الفهرس 
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Abstract
SUMMARY AND CONCLUSION
Root-knot nematodes, Meloidogyne incognita are responsible for a great economic losses in many agricultural crops and are considered one of the most difficult pests to control. Many harmful pesticides are used in their control. The aim of this study is to evaluate the nematicidal activities of plant extracts of the following plant materials:
1- Roots of Saponaria officinalis
2- Seeds of Peganum harmala
3- Seeds of Trigonella foenum-gracum
4- Leaves of Artemisia annua
5- Leaves of Nerium oleander
The powder of these plant materials were stepwise extracted by hexane then ethyl acetate then methanol and finally distilled water. This study included two objectives:
( I ): In vitro experiment: including evaluation of the plant extracts as natural nematicides against root-knot-nematode.
( II ): Greenhouse experiment: including evaluation of the ability of the most effective extracts to induce resistance in the cucumber plant against root-knot nematode.
( I ): In vitro experiment: including evaluation of the plant extracts as natural nematicides against root-knot-nematode.
(A) Effect of plants extracts on root-knot nematode mortality:
A primary in vitro experiments were carried out to determine the effect of different concentrations (5, 10, 15, and 20 mg/ml) of all plant fractions (hexane, ethyl acetate, methanol and distilled water) on larval mortality after different periods of treatments (24, 48 and 72 hrs). The results clearly indicated that the percentage of juvenile mortality increased with increasing extracts concentration and exposure time. Moreover, aqueous fractions in all plant materials and methanolic fraction of Trigonella foenum-gracum led to higher percentage of mortality. The percentage of mortality reached 100% when nematode larvae were treated
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SUMMARY AND CONCLUSION
Hanaa R. M. M. Farag (2017), Ph. D., Fac. Agr., Ain Shams Univ.
with 10mg/ml concentration of aqueous fractions of Peganum harmala seeds, Nerium oleander leaves for 72 h. while, treatment with 15mg/ml of aqueous fractions of Artemisia annua leaves, Saponaria officinalis roots and Trigonella foenum-gracum seeds caused the same mortility after 72 hours of treatments . Also, methanolic fraction of Trigonella foenum-gracum seeds (10 mg/ml) caused a complete mortality of nematode larvae after 48 h.
(B) Effect of plants extracts on nematode egg hatchability:
The effects of different concentrations (5, 10, 15, and 20 mg/ml) of aqueous fractions, mixture of all fractions and methanolic fraction of Trigonella foenum-gracum on nematode egg hatchability were investigated after different periods of treatments (24, 48 and 72 hrs). The results showed that egg hatchability was declined due to treatments of nematode eggs with the aqueous plant extracts or their mixture. The concentration of 20 mg/ml of the mixture of aqueous fractions and aqueous fractions of Saponaria officinalis, Artemisia annua, Nerium oleander, and Trigonella foenum-gracum completely prevented nematode egg hatching up to 72 hours after treatment.
(C) Effect of plant extracts on acetylcholinesterase activity in root-knot nematode:
Nematode acetylcholine esterase (AChE) activity was estimated after exposure of second stage juveniles (larvae) to 20 mg/ml of aqueous fractions of all plant materials, a mixture of all aqueous fractions and methanolic fraction of Trigonella foenum-gracum and compared to the well-known nematocide oxamyl. The results showed that AChE was inhibited due to exposure to oxamyl as well as all aqueous fractions except Peganum harmala which led to a higher activity of AChE than control. The maximum inhibition of AChE was recorded in nematode larvae treated with the aqueous fraction of Nerium oleander followed by Trigonella foenum-gracum, Saponaria officinalis and the mixture of all
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SUMMARY AND CONCLUSION
Hanaa R. M. M. Farag (2017), Ph. D., Fac. Agr., Ain Shams Univ.
aqueous extracts which exhibited higher inhibition percentage than oxamyl.
( II ): Greenhouse experiment: including evaluation of the ability of the most effective extracts to induce resistance in the cucumber plant against root-knot nematode.
To evaluate the ability of these plant extracts to induce plant resistance against root-knot nematodes, a greenhouse experiment was carried out. Nematode-infected cucumber seedlings were treated with aqueous fractions of all plant materials, mixture of all aqueous fractions and methanolic fraction of Trigonella foenum-gracum. The effects of these plant extracts on nematode infected and non-infected plant were investigated and compared to that of chemical nematocide oxamyl. The results can be summarized in the following points:
(1) Root-knot nematode population (Meloidogyne incognita).
The results showed that all treatments significantly reduced the number of galls, egg masses, and females, hatched nematode of M. inconita in infected cucumber plants as compared to infected control. Oxamyl and mixture of all aqueous fractions treatments caused the lowest reproduction factor of root-knot nematode in infected roots (93.2 and 92.1%) followed by methanolic fraction of Trigonella foenum-gracum and aqueous fraction of Saponaria officinalis with reproduction factors of 89.3 and 78.7%.
(2) Cholesterol content in nematode egg masses and females in infected root.
The results revealed that aqueous fraction of Saponaria officinalis, Artemisia annua, Trigonella foenum-gracum, mixture of all aqueous fractions and Trigonella foenum-gracum methanolic fraction led to significant reduction in cholesterol content in egg mass of M. inconita. In contrast, aqueous fraction of Peganum harmala and Nerium oleander did not cause any significant change in cholesterol content in egg mass of M.
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Hanaa R. M. M. Farag (2017), Ph. D., Fac. Agr., Ain Shams Univ.
inconita. All treatments, caused significant reduction in cholesterol content in females of M. inconita and the maximum reduction was observed with aqueous fraction of Artemisia annua and mixture of all aqueous fractions. In contrast, oxamyl treatment did not cause any significant change in cholesterol content in egg masses and female of M. inconita.
(3) Effect of plant extracts on growth parameter of cucumber plants
(A) Effect of plant extracts on root and shoot length
Nematode infection of cucumber seedlings led to significant reduction in shoot and root length of infected seedlings. Also, the results showed that aqueous fractions of Saponaria officinalis, Artimisia annua, Trigonella foenum-gracum, mixture of all fraction and Trigonella foenum gracum methanolic fraction enhanced root length of infected and non-infected plant. Also, aqueous fraction of Saponaria officinalis, mixture of all fractions and Trigonella foenum-gracum methanolic fraction enhanced shoot length of infected and non-infected plant. In addition, aqueous fraction of Nerium oleander and Trigonella foenum-gracum led to significant increase in shoot length in infected plants compared to control.
(B) Effect of plant extracts on root and shoot weight
Infection of cucumber plants with root-knot nematodes (M. inconita) led to significant reduction in shoot weight and significant increase in root weight due to galls formation. The results showed that aqueous fraction of Saponaria officinalis, Trigonella foenum-gracum, a mixture of all fractions and Trigonella foenum-gracum methanolic fraction caused significant increase in shoot weight and significant decrease in root weight of infected cucumber seedlings. While in healthy plants, aqueous fraction of Saponaria officinalis, Trigonella foenum-gracum, a mixture of all fractions and Trigonella foenum-gracum methanolic fraction caused significant increase in shoot and root weight of cucumber seedlings.
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Hanaa R. M. M. Farag (2017), Ph. D., Fac. Agr., Ain Shams Univ.
(4) Lipid peroxidation
The results showed that infection of cucumber seedlings with M. inconita led to significant increase in lipid peroxidation in roots and leaves compared with control. All treatments alleviated nematode induced lipid peroxidation in roots and leaves except treatment with aqueous fraction of Trigonella foenum-gracum which had no significant effect in the level of lipid peroxidation in cucumber roots as compared to control.
(5) Proline content
The results showed that infection of cucumber seedlings with M. inconita led to significant increase in proline content in roots. In contrast, infection of cucumber seedlings caused significant reduction in proline content in leaves compared with control. Treatments of infected seedlings with different plant extracts resulted in different changes in proline contents in roots and leaves. For instance, treatment with Trigonella foenum-gracum aqueous fraction led to increase in proline content in roots of infected cucumber seedlings compared with control. Also, treatment with Peganum harmala led to significant increase in proline in leaves of infected cucumber seedlings compared to control.
(6) Phenolic compounds
The results indicated that infection of cucumber seedlings with M. inconita led to significant increase in phenolic compounds in roots and leaves. Phenolic compounds were increased in cucumber seedlings by treatment with aqueous fraction of Trigonella foenum-gracum, Peganum harmala and mixture of all fractions in leaves of infected groups compared with control. While in roots, all treatments caused significant decrease in phenolic compounds in infected cucumber seedlings.
(7) Phenylalanine ammonia-lyase (PAL) activity
The results revealed that infection of cucumber seedlings with M. inconita led to significant increase in PAL activity in roots and leaves. In roots of infected cucumber plants, treatment with aqueous fractions of
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SUMMARY AND CONCLUSION
Hanaa R. M. M. Farag (2017), Ph. D., Fac. Agr., Ain Shams Univ.
Peganum harmala, Artemisia annua, Nerium oleander, Trigonella foenum-gracum and mixture of all aqueous fractions led to significant decrease in PAL activity, whereas there were no significant changes due to treatment with Saponaria officinalisa aqueous fraction and Trigonella foenum-gracum methanolic fraction. While in leaves of infected cucumber plants treated with aqueous fractions of Saponaria officinalis, Artemisia annua and Nerium oleander, the level in PAL activities were higher than control.
(8): Antioxidant defensive enzymes:
(A) Peroxidase activity
The results showed that infection of cucumber seedlings with M. inconita led to significant increase in POX activity in roots and shoots. Treatment of infected cucumber plants with aqueous fractions of Saponaria officinalis, Peganum harmala, Artemisia annua and Trigonella foenum-gracum led to significant increase in POX activity in roots compared to control, while there was no significant change in POX activity in plants treated with Nerium oleander aqueous fraction and mixture of all aqueous fractions. In addition, treatment with Trigonella foenum-gracum methanolic fraction caused significant decrease in POX activity. In leaves, all treatments of infected cucumber plants led to significant decrease in POX activity except aqueous fraction of Trigonella foenum-gracum and oxamyl which caused a significant increase in POX activity.
(B) Catalase activity
The results indicated that infection of cucumber seedlings with M. inconita did not cause significant change in CAT activity in roots while there was significant increas in leaves. Aqueous fractions of Peganum harmala, Nerium oleander and Trigonella foenum-graecum and methanolic fraction of Trigonella foenum-graecum led to significant increase in CAT activity in non-infected roots. Also, all treatments caused significant increases in CAT activity in infected cucumber root. In leaves,
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SUMMARY AND CONCLUSION
Hanaa R. M. M. Farag (2017), Ph. D., Fac. Agr., Ain Shams Univ.
only treatments with the mixture of all aqueous fractions and methanolic fraction of Trigonella foenum-graecum led to significant increase in CAT activity in infected cucumber seedlings.
(C) Polyphenol oxidase
The results showed that infection of cucumber seedlings with M. inconita led to significant increase in PPO activity in roots and leaves. Aqueous fraction of Saponaria officinalis, Nerium oleander and mixture of all aqueous fractions led to significant increase in PPO activity in non-infected seedlings in roots. Under root-knot nematode infection conditions, the treatment of infected cucumber seedlings with aqueous fractions of Saponaria officinalis, Artimisia annua, Nerium oleander, methanolic fractions of Trigonella foenum-graecum and oxamyl caused significant increase in PPO activity in roots. In leaves, treatment of infected cucumber seedlings with aqueous fraction of Peganum harmala, Nerium oleander and oxamyl caused significant increase in PPO activity. In contrast, treatment of infected cucumber seedlings with aqueous fraction of Saponaria officinalis, mixture of all aqueous fractions and Trigonella foenum-graecum mathanolic fraction led to significant decrease in PPO activity.
(D) Superoxide dismutase
The results revealed that infection of cucumber seedlings with M. inconita led to significant increase in SOD activity in roots and leaves. Also, the results showed that all treatments led to significant increase in SOD activity in non-infected seedlings in roots. Under root-knot nematode infection conditions, all treatments did not cause significant increase in SOD activity in roots while, the treatment with oxamyl led to significant increase in SOD activity. In leaves, treatment with aqueous fractions of Peganum harmala, Trigonella foenum-graecum, methanolic fraction of Trigonella foenum-graecum and mixture of all aqueous fractions caused significant decrease in SOD activity in infected
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SUMMARY AND CONCLUSION
Hanaa R. M. M. Farag (2017), Ph. D., Fac. Agr., Ain Shams Univ.
cucumber seedlings, while treatment with oxamyl led to significant increase in SOD activity.
In conclusion results clearly declared that aqueous fractions of these plant materials can be used as a natural nematicidal and can be an effective alternatives of chemical nematicidal. These extracts control root-knot nematodes through different biochemical pathways: (1): Their reductive effects on cholesterol accumulation in egg masses and females which led to a remarkable reduction in egg viability and hatching. (2): Their inhibitory effects on AChE which led to nematode paralysis and death. (3): Induction of plant tolerance through their effects on phenols accumulation and antioxidant defensive enzymes.