Case Reports/Kasuistiken

G. M a g g i o r e , F. Sacchi , M. M a r c o n i , E . B i anch i , A . D e i Cas , A. G . S iccard i

Functional Defects of Neutrophils in Visceral Larva Migrans Syndrome

Summary. Severe defects of neutrophil function were found in a patient suffering from larva viscera migrans syndrome. Some of the defects (metabolic activation, bactericidal activity) were transient and normalized at remission, whereas a chemotactic defect persisted throughout the observation period.

Zusammenfassung: Funktionsdefekte der Neutrophilen beim viszeralen Larva-Migrans-Syndrorn. Schwere Neu- trophilenfunktionsst0rungen wurden bei einem Patien- ten gefunden, der am Larva-viszera-migrans-Syoxtrom litt. Einige der Defekte (rnetabotische Aktivierung, bakterizide Aktivit~t) waren vortibergehend und nor- malisierten sich mit der Remission, w~rend ~ein chemo- taktischer Defekt ~tber d i e gesamte Beobachtungszeit lain bestehen blieb.

Introduction

Toxocara canis, the common dog ascaris, is the primary etiologic agent of the visceral larva migrans syndrome both in adults (1) and in children (2). The most common route of infection is ingestion of soil containing larvae or embryonated ova. Larvae migrate via lymphatics and portal circulation to various tissues and may produce fever, pulmonary infiltrates, hepatomegaly and eosinophilia. Two distinct clinical forms have been defined, the most common being a visceral form and an ocular form, characterized by severe endophthalmitis. Since the first description in 1950, many case reports on visceral larva migrans have appeared in all parts of the world. We report the case of a child with clinical, biological and serological evidence of visceral larva migrans, associated with functional defects of neutrophil leukocytes.

Case Report

A three year old boy was admitted to the Pavia University Pediatric Clinic with a three week history of cough and wheezing. He had been well until six months of age when an episode of pneumonia occurred. From one year of age he suffered from recurrent episodes of upper respiratory tract infection with cough, wheezing, vomiting, anemia and loss of weight. No systemic or visual symptoms were reported. On admission, physical examination revealed an apparently normal child without cyanosis, dyspnea, or digital clubbing. No hepatomegaly was found, and the lungs were clear.

Laboratory data revealed a white blood cell count of 19,000/cmm with a differential of 34% polymorphonu- clear leukocytes, 38% eosinophils and 27% lymphocytes. Hemoglobin was 11.1 g/dl, hematocrit 33.4%, red blood cells were slightly hypochromic with occasional basophilic stippling. Globulins were 4.22 g/dl with 1.60 g/dl of gam- maglobulins. IgG was normal (140 IU/ml) but IgA (100 IU/ml) and IgM (312 IU/ml) were moderately increased, and IgE (3,300 IU/ml) was markedly increased (normal values 0-40 IU/ml). The blood group was A Rh positive and the anti-B isohemagglutinins titer was 1/128. Delayed-type hypersensitivity skin tests (PPD, Candidin, Streptokinase-Streptodornase) demonstrated normal reactivity, blood T-lymphocytes were 58% (2975/cmm) and the complement level (assayed by CH50 assay on antibody sensitized sheep erythrodytes) was 100%, as compared to a reference pool of sera. Liver function tests were normal. Repeated stool exami- nations did not reveal parasite ova. A history of pica was recorded; the patient had often been found eating "gar- den dirt". A visceral larva migrans syndrome was suspect- ed and confirmed serologically by immunoelectrophoresis by Professor M. Gentilini (H6pital de la Piti6 Salp6tri6re, Paris). No ocular abnormalities were found and the EEG was normal. Starting in June 1979, the patient was treated with thia- bendazole (3), 25 mg/kg body weight, three days a week for two months. A moderate decrease in wheezing-asso- ciated respiratory symptoms was noted. In October 1979 the patient had a recurrence of the original symptoms, eosinophilia and hyper-IgE. He was then treated with mebendazole (two doses in two weeks) and showed com- plete recovery.

Materials and Methods

The neutrophil function tests reported in Table i were perform- ed as described by Sacchi et al. (4). Phagocytosis: The phagocytosis frequency (PHF) is the fraction of neutrophils which have phagocytized yeast particles, the normal value being 0.96 _+ 0.08 (n = 162). The phagocytosis index (PHI) is the average number of intracellular yeast particles per neutrophil, the normal value being 3.28 +_ 1.03 (n= 145). Metabolic activation: P/R, calculated from the production of 14CO 2 from 14Cl-glucose by latex-phagocyting (P) and resting

Received: 14 April 1980 Dr. G. Maggiore, Dr. F. Sacchi, Dr. M. Marconi, Dr. E. Bianchi, Clinica Pediatrica dell' Universith, Dr. A. Dei Cas, Clinica di Malattie Infettive dell' Universitg, Ospedale San Matteo, Piazzale Gotgi 5, 1-27 100 Pavia, Italy; Prof. A. G. Siccardi, Istituto di Genetica, Via S. Epifanio, 1-27 100 Pavia, Italy.

Infection 8 (1980) Nr. 4 171

G. Maggiore et al.: Visceral Larva Migrans and Neutrophil Defects

(R) neutrophils, is a measure of the phagocytosis-dependent stimulation of the hexose monophosphate shunt. The normal value at 30 rain is 5.01+1.07 (n=176). The NBT reduction frequency (NRF) is the fraction of phagocyting neutrophils which show formazan deposits, the normal value being 0.94+0.07 (n= 163). Bactericidal activity: This is evaluated as the percentage of bacteria killed in the assay using Staphylococcus aureus ATCC 6538 (BKS. a.), and Streptococcus faecalis ATCC 8043 (BKS. f.). Normal values are 87.48 + 11.08% (n= 139) and 92.22 + 8.90% (n=88) respectively. Neutrophil motility: This is assessed by means of random mobil- ity, the normal value being 34.49+ 11.98 (n=86); and chemota- xis, the normal value being 98.76+11.14 (n=87). The normal values (manuscript in preparation) were obtained from healthy controls. All variables were normally distributed. The pathological threshold values (PTV) reported in Table 1 are the values corresponding to the lower 97.5% confidence limit.

Table 1: Neutrophil function tests.

Tests a

Phagocytosis Frequency Index

Metabolic activation P/R 15 min

30 min NRF

Bactericidal activity (%) BKS.a. 60 min

90 min BKS.f. 60 min

90 rain

Motility Random Chemotaxis

A B C D ~ PTV

0.80 0.88 0.95 0.93 0.79 1.95 2.30 2.56 1.96 1.26

0.98 3.18 2.30 3.20 1.56 4.06 2.50 6.70 2.92 0.95 0.96 0.81 0.91 0.80

47.0 58.1 23.6 96.6 33.0 75.0 59.0 98.5 65.8

<10.0 31.7 <10.0 93.4 25.0 79.0 <10.0 98.8 74.8

22.0 18.9 21.3 22.3 11.0 33.1 35.3 38.8 36.6 76.8

See Methods During the course of the disease the tests were performed in four periods (A-D, see text). The data are means of at least two repeti- tions (each had two or three replicas). Experimental variations be- tween replicas and repetitions never exceeded 20%. PTV = pathological threshold; 97.5% confidence limit in a number (86-176) of healthy controls.

R e s u l t s

Leukocyte function tests were performed on the patient in four periods during the course of the disease: at first admission, before diagnosis and before therapy (A); after two months of thiabendazole therapy, when a modera te but incomplete improvement was recorded (B); at the second admission, during a full recurrence of symptoms (C); and in remission, after two weeks of mebendazole t rea tment (D). Table 1 reports the data and the control values in use in our laboratory; phagocytosis (frequency and index) was always normal as well as the frequency of NBT-reducing cells; phagocytosis-dependent activation of the hexose

monophosphate shunt and bactericidal activity were defective in periods A and C (symptomatic periods). Chemotaxis was markedly defective in all instances and random mobility was normal. The defect in chemotaxis did not appear to be the result of a serum inhibitor of leukocyte motility, since the t reatment of normal leukocytes with the serum of the patient did not impair their motility; moreover , washing the patient 's cells with normal human serum did not restore chemotactic responsiveness.

D i s c u s s i o n

Leukocyte functions are not usually studied in patients with parasitic disease, and to our knowledge there is no report in the literature of neutrophil function defects in visceral larva migrans or in other human parasitic diseases (5).

The presence of neutrophil function defects in the report- ed case raises a number of problems, especially the pro- blem of whether the defects are secondary to the parasitic infestation, or pr imary and predisposing to the infesta- tion.

Marked defects in bactericidal activity were found in the symptomatic periods, and their association with defective metabolic activation does not seem to have caused the defects, since both S. aureus (catalase-positive) and S. faecalis (catalase-negative) survived in the cells of the patient. If the bactericidal defect was due to lack of superoxide production (as in chronic granulomatous disease), S. faecalis should have been killed effectively. Such functional defects found in the symptomatic periods are similar to those encountered with " tox i c" leukocytes in the course of respiratory virus infections and certain bacterial infections, whereas in the present case these defects disappear on remission. Conversely, the defect in chemotaxis was remarkably stable and persisted at remis- sion. This could be tentatively interpreted as the result of a hyper-IgE syndrome (6); IgE levels are very high during the whole observation period and tend to remain elevated in visceral larva migrans syndrome for a long time after remission (5). Hill et al. (7) have suggested that in certain individuals high IgE levels cause defects in neutrophil chemotaxis by mediating the in vivo exposure of neutro- phils to relatively high histamine concentrations; if nor- mal neutrophils are t reated in vitro with 10 -5 M histamine (7, Sacchi, unpublished data) they show a marked chemo- tactic defect which persists after washing in histamine-free medium. Since not all patients with hyper-IgE show che- motactic defects, the nature of the difference between those who show a hyper-IgE syndrome and those who do not remains to be adcertained. As far as the chemotactic defect of our patient is concerned, one cannot distinguish at this point between an intrinsic cellular motility defect (which could have played a role in the establishment of the visceral larva migrans syndrome) and a hyper-IgE syndrome.

172 Infection 8 (1980) Nr. 4

G. Maggiore et al.: Visceral Larva Migrans and Neu t roph i l Defects

In conclusion, we think that more exhaustive studies on leukocyte functions should be carried out in human para- sitic diseases, and particularly in visceral larva migrans, to clarify the mechanism of parasitic infestation and, eventu- ally, the role of phagocytes in the pathogenesis of parasi- tic diseases.

Acknowledgements

The authors are indebted to Prof. G. R. Burgio for his constant interest and support, to Dr. A. G. Ugazio for reading the manuscript and to Prof. M. Gentilini for his help. F. Sacchi was the recipient of a fellowship from the S. P. A., Milan. The study was partially supported by a CNR grant to A. G. SiccardL

Literature

1. Shrand, H.: Visceral larva migrans: Toxacara canis infection. Lancet I (1964) 1357-1359.

2. Mok, C. H,: Visceral larva migrans: A discussion based on review of the literature. Clin. Pediatr. 7 (1968) 565-573.

3. Aur, R. J. A., Pratt, C. B., Johnson, W. W.: Thiabendazole in visceral larva migrans. Am. J. Dis. Child. 121 (197l) 226-229.

4. Sacchi, F. T., Ferrari, F. A., Fortunato, A., Maggiore, G., Marconi, M., Pagani, A., Siccardi, A. G.: A defect in neutrophil motility in two siblings with recurrent infections and a remarkable family history. Infection 7 (1979) 45-47.

5. Zinkham, W. H.: Visceral larva migrans: A review and reassessment indicating two forms of clinical expression: visceral and ocular. Am. J. Dis. Child. 132 (1978) 627-633.

6. Hill, H. R., Quie, P. G.: Raised serum IgE levels and defective neutrophil chemotaxis in three children with eczema and recurrent bacterial infections. Lancet I (1974) 183-187.

7. Hill, H. R., Estensh, R. D., Hogan, N. A., Quie, P. G.: Severe staphylococcal disease associated with allergic manifestations hyper- immunoglobulinemia E and defective neutrophil chemotaxis. J. Lab. Clin. Med. 88 (1976) 796-806.

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