Innate Immune Defense Research Articles

Molecular characterization of two newly identified glycine rich crustin genes (Mn-Gly-Cru3, Mn-Gly-Cru4) and their function in Macrobrachium nipponense innate immunity

Crustin, as an important antibacterial peptide in crustacean, plays a role in the defense against the invasion of foreign pathogens in the innate immune system. In this study, two glycine rich crustin genes (named as Mn-Gly-Cru3 and Mn-Gly-Cru4) were cloned in Macrobrachium nipponense. The full-length cDNA of Mn-Gly-Cru3 and Mn-Gly-Cru4 were 740 and 781 bp with open reading frames of 525 and 576 bp that encode 174 and 191 amino acids, respectively. The genome structure of Mn-Gly-Cru3 and Mn-Gly-Cru4 shows that both contain two exons and one intron. Both Mn-Gly-Cru3 and Mn-Gly-Cru4 protein contains a signal peptide, a cysteine-rich region, a glycine-rich region, and a whey acidic protein domain. Evolutionary analysis showed that Mn-Gly-Cru1, Mn-Gly-Cru2, Mn-Gly-Cru3 and Mn-Gly-Cru4 were clustered together. Mn-Gly-Cru3 and Mn-Gly-Cru4 were widely distributed in multiple tissues. After white spot syndrome virus (WSSV), Staphylococcus aureus or Vibrio parahaemolyticus challenge, the expression levels of Mn-Gly-Cru3 and Mn-Gly-Cru4 in the hemocytes, gills, and stomach were significantly changed. Mn-Gly-Cru3 and Mn-Gly-Cru4 knockdown remarkably increased the copy number of WSSV and the number of S. aureus and V. parahaemolyticus colonies in vivo. Purified rMn-Gly-Cru3 and rMn-Gly-Cru4 proteins could bind to a variety of bacteria and they also had binding activity toward LPS or PGN. Furthermore, rMn-Gly-Cru4 had a stronger binding activity to polysaccharides than rMn-Gly-Cru3. Our study indicates that Mn-Gly-Cru3 and Mn-Gly-Cru4 in M. nipponense are involved in the innate immune defense against bacteria or viruses.

Immune functions of C-type lectins in medical arthropods.

C-type lectins (CTLs) are a family of proteins that contain 1 or more carbohydrate-recognition domains (CRDs) and bind to a broad repertoire of ligands in the presence of calcium ions. CTLs play important roles in innate immune defenses against microorganisms by acting as pattern-recognition receptors (PRRs) for invading pathogens, such as bacteria, viruses, and parasites. After binding to pathogen-associated ligands, CTLs mediate immune responses, such as agglutination, phagocytosis, and the activation of phenol oxidase progenitors, thereby clearing pathogens. CTLs are an evolutionarily conserved family found in almost all vertebrates and invertebrates. Medical arthropods can acquire and transmit a range pathogens through various approaches, such as bloodsucking, lancing, and parasitism, thus infecting humans and animals with related diseases, some of which can be life-threatening. Recent studies have shown that lectins are important components of the arthropod immune system and are essential for the immune responses of arthropods to arthropod-borne pathogens. This article reviews the current understanding of the structure, function, and signaling pathways involved in CTLs derived from important medical arthropods.

Endoplasmic reticulum-associated protein degradation contributes to Toll innate immune defense in Drosophila melanogaster.

In Drosophila, the endoplasmic reticulum-associated protein degradation (ERAD) is engaged in regulating pleiotropic biological processes, with regard to retinal degeneration, intestinal homeostasis, and organismal development. The extent to which it functions in controlling the fly innate immune defense, however, remains largely unknown. Here, we show that blockade of the ERAD in fat bodies antagonizes the Toll but not the IMD innate immune defense in Drosophila. Genetic approaches further suggest a functional role of Me31B in the ERAD-mediated fly innate immunity. Moreover, we provide evidence that silence of Xbp1 other than PERK or Atf6 partially rescues the immune defects by the dysregulated ERAD in fat bodies. Collectively, our study uncovers an essential function of the ERAD in mediating the Toll innate immune reaction in Drosophila.

Innate immune evasion strategies of SARS-CoV-2.

SARS-CoV-2, the virus responsible for the COVID-19 pandemic, has been associated with substantial global morbidity and mortality. Despite a tropism that is largely confined to the airways, COVID-19 is associated with multiorgan dysfunction and long-term cognitive pathologies. A major driver of this biology stems from the combined effects of virus-mediated interference with the host antiviral defences in infected cells and the sensing of pathogen-associated material by bystander cells. Such a dynamic results in delayed induction of type I and III interferons (IFN-I and IFN-III) at the site of infection, but systemic IFN-I and IFN-III priming in distal organs and barrier epithelial surfaces, respectively. In this Review, we examine the relationship between SARS-CoV-2 biology and the cellular response to infection, detailing how antagonism and dysregulation of host innate immune defences contribute to disease severity of COVID-19.

Impact of Sting Pathway in Inflammation and Covid-19 Pathogenesis

The recent encounter by Sars-Cov2 (COVID-19) presented the need of having stronger immune system as the first line of body’s defense mechanism. Stronger the immune response better will be the pathogen recognition. Better understanding of immune response mechanism and functioning of its components will pave the route for defending against these pathogens. Innate immune defense being non-specific in nature identifies both DNA and RNA viruses as well as other pathogens attacking the body. Presence of foreign nucleic acids bound to the specific pattern recognition receptors (PRRs) which include toll-like receptors (TLRs), cGAS-STING, Nod-like receptors (NLR) and RNA helicase receptors (RIG-1 and MDA5) triggers a signaling cascade. These signaling channels direct to the production of type-1 interferons and other cytokine/chemokine effectors of innate immune system which in turn activate adaptive immune response. STING pathway as a prime innate detector of self and non-self-nucleic acids plays an important role in the development of inflammation followed by type-1 interferon production. COVID-19 being involved with the respiratory failure shows the main characteristic feature of cytokine storm which direct to hyper inflammation. These hyper-inflammatory responses worsen the immune compromised state of the patients which makes this disease as the leading cause of mortality worldwide. Understanding the underline mechanisms involved in and regulating the process of hyper inflammation would provide an effective treatment to reduce the mortalities. Deregulation of STING pathway is involved in various inflammatory diseases that make it a potent target to understand its mechanism of work during viral attack. Many recent studies have further supported the importance of delayed STING signaling in the pathogenesis of COVID-19 during the second phase of this disease. This review mainly focuses on the role of STING in inflammation and its function in our immune system which can be harnessed to tackle the recent pathogeneses of COVID-19.

Modulation of macrophages by biophysical cues in health and beyond.

Macrophages play a key role in tissue development and homeostasis, innate immune defence against microbes or tumours, and restoring homeostasis through tissue regeneration following infection or injury. The ability to adopt such diverse functions is due to their heterogeneous nature, which is driven largely by their developmental origin and their response to signals they encounter from the microenvironment. The most well-characterized signals driving macrophage phenotype and function are biochemical and metabolic. However, the way macrophages sense and respond to their extracellular biophysical environment is becoming increasingly recognized in the field of mechano-immunology. These biophysical cues can be signals from tissue components, such as the composition and charge of extracellular matrix or topography, elasticity, and stiffness of the tissue surrounding cells; and mechanical forces such as shear stress or stretch. Macrophages are important in determining whether a disease resolves or becomes chronic. Ageing and diseases such as cancer or fibrotic disorders are associated with significant changes in the tissue biophysical environment, and this provides signals that integrate with those from biochemical and metabolic stimuli to ultimately dictate the overall function of macrophages. This review provides a brief overview of macrophage polarization, followed by a selection of commonly recognized physiological and applied biophysical stimuli impacting macrophage activity, and the potential signalling mechanisms driving downstream responses. The effects of biophysical cues on macrophages' function in homeostasis and disease and the associated clinical implications are also highlighted.

Association of surfactant protein A2 with acute respiratory failure in children.

Interactions among single nucleotide polymorphisms (SNPs) of surfactant protein (SP) are associated with acute respiratory failure (ARF) and its short-term outcome, pulmonary dysfunction at discharge (PDAD) in children. However, genetic association studies using individual SNPs have not been conducted before. We hypothesize that SP genetic variants are associated with pediatric ARF and its short-term complications by themselves. We used available genotype and clinical data in the Floros biobank consisting of 248 children aged ≤24 months with ARF; 86 developed PDAD. A logistic regression analysis was performed for each of the 14 selected SNPs, SP-A1 and SP-A2 genotypes. A p-value less than the Bonferroni correction threshold was considered significant. A likelihood ratio test was done to compare two models (one with demographic data and another with genetic variants). Before Bonferroni correction, female sex is associated with a decreased risk of ARF. Black race and the rs721917 of the SFTPD are associated with increased risk of ARF. After Bonferroni correction, the 1A0 1A1 genotype of SFTPA2 was associated with decreased risk of ARF. The likelihood ratio test showed that the model of the genotype information with demographic data was a better fit to predict ARF risk. None of the SP SNPs and SP-A1, SP-A2 genotypes were associated with PDAD. Our results indicate that SNPs and genotypes of SPs involved in innate immunity and host defense play an important role in ARF and, in the future, may be used as biomarkers.

Neutrophil, neutrophil extracellular traps and endothelial cell dysfunction in sepsis.

Sepsis is a persistent systemic inflammatory condition involving multiple organ failures resulting from a dysregulated immune response to infection, and one of the hallmarks of sepsis is endothelial dysfunction. During its progression, neutrophils are the first line of innate immune defence against infection. Aside from traditional mechanisms, such as phagocytosis or the release of inflammatory cytokines, reactive oxygen species and other antibacterial substances, activated neutrophils also release web-like structures composed of tangled decondensed DNA, histone, myeloperoxidase and other granules called neutrophil extracellular traps (NETs), which can efficiently ensnare bacteria in the circulation. In contrast, excessive neutrophil activation and NET release may induce endothelial cells to shift toward a pro-inflammatory and pro-coagulant phenotype. Furthermore, neutrophils and NETs can degrade glycocalyx on the endothelial cell surface and increase endothelium permeability. Consequently, the endothelial barrier collapses, contributing to impaired microcirculatory blood flow, tissue hypoperfusion and life-threatening organ failure in the late phase of sepsis.

A Moonlighting Protein Secreted by a Nasal Microbiome Fortifies the Innate Host Defense Against Bacterial and Viral Infections.

Evidence suggests that the human respiratory tract, as with the gastrointestinal tract, has evolved to its current state in association with commensal microbes. However, little is known about how the airway microbiome affects the development of airway immune system. Here, we uncover a previously unidentified mode of interaction between host airway immunity and a unique strain (AIT01) of Staphylococcus epidermidis, a predominant species of the nasal microbiome. Intranasal administration of AIT01 increased the population of neutrophils and monocytes in mouse lungs. The recruitment of these immune cells resulted in the protection of the murine host against infection by Pseudomonas aeruginosa, a pathogenic bacterium. Interestingly, an AIT01-secreted protein identified as GAPDH, a well-known bacterial moonlighting protein, mediated this protective effect. Intranasal delivery of the purified GAPDH conferred significant resistance against other Gram-negative pathogens (Klebsiella pneumoniae and Acinetobacter baumannii) and influenza A virus. Our findings demonstrate the potential of a native nasal microbe and its secretory protein to enhance innate immune defense against airway infections. These results offer a promising preventive measure, particularly relevant in the context of global pandemics.

The Innate Immunity Defense against Gastrointestinal Nematodes: Vaccine Development

The nematode parasite infects both humans and animals, causing severe infections. Their unusual surface structures, in particular, pose significant challenges to the immune system. Vaccine-induced immunity, mediated by the innate immune system, could be crucial in the development of an adaptive effector response. The purpose of this paper was to provide an overview of recent research on the host's innate immune system, barriers, and cells that respond to parasitic nematodes. This study investigated the nematode-associated molecular patterns that may recognize by host. Given the innate defense is more than just a static barrier against pathogen infections. It can actively contribute as a director of the adaptive immune response, which is ultimately responsible for the rejection of invasions. The role of innate defense against pathogen infections is located in zone of researcher concentration. Some nematode parasites can actively move through tissues, they pose a challenge to the innate immune system. Furthermore, their cuticular surface, which varies with each molting, cannot be phagocytosed. The nematode's thin, carbohydrate-rich surface layer, as well as the chemicals produced by this layer, cause the first contact with the host's innate immune system. Notably, all components of the innate immune response can be activated and play an important role in the adaptive immune effector response.

Association of Early Childhood Caries with Bitter Taste Receptors: A Meta-Analysis of Genome-Wide Association Studies and Transcriptome-Wide Association Study.

Although genetics affects early childhood caries (ECC) risk, few studies have focused on finding its specific genetic determinants. Here, we performed genome-wide association studies (GWAS) in five cohorts of children (aged up to 5 years, total N = 2974, cohorts: Center for Oral Health Research in Appalachia cohorts one and two [COHRA1, COHRA2], Iowa Fluoride Study, Iowa Head Start, Avon Longitudinal Study of Parents and Children [ALSPAC]) aiming to identify genes with potential roles in ECC biology. We meta-analyzed the GWASs testing ~3.9 million genetic variants and found suggestive evidence for association at genetic regions previously associated with caries in primary and permanent dentition, including the β-defensin anti-microbial proteins. We then integrated the meta-analysis results with gene expression data in a transcriptome-wide association study (TWAS). This approach identified four genes whose genetically predicted expression was associated with ECC (p-values < 3.09 × 10-6; CDH17, TAS2R43, SMIM10L1, TAS2R14). Some of the strongest associations were with genes encoding members of the bitter taste receptor family (TAS2R); other members of this family have previously been associated with caries. Of note, we identified the receptor encoded by TAS2R14, which stimulates innate immunity and anti-microbial defense in response to molecules released by the cariogenic bacteria, Streptococcus mutans and Staphylococcus aureus. These findings provide insight into ECC genetic architecture, underscore the importance of host-microbial interaction in caries risk, and identify novel risk genes.

Proportion of Concentrate in the Diet of Early Lactation Dairy Cows Has Contrasting Effects on Circulating Leukocyte Global Transcriptomic Profiles, Health and Fertility According to Parity

The functionality of circulating leukocytes in dairy cows is suppressed after calving, with negative energy balance as a risk factor. Leukocyte transcriptomic profiles were compared separately in 44 multiparous (MP) and 18 primiparous (PP) Holstein-Friesian cows receiving diets differing in concentrate proportion to test whether immune dysfunction could be mitigated by appropriate nutrition. After calving, cows were offered either (1) low concentrate (LC); (2) medium concentrate (MC) or (3) high concentrate (HC) diets with proportions of concentrate to grass silage of 30%:70%, 50%:50% and 70%:30%, respectively. Cow phenotype data collected included circulating metabolites, milk yield and health and fertility records. RNA sequencing of circulating leukocytes at 14 days in milk was performed. The HC diet improved energy balance in both age groups. There were more differentially expressed genes in PP than MP cows (460 vs. 173, HC vs. LC comparison) with few overlaps. The MP cows on the LC diet showed upregulation of the complement and coagulation cascade and innate immune defence mechanisms against pathogens and had a trend of more cases of mastitis and poorer fertility. In contrast, the PP cows on the HC diet showed greater immune responses based on both gene expression and phenotypic data and longer interval of calving to conception. The leukocytes of MP and PP cows therefore responded differentially to the diets between age, nutrient supply and immunity affecting their health and subsequent fertility.

CfIRF8-like interacts with the TBK1/IKKε family protein and regulates host antiviral innate immunity

The interferon regulatory factor (IRF) family, a class of transcription factors with key functions, are important in host innate immune defense and stress response. However, further research is required to determine the functions of IRFs in invertebrates. In this study, the coding sequence of an IRF gene was obtained from the Zhikong scallop (Chlamys farreri) and named CfIRF8-like. The open reading frame of CfIRF8-like was 1371 bp long and encoded 456 amino acids. Protein domain prediction revealed a typical IRF domain in the N-terminus of the CfIRF8-like protein and a typical IRF3 domain in the C-terminus. Multiple sequence alignment confirmed the conservation of the amino acid sequences of these two functional protein domains. Phylogenetic analysis showed that CfIRF8-like clustered with mollusk IRF8 proteins and then clustered with vertebrate IRF3, IRF4, and IRF5 subfamily proteins. Quantitative real-time PCR detected CfIRF8-like mRNA in all tested scallop tissues, with the highest expression in the gills. Simultaneously, the expression of CfIRF8-like transcripts in gills was significantly induced by polyinosinic–polycytidylic acid challenge. The results of protein interaction experiments showed that CfIRF8-like could directly bind the TBK1/IKKε family protein of scallop (CfIKK2) via its N-terminal IRF domain, revealing the presence of an ancient functional TBK1/IKKε-IRF signaling axis in scallops. Finally, dual-luciferase reporter assay results showed that the overexpression of CfIRF8-like in human embryonic kidney 293T cells could specifically activate the interferon β promoter of mammals and the interferon-stimulated response element promoter in dose-dependent manners. The findings of this preliminary analysis of the signal transduction and immune functions of scallop CfIRF8-like protein lay a foundation for an in-depth understanding of the innate immune function of invertebrate IRFs and the development of comparative immunology. The experimental results also provide theoretical support for the breeding of scallop disease-resistant strains.

Essential immune functions of fibroblasts in innate host defense.

The term fibroblast has been used generally to describe spindle-shaped stromal cells of mesenchymal origin that produce extracellular matrix, establish tissue structure, and form scar. Current evidence has found that cells with this morphology are highly heterogeneous with some fibroblastic cells actively participating in both innate and adaptive immune defense. Detailed analysis of barrier tissues such as skin, gut, and lung now show that some fibroblasts directly sense pathogens and other danger signals to elicit host defense functions including antimicrobial activity, leukocyte recruitment, and production of cytokines and lipid mediators relevant to inflammation and immunosuppression. This review will synthesize current literature focused on the innate immune functions performed by fibroblasts at barrier tissues to highlight the previously unappreciated importance of these cells in immunity.

Avian Reovirus σA Protein Inhibits Type I Interferon Production by Abrogating Interferon Regulatory Factor 7 Activation.

Type I interferon (IFN) response is the first line of host-based innate immune defense against viral infections. However, viruses have developed multiple strategies to counter host IFN responses, so they may continue infecting hosts via effective replication. Avian reovirus (ARV), an RNA virus, causes viral arthritis or tenosynovitis in chickens. Previous studies have shown that ARV is highly resistant to the antiviral effects of IFN. However, the underlying mechanisms that enable ARV to block the IFN pathway remain unclear. In this study, we found that ectopic expression of ARV protein, σA, significantly inhibited the production of IFN-β induced by melanoma-differentiation-associated gene 5 (MDA5) and poly(I·C). Knockdown of σA during ARV infection enhances the IFN-β response and suppresses viral replication. ARV σA inhibited the MDA5-mediated IFN-β activation by targeting interferon regulatory factor 7 (IRF7). Further studies demonstrated that σA interacts with IRF7, thereby blocking IRF7 dimerization and nuclear translocation, finally leading to the inhibition of IFN-β production. These findings reveal a novel mechanism that allows ARV to evade host antiviral immunity. IMPORTANCE ARV, the causative agent of viral arthritis or tenosynovitis in chickens, has a significant economic impact as it results in poor weight gain and increased feed conversion ratios. The MDA5-mediated IFN-β signal pathway plays an important role in host antiviral defense. Therefore, RNA viruses have developed mechanisms to counter this signaling pathway and successfully establish infection. However, the strategies adopted by ARV to block MDA5-IRF7 signaling remain unclear. In the current study, we demonstrated that ARV σA inhibits this pathway by binding to IRF7, which blocked IRF7 dimerization and nuclear translocation. Our findings may provide insights into how avian reovirus counteracts the innate antiviral immunity of the host to ensure viral replication.

Cloning and characterization of type IV interferon from black carp Mylopharyngodon piceus

From mammals to fish, interferons (IFNs) play vital roles in the immune response. In this study, a newly identified type IV interferon (bcIFN-υ) from black carp (Mylopharyngodon piceus) has been cloned and characterized. The CDS of bcIFN-υ consists of 489 nucleotides, encoding 163 amino acids, with the first 20 amino acids predicted to be the signal peptide region. The immunoblot and immunofluorescence assays verified that bcIFN-υ was a secreted cytokine. qPCR analysis and reporter assay demonstrated that bcIFN-υ participated in innate immune defense and activated the transcription of fish ISRE promoter under spring viremia of carp virus (SVCV) stimulation. Additionally, compared with control group, EPC cells transfected with bcIFN-υ or incubated with the bcIFN-υ-containing conditioned media before SVCV infection showed greatly enhanced antiviral activity, and the transcription levels of MX1, PKR, ISG15 and Viperin genes were significantly increased. The subsequential co-immunoprecipitation assay identified the interaction between bcIFN-υ proteins. Collectively, our data conclude that bcIFN-υ is a kind of secretory protein with self-interaction and triggering the expression of downstream ISGs to enhance the antiviral activity of host cells.

Circadian metabolism regulates the macrophage inflammatory response

Abstract Macrophages are an integral part of the innate immune system and coordinate host defense to microbial infections, as well as shaping the remodeling response after tissue injury. Metabolism is now appreciated to be a powerful and pervasive regulator of the identity and function of macrophages. Upon exposure to microbial ligands, macrophage inflammatory activation and the associated induction of phagocytosis, inflammatory responses, and other host defense activities are supported by dynamic changes to cellular metabolism. Of note, metabolic activity is robustly regulated in a circadian fashion, with many metabolic processes displaying peak activity in one phase of the circadian cycle and trough activity in an antiphase manner. Here, we review recent findings suggesting that circadian metabolism influences macrophage activities and particularly the inflammatory response. First, we summarize macrophage activities known to display time-of-day–dependent variation and their mechanistic basis. Second, we review metabolic processes that have been shown to be rhythmically regulated in macrophages and discuss how such circadian metabolism affects or is likely to affect macrophage activities. Third, we discuss the concept of entrainment of the macrophage clock, and consider how loss of rhythmic regulation of macrophage activities may contribute to pathophysiological conditions like shift work, obesity, and aging. Finally, we propose that circadian metabolism can be used to understand the rationale and mechanistic basis of dynamic regulation of inflammatory responses during infection.

The mononuclear phagocyte system in hepatocellular carcinoma.

The mononuclear phagocyte system (MPS) consists of monocytes, dendritic cells and macrophages, which play vital roles in innate immune defense against cancer. Hepatocellular carcinoma (HCC) is a complex disease that is affected or initiated by many factors, including chronic hepatitis B virus infection, hepatitis C virus infection, metabolic disorders or alcohol consumption. Liver function, tumor stage and the performance status of patients affect HCC clinical outcomes. Studies have shown that targeted treatment of tumor microenvironment disorders may improve the efficacy of HCC treatments. Cytokines derived from the innate immune response can regulate T-cell differentiation, thereby shaping adaptive immunity, which is associated with the prognosis of HCC. Therefore, it is important to elucidate the function of the MPS in the progression of HCC. In this review, we outline the impact of HCC on the MPS. We illustrate how HCC reshapes MPS cell phenotype remodeling and the production of associated cytokines and characterize the function and impairment of the MPS in HCC.

Different Molecular Forms of TFF3 in the Human Respiratory Tract: Heterodimerization with IgG Fc Binding Protein (FCGBP) and Proteolytic Cleavage in Bronchial Secretions

The polypeptide TFF3 belongs to the trefoil factor family (TFF) of lectins. TFF3 is typically secreted from mucous epithelia together with mucins. Both intestinal and salivary TFF3 mainly exist as disulfide-linked heterodimers with IgG Fc binding protein (FCGBP). Here, we investigated bronchial tissue specimens, bronchial secretions, and bronchoalveolar lavage (BAL) fluid from patients with a chronic obstructive pulmonary disease (COPD) background by fast protein liquid chromatography and proteomics. For the first time, we identified different molecular forms of TFF3 in the lung. The high-molecular mass form represents TFF3-FCGBP oligomers, whereas the low-molecular mass forms are homodimeric and monomeric TFF3 with possibly anti-apoptotic activities. In addition, disulfide-linked TFF3 heterodimers with an Mr of about 60k and 30k were detected in both bronchial secretions and BAL fluid. In these liquids, TFF3 is partly N-terminally truncated probably by neutrophil elastase cleavage. TFF3-FCGBP is likely involved in the mucosal innate immune defense against microbial infections. We discuss a hypothetical model how TFF3 might control FCGBP oligomerization. Furthermore, we did not find indications for interactions of TFF3-FCGBP with DMBT1gp340 or the mucin MUC5AC, glycoproteins involved in mucosal innate immunity. Surprisingly, bronchial MUC5AC appeared to be degraded when compared with gastric MUC5AC.

Molecular Cloning and Characterization of a Novel Antimicrobial Peptide from the Skin of Kaloula pulchra.

Bacterial resistance to all currently available conventional antibiotics has caused a global public health crisis and led to an imperative search for new agents. Antimicrobial peptides (AMPs) are essential components of host innate immune defense against microbial invasions. The objective of this study was to report a novel AMP, brevinin-2KP, from the skin of the black Kaloula pulchra frog and describe its structural and biological characterization. The physical and chemical parameters of brevinin-2KP were predicted with the ExPASy Bioinformatics Resource Portal. The assembled sequences were aligned with ClustalW, and the phylogenetic tree was constructed using Mega. Circular dichroism (CD) experiments were carried out to identify the secondary structure and the stability of peptide in different solvent environments. The cytotoxicity of brevinin-2KP was evaluated by the MTT test. To determine antibacterial activity of brevinin- 2KP, a standard two-fold broth dilution method was used. SEM was carried out to observe the morphological change in the bacterial treated by brevinin-2KP. The live/dead bacterial viability was measured with a LIVE/DEAD® BacLight kit. Histamine release and mast cell degranulation assays were performed. The precursor of brevinin-2KP contains 72 amino acid residues, including a conserved signal peptide, acidic propeptide with KR residues, and mature peptide with a sequence of GVITDALKGAAKTVAAELLKKAHCKLTNSC. Phylogenetic analysis based on the amino acid sequences of 34 brevinin-2 peptides from 30 anuran species demonstrates that K. pulchra is genetically closely related to the genus Hylarana. The CD spectra analysis indicates that brevinin-2KP adopts random coil in the water and an organized α-helical conformation in SDS solution. Further, this secondary structure is stable under high salt and high-temperature conditions. Brevinin-2KP is weakly active towards the tested Gram-positive and Gram-negative bacteria as well as fungi due to its membranolytic action. Moreover, brevinin-2KP inhibits the proliferation of several mammal cells with IC50 values ranging from 3.27 to 59.75 μM. In addition, brevinin-2KP promotes degranulation and histamine release of mast cells, indicating that it is involved in the inflammatory response. This is the first report on AMP identified from the skin of K. pulchra. Brevinin-2KP adopts a typical amphipathic α-helix conformation in membrane mimic environment and shows antimicrobial and antitumor activities by potential membranolytic mechanism. In addition, brevinin-2KP can promote degranulation and histamine release of mast cells. Brevinin-2KP is expected to become a good drug temple molecule.