Vivax Malaria Patients Research Articles

Distribution of malaria vectors and incidence of vivax malaria at Korean army installations near the demilitarized zone, Republic of Korea.

BackgroundAs a result of the reintroduction of malaria in the Republic of Korea (ROK) in 1993 and the threat to military and civilian populations, the Korea Military National Defense (MND) increased emphasis on vector control in 2012 at ROK Army (ROKA) installations located near the DMZ, while decreasing chemoprophylaxis, fearing potential drug resistance. Mosquito surveillance demonstrated a need for continuous monitoring of disease patterns among ROKA soldiers and vector malaria infection rates to ensure positive outcomes.MethodsAnopheles spp. were collected from May–October at three ROKA installations in three locations near the DMZ. Each of the areas included one installation <2 km and two installations 11–12 km from the DMZ in Paju and Yeoncheon counties, Gyeonggi Province. Anopheles spp. were identified by polymerase chain reaction (PCR) techniques and then assayed for the presence of vivax malaria sporozoites. The ROK MND reported vivax malaria patients monthly to Korea Centers for Disease Control and Prevention. Correlations for the incidence of Plasmodium vivax patients and infected Anopheles species were analysed using the Wilcoxon rank sum test, Pearson correlation test and liner regression analysis.ResultsA total of 4282 Anopheles spp. were collected. Anopheles kleini (69.5 %) was the most commonly collected, followed by Anopheles pullus (17.3 %), Anopheles belenrae (4.5 %), Anopheles sineroides (4.2 %), Anopheles sinensis sensu stricto (2.7 %), and Anopheles lesteri (1.9 %). Overall, 21 malaria patients were reported by the ROK MND. The monthly incidence of the malaria patients correlated with the monthly number of Plasmodium vivax sporozoite positive Anopheles spp. The monthly numbers of An.kleini demonstrated the highest correlations to the numbers of ROKA malaria patients throughout the mosquito season (P < 0.01). Anopheles spp. positive for P. vivax and malaria patients at ROKA installations located <2 km from the DMZ were higher than for ROKA installations located 11–12 km from DMZ.ConclusionThe number of Anopheles spp. positive for P. vivax sporozoites correlated with the monthly number of malaria cases and exposure of ROKA soldiers from May–October to P. vivax malaria infections. Malaria vector surveillance and vector control is warranted as part of an effective malaria management program at ROKA installations located near DMZ.

Clinicopathological Analysis and Multipronged Quantitative Proteomics Reveal Oxidative Stress and Cytoskeletal Proteins as Possible Markers for Severe Vivax Malaria.

In Plasmodium vivax malaria, mechanisms that trigger transition from uncomplicated to fatal severe infections are obscure. In this multi-disciplinary study we have performed a comprehensive analysis of clinicopathological parameters and serum proteome profiles of vivax malaria patients with different severity levels of infection to investigate pathogenesis of severe malaria and identify surrogate markers of severity. Clinicopathological analysis and proteomics profiling has provided evidences for the modulation of diverse physiological pathways including oxidative stress, cytoskeletal regulation, lipid metabolism and complement cascades in severe malaria. Strikingly, unlike severe falciparum malaria the blood coagulation cascade was not found to be affected adversely in acute P. vivax infection. To the best of our knowledge, this is the first comprehensive proteomics study, which identified some possible cues for severe P. vivax infection. Our results suggest that Superoxide dismutase, Vitronectin, Titin, Apolipoprotein E, Serum amyloid A, and Haptoglobin are potential predictive markers for malaria severity.

Assessing the impact of meteorological factors on malaria patients in demilitarized zones in Republic of Korea.

BackgroundThe trend of military patients becoming infected with vivax malaria reemerged in the Republic of Korea (ROK) in 1993. The common explanation has been that infective Anopheles mosquitoes from the Democratic People’s Republic of Korea have invaded Republic of Korea’s demilitarized zone (DMZ). The aim of this study was to verify the relationship between meteorological factors and the number of malaria patients in the military in this region.MethodsThe authors estimated the effects of meteorological factors on vivax malaria patients from the military based on the monthly number of malaria cases between 2006 and 2011. Temperature, precipitation, snow depth, wind velocity, relative humidity, duration of sunshine, and cloud cover were selected as the meteorological factors to be studied. A systematic pattern in the spatial distribution of malaria cases was assessed using the Moran’s Index. Granger causality tests and cross-correlation coefficients were used to evaluate the relationship between meteorological factors and malaria patients in the military.ResultsSpatial analysis revealed significant clusters of malaria patients in the military in Republic of Korea in 2011 (Moran’s I = 0.136, p-value = 0.026). In the six years investigated, the number of malaria patients in the military in Paju decreased, but the number of malaria patients in the military in Hwacheon and Chuncheon increased. Monthly average, maximum and minimum temperatures; wind velocity; and relative humidity were found to be predicting factors of malaria in patients in the military in Paju. In contrast, wind velocity alone was not able to predict malaria in Hwacheon and Chuncheon, however, precipitation and cloud cover were able to predict malaria in Hwacheon and Chuncheon.ConclusionsThis study demonstrated that the number of malaria patients in the military is correlated with meteorological factors. The variation in occurrence of malaria cases was principally attributed to differences in meteorological factors by regions of Republic of Korea.Electronic supplementary materialThe online version of this article (doi:10.1186/s40249-016-0111-3) contains supplementary material, which is available to authorized users.

Cytokine modulation of human blood viscosity from vivax malaria patients

Malaria is a major infectious disease in several countries and is caused by protozoa of the genus Plasmodium. In vivax malaria patients, inflammatory processes occur, as well as changes in cytokines and blood flow. The present study analyzed the cytokine modulation of blood viscosity from patients infected with Plasmodium vivax (P. vivax). Blood samples were collected from 42 non-infected individuals (control group) and 37 individuals infected with P. vivax. The IL-2, IL-4, IL-6, IL-10, TNFα, TGF-β and IL-17 cytokine concentrations in the serum were assessed, and the blood rheological properties were determined. The analysis of blood viscosity for shear rates revealed that the blood viscosity of the infected patients was significantly greater than that of the non-infected individuals. The viscosity of the blood was greater in the infected individuals than in the non-infected subjects. The serum from individuals with P. vivax infections exhibited higher IFN-γ and IL-17 concentrations and lower TGF-β levels. Incubation of the blood from infected individuals with IL-17 or IL-17 associated with IFN-γ reduced the viscosity to rates equivalent to the blood from non-infected individuals. Independently of cytokine modulation, no correlation was found between the parasitemia and blood viscosity of the infected patients. These data suggest that the alterations of blood viscosity are relevant as an auxiliary tool for the clinical diagnosis of disease. In malaria, erythrocytes are more sensitive to osmotic shock, and the reduction of viscosity by IL-17 may be related to a possible immunomodulator agent during infection.

Plasmodium vivax mdr1 genotypes in isolates from successfully cured patients living in endemic and non-endemic Brazilian areas.

BackgroundPlasmodium vivax is the most widely distributed species causing the highest number of malaria cases in the world. In Brazil, P. vivax is responsible for approximately 84 % of reported cases. In the absence of a vaccine, control strategies are based on the management of cases through rapid diagnosis and adequate treatment, in addition to vector control measures. The approaches used to investigate P. vivax resistance to chloroquine (CQ) were exclusively in vivo studies because of the difficulty in keeping parasites in continuous in vitro culture. In view of the limitations related to follow-up of patients and to assessing the plasma dosage of CQ and its metabolites, an alternative approach to monitor chemo-resistance (QR) is to use molecular markers. Single nucleotide polymorphisms (SNPs) in the multidrug resistance gene pvmdr1 are putative determinants of CQ resistance (CQR), but such SNPs in P. vivax isolates from patients with good response to treatment should be further explored. The aim of this study is to investigate the mutations in the gene, supposedly associated to QR, in P. vivax isolates from successfully cured patients, living in Brazilian endemic and non-endemic areas.MethodsBlood samples were collected from 49 vivax malaria patients from endemic (Amazon Basin: 45) and non-endemic (Atlantic Forest: four) Brazilian regions and analysed for SNPs in the CQR-related P. vivax gene (pvmdr1), using PCR-based methods.ResultsAmong the 49 isolates genetically characterized for the gene pvmdr1, 34 (70 %) presented at least one mutation. T958M mutant alleles were the most frequent (73 %) followed Y976F (15 %) and F1076L (12 %). Single mutation was detected in 24 (70.5 %) isolates and double mutations in ten (29.5 %). The most common single mutant genotype was the 958M/Y976/F1076 (79 %), followed by 976F/F1076 (21 %) whereas 958M/Y976/1076L (60 %) and 976F/1076L (40 %) double mutant genotypes were detected. Single mutant profile was observed only in isolates from Amazon Basin, although double mutants were found both in the Amazon and Atlantic Forest regions. Interestingly, the genotype 958M/Y976/1076L was present in all isolates from the Atlantic Forest in the Rio de Janeiro State.ConclusionsConsidering that primaquine (PQ) efficacy is highly dependent on concurrent administration of a blood schizontocidal agent and that PQ could not circumvent CQR, together with the fact that no pvmdr1 mutation should be expected in successfully cured patients, these findings seem to indicate that the pvmdr1 gene is not a reliable marker of CQR. Further investigations are needed to define a reliable molecular marker for monitoring P. vivax CQR in P. vivax populations.

A more appropriate white blood cell count for estimating malaria parasite density in Plasmodium vivax patients in northeastern Myanmar

The conventional method of estimating parasite densities employ an assumption of 8000 white blood cells (WBCs)/μl. However, due to leucopenia in malaria patients, this number appears to overestimate parasite densities. In this study, we assessed the accuracy of parasite density estimated using this assumed WBC count in eastern Myanmar, where Plasmodium vivax has become increasingly prevalent. From 256 patients with uncomplicated P. vivax malaria, we estimated parasite density and counted WBCs by using an automated blood cell counter. It was found that WBC counts were not significantly different between patients of different gender, axillary temperature, and body mass index levels, whereas they were significantly different between age groups of patients and the time points of measurement. The median parasite densities calculated with the actual WBC counts (1903/μl) and the assumed WBC count of 8000/μl (2570/μl) were significantly different. We demonstrated that using the assumed WBC count of 8000 cells/μl to estimate parasite densities of P. vivax malaria patients in this area would lead to an overestimation. For P. vivax patients aged five years and older, an assumed WBC count of 5500/μl best estimated parasite densities. This study provides more realistic assumed WBC counts for estimating parasite densities in P. vivax patients from low-endemicity areas of Southeast Asia.

"Not so benign" Plasmodium vivax malaria: An appraisal

Introduction: Plasmodium vivax infection-associated hematological changes including severe anemia can cause significant morbidity and indirect mortality. Objective: The objective of the study is to analyze hematological changes in P. vivax malaria patients and to review pathogenesis and cause of death in P. vivax infection. Materials and Methods: During a 1-year period, patients who were screened and confirmed as P. vivax by quantitative buffy coat and peripheral smear were included in the study. The demographic details and hematological profile (hemoglobin, total white blood cell count, and platelet count) were correlated. The severity of anemia was graded based on hemoglobin and correlated with parasitemia. Results: Two hundred patients were included. Twelve of them in addition showed Plasmodium falciparum. Thrombocytopenia (74.5%) was a significant feature compared to anemia (4.5%) and leukopenia (20.5%). Only three patients had hemoglobin level <6 g%. One of them (4.3 g%) developed relapse. During relapse, he had hemolytic anemia which required transfusion. He died on follow-up. All patients were treated with the first-line antimalarial drugs. Conclusions: In view of the inherent biological characteristics, P. vivax is more difficult to eradicate than P. falciparum. The pathogenesis of anemia is multifactorial including mechanical processes and immune mechanisms which enhance removal of infected and uninfected cells. This probably leads to severe anemia in the patient who died.

Whole-genome sequencing of a Plasmodium vivax isolate from the China-Myanmar border area

Currently, there is a trend of an increasing number of Plasmodium vivaxmalaria cases in China that are imported across its Southeast Asia border, especially in the China-Myanmar border area (CMB). To date, little is known about the genetic diversity of P. vivax in this region. In this paper, we report the first genome sequencing of a P. vivaxisolate (CMB-1) from a vivax malaria patient in CMB. The sequencing data were aligned onto 96.43% of the P. vivax Salvador I reference strain (Sal I) genome with 7.84-fold coverage as well as onto 98.32% of 14 Sal I chromosomes. Using the de novo assembly approach, we generated 8,541 scaffolds and assembled a total of 27.1 Mb of sequence into CMB-1 scaffolds. Furthermore, we identified all 295 known virgenes, which is the largest subtelomeric multigene family in malaria parasites. These results provide an important foundation for further research onP. vivax population genetics.

Identification of Immunodominant B-cell Epitope Regions of Reticulocyte Binding Proteins in Plasmodium vivax by Protein Microarray Based Immunoscreening.

Plasmodium falciparum can invade all stages of red blood cells, while Plasmodium vivax can invade only reticulocytes. Although many P. vivax proteins have been discovered, their functions are largely unknown. Among them, P. vivax reticulocyte binding proteins (PvRBP1 and PvRBP2) recognize and bind to reticulocytes. Both proteins possess a C-terminal hydrophobic transmembrane domain, which drives adhesion to reticulocytes. PvRBP1 and PvRBP2 are large (> 326 kDa), which hinders identification of the functional domains. In this study, the complete genome information of the P. vivax RBP family was thoroughly analyzed using a prediction server with bioinformatics data to predict B-cell epitope domains. Eleven pvrbp family genes that included 2 pseudogenes and 9 full or partial length genes were selected and used to express recombinant proteins in a wheat germ cell-free system. The expressed proteins were used to evaluate the humoral immune response with vivax malaria patients and healthy individual serum samples by protein microarray. The recombinant fragments of 9 PvRBP proteins were successfully expressed; the soluble proteins ranged in molecular weight from 16 to 34 kDa. Evaluation of the humoral immune response to each recombinant PvRBP protein indicated a high antigenicity, with 38-88% sensitivity and 100% specificity. Of them, N-terminal parts of PvRBP2c (PVX_090325-1) and PvRBP2 like partial A (PVX_090330-1) elicited high antigenicity. In addition, the PvRBP2-like homologue B (PVX_116930) fragment was newly identified as high antigenicity and may be exploited as a potential antigenic candidate among the PvRBP family. The functional activity of the PvRBP family on merozoite invasion remains unknown.

Characterization of Plasmodium vivax Early Transcribed Membrane Protein 11.2 and Exported Protein 1.

In Plasmodium, the membrane of intracellular parasites is initially formed during invasion as an invagination of the red blood cell surface, which forms a barrier between the parasite and infected red blood cells in asexual blood stage parasites. The membrane proteins of intracellular parasites of Plasmodium species have been identified such as early-transcribed membrane proteins (ETRAMPs) and exported proteins (EXPs). However, there is little or no information regarding the intracellular parasite membrane in Plasmodium vivax. In the present study, recombinant PvETRAMP11.2 (PVX_003565) and PvEXP1 (PVX_091700) were expressed and evaluated antigenicity tests using sera from P. vivax-infected patients. A large proportion of infected individuals presented with IgG antibody responses against PvETRAMP11.2 (76.8%) and PvEXP1 (69.6%). Both of the recombinant proteins elicited high antibody titers capable of recognizing parasites of vivax malaria patients. PvETRAMP11.2 partially co-localized with PvEXP1 on the intracellular membranes of immature schizont. Moreover, they were also detected at the apical organelles of newly formed merozoites of mature schizont. We first proposed that these proteins might be synthesized in the preceding schizont stage, localized on the parasite membranes and apical organelles of infected erythrocytes, and induced high IgG antibody responses in patients.

Immunoprofiling of the Tryptophan-Rich Antigen Family in Plasmodium vivax

Tryptophan-rich antigens (TRAgs) are an antigen family that has been identified in human and rodent malaria parasites. TRAgs have been proposed as candidate antigens for potential vaccines. The Plasmodium vivax TRAg (PvTRAg) family includes 36 members. Each PvTRAg contains a tryptophan-rich (TR) domain in the C-terminal region. In this study, we recombinantly expressed all 36 PvTRAgs using a cell-free expression system, and, for the first time, profiled the IgG antibody responses against all PvTRAgs in the sera from 96 vivax malaria patients and 40 healthy individuals using protein microarray technology. The mean seropositive rate for all PvTRAgs was 60.3%. Among them, nine PvTRAgs were newly identified in this study and showed a seropositive rate of >50%. Five of them, PvTRAg_13, PvTRAg_15, PvTRAg_16, PvTRAg_26, and PvTRAg_29, produced higher levels of IgG antibody, even in low-endemicity countries. In addition, the results of an immunofluorescence analysis suggest that PvTRAgs are, at least in part, associated with caveola-vesicle complexes, a unique structure of P. vivax-infected erythrocytes. The mechanism of formation and the function of these abundant membrane structures are not known. Further investigation aimed at determining the functions of these proteins would lead to a better understanding of the blood-stage biology of P. vivax.

Surmised total leucocyte counts miscalculate the parasite index of Plasmodium vivax malaria patients of tertiary and primary care settings in South-Western India

BackgroundFor the calculation of parasite index (PI) by microscopy method, an assumed total leucocyte count (TLC) of 8,000/μL is used conventionally. However, due to obvious variation in the population and individual TLCs, use of 8,000/μL may result in either over/underestimation of the PI.MethodsThis study was aimed at ascertaining the utility of 8,000/μL TLC, as well as other assumed TLCs, with respect to measured TLC for the calculation of PI. A tertiary care hospital and five primary health centres were the base for the prospective study conducted among microscopically proven, symptomatic Plasmodium vivax mono-infection patients aged ≥18 years. PIs calculated by assumed TLCs ranging from 4,000-11,000/μL were compared with those calculated by measured TLCs. Geometric mean with 95% confidence interval, Bland-Altman plot and Wilcoxon signed rank test were used for statistical analysis.ResultsA total of 284 P. vivax mono-infection patients, including 156 from a tertiary care hospital and 128 from five primary health centres, were recruited in the study. Assumed TLCs below 5,000 cell/μL and above 5,500 cell/μL in tertiary care setting resulted in significant (p <0.05) underestimation and overestimation, respectively. However, in primary health centres, it was an assumed TLC of 5,000 cell/μL, below and above which there was significant (p <0.05) underestimation and overestimation observed, respectively.ConclusionsAssumed TLC of 8,000/μL is not suitable for the calculation of PI. Either actual TLC of the patient should be measured or a representative TLC should be derived for the population under investigation for any study requiring calculated PI by microscopy.

Clinical profile of Plasmodium falciparum and Plasmodium vivax infections in low and unstable malaria transmission settings of Colombia.

BackgroundMalaria transmission in Latin America is generally hypoendemic and unstable, with Plasmodium vivax as the most prevalent species. However, only a few studies have been carried out in areas with low and unstable transmission, whereas the clinical profile of malaria has been broadly described in hyperendemic areas. The pattern of clinical manifestations and laboratory findings in low to moderate endemic areas of Colombia is reported here.MethodsA passive surveillance study was conducted between 2011 and 2013 involving 1,328 patients with Plasmodium falciparum, P. vivax or mixed malaria infections attending malaria points-of-care of four malaria endemic-areas with distinct transmission intensities and parasite distribution. Trained physicians recorded clinical symptoms and signs as well as socio-demographic characteristics of study participants. Haematological, biochemical and urine tests were performed at the time of diagnosis.ResultsOut of 1,328 cases, 673 (50.7%) were caused by P. vivax; 650 (48.9%) were due to P. falciparum; and five (0.4%) patients had mixed infections (P. falciparum/P. vivax). Most patients (92.5%) presented with uncomplicated malaria characterized by fever, chills, headache, sweating, myalgia/arthralgia and parasitaemia ≤ 20,000 parasites/μL. Fever, tachycardia, pallor and abdominal pain on palpation were more frequent in P. falciparum patients, whereas mild hepatomegaly and splenomegaly were mostly observed with P. vivax. Non-severe anaemia (Hb 7.0-10.9 g/dL) was observed in 20% of the subjects, whereas severe anaemia (Hb < 7.0 g/dL) was present in four patients. Half of the patients presented thrombocytopaenia regardless of parasite species. Leukopaenia, neutrophilia and monocytosis were frequently observed in patients infected with P. falciparum. Mild-to-moderate biochemical alterations were present in ~25% of the patients, particularly abnormal bilirubin in those with P. falciparum and abnormal transaminases in P. vivax malaria patients. Proteinuria was present in ~50% of the patients regardless of parasite species, whereas haemoglobinuria was more common in P. vivax infections. Only 7.5% of the cases were classified as clinically severe malaria, caused by both P. vivax and P. falciparum.ConclusionsThe high prevalence of uncomplicated malaria associated with moderate parasitaemia suggests the importance of timely diagnosis and effective treatment and encourages new activities to further decrease complicated malaria cases and mortality.

Severe vivax malaria: a systematic review and meta-analysis of clinical studies since 1900.

BackgroundMalaria caused by Plasmodium vivax was long considered to have a low mortality, but recent reports from some geographical areas suggest that severe and complicated vivax malaria may be more common than previously thought.MethodsThe primary objective of this systematic review and meta-analysis was to describe the reported clinical characteristics and the geographical variation in prevalence of reported severe vivax malaria and its change over time derived from English-language articles published since 1900. Medline and Scopus databases were searched for original papers on severe vivax malaria, using as inclusion criteria modified 2010 WHO criteria for the diagnosis of severe falciparum malaria. Articles before 1949 were identified through reference lists in journals, textbooks, and personal collections of colleagues.ResultsA total of 77 studies with reported severe vivax malaria and 63 studies with no reported severe vivax malaria (totaling 46,411 and 6,753 vivax malaria patients, respectively) were included. The 77 studies with reported severe vivax malaria were mainly from India (n = 33), USA (n = 8), Indonesia (n = 6), and Pakistan (n = 6). Vivax endemic countries not reporting severe vivax malaria beyond individual case reports included: the Greater Mekong Sub-region, China, North Korea, Bangladesh, Afghanistan, Middle East (except Qatar), the horn of Africa, and Madagascar. Only 17/77 reports were from before 2000. Vivax mono-infection was confirmed by PCR in 14 studies and co-morbidities were ruled out in 23 studies. Among the 77 studies reporting severe vivax malaria, severe thrombocytopenia (<50,000/mm3) was the most common “severe” manifestation (888/45,775 with pooled prevalence of 8.6%). The case fatality was 0.3% (353/46,411). Severity syndromes varied widely between different geographical areas, with severe anaemia being most prominent in areas of high transmission and chloroquine resistance.ConclusionPlasmodium vivax can cause severe and even fatal disease, but there is a recent increase in reports over the past 15 years with larger series restricted to a limited number of geographical areas. The biological basis of these variations is currently not known. More detailed epidemiological studies are needed which dissociate causation from association to refine the definition and estimate the prevalence of severe vivax malaria.Electronic supplementary materialThe online version of this article (doi:10.1186/1475-2875-13-481) contains supplementary material, which is available to authorized users.

Spatial variation in genetic diversity and natural selection on the thrombospondin-related adhesive protein locus of Plasmodium vivax (PvTRAP).

Thrombospondin-related adhesive protein (TRAP) of malaria parasites is essential for sporozoite motility and invasions into mosquito’s salivary gland and vertebrate’s hepatocyte; thereby, it is a promising target for pre-erythrocytic vaccine. TRAP of Plasmodium vivax (PvTRAP) exhibits sequence heterogeneity among isolates, an issue relevant to vaccine development. To gain insights into variation in the complete PvTRAP sequences of parasites in Thailand, 114 vivax malaria patients were recruited in 2006–2007 from 4 major endemic provinces bordering Myanmar (Tak in the northwest, n = 30 and Prachuap Khirikhan in the southwest, n = 25), Cambodia (Chanthaburi in the east, n = 29) and Malaysia (Yala and Narathiwat in the south, n = 30). In total, 26 amino acid substitutions were detected and 9 of which were novel, resulting in 44 distinct haplotypes. Haplotype and nucleotide diversities were lowest in southern P. vivax population while higher levels of diversities were observed in other populations. Evidences of positive selection on PvTRAP were demonstrated in domains II and IV and purifying selection in domains I, II and VI. Genetic differentiation was significant between each population except that between populations bordering Myanmar where transmigration was common. Regression analysis of pairwise linearized Fst and geographic distance suggests that P. vivax populations in Thailand have been isolated by distance. Sequence diversity of PvTRAP seems to be temporally stable over one decade in Tak province based on comparison of isolates collected in 1996 (n = 36) and 2006–2007. Besides natural selection, evidences of intragenic recombination have been supported in this study that could maintain and further generate diversity in this locus. It remains to be investigated whether amino acid substitutions in PvTRAP could influence host immune responses although several predicted variant T cell epitopes drastically altered the epitope scores. Knowledge on geographic diversity in PvTRAP constitutes an important basis for vaccine design provided that vaccination largely confers variant-specific immunity.

Plasmodium falciparum, but not P. vivax, can induce erythrocytic apoptosis.

BackgroundApoptosis can occur in red blood cells (RBC) and seems to be involved in hematologic disorders related to many diseases. In malaria it is known that parasitized RBC (pRBC) is involved in the development of anemia and thrombosis; however, non-parasitized RBC (nRBC) apoptosis could amplify these malaria-associated hematologic events. In fact, in experimental malaria, increased levels of apoptosis were observed in nRBC during lethal Plasmodium yoelii 17XL infection, but in human malaria erythrocytic apoptosis has never been studied. The present study was performed to investigate if nRBC apoptosis also occurs in P. vivax and P. falciparum infections.FindingsApoptosis of nRBC was evaluated in blood samples of P. vivax malaria patients and clinically healthly individuals living in Manaus, Brazil, both ex vivo and after incubation of RBC for 24 h. Additionally, the capacity of plasma from P. vivax or P. falciparum patients was tested for induction of in vitro apoptosis of normal RBC from a clinically healthy individual living in a non-endemic malaria region. Apoptosis was detected by flow cytometry using annexin V staining. In contrast to experimental malaria that significantly increased the levels of apoptotic nRBC both ex-vivo and after 24 h of incubation, no significant alteration on apoptotic nRBC rates was detected in P. vivax infected patients when compared with non-infected control individuals. Similar results were observed when plasma of these P. vivax patients was incubated with normal RBC. Conversely, plasma from P. falciparum-infected subjects induced significant apoptosis of these cells.ConclusionsApoptosis of normal RBC can be induced by plasma from individuals with P. falciparum (but not with P. vivax) malaria. This finding could reflect the existence of erythrocytic apoptosis during infection that could contribute to the pathogenesis of hematological and vascular complications associated with falciparum malaria.

Development and Efficacy of Real-Time PCR in the Diagnosis of Vivax Malaria Using Field Samples in the Republic of Korea

The development of sensitive, rapid, and accurate diagnostic methods for vivax malaria is essential for the effective control of malaria in the Republic of Korea, where vivax malaria patients usually show low parasitemia. In this study, a TaqMan-based real-time polymerase chain reaction (PCR) method was established and compared with other PCR-based assays, including nested PCR, loop-mediated isothermal amplification, and multiplex PCR, using samples from febrile patients with suspected vivax malaria. The established real-time PCR had a high sensitivity (99.6%) and specificity (100%). Therefore, this sensitive, specific, rapid, and quantitative real-time PCR method could be used for the routine diagnosis of vivax malaria in the laboratory of the Korea National Institute of Health.

Anopheles darlingi (Diptera: Culicidae) displays increased attractiveness to infected individuals with Plasmodium vivax gametocytes.

BackgroundMost hematophagous insects use host odours as chemical cues. The odour components, some physiological parameters and host attractiveness are affected by several conditions, including infection by parasites, e.g., plasmodia and, therefore, change the epidemiological scenario. This study evaluated the attractiveness of individuals with vivax malaria before, during (7 days) and after treatment (14 days) with specific antimalarial drugs.FindingsMosquito attractiveness to vivax-infected patients was assessed using a vertical olfactometer using the foot as a source of body odour. The ratio of Anopheles darlingi mosquitoes in the lower chamber of the olfactometer was used to calculate the attractiveness, and patient temperature was measured using a digital thermometer. An increased attractiveness was found only in patients bearing vivax gametocytes during the first experiment (early infection) (P < 0.001). Patients in the first experiment tended to have a higher body temperature, but grouping patients into fever and non-fever resulted in a higher attractiveness only in the fever group of gametocyte carriers, suggesting a synergistic effect of temperature and gametocytes in the host attractiveness to A. darlingi.ConclusionsGametocyte presence and fever in vivax malaria patients increased short distance host attractiveness to An. darlingi.

Profiling the humoral immune responses to Plasmodium vivax infection and identification of candidate immunogenic rhoptry-associated membrane antigen (RAMA)

Completion of sequencing of the Plasmodium vivax genome and transcriptome offers the chance to identify antigens among >5000 candidate proteins. To identify those P. vivax proteins that are immunogenic, a total of 152 candidate proteins (160 fragments) were expressed using a wheat germ cell-free system. The results of Western blot analysis showed that 92.5% (148/160) of the targets were expressed, and 96.6% (143/148) were in a soluble form with 67.7% of solubility rate. The proteins were screened by protein arrays with sera from 22 vivax malaria patients and 10 healthy individuals to confirm their immune profile, and 44 (27.5%, 44/160) highly reactive P. vivax antigens were identified. Overall, 5 candidates (rhoptry-associated membrane antigen [RAMA], Pv-fam-a and -b, EXP-1 and hypothetical protein PVX_084775) showed a positive reaction with >80% of patient sera, and 21 candidates with 50% to 80%. More than 23% of the highly immunoreactive proteins were hypothetical proteins, described for the first time in this study. One of the top immunogenic proteins, RAMA, was characterized and confirmed to be a serological marker of recent exposure to P. vivax infection. These novel immunoproteomes should greatly facilitate the identification of promising novel malaria antigens and may warrant further study. The establishment of high-throughput cloning and expression systems has permitted the construction of protein arrays for proteome-wide study of Plasmodium vivax. In this study, high-throughput screening assays have been applied to investigate blood stage-specific immune proteomes from P. vivax. We identified 44 antigenic proteins from the 152 putative candidates, more than 23% of which were hypothetical proteins described for the first time in this study. In addition, PvRAMA was characterized further and confirmed to be a serological marker of exposure to infections. The expression of one-third of the selected antigenic genes were shifted between P. vivax and Plasmodium falciparum, suggesting that these genes may represent important factors associated with P. vivax selectivity for young erythrocytes and/or with immune evasion. These novel immune proteomes of the P. vivax blood stage provide a baseline for further prospective serological marker studies in malaria. These methods could be used to determine immunodominant candidate antigens from the P. vivax genome.

Clinical complications of G6PD deficiency in Latin American and Caribbean populations: systematic review and implications for malaria elimination programmes

BackgroundAlthough G6PDd individuals are generally asymptomatic throughout their life, the clinical burden of this genetic condition includes a range of haematological conditions, including acute haemolytic anaemia (AHA), neonatal jaundice (NNJ) and chronic non-sphaerocytic anaemia (CNSA). In Latin America (LA), the huge knowledge gap regarding G6PDd is related to the scarce understanding of the burden of clinical manifestation underlying G6PDd carriage. The aim of this work was to study the clinical significance of G6PDd in LA and the Caribbean region through a systematic review.MethodsA systematic search of the published literature was undertaken in August 2013. Bibliographies of manuscripts were also searched and additional references were identified. Only original research was included. All study designs were included, as long as any clinical information was present. Studies were eligible for inclusion if they reported clinical information from populations living in LA or Caribbean countries or about migrants from these countries living in countries outside this continent.ResultsThe Medline search generated 487 papers, and the LILACS search identified 140 papers. After applying the inclusion criteria, 100 original papers with any clinical information on G6PDd in LA were retrieved. Additionally, 16 articles were included after reading the references from these papers. These 116 articles reported data from 18 LA and Caribbean countries. The major clinical manifestations reported from LA countries were those related to AHA, namely drug-induced haemolysis. Most of the published works regarding drug-induced haemolysis in LA referred to haemolytic crises in P. vivax malaria patients during the course of the treatment with primaquine (PQ). Favism, infection-induced haemolysis, NNJ and CNSA appear to play only a minor public health role in this continent.ConclusionHaemolysis in patients using PQ seems to be the major clinical manifestation of G6PDd in LA and contributes to the morbidity of P. vivax infection in this continent, although the low number of reported cases, which could be linked to under-reporting of complications. These results support the need for better strategies to diagnose and manage G6PDd in malaria field conditions. Additionally, Malaria Control Programmes in LA should not overlook this condition in their national guidelines.