Biology subjects

Kochar, S. K.

Publications and source records attributed to Kochar, S. K..

5 recordsLinked to original sources

bioRxiv2025

Comparative clinical transcriptome of pir genes in severe Plasmodium vivax malaria

vir genes, a multigene family in Plasmodium vivax that are a part of a larger superfamily of genes called the pir (Plasmodium interspersed repeat) genes, have been reported earlier to be potentially involved in cyto-adherence and evasion of splenic clearance. Plasmodium vivax, historically characterised as a "benign" malaria parasite, has been associated with clinical outcomes including hepatic dysfunction, renal failure, and cerebral malaria in India and several global regions. It constitutes an economic burden and presents a public health challenge alongside other Plasmodium species. Here, we present a part of global transcriptomic studies using custom-designed microarrays that compare the transcriptome of the parasite responsible for severe Plasmodium vivax manifestations, specifically hepatic dysfunction and cerebral malaria from India, with an emphasis on the pir genes, some of which are reported to play a role in cyto-adherence. The RNA of the parasite isolated from 23 patients (uncomplicated group = 6, hepatic dysfunction group = 12, and cerebral malaria group = 5) was subjected to microarray hybridisation, and the data obtained showed a wide range of pir subfamilies to have been differentially expressed. We report the upregulation of 24 pir genes in the cerebral malaria group (n = 5) and 28 pir genes in the hepatic dysfunction group (n = 12), which belong to different subfamilies in at least 50% of the severe malaria patients group. Out of the upregulated pir genes in the cerebral malaria group, members of vir subfamily E (n=8 genes) and the pvpir subfamily H (n=6 genes) are expressed in a higher proportion compared to the hepatic dysfunction group, where members of vir subfamily E (n=9) and C (n=6) are expressed in a major proportion. Author SummaryPlasmodium vivax, considered to cause benign infections, has been reported over the past couple of decades as manifesting with severe symptoms like cerebral malaria, hepatic dysfunction, and others. This paper reports the analysis of a selected pir gene repertoire from microarray hybridisation performed with custom-designed 15K microarrays. Differences between pir genes, which are specifically upregulated in either hepatic dysfunction or cerebral malaria, the two manifestations considered in addition to uncomplicated vivax malaria samples, are highlighted. The latter provided the control values for comparison with the disease groups. The data presented in this study show differentially upregulated pir genes, some of which are also common to both manifestations. Literature has reported one of the VIR proteins, encoded by vir14 C gene, to be involved in adhesion to ICAM-1. The future scope of this work includes validation with a larger sample size and characterising the products of the pir gene, as depicted with reference to their role in cytoadherence or other mechanisms that could lead to severe disease. It would be possible to devise strategies based on this data that could lead to the use of some of these molecules as potential biomarkers or for therapeutic intervention.

genomics↗

bioRxiv2025

Integrative Transcriptomic and Machine Learning Approaches to decipher Mitochondrial Gene Regulation in severe Plasmodium vivax Malaria

Mitochondria in Plasmodium vivax are functionally vital despite possessing a highly reduced genome and differing substantially from the human organelle. Beyond their classical role in energy production, they dynamically coordinate processes like pyrimidine biosynthesis and heme metabolism, adapting their functions across the intra-erythrocytic development cycle (IDC). Their unique architecture and stage-specific roles enable the parasite to fine-tune mitochondrial gene expression, involving both protein-coding sense transcripts and long non-coding natural antisense transcripts (NATs). This study unveils an unprecedented regulatory complexity by integrating transcriptomic profiling with advanced machine learning to decode the role of mitochondrial sense and natural antisense transcripts (NATs) in severe P. vivax malaria. We reveal distinct, clinically relevant expression signatures, where NATs emerge not as transcriptional by-products but as potent regulators tightly linked to mitochondrial pathways and translational machinery. This dual-layered transcriptomic landscape reflects an intricate molecular strategy by which the parasite fine-tunes mitochondrial function to survive under severe disease conditions. Importantly, while these findings illuminate novel regulatory mechanisms and position mitochondrial NATs as promising targets for antimalarial drug development, they represent preliminary insights derived from a limited clinical cohort and should not be interpreted as definitive clinical indicators. Validation in larger and diverse patient populations is essential to confirm their broader biological and clinical relevance. However, these results serve as indicators for potential innovative therapeutic interventions aimed at disrupting parasite bioenergetics and regulatory networks.

genomics↗

bioRxiv2025

Differential expression of mitochondria-associated genes in clinical samples of Plasmodium falciparum showing severe manifestations

The malaria parasite mitochondrial proteins are critical targets for antimalarial drugs, however, the emergence of drug resistance against the existing protein targets necessitates novel therapeutic approaches. In this study,we profiled mitochondrial sense and natural antisense transcripts (NATs) in 22 Plasmodium falciparum clinical isolates, classified into three disease groups - uncomplicated malaria (UNC, n=6), cerebral malaria (CM, n=4), and hepatic dysfunction (HD, n=12), using a custom, strand-aware 60K microarray, and validated the transcriptome using pooled, strand-specific RNA-seq (uncomplicated pool and complicated pool). Differential gene expression in the CM and HD cohort was obtained by comparing with uncomplicated malaria (UNC) as the control cohort. The analysis revealed distinct disease-specific sense and NATs, encoded by the mitochondrial genome, along with those encoded in the nucleus and targeted to the parasite mitochondria. Although mitochondrial activity is known to be reduced in blood-stage malaria, upregulation of genes linked to tricarboxylic-acid-cycle and electron-transport in the CM cluster indicates disrupted mitochondrial bioenergetics in severe disease. Profiling of sense and antisense mitochondrial transcripts reveal a correlated expression of sense-antisense transcript pairs in both the disease manifestations, indicating a potential regulatory role of NATs in mitochondrial function. These data provide direct evidence of NATs originating from the parasite mitochondrial genome and nominate key NATs against core mitochondrial functions as potential non-conventional antimalarial targets.

genomics↗

bioRxiv2023

A Tale of Two Parasites: A Glimpse into the clinical RNA Methylome of P. falciparum and P. vivax isolates

BackgroundUnderstanding the molecular mechanisms of the malarial parasites in hosts is crucial for developing effective treatments. Epitranscriptomic research on pathogens has unveiled the significance of RNA methylation in gene regulation and pathogenesis. We present the first report investigating methylation signatures and alternative splicing events using Nanopore Direct RNA Sequencing to single-base resolution in Plasmodium falciparum and P. vivax clinical isolates with hepatic dysfunction complications. MethodologyWe performed direct RNA Sequencing using Nanopore from clinical isolates of P. falciparum and P. vivax showing hepatic dysfunction manifestation. Subsequently we performed transcriptome reconstruction using FLAIR and transcript classification using SQANTI3 followed by methylation detection using CHEUI and m6Anet to identify N6-methyladenosine (m6A) and 5-methylcytosine (m5C) methylation signatures. We also documented alternative splicing events from both the datasets. ResultsThe reference genome of Plasmodium reports >5000 genes out of which we have identified [~]50% as expressed in the two sequenced isolates, including novel isoforms and intergenic transcripts, highlighting extensive transcriptome diversity. The distinct RNA methylation profiles of m6A and m5C from the expressed transcripts were observed in sense, Natural Antisense Transcripts (NATs) and intergenic categories hinting at species-specific regulatory mechanisms. Modified transcripts originating from apicoplast and mitochondrial genomes have also been detected. These modifications are unevenly present in the annotated regions of the mRNA, potentially influencing mRNA export and translation. We have observed several splicing events, with alternative 3and 5end splicing predominating in our datasets suggesting differences in translational kinetics and possible protein characteristics in these disease conditions. ConclusionIn our data we are showing the presence of modified sense, NATs and alternatively spliced transcripts. These phenomena together suggest the presence of multiple regulatory layers which decides the post-translational proteome of the parasites in particular disease conditions. Studies like these will help to decipher the post-translational environments of malaria parasites in vivo and elucidate their inherent proteome plasticity, thus allowing the conceptualization of novel strategies for interventions.

genomics↗

bioRxiv2023

Multi-protein chimeric antigens, a novel combined approach for efficiently targeting and blocking the blood stage of Plasmodium falciparum

Plasmodium falciparum-induced malaria remains a fatal disease affecting millions of people worldwide. Mainly, the blood stage of malaria is highly pathogenic and symptomatic, rapidly damaging the host organs and occasionally leading to death. Currently, no vaccines are approved for use against the blood stage of malaria. Canonical vaccines in the past have selected the most immunodominant or essential protein to block the growth of the parasite. This strategy works efficiently for low-complexity organisms such as viruses and a few bacteria but has not shown promising results for a malaria vaccine. Plasmodium has a complex life cycle and vaccine candidates especially during blood stage are ineffective due to multiple gene families showing redundancy, immune evasion, and insufficient antibody titer. Herein, we demonstrate a novel strategy of combining multiple antigens from the blood stage of Plasmodium falciparum using only the most immunodominant peptide sequences as a way of tackling polymorphism and redundancy. We created three chimeric antigens targeting eight PfEMP1 proteins (chimeric varB) and eight merozoite surface proteins (chimeric MSP and InvP) by selecting and stitching B-cell epitopes. Our chimeric constructs show naturally circulating antibodies against individual peptides using epitope-mapping microarray as well as entire proteins in malaria-infected patients. We demonstrate that anti-varB antibodies are neutralizing in nature and significantly reduce the cytoadhesion on an organ-on-chip system with a microfluidic device mimicking physiological conditions. We have applied a Deep Learning based method to quantify the number of adhered RBCs under fluidic conditions that is used to study cytoadhesion. Furthermore, the anti-MSP and InvP antibodies show complete growth inhibition in a single cycle at a combined concentration of 0.13 mg/ml. Overall, our results show that a combination of antigenic peptides from multiple antigens can function as a next-generation vaccine and effectively block the blood stage by reducing cytoadhesion and inhibiting the parasite growth.

molecular biology↗