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There is an intricate network of relations between endophytic fungi and their hosts that affects the production of various bioactive compounds. Plant-associated endophytic fungi contain industrially important enzymes and have the potential to fulfil their rapid demand in the international market to boost business in technology. Being safe and metabolically active, they have replaced the usage of toxic and harmful chemicals and hold a credible application in biotransformation, bioremediation and industrial processes. Despite these, there are limited reports on fungal endophytes that can directly cater to the demand and supply of industrially stable enzymes. The underlying reasons include low endogenous production and secretion of enzymes from fungal endophytes which have raised concern for widely accepted applications. Hence, it is imperative to augment the biosynthetic and secretory potential of fungal endophytes. Modern state-of-the-art biotechnological technologies aiming at strain improvement using cell factory engineering as well as precise gene editing like Clustered Regularly Interspaced Palindromic Repeats (CRISPR) and its Associated proteins (Cas) systems which can provide a boost in fungal endophyte enzyme production. Additionally, it is vital to characterize optimum conditions to grow one strain with multiple enzymes (OSME). The present review encompasses various plants-derived endophytic fungal enzymes and their applications in various sectors. Furthermore, we postulate the feasibility of new precision approaches with an aim for strain improvement and enhanced enzyme production.

Abstract: Enterococcus faecalis (E. faecalis) is an opportunistic multidrug-resistant (MDR) pathogen found in the guts of humans and farmed animals. Due to the occurrence of (MDR) strain there is an urgent need to look for an alternative treatment approach. E. faecalis is a Gram-positive bacterium, which is among the most prevalent multidrug resistant hospital pathogens. Its ability to develop quorum sensing (QS) mediated biofilm formation further exacerbates the pathogenicity and triggers lifethreatening infections. Therefore, developing a suitable remedy for curing E. faecalis mediated enterococcal infections is an arduous task. Several putative virulence factors and proteins are involved in the development of biofilms in E. faecalis. Such proteins often play important roles in virulence, disease, and colonization by pathogens. The elucidation of the structure-function relationship of such protein drug targets and the interacting compounds could provide an attractive paradigm towards developing structure-based drugs against E. faecalis. This review provides a comprehensive overview of the current status, enigmas that warrant further studies, and the prospects toward alleviating the antibiotic resistance in E. faecalis. Specifically, the role of biofilm and quorum sensing (QS) in the emergence of MDR strains had been elaborated along with the importance of the protein drug targets involved in both the processes.

Nanoparticles are those agents that are made-up of single or a combination of single or multiple materials which are very small in size ranging from 1 to 100 nanometers. Several studies reveal that nanoparticles have features that interact effectively with microorganisms and can help in treating multidrug-resistant organisms. These have intrinsic antimicrobial activity and are of various types broadly divided into organic and inorganic nanoparticles. Nanoparticles can engage with bacteria and travel across the bacterial cells and host cell membranes, and help treat ESKAPE pathogens which are among the most notorious multidrug resistant superbugs. These pathogens have MDR features and have multiple types of MDR mechanisms including drug inactivation/alteration, modification of drug binding sites/targets, reduced intracellular drug accumulation and biofilm formation. For targeting different types of MDR, there are multiple types of nanoparticles such as metal nanoparticles, nanostructures, leukocyte membrane-coated nanoparticles, red blood cell membrane-coated nanoparticles, cancer cell membrane-coated nanoparticles, and platelet membrane-coated nanoparticles among others. Antimicrobial nanobiotics identified and synthesized to date harbor a vast diversity of intrinsic and modified physicochemical properties and have applications in diagnostics. No technology is without its challenges and the same is true for nanobiotics. The major challenges in this field of nanobiotic-based therapeutics are their allergic responses, assembly and pharmacokinetics. This chapter will elaborate on the mechanisms of action of various types of nanobiotics present as cost-effective solutions useful in a variety of applications in the treatment of MDR pathogens with a special focus on ESKAPE pathogens.

Sepsis is a clinical syndrome resulting from infection followed by inflammation and is one of the significant causes of mortality worldwide. The underlying reason is the host's uncontrolled inflammatory response due to an infection led to multiple organ dysfunction/failure. Neutrophils, an innate immune cell, are forerunners to reach the site of infection/inflammation for clearing the infection and resolute the inflammation during sepsis. A relatively new neutrophil effector function, neutrophil extracellular traps (NETs), have been demonstrated to kill the pathogens by releasing DNA decorated with histone and granular proteins. A growing number of pieces of shreds of evidence suggest that unregulated incidence of NETs have a significant influence on the pathogenesis of sepsis-induced multiple organ damage, including arterial hypotension, hypoxemia, coagulopathy, renal, neurological, and hepatic dysfunction. Thus, excessive production and improper resolution of NETs are of significant therapeutic value in combating sepsis-induced multiple organ failure. The purpose of this review is intended to highlight the role of NETs in sepsis-induced organ failure. Furthermore, the current status of therapeutic strategies to intersect the harmful effects of NETs to restore organ functions is discussed.

High Resolution Melting (HRM) is a homogeneous, exceptionally incredible innovation for single nucleotide polymorphism (SNP) genotyping, mutation scanning and sequence scanning in DNA samples. HRM analysis works on the principle of melting (dissociation) curve methodologies of Polymerase Chain Reaction (PCR) empowered by the new accessibility of improved double-stranded DNA (dsDNA)–binding dyes and next-generation real-time PCR instrumentation. The HRM technology portrays samples of nucleic acids on the basis of their disassociation behaviors and identifies the differences in even the short sequence in amplified PCR products, just by direct melting. Samples are further distinguished according to the length of their sequence, GC content and strand complementarity. Indeed, even a single change in the base pair in the sequences of DNA samples causes differences in the HRM curve. The difference in the melting curves of different genetic sequences at distinct rates can be observed, detected and compared using these curves. Development of the melting curves after HRM analysis is basically plotted with temperature on the X axis and fluorescence on the Y axis, which resembles the real-time PCR amplification curve but with the difference of temperature for cycle number. With the use of different DNA dyes, high-end instrumentation and sophisticated analysis software, these distinctions are detected.

World health organization (WHO) has acknowledged the problem of multi drug resistance (MDR) bacteria as an endemic and widespread problem globally. MDR in LMICs represents one of the biggest threats to global health and is one of the greatest current challenges in infectious disease research. High mortality/morbidity rates of MDR infections are mainly due to the lack of timely, rapid detection and treatment of the causative pathogen. Molecular mechanism conferring MDR against most common treatment options includes mutations in antibiotics’ susceptible genes at one or many sites. A number of methods, including culture-based, nucleic acid-based amplification of resistance conferring genes, and immunological based assays have been developed to detect MDR. Each method has defined specificity, sensitivity and time around to detect MDR infections in clinical settings.

Penicillin discovery has put forward great expectations and hope for the treatment of several infectious diseases. Inappropriate and excess use of antibiotics has led to the emergence of antibiotic-resistant (AMR) worldwide, which has become one of the greatest threats to global health. However, in the late 1940s, after approval, mass production (lead to reduced cost) and supply (lead to easy access to all people) led to the emergence of Antimicrobial Resistance (AMR). A similar behavioral pattern ensued as other classes of antibiotics were discovered (through increasing utilization to resistance). Substandard infection control practices in public healthcare settings eased the spread and transmission of resistant organisms and intensified antimicrobials' effect. The healthcare community responded with two major programs – Infection Control in the 1980s and Antimicrobial Stewardship (in the last decade). These programs depend on the end-user; however, while the importance of such global control and prevention programs cannot be disputed, these efforts alone are insufficient against the advent of AMR. Also, drug discovery has suffered from a shortage of exploitable bacterial target sites, leading to the slow evolution of novel potent drugs.

Background: Enterococcus faecalis (Ef) infections are becoming dreadfully common in hospital environments. Infections caused by Ef are difficult to treat because of its acquired resistance to different class of antibiotics, making it a multidrug resistant bacteria. Key pathogenic factor of Ef includes its ability to form biofilm on the surface of diagnostic and other medical devices. Sortase A (SrtA) is a cysteine transpeptidase which plays a pivotal role in the formation of biofilm in Ef, hence, it is considered as an important enzyme for the pathogenesis of Ef. Thus, inhibition of (SrtA) will affect biofilm formation, which will reduce its virulence and eventually Ef infection will be abridged. Objective: To find potential inhibitors of Enterococcus faecalis Sortase A (EfSrtA) through insilico and in-vitro methods. Methods: Gene coding for EfSrtA was cloned, expressed and purified. Three-dimensional model of EfSrtA was created using Swiss-Model workspace. In-silico docking studies using Autodock vina and molecular dynamics simulations of the modelled structures using Gromacs platform were performed to explore potential lead compounds against EfSrtA. In-vitro binding experiments using spectrofluorometric technique was carried out to confirm and validate the study. Results: In-silico docking and in-vitro binding experiments revealed that curcumin, berberine and myricetin bound to EfSrtA at nanomolar concentrations with high affinity. Conclusion: This is a first structural report of EfSrtA with curcumin, berberine and myricetin. Taking in account the herbal nature of these compounds, the use of these compounds as inhibitors will be advantageous. This study validated curcumin, berberine and myricetin as potential inhibitors of EfSrtA.

Enterococcus faecalis (E. faecalis) is an opportunistic multidrug-resistant (MDR) pathogen found in the guts of humans and farmed animals. Due to the occurrence of (MDR) strain there is an urgent need to look for an alternative treatment approach. E. faecalis is a Gram-positive bacterium, which is among the most prevalent multidrug resistant hospital pathogens. Its ability to develop quorum sensing (QS) mediated biofilm formation further exacerbates the pathogenicity and triggers lifethreatening infections. Therefore, developing a suitable remedy for curing E. faecalis mediated enterococcal infections is an arduous task. Several putative virulence factors and proteins are involved in the development of biofilms in E. faecalis. Such proteins often play important roles in virulence, disease, and colonization by pathogens. The elucidation of the structure-function relationship of such protein drug targets and the interacting compounds could provide an attractive paradigm towards developing structure-based drugs against E. faecalis. This review provides a comprehensive overview of the current status, enigmas that warrant further studies, and the prospects toward alleviating the antibiotic resistance in E. faecalis. Specifically, the role of biofilm and quorum sensing (QS) in the emergence of MDR strains had been elaborated along with the importance of the protein drug targets involved in both the processes.

Chronic obstructive pulmonary disease (COPD) is pulmonary emphysema characterized by blockage in the airflow resulting in the long-term breathing problem, hence a major cause of mortality worldwide. Excessive generation of free radicals and the development of chronic inflammation are the major two episodes underlying the pathogenesis of COPD. Currently used drugs targeting these episodes including anti-inflammatory, antioxidants, and corticosteroids are unsafe, require high doses, and pose serious side effects. Nanomaterial-conjugated drugs have shown promising therapeutic potential against different respiratory diseases as they are required in small quantities which lower overall treatment costs and can be effectively targeted to diseased tissue microenvironment hence having minimal side effects. Poly lactic-co-glycolic acid (PLGA) nanoparticles (NPs) are safe as their breakdown products are easily metabolized in the body. Drugs loaded on the PLGA NPs have been shown to be promising agents as anticancer, antimicrobial, antioxidants, and anti-inflammatory. Surface modification of PLGA NPs can further improve their mechanical properties, drug loading potential, and pharmacological activities. In the present review, we have presented a brief insight into the pathophysiological mechanism underlying COPD and highlighted the role, potential, and current status of PLGA NPs loaded with drugs in the therapy of COPD.

Entamoeba histolytica is the conductive agent of amoebiasis. Upon the parasite's infection, macrophages and neutrophils are activated by Interferon γ (IFN-γ), IL-13, and Tumour Necrosis Factor (TNF-α). These immune cells then carry out the amoebicidal activity by releasing Nitric Oxide Synthase (NOS) and Reactive Oxygen Species (ROS). This review talks about the protective and destructive role of ROS in Eh. E. histolytica has defense strategies against oxidative stress which is a result of excess ROS production. They possess antioxidants for their defense such as L-Cysteine, Flavodiiron proteins (FDPs), Peroxiredoxin, and Trichostatin A, which contribute to the parasite's virulence. The reactive oxygen species are harmful to the host cells as excess ROS production stimulates cell death by mechanisms like apoptosis and necroptosis. NADPH oxidase (NOX) is a key source of ROS in mammalian cells and causes apoptosis of host cells via the protein kinase transduction pathway. This review provides insights into why NOX inhibitors that could be a potent antiparasitic drug, is not effective for in vivo purposes. This paper also gives an insight into a solution that could be a potent source in generating new treatment and vaccines for amoebiasis by targeting parasite development.

The regulation of the activity of proteases by endogenous inhibitors is a common trend in almost all forms of life. Here, we review the endogenous inhibitors of cysteine proteases of three major pathogenic parasitic protozoa. The review focuses on members of the genus Plasmodium, Entamoeba, and Leishmania. Research in this domain has revealed the presence of only chagasin-like inhibitors of cysteine proteases that house a β-barrel immunoglobulin-fold and inhibit the target proteases using a 3-loop inhibitory mechanism in these pathogens. Inhibitors of cysteine proteases are highly evolvable enzymes that target a broad spectrum of pathogenic cysteine proteases with a proclivity for those involved in host-parasite interactions. A common trend reflects a limited sequence homology between cysteine proteases and their inhibitors. The inhibitors are also known to participate in other housekeeping functions of the parasites. Generalizations about their roles are thus best avoided. In this review, the reader will find comprehensive information on the cellular localization of inhibitors of cysteine proteases, their structure, function, and the associated mechanisms of action. The reader will also find a thorough analysis of the role of these inhibitors in parasite pathology and the common trends interlinking them with parasite biology and evolution.

Sepsis, which includes infection followed by inflammation, is one of the leading causes of death among neonates worldwide. The major attribute of this disease process is dysregulated host response to infection leading to organ dysfunction and potentially death. A comprehensive understanding of the host response as well as the pathogen itself are important factors contributing to outcome. Early diagnosis is paramount, as it leads to accurate assessment and improved clinical management. Accordingly, a number of diagnostic platforms have been introduced to assess the presence of blood stream pathogens in septic neonates. Unfortunately, current point-of-care (POC) methods rely on a single parameter/biomarker and thus lack a comprehensive evaluation. The emerging field of biosensing has, however, resulted in the development of a wide range of analytical devices that may be useful at POC. This review discusses currently available methods to screen the inflammatory process in neonatal sepsis. We describe POC sensor-based methods for single platform multi-analyte detection and highlight the latest advances in this evolving technology. Finally, we critically evaluate the applicability of these POC devices clinically for early diagnosis of sepsis in neonates.

Millions of people worldwide lie at the risk of parasitic protozoic infections that kill over a million people each year. The rising inefficacy of conventional therapeutics to combat these diseases, mainly due to the development of drug resistance to a handful of available licensed options contributes substantially to the rising burden of these ailments. Cysteine proteases are omnipresent enzymes that are critically implicated in the pathogenesis of protozoic infections. Despite their significance and druggability, cysteine proteases as therapeutic targets have not yet been translated into the clinic. The review presents the significance of cysteine proteases of members of the genera Plasmodium, Entamoeba, and Leishmania, known to cause Malaria, Amoebiasis, and Leishmaniasis, respectively, the protozoic diseases with the highest morbidity and mortality. Further, projecting them as targets for molecular tools like the CRISPR-Cas technology for favorable manipulation, exploration of obscure genomes, and achieving a better insight into protozoic functioning. Overcoming the hurdles that prevent us from gaining a better insight into the functioning of these enzymes in protozoic systems is a necessity. Managing the burden of parasitic protozoic infections pivotally depends upon the betterment of molecular tools and therapeutic concepts that will pave the path to an array of diagnostic and therapeutic applications.

Enterococcus faecalis (E. faecalis) is a Gram-positive coccoid, non-sporulating, facultative anaerobic, multidrug resistance bacterium responsible for almost 65% to 80% of all enterococcal nosocomial infections. It usually causes infective endocarditis, urinary tract and surgical wound infections. The increase in E. faecalis resistance to conventionally available antibiotic has rekindled intense interest in developing useful antibacterial drugs. In E. faecalis, diaminopimelate epimerase (DapF) is involved in the lysine biosynthetic pathway. The product of this pathway is precursors of peptidoglycan synthesis, which is a component of bacterial cell wall. Also, because mammals lack this enzyme, consequently E. faecalis diaminopimelate epimerase (EfDapF) represents a potential target for developing novel class of antibiotics. In this regard, we have successfully cloned, overexpressed the gene encoding DapF in BL-21(DE3) and purified with Ni-NTA Agarose resin. In addition to this, binding studies were performed using fluorescence spectroscopy in order to confirm the bindings of the identified lead compounds (acetaminophen and dexamethasone) with EfDapF. Docking studies revealed that acetaminophen found to make hydrogen bonds with Asn72 and Asn13 while dexamethasone interacted by forming hydrogen bonds with Asn205 and Glu223. Thus, biochemical studies indicated acetaminophen and dexamethasone, as potential inhibitors of EfDapF and eventually can reduce the catalytic activity of EfDapF.

Background. B.1.617.1, a variant of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) causing respiratory illness is responsible for the second wave of COVID-19 and associated with a high incidence of infectivity and mortality. To mitigate the B.1.617.1 variant of SARS-CoV-2, deciphering the protein structure and immunological responses by employing bioinformatics tools for data mining and analysis is pivotal. Objectives. Here, an in silico approach was employed for deciphering the structure and immune function of the subunit of spike (S) protein of SARS-CoV-2 B.1.617.1 variant. Methods. The partial amino acid sequence of SARS-CoV-2 B.1.617.1 variant S protein was analyzed, and its putative secondary and tertiary structure was predicted. Immunogenic analyses including B- and T-cell epitopes, interferon-gamma (IFN-γ) response, chemokine, and protective antigens for SARS-CoV 2 S proteins were predicted using appropriate tools. Results. B.1.617.1 variant S protein sequence was found to be highly stable and amphipathic. ABCpred and CTLpred analyses led to the identification of two potential antigenic B cell and T cell epitopes with starting amino acid positions at 60 and 82 (for B cell epitopes) and 54 and 98 (for T cell epitopes) having predictionscores>0.8. Further, RAMPAGE tool was used for determining the allowed and disallowed regions of the three-dimensional predicted structure of SARS-CoV-2 B.1.617.1 variant S protein. Conclusion. Together, the in silico analysis revealed the predicted structure of partial S protein, immunogenic properties, and possible regions for S protein of SARS-CoV-2 and provides a valuable prelude for engineering the targeted vaccine or drug against B.1.617.1 variant of SARS-CoV-2.

Entamoeba histolytica (Eh) is a pathogenic eukaryote that often resides silently in humans under asymptomatic stages. Upon indeterminate stimulus, it develops into fulminant amoebiasis that causes severe hepatic abscesses with 50% mortality. This neglected tropical pathogen relies massively on membrane modulation to flourish and cause disease; these modulations range from the phagocytic mode for food acquisition to a complex trogocytosis mechanism for tissue invasion. Rab GTPases form the largest branch of the Ras-like small GTPases, with a diverse set of roles across the eukaryotic kingdom. Rab GTPases are vital for the orchestration of membrane transport and the secretory pathway responsible for transporting the pathogenic effectors, such as cysteine proteases (EhCPs) which help in tissue invasion. Rab GTPases thus play a crucial role in executing the cytolytic effect of E. histolytica. First, they interact with Gal/Nac lectins required for adhering to the host cells, and then, they assist in the secretion of EhCPs. Additionally, amoebic Rab GTPases are vital for encystation because substantial vesicular trafficking is required to create dormant amoebic cysts. These cysts are the infective agent and help to spread the disease. The absence of a “bonafide” vesicular transport machinery in Eh and the existence of a diverse repertoire of amoebic Rab GTPases (EhRab) hint at their contribution in supporting this atypical machinery. Here, we provide insights into a pseudoRab GTPase, EhRabX10, by performing physicochemical analysis, predictive 3D structure modeling, protein-protein interaction studies, and in silico molecular docking. Our group is the first one to classify EhRabX10 as a pseudoRab GTPase with four nonconserved G-motifs. It possesses the basic fold of the P-loop containing nucleotide hydrolases. Through this in silico study, we provide an introduction to the characterization of the atypical EhRabX10 and set the stage for future explorations into the mechanisms of nucleotide recognition, binding, and hydrolysis employed by the pseudoEhRab GTPase family.

Entamoeba histolytica (Eh), a parasitic protozoan and the causative agent of invasive Amoebiasis, invade the host tissue through an effective secretory pathway. There are several lines of evidence suggesting that amoebic trophozoite pore-forming complex amoebapore and a large class of proteases enzymes including rhomboid proteases, cysteine proteases, and metalloproteases are implicated in host tissue invasion. For successful delivery of these molecules/cargos, trophozoites heavily rely on sorting machinery from the endoplasmic reticulum, Golgi to plasma membrane. Although, sole secretion machinery in E. histolytica is not characterized yet. Therefore, here our aim is to understand the properties of key molecules N-ethylmaleimide-sensitive fusion protein attached to protein receptors (SNAREs) in E. histolytica. SNAREs proteins are an important component of the membrane-trafficking machinery and have been associated in a range of processes including vesicle tethering, fusion as well as specificity of vesicular transport in all eukaryotic cells. SNARE proteins are architecturally simple, categorized by the presence of one copy of a homologous coiled-coil forming motif. However, the structural information and protein-protein interaction study of Eh-associated syntaxin proteins are still not known. Here, we characterize the syntaxin 1 like molecule and VAMP from Eh through physiochemical profiling, modeling, atomistic simulation, protein-protein interaction, and docking approaches on the proteins containing SNARE and synaptobrevin domain. The modeled structures and the critical residues recognized through protein interaction and docking study may provide better structural and functional insights into these proteins and may aid in the development of newer diagnostic assays.

Sepsis is one of the leading causes of mortality among critically ill patients globally. According to WHO report 2018, it is estimated to affect beyond 30 million people worldwide every year. It causes loss of human lives, which arise from infection and inflammation and long term stay in intensive care unit (ICU) in hospitals. Despite the availability of satisfactory prognostic markers contributing to the diagnosis of sepsis, millions of people die even after admission to the hospitals. Correct and early diagnosis of sepsis leads to rapid administration of appropriate antibiotics can thus potentially avert the attainment to critical stages of sepsis, thereby saving human lives. Conventional diagnostic practices are costly, time consuming and they lack adequate sensitivity and selectivity, provoking an urgent need for developing alternate sepsis diagnosis systems. Nevertheless, biosensors have the much-treasured scope for reasonable sepsis diagnosis. Advancement in nano-biotechnology has provided new paradigm for biosensor platforms with upgraded features. Here, we provide an overview of the recent advances in biosensors with a brief introduction to sepsis, followed by the conventional methods of diagnosis and bio-sensing. To conclude, a proactive role and an outlook on technologically advanced biosensor platforms are discoursed with possible biomedical applications.

Amoebiasis is a parasitic infectious disease caused by the enteric protozoan Entamoeba histolytica, a leading basis of deaths accounted to parasites, succeeding malaria and schistosomiasis. Conventional treatment methodologies used to deal with amoebiasis mainly rely on the administration of anti-amoebic compounds and vaccines but are often linked with substantial side-effects on the patient. Besides, cases of development of drug resistance in protozoans have been recorded, contributing further to the reduction in the efficiency of the treatment. Loopholes in the efficacious management of the disease call for the development of novel methodologies to manage amoebiasis. A way to achieve this is by targeting the essential metabolic processes of 'encystation' and 'excystation', and the associated biomolecules, thus interrupting the biphasic life cycle of the parasite. Technologies like the CRISPR-Cas9 system can efficiently be exploited to discover novel and essential molecules that regulate the protozoan's metabolism, while efficiently manipulating and managing the known drug targets, leading to an effective halt and forestall to the enteric infection. This review presents a perspective on these essential metabolic processes and the associated molecules that can be targeted efficaciously to prevent the transmission of amoebiasis, thus managing the disease and proving to be a fruitful endeavour.

Enterococcus faecalis (Ef) is a Gram positive multidrug resistant (MDR) bacterium contributing about 70% of total enterococcal infections. In Ef, a membrane anchored transpeptidase Sortase A plays a major role in biofilm formation. Therefore, it has been recognized as an ideal drug target against Ef. In this regard to identify the potential inhibitors of Ef Sortase A (EfSrtA∆59), we have cloned, expressed and purified EfSrtA∆59. We have also done the in-silico docking studies to identify lead molecules interacting with EfSrtA∆59. Furthermore, the binding studies of these identified lead molecules were performed with EfSrtA∆59 using fluorescence and CD spectroscopic studies. We also identified the interaction partner of EfSrtA∆59 using STRING. Protein-protein docking studies were also performed. Docking experiment revealed that benzylpenicillin, cefotaxime, pantoprazole and valsartan were bound to same site on the protein with similar interactions. Binding studies using fluorescence spectroscopic studies confirmed the binding of all the ligands to EfSrtA∆59, which was further validated by far and near-UV CD experiments. Thermo stability experiments validate the stability-activity trade-off hypothesis. Sequence based interaction studies identified that EfSrtA∆59 interact with the Ef_1091, Ef_1093 and Ef_2658 proteins. Homology model of Ef_1091 and Ef_1093 was docked with modeled EfSrtA∆59 and their interactions are also discussed.

Multiple drug resistance and treatment of contaminated water has become a serious issue in past years. Silver nanoparticles (AgNPs), being bactericidal, non-toxic, cheap and environment friendly behaviour, have drawn attention to overcome these problems. This study has been designed to synthesise AgNPs from Pseudomonas aeruginosa. AgNPs formation was confirmed by colour change and UV-vis spectroscopy. Furthermore, Fourier transform infrared spectroscopy peaks demonstrated the presence of capped proteins as reducing and stabilising agent. Transmission electron microscopy micrograph revealed spherical shape AgNPs with the size ranging between 10 and 20 nm. Antibacterial activity of AgNPs was evaluated against the most prevalent waterborne pathogens enterohaemorrhagic Escherichia coli and Salmonellae typhimurium. Moreover, the antibacterial activity of AgNPs was tested for the treatment of contaminated water which showed attenuation in bacterial load within 8 h as demonstrated by growth kinetics data. Furthermore, AgNPs did not exhibit haemolytic effects on human red blood cells (RBCs) even at 100 mg L-1 concentration of AgNPs. The results herein suggest that AgNPs synthesised by P. aeruginosa under optimised conditions exhibit microbicidal property against waterborne pathogens and having no toxic effect on human RBCs. These AgNPs could be employed for treatment of contaminated water after process optimisation.

The enteric protozoan parasite, Entamoeba histolytica (Eh), is the causative agent of amoebic dysentery and liver abscess in humans. It infects around 50 million people worldwide, which is a third general cause of death from parasitic diseases after malaria and schistosomiasis. The other prevalent form of the disease is Visceral leishmaniasis caused by Leishmania donovani which is a human blood parasite. On the other hand, the Toxoplasma gondii is an obligate intracellular protozoan parasite; it causes serious opportunistic infections in HIV-positive persons. The biological processes in all living organisms are mostly mediated by the proteins, and recognizing new target proteins and finding their function in pathogenesis will help in choosing better diagnostic markers. In eukaryotes, Rab protein plays a major role in pathogenesis. Rabs represent the largest branch in the Ras superfamily of GTPases. Among them, the Rab5 is important in the endocytosis and thus involved in pathogenesis. In this paper, we discussed the physiochemical profiling, modelling, and docking of the Rab5 protein from pathogenic species that is Entamoeba histolytica, Leishmania donovani, and Toxoplasma gondii. The modeled structures from this study and the key residues identified would give a better understanding of the three-dimensional structure and functional insights into these proteins and help in developing new drug targets.

Enterococcus faecalis is a gram-positive, rod-shape bacteria responsible for around 65% to 80% of all enterococcal nosocomial infections. It is multidrug resistant (MDR) bacterium resistant to most of the first-line antibiotics. Due to the emergence of MDR strains, there is an urgent need to find novel targets to develop new antibacterial drugs against E. faecalis. In this regard, we have identified naphthoate synthase (1,4-dihydroxy-2-naphthoyl-CoA synthase, EC: 4.1.3.36; DHNS) as an anti-E. faecalis target, as it is an essential enzyme for menaquinone (vitamin K2 ) synthetic pathway in the bacterium. Thus, inhibiting naphtholate synthase may consequently inhibit the bacteria's growth. In this regard, we report here cloning, expression, purification, and preliminary structural studies of naphthoate synthase along with in silico modeling, molecular dynamic simulation of the model and docking studies of naphthoate synthase with quercetin, a plant alkaloid. Biochemical studies have indicated quercetin, a plant flavonoid as the potential lead compound to inhibit catalytic activity of EfDHNS. Quercetin binding has also been validated by spectrofluorimetric studies in order to confirm the bindings of the ligand compound with EfDHNS at ultralow concentrations. Reported studies may provide a base for structure-based drug development of antimicrobial compounds against E. faecalis.

Background Previous study from this lab has discerned oxidative, nitrosative stress and their relationship with cytokines contributing to the severity of sepsis and organ dysfunction. Cytokines are known to induce neutrophil extracellular traps (NETs) formation via free radicals generation. Hyper-activation of neutrophil leads to the increased NETs formation or ineffective clearance of NETs would likely increase the risk of auto-antibody generation against NETs components and being partly responsible for the sepsis severity and organ dysfunction. The present study was undertaken to further assess the status of NETs formation and their correlation with severity of sepsis, with the cytokines and organ dysfunction. Methods The level of NETs formation, DNA release, elastase release, and inflammatory cytokines was determined in 80 sepsis patients and 45 healthy volunteers. Their linearity with organ parameters and associations with sepsis severity were also assessed. Results NETs formation experiment was carried out and it was significantly higher in sepsis (70%) compared to control (30%). NETs % were positively correlated with severity of sepsis and organ dysfunction. Pearson's correlation coefficient demonstrated a direct relation between NETs components and organ parameters with Sepsis severity scores. Conclusion NETs formation is significantly higher due to which it is contributing to the sepsis severity and organ failure.

Enterococcus faecalis (Ef) is one of the major pathogens involved in hospital-acquired infections. It can cause nosocomial bacteremia, surgical wound infection, and urinary tract infection. It is important to mention here that Ef is developing resistance against many commonly occurring antibiotics. The occurrence of multidrug resistance (MDR) and extensive-drug resistance (XDR) is now posing a major challenge to the medical community. In this regard, to combat the infections caused by Ef, we have to look for an alternative. Rational structure-based drug design exploits the three-dimensional structure of the target protein, which can be unraveled by various techniques such as X-ray crystallography or nuclear magnetic resonance (NMR) spectroscopy. In this review, we have discussed the complete picture of Ef infections, the possible treatment available at present, and the alternative treatment options to be explored. This study will help in better understanding of novel biological targets against Ef and the compounds, which are likely to bind with these targets. Using these detailed structural informations, rational structure-based drug design is achievable and tight inhibitors against Ef can be prepared.

Phosphorus (P), an essential macronutrient, is pivotal for growth and development of plants. Availability of phosphate (Pi), the only assimilable P, is often suboptimal in rhizospheres. Pi deficiency triggers an array of spatiotemporal adaptive responses including the differential regulation of several transcription factors (TFs). Studies on MYB TF PHR1 in Arabidopsis thaliana (Arabidopsis) and its orthologs OsPHRs in Oryza sativa (rice) have provided empirical evidence of their significant roles in the maintenance of Pi homeostasis. Since the functional characterization of PHR1 in 2001, several other TFs have now been identified in these model plants. This raised a pertinent question whether there are any likely interactions across these TFs. Clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated protein 9 (Cas9) system has provided an attractive paradigm for editing genome in plants. Here, we review the applications and challenges of this technique for genome editing of the TFs for deciphering the function and plausible interactions across them. This technology could thus provide a much-needed fillip towards engineering TFs for generating Pi use efficient plants for sustainable agriculture. Furthermore, we contemplate whether this technology could be a viable alternative to the controversial genetically modified (GM) rice or it may also eventually embroil into a limbo.

Present study utilised textile soil isolated bacterium Pseudomonas stutzeri to synthesise extracellular silver nanoparticles (AgNPs) under optimised conditions. The synthesised AgNPs were characterised using ultraviolet-visible spectroscopy, Fourier transform infrared spectroscopy (FTIR) and transmission electron microscopy (TEM). Optimisation showed AgNPs synthesis within 8 h using 2mM Ag nitrate at pH9, temperature 80°C and maximum absorbance toward 400 nm. TEM analysis revealed spherical shape AgNPs and reduction in size upto 8 nm was observed under optimised conditions. FTIR spectra confirmed presence of proteins bound to AgNPs act as reducing agent. AgNPs showed strong antibacterial activity against multi-drug resistant (MDR) Escherichia coli and Klebsiella pneumoniae as demonstrated by disc diffusion and colony forming unit assays. Zone of inhibition increased with increasing concentration of AgNPs with maximum of 19 mm against E. coli and 17 mm against K. pneumoniae at concentration of 2 μg/disc. Furthermore, AgNPs did not show any cytotoxic effects on human epithelial cells as demonstrated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay even at 2 μg/ml concentration of AgNPs. The results of the present study suggest that AgNPs can be synthesised rapidly under optimised conditions and show strong antimicrobial property against MDR pathogens without having toxicity effect on human epithelial cells.