PATHOLOGY OF MALARIA:
Malaria in humans is caused by five related protozoan (single-celled) parasites: Plasmodium falciparum, P. vivax, P. ovale, P. malariae, and P. knowlesi. The most common worldwide is P. vivax. The deadliest is P. falciparum. In 2008 P. knowlesi, which was thought to infect primarily Old World monkeys and to occur only rarely in humans, was identified as a major cause of malaria in humans in Southeast Asia, accounting for as many as 70 percent of cases in some areas. P. knowlesi was found to be easily confused with P. malariae during microscopic examination, resulting in many cases being attributed to P. malariae when in fact they may have been caused by P. knowlesi.
Plasmodium parasites are spread by the bite of infected female Anopheles mosquitoes, which feed on human blood in order to nourish their own eggs. While taking its meal (usually between dusk and dawn), an infected mosquito injects immature forms of the parasite, called sporozoites, into the person’s bloodstream. The sporozoites are carried by the blood to the liver, where they mature into forms known as schizonts. Over the next one to two weeks each schizont multiplies into thousands of other forms known as merozoites. The merozoites break out of the liver and reenter the bloodstream, where they invade red blood cells, grow and divide further, and destroy the blood cells in the process. The interval between invasion of a blood cell and rupture of that cell by the next generation of merozoites is about 48 hours for P. falciparum, P. vivax, and P. ovale. In P. malariae the cycle is 72 hours long. P. knowlesi has the shortest life cycle—24 hours—of the known human Plasmodium pathogens, and thus parasites rupture daily from infected blood cells.
Testing is performed to help diagnose malaria, to monitor for relapses, and to determine drug susceptibility of the parasite causing the infection.
THICK AND THIN BLOOD SMEARS
Diagnosis of malaria involves performing blood smears. For a blood smear, a drop of blood is applied to and spread onto a glass slide. It is then treated with a special stain and examined under a microscope for the morphology of infected blood cells and the parasite. Typically, two thick smears and two thin smears are prepared. These tests are currently the “gold standard” for malaria detection and identification. They require examination by a trained and experienced laboratorian.
The number of malaria parasites present in the blood at a given time fluctuates. Therefore, if no parasites are seen on the initial set of smears and the health practitioner still suspects malaria, then additional blood samples will be obtained to be tested. The samples may be collected at 8 to 12 hour intervals over 2 to 3 days to increase the probability of detecting the parasites. It is advantageous if the sample collection coincides with the appearance of signs and symptoms as this is the time that the parasites will most likely be detected in the blood.
Thick smears are a more sensitive test for malaria infection. A greater volume of blood is examined under the microscope and the parasites are therefore more likely to be seen. Thin smears have fewer blood cells present and allow identification of the type of Plasmodium species causing the infection. The number of infected red blood cells can also be calculated to determine the degree to which a person is infected (parasite load). This information is essential for proper treatment.
RAPID DIAGNOSTIC TESTS (ANTIGEN TESTING)
When microscopy is not readily available, rapid diagnostic tests may be used instead of blood smears. These tests detect malaria antigens (proteins) in a sample of a person’s blood (usually taken with a fingerstick) and indicate a positive result by a color change on the testing strip. They are sometimes called “dipstick” tests.
Different rapid diagnostic tests are available, and they have varying capabilities in what they detect. For example, some rapid tests may detect all four common species (P. falciparum, P. vivax, P. ovale, P. malariae) but do not distinguish between them. Others are combination tests that can detect all four common species and will identify P. falciparum specifically if it is present. The type of rapid test used is dependent on the patient population and the goals of providing a rapid test result.
The U.S. Food and Drug Administration (FDA) has approved a rapid diagnostic test for malaria. It is approved for use by hospital and reference laboratories, but not for doctors’ offices or home testing. This rapid test may allow for faster diagnosis and treatment. However, it is recommended that positive results be followed with blood smears for confirmation and to determine the extent of infection.
MOLECULAR TESTS (POLYMERASE CHAIN REACTION, PCR)
The polymerase chain reaction is a laboratory method that amplifies the parasite’s DNA and allows detection and identification of the Plasmodium species. This test can be used to confirm the diagnosis in laboratories where there is a lack of training and experience in the microscopic examination for malaria. It can also be used to determine the Plasmodium species if the results of a blood smear are unclear. Likewise, it is useful for cases in which the number of malaria parasites in the blood is low or when there are different types causing the infection (mixed) and examination using a microscope may be less accurate. The cost of these molecular testing techniques limits their use in many regions where malaria is endemic.
ANTIBODY TESTS (SEROLOGY)
Serology tests detect antibodies in the blood that are produced by the body in response to a malaria infection. They cannot diagnose an acute infection but help determine if a person was previously exposed. These tests are not routinely used in the U.S. since a diagnosis can be made sooner by detecting the parasite under the microscope or its DNA instead of waiting for an immune response to develop weeks later.
Some malarial parasites have become resistant to the drugs commonly used to treat the infections. Some specialized laboratories can test the parasites from an infected person to determine their drug susceptibility. This can be done either by growing the parasites in the presence of increasing amounts of the drug and observing the effect of the drug on the parasite or by testing the DNA of the parasite to detect markers that indicate resistance. This latter method is still being evaluated.