Open Access

Cytochrome b shows signs of adaptive protein evolution in Gerbillus species from Egypt

  • Mahmoud Amin Khalifa1Email author,
  • Mahmoud Ibrahim Younes2 and
  • Ahmed Ghazy1
The Journal of Basic and Applied Zoology201879:1

https://doi.org/10.1186/s41936-018-0014-x

Received: 25 May 2017

Accepted: 5 January 2018

Published: 16 January 2018

Abstract

Background

Amino acid polymorphisms in the mitochondrial cytochrome b (cytb) gene of four Gerbillus species have been investigated for their geographical distribution and possible functional significance. The sequences were obtained from a total of 20 specimens representing four species of genus Gerbillus collected from Siwa Oasis, Dabaa, Wadi El Natron, El Faiyum, and Baltim in Egypt.

Results

Our results identified a group of amino acid variant polymorphisms that were useful for both species taxonomic and biogeographic assignments. The results demonstrated that amino acid variants L>F173 (Leucine>Phenylalanine), A>M203 (Alanine>Methionine), and I>V221(Isoleucine>Valine) were specific to G. andersoni, while the variant V>M283 (Valine>Methionine) was only specific to G. andersoni from Baltim. The variants, L>P263 (Leucine>Proline) and M>T311 (Methionine>Threonine) were specific only to G. amoenus collected from El Faiyum. Compared to other amino acid variants, L>P263 was remarkably less frequent, and it was predicted using PROVEAN database tool to have non-neutral effects.

Conclusion

Amino acid polymorphisms within the cytochrome b gene could be assigned to specific geographic locations. They might prove suitable to track accumulated and recent environmental changes as they could represent signs of adaptive evolution.

Keywords

Cytochrome b geneAmino acid polymorphismsmtDNABiogeographic assignment Gerbillus Egypt

Background

The Egyptian gerbils of the genus Gerbillus are among the more diversified rodent genera of arid and semi-arid regions of the old world. Occupying diverse habitats, these rodents present a useful model for investigating historical environmental changes that affected the arid, old world belt and drove the distribution and morphological diversification in mammals. In Egypt, the distribution and systematics of this genus and related rodent genera have been comprehensively covered (Osborn and Helmy 1980). In this genus, the molecular assessments have brought recent insights in a group of rodents where morphological studies have always dominated (Musser and Carleton 2005; Ndiaye, Chevret, Dobigny, & Granjon, 2016; Ndiaye et al. 2016b).

At the molecular levels, the most widely used population genetic indicator in animals is mitochondrial DNA (Avise et al. 1987). One of the important mitochondrial genes for survival of organisms is the cytochrome b (cytb) gene. This gene has proved particularly useful for discerning phylogenetics and taxonomic relations (Castresana 2001; Cook et al. 1999; Irwin et al. 1991; Kuwayama and Ozawa 2000; Kocher et al. 1989; Lau et al. 1998).

The cytb gene encodes a membrane-bound molecule, a central catalytic subunit of ubiquinol cytochrome c reductase (bc1 complex or complex-III), that is present in the respiratory chain of mitochondria (Howell 1989). Given the exception of protozoans missing mitochondria, all eukaryotes need this class of reduction-oxidation enzyme, and subsequently cytb, for energy metabolism (Hauska et al. 1983; Trumpower 1990; Widget and Cramer 1991). Further studies have evaluated the physiochemical changes which resulted from the molecular evolution of cytb functional domains in pocket gophers and cetartiodactyls (McClellan and McCracken 2001).

Although many scientific studies on the assessment of cytb amino acid residues at levels of both structure and function (Esposti et al. 1993) as well as physiochemical changes (McClellan and McCracken 2001) already date back to the end of the twentieth century, it is reasonable for investigation as they become more feasible with freely available annotated collection of mtDNA sequences. Some methods have been employed to discover signs of adaptation, among them identifying local declines in diversity, indicating selective sweeps in different populations of species (Andolfatto 2001; Nielsen 2005). Possibly, recent and established online database tools as SIFT (Ng and Henikoff 2003) and PolyPhen-2 (Adzhubei et al. 2013) that have been implemented for evaluating non-synonymous amino acid changes are also helpful in this process. In consistence with previous tools, prediction and assessment of single amino acid substitution using PROVEAN (Choi et al. 2015) become more attainable.

The present study aims to address the relationships between intra- and interspecies-specific mitochondrial cytb amino acid variants in genus Gerbillus from different geographical regions in Egypt. Specific amino acid variants will be assigned to different localities of the Western Desert of Egypt.

Methods

DNA techniques

Analysis was carried out on 20 specimens from different localities in Egypt (Table 1). Specimens were classified based on morphological taxonomic characters described by Harrison and Bates (1991) and Osborn and Helmy (1980). Four species have been identified: Gerbillus campestris, G. andersoni, G. amoenus, and G. gerbillus. Molecular analyses were done in the Laboratory of Molecular Biology, Zoology Department, Faculty of Science, Al Azhar University, Cairo. Genomic DNA was isolated from femoral muscle, using QIAamp® DNA Mini Kit.
Table 1

List of specimens with their GenBank accession numbers of the sequences of cytb (italicized numbers refer to original sequences in this study), geographic origin, haplotype code, and literature source

No.

Species

Accession number

ID

Country

Locality

Geographicalcoordinates

Haplotype code

Source

1

G. amoenus

KX786153

M005255

Egypt

Faiyum

29° 34′ N–30° 54′ E

Hap1

Current study

2

G. amoenus

KX792471

M005256

Egypt

Faiyum

29° 34′ N–30° 54′ E

Hap2

Current study

3

G. amoenus

KX792472

M005257

Egypt

Faiyum

29° 34′ N–30° 54′ E

Current study

4

G. amoenus

KX792473

M005258

Egypt

Faiyum

29° 34′ N–30° 54′ E

Hap3

Current study

5

G. amoenus

KX792474

M005259

Egypt

Faiyum

29° 34′ N–30° 54′ E

Hap4

Current study

6

G. amoenus

KX786154

M005262

Egypt

Wadi El Natroun

30° 23′ N–30° 22′ E

Hap5

Current study

7

G. amoenus

KX792475

M005263

Egypt

Wadi El Natroun

30° 23′ N–30° 22′ E

Current study

8

G. andersoni

KX786151

M005265

Egypt

Baltim-Kafr El Sheikh

31° 34′ N–31° 13′ E

Hap13

Current study

9

G. andersoni

KX792465

M005266

Egypt

Baltim-Kafr El Sheikh

30° 34′ N–31° 13′ E

Hap14

Current study

10

G. andersoni

KX792466

M005267

Egypt

Baltim-Kafr El Sheikh

30° 34′ N–31° 13′ E

Current study

11

G. andersoni

KX792467

M005268

Egypt

Baltim-Kafr El Sheikh

30° 34′ N–31° 13′ E

Hap15

Current study

12

G. andersoni

KX792468

M005269

Egypt

Baltim-Kafr El Sheikh

30° 34′ N–31° 13′ E

Hap16

Current study

13

G. andersoni

KX792469

M005274

Egypt

Dabaa- Matruh

30° 59′ N–28° 27′ E

Hap17

Current study

14

G. andersoni

KX792470

M005275

Egypt

Dabaa- Matruh

30° 59′ N–28° 27′ E

Current study

15

G. andersoni

KX786152

M005277

Egypt

Dabaa- Matruh

30° 59′ N–28° 27′ E

Hap18

Current study

16

D. campestris

KX786155

M005278

Egypt

El Zaiytona, Siwa Oasis

29° 09′ N–25° 49′ E

Hap33

Current study

17

D. campestris

KX792478

M005279

Egypt

El Zaiytona, Siwa Oasis

29° 09′ N–25° 49′ E

Current study

18

D. campestris

KX792479

M005280

Egypt

El Zaiytona, Siwa Oasis

29° 09′ N–25° 49′ E

Current study

19

G. gerbillus

KX792476

M005284

Egypt

Dabaa- Matruh

30° 59′ N–28° 27′ E

Hap25

Current study

20

G. gerbillus

KX792477

M005286

Egypt

Dabaa- Matruh

30° 59′ N–28° 27′ E

Hap26

Current study

21

G. amoenus

KM236112

N3009

Niger

Ourou, Air

19° 16′ N–7° 96′ W

Hap6

Ndiaye, Tatard, et al., 2016

22

G. amoenus

AJ851270

1997016

Mauritania

Nauakchot

18° 12′ N–16° 02′ W

Hap7

Ndiaye et al. 2013

23

G. amoenus

JQ753052

1553

Libya

Sabha

27° 14′ N-14° 24′ E

Hap8

Ndiaye et al. 2013

24

G. amoenus

JQ753062

1999032

Mali

Edjerir

18° 12′ N- 1° 24′ W

Hap9

Ndiaye et al. 2013

25

G. nanus

JQ753054

Gn2

Israel

Arava

29° 51′ N-35° 03′ E

Hap10

Ndiaye et al. 2013

26

G. nanus

JQ753058

Gn6

Israel

Arava

29° 51′ N-35° 03′ E

Hap11

Ndiaye et al. 2013

27

G. nanus

JQ753063

1988007

Pakistan

Sind Desert

 

Hap12

Ndiaye et al. 2013

28

G. andersoni

KM236114

NMP48241

Jordon

Wadi Rum

29.61′ N-35.43′ E

Hap19

Ndiaye, Tatard, et al., 2016

29

G. andersoni

KF496286

NMP 48323

Libya

Sabkhat karkurah

31° 42′ N- 20° 03′ E

Hap21

Ndiaye et al. 2014

30

G. andersoni

KM236113

7044Gand

Israel

Moshav Avshalon

31° 21′ N-34° 34 ′E

Hap20

Ndiaye, Tatard, et al., 2016

31

G. andersoni

LN606673

Gand 2

Israel

Western Negev

31° 21′ N-34° 34′ E

Hap22

Ndiaye, Tatard, et al., 2016

32

G. pyramidum

JN652808

1557

Libya

Jezero Gabroon

27° 03′ N-14° 26′ E

Hap23

Ndiaye et al. 2012

33

G. pyramidum

JN652809

M5982

Mali

Oued Chacheguerene

19° 43′ N-00° 01′ W

Hap24

Ndiaye et al. 2012

34

G. gerbillus

LN606679

1999280

Mauritania

Nauakchot

18° 33′ N–15° 96′ W

Hap27

Ndiaye, Tatard, et al., 2016

35

G. gerbillus

AJ851269

 

Mauritania

Nauakchot

 

Chevret and Dobigny 2005

36

G. gerbillus

GU356564

88F

Tunisia

Faouar

33° 16′ N–08° 29′ E

Hap28

Abiadh et al. 2010

37

G. gerbillus

JN021403

LG112

Morocco

14 km N of Tarfaya

27° 57 N-12° 46 W

Hap29

Ndiaye et al. 2012

38

G. gerbillus

KF496219

M4953

Mali

Vallee Tillemsi

19° 35′ N-0° 183′ W

Hap30

Ndiaye et al. 2014

39

G. gerbillus

LN606680

2002461

Niger

Achegour

19° 01′ N 11° 71′ W

Hap31

Ndiaye, Tatard, et al., 2016

40

G. gerbillus

KR089023

CM:113823

Egypt

Giza

29° 70′ N30° 97′ E

Hap32

Alhajeri et al. 2015

41

D. campestris

KF496218

1999040

Niger

Air (Agadez)

 

Hap34

Ndiaye et al. 2014

42

D. campestris

AJ851271

1999030

Mali

Edjerir, Adrar des Iforas

18° 2′ N1° 4′ W

Hap35

Chevret and Dobigny 2005

43

D. campestris

KR089016

TK-4090

Tunisia

Mournagia Subgovt, Jebel Ain Es Seed

 

Hap36

Alhajeri et al. 2015

44

Sekeetamys calurus

AJ851276

 

Egypt

  

Hap37

Chevret and Dobigny 2005

ID refers to the identification number of the samples studied and, when available, to the corresponding museum specimen

The complete cytb was amplified using Thermocycler GeneAmp 9700, USA. Pairs of primers, L14723 (5′-ACCAAT GAC ATG AAA AAT CAT CGT T-3′) and H15915 (5′-TCT CCATTT CTG GTT TAC AAG AC-3′), as described by Ducroz (1998), were used to target the cytb gene of 1140 bp size. The polymerase chain reaction (PCR) thermal program was set as follows. Initial denaturation at 94 °C for 1 min. Then 35 cycles of denaturation at 94 °C for 1 min, annealing at 62 °C for 1 min, and extension at 72 °C for 1 min. Final extension step at 72 °C for 10 min. PCR amplifications were carried out in 25 μl reaction volume including 0.5 μl of each primer (10 pmole/μl), 12.5 μl 2× PCR Master Mix solution (i-Taq™), 10.5 H2O and 1 μl of template DNA. PCR products were purified using the QIAquick PCR purification kit and prepared for automated sequencing (3500 Genetic Analyzer, Applied Biosystems) relying on one of the primers utilized for the amplification. The acquired new sequences were all deposited in GenBank under serial accession numbers KX786151 to KX786155 and KX792465 to KX792479 after sequence quality assessments (Table 1).

Genetic data analysis

Sequence and database analysis

MEGA 7.0.14 software (Kumar et al. 2016) was used to align and proofread generated sequences from the sequencer. Conflicting DNA bases within DNA sequences were verified against the associated chromatograms. Accordingly, the cytochrome b sequences were generated for subsequent database analysis.

Basic Local Alignment Search Tool (BLAST)

To check and identify generated sequences, each was blast searched as a query through NCBI (National Center for Biotechnology Information) Blastn tool (www.ncbi.nlm.nih.gov/BLAST/). Sequences with best hits were retrieved and used as outgroups (Table 1) for further comparison to cytb sequences from the current study.

Multiple sequence alignment (MSA)

The nearly complete cytb gene sequences, 20 samples generated in this study were aligned against 24 sequences downloaded from GenBank (www.ncbi.nlm.nih.gov/genbank) of various well characterized Gerbillus species as a reference, and Sekeetamys calurus was used as an outgroup. Initial analysis, using DnaSP v.510 (Librado and Rozas 2009), of the total 44 cytb gene sequences gave 37 haplotypes represented as Hap1, Hap2, …, and Hap37 that will stand for future alignment.

Multiple sequence alignment was performed for DNA and protein sequences. The Bioinformatics Resource Portal (http://web.expasy.org/translate/) was used to translate the DNA sequence of cytb gene into its respective protein. Functional effect predictions of non-synonymous single nucleotide polymorphisms (nsSNPs) were achieved using PROVEAN (Protein Variation Effect Analyzer) (http://provean.jcvi.org/) (Kumar et al. 2014), which measures the damaging effect of variations in protein sequences (Choi et al. 2012). The prediction is based on the change in the similarity of the sequence to related protein sequences in a MSA by a delta alignment score of the reference and the variant carrying protein sequence with respect to the alignment of homologous sequences (Choi 2012). Variants with score equal to or below − 2.5 are considered as deleterious nsSNP.

Results and discussion

Multiple sequence alignment

Cytb DNA sequence

The alignment of cytb DNA sequences, from the current study and database, became more accurate after using a codon-wise approach which facilitated assuring positions of possible insertions and deletions. Codon-wise sequence alignment of cytb haplotypes from populations of different Gerbillus species, together with outgroup haplotypes, exposed most of the segregation sites which were probably enough for speciation events between different taxa.

Out of the 942 nucleotides used in the multiple alignments of cytb haplotypes, 282 variable sites compared to 660 conserved sites. From the variable sites, 38 positions were found as singletons while the remaining 244 variable sites were parsimony informative.

Cytb amino acid sequence

Multiple alignment of amino acid sequences is a standard technique for visualizing the relationships between residues in a collection of evolutionary related proteins. In the current study, multiple alignment of the cytb amino acid residues from Gerbillus species gave the chance to deepen the understanding of the evolutionary relationship between some species within the generic name Gerbillus.

Out of 313 deduced amino acid residues used in the alignment of cytb haplotypes (Fig. 1), 27 polymorphic sites were compared to 286 conserved sites. Out of these polymorphic sites, 19 sites were parsimony informative sites. Species-specific variations as well as those shared within the genus Gerbillus appear to be geographically/habitat related and could possibly be assigned to species biogeographic history. Possibly, these variations can be used to explain evolutionary relationships, as well as the type and effect of natural selection which might act differently in different geographic regions.
Fig. 1

Multiple sequence alignment of cytb protein residues haplotypes. “Hap” indicates haplotype; (.) represent the same amino acid as in the consensus sequence; (/ /) stands for sequences shared between all haplotypes. G.am: Gerbillus amoenus; G.na: Gerbillus nanus; G.an: Gerbillus andersoni; G.py: Gerbillus pyramidum; G.ge: Gerbillus gerbillus; D.ca: Dipodillus campestris (also common as Gerbillus campestris). Haplotypes with gray background are these from the current study compared to those from the database

DNA-based amino acid polymorphisms

The major species-specific and region-restricted amino acid variants within the cytb sequence of G. amoenus and its closely related congeneric species are compiled in Table 2 and Fig. 2. This evaluation of the multiple sequence alignment of cytb amino acid sequences proved helpful for exploring a group of variants, that are species specific, in more details. In addition, more understanding was achieved after using the online PROVEAN (Protein Variation Effect Analyzer) tool for filtering variants that possibly have an impact on the biological function of cytb protein.
Table 2

Intraspecific variation within different species of the Gerbillus species

Species

Collection site

Haplotype no.

Amino acid change (species specific)

Nucleotide change

Codon change

PROVEAN

Region restricted variants

Source

Predicted effect

Scores

G. amoenus

Wadi el Natroun

Hap5

V>A30

T90C

GTT>GCT

Neutral

− 2.127

V>A30

Current study

Libya

Hap8

I>N31

T93A

ATT>AAT

Neutral

− 1.930

I>N31

Outgroup

El Faiyum, Wadi el Natroun Niger, Mauritania Libya, Mali

Hap1–9

L>F223

C668T,

A670T

CTA>TTT

Neutral

− 1.553

Current study + outgroup

El Faiyum

Hap4

L>P263

T789C

CTA>CCA

Deleterious

− 4.513

L>P263

Current study

Hap1–4

M>T311

T933C

ATA>ACA

Neutral

− 0.593

M>T311

G. andersoni

Jordan

Libya

Hap19, Hap21

L>F173

C518T,

C520T

CTC>TTT

Neutral

− 1.978

Outgroup

Baltim, Dabaa, Israel

Hap13–18,

Hap20,

Hap22

C518T

CTC>TTC

Neutral

Current study + outgroup

Baltim, Dabaa, Libya,

Jordan

Israel

Hap13–22

A>M203

G608A,

C609T

GCA>ATA

Neutral

− 1.459

I>V221

A662G,

T664C

ATT>GTC

Neutral

0.127

Baltim

Hap13–16

V>M283

G848A

GTA>ATA

Neutral

− 0.015

V>M283

Current study

G. gerbillus

Tunisia, Morocco, Mali, Niger

Hap28–31

V>M30

G89A

GTA>ATA

Neutral

− 1.076

Outgroup

Dabaa, Mauritania, Tunisia, Morocco, Mali, Niger, Giza

Hap25–32

A>S203

G608T

GCA>TCA

Neutral

− 0.364

Current study + outgroup

D. campestris

Tunisia

Hap36

V>L30

G89C

GTA>CTA

Neutral

− 0.647

V>L30

Outgroup

Siwa, Niger, Mali, Tunisia

Hap33–36

F>Y217

T651A

TTC>TAC

Neutral

− 0.715

Current study + outgroup

I>T229

T687C

ATT>ACT

Neutral

1.448

V>I283

G848A,

A850T

GTA>ATT

Neutral

0.183

S. calurus

Egypt

Hap37

V>I2

T5A

GTA>GAA

Neutral

0.034

Outgroup

I>L106

A317C,

T319C

ATT>CTC

Neutral

− 1.046

I>V176

A527G

ATC>GTC

Neutral

− 0.196

L>M222

C667A

ATC>ATA

Neutral

− 0.708

F>L226

T677C,

C679A

TTC>CTA

Neutral

1.225

I>L229

A686C

ATT>CTA

Neutral

0.104

L>S237

T711C

TTA>TCA

Deleterious

− 4.159

I>T295

T885C,

T886A

ATT>ACA

Neutral

− 1.700

Fig. 2

Cytb amino acid variants within and between species. G & S indicates Gerbillus & Sekeetamys; na, ca, am, an, cam, ge, & py, represent nanus, calurus, amoenus, andersoni, campestris, gerbillus, & pyramidum, respectively. Asterisk (*) is used to indicate variants which are geographically assigned

In general, among polymorphic sites that were identified from the alignment, 8 amino acid variants V>I2, I>L106, I>V176, L>M222, F>L226, I>L229, L>S237, and I>T295 were specifically portraying the outgroup genus Sekeetamys calurus (Hap37), compared to 15 variants V>A30, I>N31, L>F223, L>P263, M>T311, L>F173, A>M203, I>V221, V>M283, V>M30, A>S203, V>L30, F>Y217, I>T229, and V>I283 assigned specifically to species members from genus Gerbillus and were enough for separating this genus from its closest outgroup S. calurus, while 8 amino acid variants were in common between both genera (Fig. 2, Table 2).

In more details, amino acid variants V>A30 (Valine>Alanine), I>N31 (Isoleucine>Asparagine), L>F223 (Leucine>Phenylalanine) L>P263 (Leucine>Proline), and M>T311 (Methionine>Threonine) showed a restriction to G. amoenus, i.e., only 5 amino acid variant positions are specifically characterizing G. amoenus members from their closest species. Further, amino acid variants and similar scale of variations that showed a specificity to the other species are assorted in Table 2 and Fig. 2. Therefore, this possibly introduces a usable utility with a respective variation scale for each species. That way, collectively with inclusion of more genes, we demand further investigation to extend this scale of variation to cover more species and genera and develop a utility as a novel trend to probably solve and/or reduce recent controversies of molecular and morphologic taxonomy of genus Gerbillus reported by Musser and Carleton (2005) and Ndiaye et al. (2016a, 2016b).

In this work, among the evaluation outputs of multiple sequence alignment of cytb protein is the assignment of a group of amino acid variants to specific biogeographic regions. The variants V>A30 in haplotype 5 (Hap5, current study), I>N31 in haplotype 8 (Hap.8, database), L>P263 and M>T311 in haplotypes 1–4 (Hap1–4, current study), V>M283 in haplotypes 13–16 (Hap13–16, current study), and V>L30 in haplotype 36 were respectively assigned to Wadi el Natroun Egypt, Libya, El Faiyum Egypt, Baltim Egypt, and Tunisia (Table 2, Fig. 1). With these different localities, characterized by certain amino acid variants, we can record signs of interaction between environmental changes and wild life of diverse habitat overtime in Egypt in order to better track future changes and answer questions relating to population level.

Database prediction of different amino acid variants using PROVEAN allowed speculation of possible effects on the biological function of cytb protein in different localities. The PROVEAN predicted that the effects of cytb amino acids were mostly of neutral biological function except for amino acid variants L>P263 in G. amoenus collected from El Faiyum (current study) and L>S237 in S. calurus from Egypt (database) which were deleterious. These amino acid variants that gave neutral PROVEAN scores are given in Table 2. We are aware that any scores of in silico prediction tools should be taken with caution and might not actually represent the true effect of variants on biological function. The predictive accuracy of these tools was recently studied by Leong et al. (2015) who concluded that they must never be relied on as a final arbiter of pathogenicity but that they should rather be assessed as raising or lowering probabilities. Also, reports by Choi et al. (2012) have underlined that low scores are given to those amino acid residues found in conserved regions or domains while high scores were found in non-conserved regions when searched against the database of related sequences. PROVEAN has a higher specificity score compared to other efficient and commonly used tools like SIFT and PolyPhen-2 (Choi et al. 2012).

Interestingly, unlike other amino acid variants that are predicted to give a neutral PROVEAN score, L>P263 (Leucine>Proline), with its relatively small prevalence as a minor allele (Fig. 2, Hap4) in and specific to the El Faiyum depression, is predicted by PROVEAN to carry a deleterious functional effect (Table 2) with a score of − 4.513. Based on this predicted effect, it may be expected that G. amoenus members carry this “P” allele as a short-lived polymorphism, which does not persist in the population long enough to become fixed and rather become lost overtime due to lack of fitness and further fixation of the alternative “L” allele, possibly because of random genetic drift as similarly explained by Ohta (1992a, 1992b) who studied amino acid polymorphisms in both Drosophila and human mtDNA. However, a relatively similar study (Rand and Kann 1996) on one of the mitochondrial genes gave a different point of view, which suggested that some pressure of adaptive divergence might be acting in conflict with forces removing or rather eliminating variation. Interestingly, this might better describe the running scene of adaptive divergence in El Faiyum Depression. With the low sample size and one out of five gerbils at that location carrying the allele with the non-neutral, and hence possibly deleterious or possibly advantageous effect on the function of the vital cytb protein, we might either have sampled by chance a very rare individual, or the allele frequency might be indeed rather high. The latter scenario would argue against a deleterious effect of L>P263 and thus favor the hypothesis of positive adaptive evolution.

Conclusions

We evaluated the importance of multiple sequence alignment of cytochrome b amino acid sequences of the genus Gerbillus from different habitat types in Egypt. This alignment proved useful for assigning specific amino acid variants within and/ or between species and to specific geographic locations of varying environmental conditions. Herewith, the provided data and analysis approach will be a useful starting point for further investigation on the geographic distribution of cytb sequence variants and their possible role in environmental adaptation of Gerbillus spp. in Egypt.

Abbreviations

A: 

Alanine

BLAST: 

Basic Local Alignment Search Tool

bp: 

Base pair

Cytb

Cytochrome b

D: 

Aspartic acid

DnaSP: 

Deoxyribonucleic acid sequence polymorphism

E: 

Glutamic acid

F: 

Phenylalanine

G: 

Glycine

H: 

Histidine

Hap: 

Haplotype

I: 

Isoleucine

L: 

Leucine

M: 

Methionine

MSA: 

Multiple Sequence Alignments

mtDNA: 

Mitochondrial deoxyribonucleic acid

N: 

Asparagine

NCBI: 

National Center for Biotechnology Information

nsSNP: 

Non-synonymous single nucleotide polymorphism

P: 

Proline

PCR: 

Polymerase chain reaction

PolyPhen2: 

Polymorphism Phenotyping 2

PROVEAN: 

Protein Variation Effect Analyzer

Q: 

Glutamine

S: 

Serine

SIFT: 

Sorting Intolerant From Tolerant

SNP: 

Single nucleotide polymorphism

Spp: 

Species

T: 

Threonine

V: 

Valine

Y: 

Tyrosine

Declarations

Acknowledgements

We are grateful to Prof. Dr. Mostafa A. Saleh from the Department of Zoology, Faculty of Science, Al Azhar University, Cairo, Egypt, for his help in collecting the study specimens and for his helpful advice. Dr. Konrad Schmidt, afrigene, Innsbruck, Austria, provided valuable help in editing the revised manuscript, both for language as well as scientific issues.

Funding

Not applicable.

Authors’ contributions

MAK is the principle investigator who designed the experiments, led the data analysis, and wrote the paper. MIY is the supervisor who identified and classified the gerbils specimens according to their morphological characteristics and shared in the writing process and revision. AG performed the main practical work. All authors read and approved the final manuscript.

Ethics approval and consent to participate

Not applicable.

Consent for publication

Not applicable.

Competing interests

The authors declare that they have no competing interests.

Publisher’s Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made.

Authors’ Affiliations

(1)
Department of Zoology, Molecular Biology Lab., Faculty of Science, Al-Azhar University, Nasr City, Egypt
(2)
Department of Zoology, Ecology Lab., Faculty of Science, Al Azhar University, Nasr City, Egypt

References

  1. Abiadh, A., Chetoui, M., Cheniti, T. L., Capanna, E., & Colangelo, P. (2010). Molecular phylogenetics of the genus Gerbillus (Rodentia, Gerbillinae): Implications for systematics, taxonomy and chromosomal evolution. Molecular Phylogenetics and Evolution, 56, 513–518.View ArticlePubMedGoogle Scholar
  2. Adzhubei, I., Jordan, D.M., Sunyaev, S.R., 2013. Predicting functional effect of human missense mutations using PolyPhen-2. Current Protocols in Human Genetics. Chapter 7, Unit7. 20.Google Scholar
  3. Alhajeri, B. H., Hunt, A. J., & Steppan, S. J. (2015). Molecular systematics of gerbils and deomyines (Rodentia: Gerbillinae, Deomyinae) and a test of desert adaptation in the tympanic bulla. Journal of Zoological Systematics and Evolutionary Research, 53, 312–330. https://doi.org/10.1111/jzs.12102.View ArticleGoogle Scholar
  4. Andolfatto, P. (2001). Adaptive hitchhiking effects on genome variability. Current Opinion in Genetics & Development, 11, 635–641.View ArticleGoogle Scholar
  5. Avise, J. C., Arnold, J., Ball, R. M., Bermingham, E., Lamb, T., Neigel, J. E., … Saunders, N. C. (1987). Intraspecific phylogeography: The mitochondrial-DNA bridge between population-genetics and systematics. Annual Review of Ecology and Systematics, 18, 489–522.View ArticleGoogle Scholar
  6. Castresana, J. (2001). Cytochrome b phylogeny and the taxonomy of great apes and mammals. Molecular Biology and Evolution, 18, 465–471.View ArticlePubMedGoogle Scholar
  7. Chevret, P., & Dobigny, G. (2005). Systematic and evolution of the subfamily Gerbillinae (Mammalia, Rodentia, Muridae). Molecular Phylogenetics and Evolution, 35, 674–688.View ArticlePubMedGoogle Scholar
  8. Choi, Y. A. (2012). Fast computation of pairwise sequence alignment scores between a protein and a set of single-locus variants of another protein in proceedings of the ACM conference on bioinformatics, computational biology and biomedicine (BCB 12), (pp. 414–417). New York, NY: ACM.Google Scholar
  9. Choi, Y. A., Chan, A. P., & PROVEAN web server (2015). A tool to predict the functional effect of amino acid substitutions and Indels. Bioinformatics, 31(16), 2745–2747.View ArticlePubMedPubMed CentralGoogle Scholar
  10. Choi, Y. A., Sims, G. E., Murphy, S., Miller, J. R., & Chan, A. P. (2012). Predicting the functional effect of amino acid substitutions and Indels. PLoS One, 7, e46688.View ArticlePubMedPubMed CentralGoogle Scholar
  11. Cook, C. E., Wang, Y., & Sensabaugh, G. (1999). A mitochondrial control region and cytochrome b phylogeny of sika deer (Cervus nippon) and report of tandem repeats in the control region. Molecular Phylogenetics and Evolution, 12, 47–56.View ArticlePubMedGoogle Scholar
  12. Ducroz, J. F. (1998). Contribution des approches cytogénétique et moléculaire à l’étude systématique et évolutive des genres de rongeurs Murinae de la «division» Arvicanthis. France: PhD thesis, Muséum National d’Histoire Naturelle.Google Scholar
  13. Esposti, M. D., De Vries, S., Crimi, M., Ghelli, A., Paternello, T., & Meyer, A. (1993). Mitochondrial cytochrome b: Evolution and structure of the protein. Biochimica et Biophysica Acta (BBA) - Bioenergetics, 1143(3), 243–271.View ArticleGoogle Scholar
  14. Harrison, D., & Bates, P. (1991). The mammals of Arabia, (2nd ed., p. 354). Sevenoaks, Kent: Harrison Zoological Museum Publ.Google Scholar
  15. Hauska, G., Hurt, E., Gabellini, N., & Lockau, W. (1983). Comparative aspects of quinol-cytochrome c/plastocyanin oxidoreductases. Biochimica et Biophysica Acta, 726, 97–133.View ArticlePubMedGoogle Scholar
  16. Howell, N. (1989). Evolutionary conservation of protein regions in the protonmotive cytochrome b and their possible roles in redox catalysis. Journal of Molecular Evolution, 29(2), 157–169.View ArticlePubMedGoogle Scholar
  17. Irwin, D. M., Kocher, T. D., & Wilson, A. C. (1991). Evolution of the cytochrome b gene of mammals. Journal of Molecular Evolution, 32, 128–144.View ArticlePubMedGoogle Scholar
  18. Kocher, T. D., Thomas, W. K., Meyer, A., Edwards, S. V., Paabo, S., Villablanca, F. X., & Wilson, A. C. (1989). Dynamics of mitochondrial DNA evolution in mammals: Amplification and sequencing with conserved primers. Proceedings of the National Academy of Sciences, 86, 6196–6200.View ArticleGoogle Scholar
  19. Kumar, A., Rajendran, V., Sethumadhavan, R., Shukla, P., Tiwari, S., & Purohit, R. (2014). Computational SNP analysis: Current approaches and future prospects. Cell Biochemistry and Biophysics, 68, 233–239.View ArticlePubMedGoogle Scholar
  20. Kumar, S., Stecher, G., & Tamura, K. (2016). MEGA7: Molecular evolutionary genetics analysis version 7.0 for bigger datasets. Molecular Biology and Evolution, 33(7), 1870–1874. https://doi.org/10.1093/molbev/msw054.View ArticlePubMedGoogle Scholar
  21. Kuwayama, R., & Ozawa, T. (2000). Phylogenetic relationships among European red deer, wapiti, and sika deer inferred from mitochondrial DNA sequences. Molecular Phylogenetics and Evolution, 15, 115–123.View ArticlePubMedGoogle Scholar
  22. Lau, C. H., Drinkwater, R. D., Yusoff, K., Tan, S. G., Hetzel, D. J., & Barker, J. S. (1998). Genetic diversity of Asian water buffalo (Bubalus bubalis): Mitochondrial DNA D-loop and cytochrome b sequence variation. Animal Genetics, 29, 253–264.View ArticlePubMedGoogle Scholar
  23. Leong, I. U., Stuckey, A., Lai, D., Skinner, J. R., & Love, D. R. (2015). Assessment of the predictive accuracy of five in silico prediction tools, alone or in combination, and two metaservers to classify long QT syndrome gene mutations. BMC Medical Genetics, 16(1), 1–13. https://doi.org/10.1186/s12881-015-0176-z.View ArticleGoogle Scholar
  24. Librado, P., & Rozas, J. (2009). DnaSP v5: A software for comprehensive analysis of DNA polymorphism data. Bioinformatics, 25, 1451–1452.View ArticlePubMedGoogle Scholar
  25. McClellan, D. A., & McCracken, K. G. (2001). Estimating the influence of selection on the variable amino acid sites of the Cytochrome b protein functional domains. Molecular Biology and Evolution, 18(6), 917–925.View ArticlePubMedGoogle Scholar
  26. Musser, G. C., & Carleton, M. D. (2005). Superfamily Muroidea. In D. E. Wilson, & D. M. Reeder (Eds.), Mammal species of the world: A taxonomic and geographic reference, (vol. 2, Third ed., pp. 894–1531). Baltimore: Johns Hopkins University Press.Google Scholar
  27. Ndiaye, A., Ba, K., Aniskin, V., Benazzou, T., Chevret, P., Konecny, A., … Granjon, L. (2012). Evolutionary systematics and biogeography of endemic gerbils from Morocco: An integrative taxonomy approach. Zoologica Scripta, 41, 11–28.View ArticleGoogle Scholar
  28. Ndiaye, A., Chevret, P., Dobigny, G., & Granjon, L. (2016a). Evolutionary systematics and biogeography of the arid habitat-adapted rodent genus Gerbillus (Rodentia, Muridae): A mostly Plio-Pleistocene African history. Journal of Zoological Systematics and Evolutionary Research. https://doi.org/10.1111/jzs.12143.
  29. Ndiaye, A., Hima, K., Dobigny, G., Sow, A., Dalecky, A., Ba, K., … Granjon, L. (2014). Integrative study of a poorly known Sahelian rodent species, Gerbillus nancillus (Rodentia, Gerbillinae). Zoologischer Anzeiger, 253, 430–439.View ArticleGoogle Scholar
  30. Ndiaye, A., Shanas, U., Chevret, P., & Granjon, L. (2013). Molecular variation and chromosomal stability within Gerbillus nanus (Rodentia, Gerbillinae): Taxonomic and biogeographic implications. Mammalia, 77, 105–111.View ArticleGoogle Scholar
  31. Ndiaye, A., Tatard, C., Stanley, W., & Granjon, L. (2016b). Taxonomic hypotheses regarding the genus Gerbillus (Rodentia, Muridae, Gerbillinae) based on molecular analyses of museum specimens. ZooKeys, 566, 145–155.View ArticleGoogle Scholar
  32. Ng, P. C., & Henikoff, S. (2003). SIFT: Predicting amino acid changes that affect protein function. Nucleic Acids Research, 31(13), 3812–3814.View ArticlePubMedPubMed CentralGoogle Scholar
  33. Nielsen, R. (2005). Molecular signatures of natural selection. Annual Review of Genetics, 39, 197–218.View ArticlePubMedGoogle Scholar
  34. Ohta, T. (1992a). The nearly neutral theory of molecular evolution. Annual Review of Ecology and Systematics, 23, 263–286.View ArticleGoogle Scholar
  35. Ohta, T. (1992b). Theoretical study of near neutrality. II. Effect of subdivided population structure with local extinction and recolonization. Genetics, 130, 917–923.PubMedPubMed CentralGoogle Scholar
  36. Osborn, D.J., Helmy, I. (1980). The contemporary land mammals of Egypt (including Sinai). Field. Zool. New series No.5.Google Scholar
  37. Rand, D. M., & Kann, L. M. (1996). Excess of amino acid polymorphism in mitochondrial DNA: Contrasts among genes from drosophila, mice and humans. Molecular Biology and Evolution, 13(6), 735–748.View ArticlePubMedGoogle Scholar
  38. Trumpower, B. L. (1990). Cytochrome bc1 complexes of microorganisms. Microbiological Reviews, 54, 101–129.PubMedPubMed CentralGoogle Scholar
  39. Widget, W. R., & Cramer, W. A. (1991). In cell culture and somatic cell genetics of plants, (vol. 7B, pp. 149–176). New York: Academic Press.Google Scholar

Copyright

© The Author(s) 2018

Advertisement