Vol.1, No.3, 73-76 (2011)
http://dx.doi.org/10.4236/oje.2011.13010
Open Journal of Ecology
C
opyright © 2011 SciRes.
Preliminary molecular variability among haplotypes of
Saudi Arabian house sparrow Passer domesticus
Sayed A. M. Amer1,2*, Metwally M. Montaser1,3, Mohammed Shobrak1
1Faculty of Science, Taif University, Taif, Kingdom of Saudi Arabia; *Corresponding Author: [e-mail]
2Department of Zoology, Faculty of Science, Cairo University, Giza, Egypt;
3Department of Zoology, Faculty of Science, Al-Azhar Uni ver sity, Cairo, Egypt.
Received 22 September 2011; revised 20 October 2011; accepted 30 October 2011.
ABSTRACT
Genetic variability of Passer domesticus from
different localities of Saudi Arabia was investi-
gated. Six hundred and fifty nine nucleotides
were sequenced from the mitochondrial cyto-
chrome b gene. There was a slight difference
among the studied haplotypes and most substi-
tutions were synonymous. In some haplotypes
at the west of Arabian Peninsula (mountainous
habitat), two transitions only were non-synony-
mous. The data were used to construct the rela-
tionship of the Arabian house sparrow to its con
specific taxa from Europe and America. Neigh-
bor-joining (NJ), maximum-parsimony (MP) and
maximum-likelihood (ML) analytical methods
were used. The three methods showed cluster-
ing of the Arabian haplotypes in one group and
their sister relationship with the haplotypes fr-
om Netherlands and America. A spanish haplo-
type came basal to both groups. Collecting
more samples and more DNA data could be of
necessary to clearly address the genetic vari-
ability of this rapidly propagated species in Ara-
bia.
Keyw ords: Saudi Arabia; House Sparrow;
Mitochondrial DNA; Genetic Variability
1. INTRODUCTION
The house sparrow, Passer domesticus is distributed
in Europe, Nor t h Af ri ca and p a rts of Asia such as Middle
East, Indian subcontinent and a narrowing band from
northern Asia toward the Pacific coast. Its large-scale
spread includes North and South America, South Africa,
Australia and New Zealand [1-3].
The genetic diversity among the avian fauna is found
to be low, probab ly due largely to the greater mobility of
birds, resulting in higher levels of gene flow [4]. There
have been several studies of the genetics of house spar-
rows on species and population level [5-11]. Moreover,
the relationships among their congeneric species and to
other finches (Passeridae and New World sparrows) have
been tackled molecularly, for the first time, by Allende et
al. [12]. Recently, the genetic relationships among pas-
serines and other related families were studied using
mitochondrial and nuclear genome’s data [13]. Dor and
Lotem [14] have studied the parentage of the house
sparrows on a molecular basis.
Quantifying and characterizing avian genetic variabil-
ity should be a priority in conservation to evaluate the
effect of recent drastic population changes, to preserve
present-day diversity and, eventually, to provide guide-
lines for future conservation plans [15]. Despite its close
proximity to man, relatively little is known about the
connectivity between house sparrow populations. Do the
various sites that has been occupied by the species form
a continuous population, or are we dealing with a more
meta population type of population structure? These
Conservation information should take into account kn-
owledge on genetic population structure. Therefore the
aim of the present study was to collect samples of the
house sparrow from around the Kingdom of Saudi Ara-
bia in order to sequence an orthologous gene from each
of these samples. Such data could aid to understand the
relatedness of these haplotypes and to get an overall pic-
ture of the genetic variability of this passerine bird pro-
viding guidelines for its conservation.
2. MATERIALS AND METHODS
We collected 17 samples of P. domesticus from the
wild of 6 localities from Saudi Arabia (Arar, Qassim,
Tabuk, Al-Madina, Taif and Jazan). See Figure 1 that
explains the exact localities. Blood and tissue samples
have been numbered and labeled immediately in the lab
and preserved in –80˚C for further molecular studies.

S. A. M. Amer et al. / Open Journal of Ecology 1 (2011) 73-7 6 74
Figure 1. Map of the Arabian Peninsula showing the Saudi
Arabian localities from which the studied samples have been
collected.
DNA was extracted from 0.5 ml blood samples with QI-
AGEN spin-column kits according to the manufacturer ’s
instruction. Extracted DNA was spectrophotmetrically
quantified at 260/280 nm and was used for polymerase
chain reaction (PCR).
PCR was performed in 50 µl total volume of reaction
buffer containing 0.2 mM dNTPs, 1.5 mM MgCl2, 2 µl of
DNA solution and 0.25U of DNA Taq-polymerase (Invi-
trogen). 0.2 µM of each of the L14841 5’-AAAAAG-
CTTCCATCCAACATCTCAGCATGATGAAA-3’ and H
15767 5’-ATGAAGGGATGTTCTACTGGTTG-3’ as de-
tailed by Edwards et al. [16] were also added. The reac-
tion mixture was put into a 0.2 ml thin-walled PCR tube
and amplification was performed in PXE 0.5 thermal
cycler (Thermo Electron Corporation Co.) with the fol-
lowing profile: 94˚C for 5min followed by 30 cycles of
94˚C for 1min, 58˚C for 1min and 72˚C for 1min. A final
strand elongation at 72˚C was done for an additional
7min.
The resultant solutions were electrophoresed on a 1.5%
agarose gel in TAE (40 mM Tris, 40 mM acetic acid and
1 mM ethylenediamine-tetra acetic acid) and the gels
were stained with ethidium bromide. 100 bp DNA Lad-
der (Biolabs) was used as a marker for the molecular
weight size. The PCR products were then purified from
gel with the use of spin column according to the Kit
Manual.
Sequencing reactions were performed in a MJ Re-
search PTC-225 Peltier Thermal Cycler using a ABI
PRISM. BigDyeTM Terminator Cycle Sequencing Kits
with AmpliTaq-DNA polymerase (FS enzyme) (Applied
Biosystems) following the protocols supplied by the ma-
nufacturer were used. A single-pass sequencing was per-
formed on each template using the last mentioned PCR-
primers. The fluorescent-labeled fragments were purified
from the unincorporated terminators with an ethanol pre-
cipitation protocol. The samples were resuspended in
distilled water and subjected to electrophoresis in an ABI
3730xl sequencer (Applied Biosystems).
Nucleotide sequences of the mitochondrial cytb gene
(659 bp) were aligned with the same fragment for other
haplotypes from the DDBJ database (1 from USA, 1
from Netherlands, 1 from Spain, 1 P. flaveolus and 1
from P. luteu s). One additional outgrou p taxan (Petronia
petron ia) was included in the alignment in order to root
the tree. The alignment was carried out by using the
DNASIS 3.5 (Hitachi) and MacClade 4.03 (Sinauer As-
sociates, Inc.) with manual adjustments. We conducted
the tree analyses by neighbor-joining (NJ), maximum-
parsimony (MP) and maximum-likelihood (ML) meth-
ods. These analyses were done in PAUP* 4.0b10 [17] by
heuristic searches with the TBR branch swapping, 10
random taxon additions and 1000 bootstrap replications
for each method. Juckes-Cantor distance model was used
to construct a neighbor-joining tree [18].
3. RESULTS AND DIS CUSSION
Unambiguous 659 sites from cytb gene for 17 samples
of the house sparrow P. domesticus were sequenced in
this study. These data were deposited in DDBJ/EMBL
GenBank database with their accession numbers (AB-
671331-AB671347). The sequences were aligned and wer e
used for an al ysis. Thes e da ta show ed b ase fr equ encies of A
= 29.3%, C = 35.1%, G = 12.8% and T = 22.8%. O f th e s e
nucleotides, 558 were constant and 101 were variables.
Sixty seven of the variable sites were parsimony-unin-
formative and 34 were informative under parsimony cri-
terion. The estimated uncorrected pair wise distance
(Table 1) was zero among the different Saudi Arabian
haplotypes except those from Jazan, which showed very
low distance to other Arabian haplotypes (D = 0.002).
Jazan and other Arabian individuals showed close dis-
tance to both American and Netherlands haplotypes. A
Spanish haplotype was equally distant from other Ara-
bians, Netherlands and American individuals (D = 0.006)
and more distant from Jazan haplotype (D = 0.008). Fig-
ure 2 depicts an NJ tree that has been constructed using
the aligned sequences and Neighbor-joining algorithm
[18] with Juckes-Cantor. Similar topology was obtained
by maximum-parsimony under similar conditions. An
optimal ML tree was also found with similar topology
and a negative log likelihood of 1369.479. The three
analytical methods (NJ, MP and ML) showed the ho-
mogeneity of the house sparrows globally (bootstrap =
100, 100, 99, respectively) and clustering of the Arabian

S. A. M. Amer et al. / Open Journal of Ecology 1 (2011) 73-7 6
7575
Ta bl e 1. Uncorrected pair wise distances determined between
haplotypes from different localities. “Others” refer to all Saudi
Arabian haplotypes except those from Jazan since they showed
zero distance when they compared.
Other localities Jazan USA Netherlands
Other localities -
Jazan 0.002 -
USA 0.003 0.005 -
Netherlands 0.003 0.005 0.000 -
Spain 0.006 0.008 0.006 0.006
Jazan
Taif
Qassim
Tabuk
Al-Madina
USA [AY030117]
Netherlands [AY495393]
Spain [AF230906]
P. flaveolus [L77904]
P. luteus [AY495394]
Petronia petronia [AF230914]
0.005 substitutions/site
Arar
96, 92
100, 100, 99
88, 77, 64
63, 72, 63
69, 72, 73House s
p
arro
w
Saudi Arabia
Figure 2. Neighbor-joining tree constructed from 659 bp of
cytb gene among the different studied haplotypes. The boot-
strap values are showed at nodes for neighbor-joining, maxi-
mum-parsimony and maximum-likelihood methods, respec-
tively, when they are more than 50%.
house sparrow (from different localities) in one group
with the American and Netherlands sparrows (bootstrap -
ping = 69, 72 and 73, respectively). The Spain sample
came basal to both groups. This relationship was trust-
able because of the reasonably strong statistical support.
One cannot discriminate any intra-population variation
for this bird in Saudi Arabia.
Nucleotide substitutions are generally considered in
terms of transitions and transversions. The sequenced fr-
agment of cytb gene showed 24 substitutions among the
different haplotypes of which two were transversions.
Among these polymorphic changes, 22 were in the third
position, and 2 were in the first and second po sitions an d
therefore, 2 of these su bstitutions were non-synonymous
and the 22 were synonymous. One of the non synony-
mous changes was found in only one haplotype from Al-
Madina and involved a substitution of phenylalanine
with Sereine at T191 ➝ C191. The position of this ami-
no acid in the frame of cytb gene is at 64. The second
substitution was found in different haplotypes inhabiting
Qassim, Tabuk, Jazan and Taif and showed a transition
for the amino acid number 116 from valine to isoleucine
at A346 ➝ G346 (Figure 3). It is notable that the non
Figure 3. The aligned translated amino acids of the sequenced
fragment of cytb gene for different haplotypes. The underlined
letters are those amino acids with polymorphism. Note that this
alignment is for the portion exhibited polymorphism (from 59
to 116).
synonymous mutations were found in the samples inhab-
iting the localities at the vicinity of west Saudi Arabia,
which is characterized by mountainous habitat. These
habitats are acquired abundance of vegetations and cha-
racterized by different climate (temperature, rainfall and
humidity). Further study is necessary in which more
statistical packages for more samples and data should be
used to relate these chang es to ecology.
Due to their ability to fly, birds can move over large
geographical scales, and their populations are therefore
often spatially more homogeneous than in some other
taxonomic groups [4,19,20] . The genetic variabili ty wit hin
house sparrow populations in Saudi Arabia were in the
same category as previously estimated for other areas in
the world [9,11,21]. Genetic variability within its popu-
lations was very limited probably due to the connection
between them [1]. We found a weak differenttiation am-
ong some of the population s, mainly in tho se at the west-
ern edge of the Kingdom, but other areas showed no
significant popu lation difference (Table 1). A ccordingly,
we could not differentiate any type of clusters of indi-
viduals within the collected data. Exten siv e homogeneity
of populations was most likely due to populations are
better connected. We, therefore, concluded that Saudi
Arabian house sparrow need s collection of more samples
and DNA data and manipulates these data by up to data
statistical programs to address its genetic diversity.
4. ACKNOWLEDGEMENTS
This work has been done under the financial support of Taif Univer-

S. A. M. Amer et al. / Open Journal of Ecology 1 (2011) 73-7 6 76
sity of Kingdom of Saudi Arabia (Project number 1/432/1066).
REFERENCES
[1] Kekkonen, J., Seppa, P., Hanski, I.K., Jensen, H., Vai-
sanen, R.A. and Brommer, J.E. (2011) Low genetic dif-
ferentiation in a sedentary bird: House sparrow popula-
tion genetics in a contiguous landscape. Heredity, 106,
183-190. doi:10.1038/hdy.2010.32
[2] Summers-Smith, J.D. (1988) The Sparrows: A study of
the genus passer. T and AD Poyser, Staffordshire.
[3] Anderson, T.R. (2006) The ubiquitous house sparrow:
From genes to populations. Oxford University Press, Ox-
ford. doi:10.1093/acprof:oso/9780195304114.001.0001
[4] Barrowclough, G.F. (1983) Biochemical studies of micro-
evolut ionary proc esses. In: Bru sh, A.H. a nd Clar k. J.G.A.
Eds., Perspectives in ornithology, University of Cam-
bridge Press, Cambridge, 223-261.
[5] Fleischer, C. (1983) A comparison of theoretical and ele-
ctrophoretic assessment of genetic structure in popula-
tions of the house sparrow (Passer domesticus). Evolu-
tion, 37, 1001-1009. doi:10.2307/2408414
[6] Parkin, D.T. and Cole, S.R. (1985) Genetic differentia-
tion and rates of evolution in some populations of the
house sparrow (Passer domesticus) in Australia and New
Zealand. Heredity, 53, 15-23. doi:10.1038/hdy.1985.4
[7] Bjordal, H., Cole, S.R. and Parkin, D.T. (1986) Genetic
differentiation among some populations of the house
sparrow Passer domesticus from south-western Norway.
Heriditas, 105, 107-114.
d oi :1 0.1111 /j .1 601 -5223.1986.tb00648.x
[8] Wetton, J.H., Parkin, T. and Carter, R.E. (1992) The use
of genetic markers for parentage analysis in Passer do-
mesticus (house sparrows). Heredity, 69, 243-254.
doi:10.1038/hdy.1992.122
[9] Neumann, K. and Wetton, J.H. (1996) Highly polymor-
phic microsatellites in the house sparrow Passer domes-
ticus. Molecular Ecology, 5, 307-309.
d oi :1 0.1111 /j .1 365 -294X.1996.tb00319.x
[10] Griffith, S.C., Stewart, I.R.K., Dawson, D.A., Owens, I.P.F.
and Burke, T. (1999) Contrasting levels of extra-pair pa-
ternity in mainland and island populations of the house
sparrow (Passer domesticus): Is there an “island effect”?
Biological Journal of the Linnaean Society, 68, 303-316.
doi:10.1006/bijl.1999.0343
[11] Griffith, S.C., Dawson, D.A., Jensen, H., Ockendon, N.,
Greig, C., Neumann, K. and Burke, T. (2007) Fourteen
polymorphic microsatellite loci characterized in the house
sparrow Passer domesticus (Passeridae, Aves). Molecu-
lar Ecology, 7, 333-336.
[12] Allende, L.M., Ruboi, I., Guillén, J., Ruiz-del-Valle, V.,
Guillen, J., Martinez-Laso, J. and Lowy, E. (2001) The
old world sparrows (Genus Passer): Phylogeography and
their relative abundance of nuclear mtDNA pseudogenes.
Journal of Molecular Evolution, 53, 144-154.
[13] Va n der Meij, M.A.A., de Bakker, M.A.G. and Bout, R.G.
(2005) Phylogenetic relationships of finches and allies
based on nuclear and mitochondrial DNA. Molecular
Phylogenetics and Evolution, 34, 97-105.
doi:10.1016/j.ympev.2004.09.006
[14] Dor, R. and Lotem, A. (2009) Heritability of nestling
begging intensity in the house sparrow (Passer domesti-
cus). Evolution, 63, 738-748.
d oi :1 0.1111 /j .1 558 -5646.2008.00598.x
[15] Negro, J.J. and Torres, M.J. (1999) Genetic variability
and differentiation of two bearded vulture Gypaetus bar-
batus populations and implications for reintroduction pro-
jects. Biological Conservation, 87, 249-254.
doi:10.1016/S0006-3207(98)00056-1
[16] Edwards, S.V. , Arctander, P. and Wilson, A.C. (1991) Mi-
tochondrial resolution of a deep branch in the genealogi-
cal tree for perching birds. Proceedings: Biological Sci-
ences, 243, 99-107. doi:10.1098/rspb.1991.0017
[17] Swofford, D.L. (2002) PAUP*. Phylogenetic analysis us-
ing parsimony (*and Other Methods). 4th Edition, Sun-
der-land, Sinauer.
[18] Saitou, N. and Nei, M. (1987) The neighbor-joining me-
thod: A new method for reconstructing phylogenetic trees.
Molecular Biology and Evolution, 4, 406-425.
[19] Evans, P.G.H. (1987) Electrophoretic variability of gene
products. In: Cooke, F. and Buckley, P.A. Eds., Av ian ge -
netics: A population and ecological approach, Academic
Press, London, 105-162.
[20] Ward, R.D., Skibinski, D.O.F. and Woodwark, M. (1992)
Protein heterozygosity, protein structure, and taxonomic
differentiation. Evolutionary Biology, 26, 73-159.
doi:10.1007/978-1-4615-3336-8_3
[21] Garnier, S., Durand, P., Arnathau, C., Risterucci, A.M., Es-
parza-Salas, R., Cellier-Holzem, E. and Sorci, G. (2009)
New polymorphic microsatellite loci in the house spar-
row, Passer dome sticus. Molecular Ecology Resources, 9,
1063-1065. d oi :10 .1111 /j. 1755-0998.2009.02552.x