Biological control is the use of natural enemies such as parasitoids, pathogens and predators to control the targeted pest by decreasing its populations and making it less damaging than it would be otherwise. It forms part of the integrated pest management (IPM) strategies. "Biological control may be the result of purposeful actions of men or may result from unassisted action of natural forces" (R.G Van Diresche, T.S Bellows, 1996). It is a mean to lower the pest population of crop and forest or to balance the ecological system affected by nonnative pests and according to J.Brodeur et al. it is also environmentally sounds and very effective.
1.2 Biological control of Diamondback moth (DBM):
For controlling of DBM biologically, the use of its natural enemies is used, particularly parasitoids which attacks different stages of its life cycle namely: DBM eggs, larvae, pupae and adults which help to maintain its population below damaging threshold. This method has had a major boost since the discovery of larval parasitoids that seem to keep the DBM population in check (Lim, 1986; Lloyd, 1940; Chua and Ooi, 1986). DBM has been pointed out to be a good example of a potentially serious pest held in control naturally by parasitoids by Xu et al. (2001).
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More than 90 parasitoids species have been recorded for DBM and an average of 60 of them appear to be important (Goodwin, 1979; Lim, 1986). However, the major ones are in the genera Diadegma, Cotesia and Microplitis.
NOTE: in this experiment cotesia plutellae has been used as the parasitoid.
Cotesia plutellae (Hymenoptera Braconidae) belongs to the family of Microgastriinae. It is solitary and a DBM -specific larval parasitoid. It is reported to be host specific to P.Xylostella by Verkerk and Wright, 1996. It occurs naturally in Taiwan but in many countries it has been introduced to control DBM. Waladde et al. (2001) found that C.Plutellae was active year round, accounting for 30-50% parasitism. It has been found that it can adapt to different region of the world very easily, it is also very aggressive and it can target it host easily, it has been found also that it is tolerant to pesticides used against DBM and it is more resistant by hyper parasites in contrast to other parasitoids of the same type (Mc Donald and Kok, 1991). This shows that it is an effective parasite of DBM.
2.2Biology of cotesia plutellae:
Its egg is always laid inside the larval body of DBM. The egg is transparent with a rounded shape end, which have a straight short peduncle (stalk).
After 5 hours of oviposition the broader anterior end of the egg measures 0.064mm and length 0.251mm with a peduncle of 0.05mm long. After 24hr the size of the egg increases from 0.12mm to 0.4 mm. The egg is found freely floating in the haemocoel (insect blood) of the DBM larva. There are sometimes more than one egg out of these eggs only one survive to pupal stage. The egg hatches and give birth to the 1st instar larva.
2.2.2 1st instar
After emergences the larva stars feeding of the blood of the DBM. It has to go through 3 instar before pupation. It is transparent and possesses a caudal appendage (tail) and mendibles . The larva measures 0.45mm long (excluding the tail) 0.1mm wide and 0.7mm long, 0.2mm wide on the first and second day respectively. It lasts between 1 and 2 days.
2.2.3 2nd instar
It is white in colour, translucent and it differ considerably from the 1st instar. During this stage the larva grows quickly, it triple its size. This instar last for 4 to 5 days.
2.2.4 3rd instar
It appear like a hymenopterous larva. It has a creamy white to yellowish green colour. Its length is 3.5mm and width is 1.1mm. The fully grown larva emerges from the DBM larva which kills the host consequently. It last for 1 to 3 days. It emerges from the DBM which is in the 4th instar. To come out the cotesia plutellea larva wake a suture on one side it make great effort to come out about 30 to 60 minutes.
The larva start to spin a white silk thread cocoon around itself. The cocoon is found below the surface of the plant leaves. It last for 2 to 7 days and on average 5 days.
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To emerge from the cocoon the adult make a round hole at one end. The females measure 2.62mm long and o.73mm wide and males 2.56mm long and 0.77mm wide. With appropriate food the adult can live more than 15 days and they can lay more than 200 eggs. They have a total life cycle (from egg to adult emergence) averages 13 days.
3.1Diamondback moth ( Plutella Xylostella)
The DBM lives through four different life stages namely: egg, larva, pupa and adults. All stages are very distinct and different from each other. For example they have different behaviour and their need in food also is different.
3.2Biology and life cycle of DBM
The egg are laid most of the time under the leaf surface randomly distributed along the edges of leaf vein of only crucifers plant. The DBM female adult lays about two 100 eggs in her lifespan from 3 to 7 days. The egg size of one egg of the DBM is very small(< 1mm). The initial colour of the egg is white and changes to brownish yellw as it attains maturity. From this stage the egg is ready to hatch itself into larvae. The eggs do not east any food and this egg stage lasts from 4 to 6 days.
The DBM goes through four instar before pupation. Nearly all the eggs laid hatched into a translucent and small larva of about 2 -3 mm long. This instar last fro about 2 days . In the second instar the caterpillar gets bigger and its length is anout 5 to 7 mm long. They have a green colour and a dark head. They get more voracious and there feeding rate increases. The second instar last for 3 to 4 days.
3.2.2 3rd instar
They get even bigger and they are about 8 to 10mm long with a green body and a dark head. They get even more voracious and there feeding rate increased more than before. During this instar they form holes of 1-2cm in the cabbage leaves. The third instar last for 3 to 4 days
3.2.3 4th instar
The caterpillar gets bigger and bigger and their feeding rate increases exponentially. By now the larva is from 10-12mm in length and is green in colour with a dark black head. They feed for 3 - 4 days and eat very slowly or not at all at the end of the 4th instar. It last for about 3 - 4 days
The fourth instar larva starts to make silken cocoon around itself and it hide inside the cocoon to form pupa. At the start the pupa is green but gradually turns brown as the larva start to change in a moth. The size of the pupa does not feed and thus is harmless. The pupation last for 4-5 days and then it emerges as a moth.
Insecticides are chemical substances used to kill pests. To kill larvae and eggs, ovicides and larvicides are used and they form part of insecticides.
The agricultural productivity has boost up in the 20th century with the use of chemicals such as insecticides. Almost all insecticides cause changes in the ecosystem; some act as toxin to mankind and others accumulates along food chains. Insecticides may be either organic or inorganic.
4.2.1 Classes of insecticides
4.2.2. Organochlorine compounds
The organochlorine insecticides have a historical importance as major class of synthetic insecticides. During the World War 2, the supplies of traditional botanical insecticides were limited by blockages and shortages. Due to this organochlorine insecticide was introduced as an alternative. Despite their great efficiency their production was stopped because of their high level of toxicity and they are not readily biodegradable.
DDT is one of the well known synthetic insecticides. According to the National Toxicology Program (NTP) of United States DDT is classified as "moderately toxic" and "moderately hazardous" by the World Health Organization (WHO), based on the rat oral LD50 of 113 mg/kg.
Organophosphates are the next most developed class of pesticides after DDT. They bind to enzymes involve in the functioning of the nervous system such as acetyl cholinesterase and other cholinesterase. Binding to those enzymes causes malfunctioning of the nervous system, disrupting the normal functioning of the insects and eventually kill the insects. Organophosphates normally accumulate in food chains and affect the wildlife. Therefore exposures to those chemicals increase the toxicity in the environment.
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The mechanisms for Carbamates insecticides have the same effect on the insects as Organophosphates. However, its action in not for long term hence it is less toxic compared to Organophosphates.
Referring to the natural compound pyrethrum that has some natural insecticidal effect, Pyrethroids have been developed. Pyrethroids are not persistent in the environment. According to Ernest Hodgson (2010) Pyrethrins have an effect on membranes of nerve cells by acting on the sodium and potassium pump, which results in the depolarization of the membranes. They are also less toxic compared to the other two insecticides mentioned above. Pyrethroids are normally used against household pests.
Neonicotinoids are derived from the natural insecticides Nicotine. These chemicals affect the nicotinic acetylcholine receptor on membranes of nerve cells."Nicotinoids is attributed to actions on post-synaptic receptors (Burkingham et al., 1997; Nagata et al., 1998) which are located in the central nervous systems. Neonicotinoids are broad-spectrum that is they can affect a range of insects and have immediate effect. Used as an alternative to Carbamates and Organophosphates, the treated insects demonstrate physical abnormality, paralysis and eventually death.
4.2.7. Biological insecticides
With the introduction of chemicals that act as insecticides, accumulation of toxins in food chains, along the ecosystems, has increased. Therefore biological insecticides have been reintroduced to reduce toxins in the environment. Nicotine and Pyrethrin were the natural insecticides used in old times by our ancestors. The bacterium Bacillus Thuringiensis that causes bacterial disease for Lepidopterans and some other insects is being in great use. Since the Bacillus Thuringiensis has little effects on other organisms, it is known to be environmentally friendly. Through the use of genetic engineering, the toxin of the bacterium has been incorporated into plants directly which avoid spraying.
Antifeedants are substances produced by plants that prevent insect from feeding upon them but do not affect the insects directly. The insects normally die from starvation. Antifeedants are not toxic therefore they may be used as an eco friendly insecticides.
4.3. Insecticidal application:
Application of insecticides is the way by which insecticides are applied to the biological targets that is pest. There are two types of insecticides namely:
Residual insecticides :
They are chemicals that are applied to provide a long term effects. They are applied on broad surfaces such as wall, floors and ceilings.
Non residual insecticides:
They chemicals are applied to kill directly the insects on the spot rather than providing a long term effects. They are applied either as space treatment or contact treatment. However, they are not as effective as residual insecticides.
4.4. Classification according to mode of entry
The toxic compounds found in the insecticides can enter the vital organs of the insects through various parts of the body. The normal behavior of the insects may simplify the entry of toxicants into their vital organs thus leading to different kind of physical and physiological disorder or may even cause death. While feeding or cleaning body parts by mouth organs, toxicants made their entry through the stomach. It can also happen through cuticle, when insects move near treated surface. Breathing through the spiracles is another way by which toxicants get access to the vital organs of the body. Therefore the toxicants can be categorized as stomach poisons, contact poisons and fumigants.
4.5. Classification according to mode of action
The groups of insecticides classified by the mode on the insect are as follows:-
4.5.1. Physical poisons
Insects are killed by the action of various chemical substances. "No direct chemical or biochemical effect is caused by these insecticides and considering these under chemical insecticidal control may appear as out of place but finds its place in the general classification of insecticides" (GHOSH M R Put the year). In order to make the effect of insecticides more efficient, some of those products can be inserted in the formulation. The effects exerted by these insecticides include:
4.5.2. Protoplasmic poisons
Precipitation of protein occurs primarily as a reaction in the insect cells. Fluorides, arsenicals, fluosilicats, borates are examples of inorganic insecticides and examples of organic insecticides are nitrophenols, nitrocresols, fatty and mineral oils, formaldehyde. Heavy metals like copper and mercury when inserted cause precipitation of protein cells in the mid gut epithelium.
4.5.3. Respiratory poisons
These types of poisons differ completely from the physical asphyxiants which are about the entry of oxygen into the breathing system in the physical ways. The poisons included in this group lead to anoxia. These insecticides mostly include fumigants such as HCN,H2S and CO that together with air enter the breathing system and disrupt completely the normal functioning of the oxygen transport. They alter the metabolism of the respiratory enzymes.
4.5.4. Nerve poisons
Neuron is responsible for the conduction of the nerve impulses in insects. Neurone are known as nerve cells. Synaptic junctions are bridged by acetylcholine. After each function of the transport of impulses through the nerve cells, the acetylcholine has to be hydrolyzed so as to bring in the remaining nerve cells. This hydrolysis is made possible by the action of enzyme (acetlycholinesterase). Due to this fact, the nerve continues to conduct impulse and these results to an increased excitation that induces the production of a new coactive substance by the central nervous system. Accumulation of this product leads to malfunctioning of the normal nerve. Tremor, convulsion, paralyses or in some extreme case death may occur.
Nerve poisons need to be liposoluble so as to pass through the final barrier. Pyrethrins and nicotine are examples of nerve poisons.
Chapter 2: Methodology:
Choice of sample:
The Brassicaceae family being large favors the choice of sample. Being a popular cultivar of the family Brassicaceae, cabbage was chosen as sample. Cabbage is available throughout the year with the introduction of new cultivars of cabbage for the winter months.
Cotesia Plutellae was chosen as the parasitoids of Plutella Xylostella
Collection of sample:
Larvae of Plutella Xylostella were captured from field of cabbages which are insecticides free.
Adult of Cotesia Plutellae were brought from AREU Division of Entomology.
Rearing of Plutella Xylostella:
Larvae of plutella xylostella were captures from cabbage fields and reared in wooden cages covered with fine nylon mesh.
Cabbage cleaned pesticides free leaves were fed to the larvae.
Cocoon was formed and uses the leaves as support.
The emerging adults were given 5% of honey solution
Fresh cabbage leaves were placed in bottle of water in the cages where the female adults can oviposit on
Once the eggs lay on the cabbage leaves they were transferred on fresh cabbage leaves to allow hatching
Once the eggs are hatched, fresh cabbage is fed to the newly hatched larvae and they were allowed to immerge into adults.
Consequently, the adults were used to breed new population of plutella xylostella.
Rearing of Cotesia Plutellae:
The coccons were placed on filter paper in Petri dishes in insect cages.
Once the adults emerged, they were fed with 10% of honey solution.
Fresh cabbage leaves with plutella xylostella of second instar were supplied to the cotesia adult.
The next day the larvae of plutella xylostella are analyzed. If the larvae are parasitized they tend to change color from green to white.
The parasitized larvae are then transferred to another cage were they are fed with fresh cabbage leaves.
New coccons are formed and used the leaves as support.
The coccons were transferred to a new cage where a new generation of cotesia plutellae is emerged. The adults were used to breed new generation, consequently increasing the population.
Coccons of cotesia plutellae was brought from AREU division of entomology.
Coccons of cotesia was brought from AREU division of enthomology.
Demarcation of plot by 10 by 10m.
The demarcation was done on the university farm where a piece of land was allocated on the 18th of October.
Manure was applied to maximize nutrient for the plants.
The holes were distanced according to " Le Guide Agricole."
Preparation of plot: weeding, manure added, holes are dug.
Branches of plants were used to provide shade to the seedlings to minimize transpiration.
Seedling was transplanted into the holes with moist soil.
Top dressing was done along with soil leveling to optimize growth of the cabbages.
Watering was carried out thrice a week.
Note: no pesticides treatment was done for the cabbages since they were used to feed the plutella xylostella which were reared.
Preparation of SORBA:
According to the instructions given on the package of the insecticides SORBA 1ml of the insecticides is diluted in 1 Litre of water.
Dilution of SORBA to different concentration:
CONCENTRATION OF SORBA.
VOLUME OF WATER ADDED/ml
VOLUME OF SORBA ADDED
Assessing effect of SORBA on adults Cotesia Plutellae:
5 adults Cotesia Plutellae were placed using a putter in a petri dish with a filter paper sprayed with SORBA.
2 drops of 5%honey solution were placed on the cover of the petri dish.
Each concentration had 3 replicates labelled R1,R2 and R3.
Results were taken every 24 hours. Mortality rate was recorded.
Assessing the effect of SORBA on coccons of Cotesia Plutella:
5 coccons of Cotesia plutellae were placed on a petri dish along with a filter paper.
Using a Potter Spray Tower (Burkard CO.), different concentration of SORBA was sprayed.
Every 24 hours, the emergence and mortaity rate of the insect was recorded.
Assessing the effect of SORBA on larvae of cotesia:
Discs of 2.5cm of cabbage leaves were made.
In a petri dish, 5 disc and parasitized plutella xylostella larvae were placed.
Three instars of Plutella Xylostella was used namely the second instar, third instar and forth instar.
Using a sprayer the different concentrations of SORBA was sprayed.
For each instar and concentration of SORBA each procedure were replicated three times.
Every 24 hours, mortality rate of Plutellae xylostella, pupation of Cotesia Plutellae and emergence were recorded.