Biology subjects

Jain, R.

Publications and source records attributed to Jain, R..

At least 73 records · Page 4Linked to original sources

bioRxiv2022

Nanoparticle-delivered TLR4 and RIG-I agonists enhance immune response to SARS-CoV-2 subunit vaccine

Despite recent success in vaccinating populations against SARS-CoV-2, concerns about immunity duration, continued efficacy against emerging variants, protection from infection and transmission, and worldwide vaccine availability, remain. Although mRNA, pDNA, and viral-vector based vaccines are being administered, no protein subunit-based SARS-CoV-2 vaccine is approved. Molecular adjuvants targeting pathogen-recognition receptors (PRRs) on antigen-presenting cells (APCs) could improve and broaden the efficacy and durability of vaccine responses. Native SARS-CoV-2 infection stimulate various PRRs, including toll-like receptors (TLRs) and retinoic-acid-inducible gene I-like receptors (RIG-I). We hypothesized that targeting the same PRRs using adjuvants on nanoparticles along with a stabilized spike (S) protein antigen could provide broad and efficient immune responses. Formulations targeting TLR4 (MPLA), TLR7/8 (R848), TLR9 (CpG), and RIG-I (PUUC) delivered on degradable polymer-nanoparticles (NPs) were combined with the S1 subunit of S protein and assessed in vitro with isogeneic mixed lymphocyte reactions (iso-MLRs). For in vivo studies, the adjuvanted nanoparticles were combined with stabilized S protein and assessed using intranasal and intramuscular prime-boost vaccination models in mice. Combination NP-adjuvants targeting both TLR and RIG-I (MPLA+PUUC, CpG+PUUC, or R848+PUUC) differentially increased proinflammatory cytokine secretion (IL-1{beta}, IL-12p70, IL-27, IFN-{beta}) by APCs cultured in vitro, and induced differential T cell proliferation. When delivered intranasally, MPLA+PUUC NPs enhanced local CD4+CD44+ activated memory T cell responses while MPLA NPs increased anti-S-protein-specific IgG and IgA in the lung. Following intramuscular delivery, PUUC-carrying NPs induced strong humoral immune responses, characterized by increases in anti-S-protein IgG and neutralizing antibody titers and germinal center B cell populations (GL7+ and BCL6+ B cells). MPLA+PUUC NPs further boosted S-protein-neutralizing antibody titers and T follicular helper cell populations in draining lymph nodes. These results suggest that SARS-CoV-2-mimicking adjuvants and subunit vaccines could lead to robust and unique route-specific adaptive immune responses and may provide additional tools against the pandemic. GRAPHICAL ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=58 SRC="FIGDIR/small/478507v1_ufig1.gif" ALT="Figure 1"> View larger version (18K): org.highwire.dtl.DTLVardef@7b8174org.highwire.dtl.DTLVardef@701721org.highwire.dtl.DTLVardef@feb860org.highwire.dtl.DTLVardef@fae094_HPS_FORMAT_FIGEXP M_FIG C_FIG

immunology↗

bioRxiv2022

Pathogenicity of SARS-CoV-2 Omicron in Syrian hamsters and its neutralization with different Variants of Concern

SARS-CoV-2 Omicron variant is rampantly spreading across the globe. Animal models are useful in understanding the disease characteristics as well as properties of emerging SARS-CoV-2 variants. We assessed the pathogenicity and immune response generated by BA.1 sub-lineage of SARS-CoV-2 Omicron variant with R346K mutation in 5 to 6-week old Syrian hamsters. Virus shedding, organ viral load, lung disease and immune response generated were sequentially assessed. The disease characteristics of Omicron were found to be similar to that of other SARS-CoV-2 variants of concerns in hamsters like high viral replication in the respiratory tract and interstitial pneumonia. The infected hamsters demonstrated lesser body weight gain in comparison to the uninfected control hamsters. Viral RNA could be detected in nasal washes and respiratory organs (nasal turbinate, trachea, bronchi and lungs) till 10 and 14 days respectively. The clearance of the virus was observed from nasal washes and lungs by day 7. Neutralizing antibody response against Omicron variant was detected from day 5 with rising antibody titers till 14 days. However, the cross-neutralization titre of the sera against other variants showed severe reduction ie., 7 fold reduction against Alpha and no titers against B.1, Beta and Delta. This preliminary data shows that Omicron variant infection can produce moderate to severe lung disease and the neutralizing antibodies produced in response to Omicron variant infection shows poor neutralizing ability against other co-circulating SARS-CoV-2 variants like Delta which necessitates caution as it may lead to increased cases of reinfection.

microbiology↗

bioRxiv2021

Protective immunity of the primary SARS-CoV-2 infection reduces disease severity post re-infection with Delta variants in Syrian hamsters

Delta variant has evolved to become dominant SARS-CoV-2 lineage worldwide and there are reports of secondary infections with varying severity in vaccinated and unvaccinated naturally recovered COVID-19 patients. As the protective immunity following the infection wanes within few months, studies of re-infection after prolonged duration is needed. Hence we assessed the potential of re-infection by Delta, Delta AY.1 and B.1 in COVID-19 recovered hamsters after 3 months of infection. Re-infection with Delta and B.1 variants in hamsters showed reduced viral shedding, lung pathology and lung viral load, whereas the upper respiratory tract viral load remained similar to that of first infection. The reduction in viral load and lung pathology after re-infection with Delta AY.1 variant was not marked. Further we assessed the disease characteristics of Delta AY.1 to understand whether it has any replication advantage over Delta variant and B.1 variant, an early isolate in Syrian hamsters. Body weight changes, viral load in respiratory organs, lung pathology, cytokine response and neutralizing antibody response were assessed. Delta AY.1 variant produced milder disease in comparison to Delta variant and the neutralizing response was similar against Delta, B.1 and B.1.351 variant in contrast to Delta or B.1 infected hamsters which showed a significant reduction in neutralization titres against B.1.351. Elevation of IL-6 levels was observed post infection in hamsters after primary infection. The prior infection could not produce sterilizing immunity but the protective effect was evident following reinfection. This indicates the importance of the transmission prevention efforts even after achieving herd immunity. Research in contextO_ST_ABSEvidence before this studyC_ST_ABSSecondary infections with Delta variant are being widely reported and there are reports of increased disease severity. Delta sub lineages with K417N substitution has caused concern worldwide due to the presence of the same substitution in Beta variant, a Variant of Concern known for its immune evasion. The information on the biological characteristics of this sub lineage is also scanty. Added value of this studyThe present study showed that the secondary infection with Delta variant does not show any evidence of increased disease severity in hamster model. Delta AY. 1 variant produces mild disease in Syrian hamsters in contrast to severe disease caused by Delta variant. Delta, B.1 and AY.1 variant infected hamster sera showed comparable cross neutralizing response against each other. In contrast to the lower neutralizing response shown by B.1 and Delta variant infected animals against B.1.351 variant, Delta AY.1 showed comparable response as that with other variants. Implications of the available evidenceSARS-CoV-2 infections do not produce sterilizing immunity but protect from developing severe disease in case of Delta variant re-infection indicating the importance of the transmission prevention efforts even after achieving herd immunity. Delta AY. 1 infection in hamsters did not show any evidence of speculated immune evasion.

molecular biology↗

bioRxiv2021

Systematic assessment of lipid profiles for the discovery of tissue contributors to the circulating lipid pool in cold exposure

Plasma lipid levels are altered in chronic conditions such as type 2 diabetes and cardiovascular disease as well as acute stresses such as fasting and cold exposure. Advances in mass spectrometry based lipidomics have uncovered the complexity of the plasma lipidome which includes over 500 lipids that serve functional roles including energy substrate and signaling molecule. The plasma lipid pool is maintained through regulation of tissue production, secretion, and uptake. A major challenge is establishing the tissues of origin and uptake for various plasma lipids, which is necessary to determine the lipid function. Using cold exposure as an acute stress, we performed global lipidomics on the plasma and nine tissues that may contribute to the circulating pool. We found that numerous species of plasma acylcarnitines (ACars) and ceramides were significantly changed with cold exposure. Through computational assessment, we identified the liver and brown adipose tissue (BAT) as major contributors and consumers of circulating ACars, in agreement with our previous work. We further identified the kidney and intestine as novel contributors to the circulating ACar pool and validated these findings with gene expression analysis. Regression analysis also identified that the BAT and kidney as regulators of the plasma ceramide pool. These studies provide an adaptable computational tool to assess tissue contribution to the plasma lipid pool. Our findings have implications in understanding the function of plasma ACars and ceramides, which are elevated in metabolic diseases. SummaryThere are over 500 identified lipids in circulating plasma, many without known origin or function. Using untargeted lipidomics on plasma and nine other tissues of cold exposed mice, we identified novel regulation of circulating acylcarnitines through the kidney and intestine, and a multiorgan system that regulates plasma ceramides. Our findings offer new targets for the study and functional characterization of circulating lipids in acute cold exposure and a computational resource for other investigators to explore multi-tissue lipidome remodeling during cold exposure. O_FIG_DISPLAY_L [Figure 1] M_FIG_DISPLAY C_FIG_DISPLAY HighlightsO_LIGlobal lipidomics atlas of 9 tissues and plasma demonstrate dynamic shift with cold exposure. C_LIO_LIAdaptive resource for the selection of extraction method, data processing, and data analysis of multi-tissue global lipidomics data. C_LIO_LIRegression analysis identified the liver, BAT, intestine, and kidney as regulators of the plasma acylcarnitine pool that are not apparent by lipid levels alone. C_LIO_LIAcute cold exposure increases plasma ceramide levels, with the BAT and kidney as major contributors C_LI

biochemistry↗

bioRxiv2021

ICOR: Improving codon optimization with recurrent neural networks

BackgroundIn protein sequences--as there are 61 sense codons but only 20 standard amino acids-- most amino acids are encoded by more than one codon. Although such synonymous codons do not alter the encoded amino acid sequence, their selection can dramatically affect the expression of the resulting protein. Codon optimization of synthetic DNA sequences is important for heterologous expression. However, existing solutions are primarily based on choosing high-frequency codons only, neglecting the important effects of rare codons. In this paper, we propose a novel recurrent-neural-network based codon optimization tool, ICOR, that aims to learn codon usage bias on a genomic dataset of Escherichia coli. We compile a dataset of over 7,000 non-redundant, high-expression, robust genes which are used for deep learning. The model uses a bidirectional long short-term memory-based architecture, allowing for the sequential context of codon usage in genes to be learned. Our tool can predict synonymous codons for synthetic genes toward optimal expression in Escherichia coli. ResultsWe demonstrate that sequential context achieved via RNN may yield codon selection that is more similar to the host genome. Based on computational metrics that predict protein expression, ICOR theoretically optimizes protein expression more than frequency-based approaches. ICOR is evaluated on 1,481 Escherichia coli genes as well as a benchmark set of 40 select DNA sequences whose heterologous expression has been previously characterized. ICORs performance is measured across five metrics: the Codon Adaptation Index, GC-content, negative repeat elements, negative cis-regulatory elements, and codon frequency distribution. ConclusionsThe results, based on in silico metrics, indicate that ICOR codon optimization is theoretically more effective in enhancing recombinant expression of proteins over other established codon optimization techniques. Our tool is provided as an open-source software package that includes the benchmark set of sequences used in this study.

synthetic biology↗

bioRxiv2021

Protein complex prediction with AlphaFold-Multimer

While the vast majority of well-structured single protein chains can now be predicted to high accuracy due to the recent AlphaFold [1] model, the prediction of multi-chain protein complexes remains a challenge in many cases. In this work, we demonstrate that an AlphaFold model trained specifically for multimeric inputs of known stoichiometry, which we call AlphaFold-Multimer, significantly increases accuracy of predicted multimeric interfaces over input-adapted single-chain AlphaFold while maintaining high intra-chain accuracy. On a benchmark dataset of 17 heterodimer proteins without templates (introduced in [2]) we achieve at least medium accuracy (DockQ [3] [≥] 0.49) on 13 targets and high accuracy (DockQ [≥] 0.8) on 7 targets, compared to 9 targets of at least medium accuracy and 4 of high accuracy for the previous state of the art system (an AlphaFold-based system from [2]). We also predict structures for a large dataset of 4,446 recent protein complexes, from which we score all non-redundant interfaces with low template identity. For heteromeric interfaces we successfully predict the interface (DockQ [≥] 0.23) in 70% of cases, and produce high accuracy predictions (DockQ [≥] 0.8) in 26% of cases, an improvement of +27 and +14 percentage points over the flexible linker modification of AlphaFold [4] respectively. For homomeric inter-faces we successfully predict the interface in 72% of cases, and produce high accuracy predictions in 36% of cases, an improvement of +8 and +7 percentage points respectively.

bioinformatics↗

bioRxiv2021

Cell type determination for cardiac differentiation occurs soon after seeding of human induced pluripotent stem cells

Cardiac directed differentiation of human induced pluripotent stem cells consistently produces a mixed population of cardiomyocytes and non-cardiac cell types even when using very well-characterized protocols. We wondered whether differentiated cell types might result from intrinsic differences in hiPS cells prior to the onset of differentiation. By associating individual differentiated cells that share a common hiPS cell precursor, we were able to test whether expression variability in differentiated cells was pre-determined from the hiPS cell state. Although within a single experiment, differentiated cells that share an hiPS cell progenitor were more transcriptionally similar to each other than to other cells in the differentiated population, when the same hiPS cells were differentiated in parallel, we did not observe high transcriptional similarity across differentiations. Additionally, we found that substantial cell death occurred during differentiation in a manner that suggested that all cells were equally likely to survive or die, suggesting that there was no intrinsic selection bias for cells descended from particular hiPS cell progenitors. These results led us to wonder about how cells grow out spatially during the directed differentiation process. Labeling cells by their expression of a few canonical cell type marker genes, we showed that cells expressing the same marker tended to occur in patches observable by visual inspection, suggesting that cell type determination across multiple cell types, once initiated, is maintained in a cell-autonomous manner for multiple divisions. Altogether, our results show that while there is substantial heterogeneity in the initial hiPS cell population, that heterogeneity is not responsible for heterogeneous outcomes, and that the window during which cell type specification occurs is likely to begin shortly after the seeding of hiPS cells for differentiation.

genomics↗

bioRxiv2021

A distinct CD115- erythro-myeloid precursor present at the maternal-embryonic interface and in the bone marrow of adult mice

During ontogeny, macrophages develop from CD115+ precursors, including erythro-myeloid progenitors (EMP). EMP arise in the embryonic yolk sac, the primary site of early haematopoiesis. In adults, CD115+ bone marrow-derived monocytes represent essential macrophage precursors. Herein, we identify a CD115- macrophage precursor within the adult bone marrow that is unrelated to the classical monocyte lineage but rather shares transcriptomic and functional characteristics of embryonic EMP. These EMPROR (for Erythro Myeloid Precursor) cells are capable of efficiently generating macrophages in disease settings. During early development, EMPROR cells were largely absent from the yolk sac but were instead found at the embryonic-maternal interface in the uterine wall. Unexpectedly, the latter site contains robust haematopoietic activity and harbours defined embryonic haematopoietic progenitor cells, including classical CD115+ EMP. Our data suggest the existence of an alternative pathway of macrophage generation in the adult. Further, we uncover a hitherto unknown site of earliest blood cell development.

immunology↗

bioRxiv2021

Middle Eastern Genetic Variation Improves Clinical Annotation of the Human Genome

Genetic variation in populations of Middle Eastern origin remains highly underrepresented in most comprehensive genomic databases. This underrepresentation hampers the functional annotation of the human genome and challenges accurate clinical variant interpretation. To highlight the importance of capturing genetic variation in the Middle East, we aggregated whole exome and genome sequencing data from 2116 individuals in the Middle East and established the Middle East Variation (MEV) database. Of the high-impact coding (missense and loss of function) variants in this database, 53% were absent from the most comprehensive Genome Aggregation Database (gnomAD), thus representing a unique Middle Eastern variation dataset which might directly impact clinical variant interpretation. We highlight 39 variants with minor allele frequency >1% in the MEV database that were previously reported as rare disease variants in ClinVar and the Human Gene Mutation Database (HGMD). Furthermore, the MEV database consisted of 281 putative homozygous loss of function (LoF) variants, or complete knockouts, of which 31.7% (89/281) were absent from gnomAD. This set represents either complete knockouts of 83 unique genes in reportedly healthy individuals, with implications regarding disease penetrance and expressivity, or might affect dispensable exons, thus refining the clinical annotation of those regions. Intriguingly, 24 of those genes have several clinically significant variants reported in ClinVar and/or HGMD. Our study shows that genetic variation in the Middle East improves functional annotation and clinical interpretation of the genome and emphasizes the need for expanding sequencing studies in the Middle East and other underrepresented populations.

genomics↗

bioRxiv2021

A Tad-like apparatus is required for contact-dependent prey killing in predatory social bacteria

Myxococcus xanthus, a soil bacterium, predates collectively using motility to invade prey colonies. Prey lysis is mostly thought to rely on secreted factors, cocktails of antibiotics and enzymes, and direct contac with Myxococcus cells. In this study, we show that on surfaces the coupling of A-motility and contact-dependent killing is the central predatory mechanism driving effective prey colony invasion and consumption. At the molecular level, contact-dependent killing involves a newly discovered type IV filament-like machinery (Kil) that both promotes motility arrest and prey cell plasmolysis. In this process, Kil proteins assemble at the predator-prey contact site, suggesting that they allow tight contact with prey cells for their intoxication. Kil-like systems form a new class of Tad-like machineries in predatory bacteria, suggesting a conserved function in predator-prey interactions. This study further reveals a novel cell-cell interaction function for bacterial pili-like assemblages.

microbiology↗

bioRxiv2021

Comparison of the pathogenicity and virus shedding of SARS CoV-2 VOC 202012/01 and D614G variant in hamster model

The emergence of SARS-CoV-2 variants has posed a serious challenge to public health system and vaccination programs across the globe. We have studied the pathogenicity and virus shedding pattern of the SARS-CoV-2 VOC 202012/01 and compared with D614G variant in Syrian hamsters. VOC 202012/01 could produce disease in hamsters characterized by body weight loss and respiratory tract tropism but mild lung pathology. Further, we also documented that neutralizing antibodies developed against VOC 202012/01 could equally neutralize D614G variant. Higher load of VOC 202012/01 in the nasal wash specimens was observed during the first week of infection outcompeting the D614G variant. The findings suggest increased fitness of VOC 202012/01 to the upper respiratory tract which could lead to higher transmission. Further investigations are needed to understand the transmissibility of new variants. One-Sentence SummarySARS-CoV-2 VOC 202012/01 infected hamsters demonstrated high viral RNA shedding through the nasal secretions and significant body weight loss with mild lung pathology compared to the D614G variant.

microbiology↗

bioRxiv2021

Ancient migrations - The first complete genome assembly, annotation and variants of the Zoroastrian-Parsi community of India

With the advent of Next Generation Sequencing, many population specific whole genome sequences published thus far, predominantly represent individuals of European ancestry. While sequencing efforts of underrepresented communities in genomes datasets, like the Yoruba West-African, Han Chinese, Tibetan, South Korean, Egyptian and Japanese have recently added to the public genomic repositories, a comprehensive understanding of human genomic diversity and discovery of trait-associated variants necessitates the need for additional population specific analysis. In this context, the genomics of the population from the Indian sub-continent, given its genetic heterogeneity needs further elucidation. In this context, the endogamous Zoroastrian-Parsi community of India, offer an exceptional insight into a homogenous population that has culturally, socially, and genetically remained intact, for 13 centuries amidst the genomic, social and cultural Indian landscape, consequent to their migration from the ancient Persian plateau. Notwithstanding longevity as a trait, this endangered community is highly susceptible to cancers, rare genetic disorders, and display a documented high incidence of neurodegenerative and autoimmune conditions. The community as a matter of cultural practice abstains from smoking. Here, we describe the assembly and annotation of the genome of an adult female, Zoroastrian-Parsi individual sequenced at a high depth of 173X using a combination of short Illumina reads (160X) and long nanopore reads (13X). Using a combination of hybrid assemblers, we created a new, population-specific human reference genome, The Zoroastrian-Parsi Genome Reference Female, AGENOME-ZPGRF, contains 2,778,216,114 nucleotides as compared to 3,096,649,726 in GRCh38 constituting 93.235% of the total genomic fraction. Annotation identified 20833 genomic features, of which 14996 are almost identical to their counterparts on GRCh38 while 5837 genomic features were covered in partial. AGENOME-ZPGRF contained 5,426,310 variants of which the majority were SNPs (4,291,601) and 960,867 SNPs were AGENOME-ZPGRF specific personal variants not listed in dbSNP. We present, AGENOME-ZPGRF as a whole reference for any genetic studies involving Zoroastrian-Parsi individuals extending their application to identify disease relevant prognostic biomarkers and variants in global population genomics studies.

genomics↗

bioRxiv2021

Assessment of immunogenicity and protective efficacy of ZyCoV-D DNA vaccine candidates in Rhesus macaques against SARS-CoV-2 infection

Vaccines remain the key protective measure to achieve herd immunity to control the disease burden and stop COVID-19 pandemic. We have developed and assessed the immunogenicity and protective efficacy of two formulations (1mg and 2mg) of ZyCoV-D (a plasmid DNA based vaccine candidates) administered through Needle Free Injection System (NFIS) and syringe-needle (intradermal) in rhesus macaques with three dose vaccine regimens. The vaccine candidate 2mg dose administered using Needle Free Injection System (NFIS) elicited a significant immune response with development of SARS-CoV-2 S1 spike region specific IgG and neutralizing antibody (NAb) titers during the immunization phase and significant enhancement in the levels after the virus challenge. In 2 mg NFIS group the IgG and NAb titers were maintained and showed gradual rise during the immunization period (15 weeks) and till 2 weeks after the virus challenge. It also conferred better protection to macaques evident by the viral clearance from nasal swab, throat swab and bronchoalveolar lavage fluid specimens in comparison with macaques from other immunized groups. In contrast, the animals from placebo group developed high levels of viremia and lung disease following the virus challenge. Besides this, the vaccine candidate also induced increase lymphocyte proliferation and cytokines response (IL-6, IL-5).The administration of the vaccine candidate with NFIS generated a better immunogenicity response in comparison to syringe-needle (intradermal route). The study demonstrated immunogenicity and protective efficacy of the vaccine candidate, ZyCoV-D in rhesus macaques.

microbiology↗

bioRxiv2020

A Fast Lysine Cross-linker DOPA Enables Mass Spectrometry Analyses of Protein Unfolding and Weak Protein-protein Interactions

Chemical cross-linking of proteins coupled with mass spectrometry analysis (CXMS) has become a widely used method for protein structure analysis. Central to this technology are chemical cross-linkers. The most popular cross-linkers are N-hydroxysuccinimide (NHS) esters, which react with protein amino groups relatively slowly over 10 minutes or more while in competition with the hydrolysis reaction of NHS esters. To improve the speed of cross-linking, we developed a new class of amine-selective and non-hydrolyzable di-ortho-phthalaldehyde (DOPA) cross-linkers. DOPA can cross-link proteins in 10 seconds under near physiological conditions, which is 60 times faster than the NHS ester cross-linker DSS. DOPA also works at low pH, low temperature, or in the presence of high concentrations of denaturants such as 8 M urea or 6 M guanidine hydrochloride. Further, DOPA-mediated pulse cross-linking captured the dynamic conformational changes associated with RNase A unfolding. Lastly, DOPA outperformed DSS at capturing weak but specific protein-protein interactions.

biochemistry↗

bioRxiv2020

Spontaneous emergence of azithromycin resistance in independent lineages of Salmonella Typhi in Northern India

BackgroundThe emergence and spread of antimicrobial resistance (AMR) pose a major threat to the effective treatment and control of typhoid fever. The ongoing outbreak of extensively drug resistant (XDR) Salmonella Typhi (S. Typhi) in Pakistan has left azithromycin as the only remaining broadly efficacious oral antimicrobial for typhoid in South Asia. Ominously, azithromycin resistant S. Typhi organisms have been subsequently reported in Bangladesh, Pakistan, and Nepal. MethodsHere, we aimed to understand the molecular basis of AMR in 66 S. Typhi isolated in a cross-sectional study performed in a suburb of Chandigarh in Northern India using whole genome sequencing (WGS) and phylogenetic analysis. ResultsWe identified seven S. Typhi organisms with the R717Q mutation in the acrB gene that was recently found to confer resistance to azithromycin in Bangladesh. Six out of the azithromycin-resistant S. Typhi isolates also exhibited triple mutations in gyrA (S83F and D87N) and parC (S80I) genes and were resistant to ciprofloxacin. These contemporary ciprofloxacin/azithromycin-resistant isolates were phylogenetically distinct from each other and from those reported from Bangladesh, Pakistan, and Nepal. ConclusionsThe independent emergence of azithromycin resistant typhoid in Northern India reflects an emerging broader problem across South Asia and illustrates the urgent need for the introduction of typhoid conjugate vaccines (TCVs) in the region. Key pointsWe identified ciprofloxacin/azithromycin-resistant Salmonella Typhi (S. Typhi) in Chandigarh in Northern India. The independent emergence of ciprofloxacin/azithromycin-resistant typhoid in Bangladesh, Pakistan, Nepal, and India and the continued spread of extensively-drug resistant (XDR) typhoid in Pakistan highlight the limitations of licensed oral treatments for typhoid fever in South Asia.

genomics↗

bioRxiv2020

Host transcriptomic profiling of COVID-19 patients with mild, moderate, and severe clinical outcomes

Characterizing key molecular and cellular pathways involved in COVID-19 is essential for disease prognosis and management. We perform shotgun transcriptome sequencing of human RNA obtained from nasopharyngeal swabs of patients with COVID-19, and identify a molecular signature associated with disease severity. Specifically, we identify globally dysregulated immune related pathways, such as cytokine-cytokine receptor signaling, complement and coagulation cascades, JAK-STAT, and TGF-{beta} signaling pathways in all, though to a higher extent in patients with severe symptoms. The excessive release of cytokines and chemokines such as CCL2, CCL22, CXCL9 and CXCL12 and certain interferons and interleukins related genes like IFIH1, IFI44, IFIT1 and IL10 were significantly higher in patients with severe clinical presentation compared to mild and moderate presentations. Moreover, early induction of the TGF-{beta} signaling pathway might be the primary cause of pulmonary fibrosis in patients with severe disease. Differential gene expression analysis identified a small set of regulatory genes that might act as strong predictors of patient outcome. Our data suggest that rapid transcriptome analysis of nasopharyngeal swabs can be a powerful approach to quantify host molecular response and may provide valuable insights into COVID-19 pathophysiology.

genomics↗

bioRxiv2020

An atlas of lamina-associated chromatin across thirteen human cell types reveals cell-type-specific and multiple subtypes of peripheral heterochromatin

Association of chromatin with lamin proteins at the nuclear periphery has emerged as a potential mechanism to coordinate cell type-specific gene expression and maintain cellular identity via gene silencing. Unlike many histone modifications and chromatin-associated proteins, lamin-associated domains (LADs) have yet to be mapped genome-wide in a diverse panel of human cell types, which has limited our understanding of the role peripheral chromatin plays in development and disease. To address this gap, we mapped LAMIN B1 (LB1) across twelve human cell types encompassing pluripotent stem cells, intermediate progenitors, and differentiated cells from all three germ layers. Integrative analyses of this atlas of peripheral chromatin with publicly available genomic data, as well as gene expression and repressive histone maps generated for this study, revealed that in all twelve cellular contexts lamin-associated chromatin is organized into at least two subtypes defined by differences in LB1 occupancy, gene expression, chromatin accessibility, transposable elements, replication timing, and radial positioning. Most genes gain or lose LB1 occupancy consistent with their cell type along developmental trajectories; however, we also identified examples where the enhancer, but not the gene body and promoter, change LAD state. Imaging of fluorescently labeled DNA in single cells validated these transitions and showed intermediate radial positioning of LADs that are gene dense, relatively accessible, and dynamic across development. This atlas represents the largest resource to date for peripheral chromatin organization studies.

developmental biology↗

bioRxiv2020

A Transcriptional Switch Governing Fibroblast Plasticity Underlies Reversibility of Chronic Heart Disease

In diseased organs, stress-activated signaling cascades alter chromatin, triggering broad shifts in transcription and cell state that exacerbate pathology. Fibroblast activation is a common stress response that worsens lung, liver, kidney and heart disease, yet its mechanistic basis remains poorly understood1,2. Pharmacologic inhibition of the BET family of transcriptional coactivators alleviates cardiac dysfunction and associated fibrosis, providing a tool to mechanistically interrogate maladaptive fibroblast states and modulate their plasticity as a potential therapeutic approach3-8. Here, we leverage dynamic single cell transcriptomic and epigenomic interrogation of heart tissue with and without BET inhibition to reveal a reversible transcriptional switch underlying stress-induced fibroblast activation. Transcriptomes of resident cardiac fibroblasts demonstrated robust and rapid toggling between the quiescent fibroblast and activated myofibroblast state in a manner that directly correlated with BET inhibitor exposure and cardiac function. Correlation of single cell chromatin accessibility with cardiac function revealed a novel set of reversibly accessible DNA elements that correlated with disease severity. Among the most dynamic elements was an enhancer regulating the transcription factor MEOX1, which was specifically expressed in activated myofibroblasts, occupied putative regulatory elements of a broad fibrotic gene program, and was required for TGF{beta}-induced myofibroblast activation. CRISPR interference of the most dynamic cis-element within the enhancer, marked by nascent transcription, prevented TGF{beta}-induced activation of Meox1. These findings identify MEOX1 as a central regulator of stress-induced myofibroblast activation associated with cardiac dysfunction. The plasticity and specificity of the BET-dependent regulation of MEOX1 in endogenous tissue fibroblasts provides new trans- and cis- targets for treating fibrotic disease.

molecular biology↗