Mosquitoes

This study was carried out from November 2018 to September 2019. Aedes aegypti larvae identified according to Harbach (1985) were collected from various water storage containers in the Jazan region of KSA, which is located at 16°54′34.8588″N, 42°34′4.4472″E along the southern Red Sea coast, just north of Yemen. Then, the larvae were reared for five generations in the Biology Department, Jazan University, according to the methods described by El-Sheikh et al. (2016). Briefly, 150 larvae were reared in 25 × 42 × 5-cm pans containing 1 liter of water at temperatures ranging from 23 to 27°C (mean 25°C) and 80 ± 6% RH.

Plant collection and preparation of crude extracts

Pulicaria jaubertii was collected from Wadi Sabia during September 2018. The plant was identified according to the descriptions in the Flora of Saudi Arabia by Migahid (1987). The leaves were washed, dried under shade, and mechanically ground, using a commercial stainless-steel blender (Philips, Model Number: HR2058). Extractions were carried out using ethanol, petroleum ether, and chloroform, according to El-Sheikh et al. (2016).

Ovicidal activity

The ovicidal activity was determined using the methods described by Subashini et al. (2017) that evaluated the ethanol, petroleum ether, and chloroform leaf extracts at concentrations ranging from 25 to 150 ppm. Thirty Ae. aegypti eggs were exposed to each extract concentration, in 300 ml of dechlorinated tap water. Hatching assessment was performed under an electrical dissecting stereomicroscope (Labo America Inc., Model: CZM6). Each test was conducted three times, along with a control. Percentages of egg hatch were calculated using the following formula:

Nonhatched eggs were removed, and larvae were observed daily until adult emergence. The parameters measured in this study included: egg hatchability, developmental period, and mortality rate of larvae, pupae, and adults.

Reproductive potential

Females that emerged from treated eggs were allowed to take a blood meal from a pigeon, and numbers of eggs produced were counted using a binocular stereomicroscope. The preoviposition period (gonotrophic cycle) was calculated as the interval between blood feeding and commencement of oviposition. Eggs were sorted into two categories, hatched and nonhatched. Nonhatched eggs were further classified into embryonated and nonembryonated. Hatched eggs and nonhatched embryonated eggs were considered as fertilized, while nonhatched, nonembryonated eggs were regarded as unfertilized (Rak and Ishii 1989).

Statistical analysis

Analysis of variance (ANOVA) was carried out to determine the differences in activity among the P. jaubertii test extracts, using Tukey's HSD (honestly significant difference) test at a 5% probability level. The statistical analyses were carried out using Statistical Package Social Science (SPSS) software (Version 11.5, SPSS 2007, Chicago, IL).

A control group of each Ae. aegypti eggs, larvae pupa, and adults were kept without treatment with the plant extract.

Ovicidal activity

The ethanolic leaf extract resulted in the highest level of Ae. aegypti ovicidal activity at all concentrations compared with other two extracts. At the highest concentration (150 ppm), complete ovicidal activity occurred (Table 1). The ethanolic and petroleum ether extracts at 100−150 ppm delayed egg duration by about 1−1.5 days compared with the chloroform extract or control.

Table 1. Mean¹ ovicidal activity of Pulicaria jaubertii extracts against Aedes aegypti in the laboratory.

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Larvicidal, pupicidal, and adulticidal activity

The cumulative mortality rates of larvae, pupae, and adults were increased by the latent toxicity of the ethanolic extract compared with the petroleum ether and chloroform extracts, where complete larval and pupal mortality was produced at 75 and 50 ppm, respectively, with no adult mortality (Table 2). The percentage of adult emergence was impacted to the greatest degree at the highest concentrations of each of the three extracts.

Table 2. Late toxic (cumulative) effect of Pulicaria jaubertii test extracts on different stages of Aedes aegypti resulting from treated eggs in the laboratory.

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Effect on developmental periods and growth index

The ethanolic extract of P. jaubertii significantly increased (P < 0.05) the larval developmental time to 15.3 and 15.6 days at 25 and 50 ppm, respectively, compared with 12.3 days for the control group (Table 3). Pupal duration was extended to 2.67 days at 25 ppm, compared with 1.7 days for the control group. The petroleum ether extract did not affect larval duration except at the highest two concentrations (75 and 100 ppm), where it was significantly prolonged (P > 0.01) to 13.3 and 14.6 days, respectively, compared with 12.3 days for the control group. In addition, the pupal duration was significantly delayed at the highest concentration (100 ppm), which was prolonged to 2.5 days, compared with 1.7 days for the control group. Larval and pupal developmental periods, however, were not adversely affected by the chloroform extract (Table 3). The ethanolic extract significantly prolonged developmental time of Ae. aegypti larvae and pupae when reared from eggs previously treated with this extract. Generally, as concentration increased for each extract, the growth index decreased (Table 3).

Table 3. Effect of Pulicaria jaubertii test extracts on developmental periods (days) of immature stages of Aedes aegypti in the laboratory.

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Preoviposition period, fecundity, fertility, and sterility index

The mean preoviposition period of females Ae. aegypti that developed from eggs previously treated with the petroleum ether extract significantly increased (P < 0.001) at concentrations of 25 and 50 ppm, respectively, compared with controls. Moreover, the preoviposition period of females from eggs treated with 50–150 ppm chloroform extract was also delayed when compared with controls. The egg abundance significantly declined (P < 0.05) from 107.8 recorded for the control group to 65.1 and 43.4% as a result of exposure to 25 and 50 ppm petroleum ether extract from females previously exposed as eggs. Furthermore, the hatchability of those eggs was subsequently reduced from 94.9% in the control to 59.4% and 34.8% at 25 and 50 ppm concentrations, respectively (Table 4). The percentage of nonhatched embryonated eggs was 15.9% in the control group, was increased to 69.3% and 78.8% at 25 and 50 ppm, respectively. A marked increase in the percentage of females that did not produce eggs (i.e., were sterile) was recorded at 25 ppm (62.2%) and at 50 ppm (85.2%) for the petroleum ether extracts, compared with the untreated females (Table 4). The preoviposition period of females produced from eggs previously exposed to chloroform extracts significantly increased at concentrations ≥75 ppm as compared with controls (Table 4). Moreover, a significant reduction in the number of eggs laid by those females occurred at the two highest concentrations. Egg hatchability was reduced from 94.9% observed in the control group to 75.2% and 62.1% at 125 and 150 ppm, respectively. The percentage of nonhatched embryonated eggs increased as extract concentration increased, with the greatest reduction occurring at 150 ppm. A marked increase in the percentage of sterile females was also observed, with the highest occurring at 150 ppm (Table 4).

Table 4. Effect of Pulicaria jaubertii petroleum ether and chloroform extracts on preoviposition period, fecundity, fertility, and sterility index of Aedes aegypti females in the laboratory.

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