Insectary
- Insectary
- Insect Repellant Evaluation
- Insecticide Evaluation
- Molecular Epidemiology and Species Indentification
INSECTARY
Access to mosquitoes with known pedigrees is vital to many research projects in the MRP and also to malaria control programmes. Establishing and maintaining such colonies of mosquitoes is the primary objective of the insectary.
Personnel:
Mr Ashok Saikoolal, a qualified animal laboratory technologist, manages the facility, assisted by Mr Joseph Shozi and Mr Mzwakhe Roald Cibane.
Acquiring the colonies:
Some of our colonies have been donated by other entomologists and are
already adapted to colony life. It is more difficult to catch
sufficient numbers of local wild mosquitoes, confirm that they are all
the same species ( see species
identification ) and then facilitate their adaptation, step
by step, to living and breeding in the insectary. We attempt to recreate
some of the essential conditions of the mosquito's natural environment:
high temperature and high humidity are maintained by air conditioners;
lights are provided with dimmers on a timer control to create an artificial
dusk and dawn (essential for successful mating). Adaptations to new food sources
are very important: adults must adapt to feeding on anaesthetised guinea
pigs, and the larvae to finely ground commercial cat pellets.
Maintaining the colonies:
A very high standard of cleanliness, careful regulation of larval
feeding (frequency and quantity), regulation of numbers of larvae per
tray, and careful egg collection are all important. Great care also
has to be taken to prevent cross-contamination of the species, many
of which will interbreed in these conditions.
Invading ants must also be kept at bay. They can decimate a mosquito colony virtually overnight.
Species kept:
- Anopheles arabiensis strain KGB
- Anopheles arabiensis strain Mamfene
- Anopheles arabiensis strain Dondotha
- Anopheles merus
- Anopheles gambiae sensu stricto
- Aedes aegypti
- Cimex lectularius
INSECT REPELLENT EVALUATION
With the spread of drug resistance and its impact
on chemoprophylaxis of malaria, the WHO have placed new emphasis on personal
protection against mosquito bites.
Field tests vs Laboratory tests?
Field tests are notoriously difficult to conduct and are subject to the vagaries of the weather, insect populations etc. While laboratory tests cannot replace field tests, they represent an effective way for preliminary testing of candidate repellents or different formulations of repellents, as they can be conducted under carefully controlled conditions with known populations of insects all at the same state of physiological development. In addition the human volunteers are not exposed to the risk of possible infection by arboviruses.
The process:
Controlled application of repellent to forearms
Bioassays: At regular intervals after the application
of repellent, treated forearms are exposed to thirty unfed female
mosquitoes for a two minute period. The mosquitoes are contained in a cage
with a gauze top and a transparent bottom. The cage is applied
to the arm gauze side down and mosquitoes bite through the gauze and
can be observed and counted through the transparent bottom. Using
the data thus generated statisticians can calculate protection times. 
Repellent trivia:
- The most widely used and most effective active ingredient is DEET
- Some alarm has been generated by reports of DEET's neurotoxicity, but compare the daily usage (estimated at millions of applications/day) with the known number of clinically reported cases of neurotoxicity (14 to date).
- Some people are more attractive to mosquitoes than others: skin temperature and personal odours play a role.
- Eating garlic or vitamin B1 won't protect you against mosquito bites.
- Beware of sonic or ultrasonic devices - none of those that have been tested work!
INSECTICIDE EVALUATION
Malaria control programmes direct efforts at two different levels: vector control and patient detection. Vector control in South Africa depends on spraying insecticide onto internal surfaces of houses in the malarious areas. Until 1995 DDT was used, but this has now been replaced by synthetic pyrethroids, which are effective against mosquitoes but more environmentally friendly.
But which one to use? There are many different synthetic pyrethroids on the market, and we know that different house construction materials interact with the insecticides in different ways.
Laboratory evaluation: This is a rapid way to test a number of different insecticides and formulations at different application rates on typical surfaces (various mud daubs, wood, cement and paint) without the expense and risk to human population of a field trial. Once suitable candidates have been identified field trials follow.
Target preparation: Mud, cement plaster formed into discs.
Insecticide application: precise doses of insecticide sprayed onto targets with Potter's Spray Tower.
Bioassay: Insectary supplies large numbers of female malaria mosquitoes ( Anopheles arabiensis ) which are exposed to the treated surfaces by placing them in conical cages attached to the targets, and recording number of deaths occurring within a 24hr period after one hour of exposure. These assays are repeated at regular intervals for up to 1 year and give a reliable indication of the insecticide's effective life.
MOLECULAR EPEDEMIOLOGY AND SPECIES IDENTIFICATION
Molecular techniques have become very important in identifying
species and genetically characterising populations. For example
some of the vectors of malaria in Africa belong to a species complex, the
members of which cannot be identified on morphological characteristics.
Predominant among the molecular techniques is the Polymerase Chain Reaction (PCR).
This technique (which earned its inventors the Nobel Prize) is highly sensitive and specific and operates by repeatedly duplicating short sections of DNA characteristic of a particular species, producing enough of the amplified product to be visible after gel electrophoresis.
PCR's used
- Anopheles gambiae sensu lato: four of the species of this complex can be identified after a single reaction sequence.
- Plasmodium species infecting man: the four species infecting man can be identified in a nested PCR reaction (sequential reamplification of a primary fragment).
- Point mutations in the Plasmodium genome associated with drug resistance: a PCR amplification of the pfmdr gene, if cut with the restriction endonuclease Nsp1 indicates the presence of a mutation thought to be associated with chloroquine resistance. Likewise a preliminary amplification of the dihydrofolate reductase gene can be reamplified with different primers to reveal point mutations associated with resistance to the drug pyrimethamine.