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Tikrit Journal of Pure Science 23 (9) 2018
ISSN: 1813 – 1662 (Print)
E-ISSN: 2415 – 1726 (On Line)
1
Tikrit Journal of Pure Science
Journal Homepage: http://main.tu-jo.com/ojs/index.php/TJPS/index
Effect of some plant extracts on the first and second larval instar
of Culex pipines pipines L.
Estabraq Mhmood Mahdi
Department of Biology , College of Science , Tikrit University , Tikrit , Iraq
A r t i c l e i n f o.
Article history:
-Received: 29 / 4 / 2018
-Accepted: 10 / 6 / 2018
-Available online: / / 2018
Keywords: Culex pipiens pipiens
L, Plant extracts, mortality
percentage.
Corresponding Author:
Name: Estabraq Mhmood Mahdi
E-mail: Abk_mm@yahoo.com
Tel:
Abstract
The present study aimed to study some alternatives chemical
pesticides and observation their effect on the life of the first and second
larva instar of Culex pipiens pipiens L. for this reason, this species of
mosquitoes was raised in the breeding cage, which was locally produced.
The plant extracts of the seeds of the Nigella sativa, Azadirachta indica,
Sesbania sesban and Lagenaria vulgaris, the larvae of the first and
second instar of mosquitoes were treated with plant extracts by
concentrations of 100, 500, 1000, 5000, ppm. The study showed different
results according to the concentration and duration of treatment. The
treatment with plant extracts showed that the Nigella sativa seed extract
is higher than the rest of the plant extracts. The mortality percentage was
93.3% in the concentration of 5000 ppm after 24 hours of treatmentThe
mortality percentage for the same extract was 100% at the concentration
of 5000 ppm within 48 hours of treatment for the first larval instar, while
the mortality percentage for the extract were 100% at 5000ppm
concentration during 24 and 48 hours of treatment for the second larval
phase.
While the Lagenaria vulgaris seed extract was the least affected from the
rest of the extracts on the larvae, where the mortality percentage 26.66,
33.33% in the concentration of 5000 ppm percentage 24, 48 hours of
treatment respectively for the first larval instar. For the second larvae, the
results were 50% and 86.66% at 5000 ppm concentration after 24, 48
hours of treatment respectively for extract of pumpkin plant
Introduction
The acquisition of disease-tolerant medicinal insects
forms a major threat to human and animal health as
well as its damage to insect control programs [1].
This problem has evolved over time, prompting
researchers to find alternatives that are non-toxic to
humans and animals [2],[3].
Mosquitoes are considered to be medical insects that
have been resistant to all pesticides in many different
places [4]. The human problem with mosquitoes has
existed since ancient times. If humans did not know
mosquitoes, An inconvenience and anxiety for his
rest is enough to be considered a problem in itself.
Culex pipines pipines L. is one of the mosquito
species known to be superior to humans and is
located close to their habitat. It favors environments
that contain a high concentration of nitrogen, such as
wastewater, sewage, rich water and ponds, with their
organic matter content [5]. The importance of
mosquitoes to humans is medically responsible for
the spread of many human and animal diseases, such
as malaria, which kill about 3 million people
annually" [6]. yllow fever, dengue fever, raftva fever
filariasis, It is caused human [7],[8] Many plant
extracts have been used as alternatives to chemical
insecticides in the killing and control of insects where
humans used plant flowers containing toxic
substances in their leaves and seeds directly through
the use of vegetable powder or after extraction with
organic solvents.Increased insect control requires
strategies to extend the use of effective vector control
compounds, so using a combination of insecticides
with other insecticides or with plant compounds is
one of these strategies for mosquito control. And that
the use of the mixture is more effective "than the use
of manufactured insecticides or plant compounds
alone [9].
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Tikrit Journal of Pure Science 23 (9) 2018
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E-ISSN: 2415 – 1726 (On Line)
2
Materials and methods
- Collecting and breeding insects:
In this study, a strain of Culex pipiens pipiens L.
larvae collected from the ponds was used for the
aquifers of the crossing area in Al-Dur city, where the
immature stages (larvae and Pupae) were collected by
means of nets consisting of a wooden arm not
exceeding a meter long with a circular metal mesh
similar to a sieve its about 10 cm. The samples were
transported by glass bottles to the breeding room,
which is a wooden cage made up of 5 transparent
plastic tubs. On the sides of the cage is a thin mesh
that does not allow mosquitoes exit. The dimensions
of the cage are 30 × 180 × 90 cm, the larvae are
placed in plastic tubing capacity to show [10]. The
method was used to place a Zajel-type bird in the
cage, tied to its legs and wings. It was used for the
purpose of feeding. For the purpose of obtaining
eggs, and removed the feathers of the chest area to
make it easier for mosquitoes to absorb blood, and
continue feeding for 12 hours in the dark, and for the
purpose of obtaining a pure farm eggs were
transferred to the breeding site until hatching and get
larvae [11].
- Collection of plant samples:
The required plant parts percentage in the present
study percentage from Tikrit and Al-Dour city, which
is the seed of the Niglla sativa, Azadirachta indica, ,
sesbania sesban and the Lagenaria vulgaris. These
plants were classified into the herbarium of the
College of Science - University of Tikrit under the
taxonomic numbers (11,10,9) respectively. This plant
was washed and cleaned of dust and dirt and dried in
the laboratory at 25 ° C. dried plants were kept in
bags until the extraction process began
Table (1) Plants seeds using in the study
English Name
The scientific name
Part used
Nigella
Nigella sativa
Seeds
Neem
Azadirachta indica
Seeds
Acacia
Sesbania sesban
Seeds
Pumpkin
Lagenaria vulgaris
Seeds
- Preparation of plant extracts:
Plant extracts were prepared in the laboratory of
College of Agriculture, University of Tikrit. The
seeds of the plants used in the current study were
grinded by Miller Electric and took 100 g of black
seed, neem, seespan and pumpkin seed powder
separately and placed in a special thimble [12]. The
500 mL flask has 250 ml of petroleum ether at 95%
concentration and the extract period lasted 48 hours
and at 55-60 °C.after extraction, the extract it was
concentrated in the vacuum rotary evaporator at the
Central Research in Tikrit university, at 55-60 ° C
temperature. to get rid of the residue of the solvent
used and to obtain a gelatinous fluid, repeat the
process was repeat for a number of times until To
obtain a sufficient amount of concentrated extract, the
extract was subsequently placed in dark glass bottles
and stored in the refrigerator to be ready for use.[13].
- Preparation required concentrations
The required concentrations were obtained by taking
1 g of the seed extract of the plant mentioned above .
each part was placed in a 100 ml glass container with
99 ml of distil water, and then Polysorbate was added
in the form of drops on the mixture Distil water and
extract) until the mixture is suspended in a way that is
easy to mix and brush, a concentration solution of 1%
or 10,000 ppm was abtained. According to the
dilution formula of C1V1 = C2V2, the other required
concentrations of 5000, 1000, 500, 100 ppm The
control treatment used only distilled water
- Studying the effect of plant extracts on the
destruction of the first and second larvae of Culex
pipiens pipiens L
I took the larvae of the first and second instar of
Culex pipiens pipiens L. were took to know the
deadly effect of plant extracts where the larvae of
these two instars before the transition to the third
instar were took and placed in glass containers of 500
ml and containing 100 ml of plant extract with
required concentrations by three replicates for each
concentration, and 10 larvae of each repeater, as well
as the case for control treatment, and should be taken
into consideration the calculation of the amount of
water preeding when preparing the required
concentrations and added to the repeaters 0.01 of feed
rabbits for the purpose of feeding the larvae, then the
larvae were observed and the results in this test were
recorded after 24 and 48 hours of treatment
- Statistical analysis
The results were statistically analyzed by using
ANOVA in the implementation of experiments. The
Duncan Multiplication test was carried out at a 5%
probability level to ascertain the differences between
the various parameters [14].
Results and discussion
1- The plant extracts efficacy of the first larval
instar after 24 hours of treatment.
The result in table 2 shows the percentage mortality
of plant extracts after24 hours the treatment of the
first larval instar of mosquitoes. Pipiens pipiens L. in
the concentrations of 100, 500, 1000, 5000, ppm,
where the results showed that there is an effect of the
plant extracts used in this study on the death of larvae
of the first instar of mosquitoes
The results of the statistical analysis showed that
there are significant differences in the toxicity of
plant extracts based on the type of plant extract used
in the study, where the percentage mortality increase
with a direct increase of concentration and that there
is a difference "between the plants to make the impact
so that the superiority of a plant on another plant, this
is due to the difference in the quality and quantity of
active compounds contained in the plants that affect
the nervous system of the insect, that paralyzes its
movement, leading to death or affect the mechanism
of action of the necessary enzymes responsible for
important biological processes, causing the cessation
of metabolism and death [15].The results in Table (2)
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Tikrit Journal of Pure Science 23 (9) 2018
ISSN: 1813 – 1662 (Print)
E-ISSN: 2415 – 1726 (On Line)
3
showed that the Nigella sativa seed extract gave
higher effect in the insect than the rest of the plant
extracts, with the highest mortality percentage of
93.3% at the concentration of 5000 ppm and the
lowest mortality percentage of 40% in concentration.
this is corresponding with [16]. among the show
cause of the larvae is due to the effect of certain plant
compounds in the killing of the epithelial cells of the
central digestive tract of the insect fed to these
compounds or that these compounds have high
toxicity affecting in the nerve tissue of the larvae and
cause paralysis and death
Also the results of Table 2 showed that the
Azadirachta indica extract came after the Nigella
sativa extract in effect on the insect. The highest
mortality percentage was 90% at the concentration of
5000 ppm and the lowest mortality percentage was
23.3% at 100 ppm. This study was agreed with a
study by [17]. On the use of Neem seed oil extract in
the control of mosquitoes and the concentration
(0.01%) led to 100% killing within 24 hours. The
results showed that the extract of Seesban was
followed the Neem extract in terms of effect. The
highest mortality percentage was 50% at 5000ppm
concentration and the lowest mortality percentage
was 10% at 100 ppmThe results showed that the
extract of the seeds of the plant pumpkin is the lowest
plant extracts impact in the insect with the highest
mortality rate 26.66% at the concentration of 5000
ppm and the lowest mortality percentage of (0) at 100
ppm concentration (Figure 1-A)
Table (2) The mortality percentage of first larval instar treated withplant extracts after 24 hours.
Extracts
Mortality percentages of concentration in ppm
Average plant
Nigella sativa
F
D
bc
a
75.8
A
Azadirachta indica
G
E
cd
90
bc
64.9
B
Sesbania sesban
H
I
G
50
cd
24.9
C
Lagenaria vulgaris
I
I
H
26.66
g
19.9
D
Average concentration
18.3
D
38.3
C
54.1
B
64.9
a
*Similar small letters in one line (horizontally) mean no significant differences
**Similar capital letters in one column (vertically) mean no significant differences.
(Using Dankin test)
2- The plant extracts efficacy of the first larval
instar after 48 hours of treatment.
The results of Table 3 showed that the Nigella sativa
extract was superior to the rest of the others in its
effect on the larvae, with the highest mortality
percentage of 100% at the concentration of 5000 ppm
and the lowest mortality percentage of 43.33% in the
concentration of 100ppm. After 48 hours of treatment
due to the duration of the exposure that caused an
increase in mortality percentage. the increasing of the
duration of exposure to the active agent, leads to
increase of mortality percentage. These results are
agreed with the results of [1]. which showed that the
duration of exposure to the active substance has the
greatest dose effectThe results indicated in Table (5)
that the extract of the Azadirachta indica came after
the Nigella sativa extract in the effect on the larvae
with the highest mortality percentage 96.66% at the
concentration of 1000 ppm and the lowest one
30.33% at 100 ppm concentration. This study is
consistent with the findings of []. in the treatment
of the Schistocerca gregaria with the Azadirachta
indica extract. The percentage of mortality increased
to 75%. The treatment of locusts with neem oil
reduced their ability to fly and the locusts could not
return to its normal state after months of treatment.
The results of Table (3) showed that the extract of
Sesbania sesban came after the Neem extract in terms
of its effect on the insect with the highest mortality
percentage 40% in the concentration 5000 ppm and
the lowest one was 16.66% in the concentration 100
ppm. The results showed that the Lagenaria vulgaris
extract is the lowest plant extracts in the larvae, with
the highest mortality percentage being 33.33% at the
concentration of 5000 ppm and the lowest mortality
percentage of 10% in the concentration of 100 ppm
(Figure 1-B).The reason of the low effect of the plant
extract on the larvae compared with the other extracts
is due to The quantity and quality of the active
substances present in the extracts, which can be
accessed through the cotyledon of the insect body,
reaching the vital target that can affect it, leading to
an end to the paralysis of the insect and then its death.
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E-ISSN: 2415 – 1726 (On Line)
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Table (3) The mortality percentage of first larval instar treated withplant extracts after 48 hours.
Extracts
Mortality percentages of concentration in ppm
Average plant
Nigella sativa
43.33
e
86.66
C
96.66
Ab
100
A
81.6
A
Azadirachta indica
30.33
f
E
90
a
96.66
Ab
66.7
B
Sesbania sesban
16.66
Hg
30.33
F
33.33
f
40
e
30.2
C
Lagenaria vulgaris
10
H
20
Fg
26.66
g
33.33
f
22.4
D
Average concentration
25.2
d
46.7
C
61.6
b
67.4
a
*Similar small letters in one line (horizontally) mean no significant differences
**Similar capital letters in one column (vertically) mean no significant differences.
(Using Dankin test)
3- The plant extracts efficacy of the second larval
instar after 24 hours of treatment.
Table (4) shows the mortality percentage of plant
extracts after 24 hours treatment for second instar
larvae. Culex Pipiens pipiens L. In the concentrations
of 100, 500, 1000, 5000, ppm, where the results
showed a mixed effect on the type and concentration
of plant extracts used in the study on the mortality of
larvae of the second instar , the results showed the
sensitivity of the larvae to the plant extracts used in
the study. It was found that there was a direct
correlation between the mortality percentage and the
concentration where the concentration is raised the
mortality percentage. The exposure of mosquito
larvae to the extracts results to paralysis in the insect
body and its inability to feed. This is due to that Plant
extracts have a high toxicity effect on the nervous
system (neurotoxins) or may interfere with it [20].
The results of table (4) showed that the plant extract
of the Nigella sativa was the most effective from the
rest of the plant extracts in terms of its effect on the
insect where the highest mortality percentage was
100% of concentration 5000 ppm and the lowest
mortality percentage 43.33% in the concentration 100
ppm, The results showed that "Neem extract come
after from the Nigella sativaextract in terms of its
effect on larvae of the second stage. The highest
mortality percentage was 96.66% at the concentration
of 5000 ppm and the lowest killing rate was 33.33%
at 100 ppm concentration. Neem in its effect on the
larvae, the highest rate of killing 60.00% at the
concentration of 5000 ppm, and the lowest mortality
percentage 20,00% at 100 ppm concentration. The
results showed that the extract of pumpkin plant is the
lowest plant extracts effect on the larvae, giving the
highest mortality percentage 50% in the concentration
of 5000 ppm and the lowest mortality percentage
10.00% in the concentration 100 ppm after 24 hours
of treatment (Figure 1-C).
Table (4) The mortality percentage of second larval instar treated withplant extracts after 24 hours.
Extracts
Mortality percentages of concentration in ppm
Average plant
Nigella sativa
f
D
bc
A
79.9
A
Azadirachta indica
g
E
cd
b
69.1
B
Sesbania sesban
fg
30.33
G
43.33
ef
60
d
38.4
C
Lagenaria vulgaris
10
H
13.33
H
23.33
hg
50
e
24.1
D
Average concentration
26.6
d
47.5
C
60.8
B
76.6
a
*Similar small letters in one line (horizontally) mean no significant differences
**Similar capital letters in one column (vertically) mean no significant differences.
(Using Dankin test)
4- The plant extracts efficacy of the second larval
instar after 48 hours of treatment.
The results of table 5 showed that the Nigella sativa
extract was the most effective on the larvae of the
second instar of mosquitoes compared with the rest of
the extracts. The highest mortality percentage was
100% at the concentration of 5000 ppm and the
lowest mortality percentage was 80% at 100 ppm
concentration The results showed that there was a
significant increase in the lethal effect of plant
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Tikrit Journal of Pure Science 23 (9) 2018
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E-ISSN: 2415 – 1726 (On Line)
5
extracts after 48 hours of larval treatment, where the
mortality percentage of the Nigella sativa extract was
higher than the mortality percentage after 24 hours of
treatment, this is due to the duration of exposure the
results were in line with the study [21]. which showed
that the highest percentage of larva death e was
66.7% at the concentration of 250 ppm and the lowest
mortality percentage of larvae reached 15.4% at 50
ppm concentration. the results showed that the
Azadirachta indica extract came after the Nigella
sativa extract in terms of the effect on the larvae.
Table (5) was the highest mortality percentage
96.66% at the concentration of 5000 ppm and the
lowest mortality percentage 66.66% at 100 ppm
concentration, the results of the study were agreed
with the results of [22]. which is used a 5% Naem
emulsion to control mosquitoes in India and it gave
good results. The results showed that the extract of
the Seesban plant was came after the Neem extract in
terms of the effect on the larvae of the second stage.
The highest mortality percentage was 93.33% at the
concentration of 5000 ppm and the lowest mortality
percentage was 50% at 100 ppm concentration. On
the larvae, giving the highest mortality rate of 86.66%
in the concentration of 5000 ppm and the lowest
mortality rate of 33.33% in the concentration of 100
ppm, The extract of the pumpkin plant has a little
effect on mosquito larvae the cause due to the amount
of active substances found in plant extracts that can
be accessed through the insect body. (Figure 1-D).our
study showed the mortality percentage of larvae
increased within 48 hours of treatment compared to
treated within 24hours. We conclude that the duration
of exposure has a significant effect on the efficacy of
the extracts used, which led to an increase in
mortality rates
Table (5) The mortality percentage of second larval instar treated withplant extracts after 48 hours.
Extracts
Mortality percentages of concentration in ppm
Average plant
Nigella sativa
80.00
f
90.00
D
100
bc
A
92.5
A
Azadirachta indica
66.66
g
80.00
E
93.33
cd
b
84.1
B
Sesbania sesban
50
fg
66.66
G
80.00
ef
93.33
d
72.4
C
Lagenaria vulgaris
33.33
H
43.33
H
66.66
hg
86.66
e
57.4
D
Average concentration
57.4
d
69.9
C
84.9
B
89.1
a
*Similar small letters in one line (horizontally) mean no significant differences
**Similar capital letters in one column (vertically) mean no significant differences.
(Using Dankin test).
Figure (1) The First and Second Larval instar Mortality Percentage treated with plant extracts.
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Tikrit Journal of Pure Science 23 (9) 2018
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Culex pipiens pipiens L.
Culex pipiens pipiens L.
ppm
ppm
ppm
ppm
ppm
ppm
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