الفهرس 
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Abstract
The purpose of this study was to survey culicine mosquito larvae from rural breeding sites in Giza and Beheira Governorates and an urban breeding site in Gharbia Governorate to determine culicine species composition and their relative abundance. Physicochemical parameters and insecticide residues of waters in these breeding sites were analyzed. In addition, the resistance levels of field-collected populations of Cx. pipiens and susceptibility to commonly used insecticides in Egypt were determined. Resistance-impact on the fitness parameters, biochemical composition as well as expression of genes relevant to insecticide resistance in the field populations of Cx. pipiens were evaluated. VI.1. Species composition and dominance of the culicine mosquitoes in rural and urban breeding sites A total of 1500 larvae were sampled from the rural breeding sites in Giza and Beheira Governorates and urban breeding site in Gharbia Governorate. Five Culex species were identified in the rural Giza Governorate breeding site. They were Culex pipiens Linnaeus comprising 95.0% of all specimens collected, Culex univittatus Theobald (2.6%), Culex antennatus (Becker) (1.2%), Culex quinqueifasciatus Say (0.8%), and Culex perexiguus Theobald (0.4%). The three Culex species identified in the rural Beheira Governorate breeding site were Cx. pipiens (97.6%), Cx. antennatus (1.6%), and Cx. univittatus (0.8%). In the urban Gharbia Governorate breeding site, five Culex species were identified namely Cx. pipiens (85.6%), Culiseta longiareolata Macquart (6.0%), Cx. univittatus (4.6%), Cx. antennatus (3.2%), and Cx. quinqueifasciatus (0.6%). In both rural and urban breeding sites surveyed, Cx. pipiens was eudominant, whereas, Cx. antennatus was minor. Based on Bray-Curtis’ dissimilarity analysis, rural and urban breeding sites tested were similar to each other in Culex species composition. VI. 2. Ecological parameters (diversity indices) in the sites Urban breeding site in Gharbia governorate had the highest value of Culex species diversity and equitability (H´: 0.58 and E: 0.33). On the other hand, the rural breeding site in Beheira governorate had the lowest diversity and equitability values (H´: 0.13 and E: 0.07). Regarding the Culex species concentration of dominance, the highest value was recorded in the rural breeding site in Beheira governorate (CD: 0.95) followed by the rural breeding site in Giza governorate (CD: 0.9) and the urban breeding site in Gharbia governorate (CD: 0.74). VI. 3. Physicochemical analysis of the water samples VI. 3. 1. Water temperature The highest water temperature was recorded in the rural Giza Governorate breeding site (32 ± 7.42 ˚C) followed by the rural Beheira Governorate breeding site (28.63 ± 5.83 ˚C). Meanwhile, the lowest temperature was recorded in the urban Gharbia Governorate breeding site (20.75 ± 4.24 ˚C). VI. 3. 2. Water pH Water pH in both rural and urban mosquito breeding sites tested was slightly alkaline. The rural Giza Governorate breeding site had the highest pH value (8.11 ± 0.42) followed by the rural breeding site in Beheira Governorate (7.85 ± 0.71). On the other hand, the urban Gharbia Governorate breeding site recorded the lowest pH value (7.79 ± 0.13). ANOVA analysis did not reveal any significant differences in pH values between rural and urban breeding sites tested. VI. 3. 3. Water salinity The highest water salinity percentage was measured in the urban breeding site in Gharbia Governorate (5.1 ± 0.61 %) followed by the rural breeding site in Giza Governorate (3.37 ± 0.45 %). The lowest salinity percentage was measured in the rural breeding site in Beheira Governorate (1.33 ± 0.25 %). ANOVA analysis showed significant differences in salinity percentage between rural and Chapter urban mosquito breeding sites. Bonferroni’s test showed that salinity percentage in rural and urban breeding sites exhibited significant differences in pair-wise comparison. VI. 3. 4. Water turbidity The mean values of water turbidity in Giza, Beheira and Gharbia breeding sites were 11.1 ± 11.5, 5.69 ± 3.84 and 3.86 ± 4.39 NTU, respectively. ANOVA analysis indicates no evidence for a statistically significant relation between water turbidity and urban and rural breeding sites tested. VI. 3. 5. Water dissolved oxygen (DO) The highest dissolved oxygen content (0.73 ± 0.12 mg/l) was recorded in the urban Gharbia Governorate breeding site. Meanwhile, similar relative contents of dissolved oxygen (0.33 ± 0.58 mg/l) and (0.3 ± 0.44 mg/l) were recorded in Giza and Beheira rural breeding sites, respectively. ANOVA results did not show any significant differences in dissolved oxygen content among urban and rural breeding sites tested. VI. 3. 6. Water dissolved nitrite The rural Beheira Governorate breeding site had the highest nitrite concentration (0.98 ± 0.40 mg/l) followed by the urban Gharbia Governorate breeding site (0.40 ± 0.35 mg/l) and the rural Giza Governorate breeding site (0.19 ± 0.09 mg/l). ANOVA results showed significant differences among the rural and urban breeding sites in dissolved nitrite content. Bonferroni’s test showed that dissolved nitrite contents in the breeding sites exhibited significant differences in pair-wise comparison only between the rural breeding sites in Giza and Beheira Governorates. VI. 4. Association between the relative abundance of Culex species and physicochemical characteristics A stepwise multiple regression analysis indicated that salinity was the most important predictor for Cx. Pipiens, Cx. univittatus, Cx. antennatus and Cx. quinqueifasciatus abundance. Cx. pipiens larvae abundance appeared negatively associated with water salinity. However, both Cx. univittatus and Cx. antennatus abundances were positively associated with water salinity Concerning Cx. quinqueifasciatus, it was inversely related to water nitrite concentration. Cx. perexiguus was positively associated with both turbidity and pH values. Regarding Cs. longiareolata, it had a positive association with both percentage salinity and nitrite concentration. VI. 5. Presence of insecticide residues in mosquito breeding sites Insecticide residue screening in the rural Giza Governorate mosquito breeding site revealed five insecticide residues namely the organic compound ortho-phenyl phenol, the organophosphorus insecticides Malathion, Dimethoate and Diazinon and the synergist piperonyl butoxide. In this breeding site, piperonyl butoxide was detected at the highest concentration (0.455 ± 0.007 mg /L) while Dimethoate was detected at the lowest concentration (0.005 ± 0.007 mg /L). However, in the other rural breeding site in Beheira Governorate, only piperonyl butoxide was detected. In the urban Gharbia Governorate breeding site, only ortho-phenyl phenol was detected. ANOVA results showed significant differences among the three breeding sites only in Malathion and piperonyl butoxide, respectively. Bonferroni test indicated that the level of Malathion residues in the rural Giza Governorate breeding site was significantly higher than those in the urban Gharbia Governorate or the rural Beheira Governorate breeding sites, while it showed a significant difference in piperonyl butoxide residues between between the urban breeding site in Gharbia Governorate and both rural breeding sites in Giza and Beheira Governorates. VI. 6. Susceptibility of field-collected Culex pipiens larvae to insecticides VI. 6. 1. Susceptibility of Culex pipiens larvae to Deltamethrin Third instar larvae of Gharbia population was more susceptible to Deltamethrin than both Beheia and Giza populations. Statistical analysis showed that the percentages of mortality of Cx. pipiens larvae were concentration-dependent for Deltamethrin. On the other hand, there was no significant difference in percentages of mortality among populations. Concerning the resistance ratios obtained at LC50, all Cx. pipiens larval populations at all sites exhibited low resistance ratios. VI. 6. 2. Susceptibility of Culex pipiens larvae to Chlorpyrifos Third instar larvae of the Gharbia population was less susceptible to chlorpyrifos than both Beheira and Giza populations. Statistical analysis revealed that the percentages of mortality of Cx. pipiens larvae were also concentration-dependent for Chlorpyrifos. However, the analysis did not show any significant differences in percentages of mortality among populations. All Cx. pipiens larvae of Gharbia, Beheira and Giza populations exhibited low resistance ratios. VI. 6. 3. Susceptibility of Culex pipiens larvae to Malathion The reference population was more susceptible to Malathion than the three field populations. Statistical analysis exhibited that the percentages of mortality of Cx. pipiens larvae were concentration-dependent for Malathion. Moreover, the analysis showed a significant difference in the percentage of mortality among populations. This significant difference could be attributed to the differences between the reference population and all the tested populations. Otherwise, intraspecific variation in the percentage of mortality was observed between observed between Gharbia population and with either Giza or Beheira populations. With respect to the resistance ratios obtained at LC50, Beheira and Giza Cx. pipiens populations showed medium resistance ratios. While Gharbia population showed low resistance. VI. 6. 4. Susceptibility of Culex pipiens larvae to Fipronil The results revealed that Cx. pipiens 3rd instar larvae of all tested populations were more susceptible to Fipronil than the other previously tested insecticides. Statistical analysis showed that the percentages of mortality of Cx. pipiens larvae were concentration-dependent for Fipronil. Furthermore, the analysis showed a significant difference in the percentage of mortality among populations. This significant difference could be attributed to the observed differences between the reference population and all the tested populations. The resistance ratios obtained at LC50 revealed that Cx. pipiens larval populations at all sites exhibited low resistance ratios. VI. 7. Evaluation of fitness cost of Malathion resistance in Culex pipiens field populations VI. 7. 1. Adult survival In males, Log-Rank test analysis showed significant differences in the survival curves among the field-collected Cx. pipiens F1 generation and the reference population. Further analysis showed that males of the F1 generation of the Gharbia Malathion-susceptible population had longer median survival (30 days) than the males of the reference population (19 days). Concerning females, there was no significant difference among the survival curves of the Gharbia Malathion-susceptible population and the Beheira Malathion-resistant F1 generations tested. VI. 7. 2. Adult fecundity and egg-hatchability The estimation of fecundity was based on the number of eggs/female/day. Statistical analysis indicated that there was no significant difference in fecundity or hatchability among the Gharbia Malathion-susceptible and Beheira Malathion-resistant field-collected and the reference Cx. pipiens populations. VI. 8. Biochemical composition of Culex pipiens larvae VI. 8. 1. Total larval proteins The ANOVA of the data showed a significant difference in the amount of total larval proteins among the Beheira Malathion-resistant and Gharbia Malathion-susceptible field-collected and the reference populations. Bonferroni’s post-hoc test indicated that the total larval proteins of the Beheira Malathion-resistant population were significantly lower than those of the Gharbia Malathion-susceptible and Reference populations. VI. 8. 2. Total larval lipids ANOVA results did not show any significant difference among the Beheira Malathion-resistant and Gharbia Malathion-susceptible field-collected and the reference populations. VI. 8. 3. Total larval carbohydrates ANOVA analysis indicated significant differences among the Beheira Malathion-resistant and Gharbia Malathion-susceptible field-collected Cx. pipines larval populations and the reference population. Bonferroni’s test showed that total larval carbohydrates of the Beheira Malathion-resistant, Gharbia Malathion-susceptible and the reference populations exhibited significant differences in pair-wise comparison. VI. 8. 4. phenol-oxidase (PO) activity Kruskal-Wallis results did not show any significant differences in the phenol-oxidase activity and specific activity between the three populations. VI. 9. Resistance gene expression in Culex pipiens field populations Analysis of variance (ANOVA) revealed significant differences in CYP6F1 and GST gene expressions among the Gharbia Malathion-susceptible, Beheira Malathion-resistant and the reference Cx. pipiens populations. Otherwise, Estα and Estβ gene expressions showed no significant differences among the Beheira Malathion-resistant and Gharbia Malathion-susceptible fieldcollected and the reference populations. Bonferroni multiple-comparison indicated that the Gharbia Malathion-susceptible population had significantly higher CYP6F1 and GST gene expression levels than both the Beheira Malathion-resistant and reference populations.