International Journal of Marine Science 2015, Vol.5, No.55: 1-9
4
Figure 2 16S rRNA PCR amplified products. Lane 1: Molecular
marker; Lane 2, 5:
F. indicus
; Lane 3:
P. monodon
; Lane 4:
P.
semisulcatus
; Lane 6:
M. monoceros
3.2 Alignment and Sequences variation of the 16S
rRNA gene
The partial nucleotide sequence of the gene was
aligned along with the published sequence of the 16S
rRNA gene region of
F. indicus
(gene bank: accession
number DQ149968) (Figure 3). Analysis was carried
on 392 bp of the amplified product. The analyzed
regions of the gene were equivalent to position
(44-436) in 16S rRNA reference sequence of (Shekhar
et al., 2005).
3.3 Base composition
The base composition among species (Table 2)
showed slight differences between them, and the base
C being the rarest (average 11.43 %) where A is the
most frequent (34.14) %. (C+G) % was found to range
between 32.3% in
M. Monoceros
to 30.8% in both of
F. indicus
and
P. semisulcatus
.
3.4 Haplotype Estimation
The alignment of 72 nucleotide sequences revealed a
total of 19 haplotypes within the four penaeid species
and among the three populations of
F. indicus
(Figure 3).
The
F. indicus
populations:
F. indicus
was found to
have 11 haplotypes (from Hap.I to Hap.XI) among the
entire population. The wild population had 8
haplotypes; Baaboud population had 4 haplotypes,
while Alkhairat aquafarm contained only three
haplotypes. It was noticed that haplotype I and IV
were shared between the three locations. Haplotype I
was found in 28 out of 42 (66.6%) individuals of
F.
indicus
, hence it is the most frequent haplotype, and it
was 100% aligned with the Gene bank reference
(DQ149968) without any insertion or deletion, while
Haplotype IV aligned with 99.7%. The wild population
appeared to have more variable sites with constant
nucleotide substitution at positions 50(G-C), 105(T-G),
285(C-A), 400(C-T), 410(T-C) and 438(T-G) respectively.
Other minor or unique substitutions were also observed.
The
P. monodon
populations: Only two haplotypes
(Hap.XII and XIII) were obtained from 18 individuals
of
P. monodon
, the frequent haplotype is Hap.XIII
which is found to be dominated through 15 sequences
(i.e. in 83.3% of population). The
P. semisulcatus
populations: 5 haplotypes were encountered from
P.
semisulcatus
.The
M. monoceros
: only one haplotype
(Hap.XIX) was recorded for
M. monoceros
species.
Indel position of a codon (ATA) at position 302, 303
and 304 respectively was observed when comparing
M. monoceros
with the other three
Penaeus
species;
and another indel of (A) nucleotide at position 319.
3.5 Genetic differentiation and distance
The genetic differentiation (Fst) between the three
populations of
F. indicus
(intra-specific population)
was 0.10957, 0.12459 and 0.14817. While the genetic
distance between
F. indicus
and other species was
tabulated in Table 3.
The genetic differentiation (Fst) between
P. monodon
populations from north (Mersa Halaka) and south
(Mersa Heidub) was recorded as 0.36585.
The gene flow (Nm) is higher between the different
populations in the same species as shown in Table 4.
While the genetic differentiation (Fst) between
F.
indicus- P. monodon, F. indicus -P. semisulcatus and F.
indicus-M. monoceros
was found to be 0.95932,
0.94655, and 0.99016 respectively.
Table 2 Base Composition (%) of 16S rRNA Gene among 4 species of Penaeid shrimps
16S rRNA gene
Species
A%
C%
G%
T%
C+G(%)
A+T(%)
Site
F. indicus
35.5
11.5
19.4
33.4
30.8
69.1
392
P. monodon
35.3
11.8
19.2
33.8
30.9
69
391
P. semisulcatus
34.4
11.5
19.7
34.4
30.8
69.1
391
M. monoceros
31.3
10.9
21.4
36.4
32.3
67.7
393
Mean
34.14
11.43
19.93
34.5
31.2
68.73
