Phylogenetics of the fruit-eating bats (Phyllostomidae: Artibeina) inferred from mitochondrial DNA sequences

2008·0.96 MB·English

by  Hoofer, Steven R

TEXAS TECH UNIVERSITY
Natural Science Research Laboratory
Occasional Papers
Museum of Texas Tech University Number 277 2 October 2008
Phylogenetics of the Fruit-eating Bats (Phyllostomidae: Artibeina)
Inferred from Mitochondrial DNA Sequences
Steven R. Hoofer, Sergio Solari, Peter A. Larsen, Robert D. Bradley, and Robert J. Baker
Abstract
Approximately 24 species classified in three groups (Artibeus, Dermanura, and Koopma¬
nia) compose Subtribe Artibeina, an assemblage of New World leaf-nosed bats (Phyllostomidae)
for which evolutionary relationships have proven difficult to resolve. We examined artibeine
systematics through broad taxonomic sampling and phylogenetic analysis of DNA sequences
for two mitochondrial genes. Analysis of 16S rRNA sequences offered an additional test of
previous genealogical hypotheses, and facilitated knowledge about the congruence in variation
between the well studied cytochrome-6 gene and the evolutionary history of this complex of
bats. Our results illustrate a high degree of congruence between these linked mitochondrial loci
that in combination offers a well resolved gene tree and robust predictions to all but a few of
the examined relationships. Highlights include: monophyly of Artibeina in contrast to previous
hypotheses of polyphyly; two main lineages within Artibeina in accordance with monophyly
of the smaller Dermanura species and larger Artibeus species; sister relationship between A.
concolor and other Artibeus species rather than with Dermanura, contrasting the argument for
recognizing A. concolor as a separate genus (Koopmania); reconfirmation of several species
formerly considered subspecies (A. planirostris, A. schwartzi, D. bogotensis, D. rava, and D.
rosenbergi)-, and further indication that A. intermedins and A. lituratus are conspecific.
Key words: 16S rRNA, Artibeus, cytochrome-6, Dermanura, DNAsequence, Koopmania,
phyllostomid bats, systematics
Resumen
Aproximadamente 24 especies pertenecientes a tres grupos (Artibeus, Dermanura, y
Koopmania) componen la subtribu Artibeina, un ensamblaje de murcielagos de hoja nasal del
Nuevo Mundo (Phyllostomidae), cuyas relaciones evolutivas han sido dificiles de resolver. Ex-
aminamos la sistematica de los artibeinos a traves de un amplio muestreo taxonomico y analisis
filogeneticos de secuencias del ADN para dos genes mitocondriales. Analisis de secuencias del
gen 16S rARN ofrecen una prueba novedosa de hipotesis genealogicas previas, facilitando el
conocimiento sobre la congruencia en variacion respecto al mejor conocido citocromo b y la
2 Occasional Papers, Museum of Texas Tech University
historia evolutiva de este complejo de especies. Nuestros resultados ilustran un alto grado de
congruencia entre estos loci mitocondriales, que en combinacion ofrecen predicciones robustas
para casi todas las relaciones examinadas. Resultados relevantes incluyen: monofilia de los Ar-
tibeina en contraste a hipotesis previas de parafilia; dos linajes mayores dentro de los Artibeina,
correspondiendo con la monofilia de especies pequenas de Dermanura y grandes de Artibeus',
la relacion cercana entre A. concolor y otras especies de Artibeus antes que con Dermanura,
en contraste con la propuesta de reconocer A. concolor como un genero distinto (Koopmania);
el reconocimiento de varias especies previamente consideradas subespecies (A. planirostris, A.
schwartzi, D. bogotensis, D. rava, y D. rosenbergi)', y el reconocimiento deA. intermedius como
un sinonimo menor de A. lituratus.
Palabras clave: 16S rARN, Artibeus', citocromo-#; Dermanura', filostomidos; Koopmania',
murcielagos; secuancias de ADN; sistematica
Introduction
Artibeine bats compose a large and diverse group In contrast, analyses of cytochrome-# DNA sequences
of fruit-eating specialists within the New World family and EcoRI-defined satellite DNA demonstrated a most
Phyllostomidae (subfamily Stenodermatinae: subtribe recent common ancestry for Artibeus, Dermanura, and
Artibeina — Baker et al. 2003). From 18 to 24 species Koopmania (monophyly of Artibeina; Van Den Bussche
are recognized (Simmons 2005; Larsen et al. 2007; et al. 1993, 1998). Based on anagenic and cladogenic
Solari et al. in prep.) and classified into three groups: interpretations of their results, coupled with morpho¬
the medium- to large-sized species of Artibeus (amplus, logical and karyotypic evidence (Andersen 1906; Baker
fimbriatus, fraterculus, hirsutus, inopinatus, interme¬ 1973; Straney et al. 1979), Van Den Bussche et al.
dius, jamaicensis, lituratus, obscurus, planirostris, (1993, 1998) recognized Artibeus and Dermanura as
and schwartzi), the small-sized species of Dermanura separate, closely related genera, and Koopmania con¬
(ianderseni, azteca, bogotensis, cinerea, glauca, gnoma, color as A. concolor. The monotypic Enchisthenes was
incomitata,phaeotis, rctva, rosenbergi, tolteca, and wat- regarded as genus distinct from Artibeina, which has
soni); and the medium-sized Koopmania {concolor). been affirmed in additional studies of morphological
Morphologically, Enchisthenes hartii shares affinities and molecular data (Baker et al. 2000, 2003; Wetterer
with Artibeus and also has been recognized as part of et al. 2000).
the artibeines (e.g., Koopman 1993, 1994).
Although the Van Den Bussche etal. (1993,1998)
Relationships among artibeine bats have proven studies are the most important and comprehensive mo¬
difficult to resolve with the characters that have lecular assessments of Artibeine relationships to date,
been examined so far (morphology, karyotypes, and their taxonomic sampling was limited at that time by
cytochrome-# DNA sequences). As a result, there are the lack of available tissue samples for Artibeus and
disagreements over rank status of Dermanura and Arti¬ Dermanura and lack of efficient methods of automated
beus and over monophyly of the group as a whole. For DNA sequencing. Tissue samples of numerous addi¬
example, Owen’s (1987, 1991) analyses of mensural tional individuals for the taxa they examined, as well as
and discrete-state morphological characters indicated several newly recognized species (A. schwartzi [Larsen
a polyphyletic origin for Artibeina: Artibeus shared et al. 2007] and D. rava and D. rosenbergi [Solari et
a most recent common ancestry with Ectophylla and al. in prep.]), are now available for molecular study.
Uroderma (his subtribe Artibeini) whereas Dermanura Also available (and feasible) now are contemporary
and Koopmania shared a most recent common ancestry phylogenetic methods that utilize objective systems for
with Enchisthenes and the white-shouldered stenoder- character weighting and efficient systems with which to
matine genera {Ametrida, Ardops, Ariteus, Stenoderma, reconcile important biological phenomena for molecu¬
Centurio, Phyllops, Pygoderma, and Sphaeronycteris). lar data (e.g., among-site rate variation, unequal base
Hoofer et al.—mtDNA Phylogeny of Artibeine Bats 3
frequencies, and nonindependence of substitutions). cytochrome-# sequences along with a complementary
Therefore, our purpose in this study was to re-assess dataset of complete 16S ribosomal RNA (rRNA) se¬
monophyly of Artibeus, Dermanura, and Artibeina, as quences. These linked genes together should increase
well as the validity of Koopmania, through broad taxo¬ the probability of detecting supported resolution to the
nomic sampling and phylogenetic analysis of complete gene tree (Moore 1995).
Materials and Methods
Specimens examined.—Specimens examined are for costs of opening and extending gaps. We viewed
listed in the Appendix, including information associ¬ alignments in MacClade software (version 4.05; Mad-
ated with museum vouchers. We generated complete dison and Maddison 2002) to ensure there were no
cytochrome-# sequences for 37 individuals and com¬ insertions, deletions, or stop codons in the cytochrome-
plete 16S rRNA sequences for 50 individuals. From # sequences and to inspect gap placement in the 16S
GenBank, we retrieved 41 cytochrome-# sequences rRNA sequences. We delimited ambiguously aligned
that were originally generated by Van Den Bussche et sites in the 16S rRNA alignment by using criteria and
al. (1993), Lim et al. (2004), Porter and Baker (2004), justification in Hoofer and Van Den Bussche (2003),
Hoofer and Baker (2006), and Larsen et al. (2007), and and performed data analysis without those sites. We
six 16S rRNA sequences that were originally generated coded nucleotides as unordered, discrete characters,
by Van Den Bussche and Hoofer (2000) and Baker et gaps as missing data, and multiple states as polymor¬
al. (2003). Lists of specimens examined including phisms. In PAUP* software (test version 4.0bl0; Swof-
voucher information are accessible in each of those ford 2002), we examined level of phylogenetic signal
publications and in the Appendix. We used sequences via the ^-statistic (Hillis and Huelsenbeck 1992) for
representing Chiroderma, Ectophylla, and Uroderma 100,000 randomly drawn trees.
as outgroups (Baker et al. 2000, 2003; Wetterer et al.
2000) and inferred relationships among ingroup taxa We inferred phylogenetic relationships by Bayes¬
representing Enchisthenes and all recognized species of ian analysis implemented in MrBayes 2.01 software
Artibeina excepting D. incomitata, for which samples (Huelsenbeck and Ronquist 2001) and by Maximum
were unavailable. Likelihood and Parsimony analyses implemented
in PAUP* software (test version 4.0b 10; Swofford
Molecular methods.—We extracted genomic 2002). The general time reversible (GTR) model with
DNAfrom skeletal muscle or organ tissue samples with allowance for gamma distribution of rate variation (T)
standard phenol methods (Longmire et al. 1997). We and for proportion of invariant sites (I) best fit both
followed previous methods to amplify and sequence cytochrome-# and 16S rRNA data based on Akaike
the entire cytochrome-# (Larsen et al. 2007) and 16S Information Criterion tests implemented in Modeltest
rRNA (Van Den Bussche and Hoofer 2000) genes. We 3.06 software (Posada and Crandall 1998).
sequenced both strands by using Big-Dye version 3.1
chain terminators, followed by electrophoresis on a For Bayesian analysis, we ran two X 106 gen¬
3100-Avant Genetic Analyzer (Applied Biosystems, erations with one cold and three incrementally heated
Foster, City, California). We assembled resulting, over¬ Markov chains, random starting trees for each chain,
lapping fragments in AssemblyLIGN™ 1.0.9 software and trees sampled (saved) every 100 generations. We
(Oxford Molecular Group PLC, Oxford, United King¬ treated model parameters as unknown variables (with
dom) and Sequencing Analysis 3.4.1 software (Applied uniform priors) to be estimated in each Bayesian analy¬
Bio systems, Inc., Foster City, California). sis (Leache and Reeder 2002). We ran three indepen¬
dent analyses with burn-in values based on empirical
Phylogenetic analysis.—We performed multiple evaluation of likelihoods converging on stable values.
sequence alignment for both data sets in Clustal X soft¬ We calculated a 50% majority-rule consensus tree from
ware (Thompson et al. 1997) with default parameters the sample of stabilized trees in PAUP* software (test
4 Occasional Papers, Museum of Texas Tech University
version 4.0bl0; Swofford 2002) and obtained branch trees by simple addition, tree-bisection-reconnection
lengths via the “sumt” option in MrBayes software branch swapping, and allowance for negative branch
(Huelsenbeck and Ronquist 2001). We assessed clade lengths. For Parsimony analysis, we treated all char¬
reliability via posterior probabilities and regarded acters and substitution types with equal probability
values > 0.95 as significant. and conducted full heuristic searches with 10 random
additions, starting trees by simple addition, and tree-
For Maximum Likelihood analyses, we used the bisection-reconnection branch swapping. We assessed
GTR + T +1 model and parameters given by Modeltest clade reliability via bootstrapping with 250 iterations
(cytochrome-#, rAC = 2.42, rAG = 19.70, rAT = 2.99, rCG for Parsimony analyses (Felsenstein 1985) and regarded
0.69. rCT = 41.75, nA= 0.31, jtC = 0.30, jtG = 0.12, a = values > 70 as support. Due to computation time, we
1.27, and P = 0.55; 16S rRNA, rAr = 3.99, rAP = 15.62, performed Maximum Likelihood bootstrapping only
rAT = 4.45, r = 0.76, rrT = 80.08, jtA= 0.37, jtC = 0.20, on the combined mitochondrial dataset and utilized
jtG = 0.18, a = 0.76, and Pinv = 0.58), performed full a “fast” stepwise-addition approach to tree searching
heuristic searches with 10 random additions, starting rather than a full-heuristic search.
Results
Cytochrome-b and 16S rRNA.—Sequence posterior probabilities for nodes and model parameters
alignment of the complete cytochrome-# gene for 37 for all sets of runs (three mns each) within data sets
specimens generated in this study (GenBank accession agreed regardless of choice of outgroup. Maximum
nos. FJ179223-FJ179259) and the 41 retrieved from Likelihood analysis resulted in a single best tree for
GenBank was unequivocal and without internal stop both cytochrome-# (Lnl = -10,611.03) and 16S rRNA
codons. Of the 1,140 characters, 697 were constant (Lnl = -8,986.50) data sets. Parsimony analysis re¬
and 380 parsimony-informative, with nucleotide varia¬ sulted in 240 most-parsimonious trees (length = 2,077,
tion distributed across codon positions as expected for Cl = 0.28, RI = 0.74) and 108 most-parsimonious trees
protein coding genes (Simon et al. 1994): 84 at first (length = 1,125, Cl = 0.46, RI = 0.77) for cytochrome-#
positions, 34 at second positions, and 325 at third and 16S rRNA data sets, respectively. For both datasets,
positions. Complete sequences of the 16S rRNA gene differences among most-parsimonious trees primarily
averaged 1,559 base pairs for the 56 taxa examined involved alternative arrangements of terminal branches
(GenBank accession nos. FJ179173-FJ179222), within species and, in a few instances, involved alter¬
ranging from 1,557 (A. fraterculus, A. inopinatus, native inter-specific relationships within Artibeus and
A. schwartzi, D. anderseni, and D. cinerea) to 1,562 Dermanura. Overall, there were some topological
(D. watsoni). Sequence alignment resulted in 1,578 differences within and between data sets and between
characters, corresponding in length and similarity to the three optimality criteria; however, none of the
other 16S rRNA sequences in GenBank. We excluded differences were supported. Statistically supported
83 characters in nine regions of the alignment (rang¬ topologies (i.e., > 70% bootstrap value, > 0.95 Bayes¬
ing from two base pairs to 46 base pairs) because of ian posterior probability) obtained from all optimality
ambiguity in assessment of positional homology. This criteria agreed within and between each data set (Figs.
left 1,495 characters for analysis, of which 1,110 were 1 and 2).
constant and 289 parsimony-informative. Levels of
phylogenetic signal were significant based on the gT Combined cytochrome-b and 16S rRNA.—We
statistic (P < 0.01—Hillis and Huelsenbeck 1992) for combined the data sets because there was high degree
cytochrome-# (-0.3335) and 16S rRNA (-0.3428). of congruence and no supported conflicts between
them (Wiens 1998). The combined data set (2,635
For cytochrome-# and 16S rRNA data sets, base pairs) included the 49 specimens shared between
Bayesian likelihoods reached stationarity before data sets. It also consists of three chimeric taxa that,
100,000 generations (i.e., bum-in = 1,000), thinning the in both cases, included cytochrome-# data from one
data points to 19,000 for each data set. Topology and specimen and 16S rRNA data from another speci-
Hoofer et al.—mtDNA Phylogeny of Artibeine Bats 5
TK15013 (DQ869424)
TK143051 (DQ869398)
TK104410 (DQ869410) A. planirostris
TK17073 (U66503)
TK18788 (U66504)
TK16633 (U66508)
ROM 107904 (AY642924) A. amplus
ROM107847 (AY642923)
A intermedius TK101996
— 0.01 substitutions/site A intermedius TK31924 (U66502)
A lituratus TK104525 (DQ869393) A. intermedius/
A lituratus TK104112 A. lituratus
A lituratus JKt 28642
A intermedius TK101993
A lituratus TK25029 (U66505)
TK17308 (U66506)
TK18787 (U66507) A. obscurus
1 00 1 TK86531
100 | TK82839 (DQ869524) A. schwartzi
100 TK82842 (DQ869525)
TK27682 (DQ869480) A. jamaicensis
TK135290
TK18991 (U66498)
TK99588 (DQ869391) A. fimbriatus
TK56670 (DQ869390)
" TK135408 (DQ869388)
' TK135760 A. fraterculus
' TK134686 (DQ869389)
97 100 L -TK16631 (U66499)
78 ioo i- [1 TTKK115500559885 A. hirsutus
100 L — NK11128 (U66500)
J TK101201
ioo r J TK101202 A. inopinatus
TK101203
“ TK40184 (U66501)
-TK11240 (U66519)
TK10378 (U66518) A. concolor
100
TK145271
100 j"lT TKK1199338810 (DQ869386) D. bogotensis
99 100 1 TK19379
100 98 100 cE ■ T■ KT1K81708491 (1U66 6513) D. gnoma
92 TK104117
100 TK104136
100 TK16636 (U66512) D. glauca
100 TK104203
93 TK104592
100 TK134526 D. rava
TK134611
TK104590
- NK14319 (U66509) D. anderseni
- TK18790(U66511) D. cinerea
100 i*I- TTKK8822899071 D. azteca
100 TK82898
TK82895
TK82896
100 TK136188 (DQ869387) D. phaeotis
89 100 TK136234
TK82894
79 100 TK5411 (U66514)
100 TK82900
P TK136023
TK82899 D. tolteca
TTKK1133I6 035
TK22579 (U66515)
100 110000 iS TK136988 TK4723 (U66510) D. watsoni
TK7877 (U66516)
97 100 r TK135691 D. rosenbergi
100 tv4 r\A cnn
100 _i TK16395 (AY157033) Ectophylla
100 ~ TK125311 (DQ312404)
■ TK22690 (U66517) Enchisthenes
■ TK25052 (DQ312414) Chiroderma
TK46006 (DQ312405) Uroderma
Figure 1. Maximum likelihood phylogram (Lnl = -10,611.03) from analysis of complete cytochrome-6 sequences
(1,140 base pairs) using best-fit model (GTR + T + I; rAC = 2.42, rAG = 19.70, rAT = 2.99, rCG = 0.69, rCT = 41.75, jtA
= 0.31, jtC = 0.30, jtG = 0.12, a = 1.27, and Pmv = 0.55). We designated Chiroderma, Ectophylla, and Uroderma as
outgroups. Numbers above branches are Bayesian posterior probabilities, whereas those below are bootstrap percentages
from Parsimony. Values are shown only for nodes supported by P > 0.95 or bootstrap percentage > 50, or both. “A ”
= Artibeus, “D. ” = Dermanura.
6 Occasional Papers, Museum of Texas Tech University
A. lituratus TK104112
A. lituratus TK128642 A. intermedius/
lituratus TK104525 A. lituratus
A. intermedius TK101993
A. intermedius TK101996
100 I— TK135290
-0.01 substitutions/site TK135905 A. jamaicensis
TK27682
TK86531 |
A. obscurus
TK17080
|
TK82842 A. schwartzi
TK82839
AMNH267998 (AF263225)
AMNH267999 (AF263226)
TK15013 A. planirostris
*- TK143051
TK104410
TK101201
TK101202 A. inopinatus
TK101203
TK150585 |
A. hirsutus
A. fraterculus
— A. concoior
D. bogotensis
|
D. gnoma
|
100 D. glauca
97 TK104203
TK82899
TK136035
D. tolteca
“ TK136023
■ TK82900
|
TK136188 D. phaeotis
TK82894
- TK134526
100 - TK134611 D. rava
100 TK104592
TK18790 — D. cinerea
|
D. anderseni
100
r- TK135691
100 nL ^ TK104509 D. rosenbergi
100 100 ioo TK104501
90 100 TK136988 — D. watsoni
I TK82898
ioo r1 t ■ TK82901 D. azteca
100 TK82897
— Enchisthenes
■ TK16395 (AY395811) — Ectophylla
- TK70524 (AY395837) — Chiroderma
TK46006 (AY395831) — Uroderma
Figure 2. Maximum likelihood phylogram (Lnl = -8,986.50) from analysis of complete 16S rRNA sequences (1,495
base pairs) using best-fit model (GTR + r +1; rAC = 3.99, rAG = 15.62, rAT = 4.45, rCG = 0.76, rCT = 80.08, jtA = 0.37, jtC
= 0.20, jtG = 0.18, a = 0.76, and Pinv = 0.58). We designated Chiroderma, Ectophylla, and Uroderma as outgroups.
Numbers above branches are Bayesian posterior probabilities, whereas those below are bootstrap percentages from
Parsimony. Values are shown only for nodes supported by P > 0.95 or bootstrap percentage > 50, or both. “A.” =
Artibeus, “D= Dermanura.
Hoofer et al.—mtDNA Phylogeny of Artibeine Bats 7
men; Artibeus obscurus comprised two individuals, of outgroup choice. Maximum Likelihood analysis
Enchisthenes hartii comprised two individuals, and resulted in a single best tree (Lnl= -15,882.18) and
Chiroderma comprised two species (C. salvini and C. Parsimony analysis resulted in two most-parsimonious
villosum). Bayesian likelihoods reached stationarity trees (length = 2,769, Cl = 0.38, RI = 0.71). Topolo¬
before 100,000 generations as above, and topology and gies and levels of nodal support obtained from all three
posterior probabilities for nodes and model parameters optimality criteria were nearly identical (Fig. 3).
for all sets of runs (three runs each) agreed regardless
Discussion
Higher-level relationships.—Few assessments of Enchisthenes (Figs. 1-3). This study therefore affirms
artibeine relationships have been undertaken that in¬ previous cladistic analyses for supporting a recent
cluded explicit phylogenetic analysis of Enchisthenes, common ancestry and monophyly of Artibeina (sensu
A. concolor (= Koopmania), and multiple representa¬ Baker et al. 2003) in contrast to Owen’s (1987, 1991)
tives of Artibeus and Dermanura. Morphological hypothesis of polyphyly. If our analyses supported the
studies by Owen (1987, 1991) and molecular studies latter hypothesis, then Artibeus would be depicted as
by Van Den Bussche et al. (1993, 1998) are the most sharing a most recent common ancestry with Ectophylla
comprehensive and reveal competing hypotheses of and the other vampyressine genera (Chiroderma, and
relationship. Whereas Owen’s analyses of essentially Uroderma), and Dermanura and Koopmania would
all stenodermatine taxa indicate independent origins be depicted as sharing a most recent common ances¬
for the small- and large-sized artibeine bats, those of try with Enchisthenes. All of our results exclude that
Van Den Bussche et al. support a recent common an¬ hypothesis.
cestry for these taxa after diverging from Enchisthenes
and other stenodermatine genera. Resolving these Although the phylogenetic position of Enchis¬
differences is key to the higher-level systematics and thenes is not fully resolved, our analyses demonstrate
taxonomy of artibeine bats. its anagenic and cladogenic distinction relative to the
artibeine bats. Thus, our results affirm previous studies
Without Owen’s hypothesis of polyphyly, which of morphological, karyotypic, allozymic, and molecular
led to him to recognize genus Artibeus (mid- to large¬ data supporting the generic distinction of Enchisthenes
sized species), elevate Dermanura (small-sized species) (Andersen 1906,1908; Miller 1907; Baker et ah 1979,
to generic rank, and describe a new genus Koopmania 2000,2003; Koop and Baker 1983; Owen 1987,1991;
(mid-sized A. concolor), rank status of the three lin¬ Van Den Bussche 1992; Van Den Bussche et ah 1993,
eages within Artibeina are arbitrary. This situation has 1998; Pumo et ah 1996; Tandler et al. 1997; Wetterer
been acknowledged by several authors, as exemplified et al. 2000) and disagree with suggestions of recogniz¬
in the most recent classificatory synthesis recognizing ing E. hartii as a congener of Artibeus (e.g., Koopman
monophyly of the group as a whole and classifying all 1985,1993,1994; Jones etal. 2002). We follow Baker
three lineages within genus Artibeus (Simmons 2005). et ah (2003) in recognizing E. hartii in its own subtribe
Further, the distinction of Enchisthenes and its distant Enchisthenina separate from subtribes Artibeina, Ecto-
relationship to the artibeine bats is well documented phyllina, and Stenodermatina.
(e.g., Andersen 1906; Van Den Bussche et al. 1993;
Baker et al. 2000; Wetterer et al. 2000). Within Artibeina, our analyses indicate two main
lineages in accordance with monophyly of the smaller
Our separate and combined analyses of cyto¬ Dermanura species and larger Artibeus species. Arti¬
chrome-^ and 16S rRNA sequences strongly support beus concolor is sister to the large species of Artibeus
a clade containing all sampled individuals referable to rather than sister to Dermanura (Figs. 1-3). Although
Artibeus, Dermanura, and Koopmania to the exclusion these relationships received different levels of statistical
of other sampled stenodermatine genera, including support in the separate analyses of cytochrome-# and
8 Occasional Papers, Museum of Texas Tech University
A. intermedius TK101996
A. lituratus TK104525
A. intermedius/
A. lituratus TK104112
100 A. lituratus
100 " A. lituratus TK128642
100 “ A. intermedius TK101993
0.01 substitutions/site £ TK15013
100 TK143051 A. planirostris
100
TK104410
100
100 1001 TK82842 |
A. schwartzi
10011 TK82839
100
100 100 L TK86531 | A. obscurus
95 100 - TK17080/TK17308
99 ,100
100 — TK135290 |
A. jamaicensis
97 100 - TK27682
97
100 90 ■TK134686
57
100 TK135760 A. fraterculus
100
■ TK135408
H ioo
100 _| TK150585 |
A. hirsutus
100 100 100 ""■tKI 50598
77 94 100
i TK101201
69 86
100 - TK101202 A. inopinatus
100
1 TK101203
100
-TK145271 — A. concolor
I TK19380
100 1 TK19381 D. bogotensis
100
100 ' TK19379
100
96 100 -TK104116 |
100 97 D. gnoma
100 - TK104117
100
99
100 100 TK104136 |
-C D. glauca
100 TK104203
100
100 TK134526
100 100 TK134611 D. rava
99 10C 100 TK104592
100
94
- NK14319 — D. anderseni
78
- TK18790 — D. cinerea
,_|TThK 82899
(P TThK 136035
D. tolteca
100 I__
TK136023
100 100 9988 - TK82900
100 100 r TK136188 |
100 D. phaeotis
100 TK82894
100
■ TK82897
100
100 - TK82901 D. azteca
89 100
100 98 100 TK82898
98 100 _r TK135691 |
90 100 100 = TK104509 D. rosenbergi
100
-TK136988 — D. watsoni
- TK16395 - Ectophylla
- TK22690 / TK55331 — Enchisthenes
-TK46006 - Uroderma
TK25052 / TK70524 — Chiroderma
Figure 3. Maximum likelihood phylogram (Lnl = -15,882.18) from analysis of combined cytochrome-# and 16S
rRNA sequences (2,635 base pairs) using best-fit model (GTR + T + I). We designated Chiroderma, Ectophylla,
and Uroderma as outgroups. Numbers above branches are Bayesian posterior probabilities, whereas those below
are bootstrap percentages from Maximum Likelihood and Parsimony, respectively. Values are shown only for nodes
supported by P > 0.95 or bootstrap percentage > 50, or both. “A” =Artibeus, “D.” = Dermanura.
Hoofer et al.—mtDNA Phylogeny of Artibeine Bats 9
16s rRNA sequences, they were depicted in all analy¬ terms of a molecular timescale of divergence of steno-
ses and highly supported in the combined sequence dermatine genera. According to Baker et al. (in litt.),
analysis (Fig. 3). As with previous morphological, Artibeus and Dermanura diverged in the Late Miocene
karyotypic, allozymic, and molecular evidence (Baker (6.3 mya) along with most of the vampyressine genera
1973; Straney etal. 1979; Van Den Busscheetal. 1993, (Chiroderma, Mesophylla, Platyrrhinus, Uroderma,
1998; Wetterer et al. 2000), our results provide no sup¬ Vampyressa, Vampyriscus, and Vampyrodes), predat¬
port to the objective argument of polyphyly that Owen ing the Pliocene divergence of the white-shouldered
(1991) used to justify recognizing A. concolor in the stenodermatine genera (Ametrida, Ardops, Ariteus,
genus Koopmania. We therefore follow the suggestion Stenoderma, Centurio, Pygoderma, and Sphaeronyc-
of Van Den Bussche et al. (1998) and the classifica¬ teris). This divergence estimate fits the criteria for
tion of Baker et al. (2003) in recognizing Koopmania genus ranking in the Age Related Classification system
concolor as Artibeus concolor. proposed for Euprimate taxa (Goodman et al. 1998).
Although the genetic distinction and sister-taxon Relationships within Artibeus and Dermanura.—
relationship between Artibeus and Dermanura is dem¬ Sister group relationships and alpha taxonomy within
onstrated in this and other studies, taxonomic status Artibeus and Dermanura continue to be conjectural,
for the two lineages as subgenera within Artibeus or and full revisions incorporating morphological and
as distinct genera is a matter of subjective ranking. molecular data are warranted for both genera. Although
Several authors have discussed this issue and ranked the not a primary focus of this study, the 16S rRNA data
lineages differently (e.g., Van Den Bussche et al. 1998; set offers robust resolution to and new insight into
Baker et al. 2000,2003; Wetterer et al. 2000; Lim et al. sister group relationships and questions of alpha tax¬
2004). Lim et al. (2004) noted that the smaller Der¬ onomy that have been debated in the morphological
manura species and larger Artibeus species cannot be and cytochrome-# literature. We briefly discuss some
diagnosed 100% on the basis of size alone because there of them.
is overlap in forearm length measurements between D.
aztecus (41-49 mm) and two species of Artibeus (con¬ Results from new cladistic analyses of morphol¬
color, 45-51 mm; inopinatus, 48-53 mm). Lacking any ogy and cytochrome-# sequences, focusing on species
diagnostic morphological characters, they recognized diversity within the enigmatic A.jamaicensis complex,
the two lineages as subgenera within Artibeus. Wet¬ have recommended species recognition for three of the
terer et al. (2000) also recognized them as subgenera 13 subspecies within A.jamaicensis (Simmons 2005):
(and Koopmania) within Artibeus because at that time planirostris (Patten 1971; Lim 1997; Guerrero et al.
there was no convenient way to refer to these taxa as a 2004; Lim et al. 2004; Larsen et al. 2007), schwartzi
monophyletic group if generic status was applied. On (Larsen et al. 2007); and triomylus (Guerrero et al.
the other hand, Solari et al. (2007) noted that Artibeus 2004; see also Larsen et al. 2007). Our analyses include
and Dermanura could be diagnosed on the basis of specimens referable to planirostris and schwartzi (but
wing coloration and dental features. not triomylus). In both cases, results from 16S rRNA
analysis mirror those from cytochrome-# in this and
We treat Artibeus and Dermanura as separate other studies, yet they provide even more robust sup¬
genera within the subtribe Artibeina following the port to the branching order. Our 16S rRNA results are
classification of Baker et al. (2003). This nomencla- best interpreted as evidence for species recognition of
tural arrangement facilitates convenient reference to A. planirostris and A. schwartzi as opposed to subspe¬
monophyly of the group as whole, recognition of both cies within A. jamaicensis. In avoiding paraphyletic
similarities and differences within it, and additional taxa, the latter would require the synonymy of at least
subgeneric classification within Artibeus and Derma¬ three other species within A. jamaicensis (amplus,
nura if warranted by future studies (see also Solari et lituratus, and obscurus). Thus, our 16S rRNA results
al. in prep, for additional arguments). Based on our affirm several studies of cytochrome-# for recognizing
results, the latter situation seems likely after contem¬ A. planirostris (Guerrero et al. 2004; Lim et al. 2004;
porary revisions are made of each genus with more Larsen et al. 2007), and affirm the suggestion by Larsen
data and taxa. Our arrangement also makes sense in et al. (2007) for recognizing A. schwartzi.
10 Occasional Papers, Museum of Texas Tech University
Our mtDNA data, along with those of Larsen graphic sampling, recommended species recognition
et al. (2007) and Lim et al. (2004), document a well for D. bogotensis and D. rosenbergi, former junior
supported sister relationship between the clade com¬ synonyms of D. glauca, and species recognition for
posed of A. fraterculus, A. inopinatus, and A. hirsutus D. rava, a former junior synonym of D. phaeotis. Our
and that of A. jamaicensis, A. lituratus, A. obscurus, analyses include specimens referable to all of these
A. planirostris, and A. schwartzi (Figs. 1, 2, 3). This taxa. In each case, our results from 16S rRNA analysis
observation has biogeographic significance, supporting mirror those from cytochrome-6 in this study and Solari
the hypothesis of Patterson et al. (1992) for an histori¬ et al. (in prep.), supporting a sister relationship between
cal connection between the biota of Middle America D. bogotensis and D. gnoma, another between D. rosen¬
and Western Andean Slope. Artibeus inopinatus and bergi and D. watsoni, and a clade containing D. rava,
A. hirsutus are distributed in xeric regions along the D. anderseni, and D. cinerea. Our 16S rRNA results
western and southern coasts of Middle America and are best interpreted as evidence for species recognition
their closest South American relative, A. fraterculus, of D. bogotensis and D. rosenbergi, rather than junior
is distributed in dry regions of southern Ecuador and synonyms oLD. glauca, and species recognition forD.
northern Peru west of the Andes Mountains. The rava, rather than a junior synonym of D. phaeotis. To
remaining species of Artibeus are sister to these xeric avoid paraphyletic taxa, the alternative classification
adapted species, and represent a South American radia¬ (Simmons 2005) would require synonymizing from
tion within the genus. one to nine other species and major taxonomic rear¬
rangement. Thus, we follow Solari et al. (in prep.) in
Results from 16S rRNA analysis also affirm recognizing 12 species within Dermanura, the nine
previous morphological (Marques-Aguiar 1994) and listed in Simmons (2005; we did not sample incomitata)
cytochrome-6 (Van Den Bussche et al. 1998; Lim et al. plus D. bogotensis, D. rava, and D. rosenbergi.
2004) analyses that suggested recognizing A. interme-
dius as a junior synonym of A. lituratus. Average 16S Our hypotheses of relationship for species di¬
rRNA sequence distance between A. intermedius and versity and species groups within Dermanura depart
A. lituratus (0.81%) is nearly equivalent to the average significantly from previous hypotheses for the genus,
distance within other Artibeus species (0.78%) and including Handley (1987). Like cytochrome-6, our
much less than that observed between species (4.62%). 16S rRNA results correspond with geographic origin
These results are of course provisional given the fact of Dermanura species better than with morphological
that we examined 16S rRNA sequences from just two similarity. Accordingly, we conclude that our system¬
individuals of intermedius (from Copan, Honduras) and atic and taxonomic hypotheses better reflect actual
three individuals of lituratus (from western Ecuador phyletic relationships rather than adaptive similar¬
and Union Island, St. Vincent and the Grenadines). ity. This is evidenced by biogeographic patterns in
However, they agree with the cytochrome-6 results the Dermanura phylogeny that correspond well with
from this and other studies that included more indi¬ diversification patterns hypothesized for Artibeus as
viduals. Therefore, we follow Marques-Aguiar (1994) well as other vertebrates (see Solari et al. in prep, for
in recognizing A. intermedius as a junior synonym of discussion of Dermanura phylogeography).
A. lituratus pending further study of combined mor¬
phological and molecular characters for populations In summary, our phylogenetic analysis of
of intermedius and lituratus, including those from the cytochrome-6 data includes fairly dense and complete
hypothesized region of sympatry in Middle America taxonomic sampling for both genera and most recog¬
(Davis 1984; Marques-Aguiar 1994). nized species within them. More importantly, analysis
of 16S rRNA sequences offers a new test of previous
Even fewer cladistic analyses have been under¬ hypotheses about shared common ancestry, sister group
taken examining species diversity within Dermanura relationships, and alpha taxonomy, thereby facilitating
(morphology, Owen 1991; cytochrome-6, Van Den knowledge about the congruence in variation between
Bussche et al. 1998). A new study by Solari et al. the well studied cytochrome-6 gene and the evolution¬
(in prep.), incorporating both morphological and ary history of bats within Artibeus and Dermanura. Our
cytochrome-6 analyses and dense taxonomic and geo¬ results illustrate a high degree of congruence between