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PubMed | Municipal Corporation of Delhi MCD and National Institute of Malaria Research NIMR
Type: Journal Article | Journal: PloS one | Year: 2016

The study is based on hypothesis that whether continuous entomological surveillance of Ae. aegypti and simultaneous appropriate interventions in key containers during non-transmission (December-May) months would have any impact on breeding of Aedes and dengue cases during the following transmission months (June-November). The impact of the surveillance and intervention measures undertaken during non-transmission months were assessed by entomological indicators namely container index (CI), house index (HI), pupal index (PI) and breteau index (BI).A total of 28 localities of West Zone of Delhi with persistent dengue endemicity were selected for the study. Out of these localities, 20 were included in study group while other 8 localities were in control group. IEC and various Aedes breeding control activities were carried out in study group in both non-transmission and transmission season whereas control group did not have any such interventions during non-transmission months as per guidelines of MCD. These activities were undertaken by a team of investigators from NIMR and SDMC, Delhi. In control group, investigators from NIMR carried out surveillance activity to monitor the breeding of Aedes mosquito in localities.Comparison of baseline data revealed that all indices in control and study group of localities were comparable and statistically non-significant (p>0.05). In both study and control groups, indices were calculated after pooling data on seasonal basis, i.e., transmission and non-transmission months for both years. The test of significance conducted on all the four indices, i.e., HI, PI, CI, and BI, revealed a significant difference (p<0.05) between the study group and control group during transmission and non-transmission months except in HI. Due to consistent intervention measures undertaken in non-transmission months in study group, reduction in CI, HI, BI and PI was observed 63%, 62%, 64% and 99% respectively during transmission months as compared to control group where increase of 59%, 102%, 73% and 71% respectively. As a result of reduction in larval indices, no dengue case (except one NS1) was observed in study group, whereas 38 dengue cases were observed in control group.Through this pilot study, it is concluded that proper intervention in non-transmission season reduces vector density and subsequently dengue cases in transmission season.

Mixson-Hayden T.,Centers for Disease Control and Prevention | Mixson-Hayden T.,Atlanta Research and Education Foundation | Jain V.,National Institute of Malaria Research NIMR Field Station | McCollum A.M.,Centers for Disease Control and Prevention | And 8 more authors.
Antimicrobial Agents and Chemotherapy | Year: 2010

Treatment of Plasmodium falciparum is complicated by the emergence and spread of parasite resistance to many of the first-line drugs used to treat malaria. Antimalarial drug resistance has been associated with specific point mutations in several genes, suggesting that these single nucleotide polymorphisms can be useful in tracking the emergence of drug resistance. In India, P. falciparum infection can manifest itself as asymptomatic, mild, or severe malaria, with or without cerebral involvement. We tested whether chloroquine- and antifolate drug-resistant genotypes would be more commonly associated with cases of cerebral malaria than with cases of mild malaria in the province of Jabalpur, India, by genotyping the dhps, dhfr, pfmdr-1, and pfcrt genes using pyrosequencing, direct sequencing, and real-time PCR. Further, we used microsatellites surrounding the genes to determine the origins and spread of the drug-resistant genotypes in this area. Resistance to chloroquine was essentially fixed, with 95% of the isolates harboring the pfcrt K76T mutation. Resistant genotypes of dhfr, dhps, and pfmdr-1 were found in 94%, 17%, and 77% of the isolates, respectively. Drug-resistant genotypes were equally likely to be associated with cerebral malaria as with mild malaria. We found evidence of a selective sweep in pfcrt and, to a lesser degree, in dhfr, indicating high levels of resistance to chloroquine and evolving resistance to pyrimethamine. Microsatellites surrounding pfcrt indicate that the resistant genotypes (SVMNT) were most similar to those found in Papua New Guinea. Copyright © 2010, American Society for Microbiology. All Rights Reserved.

Singh O.P.,National Institute of Malaria Research NIMR | Dykes C.L.,National Institute of Malaria Research NIMR | Sharma G.,National Institute of Malaria Research NIMR | Das M.K.,National Institute of Malaria Research ICMR
Journal of Medical Entomology | Year: 2015

Leucine-to-phenylalanine substitution at residue L1014 in the voltage-gated sodium channel, target site of action for dichlorodiphenyltrichloroethane (DDT) and pyrethroids, is the most common knockdown resistance (kdr) mutation reported in several insects conferring resistance against DDT and pyrethroids. Here, we report presence of two coexisting alternative transversions, A>T and A>C, on the third codon position of L1014 residue in malaria vector Anopheles subpictus Grassi (species A) from Jamshedpur (India), both leading to the same amino acid substitution of Leu-to-Phe with allelic frequencies of 19 and 67%, respectively. A single primer-introduced restriction analysis-polymerase chain reaction (PIRA-PCR) was devised for the identification of L1014F-kdr mutation in An. subpictus resulting from either type of point mutation. Genotyping of samples with PIRA-PCR revealed high frequency (82%) of L1014F-kdr mutation in the study area. © The Authors 2015. Published by Oxford University Press on behalf of Entomological Society of America.

Prajapati S.K.,National Institute of Malaria Research NIMR | Kumari P.,National Institute of Malaria Research NIMR | Singh O.P.,National Institute of Malaria Research NIMR
BMC Microbiology | Year: 2012

Background: Plasmodium vivax reticulocyte binding protein-2 (PvRBP-2) is a promising candidate for development of vaccine against parasite. DNA sequence polymorphism in pvrbp-2 which may hamper the vaccine development program has been identified in laboratory strains. Therefore, unraveling genetic polymorphism in pvrbp-2 from field isolates is a prerequisite for success in vaccine development. This study was designed with a primary aim to uncover genetic polymorphism in pvrbp-2 among P. vivax field isolates. Results: Using virtual restriction mapping of pvrbp-2 sequences, two restriction enzymes (AluI and ApoI) were selected for the development of pvrbp-2 as a PCR-RFLP marker. Restriction fragment length polymorphism (RFLP) analysis revealed a high degree of genetic polymorphism in the pvrbp-2 gene among field isolates of P. vivax. ApoI-RFLP was found to be more efficient in identifying the extent of genetic polymorphism in pvrbp-2 compared to AluI-RFLP. Combined genotyping/haplotyping of RFLP pattern revealed a total of 36 distinct RFLP patterns among 83 P. vivax isolates analyzed. DNA sequence analysis also supports high degree of genetic polymorphism among field isolates of P. vivax. Pvrbp-2 PCR-RFLP method is able to distinguish multiple infection up to 16.86% and it revealed a low level of shared genetic pool between more than two populations. Conclusion: The study suggests that pvrbp-2 is highly polymorphic genetic marker which can be used for population genetic analyses. RFLP analysis suggests presence of nearly similar proportion of Sal-1 and Belem alleles in Indian P. vivax populations. The larger extent of genetic polymorphism identified from limited samples advocates to screen genetic polymorphism in pvrbp-2 from malaria endemic geographical regions and countries for designing pvrbp-2 based anti-malarial control measures. © 2012 Prajapati et al.; licensee BioMed Central Ltd.

Prajapati S.K.,National Institute of Malaria Research NIMR | Joshi H.,National Institute of Malaria Research NIMR | Dua V.K.,National Institute of Malaria Research NIMR
Malaria Journal | Year: 2011

Background: Genetic polymorphism is an inevitable component of a multistage infectious organism, such as the malaria parasite. By means of genetic polymorphism, parasite opts particular polymorph and reveals survival advantage. Pvs25 and pvs28 are sexual stage antigen genes, expressed at the ookinete stage inside the mosquito gut, and considered as potential transmission-blocking vaccine candidates. This study presents sequence variations in two important transmission blocking antigen genes pvs25 and pvs28 in the field isolates of P. vivax from the Indian subcontinent. Methods. One hundred microscopically diagnosed P. vivax isolates were collected from five geographical regions of India. Pvs25 and pvs28 genes were PCR amplified and sequenced to assess sequence variation among field isolates. Results: A total of 26 amino acid substitutions were observed in Pvs25 (10) and Pvs28 (16) among field isolates of P. vivax. Tandem repeat polymorphism observed in pvs28 shows 3-6 tandem repeats in the field isolates. Seven and eight novel amino acid substitutions were observed in Pvs25 and Pvs28, respectively in Indian isolates. Comparison of amino acid substitutions suggests that majority of substitutions observed in global isolates were also present in Indian subcontinent. A single haplotype was observed to be major haplotype among isolates of Delhi, Nadiad, Chennai and Panna except in isolates of Kamrup. Further, population comparison analyses suggest that P. vivax isolates inhabiting in north-eastern region (Kamrup) were distantly related with the isolates from remaining parts of the country. Majority of the amino acid substitutions observed in Indian isolates were more identical to the substitutions reported from isolates of Thailand and Bangladesh. Conclusion: Study uncovered many new amino acid substitutions as well as a predominance of single haplotype in Indian subcontinent except in north-eastern region of the country. The amino acid substitutions data generated in this study from different geographical regions of the Indian subcontinent could be helpful in designing a more effective anti-malarial transmission-blocking vaccine. © 2011 Prajapati et al; licensee BioMed Central Ltd.

Sharma S.K.,National Institute of Malaria Research NIMR | Upadhyay A.K.,National Institute of Malaria Research NIMR | Haque M.A.,National Institute of Malaria Research NIMR | Tyagi P.K.,National Institute of Malaria Research NIMR | And 2 more authors.
Acta Tropica | Year: 2010

A field trial was conducted on the efficacy of Interceptor nets-a long-lasting insecticidal net (LLN) factory treated with alphacypermethrin 0.667% (w/w) corresponding to 200mg/m2, against malaria vectors Anopheles culicifacies and Anopheles fluviatilis in one of the highly endemic areas of Orissa. The study area comprised 19 villages which were randomized into three clusters and designated as Interceptor net cluster, untreated net cluster, and no net cluster. Baseline studies showed that both the vector species An. culicifacies and An. fluviatilis were 100% susceptible to alphacypermethrin. Results of wash-resistance and bio-efficacy of Interceptor nets showed 100% mortality in An. culicifacies and An. fluviatilis even after 20 washings. Bioassays on the Interceptor nets while in use in the field conditions showed a knockdown effect on 70-90% mosquitoes during different months of intervention after 3min of exposure and 100% mortality was recorded after 24h of recovery period. The median knockdown time for these species ranged between 4.10-5.25min and 4.00-5.00min respectively during intervention period. In Interceptor net study area, there was a significant reduction of 88.9, 96.3 and 90.6% in the entry rate of An. culicifacies, An. fluviatilis and other anopheline species respectively with an over all reduction of 87.5% in total mosquitoes. The overall feeding success rate of mosquitoes in the trial villages was only 12.8% in comparison to 35.0 and 78.8% in villages with untreated nets and no nets respectively. A significant reduction was also recorded in parity rate and human blood index of vector species in Interceptor net area. The results of the study showed that Interceptor nets are effective against the malaria vectors and may be used as a suitable intervention strategy in high-risk areas. © 2010 Elsevier B.V.

Prajapati S.K.,National Institute of Malaria Research NIMR | Joshi H.,National Institute of Malaria Research NIMR | Dev V.,NIMR Field Station | Dua V.K.,National Institute of Malaria Research NIMR
Malaria Journal | Year: 2011

Background: Sulphadoxine and pyrimethamine are anti-folate drugs that show synergistic anti-malarial effect. Point mutations in dihydrofolate reductase (dhfr) and dihydropteorate synthatase (dhps) cause anti-folate drug resistance phenotype in human malaria parasites. This study presents pattern of point mutations in dhfr/dhps genes among Indian sub-continent. Methods. Microscopically diagnosed one hundred Plasmodium vivax field isolates were collected from five widely separated geographical regions of India. Dhfr and dhps genes were PCR amplified and sequenced. Previously published mutations data were collected and analyzed using Chi square test to identify geographical cluster of mutant/wild type genotypes. Results: Sequence analysis revealed single (S58R), double (S58R/S117N) and quadruple (F57L/S58R/T61M/S117T/) point mutations at dhfr and single (A383G) to double (A383G/A553G) mutations at dhps in P. vivax field isolates. In addition, three new mutations were also observed at dhfr. Both, dhfr and dhps genes revealed tandem repeat variations in field isolates. Dhps revealed very low mutation frequency (14.0%) compared to dhfr (50.70%). Comparative analysis revealed a progressive increase in frequency of quadruple mutant dhfr genotype (p < 0.001) within five years in north-eastern state (Kamrup, Assam). Frequency of dhfr genotypes revealed three distinct geographical clusters of wild (northern India), double mutant (southern India), and quadruple mutant (north-eastern and island regions of India) on the Indian sub-continent. Conclusion: Study suggests that SP may be susceptible to P. vivax in India, except Andaman and north-eastern state. The distinction of geographical regions with sensitive and resistant parasite phenotypes would be highly useful for designing and administering national anti-malarial drug policy. © 2011 Prajapati et al; licensee BioMed Central Ltd.

Gupta B.,National Institute of Malaria Research NIMR | Awasthi G.,National Institute of Malaria Research NIMR | Das A.,National Institute of Malaria Research NIMR
Parasitology Research | Year: 2010

Genome scan and genotype-phenotype association study offer excellent opportunities to unearth drug/vaccine targets in human pathogens including malaria parasites. A recently conducted such study in worldwide isolates in the most devastating malaria parasite Plasmodium falciparum has reported important genomic information on genetic basis of antimalarial resistance. Several unknown genes were also found to be under strong influence of natural selection. The findings provide important insights into the malaria parasite genome evolution in general and to use this information to develop more focused malaria control strategies, in particular. © 2010 Springer-Verlag.

Sharma S.K.,National Institute of Malaria Research NIMR | Upadhyay A.K.,National Institute of Malaria Research NIMR | Haque M.A.,National Institute of Malaria Research NIMR | Tyagi P.K.,National Institute of Malaria Research NIMR | Kindo B.K.,National Institute of Malaria Research NIMR
Indian Journal of Medical Research | Year: 2012

Background & objectives: Development of insecticide resistance in malaria vectors has been a major problem for achieving effective vector control. Due to limited availability of insecticides, the only option is management of resistance by judiciously using the insecticides and rotating them to maintain their effectiveness. This study was carried out in a malaria endemic area of Sundergarh district in Orissa where synthetic pyrethroids (SP) were in use for the last couple of years. The change-over from SP to DDT was done in one arm of study, and the other two arms remained on SP and insecticide-treated nets (ITN). Entomological and parasitological monitoring was done to assess the impact. Methods: The study design comprised of three arms (i) two rounds of indoor residual spraying (IRS) with DDT 1g/m2 as a change-over insecticide in areas previously under synthetic pyrethroids; (ii) two rounds of IRS with synthetic pyrethroid (alphacypermethrin, ACM) @ 25 mg/m2; and (iii) an unsprayed area under ITN/long lasting insecticide nets (LNs). Indoor residual spraying was undertaken under strict supervision to maintain quality and coverage. Contact bioassays were conducted to know the persistence of insecticide on sprayed surfaces and adult vector density was monitored in fixed and randomly selected houses. Malaria incidence was measured through fortnightly domiciliary surveillance under primary health care system in all the study villages. Results: The insecticide susceptibility tests showed that An.culicifacies was resistant to DDT but susceptible to malathion and ACM. However, An. fluviatilis was susceptible to all the three insecticides. ACM was effective in killing An. culicifacies on mud and wooden sprayed surfaces and maintained effective bioefficacy ranging from 92 to 100 per cent up to five months, whereas DDT failed to achieve effective mortality in An.culicifacies. However, there was significant decline in the density of An.culicifacies in ACM and DDT areas in comparison to ITNs/LNs. There was 61 per cent reduction in the slide positivity rate in ACM area in comparison to 48 and 51 per cent in DDT and ITN/LNs areas, respectively. The adjusted incidence rate of malaria cases per 1000 population in three study areas also showed significant declines within each group. Interpretation & conclusions: The present findings show that the change-over of insecticide from synthetic pyrethroids to DDT brings about the same epidemiological impact as envisaged from continuing SP spray or distributing insecticide treated nets/long-lasting insecticidal nets provided there is a good quality spray and house coverage.

PubMed | Maharshi Dayanand University and National Institute of Malaria Research NIMR
Type: Journal Article | Journal: Parasitology research | Year: 2016

The rapid spread of antimalarial drug resistance in Plasmodium falciparum over the past few decades has necessitated intensive monitoring of such resistance for an effective malaria control strategy. P. falciparum dihydropteroate synthase (Pfdhps) and P. falciparum dihydrofolate reductase (Pfdhfr) genes act as molecular markers for resistance against the antimalarial drugs sulphadoxine and pyrimethamine, respectively. Resistance to pyrimethamine which is used as a partner drug in artemisinin combination therapy (ACT) is associated with several mutations in the Pfdhfr gene, namely A16V, N51I, C59R, S108N/T and I164L. Therefore, routine monitoring of Pfdhfr-drug-resistant alleles in a population may help in effective drug resistance management. Allele-specific PCR (ASPCR) is one of the commonly used methods for molecular genotyping of these alleles. In this study, we genotyped 55 samples of P. falciparum for allele discrimination at four codons of Pfdhfr (N51, C59, S108 and I164) by ASPCR using published methods and by Sangers DNA sequencing method. We found that the ASPCR identified a significantly higher number of mutant alleles as compared to the DNA sequencing method. Such discrepancies arise due to the non-specificity of some of the allele-specific primer sets and due to the lack of sensitivity of Sangers DNA sequencing method to detect minor alleles present in multiple clone infections. This study reveals the need of a highly specific and sensitive method for genotyping and detecting minor drug-resistant alleles present in multiple clonal infections.

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