الفهرس 
Effect of nematodes on different insect speciesOnly 14 pages are availabe for public view
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Abstract
Summary
The present investigation was performed to evaluate the effect of entomopathogenic nematode Heterorhabditis indica RM1 and its associated bacterium Photorhabdus luminescens toxin complex as environmentally safe control agent on 3rd instar larvae of the sand fly Phlebotomus papatasi. P.papatasi is the vector of Leishmania major, the causative agent of zoonotic cutaneous leishmaniasis (ZCL) in Egypt.
A- Susceptibility of the sand fly Phlebotomus papatasi to the entomopathogenic nematode Heterorhabditis indica RM1:
The third instar larvae of sand fly Phlebotomus papatasi were exposed to seven different concentrations of the nematode Heterorhabditis indica RM1, the infective juveniles were inoculated in the feeding medium of the larvae. The mortality of 3rd instar larvae was recorded 72h post infection for all concentrations.
The percent mortality was 14%, 28%, 30%, 44%, 62%, 86% and 88% at concentrations of 30, 60, 120, 250, 500, 1000 and 2000 IJs/ml, respectively. The lethal concentration inducing fifty percent mortality (LC50) reached 240 IJs/ml and the LC90 reached 2435 IJs/ml, these values were determined using the Probit procedure. The greater mortality was achieved with the highest concentration (2000 IJs/ml).
Inspection of cadavers for several weeks following infection revealed successful reproduction and migration of new infective juveniles for the indigenous isolate H. indica RM1. The harvested nematodes H. indica RM1 which migrate from the cadaver of 3rd instar larvae of P. papatasi 10 days post treatment were 103, 50, 52 nematodes/ larva for 30, 60, 120 IJs/ml, respectively.
B- Latent effect of the nematode Heterorhabditis indica RM1 on the reproductive parameter of Phlebotomus papatasi:
1. Effect of Heterorhabditis indica RM1on adult emergence of Phlebotomus papatasi:
A significant difference observed in adult emergence among the concentration 60, 240 847 and 2435 IJs/ml compared with control, being 30 ± 12.65, 16 ± 11.66, 8 ± 3.74, 6 ± 4 and 80 ± 9.49, respectively.
2. Effect of Heterorhabditis indica RM1 on longevity of adult emergence of Phlebotomus papatasi:
The concentrations 60, 240, 847 and 2435 IJs/ml that induced 30, 50, 75, 90 % lethality were determined and used. The life span of emerged females decreased significantly at concentration (60 and 240 IJs/ml) as compared with their control, being 9 ±1.96, 8.7 ± 1.20 and 19 ± 0.00, days, respectively. No females emerged after treatment of larvae with the concentration inducing 75% mortality (Table 2). The life span of treated male increased non significantly at concentration 60 IJs/ml, being 11.5 ±1.658, and decreased non significantly at concentration 240 IJs/ml, being 9.2 ± 2.177. There is high significant increase at concentration 847 IJs/ml, being 18 ±1.00 compared to the control (10 ± 1.29) days.
3. Effect of Heterorhabditis indica RM1on number of eggs laid per female of Phlebotomus papatasi
No significant difference was observed in the number of eggs/female at the concentration 60 IJs/ml, being 48.7 ± 10.67 while a significant increase difference was observed at concentration 240 IJs/ml compared with control being, , 82 ± 9.07 and 44.6 ± 6.14 eggs/female, respectively.
4. Effect of Heterorhabditis indica RM1 on number of hatchabilty eggs of Phlebotomus papatasi:
Besides decreasing the number of eggs laid/female in the treated groups, the exposure to EPN also led to the production of infertile eggs. There are no significant difference were observed between the concentration 60 and 240 IJs/ml comparing with the control, being 33.5 ± 6.03, 15.7 ± 2.88 and 41.8 ± 7.65, respectively.
C- Susceptibility of the sand fly Phlebotomus papatasi to Photorhabdus luminescens toxin complex:
The third instar larvae of sand fly Phlebotomus papatasi were exposed to diet treated with the original toxin complex corresponding to 11.39 mg/ml mortality and two other dilutions (1:2 and 1:4 corresponding to 11.39 mg/ml). The mortality of 3rd instar larvae was recorded 72h post treatment.
The toxin complex inducing LC50 among 3rd instar P. papatasi reached 9.5 mg/ml. Mortality increased as the concentration increased being about 60% at the concentrated toxin complex and 32% and zero at the dilutions 1:2 and 1:4, respectively.
D- Latent effect of the Photorhabdus luminescens toxin complex on the reproductive parameter of Phlebotomus papatasi:
1. Effect of Photorhabdus luminescens toxin complex on adult emergence of Phlebotomus papatasi:
The mean number of emerged adults showed significant decrease between concentrated toxin complex compared with control, being 26.7 ± 10.89 and 74.3 ± 11.64, respectively. However no significant difference occurred in the number of emerged adults due to the effect of the other two dilutions being 47.4 ± 13.06 and 46.5 ± 4.44 for 1:2 and 1:4, respectively.
2. Effect of Photorhabdus luminescens toxin complex on longevity of emerged adults of Phlebotomus papatasi:
The emerged adults were monitored for their duration (longevity): no significant differences were observed in female longevity compared with control being, 3.7 ± 0.88, 3.8 ± 1.07, 5.7 ± 1.30 and 4.7 ± 0.88 days, respectively. There are no significant difference observed in male longevity compared with control being, 5.7 ± 2.66, 2.2 ± 0.74, 5.5 ± 1.85 and 6 ± 1.35 days, respectively.
3. Effect of Photorhabdus luminescens toxin complex on eggs laid per female of Phlebotomus papatasi:
A significant decrease in number of eggs laid per female was observed due to protein toxin complex treatment, and the dilution 1:2 and 1:4 diluted concentration, compared with control, being zero eggs, 7.6 ± 1.83, 9.4 ± 1.60 and 46.4 ± 15.09, respectively.
4. Effect of Photorhabdus luminescens toxin complex on the egg hatching of Phlebotomus papatasi:
The hatchability of eggs deposited by females surviving larval treatment was greatly affected by the inoculum dose of the toxin complex compared to the control group. No egg hatching occurred concentrated toxin and 1:2 dilutions, while the dilution 1:4 did not affect hatchability being 93.3 ± 6.66 compared to the control 96.3 ± 2.4.
E- Histological studies:
1. Histopathology of the third larval instar treated with the nematode:
The larvae treated with LC50 of the H. indica RM1 were examined by microscopy, the greatest damage to the gut epithelium occurs in the anterior region of the midgut. Although there is some anterior-to-posterior progression of pathology during the 24-h observation, there is less cell disruption in the postereior midgut. Comparable histopathological changes occurred after 48 and 72h post treatment. However, the basal regions of the columnar cells exhibited marked blebbing and were partially separated from each other and the peritrophic matrix was no longer visible
2. Histopathology of the third larva instar of the sand fly treated with the toxin:
The midgut of the third larval instar of the sand fly treated with the prepared toxin complex showed marked histopathological changes. The wall of the intestine in sections examined after 12h exhibited uneven thickness. The brush border exhibited marked thickening. Besides, the wall exhibited focal rupturing. A noteworthy observation is that the peritrophic matrix disappeared. The sections examined after 24h showed similar changes. Besides, the epithelial cells in focal areas lost their stainability and exhibited very pale configuration. It seems that such change is a harbinger of cell death since in sections examined after 36h such focal areas were sloughed. In sections examined 72h post treatment the epithelial cells appeared dissociated from each others and lost their integrity. In certain areas the cells appeared pulled out of the basement membrane. Such aforementioned changes were represented all over the subsequent intervals i.e. 96h, 144h, 166h, 192h and 216h. However, in such periods the epithelial cells seemingly extruded cytoplasmic masses into the lumen of the midgut