal are discussed as potential explanations for this unexpected phylogeographic pattern.
... This has indeed been reported from M. caspica in Iran [32,33], from E. trinacris in Sicilia [34], and from M. leprosa in Spain [35,36], Morocco [37], and Tunisia [15]. While it is now well received from the investigation of molecular markers that Placobdella is a monophyletic genus, certain cryptic diversity was evidenced within several species, among which are Placobdella mexicana Moore 1898 and Placobdella ringueleti López-Jiménez and Oceguera-Figueroa 2009 [38]. Based on COI and ITS1 genetic variations, Kvist et al. [39] also illustrated seven independent lineages within P. costata, suggesting at least five unique leech species across Central and Eastern Europe, as well as in Algeria. ...
... The cytochrome c oxidase I (COI) of leeches was amplified using the forward LCO Plac 5 -AYTCAACTAATCAYAAAGAYATTGG-3 and reverse HCO Plac 5 -TADACTTCWGGRTGACCAAAAAATCA-3 primers which were designed for Placobdella spp. [38]. The COI of haemogregarines was amplified using the forward HemoFor4 5 -TGGACATTATACCCACCTTTAAG-3 and reverse HemoRev4 5 -ATACAACCCATAGCTAGTATCAT-3 primers which were designed for Haemogregarina spp. ...
Haemogregarines are blood parasites with a life-cycle involving a vertebrate as the intermediate host and an invertebrate as the definitive host and vector. Extensive phylogenetic investigations based on 18S-rRNA gene sequences have shown that Haemogregarina stepanowi (Apicomplexa: Haemogregarinidae) is able to infest a large diversity of freshwater turtle species, including the European pond turtle Emys orbicularis, the Sicilian pond turtle Emys trinacris, the Caspian turtle Mauremys caspica, the Mediterranean pond turtle Mauremys leprosa, and the Western Caspian turtle Mauremys rivulata, among others. From the same molecular markers, H. stepanowi is further considered to be a complex of cryptic species predisposed to infect the same host species. While Placobdella costata is known to be the unique vector of H. stepanowi, it is only recently that independent lineages within P. costata have been illustrated—suggesting the presence of at least five unique leech species across Western Europe. The aims of our study were therefore to investigate from mitochondrial markers (COI) the genetic diversity within haemogregarines and leeches infecting freshwater turtles of the Maghreb, in order to identify processes of parasite speciation. We showed that H. stepanowi consists of at least five cryptic species in the Maghreb, while two Placobella species were identified in the same area. Although an Eastern–Western speciation pattern was apparent for both leeches and haemogregarines, we cannot make definitive conclusions regarding co-speciation patterns between parasites and vectors. However, we cannot reject the hypothesis of a very strict host–parasite specificity within leeches.
... The other is P. rugosa (Verrill, 1874) [9,10] of the northern half of the United States and adjacent southern Canada [11][12][13]. A recent morphological study has shown that P. rugosa is barely distinguishable from P. multilineata Moore, 1953, of the southeast United States [9,14]. ...
... This finding concurs well with a parallel study of low molecular diversity of this species [8]. Parenthetically, a molecular phylogenetic study of P rugosa documents a similar low diversity [12,13]. ...
The genus Placobdella (Glossiphoniidae) has a centre of species concentration in North America. The type species P. costata is the only representative in the Palaearctic region. American Placobdella which feed on turtles are represented predominantly by two common species, P. parasitica and P. rugosa, which geographically overlap in eastern USA and southern Canada. The latter species is morphologically indistinguishable from P. multilineata of southeast USA. These two tuberculated forms are recognised herein as a clade and treated as a single ‘species’ for comparative purposes. Both P. parasitica and P. rugosa clade commonly coexist and feed on the same turtle species without host preference. This paper addresses morphological differences between them. An unexpected finding is that hatchlings of both species are very difficult to distinguish. A morphologically significant observation is that developmental divergence results in tubercle prominence in P. rugosa clade, but tubercle suppression in P. parasitica, the first example of character displacement in the Hirudinea. Morphological differences are interpreted as reflecting interspecific competition, a phenomenon not found in their Palaearctic counterpart. Why do the two American turtle leech species coexist rather than reduce competition by partitioning their food supply?
... The most conspicuous bloodsucking leech in these blackwater swamps is a relatively large tuberculated species of Placobdella typically encountered on the snapping turtle Chelydra serpentina (Linnaeus). Although this leech is common in the Albemarle region its identification is indeterminate in that it differs in detail from the more than twenty nominal species of Placobdella reported from North America (Moser et al. 2016;de Carle et al. 2017). However, designation of a new species of Placobdella is not justified at this time for the reasons discussed below. ...
... Placobdella rugosa is widely distributed, variable and frequently common in the northern United States and adjacent parts of Canada (de Carle et al. 2017;Mack et al. 2019). This species has a tortuous taxonomic history which has now been convincingly stabilised (Moser et al. 2012). ...
A tuberculated species of turtle leech is indigenous to the Great Dismal Swamp and environs of northeastern North Carolina, and differs from other known species of Placobdella. This study of hundreds of specimens for more than a decade documents its unexpected taxonomic complexity. In fact, this seemingly innocuous leech undergoes radical transformations in terms of morphology and behaviour, each adapted to a different phase of its life cycle. Biological observations reveal a progressive darkening with age which imposes taxonomic uncertainties. Furthermore, some commonly used characters are found to be unsound for taxonomic diagnosis in that they do not occur in all individuals of this species.
The primary objective of this paper is a comprehensive description of this Albemarle turtle leech. The question is then asked, what distinguishes it from allied species? Toward this end, a formal taxonomic diagnosis is proposed based on details of the proboscis complex and crop-related tuberculation. This species is allied to the northern P. rugosa (Verrill, 1874) or its southern counterpart P. multilineata Moore, 1953. However, synonymy to either of these forms is problematical due to inadequacy of type descriptions. Nonetheless, in view of habitat similarity the Albemarle leech is provisionally identified as P. multilineata pending comparable studies of variability and development of other tuberculated Placobdella.
Key words: Accessory eye, mycetome, hollensis, parasitica, rugosa, turtle parasite
... The most conspicuous bloodsucking leech in these blackwater swamps is a relatively large tuberculated species of Placobdella typically encountered on the snapping turtle Chelydra serpentina (Linnaeus). Although this leech is common in the Albemarle region its identification is indeterminate in that it differs in detail from the more than twenty nominal species of Placobdella reported from North America (Moser et al. 2016;de Carle et al. 2017). However, designation of a new species of Placobdella is not justified at this time for the reasons discussed below. ...
... Placobdella rugosa is widely distributed, variable and frequently common in the northern United States and adjacent parts of Canada (de Carle et al. 2017;Mack et al. 2019). This species has a tortuous taxonomic history which has now been convincingly stabilised (Moser et al. 2012). ...
A tuberculated species of turtle leech is indigenous to the Great Dismal Swamp and environs of northeastern North Carolina, and differs from other known species of Placobdella. This study of hundreds of specimens for more than a decade documents its unexpected taxonomic complexity. In fact, this seemingly innocuous leech undergoes radical transformations in terms of morphology and behaviour, each adapted to a different phase of its life cycle. Biological observations reveal a progressive darkening with age which imposes taxonomic uncertainties. Furthermore, some commonly used characters are found to be unsound for taxonomic diagnosis in that they do not occur in all individuals of this species. The primary objective of this paper is a comprehensive description of this Albemarle turtle leech. The question is then asked, what distinguishes it from allied species? Toward this end, a formal taxonomic diagnosis is proposed based on details of the proboscis complex and crop-related tuberculation. This species is allied to the northern P. rugosa (Verrill, 1874) or its southern counterpart P. multilineata Moore, 1953. However, synonymy to either of these forms is problematical due to inadequacy of type descriptions. Nonetheless, in view of habitat similarity the Albemarle leech is provisionally identified as P. multilineata pending comparable studies of variability and development of other tuberculated Placobdella.
... The species was originally described as a subspecies of Placobdella ornata (Verrill, 1872), subsequently elevated to full species status, reassigned to P. ornata, and finally resurrected in 2012 (Moser et al. 2012 and references therein). The species is abundant across Canada and is associated with varying climates and faunas (Klemm 1982;de Carle et al. 2017). This particular specimen was collected from the Sandy Lake ecodistrict (Table 1). ...
... Both Erpobdella and Haemopis are predaceous, with prey items ranging from oligochaetes and insect larvae, to snails, small crustaceans, and dead fish (Sawyer 1972;Barton and Metcalfe 1986;Kutschera and Wirtz 2001;Moser et al. 2006). Moreover, previous expeditions have successfully collected representatives of both genera using aquatic traps baited with beef liver and ground beef (de Carle et al. 2017). Predaceous leeches swallow their prey either whole or in pieces (Kutschera and Wirtz 2001;Pfeiffer et al. 2005;Sket and Trontelj 2008). ...
Leeches have a worldwide distribution, yet numerous geographical regions remain to be adequately surveyed. Here, we present leech species records for one of these regions: the far north of Ontario, Canada. This region is primarily wetland habitat and includes two of Ontario’s three ecozones. Morphological examinations, as well as a single instance of successful DNA amplification with subsequent molecular identification, allowed us to identify representatives of 12 species from two predatory families (Erpobdellidae and Haemopidae) and one parasitic family (Glossiphoniidae) among samples of 130 individuals. To provide a more inclusive list of species records for this remote region, our data were also augmented by 25 largely unpublished collection records (for 102 individuals) from the Canadian Museum of Nature, which revealed the presence of an additional species. We comment on finds of particular interest in our sampling with comparison to relevant literature and provide new distribution data for these species.
... Van Dam et al., 2019; Camacho et al., 2022) ． ところが，Phillips et al. Siddall and Burreson, 1998; Apakupakul et al., 1999) ．より大規模なデータセットを用いた Tessler et al. (2018) の解析でも，やはり吻蛭目の側系統性が 支持されたことから，系統関係を反映した分類体系を 構築するため，ヒラタビル科からなるヒラタビル形亜 目 Glossiphoniiformes Tessler & De Carle, 2018 ならびに エラビル科とウオビル科を含むカイヨウビル形亜目 Oceanobdelliformes Tessler & De Carle, 2018 が新設され た(Tessler et al., 2018) ．ところが，ゲノムデータに 基づく解析では，ヒラタビル形亜目がカイヨウビル形 亜目と姉妹群を形成する，つまり吻蛭目が単系統群を 形成する可能性が再び提示されている(Phillips et al., 2019; Erséus et al., 2020) ．ヒラタビル形亜目の系統的 位置については，依然として議論が続いている． その一方，吻無蛭目については一貫して単系統群で あることが支持されてきた(Apakupakul et al., 1999; Borda and Siddall, 2004) ．ただし，吻蛭目の解体に合 図 2. ヒル亜綱に属する 3 目の主要な系統仮説． わせ，吻無蛭目も従来から用いられてきたイシビル形 亜目とチスイビル形亜目，そして新設のアメリカビル 形 亜 目 Americobdelliformes Siddall, De Carle & Tessler, 2018 に分割された(Tessler et al., 2018) ．なお，最近 のヒル目での分類体系の整理に対応する形で，ケビル 目においてもケビル形亜目 Acanthobdelliformes Cios, De Carle, Świątek, Tessler & Utevsky, 2022 が設立されて いる(De Carle et al., 2022) ． ヒル目の最新の分類体系 Tessler et al. (2018) によって提唱されたヒル目を構 成する 5 亜目と，各亜目に属する分類群について以下 に 紹 介 す る(表 1, 図3) ． 各 科 の 所 属 は Tessler et al. (2018) に従ったが，解析に含まれていない科について は Nakano et al. (2018) を参考に，情報を更新した． 1. ヒラタビル形亜目 本亜目はヒラタビル科のみによって構成されるが， その既知種数は 200 を越え，ヒル目で最も多様な科と して知られている(Sket and Trontelj, 2008) ．全て淡水 棲であり，咽頭に有する吻を突き刺すことで脊椎動物 の血液または無脊椎動物の体液を吸う(Sawyer, 1986(Sawyer, ) , 1999Siddall et al., 2005;De Carle et al., 2017; Bolotov et al., Trontelj, 2004;Williams and Burreson, 2006;Utevsky et al., 2007; Bolotov et al., 2020) ．また，両科ともヒラタ ビル形亜目と同様に吻を有する．主にエラビル科はカ メ類，ウオビル科は魚類から吸血するが，後者では無 図 3. ヒル亜綱の高次系統関係．Nakano et al. (2018)，Tessler et al. (2018)，Phillips et al. (2019) を参考に作成．枝の長さは分岐 年代などには比例していない． タクサ No. 57 (2024) ...
Leeches in the order Hirudinida comprise approximately 700 species that are distributed worldwide and occupy diverse habitats, including freshwater, marine, and terrestrial habitats. Leeches have traditionally been classified into two main groups, Rhynchobdellida and Arhynchobdellida, on the basis of the presence or absence of a proboscis. However, recent molecular analyses using large datasets have suggested that Rhynchobdellida is a paraphyletic group, and this has led to calls for a new classification that better reflects phylogeny. In addition, recent studies have clarified the previously unre-solved phylogenetic relationships among leeches and two leech-like worms, Acanthobdellida and Branchiobdellida. Here, we discuss recent trends in the higher-level phylogenetic relationships and classification of leeches.
... (Phillips et al., 2019). While the COI distances for M. decora are much higher than M. mimicus, these values still fall within the range of intra-specific genetic diversity values seen for other leeches: the average COI distance of Erpobdella parva was 1.40 as seen in Anderson et al. (2020); average intraspecific COI distance for members of the genus Placobdella is 1.50 ± 1.80% (de Carle et al., 2017); and the COI distances ranged between 0% and 1.52% for Acanthobdella peledina as seen in de Carle et al. (2022). ...
In spite of their important roles in many ecosystems, data regarding population structure and biogeographic patterns of leeches are scarce. To begin to address this knowledge gap, we herein perform a phylogeographic analysis of the North American medicinal leech, Macrobdella decora (Say, 1824). A total of 224 M. decora specimens were collected from 35 localities across large swaths of USA and Canada and covering most of the known range of the species. Using four loci (mitochondrial cytochrome c oxidase subunit I [COI] and NADH dehydrogenase I [ND1], as well as nuclear 18S rRNA [18S] and 28S rRNA [28S]), we construct phylogenetic trees using several optimality criteria and superimpose geographic patterns onto the trees in order to tease out any potential structure among the populations. Rather surprisingly, given the large geographic range of the species and abundance of potential geographic barriers to gene flow, the analyses showed a conspicuous lack of structure among the different populations of M. decora . However, an AMOVA did show statistically significant differences between the genetic variation within populations and between populations (COI: FST = 0.65412, p < .00001; ND1: FST = 0.69245, p < .00001), which was largely driven by only 6 out of the 35 populations, and indicated a potential barrier for dispersal across the Appalachian Mountains. Finally, a Mantel test showed a weak, but significant, correlation between geographic distance and genetic distance (COI: r = 0.209, p = .027; ND1: r = 0.1289, p = .030); however, this correlation was primarily driven by a single locality. The overall weak structure suggests that M. decora is panmictic throughout its range, and we discuss this in light of previous population level studies in both bloodfeeding and non‐bloodfeeding species, concluding that the lack of structure in M. decora might be due to its high capacity for dispersal via hosts.
... Here, we found a similar case in which the blood-feeding Maiabdella batracophila is found nested within the liquidosomatophagous genus Helobdella. The genus Helobdella comprises more than 80 species mainly distributed in South America and it has been consistently recovered as the sister group of Haementeria, and then together, sister to Placobdella , de Carle et al. 2017). The optimization of the feeding preferences on the tree is the result of a previous study by . ...
Parental care is uncommon within Annelida. However, a remarkable exception is in the leech family Glossiphoniidae in which all its members keep their offspring attached to the ventral surface for varying lengths of time. Considered as the culmination in the evolution of parental care in leeches, two glossiphoniids, Marsupiobdella africana from South Africa and Maiabdella batracophila from South America, have a brood pouch or marsupium in which their offspring are protected. Interestingly, the structure is located in exactly the same region of the body in both species. Based on phylogenetic analysis of Glossiphoniidae, including use of nuclear and mitochondrial DNA sequences, we investigated for the first time the phylogenetic position of the two kangaroo leeches. Our results indicate that marsupium-bearing leeches belong to separate lineages, with each species more closely related to other leeches from their respective continent. We infer that the marsupium evolved independently from ancestors that maintained their eggs inside thin-walled, flexible, transparent cocoons attached directly to the ventral surface of the parents. Together with the evolution of a marsupium, both species have also ceased to produce cocoons, which are normally a characteristic of Glossiphoniidae.
... Worth mentioning, however, is that recent investigations into wide-spread leech taxa have frequently suggested that cryptic diversity is prevalent (de Carle et al. 2017;Saglam et al. 2018;Iwama et al. 2019;Anderson et al. 2020). Although some species of Theromyzon are recorded from localities throughout the Northern Hemisphere (e.g. ...
Theromyzon Philippi, 1867 is a genus of sanguivorous, freshwater leeches in the family Glossiphoniidae. The genus is broadly distributed across the globe, possibly due to the frequent feeding in the nasopharyngeal cavities of migratory waterfowl that may allow for long distance dispersal. The genus has a history of taxonomic confusion resulting from mischaracterisations of key morphological features of type specimens that have produced several re-descriptions and synonyms. Here, we bring partial order to this confusion through robust morphological investigations of newly collected North American (and a single South American) specimens, representing most of the known species diversity from this continent. We also produce the first species-level phylogeny for Theromyzon and attempt to understand species boundaries regarding both morphology and genetics. Our results demonstrate that there are at least five species of Theromyzon present in North America (T. bifarium, T. tessulatum, T. rude, T. trizonare, and a clade that needs further investigation), and a hitherto undescribed taxon that does not conform to any previously published description, and represents a unique lineage in the phylogeny; we describe this new species under the name Theromyzon tigris sp. nov. This study sheds light on the discriminatory power of select morphological characters and the distribution of phenotypes within the genus. We also provide a comprehensive classification framework for the known species within the genus designed to facilitate identification and minimise future taxonomic confusion.
... So far, the presence of decorsins/ornatins has only been confirmed in representatives of American leech species: M. decora [9], P. ornata [12], L. mexicana, and H. vizottoi (this study). The only exception is the European freshwater turtle leech Placobdella costata (Müller, unpublished data), but the genus Placobdella is of North American origin, and the presence of P. costata in the Palearctic is very likely due to a single dispersal event [45]. ...
Hematophagous leeches express a broad variety of secretory factors in their salivary glands; among them are hirudins, inhibitors of blood coagulation, and decorsins/ornatins, inhibitors of platelet aggregation. Here, we describe the identification and molecular and functional characterization of putative hirudins and decorsins/ornatins in two leech species of American origin, Limnobdella mexicana and Haementeria vizottoi. The leech species represent two orders of leeches, the proboscis-bearing Rhynchobdellida and the non-proboscis-bearing Arhynchobdellida. Members of the hirudin superfamily, such as hirudins or decorsins/ornatins, are described for the first time in the genus Haementeria. Both species expressed very potent inhibitors of platelet aggregation, but only the putative hirudins of L. mexicana displayed high thrombin-inhibitory potency, whereas the putative hirudin of H. vizottoi turned out to be a hirudin-like factor. The results of our study provide new insights into the evolutionary background of the blood-sucking lifestyle in leeches.
... Worth mentioning, however, is that recent investigations into wide-spread leech taxa have frequently suggested that cryptic diversity is prevalent (de Carle et al. 2017;Saglam et al. 2018;Iwama et al. 2019;Anderson et al. 2020). Although some species of Theromyzon are recorded from localities throughout the Northern Hemisphere (e.g. ...
Theromyzon Philippi, 1867 is a genus of sanguivorous, freshwater leeches in the family Glossiphoniidae. The genus is broadly distributed across the globe, possibly due to the frequent feeding in the nasopharyngeal cavities of migratory waterfowl that may allow for long distance dispersal. The genus has a history of taxonomic confusion resulting from mischaracterisations of key morphological features of type specimens that have produced several re-descriptions and synonyms. Here, we bring partial order to this confusion through robust morphological investigations of newly collected North American (and a single South American) specimens, representing most of the known species diversity from this continent. We also produce the first species-level phylogeny for Theromyzon and attempt to understand species boundaries regarding both morphology and genetics. Our results demonstrate that there are at least five species of Theromyzon present in North America (T. bifarium, T. tessulatum, T. rude, T. trizonare, and a clade that needs further investigation), and a hitherto undescribed taxon that does not conform to any previously published description, and represents a unique lineage in the phylogeny; we describe this new species under the name Theromyzon tigris sp. nov. This study sheds light on the discriminatory power of select morphological characters and the distribution of phenotypes within the genus. We also provide a comprehensive classification framework for the known species within the genus designed to facilitate identification and minimise future taxonomic confusion.
... Young leeches have a different body form than adults. It is a flattened "cylinder" or "tape" (Figure 4a and b), which during blood collection changes into a deceptively body-like form ("butt") similar to leeches from the Piscicolidae family of the genus Limnotrachelobdella where it is pronounced trachelosoma and urosoma ( Figure 3) (Bielecki, 1997;Cichocka & Bielecki, 2015;Cichocka et al., 2018). ...
• Four events of Placobdella costata sucking human blood are described.
• Human blood was sucked by both adult and juvenile specimens of P. costata.
• The feeding strategies of juveniles under parental care are presented.
• New data on juvenile specimens' body form are presented.
• Information on the potential role of mammals in dispersion and habitat preferences of leeches P. costata is considered.
... Another method is Bayesian Poisson Tree Processes (bPTP) [132]. It identifies significant changes in the pace of branching events on an input tree, using the number of substitutions between branching events, and it has been used in a few studies [118,131,133]. There are also a set of analyses in the Barcode of Life Database System (BOLD) [134], i.e., Barcode Gap Analysis (BGA) and the Refined Single Linkage (RESL) algorithm, the latter of which is the base of the Barcode Index Number (BIN) system [135]. ...
Methods for species delimitation using molecular data have developed greatly and have become a staple in systematic studies of clitellate worms. Here we give a historical overview of the data and methods used to delimit clitellates from the mid-1970s to today. We also discuss the taxonomical treatment of the cryptic species, including the recommendation that cryptic species, as far as possible, should be described and named. Finally, we discuss the prospects and further development of the field.
... Polymerase chain reactions were prepared using the Illustra PuRe Taq Ready-To-Go PCR Folmer et al. (1994) and Light and Siddall (1999 Table 1). Additionally, COI sequences of specimens collected from Ontario, Canada, were acquired from GenBank for inclusion in molecular comparisons (Siddall and Burreson 1998;Oceguera-Figueroa et al. 2016;de Carle et al. 2017; J.R. deWaard, unpublished GenBank sequence MG421798). Multiple sequence alignments were carried out with the MAFFT multiple sequence alignment plug-in for Geneious Prime (Katoh and Standley 2013), applying default settings. ...
Placobdella parasitica (Say, 1824) is one of the most commonly encountered turtle leeches in North America. Molecular analysis of individuals of P. parasitica representing various populations throughout its range in North America using the cytochrome c oxidase subunit I gene, revealed the presence of nine distinct groups: (1) P. parasitica sensu stricto containing members, including the neotype specimen, occurring broadly throughout the central United States westward from the Mississippi River to the Rocky Mountains and throughout southern Ontario, Canada, and the upper midwestern United States as far east as New York State; (2) West Virginia; (3) Mississippi/Alabama; (4) Northeast, including New York, Massachusetts, and Vermont; (5) New
England, including Rhode Island, Massachusetts, and Connecticut; (6) North Carolina/West Virginia; (7) South Carolina; (8) Tennessee; and (9) Florida. Both neighbor-joining and maximum likelihood analyses recovered an east-west split along the Appalachian Mountains with groups 1–3 clustering together and groups 4–9 clustering separately, with the exception of group 8 (Tennessee) that placed with the eastern groups. Group 1 includes specimens from a broad geographic distribution, yet with relatively low genetic variation, a pattern observed in other glossiphoniid species in North America. The groups with members east of the Appalachian Mountains are more tightly clustered by locality. This leech species is known to parasitize several turtle species, including Chrysemys picta, the painted turtle that originated in the central Gulf Coast region and dispersed northward representing a recolonization after Pleistocene glaciation. The neighbor-joining tree and pairwise distance data could suggest that P. parasitica has a similar phylogeographic pattern and dispersal history with its turtle hosts. In view of the morphological uniformity among the various groups, P. parasitica is provisionally considered to be a widely distributed, molecularly variable species.
... These species inhabit exclusively freshwater bodies of all continents, except for Antarctica. Despite the rising interest in this group and the emergence of studies devoted to molecular evolution and systematics of leeches (e.g., de Carle, Oceguera-Figueroa, Tessler, Siddall, & Kvist, 2017;Moser, Richardson, & Hammond, 2012;Moser, Richardson, Hammond, & Lazo-Wasem, 2011;Oceguera-Figueroa, León-Règagnon, & Siddall, 2011;Perez-Flores, Rueda-Calderon, Kvist, Siddall, & Oceguera-Figueroa, 2016), the biodiversity of this annelid group is still considered understudied, especially in the Asian part of the Palaearctic. ...
The study of biodiversity is a priority task of biological science. The structural unit of biodiversity is a species that has a clear identification in a taxonomic system. Morphological features are traditionally the main criteria for species discrimination in zoological studies. However, the presence of inter‐ and intraspecific polymorphism and phenotypic plasticity makes it difficult to identify species in many groups of invertebrates. To solve this problem, in this research, we analyzed morphological and genetic data in combination to delimit species among the Eastern Siberia Glossiphonia leeches using different approaches. Morphology analysis revealed phenetically distinct groups, suggesting the existence of at least two species in the region, G. verrucata, a rare Palaearctic species, and a potentially new species Glossiphonia sp. Moreover, sequence‐based species delimitation methods congruently supported eight distinct species groups (including two Siberian species) within the available molecular dataset of the Glossiphonia world fauna, using phylogenetic (ML and BI), coalescent (ABGD and GMYC) methods, and pairwise analysis of sequences. The detected p‐distances (modal value of 0.11) between these 8 groups and the level of genetic polymorphism (max. 0.0041) within groups indicate that the groups are 8 independent species according to the DNA barcoding. Our results once again proved the usefulness of molecular systematics. At the same time, we detected several inaccuracies in the leech species identification, as well as many ambiguous sites in sequences uploaded on GenBank, which affects the analysis and impedes progress of DNA barcoding technology.
... Previous studies have failed to take into account the genetic variation across the broad geographic ranges of erpobdellids (which can extend across the majority of North America) and, as a result, the potential cryptic diversity and level of gene flow between populations is not as well understood as it is for other groups of leeches Trontelj and Utevsky, 2012). This is especially important when considering the reported levels of cryptic diversity within Annelida (de Carle et al., 2017;De Wit and Erséus, 2010;Gustafsson et al., 2009;Hovingh, 2004), coupled with evidence of unappreciated diversity within Erpobdellidae; for example, several studies have elevated subspecies to the level of species, in recognition of this "hidden" diversity (Oceguera-Figueroa et al., 2005, 2011. ...
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.
... On the basis of molecular data, however, Siddall et al. (2005) and De Carle et al. (2017) clearly show that P. pediculata is a member of the genus Placobdella. However, the collection locality of the representative specimens of P. pediculata in the publication (GenBank AY047327, MF067120, MF067121) are not known (Light and Siddall 1999;Siddall et al. 2005;De Carle et al. 2017). In the present study, sequence data (Gen-Bank MN043903) of P. pediculata from its type locality (Lake Pepin, Minnesota) are presented for the first time. ...
Placobdella pediculata Hemingway, 1908 was originally described from individuals that were attached to Aplodinotus grunniens (freshwater drum) in Lake Pepin, Minnesota, USA. Apparently, no type material was deposited. The acquisition of contemporary specimens from its type host in the type locality facilitated redescription of P. pediculata. Placobdella pediculata is different from its congeners in that its caudal sucker is extended from the body by a pedicel (peduncle), bears digitate processes near the rim of the caudal sucker, a smooth body surface, and its anal placement (few annuli anteriad of the caudal sucker). Molecular comparison of cytochrome c oxidase subunit I sequence data from P. pediculata revealed differences of 13.8% to 17.4% among congeners.
Placobdella pediculata is a distinct species.
... DNeasyt blood and tissue kit (Qiagen, Inc., Valencia, California) was used for tissue lysis and DNA purification. Partial sequences of the mitochondrial cytochrome c oxidase subunit I (COI) and nicotinamide adenine dinucleotide dehydrogenase subunit I (ND1), as well as the nuclear ribosomal DNA of the small (18S rDNA) and large subunits (28S rDNA) of the new species were obtained with the use of the primers detailed in Phillips et al. (2010) for COI, 18S rDNA, and 28S rDNA, and from de Carle et al. (2017) for ND1, in order to investigate the phylogenetic position of the new species. Additionally, three specimens of M. decora from the Royal Ontario Museum in Toronto, Ontario, Canada (ROMIZ I10058, ROMIZ I10059, ROMIZ I10060) were sequenced and included in the present study. ...
A new species of medicinal leech, Macrobdella mimicus n. sp., is described from specimens collected in Maryland; this is the first description of a North American macrobdellid since 1975. Superficially, the new species resembles the well-known Macrobdella decora, as both species possess 4 accessory pores arranged symmetrically on the ventral surface, yet the new species is distinguished from M. decora in possessing 4-4½ annuli (rather than 3½) between the gonopores and 4 annuli (rather than 5 annuli) between the female gonopore and the first pair of accessory pores. Phylogenetic analyses, based on 2 mitochondrial and 2 nuclear loci for a set of closely related taxa, confirms the placement of the new species within the family Macrobdellidae and places it as the sister taxon to M. decora and M. diplotertia.
... Although few cox1 divergences (p-distance: 0.1-0.9%) were observed among other specimens (including both juveniles and large individuals), the values fall in the range of intraspecific cox1 divergences for other leeches (e.g., de Carle et al. 2017). ...
Torix is a leech genus containing freshwater proboscidate species, and several members of this taxon are ectoparasites specific to amphibians. Torix tukubana inhabits mountain streams in Japan, and only two frog species are known to be hosts. We collected this leech from two other amphibians, Onychodactylus japonicus (Japanese clawed salamander) and Rana ornativentris (montane brown frog), for the first time. This finding suggests that the host specificity of T. tukubana is low. The immature individuals of T. tukubana were also collected and identified based on DNA data. This is the first juvenile record of this species confirmed by its DNA barcode sequences. Several morphological characters known from large individuals and used as diagnostic characteristics in taxonomic keys were not observed in the juveniles, suggesting that these are ontogenetic traits.
... Phylogenetic methods represent a subset of the methods in de Carle et al. (2017) and are briefly detailed here. Combined matrices were aligned using MUSCLE (Edgar, 2004) with defaults. ...
Annelids possessing a posterior sucker and a fixed number of somites - most famously leeches (Hirudinida), but also crayfish worms (Branchiobdellida) and salmonid parasites (Acanthobdellida) - form a clade; however, determining the relationships between these orders has proven challenging. Here, we compile the largest molecular phylogenetic dataset yet analysed for these groups, including new sequences for key taxa. We find robust model-based support for a clade formed by Hirudinida and Acanthobdellida, contrasting the largest prior studies. We determine that conflicting prior studies included contaminant sequences for Acanthobdella peledina. In addition to this broad-scale comparison, the size of our dataset grants us invaluable insight into the internal relationships of leeches and crayfish worms. Of particular importance, a largely marine clade of leeches (Piscicolidae and Ozobranchidae) is recovered as sister to all remaining Hirudinida. This necessitates the dissolution of the paraphyletic suborder Rhynchobdellida into two new suborders (Oceanobdelliformes and Glossiphoniiformes). Likewise, we decompose Arhynchobdellida into its respective suborders: Hirudiniformes, Erpobdelliformes, and the new, monotypic, Americobdelliformes.
Species of Placobdella have been the frequent subject of revisionary and alpha-taxonomy in the past 2 decades. Recent molecular analyses introduced uncertainty about the taxonomic status of several broadly distributed and morphologically variable Placobdella species, including Placobdella picta (Verrill 1872), compounded by incomplete original descriptions reliant upon characters that are no longer unique in comparison to modern congeners. We assessed specimens of P. picta to
identify any distinct phylogenetic entities that align with our morphological observations of the type series and novel topotype specimens. Using mitochondrial COI and ND1 and nuclear 18S rDNA with Maximum Likelihood and Bayesian Inference, we evaluated species boundaries using species delimitation analyses (ABGD, mPTP, bPTP, and GMYC) and molecular phylogenetics. Our analyses revealed 2 species entities equivalent to 2 non-reciprocal monophyletic clades.
Morphological examinations revealed the lectotype was determined to be Placobdella ornata (Verrill 1872), and paralectotypes are other leech species or were poorly preserved and unidentifiable. Due to the problems with the type series, P. picta is now considered a junior synonym of P. ornata. Based on our results, we describe 2 new species: Placobdella unimaculata n. sp. from Connecticut and Placobdella desseri n. sp. from Algonquin Provincial Park, Ontario, Canada.
There is a contemporary trend in many major research institutions to de‐emphasize the importance of natural history education in favor of theoretical, laboratory, or simulation‐based research programs. This may take the form of removing biodiversity and field courses from the curriculum and the sometimes subtle maligning of natural history research as a “lesser” branch of science. Additional threats include massive funding cuts to natural history museums and the maintenance of their collections, the extirpation of taxonomists across disciplines, and a critical under‐appreciation of the role that natural history data (and other forms of observational data, including Indigenous knowledge) play in the scientific process. In this paper, we demonstrate that natural history knowledge is integral to any competitive science program through a comprehensive review of the ways in which they continue to shape modern theory and the public perception of science. We do so by reviewing how natural history research has guided the disciplines of ecology, evolution, and conservation and how natural history data are crucial for effective education programs and public policy. We underscore these insights with contemporary case studies, including: how understanding the dynamics of evolutionary radiation relies on natural history data; methods for extracting novel data from museum specimens; insights provided by multi‐decade natural history programs; and how natural history is the most logical venue for creating an informed and scientifically literate society. We conclude with recommendations aimed at students, university faculty, and administrators for integrating and supporting natural history in their mandates. Fundamentally, we are all interested in understanding the natural world, but we can often fall into the habit of abstracting our research away from its natural contexts and complexities. Doing so risks losing sight of entire vistas of new questions and insights in favor of an over‐emphasis on simulated or overly controlled studies.
Herein, we describe a new species of turtle blood-feeding leech, Placobdella nabeulensis sp. nov. from Palearctic North Africa (Tunisia and Algeria). The new species is described based on detailed morphological analyses using light and scanning electron microscopes. Apart from the detailed morphology of the atrium, morphological features alone do not sufficiently separate the species from congeners due to the absence of distinct diagnostic characters. Therefore, we turned to molecular data to better distinguish this new species from other members of the genus and establish a basis for its genetic separation. Four DNA fragments were successfully amplified, including mitochondrial COI and 12s rDNA, as well as nuclear 28S rDNA and histone H3. We then provided the molecular descriptor of the taxon, based on redundant diagnostic nucleotide combinations in DNA sequence alignment within the Folmer region. Results of the phylogenetic analysis and species delimitation methods (ABGD, ASAP, and bPTP) based on the COI locus support the species rank of the Tunisian-Algerian Placobdella . The new species is most closely related to the European species Placobdella costata (Fr. Müller, 1846) and the present study indicates that Placobdella nabeulensis sp. nov. has likely been confused with the European counterpart in several previous studies.
Leeches in the genus Hirudinaria Whitman, 1886, also known as buffalo leeches, are blood-sucking ectoparasites of vertebrates. Although they are widely distributed in Asia and had been highly abundant in the past, studies on diversity and taxonomy of this genus are still scarce. There is probably a large amount of cryptic diversity yet to be discovered, particularly from mainland Southeast Asia. In this study, we used morphology and DNA barcoding with a COI gene fragment to explored the diversity of Hirudinaria leeches in the southern region of Thailand, where a unique geographic feature could have led to the diversification of freshwater biota. Molecular phylogenetic analyses and species delimitation approaches (ABGD, bPTP, GMYC, and BOLD) revealed the presence of four putative species of Hirudinaria leeches from southern Thailand, including H. bpling, H. thailandica, and two morphologically cryptic lineages of H. manillensis. Compared to other leech genera, genetic distances of Hirudinaria leeches were relatively low (0.11–0.65% within species; 3.72–14.36% between species) and barcoding gaps were very narrow (1.54–2.88%). The species diversity, distribution pattern, and a phenomenon of low genetic divergence of Hirudinaria leeches in southern Thailand could be explained by an ancient seaway, paleo-drainage, and anthropogenic activities.
ABSTRACT
The western North America leech distributions was studied to assess this aquatic fauna diversity in widely dispersed and arid habitats during the last 10 Ka (thousand years). The leech distributions today may also reflect habitats some 10 Ma (million years) ago, with different drainages, climate, and topography, lacking precise geological events for leech barrier crossings, and correlated with other aquatic fauna distributions.
Western North America fish species have been extensively studied to determine aquatic connectivity among these dispersed habitats. The recession of the last continental glacier (10 – 15 Ka) revealed colonization from the southern Mississippi River to the mouth of the Mackenzie River and from the Columbia River and the Mackenzie River to British Columbia coastal drainages. The leeches, as fish, illustrate similar colonization of these same post-glacial habitats. Each regional section discusses the regions geology, the geological effects on aquatic fauna, and the geological effects on the leech distributions.
This study examines the geological processes to understand more ancient distributions as applied to mollusks and fish. The most classic is the Pliocene and Miocene “Fish hook” distribution between Bonneville Basin and Snake River and the western Great Basin and Pacific Coast drainages. Equally old as the “Fish hook”distributions, the southern route includes the western Great Plains, Rio Grande River and northern Mexico through southern Arizona to southern California. The Bonneville Basin, Snake River, and upper Green River was intertwined with aquatic colonization by fish with selective barriers for the fish host with mussel glochidia. The leech distributions are similar to the fish and mollusk distributions, with different sets of selectivity. Within the late Miocene time, the upper Colorado River adjoined the Gulf of California through the Grand Canyon, the upper Green and the lower Green through the Uinta Range, and the Snake adjoined the Columbia River.
With purported leech fossils found in Jurassic Europe and Silurian North America, one can suggest that leeches at one time or another have been on every continent, their distribution is a result of continental drift. Sister taxa have distributions with genetic based clade distributions on Euro-North American (Erpobdellidae) and South-North America (Helobdella) continents, suggesting isolation by continental separations. The present leech populations and distributions are a result of geological and climatic changes, with widespread abundant populations, widespread and restrictive populations, and populations isolates with possible extinctions.
The leech distributions will be discussed within these geological patterns. Leech taxonomy and Nearctic continental distributions are discussed in Section I. Postglacial mobility of the leeches to high elevations (Section II) and into northwest North America (Section III) are discussed. Section IV describes the western United States drainage basins distributions: the United States Pacific Coast and Columbia-Snake Rivers drainages (Section IVA), the Great Basin (Section IVB), the Colorado River basin (Section IVC), and the western Great Plains (Section IVD). Each section will have a discussion of the how geography and geology has affected aquatic fauna distribution. A summary discussion (Section V) of western United States leeches concludes this paper, illustrating the different distributional patterns within a geological frame.
Key Words: Hirudinida, leeches, western North America, paleogeography, drainage basin distributions, aquatic fauna
The leech genus Placobdella harbours 24 currently-recognized species, and only a handful of new species has been described in the past decade. Placobdella akahkway n. sp., a new species of ectoparasitic glossiphoniid leech is herein described from sites in the provinces of Alberta, Saskatchewan, and Manitoba, Canada. Beyond morphological description of the species, a total of 21 specimens (including the five specimens from the type series) were also employed in a phylogenetic analysis using the COI locus. The new species can be distinguished from its congeners by its possession of both compact and diffuse salivary glands. The included specimens of Placobdella akahkway n. sp. form a monophyletic group with high support, and place as the sister taxon to Placobdella kwetlumye.
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), Italy, Germany, Latvia, Montenegro, Bulgaria, Slovenia, Turkey, Azerbaijan, Tunisia, and Algeria, and uses both nuclear (Internal Transcribed Spacer [ITS] region) and mitochondrial (cytochrome c oxidase subunit I [COI]) sequence data in phylogenetic and DNA barcoding frameworks, in order to better understand species-level diversity. Seven independent lineages are present in the trees, five of which show adequate separation at the COI locus to suggest their unique species-level status (COI distances between these clades range from 4.86 to 8.10%). However, the ITS data suggest that speciation is recent or incipient in these clades, and that not enough time has passed for clear separation at this locus. We discuss the evolutionary and taxonomic implications of our findings and speculate on dispersal events that may have contributed to shaping this pattern of geographic distribution.
The only Palearctic representative of the leech genus Placobdella Blanchard, 1893 is P. costata, an ectoparasite of freshwater turtles. To date, no conclusive evidence about the possible presence of coevolutionary patterns between this leech and its turtle hosts is available due to the paucity of DNA sequence data available for P. costata; moreover, comparative host data is also mostly lacking, making any inferences more difficult. The discovery of new populations of the species in northern Italy and Sicily allowed us to generate novel mitochondrial DNA sequences and to compare the topology of the resulting phylogenetic trees with the phylogeny of the turtle hosts occurring in the study area, i.e., Emys orbicularis and E. trinacris. The branching pattern of the phylogenetic tree for P. costata is not congruent with that of its turtle hosts, thus suggesting the lack of coevolutionary or cospeciation phenomena between these taxa. The lack of a coevolutionary pattern might be ascribed to the different dispersal ability of Placobdella costata and Emys spp. and to the host generality of the leech, as confirmed by the occurrence of P. costata on aquatic turtles belonging to the genus Mauremys in areas where Emys spp. are rare or absent.
A single leech cytochrome c oxidase subunit 1 haplotype was found in each study region, and the overall nucleotide diversity was very low throughout the investigated distribution. This apparent lack of a clear phylogeographical pattern was unexpected in the P. costata populations occurring in the circum-Mediterranean areas, where the occurrence of high haplotype and nucleotide diversity is customary for most terrestrial and freshwater species. Based on the available data, we suggest a recent, post-glacial origin of the studied P. costata populations.
A new snail-eating leech, Batracobdelloides bangkhenensis sp. n., was discovered at Kasetsart University, Bangkhen Campus, Bangkok, Thailand. This species is found free living in the benthic zone of ponds; feeds on freshwater snails, including Bithynia siamensis siamensis, Indoplanorbis exustus, Radix rubiginosa, Physella acuta, and Pomacea canaliculata; and uses a shell as a shelter during the parental care period, with a colony of 7-15 juvenile individuals held on the venter inside the shell of host. Batracobdelloides bangkhenensis displays distinct morphological characters, including a rice-shaped body showing transparency, cephalization, two eye pairs merged on somite III, an anterior sucker twice as large as the cephalic region, a central mouth in the anterior sucker, seven light brown transverse rows in the neck region, absent dorsal papillae, rich green pigments on the dorsum, a male gonopore on XIIa2/XIIa3 (27-28), a female gonopore on XIIIa1/XIIIa2 (29-30), and diffuse aggregations of minute, spherical salivary glands in the neck region. Comparisons of the COI and COI-ND1 genes showed a monophyletic clade for Batracobdelloides, and the phylogenetic tree of the COI gene also indicated that B. bangkhenensis is distinct from other species in the genus, with strong support values.
Emys orbicularis (Linnaeus, 1758) was considered as a specific host of Placobdella costata (Fr. Mûller, 1846). However, since
the parasite was recorded from outside the distribution area of its host, some authors suggested a possible relationship with other
hosts. Although two accidental associations were found with another turtle, Mauremys leprosa (Schweigger, 1812), the obtained
data remain insufficient to better understand this discovered host-parasite ecological system. In this context, the present study was
carried out to evaluate the relationship between the Mediterranean pond turtle, M. leprosa, and the freshwater rhynchobdellid
leech, P. costata (Hirudinida: Glossiphoniidae), in aquatic ecosystems of Morocco. During the period from April to June 2018,
we found leeches attached to turtles in five out of 30 populations sampled with a prevalence of infection significantly higher in
adult than that in juvenile turtles. Moreover, the males are the most infested with 51% of the total, followed by females (33.3%)
and juveniles (15.7%). The obtained results indicated that 51 turtles were infested by 139 leeches with a mean intensity of
infestation of 4.17 ± 0.47 leeches/turtle (up to 10 leeches/turtle). It was higher in males than that in females in almost all sites. The
posterior limbs are the most preferred attachment site, and the body condition of turtles was not affected by the intensity of
infestation but it is rather a function of altitude. Our findings proved that M. leprosa-P. costata association ismore than accidental
and that M. leprosa is rather the main host of P. costata in aquatic ecosystems of Morocco.
New information on the occurrence of Helobdella europaea Kutschera, 1987 was obtained. Four individuals assigned to this species were found in a private aquarium in Kharkiv. Helobdella europaea has not been recorded for Ukraine so far. The taxonomic identity of the leeches was confirmed by both a morphological examination and a molecular phylogenetic analysis based on all available cox1 sequences of Helobdella leeches. Helobdella europaea was found to have extremely low genetic diversity, which has been probably caused by the founder effect. The variability of the coloration pattern of H. europaea was documented and discussed in the light of its potential usefulness for identification. The species can invade local freshwater ecosystems as has been already happened elsewhere.
With few exceptions, animal mitochondrial genomes are impressively conserved with regard to gene arrangement (synteny). For certain taxonomic groups, specific genomic “hotspot” regions present high levels of gene rearrangements. This is the case for the region between the mitochondrial genes cox2 and atp8 across Annelida, particularly in members of the blood-feeding leeches of the genus Placobdella, for which duplications and deletions of trnD have been detected. Analyses of the intergenic region between cox2 and atp8 of 21 species of Placobdella broadly collected in Canada, Mexico, Portugal, and the USA revealed numerous instances of trnD duplication restricted to the species of Placobdella, and it can be inferred based on the phylogenetic position of samples with a single trnD copy that the duplicated condition is independently lost on five occasions. In species with the duplicated trnD, great variation in the size of insertions between both copies, in the secondary structure of the trnD products, and in their respective anticodon were found. For each of three species (Placobdella rugosa, P. ringueleti, and P. lamothei), samples collected from different localities exhibit different gene arrangements, revealing an unexpected amount of intraspecific variation. The rate at which rearrangements are occurring in this mitogenome region within Placobdella has no known equivalent in the animal kingdom and we propose it as a hotspot for tRNA genes duplication-deletion. Our findings are partially compatible with a “tandem duplication and random loss” model of evolution, however in species with just one trnD, it is the second copy the one that we inferred is eliminated. This is, to our knowledge, the first study where such changes are mapped in a phylogeny looking to reveal the state of the ancestor of the group.
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.
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 re-examines 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, Norway, Iceland, England, France, Italy, Slovenia, Turkey, Russia, 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.
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.
We present the latest version of the Molecular Evolutionary Genetics Analysis (MEGA) software, which contains many sophisticated
methods and tools for phylogenomics and phylomedicine. In this major upgrade, MEGA has been optimized for use on 64-bit computing
systems for analyzing bigger datasets. Researchers can now explore and analyze tens of thousands of sequences in MEGA. The
new version also provides an advanced wizard for building timetrees and includes a new functionality to automatically predict
gene duplication events in gene family trees. The 64-bit MEGA is made available in two interfaces: graphical and command line.
The graphical user interface (GUI) is a native Microsoft Windows application that can also be used on Mac OSX. The command
line MEGA is available as native applications for Windows, Linux, and Mac OSX. They are intended for use in high-throughput
and scripted analysis. Both versions are available from www.megasoftware.net free of charge.
To date, the only species of leech reported from the American Alligator, Alligator mississippiensis is Placobdella multilineata. Seven specimens of a previously undescribed species of Placobdella were collected from the feet and lower jaw of a single female alligator from the Pascagoula River Wildlife Management Area, George County, Mississippi. The new species was named Placobdella siddalli Richardson & Moser, sp. n., in honor of the contributions of Dr. Mark Siddall to our understanding of the biology of leeches. Placobdella siddalli Richardson & Moser is similar to other papillated members of the genus Placobdella, but differs from Placobdella ali Hughes & Siddall, 2007, Placobdella rugosa (Verrill, 1874), Placobdella multilineata Moore, 1953, and Placobdella papillifera (Verrill, 1872) in coloration, papillation, ventral striping, and in the possession of a relatively large caudal sucker. In addition, molecular comparison of 626 nucleotides of CO-I between the new species and other papillated leeches (P. ali, P. multilineata, Placobdella ornata, P. papillifera, P. rugosa) revealed interspecific differences of 14.0–18.0% (88–113 nucleotides).
Mitochondrial DNA sequences, often in combination with nuclear markers and morphological data, are frequently used to unravel the phylogenetic relationships, population dynamics and biogeographic histories of a plethora of organisms. The information provided by examining complete mitochondrial genomes also enables investigation of other evolutionary events such as gene rearrangements, gene duplication and gene loss. Despite efforts to generate information to represent most of the currently recognized groups, some taxa are underrepresented in mitochondrial genomic databases. One such group is leeches (Annelida: Hirudinea: Clitellata). Herein, we expand our knowledge concerning leech mitochondrial makeup including gene arrangement, gene duplication and the evolution of mitochondrial genomes by adding newly sequenced mitochondrial genomes for three bloodfeeding species: Haementeria officinalis, Placobdella lamothei and Placobdella parasitica. With the inclusion of three new mitochondrial genomes of leeches, a better understanding of evolution for this organelle within the group is emerging. We found that gene order and genomic arrangement in the three new mitochondrial genomes is identical to previously sequenced members of Clitellata. Interestingly, within Placobdella, we recovered a genus-specific duplication of the trnD gene located between cox2 and atp8. We performed phylogenetic analyses using 12 protein-coding genes and expanded our taxon sampling by including GenBank sequences for 39 taxa; the analyses confirm the monophyletic status of Clitellata, yet disagree in several respects with other phylogenetic hypotheses based on morphology and analyses of non-mitochondrial data.
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 despite the fact that Placobdella kwetlumye only retains a single pair of salivary glands, as opposed to the 2 pairs more common in the genus. Results show that 9 different anticoagulant proteins and an additional 5 putative anti-hemostasis proteins are expressed in salivary secretions of the 3 species. In particular, an ortholog of the archetypal, single-copy, anticoagulant hirudin (not previously available as comparative data for rhynchobdellids) is present in at least 2 of 3 species examined, corroborating the notion of a single origin of blood feeding in the ancestral leech.
The freshwater turtles of the genus Emys and some leech species of the
family Glossiphoniidae are the only Palaearctic representatives of primarily
Nearctic taxa, which jointly colonised Eurasia and the Maghreb trough the
Bering Strait during the Miocene. The strict trophic relationships occurring
between the glossiphoniid parasite leech Placobdella costata and its host,
the emydid Emys orbicularis, make them a prime example of host-parasite
cophylogenetic evolution. In the light of the discovery of the Sicilian cryptic
endemic species Emys trinacris, which is the sister species to the
widespread Palaearctic E. orbicularis, the possible cophylogenetic
divergence of the turtle hosts and their leech parasites was investigated. In
spite of the deep divergence scored between the two pond turtle species
and of their allopatric distribution, their leech parasites proved to be
conspecific and indistinguishable based on the implemented molecular
marker. This unexpected decoupling might likely be ascribed to the
different dispersal abilities of the two taxa and\or to the recent, humanmediated
introduction of the leech parasites in Sicily. If this last scenario is
confirmed, the long-term effects of the introduced leech parasite on the
endemic Sicilian pond turtle Emys trinacris should be carefully monitored.
In the frame of this study, representatives of the widely-spread predatory
leech Helobdella stagnalis were observed on E. trinacris. Molecular
analyses of their stomach content allowed to rule out the possibility of the
existence of a trophic relationships between these two taxa, in contrast to
what was previously suspected, and suggests that H. stagnalis specimens
were rather attached to the turtles for non-nutritional reasons.
Specimens of the North American leech, Placobdella ornata (Verrill, 1872) were confirmed from the Donkmeer, a freshwater lake in the province of East Flanders, Belgium, by morphological and molecular analysis. Leech specimens from Belgium were morphologically consistent with the syntype series and description of P. ornata by Verrill (1872). Molecular comparison of the Belgian specimens to specimens of P. ornata from the type locality (New Haven, Connecticut, USA) using the cytochrome c oxidase subunit I (COI) gene revealed a similarity of 99.5%. Placobdella ornata from Belgium is the first documentation of a second species of the genus Placobdella in Europe.
Placobdella sophieae Oceguera-Figueroa et al., 2010 (Hirudinida: Glossiphoniidae) is reported from Oregon, California, and British Columbia for the first time. New hosts reported for P. sophieae include Taricha granulosa (rough-skinned newt), Rana pretiosa (Oregon spotted frog), and Anaxyrus boreas (western toad). Placobdella sophieae exhibits relatively low host specificity and all amphibians occurring in the Pacific Northwest are potential hosts.
Twenty freshwater, one euryhaline, one terrestrial, and six marine leeches are now known to occur in the Carolinas. Haemopis septagon, the second terrestrial leech discovered in North America, and five aquatic taxa-Placobdella nuchalis, P. translucens, Mooreobdella melanostoma, M. tetragon, and Erpobdella punctata coastalis-are previously undescribed. Specimens of Placobdella papillifera and P. multilineata, are examined in detail. Four additional species-Batracobdella phalera, B. picta, Helobdella elongata and Haemopis marmorata-have been found in the Carolinas for the first time.The distribution of leeches in the Carolinas is determined in part by the boundaries of the three physiographic provinces. Batracobdella picta, Oligobdella biannulata, and Haemopis marmorata occur only in the Appalachian Mountains, while Placobdella nuchalis, P. translucens, Glossiphonia swampina, Batracobdella phalera, Helobdella stagnalis, H. elongata and Macrobdella ditetra were found only in the coastal plain. Along with the physiographic determinants of distribution, there are also north-south factors of interest. The coastal plain species Philobdella gracilis, also found in the piedmont plateau, and Macrobdella ditetra are southern species which reach the northern limit of their range in North Carolina. On the other hand, Batracobdella picta, Erpobdella punctata punctata, Haemopis marmorata, and Macrobdella decora are common northern leeches and reach their southern limit in the area. Still other species-Placobdella nuchalis, Glossiphonia swampina, Oligobdella biannulata, Erpobdella punctata coastalis, Mooreobdella melanostoma, M. tetragon, and Haemopis septagon-appear to be highly localized and are currently known only from the Carolinas, extending into southeastern Virginia in the case of H. septagon (Shelley, 1975).Observations of locomotion and other behaviour in the biannulate leech, Oligobdella biannulata, reveal a lack of muscular activity in the mid-body regions, which may partially explain the significance of the biannulate condition. This species has been found to have more than one desmognathine salamander host in nature.A key to the hirudinifauna of the Carolinas, both known and expected, has been formulated.
In contrast to the enormous efforts made to characterize the fauna and flora from Costa Rica, except for a few isolated records, very little is known about leeches (Annelida: Clitellata) in the country. In this paper, we present the redescription of a poorly known terrestrial leech: Cylicobdella costaricae, characterizing its external and internal morphology, the latter, completely unknown to date. In addition, we report the first list of leech species for Costa Rica, including in total 12 taxa with records from the 7 provinces of the country. We provide a brief description of the leech morphology, pictures and localities for each taxon. The information shown here is the result of the study of both, specimens deposited at the Museo de Zoología of the Universidad de Costa Rica and the new collections. Finally, we provide a taxonomic key for species identification.
Oceguera-Figueroa, A., Phillips, A. J., Pacheco-Chaves, B., Reeves, W. K. & Siddall, M. E. (2010). Phylogeny of macrophagous leeches (Hirudinea, Clitellata) based on molecular data and evaluation of the barcoding locus. —Zoologica Scripta, 40, 194–203.
The phylogenetic relationships of macrophagous leech species are studied using two mitochondrial [cytochrome c oxidase subunit I (COI) and 12S rDNA] and two nuclear (28S rDNA and 18S rDNA) markers. The complete dataset analysed in this study included 49 terminals and 5540 aligned characters. Phylogenetic analyses were performed under two optimality criteria: Maximum Parsimony and Maximum Likelihood. The monophyly of the two currently recognized families (i.e. Erpobdellidae and Salifidae) is confirmed and well supported. The phylogenetic position of Gastrostomobdellidea is studied for the first time and found to be sister to family Salifidae nested well within Erpobdelliformes. Previously recognized taxonomic arrangements were evaluated and discarded through successive constraint analyses. Correlation between morphology and phylogeny was notable in Salifidae but not in Erpobdellidae. Variability of COI, the barcoding locus, was examined across species leading to the recognition of the invasive Barbronia weberi in Mexico, Costa Rica, Germany, South Africa and Taiwan.
We present a new open source, extensible and flexible software platform for Bayesian evolutionary analysis called BEAST 2. This software platform is a re-design of the popular BEAST 1 platform to correct structural deficiencies that became evident as the BEAST 1 software evolved. Key among those deficiencies was the lack of post-deployment extensibility. BEAST 2 now has a fully developed package management system that allows third party developers to write additional functionality that can be directly installed to the BEAST 2 analysis platform via a package manager without requiring a new software release of the platform. This package architecture is showcased with a number of recently published new models encompassing birth-death-sampling tree priors, phylodynamics and model averaging for substitution models and site partitioning. A second major improvement is the ability to read/write the entire state of the MCMC chain to/from disk allowing it to be easily shared between multiple instances of the BEAST software. This facilitates checkpointing and better support for multi-processor and high-end computing extensions. Finally, the functionality in new packages can be easily added to the user interface (BEAUti 2) by a simple XML template-based mechanism because BEAST 2 has been re-designed to provide greater integration between the analysis engine and the user interface so that, for example BEAST and BEAUti use exactly the same XML file format.
A written informed consent should be obtained from the patient before hirudotherapy is initiated. The patients should be treated each day of leech therapy with anti-Aeromonas antibiotics. Leeches should be applied on the darker spots of the reattached body parts or flaps. Usually 1-10 leeches are used for each treatment, while at the beginning, the patient might need two or more treatments per day. Leech therapy is used until venous capillary return is established across the wound border by angiogenesis. Usually the treatment with leeches lasts for 2-6 days. Hematologic evaluations should be performed every 4 hrs and the patient has to receive blood transfusions when the hemoglobin level is lower than 8 g/dL. Signs of regional lymphadenitis, slight swelling, and pain of regional lymph nodes on the side of leech application and subfebrile temperature can occur. Contraindications related to hirudotherapy include arterial insufficiency, hemophilia, hemorrhagic diathesis, hematological malignancies, anemia, hypotension, and sepsis. Leech therapy is not recommended in pregnancy and lactation and in patients with an unstable medical status, history of allergy to leeches or severe allergic diathesis, and disposition to keloid scar formation, as well as in those using anticoagulants and immunosuppressants.
Phylogenies are increasingly used in all fields of medical and biological research. Moreover, because of the next generation sequencing revolution, datasets used for conducting phylogenetic analyses grow at an unprecedented pace. RAxML (Randomized Axelerated Maximum Likelihood) is a popular program for phylogenetic analyses of large datasets under maximum likelihood. Since the last RAxML paper in 2006, it has been continuously maintained and extended to accommodate the increasingly growing input datasets and to serve the needs of the user community.
I present some of the most notable new features and extensions of RAxML, such as, a substantial extension of substitution models and supported data types, the introduction of SSE3, AVX, and AVX2 vector intrinsics, techniques for reducing the memory requirements of the code and a plethora of operations for conducting post-analyses on sets of trees. In addition, an up-to-date, 50 page user manual covering all new RAxML options is available.
The code is available under GNU GPL at https://github.com/stamatak/standard-RAxML.
Alexandros.Stamatakis@h-its.org.
Placobdella cryptobranchii (Johnson & Klemm, 1977) was originally described from specimens collected from Ozark Hellbenders (Cryptobranchus alleganiensis bishopi) from the North Fork of the White River in Missouri, U.S.A. Leeches collected during August 2009 to August 2011 from five localities in Missouri (including the type locality) facilitated a redescription and molecular characterization of Placobdella cryptobranchii. Placobdella cryptobranchii has a rusty, reddish-brown dorsum with 2 lateral rows of unpigmented papillae, two unpigmented nuchal bands, unpigmented patches, and pair of four pre-anal papillae. Molecular comparison of CO-I sequence data from Placobdella cryptobranchii revealed a 93-94% similarity to Placobdella ornata and 10-17% difference among other species of Placobdella.
Sequence-based methods to delimit species are central to DNA taxonomy, microbial community surveys, and DNA meta-barcoding studies. Current approaches either rely on simple sequence similarity thresholds (OTU-picking) or on complex and compute-intensive evolutionary models. OTU-picking methods scale well on large data sets, but the results are highly sensitive to the similarity threshold. Coalescent-based species delimitation approaches often rely on Bayesian statistics and MCMC sampling, and can therefore only be applied to small data sets.
We introduce the Poisson Tree Processes (PTP) model to infer putative species boundaries on a given phylogenetic input tree. We also integrate PTP with our Evolutionary Placement Algorithm (EPA-PTP) to count the number of species in phylogenetic placements. We compare our approaches to popular OTU-picking methods and the General Mixed Yule Coalescent (GMYC) model. For de novo species delimitation, the stand-alone PTP model generally outperforms GMYC as well as OTU-picking methods when evolutionary distances between species are small. PTP neither requires an ultrametric input tree, nor a sequence similarity threshold as input. In the open reference species delimitation approach, EPA-PTP yields more accurate results than de novo species delimitation methods. Finally, EPA-PTP scales on large datasets because it relies on the parallel implementations of the EPA and RAxML, thereby allowing to delimit species in high-throughput sequencing data.
The code is freely available at www.exelixis-lab.org/software.html.
Alexandros.Stamatakis@h-its.org SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics on-line.
The phylogenetic relationships of the family Erpobdellidae, a group of non-sanguivorous leeches useful as bioindicators, were investigated with the combined use of morphological characters, mitochondrial cytochrome c oxidase subunit I, mitochondrial 12S rDNA and nuclear 18S rDNA. Analyses of separate data sets and the combined data provide strong support for the contention that generic distinctions in the family are not reflective of phylogeny. The resulting hypothesis indicates the number of pairs of labial eyes as the sole morphological predictor of phylogenetic relationship. The traditionally used degree of annular subdivision is the least consistent character. In the absence of readily definable morphological synapomorphies for the resulting clades, the genera Dina, Mooreobdella, Nephelopsis and Trocheta are formally synonymised under the genus Erpobdella, the type genus of the family.
Mitochondrial inheritance in the Unionidae is complex since mitochondria can be inherited from both parents. An increased rate of recombination could lead to erroneous homology assessments, which could cause problems for phylogenetic reconstruction. For this reason we investigated the possibility of using a nuclear marker, the ribosomal internal transcribed spacer region, for phylogenetic studies in the Unionidae, as a complement and comparison to two of the most widely used mitochondrial genes today. The nuclear ribosomal internal transcribed spacer region (ITS1, 5.8S, ITS2) was sequenced from 72 specimens representing six of the seven species of Unionidae mussels occurring in NW Europe: Unio pictorum, U. tumidus, U. crassus, Anodonta anatina, A. cygnea and Pseudanodonta complanata. Sequences from Margaritifera margaritifera were used as an outgroup. The ITS sequences of all species were found to have low intragenomic and infraspecific variation. Compared to mitochondrial genes (16S and COI) they show an intermediate genetic diversity. Phylogenetic analyses produce tree topologies that are congruent with those resulting from analyses of the mitochondrial sequences. Likewise, an incongruence length difference (ILD) test showed no significant incongruence between data sets, indicating that if recombination has occurred it has not produced any conflicting patterns. Best-resolved and supported trees are obtained when gaps are treated as a fifth character state. A combined analysis of the three gene regions shows that Unio crassus and U. pictorum are more closely related than either to U. tumidus. Pseudanodonta is nested within Anodonta as sister taxon to A. cygnea. Advice on how to best preserve mussel material for DNA studies is provided.
Desserobdella phalera (Graf, 1899) n.comb. is redescribed. This glossiphoniid leech deposits cocoons directly onto the substratum and belongs, therefore, in the subfamily Glossiphoniinae. It has the following generic characters: diffuse salivary glands, two pairs of coalesced eyes, one pair of mycetomes, and subdivision of primary annuli in complete body segments. Desserobdella phalera is distinguished from its congener Desserobdella picta (Verrill, 1872) by having dorsal papillae and lateral metameric and dorsal colour markings. In addition, it has approximately 60 radiating papillae on the rim of the caudal sucker. Desserobdella phalera was found naturally on bowfin (Amia calva L.) and largemouth bass (Micropterus salmoides (Lacepède)) from Ontario, Canada, and on bowfin from Florida, U.S.A. Occurrence on bowfin is a new host record. Desserobdella phalera fed experimentally on nine species of fish.
A new genus, Desserobdella, is described to accommodate the leech Clepsine picta Verrill, 1872 which feeds exclusively on amphibians of northern and central North America. This leech belongs to the subfamily Glossiphoniinae and therefore deposits cocoons directly onto the substrate. The genus Desserobdella n.gen. has the following diagnostic characteristics: (i) two pairs of coalesced eyes; (ii) one pair of diffuse salivary glands; and (iii) a single pair of saccular mycetomes containing prokaryotic endosymbionts. Members of the new genus are distinguished from species of the closely related genus Placobdella Blanchard, 1893 on the basis of their salivary gland structure; Placobdella species have two pairs of compact salivary glands versus the single pair of diffuse salivary glands of the genus Desserobdella n.gen. Congenitors of the new genus are Desserobdella cryptobranchii (Johnson and Klemm, 1977) n.comb., Desserobdella michiganensis (Sawyer, 1972) n.comb., and Desserobdella phalera (Graf, 1899) Jones and Woo, 1990. Desserobdella picta n.gen. et comb. exhibited a stereotypical sequence of feeding behaviours on Rana catesbeiana tadpoles which involved picking at the site of proboscis insertion prior to probing, probing, and finally blood-feeding with the posterior sucker released.
The phylogenetic relationships of Glossiphoniidae, a leech family characterised by its high degree of parental care, were investigated with the combined use of morphological data and three molecular datasets. There was strong support for monophyly of most accepted genera in the group, many of which are consistent with eyespot morphology. The genera Desserobdella Barta & Sawyer, 1990 and Oligobdella Moore, 1918 are suppressed as junior synonyms of Placobdella Blanchard, 1893 and thus recognising each of Placobdella picta (Verrill, 1872) Moore, 1906, Placobdella phalera (Graf, 1899) Moore, 1906, and Placobdella biannulata (Moore, 1900), comb. nov. The species Glossiphonia elegans (Verrill, 1872) Castle, 1900 and Helobdella modesta (Verrill, 1872), comb. nov. are resurrected for the North American counterparts to European non-sanguivorous species. Glossphonia baicalensis (Stschegolew, 1922), comb. nov. is removed from the genus Torix Blanchard 1898 and Alboglossiphonia quadrata (Moore, 1949) Sawyer, 1986 is removed from the genus Hemiclepsis Vejdovsky, 1884. The biogeographic implications of the phylogenetic hypothesis are evaluated in the context of what is already known for vertebrate hosts and Tertiary continental arrangements.
Since its first release in 2001 as mainly a software package for phylogenetic analysis, DAMBE has gained many new functions that may be classified into 6 categories: 1) sequence retrieval, editing, manipulation and conversion among more than 20 standard sequence formats including MEGA, NEXUS, PHYLIP, GenBank and the new NeXML format for interoperabity, 2) motif characterization and discovery functions such as position weight matrix and Gibbs sampler, 3) descriptive genomic analysis tools with improved versions of codon adaptation index, effective number of codons, protein isoelectric point profiling, RNA and protein secondary structure prediction and calculation of minimum folding energy, and genomic skew plots with optimized window size, 4) molecular phylogenetics including sequence alignment, testing substitution saturation, distance-based, maximum parsimony and maximum likelihood methods for tree reconstructions, testing the molecular clock hypothesis with either a phylogeny or with relative-rate tests, dating gene duplication and speciation events, choosing the best-fit substitution models, and estimating rate heterogeneity over sites, 5) phylogeny-based comparative methods for continuous and discrete variables, and 6) graphic functions including secondary structure display, optimized skew plot, hydrophobicity plot as well as many other plots of amino acid properties along a protein sequence, tree display and drawing by dragging nodes to each other, and visual searching of the maximum parsimony tree. DAMBE features a graphic, user-friendly and intuitive interface, and is freely available from http://dambe.bio.uottawa.ca.
We report a major update of the MAFFT multiple sequence alignment program. This version has several new features, including
options for adding unaligned sequences into an existing alignment, adjustment of direction in nucleotide alignment, constrained
alignment and parallel processing, which were implemented after the previous major update. This report shows actual examples
to explain how these features work, alone and in combination. Some examples incorrectly aligned by MAFFT are also shown to
clarify its limitations. We discuss how to avoid misalignments, and our ongoing efforts to overcome such limitations.
AbSTrACT An assessment of the hirudinifauna of Washington State is presented. in total, 11 distinct leech species were found representing two new records for the state and two new species of the genus Placobdella, both described herein. Placobdella kwetlumye, n. sp., and Placobdella sophieae, n. sp., both collected in Squires Lake, Whatcom County, are morphologically similar to P. burresonae and P. pediculata respectively, but exhibit morphological features that readily sepa-rate them from congeners. in addition to the descriptions of the new species, here we provide a brief summary of the morphological traits possessed by each species found.
Emys orbicularis is receding from Europe, mainly due to anthropogenic habitat changes. Its parasite, Placobdella costata, is widely distributed within both the former and the present distribution range of the host. Though closely associated with the mud turtle, it may have other hosts (birds, amphibians, reptiles). Its reproductive period coincides with that of its host's migration to the breeding grounds, thus facilitating dispersal of the parasite. Based on literature data we have analyzed the geographic spread of P. costata and mud turtle to observe the possible overlap of their habitats. Observations on the population of mud turtle and the associated leech species (P. costata) were carried out in eastern Poland – Podlasie Lowland. The studies were conducted in spring and summer in 1986–1993. The leeches were collected from the turtles caught in the water and on land. Observations showed that most leeches were found on turtles inhabiting the lake or moving to a breeding area. The greatest intensity of invasion was observed in June and July and that most leeches were observed in female E. orbicularis characterized by greater length of the carapace and weight, compared with males and juvenile individuals.
Summary: The two main functions of bioinformatics are the organization and analysis of biological data using computational resources. Geneious Basic has been designed to be an easy-to-use and flexible desktop software application framework for the organization and analysis of biological data, with a focus on molecular sequences and related data types. It integrates numerous industry-standard discovery analysis tools, with interactive visualizations to generate publication-ready images. One key contribution to researchers in the life sciences is the Geneious public application programming interface (API) that affords the ability to leverage the existing framework of the Geneious Basic software platform for virtually unlimited extension and customization. The result is an increase in the speed and quality of development of computation tools for the life sciences, due to the functionality and graphical user interface available to the developer through the public API. Geneious Basic represents an ideal platform for the bioinformatics community to leverage existing components and to integrate their own specific requirements for the discovery, analysis and visualization of biological data.Availability and implementation: Binaries and public API freely available for download at http://www.geneious.com/basic, implemented in Java and supported on Linux, Apple OSX and MS Windows. The software is also available from the Bio-Linux package repository at http://nebc.nerc.ac.uk/news/geneiousonbl.Contact:
peter@biomatters.com
Understanding the evolutionary history of living organisms is a central problem in biology. Until recently the ability to infer evolutionary relationships was limited by the amount of DNA sequence data available, but new DNA sequencing technologies have largely removed this limitation. As a result, DNA sequence data are readily available or obtainable for a wide spectrum of organisms, thus creating an unprecedented opportunity to explore evolutionary relationships broadly and deeply across the Tree of Life. Unfortunately, the algorithms used to infer evolutionary relationships are NP-hard, so the dramatic increase in available DNA sequence data has created a commensurate increase in the need for access to powerful computational resources. Local laptop or desktop machines are no longer viable for analysis of the larger data sets available today, and progress in the field relies upon access to large, scalable high-performance computing resources. This paper describes development of the CIPRES Science Gateway, a web portal designed to provide researchers with transparent access to the fastest available community codes for inference of phylogenetic relationships, and implementation of these codes on scalable computational resources. Meeting the needs of the community has included developing infrastructure to provide access, working with the community to improve existing community codes, developing infrastructure to insure the portal is scalable to the entire systematics community, and adopting strategies that make the project sustainable by the community. The CIPRES Science Gateway has allowed more than 1800 unique users to run jobs that required 2.5 million Service Units since its release in December 2009. (A Service Unit is a CPU-hour at unit priority).
The program MRBAYES performs Bayesian inference of
phylogeny using a variant of Markov chain Monte Carlo.
Availability: MRBAYES, including the source code, documentation,
sample data files, and an executable, is available at http://brahms.biology.rochester.edu/software.html.
Contact: johnh{at}brahms.biology.rochester.edu
Although several commensal alphaproteobacteria form close relationships with plant hosts where they aid in (e.g.,) nitrogen fixation and nodulation, only a few inhabit animal hosts. Among these, Reichenowia picta, R. ornata and R. parasitica, are currently the only known mutualistic, alphaproteobacterial endosymbionts to inhabit leeches. These bacteria are harbored in the epithelial cells of the mycetomal structures of their freshwater leech hosts, Placobdella spp., and these structures have no other obvious function than housing bacterial symbionts. However, the function of the bacterial symbionts has remained unclear. Here, we focused both on exploring the genomic makeup of R. parasitica and on performing a robust phylogenetic analysis, based on more data than previous hypotheses, to test its position among related bacteria. We sequenced a combined pool of host and symbiont DNA from 36 pairs of mycetomes and performed an in silico separation of the different DNA pools through subtractive scaffolding. The bacterial contigs were compared to 50 annotated bacterial genomes and the genome of the freshwater leech Helobdella robusta using a BLASTn protocol. Further, amino acid sequences inferred from the contigs were used as queries against the 50 bacterial genomes to establish orthology. A total of 358 orthologous genes were used for the phylogenetic analyses. In part, results suggest that R. parasitica possesses genes coding for proteins related to nitrogen fixation, iron/vitamin B translocation and plasmid survival. Our results also indicate that R. parasitica interacts with its host in part by transmembrane signaling and that several of its genes show orthology across Rhizobiaceae. The phylogenetic analyses support the nesting of R. parasitica within the Rhizobiaceae, as sister to a group containing Agrobacterium and Rhizobium species.
A new species of rhynchobdellid leech, Placobdella ringueleti n. sp., is described based on the examination of 25 specimens. Leeches were collected from body appendages of freshwater turtles (Kinosternon leucostomum, Dermatemys mawii, and Staurotypus triporcatus ) from Chiapas, Mexico. The new species resembles other members of the genus in the ocular morphology, bilobated ovaries, and presence of 1 pair of elongated mycetomes, but it is distinguished from them by the dorsal papillar pattern, by having a continuous longitudinal mid-dorsal stripe along the dorsal surface and papillae on the dorsal surface of the posterior sucker. Placobdella ringueleti represents the third species of the genus in Mexico. The type locality of the new species is part of the “El Ocote” Biosphere Reserve, Chiapas, Mexico.
Antistasin, a 15-kDa salivary protein from the Mexican leech Haementeria officinalis, inhibits both blood coagulation and the metastasis of tumors (Tuszynski, G. P., Gasic, T. B., and Gasic, G. J. (1987) J. Biol. Chem. 262, 9718-9723). Antistasin binds to heparin-agarose, suggesting the protein interacts with sulfated glycoconjugates. The specificity of the interaction between antistasin and heparin was tested by measuring the binding of antistasin to various lipids and by comparing the ability of several charged glycoconjugates to inhibit binding. Of the lipids tested, antistasin binds with high affinity only to sulfatide (Gal(3-SO4)beta 1-1Cer) and does not bind to comparable levels of phospholipids, neutral glycosphingolipids, gangliosides, or cholesterol-3-SO4. The binding of antistasin to sulfatide is inhibited by dextran sulfate, fucoidan, and heparin, with I50 values of 1.5, 9.2, and 16 micrograms/ml, respectively. Comparable levels of chondroitin sulfates A, B, C, keratan sulfate, or hyaluronic acid do not inhibit binding. Comparisons of the amino acid sequences of antistasin and other sulfatide or heparin-binding proteins revealed a region of homology, based around the sequence Cys-Ser-Val-Thr-Cys-Gly-X-Gly-X-X-X-Arg-X-Arg, which may be a sulfated glycoconjugate binding domain. In addition, homologies were found with the alternate complement pathway protein properdin and coat proteins from malaria circumsporozoites and Herpes simplex I.
Placobdella pediculata Hemingway, 1908 was originally described from individuals that were attached to Aplodinotus grunniens (freshwater drum) in Lake Pepin, Minnesota, USA. Apparently, no type material was deposited. The acquisition of contemporary specimens from its type host in the type locality facilitated redescription of P. pediculata. Placobdella pediculata is different from its congeners in that its caudal sucker is extended from the body by a pedicel (peduncle), bears digitate processes near the rim of the caudal sucker, a smooth body surface, and its anal placement (few annuli anteriad of the caudal sucker). Molecular comparison of cytochrome c oxidase subunit I sequence data from P. pediculata revealed differences of 13.8% to 17.4% among congeners.
Placobdella pediculata is a distinct species.
Placobdella lamothei n. sp. is described based on 10 specimens collected in Tonatico, Estado de México, Mexico. The new species resembles other species of the genus in the disposition of eyespots and mycetomes connected to the oesophagus and yet can be distinguished primarily on the basis of body pigmentation pattern. This species represents the second of the genus endemic to Mexico. We also discuss previous records of Placobdella spp. in Mexico and clarify some apparently incorrect determinations.
The identification and taxonomy of Placobdella papillifera (Verrill, 1872) Moore, 1952 have been blurred by its varieties and have been compounded by the apparent lack of type material. Leeches collected from May 2008 through August 2012 from West River, New Haven, New Haven County, Connecticut, the type locality of Clepsine papillifera Verrill, 1872, were consistant with P. papillifera (Verrill, 1872) Moore 1952 as described by Verrill (1872). The acquisition of these specimens from the type locality facilitated redescription of P. papillifera. A neotype is designated to stabilize the concept of P. papillifera. Placobdella papillifera appears to exhibit relatively low host specificity, having been documented on Chrysemys picta, Chelydra serpentina, Sternotherus odoratus and Homo sapiens in this study. Molecular comparison of CO-I sequence data from P. papillifera revealed differences of 15% from Placobdella ornata, 16% from P. rugosa, 18% from P. ali, 17% from P. multilineata and 14% from P. parasitica.
Placobdella hollensis was originally named Clepsine hollensis by Whitman (1892) from specimens collected near Woods Hole, Massachusetts; however, type specimens were neither declared nor deposited. Specimens of P. hollensis were collected from Coonamessett Pond in Barnstable County (47°37′10.25″N, 70°34′20.42″W), Massachusetts, on 4 and 30 May, 2013. Their acquisition from the general locality where Whitman (1892) collected specimens facilitated redescription of P. hollensis. A neotype is designated to stabilize the concept of P. hollensis. Placobdella hollensis is unique among its congeners in its possession of accessory “eyes.” Placobdella hollensis is modestly papillated and has a dorsomedial row and a pair of paralateral rows of unpigmented papillae, two rows of three paramedial pre-anal papillae, and two rows of two paralateral pre-anal papillae. Unlike P. ali, P. multilineata, P. papillifera, and P. parasitica, the ventrum of P. hollensis does not have stripes. Placobdella hollensis does have two thin paramedial dark lines on its ventrum, which are also present in P. papillifera. Molecular comparison of CO-I sequence data from P. hollensis revealed differences of 14% from P. multilineata, 15% from P. picta, 15% to 16% from P. papillifera, 16% from P. translucens, 16% to 17% from P. rugosa, 16% to 17% from P. ornata, 17% from P. montifera, 17% to 18% from P. ali, and 18% from P. biannulata.
Placobdella parasitica (Say, 1824) was originally described from lakes of the “North-western region.” Leeches collected during May and June, 2012, from Minnesota, USA, the type locality of Hirudo parasitica Say, 1824, were consistent with P. parasitica (Say, 1824) Moore, 1901 as described by Say (1824). The acquisition of these specimens from the type locality facilitated redescription of P. parasitica. A neotype is designated to stabilize the concept of P. parasitica. In contrast to the papillated members of the genus Placobdella, P. parasitica does not have a confusing and problematic taxonomic history. Placobdella parasitica is differentiated from its congeners by its smooth dorsal surface (or sensillae present), simple (medial unpigmented yellow line) to elaborate (medial and paramedial marbled patches) pigmentation, and ventrum with 8 to 12 stripes. Molecular comparison of CO-I sequence data from P. parasitica revealed differences of 14% from Placobdella multilineata, 14% from Placobdella papillifera, 15% to 17% from Placobdella rugosa, 15% from Placobdella ornata and 16% to 17% from Placobdella ali.
The postsporogonic development of an adeleid blood parasite, Haemogregarina balli, infecting snapping turtles, Chelydra serpentina, was investigated over a period of 9 mo postfeeding (PF) in the leech Placobdella ornata. At 8 mo PF, asexually dividing stages (meronts) were observed in the circulatory system of a leech. Merozoites were observed by electron microscopy in the proboscis of another leech at 9 mo PF. A laboratory-raised snapping turtle was exposed to the bite of 2 leeches and, after a 12-week prepatency, mature gamonts were observed in the peripheral circulation. A reanalysis of the phylogeny of the Sporozoea, incorporating new data from published sources, clarified the relationship of the genus Haemogregarina with other Apicomplexans. The confirmation of asexual development in leeches for H. balli provides independent support for the relationships in the phylogeny.
The taxonomy of papillated leeches of the genus Placobdella has been extremely problematic, confounded by vague descriptions, redescriptions of species using material across a wide geographic range without reference to type material, and a plethora of dubious taxonomic acts over the course of more than a century. Leeches collected between 2008 and 2011 from the West River, New Haven, New Haven County, Connecticut—the type locality of Placobdella ornata (Verrill, 1872)—were determined to be conspecific with P. ornata (Verrill, 1872) on the basis of comparison to the syntype series. The acquisition of these specimens from the type locality facilitated re-description of P. ornata. The striking similarity of P. ornata to P. phalera (Graf, 1899) prompted comparison of the two species. Specimens conforming to the description of P. phalera were collected from Shivericks Pond in Falmouth, Massachusetts, the type locality of P. phalera. Comparison of these specimens to Graf s (1899) original description of P. phalera (particularly figure 116, which was subsequently designated as the lectotype of P. phalera in the absence of type material) revealed the specimens to be conspecific with the concept of P. phalera articulated by Graf ( 1899) and with P. ornata (Verrill, 1872). Thus, P. phalera is considered a junior synonym of P. ornata. Molecular comparison of the specimens collected from the type localities of P. ornata and P. phalera using the highly conserved gene cytochrome c oxidase subunit I revealed similarities in base sequences of greater than 99.5%, substantiating the synonymy of the species.
The identification and taxonomy of papillated leeches of the genus Placobdella have been extremely problematic, confounded by vague descriptions, redescriptions of species without reference to type material or type locality, and more than a century of numerous dubious taxonomic acts. Leeches collected on August 5 and 6, 2011, from North Dakota, USA, the type locality of Clepsine ornata rugosa Verrill, 1874, were determined to be conspecific with Placobdella rugosa (Verrill, 1874) Moore, 1901 on the basis of comparison to the syntype series. The acquisition of these specimens from the type locality facilitated resurrection and redescription of P. rugosa. Placobdella rugosa is similar to P. ali but differs in salivary gland morphology and ventral pigmentation. Molecular comparison of CO-I revealed differences of 16.1% between P. rugosa and P. ali.
