Journal of Fish Biology (2014) 85, 228–245
doi:10.1111/jfb.12408, available online at wileyonlinelibrary.com
Mitochondrial DNA phylogeography of the Labeobarbus
intermedius complex (Pisces, Cyprinidae) from Ethiopia
K. A. Beshera* and P. M. Harris
Biodiversity and Systematics, Department of Biological Sciences, Box 870345, The University
of Alabama, Tuscaloosa, AL 35487-0345, U.S.A.
(Received 7 May 2013, Accepted 14 March 2014)
Mitochondrial DNA phylogeography of populations of the Labeobarbus intermedius complex
(hexaploid barb) was investigated using 88 complete and 71 partial cytochrome b (cytb) sequences
originating from 21 localities in ive major drainages in Ethiopia and two localities in northern
Kenya. The samples included 14 of the 15 Labeobarbus species described from Lake Tana. Discrete
phylogeographic analyses of 159 cytb sequences employing Bayesian Markov Chain Monte Carlo
(MCMC) simulations using Bayesian evolutionary analysis by sampling trees (BEAST) supported
the monophyly of the L. intermedius complex, including the Lake Tana species. This analysis, in
combination with statistical parsimony analysis, identiied two mitochondrial DNA lineages within
the complex. Divergence dating employing coalescent simulations suggested that the geographic
split in the L. intermedius complex that led to the formation of these lineages occurred during the
Pleistocene (c. 0⋅5 M b.p.), consistent with the timing of volcano-tectonic events postulated to have
shaped the current landscape of East Africa.
© 2014 The Fisheries Society of the British Isles
Key words: East Africa; haplotypes; hexaploid minnow; lineages; Pleistocene; Rift Valley.
INTRODUCTION
Labeobarbus intermedius (Rüppell 1836) is a large hexaploid barb from East Africa
well known for its extensive morphological diversity (Banister, 1973). Since its
description, the taxonomic limits of L. intermedius have not been well established, and
there have been only two studies (morphology, Banister, 1973; molecular, de Graaf
et al., 2010) with relatively wider geographic sampling that examined intraspeciic
variation, although this was not the focus of the latter study. The taxonomic uncertainty
surrounding L. intermedius has prompted several authors (Nagelkerke & Sibbing,
1996; de Graaf et al., 2008, 2010) to refer to it as L. intermedius complex. The name
Labeobarbus intermedius complex is used throughout this study.
The type locality of L. intermedius complex is Lake Tana in the north western highlands of Ethiopia (Rüppell, 1836), but the redescription of the species by Banister
(1973) was based on the examination of 454 specimens collected from lakes and rivers
across Ethiopia and Lake Baringo, Kenya (0∘ 37′ N; 36∘ 04′ E). Thus far, no molecular
studies compared the nominal taxon L. intermedius with the sympatric and allopatric
*Author to whom correspondence should be addressed at present address: Wallace Community College, 1141
Wallace Dr., Dothan, AL 36303, U.S.A. Tel.: +1 3345562637; email: kabeshera@crimson.ua.edu
228
© 2014 The Fisheries Society of the British Isles
PHYLOGEOGRAPHY OF LABEOBARBUS INTERMEDIUS
229
Labeobarbus species. For most of its taxonomic history, the L. intermedius complex
was classiied in the genus Barbus. Recently, Skelton (2001, 2002) reclassiied the
large hexaploid barbs of Africa into Labeobarbus based on karyological (Golubstov &
Krysanov, 1993) and phylogenetic (Machordom & Doadrio, 2001) evidence. Throughout this study, the taxonomic recommendation of Skelton (2001, 2002) is adopted.
The existence of several morphologically distinct forms in Lake Tana led to multiple
species descriptions in the early literature (e.g. six by Rüppell, 1836; 10 by Boulenger,
1902, 1907, 1911; 10 species and 23 sub-species by Bini, 1940). The earliest comprehensive taxonomic account of L. intermedius complex was based on Banister’s (1973)
revision of the large barbs of East and Central Africa. In his revision, Banister (1973)
synonymyzed >50 nominal species and sub-species in East Africa, including those
from Lake Tana, into the L. intermedius complex (Golubstov et al., 2002). Banister
(1973) also recognized two sub-species: Labeobarbus intermedius intermedius (Rüppell 1836), widely distributed throughout Ethiopia and northern Kenya and Labeobarbus intermedius australis (Banister 1973) found in Lake Baringo, Kenya. Several
authors subsequently regarded Banister’s (1973) taxonomic conclusions as putative, at
best. Skorepa (1992) described Banister’s (1973) sub-speciic separation of L. i. australis as ‘unreal’, although he maintained Banister’s (1973) synonymy of several large
barb species and sub-species with the L. intermedius complex. Nagelkerke & Sibbing
(1997, 2000) questioned Banister’s (1973) hypothesis of a single species of large barb
and described 14 species from Lake Tana distinct from the L. intermedius complex.
Golubstov et al. (2002) suggested that populations of the L. intermedius complex from
the Ethiopian Rift Valley represent multiple evolutionary lineages.
The L. intermedius complex is widely distributed throughout Ethiopia and also enters
into northern Kenya (Banister, 1973). In Ethiopia, the L. intermedius complex is found
in all the major basins including the Rift Valley lakes, the Omo, Genale, Wabi Shebele,
Awash and Abay (including Lake Tana) River basins (Banister, 1973; Roberts, 1975).
This complex has also been reported from the Baro River system, a tributary of the
White Nile River, within the boundaries of Ethiopia (Krysanov & Golubstov, 1996).
In Kenya, the L. intermedius complex has been recorded from the Uasso Nyiro River
and Lake Turkana (3∘ 30′ N; 36∘ 00′ E) basins (Skorepa, 1992), as well as from Lake
Baringo (Banister, 1973).
H Y D R O G R A P H I C F E AT U R E S O F E T H I O P I A
The combined effects of faulting, uplifting and volcanism during the Tertiary period
were the primary geologic forces shaping the current hydrographic features within
Ethiopia (Mohr, 1966). These tectonic processes formed the Ethiopian Rift Valley, the
northernmost branch of the East African Rift System, creating a division between the
highlands of the north-west and south-east (Fig. 1). Three distinct drainage patterns
occur in the river basins of the north-western highlands. Most rivers lying on the northern and western escarpment of these highlands (Abay, Barbo and Tekeze River basins)
low to the west and form part of the Blue Nile River drainage. Rivers originating from
the eastern or north-eastern escarpment of the highlands low east into either the endoreic basins of the Rift Valley or Awash River. The headwaters of the Awash River are on
the south-eastern side of the escarpment; this river lows north-east through the Danakil
Depression into Lake Abe. Rivers on the southern side of the escarpment low south
into Lake Turkana via the Omo River system. Rivers originating from the south-eastern
© 2014 The Fisheries Society of the British Isles, Journal of Fish Biology 2014, 85, 228–245
230
K . A . B E S H E R A A N D P. M . H A R R I S
ea
dS
Re
Eritrea
Yemen
Tekeze Basin
ti
Danakil Basin
ou
Jib
G
D
Abay Basin
20
Rif
sin
t Va
lley
Ba
Ba
sin
a
Lak
es
le
be
South Sudan
13
18
14
15
16
17
GBL
e
Sh
9
Omo Basin
i
ab
W
GBL12
Og
ad
en
20
19
8
11
ma
li
7
10
6
So
Ethiopia
ANL
Barro-Akobo Basin
den
fA
o
ulf
asin
Sudan
3
1 2 4
5
Aw
ash
B
Lake Tana
Genale-Dawa Basin
Lake Turkana
Uganda
Kenya
Fig. 1. Map showing major drainages in Ethiopia, sampling localities of Labeobarbus intermedius complex (with
some points offset for clarity) and distribution of lineages ( , lineage A; , lineage B). Sampling site codes
are indicated in Table I. The geographic position of sampling localities 19, 20 and 21 were approximated.
ANL, Addis Ababa-Nekemt Lineament; GBL, Goba-Bonga Lineament.
highlands low south-east (Genale, Dawa, Wabi Shebele and Fafan Rivers) into Somalia. The major hydrographic features of the Rift Valley are a chain of endoreic lakes.
These lakes, together with the Awash River system, form three major basins within
the Ethiopian Rift Valley, i.e. (1) the Awash River drainage in the north, (2) two systems of linked lakes in the central Rift Valley (Zeway-Langano-Abiyata-Shala and
Awasa-Shallo) and waters lowing into and out of these lakes and (3) Lakes Abaya,
Chamo and Chew Bahir and their tributaries in the south. It has been hypothesized that
in the late Tertiary, these endoreic lakes formed a continuous system (Mohr, 1966) but
during the pluvial period, which lasted from Late Glacial into early Holocene, only
the four northern Rift lakes (Lakes Abiyata, Langano, Shala and Zeway) were uniied
into a larger lake, which drained to the north into the Awash River (Grove et al., 1975).
During the same period, the Chew Bahir basin had an overlow into Lake Turkana,
while the latter was connected with the White Nile system via the Baro-Akobo (Sobat)
River basin (Grove et al., 1975; Grove, 1983). Lake Turkana occurs in the northern
Kenyan Rift, which is connected with the southern Ethiopian Rift Valley by a 300 km
wide zone of overlap (Ebinger et al., 2000).
Despite its broad geographic distribution and potential value as a model organism to study the diversiication of aquatic organisms in East Africa, phylogenetic
relationships within the L. intermedius complex have not been fully clariied and
very little is known about the geographic distribution of its genetic diversity. A
previous phylogenetic analysis (de Graaf et al., 2010) of the complex was based on
limited geographic sampling and, therefore, was insuficient to draw irm conclusions
© 2014 The Fisheries Society of the British Isles, Journal of Fish Biology 2014, 85, 228–245
PHYLOGEOGRAPHY OF LABEOBARBUS INTERMEDIUS
231
on its phylogeography suggesting the need for a comprehensive phylogeographic
analysis. The aims of this study were to show the distribution of genetic variation
within the L. intermedius complex across its geographic range based on the analysis
of complete cytochrome b (cytb) gene sequences (1141 bp) and test if (1) extant
pattern of genetic variation in the L. intermedius complex relects histories of river
drainages, (2) phylogeographic structure of populations corresponds to hypothesized
previous basin connections summarized above and (3) phylogeographic structure of
the L. intermedius complex corresponds to its taxonomy.
MATERIALS AND METHODS
SAMPLING
Tissue samples of the L. intermedius complex (including the species lock of Labeobarbus
from Lake Tana) for this study were collected from 13 water bodies in four major drainages in
Ethiopia between January and May 2006. Fishes were collected using gillnets, and assistance
was provided by local isheries experts in the identiication of specimens. Additional tissue samples of the L. intermedius complex representing four water bodies were obtained from museum
collections. All tissues were preserved in 95% ethanol. Specimens examined in this study, basin
and water body sampled, sampling localities (for samples collected for this study), GenBank
accession numbers and number of individuals examined for each water body are summarized in
Table I.
D NA E X T R AC T I O N A N D S E QU E N C I N G
Total genomic DNA was extracted using the DNeasy tissue extraction kit (Qiagen;
www.qiagen.com) following the manufacturer’s instructions. The mitochondrial (mt) cytb
gene (1141 bp) was ampliied using primers and PCR protocols developed by Briolay et al.
(1998). Sequences were generated via dye terminator reactions and read on an ABI 3100 prism
sequencer (www.lifetechnologies.com). Sequences were aligned independently by eye using
Bioedit (Hall, 1999). Complete cytb sequences of 43 unique mtDNA haplotypes were deposited
in GenBank and assigned accession numbers JN886992–JN887034 (Table I).
P H Y L O G E O G R A P H I C A N A LY S I S
The present data comprises 159 cytb sequences of the L. intermedius complex from 21
localities representing ive drainages across Ethiopia and two localities in Kenya. Of these, 86
sequences (1141 bp) were generated for this study while the remaining 73 sequences [71 partial
(GQ853201–GQ853271; de Graaf et al., 2010) and two complete (AF112406 and AF780872)
cytb sequences] were retrieved from GenBank. Phylogeographic pattern within the L. intermedius complex was inferred based on cytb data employing discrete phylogeographic analysis,
which integrates spatial, temporal and genealogy inference, under the Bayesian statistical
framework implemented in Bayesian evolutionary analysis by sampling trees (BEAST; Lemey
et al., 2009). In discrete phylogeographic analysis, no outgroup taxa were used. For estimation
of time to most recent common ancestor (TMRCA), ive sequences representing three taxa,
Labeobarbus bynni bynni (Forskål 1775), Labeobarbus bynni occidentalis (Boulengar 1911)
and Labeobarbus petitjeani (Daget 1962), were chosen based on the relationships generated by
a preliminary phylogenetic analysis of a wide range of related taxa and used as outgroups.
The BEAST input ile was generated using BEAUTI 1.4.6 (Drummond & Rambaut, 2007).
Later, necessary additions for Bayesian stochastic search variable selection (BSSVS) that
enables discrete state reconstructions and inference of most parsimonious description of
phylogeographic pattern (Lemey et al., 2009) were made using text editor. The geographic
distributions of sampling locations for cytb sequences were incorporated into the input ile as
prior speciications.
© 2014 The Fisheries Society of the British Isles, Journal of Fish Biology 2014, 85, 228–245
232
Table I. List of samples of Labeobarbus spp. examined in this study along with basin and water body sampled
Water body
Species
NS
NH
Abay
Abay River
Dedessa River
Labeobarbus intermedius
L. intermedius
5
30
1, 2, 3, 4
5–9, 32, 33, 34, 41–44
Gumara River
L. intermedius
4
2, 10, 21, 12
Lake Tana
Labeobarbus acutirostris
4
11, 28,
Labeobarbus brevicephalus
4
11, 29, 31
Labeobarbus crassibarbis
4
12,
Labeobarbus dainellii
Labeobarbus gorgorensis
2
4
11, 28
12, 37, 40
Labeobarbus gorguari
3
11, 28,
L. intermedius
9
4, 11, 13, 28
Labeobarbus longissimus
Labeobarbus macrophthalmus
2
4
5, 11
12, 28, 29
Labeobarbus megastoma
4
11, 28,
Labeobarbus Negdia
4
11, 12, 29, 31
GenBank accession number
JN886992–JN886995
JN886996–JN887001,
GQ53251–GQ53266
JN886993,
JN887002–JN887004
GQ850201–GQ850203,
JN887030
GQ853204, GQ853205,
JN887033, JN887003
GQ853206–GQ853208,
JN887004
GQ853209, GQ853210
GQ853211–GQ853213,
JN887034
GQ853214, GQ853215,
JN887029
GQ853236–GQ853238,
JN887003,
JN887005–JN887007
GQ853216, JN887006
GQ853217–GQ853219,
JN887031
GQ853220–GQ853221,
JN887033
GQ853223–GQ853225,
JN887031
Site number (Fig. 1)
5
6
4
2
1, 2
1
3
1
1
1
2
2
2
K . A . B E S H E R A A N D P. M . H A R R I S
© 2014 The Fisheries Society of the British Isles, Journal of Fish Biology 2014, 85, 228–245
Basin
Basin
Awash
Rift Valley
Water body
Species
NS
Labeobarbus platydorsus
5
Awash River
Borkena River
Lake Awassa
Labeobarbus Surkis
Labeobarbus truttiformis
Labeobarbus tsanensis
L. intermedius
L. intermedius
L. intermedius
2
4
4
5
1
8
Lake Langano
Bulbula River
Lake Abaya
Lake Chamo
L. intermedius
L. intermedius
L. intermedius
L. intermedius
5
1
2
8
L. intermedius
L. intermedius
L. intermedius
L. intermedius
L. intermedius
L. intermedius
L. intermedius
L. intermedius
L. intermedius
Labeobarbus intermedius australis
Labeobarbus byinni bynni
Labeobarbus bynni occidentalis
Labeobarbus petitjeanii
6
3
5
2
4
5
3
1
1
1
1
1
1
Kulfo River
ArbaMinch Springs
Sagoe River
Darse River
Omo
Gibe River
Gilgel Gibe River
Gojeb River
Wabi Shebele Arer River
Turkana
Lake Kamnarok (Kenya)
Lake Baringo Lake Baringo (Kenya)
Outgroup taxa Nile River, Egypt
Baing River, Guinea
Baing River, Guinea
Rift Valley
NH
GenBank accession number
11, 29, 30, 39 GQ853226–GQ853228, JN887031,
JN887032
11, 12
GQ853229, JN887003
11, 29, 38
GQ853230–GQ853232, JN887031
11, 12, 28
GQ853233–GQ853235, JN887003
16
JN887008, JN887009
16
GQ853271
16, 45
JN887008, JN887010,
GQ853244–GQ853245
14, 15
JN887011–JN887013
21
GQ853246
25
JN887026, JN887028
25, 26
JN887026, JN887027,
GQ853239–GQ853242
19, 20
JN887018, JN887019
16, 17, 18
JN887014–JN887017
19
JN887018
18, 19
JN887017, JN887018
17, 18
JN887016, JN887017, JN887020
22, 23, 24
JN887021–JN887023
17, 24
JN887024, JN887025
36
GQ853247
27
AF112406
4
AF180872
AF28742
AF180829, AF287421
AF287443, AF180875
Site number (Fig. 1)
2, 3
1
2
2
7
20
9
8
19
13
16
14
15
17
18
10
11
11
21
PHYLOGEOGRAPHY OF LABEOBARBUS INTERMEDIUS
© 2014 The Fisheries Society of the British Isles, Journal of Fish Biology 2014, 85, 228–245
Table I. Continued
N S , number of specimens; N H , haplotype number.
233
234
K . A . B E S H E R A A N D P. M . H A R R I S
The best performing evolutionary model was obtained based on the Akaike information criterion (AIC; Huelsenbeck & Crandall, 1997) using MODELTEST 3.7 (Posada & Crandall,
1998). Results from MODELTEST were applied to set model parameters for discrete phylogeographic analyses and estimation of TMRCA. Analyses were performed employing BEAST via
the freely available BIOPORTAL platform (Kumar et al., 2009; www.bioportal.uio.no) with the
GTR nucleotide substitution model, an uncorrelated lognormal relaxed molecular clock model,
and a Bayesian coalescent model assuming constant population size. A generalized teleost cytb
substitution rate of 0⋅76–2⋅20% per million years was used as a uniform prior (Berendzen et al.,
2008) in all analyses. Because estimates of Labeobarbus-speciic fossil dates are not available,
no calibration points were used to root the trees.
For discrete phylogeographic analysis, ive independent Markov Chain Monte Carlo (MCMC)
simulations each with 200 million generations were run sampling trees every 20 000th generations; in TMRCA estimation, 50 million generations were run sampling every 10 000th
generations. Results from the ive runs were combined using LogCombiner 1.6.2 (Rambaut &
Drummond, 2011a). To conirm stationarity and determine the mean and 95% highest posterior
density (HPD) for TMRCA and other statistics, outputs of the runs were viewed using Tracer
1.5 (Rambaut & Drummond, 2009). TreeAnotator 1.6.2 (Rambaut & Drummond, 2011b) and
FigTree 1.3.1 (Rambaut, 2010) were used, respectively, to summarize the posterior tree distribution and visualize the annotated maximum clade credibility (MCC) tree.
The number of unique haplotypes and haplotype (H d ± s.d.) and nucleotide (pi ) diversities
were calculated in DNASP 4.5 (Rozas et al., 2003). Haplotype parsimony networks were constructed in TCS (Clement et al., 2005) using cytb haplotype dataset comprising 45 unique haplotype sequences employing statistical parsimony, a genetic algorithm introduced by Templeton
et al. (1992). In this analysis, maximum numbers of mutational steps that make parsimonious
connections between haplotype sequences were calculated with 95% conidence. A matrix of
pair-wise uncorrected p-distances among haplotypes was generated using phylogenetic analysis
using parsimony* (PAUP*) based on maximum likelihood settings generated in MODELTEST
(Posada & Crandall, 1998). Genetic divergences between and within lineages were calculated
based on this matrix.
RESULTS
S E L E C T I O N O F M O D E L O F N U C L E OT I D E S U B S T I T U T I O N
MODELTEST 3.7 (Posada & Crandall, 1998), based on AIC, determined that
the model that best described the evolution of the cytb sequence data in the L.
intermedius complex was the general time reversible (GTR-I; −lnL = 2188⋅4771,
AIC = 4394⋅9541, K = 9), proportion of invariant sites (pInv = 0⋅8322) and equal rates
for all sites.
PHYLOGEOGRAPHY
The MCC tree generated by discrete phylogeographic analyses (Fig. 2) strongly
supports the monophyly of the L. intermedius complex (posterior probability,
PP = 100%). An interesting discovery was that the L. intermedius complex was split
into two mtDNA lineages, lineage A (PP = 58%) and lineage B (PP = 96%). Of the
159 sequences examined, 118 (74%) fell into lineage A while the remaining 41
sequences (26%) fell into lineage B. Lineage A consisted of sequences geographically
originating from the Abay (Blue Nile) River including Lake Tana (localities 1–6),
Awash River (localities 7 and 20), northern Rift Valley Basins (NRV; localities 8 and
9) and Lake Baringo (Kenya). Sequences that originated from Omo River (localities
10–12), southern Ethiopian Rift Valley (SRV; localities 13–18), Wabi Shebele River
© 2014 The Fisheries Society of the British Isles, Journal of Fish Biology 2014, 85, 228–245
235
PHYLOGEOGRAPHY OF LABEOBARBUS INTERMEDIUS
100
Dedessa River
100
Awash River and
NRV lakes
Lineage A
58
Dedessa River
Lake Baringo
91
Lake Tana
100
Lake Tana
Lake Tana
Gilgelgibe River
Lineage B
96
Lakes Abaya
and Chamo
100
Lake Kamnarok
80
100
80
60
40
20
Kulfo and
Sagoe Rivers
0
Fig. 2. Maximum clade credibility tree obtained from Bayesian analysis of cytochrome b (cytb) sequences, showing two lineages (lineages A and B) and relationships among populations of Labeobarbus intermedius
complex. Branch supports (posterior probability values) are indicated above branches. Scale shows branch
length.
(locality 21), Bulbula River (locality 19) and Lake Turkana Basins comprise lineage
B (Figs 1 and 2). An unexpected result was the placement of the lone sequence from
Bulbula River within lineage B.
The TCS analysis of mitochondrial data of the L. intermedius complex produced a
single network with all haplotypes connected. In this haplotype network (Fig. 3), there
were 23 mutational steps between the most distant haplotypes. The haplotype network
consisted of two mtDNA groups (Fig. 1), consistent with the phylogeny generated from
discrete phylogeographic analyses. The irst group contained haplotypes originating
from Abay River (haplotypes 1–13, 28–35 and 37–44), Awash River and NRV (haplotypes 14, 15, 35 and 45) drainages and Lake Baringo (haplotype 4, shared with Abay
River drainage) while the second group contained haplotypes originating from SRV
© 2014 The Fisheries Society of the British Isles, Journal of Fish Biology 2014, 85, 228–245
236
K . A . B E S H E R A A N D P. M . H A R R I S
13
1
28
37
29
12
7
31
8
42
9
33
41
10
2
43
30
44
40
3
35
4
15
38
11
34
14
5
6
45
39
32
26
25
17
16
20
19
18
24
27
22
21
23
36
Fig. 3. Statistical parsimony network for cytochrome b (cytb) sequences of 45 unique Labeobarbus intermedius
complex haplotypes connected to each other with 95% c.i. Circles correspond to unique haplotypes with
numbers in circles indicating haplotype number. Circle sizes relect the frequency of each haplotype. Lines
connecting circles (including the small circles) correspond to mutation steps. Geographic origins of haplotypes are colour-coded ( , Abay River drainage; , northern Rift Valley-Awash River drainage; , Omo
River drainage; , southern Rift Valley; , Wabi Shebele River drainage; , Lake Baringo (Kenya); ,
Turkana drainage (Kenya)).
(haplotypes 16–20, 25 and 26), Omo River (haplotypes 17 and 18 shared with SRV and
21–24), Wabi Shebele River (haplotype 36) and Lake Turkana (haplotype 27) basins.
Intralineage relationships were largely unresolved with mostly poorly supported
terminal nodes (PP < 50%). Nevertheless, some monophyletic groups were apparent
within lineages A and B: (1) the group comprising populations from the Awash River
and NRV lakes (PP = 100%), (2) the Lake Chamo + Lake Abaya group (PP = 100%)
© 2014 The Fisheries Society of the British Isles, Journal of Fish Biology 2014, 85, 228–245
PHYLOGEOGRAPHY OF LABEOBARBUS INTERMEDIUS
237
and (3) Kulfo River + Sagoe River group (PP = 80%). The irst group fell within
lineage A while the last two fell within lineage B. All sequences of the species lock of
Labeobarbus from Lake Tana were placed within lineage A, but relationships among
the species were not resolved.
GENETIC DIVERSITY AND DIVERGENCE AND TMRCA
Mean sequence divergence between lineage A and lineage B, after excluding missing sites (due to partial cytb sequences retrieved from GenBank database), was 1⋅27%
while it was 0⋅93% using the whole dataset. According to Bayesian analysis (BEAST),
estimated divergence time between lineage A and lineage B was 0⋅5 million years
before present (M b.p.) [95% credible interval (c.i.): 0⋅22–0⋅98 M b.p.]. The ages of the
most recent common ancestors (TMRCAs) of lineages A and B were 0⋅24 M b.p. (95%
c.i.: 0⋅11–0⋅98 M b.p.) and 0⋅15 M b.p. (95% c.i.: 0⋅05–0⋅31 M b.p.), respectively.
A total of 45 unique mtDNA haplotypes were identiied among 159 sequences of
the L. intermedius complex: 32 haplotypes in lineage A (representing 118 sequences)
and 13 haplotypes in lineage B (representing 41 sequences). Overall, a relatively
high level of haplotype diversity (mean ± s.d. H d = 0⋅950 ± 0⋅007), but low level of
nucleotide diversity (pi = 0⋅00695) was observed. Haplotype (H d ) and nucleotide
(pi ) diversities within lineage A were 0⋅92 ± 0⋅012 and 0⋅0033 (mean ± s.d.),
respectively, while they were 0⋅87 ± 0⋅032 and 0⋅0028, respectively, in lineage
B. Within the L. intermedius complex, pair-wise haplotype sequence divergences
(expressed as per cent p-distances) varied from 0⋅08 to 1⋅65% (Table II) and average
within-lineage haplotype sequence divergences were 0⋅76% (lineage A) and 0⋅93%
(lineage B).
DISCUSSION
PHYLOGEOGRAPHY OF THE L. INTERMEDIUS COMPLEX
Discrete phylogeographic analysis of mtDNA sequence variation in the L. intermedius complex recovered two lineages: lineages A and B (Fig. 2). Lineage A was represented by sequences originating from Abay (Blue Nile) and Awash River drainages,
NRV lakes and Lake Baringo (Kenya), whereas lineage B comprised sequences from
Omo and Wabi Shebele River drainages, Lake Turkana basin and southern Ethiopian
Rift Valley populations. The geographic distributions of the two lineages of the L.
intermedius complex showed no overlap with the exception of haplotype 21, which
represents sequences originating from the Bulbula River in the NRV and Omo River
basin, suggesting historical isolation between the lineages. Haplotypes, however, were
shared extensively among drainages and populations within the two lineages. This pattern is exempliied by haplotypes 17 and 18, which were dispersed throughout the Omo
River and SRV drainages, while haplotype 14 was shared among populations of Awash
River and Lakes Awassa and Langano. There is paucity of information on the phylogeographic patterns of other co-distributed ish species of the region. Consequently, it
is not possible to determine whether the phylogeographic patterns within the L. intermedius complex relect a common pattern of evolution in aquatic taxa or are unique to
this species complex.
© 2014 The Fisheries Society of the British Isles, Journal of Fish Biology 2014, 85, 228–245
238
Table II. Pair-wise genetic distances (expressed as per cent uncorrected p-distances) in the mitochondrial cytochrome b gene between haplotypes of
Labeobarbus intermedius complex
HAP1 HAP2 HAP3 HAP4 HAP5 HAP6 HAP7 HAP8 HAP9 HAP10 HAP11 HAP12 HAP13 HAP14 HAP15 HAP16 HAP17 HAP18 HAP19 HAP20 HAP21 HAP22
–
0⋅35
0⋅44
0⋅44
0⋅79
1⋅05
0⋅61
0⋅35
0⋅53
0⋅70
0⋅44
0⋅09
0⋅44
0⋅61
0⋅70
1⋅32
1⋅23
1⋅32
1⋅40
1⋅32
1⋅40
1⋅49
1⋅49
1⋅32
1⋅40
1⋅49
1⋅40
0⋅18
0⋅35
0⋅61
0⋅18
1⋅22
0⋅68
0⋅61
0⋅09
0⋅09
0⋅44
0⋅70
0⋅26
0⋅18
0⋅18
0⋅35
0⋅09
0⋅26
0⋅44
0⋅26
0⋅35
0⋅96
0⋅88
0⋅96
1⋅05
0⋅96
1⋅05
1⋅14
1⋅14
0⋅96
1⋅05
1⋅14
1⋅05
0⋅35
0⋅18
0⋅26
0⋅35
0⋅92
0⋅29
0⋅31
0⋅18
0⋅35
0⋅61
0⋅35
0⋅26
0⋅26
0⋅44
0⋅18
0⋅35
0⋅53
0⋅35
0⋅44
0⋅88
0⋅79
0⋅88
0⋅96
0⋅88
0⋅96
1⋅05
1⋅05
0⋅88
0⋅96
1⋅05
0⋅97
0⋅44
0⋅26
0⋅35
0⋅44
0⋅82
0⋅39
0⋅20
0⋅53
0⋅79
0⋅35
0⋅26
0⋅26
0⋅44
0⋅18
0⋅35
0⋅53
0⋅35
0⋅44
1⋅05
0⋅96
1⋅05
1⋅14
1⋅05
1⋅14
1⋅23
1⋅23
1⋅05
1⋅14
1⋅23
1⋅14
0⋅44
0⋅26
0⋅35
0⋅44
1⋅02
0⋅38
0⋅41
0⋅96
0⋅70
0⋅44
0⋅61
0⋅79
0⋅53
0⋅70
0⋅88
0⋅70
0⋅79
1⋅23
1⋅14
1⋅23
1⋅32
1⋅23
1⋅32
1⋅40
1⋅40
1⋅23
1⋅32
1⋅40
1⋅32
0⋅79
0⋅61
0⋅70
0⋅79
1⋅22
0⋅58
0⋅20
0⋅96
0⋅88
0⋅88
0⋅88
0⋅79
0⋅96
1⋅14
0⋅96
1⋅05
0⋅61
0⋅53
0⋅61
0⋅70
0⋅79
0⋅70
0⋅79
0⋅79
0⋅61
0⋅70
0⋅79
0⋅70
1⋅05
0⋅88
0⋅96
1⋅05
0⋅10
1⋅07
0⋅92
0⋅44
0⋅09
0⋅61
0⋅35
0⋅53
0⋅70
0⋅53
0⋅61
1⋅23
1⋅14
1⋅23
1⋅32
1⋅23
1⋅32
1⋅40
1⋅40
1⋅23
1⋅32
1⋅40
1⋅32
0⋅61
0⋅44
0⋅53
0⋅61
1⋅22
0⋅58
0⋅61
0⋅35
0⋅53
0⋅26
0⋅26
0⋅44
0⋅44
0⋅53
1⋅14
1⋅05
1⋅14
1⋅23
1⋅14
1⋅23
1⋅32
1⋅32
1⋅14
1⋅23
1⋅32
1⋅23
0⋅35
0⋅35
0⋅44
0⋅35
1⋅12
0⋅29
0⋅31
0⋅53
0⋅26
0⋅44
0⋅61
0⋅44
0⋅53
1⋅14
1⋅05
1⋅14
1⋅23
1⋅14
1⋅23
1⋅32
1⋅32
1⋅14
1⋅23
1⋅32
1⋅23
0⋅53
0⋅35
0⋅44
0⋅53
1⋅12
0⋅48
0⋅51
0⋅26
0⋅61
0⋅79
0⋅61
0⋅70
0⋅96
0⋅88
0⋅96
1⋅05
0⋅96
1⋅05
1⋅14
1⋅14
0⋅96
1⋅05
1⋅14
1⋅05
0⋅70
0⋅53
0⋅61
0⋅53
1⋅12
0⋅67
0⋅71
0⋅35
0⋅53
0⋅35
0⋅44
1⋅05
0⋅96
1⋅05
1⋅14
1⋅05
1⋅14
1⋅23
1⋅23
1⋅05
1⋅14
1⋅23
1⋅14
0⋅44
0⋅26
0⋅35
0⋅26
1⋅02
0⋅39
0⋅41
0⋅35
0⋅53
0⋅61
1⋅23
1⋅14
1⋅23
1⋅32
1⋅23
1⋅32
1⋅40
1⋅40
1⋅23
1⋅32
1⋅40
1⋅32
0⋅09
0⋅26
0⋅53
0⋅09
1⋅12
0⋅58
0⋅51
0⋅70
0⋅79
1⋅40
1⋅32
1⋅40
1⋅49
1⋅40
1⋅49
1⋅58
1⋅58
1⋅40
1⋅49
1⋅58
1⋅49
0⋅44
0⋅44
0⋅70
0⋅44
1⋅33
0⋅67
0⋅71
0⋅09
1⋅23
1⋅14
1⋅23
1⋅32
1⋅23
1⋅32
1⋅40
1⋅40
1⋅23
1⋅32
1⋅40
1⋅32
0⋅61
0⋅44
0⋅53
0⋅61
1⋅12
0⋅58
0⋅51
1⋅32
1⋅23
1⋅32
1⋅40
1⋅32
1⋅40
1⋅49
1⋅49
1⋅32
1⋅40
1⋅49
1⋅40
0⋅70
0⋅53
0⋅61
0⋅70
1⋅22
0⋅67
0⋅61
0⋅09
0⋅18
0⋅09
0⋅18
0⋅26
0⋅35
0⋅35
0⋅18
0⋅26
0⋅35
0⋅26
1⋅14
1⋅14
1⋅23
1⋅32
0⋅61
1⋅36
1⋅23
0⋅09
0⋅18
0⋅26
0⋅18
0⋅26
0⋅26
0⋅09
0⋅18
0⋅26
0⋅18
1⋅05
1⋅05
1⋅14
1⋅23
0⋅51
1⋅27
1⋅13
0⋅09
0⋅18
0⋅26
0⋅35
0⋅35
0⋅18
0⋅26
0⋅35
0⋅09
1⋅14
1⋅14
1⋅23
1⋅32
0⋅61
1⋅36
1⋅23
0⋅09
0⋅35
0⋅44
0⋅44
0⋅26
0⋅35
0⋅44
0⋅18
1⋅23
1⋅23
1⋅32
1⋅40
0⋅71
1⋅46
1⋅33
0⋅44
0⋅53
0⋅53
0⋅35
0⋅44
0⋅53
0⋅26
1⋅14
1⋅14
1⋅23
1⋅32
0⋅81
1⋅36
1⋅23
0⋅09
0⋅09
0⋅26
0⋅35
0⋅44
0⋅35
1⋅23
1⋅23
1⋅32
1⋅40
0⋅71
1⋅45
1⋅33
0⋅18
0⋅35
0⋅44
0⋅53
0⋅44
1⋅32
1⋅32
1⋅40
1⋅49
0⋅81
1⋅55
1⋅43
K . A . B E S H E R A A N D P. M . H A R R I S
© 2014 The Fisheries Society of the British Isles, Journal of Fish Biology 2014, 85, 228–245
HAP1
HAP2
HAP3
HAP4
HAP5
HAP6
HAP7
HAP8
HAP9
HAP10
HAP11
HAP12
HAP13
HAP14
HAP15
HAP16
HAP17
HAP18
HAP19
HAP20
HAP21
HAP22
HAP23
HAP24
HAP25
HAP26
HAP27
HAP28
HAP29
HAP30
HAP31
HAP32
HAP33
HAP34
HAP1 HAP2 HAP3 HAP4 HAP5 HAP6 HAP7 HAP8 HAP9 HAP10 HAP11 HAP12 HAP13 HAP14 HAP15 HAP16 HAP17 HAP18 HAP19 HAP20 HAP21 HAP22
HAP35
HAP36
HAP37
HAP38
HAP39
HAP40
HAP41
HAP42
HAP43
HAP44
HAP45
0⋅55
1⋅62
0⋅20
0⋅59
0⋅59
0⋅58
0⋅44
0⋅68
0⋅49
0⋅68
0⋅77
0⋅22
1⋅32
0⋅39
0⋅19
0⋅19
0⋅19
0⋅22
0⋅29
0⋅10
0⋅29
0⋅38
0⋅33
1⋅22
0⋅49
0⋅29
0⋅29
0⋅29
0⋅34
0⋅39
0⋅20
0⋅39
0⋅48
0⋅33
1⋅42
0⋅48
0⋅29
0⋅29
0⋅29
0⋅33
0⋅39
0⋅19
0⋅38
0⋅48
0⋅77
1⋅62
0⋅87
0⋅68
0⋅68
0⋅68
0⋅78
0⋅78
0⋅58
0⋅77
0⋅87
0⋅99
0⋅91
1⋅17
0⋅97
0⋅97
0⋅97
1⋅11
1⋅07
0⋅88
1⋅07
1⋅16
0⋅55
1⋅62
0⋅68
0⋅49
0⋅48
0⋅48
0⋅22
0⋅20
0⋅39
0⋅19
0⋅67
0⋅44
1⋅52
0⋅38
0⋅39
0⋅39
0⋅39
0⋅33
0⋅49
0⋅29
0⋅48
0⋅57
0⋅44
1⋅52
0⋅58
0⋅39
0⋅39
0⋅39
0⋅11
0⋅10
0⋅29
0⋅10
0⋅57
0⋅65
1⋅32
0⋅77
0⋅38
0⋅38
0⋅57
0⋅55
0⋅68
0⋅48
0⋅67
0⋅76
0⋅33
1⋅42
0⋅49
0⋅10
0⋅09
0⋅29
0⋅33
0⋅39
0⋅20
0⋅39
0⋅48
0⋅44
1⋅52
0⋅10
0⋅49
0⋅49
0⋅49
0⋅33
0⋅59
0⋅39
0⋅58
0⋅67
0⋅66
1⋅73
0⋅39
0⋅58
0⋅58
0⋅58
0⋅55
0⋅68
0⋅48
0⋅67
0⋅77
0⋅11
1⋅52
0⋅68
0⋅48
0⋅48
0⋅48
0⋅44
0⋅58
0⋅39
0⋅58
0⋅10
0⋅22
1⋅62
0⋅77
0⋅58
0⋅58
0⋅58
0⋅55
0⋅68
0⋅48
0⋅67
0⋅19
1⋅31
0⋅40
1⋅46
1⋅27
1⋅27
1⋅07
1⋅34
1⋅36
0⋅97
1⋅36
1⋅45
1⋅20
0⋅30
1⋅37
1⋅17
1⋅17
0⋅97
1⋅23
1⋅27
0⋅88
1⋅27
1⋅36
1⋅31
0⋅40
1⋅47
1⋅27
1⋅27
1⋅07
1⋅34
1⋅37
0⋅98
1⋅36
1⋅46
1⋅42
0⋅50
1⋅56
1⋅36
1⋅36
1⋅17
1⋅45
1⋅46
1⋅07
1⋅46
1⋅55
1⋅31
0⋅60
1⋅47
1⋅27
1⋅27
1⋅07
1⋅34
1⋅37
0⋅98
1⋅36
1⋅46
1⋅43
0⋅10
1⋅56
1⋅36
1⋅36
1⋅16
1⋅44
1⋅46
1⋅07
1⋅45
1⋅55
1⋅54
0⋅20
1⋅65
1⋅46
1⋅46
1⋅26
1⋅56
1⋅56
1⋅17
1⋅55
1⋅65
HAP23 HAP24 HAP25 HAP26 HAP27 HAP28 HAP29 HAP30 HAP31 HAP32 HAP33 HAP34 HAP35 HAP36 HAP37 HAP38 HAP39 HAP40 HAP41 HAP42 HAP43 HAP44
0⋅35
0⋅44
0⋅53
0⋅44
1⋅32
1⋅32
1⋅40
1⋅49
0⋅81
1⋅55
1⋅43
1⋅54
0⋅20
1⋅65
1⋅45
1⋅45
1⋅26
1⋅55
1⋅55
1⋅16
1⋅55
1⋅64
0⋅26
0⋅35
0⋅09
1⋅14
1⋅14
1⋅23
1⋅32
0⋅61
1⋅36
1⋅23
1⋅31
0⋅40
1⋅47
1⋅27
1⋅27
1⋅07
1⋅34
1⋅37
0⋅98
1⋅36
1⋅46
0⋅09
0⋅35
1⋅23
1⋅23
1⋅32
1⋅40
0⋅72
1⋅46
1⋅33
1⋅43
0⋅51
1⋅57
1⋅37
1⋅37
1⋅17
1⋅45
1⋅47
1⋅08
1⋅46
1⋅56
0⋅44
1⋅32
1⋅32
1⋅40
1⋅49
0⋅82
1⋅56
1⋅44
1⋅53
0⋅61
1⋅66
1⋅47
1⋅47
1⋅27
1⋅56
1⋅57
1⋅17
1⋅56
1⋅66
1⋅23
1⋅23
1⋅32
1⋅40
0⋅72
1⋅46
1⋅33
1⋅42
0⋅51
1⋅56
1⋅37
1⋅37
1⋅17
1⋅45
1⋅47
1⋅08
1⋅46
1⋅56
0⋅35
0⋅61
0⋅18
1⋅23
0⋅68
0⋅62
0⋅55
1⋅42
0⋅20
0⋅59
0⋅59
0⋅59
0⋅45
0⋅69
0⋅49
0⋅68
0⋅77
0⋅44
0⋅35
1⋅12
0⋅48
0⋅51
0⋅44
1⋅52
0⋅39
0⋅39
0⋅39
0⋅39
0⋅44
0⋅49
0⋅29
0⋅48
0⋅58
0⋅61
1⋅12
0⋅48
0⋅51
0⋅44
1⋅52
0⋅58
0⋅39
0⋅39
0⋅39
0⋅33
0⋅49
0⋅29
0⋅48
0⋅57
1⋅22
0⋅68
0⋅61
0⋅55
1⋅62
0⋅20
0⋅39
0⋅39
0⋅58
0⋅44
0⋅68
0⋅49
0⋅68
0⋅77
1⋅22
1⋅02
1⋅09
0⋅81
1⋅21
1⋅12
1⋅12
0⋅91
1⋅21
1⋅22
0⋅81
1⋅22
1⋅22
0⋅41
0⋅56
1⋅62
0⋅67
0⋅48
0⋅48
0⋅48
0⋅44
0⋅58
0⋅39
0⋅58
0⋅67
0⋅55
1⋅42
0⋅61
0⋅51
0⋅51
0⋅51
0⋅55
0⋅61
0⋅41
0⋅61
0⋅61
1⋅53
0⋅55
0⋅45
0⋅44
0⋅44
0⋅48
0⋅55
0⋅33
0⋅55
0⋅22
1⋅62
1⋅52
1⋅52
1⋅32
1⋅54
1⋅62
1⋅22
1⋅62
1⋅62
0⋅58
0⋅58
0⋅58
0⋅44
0⋅67
0⋅48
0⋅67
0⋅77
0⋅19
0⋅39
0⋅44
0⋅48
0⋅29
0⋅48
0⋅58
0⋅39
0⋅44
0⋅48
0⋅29
0⋅48
0⋅58
0⋅44
0⋅48
0⋅10
0⋅48
0⋅58
0⋅22
0⋅33
0⋅22
0⋅55
0⋅39
0⋅19
0⋅67
0⋅39
0⋅48
0⋅67
239
HAP24
HAP25
HAP26
HAP27
HAP28
HAP29
HAP30
HAP31
HAP32
HAP33
HAP34
HAP35
HAP36
HAP37
HAP38
HAP39
HAP40
HAP41
HAP42
HAP43
HAP44
HAP45
PHYLOGEOGRAPHY OF LABEOBARBUS INTERMEDIUS
© 2014 The Fisheries Society of the British Isles, Journal of Fish Biology 2014, 85, 228–245
Table II. Continued
240
K . A . B E S H E R A A N D P. M . H A R R I S
In the MCC tree obtained from discrete phylogeographic analysis, four sequences
from Dedessa River (representing haplotypes 6 and 32) formed a clade sister to all
other sequences in lineage A and had very weak support (PP = 58%). In the haplotype
network, the two haplotypes were intermediate between the two lineages and more distant from haplotypes originating from the same river system as well as from populations
in lineage A than they are from haplotypes in lineage B. For instance, haplotypes 6 and
32 were separated from haplotype 3 (their most closely related haplotype in lineage
A) by seven and eight mutational steps, respectively, while the number of mutational
steps between these haplotypes and haplotype 17 (their most closely related haplotype
in lineage B) were six and ive, respectively. These results may suggest the presence
of undescribed biodiversity within Dedessa River, incomplete haplotype sampling or,
given the complex geologic history of the region (Beadle, 1981) and the close proximity of the headwaters of these river systems, indicate post-divergence connections
between Dedessa and Omo River systems.
It is more likely that aspects of the current phylogeographic structure of the L.
intermedius complex can be understood by considering historical geologic and climatic events of the region. According to Ebinger et al. (2000) and Bonnini et al.
(2005), Africa experienced repeated paroxysms of volcano-tectonic activities during
the Oligocene and the whole period following the Miocene. Divergence date estimates
indicate that the two lineages of the L. intermedius complex diverged c. 0⋅5 M b.p.
during the late Pleistocene. This suggests volcanic events that took place during the
Pleistocene may have greatly altered the topography and drainage patterns of the
region and thus may have signiicantly inluenced genetic differentiation in the L.
intermedius complex.
The distinctness of the two mitochondrial DNA lineages of the L. intermedius complex with no shared haplotypes strongly suggests the existence of a long-term physical
barrier to genetic exchange between drainages harbouring these lineages. Vicariant
events associated with the formation of notable structural elements such as the Addis
Ababa-Nekemt (ANL) and Goba-Bonga (GBL) tectonic lineaments may have affected
genetic divergence in the L. intermedius complex. Within the Rift Valley system, haplotypes from localities north of the GBL fall within lineage A while those from localities
south of the GBL fall within lineage B (Fig. 1). Similarly, haplotypes residing in localities on opposite sides of the ANL fall within different lineages. Volcanic activities that
led to the formation of Addis Ababa-Nekemt Line (ANL) started at c.12-10 M b.p. and
developed through three main phases: 12-7, 6-2 and <1 M b.p. (Abebe et al., 1998).
Likewise, rifting in the Ethiopian Rift Valley evolved in two different phases: an early
(Miocene-Pliocene) fault-dominated rifting stage and a later (Pleistocene) rifting stage
that involved progressive extension (Corti, 2009). Pleistocene volcano-tectonic activities responsible for the evolution of ANL and the Rift Valley appear to match the timing
of lineage divergence in the L. intermedius complex. The timing of inception of the
GBL, hypothesized by Bonnini et al. (2005) as the Oligocene, however, precedes the
estimated time of lineage divergence. The relative recency of lineage divergence (c. 0⋅5
M b.p., Pleistocene) within the L. intermedius complex suggests a later reconiguration
or formation of the GBL.
The phylogeography of the L. intermedius complex appears to correspond, at least
partly, with the evolution of the different segments of the Ethiopian Rift Valley.
Populations originating from the NRV lakes (Lakes Awassa and Langano) are more
closely related to those originating from the Abay and Awash River drainages than
© 2014 The Fisheries Society of the British Isles, Journal of Fish Biology 2014, 85, 228–245
PHYLOGEOGRAPHY OF LABEOBARBUS INTERMEDIUS
241
they are to populations from the SRV region, suggesting that populations originating
from the northern and southern segments of the Rift Valley have independent histories.
This concurs with available geologic evidence that suggests heterogeneous time-space
evolution of the different segments of the Ethiopian Rift Valley (Bonnini et al., 2005;
Corti, 2009).
Mitochondrial DNA variation in the L. intermedius complex shows that haplotypes
originating from localities 10–12 (Omo River basin) are more closely related to
those from localities l3–17 (Chamo-Abaya basin) and Lake Kamnarok (Turkana
basin). This is probably due to recent connections between these basins. Seismic and
Quaternary volcanic data (Ebinger et al., 2000) indicate that the Omo-Turkana basin
and Chamo-Abaya basin in the SRV drainage are currently linked across a 200 km
wide zone. In the late Pleistocene to early Holocene, southern Ethiopian Rift Valley
lakes had overlows into Lake Turkana (Golubstov & Redeat, 2010), suggesting
more recent faunal exchanges between the Omo-Turkana and Chamo-Abaya basins.
In addition, it is hypothesized that the Turkana basin was connected to the Nile
River in the early Holocene (Grove et al., 1975; Grove, 1983; Johnson & Malala,
2009). On the other hand, as shown in this study, Lake Baringo and Abay River
(Nile River system) share a haplotype (i.e. haplotype 4) suggesting recent genetic
exchange between the two systems. The embedment, in this study, of sequences from
Lakes Baringo and Kamnarok (Turkana basin) within lineages A and B, respectively,
suggests that the connections between Lake Baringo and the Nile River system were
severed more recently than those between the latter and Turkana basin. More samples
from Lake Baringo and Turkana basin need to be examined before making irm
conclusions.
TA X O N O M I C I M P L I C AT I O N S F O R T H E L . I N T E R M E D I U S
COMPLEX
The inding that the L. intermedius complex constitutes two divergent lineages has
signiicant taxonomic implications. Banister (1973) established a single polytypic
species, L. intermedius, with two sub-species, L. i. australis and L. i. intermedius;
the taxon distributed throughout Ethiopia and northern Kenya belonged to the former
sub-species while the latter was known only from Lake Baringo. A subsequent study
based on morphological analysis of samples from northern Kenya supported Banister’s
(1973) proposed uniication of several East African barbel species and sub-species into
the L. intermedius complex, but rejected the sub-speciic separation of L. i. australis
(Skorepa, 1992). These taxonomic suggestions, however, have never been tested
phylogenetically. The phylogeographic split of the L. intermedius complex observed
in this study is inconsistent with these hypotheses.
The estimated TMRCA (c. 0⋅5 M b.p.) for the L. intermedius complex lineages suggests that lineages A and B may have been recently isolated. The geographic isolation,
substantial level of genetic divergence and the mutually exclusive haplotypes of the
two lineages are evidence that these lineages have independent histories with little evidence of gene low. Geographic differences of karyology in the L. intermedius complex
between the Awash River population and southern populations inhabiting Lake Abaya
and Chamo basins (Golubstov & Krysanov, 1993) seem to support this dichotomy.
The results further suggest that lineage B could be distinguished as a separate species.
Comparable levels of cytb sequence divergences have been used to recognize species
© 2014 The Fisheries Society of the British Isles, Journal of Fish Biology 2014, 85, 228–245
242
K . A . B E S H E R A A N D P. M . H A R R I S
among Euro-Mediterranean barbs (Zardoya & Doadrio, 1998; Doadrio et al., 2001).
Taxonomic changes, however, cannot be advocated based on a single mtDNA gene.
Therefore, the question of whether lineage B of the L. intermedius complex merits
status as separate species deserves further study.
T H E L A K E TA N A L A B E O B A R B U S S P E C I E S F L O C K
The taxonomy and phylogenetic relationships of Lake Tana Labeobarbus species
have been the subject of much debate (Banister, 1973; Nagelkerke et al., 1994, 1995).
A recent study examining phenotypic variation among Labeobarbus forms in Lake
Tana (Nagelkerke & Sibbing, 2000) concluded that at least 15 distinct species (constituting a species lock) exist within the lake. This study recovered these species as part of
a large polytomy (lineage A) of the L. intermedius complex haplotypes. The different
clusters from Lake Tana (Fig. 1) do not correspond to the different Labeobarbus species
suggested earlier (Nagelkerke et al., 1994, 1995; Nagelkerke & Sibbing, 2000) from
the lake as these species occurred in different clusters. These results are consistent with
de Graaf et al. (2010) who also found no genetic differentiation among these species.
Nagl et al. (2000) and Verheyen et al. (2003) reported strikingly inconsistent morphological and molecular variation in the adaptive radiation of haplochromine cichlids in
Lake Victoria (East Africa, 1∘ 00′ S; 33∘ 00′ E) and hypothesized that recent radiation
may account for this discrepancy. The evolutionary diversiication of the Labeobarbus of Lake Tana seems to parallel the pattern exhibited by the haplochromines of
Lake Victoria and the very shallow genetic divergences (Table II) and lack of phylogenetic resolution (Fig. 2) observed among the Lake Tana species may be explained
by the recency of the origin of these species. The results of this study suggest that
mitochondrial cytb gene is conserved in Lake Tana Labeobarbus over the time scale
for their evolution rendering it an inappropriate marker for inferring the evolutionary
history and taxonomy of the group. Furthering the understanding on the evolutionary
diversiication and taxonomy of the Labeobarbus of Lake Tana requires more study perhaps employing fast evolving genes (e.g. microsatellite markers) or analysis of whole
genome (e.g. single nucleotide polymorphisms, SNP).
In conclusion, the present phylogeographic study shows that the diversity and distribution of genetic variation in the L. intermedius complex relects a more recent but
complex history of vicariance. The timing of divergence (c. 0⋅5 M b.p.) of the L. intermedius complex into two mitochondrial DNA lineages seems to correspond with the
timing of volcano-tectonic activities that affected the East African region during the
Pleistocene. The distinctness of the two mitochondrial DNA lineages of the L. intermedius complex suggests a long history of isolation between these lineages while
phylogenetic afinities and presence of shared haplotypes among the extant L. intermedius complex populations relect relatively recent connections between drainages
across the geographic range of the species.
We wish to thank B. Fluker, M. Sandel, N. V. Whelan, A. Teoh and E. Abebe for their helpful comments. We would like to thank Jimma University (Ethiopia) for providing a vehicle
and a driver for ieldwork. We thank the University of Kansas for providing some tissue specimens used in this study. We also gratefully acknowledge the support afforded by local isheries
experts and ishermen during ieldwork in Ethiopia. Funding for this research was provided
by National Science Foundation (NSF), Cypriniformis Tree of Life Project (DEB 0431263 to
P.M.H.).
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