Cerebral malaria or Plasmodium falciparum malaria with hypoglycaemia (original) (raw)
Letters to the Editor
Immunochromatographic test for malaria diagnosis
Site-The identification of histidine-rich protein-2 (HRP-2), synthesised by the blood stages of Plasmodium falciparum, has led to the isolation of monoclonal and polyclonal antibodies against Pf HRP-2’ and the development of an antigen-capture enzyme-linked immunosorbent assay (ELISA) 3{ }^{3} and dipstick assay. 3,4{ }^{3,4} We present our findings with a simpler and less expensive immunochromatographic test (ICT) for the rapid diagnosis of falciparum malaria.
The ICT Malaria Pf test card consists of a cardboard device with opposable faces (figure). One face has a test strip containing two antibodies specific for Pf HRP-2 antigen. One of the antibodies is conjugated to visible colloidal gold particles and impregnated into a sample pad at the base of the test strip while the second antibody is immobilised in a line above the sample pad. 10μ L10 \mu \mathrm{~L} of whole blood is added to the sample pad where lysis occurs, and any Pf HRP-2 antigen present binds to the colloidal gold-labelled antibody. On adding the running buffer to the sample pad, the blood and labelled antibody migrate up the test strip crossing the second antibody line. Blood is cleared from the membrane strip by saturating a pad on the opposing face of the ICT device with buffer, folding the card, and allowing the buffer to push the blood down the membrane. The result is then read against a clear background through the viewing window. In a positive sample, Pf HRP-2, complexed with the gold-labelled antibody, is captured by the antibody on the membrane and a pink line forms. In a negative sample, no pink line forms because there is no Pf HRP-2 antigen. A procedural control line above the test line on the membrane confirms that the test was conducted properly. The entire test procedure is completed within 6 minutes.
We evaluated the ICT Malaria Pf test by examining finger-tip specimens of blood from 251 patients with symptoms attending the Central Hospital, Honiara, Solomon Islands. After preparing thick and thin blood films, a small volume of blood was placed in edetic-acid-coated tubes and used for the ICT test within 3 hours of collection. Examination of blood films showed that 38 patients had PP falciparum, 38 had PP vivax, and one had a mixed infection of PP falciparum and PP vivax. Eight samples from patients with PP falciparum infections had a parasite density between
Figure: Photograph of two opposing faces of ICT Malaria Pf test card and sequential procedural steps of test
80 and 500/μL500 / \mu \mathrm{L} and 31 of them exceeded 500 parasites /μL/ \mu \mathrm{L}. The ICT test was performed “blind” without prior knowledge of the microscopic results. The test detected Pf HRP-2 antigen in all 39 samples that showed PP falciparum in the blood films. However, no pink line was observed in the 38 samples that showed only PP vivax in the blood films. It was also negative in 166 of the 174 samples showing no parasites by microscopy. The remaining eight samples had a faint pink line. Since seven of the eight patients had been treated recently with chloroquine or quinine, low levels of circulating antigen persisting after treatment may have been responsible for the discordant results. 3,4{ }^{3,4} It is also possible that parasitised erythrocytes may have been sequestered at the time of sampling, leaving only sub-patent infected cells in the peripheral circulation.
This preliminary field study indicates that the ICT test may prove to be a simple and practical means of making a rapid diagnosis of falciparum malaria at all levels of health care. With a sensitivity of 100%100 \% and a specificity of 96.2%96.2 \% the accuracy of the test is good, at least for parasite densities exceeding 80/μL80 / \mu \mathrm{L}. Further studies are indicated to determine the value of this test under various field conditions.
M García, S Kirimoama, D Marlborough, J Leafasia,
*K H Rieckmann
ICT Diagnostics, Brookvale, NSW, Australia; Solomon Islands Malaria Training and Research Institute, Honiara, Solomon Islands; and *Army Malaria Research Unit, University of Sydney, LMA, NSW 2174, Australia
1 Parra ME, Evans CB, Taylor DW. Identification of Plasmodium falciparum histidine-rich protein 2 in the plasma of humans with malaria. F\mathcal{F} Clin Microbiol 1991; 29: 1629-34.
2 Taylor DW, Voller A. The development and validation of a simple antigen detection ELISA for Plasmodium falciparum malaria. Trans R Soc Trop Med Hyg 1993; 87: 29-31.
3 Schiff CJ, Premji Z, Minias JN. The rapid manual Para Sight ®{ }^{\circledR}-F test. A new diagnostic tool for Plasmodium falciparum infection. Trans R Soc Trop Med Hyg 1993; 87: 646-48.
4 Beadle C, Long GW, Weiss WB, et al. Diagnosis of malaria by detection of Plasmodium falciparum HRP-2 antigen with a rapid dipstick antigen-capture assay. Lancet 1994; 343: 564-68.
Cerebral malaria or Plasmodium falciparum malaria with hypoglycaemia
Site- WHO1\mathrm{WHO}^{1} and Warrell and co-worker’s 2{ }^{2} definition of cerebral malaria in adults requires unarousable coma not attributable to any other cause and lasting at least for 30 min after a generalised convulsion, exclusion of other encephalopathies, and confirmation of acute Plasmodium falciparum infection. Patients falling within this definition show features of a diffuse symmetrical encephalopathy. From September to December, 1994, during an outbreak of malaria in Bikaner, India, we encountered some patients who fulfilled the above definition but may not be cases of cerebral malaria.
A 60-year-old man was admitted with fever for 3 days, generalised convulsions for 8 hours, and unconscious (Glasgow coma scale 5) for 2 hours. Blood film showed heavy PP falciparum parasitaemia. Blood glucose was 2⋅2mmol/L2 \cdot 2 \mathrm{mmol} / \mathrm{L}. Infusion of 200 mL of 25%25 \% glucose was started and he became conscious within 20 min . He was put on oral quinine and intravenous fluids containing 10%10 \% dextrose, and recovered completely within 7 days.
A 17-year-old woman in the second trimester of her first pregnancy had a fever for 4 days. She received paracetamol at
home and on the 5th day she became unconscious after having generalised convulsions. She was admitted to hospital after 3 hours in coma (Glasgow coma scale 4). She was sweating and had a tachycardia. Blood glucose was 2⋅0mmol/L2 \cdot 0 \mathrm{mmol} / \mathrm{L} and a blood film showed asexual forms of PP falciparum. She was given 200 mL of 25%25 \% glucose and recovered from coma within 15 min . Oral quinine was started with oral glucose. Ultrasonography revealed a single viable fetus of 25 weeks. On the 3rd day she aborted the fetus. She recovered without any complication.
These patients probably did not have cerebral malaria, but malaria with hypoglycaemia, and as such they should not be included under the heading of cerebral malaria for research purposes. White et al 1{ }^{1} reported 8%8 \% of adult patients with cerebral malaria having hypoglycaemia. They observed that hypoglycaemia is an important complication of falciparum malaria and may remain unsuspected and untreated because coma and other neurological signs and symptoms are misinterpreted as cerebral malaria. In a study of 131 children with cerebral malaria in Malawi Molyneux et al 4{ }^{4} reported hypoglycaemia (blood glucose <2⋅2mmol/L<2 \cdot 2 \mathrm{mmol} / \mathrm{L} ) in 30 patients at the time of admission. Only one patient showed clinical improvement immediately after treatment with 50%50 \% glucose, and 11%11 \% of survivors had neurological sequelae.
For research purposes the definition of cerebral malaria should include exclusion of hypoglycaemic encephalopathy.
Endocytosis by mature muscle cells of aggregates of parasitised erythrocytes and macrophages in severe malaria
Sir-To test whether vascular sequestration underlies cerebral malaria that complicates some cases of Plasmodium falciparum, 1{ }^{1} we did needle muscle biopsies (vastus lateralis) in 23 adults with clinical malaria consecutively admitted to Maputo Hospital (Mozambique) between April and June, 1995.
Of these cases, 13 had either cerebral malaria or severe malaria but without definite criteria for overt cerebral involvement. In the remaining ten patients, non-complicated malaria due to PP falciparum was diagnosed. 14 muscle samples from healthy patients matched for age and sex served as a control group. The aim of the study was to investigate the expression of endothelial cell adhesion molecules and detection of malarial antigens in muscle specimens. All specimens from patients and controls were snap-frozen for histochemistry (NADH, ATP-ase, COX) and staining (HE, Gomori’s trichrome). In addition, a small fragment of the specimen was glutaraldehyde fixed and plastic embedded for semithin sections. In four of 13 patients with severe malaria but not in the ten remaining patients or in the controls, histological muscle examination showed medium to large intracytoplasmatic inclusions composed of aggregates of erythrocytes, sometimes parasitised, plus a few nucleated cells, in otherwise normal
Figure: Two muscle fibres showing prominent aggregates of red blood cells with nucleated cells (arrow) and malarial prigment (asterisk)
Semithin sections, toluidin blue, stainings ×600\times 600.
or near normal muscle fibres (figure). Such inclusions were variable in number. Immunohistochemistry for laminin and Ulex europeus and CD31 failed to recognise basal lamina or endothelium in or around the aggregate, respectively. By contrast, positive reactions for Mac-1 (CD11b) and ICAM-1 indicated the presence of activated cells of macrophage origin within muscle fibres. Positive reactions with a polyclonal antibody against PP falciparum merozoite surface protein-1 (MSP-1) antigen demonstrated the presence of parasitised erythrocytes. 2{ }^{2}
The lack of reactivity for laminin around the inclusions argues against invagination of basal lamina and in turn the absence of Ulex and CD31 positive material makes an endothelial origin for the vacuole unlikely. The large size of the inclusions in some cases additionally argues against the possibility of internalised capillaries. 3{ }^{3} On the basis of the positive staining for Mac-1 and ICAM-1 we assume that some of the cells are macrophages somehow associated with parasitised erythrocytes within mature muscle fibres, and probably originating by an endocytosis process. Irrespective of the mechanism involved in the development of such structures, it seems they are clearly associated with severe malaria.
*F García, M Cebrián, M Corachán, M Dgedge, J M Grau
Sección de Medicina Tropical.* Servicio de Enfermedades Infecciosas, Unidad de Investigación Musicular, Servicio de Medicina Interna, and Hospital Clínic, 08036 Barcelona, Spain, and Instituto Nacional de Saude, Maputo, Mozambique
- *Dhanpat Kumar Kochar, Banshi Lal Kumawat Department of Medicine, Division of Neurology, Sardar Patel Medical College, Bikanei-224003, India
1 WHO. Severe and complicated malaria. Trans R Soc Trop Med Hyg 1990; 84 (suppl 2): 3.
2 Warrell DA, Looareesuwan S, Warrell MJ, et al. Dexamethasone proves deleterious effect in cerebral malaria: a double blind trial in 100 comatose patients. N Engl 3 Med 1982; 306: 313.
3 White NJ, Warrell DA, Chanthavanich P, et al. Severe hypoglycemia and hyperinsulinaemia in falciparum malaria. N Engl 3 Med 1983; 309: 61-66.
4 Molyneux ME, Taylor TE, Wirima JJ, et al. Clinical features and prognostic indicators in paediatric cerebral malaria: a study of 131 comatose Malawian children. Q3 Med 1989; 265: 441-59. ↩︎ - *F García, M Cebrián, M Corachán, M Dgedge, J M Grau Sección de Medicina Tropical.* Servicio de Enfermedades Infecciosas, Unidad de Investigación Musicular, Servicio de Medicina Interna, and Hospital Clínic, 08036 Barcelona, Spain, and Instituto Nacional de Saude, Maputo, Mozambique
1 Phillips RE, Solomon T. Cerebral malaria in children. Lancet 1990; 336: 1355−601355-60. ↩︎