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Background: Malaria, a prominent vector borne disease causing over a million annual cases worldwide, predominantly affects vulnerable populations in the least developed regions. Despite their preventable and treatable nature, malaria remains a global public health concern. In the last decade, India has faced a significant decline in malaria morbidity and mortality. As India pledged to eliminate malaria by 2030, this study examined a decade of surveillance data to uncover space–time clustering and seasonal trends of Plasmodium vivax and Plasmodium falciparum malaria cases in West Bengal. Methods: Seasonal and trend decomposition using Loess (STL) was applied to detect seasonal trend and anomaly of the time series. Univariate and multivariate space–time cluster analysis of both malaria cases were performed at block level using Kulldorff's space–time scan statistics from April 2011 to March 2021 to detect statistically significant space–time clusters. Results: From the time series decomposition, a clear seasonal pattern is visible for both malaria cases. Statistical analysis indicated considerable high-risk P. vivax clusters, particularly in the northern, central, and lower Gangetic areas. Whereas, P. falciparum was concentrated in the western region with a significant recent transmission towards the lower Gangetic plain. From the multivariate space–time scan statistics, the co-occurrence of both cases were detected with four significant clusters, which signifies the regions experiencing a greater burden of malaria cases. Conclusions: Seasonal trends from the time series decomposition analysis show a gradual decline for both P. vivax and P. falciparum cases in West Bengal. The space–time scan statistics identified high-risk blocks for P. vivax and P. falciparum malaria and its co-occurrence. Both malaria types exhibit significant spatiotemporal variations over the study area. Identifying emerging high-risk areas of P. falciparum malaria over the Gangetic belt indicates the need for more research for its spatial shifting. Addressing the drivers of malaria transmission in these diverse clusters demands regional cooperation and strategic strategies, crucial steps towards overcoming the final obstacles in malaria eradication. [ABSTRACT FROM AUTHOR]

This study analysed the genetic diversity of DBL1α domain of Plasmodium falciparum var gene in severe and non-severe malaria patients from Delhi and Mewat in Northern India. After confirming P. falciparum infection, samples were cloned and the var gene DBL1α domain was sequenced. Out of 377 cloned DBL sequences, 194 were from severe samples and 183 from non-severe samples. Proportion of DBL1α sequences belonging to groups 1, 4 and 5 were significantly higher in severe isolates as compared to non-severe isolates—group 1 (4.1% vs 1.09%, P = 0.0333), group 4 (69.58% vs 74.31%, P < 0.0001), and group 5 (19.58% vs 10.38%, P < 0.0001). Conversely, higher proportion of group 2 was observed in non-severe isolates (0% vs 3.82%, P = 0.0350). Highest diversity was seen in PoLV4 motif of severe and non-severe isolates and like other DBL1α sequences reported from several geographical areas (Africa, Americas, Asia, and Oceania). A total of 247 DBL1α domain haplotypes were found in this study where 139 (56.27%) haplotypes are novel and not reported from India till date. These findings could aid in developing effective malaria interventions, including vaccine and drug targets, by understanding the existing antigenic diversity and vulnerabilities in the parasite's genetic makeup. [ABSTRACT FROM AUTHOR]

Background Antimalarial drug resistance surveillance and containment are crucial for countries aiming to eliminate malaria. Monitoring resistance evolution through studies before and after treatment policy changes is essential. Methods A total of 939 Plasmodium falciparum-positive blood samples were collected between 2014 and 2015 across 10 sites in India, categorized into 4 geographic clusters. Polymerase chain reaction-amplified products were sequenced to identify point mutations at drug resistance-conferring genes (Pfdhfr , Pfdhps , Pfmdr1, and Pfk13). Results Triple Pfdhfr mutants were found only in northeast India bordering Myanmar, while the wild type (WT) was dominant in central India. Pfdhps WTs were prevalent in all areas, and no double mutants were found. Except in northwest India, Pfmdr1 WT was dominant in all clusters. Nonsynonymous double mutations were only found in northwest India. Only synonymous mutations occurred in Pfk13. These were found in central India at a low frequency. The pattern of linkage disequilibrium and principal component analysis reflects low pressure for drug resistance and heterogeneity between the geographic clusters. Conclusions Resistance levels were highest in northeast India, close to the Myanmar border, where resistance is common. Primaquine has been widely used as a gametocidal and schizonticidal drug, likely contributing to maintaining low drug resistance levels and preventing strong selection for resistance. [ABSTRACT FROM AUTHOR]

Context: Resistance to antimalarial drugs is one of the major challenges for malaria elimination. In India, artemisinin combination therapy (artesunate-sulfadoxin pyrimethamine) was introduced in place of chloroquine (CQ) for the treatment of uncomplicated falciparum malaria in 2010. Periodical monitoring of polymorphisms in antimalarial drug resistance marker genes will be useful for assessing drug pressure, mapping and monitoring of drug resistance status; and will be helpful for searching alternative treatments. Objectives: This study was conducted to study the polymorphisms in antimalarial drug resistance marker genes among clinical Plasmodium falciparum isolates collected from Kolkata after 10 years of artemisinin-based combination therapie (ACT) implementation. Materials and Methods: Blood samples were collected from P. falciparum mono-infected patients and polymorphisms in P. falciparum CQ resistance transporter (pfcrt), P. falciparum multidrug resistance (pfmdr-1), P. falciparum dihydrofolate reductase (pfdhfr), P. falciparum dihydropteroate synthetase (pfdhps), pfATPase6 and pfK-13 propeller genes were analysed by polymerase chain reaction and DNA sequencing. Results: In pfcrt gene, C72S, and K76T mutation was recorded in 100% isolates and no mutations was detected in any of the targeted codons of pfmdr-1 gene. A double mutant pfcrt haplotype SVMNT and wildtype haplotype NYD in pfmdr-1 were prevalent in 100% of study isolates. Triple mutant pfdhfr-pfdhps haplotype ANRNI-SGKAA was recorded. No polymorphism in pfK13 gene was documented in any of the isolates. Conclusions: Observed wild codon N86 along with Y184 and D1246 of pfmdr-1 gene might be an indication of the reappearance of CQ sensitivity. The absence of quadruple and quintuple haplotypes in pfdhfr-pfdhps gene along with the wild haplotype of pfK13 is evidence of ACT effectivity. Hence, similar studies with large sample size are highly suggested for monitoring the drug resistance status of P. falciparum. [ABSTRACT FROM AUTHOR]

Background: Resistance against artemisinin-based combination therapy is one of the challenges to malaria control and elimination globally. Mutations in different genes (Pfdhfr, Pfdhps, Pfk-13 and Pfmdr1) confer resistance to artesunate and sulfadoxine–pyrimethamine (AS + SP) were analysed from Mandla district, Madhya Pradesh, to assess the effectiveness of the current treatment regimen against uncomplicated Plasmodium falciparum. Methods: Dried blood spots were collected during the active fever survey and mass screening and treatment activities as part of the Malaria Elimination Demonstration Project (MEDP) from 2019 to 2020. Isolated DNA samples were used to amplify the Pfdhfr, Pfdhps, Pfk13 and Pfmdr1 genes using nested PCR and sequenced for mutation analysis using the Sanger sequencing method. Results: A total of 393 samples were subjected to PCR amplification, sequencing and sequence analysis; 199, 215, 235, and 141 samples were successfully sequenced for Pfdhfr, Pfdhps, Pfk13, Pfmdr1, respectively. Analysis revealed that the 53.3% double mutation (C59R, S108N) in Pfdhfr, 89.3% single mutation (G437A) in Pfdhps, 13.5% single mutants (N86Y), and 51.1% synonymous mutations in Pfmdr1 in the study area. Five different non-synonymous and two synonymous point mutations found in Pfk13, which were not associated to artemisinin resistance. Conclusion: The study has found that mutations linked to SP resistance are increasing in frequency, which may reduce the effectiveness of this drug as a future partner in artemisinin-based combinations. No evidence of mutations linked to artemisinin resistance in Pfk13 was found, suggesting that parasites are sensitive to artemisinin derivatives in the study area. These findings are a baseline for routine molecular surveillance to proactively identify the emergence and spread of artemisinin-resistant parasites. [ABSTRACT FROM AUTHOR]

BACKGROUND: Malaria has been one of India's most considerable health problems since 1940. The objective of our study is to determine the status of the National Malaria Elimination Programme in India by using epidemiological indicators. METHODS AND MATERIALS: The annual reports of malaria for the years 2014-2021 and monthly reports for 2020 and 2021 were collected from the official web portal and were analysed for study specific assessments. RESULTS: The API has shown a statistically significant reduction from 2017-2021 in all states along with category-1(P=0.003) and category-2(P=0.029) states/UTs, but there was no statistically significant reduction from 2017-2021 in category-3 (P=0.166) states/UTs. The zero indigenous cases had not been achieved in category-1 states/UTs. The overall percentage reduction in number of malaria cases in 2020 at the national level compared with 2014 was 83.6%. Despite states with strong health systems such as Gujarat, Maharashtra and Karnataka, have not shown zero indigenous cases in 2020 and the malaria cases noted were very far from reaching the targets. CONCLUSIONS: Although we observed a significant drop in malaria incidence from 2014 to 2020, demonstrating that the country is moving nearer to malaria elimination, it is crucial to implement the strategies to reduce Plasmodium falciparum% and re-establish surveillance programmes and execute national and state programmes in order to achieve the success of the National Malaria Elimination Programme. The recategorization of states/UTs are in accordance to the API, and implementation strategies were also needed. [ABSTRACT FROM AUTHOR]

Background: Drug resistance is a serious impediment to efficient control and elimination of malaria in endemic areas. Methods: This study aimed at analysing the genetic profile of molecular drug resistance in Plasmodium falciparum and Plasmodium vivax parasites from India over a ~ 30-year period (1993–2019). Blood samples of P. falciparum and/or P. vivax-infected patients were collected from 14 regions across India. Plasmodial genome was extracted and used for PCR amplification and sequencing of drug resistance genes in P. falciparum (crt, dhps, dhfr, mdr1, k13) and P. vivax (crt-o, dhps, dhfr, mdr1, k12) field isolates. Results: The double mutant pfcrtSVMNT was highly predominant across the country over three decades, with restricted presence of triple mutant CVIET from Maharashtra in 2012. High rates of pfdhfr-pfdhps quadruple mutants were observed with marginal presence of "fully resistant" quintuple mutant ACIRNI-ISGEAA. Also, resistant pfdhfr and pfdhps haplotype has significantly increased in Delhi between 1994 and 2010. For pfmdr1, only 86Y and 184F mutations were present while no pfk13 mutations associated with artemisinin resistance were observed. Regarding P. vivax isolates, the pvcrt-o K10 "AAG" insertion was absent in all samples collected from Delhi in 2017. Pvdhps double mutant SGNAV was found only in Goa samples of year 2008 for the first time. The pvmdr1 908L, 958M and 1076L mutations were highly prevalent in Delhi and Haryana between 2015 and 2019 at complete fixation. One nonsynonymous novel pvk12 polymorphism was identified (K264R) in Goa. Conclusions: These findings support continuous surveillance and characterization of P. falciparum and P. vivax populations as proxy for effectiveness of anti-malarial drugs in India, especially for independent emergence of artemisinin drug resistance as recently seen in Africa. [ABSTRACT FROM AUTHOR]

Blood phagocytes, such as neutrophils and monocytes, generate reactive oxygen species (ROS) as a part of host defense response against infections. We investigated the mechanism of Fcγ-Receptor (FcγR) mediated ROS production in these cells to understand how they contribute to anti-malarial immunity. Plasmodium falciparum merozoites opsonized with naturally occurring IgG triggered both intracellular and extracellular ROS generation in blood phagocytes, with neutrophils being the main contributors. Using specific inhibitors, we show that both FcγRIIIB and FcγRIIA acted synergistically to induce ROS production in neutrophils, and that NADPH oxidase 2 and the PI3K intracellular signal transduction pathway were involved in this process. High levels of neutrophil ROS were also associated with protection against febrile malaria in two geographically diverse malaria endemic regions from Ghana and India, stressing the importance of the cooperation between anti-malarial IgG and neutrophils in triggering ROS-mediated parasite killing as a mechanism for naturally acquired immunity against malaria. FcγR-mediated signaling by cytophilic IgG against Plasmodium falciparum merozoites triggers ROS generation predominantly in neutrophils as a mechanism contributing to naturally acquired immunity against malaria in endemic regions in Ghana and India. [ABSTRACT FROM AUTHOR]

Intensified Malaria Control Project (IMCP) was implemented in 2005 to control malaria in all North-Eastern and Odisha states of India. The present study aimed to investigate the impact of IMCP in reducing the malaria burden in Udalguri district, Assam state of North-East India. Malaria epidemiological data were obtained for IMCP intervention (Udalguri) and nonintervention district (West Singhbhumi, Jharkhand state). IMCP activities include introducing bi-valent rapid diagnostic kits (RDTs), Artemether-Lumefantrine drug in North-East India, long-lasting insecticidal nets (LLINs) distribution, and creating awareness programs about malaria in an intensified mode. The data revealed a significant decline in annual parasite incidence (API) from 14.94 (2005) to 2.61 (2018), −37% (95%CI: −57%, −19%, p = 001) after using LLINs in 2009 and −64% (95%CI: −116%, −14%, p = 013) after the introduction of RDTs in district Udalguri. Whereas control district showed a −28% (95%CI: −63%, 6.3%, p = 0.051) decrease in API using LLINs and a 10% (95%CI: −7.6%, 28%, p = 0.122) increase after the introduction of RDTs. Plasmodium falciparum (Pf) and P. vivax (Pv) were the major malarial parasites in Udalguri. Pv-malaria was much higher (71%) than Pf-malaria (29%) during the study period. An increasing trend of Pf cases was observed in Udalguri. Udalguri and Khoirabari BPHCs showed an overall reduction of 94% (95%CI: −143%, −45%, p = 0.001) and 84% (95%CI: −126%, −39%, p = 0.003), respectively; however, only a 10% (95%CI: −65%, −41%, p = 0.360) reduction in API was observed in Orang BPHC. An overall decrease in malaria indicates the effective implementation of vector and disease control strategies in the Udalguri district. [ABSTRACT FROM AUTHOR]

The global malaria control and elimination program faces major threats due to the emergence and transmission of the anti-malarial drug-resistant strain of Plasmodium falciparum. Monitoring of artemisinin (ART) resistance marker Kelch-13 in the malaria-endemic region is essential in mitigating the disease's morbidity and mortality. The current study aimed to generate baseline information for further surveillance in the future. The current research was designed and conducted from July 2019 to June 2021 to monitor Pfkelch13 mutation at the molecular level in the eastern region of India. We also conducted an in silico study to understand the drug-protein interactions between ART and the protein crystal of PfKelch13 (KELCH) with PDB id:4ZGC. The kelch-13 gene was amplified by nested polymerase chain reaction (PCR) and sequenced through the Sanger sequencing method. Reference 3D7 clone (PF3D7_1343700) was used to align and probe all the sequences. The sequence analysis showed the absence of validated or associated mutation in the Kelch-13 propeller domain. The absence of natural selection in drug resistance was confirmed by the Tajima test. Further, in silico interaction studies between the drug ART and the Kelch propeller domain of P. falciparum were evaluated by structure predictions, molecular docking, molecular dynamics (MD) simulations, and estimations of binding free energies for the KELCH-ART complex. The results were compared with the apoprotein (KELCH-APO). The study confirmed the favorable binding of ART with the Kelch-13 propeller domain. [ABSTRACT FROM AUTHOR]

Malaria is a major health problem in India, endemic for both Plasmodium falciparum (Pf) and Plasmodium vivax (Pv). Approximately 95% of the population reside in malaria endemic areas. Therefore, in this study long-term spatial patterns of district level annual malaria incidence data from 1995 to 2013 for both parasites have been analyzed. Broadly, three regions of higher average annual malaria incidence were identified from the analysis. These include south central, northwest, and northeast India for both parasites. Specifically, Pf cases were more widespread in the east and northeast, while Pv cases were more widespread in the northwest and districts adjacent to those for Pf cases in south central and northeast India. A majority of the districts experienced negative trends in the long-term incidence of malaria, with most of the positive trends spread across districts in northern India. In addition, the average number of deaths were mostly below 15 per year, except in Odisha and Chhattisgarh in South Central India. These lower rates from malaria may be a result of underestimation. Furthermore, the annual blood slide examination rates were above 10% for most of the country, except the northern Gangetic plains. [ABSTRACT FROM AUTHOR]

There is a paucity of literature on the association of α+-thalassemia, sickle-cell hemoglobin disorders, and malaria in India. This study aimed to understand the effect of α+-thalassemia on the severity of Plasmodium falciparum malaria in adults with respect to sickle-cell genotypes. The study subjects were categorized into 'severe-malaria' and 'uncomplicated-malaria' and age-gender matched 'control' groups. Sickle-cell and α+-thalassemia were investigated in all the recruited subjects. The effect of α+-thalassemia on the severity of malaria was analyzed in HbAA and sickle-cell genotypes (HbAS and HbSS) separately. The prevalence of α+-thalassemia in various groups ranged from 41.5% to 81.8%. The prevalence of α+-thalassemia was lower (OR = 1.64; p = 0.0013) in severe malaria (41.5%) as compared to healthy controls (53.8%) with HbAA genotype. In contrast, in HbAS genotype, the prevalence of α+-thalassemia was higher (OR = 4.11; p = 0.0002) in severe malaria (81.8%) compared to controls (52.2%). In severe malaria with HbAA genotype, there was a significantly higher hemoglobin level and low MCV and MCH level in patients with α+-thalassemia compared to the normal α-globin genotype. Further, the incidence of cerebral malaria, hepatopathy, and mortality was lower in patients (HbAA) with α+-thalassemia as compared to normal α-globin genotype (HbAA). In severe malaria with either HbAS or HbSS genotype, only a few parameters showed statistical differences with respect to α+-thalassemia. Low prevalence of α+-thalassemia in severe malaria with HbAA genotype compared to healthy controls with HbAA genotype indicates the protective effect of α+-thalassemia against severe malaria. However, the high prevalence of α+-thalassemia in patients with HbAS genotype depicts its interference in the protective effect of sickle-cell against severe malaria. [ABSTRACT FROM AUTHOR]

Background & objectives: India targets malaria elimination by 2030 in a phased manner, so malaria’s assured diagnosis is crucial. Introduction of rapid diagnostic kits in India in 2010 has revolutionized malaria surveillance. The storage temperature of rapid diagnostic tests (RDTs), kit components and handling in transportations impact the results of RDTs. Therefore, quality assurance (QA) is required before it reaches end-users. The Indian Council of Medical Research-National Institute of Malaria Research (ICMR-NIMR) has a World Health Organization (WHO) recognized lot-testing laboratory facility to assure the quality of RDTs. Methods: The ICMR-NIMR receives RDTs from different manufacturing companies as well as various agencies such as National and State Programmes and Central Medical Services Society. The WHO standard protocol is followed to conduct all the tests, including long-term and post-dispatch testing. Results: A total of 323 lots tested during January 2014-March 2021 were received from different agencies. Amongst them, 299 lots passed the quality of test and 24 failed. In long-term testing, 179 lots were tested and only nine failed. A total of 7741 RDTs were received from end-users for post-dispatch testing of which 7540 qualified the QA test with a score of 97.4 per cent. Interpretation & conclusions: RDTs received for quality testing showed compliance with QA evaluation of malaria RDTs based on the protocol recommended by the WHO. However, continuous monitoring of the quality of RDTs is required under QA programme. Quality-assured RDTs have a major role, especially in areas where low parasitaemia of parasites persists. [ABSTRACT FROM AUTHOR]

Black water fever, also called malarial haemoglobinuria occurs exclusively with Plasmodium falciparum infection and is characterized by intravascular haemolysis with severe anaemia and intermittent passage of black coloured urine. It frequently leads to acute kidney injury (AKI) with increased morbidity and mortality. Although Sri Lanka has achieved remarkable success in eliminating malaria since 2016, still Sri Lanka gets malarial cases via overseas visitors from African countries and India. Here we report a case of black water fever complicated by AKI with a brief account of its pathophysiology. [ABSTRACT FROM AUTHOR]

Context: Histidine-rich protein 2 (HRP2) detecting rapid diagnostic tests (RDTs) have played an important role in enabling prompt malaria diagnosis in remote locations. HRP2 has advantages over other biomarkers because of its abundance in the bloodstream, repetitive binding epitopes, and falciparum-specificity. Most HRP2-based RDTs also exhibit some cross-reactivity to a closely related protein (HRP3). Plasmodium falciparum parasites lacking HRP2 (pfhrp2) and 3 (pfhrp3) genes escape detection by these RDTs. Objectives: The objective of the study was to study the sensitivity and specificity of hrp2-based RDT for diagnosis of falciparum, to compare the RDT results with microscopy and polymerase chain reaction (PCR), and to determine the prevalence of HRP2 gene deletion among the RDT-negative, microscopy-positive falciparum strains. Materials and Methods: Blood samples were collected and diagnosis was done by microscopic examination, RDTs, and PCR. Results: Out of 1000 patients examined, 138 were positive for P. falciparum. Fever was the most common symptom followed by chills with rigor and headache were recorded among more than >95% of the study patients. Three microscopy-confirmed P. falciparum cases were negative by HRP2-based RDT and were found to have deletion of HRP2 and HRP3 exon 2. Conclusions: Rapid and accurate diagnosis and prompt deployment of effective antimalarial medication are essential components of appropriate case management. P. falciparum strains that evade diagnosis by RDTs represent a major threat to malaria control and elimination efforts. [ABSTRACT FROM AUTHOR]

Background: Despite significant progress in eliminating malaria from the state of Odisha, India, the disease is still considered endemic. Artesunate plus sulfadoxine-pyrimethamine (AS + SP) has been introduced since 2010 as first-line treatment for uncomplicated Plasmodium falciparum malaria. This study aimed to investigate the prevalence of mutations associated with resistance to chloroquine (CQ), sulfadoxine-pyrimethamine (SP), and artesunate (ART) in P. falciparum parasites circulating in the state. Methods: A total of 239 isolates of P. falciparum mono infection were collected during July 2018-November 2020 from the four different geographical regions of the state. Genomic DNA was extracted from 200 µL of venous blood and amplified using nested polymerase chain reaction. Mutations on gene associated with CQ (Pfcrt and Pfmdr1) were assessed by PCR amplification and restriction fragment length polymorphism, artemisinin (Pfk13) gene by DNA sequencing and SP (Pfdhfr and Pfdhps) genes by allele-specific polymerase chain reaction (AsPCR). Results: The point mutation in Pfcrt (K76T) was detected 2.1%, in Pfmdr1 (N86Y) 3.4%, and no mutations were found in Pfkelch13 propeller domain. Prevalence of Pfdhfr, Pfdhps and Pfhdfr-Pfdhps (two locus) gene mutations were 50.43%, 47.05% and 49.79% respectively. The single, double, triple and quadruple point mutations in Pfdhfr gene was 11.2%, 8.2%, 17.2% and 3.4% while, in Pfdhps gene was 10.9%,19.5%, 9.5% and 2.7% respectively. Of the total 13 haplotypes found in Pfdhfr, 8 were detected for the first time in the state and of the total 26 haplotypes found in Pfdhps, 7 were detected for the fisrt time in the state. The linked quintuple mutation Pfdhfr (N51I-C59R-S108N)-Pfdhps (A437G-K540E) responsible for clinical failure (RIII level of resistance) of SP resistance and A16V-S108T mutation in Pfdhfr responsible for cycloguanil was absent. Conclusion: The study has demonstrated a low prevalence of CQ resistance alleles in the study area. Despite the absence of the Pfkelch13 mutations, high prevalence of Pfdhfr and Pfdhps point mutations undermine the efficacy of SP partner drug, thereby threatening the P. falciparum malaria treatment policy. Therefore, continuous molecular and in vivo monitoring of ACT efficacy is warranted in Odisha. [ABSTRACT FROM AUTHOR]

Introduction: After being used vigorously for the previous two decades to treat P. falciparum, chloroquine and sulfadoxine-pyrimethamine were replaced in 2009 with an artemisinin-based combination therapy (artesunate-sulfadoxine-pyrimethamine) in an effort to combat multidrug-resistant parasites. Methods: We set out to assess the genetic variants of sulfadoxine-pyrimethamine resistance and the effectiveness of its treatment in eastern India prior to, during, and 6 to 8 years following the introduction of the new pharmacological regime. In 2008-2009, 318 P. falciparum-positive patients got the recommended doses of sulfadoxine-pyrimethamine. We used 379 additional isolates from 2015 to 2017 in addition to the 106 isolates from 2010. All 803 isolates from two study sites underwent in vitro sulfadoxine-pyrimethamine sensitivity testing and genomic characterisation of sulfadoxine-pyrimethamine resistance (pfdhfr and pfdhps). Results: In Kolkata and Purulia, we observed early treatment failure in 30.7 and 14.4% of patients, respectively, whereas recrudescence was found in 8.1 and 13.4% of patients, respectively, in 2008-2009. In 2017, the proportion of in vitro pyrimethamine and sulfadoxine resistance steadily grew in Kolkata and Purulia despite a single use of sulfadoxine-pyrimethamine. Treatment failures with sulfadoxine-pyrimethamine were linked to quintuple or quadruple pfdhfrpfdhps mutations (AICII-AGKAT, AICII-AGKAA, AICII-SGKGT, AICII-AGKAA, AICNI-AGKAA) in 2008-2009 (p < 0.001). The subsequent spread of mutant-haplotypes with higher in vitro sulfadoxine-pyrimethamine resistance (p < 0.001), such as the sextuple (dhfr-AIRNI+dhps-AGEAA, dhfr-ANRNL+dhps-AGEAA) and septuple (dhfr-AIRNI+dhps-AGEAT), mutations were observed in 2015-2017. Discussion: This successive spread of mutations with high in vitro sulfadoxine-pyrimethamine resistance confirmed the progressive increase in antifolate resistance even after an 8-year withdrawal of sulfadoxine-pyrimethamine. [ABSTRACT FROM AUTHOR]

Background Haem detoxification protein (HDP) is a significant protein in the erythrocytic stage of the Plasmodium lifecycle. HDP could be of paramount interest as a diagnostic biomarker for accurate diagnosis of malaria. We thus explored HDP genetic variation, expression levels of HDP and immune response. Methods Phylogenetic analysis was carried out using Pfhdp orthologues sequences of various Plasmodium species. Blood samples were collected from patients in central India. Pfhdp gene was amplified, and sequenced by sanger DNA sequencing. B-cell epitopes were identified in PfHDP using Bepipred Linear Epitope Prediction 2.0, and median-joining network was constructed using global Pf HDP sequences. Pfhdp expression levels during erythrocytic stage were assessed using real-time qPCR at 4-h intervals. An IgG immune response against synthetic Pf HDP peptides was analysed using ELISA. Results Phylogenetic analysis revealed the conserved nature of Pfhdp gene. Diversity analysis revealed one non-synonymous mutation (F91L) among all isolates. Neutrality tests indicated negative selection for Pfhdp gene. HDP was expressed throughout the erythrocytic cycle, and comparatively, high expression was observed in the late trophozoite and schizont stages. High IgG response against both peptides was observed, and no polymorphism was seen in any of the seven predicted B-cell epitopes. Conclusions Findings of the present study indicate the possibility of HDP being exploited as a diagnostic biomarker for Plasmodium falciparum malaria after proteomic validation studies. [ABSTRACT FROM AUTHOR]

A plethora of studies analyse the molecular markers of drug resistance and hence help in guiding the evidence-based malaria treatment policies in India. For reporting mutations, a number of techniques including DNA sequencing, restriction-fragment length polymorphism and mutation-specific polymerase chain reaction have been employed across numerous studies, including variations in the methodology used. However, there is no sufficient data from India comparing these methods as well as report the prevalence of polymorphisms in SP drug resistance molecular markers independently using such methods. Therefore, all data from Indian studies available for molecular marker studies of Plasmodium falciparum drug resistance to sulphadoxine-pyrimethamine was gathered, and a systematic review was performed. This systematic review identifies the molecular methods in use in India and compares each method for detecting sulphadoxine-pyrimethamine drug resistance marker. To delay the spread of drug-resistant parasite strains, a simplified and standardized molecular method is much needed which can be obtained by analysing the performance of each method in use and answering the necessity of newer methodological approaches. [ABSTRACT FROM AUTHOR]

Background In India, there are several malaria-endemic regions where non- falciparum species coexist with Plasmodium falciparum. Traditionally, microscopy and rapid diagnostic tests are used for the diagnosis of malaria. Nevertheless, microscopy often misses the secondary malaria parasite in mixed-infection cases due to various constraints. Misdiagnosis/misinterpretation of Plasmodium species leads to improper treatment, as the treatment for P. falciparum and Plasmodium vivax species is different, as per the national vector-borne disease control program in India. Methods Blood samples were collected from malaria-endemic regions (Jharkhand, Madhya Pradesh, Chhattisgarh, Maharashtra, Odisha, Assam, Meghalaya, Mizoram and Telangana) of India covering almost the entire country. Molecular diagnosis of Plasmodium species was carried out among microscopically confirmed P. falciparum samples collected during a therapeutic efficacy study in different years. Results The polymerase chain reaction analysis revealed a high prevalence (18%) of mixed malaria parasite infections among microscopically confirmed P. falciparum samples from malaria patients that are either missed or left out by microscopy. Conclusions Deployment of molecular tools in areas of mixed species infection may prove vital for accurate diagnosis and treatment of malaria. Further, it will help in achieving the goal of malaria elimination in India. [ABSTRACT FROM AUTHOR]

• Mutations in five drug resistance genes were analysed in Pf and Pv isolates. • 76T in crt , 108N in dhfr , 437G in dhps and 184F in mdr1 were the most prevalent Pf mutations. • The ART resistance-associated mutations in Pfk13 were absent in all Pf isolates. • 383G in dhps , 117N in dhfr and 1076L in mdr1 were the most prevalent Pv mutations. • No mutations were in observed in Pvcrt-o and Pvk12 genes in Pv samples. One of the major challenges for malaria control and elimination is the spread and emergence of antimalarial drug resistance. Mutations in Plasmodium falciparum (Pf) and Plasmodium vivax (Pv) field isolates for five drug resistance genes viz. crt, mdr1, dhps, dhfr and kelch known to confer resistance to choloroquine (CQ), sulfadoxine-pyrimethamine (SP) and artemisinin (ART) and its derivatives were analyzed. A total of 342 symptomatic isolates of P. falciparum (Pf) and P. vivax (Pv) from 1993 to 2014 were retrieved from malaria parasite repository at National Institute of Malaria Research (NIMR). Sample DNA was extracted from dried blood spots and various targeted single nucleotide polymorphisms (SNPs) associated with antimalarial drug resistance were analysed for these isolates. 72S (67.7%) and 76T (83.8%) mutations along with S VMN T haplotype (67.7%) predominated the study population for Pfcrt. The most prevalent SNPs were 108N (73.2%) and 437G (24.8%) and the most prevalent haplotypes were ACN RN I (51.9%) and SAKAA (74.5%) in Pfdhfr and Pfdhps respectively. Only two mutations in Pfmdr1 , 86Y (26.31%) and 184F (56.26%), were seen frequently in our study population. No mutations associated with Pfk13 were observed. For Pv , all the studied isolates showed two Pvdhps mutations, 383G and 553G, and two Pfdhfr mutations, 58R and 117N. Similarly, three mutations, viz. 958M, 908L and 1076L were found in Pvmdr1. No variations were observed in Pvcrt-o and Pvk12 genes. Overall, our study demonstrates an increase in mutations associated with SP resistance in both Pf and Pv , however, no single nucleotide polymorphisms (SNPs) associated with ART resistance have been observed for either species. Various SNPs associated with CQ resistance were seen in Pf ; whereas only Pvmdr1 associated resistant SNPs were observed in Pv. Therefore, molecular characterization of drug resistance genes is essential for timely monitoring and prevention of malaria by identifying the circulating drug resistant parasites in the country. [ABSTRACT FROM AUTHOR]

Purpose: Presently, India is heading towards malaria elimination. In this context, analysing malaria epidemiological data of endemic pockets is important. In the present investigation, malaria prevalence in two endemic coastal localities of India viz., Besant Nagar (Chennai district) and Pamban (Ramanathapuram district) was analysed from 2004 to 2019 and correlated with the highest maximum temperature. Methods: Malaria surveillance data and entomological data from the malaria clinics were used to investigate epidemiological parameters. The annual malaria cases were correlated with the highest maximum temperature. Results: The analysis showed that the malaria case (P.v. and P.f.) reports were directly proportional to the temperature increase. Malaria cases were remarkably increased from 2004 to 2011 and subsequently, both Plasmodium vivax and Plasmodium falciparum cases were drastically reduced till 2019. P. vivax was higher than P. falciparum and the male population was found to be more affected. Mixed infection of P. vivax and P. falciparum was observed only in the Besant Nagar site (0.3%). The most affected age group was the adult age group (15 years and above) in both Besant Nagar (76.1%) and Pamban (69.5%). Conclusion: Improved surveillance, complete treatment and integrated vector control activities correlated with a declining trend of malaria cases in both the coastal sites towards malaria elimination. [ABSTRACT FROM AUTHOR]

Background: Malaria continues to be a major public health problem in the Northeastern part of India despite the implementation of vector control measures and changes in drug policies. To develop successful vaccines against malaria, it is important to assess the diversity of vaccine candidate antigens in field isolates. This study was done to assess the diversity of Plasmodium falciparum AMA-1 vaccine candidate antigen in a malaria-endemic region of Tripura in Northeast India and compare it with previously reported global isolates with a view to assess the feasibility of developing a universal vaccine based on this antigen. Methods: Patients with fever and malaria-like illness were screened for malaria and P. falciparum positive cases were recruited for the current study. The diversity of PfAMA-1 vaccine candidate antigen was evaluated by nested PCR and RFLP. A selected number of samples were sequenced using the Sanger technique. Results: Among 56 P. falciparum positive isolates, Pfama-1 was successfully amplified in 75% (n = 42) isolates. Allele frequencies of PfAMA-1 antigen were 16.6% (n = 7) for 3D7 allele and 33.3% (n = 14) in both K1 and HB3 alleles. DNA sequencing revealed 13 haplotypes in the Pfama-1 gene including three unique haplotypes not reported earlier. No unique amino-acid substitutions were found. Global analysis with 2761 sequences revealed 435 haplotypes with a very complex network composition and few clusters. Nucleotide diversity for Tripura (0.02582 ± 0.00160) showed concordance with South-East Asian isolates while recombination parameter (Rm = 8) was lower than previous reports from India. Population genetic structure showed moderate differentiation. Conclusions: Besides documenting all previously reported allelic forms of the vaccine candidate PfAMA-1 antigen of P. falciparum, new haplotypes not reported earlier, were found in Tripura. Neutrality tests indicate that the Pfama-1 population in Tripura is under balancing selection. This is consistent with global patterns. However, the high haplotype diversity observed in the global Pfama-1 network analysis indicates that designing a universal vaccine based on this antigen may be difficult. This information adds to the existing database of genetic diversity of field isolates of P. falciparum and may be helpful in the development of more effective vaccines against the parasite. [ABSTRACT FROM AUTHOR]

Background: Information on the foci of Plasmodium species infections is essential for any country heading towards elimination. Odisha, one of the malaria-endemic states of India is targeting elimination of malaria by 2030. To support decision-making regarding targeted intervention, the distribution of Plasmodium species infections was investigated in hard-to-reach areas where a special malaria elimination drive, namely Durgama Anchalare Malaria Nirakaran (DAMaN) began in 2017. Methods: A cross-sectional survey was conducted in 2228 households during July to November 2019 in six districts, to evaluate the occurrence of Plasmodium species. The species were identified by polymerase chain reaction (PCR) followed by sequencing, in case of Plasmodium ovale. Results: Of the 3557 blood specimens tested, malaria infection was detected in 282 (7.8%) specimens by PCR. Of the total positive samples, 14.1% were P. ovale spp. and 10.3% were Plasmodium malariae infections. The majority of P. ovale spp. (75.8%) infections were mixed with either Plasmodium falciparum and/or Plasmodium vivax and found to be distributed in three geophysical regions (Northern-plateau, Central Tableland and Eastern Ghat) of the State, while P. malariae has been found in Northern-plateau and Eastern Ghat regions. Speciation revealed occurrence of both Plasmodium ovale curtisi (classic type) and Plasmodium ovale wallikeri (variant type). Conclusions: In the present study a considerable number of P. ovale spp. and P. malariae were detected in a wide geographical areas of Odisha State, which contributes around 40% of the country's total malaria burden. For successful elimination of malaria within the framework of national programme, P. ovale spp. along with P. malariae needs to be incorporated in surveillance system, especially when P. falciparum and P. vivax spp. are in rapid decline. [ABSTRACT FROM AUTHOR]

Background: A detailed analysis of household and individual level Plasmodium infection patterns in two low-endemic districts of Meghalaya was undertaken to better understand the epidemiology of malaria in northeast India. Methods: Socio-demographic and behavioural information from residents (aged 1–69 years) of households were collected through pre-tested, questionnaire conducted in 2018 and 2019. Blood samples collected from participants were tested for Plasmodium falciparum and/or Plasmodium vivax infection using rapid diagnostic test, microscopy and PCR. Plasma samples from a subset of participants were analysed for antibodies against thirteen P. falciparum and four P. vivax antigens. Associations between household and individual level risk factors, and Plasmodium infections were evaluated using multilevel logistic regression models. Results: A total of 2753 individuals from 827 households were enrolled in 2018, and 834 individuals from 222 households were enrolled in 2019. Of them, 33 (1.2%) were positive by PCR for P. falciparum in 2018 and none were positive for P. vivax. In 2019, no PCR-positive individuals were detected. All, but one, infections were asymptomatic; all 33 infections were sub-microscopic. Reported history of malaria in the past 12 months (OR = 8.84) and history of travel in the past 14 days (OR = 10.06) were significantly associated with Plasmodium infection. A significant trend of increased seropositivity with age was noted for all 17 antigens. Although adults (≥ 18 years) consistently had the highest seropositivity rates, a sizeable proportion of under-five children were also found to be seropositive. Almost all individuals (99.4%) reported sleeping under an insecticide-treated bed-net, and household indoor residual spray coverage in the 12 months preceding the survey was low (23%). Most participants correctly identified common signs and symptoms of malaria, i.e., fever (96.4%), headache (71.2%), chills (83.2%) and body-ache (61.8%). Almost all participants (94.3%) used government-provided services for treatment of malaria. Conclusion: This study explored the epidemiology of malaria in two communities in Meghalaya, India, in the context of declining transmission. The presence of widespread asymptomatic infections and seropositivity among under-five children suggest that low-level Plasmodium transmission persists in this region. Implications of the study findings for malaria elimination efforts in low-transmission settings are discussed. [ABSTRACT FROM AUTHOR]

Malaria is a major global public health problem mainly in the tropics and subtropics. Malaria control and elimination strategies mainly rely on efficacious antimalarial drugs. At present the major hurdle faced by malaria control programs is the drug resistance to antimalarials. Molecular surveillance using genetic markers associated with resistance provides a valuable tool for detecting and tracking resistance as well as providing an in-depth understanding of the development and spread of resistance. Despite numerous published literatures there are limited review articles on molecular markers of drug resistance. Hence a review was planned. An exhaustive literature search was performed on PUBMED using "malaria", "resistance", "molecular", "antimalarial", "Chloroquine", "Pfcrt", "Pfmdr", "Pvcrt", "Pvmdr"as key words. Data pertaining to India and Southeast Asia were included. This review showed the widespread presence of molecular markers of drug resistance in Plasmodium falciparum and development of resistance in Plasmodium vivax over the years in Southeast Asia and India. This can have implications on malaria elimination and treatment guidelines in this region. [ABSTRACT FROM AUTHOR]

Background: Targeting multiple key antigens that mediate distinct Plasmodium falciparum erythrocyte invasion pathways is an attractive approach for the development of blood-stage malaria vaccines. However, the challenge is to identify antigen cocktails that elicit potent strain-transcending parasite-neutralizing antibodies efficacious at low immunoglobulin G concentrations feasible to achieve through vaccination. Previous reports have screened inhibitory antibodies primarily against well adapted laboratory parasite clones. However, validation of the parasite-neutralizing efficacy against clinical isolates with minimal in vitro cultivation is equally significant to better ascertain their prospective in vivo potency.Methods: We evaluated the parasite-neutralizing activity of different antibodies individually and in combinations against laboratory adapted clones and clinical isolates. Clinical isolates were collected from Central India and Mozambique, Africa, and characterized for their invasion properties and genetic diversity of invasion ligands.Results: In our portfolio, we evaluated 25 triple antibody combinations and identified the MSP-Fu+CyRPA+RH5 antibody combination to elicit maximal parasite neutralization against P. falciparum clinical isolates with variable properties that underwent minimal in vitro cultivation.Conclusions: The MSP-Fu+CyRPA+RH5 combination exhibited highly robust parasite neutralization against P. falciparum clones and clinical isolates, thus substantiating them as promising candidate antigens and establishing a proof of principle for the development of a combinatorial P. falciparum blood-stage malaria vaccine. [ABSTRACT FROM AUTHOR]

• Resistance to artesunate + sulfadoxine-pyrimethamine (AS+SP) has been flagged for Plasmodium falciparum malaria in India. • Meta-analysis on SP-resistance markers in India reveals high likelihood of drug failure. • Malaria genetic surveillance is urgently needed to generate evidence for drug policy change. • 12 states and 58 districts identified as hot spots for surveillance of SP-resistance markers. India is on track to eliminate malaria by 2030 but emerging resistance to first-line antimalarials is a recognised threat. Two instances of rapid development, spread, and natural selection of drug-resistant mutant parasites in India (chloroquine across the country and artesunate + sulfadoxine-pyrimethamine [AS+SP] in the northeastern states) translated into drug policy changes for Plasmodium falciparum malaria in 2010 and 2013, respectively. Considering these rapid changes in the SP drug resistance-conferring mutation profile of P. falciparum , there is a need to systematically monitor the validated mutations in Pfdhfr and Pfdhps genes across India alongside AS+SP therapeutic efficacy studies. There has been no robust, systematic countrywide surveillance reported for these parameters in India, hence the current study was undertaken. Studies that reported data on WHO-validated SP resistance markers in P. falciparum across India from 2008 to January 2023 were included. Five major databases, PubMedⓇ, Web of ScienceTM, ScopusⓇ, EmbaseⓇ, and Google Scholar, were exhaustively searched. Individual and pooled prevalence estimates of mutations were obtained through random- and fixed-effect models. Data were depicted using forest plots created with a 95% confidence interval. The study is registered with PROSPERO (CRD42021236012). A total of 37 publications, and 533 Pfdhfr and 134 Pfdhps National Centre of Biotechnology Information (NCBI) DNA sequences were included from >4000 samples. The study included information from 80 districts, 21 states and 3 union territories (UTs) from India. The two PfDHFR mutations, C59R (62%) and S108N (74%), were the most prevalent mutations (pooled estimates 61% and 71%, respectively) and appeared to be stabilised/fixed. Although rarest overall, the prevalence of I164L was observed to be as high as 32%. The Pf DHFR double mutants were the most prevalent overall (51%; pooled 42%). The prevalence of triple and quadruple mutations was 6% and 5%, respectively, and is an immediate concern for some states. The most prevalent Pf DHPS mutation was A437G (39%), followed by K540E (25%) and A581G (12%). There was a low overall prevalence of Pf DHFR/ Pf DHPS quintuple and sextuple mutations but surveillance for these mutations is critical for some areas. The analyses span the two critical policy changes, highlight the areas of concern, and guide policymakers in strategising and refining the anti-malaria drug policy for malaria elimination. The results of the analyses also highlight the SP-resistance hot spots, critical gaps and challenges, and indicate that focal and local malaria genetic surveillance (including drug-resistance markers) is needed until malaria is successfully eliminated. [Display omitted] [ABSTRACT FROM AUTHOR]

Artemisinin is the frontline fast-acting anti-malarial against P. falciparum. Emergence and spread of resistant parasite in eastern-India poses a threat to national malaria control programs. Therefore, the objective of our study is to evaluate the artesunate-sulfadoxine-pyrimethamine efficacy in Central India. 180 monoclonal P. falciparum-infected patients received standard ASSP therapy during August 2015–January 2017, soon after diagnosis and monitored over next 42-days. Artemisinin-resistance was assessed through in-vivo parasite clearance half-life (PC1/2), ex-vivo ring-stage survivability (RSA), and genome analysis of kelch13 and other candidate gene (pfcrt, pfmdr1, pfatpase 6, pfdhfr and pfdhps). Of 180 P. falciparum positive patients, 9.5% showed increased PC1/2 (> 5.5 h), among them eleven isolates (6.1%) showed reduced sensitivity to RSA. In 4.4% of cases, parasites were not cleared by 72 h and showed prolonged PC1/2(5.6 h) (P < 0.005) along with significantly higher RSA (2.2%) than cured patients (0.4%). None of day-3 positive isolates contained the pfkelch13 mutation implicated in artemisinin resistance. Parasite recrudescence was observed in 5.6% patients, which was associated with triple dhfr–dhps (A16I51R59N108I164–S436G437K540G581T613) combination mutation. Emergence of reduced sensitivity to artesunate-sulfadoxine-pyrimethamine, in central India highlighted the risk toward spread of resistant parasite across different parts of India. Day-3 positive parasite, featuring the phenotype of artemisinin-resistance without pfkelch13 mutation, suggested kelch13-independent artemisinin-resistance. [ABSTRACT FROM AUTHOR]

Introduction: There are several tropical mosquito borne infections, such as Malaria and Dengue, these are the two major and common arthropod borne infections that cause high morbidity and mortality in many patients and are major public health concern worldwide. Concurrent malaria and dengue infection is an important condition that is infrequently reported. Aim: To find out the prevalence rate of co-infection for both dengue and malaria and also to associate the severity of such co-infections with symptoms and haematological parameters. Materials and Methods: In observational cross-sectional study, samples were collected from 604 febrile patients clinically suspected for malaria and dengue attending the Medicine Out Patient Department (OPD) of Dhiraj Hospital, Gujarat, India. The samples were diagnosed for malaria by using rapid malaria antigen test kit and peripheral blood smear microscopy for the identification of Plasmodium spp. The dengue NS-1 antigen, (Immunoglobulin) IgM and IgG antibodies rapid kit were use to rule out dengue infection. The statistical analysis done by software EPI info. Results: In the present study, out of 604, 58 patients (9.6%) were positive for malaria and 80 patients (13.24%) were positive for dengue, while 21 patients (3.47%) were having concomitant infection with both dengue and malaria. The most affected age group was 31-60 years, 45.53%. The study revealed male preponderance (61.59%), hepatomegaly and jaundice were seen in 52.38% patients, haemorrhagic manifestations in 23.80%, kidney failure in 4.76% (1), and thrombocytopenia (platelet count <150,000/cubic millimeter (cmm)) were noted in 95.23% patients. Conclusion: It is mandatory to test any febrile patient for both malaria and dengue so that the diagnosis of the patient is not held back by missing any one. The clinicians who treat febrile patients in or from endemic areas must thoroughly examine and diagnose for both malaria and dengue, still one or the other is positive. Vector control, health education and good hygiene are community based preventive measures that are needed to control both the diseases. [ABSTRACT FROM AUTHOR]

Antibiotic resistance in the context of treating malarial infections is a major challenge in India. Home remedies such as thulasi leaves (Ocimum tenuiflorum), black pepper seeds (Piper nigrum), clove buds (Syzygium aromaticum), cinnamon bark (Cinnamomum verum), and nilavembu whole plant powder (Andrographis paniculata) were taken to explore antimalarial and methicillin-resistant Staphylococcus aureus (MRSA) activity. Among the five extracts, the best two extracts, C. verum and P. nigrum extract, showed the presence of Quercetin. Phytoniosomes were prepared by simple probe sonication with the two extracts and the resultant vesicles were in the size range of (319.7 nm). They showed significant (P < 0.001) antimalarial potency IC50 at 5.25 µg/ml against P. falciparum 3D7. In addition, their cytotoxicity (TC50) against Vero cell line was found to be > 100 µg/ml. The therapeutic index was found to be > 32 µg/ml. Phytoniosomes were converted to a capsule dosage form by lyophilization and this capsule was stable up to 90 days. [ABSTRACT FROM AUTHOR]

Hematological abnormalities such as anemia and coagulopathies are well-established complications in malaria. Other well-recognized complications of malaria are cerebral malaria and kidney injury. In this case report, we present a man that presented with a gastrointestinal bleeding as a complication of a malaria infection, which was found through histopathological investigations of the resected colon. [ABSTRACT FROM AUTHOR]

Background: Despite declining incidence over the past decade, malaria remains an important health burden in India. This study aimed to assess the village-level temporal patterns of Plasmodium infection in two districts of the north-eastern state of Meghalaya and evaluate risk factors that might explain these patterns. Methods: Primary Health Centre passive malaria case data from 2014 to 2018 were analysed to characterize village-specific annual incidence and temporal trends. Active malaria case detection was undertaken in 2018 and 2019 to detect Plasmodium infections using PCR. A questionnaire collected socio-demographic, environmental, and behavioural data, and households were spatially mapped via GPS. Adult mosquitoes were sampled at a subset of subjects' houses, and Anopheles were identified by PCR and sequencing. Risk factors for Plasmodium infection were evaluated using bivariate and multivariate logistic regression analysis, and spatial cluster analysis was undertaken. Results: The annual malaria incidence from PHC-based passive surveillance datasets in 2014–2018 was heterogenous but declining across villages in both districts. Active surveillance in 2018 enrolled 1468 individuals from 468 households (West Jaintia Hills) and 1274 individuals from 359 households (West Khasi Hills). Plasmodium falciparum prevalence per 100 people varied from 0 to 4.1% in the nine villages of West Jaintia Hills, and from 0 to 10.6% in the 12 villages of West Khasi Hills. Significant clustering of P. falciparum infections [observed = 11, expected = 2.15, Relative Risk (RR) = 12.65; p < 0.001] was observed in West Khasi Hills. A total of 13 Anopheles species were found at 53 houses in five villages, with Anopheles jeyporiensis being the most abundant. Risk of infection increased with presence of mosquitoes and electricity in the households [Odds Ratio (OR) = 1.19 and 1.11], respectively. Households with reported animals had reduced infection risk (OR = 0.91). Conclusion: Malaria incidence during 2014–2018 declined in all study villages covered by the passive surveillance data, a period that includes the first widespread insecticide-treated net campaign. The survey data from 2018 revealed a significant association between Plasmodium infection and certain household characteristics. Since species of Plasmodium-competent mosquito vectors continue to be abundant, malaria resurgence remains a threat, and control efforts should continue. [ABSTRACT FROM AUTHOR]

• "cys2/MFK*/*REY" motif with short length were restricted to cerebral and severe malaria. • Higher expression level of var group A was associated with malaria severity. • Peptide VAR P5 (DIGDIVRGKDLY) appeared to be a good marker for severity. Plasmodium falciparum erythrocyte membrane protein is encoded by a highly variable multicopy var gene family known to play a key role in malaria pathogenicity. Therefore, we investigated sequence variation, expression profile and immune response of the Duffy binding-like domain (DBLα) region of the var gene. Blood samples were collected from patients with cerebral, severe and mild malaria in Chhattisgarh, India, a region with endemic malaria. Polymerase chain reaction amplicons were cloned and sequenced to determine sequence variation. The expression level was analyzed targeting the upstream region of var gene using the Delta-Delta-Ct method. Immunoglobulin G (IgG) level was determined against the 6 synthetic peptides of the DBLα region. The study identified that group 1 and group 5 sequences (cysteine/position of limited variability (cys/PoLV) classification) along with cys2/cys4 and MFK*/REY motifs and short amino acid length were significantly associated with malaria severity. The specific PoLV (MFKS, LREA, PTNL) were restricted to cerebral malaria. The expression level of var group A was higher than var groups B and C, demonstrating its prognostic characteristic. All peptides showed high-quality IgG response, while VAR P5 appeared to be a good marker for severity. The present study illustrates the presence of specific sequences of DBLα tags involved in severe malaria that could be targeted in future interventions for malaria control and elimination. [ABSTRACT FROM AUTHOR]

Context: Malaria is one of the major public health problems worldwide with increased morbidity and mortality. India has the fourth highest malaria burden in the world. Malaria parasite affects multiple organs of the body including liver, spleen, brain, kidney, blood vessels etc. Organ dysfunction predominantly seen in Plasmodium falciparum malaria, is not in Plasmodium vivax infections. Aims: The present study investigated the abnormalities of liver function test in acute cases of Plasmodium Falciparum infection. Settings and Design: A hospital-based case--control study conducted in a tertiary care center in West Bengal, India, for a duration of 7 months and included 53 acute falciparum malaria cases attending the outpatient department (OPD) and 47 age and sex matched patients, with complaints unrelated to the present study, as controls. Materials and Methods: Serum levels of bilirubin, serum glutamic oxaloacetic transaminase (SGOT), serum glutamic pyruvate transaminase (SGPT), alkaline phosphatase, total protein and albumin were assayed using autoanalyzer (Konelab 60i autoanalyser). Statistical analysis used: SPSS software version 20 (IBM, New York, USA). Results: In this study serum bilirubin (total, conjugated and unconjugated ) SGOT, SGPT, alkaline phosphatase levels were highly significantly raised in cases with respect to controls. Levels of total protein, albumin and globulin in cases was not significantly different from the corresponding levels in controls. Conclusions: Hepatic function impairment in malaria is an important and potentially dangerous complication. Data from the present study can be useful for future research and also help in determining the prognosis and treatment. [ABSTRACT FROM AUTHOR]

Background: One of the most common infections caused by protozoa in India is malaria. Malaria cases have different geographic distribution in different areas. So it is important that hospitals in different areas draw their own set of data regarding clinical and hematological characteristics of malaria. The present study was conducted to evaluate the spectrum of clinical and hematological changes in patients of malaria. Material and method: We conducted retrospective study at Pathology department, Sumandeepvidyapeeth, with 55 patients aged between 5 to 75 years. Results: Out of 55, 40 were males and 15 were females. PV constituted majority of the cases, followed by PF. We reported anemia in 34 of malaria cases. Maximum cases of anemia was seen in PF. Thrombocytopenia was seen in 42 of malaria cases. Maximum cases was seen in PV. Leucocyte count was within reference range in majority of our cases 42. Conclusion: Fever with rigors and chills, headache and splenomegaly are the most common clinical alteration and anemia and thrombocytopenia are most common hematological alteration in cases of malaria. Presence of these classic picture in a patient ever, helps diagnosing malaria at the earlier stages. In that way, early treatment can be introduced. [ABSTRACT FROM AUTHOR]

India is a major contributor to the global burden of malaria, especially Plasmodium vivax infection. Understanding the spatiotemporal trends of malaria across India over the last two decades may assist in targeted intervention. The population-normalized spatiotemporal trends of malaria epidemiology in India from 2007 to 2022 were analyzed using a geographic information system with the publicly available "malaria situation" report of the National Vector Borne Disease Control Program (NVBDCP). The NVBDCP data showed malaria cases to have steeply declined from 1.17 million in 2015 to 0.18 million cases in 2022; this is 10.1 and 18.7 fold lower than the WHO's estimate of 11.93 million and 3.38 million cases in 2015 and 2022, respectively. From 2007 to 2022, Mizoram, Meghalaya, Tripura, Odisha, Chhattisgarh, and Jharkhand consistently reported high caseloads of Plasmodium falciparum. In the same period, the P. vivax caseload was high in Arunachal Pradesh, Mizoram, Nagaland, Jharkhand, Odisha, Chhattisgarh, Goa, Daman and Diu, Dadra and Nagar Haveli, and Andaman and Nicobar Islands. The distribution of forest cover, annual rainfall, and proportion of the Scheduled Tribe population (the most underprivileged in Indian society) spatially correlated with malaria cases and deaths. Mizoram is the only state where cases were higher in 2022 than in 2007. Overall, India has made tremendous progress in controlling malaria and malaria-related deaths in the last decade. The decline could be attributed to the effective vector and parasite control strategies implemented across the country.

Traditionally attributed only to Plasmodium falciparum, Plasmodium vivax has recently been reported to cause a significant burden of complicated malaria cases. The present study aimed to delineate the clinical spectrum and identify predictors for severe disease. This was a prospective observational cohort study conducted at a tertiary care hospital in North India. Patients with acute febrile illness (AFI) aged at least 14 years were included if they were diagnosed with vivax malaria based on rapid kits or peripheral smears. Clinical data and investigations during hospital stay was recorded. 439 cases of acute febrile illness were screened, of whom 50 (11%) were diagnosed with malaria including eight P. falciparum infections. Forty-two vivax malaria cases, 22 (52%) of whom were severe, were followed till discharge or death. The median age of the cohort was 24.5 years (Q1-Q3, 19-36 years), including a total of 29 males (69%). Severe malaria was more frequently associated with historical complaints of oliguria or dyspnea, and examination findings of pallor, splenomegaly or altered sensorium. The following five factors were identified to predict severe disease: prolonged illness over 7 days, symptoms of oliguria or dyspnea, examination findings of pallor or crepitations on auscultation. Malaria accounts for 1 in 10 cases of AFI at our North Indian tertiary care center and approximately half of them present with severe disease. Prolonged duration of disease prior to presentation is a modifiable predictor for severe disease and should be targeted for reducing morbidity. [ABSTRACT FROM AUTHOR]

Image, graphical abstract • 5 compounds were isolated from Xanthium strumarium L. by bioassay guided method. • Antimalarial efficacy was studied against Plasmodium falciparum 3D7 strain. • Two compounds showed excellent antimalarial activity with IC 50 = 27.25 and 7.14 μM. • Results supported the traditional use of X. strumarium L. for the cure of malaria. Malaria is a major threat to global health and continues to claim lives of many people each year, especially in developing countries. Xanthium strumarium L., is used by traditional health practitioners in the management of malaria fever in North East India. Bioassay guided fractionation of X. strumarium L. extracts, led to the isolation of five compounds from the aerial part and fruit of Xanthium strumarium , namely, stigmasta-5,22-dien-3β-ol (1), xanthinosin (2), stigmasterol-3- O - β -D-glucopyranoside (3), oleic acid (4) and (E)-2, 3-dihydroxypropyl-octadec-9-enoate (5). Antimalarial potential of isolated compounds were evaluated against 3D7 strain of Plasmodium falciparum by schizont maturation inhibition assay method. The structures of isolated compounds were established by HR-MS, NMR experiments and comparison from literature data. Compounds 2 and 3 showed significant antimalarial properties against 3D7 strain of P.falciparum with IC 50 value 27.25 and 7.14μM respectively. The antimalarial activities of xanthinosin (2) and stigmasterol-3-O-β-D-glucopyranoside (3) lend credence to the application of X. strumarium against malaria and thus provides new template for development of antimalarial molecules. X. strumarium has also scope for the development of antimalarial phyto-pharmaceutical with five identified compounds with P.falciparum inhibitory activity as per the drugs and cosmetics act of India (1940, amended 31st December 2016). [ABSTRACT FROM AUTHOR]

Introduction: Drug resistant malaria represents the most challenging health problem worldwide. The major threat to malaria control is the increasing incidence of complicated and drug-resistant cases. Aim: To determine the prevalence of clinical artesunate resistance among complicated malaria cases by invivo method. Materials and Methods: The study included 241 patients with complicated malaria, which showed trophozoites of malarial parasite Plasmodium in the blood smear. They were administered with artesunate monotherapy. They were then followed-up for clinical response to therapy and quantification of parasitaemia levels on day 1, day 2, day 3, day 7, day 14 and day 28. Clinical examination included core body temperature recording, assessment of associated complications like vomiting, altered sensorium, anaemia, bleeding tendencies and investigation of renal parameters, liver function test and hypoglycaemia. Results: Out of 241 complicated malaria cases showing malarial parasite in peripheral blood smear, 204 cases were of Plasmodium falciparum, 32 cases were of Plasmodium vivax and five were cases of mixed Plasmodium falciparum and Plasmodium vivax infection. Among the 241 malaria positive cases, the most commonly associated complication was severe anaemia in 87 (36.1%) cases. All the 32 cases of Plasmodium vivax malaria and the five cases of mixed infection responded and recovered after administration of artesunate monotherapy. But among the 204 cases of Plasmodium falciparum, clinical artesunate resistance was detected in 6 (2.94%) cases and the rest 198 (97.06%) cases were sensitive to artesunate monotherapy. Conclusion: Clinical artesunate resistance was noticed in 2.94% cases of Plasmodium falciparum malaria. Hence, continuous monitoring of antimalarial therapy is required, especially in complicated cases of Plasmodium falciparum malaria, to prevent the emergence of artesunate resistance. [ABSTRACT FROM AUTHOR]

Background: Artesunate plus sulfadoxine-pyrimethamine (ASP) is first-line treatment for uncomplicated Plasmodium falciparum malaria in most of India, except for six North-eastern provinces where treatment failure rates were high. In Ujjain, central India, the frequency of mutations associated with increased drug tolerance, but not overt resistance to sulfadoxine and pyrimethamine were 9% and > 80%, respectively, in 2009 and 2010, just prior to the introduction of ASP. The frequency of drug resistance associated mutations in Ujjain in 2015-2016 after 3-4 years of ASP use, are reported. Methods: Blood samples from patients with P. falciparum mono-infection verified by microscopy were collected on filter-paper at all nine major pathology laboratories in Ujjain city. Codons pfdhfr 16-185, pfdhps 436-632 and K13 407-689 were identified by sequencing. Pfcrt K76T and pfmdr1 N86Y were identified by restriction fragment length polymorphism. Results: Sulfadoxine-pyrimethamine resistance-associated pfdhfr 108 N and 59R alleles were found in 100/104 (96%) and 87/91 (96%) samples, respectively. Pfdhps 437G was found in 10/105 (10%) samples. Double mutant pfdhfr 59R + 108 N were found in 75/81 (93%) samples. Triple mutant pfdhfr 59R + 108 N and pfdhps 437G were found in 6/78 (8%) samples. Chloroquine-resistance-associated pfcrt 76T was found in 102/102 (100%). Pfmdr1 N86 and 86Y were identified in 83/115 (72%) and 32/115 (28%) samples, respectively. Conclusion: The frequency of P. falciparum with reduced susceptibility to sulfadoxine-pyrimethamine remained high, but did not appear to have increased significantly since the introduction of ASP. No polymorphisms in K13 associated with decreased artemisinin susceptibility were found. ASP probably remained effective, supporting continued ASP use. [ABSTRACT FROM AUTHOR]

Context: The arid climate of Western Rajasthan is challenging for malaria transmission, with the number of cases correlating directly with the annual rainfall pattern. Moreover, >90% of the cases in this region are caused by Plasmodium vivax, which has recently been shown to cause a similar degree of thrombocytopenia as Plasmodium falciparum. Aims and Objectives: The aim of the study was to determine the degree of thrombocytopenia in malaria patients and its association with different species of malaria in this region with an unstable malaria epidemiology. Materials and Methods: This retrospective study was conducted on all microbiologically confirmed malaria patients with documented platelet counts from August 2017 to October 2018. Microbiological diagnosis was established by rapid diagnostic tests and peripheral blood film examination. Platelet counts were used to assess the degree of thrombocytopenia. Results: A total of 130 cases were included in the study, of which 118 (91%) were caused by P. vivax, while the rest 12 (9%) were caused by P. falciparum. Thrombocytopenia was present in 108 (83%) cases, and the mean values of platelets in thrombocytopenic patients with P. vivax and P. falciparum infection were 72600/μL and 48500/μL, respectively. Although P. falciparum infection was significantly associated with severe thrombocytopenia (odds ratio: 4.7, [95% confidence interval 1.3–16.1]), extremely low platelet counts (n = 5) warranting platelet transfusions (n = 1) were seen only in P. vivax cases. Only one patient required platelet transfusions in these patients suggesting good tolerance to thrombocytopenia. Conclusions: Avoiding unnecessary transfusions in febrile thrombocytopenic patients with an established malaria diagnosis can help in reducing transfusion-transmitted infections. [ABSTRACT FROM AUTHOR]

Context: Both malaria and lymphatic filariasis (LF) are mosquito-borne diseases caused by protozoal and nematode parasites, respectively, and are serious public health problem in India. Although the vectors of the diseases are different, they can coexist in favorable conditions. Fever is the common symptom for both the diseases, but the emphasis is given for diagnosis and treatment of malaria due to its life-threatening severity, LF remained neglected. Detection and management of microfilaria are equally important. During the diagnosis of malaria, a few microfilaria were detected, which prompted us to undertake this study with following objectives. Objectives: The objective of the study was to determine the incidence of microfilaremia among the febrile patients attending for malaria diagnosis. Subjects and Methods: Thick and thin peripheral blood smears from all patients attended were examined following Giemsa staining. Different malarial indexes were analyzed. Results: Out of 8681 patients examined, 1778 were positive for Plasmodium vivax and 328 for Plasmodium falciparum with a slide positivity rate 20.48%. Twenty-six patients were positive for microfilaria of Wuchereria bancroftii among which five were coinfected with P. vivax and one with P. falciparum. Most of the microfilaria-positive patients were adult and originally from northern districts of Bihar. Conclusions: High incidence of microfilaria among febrile patients attending for malaria is alarming for urban Kolkata. Although the patients were originally from Bihar, they are staying in Kolkata for a long time, might be a source for transmission. Epidemiological study by collecting night blood samples and entomological survey is highly suggestive to explore local transmission if any. [ABSTRACT FROM AUTHOR]

Ozonide antimalarials, OZ277 (arterolane) and OZ439 (artefenomel), are synthetic peroxide-based antimalarials with potent activity against the deadliest malaria parasite, Plasmodium falciparum. Here we used a "multi-omics" workflow, in combination with activity-based protein profiling (ABPP), to demonstrate that peroxide antimalarials initially target the haemoglobin (Hb) digestion pathway to kill malaria parasites. Time-dependent metabolomic profiling of ozonide-treated P. falciparum infected red blood cells revealed a rapid depletion of short Hb-derived peptides followed by subsequent alterations in lipid and nucleotide metabolism, while untargeted peptidomics showed accumulation of longer Hb-derived peptides. Quantitative proteomics and ABPP assays demonstrated that Hb-digesting proteases were increased in abundance and activity following treatment, respectively. Ozonide-induced depletion of short Hb-derived peptides was less extensive in a drug-treated K13-mutant artemisinin resistant parasite line (Cam3.IIR539T) than in the drug-treated isogenic sensitive strain (Cam3.IIrev), further confirming the association between ozonide activity and Hb catabolism. To demonstrate that compromised Hb catabolism may be a primary mechanism involved in ozonide antimalarial activity, we showed that parasites forced to rely solely on Hb digestion for amino acids became hypersensitive to short ozonide exposures. Quantitative proteomics analysis also revealed parasite proteins involved in translation and the ubiquitin-proteasome system were enriched following drug treatment, suggestive of the parasite engaging a stress response to mitigate ozonide-induced damage. Taken together, these data point to a mechanism of action involving initial impairment of Hb catabolism, and indicate that the parasite regulates protein turnover to manage ozonide-induced damage. Author summary: The ozonides are a novel class of fully synthetic antimalarial drugs with potent activity against all parasite species that cause malaria, including the deadliest, Plasmodium falciparum. With the emergence of resistance to current frontline artemisinin-based antimalarials, new drugs are urgently needed and a clear understanding of their mechanism of action is essential so that they can be optimally deployed in the field. Here, we studied the biochemical effects of two ozonides, OZ277 (marketed in India in combination with piperaquine) and OZ439 (in Phase IIb clinical trials) in P. falciparum parasites using an untargeted multi-omics approach consisting of proteomics, peptidomics and time-dependent metabolomics, along with activity-based protease profiling. We found that the ozonides initially disrupt haemoglobin metabolism and that they likely engage the parasite proteostatic stress response. Furthermore, when the duration of ozonide exposure was extended beyond 3 hours to reflect clinically-relevant exposure periods, additional parasite biochemical pathways were perturbed. This comprehensive analysis provides new insight into the antimalarial mode of action of ozonides and provides new opportunities for interventions to enhance their antimalarial efficacy. [ABSTRACT FROM AUTHOR]

Background: India has launched the malaria elimination initiative in February 2016. Studies suggest that estimates of malaria are useful to rationalize interventions and track their impact. Hence, a national study was launched to estimate burden of malaria in India in 2015. Methods: For sampling, all 624 districts of India were grouped in three Annual Parasite Incidence (cases per thousand population) categories, < two (low); two-five (moderate) and > five (high) API. Using probability proportional to size (PPS) method, two districts from each stratum were selected covering randomly 200,000 persons per district. Active surveillance was strengthened with 40 trained workers per study district. Data on malaria cases and deaths was collated from all health care providers i.e. pathological laboratories, private practitioners and hospitals in private and public health sectors and was used for analysis and burden estimation. Results: Out of 1215,114 population under surveillance, 198,612 (16.3%) tests were performed and 19,386 (9.7%) malaria cases were detected. The malaria cases estimated in India were 3875,078 (95% confidence interval 3792,018–3958,137) with API of 3.05 (2.99–3.12) including 2789,483 (2740,577–2838,389) Plasmodium falciparum with Annual Falciparum Incidence of 2.2 (2.16–2.24). Out of 8025 deaths investigated, 102 (1.27%) were attributed to malaria. The estimated deaths in India were 29,341 (23,354–35,327) including 19,067 (13,665–24,470) confirmed and 10,274 (7694–12,853) suspected deaths in 2015–2016. Conclusions: Estimated malaria incidence was about four folds greater than one million reported by the national programme, but three folds lesser than thirteen million estimated by the World Health Organization (WHO). However, the estimated deaths were 93 folds more than average 313 deaths reported by the national malaria programme in 2015–2016. The 29,341 deaths were comparable with 24,000 deaths in 2015 and 22,786 deaths in 2016 estimated by the WHO for India. These malaria estimates can serve as a benchmark for tracking the success of malaria elimination campaign in India. [ABSTRACT FROM AUTHOR]

Background: In 2017, nearly 80% of malaria morbidity and mortality occurred in sub-Saharan African (SSA) countries and India. Rapid diagnostic tests (RDTs), especially those targeting histidine-rich protein 2 (PfHRP2) of Plasmodium falciparum, have become an important diagnostic tool in these malaria-endemic areas. However, the chances of RDT-oriented successful treatment are increasingly jeopardized by the appearance of mutants with deletions in pfhrp2 and pfhrp3 genes. This systematic review and meta-analysis determines the prevalence of field P. falciparum isolates with deletion in pfhrp2 and/or pfhrp3 genes and their proportion among false-negative results in the PfHRP2-based RDTs in SSA and India. Methods: Eight electronic databases were used for searching potentially relevant publications for the systematic review analysis, wherein the main methodological aspects of included studies were analysed and some missing links in the included studies were identified. Results: A total of 19 studies were included, 16 from SSA and 3 from India. The pooled prevalence of pfhrp2 deletions was 8 and 5% while 16 and 4% for pfhrp3 gene deletions in Africa and India, respectively. The pooled proportion of pfhrp2 gene deletions found among false negative PfHRP2-based RDTs results was about 27.0 and 69.0% in Africa and India, respectively. Conclusions: This review study indicates a relatively high proportion of both pfhrp2/3 genes deletions in P. falciparum isolates and among false-negative malaria cases using PfHRP2-based RDT results in SSA and India. Recently the deletions in pfhrp2/3 genes have also been reported from two African countries (Nigeria and Sudan). This review emphasizes the importance of more extensive studies and standardization of studies addressing the pfhrp2/3 gene deletions in malarious areas. [ABSTRACT FROM AUTHOR]

Background: Deltamethrin 62.5 polymer-enhanced suspension concentrate (SC-PE) is one of the World Health Organization-approved insecticides for indoor residual spraying and was recommended to evaluate its residual activity for determination of appropriate spray cycles in different eco-epidemiologic settings. In the current study, efficacy of deltamethrin 62.5 SC-PE was evaluated against vectors of malaria and its impact on malaria incidence in a Plasmodium falciparum hyper-endemic area in Koraput district, Odisha State, India. Methods: The trial had two comparable arms, arm 1 with residual spraying of deltamethrin 62.5 SC-PE and arm 2 with deltamethrin 2.5% WP (positive control). Comparative assessment of the impact of each intervention arm on entomological (density, parity, infection and human blood index), epidemiological (malaria incidence) parameters, residual efficacy and adverse effects were evaluated. Results: Both the arms were comparable in terms of entomological and epidemiological parameters. While, deltamethrin 62.5 SC-PE was found to be effective for 150 days in mud and wood surfaces and 157 days in cement surfaces; deltamethrin 2.5% was effective only for 105 days on mud surfaces and 113 days on cement and wood surfaces. Conclusions: Deltamethrin 62.5 SC-PE had prolonged killing effectiveness up to 5 months. Hence, one round of IRS with deltamethrin 62.5 SC-PE would be sufficient to cover two existing malaria peak transmission seasons (July–August and October–November) in many parts of India. [ABSTRACT FROM AUTHOR]

Background: In India, Accredited Social Health Activists (ASHAs) deliver services for diagnosis and treatment of malaria, although unlicensed medical practitioners (UMPs) (informal health providers) are most preferred in communities. A cross sectional survey was conducted to: (i) assess knowledge and treatment-seeking practices in the community, and (ii) explore the diagnosis and treatment practices related to malaria of UMPs working in rural and tribal-dominated high malaria endemic areas of central India, and whether they adhere to the national guidelines. Methods: A multi-stage sampling method and survey technique was adopted. Heads of the households and UMPs were interviewed using a structured interview schedule to assess knowledge and malaria treatment practices. Results: Knowledge regarding malaria symptoms was generally accurate, but misconceptions emerged related to malaria transmission and mosquito breeding places. Modern preventive measures were poorly accessed by the households. UMPs were the most preferred health providers (49%) and the first choice in households for seeking treatment. UMPs typically lacked knowledge of the names of malaria parasite species and species-specific diagnosis and treatment. Further, irrational use of anti-malarial drugs was common. Conclusions: UMPs were the most preferred type of health care providers in rural communities where health infrastructure is poor. The study suggests enhancing training of UMPs on national guidelines for malaria diagnosis and treatment to strengthen their ability to contribute to achievement of India's malaria elimination goals. [ABSTRACT FROM AUTHOR]