BackgroundBoth Plasmodium vivax and Plasmodium falciparum are prevalent in Pakistan, yet up-to-date data on the epidemiology of malaria in Pakistan are not available. This study was undertaken to determine the current prevalence and distribution of Plasmodium species across the country.MethodsA malariometric population survey was conducted in 2011 using blood samples collected from 801 febrile patients of all ages in four provinces and the capital city of Islamabad. Microscopically confirmed Plasmodium-positive blood samples were reconfirmed by polymerase chain reaction (PCR). Confirmed parasite-positive samples were subjected to species-specific PCR capable of detecting four species of human malaria.ResultsOf the 707 PCR-positive samples, 128 (18%) were P. falciparum, 536 (76%) were P. vivax, and 43 (6%) were mixed P. falciparum and P. vivax. Ninety-four microscopy-positive samples were PCR-negative, and Plasmodium malariae and Plasmodium ovale were not detected. Prevalence of P. vivax ranged from 2.4% in Punjab Province to 10.8% in Sindh Province and prevalence of P. falciparum ranged from 0.1% in Islamabad to 3.8% in Balochistan.ConclusionsPlasmodium infections in Pakistan are largely attributed to P. vivax but P. falciparum and mixed species infections are also prevalent. In addition, regional variation in the prevalence and species composition of malaria is high.
BackgroundMalaria is the fifth leading cause of death worldwide. Pakistan is considered as a moderate malaria-endemic country but still, 177 million individuals are at risk of malaria. Roughly 60% of Pakistan’s population, live in malaria-endemic regions. The present study is based upon the survey of various health care centers in 10 major cities of Northern and Southern Punjab to find out the malarial infection patterns in 2015. The diagnosis, seasonal variations, age and gender-wise distribution of Plasmodium spp. circulating in the study area were also included in the objectives.MethodsThe malaria-suspected patients ‘16075’ were enrolled for malaria diagnosis using microscopy, out of which 925 were malaria positive which were processed for molecular analysis using nested PCR. The 18S rRNA genes of Plasmodium species were amplified, sequenced, blast and the phylogenetic tree was constructed based on sequences using online integrated tool MEGA7.ResultsThe 364 cases recruited from Northern Punjab with the highest incidence in Rawalpindi (25.5%) and lowest in Chakwal (15.9%). From Southern Punjab 561 cases were enlisted Rajanpur (21.4%) maximum and lowest from Multan and Rahim Yar Khan (18%). The slide positivity rate, annual parasite incidence, and annual blood examination rates were 5.7 per 1000 population, 0.1, and 0.2% respectively. The only P. vivax (66.7%), P. falciparum (23.7%) and mixed infection by these two species (9.6%) were diagnosed. The same trend (P. vivax > P. falciparum > mixed infection) in species identification %age was confirmed from molecular analysis. However, the occurrence of malaria was higher in Southern Punjab (5.5%) as compared to the Northern Punjab (4.0%). The overall malaria percentage occurrence of treatment-seeking patients in all recruited cities of Punjab was 4.9%. The age-group of 1–20 and males among genders were more affected by malaria. The comparison of different seasons showed that the malaria infection was at a peak in Summer and post-monsoon.ConclusionThe incidence of malaria was high in the flood infected rural areas of Southern Punjab, Summer, and post-monsoon. The age group (1–20) and gender-wise males were more affected by malaria.
BackgroundPlasmodium vivax is the most prevalent malaria species in Pakistan, with a distribution that coincides with Plasmodium falciparum in many parts of the country. Both species are likely exposed to drug pressure from a number of anti-malarials including chloroquine, sulphadoxine-pyrimethamine (SP), and artemisinin combination therapy, yet little is known regarding the effects of drug pressure on parasite genes associated with drug resistance. The aims of this study were to determine the prevalence of polymorphisms in the SP resistance-associated genes pvdhfr, pvdhps and chloroquine resistance-associated gene pvmdr1 in P. vivax isolates collected from across the country.MethodsIn 2011, 801 microscopically confirmed malaria-parasite positive filter paper blood samples were collected at 14 sites representing four provinces and the capital city of Islamabad. Species-specific polymerase chain reaction (PCR) was used to identify human Plasmodium species infection. PCR-positive P. vivax isolates were subjected to sequencing of pvdhfr, pvdhps and pvmdr1 and to real-time PCR analysis to assess pvmdr1 copy number variation.ResultsOf the 801 samples, 536 were determined to be P. vivax, 128 were P. falciparum, 43 were mixed vivax/falciparum infections and 94 were PCR-negative for Plasmodium infection. Of PCR-positive P. vivax samples, 372 were selected for sequence analysis. Seventy-six of the isolates (23%) were double mutant at positions S58R and S117N in pvdhfr. Additionally, two mutations at positions N50I and S93H were observed in 55 (15%) and 24 (7%) of samples, respectively. Three 18 base pair insertion-deletions (indels) were observed in pvdhfr, with two insertions at different nucleotide positions in 36 isolates and deletions in 10. Ninety-two percent of samples contained the pvdhps (S382/A383G/K512/A553/V585) SAKAV wild type haplotype. For pvmdr1, all isolates were wild type at position Y976F and 335 (98%) carried the mutation at codon F1076L. All isolates harboured single copies of the pvmdr1 gene.ConclusionsThe prevalence of mutations associated with SP resistance in P. vivax is low in Pakistan. The high prevalence of P. vivax mutant pvmdr1 codon F1076L indicates that efficacy of chloroquine plus primaquine could be in danger of being compromised, but further studies are required to assess the clinical relevance of this observation. These findings will serve as a baseline for further monitoring of drug-resistant P. vivax malaria in Pakistan.
BackgroundFew studies have been conducted in Pakistan to determine the efficacy of chloroquine and sulphadoxine-pyrimethamine (SP), which remain in use as treatment for Plasmodium vivax and in combination with artesunate to treat Plasmodium falciparum, respectively. In this study, samples from several sites across Pakistan were characterized to determine prevalence of molecular resistance markers in the P. falciparum chloroquine resistance transporter (pfcrt), multidrug resistance (pfmdr1), dihydrofolate reductase (pfdhfr) and dihydropteroate synthase (pfdhps) genes and the origin of chloroquine-resistant P. falciparum parasites.MethodsMicroscopy-confirmed malaria parasite-positive blood samples from 801 patients across the country were collected in 2011. Of these, 171 infections were identified by polymerase chain reaction (PCR) as P. falciparum and analysed by pyrosequencing for mutations conferring chloroquine resistance (pfcrt codons 72–76), multidrug resistance (pfmdr1 N86Y, Y184F, S1034C, N1042D and D1246Y), pyrimethamine resistance (pfdhfr, C50R, N51I, C59R, S108N and I164L) and sulphadoxine resistance (pfdhps, S436A, A437G, K540E, A581G and A613T/S). pfmdr1 gene copy number variation was determined by real-time PCR, and microsatellites flanking the pfcrt locus were typed to determine the origin of the chloroquine-resistant haplotype.ResultsThe pfcrt K76T mutation was found in all samples as part of the S72/V73/M74/N75/T76 (SVMNT) haplotype. Microsatellites flanking pfcrt showed high similarity to the signature found in India and Papua New Guinea. pfmdr1 N86Y was found in 20% of samples and all samples harboured a single copy of the pfmdr1 gene. The pfdhfr double mutation C59R + S108N was present in 87% of samples while the pfdhfr triple mutant (N51I + C59R + S108N) was not detected. Pfdhps A437G was found in 60% of samples. Pure pfdhps K540E was rare, at 4%, but mixed genotype 540 K/E was found in 77% of samples. Similarly, pure pfdhps A581G was found in 4% of the isolates while mixed 581A/G was found in 39% of samples.ConclusionsThese results suggest an emerging problem with multidrug resistant P. falciparum in Pakistan. The chloroquine resistance genotype has reached complete fixation in the population, with a microsatellite pattern indicative of a selective sweep. Moreover, the prevalence of mutations in both pfdhfr and pfdhps, albeit without the presence of the pfdhfr triple mutant, indicates that continued monitoring is warranted to assess whether SP remains efficacious as a partner drug for artesunate for the treatment of P. falciparum.
Chikungunya virus (CHIKV) is one of the emerging viruses around the globe. It belongs to the family Togaviridae and genus Alphavirus and is an arthropod borne virus that transmits by the bite of an infected mosquito, mainly through Aedes aegypti and Aedes albopcitus. It is a spherical, enveloped virus with positive single stranded RNA genome. It was first discovered during 1952-53 in Tanganyika, after which outbreaks were documented in many regions of the world. CHIKV has two transmission cycles; an enzootic sylvatic cycle and an urban cycle. CHIKV genome contains 11,900 nucleotides and two open reading frames and shows great sequence variability. Molecular mechanisms of virus host-cell interactions and the pathogenesis of disease are not fully understood. The disease involves three phases; acute, post-acute and chronic with symptoms including high-grade fever, arthralgia, macupapular rashes and headache. There is no licensed vaccine or specific treatment for CHIKV infection. This lack of specific interventions combined with difficulties in making a precise diagnosis together make the disease difficult to manage.In this review we aim to present the current knowledge of global epidemiology, transmission, structure, various aspects of diagnosis as well as highlight potential antiviral drugs and vaccines against CHIKV.
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