In vitro atovaquone/proguanil susceptibility and characterization of the cytochrome b gene of Plasmodium falciparum from different endemic regions of Thailand

<p>Abstract</p> <p>Background</p> <p>The emergence of <it>Plasmodium falciparum </it>resistant to most currently used antimalarial drugs is the major problem in malaria control along the Thai-Myanmar and Thai-Cambodia borders. Although artemisinin-based combination therapy has been recommended for the treatment of multidrug-resistant falciparum malaria, these combinations are not available for some people, such as travelers from North America. A fixed-dose combination of atovaquone and proguanil (Malarone) has been proved to be effective for the treatment and prophylaxis of malaria which is already approved by countries in North America and Europe. Determination of the phenotypes and genotypes related to atovaquone/proguanil response in Thai isolates of <it>P. falciparum </it>will be useful for rationale drug use. The main purpose of this study was to explore the <it>in vitro </it>atovaquone/proguanil susceptibility of recently adapted Thai isolates of <it>P. falciparum</it>. Genotypic characterization of the <it>cytb </it>gene of these isolates was also determined since it has been reported that point mutations, particularly codon 268 in the cytochrome b gene (<it>cytb</it>) have been linked to atovaquone/proguanil treatment failure.</p> <p>Methods</p> <p>Eighty three <it>P. falciparum </it>isolates collected during 1998 to 2005 from four different multidrug resistance areas of Thailand were determined for the <it>in vitro </it>atovaquone/proguanil susceptibilities using radioisotopic assay. Mutations in the <it>cytb </it>gene were determined by PCR-RFLP and sequence analysis.</p> <p>Results</p> <p>The mean atovaquone and proguanil IC₅₀was 3.4 nM and 36.5 μM, respectively. All 83 Thai isolates were atovaquone sensitive. None of the 83 isolates contained the mutations at codon 268 of the <it>cytb </it>gene. DNA sequencing of the <it>cytb </it>gene of 20 parasite isolates showed no other mutations.</p> <p>Conclusion</p> <p>In agreement with a recent efficacy study of atovaquone/proguanil, the present information indicates that atovaquone/proguanil can be one of the drugs of choice for the treatment and prophylaxis of multidrug-resistant falciparum malaria in Thailand.</p

Box plots of atovaquone and proguanil ICof parasite isolates collected from different endemic areas

<p><b>Copyright information:</b></p><p>Taken from "atovaquone/proguanil susceptibility and characterization of the gene of from different endemic regions of Thailand"</p><p>http://www.malariajournal.com/content/7/1/23</p><p>Malaria Journal 2008;7():23-23.</p><p>Published online 28 Jan 2008</p><p>PMCID:PMC2265725.</p><p></p

Influence of the pfmdr1 Gene on In Vitro Sensitivities of Piperaquine in Thai Isolates of Plasmodium falciparum.

Piperaquine combined with dihydroartemisinin is one of the artemisinin derivative combination therapies, which can replace artesunate–mefloquine in treating uncomplicated falciparum malaria in Thailand. The aim of this study was to determine the in vitro sensitivity of Thai Plasmodium falciparum isolates against piperaquine and the influence of the pfmdr1 gene on in vitro response. One hundred and thirty-seven standard laboratory and adapted Thai isolates of P. falciparum were accessed for in vitro piperaquine sensitivity. Polymorphisms of the pfmdr1 gene were determined by polymerase chain reaction methods. The mean and standard deviation of the piperaquine IC50 in Thai isolates of P. falciparum were 16.7 ± 6.3 nM. The parasites exhibiting chloroquine IC50 of ≥ 100 nM were significantly less sensitive to piperaquine compared with the parasite with chloroquine IC50 of < 100 nM. No significant association between the pfmdr1 copy number and piperaquine IC50 values was found. In contrast, the parasites containing the pfmdr1 86Y allele exhibited significantly reduced piperaquine sensitivity. Before nationwide implementation of dihydroartemisinin–piperaquine as the first-line treatment in Thailand, in vitro and in vivo evaluations of this combination should be performed especially in areas where parasites containing the pfmdr1 86Y allele are predominant such as the Thai–Malaysian border

Polymorphisms of the <it>pfmdr1 </it>but not the <it>pfnhe-1 </it>gene is associated with <it>in vitro </it>quinine sensitivity in Thai isolates of <it>Plasmodium falciparum</it>

Abstract Background The emergence of Plasmodium falciparum resistance to most currently used anti-malarial drugs is a major problem in malaria control along the Thai-Myanmar and Thai-Cambodia borders. Quinine (QN) with tetracycline/doxycycline has been used as the second-line treatment for uncomplicated falciparum malaria. In addition, QN monotherapy has been the first-line treatment for falciparum malaria in pregnant women. However, reduced in vitro and in vivo responses to QN have been reported. To date, a few genetic markers for QN resistance have been proposed including Plasmodium falciparum chloroquine resistance transporter (pfcrt), P. falciparum multidrug resistance 1 (pfmdr1), and P. falciparum Na+/H+ exchanger (pfnhe-1). This study was to investigate the role of the pfmdr1 and pfnhe-1 gene on in vitro QN sensitivity in Thai isolates of P. falciparum. Methods Eighty-five Thai isolates of P. falciparum from the Thai-Myanmar and Thai-Cambodia borders from 2003-2008 were determined for in vitro QN sensitivity using radioisotopic assay. Polymorphisms of the pfmdr1 and pfnhe-1 gene were determined by PCR-RFLP and sequence analysis. Associations between the in vitro QN sensitivity and the polymorphisms of the pfmdr1 and pfnhe-1 gene were evaluated. Results The mean QN IC50 was 202.8 nM (range 25.7-654.4 nM). Only four isolates were QN resistant when the IC50 of >500 nM was used as the cut-off point. Significant associations were found between the pfmdr1 mutations at codons N86Y and N1042D and in vitro QN sensitivity. However, no associations with the number of DNNND, DDNNNDNHNDD, and NHNDNHNNDDD repeats in the microsatellite ms4760 of the pfnhe-1 gene were identified. Conclusion Data from the present study put doubt regarding the pfnhe-1 gene as to whether it could be used as the suitable marker for QN resistance in Thailand. In contrast, it confirms the influence of the pfmdr1 gene on in vitro QN sensitivity.</p

Association between the pfmdr1 gene and in vitro artemether and lumefantrine sensitivity in Thai isolates of Plasmodium falciparum.

We evaluated the influence of pfmdr1 mutations and copy number on in vitro artemether and lumefantrine sensitivity in 101 laboratory and adapted Thai isolates of Plasmodium falciparum. Approximately one-fourth of these isolates exhibited reduced lumefantrine susceptibility. We found that both mutations and amplification of the pfmdr1 gene influenced in vitro artemether and lumefantrine sensitivity. Using multivariate analysis, 184F or 1042N alleles and a copy number of ≥ 4 were identified as the independent markers for decreased lumefantrine susceptibility. Separate analysis also indicated that parasites from different geographical areas were influenced by different genetic markers

Transcriptome analysis reveals pathogenicity and evolutionary history of the pathogenic oomycete Pythium insidiosum

Oomycetes form a unique group of microorganisms that share hyphal morphology with fungi. Most of pathogenic oomycetes infect plants, while some species are capable of infecting animals. Pythium insidiosum is the only oomycete that can infect both humans and animals, and causes a life-threatening infectious disease, called 'pythiosis'. Controlling an infection caused by P. insidiosum is problematic because effective antimicrobial drugs are not available. Information on the biology and pathogenesis of P. insidiosum is limited. We generated a P. insidiosum transcriptome of 26735 unigenes, using the 454 sequencing platform. As adaptations to increased temperature inside human hosts are required for a successful pathogen, we generated P. insidiosum transcriptomes at 28°C and 37°C and identified 625 up-regulated and 449 down-regulated genes at 37°C. Comparing the proteomes of oomycetes, fungi, and parasites provided clues on the evolutionary history of P. insidiosum. Potential virulence factors of P. insidiosum, including putative effectors, were identified. Pythium insidiosum harbored an extensive repertoire of ~300 elicitin domain-containing proteins. The transcriptome, presented herein, provides an invaluable resource for exploring P. insidiosum's biology, pathogenesis, and evolution.14 page(s

Autochthonous disseminated dermal and visceral leishmaniasis in an AIDS patient, southern Thailand, caused by Leishmania siamensis

We report the first establishment of in vitro cultivation and genotypic characterization of Leishmania siamensis isolated from an autochthonous disseminated dermal and visceral leishmaniasis in a Thai acquired immunodeficiency syndrome (AIDS) patient. The molecular identification has shown that the parasite was identical to L. siamensis, a recently described Leishmania species reported in the southern provinces of Thailand. The phylogenetic analysis has confirmed L. siamensis as closely related to the zoonotic Leishmania species L. enrietti