Validation of micro-chip based PCR assays for diagnosis of both Plasmodium falciparum and Plasmodium vivax

Background & objectives: Microscopy is considered as the gold standard for malaria diagnosis, however sub-microscopic infections can only be detected by Polymerase chain reaction, which demands high cost and elaborate laboratory setup. The Micro-chip PCR based Truenat Malaria Pv-Pf and Pf assay is a portable solution for detection of sub-microscopic/asymptomatic cases of malaria in the field, three lots of which were evaluated for P. falciparum and P. vivax malaria. Methods: Three lots of Truenat® Malaria Pv-Pf and Pf assay (kits) were assessed using blood samples of P. vivax and P. falciparum as well as malaria negative blood samples. DNA was extracted from the blood samples using the Trueprep Auto v2 Universal Cartridge based sample prep device and real time qPCR was performed using Truelab DUO micro PCR Analyzer with three lots of Truenat® Malaria Pv-Pf and Pf Assays. Mean, Standard deviation and one-way analysis of variance (ANOVA) was used to assess the significance of inter-lot variability in Cycle threshold values. Results: The Truenat® Malaria Pv-Pf and Pf assays identified the malaria parasites with 100% accuracy. Based on the test for variance (ANOVA) the inter-lot variability in cycle threshold values were not significant, indicating a high degree of precision. Interpretation & conclusion: Based on high accuracy and precision between different lots, the Truenat® Malaria Pv-Pf and Pf assays were found to be suitable for the diagnosis of sub-microscopic infections in field conditions to provide support in elimination of malaria.

PCR-RFLP Based genetic diversity of Plasmodium vivax genotypes in district Mardan, Pakistan / Diversidade genética baseada em PCR-RFLP de genótipos de Plasmodium vivax no distrito de Mardan, Paquistão

Plasmodium vivax is the most common human malaria parasite in Asian countries including Pakistan. Present study was designed to explore the genetic diversity of plasmodium vivax genotypes based on Pvmsp-3α and Pvmsp-3ßgenes using allelic specific nested PCR and RFLP assays markers from field isolates in district Mardan, Pakistan. Blood samples of 200 P. vivax malarial patients were collected after taking their written informed consent. Genetic diversity in nested PCR products was determined by Restriction Fragment Length Polymorphism (RFLP) utilizing Alu1 and PstI restriction enzymes for alpha and beta gene products digestion, respectively. For analysis the genetic diversity of the sub allelic variants of Pvmsp3α and Pvmsp3ß genes, Chi-Square test was performed by utilizing Minitab programming software 18. The P value 0.05 was considered as statistically significant. For Pvmsp3α genes after gel electrophoresis of digested products, four distinct genotypes were obtained from total of 50 samples; type A: 35 (70%) (1.5-2.0 kb), 12 of type B (24%) (1.5-1.7 kb), 2 of type C (4%) (0.5-1.5) and one for type D (2%) (0.5-0.65 kb) which could be characterized into 9 allelic pattern (A1-A4, B1-B3, C1, D), in which A3 remained the most predominant. For Pvmsp-3ßgenes, three distinct genotypes were obtained from 50 samples; 40(80%) of type A (1.5-2.5 kb), 9 (18%) of type B (1.0-1.5kb) and 1(2%) of type C (0.65 kb) which could be characterized into 6 allelic patterns (A1-A3, B1-B2, and C1). Most dominant one in Type A was A1 alleles which were noted (46%), while in Type B, the most dominant were B1 (10%).This study is the first ever report of molecular epidemiology and genetic variation in Pvmsp-3α and Pvmsp-3ß genes of P. vivax isolates by using PCR/RFLP from District Mardan and showed a remarkable level of genetic diversity in the studied genes of circulating parasites in the study area. The results of this study will contribute in future studies about the genetic structure of parasite and vaccine development against the malaria.

O Plasmodium vivax é o parasita da malária humana mais comum nos países asiáticos, incluindo o Paquistão. O presente estudo foi desenhado para explorar a diversidade genética de genótipos de Plasmodium vivax baseados nos genes Pvmsp-3α e Pvmsp-3ß, usando marcadores de ensaios alélicos nested PCR e RFLP de isolados de campo no distrito de Mardan, Paquistão. Amostras de sangue de 200 pacientes com malária por P. vivax foram coletadas após assinatura do termo de consentimento livre e esclarecido. A diversidade genética em produtos de PCR nested foi determinada por polimorfismo de fragmento de restrição (RFLP) utilizando as enzimas de restrição Alu1 e PstI para a digestão dos produtos dos genes alfa e beta, respectivamente. Para análise da diversidade genética das variantes subalélicas dos genes Pvmsp3α e Pvmsp3ß, o teste Qui-quadrado foi realizado utilizando o software de programação Minitab 18. O valor P = 0,05 foi considerado estatisticamente significativo. Para os genes Pvmsp3α, após eletroforese em gel de produtos digeridos, quatro genótipos distintos foram obtidos de um total de 50 amostras; tipo A: 35 (70%) (1,5-2,0 kb), 12 do tipo B (24%) (1,5-1,7 kb), 2 do tipo C (4%) (0,5-1,5) e um para o tipo D (2%) (0,5-0,65 kb), que podem ser caracterizados em nove padrões alélicos (A1-A4, B1-B3, C1, D), em que A3 permaneceu como o mais predominante. Para Pvmsp-3ßgenes, três genótipos distintos foram obtidos a partir de 50 amostras; 40 (80%) do tipo A (1,5-2,5 kb), 9 (18%) do tipo B (1,0-1,5 kb) e 1 (2%) do tipo C (0,65 kb), que podem ser caracterizados em seis padrões alélicos (A1-A3, B1-B2 e C1). Os mais dominantes no tipo A foram o alelo A1, observados em 46%, enquanto, no tipo B, os mais dominantes foram B1 (10%). Este estudo é o primeiro relato de epidemiologia molecular e variação genética em Pvmsp-3α. Os genes Pvmsp-3ß de isolados de P. vivax utilizando PCR/RFLP do Distrito Mardan mostraram um nível notável de diversidade genética nos genes estudados de parasitas circulantes na área de estudo. Os resultados desse estudo contribuirão em estudos futuros sobre a estrutura genética do parasita e o desenvolvimento de vacinas contra a malária.

PCR-RFLP Based genetic diversity of Plasmodium vivax genotypes in district Mardan, Pakistan / Diversidade genética baseada em PCR RFLP de genótipos de Plasmodium vivax no distrito de Mardan, Paquistão

Plasmodium vivax is the most common human malaria parasite in Asian countries including Pakistan. Present study was designed to explore the genetic diversity of plasmodium vivax genotypes based on Pvmsp-3α and Pvmsp-3βgenes using allelic specific nested PCR and RFLP assays markers from field isolates in district Mardan, Pakistan. Blood samples of 200 P. vivax malarial patients were collected after taking their written informed consent. Genetic diversity in nested PCR products was determined by Restriction Fragment Length Polymorphism (RFLP) utilizing Alu1 and PstI restriction enzymes for alpha and beta gene products digestion, respectively. For analysis the genetic diversity of the sub allelic variants of Pvmsp3α and Pvmsp3β genes, Chi-Square test was performed by utilizing Minitab programming software 18. The P value 0.05 was considered as statistically significant. For Pvmsp 3α genes after gel electrophoresis of digested products, four distinct genotypes were obtained from total of 50 samples; type A: 35 (70%) (1.5-2.0 kb), 12 of type B (24%) (1.5-1.7 kb), 2 of type C (4%) (0.5-1.5) and one for type D (2%) (0.5-0.65 kb) which could be characterized into 9 allelic pattern (A1-A4, B1-B3, C1, D), in which A3 remained the most predominant. For Pvmsp-3βgenes, three distinct genotypes were obtained from 50 samples; 40(80%) of type A (1.5-2.5 kb), 9 (18%) of type B (1.0-1.5kb) and 1(2%) of type C (0.65 kb) which could be characterized into 6 allelic patterns (A1-A3, B1-B2, and C1). Most dominant one in Type A was A1 alleles which were noted (46%), while in Type B, the most dominant were B1 (10%).This study is the first ever report of molecular epidemiology and genetic variation in Pvmsp-3α and Pvmsp-3β genes of P. vivax isolates by using PCR/RFLP from District Mardan and [...].

O Plasmodium vivax é o parasita da malária humana mais comum nos países asiáticos, incluindo o Paquistão. O presente estudo foi desenhado para explorar a diversidade genética de genótipos de Plasmodium vivax baseados nos genes Pvmsp-3α e Pvmsp-3β, usando marcadores de ensaios alélicos nested PCR e RFLP de isolados de campo no distrito de Mardan, Paquistão. Amostras de sangue de 200 pacientes com malária por P. vivax foram coletadas após assinatura do termo de consentimento livre e esclarecido. A diversidade genética em produtos de PCR nested foi determinada por polimorfismo de fragmento de restrição (RFLP) utilizando as enzimas de restrição Alu1 e PstI para a digestão dos produtos dos genes alfa e beta, respectivamente. Para análise da diversidade genética das variantes subalélicas dos genes Pvmsp3α e Pvmsp3β, o teste Qui-quadrado foi realizado utilizando o software de programação Minitab 18. O valor P = 0,05 foi considerado estatisticamente significativo. Para os genes Pvmsp 3α, após eletroforese em gel de produtos digeridos, quatro genótipos distintos foram obtidos de um total de 50 amostras; tipo A: 35 (70%) (1,5-2,0 kb), 12 do tipo B (24%) (1,5-1,7 kb), 2 do tipo C (4%) (0,5-1,5) e um para o tipo D (2%) (0,5-0,65 kb), que podem ser caracterizados em nove padrões alélicos (A1-A4, B1-B3, C1, D), em que A3 permaneceu como o mais predominante. Para Pvmsp-3βgenes, três genótipos distintos foram obtidos a partir de 50 amostras; 40 (80%) do tipo A (1,5-2,5 kb), 9 (18%) do tipo B (1,0-1,5 kb) e 1 (2%) do tipo C (0,65 kb), que podem ser caracterizados em seis padrões alélicos (A1-A3, B1-B2 e C1). Os mais dominantes no tipo A foram o alelo A1, observados em 46%, enquanto, no tipo B, os mais dominantes foram B1 (10%). Este estudo é o primeiro relato de epidemiologia molecular e variação genética em Pvmsp-3α. Os genes Pvmsp-3β de isolados de P. vivax utilizando PCR/RFLP do Distrito Mardan mostraram um nível notável de diversidade genética nos genes estudados [...].

PCR-RFLP Based genetic diversity of Plasmodium vivax genotypes in district Mardan, Pakistan

Abstract Plasmodium vivax is the most common human malaria parasite in Asian countries including Pakistan. Present study was designed to explore the genetic diversity of plasmodium vivax genotypes based on Pvmsp-3 and Pvmsp-3genes using allelic specific nested PCR and RFLP assays markers from field isolates in district Mardan, Pakistan. Blood samples of 200 P. vivax malarial patients were collected after taking their written informed consent. Genetic diversity in nested PCR products was determined by Restriction Fragment Length Polymorphism (RFLP) utilizing Alu1 and PstI restriction enzymes for alpha and beta gene products digestion, respectively. For analysis the genetic diversity of the sub allelic variants of Pvmsp3 and Pvmsp3 genes, Chi-Square test was performed by utilizing Minitab programming software 18. The P value 0.05 was considered as statistically significant. For Pvmsp-3 genes after gel electrophoresis of digested products, four distinct genotypes were obtained from total of 50 samples; type A: 35 (70%) (1.5-2.0 kb), 12 of type B (24%) (1.5-1.7 kb), 2 of type C (4%) (0.5-1.5) and one for type D (2%) (0.5-0.65 kb) which could be characterized into 9 allelic pattern (A1-A4, B1-B3, C1, D), in which A3 remained the most predominant. For Pvmsp-3genes, three distinct genotypes were obtained from 50 samples; 40(80%) of type A (1.5-2.5 kb), 9 (18%) of type B (1.0-1.5kb) and 1(2%) of type C (0.65 kb) which could be characterized into 6 allelic patterns (A1-A3, B1-B2, and C1). Most dominant one in Type A was A1 alleles which were noted (46%), while in Type B, the most dominant were B1 (10%).This study is the first ever report of molecular epidemiology and genetic variation in Pvmsp-3 and Pvmsp-3 genes of P. vivax isolates by using PCR/RFLP from District Mardan and showed a remarkable level of genetic diversity in the studied genes of circulating parasites in the study area. The results of this study will contribute in future studies about the genetic structure of parasite and vaccine development against the malaria.

Resumo O Plasmodium vivax é o parasita da malária humana mais comum nos países asiáticos, incluindo o Paquistão. O presente estudo foi desenhado para explorar a diversidade genética de genótipos de Plasmodium vivax baseados nos genes Pvmsp-3 e Pvmsp-3, usando marcadores de ensaios alélicos nested PCR e RFLP de isolados de campo no distrito de Mardan, Paquistão. Amostras de sangue de 200 pacientes com malária por P. vivax foram coletadas após assinatura do termo de consentimento livre e esclarecido. A diversidade genética em produtos de PCR nested foi determinada por polimorfismo de fragmento de restrição (RFLP) utilizando as enzimas de restrição Alu1 e PstI para a digestão dos produtos dos genes alfa e beta, respectivamente. Para análise da diversidade genética das variantes subalélicas dos genes Pvmsp3 e Pvmsp3, o teste Qui-quadrado foi realizado utilizando o software de programação Minitab 18. O valor P = 0,05 foi considerado estatisticamente significativo. Para os genes Pvmsp-3, após eletroforese em gel de produtos digeridos, quatro genótipos distintos foram obtidos de um total de 50 amostras; tipo A: 35 (70%) (1,5-2,0 kb), 12 do tipo B (24%) (1,5-1,7 kb), 2 do tipo C (4%) (0,5-1,5) e um para o tipo D (2%) (0,5-0,65 kb), que podem ser caracterizados em nove padrões alélicos (A1-A4, B1-B3, C1, D), em que A3 permaneceu como o mais predominante. Para Pvmsp-3genes, três genótipos distintos foram obtidos a partir de 50 amostras; 40 (80%) do tipo A (1,5-2,5 kb), 9 (18%) do tipo B (1,0-1,5 kb) e 1 (2%) do tipo C (0,65 kb), que podem ser caracterizados em seis padrões alélicos (A1-A3, B1-B2 e C1). Os mais dominantes no tipo A foram o alelo A1, observados em 46%, enquanto, no tipo B, os mais dominantes foram B1 (10%). Este estudo é o primeiro relato de epidemiologia molecular e variação genética em Pvmsp-3. Os genes Pvmsp-3 de isolados de P. vivax utilizando PCR/RFLP do Distrito Mardan mostraram um nível notável de diversidade genética nos genes estudados de parasitas circulantes na área de estudo. Os resultados desse estudo contribuirão em estudos futuros sobre a estrutura genética do parasita e o desenvolvimento de vacinas contra a malária.

In vitro and ex vivo antiplasmodial activity of 1-(3-benzyloxy-4-methoxy-phenyl)-3-(3,4,5-trimethoxy-phenyl)-propan-1-one) against circulating strains of Plasmodium spp. in the state of Rondônia, Brazil

Abstract Malaria is a disease caused by Plasmodium spp. protozoa. The ability of Plasmodium to develop resistance to current antimalarial drugs makes the study of chemotherapeutic alternatives extremely important. This study aimed to evaluate the antimalarial activity of compound 3286938 (1-(3-benzyloxy-4-methoxy-phenyl)-3-(3,4,5-trimethoxy-phenyl)-propan-1-one), which presents in its structure a 3,4,5-trimethoxyphenyl group, in vitro, using the W2 strain of P. falciparum and against circulating strains of P. vivax and P. falciparum from the state of Rondônia. The compound 3286938 obtained an IC50 of 24.4 µM against the W2 strain of P. falciparum, and against the circulating strains, it presented a median (MD)=38.7 µM for P. vivax and MD=6.7 µM for P. falciparum. As for toxicity, 3286938 showed CC50 > 500 µM for VERO and HepG2 strains with a selectivity index greater than 12.9, a ratio calculated for P. falciparum and P. vivax regarding Vero and HepG2 cells. The compound was not considered hemolytic in in vitro assays, thus indicating the specificity of its antiplasmodial action. Based on the results presented, and considering the unprecedented character of the compound, it can be concluded that 3286938 was shown to be promising for complementary in vitro and in vivo studies aiming to produce effective antiplasmodial action.

Plasmodium vivax metacaspase 1 (PvMCA1) catalytic domain is conserved in field isolates from Brazilian Amazon

In the present study, we investigated the genetic diversity of Plasmodium vivax metacaspase 1 (PvMCA1) catalytic domain in two municipalities of the main malaria hotspot in Brazil, i.e., the Juruá Valley, and observed complete sequence identity among all P. vivax field isolates and the Sal-1 reference strain. Analysis of PvMCA1 catalytic domain in different P. vivax genomic sequences publicly available also revealed a high degree of conservation worldwide, with very few amino acid substitutions that were not related to putative histidine and cysteine catalytic residues, whose involvement with the active site of protease was herein predicted by molecular modeling. The genetic conservation presented by PvMCA1 may contribute to its eligibility as a druggable target candidate in vivax malaria.

Clinical Profile of Severe Plasmodium falciparum and P. vivax Malaria in Jazan Region, Saudi Arabia

@#Introduction: Malaria is the most common and widely prevalent life-threatening infectious disease in tropical and subtropical regions where it causes high rates of morbidity and mortality. Despite the successful elimination of malaria transmission in most of Saudi Arabia, the Jazan region is still considered to be a malaria-endemic area. Therefore, this study aimed at investigating the clinical profile of severe Plasmodium falciparum and P. vivax malaria in Jazan region, southwestern Saudi Arabia. Methods: A total of 25 febrile patients who presented at Sabya General Hospital and were suspected of having severe malaria were included in this study. Blood samples were collected and examined for malaria by rapid diagnostic test and by Giemsa-stained thin and thick blood films. Clinical and laboratory analyses to identify severe malaria complications were performed at the hospital. Results: In all, 76% (19/25) of the participants were infected with P. falciparum and 24% (6/25) had P. vivax. The most common complications in both groups were prostration (48%) followed by jaundice (44%) and liver dysfunction (36%). The prevalence of severe anaemia was significantly higher (P = 0.031) among P. vivax-infected patients (50%) compared with P. falciparum-infected patient (5.3%). In falciparum-infected patients, those with severe parasitaemia had a higher risk of developing prostration compared with those with low-to-moderate parasitaemia (P = 0.038). Conclusion: Most cases were caused by P. falciparum which required proper monitoring and treatment. However, those with P. vivax exhibited severe symptoms similar to those with P. falciparum.

A cross sectional study of thrombocytopenia in malaria positive cases in a tertiary care hospital of Bareilly

Background: To find out correlation of thrombocytopenia with malaria. Malaria is a protozoal disease caused by infection with parasite of genus Plasmodium. Thrombocytopenia is a common and early sign of malarial infection and 60-80% thrombocytopenia is observed in malarial cases and present more frequently and severe in complicated P. falciparum malaria.Methods: A cross sectional study done in Central Pathological Lab of Department of Pathology, RMCH, Bareilly. Blood samples collected in ethylenediaminetetraacetic acid vial and blood smear was examined for malaria parasite within red blood cells. Malaria rapid test was done for detection of Plasmodium species and platelet count was done.Results: 780 cases of malaria was studied from September 2018 to December 2018, male predominance of 54.5%, maximum malarial positive cases 26.92% in the age group of 21-30 years, maximum 86.28% cases were of P. vivax, and thrombocytopenia was observed in 91.54% cases.Conclusions: Mostly developing countries with limited resources and trained health manpower are malaria-endemic region of world. Thrombocytopenia is associated with both P. vivax and P. falciparum infections. In our study significance association between malaria and thrombocytopenia has been observed. We suggest malaria should be a consideration in all patients with fever and thrombocytopenia.

Analysis of clinical profile and prescription pattern of malaria in a tertiary care hospital in Karnataka, India

Background: Malaria is one of the leading causes of morbidity and mortality in developing countries like India. Plasmodium falciparum and Plasmodium vivax are the commonest species implicated for an increased incidence of malaria in India. The pattern of disease, signs, and symptoms vary from place to place, region to region due to demographic variations. The current study was undertaken to study the differences in the clinical profile of malaria, particularly signs and symptoms, complications and response to treatment in malaria.Methods: A retrospective, single center, surveillance study was carried out at a tertiary health care center in Mangalore. All patients aged above 18 years diagnosed as malaria by peripheral smear method and rapid diagnostic tests were included in the study. The clinical features, complications, and response to treatment were noted.Results: Fifty eight patients diagnosed as malaria were included in the study. Compared to other studies and nationwide incidences, here P. vivax emerged as the leading cause of malaria. All patients presented with fever varying from 3-20 days. About 30 patients complained of headache and 21 patients presented with malaise. In about 6 patient’s complications were seen. Majority of patients received artemisinin derivatives followed by chloroquine for treatment of malariaConclusions: Previous thinking that complications are only seen with P. falciparum has to be changed. Now many complications, mild as well as severe type are seen in P. vivax malaria. Drug resistance is another global problem which needs to be tackled wisely by systematic usage of antimalarials.

Liver dysfunction in malaria – An observational study

Introduction: Malaria is associated with significant mortality and morbidity in India. Hepatic dysfunction in malaria is often under-diagnosed. Early identification of hepatic dysfunction is crucial for preventing complications. Materials and methods: A prospective observational study was conducted in NRI Medical College and General Hospital, Chinakakani, Guntur District in Andhra Pradesh. A total of 50 patients with malaria were studied. Liver function tests were performed to assess the type of jaundice. The collected data was analysed. Results: The incidence was P. Vivax and P. falciparum malaria were 64% and 34% respectively. The incidence of jaundice was 26%. All of them had predominantly conjugated hyperbilirubinemia. Around 14% had mixed jaundice, and 22% had hepatic jaundice. Only one case expired which had the highest level of serum bilirubin. Conclusion: Hepatic involvement is more common among those with malaria. The incidence in vivax and falciparum malaria is comparable. Conjugated bilirubinemia with elevated liver enzymes is the most common manifestation. Early screening and identification of hepatic involvement in malaria are crucial. Initiation of treatment on time will aid in reducing mortality and morbidity

Lack of quadruple and quintuple mutant alleles associated with sulfadoxine-pyrimethamine resistance in Plasmodium vivax isolates from Brazilian endemic areas

BACKGROUND AND OBJECTIVE Brazil is responsible for a large number of Plasmodium vivax cases in America. Given the emergence of P. vivax parasites resistant to chloroquine and the effectiveness of antifolates in vivax malaria treatment together with a correlation between mutations in P. vivax dhfr and dhps genes and SP treatment failure, the point mutations in these genes were investigated. METHODS Blood samples from 54 patients experiencing vivax malaria symptomatic episodes in the Amazonian Region were investigated. Genomic DNA was extracted using a DNA extraction kit (QIAGENTM). Nested polymerase chain reaction (PCR) amplification was carried out followed by Sanger sequencing to detect single nucleotide polymorphisms (SNPs). FINDINGS All tested isolates showed non-synonymous mutations in pvdhfr gene: 117N (54/54, 100%) and 58R (25/54, 46%). Double mutant allele 58R/117N (FRTNI, 28%) was the most frequent followed by triple mutant alleles (58R/117N/173L, FRTNL, 11%; 58R/61M/117N, FRMNI, 5% 117N/173L, FSTNL, 4%) and quadruple mutant allele (58R/61M/117N/173L, FRMNL, 2%). A single mutation was observed at codon C383G in pvdhps gene (SGKAV, 48%). CONCLUSION No evidence of molecular signatures associated with P. vivax resistance to SP was observed in the Brazilian samples.

Changing clinical profile of malaria at a tertiary care hospital

Background: There is a widespread range of diverse typical and atypical manifestations of malaria. The diagnosis of malaria may escape the attention of treating physician due to its unusual and vague presentations.The morbidity and mortality due to malaria is increased due to lack of early diagnosis and right treatment. The Aim of the present study was to examine the changing clinical pattern of malaria with special attention to atypical presentations.Methods: The present study comprised of 630 cases of definitively diagnosed malaria.Diagnostic methods used were conventional thick and thin peripheral smear stained with Leishman stain and rapid malarial antigen test.Results: This study revealed atypical symptoms like lack of taste (1.3%), throat discomfort (13.33%) and cough (24.0%) and vomiting (52.4%) as presenting complaints. These were significantly more in patients with P. vivax infestations.Conclusions: A high degree of suspicion is necessary for early detection and treatment of malaria, especially of unusual presentations.

Inespecificidad persistente en amplificación isotérmica mediada por loop para diagnóstico de malaria / Persistent non-specificity in loop-mediated insothermal amplication for malaria diagnosis

Objetivo. Describir el ensayo realizado para la estandarización de la técnica de diagnóstico para malaria, denominada amplificación isotérmica mediada por loop (LAMP). Métodos. Se extrajeron 48 muestras divididas en 22 muestras para P. falciparum, 20 de P. vivax y 6 negativas para las mismas especies. Resultados. Se observaron falsos positivos, amplificaciones específicas que evidencian la inespecificidad de la técnica. Se propone estudiar los amplicones obtenidos por medio de restricción de fragmentos y secuenciación para elucidar el origen del problema.

Objetive. The present study describes the assay performed to standardize a malaria diagnostic DNA test, called loop-mediated isothermal amplification (LAMP). Methods. Extractions from 48 Plasmodium-infected blood samples (22 P.falciparum-positive samples, 20 P. vivax-positive samples and 6 negative control samples). Results. The data show false positives, unspecific amplifications that demonstrate the low specificity of the test. We propose to study the amplicons obtained by restriction fragment and sequencing to investigate the underlying cause of the problem.

Malarial Acute Kidney Injury: 25 Years Experience from a Center in an Endemic Region.

Background: While malaria rarely occurs in many parts of the world, it still causes serious complications like acute kidney injury (AKI) in endemic areas and needs to be reported. Methods: This study was carried out at Sindh Institute of Urology and Transplantation, Karachi, Pakistan. From January 1990 – December 2014, 5623 patients with acute kidney injury (AKI) were registered at this institution. AKI was defined as sudden rise in creatinine or decline in urine output or both. All patients had normal sized non obstructed kidneys on ultrasonography, with no previous co morbidity. Malaria parasite was seen on blood peripheral film in all patients. Results: Among total patients with AKI, 671 (11.93%) developed AKI in association with malarial infection. Average age of patients was 33.70±16.426 (range 4-98 years) with M: F ratio of 3:1. The causes were plasmodium falciparum in 59%, vivax in 15.2%, dual infection in 3.57% and undefined species in the rest. Oligo-anuria and vomiting were the most common associated symptoms along with fever. Renal replacement therapy was required in 76.6% of patients. Complete recovery was seen in 64.82%, while 21.2% died during the acute phase of illness. Jaundice, old age, altered level of consciousness, raised total leukocyte count, oliguria, hyperkalemia and falciparum malaria were the independent risk factors associated with high mortality. Conclusion: Malaria still causes significant morbidity and mortality in our part of the world. Vivax malaria which was thought to be ‘benign’ can present with hemolysis, thrombocytopenia and kidney failure, though risk of death is 2.36 fold higher with falciparum malaria.

Field performance of malaria rapid diagnostic test for the detection of Plasmodium falciparum infection in Odisha State, India.

Background & objectives: Rapid diagnostic tests (RDTs) have become an essential surveillance tool in the malaria control programme in India. The current study aimed to assess the performance of ParaHIT-f, a rapid test in diagnosis of Plasmodium falciparum infection through detecting its specific antigen, histidine rich protein 2 (PfHRP-2), in Odisha State, India. Methods: The study was undertaken in eight falciparum malaria endemic southern districts of Odisha State. Febrile patients included through active case detection, were diagnosed by Accredited Social Health Activists (ASHAs) for P. falciparum infection using the RDT, ParaHIT-f. The performance of ParaHIT-f was evaluated using microscopy as the gold standard. Results: A total of 1030 febrile patients were screened by both microscopy and the RDT for P. falciparum infection. The sensitivity of ParaHIT-f was 63.6% (95% CI: 56.0-70.6) and specificity was 98.9% (95% CI: 97.9-99.5), with positive and negative predictive values (PPV and NPV) of 92.6% (95% CI: 86.0-96.3) and 93.0% (95% CI: 91.0-94.5), respectively. When related to parasitaemia, the RDT sensitivity was 47.8% at the low parasitaemia of 4 to 40 parasites/μl of blood. Interpretation & conclusions: The results showed that the performance of the RDT, ParaHIT-f, was not as sensitive as microscopy in detecting true falciparum infections; a high specificity presented a low frequency of false-positive RDT results. The sensitivity of ParaHIT-f was around 60 per cent. It is, therefore, essential to improve the efficiency (sensitivity) of the kit so that the true falciparum infections will not be missed especially in areas where P. falciparum has been the predominant species causing cerebral malaria.

Congenital and neonatal malaria in Asian Indian population.

Background: Congenital malaria is defined as malarial parasites demonstrated in the peripheral blood smear of the newborn from twenty four hours to seven days of life. Malaria is endemic in India, neonatal disease is considered rare. Routine screening for malaria is essential for all neonates with fever in endemic areas. Early diagnosis and treatment of malaria could effectively prevent infant mortality. The aim of the present observational prospective study is to describe the occurrence and clinical spectrum of congenital vivax malaria in admitted neonates in Bikaner, India (low endemic region). Congenital malaria has been predominantly reported for P. falciparum from different parts of the world but the reports with P. vivax are very scanty. Methods: This prospective study was conducted on admitted neonate from January 2011 to December 2012. The species diagnosis was done by peripheral blood smear examination and rapid diagnostic test. The possibilities of other disease/infections causing similar illness were investigated thoroughly and stringently. A structured questionnaire was used to collect clinical data on newborn and maternal health during pregnancy. Results: A total of 1168 new born admitted in first week of life were screened. Out of them 23 (1.97%) had evidence of parasitaemia (P. vivax 17 and P. falciparum 6). The criteria for admission in these 17 neonates with congenital vivax malaria were LBW and prematurity (41.18%), septicemia (35.29%), perinatal asphyxia (17.65%), jaundice (17.65%) and seizures (5.88%). Conclusions: This study emphasizes the occurrence of P. vivax congenital malaria even in neonates in low transmission area and without typical manifestations. The emphasis is also on the relevance even in very low transmission areas of not only maintaining, but even increasing clinical and epidemiological awareness of this preventable and treatable disease in pregnancy and in the neonate.

Diferencias entre síntomas clínicos y paraclínicos al ingreso a urgencias de pacientes militares colombianos con malaria severa y no severa / Differences between clinical and paraclinical symptoms at admission to the emergency department of colombian military patients with severe and non-severe malaria

Antecedentes: en los últimos años ha sido de interés el reconocimiento de manifestaciones clínicas con que cursan los pacientes que desarrollarán malaria severa (MS), y la población militar es susceptible de adquirir la infección con el desarrollo de complicaciones asociado al retraso en el reconocimiento de sus manifestaciones clínicas. Material y métodos: se realizó un análisis de una cohorte retrospectiva de sujetos con malaria atendidos en un hospital de referencia militar y el desarrollo de complicaciones según criterios de OMS durante observación hospitalaria en mayores de 18 años; se estudiaron variables demográficas, clínicas, al examen físico, resultados paraclínicos y de manejo, los datos fueron analizados en el programa estadístico SPSS. Resultados: se analizaron 533 registros, se encontraron diferencias al ingreso en disnea 42.2% MS vs 16.88% malaria no complicada (MNC) (p<0.001), vómito 65.6% MS vs 7% MNC (p=0.002), antecedente de malaria 30% MS vs 44% MNC (p=0,006), frecuencia respiratoria 20.62 (de: 5.2) resp/min MS vs 19.62 (5.2) lat/min MNC, glasgow <15.8% MS vs 0.4% MNC, Hb 11.06 (3,25) g/dL MS vs 13.62 (2.16) MNC (p<0,001), Hto 33.48 (8.15)% MS vs 39.70 (6.68)% MNC(p<0.001), leucocitos 7390 (5601) cel/mL MS vs 6319 (4862) cel/mL MNC (p=0.027), Bilirrubina total 6 (7.55) mg/dL MS vs 2.6 (2.55) mg/dL MNC, creatinina 1.98 (3.61) mg/dL MSvs 1.03 (0.22) mg/dL (p=0.029), glicemia 95.09 (21.96) mg/dL vs 103 (22.06) mg/dL (p=0,001), P. falciparum 45.6% MS vs 28.3% MNC (p=0.047). Conclusiones: el comportamiento de la malaria en población militar es similar a la población general, sin embargo, los antecedentes de malaria, características clínicas y de laboratorio pueden ser útiles para predicción de complicaciones, se requieren más estudios para corroborar estos hallazgos. (Acta Med Colomb 2015; 40: 202-208).

Background: in recent years the recognition of clinical manifestations that occur with patients who develop severe malaria has raised interest, and the military population is susceptible of acquiring the infection with the development of complications associated with delayed recognition of its clinical manifestations. Materials and Methods: A retrospective analysis of a cohort of patients with malaria treated at a reference military hospital and development of complications according to WHO criteria during hospital observation in patients over 18 years was conducted; demographic, clinical, physical examination, paraclinical and clinical management results variables were studied. Data were analyzed in SPSS statistical progam. Results: 533 records were analyzed; differences found at hospital admission were dyspnea42.2% in severe malaria vs.16.88% in uncomplicated malaria (p <0.001), vomiting 65.6% insevere malaria vs. 7% in uncomplicated malaria (p = 0.002), history of malaria 30% in severe malaria vs. 44% in uncomplicated malaria (p = 0.006), respiratory rate 20.62 breaths/min (5.2) in severe malaria vs. 19.62 (5.2) breaths/min in uncomplicated malaria. Glasgow <15.8% in severe malaria vs. 0.4% in uncomplicated malaria, Hgb 11.06 g/dL (3.25) in severe malaria vs. 13.62g/dL (2.16) in uncomplicated malaria (p<0,001), hematocrit 33.48 (8.15) in severe malaria vs.39.70% (6.68)% in uncomplicated malaria (p <0.001), leukocytes 7390 (5601) cells/mL in severe malaria vs. 6319 (4862) cells/mL in uncomplicated malaria (p = 0.027), total bilirubin 6 (7.55)mg/dL in severe malaria vs. 2.6 (2.55) mg/dL in uncomplicated malaria, creatinine 1.98 (3.61)mg/dL in severe malaria vs. 1.03 (0.22) mg/dL in uncomplicated malaria (p = 0.029), glucose 95.09 (21.96) mg/dL vs. 103 (22.06) mg/dL (p = 0.001), P. falciparum 45.6% in severe malaria vs. 28.3% in uncomplicated malaria (p = 0.047). Conclusions: The behavior of malaria in military population is similar to that in the general population; however, the history of malaria, clinical and laboratory features may be useful for prediction of complications; further studies are needed to confirm these findings. (Acta Med Colomb2015; 40: 202-208).

Malaria situation in India with special reference to tribal areas.

Background & objectives: In India, malaria is a major public health problem in states having predominantly tribal population. The objective of this analysis was to find out the incidence of malaria in various states/districts having varied proportions of tribal population using National Vector Borne Disease Control Programme (NVBDCP) data. Methods: States and districts were classified into three categories based on proportions of Scheduled Tribes (ST) population as <10, 10-29.9 and 30 per cent + ST population. Five year average (2008-2012) of all important malaria indicators collected by NVBDCP was taken to normalize the effect of annual fluctuations in malaria incidence. Results: State level analysis revealed that ten states/UTs with 30 per cent or more tribal population comprising only three per cent of total population, contributed 14 per cent of total malaria, 21 per cent Plasmodium falciparum and 29 per cent of deaths due to malaria. Similarly, district level analysis showed that districts with 30 per cent or more tribal population comprising about eight per cent country’s population contributed to 46 per cent of total malaria cases, 70 per cent P. falciparum and 47 per cent malarial deaths in the country. Interpretation & conclusions: our analysis showed that the neglect of the ethnic communities in tribal areas would be detrimental to the overall reduction of morbidity and mortality due to malaria. The fight against the increasing burden of malaria in tribal belt requires adoption of multiple approaches and socio-economic development of the tribal communities.

Associação entre bimarcadores imunogenéticos e a gravidade ou resistência na malária por plasmodium vivax / Association between immunogenic markers and the severity and resistance in plasmodium vivax malaria

INTRODUÇÃO: A malária é uma das doenças infecto-parasitárias mais incidentes no mundo com grande morbimortalidade. Dentre as espécies infectivas ao ser humano, o Plasmodium vivax é a espécie predominante no Brasil, quase que exclusivamente na Região Amazônica. O espectro clínico da malária abrange desde uma infecção assintomática até casos moderados,com hiperbilirrubinemia isolada ou graves. A produção de mediadores inflamatórios pelo sistema imune, a via de metabolização do heme e os níveis sistêmicos de hepcidina são importantes mecanismos associados afisiopatologia dos diferentes desfechos clínicos da malária. Além disso, coinfecções podem modular ou intensificar a resposta imune de indivíduos infectados pelo plasmódio. OBJETIVO: Neste ínterim, a identificação de biomarcadores confiáveis tanto de gravidade ou resistência são indispensáveis para o auxílio no seguimento, diagnóstico e terapêutica da malária...

INTRODUCTION: Malaria is one of the most frequent infectious diseases inthe world with high morbidity and mortality. Among the infective species tohumans, Plasmodium vivax is the most predominant species in Brazil, withdisease incidence almost exclusively observed in the Amazon Region. Theclinical spectrum of malaria can range from asymptomatic infection to mildcases, malaria with isolated hyperbilirubinaemia or severe infection. Theimmune system production of inflammatory mediators, the heme metabolismpathway and systemic levels of hepcidin are important mechanismsassociated with pathophysiology of different malaria clinical outcomes. Inaddition, co-infections can modulate or enhance the immune response ofindividuals infected with P. vivax. OBJECTIVE: In this context, the identification of reliable biomarkers for disease severity and resistance are essential for the diagnosis, treatment and follow-up of malaria...

Study of sodium and potassium ion disturbances in malaria at Ahmedabad, Gujarat, India.

Background: Malaria is an endemic disease in Gujrat, caused by the bite of the female anopheles mosquito. Electrolyte disturbances particularly hyponatremia is a common complication in malaria. Aim: To determine sodium and potassium ion disturbances in malaria cases at Ahmedabad, Gujarat Material and methods: Total 200 indoor and outdoor patients of malaria who came to tertiary care hospital, Ahmedabad during period of July – 2013 to June – 2014 were included in present study. Patients were diagnosed for malaria after the examination of both thick and thin film peripheral smears and malarial antigen detection rapid card test. All patients were subjected to relevant laboratory investigations like complete blood count and serum electrolyte (sodium and potassium ion). Observation: Hyponatremia and hypokalemia were more common in both P. Falciparum and P. Vivax malaria. Severity of hyponatremia and hypokalemia were more in P. Falciparum than P. Vivax Conclusion: Serum electrolytes must be estimated in each malaria case to prevent grave complications.