Biology subjects

Jain, M.

Publications and source records attributed to Jain, M..

At least 55 records · Page 3Linked to original sources

bioRxiv2025

Deep learning-enabled scaffolding of spatial arrays of PfCSP epitopes

Malaria is a leading cause of disease in developing countries. The licensed malaria vaccine RTS,S/AS01 confers partial protection in part due to the elicitation of circumsporozoite protein (CSP) antibodies, of which those to the CSP repeat and junctional regions offer the most potent protection. Anti-repeat region antibodies, including the protective antibody L9, frequently develop mutations that promote inter-Fab contacts when bound to CSP in "spiral" quaternary structures. As a first step toward the design of immunogens that elicit L9-like antibodies, we utilized generative deep learning models to design epitope-scaffolds that incorporated up to three junctional repeat epitopes with structural conformations and relative spatial orientations matching those of the multivalent complex of CSP bound to three copies of L9. Affinity and structural studies demonstrated accurate scaffolding of two epitopes with the intended relative orientation, and less accurate positioning of the third epitope. This study demonstrates proof of principle for design of multi-epitope scaffolds with pre-determined relative epitope spatial positioning. The study also represents an initial step toward development of multi-epitope immunogens to elicit antibodies that utilize homotypic interactions to bind pathogen in multivalent clusters.

immunology↗

bioRxiv2025

Sounds of silence: electric mobility promises a quieter soundscape for wildlife, but may challenge ultrasonically sensitive species globally

Expanding transportation networks generate anthropogenic noise, a stressor for wildlife. The rapid transition to electric mobility, with projections of 40-45 million new electric vehicles (EVs) annually by 2030 (two in five new light-duty vehicles), promises quieter audible environments. However, a critical gap exists in our understanding of the acoustic trade-offs, especially of vehicle-generated ultrasound (frequencies >20 kHz). While ultrasound is known to be biologically potent, used for instance in wildlife repellents, its presence and potential impacts as an emission from traffic on sensitive species, such as bats, remain unaddressed. While the shift to EVs may reduce overall audible noise emissions, offering a green alternative, potential challenges for species with auditory systems tuned to ultrasound, and also actively using it via echolocation, could offset these benefits. This may set up a green-green dilemma, as the objective of decarbonising transport via e-mobility may undermine efforts to restore urban biodiversity. Our systematic review of 49 studies on anthropogenic/vehicular noise impacts on 114 bat species/species groups revealed that road noise, especially when overlapping with bat echolocation and communication frequencies, consistently elicited negative responses in playback experiments, whereas field observations were more variable. A meta-analysis from this dataset showed that the odds of bats being affected significantly increased with both noise intensity and frequency. Building on this evidence, this review explores the implications for a future dominated by EVs. We highlight the urgent need for further research to define acoustic thresholds and develop conservation strategies for wildlife in rapidly urbanising and electrifying landscapes.

ecology↗

bioRxiv2025

Haplotype-Resolved DNA Methylation at the APOE Locus identifies Allele-Specific Epigenetic Signatures Relevant to Alzheimer's Disease Risk

The APOE gene encodes a key lipid transport protein and plays a central role in Alzheimers disease (AD) pathogenesis. Three common APOE alleles, {varepsilon}2 (rs7412(C>T), {varepsilon}3 (reference), and {varepsilon}4 (rs429358(T>C)), arise from two coding variants in exon 4 and confer distinct AD risk profiles, with {varepsilon}4 increasing risk and {varepsilon}2 providing protection. The {varepsilon}3-linked APOE variant rs769455[T] has also been associated with elevated AD risk in individuals of African ancestry carrying both rs769455[T] and {varepsilon}4 alleles. These single nucleotide variants (SNVs) reside in a cytosine-phosphate-guanine (CpG) island, which is a region with a higher frequency of CpG sites compared to the rest of the genome. CpG sites are subject to 5-methylcytosine (5mC) methylation by DNA methyltransferases which add a methyl group to the fifth carbon on the cytosine residue of a CpG site. The presence of SNVs can disrupt this process, making these regions prime targets for differential methylation; however, allele-specific methylation patterns in APOE remain poorly resolved due to technical limitations of conventional bisulfite and methylation array based methods, including degraded DNA quality, sparse CpG coverage, and lack of haplotype phasing. Here, we leverage high-accuracy long-read sequencing data to generate haplotype-resolved methylation profiles of the APOE locus in 332 postmortem brain samples from two ancestrally different cohorts. This includes 201 individuals of European ancestry from the North American Brain Expression Consortium (NABEC), comprising 402 haplotypes (48 {varepsilon}2 and 58 {varepsilon}4 alleles), and 131 individuals of African and African admixed ancestry from the Human Brain Core Collection (HBCC), comprising 262 haplotypes (25 {varepsilon}2, 64 {varepsilon}4, and 7 rs769455 alleles). A linear regression analysis identified 18 novel differentially methylated CpG sites (DMCs) associated with APOE {varepsilon}2, {varepsilon}4, and rs769455 within a gene cluster spanning TOMM40, APOE, APOC1, and APOC4-APOC2. This represents the most comprehensive haplotype-resolved methylation study of APOE in human brain tissue to date. Our results uncover distinct allele-specific methylation signatures and demonstrate the power of long-read sequencing for resolving epigenetic variation relevant to AD risk.

genomics↗

bioRxiv2025

Population Pharmacokinetics and Probability of Target Attainment Analysis of Vancomycin Following Intermittent and Continuous Infusion in Adults with Cystic Fibrosis

Vancomycin is the drug of choice for treating pulmonary infections caused by methicillinresistant Staphylococcus aureus (MRSA) in people with cystic fibrosis (PwCF). This study characterized the pharmacokinetics (PK) of continuous and intermittent vancomycin infusions following a loading dose using population PK (PopPK) modeling to inform dosing in PwCF. The PopPK model was developed using therapeutic drug monitoring (TDM) data from adult PwCF who received a vancomycin loading dose followed by intermittent, continuous, or both infusion types for MRSA-related pulmonary exacerbations. A total of 212 samples were collected following 90 intermittent and 42 continuous infusions in 21 patients. The final model was a two-compartment model with first-order elimination, incorporating creatinine clearance (CrCL) as a covariate on vancomycin clearance (CL). The estimated CL and volume of distribution were 4.05 L/h/70 kg and 22.5 L/70 kg, respectively. The model was used to predict the probability of target attainment (PTA) following a single intermittent loading dose (500-1500 mg) and continuous infusion (500-6000 mg) over 24 hours. PTA was assessed using efficacy and toxicity thresholds defined by Area Under the Curve0-24 (AUC0-24)/Minimum Inhibitory Concentration (MIC) ratios [≥]400 mg{middle dot}h/L and <650 mg{middle dot}h/L, respectively. At a MIC of 1 {micro}g/mL, a loading dose of 500 mg followed by a 3750 mg continuous infusion achieved PTA targets for efficacy (66.7%) and safety (82.7%). These findings support the use of PopPK modeling to guide vancomycin dosing strategies for MRSA pulmonary infections in PwCF.

pharmacology and toxicology↗

bioRxiv2025

Evaluation of Nanopore direct RNA sequencing updates for modification detection

Nanopore technology can directly sequence intact RNA molecules, offering a unique capability to read native modifications. Oxford Nanopore Technologies recently updated its direct RNA sequencing technology from RNA002 to RNA004 chemistry. This update included an improved basecaller (Dorado) for increased sequencing accuracy, and ionic current models for de novo detection of four RNA modifications. Using a single RNA extraction from GM12878 B-lymphocyte cell line, we compared RNA002 and RNA004 sequencing chemistries and evaluated Dorado modification calling accuracy. We computed U-to-C mismatches, previously used to identify putative pseudouridine sites, and ran m6anet for identifying putative N6-methyladenosine sites. Dorado results for each respective modification showed both global and site-specific differences when compared to RNA002 results. We used DRS data from in vitro transcription of GM12878 genomic DNA as well as synthetic oligonucleotides to evaluate Dorado modification calling performance. Dorados pseudouridine model achieved 96-98% for both accuracy and F1-score. Similarly, Dorados N6-methyladenosine model achieved 94-98% accuracy, 96-99% F1-score. Our results demonstrate that Nanopore Direct RNA sequencing could simultaneously detect pseudouridine, N6-methyladenosine, 5-methylcytosine, and inosine modifications on individual mRNA strands. It is critical to validate these calls using orthogonal methods (e.g., Liquid Chromatography with Tandem Mass Spectrometry) for increased confidence. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=81 SRC="FIGDIR/small/651717v1_ufig1.gif" ALT="Figure 1"> View larger version (25K): org.highwire.dtl.DTLVardef@1215534org.highwire.dtl.DTLVardef@160f38forg.highwire.dtl.DTLVardef@164a9corg.highwire.dtl.DTLVardef@17c72e5_HPS_FORMAT_FIGEXP M_FIG C_FIG

genomics↗

bioRxiv2025

Genomic in vitro transcription and Nanopore direct RNA sequencing of a human B-Lymphocyte cell line

Genomic DNA used as a template for in vitro transcription of RNA can serve as a true negative control for benchmarking RNA modification detection by Nanopore direct RNA sequencing (DRS) models. We generated DRS data for in vitro transcribed (IVT) RNA composed of canonical nucleotides using genomic DNA from a human cell line. We applied Dorado modification calling models to these data, and calculated 9-mer specific false-positive rates for eight RNA modifications as a comparison point for future development of RNA modification models. O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=119 SRC="FIGDIR/small/650674v2_ufig1.gif" ALT="Figure 1"> View larger version (23K): org.highwire.dtl.DTLVardef@962feorg.highwire.dtl.DTLVardef@4237aborg.highwire.dtl.DTLVardef@154d346org.highwire.dtl.DTLVardef@1fad8f2_HPS_FORMAT_FIGEXP M_FIG C_FIG

genomics↗

bioRxiv2025

Isotope tracing-based metabolite identificationfor mass spectrometry metabolomics

Modern mass spectrometry-based metabolomics is a key technology for biomedicine, enabling discovery and quantification of a wide array of biomolecules critical for human physiology. Yet, only a fraction of human metabolites have been structurally determined, and the majority of features in typical metabolomics data remain unknown. To date, metabolite identification relies largely on comparing MS2 fragmentation patterns against known standards, related compounds or predicted spectra. Here, we propose an orthogonal approach to identification of endogenous metabolites, based on mass isotopomer distributions (MIDs) measured in an isotope-labeled reference material. We introduce a computational measure of pairwise distance between metabolite MIDs that allows identifying novel metabolites by their similarity to previously known peaks. Using cell material labeled with 20 individual 13C tracers, this method identified 62% of all unknown peaks, including previously never seen metabolites. Importantly, MID-based identification is highly complementary to MS2-based methods in that MIDs reflect the biochemical origin of metabolites, and therefore also yields insight into their synthesis pathways, while MS2 spectra mainly reflect structural features. Accordingly, our method performed best for small molecules, while MS2-based identification was stronger on lipids and complex natural products. Among the metabolites discovered was trimethylglycyl-lysine, a novel amino acid derivative that is altered in human muscle tissue after intensive lifestyle treatment. MID-based annotation using isotope-labeled reference materials enables identification of novel endogenous metabolites, extending the reach of mass spectrometry-based metabolomics.

biochemistry↗

bioRxiv2025

Edited Filamin A in myeloid cells reduces intestinal inflammation and protects from colitis

Patho-mechanistic origins and disease dynamics of ulcerative colitis are still poorly understood. The actin-crosslinker Filamin A (FLNA) impacts cellular responses through interaction with cytosolic proteins. FLNA exists in two forms that differ only in one amino acid: genome-encoded FLNAQ and FLNAR - generated by post-transcriptional A-to-I editing. FLNA is edited in fibroblasts, smooth muscle- and endothelial cells in the colon. We identified the FLNA editing status as a key determinant of colitis severity. FLNA editing was highest in healthy colons and reduced during acute murine and human colitis. Mice that exclusively express edited FLNAR and do not downregulate editing upon challenge were highly resistant to DSS-induced colitis, whereas fully unedited FLNAQ animals developed severe inflammation. While the genetic induction of FLNA editing influenced transcriptional states of structural cells and the microbiome composition, we found that FLNAR exerts protection specifically via its influence on myeloid cells, which are not edited under physiological conditions. Introducing fixed, fully edited FLNAR did not hamper normal cell migration but reduced macrophage inflammation and rendered neutrophils less prone to NETosis. In conclusion, loss of FLNA editing correlates with colitis severity, and targeted FLNA editing of myeloid cells might serve as a novel therapeutic approach in intestinal inflammation. SummaryIn this study, Gawish et al. show that RNA editing of the actin cross-linker FLNA is similarly regulated in mice and humans and that the targeted induction of edited FLNAR in myeloid cells governs resistance to DSS-induced colitis, revealing its potential in IBD therapy.

molecular biology↗

bioRxiv2025

The Th1/Th17 axis regulates chimeric antigen receptor (CAR) T cell therapy toxicities

CAR-T therapy has led to significant improvements in patient survival. However, a subset of patients experience high-grade toxicities, including cytokine release syndrome (CRS) and immune cell-associated hematologic toxicity (ICAHT). We utilized IL-2R knockout mice to model cytokine toxicities with elevated levels of IL6, IFN{gamma}, and TNF and increased M1-like macrophages. Onset of CRS was accompanied by a reduction in peripheral blood neutrophils due to disruption of bone marrow neutrophil homeostasis characterized by an increase in apoptotic neutrophils and a decrease in proliferative and mature neutrophils. Both non-tumor-bearing and E-ALL tumor-bearing mice recapitulated the co-occurrence of CRS and neutropenia. IFN{gamma}-blockade alleviated CRS and neutropenia without affecting CAR-T efficacy. Mechanistically, a Th1-Th17 imbalance was observed to drive co-occurrence of CRS and neutropenia in an IFN{gamma}-dependent manner leading to decreased IL-17A and G-CSF, neutrophil production, and neutrophil survival. In patients, we observed an increase in the IFN{gamma}-to-IL-17A ratio in the peripheral blood during high-grade CRS and neutropenia. We have uncovered a biological basis for ICAHT and provide support for the use of IFN{gamma}-blockade to reduce CRS and neutropenia. Statement of SignificanceDespite clinical success of CAR-T therapy, patients develop toxicities such as cytokine release syndrome and neutropenia, whose co-occurrence impacts their survival and quality-of-life. We recapitulate these toxicities in mice to discover their co-occurrence is driven by Th1-Th17 imbalance following CAR-T administration, which can be prevented via IFN{gamma} blockade.

cancer biology↗

bioRxiv2025

The T Cell-Specific miRNA-Target Network in Psoriasis: A Systematic Review

Psoriasis is a chronic skin disease characterized by keratinocyte hyperproliferation, involving T cells as the major players in the manifestation of lesional and systemic inflammation. With the extensive exploration of keratinocyte dysfunctionality, T cell-centric deregulation has gained much focus in the context of disease etiology. miRNAs, the molecular switches known to drive disease manifestation via altering keratinocyte biology, have not been much studied in the context of T cell alterations, with only a few reports. This study aims to explore T cell-associated miRNAs and their target signaling networks to understand their role in modulating psoriasis-specific T cell changes. miRNAs with altered expression in psoriatic T cells were identified through a meta-analysis of the existing literature and datasets. These miRNAs were then used to identify cognate targets and T cell signaling pathways to elucidate T cell-specific cascades influenced by miRNAs, potentially explaining miRNA driven T cell dysregulation in psoriasis. The study identified 14 miRNAs with altered expression in psoriatic T cells and 256 downstream targets involved in 15 T cell-associated signaling pathways. The miRNA-driven targets and pathways identified may dysregulate T cell activation, proliferation, survival, and subset differentiation. These findings suggest multiple miRNA-target signaling axes that require experimental validation to better understand T cell-mediated disease etiology.

systems biology↗

bioRxiv2025

Neutrophil KLF2 regulates inflammasome-dependent neonatal mortality from endotoxemia

Preterm neonates die at a significantly higher rate from sepsis than full-term neonates, attributable to their dysregulated immune response. In addition to tissue destruction caused directly by bacterial invasion, an overwhelming cytokine response by the immune cells to bacterial antigens also results in collateral damage. Sepsis leads to decreased gene expression of a critical transcription factor, Kruppel-like factor-2 (KLF2), a tonic repressor of myeloid cell activation. Using a murine model of myeloid-Klf2 deletion, we show that loss of KLF2 is associated with decreased survival after endotoxemia in a developmentally dependent manner, with increased mortality at postnatal day 4 (P4) compared to P12 pups. This survival is significantly increased by neutrophil depletion. P4 knockout pups have increased pro-inflammatory cytokine levels after endotoxemia compared to P4 controls or P12 pups, with significantly increased levels of IL-1{beta}, a product of the activation of the NLRP3 inflammasome complex. Loss of myeloid-KLF2 at an earlier postnatal age leads to a greater increase in NLRP3 priming and activation and greater IL-1{beta} release by BMNs. Inhibition of NLRP3 inflammasome activation by MCC950 significantly increased survival after endotoxemia in P4 pups. Transcriptomic analysis of bone marrow neutrophils showed that loss of myeloid-KLF2 is associated with gene enrichment of pro-inflammatory pathways in a developmentally dependent manner. These data suggest that targeting KLF2 could be a novel strategy to decrease the pro-inflammatory cytokine storm in neonatal sepsis and improve survival in neonates with sepsis. Summary sentenceKLF2 regulates the developmental response to endotoxin in neonatal mice through the NLRP3 inflammasome signaling pathway.

immunology↗

bioRxiv2025

State or personality trait: Determinants of boldness and shoal size preference in adult zebrafish

Animals adjust their behavior in response to physiological states to optimize the trade-off between energy acquisition and survival. A key question in animal personality research is whether behavioral variation is driven by consistent physiological state differences or stable personality traits. This study evaluates boldness and shoal size discrimination in adult male zebrafish (Danio rerio) across three hunger states: same-day feeding (H0), 24-hour food deprivation (H1), and 48-hour food deprivation (H2). Behavioral assessments were conducted over two testing cycles, with each individual undergoing every test twice for each hunger state within each cycle. The assays comprised the Open Field Test (OFT), Emergence Test (ET), and Social Preference Test (SPT), with the SPT conducted under binary (4 vs. 2 fish) and ternary (4 vs. 2 vs. 1 fish) choice conditions. Hunger significantly reduced emergence latency, but did not affect exploratory behavior in the OFT. Hunger also enhanced preference for larger shoals above chance in the binary SPT during the second testing cycle, though this effect was absent in ternary choices. Testing cycle effects revealed habituation-driven increases in boldness in both the tests and a progressive shift toward larger shoal preferences in the binary SPT. Among all assays, only the OFT showed significant repeatability, making it a reliable measure of boldness. However, it did not show consistent individual differences in plasticity due to hunger. These results highlight the crucial role of labile physiological states, such as hunger and habituation, in driving behavioral variation and underscore the importance of designing studies that disentangle state-dependent behaviors from stable personality traits in animal personality research.

animal behavior and cognition↗

bioRxiv2025

Pancreatic injury induces β-cell regeneration in axolotl

BackgroundDiabetes is a condition characterized by a loss of pancreatic {beta}-cell function which results in the dysregulation of insulin homeostasis. Using a partial pancreatectomy model in axolotl, we aimed to observe the pancreatic response to injury. ResultsHere we show a comprehensive histological assessment of pancreatic islets in axolotl. Following pancreatic injury, no apparent blastemal structure was observed. We found a significant, organ-wide increase in cellular proliferation post-resection in the pancreas compared to sham-operated controls. This proliferative response was most robust at the site of injury. We found that {beta}-cells actively contributed to the increased rates of proliferation upon injury. {beta}-cell proliferation manifested in increased {beta}-cell mass in injured tissue at two weeks post injury. At four weeks post injury, we found organ-wide proliferation to be extinguished while proliferation at the injury site persisted, corresponding to pancreatic tissue recovery. Similarly, total {beta}-cell mass was comparable to sham after four weeks. ConclusionsOur findings suggest a non-blastema-mediated regeneration process takes place in the pancreas, by which pancreatic resection induces whole-organ {beta}-cell proliferation without the formation of a blastemal structure. This process is analogous to other models of compensatory growth in axolotl, including liver regeneration.

developmental biology↗

bioRxiv2024

Long-read sequencing of hundreds of diverse brains provides insight into the impact of structural variation on gene expression and DNA methylation

Structural variants (SVs) drive gene expression in the human brain and are causative of many neurological conditions. However, most existing genetic studies have been based on short-read sequencing methods, which capture fewer than half of the SVs present in any one individual. Long-read sequencing (LRS) enhances our ability to detect disease-associated and functionally relevant structural variants; however, its application in large-scale genomic studies has been limited by challenges in sample preparation and high costs. Here, we leverage a new scalable wet-lab protocol and computational pipeline for whole-genome Oxford Nanopore Technologies sequencing and apply it to neurologically normal control samples from the North American Brain Expression Consortium (NABEC) (European ancestry) and Human Brain Collection Core (HBCC) (African or African admixed ancestry) cohorts. Through this work, we present a publicly available long-read resource from 351 human brain samples (median N50: 27 Kbp and at an average depth of ~40x genome coverage). We discover approximately 234,905 SVs and produce locally phased assemblies that cover 95% of all protein-coding genes in GRCh38. To resolve cis-regulatory effects, we develop ASM-LR, a method for allele-specific methylation analysis from long-read data, revealing both strong and subtle regulatory effects, including numerous novel methylation QTLs masked in unphased models. Our results highlight the power of haplotype-resolved methylation to uncover regulatory mechanisms and establish a foundational resource for exploring how genetic variation shapes gene expression and epigenetic architecture across diverse ancestries.

genomics↗

bioRxiv2024

Branched DNA Processing by a Thermostable CAS-Cas4 from Thermococcus onnurineus: Expanding Biochemical Landscape of Nuclease Activity

The adaptive immune function of CRISPR-Cas systems in bacteria and archaea is mediated through CRISPR-Associated Proteins (Cas). The adaptation module, typically involving Cas1, Cas2, and Cas4, helps integrate viral "spacer" sequences into the host genome. Cas4 proteins are classified into two types based on neighboring genes: CAS-Cas4, flanked by other cas genes, and Solo-Cas4, which exist independently. While CAS-Cas4 proteins are implicated in adaptation, they remain biochemically uncharacterized in archaea, unlike archaeal Solo-Cas4 proteins. This study biochemically characterizes TON_0321, a CAS-Cas4 protein from the Type IV-C CRISPR cassette of Thermococcus onnurineus. TON_0321 exhibits 5' to 3' exonuclease activity and unique structure-dependent endonuclease activity, shedding light on CAS-Cas4 functional diversity. A distinct spatial organization of the catalytic site, angled with the positively charged patch on the protein surface, enables TON_0321 to recognize branching points in DNA substrates. Furthermore, this spatial arrangement facilitates cleavage 2 to 3 nucleotides away from the branch point in the 5' direction, demonstrating structure-specific endonuclease activity.

biochemistry↗

bioRxiv2024

The impact of fold-dilution and diluent volume on the estimation of colony forming units in a bacterial culture

Considering the non-uniform distribution of bacteria in a liquid culture, accurate enumeration of bacteria by serial dilution is important yet challenging in microbiological research. The deviation of estimated population size from the actual population size is anticipated in view of the sampling error arising at dilution, which is known to increase at each successive dilution; and the sampling error associated with the sample volume, where it is inversely proportional. In the dilution process of such population following a Poisson distribution, the relation of sampling error with the sequentially diluted suspensions is yet to be empirically demonstrated. Also, as a given-fold dilution can be achieved by passaging of either a higher or a lower sample volume, the sampling error associated in this case with the volume being passaged also remains to be evaluated. In view of that, the current experimental study was undertaken to further understand bacterial distribution and the underlying sampling error in dimensions previously not demonstrated. We demonstrated that sampling error in the enumeration process is independent of the bacterial cell densities of the diluted suspensions, which was on contrary to the theoretical assumptions. Moreover, sampling error is also not dependent on the number of passages in the dilution process. Importantly, we also demonstrated that passaging either a higher or lower sample volumes during serial dilution yielded similar enumeration, which was exceptional with regards to the statistical fluctuations inherent to a Poisson distribution.

microbiology↗

bioRxiv2024

Tissue-independent nature of plant susceptibility: a comparative pathogenicity study of the systemic phytopathogen Ralstonia pseudosolanacearum in eggplant and tomato seedlings through root and leaf

Ralstonia pseudosolanacearum causes a lethal bacterial wilt disease in many plant species, posing significant economic challenges. Although tomato has been a primary model host for investigating the pathogenicity and systemic infection of this bacterium, this manuscript presents a comparative pathogenicity study between two closely related solanaceous hosts, tomato and eggplant, revealing differential host responses to the same pathogen, specifically through leaves and roots. Interestingly, eggplant seedlings exhibited a significantly higher susceptibility to cotyledon leaf inoculation than tomato seedlings. In the case of tomato, a few seedlings escaped wilting (called escapees), which was usual at a high pathogen load, unlike the case of eggplant seedlings where escapees could not be observed even at a 100-fold lower pathogen concentration. The greater susceptibility of eggplant was further demonstrated by performing both single- and double-leaf inoculations in the seedlings. Surprisingly, root inoculations resulted in a significantly lower mortality for eggplant than for tomato seedlings. The contrasting susceptibility between the two hosts regarding root and leaf regions indicates the tissue-dependent nature of susceptibility. The study underscores the value of employing multiple host species to uncover new insights into pathogen behaviour and host-pathogen interactions.

pathology↗

bioRxiv2024

Clair3-RNA: A deep learning-based small variant caller for long-read RNA sequencing data

Variant calling using long-read RNA sequencing (lrRNA-seq) can be applied to diverse tasks, such as capturing full-length isoforms and gene expression profiling. It poses challenges, however, due to higher error rates than DNA data, the complexities of transcript diversity, RNA editing events, etc. In this paper, we propose Clair3-RNA, the first deep learning-based variant caller tailored for lrRNA-seq data. Clair3-RNA leverages the strengths of the Clair series pipelines and incorporates several techniques optimized for lrRNA-seq data, such as uneven coverage normalization, refinement of training materials, editing site discovery, and the incorporation of phasing haplotype to enhance variant-calling performance. Clair3-RNA is available for various platforms, including PacBio and ONT complementary DNA sequencing (cDNA), and ONT direct RNA sequencing (dRNA). Our results demonstrated that Clair3-RNA achieved a [~]91% SNP F1-score on the ONT platform using the latest ONT SQK-RNA004 kit (dRNA004) and a [~]92% SNP F1-score in PacBio Iso-Seq and MAS-Seq for variants supported by at least four reads. The performance reached a [~]95% and [~]96% F1-score for ONT and PacBio, respectively, with at least ten supporting reads and disregarding the zygosity. With read phased, the performance reached [~]97% for ONT and [~]98% for PacBio. Extensive evaluation of various GIAB samples demonstrated that Clair3-RNA consistently outperformed existing callers and is capable of distinguishing RNA high-quality editing sites from variants accurately. Clair3-RNA is open-source and available at (https://github.com/HKU-BAL/Clair3-RNA).

bioinformatics↗