The American Journal of Tropical Medicine and Hygiene - Volume 29, Issue 2, 1980

A cohort of 113 women and their newborns from the coastal area of El Salvador were studied longitudinally to estimate malaria incidence and indirect fluorescent antibody (IFA) response to malaria infection. The district in which the study was conducted had an estimated annual parasite index of 600/1,000 inhabitants, and all malaria infections were treated immediately with a 4-aminoquinoline. In the third trimester of pregnancy, the IFA response to Plasmodium falciparum was significantly depressed. As a result of antimalarial therapy and depressed immune responsiveness, 49% (P. vivax) and 53% (P. falciparum) of the pregnant subjects had a malaria IFA titer <1:20 at the time of delivery. Malaria IFA crossed the placenta to the fetus with a step-down of approximately a 4-fold dilution, except for the step-up noted in the P. falciparum titer for 17 of 116 newborns. Due to the overall low prevalence and intensity of maternal IFA, a titer of at least 1:20 was passed to only 23% (P. vivax) and 45% (P. falciparum) of newborns. Passively-acquired malaria IFA degraded with a half-life estimated between 43 and 52 days. During follow-up of infants to 6 months of age, no protection from malaria resulting from passively-acquired antibody could be demonstrated. Because of the limited transplacental immunization of these newborns with antimalarial antibody, it appears that passive immunity can exert little effect on the incidence of infant malaria in coastal El Salvador.

This report deals with the major features of untreated infections with Plasmodium inui as exhibited in a group of 31 rhesus monkeys (27 inoculated with trophozoites, four with sporozoites). Infections in nine monkeys were followed to self-cure. In 22 subjects they were interrupted prematurely: in 17 by chemotherapy 454 to 3,931 days after onset of patency; in 5 by death 491 to 1,025 days after onset of patency. Substantial monkey-to-monkey variations were encountered in the intensity of the parasitemia and its cyclic undulations, in the morbid features of the disease, and in the duration of infection. Thus initial peak parasitemias varied by as much as 15-fold. Some post-peak parasitemias were sustained at relatively constant and moderately high levels for many months; others fluctuated cyclically between barely detectable and readily countable numbers. In most subjects, there was no evidence of morbidity attributable to malarial infection; in two, reactions were severe enough to lead to death. The duration of untreated infections could be as brief as 14 months or could approach 14 years. With due allowance for premature interruptions, the majority of infections could be expected to persist for 4–6 years.

Anti-Trypanosoma cruzi agglutinins were studied by the direct agglutination test (DA) with or without previous treatment of the sera with 2-mercaptoethanol (2-ME) (DA and 2-MEDA, respectively), in serial serum samples obtained from 24 patients with acute Chagas' disease. Their agglutinin titers were compared with those found in 25 patients of similar age with other acute infections and 50 healthy children. All of the controls were negative by complement-fixation test for T. cruzi infection. All sera were also tested by indirect hemagglutination (IHA) and indirect immunofluorescence (IIF) tests, using anti-total human globulin (IIF-Ig) and anti-IgM human globulin (IIF-IgM). In the first serum sample obtained after infection from the chagasic patients, DA and 2-MEDA detected IgM antibodies in seven patients who were also positive by IIF-IgM. However, in 13 early sera from other patients antibodies were demonstrated by IIF-IgM only. Four patients were negative by both techniques. Early serum samples were usually negative in the IHA test. Further samples obtained from those patients in whom the infection became chronic were reactive in the IHA test. In these sera, agglutinin titers were usually not affected by 2-ME treatment, indicating a change in the class of Ig reactivity (from IgM to IgG). This observation was coincident with decreasing titers established by IIF-IgM, whereas antibody titers detected by IIF-Ig remained positive. However, in four patients IIF-IgM antibodies either persisted or reappeared following a period during which they were negative. During the follow-up of four patients in whom serologic reactivity became negative after treatment, antibodies were detected later. In two of these cases a positive IIF-IgM was observed.

A micro enzyme-linked immunosorbent assay (ELISA) for antibodies to Trypanosoma cruzi was evaluated and the results obtained by ELISA were compared with those obtained by the complement fixation test (CF) and indirect fluorescent antibody test (IFA). Fifty sera collected from residents of the southeastern United States all had reciprocal ELISA titers ⩽320. Similarly, serum samples from 17 patients with T. cruzi infection proven by xenodiagnosis had reciprocal ELISA titers of ⩾1,280. Specimens from 302 El Salvador Army recruits were tested by ELISA, IFA, and CF. Excellent correlation was observed between results obtained by the three serologic tests; 62.9% of the samples were negative by each of the three tests and 24.5% were positive by all. Overall, 29.5% of the sera were positive for antibodies to T. cruzi by ELISA, 29.5% by IFA, and 31.5% by CF. The data suggest that the micro ELISA is a promising serologic test for measuring antibodies to T. cruzi in individuals and in populations.

An immunoenzymatic diagnostic technique applicable to cutaneous leishmaniasis is described. The antigen used (Leishmania tropica major) was equally useful in diagnosing visceral and mucocutaneous forms of the disease. The criteria for positivity were defined by using groups of negative controls, and the specificity of the reaction was evaluated by using sera from patients with various diseases. Among these, sera from patients with lepromatous leprosy, tuberculosis, or African trypanosomiasis strongly cross-reacted with leishmania antigen. Examining serial dilutions of the sera facilitated the interpretation of the results and eliminated a significant percentage of false positives.

Of 21 confirmed cases of New World leishmaniasis, 16 exhibited antibody to antigens of the promastigote of Leishmania braziliensis panamensis by the enzyme-linked immunosorbent assay (ELISA). Comparison of antibody titers obtained by ELISA with titers obtained by indirect immunofluorescence using an amastigote substrate confirmed that the sensitivities of the two techniques were within the same range (r = 0.80). Although sera from patients with New World leishmaniasis failed to react with antigens extracted from epimastigotes of Trypanosoma cruzi, sera from 39 cases of Chagas' disease were reactive with promastigotes of L. braziliensis panamensis. This apparent unidirectional cross-reactivity has been attributed to differences in potency of the antigenic stimulus presented in the two diseases.

Circulating immune complexes (CIC), anti-IgG, anti-DNA, and anti-collagen autoantibodies (Ab) were investigated in sera from patients with South American leishmaniasis. No significant levels of anti-DNA or anti-collagen autoantibodies were observed. Only a few patients with cutaneous leishmaniasis or with only one mucocutaneous lesion showed values for CIC and anti-IgG Ab higher than those in the control group. In contrast, both CIC and anti-IgG Ab were demonstrated in most patients with several lesions due to mucocutaneous leishmaniasis (MMC). Moreover, a close correlation was noticed between the detection of CIC and anti-IgG Ab in MMC patients. This relationship suggested that part of the detected CIC could have been formed by IgG-anti-IgG complexes. The involvement of these immunopathologically active substances in the clinical evolution of mucocutaneous leishmaniasis is discussed.

In this study, isozyme patterns for 14 different enzymes were compared for culture strains of Leishmania braziliensis, L. hertigi, L. mexicana, L. donovani, L. tropica, and L. adleri. The isozyme separation was made by means of cellulose acetate electrophoresis. Each of the species had distinct isozyme patterns for aspartate aminotransferase, glucose 6-phosphate dehydrogenase, 6-phosphogluconate dehydrogenase, and fructokinase. For other enzymes, two or more species had identically migrating bands; however, by using combinations of the other 10 enzymes it was possible to separate any one of the six species. In addition to these interspecific differences the Panama strains of L. braziliensis had two different malic dehydrogenase isozyme patterns; therefore, they fell into two distinct groups. These strains otherwise had identical isozyme patterns.

In in vitro assays the hemolytic activity of total homogenates of trophozoites of seven strains of amebae were tested for specificity and potency against the erythrocytes of eight mammalian species. The species of Entamoeba included in the tests were E. histolytica, E. invadens, E. moshkovskii, and the Laredo type of E. histolytica. The hemolytic activity was found to be dose-dependent and strain-specific, with E. histolytica being generally the species having the highest hemolytic potency.

The uptake of amino acids and glucose by microfilariae of Dirofilaria immitis maintained for 8 days in vitro was studied. The quantity of glutamic acid, histidine, lysine, alanine, tryptophan, and cystine in the medium did not change. Isoleucine, serine, valine, phenylalanine, tyrosine, methionine, and cystathionine decreased slightly; both asparatic acid and leucine decreased considerably. Glutamine, threonine, and proline decreased markedly, while glycine increased to some extent. Ornithine increased and arginine decreased; however, their changes were smaller than the changes of respective amino acids in the control medium free of microfilariae. Glucose decreased markedly.

The 50-mg Kato thick smear was modified by using stainless steel templates which deliver 20 mg of stools accurately; fecal material can thus be processed in the field and the smears cleared within 15 minutes. The counts obtained from the 20-mg quick Kato were proportional to those obtained by the 50-mg thick smear.

Schistosomes obtained by perfusion from host animals as early as 2 hours after in vivo treatment with hycanthone and transferred into untreated recipient hamsters died in the recipient host. In contrast, unexposed schistosomes transferred into recipient hamsters treated from 7 days to 36 hours previously showed a normal survival. In vitro treatment of schistosomes with hycanthone concentrations comparable to those used in in vivo studies, followed by transfer of the parasites into normal hamsters, resulted in death of the worms. The time of lethal hycanthone exposure in vitro could be as short as 15 minutes. Hycanthone-resistant schistosomes or immature worms were not affected under similar in vitro conditions. Our data suggest that the schistosomicidal effect of hycanthone is not caused by a host-derived metabolite.

The effect of unisexual schistosome infection on the activities of several hepatic enzymes was studied in mice. The activities of hepatic drug-metabolizing enzymes in mice infected with either female or male schistosomes were not significantly different from those of noninfected control animals. However, the total amount of heme pigment in the liver of infected mice was 2.7 (female infection) and 8.9 (male infection) times greater than that of control animals. The durations of hexobarbital sleeping times and of zoxazolamine paralysis in unisexual schistosome infections did not differ from those of uninfected controls. Therefore, an accumulation of schistosome pigment without egg deposition, as in this unisexual infection study, does not result in a severe reduction of hepatic drug-metabolizing capacity.

The effects of infection with Schistosoma mansoni on the activities of several hepatic drug-metabolizing enzymes were investigated in congenitally athymic homozygotic nude mice and in a heterozygotic strain of BALB/c derived mice. In athymic nude mice, infection with schistosomes of the same duration and intensity (in terms of the number of eggs in the liver) as in heterozygotic mice resulted in a much smaller reduction in hepatic drug-metabolizing enzyme activities. Therefore, the severe reductions of the hepatic drug-metabolizing function in this infection occur only in mice that are immunologically competent and, thus, are dependent on the host's response to the parasite eggs.

The presence of a P1-active substance in hydatid cyst fluid has been known since 1957. Recent work has revealed the presence of high-titer anti-P1 antibodies in blood group P2 patients with Fasciola hepatica infection (distomiasis). These findings, together with the observation of a high percentage of cross-reactions in the hydatid indirect hemagglutination test (IHA) among distomiasis patients, led us to study the possible role played by anti-P1 antibodies in the occurrence of these nonspecific reactions. Of the 51 sera from patients with distomiasis studied, seven were IHA-positive. Of these, six presented high titer anti-P1 antibodies. Absorption of these sera with P1 erythrocytes eliminated their anti-P1 activity; at the same time, four of the sera became negative in the IHA test and the IHA titers of the two remaining sera decreased without disappearing entirely. Results with two reference animal anti-P1 sera confirmed these findings. The agar gel diffusion reaction revealed cross-reactions in addition to the P1-anti-P1 precipitin line. The negative results obtained with seven high titer anti-P1 distomiasis sera and 21 control sera from patients without hydatidosis, which showed a weak “naturally-occurring” anti-P1 activity, led us to discuss the conditions necessary for the reaction between anti-P1 and P1 antigen of hydatid cyst fluid to be detectable by IHA at levels significant for serodiagnosis of hydatidosis. Finally, the importance of further work to improve the specificity of the quantitative methods for serodiagnosis of hydatid disease without impairing their sensitivity is emphasized.

To determine the role of rotavirus, enterotoxigenic Escherichia coli and enteropathogenic E. coli in diarrheal disease of non-hospitalized children and adults living in rural El Salvador, stool specimens were collected from 156 persons with diarrhea and 134 age- and sex-matched controls over a 1-year period. Enterotoxigenic Escherichia coli (ETEC) were isolated as frequently from controls (13.4%) as from diarrhea cases (12.2%). Enteropathogenic E. coli were isolated from 13 cases (8.3%) and 10 (7.7%) controls. Rotavirus was demonstrated in only five of the 129 specimens from cases examined; the five persons infected were ≤3 years of age. No invasive E. coli were found. Serotyping of ETEC revealed serogroups of ETEC previously associated with enterotoxigenicity but was not helpful in separating infection from disease. The etiology of diarrhea in this rural, non-hospitalized population was complex. Isolation of a known pathogen did not prove etiology. The rotaviruses, which have been isolated frequently from hospitalized persons, were rare. Further laboratory and epidemiologic studies in such populations are needed to identify those factors that determine pathogenicity.

Vampire bats were inoculated intramuscularly and subcutaneously with varying doses of rabies virus to simulate bites by rabid animals in nature. Daily saliva samples were then taken from these animals to determine whether they excreted virus and for how long. Vampire bats appear to react to rabies virus as do other animals, with variable incubation periods, some excretion of virus in the saliva, but no prolonged excretion “carrier state.”

A mathematical model of Crimean hemorrhagic fever (CHF) which satisfactorily describes the real epidemic process has been developed. Estimates of the intensity of infection and probability of disease have been obtained on the basis of the model. The probability of disease for subjects who had been infected was found to be a stable value characterizing the infection. In CHF, this value is 0.2153; in other words, the ratio of inapparent to clinically overt forms is approximately 5:1. By means of the model the causes of the lack of a stable immune portion among the human populations in CHF foci have been elucidated.

Patients with St. Louis encephalitis may have urinary tract symptoms when they develop manifestations related to involvement of the central nervous system. During 1976, 12 patients with St. Louis encephalitis were studied for the presence of antigenuria, and compared to controls. Although virus isolation attempts were negative indirect immunofluorescence, electron microscopy, and immune electron microscopy demonstrated the presence of viral antigen in urine. Further study of antigenuria in St. Louis encephalitis is needed to delineate the pathophysiology of the lower urinary tract symptoms and to determine whether this phenomenon might furnish the basis for a rapid diagnostic test for the disease.

All of 13 species of northward migrating shorebirds and 7 species of songbirds captured on the Pacific coast of Guatemala during April and May of 1974–1976 were susceptible to infection with small doses of either an epizootic or an enzootic strain of Venezuelan encephalitis (VE) virus. They produce moderate to high levels of viremia for 2–4 days post-inoculation; levels high enough to infect both epizootic and enzootic vector mosquitoes. Viremias were often sufficient even on the 3rd day after inoculation, a time that might represent the end of a migratory flight, assuming that the physiological state of the birds after capture reflected that during migratory flight. Birds of many taxa react similarly to infection with strains of VE virus, and have the potential for being moderately to highly effective amplifying hosts. However, whether northward migrating birds could have been the agents for the introduction of the epizootic Ecuadorian strain that initiated the middle-American epizootic of 1969–1971 is less clear. Data are not available for the extent, rates or routes of migration between the region of Ecuador and Central America, but the best information on the real speed of migration from elsewhere indicates that even warblers that fly more slowly than shorebirds could make the flight in 72 hours or less. Still, that there are geographically segregated subtypes of VE virus suggests that avian transport has been of minimal importance over long time spans. The role of inactivated vaccines in the middle-American epizootic remains an open question.

From January 1976 through January 1979 serum specimens from 1,575 individuals were received at the Center for Disease Control and tested for antibodies to rickettsiae. Of these, sera from eight persons gave serological results indicative of recent infections with epidemic typhus rickettsiae (Rickettsia prowazekii). Five of the persons were from Georgia, and one each was from Tennessee, Pennsylvania and Massachusetts. The illnesses occurred during the winter, chiefly in persons living in a rural environment. The clinical picture was compatible with louse-borne epidemic typhus. There was no apparent contact with human body or head lice, and no cases occurred in patient contacts, indicating that infection was not associated with the classic man-louse-man cycle of epidemic typhus. Two of the eight patients had contact with flying squirrels, suggesting that they became infected from this known extrahuman reservoir of R. prowazekii.

The distribution of sucrose-electrolyte oral therapy packets (1 liter) by community-based workers in a rural Bangladesh population of 157,000 was evaluated. A similar population of 134,000 served as a comparison group. The locally-produced packets showed satisfactory chemical composition with a shelf-life of up to 3 months and a cost of U.S. $0.05. After 4 months the workers were distributing an average of 70 packets/1,000 population per month. Most patients used one packet for each episode of diarrhea; 13% of children used two packets, and 15% and 8% of adults used, respectively, two and three packets. The electrolyte composition of the oral fluids prepared by field workers and mothers showed substantial variation, but no hyperconcentrated solutions were noted. A comparison of the hospitalization rate from the two study areas suggested a 29% reduction in hospitalization for diarrhea during the 4 months of distribution.

A sudden outbreak of skin ulcers occurred in the Cook Islands beginning in early 1976. Information from the outbreak indicated that ulcers were most consistent with the clinical entity of tropical ulcer. A retrospective analysis was undertaken to determine the clinical and epidemiological features of the outbreak. The ulcers occurred most commonly in males in the 10–14 and 15–19 age groups. Antecedent trauma appeared to initiate the ulcer. Most ulcers were solitary, occurred on the lower limbs, and healed leaving a pigmented scar. The ulcers responded well to penicillin. The cause of the ulcers was not established, but gram-negative, pleomorphic bacilli were consistently observed in ulcer biopsies.

The Waorani Indians of eastern Ecuador provide a unique opportunity for studying exposure of an isolated human population to various infectious disease agents. Using serologic tests to determine antibody prevalence, skin test data, and stool examination for parasites, we have been able to construct a profile of infectious diseases which are endemic, and others which have been introduced into the Waorani population. These findings are compared with similar data reported from elsewhere in the Amazon. Serologic studies demonstrating the presence of antibody to measles and poliovirus type 3 after vaccination indicate that the Waorani respond normally to viral challenge with these agents. The question of genetic inability among aboriginal Amerindians to respond to viral agents is discussed. Finally, general recommendations are made regarding the future health care of the Waorani.

Microfilariae of Mansonella ozzardi, concentrated from human blood, frozen with a cryoprotectant chemical, and stored at liquid or vapor-phase liquid nitrogen temperatures were motile when thawed after 2.5 years of preservation.

Frogs and toads, as paratenic hosts, are known to be sources of human angiostrongyliasis. The present investigation was carried out to examine the role of the tadpole as an experimental intermediate host. First-, second-, and third-stage larvae were found in tadpoles of Xenopus laevis from the 1st, 14th, and 18th days, respectively, after exposure to the first-stage larva of Angiostrongylus cantonensis. Third-stage larvae collected from a frog which had metamorphosed from an exposed tadpole reached maturity and oviposited in an albino rat. This is the first demonstration that a vertebrate, the tadpole of X. laevis, can serve as an experimental intermediate host of A. cantonensis.

A lampyrid larva (Coleoptera) was found during snail collecting in Liberia, West Africa to be a natural predator of Biomphalaria pfeifferi and Bulinus globosus.

One hundred and fifty-five Hymenolepis nana-infected children were treated with a single oral dose of praziquantel, a new and effective isochinolinpyrazin compound with a broad spectrum of activity against cestodes. The children were distributed in three groups which received different dosages of the drug. Parasitological cures were obtained in 64 (98.5%) of 65 children given 25 mg/kg body weight, in 61 (93.8%) of 65 given 15 mg/kg, and in 19 (76.0%) of 25 given 10 mg/kg. Durg tolerance was good, and no clinical side effects were observed. A series of blood and urine tests performed before and after treatment in 30 patients who received 25 mg/kg remained within normal values.

Biochemistry and Physiology of Protozoa, Volume 1, edited by M. Levandowsky and S. H. Hutner. xvi + 462 pages, illustrated. Academic Press, 111 Fifth Avenue, New York, N.Y. 10003. 1979. $45.00.

Companion to Clinical Medicine in the Tropics and Subtropics, by E. B. Adams. x + 210 pages, illustrated. Oxford University Press, 200 Madison Avenue, New York, N.Y. 10016.

In writing this manual the authors' stated objective was to present diagnostic parasitology in a clear, concise manner, for use as a bench manual to be supplemented by other sources. Garcia and Ash have succeeded in meeting their objective. This second edition is a thorough revision and expansion of the first edition. The manual is arranged in 12 chapters, an appendix, glossary and references, general and supplemental.

Chapters 1–3 present standard methodologies for collection, preservation, macro- and microscopic examination of fecal specimens and permanent staining of smears. Advantages and disadvantages of methods cited are discussed and the reader is informed why it is necessary to do specific procedures. References are cited for each technique as an aid if problems are encountered or detailed information is desired.

Chapters 4–6 detail methodologies for culture, concentration and recovery of larval nematodes, estimation of worm burden, methodology for collection and examination of specimens from the digestive and urogenital systems, sputum aspirates and biopsy material.

3 September 1979

To the Editor:

We have read the Presidential Address in the May 1979 issue of the American Journal of Tropical Medicine and Hygiene, and would like to comment on it. We make these comments because we believe some of the statements in the Address, especially when made by a person of Dr. Kagan's international stature, reflect unfavorably on one of the most important tools for schistosomiasis transmission control.

In the Address, Dr. Kagan makes the assertion that “The control of schistosomiasis by mollusciciding, however, has proven to be essentially ineffective.” If he had added the qualifying statement: “where used in a haphazard manner, without appropriate precontrol studies,” then we might have agreed with him. Obviously, in any situation in which pesticides are used without proper knowledge of the local epidemiology of any disease, then environmental impact on non-target species, lack of cost-effectiveness, development of target-species resistance, and failure to attain interruption of transmission may indeed occur.

27 November 1979

To the Editor:

In my Presidential Address, when I was discussing the control of schistosomiasis by drugs, I stated: “The control of schistosomiasis by mollusciciding, however, has proven to be essentially ineffective.” Doctors Christie, Prentice, and Barnish took exception to this remark because I did not qualify my statement. In their letter they list some of the reasons why mollusciciding has been ineffective—because of use “in a haphazard manner, with inadequate precontrol studies” and “without proper knowledge” with regard to application, etc. In addition to these technical aspects, there are other reasons why the control of snail populations by mollusciciding is so difficult. Snail populations can be and have been drastically reduced by the application of chemicals in an endemic area. However, as soon as the program is terminated or relaxed for financial, logistic, or political reasons, the snail population usually returns to what it was before the chemicals were applied. There can he no short-term control of schistosomiasis by the use of molluscicides.