Tuberculosis (TB), caused by Mycobacterium tuberculosis ( M.tb ), continues to pose a critical global health threat as a leading infectious cause of mortality. Therapeutic efficacy is increasingly compromised by the emergence of multidrug-resistant strains and the limitations of existing regimens, which necessitate treatment durations of six months or longer. Protein tyrosine phosphatase B from Mtb (PtpB- Mtb ) has been recognized as a critical virulence factor, representing a promising target for novel antitubercular therapies due to its unique structural and functional properties. In this study, a comprehensive structure-based virtual screening approach was employed to identify novel small-molecule scaffolds with inhibitory potential against PtpB- Mtb . The ChemBridge compound library was curated and filtered for drug-like properties, followed by hierarchical molecular docking and molecular dynamics simulations to prioritize candidates with high predicted affinity and stability within the PtpB- Mtb active site. Quantum mechanical calculations further characterized the electronic properties of top hits. Recombinant PtpB- Mtb was expressed and purified to homogeneity, and in vitro enzymatic assays were performed to evaluate the inhibitory potency and selectivity of shortlisted compounds. Two derivatives bearing pyrazolo[4,3-c]pyridine and 1,4-diazepane ring nuclei demonstrated significant inhibition of PtpB- Mtb activity, exhibiting IC₅₀ values of 14.4 µM and 32.6 µM, respectively. Biolayer interferometry confirmed strong and specific binding to PtpB-Mtb, with dissociation constants (K d ) of 0.012 µM and 0.57 µM. The integrated workflow presented herein highlights the potential of these novel scaffolds as starting points for the development of selective, cell-permeable PtpB- Mtb inhibitors, offering a promising avenue for next-generation anti-tubercular drug discovery.

Mycobacterium indicus pranii (MIP), an atypical mycobacterium originally developed as an anti-leprosy vaccine, has emerged as a potent immunomodulator with diverse therapeutic applications. Despite its clinical significance, molecular mechanisms underlying MIP’s immunomodulatory properties remain largely unexplored. Bacterial phosphatases are recognized as crucial virulence factors that enable pathogens to evade host defenses by modulating host immune signaling pathways, including phosphoinositide metabolism. MIP_07528 was identified as a putative protein tyrosine phosphatase B (PtpB) ortholog through in silico analysis, with significant sequence conservation observed within catalytic domains of pathogenic mycobacterial PtpB proteins. Phosphatase activity was detected in both cell lysate and culture filtrate fractions, revealing differential expression patterns between MIP and M. tuberculosis. Upregulation of MIP_07528 was demonstrated under oxidative stress, suggesting involvement in stress adaptation. The recombinant protein exhibited distinctive kinetic properties, characterized by higher substrate affinity yet increased susceptibility to oxidative inactivation compared to its M. tuberculosis counterpart. In macrophages, MIP_07528 suppressed pro-inflammatory cytokines while enhancing anti-inflammatory IL-10 production. These findings establish MIP_07528 as a functional phosphatase that may contribute to MIP’s immunomodulatory properties. This work advances understanding of phosphatase function in non-pathogenic mycobacteria while providing insights into virulence factor evolution and establishing a foundation for novel antimicrobial strategies.

Anthrax, caused by Bacillus anthracis, remains a critical zoonotic threat, with treatment efficacy increasingly compromised by advanced infection progression and rising antibiotic resistance. This study leverages integrative computational strategies to identify and characterize novel therapeutic targets among previously uncharacterized molecular chaperones-Trigger Factor (BASTig) and peptidyl-prolyl cis-trans isomerase B (BASPpiB)-from B. anthracis Sterne. Structural elucidation using homology modelling and AlphaFold revealed distinctive architectures for BASTig (425 residues) and BASPpiB (145 residues). High-throughput virtual screening of diverse chemical libraries pinpointed compounds 51002 and 50423 as promising inhibitors, with strong binding affinities of -52.58 and -66.4 kcal/mol, respectively. ADME profiling confirmed favourable drug-like properties, and molecular dynamics simulations demonstrated stable protein-ligand interactions. Quantum mechanical calculations further supported the electronic complementarity and thermodynamic stability of these complexes. Electrostatic surface potential (ESP) analysis revealed that compound 51002 features predominantly positive charge distributions, favouring interactions with acidic residues in BASTig, while compound 50423 displays heterogeneous electrostatic regions, enabling adaptive binding to BASPpiB's dynamic pocket. Toxicity predictions indicated acceptable safety profiles for both leads. Immunogenicity assessment showed differential antigenic potential (BASPpiB: 100 %, BASTig: 66 %). Epitope mapping with ABCpred identified multiple high-scoring, spatially distributed B-cell epitopes in both proteins, with substantial concordance between predictive algorithms. These results highlight the therapeutic promise of targeting molecular chaperones in B. anthracis and provide a foundation for both small-molecule drug discovery and rational immunogen design, addressing urgent needs in anthrax intervention and antimicrobial resistance.

Toxin–antitoxin (TA) modules represent sophisticated regulatory networks that have evolved from simple plasmid maintenance factors into multifunctional genetic modules orchestrating bacterial stress responses, pathogenesis, and ecological adaptation. This review highlights a compelling correlation between the abundance of toxin–antitoxin (TA) modules and bacterial pathogenicity, as exemplified by Mycobacterium tuberculosis (M.tb), which encodes 118 TA loci—significantly more than the fewer than 10 found in closely related saprophytic species. The clinical significance of TA modules extends beyond traditional stress response roles to encompass antimicrobial persistence, where systems like VapBC and MazEF facilitate dormant subpopulations that survive antibiotic therapy while maintaining chronic infections. Recent discoveries have revealed TA modules as sophisticated bacterial defense mechanisms against bacteriophage infection, with DarTG and ToxIN systems representing novel antiviral immunity components that complement CRISPR-Cas and restriction–modification systems. The immunomodulatory capacity of TA modules demonstrates their role in host–pathogen interactions, where systems such as VapC12 in M.tb promote macrophage polarization toward permissive M2 phenotypes while inducing anti-inflammatory cytokine production. Large-scale genomic analyses reveal that TA modules function as drivers of horizontal gene transfer networks, with their signatures enabling accurate prediction of plasmid community membership and serving as determinants of microbial community structure. The biotechnological applications of TA modules have expanded to include genetic circuit stabilization, biocontainment device construction, and multi-species microbial community engineering, while therapeutic strategies focus on developing multi-target inhibitors against conserved TA protein families as promising approaches for combating drug-resistant bacterial infections. The evolutionary conservation of TA modules across diverse bacterial lineages underscores their fundamental importance as central organizing principles in bacterial adaptation strategies, where their multifunctional nature reflects complex selective pressures operating across environmental niches and host-associated ecosystems. This review provides an integrated perspective on TA modules as dynamic regulatory elements that support bacterial persistence, immune evasion, and ecological versatility, establishing them as genetic elements with truly “many faces and functions” in prokaryotic biology.

Mycobacterium tuberculosis (M.tb) exhibits remarkable adaptability and persistence within host micro-environments, making tuberculosis a persistent global health challenge. Finding safe and relevant model organisms to study M.tb pathobiology is essential for augmenting research in this field. Mycobacterium indicus pranii (MIP), a non-pathogenic mycobacterial species with known immunomodulatory properties and established safety in human applications, represents a promising yet underutilized research model. Within hosts, M.tb encounters diverse stress conditions including oxidative and nitrosative challenges, nutrient limitation, pH fluctuations, and immune cell-mediated pressures, all of which shape its survival strategies. This investigation presents a thorough evaluation of MIP as an alternative research model through systematic comparative analyses with the conventionally utilized Mycobacterium smegmatis (M.smeg). Genomic investigation revealed MIP possesses a significantly higher number of M.tb-homologous virulence-associated genes and conserved drug targets as compared to M. smeg, while sharing equivalent human-homologous gene content with M.tb. Functional assays demonstrated MIP's superior tolerance to multiple stress conditions relevant to host micro-environments, including SDS-mediated envelope stress, nitrosative stress, acidic and alkaline pH extremes, copper toxicity, and elevated temperature—characteristics supported by the presence of key M.tb stress regulator homologs (sigE, sigH, mprAB, and Rv2745c). In macrophage infection models, MIP exhibited enhanced intracellular persistence as compared to M.smeg and induced a balanced cytokine profile resembling M.tb infection. The distinctive genomic and physiological characteristics of MIP establish its biological relevance as a superior surrogate model for investigating mycobacterial stress adaptation mechanisms, virulence determinants, and host-pathogen interactions, potentially accelerating discovery of novel therapeutic strategies against tuberculosis.

ABSTRACT In contrast to adaptive immunity, which relies on memory T and B cells for long-term pathogen-specific responses, trained immunity involves the enhancement of innate immune responses through cellular reprogramming. Experimental evidence from animal models and human studies supports the concept of trained immunity and its potential therapeutic applications in the development of personalized medicine. However, there remains a huge gap in understanding the mechanisms, identifying specific microbial triggers responsible for the induction of trained immunity. This underscores the importance of investigating the potential role of trained immunity in redefining host defense and highlights future research directions. This minireview will provide a comprehensive summary of the new paradigm of trained immunity or innate memory pathways. It will shed light on infection-induced pathways through non-specific stimulation within macrophages and natural killer cells, which will be further elaborated in multiple disease perspectives caused by infectious agents such as bacteria, fungi, and viruses. The article further elaborates on the biochemical and cellular basis of trained immunity and its impact on disease status during recurrent exposures. The review concludes with a perspective segment discussing potential therapeutic benefits, limitations, and future challenges in this area of study. The review also sheds light upon potential risks involved in the induction of trained immunity.

Trigger factor, as a chaperone protein, is required for survival of Mycobacterium tuberculosis (M.tb) in a stressed environment. This protein interacts with various partners in both the pre- and the post-translation processes, yet the crystal structures of the M.tb trigger factor remain unresolved. In this study, we developed a homology model of M.tb trigger factor to facilitate the discovery and design of inhibitors. To validate the model, we employed several methodologies, including Ramachandran plot and molecular dynamics simulations. The simulations showed a stable trajectory, indicating the accuracy of the model. The active site of M.tb Trigger Factor was identified based on site scores, and virtual screening of over 70,000 compounds led to the identification of two potential hits: HTS02984 (ethyl 2-(3-(4-fluorophenyl)ureido)-6-methyl-4,5,6,7-tetrahydrothieno[2,3-c]pyridine-3-carboxylate) and S06856 ((E)-N-(4-((2-(4-(tert-butyl)benzoyl)hydrazono)methyl)phenyl) acetamide). These compounds showed strong binding affinity and energy scores, and their chemical descriptors were evaluated. Our study provides a reliable computational model for M.tb Trigger Factor and identifies two potential inhibitors for this crucial protein, which could aid in the development of novel therapies against tuberculosis.Communicated by Ramaswamy H. Sarma

Peptidyl prolyl cis/trans isomerases (PPIases), a ubiquitously distributed superfamily of enzymes, associated with signal transduction, trafficking, assembly, biofilm formation, stress tolerance, cell cycle regulation, gene expression and tissue regeneration, is a key regulator of metabolic disorders and microbial virulence. This review assumes an integrative approach, to provide a holistic overview of the structural and functional diversity of PPIases, examining their conformational dynamics, cellular distribution, and physiological significance. We explore their intricate involvement in cellular processes and virulence modulation in both eukaryotic and prokaryotic systems. Additionally, we evaluate the potential of these molecular chaperones as drug targets and vaccine candidates, emphasizing their relevance in therapeutic development. By synthesizing recent findings and providing a broader perspective on these proteins, this review aims to enhance our understanding of their multifaceted roles in biology and their potential applications in medicine.

Mycobacterium indicus pranii (MIP), a benign saprophyte with potent immunomodulatory attributes, holds a pivotal position in mycobacterial evolution, potentially serving as the precursor to the pathogenic Mycobacterium avium complex (MAC). Despite its established immunotherapeutic efficacy against leprosy and notable outcomes in gram-negative sepsis and COVID-19 cases, the genomic and biochemical features of MIP remain largely elusive. This study explores the uncharted territory of toxin-antitoxin (TA) systems within MIP, hypothesizing their role in mycobacterial pathogenicity regulation. Genome-wide screening, employing diverse databases, unveils putative TA modules in MIP, setting the stage for a comparative analysis with known modules in Mycobacterium tuberculosis, Mycobacterium smegmatis, Escherichia coli, and Vibrio cholerae. The study further delves into the TA network of MAC and Mycobacterium intracellulare, unraveling interactive properties and family characteristics of identified TA modules in MIP. This comprehensive exploration seeks to illuminate the contribution of TA modules in regulating virulence, habitat diversification, and the evolutionary pathogenicity of mycobacteria. The insights garnered from this investigation not only enhance our understanding of MIP's potential as a vaccine candidate but also hold promise in optimizing tuberculosis drug regimens for expedited recovery.

Mycobacterium tuberculosis (Mtb) is an extremely pathogenic bacterium which is responsible for causing tuberculosis. M. tuberculosis generally causes infection in lungs and other organs, can survive different physiological conditions inside the host and can also cause latent infection. Mtb, for its survival and pathogenesis, has to modulate various host pathways. The common pathways that are altered by Mtb include modulation of glycolytic flux, endoplasmic reticulum (EPR) stress, mitochondrial metabolism, apoptosis, and necrosis, inhibition of phagosome maturation and autophagy. Mtb harbors various mechanisms to evade host defense pathways for its survival. There are several types of antiinflammatory microRNA-21 which can help in cadence of glycolytic flux and also regulate the levels of phosphofructokinase by decreasing the biosynthetic precursors which are required for inflammatory responses. Further, the EPR stress pathway can be modulated by Mtb with the help of unfolded protein response–inducible transcription factor C/EBP homologous protein. The necrosis is regulated by inhibiting reactive oxygen species (ROS) production with the help of prostaglandin E2. With the help of extracellular regulated kinase 1/2, signal transducer and activator of transcription 3, and mitogen-activated protein kinase p38, interleukin -10 executes apoptosis pathway. Further, autophagy modulation synthesizes phenolic glycolipid with the help of polyketide synthase, encoded by intact pks 1–15 and by manipulating Ca2+ signaling.

Contrary to common understanding, bacterial responses are not standalone. External stimuli trigger bacterial communities to respond in consorted manner. Mega habitats on earth predominantly support biofilms form of growth. Heterogeneous microbial consortia stabilize over matrix and grow on the interface between any two of solid, liquid, or gas. This biofilm is advantageous to bacterial population by supporting nutrient acquisition, its use and recycling along with retention of aqueous microenvironment, enhanced cell-to-cell communication and orchestrated response against stresses like biocides and toxins. Chemical sensing of bacterial quorum, coordinated response in bacterial biofilms, different roles and spatial positioning of various bacterial populations makes them invincible. Biofilms also impact the bacterial virulence capacity, pathogenicity, antibiotic release, sporulation, etc. Bacterial communities in host-microbiome impact the host pathology or may have a causal association. Also, beneficial microflora at host surfaces, inner tracks and cavities and tissue linings is protective against continuous environmental stresses and dysbiosis in the normal microflora associated with disease onset. This chapter aims to describe the bacterial stress adaptation mechanisms and pathological impact of bacterial communities and summarize the new development in the area in recent times.

Mycobacterium tuberculosis (M.tb)-encoded factors protect it against host-generated stresses and support its survival in the hostile host environment. M.tb possesses two peptidyl-prolyl cis-trans isomerases and a probable trigger factor encoded by Rv2462c which has an FKBP-like PPIase domain. PPIases are known to assist the folding of peptidyl-prolyl bonds and are involved in various cellular processes important for bacterial survival in host-generated stresses. In this study, we aim to functionally characterize Rv2462c of M.tb. Our data suggest that the trigger factor of M.tb exhibits chaperone activity both in vitro and in vivo. Heterologous expression of M.tb-Rv2462c locus into Mycobacterium smegmatis enhanced its survival within macrophages, adaptation to oxidative stress and biofilm formation. M.tb-trigger factor has strong immunomodulatory potential and modifies the cytokine profile of the host towards the proinflammatory axis.

Mycobacterium tuberculosis, a pathogen that causes tuberculosis encounters a variety of stresses and extreme conditions such as acidic conditions, hypoxic and immune system stress, metal, and heat shocks during host infection. It withstands these hostile environments by employing various survival strategies. The lung macrophages are the primary immune cells that interact with M. tuberculosis after infection. Various proteins of M. tuberculosis are responsible for the survival of M. tuberculosis in acidic and hypoxic conditions inside the host. M. tuberculosis tolerates temperature variations with the help of various heat shock proteins (Hsp) such as Hsp70, Hsp22.5, HspR, and the protein Acr2, which is an active member of α-crystalline family of molecular chaperones. M. tuberculosis also overcomes toxic concentrations of various metal ions. M. tuberculosis fulfills iron requirements by the acquisition of iron using siderophores such as mycobactins and carboxymycobactins.

Contaminated wastewater is a serious threat to different fragile ecosystems and related life forms. The presence of microorganisms, such as bacteria, fungi, yeast, and viruses in water is harmful for humans. Contaminated water containing several pathogenic bacteria act as the vehicle for several infectious diseases. Neutralization and removal of these pathogens before discharging into stream water or using them for other purposes is inevitable. Several molecular biology techniques are employed for detection of these pathogens. Specific biomolecules at the microbial surface may be used for the detection of pathogens by tagging sensors molecules such as antibodies, aptamers, carbohydrates, and antimicrobial peptides with different nanoparticles.

The nanotechnology has gifted humankind with so many types of nanoparticles, which are highly used in industrial, biochemical, chemical, physicochemical and in many other fields. Metal oxide nanoparticles, such as Zinc oxide, Titanium dioxide, Magnesium oxide, Aluminium oxide, etc. have unique physiochemical properties. In the emergence of antibiotic resistance, the development of antimicrobial agents has become a matter of prime interest. Nanotechnology development has received great attention for targeting pathogens. Many metal oxide nanoparticles are antibacterial agents and are also called "nanoantibiotics." Nanoparticles have conductivity, solubility, magnetic, mechanical, and antibacterial characteristics. This technology has potential in wastewater treatment and improving treatment efficiency. The present chapter summarizes the antimicrobial properties of nanoparticles employed for decontaminating wastewater and their mechanisms of action in detail.

Bacterial communities respond to stimuli in a consorted manner, quite differing from planktonic bacteria. Bacterial growth in biofilm form is the most predominant in all mega and micro-habitats. This matrix stabilized microbial consortia develops in the heterogenic micro-environment creating an interface of any two of solid, liquid or gas where biofilm forms. Biofilm formation offers nutrient acquisition by sorption, synergistic use and recycling. It also benefits by retention of aqueous support and enhanced cell to cell communication. But the most important ones are coordinated behavior and tolerance to stress like biocides and disinfectants. Another mechanism of coordinated bacterial behavior is quorum sensing. These are the well-orchestrated actions that are directed through differential gene expression and regulated by molecular communication among bacteria. The processes of quorum sensing not only affect the biofilm formation but virulence, pathogenicity, antibiotic release, spore formation, etc. In a similar context, papulation of bacteria shows the presence of persister cells that differ in physiological status causing enhanced stress tolerance. Very often, persister cells show growth arrest and antibiotic tolerance upon stress stimulation. The name itself justifies the difficult clearance of these cells from the infected host upon routine clinical interventions. Lastly, the bacterial communities appear as microbiome in normal and disease conditions in humans and other hosts. The composition of the bacterial milieu in the outer and inner surfaces, cavities, and cell linings helps to sustain the host in normal form against continuous environmental stresses. Also, any dysbiosis of the microbiome may have a causal or supportive association with disease conditions. Thus, the chapter aims to capture the holistic snapshot of the struggle of bacterial communities against chemical, antibiotic and host generated stresses and the underlined mechanisms for their adaptation.

Macrophages are key arsenals of the immune system against invaders. After compartmental isolation of a pathogen in phagosomes, the host immune response attempts to neutralize the pathogen. However, pathogens possess the ability to subvert these assaults and can also convert macrophages into their replicative niche. The multiple host defense evasion mechanisms employed by these pathogens include phagosome maturation arrest, molecular mimicry through secretory antigens, interference with host signaling, active radical neutralization, inhibition of phagosome acidification, alteration of programmed cell death, and other mechanisms. Macrophage biology as a part of the host-pathogen interaction has expanded rapidly in the past decade. The present review aims to shed some light upon the macrophage defense evasion strategies employed by pathogens. We have also incorporated recent knowledge in the field of macrophage dynamics during infection and evolutionary perspectives of macrophage dynamics.

Water bodies get contaminants from natural and anthropogenic sources. Infectious ingredients of wastewater are released from hospitals, veterinary clinics, households, and industries and are harmful to the aquatic ecosystem. Aquatic animals are more vulnerable to these contaminants because of direct exposure to harmful to the water bodies. Different approaches, such as Ames bioassay, comet assay, and luminescent biosensor, are routine monitoring tools to assay infectious contaminants in wastewater effluents. The proposed chapter aims to review the classical and emerging approaches to assay the contaminants present in the wastewater. The source and type of contaminants will also be described. The emerging "-omics" technology including metagenomics, transcriptomics, metatranscriptomics, and metabolomics for toxicological assessment of water and other environmental samples will also be discussed. The discussion will cover the assay method for both culturable and nonculturable microbial contaminants for detection. All emerging methods such as high-throughput next-generation sequencing as well as fluorescence in situ hybridization, denaturant gradient gel electrophoresis, and biosensors will also be detailed in the current chapter.