Abstract:
The present invention discloses a composition for use in luring  Aedes aegypti  mosquitoes to lay their eggs in a target site, which comprises of an effective amount n-heneicosane in a suitable organic solvent. The composition may also contain an effective amount of insecticide capable of killing the mosquito larvae soon after they hatch out of their eggs. measurement switch, a transformer (low voltage/medium voltage), a DC/AC inverter, a capacitor, a voltage dip detection circuit, and a control circuit.

Description:
FIELD OF THE INVENTION 
       [0001]    The present invention relates to compositions for use in the control of dengue transmitting mosquito. In particular, the present invention relates to a composition for use in the control of dengue transmitting mosquito comprising of an oviposition attractant to attract  Aedes aegypti  mosquito to lay eggs at a predetermined site. The present invention also relates to a composition for use in the control of dengue transmitting mosquito comprising of an oviposition attractant to attract  Aedes aegypti  mosquito to lay eggs at a predetermined site and a growth regulator, which inhibits the metamorphosis of larvae to adult mosquitoes. The present invention also relates to processes for preparation of such compositions. The present invention also relates to methods for controlling  Aedes aegypti  mosquito and use of the compositions of the present invention for controlling  Aedes aegypti  mosquitoes. 
       DESCRIPTION OF THE PRIOR ART 
       [0002]    Mosquitoes are medically important vectors, both in number of diseases they transmit and the magnitude of health problems world-wide. About 400 species of mosquitoes can spread disease to humans. These include yellow fever and dengue hemorrhagic fever for which the mosquitoes,  Aedes aegypti  and  Aedes albopictus  are the important vectors. According to the World Health Organization, at least 700 million people each year are infected with disease by mosquitoes. 
         [0003]    Dengue fever is a serious disease of Asia and Africa. In India, public is aware of dengue haemorrhagic fever due to the epidemics in 1996 particularly in Delhi and surrounding regions. Classic dengue, known for its low mortality but very uncomfortable symptoms, has become more serious, both in frequency and mortality, in recent years.  Aedes aegypti  mosquito is the most prominent vector of dengue throughout the world. World wide at present there is no dengue vaccine. Therefore, mosquito control measures are the only remedy to this problem. 
         [0004]    A variety of methods have been practised in the art in an attempt to control  Aedes  mosquitoes. 
         [0005]    One of the methods used for the control of mosquito involves attract and kill devices which uses different combinations such as ultraviolet light, Carbondioxide, and 1-octen-3-ol to attract mosquitoes. 
         [0006]    The main disadvantage of this method is that these devices are not efficient. 
         [0007]    Yet another disadvantage of this method is that these devices are not suitable for control of mosquitoes in the field conditions. 
         [0008]    Still another disadvantage of this method is that it is an expensive method for its public use. 
         [0009]    Another method known in the art for controlling mosquitoes is the use of insecticides such as organophosphorus pesticides (malathion or fenitrothion), carbamates and synthetic pyrethroids by residual application or fogging to kill flying adult mosquitoes and, presumably, some mosquitoes resting on vegetation at that site. 
         [0010]    The main drawback of this method is that it is suitable for control of mosquitoes in the field areas only as they are highly toxic in nature thus causing an environmental hazard. 
         [0011]    Yet another drawback of this method is that more often mosquitoes develop insecticidal resistance. 
         [0012]    Still another drawback of this method is that the adult mosquitoes which come into contact with the insecticides are only killed whereas the eggs/larvae/pupae are unaffected by this method. 
         [0013]    Still another drawback of this method is that non-target organisms are also killed along with mosquitoes such as aquatic fish, etc. Thus it is not environment and eco-friendly. 
         [0014]    Another method known in the art of control of mosquitoes involves larval management. This includes the use of chemical larvicides, microbial insecticides ( Bacillus thuringiensis B thuringiensis israeliensis ), larvicidal oils, juvenile hormone (methoprene), and mosquito predator fish to control mosquito larvae. 
         [0015]    The main disadvantage of these methods is that they are suitable for use in ponds, canals, irrigated fields and some other freshwater sources for mosquito larval control only. 
         [0016]    Another disadvantage of these methods is that closer surveillance is required to find out the larval breeding sites for the application of larvicides. 
         [0017]    Still another disadvantage of microbial control and mosquito predator fish control methods are that they cannot be used in a container breeding mosquitoes like  Aedes aegypti.    
         [0018]    Further another disadvantage of these methods is that the mosquitoes develop resistance to microbial agents. 
         [0019]    Yet another disadvantage of microbial larvicide is that it degrades faster in presence of sun light. 
         [0020]    Another disadvantage of these methods, which are based on larvicides, is that they are not economical to use for the control of dengue transmitting mosquitoes. 
         [0021]    Another disadvantage of the method using mosquito predator fish to eliminate mosquito larvae is that the life-period of the mosquito predator fish depends on the specific characteristics of the water like salinity. Maintenance of such conditions poses limitations for the practical application of this method. 
         [0022]    Still another disadvantage of this method is that it cannot be used for monitoring the vector population. 
         [0023]    Yet another disadvantage of these methods is that they are not cost effective. 
         [0024]    Thus, there is an urgent need for a new mosquito control composition and method, which is simple to use, cost-effective, and environment and eco-friendly. 
       OBJECTS OF THE PRESENT INVENTION 
       [0025]    It is therefore, one of the main objects of the present invention to provide a composition for the control of dengue transmitting mosquito  Aedes aegypt , which overcomes the disadvantages of the prior art. 
         [0026]    Another object of the present invention is to provide an environment and eco-friendly composition for the control of dengue transmitting mosquito  Aedes aegypt , which does not contain any toxic chemical. 
         [0027]    Another object of the present invention is to provide a composition, which can be used easily in ovitraps, coolers, overhead tanks and other dengue transmitting mosquito breeding aquatic habitats. 
         [0028]    Still another object of the present invention is to provide a novel chemical composition, which attracts dengue transmitting mosquitoes to lay eggs at a given site and control mosquito population by blocking the larval growth to adults. 
         [0029]    Yet another object of the present invention is to provide a novel chemical composition which can be used for the control of breeding of dengue transmitting mosquitoes in indoor as well as outdoor applications. 
         [0030]    Still another object of the present invention is to provide a novel chemical composition for the control of dengue transmitting mosquito  Aedes aegypt , which does not affect the other beneficial insects in the same aquatic habitat. 
         [0031]    Yet another object of the present invention is to provide a novel chemical composition for nearly 100% control of dengue transmitting mosquito  Aedes aegypt , within 2 days of use. 
       SUMMARY OF THE INVENTION 
       [0032]    The present invention is based on the unexpected finding that an oviposition attractant pheromone of the dengue transmitting mosquito,  Aedes aegypti  in combination with a suitable growth regulator in proper composition prevents the development of  Aedes aegypti  at larval stage itself, thus controlling the mosquito population by ‘lure and kill technique’ 
         [0033]    Accordingly the present invention provides, a chemical composition, comprising of an oviposition attractant pheromone n-heneicosane and optionally an insect growth regulator in an inert carrier. The said chemical composition controls dengue transmitting mosquito population by attracting the females to lay eggs in water where it is applied and stops further development of larvae to adults. This chemical composition can be applied to any  Aedes aegypti  mosquito breeding in aquatic habitat. 
         [0034]    The composition can be used in different formulations such as spray, tablets and lotions and it kills the larvae within two days after hatching from the egg. The mortality rate is 100%. This method of  Aedes aegypti  mosquito control is simple, cost effective, efficient and eco-friendly. 
         [0035]    Accordingly, the present invention provides a composition for use in luring  Aedes aegypti  mosquitoes to lay their eggs in a target site which comprises of an effective amount n-heneicosane in a suitable organic solvent. 
         [0036]    In a preferred feature, said solvent is selected from as petroleum ether, n-pentane, n-hexane, methanol or dichloromethane. 
         [0037]    In a preferred feature, the ratio of n-heneicosane to said solvent is preferably, from 1:20 to 1:70. 
         [0038]    In a preferred feature, said composition is a lure and kill composition and further comprises an effective amount of an insect growth regulator or insecticide. 
         [0039]    In a preferred feature, said insecticide is a larvicide. 
         [0040]    In a preferred feature, said insect growth regulator is selected from diflubenzuron, neemazal and triflumaron. 
         [0041]    In a preferred feature, said insect growth regulator comprises 100-120 mg in diflubenzuron 500 ml of an organic solvent selected from acetone or dichloromethane. 
         [0042]    In a preferred feature, said n-heneicosane concentration is 8-20 g preferably 10-12 g per litre of the composition. 
         [0043]    The resent invention also provides a process for the preparation of a composition for the control of dengue transmitting mosquito having lure and kill property comprising of the steps of: 
         [0000]    (a) preparing oviposition attractant solution by dissolving n-heneicosane an organic solvent;
 
(b) preparing insect regulator solution by dissolving an insect growth regulator in an organic solvent;
 
(c) mixing the above two solutions thoroughly.
 
         [0044]    In a preferred feature, the ratio of n-heneicosane to said solvent is preferably, from 1:20 to 1:70. 
         [0045]    In a preferred feature, said insect growth regulator comprises 100-120 mg of diflubenzuron in 500 ml of an organic solvent selected from acetone or dichloromethane. 
         [0046]    In a preferred feature, the process comprises preparing oviposition attractant solution by dissolving 8 to 20 g of n-heneicosane preferably 10-12 g in 500 ml of organic solvents like n-pentane, n-hexane or dichloromethane, preparing insect regulator solution by dissolving 800-2000 mg of insect growth regulator like diflubenzuron preferably 100-120 mg in 500 ml of organic solvents like acetone or dichloromethane and mixing the above two solutions thoroughly. 
         [0047]    The present invention also provides use of n-heneicosane for luring  Aedes aegypti  mosquitoes to lay their eggs in a target site. 
         [0048]    The present invention also provides use of n-heneicosane in the preparation of a lure and kill composition for luring  Aedes aegypti  mosquitoes to lay their eggs in a target site and killing them at the larval stage. 
         [0049]    The present invention also provides a method for luring  Aedes aegypti  mosquitoes to lay their eggs in a target site which comprises treating said target site with an effective amount of a composition containing n-heneicosane. 
         [0050]    The present invention also provides a method for luring  Aedes aegypti  mosquitoes to lay their eggs in a target site which comprises treating said target site with an effective amount of a composition as claimed in any one of claims  1  to  8 . 
     
    
     DETAILED DESCRIPTION 
       [0051]    The present invention relates to the control of  Aedes aegypti  mosquitoes based on “lure and kill” technique. These mosquitoes are lured chemically to lay eggs only at a specific site. When the eggs are hatched, the entire larvae population can be destroyed. According to the invention the composition of the present invention ensures that the eggs are laid only in a specific site and not anywhere. 
         [0052]    The present invention is based on the unexpected finding that heneicosane, a C-21 hydrocarbon, is capable of acting as a most effective oviposition attractant. Thus, any standing water containing a dose of heneicosane will attract and lure  Aedes aegypti  mosquitoes to lay their eggs only in such water and no other. Subsequently, either during the larval stage or just before the eggs are hatched, an effective dosage of a conventional insecticide may be added to completely destroy the larvae. 
         [0053]    In an alternative embodiment, heneicosane may be synergistically premixed with an insecticide so that the composition itself acts as a “lure and kill” composition. 
         [0054]    The present invention also relates to a process for the preparation of compositions for use in lure and/or killing of  Aedes aegypti  mosquitoes. The present invention is based on the unexpected finding that  Aedes aegypti  mosquitoes tend to lay eggs in stagnant water where eggs of the same species of mosquitoes had been laid previously. This led the inventors to analyse various samples of water which in turn led to the finding that water in which eggs had previously been laid by  Aedes aegypti  mosquitoes and which had hatched into larval stage contained various semio-chemicals. Analysis of these semio-chemicals identified them as heneicosane, nonacosane, octadecane, isopropyl myristate and docosane. Of these five chemicals, heneicosane was found to be most effective oviposition attractant which chemically “invites” the mosquitoes to lay their eggs in the same place again and again. When, stagnant water in which  Aedes aegypti  mosquitoes had bred earlier was left untouched the incidence of further egg laying in the same water and thriving of larval population increased. However, when such water was replaced by fresh water the incidence of egg laying decreased considerably and thereafter, gradually increased over a period of time. However, when stagnant water was kept alongside water containing heneicosane, the entire egg laying took place only in water containing heneicosane thereby clearly establishing that the  Aedes aegypti  mosquitoes were attracted to water containing heneicosane for the purposes of laying their eggs. Thus, the present invention makes it possible to lure  Aedes aegypti  mosquitoes to lay their eggs only at specific chosen sites making it possible to thereafter to destroy the entire larval population and control further breeding of the mosquitoes. This has a tremendous industrial application. It minimises the use of insecticides and therefore, wastage thereof, since in accordance with the present invention, the insecticides are required to be used only at specific target sites instead of over a large area. It also ensures that the insects do not escape. Spraying of in insecticides over a large area offers tremendous opportunities to an adult mosquito to escape. On the contrary, present invention ensures that the entire larval population is congregated at a specific site where 100% killing thereof can be achieved. 
         [0055]    The present invention also relates to a process for the preparation of the composition of the present invention. In the first aspect, the invention relates to preparation and isolation of heneicosane. In this embodiment,  Aedes aegypti  mosquitoes are bred and allowed to lay eggs in stagnant water. Once the eggs are hatched and during the larval stage thereof, the water is collected. This water is referred to as larval conditioned water (LCW). The semio-chemicals are thereafter extracted from the LCW. Conventional organic solvents such as petroleum, ether, hexane, dichloromethane, or methanol are used for the extraction of the semio-chemicals. The most preferred solvent is hexane. The extracted solution comprised of the following five semio-chemicals: heneicosane, nonacosane, octadecane, isopropyl myristate and docosane. From the mixture eof such five semio-chemicals, henesicosane is separated. 
         [0056]    A composition comprising or containing heneicosane is sprayed into water referred to herein as target site. The composition may include any insecticide. In the next few days, there is a tremendous incidence of egg laying activity by  Aedes aegypti  mosquitoes at the target site. The composition may already contain a conventional insecticide, which is effective against  Aedes aegypti  larvae. Alternatively, the target site is treated with a conventional insecticide or insect growth regulator such as diflubenzuron, neemazal, triflumaron. Diflubenzuron is the preferred insect growth regulator. 
         [0057]    In a preferred embodiment, solutions of oviposition attractant containing heneicosane and insect growth regulator are prepared and kept separately. These two compositions are subsequently mixed in a predetermined ratio, preferably at a ratio of 1:1. The two compositions may be mixed either in advance or just prior to use. 
         [0058]    The composition of the present invention has been found to be extremely effective in “lure and kill” technique against  Aedes aegypti  mosquitoes. 
         [0059]    In a preferred embodiment, the process in accordance with the present invention for the preparation of new chemical composition for the control of dengue transmitting mosquitoes comprises of following steps: 
       Step (a): Preparation of Oviposition Attractant Solution: 
       [0060]      Aedes aegypti  mosquitoes are bred and the water is collected for isolation where the mosquitoes are at larval stage. This water is called as larval conditioned water (LCW). The semio-chemicals are extracted from the LCW by organic solvents such as petroleum ether, hexane, dichloromethane or methanol preferably hexane. The extractant is analysed by various spectral techniques like IR, NMR and GCMS and the five semio-chemicals identified are heneicosane, nonacosane, octadecane, isopropyl myristate and decosane. Amongst these heneicosane (C 21 hydrocarbon) is found to be the most effective oviposition attractant. 
         [0061]    In a stoppered bottle, 1.6 to 2% w/v preferably 1.0-1.2% w/v of n-heneicosane is dissolved completely in n-pentane or n-hexane HPLC grade preferably n-hexane. 
       Step (b): Preparation of Insect Growth Regulator Solution: 
       [0062]    In a stoppered bottle, 0.16 to 0.2% w/v, preferably, 0.1 to 0.12% w/v of insect growth regulator like diflubenzuron, neemazal, triflumaron preferably diflubenzuron is dissolved completely in acetone HPLC grade. 
       Step (c): Preparation of the Chemical Composition Having Lure and Kill Property: 
       [0063]    The oviposition attractant solution prepared in step (a) and insect growth regulator solution prepared in step (b) are taken in a ratio of 1:1 and the mixture is shaken thoroughly. This composition can readily be used to control the dengue transmitting mosquitoes breeding by “lure and kill” technique. 
         [0064]    The above chemical composition can be used in different formulations such as spray, tablets and lotions. Extensive field trials with this chemical composition conducted after adequate precaution of confidentiality indicated that larvae after hatching from the eggs die within 2 days and the mortality rate is 100%. The water in ovitraps containing this chemical composition is found to be effective for 7 to 10 days to attract the  Aedes aegypti  to lay eggs and kill the larvae a second time after hatching from the eggs relaid in the same water. This method of mosquito control is simple, cost effective, efficient and eco-friendly. 
         [0065]    In a preferred embodiment, the process for the preparation of the composition of the present invention comprises comprising of the steps of:
       (a) preparing oviposition attractant solution by dissolving 8 to 20 g of n-heneicosane preferably 10-12 g in 500 ml of organic solvents like n-pentane, n-hexane or dichloromethane.   (b) preparing insect regulator solution by dissolving 800-2000 mg of insect growth regulator like diflubenzuron preferably 100-120 mg in 500 ml of organic solvents like acetone or dichloromethane.       
 
         [0068]    The present invention will now be described in greater detail with the following examples. It be understood by a person skilled in the art that the examples are for illustrative purposes only and various modifications of the present invention are possible without departing from the spirit and scope thereof. 
       EXAMPLE 1 
     The Following Examples Illustrate the Process for Preparing the Composition of the Present Invention 
       [0069]    To a 1 L stoppered bottle, 250 ml of or n-pentane (HPLC grade) is taken. To this 13 g of n-heneicosane g is added and the flask is shaken well to dissolve it completely. 
         [0070]    250 ml of acetone HPLC grade is taken in another 500 ml of stoppered bottle. To this 100 mg of diflubenzuron is added and the flask is shaken well to dissolve it completely. This is transferred to the bottle containing the heneicosane solution and the mixture is shaken thoroughly. This composition is ready for use for the control of dengue transmitting mosquito breeding. 
       EXAMPLE 2 
       [0071]    To a L stoppered bottle, 250 ml of n-hexane HPLC grade is taken. To this 10 g of n-heneicosane g is added and the flask is shaken well to dissolve it completely. 
         [0072]    250 ml of acetone HPLC grade is taken in another 500 ml of stoppered bottle. To this 100 mg of diflubenzuron is added and the flask is shaken well to dissolve it completely. This is transferred to the bottle containing the heneicosane solution and the mixture is shaken thoroughly. This composition is ready for use for the control of dengue transmitting mosquito breeding. 
       EXAMPLE 3 
       [0073]    To a L stoppered bottle, 250 ml of or n-pentane HPLC grade is taken. To this 20 g of n-heneicosane is added and the flask is shaken well to dissolve it completely. 
         [0074]    250 of acetone HPLC grade is taken in another 500 ml of stoppered bottle. To this 150 mg of diflubenzuron is added and the flask is shaken well to dissolve it completely. This is transferred to the bottle containing the heneicosane solution and the mixture is shaken thoroughly. This composition is ready for use for the control of dengue transmitting mosquito breeding. 
       EXAMPLE 4 
     The Following Examples Illustrate the Effectiveness of the “Luring” Effect 
       [0075]    The following experiment was conducted under field conditions in a specially cordoned area to which public did not have any access. Fifteen open tanks with water filled to the top were kept in the open. All the tanks were made of same material, were of the same size and contained same amount of water from the same source. The tanks were numbered consecutively and allowed to remain for a week. After fifteen days all the tanks were examined and it was found that all the tanks contained nearly the same amount of larval population of  Aedes aegypti  mosquitoes. This establishes that the mosquitoes did not have any preference for any particular water tank. 
         [0076]    All the fifteen tanks were drained of water and thoroughly cleaned. Subsequently, they were filled with water from the same source. In tank No. 1, a predetermined amount of a solution of heneicosane in n-hexane (1% w/v) was added. After fifteen days, water from each tank was analysed. It was found that only tank No. 1 containing the heneicosane contained mosquito larvae, thereby establishing that  Aedes aegypti  mosquitoes were clearly attracted and lured to lay eggs in water containing heneicosane. 
       EXAMPLE 5 
       [0077]    Example No. 4 was repeated 14 times with heneicosane being added in tank Nos. 2 to 15 consequentially. It was found that each time the tank containing heneicosane attracted  Aedes aegypti  mosquitoes to lay their eggs therein. 
       EXAMPLE 6 
       [0078]    Example Nos. 4 and 5 were repeated with heneicosane dissolved in n-pentane. The results were exactly the same. 
       EXAMPLE 7 
       [0079]    The above example was repeated with varying concentrations of heneicosane in the solvent. The concentration of 1.0 to 2% w/v was found to be most effective.