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Tribal phylogenetic relationships within Vespertilioninae (Chiroptera: Vespertilionidae) based on mitochondrial and nuclear sequence data

Zachary P. Roehrs, Justin B. Lack, Ronald A. Van Den Bussche
DOI: http://dx.doi.org/10.1644/09-MAMM-A-325.1 1073-1092 First published online: 15 October 2010


A paucity of useful characters, morphological convergence, and potential rapid radiation has hindered systematists in elucidating evolutionary relationships within Vespertilioninae. In this study >8,500 base pairs of digenomic DNA for 111 taxa were sequenced and analyzed using maximum-parsimony and Bayesian phylogenetic methods to construct trees and reexamine hypotheses of supergeneric evolutionary relationships in Vespertilioninae. Results of these analyses validate monophyly of Vespertilioninae with the exclusion of Myotis and support recognition of 6 tribes: Antrozoini, Lasiurini, Scotophilini, Vespertilionini, and 2 new unnamed tribal clades, the perimyotine group and the hypsugine group. Tree topologies indicate a Nycticeiini–Eptesicini group, but this clade is not supported. The heuristically pleasing tribe Plecotini also is unresolved in these analyses. These results provided further support and greater resolution for previously proposed hypotheses of Vespertilioninae evolution based on mitochondrial DNA, and although deep branching patterns are not fully resolved, these data increase our understanding of the evolution of this ecologically important and diverse group of bats.

Key words
  • Antrozoini
  • digenomic sequence data
  • Eptesicini–Nycticeiini
  • Lasiurini
  • mitochondrial DNA
  • nuclear DNA
  • Scotophilini
  • systematics
  • Vespertilioninae
  • Vespertilionini

Understanding the evolutionary relationships within the subfamily Vespertilioninae (Mammalia: Chiroptera: Vespertilionidae) has been difficult for systematists because of the evolutionary and ecological success (in terms of species richness and biogeography) and constrained circumscription (in terms of morphological diversification) of this subfamily. Approximately 240 species have been described and placed in this subfamily (Simmons 2005). However, few useful synapomorphic morphologic character states exist that unambiguously define taxa belonging to Vespertilioninae (Hill and Harrison 1987; Koopman 1994; Miller 1907; Simmons 1998; Tate 1942; Wallin 1969). The significance any 1 of these characters receives in relation to the divergence of these taxa is in debate (lumper or splitter—Ellerman and Morrison-Scott 1951; Hill and Harrison 1987; Simpson 1945; Zima and Horáček 1985). Furthermore, it seems likely that parallel or convergent evolution of some of these characters (e.g., number of incisors, cusp pattern, and I2 size; number of anterior upper premolars; and pelage color) has led to classifications incongruent with evolutionary history within Vespertilioninae (Ärnbäck-Christie-Linde 1909; Ellerman and Morrison-Scott 1951; Heller and Volleth 1984; Hill 1966; Hill and Harrison 1987; Hill and Top73x00E1;l 1973; Horáček and Zima 1978; Koopman 1975; Rosevear 1962; Tate 1942; Volleth and Heller 1994b; Zima and Horáček 1985). These limitations have led to ambiguity in our understanding of evolutionary relationships within this diverse subfamily, which has hindered development of a generally agreed-upon classification.

Of particular interest in this study are supergeneric relationships of bats within Vespertilioninae. Although Miller (1907) set the foundation for our modern classification of these bats (without downplaying work of his predecessors— Dobson 1875, 1878; Gill 1885; Gray 1821, 1866) and drew attention to similarities between genera (e.g., “Eptesicus-like” or “Pipistrellus-like”), he did not formally elucidate evolutionary relationships or provide taxonomic names to any rank above genus within Vespertilioninae. It was not until the work of Tate (1942) that a testable hypothesis for classification of bats within Vespertilioninae was described (Table 1). This is in stark contrast to Simpson (1945), who rejected a tribal classification rank for Vespertilioninae. and synonomized many genera. Most authors since these classic works have followed the classification of Tate (1942), using a tribal rank, but followed Simpson (1945) in identifying fewer genera for their classifications (Koopman 1984, 1994; Koopman and Jones 1970; McKenna and Bell 1997). Although more recent studies based on bacular morphology and cytogenetics have provided insight into evolutionary relationships of Vespertilioninae, many relationships remain unresolved, many taxa remain unstudied, and some of these findings contradict previous hypotheses about evolution of Vespertilioninae (Ao et al. 2006; Hill and Harrison 1987; Volleth and Heller 1994a, 1994b; Volleth et al. 2001, 2006). Excluding Myotini, which has been elevated to its own subfamily (Hoofer and Van Den Bussche 2003; Lack et al. 2010; Stadelmann et al. 2004), historically 9 tribes have been proposed in various classifications to organize the systematics of Vespertilioninae, including Antrozoini (Miller 1897), Eptesicini (Volleth and Heller 1994b), Lasiurini (Tate 1942), Nycticeiini (Gervais 1855), Nyctophilini (Peters 1865), Pipistrellini (Tate 1942), Plecotini (Gray 1866), Scotophilini (Hill and Harrison 1987), and Vespertilionini (Gray 1821). The validity of these tribes has been accepted or discredited to various degrees, and their exact rank, position, circumscription, and composition are subjects of continuing debate (Hill and Harrison 1987; Hoofer and Van Den Bussche 2003; Table 1).

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Table 1

Historic classifications of Vespertilioninae. Taxa marked with an asterisk (*) are currently recognized taxa that would have been synonyms in authors’ taxonomic system. A dagger (†) denotes these taxa as incertae sedis.

Tate (1942)Simpson (1945)Hill and Harrison (1987)KoopmanaVollethbMcKenna and Bell (1997)Hoofer and Van Den Bussche (2003)Simmons (2005)
MyotisMyotis*Cistugo* Cistugo*CistugoOtonycteris†Myotis
Chalinolobus* GlauconycterisEudiscopusPipistrellusNyctalusPhiletor*NyctalusNyctalus
Hesperoptenus*IaLaephotis*Hypsugo*Perimyotis* Falsistrellus*Parastrellus
*Perimyotis*ParastrellusScotorepensUnnamed genusLaephotis
*Vespadelus* VespadelusTylonycterisVespadelusMimetillus
  • a This arrangement can be found in Koopman and Jones (1970), Koopman (1984), and Koopman (1994), but the latter provides the most information and is the basis for depicted classification.

  • b This is a combination of results taken from Heller and Volleth (1984), Kearney et al. (2002), Volleth and Tidemann (1991), Volleth and Heller (1994b), and Volleth et al. (2001), with most-recent papers taking precedence.

With the development of modern techniques in polymerase chain reaction, DNA sequencing, and molecular data analysis, researchers are reevaluating evolutionary relationships of bats in this family, bringing to bear the advantages of the enormous number of characters provided by molecular data (Bickham et al. 2004; Gu et al. 2008; Hoofer and Van Den Bussche 2001; Hoofer et al. 2003, 2006; Lack et al. 2010; Miller-Butterworth et al. 2007; Ruedi and Mayer 2001; Stadelmann et al. 2004, 2007). Mayer and von Helversen (2001) and Mayer et al. (2007) sequenced the ND1 mitochondrial coding gene of western Palearctic vespertilionids, Kawai et al. (2002) examined ND1, the nuclear exon vWF, and short interspersed elements (SINEs) of mainly eastern Palearctic bats, and Hoofer and Van Den Bussche (2003) used 2.6 kilobases of the ribosomal mitochondrial genome from 120 globally sampled vespertilionids to evaluate evolutionary relationships within Vespertilionidae. However, as in previous studies, results of these studies provided insufficient resolution to explicate the deep branching patterns within Vespertilioninae.

Potentially convergent or uninformative characters, rapid diversification of vespertilionids leading to deep branching patterns, and subsequent lack of genetic resolution have left our understanding of evolutionary relationships relatively ambiguous for the last 100 years. The purpose of this study was to elucidate polygenetic relationships within Vespertilioninae using both coding and noncoding regions of nuclear and mitochondrial genomes with the focus on resolving tribal composition and intertribal systematic relationships. Furthermore, these digenomic data were used to assess the validity of previously proposed tribes (Antrozoini, Eptesicini, Lasiurini, Nycticeiini, Nyctophilini, Pipistrellini, Plecotini, Scotophilini, and Vespertilionini) within Vespertilioninae. Production of a resolved and supported phylogeny for Vespertilioninae would enhance our understanding of the evolution of one of the most taxonomically diverse, geographically widespread, and ecologically successful groups of mammals and would increase our abilities to answer important ecological, evolutionary, and biogeographical questions.

Materials and Methods

Taxonomic sampling.—Included in this study are samples from 31 (70%) of the 44 currently recognized genera, 77 (32%) of the 241 species within Vespertilioninae, and 21 species of Myotinae (Simmons 2005; see Appendix I for list of taxa, general collecting locality, and voucher information). Taxa were included based on availability with the intent of representing distributional and ecological diversities of its members. Representatives of the subfamilies Kerivoulinae and Murininae were included as out-groups to polarize character-state transformations. Tissue samples were provided by several natural history collections, and most tissues are represented by voucher specimens (Ruedas et al. 2000) in the following institutions: Abilene Christian University, American Museum of Natural History, Carnegie Museum of Natural History, Colecci0ón Mamíferos Lilto, Universidad Nacional de Tucuman, Durban Natural Science Museum, Field Museum of Natural History, Indiana State University Vertebrate Collection, Muséum d'Histoire Naturelle de Genéve, Museum of Southwestern Biology at the University of New Mexico, Museum of Texas Tech University, Natural History Museum of Bern, Oklahoma State University Collection of Vertebrates, Royal Ontario Museum, Sam Noble Oklahoma Museum of Natural History, Texas Cooperative Wildlife Collection at Texas A&M University, Universidad Autónoma Metropolitana–Iztapalapa, Universidad Nacional Autónoma de México, and University of Lausanne, Institut de Zoologie et d'Ecdlogie Animale (Appendix I). The acquisition of tissues samples and voucher specimens by the authors were conducted following the guidelines of the American Society of Mammalogists (Gannon et al. 2007). However, the majority of tissue samples came from preexisting collections housed in previously mentioned collections. Identifications of many specimens were verified by Steven R. Hoofer (Hoofer and Van Den Bussche 2003) and Manuel Ruedi (Muséum d'Histoire Naturelle de Genève, pers. comm.); otherwise, we relied on the identifications of the above collections.

Extraction, amplification, and sequencing.—Whole genomic DNA was isolated from skeletal muscle or organ tissue samples from 111 individuals following procedures of Longmire et al. (1997) or the DNeasy Tissue Kit (Qiagen, Austen, Texas). Previously designed primers were used to target 3 exons, apolipoprotein B (APOB), dentin matrix acidic phosphoprotein I (DMP1), and recombination activating gene II (RAG2), and intron regions of 3 other genes, protein kinase C, iota (PRKCI), signal transducer and activator of transcription 5A (STAT5A), and thyrotropin (THY—Lack et al. 2010). These nuclear markers were chosen because they have resolved deep branching patterns in Chiroptera and other mammalian taxa (Amrine-Madsen et al. 2003; Baker et al. 2000; Eick et al. 2005; Matthee and Davis 2001; Matthee et al. 2001, 2004, 2007; Van Den Bussche et al. 2003). Polymerase chain reaction amplifications were conducted using 200–500 ng of DNA, 1 unit of Taq polymerase, 0.14 mM of each deoxynucleoside triphosphate, 5 µl of 10× buffer, 3.5 mM of MgCl2, 0.8 mg/ml of bovine serum albumin, and 0.15 µl of each primer in a 30-µl total volume reaction. The general polymerase chain reaction thermal profile used for these reactions began with an initial 3-min denaturing of 94–95°C, followed by 35–40 cycles of 94–95°C for 30 s, 40–62°C for 1.5 min, and 72°C for 1 min (Lack et al. 2010). Amplification ended with a final elongation at 72° C for 10 min to ensure all reactions were completed. Polymerase chain reaction products were filtered to remove excess reactants using Wizard SV Gel and PCR Clean-Up System (Promega, Madison, Wisconsin). Sequencing reactions were conducted in both directions using Big Dye chain terminator and a 3130 Genetic Analyzer (Applied Biosystems, Inc., Foster City, California).

In addition to the sequence data generated for this study, we included previously published mitochondrial ribosomal DNA (mtDNA; comprising 12S rRNA, tRNAVal, and 16S rRNA) for 100 individuals, DMP1 for 3 individuals, and RAG2 for 6 individuals (Hoofer et al. 2003; Hoofer and Van Den Bussche 2001, 2003; Lack et al. 2010; Van Den Bussche and Hoofer 2000, 2001; Van Den Bussche et al. 2003). Amplifications and sequencing of the mtDNA gene regions were conducted for 11 additional individuals using primers and methods outlined in Van Den Bussche and Hoofer (2000). Sequence data for the nuclear DNA (nDNA) also were supplemented for 4 individuals with sequences of PRKCI, STAT5A, and THY published by Eick et al. (2005) and deposited in GenBank (http://www.ncbi.nlm.nih.gov/).

Phylogenetic analysis.73x2014;Forward and reverse sequences for each gene region were assembled using the program Geneious 4.5.4 (Biomatters Ltd., Auckland, New Zealand). Alignment of sequence contigs was performed using ClustalW 1.83.XP (Thompson et al. 1994) through Geneious 4.5.4 and then assessed and manually optimized using MacClade 4.05 (Maddison and Maddison 2002). Regions appearing to violate the assumption of positional homology were recognized and excluded from phylogenetic analyses based on the procedures of Lutzoni et al. (2000). The mtDNA and each of the nDNA gene regions were analyzed independently using maximum parsimony in PAUP* version 4.0b10 (Swofford 2002) and Bayesian phylogenetic methods in MRBAYES version 3.1.2 (Huelsenbeck and Ronquist 2001). An unweighted nucleotide substitution model, a heuristic search with 25 random additions of taxa, a tree-bisection-reconnection branch exchanging algorithm, and 1,000 bootstrap replicates were parameters used in maximum-parsimony analysis. Bayesian analysis employed a 4-chain (3 hot and 1 cold) parallel Metropolis-coupled Markov chain Monte Carlo, which was run for 1 × 107 generations, with sampling every 1,000 generations, and a temperature parameter of T = 0.02. Data were partitioned by codon for exons (APOB, DMP1, and RAG2), by marker for introns (PRKCI, STAT5A, and THY), and as a separate partition for mtDNA data. ModelTest version 3.06 (Posada and Crandall 1998) was used to identify the most appropriate nucleotide substitution model for Bayesian analysis resulting in the implementation of a General Time Reversible model with gamma-distributed rate variation among sites and inclusion of a proportion of invariable sites (GTR + Γ + I—Rodríguez et al. 1990). Model parameters were not defined a priori in Bayesian analysis but were treated as unknown variables with uniform priors. A random unconstrained starting tree with uniform priors was used for Bayesian analysis, and the burn-in values were determined by plotting likelihood scores per 1,000 generations and locating the region at which model parameters and tree scores reach stationarity. Nodes in the resulting trees were considered supported if they had ≥70% maximum-parsimony bootstrap support and ≥0.95 Bayesian posterior probabilities.

To examine incongruencies between gene regions and evaluate appropriateness of combining gene regions, each was analyzed independently, and resulting gene trees were compared. We used a 90% concordance criterion to evaluate appropriateness of concatenation where resulting gene trees must be in concordance at a minimum of 90% of nodes before concatenation of these data for further analysis (De Queiroz 1993). Based on results of these concordance tests (described in “Results”), data were concatenated into 3 data sets: mtDNA, nDNA, and combined (mtDNA + nDNA) data sets for maximum-parsimony and Bayesian phylogenetic analysis. The program TREEPUZZLE 5.2 (Schmidt et al. 2002) was used to conduct likelihood-mapping (Strimmer and von Haeseler 1997) with a GTR + Г + I model of nucleotide substitution to examine the phylogenetic potential of each independent gene region and combined data partitions.


Independent gene regions and concordance.—All sequences generated in this study have been submitted to GenBank (see Appendix I for GenBank accession numbers). The nuclear gene regions analyzed were relatively short, 280–1,240 base pairs (bp; Table 2), and independently contained few phylo-genetically informative positions (129–415 bp). Likelihood-mapping demonstrated that for these independent nDNA gene regions the number of positions analyzed is correlated positively to quartet resolution (Strimmer and von Haeseler 1997; Fig. 1). The mtDNA, nDNA, and combined data sets showed the same trend, but the slope was less positive. However, while accounting for this general size-resolvability relationship, the exons (especially DMP1 and RAG2) outperformed introns in their ability to resolve quartets, possibly indicating greater systematic error caused by signal saturation due to a potentially higher substitution rate in these introns. Analysis of each of nDNA gene regions independently and comparison of these gene trees (not shown) provided a high level of topology concordance (>90% concordance in supported topology). The only repeatedly supported incongruencies were in the variable position of Baeodon and in a few Vespertilioninae taxa embedded in the Myotis clades for APOB and PRKCI. The combined data set was analyzed twice, once excluding APOB and once excluding PRKCI, resulting in no effect to topology and relatively few clades becoming unsupported (posterior probabilities ≥ 0.95; e.g., support for inclusion of Baeodon in Antrozoini). Therefore, the independent nDNA gene regions were concatenated for further analysis.

Fig. 1

Scatter plot of the percentage of resolved quartets from likelihood-mapping by number of analyzed positions for each individual nuclear DNA (nDNA) gene region and the mitochondrial DNA, nDNA, and combined data sets. See Table 2 for data and Strimmer and von Haeseler (1997) for discussion of likelihood-mapping.

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Table 2

Characteristics of individual gene regions and combined data partitions. Aligned positions constitute the full aligned length including indel regions. Excluded positions are those that potentially violate positional homogeneity. Analyzed positions are aligned minus excluded positions. Percent resolved and percent unresolved refers to the percent of quartets resolved and unresolved in likelihood-mapping analysis (Strimmer and von Haeseler 1997). APOB = apolipoprotein B; DMP1 = dentin matrix acidic phosphoprotein I; RAG2 = recombination activating gene II; PRKCI = protein kinase C, iota; STAT5A = signal transducer and activator of transcription 5 A; THY = thyrotropin; mtDNA = mitochondrial DNA; and nDNA = nuclear DNA.

Markertaxapositionspositionspositionspositionsinformative positionsresolvedaunresolvedb
  • a Percent resolved = percent occupancy of P vectors in attraction basins for fully resolved topologies [A1 + A2 + A3].

  • b Percent unresolved = percent occupancy of P vectors in attraction basins for unresolved topologies [A 13 + A12 + A23 + A*].

Mitochondrial DNA sequences.—New ribosomal mtDNA sequence data were generated for 11 individuals of 9 taxa: 3 individuals of Eptesicus macrotus, and 1 individual each of Arielulus aureocollaris, E. magellanicus, E. serotinus, Lasiurus intermedius, Pipistrellus hesperidus, P. paterculus, P. pipistrellus, and Tylonycteris robustula, which supplemented 100 mtDNA sequences previously generated (Hoofer and Van Den Bussche 2003; Lack et al. 2010). These 111 sequences were aligned to provide 2,940 aligned positions, of which 968 were excluded prior to analysis for potential violation of positional homology (Table 2). Of the remaining 1,972 positions, 861 were variable and 671 were phylogenetically informative. Maximum-parsimony analysis resulted in 718 parsimonious trees of 5,713 steps, with 43 supported clades (Fig. 2), a consistency index excluding uninformative characters (CI) of 0.1978, and a retention index (RI) of 0.5727. Bayesian analysis had a burn-in value of 882 generations and resulted in 58 supported clades (Fig. 2).

Fig. 2

Phylogram from Bayesian analysis of the 12S rRNA, tRNAVal, and 16S rRNA mitochondrial DNA genes, with supported phylogenetic relationships from both maximum-parsimony and Bayesian analysis depicted. Circles indicate clades supported by maximum parsimony (≥70% bootstrap values), whereas asterisks indicate clades supported by Bayesian analysis (≥0.95 posterior probability). Taxonomic abbreviations include: A. = Antrozous, Ch. = Chalinolobus, C. = Corynorhinus, E. = Eptesicus, G. = Glauconycteris, H. = Hypsugo, L. = Lasiurus, N. = Neoromicia, Ny. = Nyctalus, P. = Pipistrellus, PI. = Plecotus, R. = Rhogeessa, S. = Scotophilus, T. = Tylonycteris, and V. =Vespadelus. For species with more than 1 representative, general locality information is provided in parentheses following the species name. Locality abbreviations follow United States postal codes or include: Arg. = Argentina; Ca. = Catamarca Province, Argentina; Eu. = Europe; Ne. = Neuquén Province, Argentina; Sa. = Salta Province, Argentina; and Tu. = Tunisia. Scale is in number of substitutions per site.

Nuclear DNA sequences.—Sequence data for the concatenated nDNA partition were generated for 111 taxa, of which 18 are missing ≥1 gene region (13–23% of nDNA data set). Vespadelus vulturnus was missing the most sequence data because we were unable to amplify or sequence the APOB or DMP1 gene regions successfully for this taxon, and Baeodon alleni was missing the least with the last 470 bp of RAG2 missing. In most cases missing data were from the STAT5A gene region, which proved to be the most difficult to amplify and was not generated for the following 16 taxa: Eptesicus magellanicus, Glauconycteris beatrix, G. egeria, Hypsugo cadornae, H. savii, Nyctalus leisleri, N. noctula, Pipistrellus coromandra, P. hesperidus, P. javanicus, P. nathusii, P. tenuis, Scotoecus hirundo, Tylonycteris pachypus, T. robustula, and Vespertilio murinus. No changes in clade support or topological resolution were observed when the data set was analyzed excluding STAT5A (data not shown).

Concatenated alignment of the nDNA gene regions provided 5,570 aligned positions (Table 2). With the exclusion of 766 positions for possible violations of positional homology prior to analysis, the remaining 4,804 positions included 2,232 variable positions and 1,654 phylogenetically informative positions. The maximum-parsimony analysis resulted in 24 most-parsimonious trees of 7,941 steps, 52 supported clades, and a CI of 0,4574 and an RI of 0.7152, excluding uninformative characters (Fig. 3). The majority of differences between the 24 most-parsimonious trees involved the relationship between the clades comprising the Antrozoini, Plecotini, Lasiurini, Scotophilini, New World pipistrelles, and a clade including the remaining Pipistrellus-like bats. Also variable was the position of Arielulus and Lasionycteris within Nycticeiini (sensu Hoofer and Van Den Bussche 2003, excluding Nycticeius) and the intrarelationships of some member of the Pipistrellus. Finally, some variation in topologies was attributed to variability within the genus Scotophilus. A burn-in value of 1,001 generations was used for the Bayesian analysis, which resulted in a tree with 57 supported clades (Fig. 3).

Fig. 3

Phylogram from Bayesian analysis of the concatenated nuclear DNA gene regions apolipoprotein B (APOB), dentin matrix acidic phosphoprotein I (DMP1), recombination activating gene II (RAG2), protein kinase C, iota (PRKCI), signal transducer and activator of transcription 5A (STAT5A), and thyrotropin (THY), with supported phylogenetic relationships from both maximum-parsimony (≥70% bootstrap values) and Bayesian analysis (≥0.95 posterior probability) depicted. Scale is in number of substitutions per site. Symbols and abbreviations as in Fig. 2.

Combined sequences.—More than 90% of clades were in concordance between mtDNA and nDNA trees, and those data sets were concatenated for the combined analysis. Despite this high level of concordance, 2 areas with supported discrepancies between the mtDNA and nDNA trees were found. These supported discrepancies were found toward clade tips and fell outside the focus of this study. The 1st discrepancy related to the sister taxon of P. coromandra, which was P. tenuis in the mtDNA tree (Fig. 2) and P. javanicus in the nDNA tree (Fig. 3). The 2nd difference involved relationships within Lasiurus, which formed a well-supported clade in both analyses. Concatenation of the mtDNA and nDNA data sets resulted in 8,510 aligned positions. Because of possible violation of positional homology, 1,734 positions were excluded prior to analysis leaving 6,776 positions for phylogenetic analysis (Table 2). Of those remaining positions, 3,093 were variable and 2,344 were phylogenetically informative. The maximum-parsimony analysis resulted in 4 most-parsimonious trees, with 13,885 steps and 57 supported clades (Fig. 4). Excluding uninformative characters, the CI was 0.3380 and the RI was 0.6439. Differences among the 4 most-parsimonious trees related to relationships among taxa in the New World Myotis and the position of Otonycteris73x2013;Barbastella basal to either the genus Plecotus or Corynorhinus. For the Bayesian analysis a burn-in of 1,000 generations was used and resulted in a tree with 69 supported clades (Fig. 4).

Fig. 4

Phylogram from Bayesian analysis of the combined ribosomal mitochondrial DNA (12S rRNA, tRNAVal, and 16S rRNA) and nuclear DNA (apolipoprotein B [APOB], dentin matrix acidic phosphoprotein I [DMP1], recombination activating gene II [RAG2], protein kinase C, iota [PRKCI], signal transducer and activator of transcription 5A [STAT5A], and thyrotropin [THY]) gene regions, with supported phylogenetic relationships from both maximum-parsimony (≥70% bootstrap values) and Bayesian analysis (≥0.95 posterior probability) depicted. Scale is in number of substitutions per site. Symbols and abbreviations as in Fig. 2.


Elucidating evolutionary relationships within Vespertilioninae historically has been problematic. A paucity of useful characters, possible convergence among these character states, and a rapid radiation of major lineages within this subfamily have hindered efforts to understand evolutionary relationships of these taxa for >100 years (Ellerman and Morrison-Scott 1951; Heller and Volleth 1984; Hill 1966; Hill and Harrison 1987; Hill and Topál 1973; Horáček and Zima 1978; Koopman 1975,1994; Lack et al. 2010; Miller 1907; Rosevear 1962; Simmons 1998; Tate 1942; Volleth and Heller 1994b; Zima and Horáček 1985). Efforts over the last 20 years provided some refined hypotheses but were incomplete, were incongruent with historic hypotheses, or did not clarify all relationships within Vespertilioninae (Hill and Harrison 1987; Volleth and Heller 1994b; Volleth et al. 2001, 2006). Recent molecular analyses (Hoofer et al. 2003; Hoofer and Van Den Bussche 2001, 2003; Kawai et al. 2002; Mayer et al. 2007; Mayer and von Helversen 2001) have tested previous hypotheses with new informative characters using phylogenetic methods. Using ribosomal mtDNA sequence data, Hoofer and Van Den Bussche (2003) completed the most comprehensive phylogenetic study of Vespertilionidae and provided a sound hypothesis for the evolutionary relationships for many of these bats. However, they were still unable to resolve many of the supergeneric relationships within Vespertilioninae and presented new evolutionary hypotheses that require further testing. To resolve these relationships >5,500 bp of coding and noncoding sequence data from the nDNA genome were analyzed in combination with the previously sequenced mtDNA data to reevaluate hypotheses of the evolutionary relationships within Vespertilioninae. Because of the stochastic nature of lineage sorting the inclusion of data from the nuclear and mitochondrial genomes is important in fully understanding evolutionary relationships (Avise 1994).

Tribes of Vespertilioninae.—This study provides phylogenetic information for 8 of the 10 tribes previously proposed in various classifications of Vespertilioninae (Antrozoini, Eptesicini, Lasiurini, Myotini, Nycticeiini, Nyctophilini, Pipistrellini, Plecotini, Scotophilini, and Vespertilionini). We were unable to obtain tissue samples from either Nyctophilus or Pharotis Nyctophilini sensu (Koopman 1994; Simmons 2005) and therefore were unable to address phylogenetic affinities of these taxa. Accumulating evidence of the affinities of Myotis to Kerivoulinae and Murininae has required removal of Myotini (excluding Lasionycteris) from Vespertilioninae and elevation of Myotis to subfamily rank (Myotinae—Hoofer and Van Den Bussche 2003; Kawai et al. 2002; Lack et al. 2010; Stadelmann et al. 2004; Volleth and Heller 1994b). Although Myotis taxa were included in this study and are supported as a monophyletic group, this study was not designed to examine the affinities of the Myotis.

With regard to the remaining 8 traditionally recognized tribes, the combined tree provided support for 6 tribes, Antrozoini, Lasiurini, Scotophilini, Vespertilionini, and 2 unnamed tribes hereafter referred to as the hypsugine group and the perimyotine group (Fig. 4). Lasiurus has been recognized as a unique group within Vespertilioninae since the genus was 1st described (Gray 1831), and classification of Lasiurus into its own tribe by Tate (1942) has not been challenged (Bickham 1979, 1987; Hall and Jones 1961; Handley 1960; Hill and Harrison 1987; Hoofer and Van Den Bussche 2003; Koopman 1994; Miller 1907). Results from the combined analysis also support monophyly of Lasiurini (Fig. 4). The combined tree is not fully resolved with respect to interspecific relationships within Lasiurus, but a supported red bat clade (L. atratus, L. seminolus, L. blossevillii, and L. borealis) is present. However, without full resolution within Lasiurus, previous hypotheses about relationships of red bats to proposed lineages of yellow bat (Dasypterus) and hoary bat (Lasiurus cinereus) cannot be tested.

Scotophilini was the 2nd tribe supported by the combined analysis (Fig. 4). The genus Scotophilus historically has been included in the tribe Nycticeiini (Koopman 1994; McKenna and Bell 1997; Simmons 2005; Tate 1942). This position has been contradicted by bacular morphology (Hill and Harrison 1987), cytogenetics (Volleth et al. 2006), and ribosomal mtDNA (Hoofer and Van Den Bussche 2003) and was rejected in this study by the combined mtDNA and nDNA analysis and by each independently (Fig. 4). These results are congruent with phylogenetic analysis of a combined mtDNA and nDNA supermatrix targeted at assessing Nycticeiini and Scotophilini monophyly (Roehrs 2009).

Antrozoini is the 3rd supported clade in the combined analysis (Fig. 4). The group consisting of Antrozous and Bauerus (often a synonym of Antrozous—cf. Engstrom and Wilson 1981) was 1st described as subfamily Antrozoinae (Miller 1897; Simmons 2005) and has since been unstable in position and rank. Miller (1907) grouped Antrozous and Bauerus in subfamily Nyctophilinae with Nyctophilus and Pharotis, a classification supported by Tate (1941) and Simpson (1945). Koopman and Jones (1970) were 1st to place Antrozous and Bauerus into tribe Antrozoini, but its position remained within Nyctophilinae. This position of Antrozoini within Nyctophilinae was questioned by Koopman (1970) based on zoogeography and Pine et al. (1971) based on bacular morphology. Antrozoini has since been placed within Vespertilioninae by most authors, with various affinities (Hill and Harrison 1987; Koopman 1994; McKenna and Bell 1997). The most divergent exception to this hypothesis is the elevation of Antrozoini to its own family, Antrozoidae, aligned closely to Molossidae (Simmons 1998; Simmons and Geisler 1998). However, this hypothesis has not been supported by phylogenetic analysis of mtDNA (Hoofer and Van Den Bussche 2003) or nDNA (Miller-Butterworth et al. 2007). Hoofer and Van Den Bussche (2003) redefined Antrozoini by including Rhogeessa and Baeodon into the tribe. Their arrangement is supported largely by the combined tree with a monophyletic Rhogeessa sister to an Antrozous-Bauerus clade; however, the position of Baeodon was unresolved (Fig. 4). As in Hoofer and Van Den Bussche (2003), our mtDNA gene tree supports the inclusion of Baeodon in Antrozoini (Fig. 2), but results of the nDNA analysis place Baeodon basal to the Lasiurini (Fig. 3). This relationship is not supported, and it is possible that the incomplete nDNA data set for Baeodon may cause instability at this node resulting in a lack of resolution.

The 4th supported group consisted of New World pipistrelles, Parastrellus hesperus and Perimyotis subflavus (Fig. 4), and would constitute a new, yet-to-be-named, tribe referred to here as the perimyotine group. These results support Hoofer and Van Den Bussche (2003) placing each species in its own genus, but their phylogeny was unresolved relative to the position of these taxa within Vespertilioninae and their relationship to each other. Although affinities for this perimyotine group are not clear, these 2 taxa are supported in a deeply diverging clade. Furthermore, the combined analysis demonstrates that these taxa are distinct from Pipistrellus and fall outside of the other Pipistrellus-like bats. The inclusion of Parastrellus and Perimyotis into their own tribe initially seems counterintuitive based on previous research (Baker and Patton 1967; Hamilton 1949; Hill and Harrison 1987; Tate 1942). However, these taxa were problematic to place within Pipistrellus (sensu Koopman 1994), and many other taxa (Arielulus, Falsistrellus, Hypsugo, Neoromicia, and Vespadelus) previously included in Pipistrellus are today considered valid genera with different affinities than to Pipistrellus. Furthermore, a single colonization of the Nearctic by the most recent common ancestor of these taxa is more parsimonious than multiple colonization events, and their deep divergence allows for the morphological and chromosomal divergence separating them. Considering New World pipistrelles as a separate tribe preserves their generic and deeply divergent differences (Hamilton 1949) while maintaining their apparent common ancestry (Fig. 4). However, this tribal-level peri-myotine group should be considered tentative until further research corroborates this relationship and resolves their position within Vespertilioninae.

The last 2 supported tribes form a sister relationship in the combined tree and include most taxa historically considered Pipistrellus-like (Fig. 4). The 1st of these tribes is composed of Nyctalus, Pipistrellus, Scotoecus, and Vespertilio. Because of inclusion of Vespertilio in this tribe and Vespertilio having priority, the most appropriate name for this tribe is Vespertilionini. The other tribe consisted of Chalinolobus, Hypsugo, Laephotis, Neoromicia, Nycticeinops, Tylonycteris, and Vespadelus. This tribe is currently unnamed, but because Hypsugo has priority, this group will be referred to as the hypsugine group. These results are congruent with the results of Roehrs (2009), who addressed intergeneric relationships of Pipistrellus-like bats using a digenomic data set with reduced taxon sampling.

Two other previously documented tribes, Nycticeiini and Plecotini, deserve mention. The combined phylogram presented here (Fig. 4) corroborates recent research (Hill and Harrison 1987; Hoofer and Van Den Bussche 2003; Roehrs 2009; Volleth et al. 2006) in rejecting Nycticeiini (sensu Tate 1942). However, with regard to Nycticeiini (sensu Hoofer and Van Den Bussche 2003), the combined analysis was in congruence topologically, but the clade lacked statistical support (Fig. 4). This lack of support likely stems from a difference between the mtDNA and nDNA tree topologies. The mtDNA gene tree from this study is in agreement with Hoofer and Van Den Bussche (2003) with only the Bayesian analysis supporting Nycticeiini. The nDNA tree includes an unsupported Nycticeiini clade excluding Nycticeius making this clade more appropriately named Eptesicini. As discussed by Roehrs (2009), it is apparent that Arielulus, Eptesicus (including Histiotus), Glauconycteris, Lasionycteris, and Scotomanes form a tribal-level clade, but more effort will be required to resolve the position of Nycticeius and will have an impact on the nomenclature of this clade.

Although taxa included in Plecotini have not been completely stable, this tribe has been consistently included in Vespertilioninae classification since it was described by Gray (1866) as Plecotina (Table 1). Handley (1959) is responsible for establishing the core Plecotini genera currently recognized: Barbastella, Corynorhinus, Euderma, Idionycteris, and Plecotus. Other taxa also have been included in Plecotini: Baeodon, Nycticeius, Otonycteris, Rhogeessa, Nyctophilus, and Histiotus (Bogdanowicz et al. 1998; Dobson 1878; Hill and Harrison 1987; Kawai et al. 2002; Pine et al. 1971; Qumsiyeh and Bickham 1993). Although morphologic and cytogenetic data support monophyly of the core Plecotini (Bogdanowicz et al. 1998; Frost and Timm 1992; Handley 1959; Leniec et al. 1987; Tate 1942; Tumlison and Douglas 1992; Volleth and Heller 1994a, 1994b), monophyly of this tribe only recently has been tested explicitly (Hoofer and Van Den Bussche 2001,2003). Hoofer and Van Den Bussche (2003) were unable to unambiguously support monophyly of the core Plecotini or their relationship to other previously proposed closely related genera. The combined analysis of this study, and the mtDNA and nDNA trees independently, also were unable to resolve Plecotini, leaving this tribe neither supported nor rejected (Fig. 4). These taxa could be extant members of one of the earliest radiations from Vespertilioninae ancestral stock and appear to have rapidly diverged, not allowing time for these gene regions to accumulate sufficient synapomorphic characters to clarify their evolutionary histories. Finally, despite a general lack of resolution of deep phylogenetic relationships within Vespertilioninae, the subfamily is supported as a monophyletic group to the exclusion of Myotis, which is congruent with current hypotheses.

Usefulness of nDNA and combined data.—The nuclear gene regions included in this study were individually relatively short (averaging ∼800 bp), had few variable positions (173– 570 bp), and included even few potential phylogenetically informative positions (129–415 bp; Table 2). For any 1 nDNA gene region relatively few potentially informative positions per taxon were found, resulting in topologies that were not fully resolved and less informative of true evolutionary relationships. Results of likelihood-mapping tended to support this supposition, with most independent nDNA gene regions resolving <80% of quartets and all independent nDNA gene regions resolving <90% of quartets (Fig. 1). Furthermore, because it is difficult to predict whether a particular gene tree reflects true evolutionary relationships, most studies currently use a suite of gene regions from multiple genomes to overcome potential problems with nonphylogenetic signal within any 1 particular gene region (Philippe and Telford 2006; Rodríguez-Ezpeleta et al. 2007). Gene regions included in this study have been used successfully in various combinations in previous studies of bats and other mammals (Amrine-Madsen et al. 2003; Baker et al. 2003; Eick et al. 2005; Matthee and Davis 2001; Matthee et al. 2001, 2004, 2007; Murphy et al. 2001; Van Den Bussche et al. 2003), and all of these markers have been included in a recent study of the phylogenetic relationships of Miniopteridae, Cistugo, Myotinae, Kerivoulinae, Murininae, and Vespertilioninae (Lack et al. 2010).

Although results presented here provide a more resolved hypothesis of evolutionary relationships of Vespertilioninae than previous phylogenetic studies, it appears that more sequence data and more taxa will be necessary to overcome stochastic error and fully resolve deep evolutionary patterns within this subfamily. However, these studies will need to overcome potential systematic errors that tend to increase with increasing amounts of sequence data by excluding taxa, genes, and possibly even codon positions that exhibit relatively rapid rates of evolution (Baurain et al. 2007; Brinkmann and Philippe 2008; Philippe and Telford 2006; Rodríguez-Ezpeleta et al. 2007).


This project would not have been possible without the support and loan of tissues from individuals and institutions including: R. J. Baker, Museum of Texas Tech University; N. B. Simmons, American Museum of Natural History; B. D. Patterson, L. R. Heaney, and W. T. Stanley, Field Museum of Natural History; S. B. McLaren, Carnegie Museum of Natural History; M. D. Engstrom and B. Lim, Royal Ontario Museum; M. Ruedi, Musé;um d'Histoire Naturelle de Genève and the University of Lausanne, Institut de Zoologie et d'Ecologie Animale; P. J. Taylor, Durban Natural Science Museum; J. K. Braun and M. A. Mares, Sam Noble Oklahoma Museum of Natural History; T. L. Yates, Museum of Southwestern Biology at the University of New Mexico; R. L. Honeycutt and D. Schlitter, Texas Cooperative Wildlife Collection at Texas A&M University; J. O. Whitaker, Jr., and D. S. Sparks, Indiana State University Vertebrate Collection; and T. E. Lee, Jr., Abilene Christian University. We thank C. E. Stanley, Jr., for generating a portion of the sequence data included in this project. G. Eick was helpful in providing primers and advice on polymerase chain reaction profiles for the nuclear introns used in this study. Thanks go to the Oklahoma State University Recombinant DNA/Protein Core Facility for use of its equipment and assistance in troubleshooting. J. K. Braun, M. J. Hamilton, D. M. Leslie, M. A. Magnuson, R. J. Tyrl, and 2 anonymous reviewers reviewed drafts of this document. National Science Foundation grants DEB-9873657 and DEB-0610844 to RAVDB funded this research. Any opinions, findings, and conclusions or recommendations expressed in this paper are those of the authors and do not necessarily reflect the views of the National Science Foundation.

Appendix I

View this table:
Appendix I

Taxonomic samples included in this study with tissue collection number, voucher specimen catalog number, general locality, and GenBank accession numbers (http://www.ncbi.nlm.nih.gov/). Specimens and tissue samples are housed in the following institutions: Abilene Christian University (ACU), American Museum of Natural History (AMNH), Carnegie Museum of Natural History (CM, SP), Colecció;n Mamíferos Lillo, Universidad Nacional de Tucumán (CML), Durban Natural Science Museum (DM), Field Museum of Natural History (FMNH), Indiana State University Vertebrate Collection (ISUV), Muséum d'Histoire Naturelle de Genève (MHNG), Museum of Southwestern Biology at the University of New Mexico (MSB, NK), Museum of Texas Tech University (TTU, TK), Natural History Museum of Bern (NHMB), Oklahoma State University Collection of Vertebrates (OSU, OK), Royal Ontario Museum (ROM, F), Sam Noble Oklahoma Museum of Natural History (OMNH, OCGR), Texas Cooperative Wildlife Collection at Texas A&M University (TCWC), Universidad Autónoma Metropolitana–Iztapalapa (UAMI), Universidad Nacional Autónoma de México (UNAM), and University of Lausanne, Institut de Zoologie et d'Ecologie Animale (IZEA) Mitochondrial DNA (mtDNA) for a specific taxon with GenBank accession number starting in AF or AY may not have been amplified from the specific specimen indicated. A dash (—) denotes information unavailable and therefore missing. A few specimens came from the personal collections of Dale W. Sparks (DWS), Manuel Ruedi (M), and Rodney L. Honeycutt (RLH and 05M3). GenBank accession numbers for sequences generated in this study are indicated in boldface type; all others were published previously (Eick et al. 2005; Hoofer and Van Den Bussche 2003; Lack et al. 2010). APOB = apolipoprotein B; DMP1 = dentin matrix acidic phosphoprotein I; RAG2 = recombination activating gene II; PRKCI = protein kinase C, iota; STAT5A = signal transducer and activator of transcription 5A; and THY = thyrotropin.

GenBank accession no.
Tissue collection no.Museum catalog no.
Kerivoula hardwickiiF44154ROM110829Đồ ng Nai Province, VietnamAF345928GU328143AY141893AY141034GU328304GU328378GU328447
Kerivoula lenis (analyzed previously as K. papillosa)F44175ROM110850Đồng Nai Province, VietnamAF345927GU328144GU328229AY141035GU328305GU328379GU328448
Kerivoula pellucidaF35987ROM102177East Kalimantan Province, IndonesiaAY495476GU328145GU328230GU328064GU328306GU328380GU328449
Harpiocephalus harpiaTK21258CM88159Uthai Thani Province, ThailandAF263235GU328139AY141892AY141031GU328300GU328375GU328443
Murina cyclotisM1209MHNG1826.033Phôngsaly Province, LaoGU952767GU328155GU328238GU328072GU328313GU328386GU328456
People–s Democratic Republic
Murina huttoniF42722ROM107739Đắk Lắk Province, VietnamAY495490GU328156GU328239GU328073GU328314GU328387GU328457
Murina tubinarisM1179MHNG1926.034Phôngsaly Province, LaoGU952768GU328157GU328240GU328074GU328315GU328388GU328458
People's Democratic Republic
Myotis albescensTK17932CM77691Marowijne District, SurinameAY495492GU328159GU328241GU328076GU328317GU328390GU328460
Myotis bocagiiFMNH150075FMNH150075Tanga Region, TanzaniaAF326096GU328160GU328242GU328077GU328318GU328391GU328461
Myotis cf. browni (analyzed previously as M. muricola)FMNH147067FMNH147067Mindanao Island, Philippine IslandsAY495504GU328169GU328251GU328086GU328327GU328400GU328470
Myotis californicusTK78797TTU79325TexasAY495495GU328161GU328243GU328078GU328319GU328392GU328462
Myotis capacciniiTK25610TTU40554Northern Province, JordanAY495494GU328162GU328244GU328079GU328320GU328393GU328463
Myotis ciliolabrumTK83155TTU78520TexasAY495497GU328163GU328245GU328080GU328321GU328394GU328464
Myotis dominicensisTK15613TTU31503St. Joseph Parish, DominicaAY495500GU328164GU328246GU328081GU328322GU328395GU328465
Myotis fortidensTK43186UAMIMichoacán, MexicoAY495502GU328165GU328247GU328082GU328323GU328396GU328466
Myotis keaysiTK13532Yucatán, MexicoAY495503GU328166GU328248GU328083GU328324GU328397GU328467
Myotis latirostrisM606MHNGMiao-Li County, TaiwanGU952769GU328167GU328249GU328084GU328325GU328398GU328468
Myotis levisFMNH141600FMNH141600São Paulo, Brazil AustraliaAF326097GU328168GU328250GU328085GU328326GU328399GU328469
Myotis moluccarum (analyzed previously as M. adversus)RLH62TCWCAY495491GU328158(not sequenced)GU328075GU328316GU328389GU328459
Myotis myotisIZEA3790MHNG1805.062Canton of Bern, SwitzerlandAF326098GU328170GU328252GU328087GU328328GU328401GU328471
Myotis nigricansFMNH129210FMNH129210Amazonas, PeruAF326099GU328171GU328253GU328088GU328329GU328402GU328472
Myotis ripariusAMNH268591AMNH268591Paracou, French GuianaAF263236GU328172GU328254GU328089GU328330GU328403GU328473
Myotis septentrionalisDWS609ISUV6454IndianaAY495507GU328173GU328255GU328090GU328331GU328404GU328474
Myotis thysanodesTTU79327TK78796Texas(not sequenced)(not sequenced)(not sequenced)GU328091(not sequenced)(not sequenced)GU328475
Myotis thysanodesTK78802TTU79330TexasAF326100GU328174GU328256(not sequenced)GU328332GU328405(not sequenced)
Myotis veliferTK79170TTU78599TexasAF263237GU328175GU328257AY141033GU328333GU328406GU328476
Myotis volansTK78980TTU79545TexasAY495510GU328176GU328258GU328092GU328334GU328407GU328477
Myotis welwitschiiFMNH144313FMNH144313Kasese District, Uganda OklahomaAY495511GU328177GU328259GU328093GU328335GU328408GU328478
Myotis yumanensisTK28753TTU43200AY495512GU328178GU328260GU328094GU328336GU328409GU328479
Antrozous pallidusNK506MSB40576CaliforniaGU328037GU328120GU328209GU328045GU328285GU328360GU328428
Antrozous pallidusNK39195MSBArizonaGU328038GU328121GU328210GU328046GU328286GU328361GU328429
Antrozous pallidusTK49646TTU71101TexasAF326088GU328122GU328211GU328047GU328287GU328362GU328430
Baeodon alleniTK45023UN AMMichoacán, MexicoAF326108HM561577HM561677HM561632HM568371HM568338HM593057
Bauerus dubiaquercusF33200ROM97719Campeche, MexicoAY395863GU328125GU328214GU328050GU328289GU328364GU328432
Rhogeessa aeneusTK20712TTU40012Belize District, BelizeAY495530HM561578HM561678HM561633HM568372HM568339HM593058
Rhogeessa miraTK45014UN AMMichoacán, MexicoAY495531HM561579HM561679HM561634HM568373HM568340HM593059
Rhogeessa parvulaTK20653TTU36633Sonora, MexicoAF326109GU328196GU328274GU328108GU328350GU328419GU328492
Rhogeessa tumidaTK40186TTU61231Valle Department, HondurasAF326110GU328197GU328275GU328109GU328351GU328420GU328493
Hypsugine group
Chalinolobus gouldiiRLH27TCWCAustraliaAY495461HM561610HM561710HM561665HM568404HM568363HM593090
Chalinolobus morio05M3TCWCAustraliaAY495462GU328129GU328218GU328054GU328292GU328367GU328435
Hypsugo cadornaeM1183MHNG1926.050Phôngsaly Province, Lao People's Democratic RepublicGU328041GU328140GU328226GU328061GU328301(not sequenced)GU328444
Hypsugo eisentrautiF34348ROM100532Côte d'IvoireAY495473HM561611HM561711HM561666HM568405HM568364HM593091
Hypsugo saviiIZEA3586MHNG1804.100Canton of Valais, SwitzerlandAY495475HM561612HM561712HM561667HM568406(not sequenced)HM593092
Laephotis namibensisSP4160CM93187Maltahöhe District, NamibiaAY495477HM561613HM561713HM561668HM568407HM568365HM593093
Neoromicia brunneaTK21501CM90802Estuaire Province, GabonAY495514HM561614HM561714HM561669HM568408HM568366HM593094
Neoromicia nanaDM7542DM7542KwaZulu-Natal Province, South AfricaAY495474GU328141GU328227GU328062GU328302GU328376GU328445
Neoromicia rendalliTK33238CM97977Coastal Province, KenyaAY495515HM561615HM561715HM561670HM568409HM568367HM593095
Neoromicia somalicaTK33214CM97978Coastal Province, KenyaAY495516HM561616HM561716HM561671HM568410HM568368HM593096
Nycticeinops schliejfeniTK33373CM97998Eastern Province, KenyaAF326101GU328183GU328261GU328095AJ866330AJ865440AJ865685
Tylonycteris pachypusF38442ROM106164Tuyen Quang Province, VietnamAY495538HM561617HM561717HM561672HM568411(not sequenced)HM593097
Tylonycteris robustulaM1203MHNG1926.059Phôngsaly Province, Lao People's Democratic RepublicHM561631HM561618HM561718HM561673HM568412(not sequenced)HM593098
Vespadelus regulusRLH30TCWCAustraliaAY495539HM561619HM561719HM561674HM568413HM568369HM593099
Vespadelus vulturnusRLH16TCWCAustraliaAY495499(not sequenced)(not sequenced)HM561675HM568414HM568370HM593100
Lasiurus atratusF39221ROM107228Potaro-Siparuni, GuyanaAY495478HM561580HM561680HM561635HM568374HM568341HM593060
Lasiurus blossevilliiF38133ROM104285Chiriquí Province, PanamaAY495479HM561581HM561681HM561636HM568375HM568342HM593061
Lasiurus borealisTK49732TTU71170TexasAY495480HM561582HM561682HM561637HM568376HM568343HM593062
Lasiurus cinereusTK78926TTUTexasAY495482HM561583HM561683HM561638HM568377HM568344HM593063
Lasiurus egaTK43132UNAMMichoacán, MexicoAY495483HM561584HM561684HM561639HM568378HM568345HM593064
Lasiurus intermediusTK20513TTU36631Oaxaca, Mexico(not sequenced)HM561585(not sequenced)HM561640HM568379HM568346HM593065
Lasiurus intermediusTK84510TTU80739TexasHM561627(not sequenced)HM561685(not sequenced)(not sequenced)(not sequenced)(not sequenced)
Lasiurus seminolusTK90686TTU80699TexasAY495484HM561586HM561686HM561641HM568380HM568347HM593066
Lasiurus xanthinusTK78704TTU78296TexasAY495485HM561587HM561687HM561642HM568381HM568348HM593067
Arielulus aureocollarisF38447ROM106169Tuyen Quang Province, VietnamHM561621HM561588HM561688HM561643HM568382HM568349HM593068
Eptesicus brasiliensisTK17809CM76812Nickerie District, SurinameAY495464HM561589HM561689HM561644HM568383HM568350HM593069
Eptesicus diminutusTK15033TTU48154Guarico, VenezuelaAY495465GU328133GU328220GU328056GU328295GU328370GU328438
Eptesicus furinalisAMNH268583AMNH268583Paracou, French GuianaAF263234GU328135GU328222AY141030GU328297GU328372GU328440
Eptesicus fuscusSP844CM102826West VirginiaAF326092GU328136GU328223GU328058GU328298GU328373GU328441
Eptesicus hottentotusTK33013CM89000Rift Valley Province, KenyaAY495466GU328137GU328224GU328059AJ866329AJ865438AJ865683
Eptesicus macrotusOCGR2301CML3230Neuquán Province, ArgentinaHM561622HM561590HM561690HM561645HM568384HM568351HM593070
Eptesicus macrotusFMNH129207FMNH129207Ancash Region, PeruAY495472HM561591HM561691HM561646HM568385HM568352HM593071
Eptesicus macrotusOCGR4227OMNH27925Salta Province, ArgentinaHM561623HM561592HM561692HM561647HM568386HM568353HM593072
Eptesicus macrotusOCGR3806OMNH32879Catamarca Province, ArgentinaHM561624HM561593HM561693HM561648HM568387HM568354HM593073
Eptesicus magellanicusOCGR2303OMNH23500Neuquán Province, ArgentinaHM561625HM561594HM561694HM561649HM568388(not se quenced)HM593074
Eptesicus serotinusM816MHNG1807.065GreeceHM561626HM561595HM561695HM561650HM568389HM568355HM593075
Eptesicus serotinusTK40897TTU70947Sidi Bou Zid Governorate, TunisiaAY495467HM561596HM561696HM561651HM568390HM568356HM593076
Glauconycteris argentataFMNH15119FMNH15119Kilimanjaro Region, TanzaniaAY495468HM561597HM561697HM561652HM568391HM568357HM593077
Glauconycteris beatrixFMNH149417FMNH149417Haute Zaire, ZaireAY495469HM561598HM561698HM561653HM568392(not sequenced)HM593078
Glauconycteris egeriaAMNH268381AMNH268381Central African RepublicAY495470HM561599HM561699HM561654HM568393(not sequenced)HM593079
Glauconycteris variegataTK33545CM97983Western Province, KenyaAY495471HM561600HM561700HM561655HM568394HM568358HM593080
Lasionycteris noctivagansTK24216TTU56255TexasAF326095GU328146(not sequenced)GU328065(notGU328381(not sequenced)
Lasionycteris noctivagansTK24889Oklahoma(not sequenced)(not sequenced)GU328231(not sequenced)GU328307(not sequenced)GU328450
Nycticeius humeralisTK26380TTU49536TexasAF326102(not sequenced)(not sequenced)GU328096(not sequenced)(not sequenced)(not sequenced)
Nycticeius humeralisTK90649TTU80664Texas(not sequenced)GU328184GU328262(not sequenced)GU328338GU328411GU328481
Scotomanes ornatusF42568ROM107594Tuyen Quang Province, VietnamAY495537HM561601HM561701HM561656HM568395HM568359HM593081
Scotophilus borbonicusTK33267CM98041Coastal Province, KenyaAY495532GU328199GU328276GU328110GU328352GU328421GU328494
Scotophilus dinganiiFMNH147235FMNH147235Tanga Region, TanzaniaAY495533GU328200GU328277GU328111AJ866332AJ865441AJ865686
Scotophilus heathiiF42769ROM107786Đắk Lắic Province, VietnamAY495534GU328201GU328278GU328112GU328353GU328422GU328495
Scotophilus kuhliiFMNH145684FMNH145684Sibuyan Island, Philippine IslandsAF326111GU328202GU328279GU328113GU328354GU328423GU328496
Scotophilus leucogasterTK33359CM90854Eastern Province, KenyaAY395867GU328203GU328280GU328114GU328355GU328424GU328497
Scotophilus nuxTK33484Western Province, KenyaAY495535GU328204GU328281GU328115GU328356GU328425GU328498
Scotophilus viridisFMNH150084FMNH150084Tanga Region, TanzaniaAF326112GU328206GU328283GU328117GU328357GU328426GU328499
Perimyotine group
Parastrellus hesperusTK78703TTU79269TexasAY495522GU328187GU328265GU328099GU328341GU328413GU328483
Perimyotis subflavusTK90671TTU80684TexasAY495523GU328191GU328269GU328103GU328345GU328416GU328487
Barbastella barbastellusIZEA3590MHNG1804.094Canton of Valais, SwitzerlandAF326089GU328124GU328213GU328049GU328288GU328363GU328431
Corynorhinus mexicanusTK45849UAMIMichoacán, MexicoAF326090GU328128GU328217GU328053GU328291GU328366GU328434
Corynorhinus rafinesquiiTK5959TTU45380ArkansasAF326091GU328130GU328219GU328055GU328293GU328368GU328436
Corynorhinus townsendiiOKI 1530OSU13099Oklahoma(not sequenced)GU328131(not sequenced)(not sequenced)GU328294GU328369GU328437
Corynorhinus townsendiiTK83182TTU78531TexasAF263238(not sequenced)AY141891AY141029(not sequenced)(not sequenced)(not sequenced)
Euderma maculatumNK36260MSB 121373UtahAF326093GU328138GU328225GU328060GU328299GU328374GU328442
Idionycteris phyllotisACU736ACU736(not sequenced)GU328142GU328228(not sequenced)GU328303GU328377(not sequenced)
Idionycteris phyllotisNK36122MSB120921UtahAF326094(not sequenced)(not sequenced)GU328063(not sequenced)(not sequenced)GU328446
Otonycteris hemprichiiSP7882Maan Government, JordanAF326103GU328186GU328264GU328098GU328340GU328412GU328482
Plecotus auritusIZEA2693(not sequenced)(not sequenced)GU328266(not sequenced)GU328342GU328414(not sequenced)
Plecotus auritusIZEA2694MHNG1806.047Canton of Valais, SwitzerlandAF326106GU328188(not sequenced)GU328100(not sequenced)(not sequenced)GU328484
Plecotus austriacusIZEA3722MHNG1806.042Canton of Valais, SwitzerlandAF326107GU328189GU328267GU328101GU328343GU328415GU328485
Plecotus gaisleriIZEA4780MHNG1806.051Meknés-tafilalet, MoroccoGU328043GU328192GU328270GU328104GU328346GU328417GU328488
Nyctalus leisleriFMNH140374FMNH140374Malakand Division, PakistanAY495517HM561602HM561702HM561657HM568396(not sequenced)HM593082
Nyctalus noctulaNHMB 209/87NHMB 209/87Canton of Berne, SwitzerlandAY495518HM561603HM561703HM561658HM568397(not sequenced)HM593083
Pipistrellus coromandraFMNH140377FMNH140377Malakand Division, PakistanAY495524GU328190GU328268GU328102GU328344(not sequenced)GU328486
Pipistrellus hesperidusDM8013DM8013KwaZulu-Natal, South AfricaHM561628HM561604HM561704HM561659HM568398(not sequenced)HM593084
Pipistrellus javanicusFMMNH147069FMMNH147069Mindanao Island, PhilippinesAY495525GU328193GU328271GU328105GU328347(not sequenced)GU328489
Pipistrellus nathusiiIZEA2830MHNG1806.003Canton of Vaud, SwitzerlandAF326104HM561605(not sequenced)HM561660(not sequenced)(not sequenced)(not sequenced)
Pipistrellus nathusiiIZEA3406MHNG1806.001Canton of Vaud, Switzerland(not sequenced)(not sequenced)HM561705(not sequenced)HM568399(not sequenced)HM593085
Pipistrellus paterculusM1181MHNG1926.045Phôngsaly Province, Lao People's Democratic RepublicHM561629HM561606HM561706HM561661HM568400HM568360HM593086
Pipistrellus pipistrellusM1439MHNG1956.031Canton of Genève, SwitzerlandHM561630HM561607HM561707HM561662HM568401HM568361HM593087
Pipistrellus pygmaeusIZEA3403MHNG1806.032Barcelona Province, SpainAF326105GU328195GU328273GU328107GU328349HM568362GU328491
(analyzed previously as P. pipistrellus)
Pipistrellus tenuisFMNH137021FMNH137021Sibuyan Island, PhilippineAY495529HM561608HM561708HM561663HM568402(not sequenced)HM593088
Scotoecus hirundoFMNH151204FMNH151204Kilimanjaro Region, TanzaniaAY495536HM561609HM561709HM561664HM568403(not sequenced)HM593089
Vespertilio murinusIZEA3599MHNG1808.017Canton of Valais, SwitzerlandAY395866HM561620HM561720HM561676HM568415(not sequenced)HM593101


  • Associate Editor was David L. Reed.

Literature Cited

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