ANNALS O F CLINICAL AND LABORATORY SCIEN CE, Vol. 15, No. 6Copyright © 1985, Institu te for Clinical Science, Inc.

Schizonts, M erozoites, and Phagocytosis in Falciparum Malaria

TSIEH SUN, and CHHAYA CHAKRABARTI, M .D .t

*Department of Laboratories, North Shore University Hospital,

Manhasset, NY 11030 and

*Department of Pathology, Cornell University Medical College,

New York, NY 10021 and

tDepartment o f Pediatrics, Jamaica Hospital,

Jamaica, NY 11418

ABSTRACT

Two Nigerian siblings, ages 10 and 4 years, respectively, were infected with Plasmodium falciparum and w ere adm itted to the hospital on the same day. The younger child died on the day of admission, bu t the older ch ild su rv ived . The p e rip h e ra l b lood sm ears of the younger p a tie n t showed the ring forms, schizonts, free merozoites, and phagocytosis of malarial parasites by both monocytes and polymorphonuclear leukocytes, whereas the sm ear from the older patient revealed only ring forms. The prognostic significance of this unusual observation and the host factors that affect the survival of the patients are discussed. This is the first docu­m ented case in which phagocytosis of malarial parasites by polymorpho­nuclear leukocytes is observed.

Introduction

Malaria is not a common disease in this country. However, because of a world­wide resurgence and an influx of refu­gees into this country, a further increase in th e in c id en ce of m alaria shou ld be e x p e c te d .11 Two cases are re p o rte d of malaria, owing to Plasmodium fa lc ipa ­rum, in two siblings who contracted the

t Address reprint requests to: Dr. T. Sun, Depart­ment of Laboratories, North Shore University Hos­pital, 300 Community Drive, Manhasset, NY 11030.

infection at the same time. The boy died, and th e girl su rv ived . T he p e rip h e ra l blood smears of the boy showed phago­cytosis of the parasites and the presence of schizonts and free merozoites. These p h en o m e n a are uncom m on and are of prognostic significance.

Case H istory

Patient 1. A 4-year-old black Nigerian boy was admitted to Jamaica Hospital. His chief complaint was high fever, chills, nausea, and vomiting on the day of admission. Two days prior to admission, he had a low fever and headache.

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466 SUN AND CHAKRABARTI

The patient had been in the United States for one year but had travelled back to Nigeria four to five times during that period. His last visit to Nigeria was two weeks prior to admission.

Physical examination on admission showed a tem ­perature of 105°F and respiration rate o f 38 per min­ute. The patient was in acute stress, lethargic, weak and dehydrated but not emaciated (weight 36.5 lbs.). His liver was 3 cm below the right costal margin, and the spleen tip was palpable.

Laboratory tests for electrolytes showed potas­sium, 3.8 m Eq per L; sodium, 129 m Eq per L; C 0 2, 11 mEq per L; BUN, 29 mg per dl; and glucose, 93 mg per dl. Hematologic examination revealed hem o­globin, 9 .8 g per dl and W BC, 3 ,500 per m m 3; including lymphocytes, 54 percent; atypical lympho­cytes 7 percent; neutrophils, 20 percent; band form, 17 percent; monocytes, one percent; and myelocytes, one percent. Platelets were moderately decreased.

The peripheral blood smears were examined by the senior author. M icroscopic examination of the smear showed approximately seven percent infected erythrocytes. Most o f the parasites were early tro­phozoites (ring forms). The rings were small and del­icate, showing frequent double chromatin dots. Some erythrocytes contained two to three ring forms. Infected erythrocytes were normal-sized with no Schüffner’s dots. Although no gam etocytes were detected, the blood picture was consistent with Plas­modium falciparum infection. Unusual features were the presence o f considerable numbers of schizonts, containing approximately 16 to 26 m erozoites and black pigment (figures 1 and 2), and occasional free m erozoites (figure 3). Phagocytosis o f parasites by monocytes (figures 3 and 4) and polymorphonuclear leukocytes (figures 5 and 6) were frequently encoun­tered. Some phagocytized parasites still maintained a rosette pattern (figure 5), while others were partly digested (figure 6).

Initially the patient was given intravenous fluids to correct his dehydration. After the detection of P.

F i g u r e 1. An immature schizont in a peripheral blood smear. Wright-Giemsa ( x 2,000).

F i g u r e 2. A mature schizont showing prominent segmentation and occupying the entire erythrocyte. Wright-Giemsa ( x 2,000).

fa lc ip a ru m in the peripheral blood sm ears, the patient received 80 mg of chloroquine dihydrochlo­ride i.m. four and a half hours after admission and 70 mg one hour later. Within 10 minutes, the patient convulsed and was given 5 mg of Valium, but cardiac arrest ensued. Vigorous resuscitation was unsuccess­ful and he was pronounced dead 30 minutes later. An autopsy request was refused.

Patient 2. A 10-year-old black Nigerian girl, who presented with headache, high fever and vomiting for four days, was admitted to the Jamaica Hospital on the sam e day. The patient had b een seen in another hospital the previous day and was given amoxicillin with no obvious effect.

The patient had “malaria” one year previously and was hospitalized in Nigeria. The date o f her last visit to Nigeria was also two weeks previously, at the same time as her brother’s (Case 1).

On admission, her temperature was 104°F. Physi­cal examination revealed nuchal rigidity and menin­geal signs. Her liver was 3 cm below the right costal margin with right upper quadrant tenderness. The sp leen was not palpable. No scleral icterus was noted.

Laboratory tests show ed norm al e lectro lytes. Hematologic examination revealed hemoglobin 9.9 g per dl; WBC, 6,400 per mm3 with 7 percent lym­phocytes; 4 percent atypical lymphocytes; 5 percent m onocytes; 81 percent polym orphonuclear leuko­cytes, and 3 percent band form. Platelet count was35,000 per mm3. Liver function tests showed slightly elevated liver enzymes. Bilirubin was 1.6 mg per dl. Prothrombin time and partial thromboplastin time were normal. Sickle cell preparation was negative. G lucose-6-phosphate dehydrogenase (G-6-PD) was normal. Cerebrospinal fluid examination and blood culture were also negative. Urinalysis showed 3 + albumin, but no hemoglobinuria was demonstrated.

The peripheral blood smears were examined by the senior author. About 12 percent of the erythro­cytes contained single and double delicate ring forms with the frequent presence of double chromatin dots in rings. Infected erythrocytes were of normal size and contained no Schiiffner’s dots. Gam etocytes, schizonts, and merozoites were not found. No phago­cytosis was demonstrated.

The patient was started initially on intravenous fluids and ampicillin. After malarial parasites were found in the blood smear, she was treated with chlo­roquine dihydrochloride (150 mg) by muscular injec­tion eight hours later. Chloroquine was continued until the fifth hospital day. She was discharged eight days after admission.

Discussion

The simultaneous malarial infection of two siblings with such m arked difference in prognosis provides an opportunity for analysing the host factors in resistance to this disease. As young ch ildren of the same family, their genetic composition,

MORPHOLOGY O F FALCIPARUM MALARIA 467

F i g u r e 3. A peripheral blood smear showing a cluster o f free merozoites (arrow head) and a few merozoites (arrow) being phagocytized by a monocyte. Wright-Giemsa (x 2,000).

F i g u r e 4. A cluster of malarial parasites residing within a parasitophorous vacuole o f a monocyte. Wright- Giemsa (x 2,000).

nutritional background, and the tim e of exposure to the infection are probably similar. The tim e of treatm ent of both cases was also identical.

The question is why did the boy die and the girl survive? A study of a group of young adults showed no associations betw een age and course of infection,4 but cerebral malaria occurs most frequently in children of pre-school age in Africa.10 A re c e n t s tudy in T hailand , how ever, show ed th a t m ost cases of ce reb ra l malaria occured in adults rather than in children.7 It was found in that study that 65 p e rc e n t of th e c e re b ra l m alaria p a tien ts w ere im m igran ts from n o n ­endem ic areas.7 Obviously, it is the pre­vious exposure rather than age alone that d e te rm in es th e im m une sta tus of the host. In this report, the sister had a his­tory of previous infection, which proba­bly accoun ted for th e re la tiv e ly b e ­

nign course; w hereas the non-im m une b ro th e r rap id ly d ev e lo p ed c e reb ra l malaria. It is also interesting to note that the survivor did not have sickle cell dis­ease and G-6-PD deficiency which might have c o n tr ib u te d to h e r res is tan ce to malaria.2

In m ost cases of falciparum m alaria, the o lder trophozoites, schizonts, and seg m en ters are se q u e s te re d from the peripheral circulation and thus they are seldom seen in th e p e rip h e ra l b lood sm ear.1,5 In the light of u ltrastructu ra l studies, it has been found that the for­m ation of e lec tron-dense excrescences (knobs) on the erythrocytic m em brane is responsib le for th e a ttach m en t of infected red blood cells to th e venous en d o th e liu m of th e in te rn a l organs, re su ltin g in th e ir re te n tio n .3'8 W hile most textbooks emphasize the character­istic p rese n c e of only ring form s and

468 SUN AND CHAKRABARTI

F i g u r e 5. A schizont with an intact rosette pattern is seen inside the cytoplasm of a polymorphonuclear leuocyte. Another polymorphonuclear leukocyte present in the upper field showing no phagocytosis. Wright- Giemsa (X 2,000).

F i g u r e 6 . A p a r t ly d ig e s t e d c lu s t e r o f m a la r ia l p a ra s i t e s r e s id in g in s id e t h e c y to p la sm o f a p o ly m o r p h o ­n u c le a r le u k o c y te . W r ig h t - G ie m s a ( x 2 ,0 0 0 ).

gametocytes in the peripheral blood of p a tien ts in fec ted w ith P. fa lc ip a ru m , Beaver et al point out that the existence of schizonts and other developing forms does not automatically rule out P. fa lc i­parum infection.1 It is rather a sign of grave prognosis because these forms are usually seen in heavily infected or m or­ibund patien ts.1-6 However, the presence of free merozoites as seen in this case is extremely rare since merozoites invade o ther ery throcytes alm ost im m ediately after their being released. In vitro hemo­lysis in an old blood specimen may lead to the ex istence of ex traery th rocy tic merozoites; however, in our cases, blood sm ears w ere m ade im m edia te ly after specim en co llection and thus in v itro hemolysis was unlikely.

T he p rognostic significance of the presence of the schizonts and/or m ero­

zoites in p e rip h e ra l blood sm ears is obvious w hen com paring th ese two cases, because the surv ivor had even higher parasite counts than her brother who d ied . P arasite coun t has been regarded as the most im portant indicator for th e p rognosis of p a tien ts w ith m alaria.4 However, when the developing forms are seen in the peripheral blood of a falciparum malaria case, a poor prog­nosis is indicated even though the para­site count is not very high, as seen in our Patient 1.

A lthough phagocytosis of P. fa lc ip a ­rum was first d o cu m en ted in hum an cases as recently as in 1980,9 it is prob­ably not a very ra re phenom onen in heavily infected cases.* The unique fea­ture in this observation is that the para­

* W. A. Krotoski, personal communication.

M O R PH O L O G Y O F FA LCIPA RU M M ALARIA 469

sites w ere n o t only d e m o n s tra ted in m onocytes b u t also in p o ly m o rp h o n u ­clear leukocytes.

Acknowledgm ent

The authors wish to thank W. A. Krotoski, M .D ., P h.D ., Chief, Tropical Infectious D isease Research Program and Clinical Research D epartm ent, U .S. Public Health Service Hospital, N ew Orleans, LA, for reviewing the blood smears o f the first case, and M arlene S ie g e l, M .S ., M .T. (ASCP) and Mary Brown, M.T. (ASCP) for technical assistance.

References

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2. F r i e d m a n , M. J. and T r a c e r , W .: The biochem­istry of resistance to malaria. Sci. Am. 244:154- 164, 1981.

3. L a n g r e t h , S. G ., J e n s e n , J. B ., R e e s e , R . T . , and T r a c e r , W.: Fine structure of human malaria in vitro. J. Protozool. 25:443-452, 1978.

4. P a z z a g l ia , G. and W o o d w a r d , W . E.: An anal­ysis o f the relationship of host factors to clinical falciparum malaria by multiple regression tech­

niques. Am. J. Trop. M ed. Hyg. 31:202-210,1982.

5. S p e n c e r , H. C. and S t r i c k l a n d , G. T . : Malaria. Hunter’s Tropical M edicine. Strickland, G. T., ed. Philadelphia, W. B. Saunders Co., 1984, pp. 516-552 .

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7. T h a r a v a n ij , S., W a r r e l , M. J., T a n t i v a n i c h , S., T a p c h a i , P . , C h o n g s a - N g u a n , M., P r a s e r t s i r i - r o j , V . , and P e t a r a p o t i k u l , J.: Factors contrib­uting to the developm ent of cerebral malaria. I. Humoral immune responses. Am. J. Trop. Med. Hyg. 33:1 -11 , 1984.

8 . U d e i n y a , I. J., S c h m i d t , J. A . , A i k a w a , M . ,

M i l l e R ; L. H ., and G r e e n , I: F aliciparum malaria-infected erythrocytes specifically bind to cu ltu red hum an e n d o th e lia l c e ils . S c ien ce 213:555-557, 1981.

9. V e r n e s , A. : Phagocytosis o f P. falciparum par­asitised erythrocytes by peripheral monocytes. Lancet ii: 1297-1298, 1980.

10. W o r l d H e a l t h O r g a n i z a t i o n : Developm ents in Malaria Immunology, Tech. Rep. Ser. No. 579, Geneva, World Health Organization, 1975, p.45.

11. W y l e r , D. J.: Malaria— Resurgence, resistance and research (in two parts). New Engl. J. Med. 308:875-878; 308:93 4 -9 4 0 , 1983.