Chloroquine-resistant Plasmodium vivax in transmigration settlements of West Kalimantan, Indonesia.

Malariometric surveys were conducted during July 1996 in native Dayak villages and predominantly Javanese transmigration settlements in Ketapang district of West Kalimantan, Indonesia. Malaria prevalence ranged from 0.9% to 2.7% in Dayak villages and from 1% to 20% in the transmigration settlements. Plasmodium falciparum accounted for 67% of the cases among Dayaks but P. vivax was dominant among transmigrants, accounting for more than 72% of the infections. Chloroquine sensitivity/resistance was assessed by 28-day in vivo testing of uncomplicated malaria infections and measurement of chloroquine blood levels in cases where parasitemias reappeared within the 28-day test period. Resistance was based on the appearance of asexual parasites against chloroquine plus desethylchloroquine levels exceeding the minimally effective whole blood concentrations proposed for sensitive parasite strains (P. vivax, 100 ng/ml; P. falciparum, 200 ng/ml). All parasitemias cleared initially within four days of beginning supervised chloroquine therapy (25 mg base/kg over a 48-hr period), but asexual parasites reappeared within 28 days in 27 of 52 P. vivax and three of 12 P. falciparum cases. Chloroquine blood levels at the time of recurrent parasitemias revealed resistance in 12 of the 27 P. vivax cases and in one of the three P. falciparum cases. Genotypes of nine of the 12 recurrent P. vivax isolates matched with their primary isolates and ruled out reinfection. These findings establish the presence of chloroquine-resistant P. vivax on the island of Borneo. The pattern of malaria and the high frequency of chloroquine resistance by P. vivax at the West Kalimantan location may relate to demographic, ecologic, agricultural, and socioeconomic changes associated with transmigration.

Survey of in vivo sensitivity to chloroquine by Plasmodium falciparum and P. vivax in Lombok, Indonesia.

A malariometric survey was conducted in 14 villages of Sekotong district, in Lombok, Indonesia during October 1994. Point prevalence of malaria ranged from 0% to 15% in the surveyed villages, averaging 6% overall, and Plasmodium falciparum accounted for 63% of the infections. Forty-nine patients with uncomplicated malaria and parasite counts ranging from 40 to 10,800 asexual forms/microliter were enrolled in a 28-day in vivo test of chloroquine sensitivity. All subjects received a supervised therapeutic regimen of chloroquine (25 mg base/kg over a 48-hr period) and parasitemia and symptoms were closely monitored for 28 days. Asexual parasites were eliminated within four days in the 29 P. falciparum and 20 P. vivax study patients enrolled. The cumulative incidence of therapeutic failure (recurrent symptomatic parasitemia) among P. falciparum cases at days 7, 14, and 28 was 7%, 10%, and 14% (4 of 29), respectively. However in all four cases, parasitemias recurred against chloroquine blood levels below the minimally effective concentration (MEC) of 200 ng/ml and do not confirm chloroquine resistance. All 20 P. vivax parasitemias were sensitive to chloroquine and the blood remained clear, with the exception of one case in which an asymptomatic parasitemia appeared on day 28. Parasitemias by P. falciparum and P. vivax that were observed before supervised therapy, but in the presence of whole blood chloroquine above normally suppressive MEC levels, suggest resistance to suppressive or prophylactic regimens of chloroquine.

Comparative performance of the ParaSight F test for detection of Plasmodium falciparum in malaria-immune and nonimmune populations in Irian Jaya, Indonesia.

A comparison was made of the performance of the ParaSight F test (F test) for detection of Plasmodium falciparum in blood from malaria-immune (410 native Irianese) and nonimmune (369 new transmigrants) populations in Irian Jaya, Indonesia, where malaria is hyperendemic and all four species of human malaria occur. There were highly significant differences between populations in the sensitivity (Irianese, 60% versus transmigrants, 84%; P < 0.001) and specificity (Irianese, 97% versus transmigrants, 84%; P < 0.001) of the F test. The test had comparably high levels of sensitivity for Irianese children aged < or = 10 years, both age groups of transmigrants (76-85%), but low sensitivity for Irianese aged > 10 years (40%), among whom only 7% of parasitaemias < 120 per microliter and 69% of those > 120 per microliter were detected. Specificity was comparably high for transmigrant children aged < or = 10 years and both age groups of Irianese (93-98%). The low specificity for transmigrants aged > 10 years (79%) was due to a preponderance of false positives, frequently identified by microscopy as P. vivax. The results suggest that comparison based on microscopy underestimated the performance of the ParaSight F test and that malaria immune status, irrespective of P. falciparum density, may influence the test's sensitivity.

PIP: The ParaSight F test uses a nonmicroscopic dipstick approach to the rapid detection of Plasmodium falciparum in blood. The performance of this test was assessed in serum samples collected in Irian Jaya, Indonesia, from 410 native Irianese (malaria-immune) and 369 new transmigrants (nonimmune). Of particular interest was the capability of the F test to detect P. falciparum prevalence among children, whose immunity is less than that of adults. There were highly significant differences by population in the F test's sensitivity (60% for Irianese vs. 84% for transmigrants) and specificity (97% for Irianese vs. 84% for transmigrants). The test had high sensitivity levels (76-85%) for Irianese children 10 years of age and under and both child and adult transmigrants, but low sensitivity (40%) for Irianese over 10 years of age. Specificity was comparably high (93-98%) for transmigrant children and both age groups of Irianese. The low specificity (79%) for transmigrants over 10 years of age reflected a preponderance of false positives, frequently identified by microscopy as P. vivax. These findings suggest that microscopy comparisons underestimate the performance of the ParaSight F test and that malaria immune status, regardless of P. falciparum density, may influence the test's sensitivity.

Humoral immune responses in hamsters infected with Opisthorchis viverrini.

The kinetics and nature of humoral immune responses to somatic and excretory-secretory (ES) antigens were investigated in hamsters experimentally infected with different numbers of Opisthorchis viverrini. ES antigens were obtained from the in vitro culture of adult flukes and somatic antigens were aqueous extracts of adult flukes. Antibodies in the serum and bile of infected animals were determined by the microhaemagglutination technique, using glutaraldehyde fixed sheep red blood cells sensitized with these parasite antigens. Antibody responses to both somatic and ES antigens were detected in the serum from the second week of infection onward. The peak response was noted at the end of the second month and declined slowly thereafter. Antibody levels in animals with heavy infections (100 metacercariae) appeared earlier but declined more rapidly than in animals with light infections (25 metacercariae). The serum antibodies were highly sensitive to mercaptoethanol throughout the course of infection (23 weeks). Antibodies also appeared in the bile obtained at the time of sacrifice but their titres were rather low compared with those in the serum. Like serum antibodies, biliary antibodies were reactive with both somatic and ES antigens. Biliary antibodies were of the secondary IgA type. These findings are discussed in relation to pathogenesis of the disease process and to the possible usefulness in immunodiagnosis.

Attempts to induce protective immunity in hamsters against infection by a liver fluke of man (Opisthorchis viverrini).

The development of acquired resistance in opisthorchiasis was studied in hamsters experimentally infected with Opisthorchis viverrini. The induction of protective immunity was attempted by first exposing adult female golden Syrian hamsters to 1, 2 or 3 doses of infective metacercariae obtained from naturally infected cyprinoid fishes and then reinfecting them with 80 metacercariae. In other experiments, animals that were infected with 50 metacercariae were treated with praziquantel prior to being rechallenged in order to eliminate the flukes that had developed from the first infection. The effect of long-term chronic infections was also studied. Faecal egg counts were determined at weekly intervals from 4-5 weeks onwards. The animals were killed 2-3 months after the last infection for worm recovery, and terminal faecal egg output/g faeces/worm was calculated. The data showed that prior infection of animals with O. viverrini did not induce significant protective immunity against reinfection by the same parasite. Lack of protection was also noted in animals reinfected several times with small doses of metacercariae. However, under certain circumstances, prior infection could result in a significant reduction in the faecal egg output due to subsequent infection.

Effect of culture media on production of excretory-secretory products and egg output of Opisthorchis viverrini in vitro.

Adult Opisthorchis viverrini were maintained metabolically active in vitro for an extended period in Earle's basal medium (BME). The worms were most active during the first 7 to 10 days of incubation and the culturing fluids were found to react in immunodiffusion with antisera. The mean survival period and egg output per worm per day could be enhanced when BME was supplemented with 5% normal, human bile but not with the same concentration of bile ultrafiltrate. A similar degree of enhancement was obtained when the medium was supplemented with 1% normal human or hamster sera. The enrichment effect of human serum could be partially replaced by albumin. However, the results obtained suggested that another serum or biliary macromolecular component(s) was required to maintain the flukes metabolically active in vitro for an extended period.

Effect of treatment with diethylcarbamazine on immune responses to filarial antigens in patients infected with Brugia malayi.

We studied the effect of treatment with diethylcarbamazine (DEC) on immune responses to parasite antigens in humans infected with Brugia malayi. In vitro lymphocyte proliferative responses to microfilarial antigens increased in patients who became amicrofilaremic after treatment with DEC. No changes in reactivity were observed in amicrofilaremic individuals who were given DEC or in a small number of patients who remained microfilaremic after treatment. Reactions to other antigens (PPD and SKSD) were not affected by drug treatment. Serum titers of antibodies to the sheath of B. malayi microfilariae did not significantly change during the period of observation. These findings indicate that DEC partially reverses the state of cellular unresponsiveness to parasite antigens associated with patient filarial infections.

Brugia malayi: relationship between anti-sheath antibodies and amicrofilaremia in natives living in an endemic area of South Kalimantan, Borneo.

A seroepidemiological approach was taken to elucidate the relationship between anti-microfilarial antibodies and amicrofilaremia in humans living under natural conditions of exposure to Brugia malayi. Entomological observations indicated that all of the people in the study population in South Kalimantan, Borneo, were exposed repeatedly to filarial infection. A third of the population had antibodies to the sheath of microfilariae. The prevalence and titer of anti-sheath IgM was higher than anti-sheath IgG or IgA. There was a statistically significant correlation between anti-sheath antibody and amicrofilaremia and these antibodies may play a role in regulating peripheral microfilaremia.

Antigen-specific suppressor cells and suppressor factors in human filariasis with Brugia malayi.

We investigated the mechanisms of specific immune unresponsiveness to microfilarial antigens. The blood of patients with obvious Brugia malayi infections contains an adherent cell type that specifically suppresses reactions to microfilarial antigens but not to other antigens. In the absence of continued stimulation by parasite antigens, this suppressor cell loses its functional activity after overnight culture in vitro. Furthermore, serums from patients with and without microfilaremia contain factors that also suppress reactions to filarial antigens in vitro. These results suggest that immune unresponsiveness in human beings with patent filarial infections is due to active suppression of immune responses directed against the parasite and not to an intrinsic inability of infected patients to react to parasite antigens.