IgG subclasses pattern and high-avidity antibody to the C-terminal region of merozoite surface protein 1 of Plasmodium vivax in an unstable hypoendemic region in Iran

The C-terminal region of Plasmodium vivax merozoite surface protein 1 (PvMSP-1₁₉) is a leading vaccine candidate for inclusion in a polyvalent malaria vaccine. In the present study, the IgG subclasses profile and the avidity of IgG to PvMSP-1₁₉ were evaluated in individuals (n = 94) naturally exposed to P. vivax parasite in malaria endemic areas in Chabahar districts, Iran. In individuals with patent P. vivax malaria, 86.1% was sero-positive to PvMSP-1₁₉ and IgG1 (81.9%) was the predominant subclass. In addition, to determine the persistence of specific IgG, IgG1 and IgG3 antibodies to PvMSP-1₁₉, the frequency of antibodies was determined in the infected subjects (n = 74) after treatment with standard chloroquine and it was detected that the frequency of responders was significantly reduced to 51.3%, 51% and 16.2%, respectively. The antigen-binding avidity of IgG antibodies to PvMSP-1₁₉ was measured in sero-positive sera and the high-avidity of IgG, IgG1 and IgG3 was found in 66.6%, 61% and 47% of the infected subjects with P. vivax, respectively. The present result shows that individuals who exposed to vivax malaria in the endemic region in Iran develop antibodies with high-avidity to PvMSP-1₁₉. These results could help to understand the interactions between the host and P. vivax parasite in development of MSP-1₁₉-based vaccine.     

High prevalence of PfCRT K76T mutation in Plasmodium falciparum isolates in Ghana

Plasmodium falciparum has successfully developed resistance to almost all currently used antimalarials. A single nucleotide polymorphism in the P. falciparum chloroquine resistance transporter (Pfcrt) gene at position 76 resulting in a change in coding from lysine to threonine (K76T) has been implicated to be the corner stone of chloroquine resistance. Widespread resistance to chloroquine in endemic regions led to its replacement with other antimalarials. In some areas this replacement resulted in a reversion of the mutant T76 allele to the wild-type K76 allele. This study was conducted to determine the prevalence of the K76T mutation of the Pfcrt gene eight years after the ban on chloroquine sales and use. A cross-sectional study was conducted in 6 regional hospitals in Ghana. PCR-RFLP was used to analyse samples collected to determine the prevalence of Pfcrt K76T mutation. Of the 1318 participants recruited for this study, 246 were found to harbour the P. falciparum parasites, of which 60.98% (150/246) showed symptoms for malaria. The prevalence of the Pfcrt T76 mutant allele was 58.54% (144/246) and that of the K76 wild-type allele was 41.46% (102/246). No difference of statistical significance was observed in the distribution of the alleles in the symptomatic and asymptomatic participants (P = 0.632). No significant association was, again, observed between the alleles and parasite density (P = 0.314), as well as between the alleles and Hb levels of the participants (P = 0.254). Notwithstanding the decline in the prevalence of the Pfcrt T76 mutation since the antimalarial policy change in 2004, the 58.54% prevalence recorded in this study is considered high after eight years of the abolishment of chloroquine usage in Ghana. This is in contrast to findings from other endemic areas where the mutant allele significantly reduced in the population after a reduction chloroquine use.     

Association between chloroquine resistance phenotypes and point mutations in pfcrt and pfmdr1 in Plasmodium falciparum isolates from Thailand

The relationship between the in vitro susceptibility of Plasmodium falciparum isolates to the quinoline antimalarials chloroquine (CQ), mefloquine (MQ), and quinine (QN), and pfcrt and pfmdr1 gene polymorphisms were investigated. Field isolates (110 samples) were collected from various endemic areas of Thailand throughout 2002-2004. The pfcrt 76T allele was identified in 109 isolates (99.1%) while pfcrt 76K was found in a single (0.9%) isolate. The pfmdr 86N, 86Y, and the combination (86N + 86Y) alleles were identified in 83 (75.5%), 22 (20%), and 5 (4.5%) isolates, respectively. The pfmdr1 1042N, 1042D alleles and a mixture (1042N + 1042D) of the alleles were found in 94 (85.5%), 12 (10.9%) and 4 (3.6%) isolates, respectively. The pfmdr1 1246Y allele was detected in a single (0.9%) isolate. The pfmdr1 gene polymorphisms (86-1042-1246) was grouped into seven haplotypes as follows: N-N-D (68 isolates; 61.2%), Y-N-D (22 isolates; 19.8%), N-D-D (11 isolates; 9.9%), N-D-Y (1 isolate; 0.9%), N/Y-N-D (4 isolates; 3.6%), N-N/D-D (3 isolates; 2.7%), and N/Y-N/D-D (1 isolate; 0.9%). Eight different combinations of pfcrt-pfmdr1 genotypes were observed. Only one CQ-, MQ- and QN-sensitive isolate was found at the Thai-Laos border and no cases of QN resistance were found in this study.     

Plasmodium falciparum GPI toxin: A common foe for man and mosquito

The glycosylphosphatidylinositol (GPI) anchor of the malaria parasite, Plasmodium falciparum, which can be regarded as an endotoxin, plays a role in the induced pathology associated with severe malaria in humans. However, it is unclear whether the main mosquito vector, Anopheles gambiae, can specifically recognize, and respond to GPI from the malaria parasite. Recent data suggests that the malaria vector does mount a specific response against malaria GPI. In addition, following the strong immune response, mosquito fecundity is severely affected, resulting in a significant reduction in viable eggs produced. In this mini-review we look at the increased interest in understanding the way that malaria antigens are recognized in the mosquito, and how this relates to a better understanding of the interactions between the malaria parasite and both human and vector.     

Detection of Plasmodium vivax and Plasmodium falciparum DNA in human saliva and urine: Loop-mediated isothermal amplification for malaria diagnosis

This study investigated loop-mediated isothermal amplification (LAMP) detection of Plasmodium falciparum and Plasmodium vivax in urine and saliva of malaria patients. From May to November 2011, 108 febrile patients referred to health centers in Sistan and Baluchestan Province of south-eastern Iran participated in the study. Saliva, urine, and blood samples were analyzed with nested PCR and LAMP targeting the species-specific nucleotide sequence of small subunit ribosomal RNA gene (18S rRNA) of P. falciparum and P. vivax and evaluated for diagnostic accuracy by comparison to blood nested PCR assay. When nested PCR of blood is used as standard, microscopy and nested PCR of saliva and urine samples showed sensitivity of 97.2%, 89.4% and 71% and specificity of 100%, 97.3% and 100%, respectively. LAMP sensitivity of blood, saliva, and urine was 95.8%, 47% and 29%, respectively, whereas LAMP specificity of these samples was 100%. Microscopy and nested PCR of saliva and LAMP of blood were comparable to nested PCR of blood (к = 0.95, 0.83, and 0.94, respectively), but agreement for nested PCR of urine was moderate (к = 0.64) and poor to fair for saliva LAMP and urine LAMP (к = 0.38 and 0.23, respectively). LAMP assay showed low sensitivity for detection of Plasmodium DNA in human saliva and urine compared to results with blood and to nested PCR of blood, saliva, and urine. However, considering the advantages of LAMP technology and of saliva and urine sampling, further research into the method is worthwhile. LAMP protocol and precise preparation protocols need to be defined and optimized for template DNA of saliva and urine.     

Transplacentally transferred functional antibodies against Plasmodium falciparum decrease with age

Transplacental transfer of antibodies from clinically malaria immune pregnant women to their fetuses is thought to provide passive protection against malaria during infancy. However, the presences and duration of functional antibodies against Plasmodium falciparum (Pf) in newborns has not been described. We used growth inhibition assays (GIA) to measure total anti-malaria functional antibodies present at birth and over the following year. Samples were drawn from cord blood (n = 86) and in infants at six and 12 months of life (n = 86 and 65 respectively). Three laboratory Pf strains (D10, W2mef, 3D7) and a field isolate (Msambweni 2006) were used in the assays. Median (ranges) GIA levels for cord plasma differed between laboratory parasite strains: D10, 0% (0–81); W2mef, 6% (0–80); 3D7, 18% (0–88); Msambweni 2006, 6% (0–43) (P < 0.001, Wilcoxon signed-rank test). GIA levels against all Pf strains were found to decline in infants from birth to six months (P < 0.01, Wilcoxon, signed-rank test). Functional antibodies as measured by GIA are transferred to the fetus and wane in the infants over time. Infant protection from clinical malaria disease may in part be mediated by these functional anti-malaria antibodies.     

The periodicity of Plasmodium vivax and Plasmodium falciparum in Venezuela

We investigated the periodicity of Plasmodium vivax and P. falciparum incidence in time-series of malaria data (1990–2010) from three endemic regions in Venezuela. In particular, we determined whether disease epidemics were related to local climate variability and regional climate anomalies such as the El Niño Southern Oscillation (ENSO). Malaria periodicity was found to exhibit unique features in each studied region. Significant multi-annual cycles of 2- to about 6-year periods were identified. The inter-annual variability of malaria cases was coherent with that of SSTs (ENSO), mainly at temporal scales within the 3–6 year periods. Additionally, malaria cases were intensified approximately 1 year after an El Niño event, a pattern that highlights the role of climate inter-annual variability in the epidemic patterns. Rainfall mediated the effect of ENSO on malaria locally. Particularly, rains from the last phase of the season had a critical role in the temporal dynamics of Plasmodium. The malaria–climate relationship was complex and transient, varying in strength with the region and species. By identifying temporal cycles of malaria we have made a first step in predicting high-risk years in Venezuela. Our findings emphasize the importance of analyzing high-resolution spatial–temporal data to better understand malaria transmission dynamics.     

Extensive heterozygosity in flanking microsatellites of Plasmodium falciparum Na/H exchanger (pfnhe-1) gene among Indian isolates

Plasmodium falciparum Na⁺/H⁺ exchanger-1 (pfnhe-1) gene has been proposed to be a possible marker for quinine resistance. Here, we describe the sequence analysis of the flanking microsatellites of the pfnhe-1 gene among 108 Indian P.falciparum isolates. Among the parasite population, a high degree of polymorphism was observed at all the 10 microsatellite loci within ±40 kb region of the pfnhe-1 gene where the number of alleles varied from 2 to 16 with a high expected heterozygosity ranging from 0.43 to 0.91 at these loci. Also, higher levels of heterozygosity have been observed in P.falciparum isolates collected from both low and high transmission and drug resistant areas. Furthermore, there was no association between QN resistance associated DNNND repeats in PFNHE-1 and the flanking microsatellite haplotypes. In conclusion, the observed high level of microsatellite polymorphism and absence of selective sweep in the flanking ±40 kb region of the pfnhe-1 gene could be an indication that there is no strong selection pressure on this target gene.     

Near-fixation of a Pfmsp1 block 2 allelic variant in genetically diverse Plasmodium falciparum populations across Western Colombia

Assessment of the genetic diversity of Plasmodium falciparum in 110 Colombian isolates revealed that nearly all the parasites in the 97 isolates collected in endemic regions west of the Andes shared the same Pfmsp1 block 2 MAD20-type allelic variant, despite showing high diversity for other genetical markers. Analysis of published data indicated that the prevalence of this allelic variant of a major vaccine candidate antigen was already dominant since 1998. This phenomenon, which had not been hitherto recorded for a malaria blood stage antigen, is of biological and immunological interest but remains unexplained.     

Monitoring of in vitro susceptibilities and molecular markers of resistance of Plasmodium falciparum isolates from Thai-Myanmar border to chloroquine, quinine, mefloquine and artesunate

Malaria is one of the major causes of morbidity and mortality worldwide. The major factor which has aggravated the situation is the emergence of multidrug resistant Plasmodium falciparum malaria. To successfully deal with the problem, thorough understanding of the molecular bases for reduced parasite sensitivity to existing antimalarial drugs is of considerable importance. The objective of this work was to broaden the insight into the molecular mechanisms of resistance of P. falciparum to quinoline-containing antimalarials and artemisinin derivatives. Polymorphisms of the candidate genes pfmdr1 and pfcrt were investigated in relation to the susceptibility (in vitro sensitivity) of P. falciparum isolates to chloroquine (CQ), mefloquine (MQ), quinine (QN) and the artemisinin derivative - artesunate (AS). A total of 26 P. falciparum isolates were successful cultured. In vitro sensitivity results indicate the increase in susceptibility of P. falciparum strains in Thailand to CQ, while the susceptibility to MQ and QN was markedly declined. The pattern of cross-resistance was observed between MQ vs QN vs AS. Only one point mutation in the pfmdr1 gene, i.e., N86Y was observed with low prevalence of 7.7% (2/26). In contrast, the mutations at positions 76T, 220S, 271E, 326S, 356T and 371I in the pfcrt gene were identified in almost all isolates (25 isolates, 96.2%). The association between polymorphisms of the pfmdr1 and susceptibility of the parasite to MQ and QN was observed (increased susceptibilities to MQ and QN in isolates with mutations). Moreover, the correlation between pfmdr1 gene amplification and susceptibility of the parasite to MQ, QN and AS was observed (decreased susceptibilities to MQ, QN and AS in isolates with increased pfmdr1 copy number).     

Novel point mutations in sulfadoxine resistance genes of Plasmodium falciparum from India

Point mutations in the dhfr and dhps genes of Plasmodium falciparum are associated with pyrimethamine and sulfadoxine resistance respectively. In this study we have analyzed these genes from Bikaner (situated in North-West region of India), where both uncomplicated and severe manifestations of P. falciparum malaria are seen. A majority of isolates showed double mutant allele for DHFR. In contrast, the only reported mutation present in DHPS was A437G in few isolates. In addition, three novel non-synonymous mutations were observed in the PfDHPS from this region viz., S587F, N666K and C668W. The mutations at the 666 and 668 codon seem to form a bend in the big loop region of the DHPS enzyme and may affect the binding of the drug to the enzyme. Molecular docking of sulfadoxine to this mutated structure indicates reduction in its binding affinity to this enzyme.     

A better resolution for integrating methods for monitoring Plasmodium falciparum resistance to antimalarial drugs

Effective chemotherapy is the mainstay of malaria control. However, resistance of falciparum malaria to antimalarial drugs compromised the efforts to eliminate the disease and led to the resurgence of malaria epidemics. Three main approaches are used to monitor antimalarial drug efficacy and drug resistance; namely, in vivo trials, in vitro/ex vivo assays and molecular markers of drug resistance. Each approach has its implications of use as well as its advantages and drawbacks. Therefore, there is a need to use an integrated approach that would give the utmost effect to detect resistance as early as its emergence and to track it once spread. Such integration becomes increasingly needed in the era of artemisinin-based combination therapy as a forward action to deter resistance. The existence of regional and global networks for the standardization of methodology, provision of high quality reagents for the assessment of antimalarial drug resistance and dissemination of open-access data would help in approaching an integrated resistance surveillance system on a global scale.     

Plasmodium vivax in the Democratic Republic of East Timor: Parasite prevalence and antifolate resistance-associated mutations

In the Democratic Republic of East Timor, Plasmodium falciparum and Plasmodium vivax malaria coexist, but limited information is available about the latter species. Consequently, the prevalence of P. vivax and of its corresponding antifolate resistance-associated mutations in the pvdhfr and pvdhps genes was assessed here. Blood samples were collected from 650 individuals distributed among six districts, over two different periods, by either passive case detection (PCD) or active case detection (ACD). As expected, malaria was over-represented in the PCD sample (26% PCD vs 5% ACD), because the infection increases medical care seeking. Additionally, the relative frequency of P. vivax infections in symptomatic individuals (37%) was twice as high as the one in the asymptomatic sampling group (18%), suggesting that that this parasite is accounting for a significant proportion malaria-attributed morbidity. The frequency of specific sulfadoxine-pyrimethamine resistance-associated mutations genes was ascertained in P. vivax positive samples by PCR-RFLP. Although no mutants were detected in codons 383 and 553 of pvdhps, 48%, 76% and 82% of P. vivax-infected samples harbored the dhfr 33L, 58R and 117N mutations, respectively. Additionally, the frequency of parasites carrying both pvdhfr 58R and 117N mutant alleles accounted for a third of all genotypes analyzed, most likely due to inadvertent SP use in the past. In conclusion, evidence-based information is provided to promote optimized drug deployment and limit the evolution of resistance to antifolate resistance in P. vivax from East Timor.     

Naturally acquired antibody responses to the components of the Plasmodium falciparum merozoite surface protein 1 complex

Merozoite surface protein 6 (MSP6) and 7 (MSP7) of Plasmodium falciparum are peripheral membrane proteins whose cleaved products, MSP636, MSP722 and MSP719, are found on the merozoite surface as components of a non-covalently bound complex which also contains four polypeptides derived from merozoite surface protein 1 (MSP1). We have expressed both the precursor regions and the processed mature products of MSP6 and MSP7 in Escherichia coli and showed that these recombinant proteins react with human immune sera. In a set of sera collected from individuals living in malaria-endemic areas of Southern-central Vietnam, antibodies to the mature polypeptides of MSP636 and MSP722 were detected in 50.6 and 85.6% of the serum samples, whereas antibodies to the precursor regions of MSP6 and MSP7 were detected in only 12.1 and 42.5% of the serum samples, respectively. The predominant subclass of anti-MSP6 antibodies was IgG1, whereas the predominant subclass of anti-MSP7 antibodies was IgG3. In the same set of serum samples, the antibody responses to MSP119 are predominantly IgGI, whereas antibodies to merozoite surface protein 4 (MSP4) are mainly IgG3. This data is consistent with the proposition that, during malaria infection, variable proteins induce responses that are predominantly of the IgG3 isotype, and conserved proteins induce responses that are predominantly IgG1. The antibodies to MSP6, MSP7 and MSP119 all decreased at the time of infection, but increased during the convalescent period. No correlation was observed between the antibodies at the commencement of the study and absence of parasitaemia during surveillance in this population.     

Subtle changes in Plasmodium falciparum infection complexity following enhanced intervention in Malawi

With support from the Global Fund, the United States President's Malaria Initiative (PMI) and other cooperating partners, Malawi is implementing a comprehensive malaria control programme involving indoor residual spraying in targeted districts, universal coverage with insecticide-treated bed nets, use of rapid diagnostic tests to confirm the clinical diagnosis of malaria and use of the highly effective artemisinin-based combination therapy, artemether-lumefantrine (AL), as the first-line treatment for malaria. We genotyped 24 genome-wide single nucleotide polymorphisms (SNPs) in Plasmodium falciparum infections (n = 316) sampled from a single location in Malawi before (2006 and 2007) and after enhanced intervention (2008 and 2012). The SNP data generated were used to examine temporal changes in the proportion of multiple-genotype infections (MIs), mean number of heterozygous SNPs within MIs, parasite genetic diversity (expected heterozygosity and genotypic richness), multilocus linkage disequilibrium and effective population size (N_e). While the proportion of MIs, expected heterozygosity, genotypic richness, multilocus linkage disequilibrium and N_e were unchanged over time, the mean number (±standard deviation) of heterozygous SNPs within MIs decreased significantly (p = 0.01) from 9(±1) in 2006 to 7(±1) in 2012. These findings indicate that the genetic diversity of P. falciparum malaria parasites in this area remains high, suggesting that only subtle gains, if any, have been made in reducing malaria transmission. Continued surveillance is required to evaluate the impact of malaria control interventions in this area and the rest of Malawi, and to better target control interventions.     

Origins and spread of pfdhfr mutant alleles in Plasmodium falciparum

The emergence and spread of Plasmodium falciparum parasite resistant to sulfadoxine and pyrimethamine (SP) poses a serious public health problem. Resistance is caused by point mutations in dihydrofolate reductase (pfdhfr) and dihydropteroate synthase (pfdhps), the two key enzymes in the folate biosynthetic pathway. The use of microsatellite markers flanking pfdhfr has recently shown that the invasion of limited resistant lineages may explain the widespread SP resistance in many endemic regions. In Africa, however, multiple indigenous origins of pfdhfr triple mutants have been demonstrated. More new independent lineages and routes of geographical spread of resistance may be found by further molecular evolutionary analyses using samples from various endemic regions. Here, I review recent studies about the history of SP usage and the evolution and spread of resistant lineages while addressing the technical issue of microsatellite analysis.     

Mutant Plasmodium falciparum chloroquine resistance transporter in Hodeidah,Yemen: Association with parasitologic indices and treatment-seeking behaviors

Malaria still represents a major health problem in Yemen, particularly in Hodeidah, despite continuing efforts to eliminate it. With the absence of clinically proven vaccines, chemotherapy with antimalarials is still greatly needed. Chloroquine (CQ) has been popular as the drug of choice for malaria control. However, Plasmodium falciparum resistance to CQ has been one of the main obstacles in malaria control and elimination. Although CQ is no longer the recommended antimalarial chemotherapy, it has remained the number one over-the-counter antimalarial drug in many endemic areas, including Yemen, and is still used for self-medication. In addition, promising reports on CQ efficacy reversal in many African countries brought it again into the scene. This has led to a growing interest in the possibility of its re-introduction, particularly with the concerns raised about the parasite resistance to artemisinin-based combination therapies. Therefore, the present study aimed at analyzing the CQ-associated pfcrt 76T mutation in P. falciparum isolates from patients with uncomplicated falciparum malaria in Hodeidah, west of Yemen. The association of treatment-seeking behaviors and antimalarial drug use with the pfcrt 76T mutant allele was also studied. It was revealed that there is still a sustained high frequency of this molecular marker among parasite isolates associated with younger age, decreased parasite density and the presence of gametocytes in blood. Delay in seeking treatment and frequent use of antimalarials were the behaviors significantly associated with the presence of the pfcrt 76T mutant allele among patients reporting a history of malaria treatment.     

恶性疟原虫主要裂殖子表面蛋白1C末端19ku片段(MSP1-19)的表达及免疫效应检测

目的 探讨恶性疟原虫主要裂殖子表面蛋白1C末端19ku片段（MSP1-19）重组蛋白的免疫活性。方法 利用毕赤酵母高效表达系统分泌表达MSP1-19，表达产物纯化后免疫新西兰兔，3次免疫后ELISA检测其血清中IgG滴度的变化，观测重组MSP“。免疫效应。结果 MSP1-19在毕赤酵母高效表达；重组MSP1-19免疫后，兔血清中IgG滴度的变化与对照组有显著差异。结论 MSP1-19重组蛋白具有较好的免疫原性。     

Antibody specificities of children living in a malaria endemic area to inhibitory and blocking epitopes on MSP-119 of Plasmodium falciparum

Merozoite surface protein-1₁₉ (MSP-1₁₉) specific antibodies which include processing inhibitory, blocking and neutral antibodies have been identified in individuals exposed to Plasmodium falciparum. Here we intend to look at the effect of single and multiple amino acid substitutions of MSP-1₁₉ on the recognition by polyclonal antibodies from children living in Igbo-Ora, Nigeria. This would provide us with information on the possibility of eliciting mainly processing inhibitory antibodies with a recombinant MSP-1₁₉ vaccine. Blood was collected from children in the rainy season and binding of anti-MSP-1₁₉ antibodies to modified mutants of MSP-1₁₉ was analysed by ELISA. The MSP-1₁₉ mutant proteins with single substitutions at positions 22 (Leu → Arg), 43 (Glu → Leu) and 53 (Asn → Arg) and the MSP-1₁₉ mutant protein with multiple substitutions at positions 27 + 31 + 34 + 43 (Glu → Tyr, Leu → Arg, Tyr → Ser, Glu → Leu); which had inhibitory epitopes; had the highest recognition. Children recognised both sets of mutants with different age groups having different recognition levels. The percentage of malaria positive individuals (32-80%) with antibodies that bound to the mutants MSP-1₁₉ containing epitopes that recognise only processing inhibitory and not blocking antibodies, were significantly different from those with antibodies that did not bind to these mutants (21-28%). The amino acid substitutions that abolished the binding of blocking antibodies without affecting the binding of inhibitory antibodies are of particular interest in the design of MSP-1₁₉ based malaria vaccines. Although these MSP-1₁₉ mutants have not been found in natural population, their recognition by polyclonal antibodies from humans naturally infected with malaria is very promising for the future use of MSP-1₁₉ mutants in the design of a malaria vaccine.     

大肠埃希菌表达的恶性疟原虫裂殖子表面蛋白MSP1-42的免疫原性

目的纯化大肠埃希菌Rosettagami（DE3）表达的可溶性裂殖子表面蛋白1C末端蛋白（MSP1-42,3D7株）,检测其体外抑制恶性疟原虫生长的效果。方法利用大肠埃希菌Rosettagami（DE3）为宿主菌诱导表达MSP1-42,用带组氨酸标签的镍柱纯化可溶性的MSP1-42。6只新西兰白兔随机分为免疫组和对照组,每组3只。用MSP142与弗氏佐剂乳化后,皮下注射,抗原免疫剂量每次为200“g／只,共免疫4次,每次间隔2周。佐剂对照组以PBS代替抗原同法免疫。免疫前及末次免疫2周后取血,用酶联免疫吸附试验和间接荧光抗体试验检测血清中特异性抗体及其与天然抗原的反应,用含10％和20％免疫血清的培养基体外培养恶性疟原虫（海南分离株,Fcc1／HN）,检测免疫血清体外抑制恶性疟原虫生长的效果。结果经镍柱纯化后获得纯度为95％以上的MSP1-42蛋白;免疫组个体在第4次免疫后血清特异性抗体的滴度依次为1：640000、1：640000、1：160000,间接荧光抗体试验检测表明MSP1-42免疫兔血清与恶性疟原虫表面蛋白有阳性反应;免疫血清能抑制恶性疟原虫在体外生长,在10％浓度时,3只免疫兔血清的抑制率依次为（51．94-24．2）％、（29．4±8．6）％和（86．7±7．4）％,在20％浓度时,抑制率分别为（93．3±7．5）％、（65．3±10．6）％和（96．4±1．0）％。结论大肠埃希菌Rosettagami（DE3）表达的MSP142蛋白免疫血清能识别恶性疟原虫天然抗原,体外培养能抑制恶性疟原虫的生长。