Devices for rapid diagnosis of Malaria: evaluation of prototype assays that detect Plasmodium falciparum histidine-rich protein 2 and a Plasmodium vivax-specific antigen

The ParaSight F test was developed as a pioneer industry effort in the large-scale, process-controlled production of a device for the rapid diagnosis of malaria. This device performed well in field settings but was limited to the detection of a single malaria species, Plasmodium falciparum. The ParaSight F+V assay advanced upon the ParaSight F test format by incorporating a monoclonal antibody directed against a proprietary Plasmodium vivax-specific antigen, in addition to the antibody directed against P. falciparum histidine-rich protein 2, which was used in the ParaSight F assay. The modified assay was developed to add the capability to detect P. falciparum and P. vivax in a single-test-strip format. The present study evaluated three distinct ParaSight F+V prototypes with samples from symptomatic patients in regions of Thailand and Peru where malaria is endemic. Over a 2-year enrollment period (1998 and 1999), a total of 4,894 patients consented to participation in the study. Compared with the results for duplicate microscopic examinations of Giemsa-stained blood smears as the reference diagnostic standard, each successive prototype showed substantial improvement in performance. The final ParaSight F+V prototype, evaluated in 1999, had an overall sensitivity for detection of asexual P. falciparum parasites of 98%. The sensitivity of the device was 100% for P. falciparum densities of >500 parasites/ micro l, with a sensitivity of 83% for parasite densities of 5,000/ micro l, 92% for parasite densities of 1,001 to 5,000/ micro l, 94% for parasite densities of 501 to 1,000/ micro l, and 55% for parasite densities of 1 to 500/ micro l. The specificity for the exclusion of P. vivax was 87%. The areas under the receiver operating characteristic curves for the diagnostic performance of the assay for the detection of P. falciparum and P. vivax were 0.8907 and 0.8522, respectively. These findings indicate that assays for rapid diagnosis have the potential to enhance diagnostic capabilities in those instances in which skilled microscopy is not readily available.     

Emerging rickettsioses of the Thai-Myanmar border

To investigate the presence of rickettsioses in rural residents of the central Thai-Myanmar border, we tested the blood of 46 patients with fever. Four patients had murine typhus, three patients had scrub typhus, and eight patients had spotted fever group rickettsioses, including the first case of Rickettsia felis infection reported in Asia.     

Drug-Resistant Malaria: The Era of ACT

As drug-resistant falciparum malaria has continued to evolve and spread worldwide, artemisinin-based combination therapies (ACT) have become the centerpiece of global malaria control over the past decade. This review discusses how advances in antimalarial drug resistance monitoring and rational use of the array of ACTs now available can maximize the impact of this highly efficacious therapy, even as resistance to artemisinins is emerging in Southeast Asia.     

In vivo sensitivity monitoring of mefloquine monotherapy and artesunate-mefloquine combinations for the treatment of uncomplicated falciparum malaria in Thailand in 2003

OBJECTIVE: To monitor the efficacy of anti-malarial treatments in Thailand. METHOD: A 28-day in vivo study in nine provinces along international borders in 2003. The first group comprised 164 patients from four provinces: Mae Hong Son, Chiang Mai, Ratchaburi and Ubon Ratchathani. These patients received 15 mg/kg mefloquine as a single dose. The second group, 58 patients from Kanchanaburi, were treated with 15 mg/kg mefloquine plus artesunate (12 mg/kg). The third group, 196 patients from provinces with high-level mefloquine resistance (Tak, Ranong, Chanthaburi and Trat), received 25 mg/kg of mefloquine plus 12 mg/kg artesunate. In all arms, follow-up blood smears were scheduled for days 1, 2, 3, 7, 14, 21 and 28. All patients tolerated the regimens well. RESULTS: The percentage of adequate clinical and parasitological response to mefloquine monotherapy was 62.0% in Mae Hong Son, 75.0% in Chiang Mai, 94.0% in Ratchaburi and 89.7% in Ubon Ratchathani. In Kanchanaburi, the percentage of adequate clinical and parasitological response to the artesunate-mefloquine combination was 94.2%. In the third group, this response exceeded 90%, except in Trat, where it was only 78.6% (44 patients). CONCLUSION: Mefloquine monotherapy must urgently be replaced in Mae Hong Son and Chiang Mai. The markedly reduced efficacy of the artesunate-mefloquine combination used in Trat raises questions about the future of this therapy on the southeastern border of Thailand with Cambodia. It is very worrying because no practical and affordable alternative is yet available.     

Sources of variability in determining malaria parasite density by microscopy

Enumeration of parasites by microscopic examination of blood smears is the only method available for quantifying parasitemia in infected blood. However, the sources and scale of error inherent in this technique have not been systematically investigated. Here we use data collected in outpatient clinics in Peru and Thailand to elucidate important sources of variation in parasite density measurements. We show that discrepancies between readings from two independent microscopists and multiple readings from a single microscopist are inversely related to the density of the infection. We present an example of how differences in reader technique, specifically the number of white blood cells counted, can contribute to the differences between readings. We discuss the implications of this analysis for field studies and clinical trials.     

Rapid diagnostic testing for malaria

Malaria rapid diagnostic devices (MRDD) have been developed with the hope that they would offer accurate, reliable, rapid, cheap and easily available alternatives to traditional methods of malaria diagnosis. The results from early malaria rapid diagnostic studies were quite promising, especially for detecting Plasmodium falciparum at densities of more than 100-500 parasites/microl. Despite the introduction of these devices over a decade ago, only a few target antigens have been introduced. Of greater concern, these devices have shown limitations in sensitivity, ability to differentiate species and robustness under field conditions in the tropics. Recent trials have revealed wide variability in sensitivity both within and between products. We review the recent trials assessing MRDD use for the diagnosis of P. falciparum and non-P. falciparum infections in endemic and non-endemic countries and describe the various aspects of these devices which need further improvement. High quality, accurate, rapid and affordable diagnostic tools are urgently needed now that new antimalarial regimens, characterized by higher cost and increased toxicity, have been introduced more widely in response to emerging multi-drug resistance.     

Bartonella species in rodents and shrews in the greater Jakarta area

In February 2004, we captured 221 rodents and shrews in the Greater Jakarta area as part of a study to determine the prevalence of rodent-associated vector-borne infections. Microscopic examination of blood smears revealed 6% (13/218) to be positive for Bartonella spp. The corresponding DNA samples, either from blood blots or frozen spleen pieces and from fleas collected on these animals, were tested for evidence of Bartonella infection by PCR, targeting the portions: 378bp and 930bp of the citrate synthase gene (g/tA). The sequences from our sample clusters with a Peruvian entity, B. phoceensis, B. rattimassiliensis and B. elizabethae, the latter species has been associated with endocarditis and neuroretinitis in humans. As previous analyses have shown, there appears to be little geographic or host consistency with phylogenetic placement. The public health significance of these findings remains to be determined.     

Efficacy of the merozoite surface protein 1 of Plasmodium vivax as an antigen for ELISA to diagnose malaria

Malaria is still a major health problem in Thailand and its incidence is currently rising in Korea. To identify a useful antigen for the diagnosis of malaria patients, a cDNA expression library from malaria parasites was constructed and screened out immunologically. One clone was selected in view of its predominant reactivity with the patient sera. The recombinant malaria parasite antigen (Pv30) with 27 kDa as a C-terminal His-tag fusion protein that was produced in Escherichia coli was identified through immunoblot analysis. The deduced amino acid sequence had the sequence homology with the merozoite surface protein 1 (MSP1) genes of Plasmodium falciparum and P. yoelii, each by 41% and 42%, respectively. Measurement of serum IgG and IgM antibody to Pv30 by enzyme-linked immunosorbent assay (ELISA) was evaluated as a serodiagnostic test for malaria patients in Thailand (endemic area) and Korea (recently reemerging area). The sensitivity of P. vivax, P. falciparum, and P. malariae was 96.3% (26 /27), 90.6% (29/32), and 100% (6/6), respectively, and the specificity was 63.5% (40/63) in Thailand samples. The sensitivity of P. vivax was 98.8% (88/89), and the specificity was 96.6% (86/89) in Korean samples. Pv30 appears to be a good and reliable recombinant antigen for serodiagonosis of malaria in a nonendemic area.     

Hematologic and clinical indices of malaria in a semi-immune population of western Thailand

This study examines hematologic profiles of persons with acute Plasmodium falciparum or P. vivax infection in Maesod on Thailand's western border with Myanmar compared with febrile, non-parasitemic persons also reporting to malaria clinics. Nine hundred seventy-nine subjects were malaria-negative, 414 were infected with P. falciparum, and 646 were infected with P. vivax. Persons with patent parasitemia tended to have significantly lower white blood cell, red blood cell, platelet, and hemoglobin levels than those who were malaria-negative. For the first time, a parallel trend in thrombocytopenia with parasitemia was found to be associated with both P. falciparum, and P. vivax infection. Using logistic regression, persons with platelet counts < 150,000/microL were 12-15 times more likely to have malaria than persons with platelet counts > or = 150,000/microL. This study supplements previous literature on the hematologic effects of malaria and helps define those alterations for a semi-immune population. Thrombocytopenia is identified as a key indicator of malaria in these febrile patients.