Project description:We analyzed bone marrow aspirates from seven patients with vivax malaria and RNAseq analysis identified genes assoviated with erythropoietic defects
Project description:Plasmodium vivax is the most geographically widespread human malaria parasite causing approximately 130-435 million infections annually. It is an economic burden in many parts of the world and poses a public health challenge along with the other Plasmodium sp. The biology of this parasite is very little understood. Emerging evidences of severe complications due to infections by this parasite provides an impetus to focus research on the same. Investigating this parasite directly from the infected patients is the most feasible way to study its biology and any pathogenic mechanisms which may exist. Gene expression studies of this parasite directly obtained from the patients has provided evidence of gene regulation resulting in varying amount of transcript levels in the different blood stages. However, the mechanisms regulating gene expression in malaria parasites are not well understood. Discovery of natural antisense transcripts (NATs) in P. falciparum has suggested that these might play an important role in regulating gene expression. We report here the genome-wide occurrence of NATs in P. vivax parasites from patients with differing clinical symptoms. A total of 1348 NATs against annotated gene loci have been detected using a custom designed strand specific microarray. Majority of NATs identified from this study shows positive correlation with the expression pattern of the sense transcript. Our data also shows condition specific expression patterns of varying S and AS transcript levels. Genes with AS transcripts enrich to various biological processes. This is the first report detailing the presence of NATs from clinical isolates of P. vivax. The data suggests differential regulation of gene expression in diverse clinical conditions and would lead to future detailed investigations of genome regulation. Plasmodium vivax isolates were collected from patients (n = 8) with differing clinical conditions.The patients exhibited symptoms categorized as un-complicated (n =1) or complicated malaria (n = 7). Criteria for determination of complicated disease were based on World Health Organization year 2010 guidelines. Microarray array based transcriptional profiling was carried out to detect prevalence of natural antisense transcripts.
Project description:High density oligonucleotide microarrays have been used on Plasmodium vivax field isolates to estimate whole genome expression. However, no microarray platform has been experimentally optimized for studying the transcriptome of field isolates. In the present study, we adopted both bioinformatics and experimental testing approaches to select best optimized probes suitable for detecting parasite transcripts from field samples and included them in designing a custom 15K P. vivax microarray. This microarray has long oligonucleotide probes (60mer) that were in-situ synthesized onto glass slides using Agilent SurePrint technology and has been developed into an 8X15K format (8 identical arrays on a single slide). Probes in this array were experimentally validated and represents 4180 P. vivax genes in sense orientation, of which 1219 genes have also probes in antisense orientation. Validation of the 15K array by using field samples (n=14) has shown 99% of parasite transcript detection from any of the samples. Correlation analysis between duplicate probes (n=85) present in the arrays showed perfect correlation (r2=0.98) indicating the reproducibility. Multiple probes representing the same gene exhibited similar kind of expression pattern across the samples (positive correlation, r≥0.6). Comparison of hybridization data with the previous studies and quantitative real-time PCR experiments were performed to highlight the microarray validation procedure. This array is unique in its design, and results indicate that the array is sensitive and reproducible. Hence, this microarray could be a valuable functional genomics tool to generate reliable expression data from P. vivax field isolates.
Project description:Plasmodium vivax is the most geographically widespread human malaria parasite causing approximately 130-435 million infections annually. It is an economic burden in many parts of the world and poses a public health challenge along with the other Plasmodium sp. The biology of this parasite is very little understood. Emerging evidences of severe complications due to infections by this parasite provides an impetus to focus research on the same. Investigating this parasite directly from the infected patients is the most feasible way to study its biology and any pathogenic mechanisms which may exist. Gene expression studies of this parasite directly obtained from the patients has provided evidence of gene regulation resulting in varying amount of transcript levels in the different blood stages. However, the mechanisms regulating gene expression in malaria parasites are not well understood. Discovery of natural antisense transcripts (NATs) in P. falciparum has suggested that these might play an important role in regulating gene expression. We report here the genome-wide occurrence of NATs in P. vivax parasites from patients with differing clinical symptoms. A total of 1348 NATs against annotated gene loci have been detected using a custom designed strand specific microarray. Majority of NATs identified from this study shows positive correlation with the expression pattern of the sense transcript. Our data also shows condition specific expression patterns of varying S and AS transcript levels. Genes with AS transcripts enrich to various biological processes. This is the first report detailing the presence of NATs from clinical isolates of P. vivax. The data suggests differential regulation of gene expression in diverse clinical conditions and would lead to future detailed investigations of genome regulation.
Project description:Unlike in Asia and Latin America, Plasmodium vivax infections were rare in Sub-Saharan Africa due to the absence of the Duffy blood group antigen (Duffy Antigen), the only known erythrocyte receptor for the P. vivax merozoite invasion ligand, Duffy Binding Protein 1 (DBP1). However, P. vivax infections have been documented in Duffy-negative individuals throughout Africa, suggesting that P. vivax may use ligands other than DBP1 to invade Duffy-negative erythrocytes through other receptors. To identify potential P. vivax ligands, we compared parasite gene expression in Saimiri and Aotus monkey erythrocytes infected with P. vivax Salvador I (Sal I). DBP1 binds Aotus but does not bind to Saimiri erythrocytes, and thus P. vivax Sal I must invade Saimiri erythrocytes independently of DBP1. Comparing RNA sequencing (RNAseq) data for late stage infections in Saimiri and Aotus erythrocytes when invasion ligands are expressed, we identified genes that belong to tryptophan-rich antigen and MSP3 families that were more abundantly expressed in Saimiri infections as compared to Aotus infections. These genes may encode potential ligands responsible for P. vivax infections of Duffy-negative Africans.
Project description:Transcription profile of the Plasmodium vivax intraerythrocytic cycle Total RNA in Plasmodium vivax strain at every 6 hour of intraerythrocytic cycle using RNA-seq