Biology subjects

Kalia, V.

Publications and source records attributed to Kalia, V..

At least 19 recordsLinked to original sources

bioRxiv2026

Hypoxia versus immune depletion - immune profiling and treatment cessation provide mechanistic insights and considerations for translation in Leigh syndrome

Genetic mitochondrial diseases (GMDs) are major challenges to human health accounting for a significant fraction of heritable neurologic diseases, myopathies, and inborn errors of metabolism. Leigh syndrome (LS) is the most common clinical presentation of GMD in pediatric patients. LS is a severe and complex disease for which effective clinical therapies are currently lacking. Preclinical therapies identified in the Ndufs4(-/-) mouse model of LS include immune-targeting interventions and chronic mild hypoxia (11% oxygen). Immune-targeting interventions include rapamycin and high-dose pexidartinib, the latter appearing to fully suppress disease. The mechanisms underlying the benefits of hypoxia remain unclear, and the relationship between hypoxia and immune interventions have not been assessed. Here, we report the immune profile of brainstem of the Ndufs4(-/-) mouse model prior to and after disease onset and the impact of pexidartinib treatment. We provide evidence that macrophages/monocytes drive pathology, consistent with recent genetic studies. We additionally find that pre-disease onset animals lack signs of inflammation, and that the elimination of leukocytes fully suppresses the molecular signature of disease. Finally, using distinct post-developmental periods of treatment, we find pexidartinib and rapamycin provide benefits which persist long beyond treatment cessation, while cessation of hypoxia results in rapid disease onset and an acceleration of disease progression. These findings are consistent with hypoxia acting upstream of immune cell activation and have major implications for the therapeutic translation of both hypoxia and immune targeting interventions. Our findings establish hypoxia-cessation as a novel method for synchronizing inflammatory disease onset in the Ndufs4(-/-) model which will be useful in future mechanistic studies.

pathology↗

bioRxiv2026

SARS-CoV-2 S1 spike protein induces a temporal systemic immune response and promotes long-term anxiety-like behaviors

The SARS-CoV-2 S1 protein is associated with immune cell activation and persistent neurological symptoms, yet the underlying mechanisms remain unclear, posing a major challenge in elucidating Long COVID pathophysiology. To investigate how circulating S1 contributes to long-term neurological alterations, we intravenously injected hACE2 mice with varying doses of S1 (5, 10, and 20 {micro}g) and observed temporally dysregulated systemic inflammatory responses accompanied by sub-acute CD4+ T cell infiltration into central limbic regions. This immune response induced mild sustained increases in cFos+ cells in the amygdala and mild neuroinflammation in the hippocampal CA1 region, resulting in both acute and long-term anxiety-like behaviors, while working memory remained unaffected. Together, these findings suggest that systemic S1 protein induces a sustained proinflammatory response that promotes lasting neurological alterations through immune-to-brain signaling pathways. Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=136 SRC="FIGDIR/small/733010v1_ufig1.gif" ALT="Figure 1"> View larger version (36K): org.highwire.dtl.DTLVardef@10982caorg.highwire.dtl.DTLVardef@169a275org.highwire.dtl.DTLVardef@28e66aorg.highwire.dtl.DTLVardef@12f6c85_HPS_FORMAT_FIGEXP M_FIG C_FIG

Molecular Biology↗

bioRxiv2026

Sex-specific roles for yawning in the Emperor cichlid Boulengerochromis microlepis

Yawning is a poorly understood, yet spontaneous and ubiquitous behaviour. Three main hypotheses explain the function of yawns, most notably stimulating arousal. Yawns can also play a role in social cohesion through their "contagious" nature, eliciting an unconscious yawn in an observer. Yawn contagion has been observed in mammals and birds, and has been suggested in fish. Yawning also contributes to sexual dimorphisms, whereby males and females display differences in yawn frequency or function. Here, we examine these hypotheses in juveniles of the African cichlid, Boulengerochromis microlepis. By re-analyzing hundreds of hours of video recordings and nearly 3000 observed yawns we find support for all three yawn hypotheses in fish and two temporally and sexually distinct effects for yawns. First, our results suggest yawns at night and dawn induce arousal by delaying transitions into sleep. In contrast, daytime yawns induced rapid yawn contagion in neighbouring fish. Our analyses also demonstrate clear differences between sexes, with increased yawning in males at night and dawn associated with less sleep, whereas females displayed more robust social yawning. Together, our results expand the evolutionary context for yawning, and raise further questions about its functions in sleep and social cohesion across animals, and across sexes.

animal behavior and cognition↗

bioRxiv2026

MyD88 deficiency modestly attenuates disease in a Leigh syndrome mouse model while enrofloxacin accelerates disease

Primary genetic mitochondrial diseases (GMDs) are a clinically and genetically diverse group of diseases estimated to impact over 1 in 4,000 individuals. Leigh syndrome (LS) is the most common pediatric presentation of GMD. LS typically presents within the first years of life and is a severe progressive multi-system disorder. Symmetric progressive inflammatory brain lesions are a defining feature of the disease. Patients can also present with seizures, metabolic dysfunction, muscle weakness, and other symptoms. No effective clinical treatments currently exist. Recent data from the Ndufs4(-/-) mouse model shows that peripheral macrophages contribute to brain lesions in LS, that disease is causally driven by innate immune populations, and that depletion of innate immune cells prevents LS disease. However, the precise mechanisms underlying immune activation remain unknown. Certain mitochondrial macromolecules retain bacterial signatures and can act as potent agonists for innate immune pathways. For example, cytoplasmic mitochondrial RNA and mitochondrial DNA are detected by Toll-like receptors (TLRs) 7 and 9, respectively, at the endosome. Accordingly, these are considered strong candidates for mediating innate immune activation in LS. Here, we generated TLR signaling deficient Ndufs4(-/-)/MyD88(-/-) animals to assess whether TLR signaling plays a role in disease onset or progression in LS. Loss of MyD88 in Ndufs4(-/-) animals statistically significantly increased survival and delayed the onset of some symptoms, but the benefits were modest compared to CSF1R inhibition from prior work. We conclude that Myd88-mediated immune signaling is not a primary driver of LS. Notably, prophylactic enrofloxacin treatment, which was necessary for production of innate immune deficient MyD88(-/-) animals, modestly decreased survival and accelerated disease. The impact of enrofloxacin and similar drugs in the context of mitochondrial disease warrants further investigation.

neuroscience↗

bioRxiv2026

Querying functional drivers in primary murine naive and memory T cells using RNP-mediated CRISPR-Cas9 Gene Deletion

Cytotoxic CD8 T lymphocytes (CTL) are critical for the clearance of pathogenic cells via antigen-dependent cell lysis, thus providing protection against infectious diseases and cancers. CTLs represent one of the key targets of immunotherapies and vaccine design. While microarray and single-cell RNA sequencing of effector and memory CD8 T cells have identified several promising gene targets that may modulate CD8 T cell responses, their development is slowed down by the financial and time constraints related to the generation of germline knockout mice for further validation studies. Here we present a protocol for conducting efficient deletion of genes from activated effector, as well as resting naive and memory primary murine CD8 T cells using RNP-based Crispr/Cas9 technology. This CRISPR modification of CD8 T cells was then adapted to study the effects of gene deletion in the context of acute memory differentiation as well as chronic exhaustion of CD8 T cells. To this end, we have titrated the necessary dose of antigen-specific CD8 T cells to study their differentiation in acute and chronic infections and confirmed the model by demonstrating rapid expansion of CRISPR mediated PD-1 ablated CD8 T cells in a chronic viral infection. Finally, by combining this methodology with a murine model of subcutaneous tumor challenge, this study provides a unique screening system for genes critical for mediating clearance of malignant tumor cells by CTLs. This study expands current technical capabilities for rapid evaluation of functional role of any candidate gene in CTL responses to infections and cancer through targeted gene deletion at any stage of CD8 T cell differentiation without the need for germline gene deletion mouse models.

immunology↗

bioRxiv2026

Suppression of CD4 T cells by Tregs dictates metabolic remodeling of CD8 T cells during effector to memory transition

Metabolic transitions between naive, effector and memory T cell states are largely orchestrated by TCR, costimulatory and cytokine signals along with nutrient availability in the immune microenvironment. Treg cells have been shown to play a critical role in the effector-to-memory (E-M) transition of virus-specific CD8 T cells through regulation of proliferation and cytotoxic functional programs. However, the precise Treg-dependent metabolic changes that occur in the microniches and, the underlying molecular and cellular mediators of E-M transition remain undefined. Here we show that Treg cells promote the metabolic remodeling of memory-precursor effector CD8 T cells (MPEC) from aerobic glycolysis to fatty acid oxidation as they enter quiescence after antigen clearance. Our data implicate the anatomic microniche of the splenic white pulp as a site for Treg-MPEC interactions. We further show that optimal E-M metabolic transition requires regulation of effector CD4 T cells and inflammatory myeloid cells through inhibitory CTLA4 signals from Treg cells. Moreover, antagonism of inflammatory cytokine interferon-{gamma} (IFN-{gamma}) signals partially rescues the memory defects associated with absence of Treg cells. Together, these findings support a metabolic triad model of memory CD8 T cell differentiation where Treg-dependent regulation of inflammation from effector CD4 T cells promotes the transition of CD8 T cells from cytotoxic effector to quiescent memory metabolic programs. These studies define novel molecular targets that may be exploited to manipulate metabolism, migration and memory function during vaccination.

immunology↗

bioRxiv2026

MicroRNA-17~92 drives metabolic programming of virus-specific effector CD4 and CD8 T cell responses

The miR-17[~]92 microRNA cluster drives oncogenesis by inducing proliferation and survival of cancer cells. A similar pro-proliferative role of miR-17[~]92 has also been identified in pathogen- and tumor-reactive T cells. However, the metabolic underpinnings of miR-17[~]92-drivenT cell expansion and effector differentiation remain undefined. Using a murine model of conditional miR-17[~]92 deletion or constitutive overexpression in viral antigen-specific CD4 or CD8 T cells, here we show that miR-17[~]92 drives terminal differentiation of CD4 and CD8 T cells through sustained activation of mTOR and glycolytic bioenergetics pathways. Constitutively increased expression of miR-17[~]92 led to a universal increase in the numbers of antigen-specific T follicular helper (TFH), T helper 1 (TH1) and cytotoxic T lymphocyte (CTL) effector subsets. During early stages of T cell activation, miR-17[~]92 overexpression was associated with increased glucose uptake and heightened glycolytic and oxidative metabolism to sustain increased proliferation in both CD4 and CD8 T cells. However, prolonged overexpression of miR-17[~]92 led to a loss of polyfunctionality and compromised metabolic fitness (glycolysis and mitochondrial respiration) by the peak of the effector responses. These studies establish miR-17[~]92 as a critical driver of T cell proliferation and effector differentiation through metabolic regulation of both effector CTL and CD4 T cell subsets (TFH and TH1) that are critical for combating viral infections. These studies form the basis for future manipulation of miR-17[~]92 gene/metabolic regulatory network to commandeer T cell immunity during infection, vaccination or cancer immunotherapy.

immunology↗

bioRxiv2025

Transgenic Cowpea Expressing Synthetic BtCry1Ab Confers High Resistance to Legume Pod Borer (Maruca vitrata)

Cowpea, an important food legume crop, suffers significant yield losses due to insect pests, particularly the legume pod borer (Maruca vitrata). Narrow genetic base and lack of sufficient resistance to M. vitrata in existing cowpea germplasm, make them less effective in managing M. vitrata populations. To address this challenge, a transgenic approach is explored. In this study, we develop transgenic cowpea with enhanced resistance to M. vitrata by overexpressing a synthetic cry1Ab gene, driven by constitutive CaMV35S viral promoter, in an Indian cowpea cultivar. High expression of Cry1Ab was detected in leaves and pods. Bioassays with M. vitrata and Helicoverpa armigera larvae demonstrated significant resistance of transgenic lines to these lepidopteran pests. The transgenic lines exhibited reduced pod damage, decreased larval feeding, and increased insect mortality compared to non-transgenic controls. NMR analysis demonstrated absence of any new metabolites in transgenic plants. Furthermore, these transgenic lines showed no penalty on growth and development. The transgenic lines exhibited no observable phenotypic drawbacks, indicating their promise for sustainable agricultural practices. Our findings suggest that these transgenic cowpea lines could offer broad-spectrum protection against these pests, providing an effective pest management solution.

plant biology↗

bioRxiv2025

Divergent effects of a Treg selective IL-2 mutein on Influenza-specific T cell responses

Enhancing regulatory T cell (Treg) function offers a compelling therapeutic strategy for autoimmune disease. Engineered IL-2 muteins selectively expand functional Tregs with minimal impact on other immune cells, but their potential to compromise antiviral immunity remains largely unexplored. Here, we used a murine model of Influenza A virus (Flu) infection to determine how IL-2 mutein shapes T cell responses to respiratory virus infection. IL-2 mutein administration prior to infection suppressed Flu-specific (Flu-sp) CD8 T cell responses and altered their localization and phenotype within the lungs, without affecting bystander CD8 T cells. This suppression correlated with reduced antigen presentation molecule expression on conventional dendritic cells (cDCs) early after infection but did not impact Flu-sp CD8 T cell priming. In contrast, administering IL-2 mutein during infection exacerbated disease and drove CD25-dependent expansion of Flu-sp CD8 T cells. Despite these opposing effects on effector responses, Fc.Mut24-treated mice generated robust antibody responses and protective T cell memory which were maintained for at least 170 days. These findings reveal that Fc.Mut24 has temporally distinct effects on antiviral immunity, dampening early effector responses when given before infection, but enhancing effector expansion and disease severity when delivered during infection. Our results provide critical context for the therapeutic application of IL-2 muteins and highlight the importance of treatment timing in balancing immune modulation with protective immunity.

immunology↗

bioRxiv2024

Potent antitumor activity of a designed interleukin-21 mimic

Long-standing goals of cancer immunotherapy are to activate cytotoxic antitumor T cells across a broad range of affinities while dampening suppressive regulatory T (Treg) cell responses, but current approaches achieve these goals with limited success. Here, we report a de novo IL-21 mimic, 21h10, designed to have augmented stability and high signaling potency in both humans and mice. In multiple animal models and in ex vivo human melanoma patient derived organotypic tumor spheroids (PDOTS), 21h10 showed robust antitumor activity. 21h10 generates significantly prolonged STAT signaling in vivo compared with native IL-21, and has considerably stronger anti-tumor activity. Toxicities associated with systemic administration of 21h10 could be mitigated by TNF blockade without compromising antitumor efficacy. In the tumor microenvironment, 21h10 induced highly cytotoxic antitumor T cells from clonotypes with a range of affinities for endogenous tumor antigens, robustly expanding low-affinity cytotoxic T cells and driving high expression of interferon- (IFN-) and granzyme B compared to native IL-21, while increasing the frequency of IFN-+ Th1 cells and reducing that of Foxp3+ Tregs. As 21h10 has full human/mouse cross-reactivity, high stability and potency, and potentiates low-affinity antitumor responses, it has considerable translational potential.

biochemistry↗

bioRxiv2023

Glial swip-10 expression controls systemic mitochondrial function, oxidative stress, and neuronal viability via copper ion homeostasis

Cuprous copper (Cu(I)) is an essential cofactor for enzymes supporting many cellular functions including mitochondrial respiration and suppression of oxidative stress. Neurons are particularly dependent on these pathways, with multiple neurodegenerative diseases, including Alzheimers disease (AD), Parkinsons disease, associated with their dysfunction. Key features of Cu(I) contributions to neuronal health in vivo remain to be defined, owing largely to the complex processes involved in Cu(I) production, intracellular transport, and systemic redistribution. Here, we provide genetic and pharmacological evidence that swip-10 is a critical determinant of systemic Cu(I) levels in C. elegans, with deletion leading to systemic deficits in mitochondrial respiration, production of oxidative stress, and neurodegeneration. These phenotypes can be reproduced in wild-type worms by Cu(I)-specific chelation and offset in swip-10 mutants by growth on the Cu(I) enhancing molecule elesclomol, as well as by glial expression of wildtype swip-10. MBLAC1, the most closely related mammalian ortholog to swip-10, encodes for a pre-mRNA processing enzyme for H3 histone, a protein whose actions surprisingly include an enzymatic capacity to produce Cu(I) via the reduction of Cu(II). Moreover, genome-wide association studies and post-mortem molecular studies implicate reductions of MBLAC1 expression in risk for AD with cardiovascular disease comorbidity. Consistent with these studies, we demonstrate that the deposition of {beta}-amyloid plaques, an AD pathological hallmark, in worms engineered to express human A{beta}1-42, is greatly exaggerated by mutation of swip-10. Together, these studies identify a novel glial-expressed, and pathway for Cu(I) production that may be targeted for the treatment of AD and other neurodegenerative diseases. Significance StatementDevastating neurodegenerative diseases such as Alzheimers disease, and Parkinsons disease are associated with disruptions in copper (Cu) homeostasis. Alterations in Cu(I) give rise to increased oxidative stress burden, mitochondrial and metabolic dysfunction, and can accelerate production and/or potentiate toxicity of disease-associated protein aggregates. Here, using the model system Caenorhabditis elegans, we establish a role for the gene swip-10 in systemic Cu(I) homeostasis. Perturbation of this pathway in worms recapitulates biochemical, histological, and pathological features seen in human neurodegenerative disease. We reveal that these changes can be suppressed pharmacologically and arise when swip-10 expression is eliminated from glial cells. Our work implicates swip-10 and orthologs as key players in Cu(I) homeostasis that may be exploitable to treat multiple neurodegenerative diseases.

neuroscience↗

bioRxiv2023

Versatile Tissue-Injectable Hydrogels with Extended Hydrolytic Release of Bioactive Protein Therapeutics

Hydrogels generally have broad utilization in healthcare due to their tunable structures, high water content, and inherent biocompatibility. FDA-approved applications of hydrogels include spinal cord regeneration, skin fillers, and local therapeutic delivery. Drawbacks exist in the clinical hydrogel space, largely pertaining to inconsistent therapeutic exposure, short-lived release windows, and difficulties inserting the polymer into tissue. In this study, we engineered injectable, biocompatible hydrogels that function as a local protein therapeutic depot with a high degree of user-customizability. We showcase a PEG-based hydrogel functionalized with bioorthogonal strain-promoted azide-alkyne cycloaddition (SPAAC) handles for its polymerization and functionalization with a variety of payloads. Small-molecule and protein cargos, including chemokines and antibodies, were site-specifically modified with hydrolysable "azidoesters" of varying hydrophobicity via direct chemical conjugation or sortase-mediated transpeptidation. These hydrolysable esters afforded extended release of payloads linked to our hydrogels beyond diffusion; with timescales spanning days to months dependent on ester hydrophobicity. Injected hydrogels polymerize in situ and remain in tissue over extended periods of time. Hydrogel-delivered protein payloads elicit biological activity after being modified with SPAAC-compatible linkers, as demonstrated by the successful recruitment of murine T-cells to a mouse melanoma model by hydrolytically released murine CXCL10. These results highlight a highly versatile, customizable hydrogel-based delivery system for local delivery of protein therapeutics with payload release profiles appropriate for a variety of clinical needs. Translational ImpactWe developed injectable hydrogels that provide loco-regional, controlled release of protein therapeutics. Local delivery is especially suitable for potent, protein-based drugs like chemokines and monoclonal antibodies whose systemic toxicities can be life threatening. Our hydrogel is equipped with slowly hydrolyzing linkers for transient payload coupling, prolonging its therapeutic window and minimizing the need for repeat surgeries in a clinical setting. A range of release profiles, spanning days to over a month, and a broad compatibility to therapeutically relevant, recombinant proteins provide clinicians with flexible therapeutic options to suit a variety of circumstances.

bioengineering↗

bioRxiv2023

IFNγ modestly contributes to disease progression in the Ndufs4(-/-) model of Leigh syndrome while IP10 is dispensable

Leigh syndrome (LS) is the most common pediatric presentation of genetic mitochondrial disease. LS is a multi-system disease characterized by severe neurologic and metabolic abnormalities. The defining feature of the disease is the presence of symmetric, bilateral, progressive necrotizing lesions in the brain stem, cerebellum, and basal ganglia. The pathogenic mechanisms underlying disease initiation and progression in LS have yet to be elucidated. Recent evidence demonstrates that the immune system plays a key role in LS pathogenesis. Treatment with the macrophage-depleting Csf1r inhibitor pexidartinib prevents disease in the Ndufs4(-/-) mouse model of LS, but the mechanisms leading to immune activation and governing disease progression remain to be elucidated. In recent work, the cytokines IFN{gamma} and IFN{gamma}-induced protein 10 (IP10) were found to be significantly elevated in Ndufs4(-/-) brainstem. Given their role as macrophage-activating factors, here we sought to assess the role of IFN{gamma} and IP10 in LS using by generating Ndufs4(-/-)/Ifng(-/-) and Ndufs4(-/-)/IP10(-/-) double knockout lines. We find that IP10 alone does not significantly impact the onset or progression of disease in the Ndufs4(-/-) model, while IFN{gamma} loss significantly, but modestly, improves survival. These data indicate that IFN{gamma} contributes to pathology, but that IFN{gamma} and IP10 are both dispensable for overall disease course of LS. Our findings support some role for IFN{gamma} targeting therapies in the management of mitochondrial disease, but suggest they may provide only modest benefits, at least in LS.

pathology↗

bioRxiv2023

Tempered IL-2 Signals Program PD-1 Checkpoint Blockade-Responsive Stem-like Exhausted T Cells During Priming

Stem-like progenitor exhausted CD8 T cells are critical for maintaining long-term resistance during chronic infections and cancer, and represent an important checkpoint blockade immunotherapy target for functional reinvigoration and disease control. Hence, there is vigorous interest in understanding the ontogenesis of TCF-1Hi stem-like exhausted CD8 T cells, and in defining the signals that promote their development. Here, we show that virus-specific TCF1HiGzmBLo CD8 T cells develop during early stages of chronic viral infection, and are progressively deprogrammed from the stem-like lineage towards terminal differentiation by strong IL-2 signals. In vivo fate-tracking studies show that strong T cell-intrinsic IL-2 signals through the high affinity heterotrimeric IL-2R/{beta}/{gamma} receptor drive skewed development of terminally differentiated TCF-1LoGzmBHi cells, which largely die in chronic antigenic environment. In contrast, tempered IL-2 signals through the intermediate affinity IL-2R{beta}/{gamma} heterodimer, or delayed priming in diminished IL-2 milieu support preferential development of TCF-1Hi stem-like exhausted CD8 T cells, capable of long-term persistence and potent responsiveness to PD-1 therapy in later stages of chronic viral infection. In human tumors as well, single cell RNA-seq analyses of tumor infiltrating lymphocytes from melanoma, human papillomavirus+ head and neck cancer and lung cancer patients revealed an inverse relationship between IL-2 signaling signature and T cell stemness. Moreover, melanoma patients with enriched IL-2 signaling signature showed poor responses to checkpoint blockade immunotherapy. Collectively, these findings bear relevance to clinical immunotherapies of chronic viral infections and cancers, and support exogenous IL-2 signal manipulation prior to therapy as a strategy for augmenting long-term clinical outcomes. ONE SENTENCE SUMMARYProgramming of exhausted T cell fates by differential IL-2

immunology↗

bioRxiv2023

Peripheral macrophages causally contribute to disease onset and progression in the Ndufs4(KO) model of Leigh syndrome

Subacute necrotizing encephalopathy, or Leigh syndrome (LS), is the most common paediatric presentation of genetic mitochondrial disease. LS is a multi-system disorder with severe neurologic, metabolic, and musculoskeletal symptoms. The presence of progressive, symmetric, necrotizing lesions in the brainstem are a defining feature of the disease, and the major cause of morbidity and mortality, but the mechanisms underlying their pathogenesis have been elusive. Recently, we demonstrated that high-dose pexidartinib, a CSF1R inhibitor, prevents LS CNS lesions and systemic disease in the Ndufs4(-/-) mouse model of LS. While the dose-response in this study implicated peripheral immune cells, the immune populations involved have not yet been elucidated. Here, we used a targeted genetic tool, deletion of the colony stimulating factor 1 receptor (CSF1R) macrophage super-enhancer FIRE (Csf1r{Delta}FIRE), to specifically deplete microglia and define the role of microglia in the pathogenesis of LS. Homozygosity for the Csf1r{Delta}FIRE allele ablates microglia in both control and Ndufs4(-/-) animals, but onset of CNS lesions and sequalae in the Ndufs4(-/-), including mortality, are only marginally impacted by microglia depletion. The overall development of necrotizing CNS lesions is not altered, though microglia remain absent. Finally, histologic analysis of brainstem lesions provides direct evidence of a causal role for peripheral macrophages in the characteristic CNS lesions. These data demonstrate that peripheral macrophages play a key role in the pathogenesis disease in the Ndufs4(-/-) model.

pathology↗

bioRxiv2022

A novel fluorobenzothiazole RBx 10080758 as a dual inhibitor against bacterial GyraseB (GyrB) and Topoisomerase IV (parE) of Gram-positive pathogens causing skin and respiratory infections

Development of a novel inhibitor targeting Gyrase B and Topoisomerase IV offers a potential opportunity to combat the drug resistance. In the present study, we extensively investigated the efficacy of RBx 10080758, a novel fluorobenzothiazole, against skin and respiratory infections caused by Staphylococci and Streptococci in in vitro and in vivo models. RBx 10080758 showed a potent IC50 of 0.06 M against Gyrase and Topoisomerase IV and also exhibited a strong whole cell in vitro activity with MIC ranges of 0.015-0.06, 0.015-0.03, 0.008-0.03, 0.008-0.03, 0.008-0.0.06, 0.015-0.06 g/ml against Staphylococcus. aureus, Streptococcus pneumoniae, Coagulase negative Staphylococci, Streptococcus viridans, Streptococcus pyogenes and Enterococcus, respectively. As expected from the novel class of molecule, it retains potent even against linezolid and vancomycin resistant strains. Interesting, in mouse systemic infection 10 mg/kg, IV dose protected 100% mice from lethal infections. In rat thigh infection model with MRSA WCUH29 at 45 mg/kg exhibited >3-log10 cfu reduction in thigh muscles. RBx 10080758 displayed potent in vitro and in vivo activity against a panel of MDR Gram-positive bacteria. As a novel chemical class, the fluorobenzothiazoles have the potential to become clinically viable antibiotics, to address the drug resistance problem by its unique dual targeting mechanism of action.

microbiology↗

bioRxiv2022

Novel modification of Luminex assay for characterization of extracellular vesicle populations in biofluids.

Most approaches to extracellular vesicle (EV) characterization focus on EV size or density. However, such approaches provide few clues regarding EV origin, molecular composition, and function. New methods to characterize the EV surface proteins may aid our understanding of their origin, physiological roles, and biomarker potential. Recently developed immunoassays for intact EVs based on ELISA, NanoView, SIMOA and MesoScale platforms are highly sensitive, but have limited multiplexing capabilities, whereas MACSPlex FACS enables the detection of multiple EV surface proteins, but requires significant quantities of purified EVs, which limits its adoption. Here, we describe a novel Luminex-based immunoassay, which combines multiplexing capabilities with high sensitivity and, importantly, bypasses the enrichment and purification steps that require larger sample volumes. We demonstrate the methods specificity for detecting EV surface proteins using multiple EV depletion techniques, EVs of specific cellular origin isolated from culture media, and by co-localization with established EV surface markers. Using this novel approach, we elucidate differences in the tetraspanin profiles of the EVs carrying erythrocyte and neuron markers. Using size exclusion chromatography, we show that plasma EVs of putative neuronal and tissue macrophage origin are eluted in fractions distinct from those derived from erythrocytes, or from their respective cultured cells. In conclusion, our novel multiplexed assay differentiates between EVs from erythrocytes, macrophages, and neurons, and offers a new means for capture, classification, and profiling of EVs from diverse sources.

cell biology↗

bioRxiv2022

Evidences for newer insecticides and growth regulators resistance in field populations of Helicoverpa armigera (Lepidoptera: Noctuidae) in Karnataka, India

The prominence of Helicoverpaarmigera(Lepidoptera: Noctuidae)averse insecticide resistance was traversed in the course of 2017 in Karnataka,India. The results divulged typical resistance level prostrating in selected newer insecticides, even though exiguous higher resistance airing in insect growth regulator Novaluranwith LC 50of and 13.02 to 18.07 ppm and 1.17 to 1.95 folds resistance) compared to newer group insecticides Spinasad, Indoxacarb, Flubendiamide and Rynaxypyr(3.57 to 10.19 ppm, 1.01 to 1.27 fold). Raichur and Kalaburgi strains comprehend higher resistance to Novaluran and newer insecticides with exception of Flubendiamide (Raichur and Vijayapura strains), respectively and Spinosad (Kalaburgi and Raichur strains), respectively. The morphometric parameters of larval length, pupal length, and weight were most in RCH stain (2.75{+/-}0.48 cm, 1.76{+/-}0.18 cm, and 0.511{+/-}0.04 g, 0.309{+/-}0.05 g) respectively, which was pursued by Kalaburgi strain. The morphometric correlation revealed that larval length was a significant positive relation with insecticide resistance which might be an influence of resistance but not merely responsible. Among newer insecticides, a significant positive correlation between Rynaxypyr and Indoxacarb was evident, similarly, Nuvaluran with Indoxacarb and Rynaxypyr as well. Usage pattern revealed that 81.67 % of farmers found to use insecticides more than the recommended dose and 70.83% have habit consecutive applications of products from the same chemical group which bear witness to developing resistance.

ecology↗