Impact of IRS: Four-years of entomological surveillance of the Indian Visceral Leishmaniases elimination programme

BACKGROUND In 2005, Bangladesh, India and Nepal agreed to eliminate visceral leishmaniasis (VL) as a public health problem. The approach to this was through improved case detection and treatment, and controlling transmission by the sand fly vector Phlebotomus argentipes, with indoor residual spraying (IRS) of insecticide. Initially, India applied DDT with stirrup pumps for IRS, however, this did not reduce transmission. After 2015 onwards, the pyrethroid alpha-cypermethrin was applied with compression pumps, and entomological surveillance was initiated in 2016. METHODS Eight sentinel sites were established in the Indian states of Bihar, Jharkhand and West Bengal. IRS coverage was monitored by household survey, quality of insecticide application was measured by HPLC, presence and abundance of the VL vector was monitored by CDC light traps, insecticide resistance was measured with WHO diagnostic assays and case incidence was determined from the VL case register KAMIS. RESULTS Complete treatment of houses with IRS increased across all sites from 57% in 2016 to 70% of houses in 2019, rising to >80% if partial house IRS coverage is included (except West Bengal). The quality of insecticide application has improved compared to previous studies, average doses of insecticide on filters papers ranged from 1.52 times the target dose of 25mg/m2 alpha-cypermethrin in 2019 to 1.67 times in 2018. Resistance to DDT has continued to increase, but the vector was not resistant to carbamates, organophosphates or pyrethroids. The annual and seasonal abundance of P. argentipes declined between 2016 to 2019 with an overall infection rate of 0.03%. This was associated with a decline in VL incidence for the blocks represented by the sentinel sites from 1.16 per 10,000 population in 2016 to 0.51 per 10,000 in 2019. CONCLUSION Through effective case detection and management reducing the infection reservoirs for P. argentipes in the human population combined with IRS keeping P. argentipes abundance and infectivity low has reduced VL transmission. This combination of effective case management and vector control has now brought India within reach of the VL elimination targets

Alternative splicing is a major mechanism of gene regulation in diffuse large B cell lymphoma

Open Access JournalOral Session - Non-Hodgkin Lymphoma - Biology, excluding Therapy: Advanced Technology in Non-Hodgkin Lymphoma GeneticsThis journal issue is proceedings of ASH Conference 2010Diffuse large B-cell lymphomas (DLBCL) is the most common form of lymphoma and these tumors demonstrate a striking molecular and clinical heterogeneity. Gene expression profiling has shown that these tumors can be divided into at least two groups, activated B cell-like (ABC) and germinal B cell-like (GCB), as well as a number of other cellular processes underlying survival and tumor biology. However, the molecular mechanisms which lead to the differential gene expression patterns are not well understood. Alternative splicing is a process through which individual exons that comprise genes that are assembled into different gene isoforms with potentially different function. Alternative splicing has been shown to be a ubiquitous mechanism of gene regulation in eukaryotes and a number of cancers. The role of alternative splicing in DLBCL is unknown. We hypothesized that alternative splicing might play an important role in DLBCL. We measured genome-wide expression of over 1 million exons in 106 primary DLBCL tumors using Affymetrix Exon 1.0 ST microarrays. The same cases were also profiled for gene expression using a conventional Affymetrix Gene microarray for comparison, and further sub-classified as the molecular subgroups of DLBCL and for independent assessment of gene expression associated with known biological processes. We identified those genes as alternatively spliced which had at least one exon that was significantly different (P<0.01) compared with the comparison group. Through examination of exon-level expression data, we found evidence for splicing events in over 10,000 genes that affect at least 10% of DLBCL cases. We identified over 200 genes that have differential exon usage between ABC and GCB DLBCL. The expression of selected alternatively spliced exons was confirmed by real-time PCR. We further examined the occurrence of alternative splicing in a number of cellular processes including each of the survival associated gene expression signatures. We found that alternative splicing regulates a significant number of genes underlying the survival-associated proliferation, stromal response and germinal center differentiation gene expression signatures (P<10–6 in all cases). In addition, a number of processes that are known to be important in oncogenesis appeared to be highly regulated by alternative splicing including transcription factor activity, DNA-repair and apoptosis (P<0.001). These data confirm that alternative splicing plays a significant role in regulating genes that are important mediators of DLBCL biology. We further investigated whether lineage-specific effects were responsible for some of the observed differences in splicing in the molecular subgroups of DLBCLs. We obtained normal resting B cells from healthy donors and stimulated them with IgM and CD40-ligand to generate activated B cells. We found that the gene-isoforms expressed highly in activated normal B cells were significantly enriched in ABC-DLBCLS (P<0.01), suggesting that lineage-derived differences in isoform splicing are preserved in their malignant counterparts. Our data indicate that alternative splicing provides an additional and significant component of regulation that encompasses nearly every biological process that is known to be important in DLBCL. Future studies that examine the role gene expression will need to recognize the specific isoforms that result in proteins with altered function.link_to_OA_fulltex

A comprehensive identification of the microrna transcriptome and its application in B cell malignancies

Poster Board 2-380: abstract 2403BACKGROUND: MicroRNAs are 18-22 nucleotide-long RNA molecules that regulate expression of genes. We and others have previously demonstrated a role for microRNAs in the pathogenesis of B cell malignancies. Computational predictions suggest that the human genome encodes several thousand microRNAs. Thus far, about 700 microRNAs have been discovered in humans, including over 200 new microRNAs in the past year alone. The ongoing discovery of microRNAs makes it difficult to comprehensively study their role in a disease group. The advent of high throughput sequencing allows the simultaneous identification of millions of transcripts, thereby providing a sensitivity that is several orders of magnitude higher than conventional methods. We hypothesized that high throughput sequencing would be an effective tool to comprehensively identify microRNAs in normal and malignant B cells …link_to_OA_fulltex

ID3 is a novel tumor suppressor gene in Burkitt lymphoma

Open Access JournalOral Sessions - 622. Non-Hodgkin Lymphoma - Biology, excluding Therapy: Highly Aggressive LymphomasThis journal issue is proceedings of ASH Conference 2012Burkitt Lymphoma (BL) is a highly proliferative form of non-Hodgkin lymphoma and is characterized by translocation of the C-MYC gene to the immunoglobulin gene loci resulting in deregulation. The role of collaborating gene mutations in BL is largely unknown. We performed whole exome sequencing and gene expression profiling of 57 Burkitt lymphoma and 94 DLBCL exomes. Mutational analysis revealed that ID3 is recurrently mutated in 38% of Burkitt lymphoma samples. ID3 mutations did not occur in any of the 94 DLBCL cases. ID3 gene expression was also found to be a distinguishing feature of Burkitt lymphomas (P<10–6), compared to DLBCL. We found a total of 27 distinct mutations in the ID3 genes among the 22 BL cases. These included five frameshift, four nonsense, and 18 missense mutations. We validated 16 of these events with Sanger sequencing with over 90% concordance. All of these mutations were located in the highly conserved helix-loop-helix region located on Exon 1. We explored the biological significance of ID3 mutations by initially comparing the gene expression profiles of BL cases that had mutated and wild-type ID3. Gene set enrichment analysis showed that those samples with mutated ID3 had higher expression of genes that were involved in cell cycle regulation, specifically those involved in the G1-S transition (P=0.01). In order to experimentally investigate the functional consequences of ID3 mutation, we generated mutant constructs corresponding to six different ID3 mutations observed in BLs. These mutant constructs were cloned into lentiviral vectors and overexpressed in BL cells that were wild type for ID3. We then performed cell cycle analysis for these wild type cells expressing GFP controls or the mutant constructs. We found that BL cells expressing each of the six mutant constructs demonstrated significant cell cycle progression from G1 to S phase compared to wild-type (P=0.01). Separately, we tested the effects of expressing mutant ID3 in cell proliferation assays and found that cells expressing mutant ID3 were considerably more proliferative than those expressing wild type (P=0.03). Conversely, we over-expressed the wild type form of ID3 in BL cells that had mutated ID3. These experiments completely rescued the observed phenotypes of the mutant ID3 constructs, with reduced cell cycle progression through increased G1 phase and decreased S-phase (P=0.04). We also noted decreased cell proliferation in these cells (P=0.03). These experiments support a role for ID3 as a novel tumor suppressor gene in Burkitt lymphoma. ID3 is a basic helix loop helix (bHLH) protein that binds to other E-proteins, blocking their ability to bind DNA. ID3 has been shown to be involved in a variety of biological processes including development and T and B cell differentiation. ID3 knockout mice have been shown to develop T cell as well as B cell lymphomas. Our data implicates this gene for the first time as a tumor suppressor in human cancer.link_to_OA_fulltex