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Biology of Malaria Vectors Biology of Malaria Vectors and Parasite-Vector and Parasite-Vector
Relationships Relationships
Dawn WessonDawn Wesson
Tulane Department of Tulane Department of Tropical MedicineTropical Medicine
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Malaria Vector BiologyMalaria Vector Biology
Anopheline Life Cycle – habitat Anopheline Life Cycle – habitat preferences, types of habitat, preferences, types of habitat, unpolluted waterunpolluted water
Effect of human activities on habitat Effect of human activities on habitat creation – agriculture, irrigation, etc.creation – agriculture, irrigation, etc.
Biology of Malaria Vectors – General Biology of Malaria Vectors – General and Specificand Specific
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Family Culicidae > 3500 species 3 subfamilies:
Anophelinae - Anopheles, Bironella and Chagasia, ~ 500 species
Toxorhynchitinae - Toxorhynchites, 70+ species (all non-bloodfeeding)
Culicinae - Aedes, Culex, Haemagogus, Mansonia, and all other genera, > 3000 species
Anophelinae
Toxorhynchitinae
Culicinaetime
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Anopheles mosquito life cycle
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eggs
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Anopheline Culicine
Adult
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Genus Anopheles6 subgenera:
Cellia - >230 species, most important Old World malaria vectors (Africa and Asia)
Anopheles - >180 sp., were the most important malaria vectors in Europe and N. America
Nyssorhynchus - >40 sp., most important New World malaria vectors
Kertezia - >10 sp., NW, bromeliads
Lophopodomyia – 6 sp., NW tropics
Stethomyia – 5 sp., NW tropics
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AnophelesAnopheles Habitat Habitat PreferencesPreferences
Effects of human activitiesEffects of human activities Major malaria vectors tend to be Major malaria vectors tend to be
colonizing species in temporary colonizing species in temporary habitats free of established predatorshabitats free of established predators
They have evolved with humans to They have evolved with humans to take advantage of these take advantage of these
environmentsenvironments
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LARVAL HABITAT - LARVAL HABITAT - An. albimanusAn. albimanus in Cuba in Cuba
WHO/TDR/Service, 1992
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LARVAL HABITAT - LARVAL HABITAT - An. bellator An. bellator in Brazil in Brazil from bromeliadesfrom bromeliades
WHO/TDR/Service, 1992
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LARVAL HABITAT - LARVAL HABITAT - An. pseudopunctipennisAn. pseudopunctipennis in Mexico in Mexico
WHO/TDR/Service, 1992
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LARVAL HABITAT - LARVAL HABITAT - An. stephensiAn. stephensi from from water tanks on rooftops in Dubaiwater tanks on rooftops in Dubai
WHO/TDR/Service, 1992
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LARVAL HABITAT - LARVAL HABITAT - Standing water Standing water created by road created by road
building in Beninbuilding in Benin
WHO/TDR/Olliaro, 1988
LARVAL HABITAT - LARVAL HABITAT - Irrigation ditches Irrigation ditches provide provide AnophelesAnopheles breeding sites in the breeding sites in the GambiaGambia
WHO/TDR/Lindsay, 1991
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WHO/TDR/Lindsay, 1991
WHO/TDR/Martel, 1994
LARVAL HABITAT - Rice fields and irrigated areas provide LARVAL HABITAT - Rice fields and irrigated areas provide AnophelesAnopheles breeding sites in Viet Nam and the Gambia breeding sites in Viet Nam and the Gambia
1515WHO/TDR/Service, 1992
WHO/TDR/Ragavoodoo, 1992
Roof water breeding Roof water breeding site of site of An. arabiensisAn. arabiensis in Mauritiusin Mauritius
Water storage Water storage pots, breeding site pots, breeding site of of An. gambiaeAn. gambiae and other and other mosquitoes in mosquitoes in NigeriaNigeria
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WHO/TDR/HOLT Studios, 1992
Anopheles gambiae
Biology of Biology of Anopheles Anopheles gambiaegambiae
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Anopheles gambiaeAnopheles gambiae Major malaria vector in sub-Saharan Major malaria vector in sub-Saharan
Africa Africa Typical anopheline life cycle, but Typical anopheline life cycle, but
extreme preference for living around extreme preference for living around and feeding on humansand feeding on humans
Preferred Preferred ovipositionoviposition sites – small sites – small temporary pools in full sunlighttemporary pools in full sunlight
Seasonal abundance correlates with Seasonal abundance correlates with rainfallrainfall
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Anopheles gambiaeAnopheles gambiae – life – life cyclecycle
Other sites – irrigated areas (rice Other sites – irrigated areas (rice fields); drying streams in dry season; fields); drying streams in dry season; habitats created by humans habitats created by humans
EggsEggs laid on water or damp soil; hatch laid on water or damp soil; hatch 48 hr. – 2 weeks48 hr. – 2 weeks
LarvaeLarvae can crawl across damp soil can crawl across damp soil from drying pool to another with waterfrom drying pool to another with water
Larval development - <week with Larval development - <week with sufficient temperature and foodsufficient temperature and food
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Anopheles gambiaeAnopheles gambiae – life – life cyclecycle
Larvae are filter feeders on surface Larvae are filter feeders on surface film – algae and bacteria film – algae and bacteria
PupationPupation in full sunlight – can be in full sunlight – can be induced in laboratory with lightinduced in laboratory with light
Pupal development in 24 hr. – 3 Pupal development in 24 hr. – 3 days; temperature dependentdays; temperature dependent
Adult emergenceAdult emergence at night at night Both sexes need 24 hr. to reach Both sexes need 24 hr. to reach
sexual maturity – male terminalia sexual maturity – male terminalia (genitalia) rotate 180(genitalia) rotate 180..
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Mosquito Emerging from Pupal Exuvia
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Anopheles gambiaeAnopheles gambiae – adult – adult behaviorbehavior
Male mosquito swarming behavior – Male mosquito swarming behavior – females fly into swarm to females fly into swarm to mate mate (not (not well documented in wild well documented in wild An. gambiaeAn. gambiae but does occur in lab colonies). but does occur in lab colonies).
Male activity increases at sundown. Male activity increases at sundown. Changes in antennae (plumes folded Changes in antennae (plumes folded up during day – open to detect female up during day – open to detect female flight sound; Johnston's organ)flight sound; Johnston's organ)
Males attracted to females and mate Males attracted to females and mate in flight – females probably mate only in flight – females probably mate only once (?) – store sperm in once (?) – store sperm in spermathecae spermathecae
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Anopheles gambiaeAnopheles gambiae – host – host seekingseeking
Mated Mated An. gambiaeAn. gambiae females females seek seek bloodblood at night (after sundown) - at night (after sundown) - ~90% of bloodmeals taken from ~90% of bloodmeals taken from sleeping human hosts and they sleeping human hosts and they usually rest on the inside walls of the usually rest on the inside walls of the house to digest the meal house to digest the meal
Egg development takes about 48 hrs Egg development takes about 48 hrs during warm season – longer in cooler during warm season – longer in cooler weather weather
Oviposition occurs at night – usually Oviposition occurs at night – usually the 2nd night after a bloodmeal the 2nd night after a bloodmeal
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Anopheles gambiaeAnopheles gambiae – host – host seekingseeking
The female then searches for another The female then searches for another bloodmeal - during warm season, a female bloodmeal - during warm season, a female is capable of ovipositing every other night is capable of ovipositing every other night
This behavior has implications for the This behavior has implications for the timing of timing of host seekinghost seeking by by An. gambiaeAn. gambiae females – early evening blood-seeking females – early evening blood-seeking females are probably feeding for the first females are probably feeding for the first time (they have not laid eggs yet – time (they have not laid eggs yet – nulliparous), while older (parous) females nulliparous), while older (parous) females tend to seek blood later at night (they tend to seek blood later at night (they have to oviposit first)have to oviposit first)
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Anopheles gambiaeAnopheles gambiae – host – host seekingseeking
Extrinsic incubation periodExtrinsic incubation period (minimum) (minimum) of of Plasmodium falciparumPlasmodium falciparum in the mosquito in the mosquito is 8-10 days – so under ideal conditions, is 8-10 days – so under ideal conditions, the female would take 5-6 bloodmeals in the female would take 5-6 bloodmeals in the process of acquiring parasites and the process of acquiring parasites and living long enough to transmit them (about living long enough to transmit them (about 2 weeks) 2 weeks)
In real life…environmental factors will In real life…environmental factors will usually affect time line – temperature, usually affect time line – temperature, rainfall, wind will interfere with the ability rainfall, wind will interfere with the ability to oviposit and blood-feed at will. Most to oviposit and blood-feed at will. Most field collected field collected An. gambiaeAn. gambiae females with females with P. P. falciparumfalciparum sporozoites in their salivary sporozoites in their salivary glands have taken 3-4 blood mealsglands have taken 3-4 blood meals
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• If, after locating host and ingesting blood, the blood meal is large, distention-induced host seeking inhibition is triggered
• This tapers off as the blood is assimilated and excreted
• Eggs mature producing oocyte-induced host-seeking inhibition, which gradually develops and then fades
• Mature eggs induce preovipostion behavior, leading to oviposition
Physiology Physiology of of Gonotrophic Gonotrophic CycleCycle
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Other factors influencing host seeking…Other factors influencing host seeking… Host defensive behavior Host defensive behavior Mosquito age – older mosquitoes more likely to Mosquito age – older mosquitoes more likely to
seek blood even when gravidseek blood even when gravid Larval nutrition – if poor, blood may go to support Larval nutrition – if poor, blood may go to support
adult metabolismadult metabolism Mating status – unmated less likely to host seekMating status – unmated less likely to host seek Nutritional status of male with which female Nutritional status of male with which female
mated – poor nutrition in male results in more mated – poor nutrition in male results in more host seekinghost seeking
Mosquito species – some, such as Mosquito species – some, such as An. gambiaeAn. gambiae, , host seek every 24 hrs. until replete (even if host seek every 24 hrs. until replete (even if gravid!)gravid!)
All of these factors potentially contribute to All of these factors potentially contribute to multiple bloodmeals per gonotrophic cycle, multiple bloodmeals per gonotrophic cycle, increasing the potential for malaria transmissionincreasing the potential for malaria transmission
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Malaria Parasite-Vector Malaria Parasite-Vector Relationships Relationships
Malaria Transmission CycleMalaria Transmission Cycle
Parasite Infection SpecificityParasite Infection Specificity
Mosquito Immune DefensesMosquito Immune Defenses
ookinete
oocyst
gametocytes
macrogametocyte
microgametocyte
zygotesalivary glands
oocyst with sporozoites
sporozoites
salivary glands
sporozoites
midgut infected with oocysts
cross section of oocyst Plasmodium Development in Anopheles
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Alimentary Canal
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Alimentary CanalAlimentary Canal Within the alimentary canal, the malaria parasite encounters Within the alimentary canal, the malaria parasite encounters
various structural and physiological/ biochemical characteristics various structural and physiological/ biochemical characteristics that can influence its survivalthat can influence its survival
The noncellular (chitinous) peritrophic membrane (PM) can be an The noncellular (chitinous) peritrophic membrane (PM) can be an effective physical barrier, preventing midgut infectioneffective physical barrier, preventing midgut infection
Vector specificity for malaria pathogens may be linked to the Vector specificity for malaria pathogens may be linked to the rate of PM formation versus the rate of ookinete production in rate of PM formation versus the rate of ookinete production in bloodmealbloodmeal
Adult mosquitoes secrete PM1, while larvae secret PM2Adult mosquitoes secrete PM1, while larvae secret PM2 PM1 secretion is triggered by dramatic extension of the midgut PM1 secretion is triggered by dramatic extension of the midgut
epithelium during ingestion of a bloodmealepithelium during ingestion of a bloodmeal
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Alimentary CanalAlimentary Canal After ingestion, the gametocytes go through a After ingestion, the gametocytes go through a
complete sexual cycle in the midgut lumen and complete sexual cycle in the midgut lumen and develop into motile ookintes (~16-24 hrs)develop into motile ookintes (~16-24 hrs)
Invasion of gut epithelilal cells occurs about 30 Invasion of gut epithelilal cells occurs about 30 hrs after bloodmealhrs after bloodmeal
In In P. gallinaceum P. gallinaceum / / Ae. aegypti Ae. aegypti , , Plasmodium Plasmodium secretes a chitinase in order to penetrate the PM secretes a chitinase in order to penetrate the PM (inhibiting chitinase blocks transmission). (inhibiting chitinase blocks transmission). Trypsin, secreted by the mosquito, activates Trypsin, secreted by the mosquito, activates parasite chitinase.parasite chitinase.
This system may vary in different mosquitoes – This system may vary in different mosquitoes – PM formation in PM formation in An. stephensiAn. stephensi variably detected variably detected
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Bloodmeal processing - stepsBloodmeal processing - steps
1.1. Removal of excess water from the Removal of excess water from the bloodmealbloodmeal
2.2. Breakdown of vertebrate blood cells Breakdown of vertebrate blood cells (hemolysis)(hemolysis)
3.3. Hydrolytic degradation of macromolecules Hydrolytic degradation of macromolecules in the bloodmeal (digestion)in the bloodmeal (digestion)
4.4. Absorption of small molecules into the Absorption of small molecules into the midgut epithelial cells and subsequently into midgut epithelial cells and subsequently into the hemocoelthe hemocoel
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Hemolysis of BloodmealHemolysis of Bloodmeal
Hemolysis breaks down cells to release proteins Hemolysis breaks down cells to release proteins and other nutrients, making them accessible to and other nutrients, making them accessible to the digestive enzymesthe digestive enzymes
Hemolysis may be achieved mechanically Hemolysis may be achieved mechanically (cibarial armature) or biochemically (hemolytic (cibarial armature) or biochemically (hemolytic factors including small peptides and free fatty factors including small peptides and free fatty acids)acids)
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Absorption of Bloodmeal NutrientsAbsorption of Bloodmeal Nutrients
Differences between insects that show continuous Differences between insects that show continuous digestion (eg, tsetse flies -- absorption occurs through digestion (eg, tsetse flies -- absorption occurs through specialized cells) vs those that show batch digestion specialized cells) vs those that show batch digestion (eg, mosquitoes -- same cells that secrete enzymes (eg, mosquitoes -- same cells that secrete enzymes also carry out absorption)also carry out absorption)
Processes range from simple diffusion (eg, absorption Processes range from simple diffusion (eg, absorption of sugar into the hemolymph) to active transport of sugar into the hemolymph) to active transport (amino acids); little is known about absorption of other (amino acids); little is known about absorption of other molecules like lipids, vitamins, and mineralsmolecules like lipids, vitamins, and minerals
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Peritrophic Matrix (PM) Peritrophic Matrix (PM)
The peritrophic matrix is a layer of acellular material The peritrophic matrix is a layer of acellular material separating ingested food from epithelial cellsseparating ingested food from epithelial cells
““peritrophic” comes from the Greek word peri for peritrophic” comes from the Greek word peri for around; trophic is the Greek word for food. The PM around; trophic is the Greek word for food. The PM surrounds the food bolus.surrounds the food bolus.
Peritrophic membrane was termed >100 years ago Peritrophic membrane was termed >100 years ago but membrane implies lipid bilayer. The PM is not -- but membrane implies lipid bilayer. The PM is not -- it is a sheath of cheesy material of amorphous it is a sheath of cheesy material of amorphous appearance. The word matrix is more suitable!appearance. The word matrix is more suitable!
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Other important points -- PMOther important points -- PM
The signal that activates PM secretion is the The signal that activates PM secretion is the physical distention of the midgut epithelium; eg, physical distention of the midgut epithelium; eg, ingestion of partial bm does not trigger PM formationingestion of partial bm does not trigger PM formation
Mosquitoes, blackflies, and sandflies secrete Mosquitoes, blackflies, and sandflies secrete different type of PM during larval lifedifferent type of PM during larval life
PM is permeable to digestive enzymesPM is permeable to digestive enzymes Possible barrier to pathogen infectionPossible barrier to pathogen infection
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Structure of salivary glandsStructure of salivary glands
Structure varies among insect phylaStructure varies among insect phyla In mosquitoes, salivary glands of both sexes are In mosquitoes, salivary glands of both sexes are
paired organs located in the thorax, and each paired organs located in the thorax, and each gland consists of 3 lobes connected to a main gland consists of 3 lobes connected to a main salivary gland duct (male sg’s small)salivary gland duct (male sg’s small)
Female sg’s have two identical lateral lobes and Female sg’s have two identical lateral lobes and one shorter medium lobe. Lateral lobes can be one shorter medium lobe. Lateral lobes can be divided according to proximal and distal regions divided according to proximal and distal regions (different regions secrete different proteins)(different regions secrete different proteins)
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Function of the salivary glandsFunction of the salivary glands
Saliva contains enzymes that digest Saliva contains enzymes that digest sugarssugars
Salivary gland secretions play a role in the Salivary gland secretions play a role in the maintenance of feeding mouthparts - maintenance of feeding mouthparts - saliva acts as a lubricantsaliva acts as a lubricant
In ticks, water in ingested blood is cycled In ticks, water in ingested blood is cycled back through the sg’s where it is returned back through the sg’s where it is returned to the host to the host
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Salivary Glands and Salivary Glands and BloodfeedingBloodfeeding
Salivary glands produce a saliva that facilitates Salivary glands produce a saliva that facilitates rapid and efficient feeding (hemagglutinin, rapid and efficient feeding (hemagglutinin, anticoagulant, antiplatelet activity, vasodilators)anticoagulant, antiplatelet activity, vasodilators)
Parasites can increase the probability of their Parasites can increase the probability of their transmission by modifying arthropod salivary transmission by modifying arthropod salivary activitiesactivities
Malaria sporozoites infect the female-specific Malaria sporozoites infect the female-specific salivary gland lobes (distal-lateral and medial)salivary gland lobes (distal-lateral and medial)
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Salivary Glands and Bloodfeeding -Salivary Glands and Bloodfeeding -22
Parasite invasion causes cellular damage in the Parasite invasion causes cellular damage in the glands – 4-5x reduction in apyrase activityglands – 4-5x reduction in apyrase activity
The salivary apyrases of blood-feeding The salivary apyrases of blood-feeding arthropods are nucleotide hydrolysing enzymes arthropods are nucleotide hydrolysing enzymes and have been implicated in the inhibition of host and have been implicated in the inhibition of host platelet aggregation through the hydrolysis of platelet aggregation through the hydrolysis of extracellular ADP.extracellular ADP.
Sporozoite-infected mosquitoes take longer to Sporozoite-infected mosquitoes take longer to probe – more sporozoites releasedprobe – more sporozoites released
Also, more interrupted feedings – bite more Also, more interrupted feedings – bite more frequently before achieving successful bloodmealfrequently before achieving successful bloodmeal
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Immune responses of vectorsImmune responses of vectors
Arthropod immune responses are not like Arthropod immune responses are not like vertebrate antigen-antibody reactions but vertebrate antigen-antibody reactions but the internal defense mechanisms are still the internal defense mechanisms are still specific and effective in destroying specific and effective in destroying pathogens and parasites.pathogens and parasites.
Much of what we know comes from Much of what we know comes from immune studies of lepidopteran larvae.immune studies of lepidopteran larvae.
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Cuticular and gut barriersCuticular and gut barriers
The arthropods possess a rigid cuticle that The arthropods possess a rigid cuticle that functions as a barrier to potential pathogens. functions as a barrier to potential pathogens. Microorganisms do not penetrate the Microorganisms do not penetrate the exoskeleton unless there is a wound.exoskeleton unless there is a wound.
Many potential pathogens are ingested. Many potential pathogens are ingested. Some are passed on through the feces or Some are passed on through the feces or through regurgitation. Some are walled off through regurgitation. Some are walled off by the peritrophic matrix (barrier?).by the peritrophic matrix (barrier?).
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Possible outcomes following exposure Possible outcomes following exposure of an arthropod to a parasiteof an arthropod to a parasite
susceptible arthropodsusceptible arthropod: the parasite receives : the parasite receives appropriate stimuli from the biochemical appropriate stimuli from the biochemical environment and develops successfullyenvironment and develops successfully
resistant arthropodresistant arthropod: some or all of the parasites are : some or all of the parasites are recognized as foreign by the cellular/humoral recognized as foreign by the cellular/humoral components in the hemolymph, and the arthropod components in the hemolymph, and the arthropod immune response sequesters and destroys parasiteimmune response sequesters and destroys parasite
refractory arthropodrefractory arthropod: the parasites do not elicit an : the parasites do not elicit an immune response but they fail to develop due to immune response but they fail to develop due to physiological or biochemical incompatabilityphysiological or biochemical incompatability
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Cellular immunity in insectsCellular immunity in insects
PhagocytosisPhagocytosis. In mosquitoes, . In mosquitoes, phagocytosis activity is a function of the phagocytosis activity is a function of the numbers of hemocytes presentnumbers of hemocytes present
EncapsulationEncapsulation. The main defense . The main defense mechanism of insects against invaders too mechanism of insects against invaders too large to be phagocytosed is encapsulation. large to be phagocytosed is encapsulation. Phenol oxidase enzymes are involved in Phenol oxidase enzymes are involved in melanotic encapsulation of parasites melanotic encapsulation of parasites (worms and malaria parasites)(worms and malaria parasites)
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SummarySummary
• Anopheles gambiaeAnopheles gambiae is well adapted to take is well adapted to take advantage of temporary aquatic habitat advantage of temporary aquatic habitat associated with human activities (farming, associated with human activities (farming, construction, etc.)construction, etc.)
• Behaviors such as preferential feeding on Behaviors such as preferential feeding on humans and resting in homes keep it closely humans and resting in homes keep it closely associated with us.associated with us.
• The association between The association between Anopheles Anopheles mosquito and mosquito and Plasmodium Plasmodium parasite is controlled by a series of parasite is controlled by a series of physical, physiological and biochemical physical, physiological and biochemical interactions, which may lead to a successful interactions, which may lead to a successful infection followed by transmission to a new host.infection followed by transmission to a new host.
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Additional Reading for More Detail:Additional Reading for More Detail:
Biology of Biology of AnophelesAnopheles mosquitoes – general mosquitoes – generalMedical Entomology for Students, 4Medical Entomology for Students, 4thth Edition – pp. 33-51 Edition – pp. 33-51
Biology of Biology of Anopheles gambiaeAnopheles gambiae mosquitoes mosquitoes Biology of Disease Vectors, 1Biology of Disease Vectors, 1stst Edition – pp. 75-77 Edition – pp. 75-77
Host seeking behavior in mosquitoes – generalHost seeking behavior in mosquitoes – generalBiology of Disease Vectors, 2Biology of Disease Vectors, 2ndnd Edition (BODV) – pp. 277- Edition (BODV) – pp. 277-
287287
Midgut structure and Peritrophic MatrixMidgut structure and Peritrophic MatrixBODV – pp. 289-310BODV – pp. 289-310
Bloodmeal Processing, Egg Development and Osmotic RegulationBloodmeal Processing, Egg Development and Osmotic Regulation BODV – pp. 329-362BODV – pp. 329-362
Immune Response in VectorsImmune Response in Vectors BODV – pp. 363-376BODV – pp. 363-376
Salivary Glands and Saliva in Bloodfeeding InsectsSalivary Glands and Saliva in Bloodfeeding Insects BODV – pp. 377-386BODV – pp. 377-386
