Serologic Markers for Detecting Malaria in Areas of Low Endemicity,Somalia, 2008

Areas in which malaria is not highly endemic are suitable for malaria elimination, but assessing transmission is difficult because of lack of sensitivity of commonly used methods. We evaluated serologic markers for detecting variation in malaria exposure in Somalia. Plasmodium falciparum or P. vivax was not detected by microscopy in cross-sectional surveys of samples from persons during the dry (0/1,178) and wet (0/1,128) seasons. Antibody responses against P. falciparum or P. vivax were detected in 17.9% (179/1,001) and 19.3% (202/1,044) of persons tested. Reactivity against P. falciparum was significantly different between 3 villages (p<0.001); clusters of seroreactivity were present. Distance to the nearest seasonal river was negatively associated with P. falciparum (p = 0.028) and P. vivax seroreactivity (p = 0.016). Serologic markers are a promising tool for detecting spatial variation in malaria exposure and evaluating malaria control efforts in areas where transmission has decreased to levels below the detection limit of microscopy.     

Vaxfectin enhances both antibody and in vitro T cell responses to each component of a 5-gene Plasmodium falciparum plasmid DNA vaccine mixture administered at low doses

We previously reported the capacity of the cationic lipid-based formulation, Vaxfectin^®, to enhance the immunogenicity and protective efficacy of a low dose plasmid DNA vaccine against Plasmodium yoelii malaria in mice. Here, we have extended this finding to human Plasmodium falciparum genes, evaluating the immune enhancing effect of Vaxfectin^® formulation on a mixture, designated CSLAM, of five plasmid DNA vaccines encoding antigens from the sporozoite (PfCSP, PfSSP2/TRAP), intrahepatic (PfLSA1), and erythrocytic (PfAMA1, PfMSP1) life cycle stages of P. falciparum administered at 2, 10 or 50 μg doses. Vaxfectin^® formulation enhanced both antibody and cellular immune responses to each component of the multi-antigen vaccine mixture, as assessed by ELISA, IFAT, and IFN-γ ELIspot, respectively. There was no apparent antigenic competition, as indicated by comparison of responses induced in mice immunized with PfCSP vs. CSLAM. These data showing that Vaxfectin^® can enhance the immunogenicity of plasmid DNA vaccines administered at low doses per body weight, and in combinations, has important clinical implications for the development of a vaccine against malaria, as well as against other public health threats.     

Differences in automated depolarization patterns of Plasmodium faiciparum and P. vivax malaria infections defined by the Cell-Dyn® CD4000 haematology analyser

Of 1014 samples submitted for full blood count analysis and malaria screening, 854 were designated malaria-negative by blood film microscopy, 79 were unequivocally identified as Plasmodium vivax and 81 as P. falciparum. All samples were additionally analysed with the Abbott Cell-Dyn® CD4000 haematology instrument, and leucocyte differential plots of 90° polarized vs. 90° depolarized (NEU-EOS plot) and 90° depolarized vs. 0° light (EOS I plot) scatter were specifically examined for abnormal depolarization patterns. Depolarization pattern types were correlated with microscopy (species) results, and these correlations were consolidated by polymerase chain reaction analysis. All 854 microscopically-designated malaria-negative samples showed a type 1 (normal) CD4000 depolarization pattern. Abnormal pattern types 2, 3a and 3b were entirely restricted to one of the two malaria categories. Plasmodium falciparum malaria showed two CD4000 pattern types only; a ‘normal’ type 1 pattern was seen in 36/75 (48%) cases and the remaining 39 cases were all abnormal pattern type 3a. In contrast, most (79/85) P. vivax malaria cases showed a distinctive clustered EOS I population (types 2 and 3b patterns) that was not seen with P. falciparum. Automated depolarization analysis provides an effective means of detecting malaria-associated haemozoin, and the patterns of intracellular haemozoin further appear to provide species differentiation between P. falciparum and P. vivax.     

DNA Sequence Polymorphism of the Lactate Dehydrogenase Genefrom Iranian Plasmodium vivax and Plasmodium falciparum Isolates

Background:

Parasite lactate dehydrogenase (pLDH) is extensively employed as malaria rapid diagnostic tests (RDTs). Moreover, it is a well-known drug target candidate. However, the genetic diversity of this gene might influence performance of RDT kits and its drug target candidacy. This study aimed to determine polymorphism of pLDH gene from Iranian isolates of P. vivax and P. falciparum.

Methods:

Genomic DNA was extracted from whole blood of microscopically confirmed P. vivax and P. falciparum infected patients. pLDH gene of P. falciparum and P. vivax was amplified using conventional PCR from 43 symptomatic malaria patients from Sistan and Baluchistan Province, Southeast Iran from 2012 to 2013.

Results:

Sequence analysis of 15 P. vivax LDH showed fourteen had 100% identity with P. vivax Sal-1 and Belem strains. Two nucleotide substitutions were detected with only one resulted in amino acid change. Analysis of P. falciparum LDH sequences showed six of the seven sequences had 100% homology with P. falciparum 3D7 and Mzr-1. Moreover, PfLDH displayed three nucleotide changes that resulted in changing only one amino acid. PvLDH and PfLDH showed 75%–76% nucleotide and 90.4%–90.76% amino acid homology.

Conclusion:

pLDH gene from Iranian P. falciparum and P. vivax isolates displayed 98.8–100% homology with 1–3 nucleotide substitutions. This indicated this gene was relatively conserved. Additional studies can be done weather this genetic variation can influence the performance of pLDH based RDTs or not.     

Severe Plasmodium vivax Malaria in Pakistan

To compare the severity of Plasmodium vivax malaria with that of P. falciparum malaria, we conducted a retrospective cross-sectional study of 356 adults hospitalized with malaria (2009–2011) in Pakistan. P. vivax and P. falciparum accounted for 83% and 13% of cases, respectively; 79.9% of patients with severe malaria were infected with P. vivax.     

Molecular Detection of Malaria in South Punjab with Higher Proportion of Mixed Infections

Background

Malaria is well known for its fatalities worldwide, Plasmodium vivax and the Plasmodium falciparum are the two important species of malaria reported from Pakistan and creating lots of morbidities across the country.

Method

Study was conducted to determine the Surveillance of malaria in South Punjab by microscopy and Polymerase chain reaction (PCR).

Result

samples out of 100 patients were found positive for malarial parasites. One patient was found with mixed infection, whereas P. falciparum and P. vivax infections were detected in 17 and 22 patients, respectively. In nested PCR, genus-specific primers for Plasmodium species. in round 1 and species-specific primers for P. falciparum and P. vivax in round 2 were used. By the application of PCR 41% were found to be infected by Plasmodium spp. Among Plasmodium positive patients: mixed, P. falciparum and P. vivax infection were detected in 10, 15 and 16 patients, respectively. Thirty nine microscopically positive patients confirmed to have Plasmodium spp. One negative by PCR, 2 microscopically negative patients had shown Plasmodium spp. infection (P. falciparum and P. vivax) by PCR. In total samples, P. falciparum, P. vivax and mixed infection accounted for 36.6%, 39.0% and 24.3%, respectively.

Conclusion

Microscopy was found deficient for interpretation of mixed infections, low parasitaemia, and species specific diagnosis. The sensitivity, specificity and efficacy of nested PCR was calculated 95%, 98% and 97%, respectively, showing PCR as a more effective and efficient diagnostic tool for malaria.     

Assessment of routine malaria diagnosis in the Venezuelan Amazon

The quality of routine malaria diagnosis is a crucial topic of malaria control. The aim of this assessment was to monitor and evaluate the quality of routine malaria diagnosis in Amazonas (Venezuela) and to improve the quality control system. The traditional non-blinded quality control system was found to be overburdened with diagnostic samples. A modified sampling system with fewer samples to be tested was proposed. Expert microscopists blindly double-checked 1000 slides and 550 rapid diagnostic tests (RDT) (OptiMAL-IT®) from health posts (HP). For Plasmodium vivax, HP microscopy and OptiMAL-IT® showed sensitivies of 86% and 63%, respectively. For P. falciparum, HP microscopy and OptiMAL-IT® showed sensitivities of 68% and 89%, respectively. Both methods lost accuracy when fewer parasites occurred in the sample. HP microscopists from different municipalities displayed significant differences in diagnostic quality. Overall, quality of routine malaria diagnosis in the Venezuelan Amazon is good but not optimal. The change from the traditional non-blinded quality control system to blinded cross-checking of a minimal selection of samples is - comparatively - a low cost intervention with possibly high impact on the quality of routine malaria diagnosis. The introduction of RDTs should be discussed carefully in order not to displace an existing network of HP microscopists.     

Detection of malaria in relation to fever and grade of malnutrition among malnourished children in Ethiopia

Setting:

Forty-eight nutritional rehabilitation centres in southern Ethiopia.

Objective:

To determine 1) the frequency of temperature recording under programme conditions, 2) the proportion of malnourished children with and without fever who had falciparum malaria and 3) the association between malaria and grade of malnutrition.

Design:

This was a retrospective analysis of routine programme data.

Results:

Of 19 200 malnourished children, 16 716 (mean age 4.4 years, 7412 males) underwent a rapid malaria diagnostic test (Paracheck Pf ^®). Malnutrition was graded as severe (38%), moderate (35%) and mild (27%). Temperature was not recorded in 15 248 (91%) children. Malaria was diagnosed in 57 (28%) children with fever (n = 206) and 122 (10%) children with no fever (n = 1262). The prevalence of falciparum malaria was 9%. Malaria prevalence was significantly associated with grade of malnutrition: Paracheck Pf was positive in respectively 5%, 8% and 10% of children with mild, moderate and severe malnutrition (χ² for trend 78, P < 0.001).

Conclusions:

This study shows the value of routine malaria screening in malnourished children, especially those with more severe grades of malnutrition, irrespective of fever. Operational shortcomings are highlighted and ways forward to address these problems are discussed.     

Clinical Malaria along the China–Myanmar Border,Yunnan Province,China, January 2011–August 2012

Passive surveillance for malaria cases was conducted in Yunnan Province, China, along the China–Myanmar border. Infection with Plasmodium vivax and P. falciparum protozoa accounted for 69% and 28% of the cases, respectively. Most patients were adult men. Cross-border travel into Myanmar was a key risk factor for P. falciparum malaria in China.     

Immunization with genetically attenuated P. falciparum parasites induces long-lived antibodies that efficiently block hepatocyte invasion by sporozoites

Whole-parasite malaria vaccines have shown promise in clinical trials. We recently reported the first human trial of a malaria vaccine based on Plasmodium falciparum genetically attenuated parasites (PfGAP). Herein we report for the first time that PfGAP induces prolonged functional humoral responses in humans. Six volunteers were exposed to 5 bites of PfGAP-infected mosquitoes followed by approximately 200 bites. Plasma collected from all volunteers 3 months after the last exposure efficiently inhibits invasion of hepatocytes by P. falciparum sporozoites. The level of inhibition observed is comparable to that attained using plasma collected after 4–5 intravenously administrations of high numbers of irradiated sporozoites, validating the potential of PfGAP malaria vaccines. Our data highlight the role of antibody responses in pre-erythrocytic stages of human malaria, and suggests that to be protective, malaria vaccines might need to elicit long-lasting functional antibodies in addition to cellular responses.     

The efficiency of sporozoite transmission in the human malarias, Plasmodium falciparum and P. vivax

Reported are malaria sporozoite and inoculation rates over a 1-year period in eight epidemiologically defined villages of different endemicity in Madang Province, Papua New Guinea. In the study, more than 41 000 wild-caught mosquitos were analysed for Plasmodium falciparum and P. vivax sporozoites by ELISA. In a given village the entomological inoculation rates correlated strongly with the prevalences of both these malarial parasites in children. However, the prevalence of P. falciparum infections in children was much higher than that of P. vivax, despite similar inoculation rates for the two species. These data suggest that in Papua New Guinea P. falciparum is more efficiently transmitted than P. vivax from mosquito to man. The increased efficiency of transmission of P. falciparum may be due to the heavier sporozoite densities in wild-caught mosquitos naturally infected with P. falciparum sporozoites that were tenfold greater than the sporozoite densities in mosquitos infected with P. vivax.     

Plasmodium cynomolgi Co-infections among Symptomatic Malaria Patients,Thailand

Among 1,180 symptomatic malaria patients, 9 (0.76%) infected with Plasmodium cynomolgi were co-infected with P. vivax (n = 7), P. falciparum (n = 1), or P. vivax and P. knowlesi (n = 1). Patients were from Tak, Chanthaburi, Ubon Ratchathani, Yala, and Narathiwat Provinces, suggesting P. cynomolgi is widespread in this country.     

Aspects of imported malaria at a district general hospital in non-endemic Kuwait,Arabian Gulf

There is no indigenous mosquito-borne transmission of malaria in Kuwait. However, in a five year period at a district general hospital, the number of laboratory-diagnosed cases of malaria increased annually from 25 to 84, a rise of 336%. Except for two induced infections, all were imported, mainly from the Indian subcontinent. Plasmodium vivax was responsible for 87.29% of the cases; P. falciparum (12.05%), a mixed infection of P. vivax and P. falciparum (0.33%) and a case of P. ovale (0.33%) were also identified. Rapid preparation of acetone-fixed, Giemsa-stained thick blood films, a heightened awareness of the infection, examination of multiple samples of blood from patients and the general resurgence of malaria in endemic areas were some of the factors responsible for the high number of cases diagnosed. Most patients were young males and presented with clinical malaria due to P. vivax between May and October each year, an apparent seasonal peak. However, many were already resident in the country for a variable period. Patients with P. falciparum though, presented clinically within two weeks of arrival in the country. Parasite densities were calculated to monitor the progress of treatment and identify quickly any possible chloroquine-resistant P. falciparum strains. A policy of active prophylaxis is suggested to stem the tide of imported malaria.Corresponding author.     

A focus of hyperendemic Plasmodium malariae��P. vivax with no P. falciparum in a primitive population in the Peruvian Amazon jungle: Studies by means of immunofluorescence and blood smear

Findings in a sample population in southeastern Peru with a very high rate of malaria infection, due to Plasmodium malariae and P. vivax with apparently no P. falciparum, are described. The proportion of persons with P. malariae in this sample population, as determined by slide examination, appears to be the greatest ever reported for any area before the introduction of control measures. Although very few P. vivax were found on stained slides, results of the indirect immunofluorescence test indicated that this species was probably as prevalent as P. malariae; the absence of P. falciparum was supported by results of serologic tests. Possible reasons for this focus of malaria with no P. falciparum are discussed.     

In vivo Susceptibility of Plasmodium vivax to Chloroquine in Southeastern Iran

Background

Plasmodium vivax is the predominant species causes of malaria with about 90% total annual reported malaria in Iran. This study conducted to determine the susceptibility of Plasmodium vivax isolates to chloroquine in Sistan and Balochistan Province, southeastern Iran.

Methods

A total 270 subjects with symptomatic malaria and confirmed P. vivax infection completed the designed 28-day in vivo study. The thick and thin film blood smears were screened for malaria parasites by microscopy. The nested PCR was applied using the Plasmodium 18 subunit ribosomal ribonucleic (Ssr RNA) genes for detecting mixed infections and diagnosis of parasites in the samples with low parasite on days 0, 5, 6, 7, and 28.

Results

P. vivax was cleared in 15%, 50%, 95%, and 100% of patients on days 1, 2, 3, 4 respectively by microscopy assessment. Six patients were exhibited specific P. vivax band in nested PCR on day 5. No recurrence was observed on days 7, 14 and 28. Mean (±standard deviation) parasite clearance time was 2.41 (±0.8) days.

Conclusion

P. vivax is still susceptible to chloroquine in Southeatern Iran. This finding is compatible with results of neighboring countries Pakistan and Afghanistan.     

Antibodies to Pf155, a major antigen of Plasmodium falciparum: seroepidemiological studies in Haiti

The presence of malaria parasites and the serological antibody responses against whole Plasmodium falciparum and the Pf155 antigen were studied in the population of a small rural locality in Haiti in December 1985. Only 7 (1.5%) of the individuals were found to be infected with P. falciparum, the only species observed. Antibodies to P. falciparum were detected in an ELISA in 38.2% of the sera, the positivity rates being age-related. Anti-Pf155 antibodies were detected in 12.5% and 13.6% of individuals by two different techniques used. The anti-Pf155 positivity rates increased only after 25 years of age. No trends were detected for a clear-cut protective value of Pf155 antibodies against clinical malaria and further longitudinally conducted field surveys are needed to satisfactorily assess the potential protective effect of Pf155 antibodies.     

Co-infections with Plasmodium knowlesi and Other Malaria Parasites, Myanmar

To determine the frequency of co-infections with Plasmodium species in southern Myanmar, we investigated the prevalence of P. knowlesi. More than 20% of patients with malaria had P. knowlesi infection, which occurred predominantly as a co-infection with either P. falciparum or P. vivax.     

Plasmodium vivax Malaria among Military Personnel, French Guiana, 1998�C2008

We obtained health surveillance epidemiologic data on malaria among French military personnel deployed to French Guiana during 1998–2008. Incidence of Plasmodium vivax malaria increased and that of P. falciparum remained stable. This new epidemiologic situation has led to modification of malaria treatment for deployed military personnel.     

Paracheck-pf® Test Versus Microscopy in the Diagnosis of Falciparum Malaria in Arbaminch Zuria Woreda of South Ethiopia

Background

Malaria is a major cause of morbidity and mortality in Ethiopia. Rapid diagnostic tests such as Paracheck Pf are the major tools for falciparum malaria diagnosis as an alternative to microscopy in peripheral health facilities. The objective of this study was to evaluate the sensitivity and specificity of Paracheck Pf against microscopy for diagnosis of P.falciparum infection and observe the persistence of the antigen for an elongated period.

Methods

Cross sectional study was undertaken in Arbaminch Zuria at Shele health center from October 2008 to January 2009. Paracheck-Pf versus microscopy comparison was done in conjunction with an artemisinin-based combination therapy efficacy monitoring for a period of 28 days. Standard microscopic procedures were done by experienced laboratory technicians and paracheck-Pf was performed in accordance with the manufacturer''s instruction.

Results

out of 1293 examined blood films, 400(31%) were found to be malaria positive. Considering microscopy as the gold standard, paracheck-pf showed sensitivity of 94.1 %( 95%CI: 89.9–98.3%) and specificity of 80.0% (95%CI: 67.6–92.4%). The positive and negative predictive values were 93.3 %( 95%CI: 88.8–97.8%) and 82.1% (95%CI: 70–94.1%), respectively. Comparing microscopy results 98.7 % (79/80), 60% (48/80), 48.1% (37/77), and 44.6 %( 33/74) were also found to be positive by paracheck-pf at days7, 14, 21, and 28, respectively.

Conclusion

Paracheck Pf® has a comparable diagnostic performance in detecting P. falciparum infections through the persistence of frequent false positivity is a limitation. Thus, this diagnostic test is not appropriate for monitoring of treatment effect.     

Process development for the production of an E. coli produced clinical grade recombinant malaria vaccine for Plasmodium vivax

The global eradication of malaria will require the development of vaccines to prevent infection cause by Plasmodium vivax in addition to Plasmodium falciparum. In an attempt to contribute to this effort we have previously reported the cloning and expression of a vaccine based on the circumsporozoite protein of P. vivax. The synthetic vaccine encodes for a full-length molecule encompassing the N-terminal and C-terminal regions flanking a chimeric repeat region representing VK210 and VK247, the two major alleles of P. vivax CSP. The vaccine, designated vivax malaria protein 001 (VMP001), was purified to >95% homogeneity using a three-column purification scheme and had low endotoxin levels and passed the rabbit pyrogenicity assay. The protein is recognized by monoclonal antibodies directed against the two repeat motifs, as well as polyclonal antibodies. Immunization with VMP001 induced high titer antibodies in mice using Montanide ISA 720. We currently have more than 10,000 doses of purified bulk and 1800 vials of formulated bulk vaccine available for clinical testing and VMP001 is currently undergoing further development as a candidate vaccine to prevent malaria in humans.