Plasmodium vivax resistance to chloroquine in Dawei, southern Myanmar

Objective  To assess the efficacy of chloroquine in the treatment of Plasmodium vivax malaria in in Dawei District, southern Myanmar.
Methods  Enrolled patients at Sonsinphya clinic >6 months of age were assessed clinically and parasitologically every week for 28 days. To differentiate new infections from recrudescence, we genotyped pre- and post-treatment parasitaemia. Blood chloroquine was measured to confirm resistant strains.
Results  Between December 2002 and April 2003, 2661 patients were screened, of whom 252 were included and 235 analysed. Thirty-four per cent (95% CI: 28.1–40.6) of patients had recurrent parasitaemia and were considered treatment failures. 59.4% of these recurrences were with a different parasite strain. Two (0.8%) patients with recurrences on day 14 had chloroquine concentrations above the threshold of 100 ng/ml and were considered infected with chloroquine resistant parasites. 21% of failures occurred during the first 3 weeks of follow-up: early recurrence and median levels of blood chloroquine comparable to those of controls suggested P. vivax resistance.
Conclusions  Plasmodium vivax resistance to chloroquine seems to be emerging in Dawei, near the Thai-Burmese border. While chloroquine remains the first-line drug for P. vivax infections in this area of Myanmar, regular monitoring is needed to detect further development of parasite resistance.     

缅甸拉咱市氯喹治疗间日疟的敏感性评价

缅甸拉咱市单纯间日疟原虫感染者48例分为氯喹A方案组（26例）和B方案组（22例）,分别采用成人总剂量1 200 mg （第1天顿服600 mg,第2、3天300 mg/d）和1 500 mg（第1天顿服750 mg,第2、3天375 mg/d）的三天疗法治疗。于服药当天、服药后第1、2、3、7、14、21和28天采集指血或耳垂血制作厚、薄血涂片检查疟原虫,测量体温和观察药物不良反应。氯喹治疗后,两组病例血内无性体原虫3 d内全部阴转。第28天随访治愈率为100%。结果表明,缅甸拉咱市的间日疟病例对氯喹治疗均敏感,该边境地区间日疟病例可采用氯喹进行临床治疗。     

复式PCR检测间日疟原虫和恶性疟原虫的现场应用

目的探讨复式PCR方法在疟疾诊断中的现场应用价值。方法根据疟原虫18 S rRNA基因序列,合成疟原虫属特异性上游引物和间日疟原虫、恶性疟原虫种特异性下游引物,优化PCR反应体系,建立在同一PCR反应体系中同时检测间日疟原虫、恶性疟原虫基因组特异性DNA片段的疟疾诊断方法,并评价其现场应用价值。结果间日疟原虫和恶性疟原虫基因组DNA经过复式PCR后,分别扩增出1 451 bp和833 bp的特异性条带,而伯氏疟原虫、食蟹猴疟原虫及健康人血样均无扩增带出现,用该反应体系可检出原虫血症为1.1×10-6和5.6×10-7的间日疟原虫和恶性疟原虫感染。与镜检法相比,复式PCR检测119份现场样本,112份与镜检结果相同,阳性率为54%,漏诊率为0.8%,误诊率为0,而镜检法依次分别为53%、1.7%和3.4%。结论复式PCR方法检测疟原虫具有简便、快速、特异、敏感等优点,在疑似病例的鉴别诊断和分子流行病学调查中具有良好的应用价值。     

Plasmodium vivax: recent world expansion and genetic identity to Plasmodium simium

Plasmodium vivax causes the most geographically widespread human malaria, accounting annually for 70-80 million clinical cases throughout the tropical and subtropical regions of the world's continents. We have analyzed the DNA sequences of the Csp (circumsporozoite protein) gene in 24 geographically representative strains of P. vivax and 2 of P. simium, which parasitizes several species of New World monkeys. The Csp sequences are of two types, VK210 and VK247, which differ by three diagnostic amino acid replacements, one in each of the 5' and 3' terminal regions [5' nonrepeat (NR) and 3' NR] of the gene and in an insertion sequence that precedes the 3' NR region. The central region of the gene consists of approximately 38 repetitive "motifs," which are alternatively four and five amino acids long, which also are diagnostically different between the VK210 and VK247 types. There are very few synonymous substitutions within and between the two types of strains, which we hypothesize reflects that the worldwide spread of P. vivax is very recent. The two P. simium Csp sequences belong one to each of the two VK types and are genetically indistinguishable from the corresponding P. vivax strains, suggesting that at least two host transfers have occurred between humans and New World monkeys. We exclude as unlikely the possibility that the two types of sequences could have independently arisen in humans and platyrrhines by natural selection. There are reasons favoring each of the two possible directions of host transfer between humans and monkeys.     

Resistance to antimalarials by Plasmodium falciparum in Arso PIR, Irian Jaya, Indonesia

Between 1987 and 1990, susceptibility of Plasmodium falciparum to chloroquine and to Fansidar was measured in vivo in 151 volunteers using the standard 7-day test. All volunteers lived in Arso PIR, Irian Jaya. A 25 mg/kg dose of chloroquine base was administered over a three-day period to 92 volunteers positive for P. falciparum rings (greater than 10 rings/200 white blood cells). Fifty volunteers (54%) showed results consistent with resistance. Twenty-nine were classified RII, and 21 RIII. In November 1989, a single curative dose of Fansidar was administered to 59 volunteers divided among three groups with 18 months, four years, and life-long exposure to endemic malaria. The proportion of volunteers in each group still positive for P. falciparum on day 7 of followup was 54%, 0%, and 14%, respectively. Thus, immune status profoundly effected clinically response to Fansidar. Standard in vitro microtests were also performed on parasites from 11 volunteers against chloroquine, amodiaquine, quinine, pyrimethamine/sulfadoxine, and and mefloquine. Nine of ten isolates showed in vitro growth consistent with resistance to chloroquine. Tests with other drugs showed few isolates with results considered indicative of susceptibility. Arso PIR has a severe drug resistance problem.     

Distribution of two species of malaria, Plasmodium falciparum and Plasmodium vivax, on Lombok Island, Indonesia

Medical and entomological surveys were conducted to determine the risk factors of Plasmodium falciparum and P. vivax infections on Lombok Island, Indonesia, to find the risk factors and the main mosquito vectors for each malaria. Multivariate longitudinal analysis demonstrated two significant risk factors for infection with P. falciparum: disappearance of P. vivax parasitemia (p<0.001) and a specific study site (p<0.001). In contrast, younger age (p=0.024) and the interpolated virtual density of An. subpictus (p=0.041) were significantly associated with increased risk of infection with P. vivax. Thus, it seems that the distribution of P. vivax was determined largely by the presence of An. subpictus, whilst that of P. falciparum was influenced by antagonism with P. vivax. This result shows the importance of following-up treated P. vivax patients to identify recrudescence of P. falciparum in this area.     

Host immune response to Plasmodium.

Malaria is the most widely spread infectious disease of man affecting almost half of the world's population. Control of malaria remains one of the world's biggest health challenges. Development of vaccines has been considered a valid and necessary complement to control malaria in addition to the control measures of the vectors. The Plasmodium parasites that cause the disease have many stages in their cycle, each with distinct morphology and antigenicity. Understanding the activation, interaction and effector function of the different components of the immune system in relation to target antigens on different stages of malaria parasites is necessary to achieve complete protection by vaccination.     

Sensitivity of Plasmodium vivax to chloroquine in Sa Kaeo Province,Thailand

Following a recent, abrupt local increase in the incidence of vivax malaria, a study was conducted in order to evaluate the efficacy of chloroquine for the treatment of 26 adult patients with acute vivax malaria in Sa Kaeo Province, Thailand. The chloroquine sensitivity of Plasmodium vivax has been assessed in parallel, using a growth inhibition method. Blood samples for the in vitro tests were taken prior to the administration of the standard treatment with chloroquine--in total 25 mg base/kg over 3 days--and primaquine 0.25 mg base/kg once daily for 14 days. The efficacy has been assessed according to the WHO standard in vivo test. The cure rate was 100%. No recrudescence was observed during the follow-up period of 28 days. The mean fever clearance time (FCT) was 40 h, the mean parasite clearance time (PCT) was 49 h. Mean IC(50) and IC(90) of the parasites were 28 and 171 nM, respectively. These results show that local P. vivax is still sensitive to chloroquine. The epidemic outbreak was therefore obviously not due to the presence of chloroquine-resistant P. vivax.     

Genetic diversity of Plasmodium vivax and Plasmodium falciparum in Kohat District, Pakistan

Malaria is one of the serious diseases threatening human health in Pakistan and contributes to a large proportion of the total malaria deaths in South Asia. However, little is known about the nature and extent of genetic diversity of the malarial parasites circulating in Pakistan. This study was designed to assess the infection status of Plasmodium and the genetic diversity of Plasmodium vivax and Plasmodium falciparum by analyzing msp-3α, msp-3β and msp-1, msp-2 genes respectively using allele specific nested PCR and RFLP assays. For this purpose, 130 field isolates were collected from the individuals who exhibited clinical symptoms associated with malaria in the Kohat region of Khyber Pakhtoonkhwa (KPK), Pakistan. Among 130 blood samples collected, P. vivax was detected in 105/130 (80.8%) and P. falciparum in 21/130 (16.2%). Mixed infections with both parasites were detected in 4/130 (3%) of the isolates. A large number of distinguishable alleles were found for msp genetic markers: 10 alleles for msp-3α and seven for msp-3β with one mixed infection in case of msp-3β. The genotyping of P. falciparum showed that K1+MAD20 mixed genotype was dominant in msp-1 and FC27 in msp-2. The results collectively suggest that P. vivax and P. falciparum populations in this region are highly polymorphic and mixed infections are prevalent.     

Resistance to chloroquine by Plasmodium vivax in Irian Jaya, Indonesia.

Evidence of emerging resistance to chloroquine by Plasmodium vivax is described from Irian Jaya (Indonesian New Guinea). Sixteen of 24 residents in the village of Arso PIR II taking supervised weekly chloroquine prophylaxis (5 mg base/kg) had asexual parasitemia with P. vivax at least once during eight weeks of surveillance. An American working in the same village developed symptomatic P. vivax parasitemia despite chloroquine prophylaxis. Five days after therapy with 600 mg chloroquine base, the asexual parasitemia in the American increased 40-fold, but cleared after treatment with 1,500 mg chloroquine base. Serum samples were not available from many of the cases, but six local residents and the American had serum levels of chloroquine in excess of the ordinarily suppressive 15 ng/ml at the time of their asexual parasitemias (16-70 ng/ml). The weekly 300 mg base tablet of chloroquine, which has been the standard for prophylaxis against malaria for more than 40 years, was not effective against P. vivax in Arso PIR, Irian Jaya.     

Autoimmune thrombocytopenia in recurrent polietiological malaria (Plasmodium falciparum, Plasmodium vivax)

Thrombocytopenia frequently appear in severe malaria. The reasons of low blood platelets count are different and its results of hypersplenism, subclinical course of intravascular coagulation (DIC). Thrombocytopenia from "consumption" is consequence of sequestration of blood platelets in blood vessels of lungs and cerebral. We examination 29 years old men, who was as forest worker in islands on Indonesia. He was treated with recurrent, poliethiological malaria (Plasmodium falciparum, Plasmodium vivar) and severe thrombocytopenia (17.0 G/L) without hepatosplenomegalia. Antiplatelet antibody was examined in blood serum by ELISA methods (GTI - PAKPLUS. In blood serum was detected IgG antibody agai nstglicoprotein receptors on surface of blood platelets GPIIb/IIIa, GPIV, GPIb/IX, GPV, GPIa/IIa. Chronic infections of Plasmodium may conduct to autoimmune destruction of blood platelets.     

Plasmodium falciparum and Plasmodium vivax infections in the owl monkey (Aotus trivirgatus). II. Responses to chloroquine, quinine, and pyrimethamine.

The studies described in this report were designed to determine the responses of established infections with eight strains of Plasmodium falciparum and two strains of P. vivax in owl monkeys to treatment with chloroquine, quinine, and pyrimethamine. Responses with these different strains ranged from cure via application of well-tolerated doses of two of the above drugs and refractoriness to treatment with maximally tolerated doses of the third, to complete resistance to maximally tolerated doses of all three compounds. The results of treatment exhibited in infected owl monkeys correlated well in two respects with those reported in humans infected with the same plasmodial species. First, calculated on a milligram per M2 basis, the doses of chloroquine, quinine, or pyrimethamine required for a CD90 response in owl monkeys infected with strains susceptible to these drugs were remarkably similar to the doses required and/or employed for cure of infections with so-called drug-susceptible strains in human patients. Secondly, with few exceptions, the responses to the above drugs in owl monkeys infected with the ten specially selected strains were essentially identical with those exhibited by human volunteers or patients infected with the same strains. Together, these findings and correlations provide strong support for use of owl monkeys infected with appropriate strains of P. falciparum and P. vivax in the search for more broadly effective antimalarial drugs.     

In vitro susceptibility of Plasmodium falciparum isolates from Jilore, Kenya, to antimalarial drugs

Twenty-six Plasmodium falciparum isolates obtained during a prophylaxis study at Jilore primary school, Malindi, Kenya, were adapted to in vitro culture and their susceptibility to 13 antimalarial drugs was tested by a modified radioisotopic method. Pyrimethamine, chloroquine, amodiaquine, cycloguanil, chlorcycloguanil, quinine, quinidine and sulfadoxine, and the experimental compounds MB 35769, mefloquine, WR 184806, parvoquone, and menoctone were used. The isolates could be divided into two groups with significantly different susceptibility to pyrimethamine, shown by a 755-fold difference in the mean ID50 values (2.77 +/- 1.98 x 10(-10) mol/l and 2.09 +/- 1.64 x 10(-7) mol/l). The mean susceptibility of the two groups differed 7.7-fold for chlorcycloguanil and 14.6-fold for cycloguanil, but were not significantly different for the other drugs. All isolates were more sensitive to amodiaquine than to chloroquine in vitro. The ratio of the geometric mean ID50 values of chloroquine to amodiaquine was 3.13. The ratio for the chemically related compounds parvoquone to menoctone was 5.63, quinine to quinidine was 5.58, and mefloquine to WR 184806 was 12.16.     

Age-acquired immunity to a Plasmodium vivax invasion ligand,the duffy binding protein

To investigate the interactions of glycoconjugates with the innate immune system, peripheral blood mononuclear cells were stimulated with glycolipids derived from Schistosoma mansoni eggs and worms and with biochemically synthesized neoglycoconjugates. Egg glycolipids stimulated the production of interleukin (IL)--10, IL-6, and tumor necrosis factor--alpha in monocytes, whereas worm glycolipids failed to do so. When monoclonal antibodies that specifically recognize defined carbohydrate epitopes were used, the binding of a GalNAc beta 1-4(Fuc alpha 1-2Fuc alpha 1-3)GlcNAc (LDN-DF) reactive antibody was pronounced on egg glycolipids but was absent on worm glycolipids. The binding of antibodies that recognize Gal beta 1-4(Fuc alpha 1-3)GlcNAc (LewisX), GalNAc beta 1-4GlcNAc (LDN), and GalNAc beta 1-4(Fuc alpha 1-3)GlcNAc (LDN-F) was comparable for both preparations. Cytokine production in response to neoglycoconjugates containing enzymatically synthesized glycans also was measured. The LDN-DF neoglycoconjugate was the most potent cytokine inducer, which indicates that this difucosylated glycan can act at the host-parasite interface and can trigger innate immune responses.     

In vivo responses to antimalarials by Plasmodium falciparum and Plasmodium vivax from isolated Gag Island off northwest Irian Jaya, Indonesia.

There is renewed interest in the rich nickel and cobalt deposits of Pulau Gag, an isolated but malarious island off the northwest coast of Irian Jaya. In preparation for an expanded workforce, an environmental assessment of malaria risk was made, focusing upon malaria prevalence in the small indigenous population, and the in vivo sensitivity of Plasmodium falciparum and P. vivax to chloroquine (CQ) and sulfadoxine/pyrimethamine (S/P), the respective first- and second-line drugs for uncomplicated malaria in Indonesia. During April-June 1997, mildly symptomatic or asymptomatic malaria infections were found in 24% of 456 native residents. Infections by P. falciparum accounted for 60% of the cases. Respective day 28 cure rates for CQ (10 mg base/kg on days 0 and 1; 5 mg/kg on day 2) in children and adults were 14% and 55% (P < 0.005). Type RII and RIII resistance characterized only 5% of the CQ failures. Re-treatment of 36 P. falciparum CQ treatment failures with S/P (25 mg/kg and 1.25 mg/kg, respectively) demonstrated rapid clearance and complete sensitivity during the 28-day follow-up period. More than 97% of the P. vivax malaria cases treated with CQ cleared parasitemia within 48 hr. Three cases of P. vivax malaria recurred between days 21 and 28, but against low drug levels in the blood. The low frequency of RII and RIII P. falciparum resistance to CQ, the complete sensitivity of this species to S/P, and the absence of CQ resistance by P. vivax are in contrast to in vivo and in vitro test results from sites on mainland Irian Jaya.     

Sensitivity of Plasmodium falciparum isolates to chloroquine in Kisumu and Malindi, Kenya

The chloroquine sensitivity of Plasmodium falciparum isolates from infected persons living in Kisumu and Malindi, Kenya, was determined in vivo and vitro. There was no evidence of chloroquine resistance in 217 patients with P. faliparum infections who underwent standard W.H.O. 7-day in vivo tests. In 71 extended 35-day in vivo tests parasitemia recurred in 14 patients on days 21, 28, or 35. Parasites isolated from these 14 persons during the following period were tested in vitro. Eight tests were successful and showed the isolates to be chloroquine sensitive in vitro, suggesting that the recurrence of parasitemia resulted from reinfection rather than resistance. Macro in vitro tests were done on an additional 67 infected persons, 11 of whom also had sensitive 7-day in vivo tests. Chloroquine resistance was not demonstrated in vitro. In Malindi 100% of isolates were inhibited by a chloroquine concentration of less than or equal to 0.75 nmol/ml blood and 80% by less than or equal to 0.5 nmol as compared with 69% and 27.3% respectively of those from Kisumu. These data from individuals living in malarious areas of Kenya contrast with continuing reports of proven chloroquine-resistant P. falciparum malaria in non-immune visitors who acquired their infections in Kenya.     

Susceptibility of Anopheles hermsi to Plasmodium vivax

Two outbreaks of Plasmodium vivax malaria have occurred recently in southern California, and Anopheles hermsi, a newly described species closely related to A freeborni, has been implicated as the vector. To assess the competence of A. hermsi as a vector, its susceptibility to P. vivax was compared with that of the efficient vector A. freeborni by allowing 150 females of each species to feed to repletion on infected squirrel monkeys. Oocyst density was greater in A. hermsi than in A. freeborni, and the frequency and density of sporozoites were similar. A. hermsi was susceptible to P. vivax and readily supported development to the sporozoite stage.