Plasmodium malaria in the blood
Plasmodium in the blood smear in healthy people is absent. Malarial plasmodia alternately parasitize in 2 hosts: in the body of a female mosquito of the genus Anopheles, where sexual reproduction occurs, sporogony, and in the human body where asexual reproduction occurs, schizogony. The initial phase of schizogony occurs in hepatocytes( extra-eritritic-schizogony), and the subsequent phase in erythrocytes( erythrocytic schizogonia).Developing in erythrocytes, plasmodia feed on Hb and destroy the affected erythrocytes. All pathological manifestations of malaria [fever attacks, anemia, splenomegaly, central nervous system( CNS) damage in the tropical form of malaria] are associated with erythrocytic schizogony.
Parasitological diagnosis of malaria is based on the detection of asexual and sexual forms of the pathogen in a microscopic study of blood, which is possible only during its development in the red blood cell. To detect plasmodia and determine their type, blood preparations prepared by the method of "thin smear" and "thick drop", painted according to Romanovsky-Giemsa, are used. Both methods, which have their advantages and disadvantages, are complementary. Detection in the blood smear of
or a thick drop of any stages of the plasmodium( even 1 parasite) developing in red blood cells( trophozoites - young and adult, schizonts - immature and mature, and sexual forms of gametocytes - male and female) is the only indisputable proof of malaria. It should be borne in mind that the volume of blood in a thick drop is 20-40 times higher than in a thin smear, so a positive response can be given even after smear examination, and negative - only after the study of a thick drop with an immersion objective for at least 5 min, with viewing at least 100 fields of vision( WHO standard) [Lysenko A.Ya., Krasilnikov AA, 1999].The sensitivity of the "thick drop" method is such that when viewing 100-150 fields of vision, approximately 8 parasites per 1 μl of blood can be detected. Care must be taken to detect a single formation resembling ring-shaped trophozoite in a thick drop, since the appearance of this stage of the parasite can be simulated by various artifacts. If it is not possible to detect plasmodia in the presence of a suspected malaria in a single study, it is sometimes necessary to conduct multiple studies( in tropical malaria, blood smears should be taken every 6 hours throughout the attack).There are 4 types of plasmodia.
■ P. falciparum is the causative agent of tropical fever, the most dangerous form of malaria, which requires urgent treatment. In P. falciparum erythrocytic schizogony begins in the peripheral bloodstream, and ends in the central, due to the delay of the affected erythrocytes in the capillaries of internal organs. As a result, at the onset of infection, only young trophozoites( "rings") are present in blood preparations. The gametocytes after maturation in the capillaries of internal organs are found in the peripheral blood on the 10th -12th day of the disease. Detection in the peripheral blood of adult trophozoites or schizonts of any age indicates the onset of a malignant course of tropical malaria and a near fatal outcome, unless urgent measures are taken. In other types of malaria, erythrocytic schizogony flows entirely in the peripheral blood stream. The gametocytes of P. falciparum, in contrast to other species of plasmodia, are not round but oblong in form and differ in a long period of life. They die within 2-6 weeks( other types - for 1-3 days), so the detection of gametocytes P. falciparum for many days after curing the patient( cessation of erythrocytic schizogony) due to the action of schizoticidal drugs is a common phenomenon that is not considered an indicatorinefficiency of therapy.
■ P. vivax is the causative agent of a three-day malaria.
■ P. malariae - causative agent of four-day malaria
■ P. ovale - causative agent of malaria oval( type of three-day).
The cycle of erythrocytic schizogony is repeated in P. falciparum, P. vivax and P. ovale every 48 hours, in P. malariae - 72 hours. Malarial attacks develop in that phase of the cycle of erythrocytic schizogony, when the bulk of the infected red blood cells is destroyed and the daughterindividuals of plasmodia( merozoites) invade intact erythrocytes.
To establish the species belonging to malarial parasites, the following matters: the presence of polymorphism of the age stages or one leading, their combination with gametocytes;morphology of different age stages, their size in relation to the affected erythrocyte;nature, size of the nucleus and cytoplasm;intensity of pigment, its shape, size of grains / granules;the number of merozoites in mature schizonts, their size and location with respect to the accumulation of pigment;the propensity of the parasite to attack the erythrocytes of a certain age( tropism);propensity to multiple lesions of individual red blood cells by several parasite individuals and its intensity;the size of the affected erythrocytes in relation to the uninjured, the shape of the affected erythrocytes, the presence of azurophilic granularity in the affected erythrocytes;form of gametocytes.
In acute attacks of malaria, there is a certain pattern of blood changes. During the chill appears neutrophilic leukocytosis with a shift to the left. During the period of fever, the number of leukocytes decreases somewhat. When sweat appears and with apyrexia, monocytosis builds up. Later, after 2-4 attacks, anemia develops, which is especially early and rapidly developing with tropical fever. Anemia is mainly hemolytic in nature and is accompanied by an increase in the content of reticulocytes. In blood smears, there are poikilo-cytosis, anisocytosis, and polychromatophilia of erythrocytes. With the addition of bone marrow depression, the amount of reticulocytes decreases. Sometimes there is a picture of pernicious anemia. ESR in malaria is significantly increased.
In the interictal( febrile) period in the blood for all forms of malaria, in addition to the tropical, adult trophozoites predominate. During this period of the disease, certain stages of the plasmodia are present in the blood constantly, up to the complete cessation of erythrocytic schizogony. In this regard, there is no need to take blood for research only at the height of a malarial attack, but you can examine it at any time. The absence of plasmodium of malaria in blood smears and a thick drop of a patient with malaria reflects only the thoroughness of the research conducted and the professional competence of the laboratory specialist.
When assessing the intensity of parasitemia, the total asexual and sexual forms are taken into account, with the exception of P. falciparum. The intensity of the parasite is estimated by a "thick drop" in the count of 1 μl of blood. Counts the number of parasites in relation to a certain number of leukocytes. When 10 or more parasites are detected for 200 white blood cells, the count is over. If 9 or less parasites are detected per 200 leukocytes, counting is continued to determine the number of parasites per 500 leukocytes. When single parasites are detected in a "thick drop" of blood, their number per 1000 leukocytes is counted. Determination of the number of parasites in 1 μl of blood is carried out according to the following formula: X = A ×( B / C), where: X - number of parasites in 1 μl of blood;A - counted number of parasites;B - the number of leukocytes in 1 μl of blood;C - counted number of leukocytes.
In cases where it is not possible to determine the number of leukocytes in a given patient, their number in 1 μl is conventionally taken as 8,000 according to the WHO recommendation.
Monitoring of the effectiveness of treatment is carried out by examining a thick drop of blood with counting parasites in 1 μl of blood. The study should be performed daily from the 1st to the 7th day from the beginning of chemotherapy. With the disappearance of parasites during this period, further blood tests are performed on the 14th, 21st and 28th days from the start of treatment. If resistance is detected( assessed by the level of parasitemia) and accordingly ineffective treatment, the antimalarial drug is replaced by a specific drug of another group and the blood test is carried out according to the same scheme [WHO recommendations, 1994].
For patients who have suffered tropical malaria, establish a follow-up for 1-2 months, with a parasitological examination of blood at intervals of 1-2 weeks. Clinical examination of patients with malaria caused by P. vivax, P. ovale and P. malariae should be performed within 2 years. With any increase in body temperature, these individuals need a laboratory blood test to detect malarial plasmodia.