Adulticidal, repellent, and ovicidal properties of indigenous plant extracts against the malarial vector, Anopheles stephensi (Diptera: Culicidae)

Mosquito-borne diseases with an economic impact create loss in commercial and labor outputs, particularly in countries with tropical and subtropical climates. Mosquito control is facing a threat because of the emergence of resistance to synthetic insecticides. Extracts from plants may be alternative sources of mosquito control agents because they constitute a rich source of bioactive compounds that are biodegradable into nontoxic products and potentially suitable for use to control mosquitoes. Insecticides of botanical origin may serve as suitable alternative biocontrol techniques in the future. In view of the recently increased interest in developing plant origin insecticides as an alternative to chemical insecticide, this study was undertaken to assess the adulticidal, repellent, and ovicidal potential of the crude hexane, ethyl acetate, benzene, aqueous, and methanol solvent extracts from the medicinal plants Andrographis paniculata, Cassia occidentalis, and Euphorbia hirta against the medically important mosquito vector, Anopheles stephensi (Diptera: Culicidae).The adult mortality was observed after 24 h of exposure. All extracts showed moderate adulticide effects; however, the highest adult mortality was found in methanol extract of A. paniculata followed by C. occidentalis and E. hirta against the adults of A. stephensi with LC₅₀ and LC₉₀ values of 210.30, 225.91, and 263.91 ppm and 527.31, 586.36, and 621.91 ppm, respectively. The results of the repellent activity of hexane, ethyl acetate, benzene, aqueous, and methanol extract of A. paniculata, C. occidentalis, and E. hirta plants at three different concentrations of 1.0, 3.0, and 6.0 mg/cm² were applied on skin of forearm in man and exposed against adult female mosquitoes. In this observation, these three plant crude extracts gave protection against mosquito bites without any allergic reaction to the test person, and also, the repellent activity is dependent on the strength of the plant extracts. Mean percent hatchability of the ovicidal activity was observed 48 h post-treatment. The percent hatchability was inversely proportional to the concentration of extract and directly proportional to the eggs. Mortality of 100 % with methanol extract of A. paniculata exerted at 150 ppm and aqueous, methanol extract of C. occidentalis and E. hirta were exerted at 300 ppm. These results suggest that the leaf extracts of A. paniculata, C. occidentalis, and E. hirta have the potential to be used as an ideal eco-friendly approach for the control of the A. stephensi. Further detailed research is needed to identify the active ingredient in the extracts and implement the effective mosquito management program.     

Evaluation of indigenous plant extracts against the malarial vector, Anopheles stephensi (Liston) (Diptera: Culicidae)

Since ancient times, plant and microbial products were used in various aspects. However, their use against insects decreased when chemical products became developed. Recently, concerns increased with respect to public health and environmental security requiring detection of natural products that may be used against insects. In this study, mosquito Larvicidal and ovicidal activity of crude hexane, ethyl acetate, benzene, chloroform, and methanol extracts of the leaf of three plants, Eclipta alba, Cardiospermum halicacabum, and Andrographis paniculata, were tested against the early third-instar larvae of Anopheles stephensi (Liston) (Diptera: Culicidae). The larval mortality was observed after 24 h of exposure. All extracts showed moderate larvicidal effects; however, the highest larval mortality was found in methanol extract of A. paniculata, E. alba, and C. halicacabum against the larvae of A. stephensi (LC₅₀ = 79.68, 112.56, and 133.01 ppm; LC₉₀ = 154.66, 220.68, and 270.72 ppm), respectively. Mean percent hatchability of the ovicidal activity was observed 48 h post-treatment. The percent hatchability was inversely proportional to the concentration of extract and directly proportional to the eggs. Mortality of 100% with methanol and ethyl acetate extract of A. paniculata and methanol extract of E. alba were exerted at 200 ppm and methanol and benzene extract of C. halicacabum exerted at 150 ppm. This is an ideal eco-friendly approach for the control of the malaria vector, A. stephensi. Therefore, this study provides first report on the larvicidal and ovicidal activities against malaria vector, A. stephensi of E. alba plant extracts.     

Studies on effect of Acalypha indica L. (Euphorbiaceae) leaf extracts on the malarial vector, Anopheles stephensi Liston (Diptera:Culicidae)

The leaf extract of Acalypha indica with different solvents viz, benzene, chloroform, ethyl acetate and methanol were tested for larvicidal, ovicidal activity and oviposition attractancy against Anopheles stephensi. The larval mortality was observed after 24 h exposure. The LC(50) values are 19.25, 27.76, 23.26 and 15.03 ppm, respectively. Mean percent hatchability of the ovicidal activity was observed 120 h after treatment. The percent hatchability was inversely proportional to the concentration of extract and directly proportional to the eggs. The highest effective attractancy of 90.09%, 94.20%, 85.43% and 95.75% were observed at 100 ppm concentration viz, benzene, chloroform, ethyl acetate and methanol, respectively. The lowest effective attractancy of 47.17%, 61.94%, 49.28% and 68.12% were observed at 25 ppm concentration viz, benzene, chloroform, ethyl acetate and methanol, respectively. The results that the leaf extract of A. indica is promising as larvicidal and ovicidal activity and oviposition attractancy against malaria vector A. stephensi.     

Rubidium marking of Anopheles stephensi (Diptera: Culicidae)

Immature Anopheles stephensi Liston were reared in untreated water and water containing eight 2-fold dilutions of rubidium (Rb) from 1,000 to 7.8 ppm to determine the concentration that allowed reliable detection and produced the least toxic effects as measured by adult emergence, weight, and survival. The amount of Rb detected in mosquitoes increased positively with increasing concentrations in the rearing water. Concentrations > or = 31.2 ppm Rb in the rearing water provided high and consistent detection levels of > or = 3,500 ppm Rb/mg of adult mosquito. There were no adverse effects of Rb on the weights of mosquitoes. However, increased Rb concentrations in the rearing water were associated with decreased emergence and survival. After 8 d, percentage emergence from Rb concentrations of 0-31.2 ppm was > or = 50%. At day 21, Rb concentrations of 0-31.2 ppm showed < or = 29% reduction of female survival compared with controls. The recommended concentration for reliable Rb detection with minimal toxic effects in An. stephensi was 32 ppm.     

Antimalarial efficacy of dynamic compound of plumbagin chemical constituent from Plumbago zeylanica Linn (Plumbaginaceae) against the malarial vector Anopheles stephensi Liston (Diptera: Culicidae)

In the present investigation, the effective root compound of plumbagin of Plumbago zeylanica (Plumbaginaceae) was evaluated for chemical constituent and antimalarial effect against the fourth instar larvae of Anopheles stephensi Liston (Diptera). In the chromatographic analyses of root compound with Rf value of 0.788 and NMR analyses also revealed that the effective compound contain naphthoquinone plumbagin were identified as the major chemical constituent. Larval mortality was observed after 3 h of exposure period. The plumbagin compound showed remarkable larvicidal activity against A. stephensi (LC₅₀ 32.65 and LC₉₀72.27 ppm). Histopathological effects of compound was observed in the treated larvae. Based on the results, the plumbagin compound of P. zeylanica can be considered as a new source of natural larvicide for the control of malarial vector.     

Evaluation of larvicidal activity of Acalypha alnifolia Klein ex Willd. (Euphorbiaceae) leaf extract against the malarial vector, Anopheles stephensi, dengue vector, Aedes aegypti and Bancroftian filariasis vector, Culex quinquefasciatus (Diptera: Culicidae)

The leaf extract of Acalypha alnifolia with different solvents — hexane, chloroform, ethyl acetate, acetone and methanol — were tested for larvicidal activity against three important mosquitoes such as malarial vector, Anopheles stephensi, dengue vector, Aedes aegypti and Bancroftian filariasis vector, Culex quinquefasciatus. The medicinal plants were collected from the area around Kallar Hills near the Western Ghats, Coimbatore, India. A. alnifolia plant was washed with tap water and shade dried at room temperature. The dried leaves were powdered mechanically using commercial electrical stainless steel blender. The powder 800 g of the leaf material was extract with 2.5 litre of various each organic solvents such as hexane, chloroform, ethyl acetate, acetone, methanol for 8 h using Soxhlet apparatus, and filtered. The crude plant extracts were evaporated to dryness in a rotary vacuum evaporator. The yield of extracts was hexane (8.64 g), chloroform (10.74 g), ethyl acetate (9.14 g), acetone (10.02 g), and methanol (11.43 g). One gram of the each plant residue was dissolved separately in 100 ml of acetone (stock solution) from which different concentrations, i.e., 50, 150, 250, 350 and 450 ppm, was prepared. The hexane, chloroform, ethyl acetate, acetone was moderate considerable mortality; however, the highest larval mortality was methanolic extract observed in three mosquito vectors. The larval mortality was observed after 24 h exposure. No mortality was observed in the control. The early fourth-instar larvae of A. stephensi had values of LC₅₀ = 197.37, 178.75, 164.34, 149.90 and 125.73 ppm and LC₉₀ = 477.60, 459.21, 435.07, 416.20 and 395.50 ppm, respectively. The A. aegypti had values of LC₅₀ = 202.15, 182.58, 160.35, 146.07 and 128.55 ppm and LC₉₀ = 476.57, 460.83, 440.78, 415.38 and 381.67 ppm, respectively. The C. quinquefasciatus had values of LC₅₀ = 198.79, 172.48, 151.06, 140.69 and 127.98 ppm and LC₉₀ = 458.73, 430.66, 418.78, 408.83 and 386.26 ppm, respectively. The results of the leaf extract of A. alnifloia are promising as good larvicidal activity against the mosquito vector, A. stephensi, A. aegypti, C. quinquefasciatus. Therefore, this study provides first report on the larvicidal activities against three species of mosquito vectors of this plant extracts from Southern India.     

Mosquito larvicidal, ovicidal, and repellent properties of botanical extracts against Anopheles stephensi, Aedes aegypti, and Culex quinquefasciatus (Diptera: Culicidae)

Mosquito-borne diseases have an economic impact, including loss in commercial and labor outputs, particularly in countries with tropical and subtropical climates; however, no part of the world is free from vector-borne diseases. In mosquito control programs, botanical origin may have the potential to be used successfully as eggs, larvae, and adult. The larvicidal, ovicidal, and repellent activities of crude benzene and ethyl acetate extracts of leaf of Ervatamia coronaria and Caesalpinia pulcherrima were assayed for their toxicity against three important vector mosquitoes, viz., Anopheles stephensi, Aedes aegypti, and Culex quinquefasciatus (Diptera: Culicidae). The larval mortality was observed after 24 h of exposure. All extracts showed moderate larvicidal effects; however, the highest larval mortality was found in benzene extract of E. coronaria against the larvae of Anopheles Stephensi, Aedes aegypti, and Culex quinquefasciatus with the LC₅₀ and LC₉₀ values were 79.08, 89.59, and 96.15 ppm and 150.47, 166.04, and 174.10 ppm, respectively. Mean percent hatchability of the ovicidal activity was observed 48 h posttreatment. The percent hatchability was inversely proportional to the concentration of extract and directly proportional to the eggs. The leaf extract of E. coronaria was found to be most effective than Caesalpinia pulcherrima against eggs/egg rafts of three vector mosquitoes. For E. coronaria, the benzene extract exerted 300, 250, and 200 ppm against Anopheles stephensi, Aedes aegypti, and Culex quinquefasciatus, respectively. The results of the repellent activity of benzene and ethyl acetate extract of E. coronaria and Caesalpinia pulcherrima plants at three different concentrations of 1.0, 2.5, and 5.0 mg/cm² were applied on skin of fore arm in man and exposed against adult female mosquitoes. In this observation, these two plant crude extracts gave protection against mosquito bites without any allergic reaction to the test person, and also, the repellent activity is dependent on the strength of the plant extracts. These results suggest that the leaf solvent plant extracts have the potential to be used as an ideal ecofriendly approach for the control of mosquitoes. This is the first report on the mosquito larvicidal, ovicidal, and repellent activities of the reported E. coronaria and Caesalpinia pulcherrima plants.     

Larvicidal, pupicidal, repellent and adulticidal activity of Citrus sinensis orange peel extract against Anopheles stephensi, Aedes aegypti and Culex quinquefasciatus (Diptera: Culicidae)

Mosquitoes are the carriers of severe and well-known illnesses such as malaria, arboviral encephalitis, dengue fever, chikunguniya fever, West Nile virus and yellow fever. These diseases produce significant morbidity and mortality in humans and livestock around the world. The present study explored the effects of orange peel ethanol extract of Citrus sinensis on larvicidal, pupicidal, repellent and adulticidal activity against Anopheles stephensi, Aedes aegypti and Culex quinquefasciatus. The orange peel material was shade dried at room temperature and powdered coarsely. From orange peel, 300?g powdered was macerated with 1?L of ethanol sequentially for a period of 72?h each and filtered. The yields of the orange peel ethanol crude extract of C. sinensis 13.86?g, respectively. The extracts were concentrated at reduced temperature on a rotary vacuum evaporator and stored at a temperature of 4?°C. The larvicidal, pupicidal and adult mortality was observed after 24?h of exposure; no mortality was observed in the control group. For C. sinensis, the median lethal concentration values (LC(50)) observed for the larvicidal and pupicidal activities against mosquito vector species A. stephensi first to fourth larval instars and pupae were 182.24, 227.93, 291.69, 398.00 and 490.84?ppm; A. aegypti values were 92.27, 106.60, 204.87, 264.26, 342.45, 436.93 and 497.41?ppm; and C. quinquefasciatus values were 244.70, 324.04, 385.32, 452.78 and 530.97?ppm, respectively. The results of maximum repellent activity were observed at 450?ppm in ethanol extracts of C. sinensis and the mean complete protection time ranged from 150 to 180?min was tested. The ethanol extract of C. sinensis showed 100?% repellency in 150?min and showed complete protection in 90?min at 350?ppm against A. stephensi, A. aegypti and C. quinquefasciatus, respectively. The adult mortality was found in ethanol extract of C. sinensis with the LC(50) and LC(90) values of 272.19 and 457.14?ppm, A. stephensi; 289.62 and 494.88?ppm, A. aegypti; and 320.38 and 524.57?ppm, respectively. These results suggest that the orange peel extracts of C. sinensis have the potential to be used as an ideal eco-friendly approach for the control of the vector control programmes.     

Intraspecific variation in the reproductive capacity of Anopheles stephensi (Diptera: Culicidae)

Intraspecific variation in the reproductive capacity of Anopheles stephensi Liston females was studied under constant laboratory conditions. Of 421 engorged females examined individually throughout their lifetimes, 260 laid a total of 479 egg batches with a maximum of nine ovipositions per female. The number of eggs per oviposition varied from 5 to 247 (average 96.8). The number of ovipositions per female were correlated positively with the average number of eggs per batch and exhibited a negative binomial distribution among females, indicating that a small portion of the population exhibited a particularly high fecundity. Among the ovipositing females, the total fecundity and fertility ranged, respectively, from 5 to 1,084 eggs and from 0 to 1,036 larvae per female. The estimated maximal number of female progeny produced by a female in her lifetime was 317, and the estimates of net reproductive rate (Ro) and capacity for increase (rc) were 25.7 and 0.17, respectively. The number of eggs in the first oviposition was predictive of the total fecundity of a female. The wide range of individual variation in the reproductive capacity of An. stephensi emphasized the requirement for a large sample size for reliable estimation.     

Laboratory evaluation of traditionally used plant-based insect repellent against the malaria vector Anopheles arabiensis Patton (Diptera: Culicidae)

A laboratory study was carried out to evaluate the repellent efficacy of a methanol-leaf extract of Ethiopian traditionally used insect repellent plant viz., Lomi sar [vernacular name (local native language, Amharic); Cymbopogon citratus (DC) Stapf. (Poaceae)] against Anopheles arabiensis at four different concentrations viz., 1.0, 1.5, 2.0, and 2.5 mg/cm². The percentage protection in relation to the dose method was performed. C. citratus extract has shown various degrees of repellency impact against A. arabiensis. It provided the maximum total percentage protection of 78.83% at 2.5 mg/cm² and followed 68.06% at 2.0 mg/cm² for 12 h. All four tested concentrations of C. citratus extract offered significant protection and Student's t test results shows statistically significant (p value = 0.001) [1.0 mg/cm² (t = 22.89; df = 4); 1.5 mg/cm² (t = 24.03; df = 4); 2.0 mg/cm² (t = 36.92; df = 4); 2.5 mg/cm² (t = 22.31; df = 4)] difference between treated and control groups. The result suggests that it could serve as a potent insect repellent against vectors of disease. Globally, C. citratus is renowned for its therapeutic values. Above and beyond, due to its user- as well as environmental-friendly nature, it should be promoted among the marginalized populations in order to reduce man-vector contact. In addition, this appropriate strategy affords the opportunity to minimize chemical repellent usage and the risks associated with adverse side effects. At the end of the day, traditionally used plant-based insect repellents could be viable safer alternative sources for chemical insect repellents.     

Mosquitocidal properties of nereistoxin against Anopheles stephensi, Aedes aegypti and Culex quinquefasciatus (Diptera: Culicidae)

Emergence of resistance among mosquitoes is a recent problem. Safe and eco-friendly agents from biological origins are the need of the hour. Nereistoxin, a naturally occurring substance, was first isolated from the marine annelid Lumbriconereis heteropoda and stored in the freezer. In the present study, the toxicity of nereistoxin was evaluated against vector mosquitoes. The larvicidal, ovicidal and adulticidal activities of nereistoxin were assayed for their toxicity against three important vector mosquitoes, viz., Anopheles stephensi, Aedes aegypti and Culex quinquefasciatus (Diptera: Culicidae). The nereistoxin was inversely proportional to the concentration and directly proportional to the mosquitoes. The larvicidal activity after 24 h LC₅₀ value was observed at 0.467, 0.535 and 0.601 ppm for A. stephensi, A. aegypti and C. quinquefasciatus, respectively. The ovicidal activity after 120 h zero percentage of egg hatchability was observed at a concentration of 0.8 ppm for A. stephensi and 1.0 ppm for A. aegypti and C. quinquefasciatus. The results of the adulticidal activity after 24 h LD₅₀ value were observed at 0.022, 0.028 and 0.034 ppm for A. stephensi, A. aegypti and C. quinquefasciatus, respectively. The extracted nereistoxin was characterized and identified by ultraviolet, infrared, nuclear magnetic resonance, mass spectroscopic methods and high pressure liquid chromatography. These results clearly reveal that the nereistoxin served as a potential larvicidal, ovicidal and adulticidal activity against vector mosquitoes.     

Blood vessel location time by Anopheles stephensi (Diptera: Culicidae).

Probing behavior of Anopheles stephensi Liston is characterized by the following: the first probe is longer than subsequent ones, the probability of blood location rises initially and then falls, and blood vessel location is deterministic. The overall probing behavior of An. stephensi, therefore, is similar to that of Aedes aegypti (L.); i.e., differences between them were quantitative and may be accounted for by different levels of salivary apyrase and different experimental vertebrate hosts.     

Mosquito larvicidal, pupicidal, adulticidal, and repellent activity of Artemisia nilagirica (Family: Compositae) against Anopheles stephensi and Aedes aegypti

Mosquito-borne diseases have an economic impact, including loss in commercial and labor outputs, particularly in countries with tropical and subtropical climates; however, no part of the world is free from vector-borne diseases. The aim of the present study, to evaluate the larvicidal, pupicidal, repellent, and adulticidal activities of methanol crude extract of Artemisia nilagirica were assayed for their toxicity against two important vector mosquitoes, viz., Anopheles stephensi and Aedes aegypti (Diptera: Culicidae). The fresh leaves of A. nilagirica were washed thoroughly in tap water and shade dried at room temperature (28?±?2?°C) for 5?C8?days. The air-dried materials were powdered separately using commercial electrical blender. From the plants, 500?g powdered was macerated with 1.5?L organic solvents of methanol sequentially for a period of 72?h each and filtered. The larval and pupal mortality was observed after 24?h of exposure; no mortality was observed in the control group. The first- to fourth-instar larvae and pupae of A. stephensi had values of LC₅₀?=?272.50, 311.40, 361.51, 442.51, and 477.23?ppm, and the LC₉₀?=?590.07, 688.81, 789.34, 901.59, and 959.30?ppm; the A. aegypti had values of LC₅₀?=?300.84, 338.79, 394.69, 470.74, and 542.11?ppm, and the LC₉₀?=?646.67, 726.07, 805.49, 892.01, and 991.29?ppm, respectively. The results of the repellent activity of plant extract of A. nilagirica plants at five different concentrations of 50, 150, 250, 350, and 450?ppm were applied on skin of fore arm in man and exposed against adult female mosquitoes. In this observation, the plant crude extract gave protection against mosquito bites without any allergic reaction to the test person, and also, the repellent activity is dependent on the strength of the plant extracts. The adult mortality was found in methanol extract of A. nilagirica, with the LC₅₀ and LC₉₀ values of 205.78 and 459.51?ppm for A. stephensi, and 242.52 and 523.73?ppm for A. aegypti, respectively. This result suggests that the leaf extract have the potential to be used as an ideal eco-friendly approach for the control of vector mosquito as target species.     

Adulticidal and repellent properties of Cassia tora Linn. (Family: Caesalpinaceae) against Culex quinquefasciatus, Aedes aegypti, and Anopheles stephensi

Mosquitoes have developed resistance to various synthetic insecticides, making its control increasingly difficult. Insecticides of botanical origin may serve as suitable alternative biocontrol techniques in the future. The adulticidal and repellent activities of crude hexane, chloroform, benzene, acetone, and methanol extracts of the leaf of Cassia tora were assayed for their toxicity against three important vector mosquitoes, viz., Culex quinquefasciatus, Aedes aegypti, and Anopheles stephensi. The adult mortality was observed after 24?h of exposure. All extracts showed moderate adulticidal effects; however, the highest adult mortality observed was found in methanol extract. The LC₅₀ and LC₉₀ values of C. tora leaf extracts against adulticidal activity of (hexane, chloroform benzene, acetone, and methanol) C. quinquefasciatus, A. aegypti, and A. stephensi were the following: C. quinquefasciatus LC₅₀ values were 338.81, 315.73, 296.13, 279.23, and 261.03?ppm and LC₉₀ values were 575.77, 539.31, 513.99, 497.06, and 476.03?ppm; A. aegypti LC₅₀ values were 329.82, 307.31, 287.15, 269.57, and 252.03?ppm and LC₉₀ values were 563.24, 528.33, 496.92, 477.61, and 448.05?ppm; and A. stephensi LC₅₀ values were 317.28, 300.30, 277.51, 263.35, and 251.43?ppm and LC₉₀ values were 538.22, 512.90, 483.78, 461.08, and 430.70?ppm, respectively. The results of the repellent activity of hexane, chloroform, benzene, acetone, and methanol extracts of C. tora plant at three different concentrations of 1.0, 2.5, and 5.0?mg/cm² were applied on skin of forearm in man and exposed against adult female mosquitoes. In this observation, this plant crude extracts gave protection against mosquito bites without any allergic reaction to the test person, and also, the repellent activity is dependent on the strength of the plant extracts. These results suggest that the leaf solvent plant extracts have the potential to be used as an ideal eco-friendly approach for the control of mosquitoes. This is the first report on mosquito adulticidal and repellent activities of the reported C. tora against mosquito vectors from Southern India.     

Chemical composition and larvicidal activity of the essential oil of Plectranthus amboinicus (Lour.) Spreng against Anopheles stephensi: a malarial vector mosquito

Essential oil of Plectranthus amboinicus was studied for its chemical composition and larvicidal potential against the malarial vector mosquito Anopheles stephensi. Totally 26 compounds were identified by GC and GC-MS. The major chemical compounds were carvacrol (28.65%) followed by thymol (21.66%), α-humulene (9.67%), undecanal (8.29%), γ-terpinene (7.76%), ρ-cymene (6.46%), caryophyllene oxide (5.85%), α-terpineol (3.28%) and β-selinene (2.01%). The larvicidal assay was conducted to record the LC₅₀ and LC₉₀ values and the larval mortality was observed after 12 and 24 h of exposure period. The LC₅₀ values of the oil were 33.54 (after 12 h) and 28.37 ppm (after 24 h). The LC₉₀ values of the oil were 70.27 (after 12 h) and 59.38 ppm (after 24 h). The results of the present study showed that the essential oil of P. amboinicus is one of the inexpensive and eco-friendly sources of natural mosquito larvicidal agent to control/reduce the population of malarial vector mosquito.     

Salivary gland proteome of the human malaria vector, Anopheles campestris-like (Diptera: Culicidae)

Anopheles campestris-like is proven to be a high-potential vector of Plasmodium vivax in Thailand. In this study, A. campestris-like salivary gland proteins were determined and analyzed by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), two-dimensional gel electrophoresis, and nano-liquid chromatography-mass spectrometry. The total amount of salivary gland proteins in the mosquitoes aged 3–5 days was approximately 0.1?±?0.05 μg/male and 1.38?±?0.01 μg/female. SDS-PAGE analysis revealed at least 12 major proteins found in the female salivary glands and each morphological region of the female glands contained different major proteins. Two-dimensional gel electrophoresis showed approximately 20 major and several minor protein spots displaying relative molecular masses from 10 to 72 kDa with electric points ranging from 3.9 to 10. At least 15 glycoproteins were detected in the female glands. Similar electrophoretic protein profiles were detected comparing the male and proximal-lateral lobes of the female glands, suggesting that these lobes are responsible for sugar feeding. Blood-feeding proteins, i.e., putative 5′-nucleotidase/apyrase, anti-platelet protein, long-form D7 salivary protein, D7-related 1 protein, and gSG6, were detected in the distal-lateral lobes (DL) and/or medial lobes (ML) of the female glands. The major spots related to housekeeping proteins from other arthropod species including Culex quinquefasciatus serine/threonine-protein kinase rio3 expressed in both male and female glands, Ixodes scapularis putative sil1 expressed in DL and ML, and I. scapularis putative cyclophilin A expressed in DL. These results provide information for further study on the salivary gland proteins of A. campestris-like that are involved in hematophagy and disease transmission.     

A standard cytogenetic photomap for the mosquito Anopheles stephensi (Diptera: Culicidae): application for physical mapping

To facilitate physical genome mapping, we have developed a new cytogenetic photomap for Anopheles stephensi (Liston) (Diptera: Culicidae), an important malaria vector in Asia. The high-resolution images of the ovarian polytene chromosomes have been straightened and divided by numbered divisions and lettered subdivisions. The exact chromosomal locations of eight DNA probes have been determined by fluorescent in situ hybridization. Using the DNA sequences, we have established correspondence between chromosomal arms among An. stephensi, Anopheles gambiae (Patton), and Anopheles funestus (Giles). The results support previous cytogenetic observations of arm translocations taking place during diversification of the species. To make the cytogenetic map useful for population genetics studies, we have indicated the chromosomal positions for the breakpoints of 19 polymorphic inversions.