” ال مكافحة ال بيولوجية لسوسة النخيل الحمراء “ “Biocontrol of the Red Palm Weevil” Presented by

1. ”المكافحة البيولوجية لسوسة النخيل الحمراء“ “Biocontrol of the Red Palm Weevil” Presented by Muhammad Mostafa Shamseldean Proferssor of Entomonematology and Agricultural Zoology, Head of the Applied Center for Entomonematodes, (ACE), Department of Agricultural Zoology and Nematology, Faculty of Agriculture, Cairo University, Giza 12613, Egypt

2. What are Nematodes? Nematodes are round or ell worms. They are unsegmented pseudocoelomate cylindrical animals which have been able to adapt to almost every habitat available to animal life. النيماتودا ديدان خيطية أوثعبانية، غير مقسمة، أسطوانية ذات تجويف جسم كاذب، وتستطيع النيماتودا التأقلم للعيش تقريباً في كل البيئات المتاحة لحياة الحيوان.

3. Heterorhabditidae Steinernematidae Neosteinernematidae Entomopathogenic nematodes

5. Why Entomopathogenic nematodes? • High pathogenicity against diverse insect pests. • Motility of infective juveniles and host finding behavior either ambusher or cruiser. • Exemption from registration according to EPA and numerous developed countries in the world. • Efficient mass production. • Ease of field application. • Safety against non target organisms. • Inexpensive and fast to develop as a product.

6. Process Heterorhabditidae and Steinernematidae • Dauer stage (infective 3rd juvenile stage) enters insect. • Releases symbiotic bacteria through the mouth. • Bacteria multiply; kill the host within 48 hours. • Nematodes feed on bacteria and bacterial products. • Develop to adults. • Produce eggs. • At second or third generation, dauers exit. • Dauer stages search for new host.

7. Field utilization against target insect pest Mass culture Survey and identification

8. Survey and identification

9. Survey of EPN • 1. Sampling Scheme • 2. Isolation of EPN

10. Soil sampling for EPN

11. Steps of the baiting technique

12. Dominant vegetations • Behera governorate • Mango, 47.4% Citrus, 33.3 % Guava, 5.3 % Apple, 13.2 % Grape, 7.9% Olive, 5.3 %

13. Mass production of EPN In vivo mass culture In vitro mass culture

14. Harvesting in the White trap (2D method)- White 1927

15. In vivo mass culture Main scenario depends on: Developing a novel technology for the in vivo mass culture LOTEK (Low cost technology) (3D Method)

16. Inoculation Ideal scenario for the in vivo technology • Conditioning Separation • Nematode • harvesting Clean-Up

17. 1) Infection: Spraying IJs is better than dipping

18. 2) Conditioning:

19. 3) Harvester of LOTEK Water drain of LOTEK

20. I 4) Separator: A • Water reservoir • Control valve • Distribution manifold • Separation screen • Concentrate collector • Waste deflector • Waste collector • To storage tank • From storage • To drain B C H D E F J G

21. Diluted & Dirty Nematode Suspension From Tap To Drain Clean & Concentrated Nematode Suspension Nematode separator

22. b a 5) The storage tank (a) and the storage tank from inside (b) is provided with a stirrer and an outer air pump.

23. In vitro mass culture • Isolation of bacteria • 2. Isolation of nematode eggs • 3. Preparation of monoxenic culture • 4. Nematode media preparation and IJs inocula

24. Nematode growth In vitro mass culture

25. Quality control of the nematode product • 1. Viability (% living nematodes). • 2. Virulence. Use the right nematode. • 3. Storage ability. Produce clean nematodes. • 4. Morphological examination • (presence of double cuticle & bacteria). • 5. Limited field tests if possible. Use the • right application method in the field.

28. Insect pests in cryptic habitats • Field utilization of EPN Such as the Red palm weevil on palm trees

29. Red Palm Weevil (RPW), Rhynchophorus ferrugineus (Olivier) is the most dangerous and deadly pest of date, coconut, oil, sago and other palms.  Classification Order:    Coleoptera Family:    Curculionidae Scincetific name:Rhynchophorus ferrugineus(Olivier) Common name: Red Palm Weevil and/or Asian Palm Weevil and Indian palm weevil. There are seven other species of this genus as follows: 1) Rhynchophorus bilineatus 2) Rhynchophorus cruentatus 3) Rhynchophorus palmarum 4) Rhynchophorus papuanus 5) Rhynchophorus phoenicis 6) Rhynchophorus schach 7) Rhynchophorus vulneratus. Target insect pest

30. 1-2 weeks 2-3 months 2-3 months Life Cycle of the Red Palm Weevil 2-3 Weeks

31. Cutting shoots leads to kairomones secretion which attract adult females of the red palm weevil to lay eggs

33. Symptoms of palm infection with RPW The symptoms due to the palm weevil infection are obvious and can be summarized in a leaky dark gummy material with a foul odor oozing out of the initial insect penetration spot and the presence of dead lower leaves due to the development and activities of the insects inside the palm tree.

34. Larvae of RPW inside and outside the tree trunck

35. Application technique (important) Nematode suspensions were injected through artificial tunnels made in an angle into the core of an infected palm tree at the top of the insect infection level. These tunnels were made by a 830 W electrical hammer drill equipped with a 45-cm long and 13-mm thick screw. A perforated PVC tube should be inserted through the artificial tunnels.  

36. The nematode juveniles were applied through two injections of 60 ml. each of nematode suspension with a disposable plastic syringe. The concentration of the nematode suspension was 3000 IJs/ml., which is equal to 1,440,000 IJs/palm tree injected through 4 tunnels in each treatment. After the nematode injections the external end of the perforated PVC tube was sealed with a tight plastic button. The injection was repeated on a weekly basis for 2 consecutive weeks. The results were recorded in the third week after the first treatment with nematodes.

37. RESULTS Table 1. Percent mortality of larvae and adults red palm weevil treated in the laboratory with local nematode isolates (1000 IJs/insect) belong to the genus Steinernema.

38. Table 2. Field efficacy of local isolates of Steinernema spp. on the red palm weevil throughout the years 2001, 2002, and 2003.

39. Table 3. Field efficacy of local isolates of Steinernema spp. against The red palm weevil from the year 2001 to 2004. Season % of tree recovery 2001 2002 2003 2004 EGG4 EBNE EGG4 EBNE EGG4 EIKF EGG4 Winter 91.7 72.3 100 83.3 86.7 78.3 87.3 Spring 88.0 76.0 83.3 79.3 88.6 79.3 89.6 Summer 83.3 75.0 83.3 75.0 79.2 76.3 82.3 Fall 100.0 77.0 100.0 75.0 82.3 83.3 100.0 Total numbers of treated trees 54 54 49 48 42 38 171 Mean percent of tree recovery + SE 88.9 + 2.24 77.8 + 4.18 91.8 + 3.17 77.1 + 2.73 83.3 + 4.05 72.2 + 3.14 90.8 + 3.12

40. Bulk of nematode juveniles emerging from cadavers of adults RPW

41. Percent tree recovery

42. Conclusions 1) Field tests have shown that Entomopathogenic nematodes (EPN) caused tree recovery rate ranging from 75 to 98% throughout the 3 years of nematode applications. 2) Variations in recovery rate can be attributed to different levels and intensity of palm tree infection with the Red Palm Weevil (RPW) and timing of nematode treatment affected by the weather conditions and activity of the pest in different seasons.

43. Commercial Jasmine oil as a novel and safe approach to control the red palm weevil Rhynchophorus ferrugineus (Oliv.) Field application The emulsion oil was used as preventive and curative methods against RPW from the season 2001 until now. Injection technique The infestation was divided to three groups (New, medium and advanced) and treated with commercial jasmine oil (table 4).

44. Table 4. Field application of commercial jasmine essential oil (10%) against red palm weevil (dosage applied per tree )

45. Date No. Of treated trees Level of infection in palm trees No. of recovered trees % of recovered trees Field application: Table 5. Evaluation of field application using oil emulsion against the red palm weevil at Ismailia governorate during the year 2003. New Medium Old New Medium Old January 17 5 10 2 4 8 2 82.35 February 12 6 1 5 4 1 3 66.66 March 24 5 10 9 3 7 7 70.83 April 25 0 19 6 0 15 5 80 May 12 3 6 3 3 5 3 91.66 June 25 4 14 7 4 12 4 80.95 July 31 5 18 8 4 16 7 87.09 August 53 11 32 10 8 27 6 77035 September 38 17 15 6 14 12 4 78.94 October 40 6 30 4 6 25 4 87.5 November 34 5 25 4 5 23 3 91.17 December 21 0 12 9 0 10 7 80.95 Total 332 67 192 73 55 161 55 81.63

46. Figure 2 . Effect of commercial jasmine oil on tree recovery from the red palm weevil infestation in the field throughout the years 2001 to 2004. 100 90 80 70 60 %of tree recovery 50 40 30 20 10 0 2001 2002 2003 2004 Years

47. CONCLUSIONS 1) The use of 10% commercial jasmine oil was lethal to immature and adult stages of the red palm weevil in the laboratory. 2) An 8% of commercial jasmine oil has a repellant effect to larvae and adults and works as an anti-feeding on larvae in the laboratory. 3) The 10% commercial jasmine oil has a curative effect when injected into infested palm trees in the field and 78 to 92% of the treated palm trees have recovered (more than 1000 trees were tested). 4) The same 10% commercial jasmine oil has a protection effect when sprayed on healthy palm trees in the field.

50. Egyptus rhynchophorus A RPW adult carrying mites Deformed Adult Red Plam Weevil Other Biocontrol Agents An ectoparasitic mite on the pupae of the red palm weevil.