In silico antiplasmodial effects of phytocompounds derived from Andrographis paniculata on validated drug targets of different stages of Plasmodium falciparum

Background:Andrographis paniculata has been widely reported as an herbal plant for malaria treatment.The increasing rate of resistance to recommended antimalarial drugs has justified the need for a continuous search for new and more potent drugs that target all stages of the Plasmodium falciparum life cycle from natural plant sources.This study aimed to determine the antiplasmodial effect of phytocompounds derived from A.paniculata on the stages of plasmodium falciparum.Methods:Phytocompounds from A.paniculata were identified by Gas Chromatography-Mass Spectrophotometry(GCMS)analysis.The phytocompounds were screened for their druggability using Lipinski’s rule of five and subjected to Absorption,Distribution,Metabolism,Excretion,Toxicity(ADMET)and druglikeness analysis.The phytocompounds were docked against some validated drug targets at different stages of Plasmodium falciparum(hepatic,asexual,sexual,and vector targets)using PyRx software to analyze the inhibitory potential and protein-ligand interaction.Thereafter,the stability and flexibility of the best complexes were assessed through molecular dynamics simulations at 50ns using WebGRO.Result:The 7a-Isopropenyl-4,5-dimethyloctahydroinden-4-yl exhibited a higher binding affinity and better stability throughout the simulation period with P.falciparum dihydrofolate reductase-thymidylate synthase and Plasmodium falciparum M1 alanyl aminopeptidase for asexual blood stage and gametocyte stage of Plasmodium falciparum,respectively than the existing drugs.Meanwhile,N-Ethyl-3-methoxy-4-methylphenethylamine was also found to have a higher binding affinity and more stability throughout the simulation period with P.falciparum purine nucleoside phosphorylase and Plasmodium falciparum gametocyte surface protein for Hepatic schizonts stage of Plasmodium falciparum and gametocyte transmission blocking stage,respectively,than the existing drugs.Conclusion:The 7a-Isopropenyl-4,5-dimethyloctahydroinden-4-yl and N-Ethyl-3-methoxy-4 methylphenethylamine from A.paniculata are predicted as an antimalarial drug candidate.Thus,it is recommended that in vitro and in vivo bioassays be conducted on these hit compounds to validate these predictions.

Selective Effect of Qinghaosu on Different Stages of Plasmodium falciparum in Vitro

Using highly synchronous cultures of Plasmodium falciparum in vitro,the susceptibi- lity of the different stages of the intraerythrocytic parasites to Qinghaosu (QHS) was assessed.The anti- parasitic effect of QHS was measured by comparing the changes of irradiation of^3 H-hypoxanthine in- corporated into the nucleic acids of parasites exposed to various concentrations of QHS at different stages of growth.It was found that the trophozoite stage of the parasite was the most sensitive to QHS, whereas the early ring stage was the least sensitive,and the sensitivities of the late ring and schizont stages fell between those of the early ring and trophozoite stages.The results revealed the correlation of stage-dependent effects of QHS with the blockade of the protein metabolism of the parasite.

Polymorphisms of the oxidant enzymes glutathione S-transferase and glutathione reductase and their association with resistance of Plasmodium falciparum isolates to antimalarial drugs

Objective:To investigate the association between amplification of the two regulatory genes controlling glutathione(GSH) levels,glutathione reductase(PfGR) and glutathione S-transferase (PfGST) genes and sensitivity of Plasmodium falciparum(P.falciparum) isolates collected from different malaria endemic areas of Thailand to standard antimalarial drugs.Methods:A total of 70 P.falciparum isolates were collected from endemic areas of multi-drug resistance (Tak,Chantaburi and Ranong Provinces) during the year 2008-2009.The in vitro assessment of antimalarial activity of P.falciparum clones(K1- and Dd2 chloroquine resistant and 3D7- chloroquine sensitive) and isolates to chloroquine,quinine,mefloquine and arteusnate was performed based on SYBR Green modified assay.Results:68(97.14%),11(15.71%) and 28(40%) isolates respectively were classified as chloroquine-,quinine- and mefloquine-resistant isolates. With this limited number of P.falciparum isolates included in the analysis,no significant association between amplification of PfGST gene and sensitivity of the parasite to chloroquine, quinine,mefloquine and quinine was found.Based on PCR analysis,Dd2,Kl and 3D7 clones all contained only one copy of the PfGST gene.All isolates(70) also carried only one copy number of PfGST gene.There appears to be an association between amplification of PfGR gene and chloroquine resistance.The 3D7 and Dd2 clones were found to carry only one PfGR gene copy, whereas the K1 clone carried two gene copies.Conclusions:Chloroquine resistance is likely to be a consequence of multi-factors and enzymes in the GSH system may be partly involved. Larger number of parasite isolates are required to increase power of the hypothesis testing in order to confirm the involvement of both genes as well as other genes implicated in glutathione metabolism in conferring chloroquine resistance.

Predicted essential proteins of Plasmodium falciparum for potential drug targets

Objective:To identify novel drug targets for treatment of Plasmodium falciparum.Methods: Local BT.ASTP were used to find the proteins non-homologous to human essential proteins as novel drug targets.Functional domains of novel drug targets were identified by InterPro and Pfam.3D structures of potential drug targets were predicated by the SWISS-MODEL workspace. Ligands and ligand-binding sites of the proteins were searched by Ef-seek.Results:Three essential proteins were identified that might be considered as potential drug targets.AAN37254.1 belonged to 1-deoxy-D-xylulose 5-phosphate reductoisomerase,CAD50499.1 belonged to chorismale synthase,CAD51220.1 belonged to FAD binging 3 family,but the function of CAD51220.1 was unknown.The 3D structures,ligands and ligand-binding sites of AAM37254.1 and CAD50499.1 were successfully predicated.Conclusions:Two of these potential drug targets are key enzymes in 2-C-methyl-d-erythritol 4-phosphate pathway and shikimate pathway, which are absent in humans,so these two essential proteins are good potential drug targets.The function and 3D structures of CAD50499.1 is still unknown,it still need further study.

Comparative studies of serum-free media and detection techniques for <i>in vitro</i>drug sensitivity assessment of <i>Plasmodium falciparum</i>

Malaria continues to be a devastating disease. In a previous study, we formulated a chemically defined culture medium that is able to sustain the complete intraerythrocytic growth of Plasmodium falciparum. We tested the feasibility of using the medium (CDRPMI) as well as human serum-free media enriched with commercially available human-serum substitutes (GFSRPMI and ALBRPMI) to assess the drug sensitivity of P. falciparum, using chloroquine diphosphate (CQ) and dihydroartemisinin (DHART) as conventional antimalarial drugs. Growth inhibition was measured by four different methods: flow cytometry with SYBR Green I (FCM), microscopy (Giemsa method), enzymatic estimation of parasite lactate dehydrogenase (pLDH), and histidine-rich protein 2 (HRPII) determination. In drug sensitivity tests on asynchronous parasites cultured for 96 h in the presence of drugs, the dose-response curves were similar and differences in the 50% growth inhibition concentrations for the drugs, which were estimated by the four methods, were not statistically significant for the three culture media. The effect of the drugs on the growth of synchronous parasites at the ring stage was also assessed in micro-volume tests by three different methods of FCM: tracking fluorescent erythrocytes, schizont test, and merozoite test. Dose-response curves for the drugs were similar, and differences in the 50% growth inhibition concentrations were not statistically significant for CDRPMI and GFSRPMI. Thus CDRPMI as well as GFSRPMI and ALBRPMI can be similarly useful media for drug sensitivity testing of P. falciparum. The FCM, pLDH and HRPII estimations were fast and reliable detection methods, with FCM allowing schizont and merozoite tests to be performed with shorter periods of culture.

Indian Anti-Malaria OMARIA Is Effective Against African Drug Resistant <i>P. falciparum</i>Field Isolates and Laboratory Strains;without Toxicity

OMARIA which is used to treat malaria in Odisa province, India, was investigated in Africa. The in-vitro anti-malarial activity of OMARIA was tested on P. falciparum strains FCB (chloroquine-resistant) and 3D7 (chloroquine-sensitive) and on fresh clinical isolates from Gabon, using the DELI method. Host cell toxicity was analysed with the MTT test. Interesting activity was observed. Inhibition concentrations (IC50) were 20.6 ± 5.2 μg/ml and 14.1 ± 4.3μg/ml respectively on FCB and 3D7 strains. On clinical isolates, the mean of IC50 was 10.65 ± 4.8μg/ml. OMARIA is highly potent against all field isolates tested by us (Gabon includes Pfmdr1 N86). Lethal dose on Vero cells being 165 ± 10.7μg/ml indicate a selective index of 13 for FCB, i.e., non-toxic. Data substantiates scientific rationale for use of OMARIA. This information and such understanding can be used in searching African phyto parables (for use in Africa with similar results as in India) and in new drug design. With Indian assistance, Punica granatum can also be cultivated in Central Africa, and OMARIA can be made, with an aim to Fight Malaria at Home.

Bioinformatics analysis for structure and function of CPR of Plasmodium falciparum

Objective:To analyse the structure and function of NADPH-cytochrome p450 reductase(CYPOR or CPR) from Plasmodium falciparum(Pf),and to predict its’ drug target and vaccine target. Methods:The structure,function,drug target and vaccine target of CPR from Plasmodium falciparum were analyzed and predicted by bioinformatics methods.Results:PfCPR,which was older CPR,had close relationship with the CPR from other Plasmodium species,but it was distant from its hosts,such as Homo sapiens and Anopheles.PfCPR was located in the cellular nucleus of Plasmodium falciparum.335aa-352aa and 591aa - 608aa were inserted the interior side of the nuclear membrane,while 151aa-265aa was located in the nucleolus organizer regions.PfCPR had 40 function sites and 44 protein-protein binding sites in amino acid sequence.The teriary structure of laa-700aa was forcep-shaped with wings.15 segments of PfCPR had no homology with Homo sapien CPR and most were exposed on the surface of the protein.These segments had 25 protein-protein binding sites.While 13 other segments all possessed function sites. Conclusions:The evolution or genesis of Plasmodium falciparum is earlier than those of Homo sapiens.PfCPR is a possible resistance site of antimalarial drug and may involve immune evasion, which is associated with parasite of sporozoite in hepatocytes.PfCPR is unsuitable as vaccine target,but it has at least 13 ideal drug targets.

Comparison of protein patterns between Plasmodium falciparum mutant clone T9/94-M1-1(b3) induced by pyrimethamine and the original parent clone T9/94

Objective:To compare the protein patterns from the extracts of the mutant clone T9/94-M1-1(b3)induced by pyrimethamine,and the original parent clone T9/94 following separation of parasite extracts by two-dimensional electrophoresis(2-DE).Methods:Proteins were solubilized and separated according to their charges and sizes.The separated protein spots were then detected by silver staining and analyzed for protein density by the powerful image analysis software.Results:Differentially expressed protein patterns(up—or down-regulation)were separated from the extracts from the two clones.A total of 223 and 134 protein spots were detected from the extracts of T9/94 and T9/94-M1-1(b3)clones,respectively.Marked reduction in density of protein expression was observed with the extract from the mutant(resistant)clone compared with the parent(sensitive)clone.A total of 25 protein spots showed at least two-fold difference in density,some of which exhibited as high as ten-fold difference.Conclusions:These proteins may be the molecular targets of resistance of Plasmodium falciparum to pyrimethamine.Further study to identify the chemical structures of these proteins by mass spectrometry is required.

Insights into the pyrimidine biosynthetic pathway of human malaria parasite Plasmodium falciparum as chemotherapeutic target

Malaria is a major cause of morbidity and mortality in humans. Artemisinins remain as the first-line treatment for Plasmodium falciparum(P. falciparum) malaria although drug resistance has already emerged and spread in Southeast Asia. Thus, to fight this disease, there is an urgent need to develop new antimalarial drugs for malaria chemotherapy. Unlike human host cells, P. falciparum cannot salvage preformed pyrimidine bases or nucleosides from the extracellular environment and relies solely on nucleotides synthesized through the de novo biosynthetic pathway. This review presents significant progress on understanding the de novo pyrimidine pathway and the functional enzymes in the human parasite P. falciparum. Current knowledge in genomics and metabolomics are described, particularly focusing on the parasite purine and pyrimidine nucleotide metabolism. These include gene annotation, characterization and molecular mechanism of the enzymes that are different from the human host pathway. Recent elucidation of the three-dimensional crystal structures and the catalytic reactions of three enzymes: dihydroorotate dehydrogenase, orotate phosphoribosyltransferase, and orotidine 5'-monophosphate decarboxylase, as well as their inhibitors are reviewed in the context of their therapeutic potential against malaria.

Modeling and targeting an essential metabolic pathway of Plasmodium falciparum in apicoplast using Petri nets

Petri net(PN) is one of the promising computational and mathematical formalisms used to represent and study the behavior of complex metabolic networks. The various available analysis techniques of PN could be used to validate and analyze the network in different scenarios. Plasmodium falciparum is one of the threatening parasites which causes malaria, a deadly disease affecting a large number of today’s world population. The development of antimalarial drug resistance is an emerging global threat, highlighting the need to discover novel antimalarial targets. The fatty acid biosynthesis of malarial parasite is one of the essential metabolic pathways required for its growth and is present in apicoplast, a non-photosynthetic plastid. The malarial parasite obtains fatty acids by using type two fatty acid synthase(FAS II) enzyme,which is different from type one enzyme used by human host, making it an ideal drug target.This article proposes and studies the PN model of the parasite’s FAS II pathway to analyze the mechanism of potential drug targets in this pathway. The proposed PN model can serve as a base for further findings in the field of antimalarial drug targets to decrease the malaria mortality rate.

Prioritised identification of structural classes of natural products from higher plants in the expedition of antimalarial drug discovery

The emergence and spread of drug-recalcitrant Plasmodium falciparum parasites threaten to reverse the gains made in the fight against malaria.Urgent measures need to be taken to curb this impending challenge.The higher plant-derived sesquiterpene,quinoline alkaloids,and naphthoquinone natural product classes of compounds have previously served as phenomenal chemical scaffolds from which integral antimalarial drugs were developed.Historical successes serve as an inspiration for the continued investigation of plant-derived natural products compounds in search of novel molecular templates from which new antimalarial drugs could be developed.The aim of this study was to identify potential chemical scaffolds for malaria drug discovery following analysis of historical data on phytochemicals screened in vitro against P.falciparum.To identify these novel scaffolds,we queried an in-house manually curated database of plant-derived natural product compounds and their in vitro biological data.Natural products were assigned to different structural classes using NPClassifier.To identify the most promising chemical scaffolds,we then correlated natural compound class with bioactivity and other data,namely(i)potency,(ii)resistance index,(iii)selectivity index and(iv)physicochemical properties.We used an unbiased scoring system to rank the different natural product classes based on the assessment of their bioactivity data.From this analysis we identified the top-ranked natural product pathway as the alkaloids.The top three ranked super classes identified were(i)pseudoalkaloids,(ii)naphthalenes and(iii)tyrosine alkaloids and the top five ranked classes(i)quassinoids(of super class triterpenoids),(ii)steroidal alkaloids(of super class pseudoalkaloids)(iii)cycloeudesmane sesquiterpenoids(of super class triterpenoids)(iv)isoquinoline alkaloids(of super class tyrosine alkaloids)and(v)naphthoquinones(of super class naphthalenes).Launched chemical space of these identified classes of compounds was,by and large,distinct from that of‘legacy’antimalarial drugs.Our study was able to identify chemical scaffolds with acceptable biological properties that are structurally different from current and previously used antimalarial drugs.These molecules have the potential to be developed into new antimalarial drugs.

Molecular markers for artemisinin and partner drug resistance in natural Plasmodium falciparum populations following increased insecticide treated net coverage along the slope of mount Cameroon:cross-sectional study

Background:Drug resistance is one of the greatest challenges of malaria control programmes,with the monitoring of parasite resistance to artemisinins or to Artemisinin Combination Therapy(ACT)partner drugs critical to elimination efforts.Markers of resistance to a wide panel of antimalarials were assessed in natural parasite populations from southwestern Cameroon.Methods:Individuals with asymptomatic parasitaemia or uncomplicated malaria were enrolled through crosssectional surveys from May 2013 to March 2014 along the slope of mount Cameroon.Plasmodium falciparum malaria parasitaemic blood,screened by light microscopy,was depleted of leucocytes using CF11 cellulose columns and the parasite genotype ascertained by sequencing on the Illumina HiSeq platform.Results:A total of 259 participants were enrolled in this study from three different altitudes.While some alleles associated with drug resistance in pfdhfr,pfmdr1 and pfcrt were highly prevalent,less than 3%of all samples carried mutations in the pfkelch13 gene,none of which were amongst those associated with slow artemisinin parasite clearance rates in Southeast Asia.The most prevalent haplotypes were triple mutants PfdhfrI51R59N108I164(99%),pfcrt-C72V73I74E75T76(47.3%),and single mutants PfdhpsS_(436)G_(437)K_(540)A_(581)A_(613)(69%)and Pfmdr1 N_(86)F)(184)D_(1246)(53.2%).Conclusions:The predominance of the Pf pfcrt CVIET and Pf dhfr IRN triple mutant parasites and absence of pfkelch13 resistance alleles suggest that the amodiaquine and pyrimethamine components of AS-AQ and SP may no longer be effective in their role while chloroquine resistance still persists in southwestern Cameroon.

双氢青蒿素和奎宁对恶性疟原虫早期配子体作用的随机比较

【目的】研究双氢青蒿素对恶性疟原虫早期配子体的抑杀作用。【方法】仅骨髓带恶性疟原虫早期配子体而骨髓与外周血液均无成熟配子体的患者 11例 ,随机分为A、B 2组。A组 6例口服双氢青蒿素片 7d总量 4 80mg ;B组 5例口服硫酸奎宁片 7d总量 10 50 0mg ,定时取骨髓和外周血液涂片 ,观察两组配子体密度的变化。【结果】A组骨髓早期配子体药后 10d全部转阴 ;而B组全部阳性 ,至药后 14d仍有 2 / 5例阳性。外周血液配子体转阴时间 ,A组为 ( 4 .8± 0 .9)d ;B组为 ( 2 2 .0± 5.8)d。【结论】双氢青蒿素能杀灭恶性疟原虫早期配子体 。

青蒿琥酯片治疗恶性疟的剂量再探索

为探索青蒿琥酯治疗恶性疟的最佳剂量和疗程，本研究在中国海南岛抗性恶性疟流行地区，采用青蒿琥酯片５ｄ疗程总量６００ｍｇ和７ｄ疗程总量８００ｍｇ治疗无并发症恶性疟疾，进行开放性随机比较。每组治疗５０例，全部病例住院２８ｄ观察原虫复燃情况。结果：两组病例临床症状均被迅速控制。５ｄ疗程组和７ｄ疗程组退热时间分别为（２４．０±１３．６）ｈ和（２０．０±１０．１）ｈ，原虫转阴时间分别为（６１．７±２１．１）ｈ和（５５．８±１５．８）ｈ，两组无显著性差异。２８ｄ内原虫复燃率７ｄ组为４．２％（２／４８），５ｄ组为１９．１％（９／４７），治愈率分别为９５．８％和８０．９％。两组比较有非常显著性差异（χ２＝５．２１，Ｐ＜０．０１），表明７ｄ疗程可使复燃率明显下降。两组皆未见临床不良反应。提示青蒿琥酯７ｄ疗程总量８００ｍｇ是一个高治愈率的给药方案，可作为标准疗程剂量。

青蒿琥酯治疗脑型疟的临床疗效

在多重抗药性恶性疟流行区的越南南方采用青蒿琥酯３６０～４２０ｍｇ／５ｄ静脉注射治疗１６８例脑型疟，配合积极的对症处理和支持疗法，治愈１５２例（９０．５％）、死亡１６例（９．５％）；平均原虫转阴时间为（５８．５±２９．０）ｈ，平均退热时间为（５４．３±３８．３）ｈ，平均昏迷清醒时间为（３７．５±３３．８）ｈ。全部病人在治疗过程中均未发现与药物有关的毒副反应。结果表明：在多重抗药性恶性疟流行区治疗重症疟疾患者，青蒿琥酯是一个理想的速效。

我国恶性疟原虫对氯喹抗性的消长

目的 监测停止或减少使用氯喹防治恶性疟后恶性疟原虫对氯喹抗性的变化。 方法 采用世界卫生组织 (WHO)制定的体外微量法和体内四周法 ,在停用氯喹后不同时间测定恶性疟原虫对氯喹的敏感性。 结果 海南省乐东县抱由镇体外法测定抗性率由 1981年的 97 9%降至 1997年的 2 6 7% (P <0 0 1) ,完全抑制裂殖体形成的平均药浓度由 10 46± 7 14 pmol/μl血 降至 1 63± 1 47pmol/μl血 (P <0 0 1) ,用较高药浓度 (>6 4pmol/μl血)才能完全抑制裂殖体形成的病例所占比例由 83 3 %降为 6 7% (P <0 0 1)。体内法测定抗性率由 1981年的 84 2 %降为 1997年的 18 4% (P <0 0 1) ,三级抗性 (RⅢ )占抗性病例的比例由 5 3 1%降为 14 3 % (P <0 0 1) ,血中无性体疟原虫平均消失时间由 72 0± 2 1 6h变为 5 0 7± 16 1h。2 0 0 1年三亚市雅亮乡体外法测定抗性率为 5 9 8% ,平均抑制药浓度 3 5 6± 2 12 pmol/μl血 。 2 0 0 3年乐东县福抱乡体内法测定抗性率为 62 5 % ,RI、RⅡ和RⅢ分别占抗性病例 5 0 %、 3 0 %和 2 0 % ,无性体疟原虫平均消失时间 5 6 9± 17 2h。云南省勐腊县体外法测定抗性率由 1981年的97 4%降至 1999年的 77 8% (P <0 0 1) ,完全抑制裂殖体形成的平均药物浓度由 17 2± 12

双氢青蒿素对恶性疟原虫有性生殖期的影响

１８ 例带配子体的恶性疟患者随机分为２ 组： 双氢青蒿素组１０ 例， 口服５ ｄ 疗程总量３６０ ｍｇ ； 奎宁组８ 例， 口服７ ｄ 疗程总量１０ ５００ ｍｇ 。全部患者每天涂血片计算配子体密度， 于给药当天（Ｄ０） ， 药后４ 、７ 、１０ 、１４ 、２１ 和２８ ｄ （Ｄ４ 、Ｄ７ 、Ｄ１０ 、Ｄ１４ 、Ｄ２１ 、和Ｄ２８） 取血作大劣按蚊感染试验。患者血中配子体转阴时间， 双氢青蒿素组为２１ ．８ ±５ ．２ ｄ ； 奎宁组为３１ ．５ ±８ ．７ ｄ 。按蚊感染试验结果： 双氢青蒿素组有４ 例在Ｄ０Ｄ４ 、Ｄ７ 、Ｄ１０ 、Ｄ１４ 均未能感染按蚊； 其余６ 例于Ｄ０ 、Ｄ４ 、Ｄ７ 、Ｄ１０ 、Ｄ１４ 、Ｄ２１ 按蚊感染阳性例数为６／６ 、６／６ 、６／６ 、２／６ 、０／６ 和０／６ 。奎宁组Ｄ０ 、Ｄ４ 、Ｄ７ 、Ｄ１０ 、Ｄ１４ 、Ｄ２１ 和Ｄ２８ 按蚊感染阳性例数为４／８ 、８／８ 、８／８ 、８／８ 、８／８ 、２／８ 和０／８ 。这表明双氢青蒿素５ ｄ 总量３６０ ｍｇ 对恶性疟原虫有性生殖期发育有明显抑制作用， 对生理上未成熟的配子体， 可中断其发育。

青蒿琥酯对恶性疟原虫早期配子体的作用

为了研究青蒿琥酯对恶性疟原虫早期配子体（ＰＦＧｅ）的作用，选择经骨髓涂片查到ＰＦＧｅ，但未有成熟配子体（ＰＦＧｍ）的脑型疟患者３４例，对其治疗前后ＰＦＧｅ的改变进行了专门观察。方法是采用青蒿琥酯静脉给药，５ｄ疗程总量３６０～４２０ｍｇ。治疗后，于当天（Ｄ０）和第７天（Ｄ７）或加第１０天（Ｄ１０）作骨髓涂片，观察ＰＦＧｅ的变化，每天取外周血片计算ＰＦＧｅ和ＰＦＧｍ数，连续２８ｄ或直至ＰＦＧ消失。结果３４例治疗后，Ｄ７骨髓ＰＦＧｅ消失２７例，Ｄ１０消失７例。２９例外周血查到ＰＦＧｅ者，全部于治疗后２～８ｄ消失，平均（４．５±１．９）ｄ。３３例骨髓和外周血均一直未查到ＰＦＧｍ，仅１例于首剂给药后１０ｈ，外周血查到少量ＰＦＧｍ，持续至Ｄ１１消失。结果表明，青蒿琥酯对Ⅰ～Ⅳ期ＰＦＧｅ都有杀灭作用。只有极少数接近成熟的配子体未被控制而出现于外周血液。

我国海南省抗药性恶性疟流行情况的变迁

由于恶性疟原虫多种抗药性的不断产生和蔓延，给疟疾防治带来了困难。本文以调查研究和科学实验相结合的方法，对我国高疟区海南省抗药性恶性疟流行情况的变迁作一系统报导。１９５５年该岛２７７万人口患疟疾者有２８６７４５人，因疟疾而致死者２８７人。１９５９年进行了Ｄ．Ｄ．Ｔ．滞留喷洒，效果良好，主要传疟媒介微小按蚊由原占蚊群组成６７．５％下降到０．９％。此后疟疾发病率逐年下降，至１９６５年带虫率已低于０．９％和０．２％。１９７３年作者等在该岛三亚市发现了抗氯喹恶性疟病例，至１９７８年岛内各地先后发现抗氯喹恶性疟存在，其中约１／３属于ＲⅡ＿ＲⅢ级抗性。１９７９年该岛以哌喹全面取代氯喹应用于疟疾防治。但到了１９８２年抗哌喹恶性疟病例又不断出现，抗性率为１０％～２０％，其中有５％为ＲⅡ＿ＲⅢ级抗性。１９７８～１９９０年作者等在三亚市建立了体外测试方法，以监测恶性疟原虫的敏感性。试验证明，恶性疟原虫分离株抗氯喹程度已下降，对哌喹则具有抗性，但在哌喹与氯喹之间未见明显交叉抗性，而对蒿甲醚和咯萘啶则显示敏感。近年来海南省恶性疟发病率已大幅度下降，１９９６年仅为５４．９３／１０万人口，但对疟原虫抗药性问题仍需继续监测。

双氢青蒿素与咯萘啶伍用治疗抗性恶性疟的研究

目的 寻找更理想的治疗抗药性恶性疟的联合用药方案。 方法 双氢青蒿素、咯萘啶单用及伍用分别治疗 2 4、2 5和 32例恶性疟现症患者 ,分别于服药后第 14、2 1及 2 8天随访 ,以退热时间、原虫消失时间、复燃时间、复燃率、治愈率、配子体携带率、药物副作用发生率等为指标 ,以双氢青蒿素、咯萘啶标准疗法为对照 ,进行临床双盲试验 ,综合评估双氢青蒿素合并咯萘啶疗法。 结果 伍用组退热时间 35 .7± 2 4 .7h与咯萘啶单用组 35 .8± 16 .5 h相似 (P>0 .0 5 ) ,显著快于双氢青蒿素单用组 5 2 .6± 38.9h(P<0 .0 1)。伍用组无性体原虫消失时间 2 3.8± 10 .1h与双氢青蒿素单用组 2 2 .9± 6 .5 h相似 (P>0 .0 5 ) ,显著快于咯萘啶单用组 4 9.4± 2 0 .3h (P<0 .0 1)。伍用组治后配子体出现率、持续时间及密度分别为 2 0 .0 %、5 .7d和 4个 /μl血 ,与双氢青蒿素单用组 16 .7%、3.5 d和 3个 /μl血相似 (P>0 .0 5 ) ,明显好于咯萘啶单用组 6 0 .9%、11.5 d和 12个 /μl血 (P<0 .0 1)。 3种方案均无明显药物副反应。 结论 双氢青蒿素与咯萘啶伍用保持了两药的优点 ,克服了两药的不足 。