The American Journal of Tropical Medicine and Hygiene - Volume 58, Issue 4, April 1998

The neutralization of Japanese encephalitis virus (JEV) was studied using JEV-specific neutralizing (NT) monoclonal antibody (MAb) 503 that recognizes the envelope glycoprotein. Analysis using radiolabeled JEV and observations by confocal laser microscopy and electron microscopy indicated that the NT and protection activities of MAb 503 did not result from the prevention of the first step of JEV infection, binding of virus to the cell surface. Treatment with MAb 503 strongly inhibited JEV-induced cell fusion and internalization of JEV into the cells, and resulted in enhanced release of JEV-RNA from the cells. These observations suggested that the NT activity of MAb 503 is involved in the later steps of JEV infection.

Agglutination and rosette formation are in vitro characteristics of Plasmodium falciparum-infected erythrocytes, which have been associated with host protective immune responses and also with parasite virulence. The present study was carried out in an area of seasonal and unstable malaria transmission in eastern Sudan. Plasma samples were obtained before, during, and after the transmission season from a volunteer cohort of 64 individuals seven years of age and older. These plasmas were assayed for their ability to agglutinate cultured parasitized erythrocytes originally obtained from acute malaria infection samples taken from five of the cohort members. Our data show that the capacity of donor plasma samples to agglutinate parasitized cells depended largely on the time of sampling relative to the transmission season, at least within this epidemiologic setting. Thus, although less than half of the pretransmission season samples could agglutinate any of the five lines of cultured parasites, all post-transmission season samples could agglutinate at least one of the parasite lines, with 74% agglutinating two or more lines. This increase in the agglutination capacity of individual plasma samples after the transmission season occurred essentially regardless of whether an individual had experienced a clinical malaria attack during the transmission season. The study thus confirms the acquisition of agglutinating antibodies following episodes of clinical malaria, but also demonstrates that such acquisition can take place in the absence of disease, presumably as a consequence of subclinical infection. This is the first demonstration of marked seasonal fluctuations in the capacity of individuals' sera to agglutinate parasitized red blood cells. Possible explanations for this effect include a decrease in the levels of agglutinating antibodies between seasons, or shifts in the antigens being recognized by such antibodies from one transmission season to the next. Finally, we showed the existence of marked seasonal fluctuation in the levels of agglutinating antibodies, either because levels of such antibodies are not sustained between seasons or because the antigens recognized change from one season to the next.

In a cross-sectional survey carried out in west Africa (The Gambia), where Plasmodium falciparum malaria is endemic with seasonal transmission, 178 individuals 1-75 years of age were assessed for their antibody response to the malaria vaccine candidate, merozoite surface protein 2 (MSP2). Total IgG to recombinant antigens representing full-length, repetitive, and group-specific domains of both allelic families of MSP2 was determined by ELISA. The IgG-subclass profile of IgG-positive sera was assessed. Antibody prevalence was age-dependent, reaching a peak during adolescence. In MSP2-seropositive individuals, there was a predominance of cytophilic antibodies (IgG1 and IgG3); IgG1 antibodies were prevalent in children less than 10 years of age, whereas in adolescents and adults MSP2-specific antibodies were predominantly IgG3. In parallel, we conducted a longitudinal study of children (3-8 years of age) from the same community; sera collected before the malaria transmission season were tested for the presence of anti-MSP2 antibodies. The subsequent susceptibility of these children to clinical malaria was monitored and the association between anti-MSP2 antibodies of different IgG subclasses and resistance to clinical malaria was tested. The presence of IgG3 antibodies to MSP2 serogroup A was negatively associated with the risk of clinical malaria whereas IgG1 antibodies to MSP2 serogroup B were associated with an increased risk of clinical infection. Our data suggest that age/exposure-related acquisition of IgG3 antibodies to MSP2 may contribute to the development of clinically protective immunity to malaria.

Serology is a critical component in the diagnosis of amebic liver abscess. However, in areas endemic for amebiasis there is a high background level of seropositivity for amebiasis (owing to previous infection with Entamoeba histolytica), which may complicate the interpretation of a positive serologic test result. Recently, we reported that serologic tests based on recombinant E. histolytica antigens might offer improved diagnosis of current invasive amebiasis because they apparently differentiated active infection from past exposure to the parasite. To confirm this finding, we have performed a longitudinal study on 20 patients with amebic liver abscess by examining their seroreactivity over time with recombinant versions of two major E. histolytica proteins, the serine rich E. histolytica protein (SREHP), and the 170-kD subunit of the galactose-specific adhesin. We found that more than 50% of the patients examined had become seronegative by one or both recombinant tests within 180 days of their diagnosis of amebic liver abscess. In the case of the recombinant SREHP-based tests, 12 patients had become seronegative 90 days after presentation. In contrast, all patients remained seropositive by a standard conventional test, an indirect hemagglutination test, at more than six months after presentation. Our study shows that patients lose seroreactivity with the recombinant SREHP or 170-kD antigen-based tests more rapidly than with a conventional serologic test; this may make them useful for the serologic diagnosis of amebiasis in endemic areas.

Cathepsin L1 (CL1), an immunogenic cysteine proteinase secreted by juvenile and adult Fasciola hepatica, was assessed for its potential as a diagnostic agent for the serologic detection of human fascioliasis. Using ELISAs, we compared the ability of liver fluke homogenates (LFH), excretory/secretory (ES) products, and CL1 to discriminate between seropositive (infected) and seronegative (noninfected) individuals within a population of 95 patients from the Bolivian Altiplano. A high prevalence of human fascioliasis has been reported in this region. The division between the seropositive and seronegative individuals was poorly defined when LFH was used as the antigen. A greater discrimination between these populations was achieved with both ES and CL1. A K-means cluster analysis using the combined ES and CL1 ELISA data identified a cluster of seropositive individuals. Cathepsin L1 detected a subset (20) of these seropositive individuals while ES detected all 26; however, ES detected nine additional individuals that were in the seronegative cluster. The ratio of the mean absorbance readings between seropositive and seronegative individuals was markedly improved by using conjugated second antibodies to IgG4, the predominant isotype elicited by infection. In these IgG4-ELISAs, CL1 again identified fewer individuals as seropositive than did ES, but improved the discrimination between the seropositive and seronegative individuals and thus provided a more conclusive diagnosis. Sera obtained from patients infected with schistosomiasis mansoni, cysticercosis, hydatidosis, and Chagas' disease were negative in these assays, which demonstrated the specificity of the IgG4-ELISA for detecting fascioliasis. Twenty of the 95 patients (21%) were seropositive for fascioliasis by the CL1 IgG4-ELISA, confirming the earlier reports of the high prevalence of disease in this region. A standardized diagnostic test for human fascioliasis, based on an ELISA that detects IgG4 responses to CL1, could be available to all diagnostic centers if sufficient quantities of recombinant CL1 can be produced.

Pulmonary fibrosis was induced following inoculation of Paracoccidioides brasiliensis conidia intranasally in BALB/c mice. Fibrosis was associated with formation of granulomas, increase in lung hydroxyproline, and sustained increases in tissue tumor necrosis factor-alpha and transforming growth factor-beta. This study suggests a role for these cytokines in generation of pulmonary fibrosis associated with chronic granulomatous infectious diseases.

Honduras has at least five-times more human immunodeficiency virus (HIV)-infected individuals than any other country in Central America. The relationship between HIV status and the presence of intestinal parasites in this part of the world is unknown. This study presents the results from a prospective, comparative study for the presence of parasites in 52 HIV-positive and 48 HIV-negative persons in San Pedro Sula, Honduras. Infection with HIV was determined by microagglutination and confirmed by Western blot analysis. Parasites were detected in stools using formalin-ether concentration, and Kinyoun and trichrome staining. Age, sex, and clinical state of HIV infection were recorded for each study participant. Our results indicate that Cryptosporidium parvum and Strongyloides stercoralis, which are intracellular or live in the mucosa, were found exclusively in persons infected with HIV. In comparison, the prevalence of the extracellular parasites Giardia lamblia, Ascaris lumbricoides, and Trichuris trichiura was significantly higher (P < 0.05) in persons who were HIV-negative. Trichuris worms are in contact with the gut epithelium and less so with the mucosa, whereas Strongyloides lives within the gut mucosa. It is possible that changes in the gut epithelium due to HIV infection do not affect the mucosa and therefore would not affect Strongyloides. We conclude that infection with HIV may selectively deter the establishment of certain intestinal parasites. This may be due to the fact that HIV-induced enteropathy does not favor the establishment of extracellular parasites. Intracellular and mucosal dwelling organisms, however, may benefit from pathologic changes and reduced local immune responses induced by the virus, which, in turn, may lead to higher prevalence among HIV-infected individuals.

From 1987 to 1995, a retrospective case study was conducted at the Ramon y Cajal Hospital in Madrid, Spain, a public teaching hospital with 1,100 beds, to determine the clinicoepidemiologic characteristics, survival, and prognostic factors of patients with visceral leishmaniasis (VL) and human immunodeficiency virus (HIV) infection. The prevalence of VL in HIV+ patients compared with HIV- patients was studied. Epidemiologic, clinical, and parasitologic characteristics, as well as the effects of treatment, prognosis, and survival in 54 HIV+ patients (90 episodes) with VL were defined. Comparative survival studies among patients with and without acquired immunodeficiency syndrome (AIDS)-defining criteria and multivariate analysis of survival risk factors were performed. The prevalence of VL in patients with AIDS was much higher than in immunocompetent individuals. In spite of a good initial response to treatment for VL, 60.6% of the patients had relapsed by the end of one year. Mortality from the first episode was 18.5%, and 24% died in the first month after diagnosis of any VL episode. The mean survival of the 29 patients who died was 10.27 months. Survival in patients with and without AIDS at the time of the first episode of VL was compared at 30 months: 53.7% versus 20.5% (P = 0.00149). We found no significant difference (P = 0.24) in the survival of HIV+ patients who had died of VL without AIDS at the time of the first episode of VL compared with those of a control group of 413 dead patients with AIDS without VL. A diagnosis of AIDS at the time of the first episode of VL and thrombocytopenia were the only risk factors found related to survival. We conclude that in AIDS patients, VL is a recurrent disease that is highly prevalent and whose clinical course is modified by HIV.

Seventy-five isolates from the State of Campeche, Mexico, an area endemic for localized cutaneous leishmaniasis (LCL), were characterized by isoenzyme markers (glucose phosphate isomerase, mannose phospate isomerase, nucleoside hydrolase, phosphoglucomutase, 6-phosphogluconate dehydrogenase, and glucose-6-phosphate dehydrogenase). Seventy (93.3%) were identified as Leishmania (Leishmania) mexicana and 5 (6.7%) as L. (Viannia) braziliensis. This is the first report of authochthonus human LCL caused by L. (V.) braziliensis in the State of Campeche, Yucatan Peninsula, Mexico.

Three groups of three, six, and 12 dogs with parasitologically proven clinical visceral leishmaniasis (Leishmania chagasi infection) were treated with intramuscular aminosidine sulfate at doses of 20 mg/kg/day for 15 days; 80 mg/kg/day for 20 days, and 40 mg/kg/day for 30 days, respectively. Follow-up was by parasitologic examination of bone marrow and skin, serology using the indirect immunofluorescent antibody test, and clinical examination for signs of visceral leishmaniasis or adverse effects of treatment. In animals treated with 20 mg/kg/day, for 15 days, there was dramatic clinical improvement with disappearance of conjunctivitis, increase in appetite, weight gain, and recovery of normal skin condition and a healthy coat, but parasitologic relapse occurred between 50 and 100 days after initiation of treatment. Adverse effects were seen with treatment with 80 mg/kg/day for 20 days; three dogs died during or just after treatment, two showed temporary recovery, and one showed total clinical and parasitologic cure that was maintained for four years. Although adverse effects and relapses were seen in some dogs treated with 40 mg/kg/day for 30 days, three of 12 dogs showed complete parasitologic and clinical cure that was sustained for at least four years. Aminosidine treatment cannot be recommended as an alternative to the humane destruction of dogs for the control of canine visceral leishmaniasis because ineffective treatment may prolong carrier status or encourage development of drug resistance. This drug may be a therapeutic option if there is no danger of a dog acting as a reservoir of infection. Achievement of clinical recovery and limited cure with aminosidine suggests that further trials would be of value, possibly in combination with other anti-leishmanial drugs and with supportive measures to reduce adverse effects.

The emergence of chloroquine resistance, and a world-wide scarcity of quinine, have resulted in a search for newer antimalarial drugs directed against falciparum malaria. Allopurinol causes virtually complete inhibition of purine biosynthesis of malaria parasites, which may prove lethal to the parasites. This study was designed to examine if allopurinol is additive to quinine in the treatment of acute falciparum malaria. Forty-seven Asian-Indian adults with acute complicated falciparum malaria were assigned to a treatment period of five days. They were randomly assigned to receive either oral allopurinol (12 mg/kg in three divided doses for five days) plus quinine (600 mg intravenously every 8 hr for two days, followed by 600 mg orally every 8 hr for three days ) (n = 24), or quinine alone (600 mg intravenously every 8 hr for two days, followed by 600 mg orally every 8 hr for three days) (n = 23). The responses were assessed by parasite clearance time, defervescence time, splenomegaly disappearance time, and cure rate. In the allopurinol-quinine (ALLQUIN)-treated group, all the durations were significantly shorter than those in the quinine alone (QUIN)-treated group. They were ALLQUIN versus QUIN (mean +/- SD = 65.33 +/- 11.47 hr versus 76.78 +/- 18.20 hr; P = 0.0214; 57.66 +/- 13.01 hr versus 82.52 +/- 23.55 hr, P = 0.0002; 10 +/- 1.64 days versus 14.65 +/- 2.4 days; P = 0.0002), respectively. The cure rate was higher in the ALLQUIN group (91.7%) than in the QUIN group (87%). However, this difference was not statistically significant. Therefore, this study indicates that allopurinol can be an additive to quinine to bring about both faster eradication of Plasmodium falciparum and clinical remission than with quinine alone.

One hundred nine Gabonese patients infected with Loa loa microfilariae were treated with ivermectin (200 microg/kg of body weight) at the Parasitology, Mycology and Tropical Medicine Department (Faculte de Medecine et des Sciences de la Sante, Libreville, Gabon). Each was given one dose per month for six consecutive months. The peripheral blood microfilaria (mf) count before and after each dose showed an average decrease in the microfilaremia of 87.3% (short-term-single dose). An annual single-dose mass treatment with 200 microg/kg of ivermectin was sufficient to control the parasite in populations with low (< 400/ml) L. loa mf counts. One month after the sixth dose (short-term-multiple doses), the average microfilaremia rate had decreased by 99.2% compared with the initial infection (35 patients). Samples were taken from 28 patients one month after the first dose and one month after the sixth dose. The average mf count decreased by 96.4% after the first dose and by 99.6% after the sixth dose (average residual mf counts = 13.7 and 1.5 mf/ml, respectively). The mf count after the sixth dose was only 11.2% of the count after the first dose. The low mf count persisted for more than six months after the sixth treatment (long-term-multiple doses). Thus, mass treatment with multiple doses is more appropriate for areas where the blood mf count is very high. These results show that the number of the annual treatments used in mass chemotherapy with ivermectin can be adapted to each population to provide efficient protection.

Over the past nine years, more than 12 million people exposed to Onchocerca volvulus infection have received at least one dose of ivermectin, almost all without serious adverse reactions. Since 1991, however, several cases with neurologic manifestations, including coma, have been reported after ivermectin treatment of persons infected with O. volvulus who also had concomitant Loa loa infection with very high microfilaremia (> 50,000 microfilariae/ml of blood). In 1995, four criteria were established to define probable cases of Loa encephalopathy temporally related to treatment with ivermectin (PLERI). The present paper describes three PLERI cases recorded in Cameroon and compares them with two others reported previously. Disorders of consciousness began 3-4 days after treatment. The objective neurologic signs were variable. The conditions improved favorably in three patients who benefited from early hospitalization and good nursing; their disorders of consciousness lasted only 2-3 days; the results of clinical examination became normal after one month and electroencephalographic abnormalities disappeared after 5-7 months. Conversely, late diagnosis and delay in proper management in two others probably led to worsening of the condition and to fatal outcome related to the usual complications of coma. In addition to these cases, patients w with high Loa microfilaremia also developed milder neurologic manifestations causing functional impairment lasting for at least one week after treatment. Before launching mass ivermectin distribution programs to control onchocerciasis in central Africa, communities in which the intensity of concomitant L. loa microfilaremia is high need to be identified, and specific educational measures and monitoring strategies should be developed and applied before they are treated.

The correlations between malnutrition, parasitosis (especially helminth infections), and child development are complex, and studies of these interrelationships will allow health agencies to maximize screening and intervention strategies for developing countries. We examined these correlations in a cross-sectional program in Carazo State, Nicaragua. Nine hundred sixty-one children in two age strata (ages 0-24 months and ages 2-10 years) from one urban and three rural communities were screened for intestinal parasites (direct smear and ZnSO4 flotation), malnutrition, and developmental delays. Nutritional status was determined as weight-for-age (WFA), weight-for-height (WFH), and height-for-age (HFA). Developmental status (normal, suspect) was determined for the four subtests of the Denver II Screening Test. The prevalence of malnutrition was 14.6% (WFA), 8.4% (WFH), and 36.3% (HFA). Parasitosis was more prevalent in children less than 24 months of age with low HFA, whereas in older children low WFA was more closely associated with parasitic infections. Ascaris and Trichuris were more prevalent in malnourished children. On the Denver II, suspect test results in all four categories (language, social, gross motor, and fine motor) were associated with low WFA, and suspect language tests were associated with both intestinal parasites (P = 0.0003) and Ascaris infection in particular (P = 0.044). Developmental disabilities are a significant and frequently undetected health problem in developing countries, and malnutrition associated with intestinal helminth infections may be an important contributory factor for these disabilities.

Studies of the correlations between malnutrition, parasitosis, and child development will help health agencies to maximize screening and intervention strategies for developing countries. Such correlations were examined in a cross-sectional study in Carazo State, Nicaragua. 961 children aged 0-24 months and 2-10 years from 1 urban and 3 rural communities were screened for infection with intestinal parasites, malnutrition, and developmental delays. Developmental status was determined for the 4 subtests of the Denver II Screening Test. The prevalence of malnutrition was 14.6% according to weight-for-age (WFA), 8.4% for weight-for-height (WFH), and 36.3% for height-for-age (HFA). Parasitosis was more prevalent among children under age 2 years with low HFA, while low WFA was more closely associated with parasitic infections in older children. Ascaris and Trichuris were more prevalent in malnourished children. On the Denver II, suspect test results in language, social, gross motor, and fine motor were associated with low WFA, while suspect language tests were associated with both intestinal parasites, and Ascaris infection in particular. Developmental disabilities are a significant and often undetected health problem in developing countries. Malnutrition associated with intestinal helminth infections may be an important contributory factor for such disabilities.

The aim of the present study was to determine whether an increase in resting energy expenditure (REE) contributes to the impaired nutritional status of Gambian children infected by a low level of infection with pathogenic helminths. The REE of 24 children infected with hookworm, Ascaris, Strongyloides, or Trichuris (mean +/- SEM age = 11.9 +/- 0.1 years) and eight controls without infection (mean +/- SEM age = 11.8 +/- 0.1 years) were measured by indirect calorimetry with a hood system (test A). This measurement was repeated after treatment with 400 mg of albendazole (patients) or a placebo (controls) (test B). When normalized for fat free mass, REE in test A was not different in the patients (177 +/- 2 kJ/kg x day) and in the controls (164 +/- 7 kJ/kg x day); furthermore, REE did not change significantly after treatment in the patients (173 +/- 3 kJ/kg x day) or in the controls (160 +/- 8 kJ/kg x day). There was no significant difference in the respiratory quotient between patients and controls, nor between tests A and B. It is concluded that a low level of helminth infection does not affect significantly the energy metabolism of Gambian children.

Findings are presented from a study conducted to determine whether an increase in resting energy expenditure (REE) contributes to the impaired nutritional status of Gambian children infected by a low level of infection with pathogenic helminths. The REE of 24 children infected with hookworm, Ascaris, Strongyloides, or Trichuris and 8 uninfected controls was measured by indirect calorimetry with a hood system. That measurement was then repeated after treatment with 400 mg of albendazole or a placebo. When normalized for fat free mass, REE in the indirect calorimetry test was not different in the patients and the controls. Furthermore, REE did not significantly change after treatment in the patients or in the controls. No significant difference was observed in the respiratory quotient between patients and controls, nor between the indirect calorimetry test and the measurement repeated after treatment with 400 mg of albendazole or a placebo. These findings suggest that a low level of helminth infection does not significantly affect the energy metabolism of Gambian children.

There is now accumulating evidence for the involvement of genetic factors in the control of immune response against malaria. These arguments come from numerous animal models, from population studies showing associations of red blood cell genetic defects as well as HLA antigens with severe malaria, and from familial studies including a recent segregation analysis, which led to detection of a major gene effect predisposing to high infection levels. The heterogeneity and complexity of this genetic control is one of the main findings of these previous studies, and probably a major cause of the difficulty in developing an effective malaria vaccine. A segregation analysis of blood infection levels is performed here in 44 pedigrees living in the tropical rain forest of southern Cameroon and exposed to high vectorial transmission intensity. The results confirm the existence of complex genetic factors controlling blood infection levels in human malaria but are not consistent with the parent-offspring transmission of a single Mendelian gene. This study also shows the dramatic effect of age on infection levels and its interaction with a putative major gene suggesting that genetic related differences are much more important in children than in adults. Further genetic studies focused on children may help to identify the nature of the genetic factors involved in the expression of human malaria, by means of linkage analyses using both familial information and genetic markers.

Females of laboratory-reared Culex quinquefasciatus were allowed to take blood meals on individuals with low (1-100 microfilariae/ml of peripheral blood), moderate (101-500 microfilariae/ml), and high (> 500 microfilariae/ml) microfilaremia. The mosquitoes ingested 1.39-3.80 microl of blood and infective third-stage (L3) larvae were first recorded 13-14 days after the infecting blood meal. The number of microfilariae ingested by mosquitoes was proportional to the density of microfilariae in the peripheral blood of the human subjects, but with a concentration factor of up to 6.5 times the expected number. Survival of mosquitoes was not influenced by the density of microfilariae in the peripheral blood of infected individuals. Infectivity indices were proportional to microfilaremia in human subjects. The number of L3 larvae/female (intensity of infection) was not influenced by individual microfilaremia. The highest vector efficiency and the best experimental infection index were recorded in mosquitoes that blood fed on individuals with moderate microfilaremia. The results are discussed in relation to the transmission on Bancroftian filariasis in the study area.

Persons with human immunodeficiency virus (HIV) infection who subsequently develop an acute sexually transmitted disease have an increased probability of transmitting HIV. Therefore, characterizing such persons can help direct prevention efforts to a group who are likely to be continuing sources of HIV transmission. We assessed the incidence and factors associated with trichomoniasis in a cohort of HIV-infected women receiving care at a public clinic in Los Angeles County, California from 1992 through 1995. Demographic, clinical, and behavioral data were available from medical records and from patient interviews. Trichomonas infection was the most frequently identified sexually transmitted disease and was found in 37 (17.4%) of 212 women representing a crude incidence rate of 14.1 per 100 person-years experience. The crude rate of trichomoniasis was highest in black women (69.0 per 100 person-years), women with a history of trading sex for drugs or money (51.0 per 100 person-years), those using crack or cocaine (35.5 per 100 person-years), women with four or more sex partners (43.0 per 100 person years), and those born in the United States (23.3 per 100 person-years). Among women with severe immunosuppression (CD4+ count < 200), 18.4% (18 of 98) were diagnosed with trichomoniasis. After multivariate analysis using a Cox proportional hazards approach, black race (adjusted rate ratio [RR] = 5.6, 95% confidence interval [CI] = 2.3, 13.3) continued to be strongly associated with Trichomonas infection. Trading sex for money or drugs (adjusted RR = 25.2, 95% CI = 4.3, 148.6) and single marital status (adjusted RR = 3.7, 95% CI = 1.1, 13.0) were independent risk factors for trichomoniasis in nonblack women but not among black women. Data from this study indicate that Trichomonas may be a frequently acquired infection in HIV-positive women. Our findings suggest that HIV-infected women who are black, and nonblack women who trade sex for money or drugs or are unmarried, are at increased risk of trichomoniasis and therefore may be more likely to transmit HIV infection. Local HIV prevention strategies should target such women for intervention efforts.

Human echinostomiasis, endemic to southeast Asia and the Far East, is a food-borne, intestinal, zoonotic parasitosis attributed to at least 16 species of digenean trematodes transmitted by snails. Two separate life cycles of echinostomes, human and sylvatic, efficiently operate in endemic areas. Clinical symptoms of echinostomiasis include abdominal pain, violent watery diarrhea, and anorexia. The disease occurs focally and transmission is linked to fresh or brackish water habitats. Infections are associated with common sociocultural practices of eating raw or insufficiently cooked mollusks, fish, crustaceans, and amphibians, promiscuous defecation, and the use of night soil (human excrement collected from latrines) for fertilization of fish ponds. The prevalence of infection ranges from 44% in the Philippines to 5% in mainland China, and from 50% in northern Thailand to 9% in Korea. Although the patterns of other food-borne trematodiases have changed in Asia following changes in habits, cultural practices, health education, industrialization, and environmental alteration, human echinostomiasis remains a health problem. The disease is most prevalent in remote rural places among low-wage earners and in women of child bearing age. Echinostomiasis is aggravated by socioeconomic factors such as poverty, malnutrition, an explosively growing free-food market, a lack of supervised food inspection, poor or insufficient sanitation, other helminthiases, and declining economic conditions. Furthermore, World Health Organization control programs implemented for other food-borne helminthiases and sustained in endemic areas are not fully successful for echinostomiasis because these parasites display extremely broad specificity for the second intermediate host and are capable of completing the life cycle without involvement of the human host.

Paracoccidioides brasiliensis, the causative agent of paracoccidioidomycosis (PCM), was first isolated from armadillos from the Amazonian region where the mycosis is uncommon. In the present study, we report on the high incidence of PCM infection in armadillos from a hyperendemic region of the disease. Four nine-banded armadillos (Dasypus novemcinctus) were captured in the endemic area of Botucatu, Sao Paulo, Brazil, killed by manual cervical dislocation and autopsied under sterile conditions. Fragments of lung, spleen, liver, and mesenteric lymph nodes were processed for histology, cultured on Mycosel agar at 37 degrees C, and homogenized for inoculation into the testis and peritoneum of hamsters. The animals were killed from week 6 to week 20 postinoculation and fragments of liver, lung, spleen, testis, and lymph nodes were cultured on brain heart infusion agar at 37 degrees C. Paracoccidioides brasiliensis was isolated from three armadillos both by direct organ culture and from the liver, spleen, lung, and mesenteric lymph nodes of hamsters. In addition, one positive armadillo presented histologically proven PCM disease in a mesenteric lymph node. The three armadillos isolates (Pb-A1, Pb-A2, and Pb-A4) presented thermodependent dimorphism, urease activity, and casein assimilation, showed amplification of the gp43 gene, and were highly virulent in intratesticularly inoculated hamsters. The isolates expressed the gp43 glycoprotein, the immunodominant antigen of the fungus, and reacted with a pool of sera from PCM patients. Taken together, the present data confirm that armadillos are a natural reservoir of P. brasiliensis and demonstrate that the animal is a sylvan host to the fungus.