Renal-resident macrophages (RMs) are essential regulators of kidney homeostasis and repair, yet the mechanisms governing RM niche regeneration after acute depletion remain poorly defined. To overcome these limitations, we have developed an inducible human CD59- intermedilysin (hCD59-ILY) ablation system, enabling rapid, specific, and reversible depletion of targeted macrophage populations, and subsequent replenishment of RMs, followed by longitudinal scRNA-seq analysis of kidneys at baseline and days 1, 3, and 7 post-ablation. RM ablation triggered a rapid and sustained upregulation of Cx3cl1, predominantly in proximal tubular epithelial cells (PTC1/PTC2), establishing a persistent chemotactic niche signal that coincided with macrophage repopulation. Regenerating RMs transitioned from inflammatory/stress-associated states toward metabolically active and proliferative phenotypes enriched in glycolysis, oxidative phosphorylation, MYC, and cell-cycle programs, with attenuation of canonical inflammatory pathways. Cell-cell communication analysis revealed an early burst of intercellular signaling at day 1, followed by progressive normalization, with fibronectin (Fn1), osteopontin (Spp1), chemokine (Ccl), and amyloid precursor protein (App) axes emerging as key mediators of niche restoration. Transcriptional network analysis identified a conserved regulatory module (Tfe3, Mitf, Hif1a, Myc, Gabpa, Rcor1) coordinating macrophage differentiation and regenerative programming, linking metabolic adaptation to lineage reconstitution. Sub-clustering revealed five dynamically shifting RM subsets with distinct inflammatory, remodeling, proliferative, and surveillance states, reflecting a hierarchical regeneration process. Functional validation using clodronate-mediated depletion in Secreted Phosphoprotein 1 (Spp1) (Opn)-deficient mice demonstrated impaired macrophage repopulation, establishing osteopontin as a critical regulator of RM regeneration. Together, these data define a coordinated epithelial-immune circuit in which Cx3cl1-driven chemotaxis, Spp1-dependent signaling, and a core transcriptional network orchestrate macrophage niche reconstitution and kidney repair following acute immune cell ablation.

Introduction:Chikungunya virus (CHIKV) infection causes acute febrile illness and severe joint inflammation, with some patients progressing to chronic arthritis. Early isotype-switched neutralizing antibody (nAb) responses have been associated with protection from chronic arthritis, but the cellular and molecular features of these early responses remain poorly defined. Methods:We performed single-cell sorting of plasmablasts (PBs) from patients with acute CHIKV infection and generated 94 human monoclonal antibodies (mAbs). These were evaluated for binding to CHIKV-infected Vero cells and to purified CHIKV, neutralizing activity, isotype distribution, somatic hypermutation (SHM), and immunoglobulin gene usage. PB-derived antibodies were compared with CHIKV-specific memory B cell (MBC)-derived antibodies from seropositive individuals. Results:Over one-third of PB-derived mAbs recognized CHIKV-infected Vero cells, and most were class-switched. Among these, 12 bound to purified CHIKV, and 6 of them showed neutralizing activity. All PB-derived nAbs had germline-like sequences with little to no SHM, and four displayed shared clonotypic features, including a conserved `WEL' motif in the CDRH3 region. Conversely, CHIKV-specific MBC-derived nAbs showed diverse but often higher levels of SHM, consistent with affinity maturation. Discussion:Together, our findings suggest that early nAb responses generated during acute CHIKV infection are germline-biased with overlapping features suggestive of convergent clonotype responses, providing a cellular and molecular basis of the early protective humoral immunity.

Abstract Influenza A virus (IAV) remains a major cause of respiratory morbidity and mortality, yet the role of Toll-like receptor 7 (TLR7), an RNA sensor, and its downstream signaling events, such as interferon regulatory factor 7 (IRF7), in IAV infection remain unclear. To address this question, we used single-cell RNA sequencing, genetic mouse models, and immunological analysis. Single -cell transcriptomic profiling of the infected lungs revealed robust upregulation of Tlr7 and genes associated with interferon pathways in dendritic cells and B cells, alongside widespread induction of Irf7 across immune and non-immune compartments. Tlr7 -deficient mice exhibited normal viral control, lung pathology, and survival following IAV challenge. In contrast, Irf7 deficiency resulted in significantly increased disease severity, impaired early interferon responses, exacerbated bronchial epithelial hyperplasia, and defective early humoral priming. In assessing adaptive immunity, both Irf7 -deficient and Tlr7 -deficient mice had reduced antihemagglutinin antibody production. Mechanistically, IRF7 protein expression and downstream signaling were largely preserved in TLR7-deficient mice, indicating that IRF7 activation during IAV infection occurs independently of TLR7. Collectively, these findings identify IRF7 as a non-redundant determinant of innate immunity and disease outcomes during IAV infection, while positioning TLR7 as a modulatory factor primarily influencing adaptive immune maturation. Our study refines current models of antiviral sensing by uncoupling receptor induction from functional necessity and highlights IRF7 as a critical downstream regulator dictating host defense against acute influenza A infection. Importance Influenza A virus is a respiratory pathogen that remains a major threat to global health as a seasonal disease and a source of periodic pandemics. The outcomes of the infection can range from mild illness to severe pneumonia and death, particularly in vulnerable populations, yet the reasons why some individuals develop more severe disease are not fully understood. Early immune defenses in the lungs are critical for controlling the virus, but they can also contribute to harmful inflammation if not properly regulated. In particular, key sensors that detect viral genetic material and the signaling pathways that activate antiviral responses play an essential role in shaping these outcomes. The significance of our study lies in defining how these early immune mechanisms influence the course of influenza A infection, providing insight that may guide the development of improved therapies for influenza and related respiratory viruses.

Toll-like receptor 7 (TLR7) is a key endosomal sensor that detects single-stranded RNA, linking innate and adaptive immunity through the induction of type I interferons and proinflammatory cytokines. Recent studies have underscored the pivotal role of TLR7 in shaping immune responses to respiratory viral infections, including SARS-CoV-2, influenza A virus, and respiratory syncytial virus (RSV), as well as in the pathogenesis of systemic autoimmune diseases such as systemic lupus erythematosus (SLE), which can be triggered by the respiratory viral infections. In COVID-19, TLR7 deficiency is associated with severe disease, particularly in males, due to impaired interferon responses and antibody production. In influenza, TLR7 enhances humoral and cytotoxic responses, though its overactivation may contribute to immunopathology. The role of TLR7 in RSV remains controversial, with both protective and detrimental effects reported depending on host and experimental context. In contrast, TLR7 plays a pathogenic role in SLE by amplifying type I interferon signaling and promoting autoreactive B cell activation. This review synthesizes current knowledge on TLR7-mediated signaling across these diseases, highlighting its context-dependent functions and dualistic nature in immunity and disease. We will discuss mechanistic insights, clinical relevance, and emerging therapeutic strategies targeting TLR7, emphasizing the need for precision modulation of this pathway in the treatment of viral infections and autoimmune disorders.

Introduction:SARS-CoV-2 and, to a lesser extent, influenza A can lead to long-term complications in the respiratory and nervous systems. However, the mechanisms driving post-viral sequelae remain poorly understood. Methods:To address this gap, we longitudinally characterized C57BL/6 mice infected with sublethal doses of mouse-adapted SARS-CoV-2 (MA30) or influenza A (PR8). Lung and brain tissues were analyzed at 14-, 21-, and 28-days post-infection (DPI) using histological analysis and bulk-RNA sequencing. Results:In the lungs, both infections caused prolonged inflammation and fibrosis. MA30-infected lungs showed persistent upregulation of inflammation, coagulation, complement, as well as fibrotic, and extracellular matrix (ECM) remodeling pathways at 21 DPI, alongside downregulation of epithelial junction and metabolic program pathways. In contrast, PR8-infected lungs exhibited a strong acute interferon response and chronic upregulation of basal epithelial markers (e.g., Krt5, Krt14), consistent with epithelial regeneration. Notably, only PR8-infected mice displayed KRT5+ progenitor cell migration into damaged lung regions, indicating divergence in repair mechanisms. Neither MA30-infected, nor PR8-infected mice had detectable brain infection. However, MA30 mice, but not PR8-infected mice exhibited an elevated frequency of microhemorrhages at early timepoints and marked neuroinflammation at all timepoints. Transcriptomic profiling of MA30-infected brains showed enrichment for up-regulation of ECM remodeling, vascular dysfunction, IL6-signaling pathways along with a virus-specific disruption of the hypothalamic-pituitary axis with MA30 infection not seen in PR8-infected brains. These included genes linked to neuroinflammation, sensory processing disruption, and microvascular injury, mirroring clinical features of Long COVID. Discussion:Together, these findings establish distinct tissue-specific trajectories of long-term pathology following SARS-CoV-2 and influenza infection and provide a foundation for dissecting the mechanisms of post-viral lung and brain disease.

Objective:Systemic sclerosis (SSc) is a severe autoimmune disease characterized by immune dysregulation, fibrosis, and substantial morbidity and mortality. Although type I interferon-related pathways have been implicated in SSc, the contribution of Toll-like receptor 7 (TLR7) and its downstream signaling, including the transcription factor interferon regulatory factor 7 (IRF7) and effector molecules such as CCL2 and CCL12, to disease pathogenesis remains unclear. Methods:We used a bleomycin (BLM)-induced mouse model of SSc. Male wild-type (WT), Tlr7-deficient (Tlr7-/- ), and Irf7-deficient (Irf7-/- ) mice (10-12 weeks old) received daily subcutaneous BLM (2.5 mg/kg/day) for 4 weeks. In a separate experiment, BLM-treated WT mice were treated with the CCR2 antagonist (RS504393). Disease features were evaluated by histopathology, ELISA, flow cytometry, western blotting, RT-qPCR, and Olink proteomics. Results:BLM-treated Tlr7-/- and Irf7-/- mice showed less body weight loss, reduced pulmonary interstitial inflammation, fewer inflammatory monocytes in the spleen, lower pulmonary type I interferon-related gene expression, and lower serum anti-topoisomerase I autoantibody (anti-Scl-70) levels. Pharmacologic CCR2 antagonism attenuated BLM-induced body weight loss, lung injury, and reduced dermal collagen deposition. Conclusion:These findings support a pathogenic role for TLR7-IRF7-IFN-I signaling in anti-Scl-70 autoantibody production and pulmonary inflammation in experimental SSc, and suggest that downstream chemokine pathways may represent therapeutic targets.

Background:Renal-resident macrophages (RMs) are essential regulators of kidney homeostasis and repair, yet the cellular and molecular mechanisms governing RM niche regeneration after acute depletion remain poorly defined. How epithelial-immune interactions coordinate RM repopulation is particularly unclear. Methods:We employed an inducible hCD59 intermedilysin (ILY) ablation system to achieve rapid and specific depletion, and subsequent replenishment of RMs, followed by longitudinal single-cell RNA sequencing (scRNA-seq) of kidneys at baseline and days 1, 3, and 7 post-ablation. Integrated transcriptomic, pathway, transcription factor, and cell-cell communication analyses were combined with functional validation using clodronate-mediated macrophage depletion in Spp1 (Opn)-deficient mice. Results:Acute ILY-mediated ablation resulted in rapid and selective RM depletion, followed by robust regeneration reaching ~ 75% of baseline by day 7. scRNA-seq faithfully captured RM loss and recovery and revealed a sustained epithelial-derived chemotactic response, with proximal tubule epithelial cells identified as the dominant source of CX3CL1 driving RM recruitment and maintenance. Regenerating macrophages adopted a transient injury-adaptive transcriptional program characterized by metabolic activation, proliferation, and stress-response pathways, with relative attenuation of canonical inflammatory signaling. Cell-cell communication analysis identified macrophages as dominant signaling hubs, coordinating immune and epithelial responses through temporally regulated Spp1, Fn1, Ccl, and App-mediated networks. Functional studies demonstrated that Spp1/osteopontin is required for efficient RM regeneration following depletion. SCENIC-STRING analysis connected 18 upregulated transcription factors (TFs) to IL-1 signaling, myeloid differentiation, and tissue remodeling, indicating a coordinated transcriptional program driving RM regeneration. Sub-clustering uncovered five RM subsets and ten proximal tubule cell states with dynamic, time-dependent shifts, revealing a hierarchical macrophage-epithelial communication program that orchestrates niche restoration and tubular repair. Conclusion:Our study defines RM regeneration as a transcriptionally regulated, communication-driven process orchestrated by epithelial-derived chemokines, macrophage metabolic reprogramming, and subtype-specific signaling networks. These findings revealed a hierarchical macrophage-epithelial communication program coordinating RM niche restoration and tubular repair.

It remains unclear whether podocyte loss directly causes acute renal tubular cell (RTC) damage and interstitial fibrosis, thereby leading to renal failure. Here, we applied intermedilysin (ILY)-mediated human CD59 (hCD59) cell ablation to generate an acute, specific podocyte-ablation mouse model. Cre-induced hCD59 transgenics (ihCD59) were crossed with Nphs2Cre to generate ihCD59 +/- /Nphs2Cre +/- mice. The specific and rapid podocyte-ablation mediated by ILY injection directly caused RTC necrosis, leading to renal failure and even death within 2-3 days in a dose-dependent manner. Treating mice that received an ILY lethal dose with peritoneal dialysis or administering a non-lethal dose, we extended their survival beyond six weeks and found that mice developed interstitial fibrosis and glomerulosclerosis with persistent proteinuria and tubule damage. Podocyte-ablation caused massive disruption of glomerular function at week 1, and then partial recovery by week 2. Genes and pathways of TLRs and apoptosis, and mitochondrial functions were respectively upregulated and downregulated in both ablated-podocyte mouse and biopsied-glomerulonephritis patient kidney samples. Together, this rapid podocyte-ablation causes acute RTC necrosis that progresses to interstitial fibrosis in this mouse model, which is applicable for dissecting mechanisms underlying podocyte injury-mediated tubular damage and glomerular repair, with the potential to reveal novel therapeutic targets for kidney diseases.

Renal macrophages (RMs) are essential for kidney health, orchestrating immune surveillance, tissue homeostasis, and responses to injury. Previously, we reported the use of a human CD59 (hCD59)/intermedilysin (ILY) cell ablation tool to study the distinct fate, dynamics, and niches of RMs of bone marrow or embryonic origin. RMs originate from yolk sac-derived macrophages, fetal liver monocytes, and bone marrow-derived monocytes and are maintained in adulthood through local proliferation and recruitment of circulating monocytes. Here, we report a detailed protocol for the selective ablation of RMs to study their regeneration, including 1) generation and characterization of the Cre-inducible expression of hCD59 in mouse RMs, 2) purification of ILY and characterization of ILY activity, 3) induction of hCD59 expression on RMs in compound mice, and 4) characterization of regeneration after ILY-mediated RM ablation. ILY specifically and rapidly depletes RMs in compound mice, with efficient macrophage ablation within 1 day of ILY administration. Renal macrophage regeneration began by day 3 post-ablation, with similar to 88% recovery by day 7. This model offers a powerful tool for studying macrophage biology and can be used for selectively ablating other cell populations in the kidney, liver, and fatty tissues to investigate their function and regeneration.

A mouse model of HIV-associated atherosclerosis (Tg26+/-ApoE-/-) exhibited increased plaque area compared with the ApoE-/- mouse, linked to elevated indoleamine 2,3-dioxygenase (IDO) activity. IDO catalyses the conversion of tryptophan (TRP) into kynurenine (KYN), measured by the KYN-to-TRP ratio. As a biomarker of inflammation, IDO has been implicated as a risk factor for cardiovascular disease. To investigate the effect of exercise training on atherogenesis and IDO activity in Tg26+/-ApoE-/- mice, nine Tg26+/-ApoE-/- and 18 ApoE-/- male mice were fed an atherogenic diet and randomized into exercised or control groups. The exercised groups underwent an 8-week treadmill protocol at moderate intensity (five times per week at 60% maximum velocity). Concentrations of KYN, TRP and cytokines were measured using ELISA, immune expression by flow cytometry, and lipid profile by a biochemistry analyser. Aortas were harvested post mortem for en face analysis. Tg26+/-ApoE-/- mice showed ∼40% larger plaques than ApoE-/- mice (P = 0.01), with slightly higher neutrophil (P = 0.05) and monocyte expression (P = 0.06). Plaque area was reduced by 40% in exercised ApoE-/- mice (P = 0.04), but by only 12% in exercised Tg26+/-ApoE-/- animals (P = 0.85). Exercised Tg26+/-ApoE-/- mice showed higher IDO activity than exercised ApoE-/- mice (58.57% ± 6.88% vs. -4.62% ± 17.20%, P = 0.01), which was positively correlated with plaque area (R = 0.99, P = 0.02). Exercised ApoE-/- mice showed significantly lower triglyceride levels compared with exercised Tg26+/-ApoE-/- mice (75.8 ± 14.8 vs. 165.2 ± 43.6 mg/dL; P = 0.02). Unlike ApoE-/- mice, moderate-intensity aerobic training did not reduce plaque area in mice with HIV-associated atherosclerosis. Moreover, exercise training appeared to increase inflammation in Tg26+/-ApoE-/- mice, as indicated by elevated IDO activity.

Renal macrophages (RMs) are essential for kidney health, orchestrating immune surveillance, tissue homeostasis, and responses to injury. Previously, we reported the use of a human CD59 (hCD59)/intermedilysin (ILY) cell ablation tool to study the distinct fate, dynamics, and niches of RMs of bone marrow or embryonic origin. RMs originate from yolk sac-derived macrophages, fetal liver monocytes, and bone marrow-derived monocytes and are maintained in adulthood through local proliferation and recruitment of circulating monocytes. Here, we report a detailed protocol for the selective ablation of RMs to study their regeneration, including 1) generation and characterization of the Cre-inducible expression of hCD59 in mouse RMs, 2) purification of ILY and characterization of ILY activity, 3) induction of hCD59 expression on RMs in compound mice, and 4) characterization of regeneration after ILY-mediated RM ablation. ILY specifically and rapidly depletes RMs in compound mice, with efficient macrophage ablation within 1 day of ILY administration. Renal macrophage regeneration began by day 3 post-ablation, with ~88% recovery by day 7. This model offers a powerful tool for studying macrophage biology and can be used for selectively ablating other cell populations in the kidney, liver, and fatty tissues to investigate their function and regeneration.

Emerging evidence indicates that activation of complement system leading to the formation of the membrane attack complex (MAC) plays a detrimental role in COVID-19. However, their pathogenic roles have never been experimentally investigated before. We used three knock out mice strains (1. C3-/-; 2. C7-/-; and 3. Cd59ab-/-) to evaluate the role of complement in severe COVID-19 pathogenesis. C3 deficient mice lack a key common component of all three complement activation pathways and are unable to generate C3 and C5 convertases. C7 deficient mice lack a complement protein needed for MAC formation. Cd59ab deficient mice lack an important inhibitor of MAC formation. We also used anti-C5 antibody to block and evaluate the therapeutic potential of inhibiting MAC formation. We demonstrate that inhibition of complement activation (in C3-/-) and MAC formation (in C3-/-. C7-/-, and anti-C5 antibody) attenuates severe COVID-19; whereas enhancement of MAC formation (Cd59ab-/-) accelerates severe COVID-19. The degree of MAC but not C3 deposits in the lungs of C3-/-, C7-/- mice, and Cd59ab-/- mice as compared to their control mice is associated with the attenuation or acceleration of SARS-CoV-2-induced disease. Further, the lack of terminal complement activation for the formation of MAC in C7 deficient mice protects endothelial dysfunction, which is associated with the attenuation of diseases and pathologic changes. Our results demonstrated the causative effect of MAC in severe COVID-19 and indicate a potential avenue for modulating the complement system and MAC formation in the treatment of severe COVID-19.

This study investigates the roles of T, B, and Natural Killer (NK) cells in the pathogenesis of severe COVID-19, utilizing mouse-adapted SARS-CoV-2-MA30 (MA30). To evaluate this MA30 mouse model, we characterized MA30-infected C57BL/6 mice (B6) and compared them with SARS-CoV-2-WA1 (an original SARS-CoV-2 strain) infected K18-human ACE2 (K18-hACE2) mice. We found that the infected B6 mice developed severe peribronchial inflammation and rapid severe pulmonary edema, but less lung interstitial inflammation than the infected K18-hACE2 mice. These pathological findings recapitulate some pathological changes seen in severe COVID-19 patients. Using this MA30-infected mouse model, we further demonstrate that T and/or B cells are essential in mounting an effective immune response against SARS-CoV-2. This was evident as Rag2−/− showed heightened vulnerability to infection and inhibited viral clearance. Conversely, the depletion of NK cells did not significantly alter the disease course in Rag2−/− mice, underscoring the minimal role of NK cells in the acute phase of MA30-induced disease. Together, our results indicate that T and/or B cells, but not NK cells, mitigate MA30-induced disease in mice and the infected mouse model can be used for dissecting the pathogenesis and immunology of severe COVID-19.

Toll-like receptor 7 (Tlr7) deficiency-accelerated severe COVID-19 is associated with reduced production of interferons (IFNs). However, the underlying mechanisms remain elusive. To address these questions, we utilize Tlr7 and Irf7 deficiency mice, single-cell RNA analysis together with bone marrow transplantation approaches. We demonstrate that at the early phase of infection, SARS-CoV-2 causes the upregulation of Tlr7, Irf7, and IFN pathways in the lungs of the infected mice. The deficiency of Tlr7 and Irf7 globally and/or in immune cells in mice increases the severity of COVID-19 via impaired IFN activation in both immune and/or non-immune cells, leading to increased lung viral loads. These effects are associated with reduced IFN alpha and gamma levels in the circulation. The deficiency of Tlr7 tends to cause the reduced production and nuclear translocation of interferon regulatory factor 7 (IRF7) in the lungs of the infected mice, indicative of reduced IRF7 activation. Despite higher amounts of lung viral antigen, Tlr7 or Irf7 deficiency resulted in substantially reduced production of antibodies against SARS-CoV-2, thereby delaying the viral clearance. These results highlight the importance of the activation of TLR7 and IRF7 leading to IFN production on the development of innate and adaptive immunity against COVID-19.

Macrophages are exceptionally diversified cell types and perform unique features and functions when exposed to different stimuli within the specific microenvironment of various kidney diseases. In instances of kidney tissue necrosis or infection, specific patterns associated with damage or pathogens prompt the development of pro-inflammatory macrophages (M1). These M1 macrophages contribute to exacerbating tissue damage, inflammation, and eventual fibrosis. Conversely, anti-inflammatory macrophages (M2) arise in the same circumstances, contributing to kidney repair and regeneration processes. Impaired tissue repair causes fibrosis, and hence macrophages play a protective and pathogenic role. In response to harmful stimuli within the body, inflammasomes, complex assemblies of multiple proteins, assume a pivotal function in innate immunity. The initiation of inflammasomes triggers the activation of caspase 1, which in turn facilitates the maturation of cytokines, inflammation, and cell death. Macrophages in the kidneys possess the complete elements of the NLRP3 inflammasome, including NLRP3, ASC, and pro-caspase-1. When the NLRP3 inflammasomes are activated, it triggers the activation of caspase-1, resulting in the release of mature proinflammatory cytokines (IL)-1β and IL-18 and cleavage of Gasdermin D (GSDMD). This activation process therefore then induces pyroptosis, leading to renal inflammation, cell death, and renal dysfunction. The NLRP3–ASC–caspase-1–IL-1β–IL-18 pathway has been identified as a factor in the development of the pathophysiology of numerous kidney diseases. In this review, we explore current progress in understanding macrophage behavior concerning inflammation, injury, and fibrosis in kidneys. Emphasizing the pivotal role of activated macrophages in both the advancement and recovery phases of renal diseases, the article delves into potential strategies to modify macrophage functionality and it also discusses emerging approaches to selectively target NLRP3 inflammasomes and their signaling components within the kidney, aiming to facilitate the healing process in kidney diseases.

Introduction Chikungunya is caused by an alpha virus transmitted to humans by an infected mosquito. Infection is generally considered to be self-limiting and non-critical. Chikungunya infection may be diagnosed by severe joint pain with fever, but it is difficult to diagnose because the symptoms of chikungunya are common to many pathogens, including dengue fever. Diagnosis mainly depends on viral culture, reverse transcriptase polymerase chain reaction (RT-PCR), and IgM ELISA. Early and accurate diagnosis of the virus can be achieved by the application of PCR methods, but the high cost and the need for a thermal cycler restrict the use of such methods. On the other hand, antibody-based IgM ELISA is considered to be inexpensive, but antibodies against chikungunya virus (CHIKV) only develop after 4 days of infection, so it has limited application in the earlier diagnosis of viral infection and the management of patients. Because of these challenges, a simple antigen-based sensitive, specific, and rapid detection method is required for the early and accurate clinical diagnosis of chikungunya. Methods The amino acid sequence of CHIKV ectodomain E1 and E2 proteins was analyzed using bioinformatics tools to determine the antigenic residues, particularly the B-cell epitopes and their characteristics. Recombinant E2-E1 CHIKV antigen was used for the development of polyclonal antibodies in hamsters and IgG was purified. Serological tests of 96 CHIKV patients were conducted by antigen-capture ELISA using primary antibodies raised against rCHIKV E2-E1 in hamsters and human anti-CHIKV antibodies. Results We observed high specificity and sensitivity, of 100% and 95.8%, respectively, and these values demonstrate the efficiency of the test as a clinical diagnostic tool. There was no cross-reactivity with samples taken from dengue patients. Discussion Our simple and sensitive sandwich ELISA for the early-phase detection of CHIKV infection may be used to improve the diagnosis of chikungunya.

The novel coronavirus SARS-CoV-2, responsible for the COVID-19 outbreak, has become a pandemic threatening millions of lives worldwide. Recently, several vaccine candidates and drugs have shown promising effects in preventing or treating COVID-19, but due to the development of mutant strains through rapid viral evolution, urgent investigations are warranted in order to develop preventive measures and further improve current vaccine candidates. Positive-sense-single-stranded RNA viruses comprise many (re)emerging human pathogens that pose a public health problem. Our innate immune system and, in particular, the interferon response form an important first line of defense against these viruses. Flexibility in the genome aids the virus to develop multiple strategies to evade the innate immune response and efficiently promotes their replication and infective capacity. This review will focus on the innate immune response to SARS-CoV-2 infection and the virus' evasion of the innate immune system by escaping recognition or inhibiting the production of an antiviral state. Since interferons have been implicated in inflammatory diseases and immunopathology along with their protective role in infection, antagonizing the immune response may have an ambiguous effect on the clinical outcome of the viral disease. This pathology is characterized by intense, rapid stimulation of the innate immune response that triggers activation of the Nod-like receptor family, pyrin-domain-containing 3 (NLRP3) inflammasome pathway, and release of its products including the pro-inflammatory cytokines IL-6, IL-18, and IL-1β. This predictive view may aid in designing an immune intervention or preventive vaccine for COVID-19 in the near future.

Visceral leishmaniasis (VL) or kala-azar is a vector-borne dreaded protozoal infection that is caused by the parasite Leishmania donovani. With increases in the dramatic infection rates, present drug toxicity, resistance, and the absence of an approved vaccine, the development of new antileishmanial compounds from plant sources remains the keystone for the control of visceral leishmaniasis. In this study, we evaluated the leishmanicidal effect of thymoquinone against L. donovani with an in vitro and ex vivo model. Thymoquinone exhibited potent antipromastigote activity with IC50 and IC90 concentrations achieved at 6.33 ± 1.21 and 20.71 ± 2.15 μM, respectively, whereas the IC50 and IC90 concentrations were found to be 7.83 ± 1.65 and 27.25 ± 2.20 μM against the intramacrophagic form of amastigotes, respectively. Morphological changes in promastigotes and growth reversibility study following treatment confirmed the leishmanicidal effect of thymoquinone. Further, thymoquinone exhibited leishmanicidal activities against L. donovani promastigote through cytoplasmic shrinkage, membrane blebbing, chromatin condensation, cellular and nuclear shrinkage, and DNA fragmentation, as observed under scanning and transmission electron microscopy analyses. The antileishmanial activity was exerted via programmed cell death as proved by exposure of phosphatidylserine, DNA nicking by TUNEL assay, and loss of mitochondrial membrane potential. Thymoquinone at a concentration of 200 μM was devoid of any cytotoxic effects against mammalian macrophage cells. Thymoquinone showed strong leishmanicidal activity against L. donovani, which is mediated via an apoptosis mode of parasitic cell death, and accordingly, thymoquinone may be the source of a new lead molecule for the cure of VL.

The re-emergence of Chikungunya virus (CHIKV) infection in humans with no approved antiviral therapies or vaccines is one of the major problems with global significance. In the present investigation, we screened 80 in-house quinoline derivatives for their anti-CHIKV activity by computational techniques and found 4-hydroxy-1-methyl-3-(3-morpholinopropanoyl)quinoline-2(1H)-one (QVIR) to have potential binding affinities with CHIKV nsP2 and E2 glycoproteins. QVIR was evaluated in vitro for its anti-CHIKV potential. QVIR showed strong inhibition of CHIKV infection with an EC50 (50% effective concentration) value of 2.2 ± 0.49 μM without significant cytotoxicity (CC50 > 200 μM) and was chosen for further elucidation of its antiviral mechanism. The infectious viral particle formation was abolished by approximately 72% at a QVIR concentration of 20 μM during infection in the BHK-21 cell line, and the CHIKV RNA synthesis was diminished by 84% for nsP2 as well as 74% for E2, whereas the levels of viral proteins were decreased by 69.9% for nsP2 and 53.9% for E2. Flow cytometry analysis confirmed a huge decline in the expression of viral nsP2 and E2 proteins by 71.84 and 67.7%, respectively. Time of addition experiments indicated that QVIR inhibited viral infection at early and late stages of viral replication cycle, and the optimal inhibition was observed at 16 h post infection. The present study advocates for the first time that QVIR acts as a substantial and potent inhibitor against CHIKV and might be as an auspicious novel drug candidate for the development of therapeutic agents against CHIKV infections.

Visceral leishmaniasis (VL) is caused by a protozoan parasite, Leishmania donovani (L. donovani). It affects around 1–2 million people around the world annually. There is an urgent need to investigate new medicament of it due to difficult method of drug administration, long period of treatment, high cost of the drug, adverse side-effects, low efficacy and development of parasite resistance to the available drugs. Medicinal plants have also been used for the treatment of different diseases in traditional system of medicines due to their holistic effects. The Drugs for Neglected Diseases initiative (DNDi), Geneva, Switzerland has already started the program for identification of potential medicinal plant and plant products having antileishmanial potential. Keeping all these in consideration, we planned to study the antileishmanial activity of one of the medicinal plant, Embilica officinalis L. (EO) fruit extract. EO fruit extract inhibited the growth and proliferation of promastigotes as well as intra-macrophagic amastigotes in dose-dependent manner. EO fruit extract induced morphological and ultrastructural changes in parasites as observed under Electron Microscope. It also induced the oxidative stress, mitochondrial dysfunction, DNA laddering and apotosis-like cell death in parasites. Here, we for the first time reported such a detailed mechanism of action of antileishmanial activity of EO fruit extract. Our results suggested that EO fruit extract could be used for the development of new phytomedicine against leishmaniasis.