Two novel mutations of pfdhps K540T and I588F, affecting sulphadoxine-pyrimethamine-resistant response in uncomplicated falciparum malaria at Banjar district, South Kalimantan Province, Indonesia

Background: Mutations in pfdhfr and pfdhps genes have been shown to associate with sulphadoxine-pyrimethamine (SP) resistance of Plasmodium falciparum parasites. However, pfdhfr, pfdhps genotypes and the correlations to SP treatment outcome in Indonesia has not yet been well analysed. Methods. After obtaining informed consent, 61 uncomplicated falciparum malaria patients were recruited in Banjar district, South Kalimantan Province, Indonesia, from October 2009 to August 2010. They were treated by a single oral dose of SP and its effects on clinical and parasitological status were followed until day 28 after treatment. Occasionally, a thick smear blood film for microscopy observation and blood spot on a filter paper for pfdhfr and pfdhps genotype analysis were collected. Results: Pfdhfr and pfdhps genotypes from 24 P. falciparum-infected patients consisting of adequate clinical parasitological response (ACPR) (n = 6; 25.0%) and early treatment failure (ETF) (n = 10; 41.7%) or late parasitological failure (LPF) (n = 8; 33.3%) were obtained by sequencing. Two novel mutations of pfdhps gene, K540T and I588F, were determined in ten and five isolates, respectively. These mutations were present in the pfdhfr/pfdhps combined haplotypes of ANRNI/SGTGA (n = 6), ANRNL/SGTGA (n = 4), and ANRNI/SGEAA(588F) (n = 5), (mutation codons are bold typed); these haplotypes were mostly belonging to parasitological failure (ETF or LPF). The parasites acquiring five mutations in pfdhfr/pfdhps haplotypes and four mutations with additional I588F did not respond adequately to SP treatment. Conclusion: Many of Plasmodium falciparum infected patients in Banjar district, South Kalimantan, Indonesia did not respond adequately to SP treatment and these low ineffectiveness of SP in this area was associated with two novel mutations of pfdhps, K540T and I588F

Polymorphism of Plasmodium Falciparum Dihydrofolate Reductase and Dihydropteroate Synthase Genes Among Pregnant Women with Falciparum Malaria in Banjar District, South Kalimantan Province, Indonesia

Pregnant women are highly vulnerable to malaria infection in its endemic areas, particularly infection by Plasmodium falciparum that can cause premature, low birth weight, severe anemia in pregnant women, and death. Sulfadoxine-pyrimethamine (SP) for Intermittent Preventive Treatment for pregnant (IPTp) is used for malaria control in pregnancy recommended by the World Health Organization that has already been implemented in Africa. The P. falciparum resistance to SP has been reported in several malarial endemic areas, and mutations in the genes of Plasmodium falciparum Dihydrofolate Reductase (Pfdhfr) and Dihydropteroate Synthase (Pfdhps) are shown to be associated with parasite resistance to SP treatment. Genetic analysis of Pfdhfr and Pfdhps genes in pregnant women infected with P. falciparum has not yet been examined in Indonesia. The cross-sectional study was conducted at two subdistricts, Sungai Pinang and Peramasan, in Banjar district of South Kalimantan Province, where 127 pregnant women were recruited from 2008 to April 2010. Two important mutations in Pfdhfr gene (amino acid positions at N51 and S108) and three in Pfdhps gene (A437, K540 and A581) were analyzed by nested PCR-RFLP method. All of the seven pregnant women samples infected with P. falciparum presented PfDHFR 108N and PfDHPS 437G mutations. One of the samples had the additional mutation at PfDHPS 540, in which Lys is substituted by Glu. These results suggested that P. falciparum might present only some resistance to SP at Sungai Pinang and Peramasan subdistricts, Banjar District, South Kalimantan province, Indonesia. Although there were limited number of samples, this study showed only few mutations of Pfdhfr and Pfdhps genes in P. falciparum at Banjar district, South Kalimantan Province, that suggests SP might be effective for IPTp in this area. Thus, further analysis of the other mutation sites in Pfdhfr and Pfdhps genes and in vivo efficacy study of SP with more sufficient sample numbers will be necessary to confirm this preliminarily result

Incidence and mutation analysis of glucose-6-phosphate dehydrogenase deficiency in eastern indonesian populations

We conducted a field survey of glucose-6-phosphate dehydrogenese (G6PD) deficiency in the eastern Indonesian islands, and analyzed G6PD variants molecularly. The incidence of G6PD deficiency in 5 ethnic groups (Manggarai, Bajawa, Nage-Keo, Larantuka, and Palue) on the Flores and Palue Islands was lower than that of another native group, Sikka, or a nonnative group, Riung. Molecular analysis of G6PD variants indicated that 19 cases in Sikka had a frequency distribution of G6PD variants similar to those in our previous studies, while 8 cases in Riung had a different frequency distribution of G6PD variants. On the other hand, from field surveys in another 8 ethnic groups (Timorese, Sumbanese, Savunese, Kendari, Buton, Muna, Minahasa, and Sangirese) on the islands of West Timor, Sumba, Sulawesi, Muna and Bangka, a total of 49 deficient cases were detected. Thirty-nine of these 49 cases had G6PD Vanua Lava (383T＞C) of Melanesian origin. In our previous studies, many cases of G6PD Vanua Lava were found on other eastern Indonesian islands. Taken together, these findings may indicate that G6PD Vanua Lava is the most common variant in eastern Indonesian populations, except for Sikka

Ekspresi MRNA Interleukin-10 (IL-10) dalam Kaitannya dengan Patogenesis Malaria Berat pada Mencit Strain BALB/C yang Diinfeksi Plasmodium Yoelli 17XL

Patogenesis malaria berat masih belum diketahui menyeluruh. Episode malaria berat dapat disebabkan oleh dua keadaan, produksi tinggi pada Fase awal IL-10 dan atau kurangnya produksi IL-10 pada Fase transisi. Peran IL-10 pada malaria masih belum jelas diketahui. Penelitian ini bertujuan untuk menganalisis ekspresi IL-10 dalam kaitannya dengan patogenesis malaria berat pada mencit strain BALB/c yang diinfeksi P. yoelii 17XL.Penelitian eksperimental laboratoris dengan rancangan “post test only control group design” dilakukan, dan menggunakan 24 mencit strain BALB/c, betina, usia 7-8 minggu, yang dibagi menjadi 4 kelompok. Mencit BALB/c diinfeksi secara intraperitoneal dengan 1x105 P. yoelii 17XL, dan dikorbankan pada hari ke-3 dan ke-6 pasca infeksi. Parasitemia dan kadar hemoglobin diperiksa setiap hari. Jaringan limpa diambil untuk isolasi RNA. Ekspresi mRNA IL-10, TNFα, dan IFNγ dianalisis dengan RT-PCR. Mencit BALB/c dinfeksi dengan 1x105 P. yoelii 17XL menunjukan infeksi letal, yang ditandai dengan peningkatan parasitemia sejalan dengan penurunan kadar hemoglobin, terjadi setelah hari ke-3 pasca infeksi. Ekspresi mRNA IL-10, begitu pula dengan TNFα dan IFNγ pada hari ke-3 paska infeksi menunjukan peningkatan dibandingkan pada kontrol dan hari ke-6 paska infeksi. Tidak adanya ekspresi IL-10 pada H6, menunjukkan kemungkinan adanya kegagalan regulator mengontrol malaria berat pada infeksi P. yoelii 17XL pada mencit BALB/c. Kata kunci— mRNA IL-10, RT-PCR, patogenesis malaria berat, mencit BALB/c yang diinfeksi P. yoelii 17X

Experimental Models Point Mutations in Plasmodium Falciparum Pfatpase6 Gene Exposed to Recuring Artemisinin in Vitro

The aims of this research to prove that repeated exposure of artemisinin can cause pfatpase6 gene mutation on Plasmodium falciparum in vitro. The research methods used culture In Vitro Plasmodium falciparum of strain 2300 IC50 value determination test artemisinin, artemisinin repeated exposure test (PO1, PO2, PO3 dan PO4) dose IC50, DNA extraction, gene amplification of pfatpase6 using Polymerase Chain Reaction (PCR) technique, electrophoresis, PCR product purification, labeling DNA from PCR results, DNA precipitation of PCR product, application of product labeling on the sequencing machines, analysis of the results of sequencing, and Data Analysis. The results of PCR pfatpase6 gene amplification include region 6 – 3216 for codon 89-1031 located in exon 1 and 2 Plasmodium falciparum 2300 by using five pairs of primers. Primer pair 1FR produce a long amplicon of 737 bp which covers of codon 89; primer pair 2FR produce a long amplicon of 813 bp which covers of codon 263, 431; primer pair 4FR produce a long amplicon of 700 bp which covers of codon 460, 465, 623; primer pair 5FR produce a long amplicon of 550 bp which includes of codon 683, 769; and primer pair 6FR produce a long amplicon of 876 bp which covers of codon 898, 1031.Multialigment pfatpase6 gene Plasmodium falciparum of strains Papua 2300 point mutations are obtained in the form of transition and transversion in treatment groups at the same nucleotide region 123, 2035, 2043, 2138 dan 2148. Conclusion of this research Artemisinin repeated exposure can cause point mutations in pfatpase6 genes Plasmodium falciparum of strains 2300 in vitr

SCIENTIFIC NOTE FIRST RECORD OF ANOPHELES BALABACENSIS FROM WESTERN SUMBAWA ISLAND, INDONESIA

ABSTRACT. An anopheline mosquito surveillance was conducted in the malaria endemic areas of Utan Rhee and Lunyuk counties, eastern Sumbawa Island, in 2004 and 2005. Eight species of Anopheles were collected, including a new record of An. balabacensis on the island

インドネシア・ロンボク島のマラリア浸淫地におけるハマダラカ蚊相とマラリア媒介蚊の究明

A systematic mosquito collection was carried out for 3 years from November 2001 to September 2004 in western Lombok Island, Indonesia to clarify Anopheles fauna, and to confirm vector species in malaria endemic areas. Adult mosquitoes were collected at 14 sites in the study area by using double-walled mesh nets with human or cow bait. A total of 11 species were encountered. Anopheles vagus was the most predominant. The second most abundant species differed among the sub-study areas; An. sundaicus was abundant in the coastal plain area, and An. balabacensis in the mountainous area. Anopheles balabacensis showed high anthropophily and exophagy and An. sundaicus moderate anthropophily and exophagy. Malaria parasite detection from the collected mosquitoes was also carried out through the detection of circumsporozoite protein by the VecTest^. Fourteen and 4 samples, which were positive for Plasmodium falciparum and P. vivax antigen respectively, were found from An. subpictus, An. sundaicus and An. balabacensis. We conclude that malaria in the coastal plain area is transmitted by An. sundaicus and An. subpictus, whereas An. balabacensis is the primary vector in the mountainous area of Lombok Island.2001年11月から2004年9月までインドネシア・ロンボク島・ムニンティング郡においてマラリア媒介蚊調査を行った.合計11種のハマダラカを沿岸,丘陵地,平地,山間部の異なる4地域で採取した.沿岸と山間部で採取した10種について,Vectestを用いてマラリア原虫抗原検出試験を行った結果,森林部で採取したAn.balabacensis,沿岸部で採取したAn.sundaicusとAn.subpictusから陽性反応が得られた.特にAn.balabacensisから多くの陽性反応を得た.これら3種は屋外吸血性,人嗜好性が認められ,マラリア患者との発生時期や分布地域とも一致したため,マラリア媒介蚊と判断した.ロンボク島でのAn.balabacensisによるマラリア媒介は,本研究が初めて明らかにした