Supplementary materials
Supplementary Table 1. State:step table based on maximum parsimony optimization of all
morphological characters (see also Figure 2).
Supplementary Table 2. Amplification and sequencing primers used in this study.
Gene
Forward primer
Reverse primer
thy
GGGTATGTAGTTCATCTTACTTC
GGCATCCTGGTATTTCTACAGTCTTG
550
stat5a
CTGCTCATCAACAAGCCCGA
GGCTTCAGGTTCCACAGGTTGC
580
plcb4
GTGAAATTGGAAGCCGAGAT
CACCAAGCTCATTTACTTGTGA
369
bdnf
CATCCTTTTCCTTACTATGGTT
TTCCAGTGCCTTTTGTCTATG
600
bdnf
*CCAGGTGAGAAGAGTGATGAC
*AGTGCCGAACTACCCAGTCGTA
600
ttn6 first half
CTTGACCTKGATTCYGAAYTAAGGAAAGGG †ACCTTTGCACCTCCATCRATGATGG
700
ttn6 first half
†CACCTCTCTTGTTCTTGACAATG
*ACCTTTGCACCTCCATCRATGATGG
700
ttn6 second half
†GARATCASTTGGTCTCGAGAAGAAGG
GCATTACAGACTTTGGATTCAGC
600
ttn6 second half
†GARATCASTTGGTCTCGAGAAGAAGG
*GATAAACCTGATCCTCCAAAAGG
600
rag2
TCATGGAGGGAAAACACCAAA
TGCACTGGAGACAGAGATTC
485
atp7a
TCCCTGGACAATCAAGAAGC
AAGGTAGCATCAAATCCCATGT
700
cytb first half
CGACYAATGACATGAAAAATCACCGTTG
GTAGGGGTGRAATGGAATTAKGTC
720
TTCCCTTTKCYGGTTTACAAGAC
588
cytb second half CACGAAACAGGATCYAACAAYCC
Expected Length (bp)
*These primers were designed for this study.
† Since the lengths of the amplification targets for genes ttn6 is ~1300 bp, internal primers were designed to make bidirectional
sequencing possible using Sanger technology.
Supplementary Table 3. Touchdown PCR amplification conditions used for amplifying all
loci except thy.
Step
Temperature (oC) Time (s) Number of cycles
Step
Initial denaturation
95
180
Touchdown amplification
2/temperature
Denaturation
95
45
Annealing
60/58/56/54
90
Extension
72
90
Amplification
30
Denaturation
95
45
Annealing
52
90
Extension
72
90
72
600
Final Extension
Supplementary Table 4. PCR amplification conditions used for amplifying thy.
Step
Step
Initial Denaturation
Temperature (oC) Time (s) Number of cycles
95
180
Amplification
Final extension
30
Denaturation
95
45
Annealing
50
90
Extension
72
90
72
600
Supplementary Table 5. PCR amplification reagent mix used in this study.
Reagent
Volume (ul)
PCR buffer 10X
3.0
MgCl2 (25mM)
3.0
Taq polymerase
0.3
dNTPs (10mM each)
0.5
Nuclease-free H20
14.2
Forward Primer
3.0
Reverse Primer
3.0
Template DNA
3.0
Total
30.0
Supplementary Table 6. Node, identity and description of characters that strongly supported nodes inferred to be incorrect.
Node
Character in Description
morphological
matrix
Nectar-feeding
21 Upper C crown proportions not (0); anteroposteriorly elongated (1).
clade
Neonycteris in
Micronycteris
23 Upper C anteromedial accessory cusp no raised cingulum or cusp (0); raised cingulum
present but cusp absent (1); cusp present but low (2); cusp tall, greater than half height of
main cusp (3).
Lonchorhina in
Phyllostominae
Lonchorhina in
25 Upper C lingual cingulum lingual cingulum absent (0); faint lingual cingulum present (1);
narrow well-developed cingulum present (2); wide well-developed cingulum present (3).
31 Upper C distal cingulum absent (0); cingulum small (1); large (2).
Phyllostominae
Neonycteris in
Micronycteris
46 Upper P4 number of roots three roots (0); two roots (1); one root (2).
Neonycteris in
Micronycteris
52 Upper P4 postparacrista extent postparacrista decreases sharply in height near paracone
and does not extend to distal edge of tooth (0); postparacrista extends as a continuous
crest to the distal aspect of the tooth (1).
Neonycteris in
Micronycteris
Nectar-feeding
53 Upper P4 distal accessory cusps distal accessory cusps absent (0); one distal accessory
cusp present (1); two distal accessory cusps (2).
58 Upper P5 anterior margin narrow (0); wide (1).
clade
Neonycteris in
Micronycteris
59 Upper P5 lingual cingular cusp cingulum with raised edge but no cingular cusp (0);
lingual cusp weakly developed (1); lingual cusp well developed (2); two well developed
lingual cusps (3).
Neonycteris in
62 Upper P5 anterolabial cusp absent (0); weakly developed (1); strongly developed (2).
Micronycteris
Neonycteris in
Micronycteris
Neonycteris in
66 Upper P5 proportions in occlusal view P5 length less than width (0); length and width
subequal (1); P5 length greater than width (2).
70 Upper molars ectocingulum development strongly developed (0); present but weak (1);
Micronycteris
Lonchorhina in
absent (2).
73 Upper M1 and M2 lingual cingulum absent (0); present (1).
Phyllostominae
Lonchorhina in
Phyllostominae
Lonchorhina in
77 Upper M1 and M2 metacone and paracone relative height metacone taller than paracone
(0); metacone and paracone subequal in height (1); metacone shorter than paracone (2).
82 Upper M1 and M2 trigon width and depth wide and shallow (0); narrow and deep (1).
Phyllostominae
Lonchorhina in
Phyllostominae
109 Upper M1 hypocone shelf development hypocone shelf very large, maximum mesiodistal
lenght equal to or greater than distance between paracone and metacone (0); hypocone
shelf large, maximum mesiodistal length greater than half the distance between paracone
and metacone but not 100% of that distance (1); hypocone shelf small, well developed
and clearly distinct from trigon, maximum mesiodistal length less than half the distance
between the paracone and metacone (2); hypocone shelf very small, present as a cingulum
but may not be clearly distinguished from the trigon (3); hypocone shelf absent (4).
Nectar-feeding
138 Upper M3 premetacrista development premetacrista longer than postparacrista (0);
clade
premetacrista and postparacrista length subequal (1); premetacrista one-half to three
quarters the length of the postparacrista (2); premetacrista one-third to slightly less than
one-half the length of the postparacrista (3); premetacrista present but less than one-third
the length of the postparacrista (4); premetacrista absent (5).
Nectar-feeding
clade
Nectar-feeding
clade
Nectar-feeding
clade
139 Upper M3 metacone metacone present as a disinct cusp (0); metacone indistinct,
incorperated into premetacrista (1); metacone entirely absent (2).
158 Lower c anterior diastema absent (0); diastema present between canine and last lower
incisor (i1, i2, or i3) (1).
159 Lower c tip divergence lower canines not laterally divergent (0); canines laterally
divergent, shafts slanted outward so that the tip of each tooth lies dorsal or dorsal and
lateral to the lateral edge of the root where the tooth enters the alveolus (1).
Neonycteris in
174 Lower p4 number of roots three roots (0); two roots (1); one root (2).
Micronycteris
Neonycteris in
Micronycteris
178 Lower p4 overall morphology p4 tribosphenic with a distinct trigonid and talonid (0); p4
premolariform, with cusps but not arranged in tribosphenic pattern (1); p4 peglike or
nearly featureless (2).
Nectar-feeding
179 Lower p4 anterolingual cuspule absent (0); cuspule present (1).
clade
Nectar-feeding
181 Lower p4 distal cuspule absent (0); cuspule present (1).
clade
Nectar-feeding
clade
Neonycteris in
Micronycteris
Lonchorhina in
183 Lower p4 occlusal outline long and strongly laterally compressed (length/width > or =
1.7) (0); short and broad (length/width < 1.7) (1).
187 Lower p4 and p5 crown morphology crown morphology of p4 and p5 similar (0); p4 and
p5 with different crown morphologies (1).
202 Lower p5 protocristid absent (0); one protocristid (1); two protocristids (2).
Phyllostominae
Lonchorhina in
Phyllostominae
Lonchorhina in
Phyllostominae
205 Lower p5 protoconid development less robust, taller, sharper (0); more robust, shorter,
more blunt (1); extremely short (2).
250 Lower m1 hypoflexid depth deep (0); shallow (1); absent (2).
Lonchorhina in
Phyllostominae
Neonycteris in
267 Lower m3 labial cingulid development strongly developed (0); weakly developed (1);
absent (2).
276 Lower m3 hypoconulid presence present (0); absent (1).
Micronycteris
Nectar-feeding
clade
278 Lower m3 talonid width talonid wider than trigonid (0); talonid and trigonid subequal (1);
talonid narrower than trigonid (2).
Supplementary Figure 1. Summary phylograms from maximum parsimony, maximum
likelihood, and Bayesian analyses of morphological data. Branches collapsed into triangles
had maximum support, † identifies fossil genera or species, non-monophyletic genera are in
bold and outgroups are shown in gray font. Percent bootstrap support or posterior
probabilities <100 are shown above or below internal branches, branch and number colors
are scaled to branch support. A. Majority-rule consensus of 411 most parsimonious trees
(length = 2888 steps). B. Maximum likelihood phylogeny all morphological characters (loglikelihood = -13800). Dashed line indicates branch is not shown to scale. C. Consensus
phylogram of all compatible bipartitions in Bayesian posterior trees from 4 independent runs
(6 chains total). The harmonic mean of the posterior log-likelihood of the model given the
data was -10583.
Supplementary Figure 2. Summary phylogram of posterior trees from 4 independent runs
(8 chains total) of Bayesian analyses of DNA sequence data. The harmonic mean of the
posterior log-likelihood of the model given the data was -103597. Posterior probabilities
<1.00 are shown below or above internal branches, branch and number colors are scaled to
Bayesian posterior probability. Non-monophyletic genera are in bold, outgroups are shown
in gray font.