High-resolution melt analysis for the detection of a mutation associated with permethrin resistance in a population of scabies mites.

Permethrin as a topical acaricide cream is widely used to treat scabies. The neuronal voltage-sensitive sodium channel (Vssc), necessary for the generation of action potentials in excitable cells, is the target of pyrethroid acaricides such as permethrin. Pyrethroid resistance has been linked to specific mutations in the Vssc gene. Following the partial sequencing of the Vssc gene in the scabies mite Sarcoptes scabiei (L.) (Astigmata: Sarcoptidae), we compared Vssc gene sequences from permethrin-sensitive and -tolerant S. scabiei var. canis Gerlach mites, and identified a G to A single nucleotide polymorphism (SNP) in permethrin-tolerant mites resulting in an amino acid change from glycine to aspartic acid in domain III S6. The mutation is in a region of the gene where mutations have been identified in a range of pyrethroid-resistant arthropods. Results of in vitro permethrin exposure assays showed that survival rates for mites bearing the mutation were similar to those previously reported for mites from human subjects where clinical tolerance to permethrin had been observed. A real-time polymerase chain reaction-high-resolution melt (PCR-HRM) assay was developed to detect this SNP. This assay provides a useful methodology for screening for this and other mutations associated with permethrin resistance in scabies mite populations and thus facilitates surveillance for acaricide resistance.

Field evaluation of malaria rapid diagnostic tests for the diagnosis of P. falciparum and non-P. falciparum infections.

The objective of this study was to evaluate various malaria rapid diagnostic tests as a tool in the detection of P. falciparum and non-P. falciparum infections in field conditions. Four field surveys were conducted in malaria-endemic areas of Palawan and Davao del Norte, Philippines to validate the various rapid diagnostic tests, namely Diamed OptiMAL 48 (DiaMed AG, Switzerland), ParaHIT f (Span Diagnostics, India), Orchid OptiMAL, and Paracheck Pf (both from Orchid Biomedical Systems, India). The results of the various rapid diagnostic tests were compared to those of microscopy. Sensitivity, specificity and detection rates according to the level of parasitemia were used as parameters to describe the performance of the various rapid diagnostic tests in the field. Practical and operational assessments were also done. The results of the study show that the sensitivity and detection rates were generally lower than previously reported, with sensitivities ranging from 4.8% to 20.6%, except for Diamed OptiMAL 48, which had sensitivities of 78.8% to 96.8%, and detection rates of 50.0% to 96.8%. The rest had detection rates ranging from 0.0% to 50.0%. All the specificities ranged from 18.2% to 100.0%. Improper conditions at the time of manufacturing, storage, transport, and utilization may affect the validity of the results. Rapid diagnostic tests for malaria provide practical means of detecting malarial infections, especially in endemic areas. However, issues regarding variability in performance must to be addressed before they can be used as mainstream diagnostic tools.

Plasmodium vivax synonymous substitution frequencies, evolution and population structure deduced from diversity in AMA 1 and MSP 1 genes.

Polymorphic regions of the genes encoding Plasmodium vivax apical membrane antigen 1 (PvAMA1) and P. vivax merozoite surface protein 1 (PvMSP1) were sequenced to examine population diversity both within and between geographical areas. Sequences were obtained for 219 isolates for PvAMA1 and for 175 isolates for PvMSP1 from Africa, China, India, Indonesia, Philippines, Papua New Guinea, Solomon Islands and Thailand. Over half of the isolates were obtained from different regions within the Philippines, and this was used to look at the diversity within a country. Sixty nine haplotypes and 22 polymorphic sites in a 414-bp region of PvAMA1 and 41 haplotypes and 34 polymorphic sites in a 249-bp fragment of PvMSP1 were detected. For both PvAMA1 and PvMSP1, four previously unreported polymorphic nucleotide positions were identified. Population analysis indicated that there were significant differences in allele frequencies between different regions but these differences were small compared to the diversity within populations (Fixation index, F(ST), of 0.126 and 0.078 for PvAMA1 and PvMSP1, respectively). PvAMA1 and PvMSP1 had similar nonsynonymous substitution frequencies but surprisingly, the synonymous substitution frequency for PvMSP1 was eight times the frequency for PvAMA1 suggesting that synonymous substitutions in at least PvAMA1 are not neutral.

Field epidemiological studies on malaria in a low endemic area in the Philippines.

Field epidemiological studies were conducted to examine factors affecting endemicity in an area with a low prevalence of malaria. Two annual cross sectional surveys were done to estimate parasite prevalence rates at two periods in time, to determine the distribution of the parasitemic population and to describe the serological status of the population. A longitudinal study of a sample of infected people was used to measure reinfection rates and antibody dynamics. A 2 year passive case detection was done to estimate the number and distribution of people with symptomatic infections. Malaria was found in all age groups, with marked clustering of cases. Active and passive case detection and serological surveys all gave a similar pattern of malaria distribution: generally low prevalence with small foci of relatively high endemicity. The infection frequencies were generally similar in all age groups, measured by both active and passive case detection. There was a high frequency of P. falciparum gametocytemic infections in the asymptomatic cases found through active case detection. Twenty to 39 year old males had the highest frequency of infection by active case detection, and 10-19 year old males by passive case detection. These two groups were also more likely to be gametocyte positive than their female counterparts, suggesting that in this community, this portion of the population acts as the main reservoir of infection.