Development Of Novel Antibacterial Drugs Research Articles

Riboswitches as therapeutic targets: promise of a new era of antibiotics

ABSTRACT Introduction The growth of antibiotic resistance among bacterial pathogens is an impending global threat that can only be averted through the development of novel antibacterial drugs. A promising answer could be the targeting of riboswitches, structured RNA elements found almost exclusively in bacteria. Areas covered This review examines the potential of riboswitches as novel antibacterial drug targets. The limited mechanisms of action of currently available antibiotics are summarized, followed by a delineation of the functional mechanisms of riboswitches. We then discuss the potential for developing novel approaches that target paradigmatic riboswitches in the context of their bacterial gene expression machinery. Expert opinion We highlight potential advantages of targeting riboswitches in their functional form, embedded within gene expression complexes critical for bacterial survival. We emphasize the benefits of this approach, including potentially higher species specificity and lower side effects.

The usnic acid derivative peziculone targets cell walls of Gram-positive bacteria revealed by high-throughput CRISPRi-seq analysis

Usnic acid, a representative dibenzofuran metabolite, is known to have antimicrobial properties. However, despite considerable interest as an antimicrobial agent, the mechanism by which usnic acid and its derivatives exert their action is not fully characterized. This article describes the synthesis of peziculone, a 5:1 equilibrium mixture of two inseparable usnic acid derivatives: peziculone A and peziculone B. The antibacterial activity of peziculone against several Gram-positive bacterial pathogens was found to be significantly better compared with usnic acid. Clustered regularly interspaced short palindromic repeats interference sequencing analysis and membrane fluorescent staining were used to demonstrate that peziculone destabilizes the cell walls of Gram-positive bacteria. Additionally, peziculone 2.5 and 3.5 µg/mL impaired cell surface appendages and biofilm formation by Staphylococcus aureus. Taken together, these data demonstrate that peziculone, a derivative compound of usnic acid, has significant antimicrobial activity against Gram-positive bacteria by targeting the cell walls; this provides a platform for development of novel antibacterial drugs.

Statistical Medium Optimization for the Production of Anti-Methicillin-Resistant Staphylococcus aureus Metabolites from a Coal-Mining-Soil-Derived Streptomyces rochei CMB47

The development of novel antibacterial drugs needs urgent action due to the global emergence of antibiotic resistance. In this challenge, actinobacterial strains from arid ecosystems are proving to be promising sources of new bioactive metabolites. The identified Streptomyces rochei strain CMB47, isolated from coal mine Saharan soil, provided an ethyl acetate extract which tested against a series of pathogens. It displayed a minimum inhibitory concentration of <0.439 µg/mL against MRSA. A statistical experimental design using a response surface methodology (RSM) based on the second-order rotatable central composite design (RCCD) was planned to develop an efficient fermentation process able to improve the bioactive metabolite production. The optimal conditions were determined for starch and NaNO3 concentrations, incubation time and the initial pH value, reaching the inhibition zone diameter of 20 mm, close to the experimental value, after validation of the model. A bioassay-guided fractionation of the crude extract provided the most active fractions, which were analyzed by HPLC equipped with a photodiode array detector and coupled online with an electrospray mass spectrometer (HPLC-DAD/ESI-MS), obtaining preliminary indications on the molecular structures of the metabolites. These results support the potential interest in further investigations into the purification and full characterization of the metabolites responsible for the biological activity observed so far.

Recent Advances in Using Natural Antibacterial Additives in Bioactive Wound Dressings.

Wound care is a global health issue with a financial burden of up to US $96.8 billion annually in the USA alone. Chronic non-healing wounds which show delayed and incomplete healing are especially problematic. Although there are more than 3000 dressing types in the wound management market, new developments in more efficient wound dressings will require innovative approaches such as embedding antibacterial additives into wound-dressing materials. The lack of novel antibacterial agents and the misuse of current antibiotics have caused an increase in antimicrobial resistance (AMR) which is estimated to cause 10 million deaths by 2050 worldwide. These ongoing challenges clearly indicate an urgent need for developing new antibacterial additives in wound dressings targeting microbial pathogens. Natural products and their derivatives have long been a significant source of pharmaceuticals against AMR. Scrutinising the data of newly approved drugs has identified plants as one of the biggest and most important sources in the development of novel antibacterial drugs. Some of the plant-based antibacterial additives, such as essential oils and plant extracts, have been previously used in wound dressings; however, there is another source of plant-derived antibacterial additives, i.e., those produced by symbiotic endophytic fungi, that show great potential in wound dressing applications. Endophytes represent a novel, natural, and sustainable source of bioactive compounds for therapeutic applications, including as efficient antibacterial additives for chronic wound dressings. This review examines and appraises recent developments in bioactive wound dressings that incorporate natural products as antibacterial agents as well as advances in endophyte research that show great potential in treating chronic wounds.

The Multifaceted MEP Pathway: Towards New Therapeutic Perspectives.

Isoprenoids, a diverse class of natural products, are present in all living organisms. Their two universal building blocks are synthesized via two independent pathways: the mevalonate pathway and the 2-C-methyl-ᴅ-erythritol 4-phosphate (MEP) pathway. The presence of the latter in pathogenic bacteria and its absence in humans make all its enzymes suitable targets for the development of novel antibacterial drugs. (E)-4-Hydroxy-3-methyl-but-2-enyl diphosphate (HMBPP), the last intermediate of this pathway, is a natural ligand for the human Vγ9Vδ2 T cells and the most potent natural phosphoantigen known to date. Moreover, 5-hydroxypentane-2,3-dione, a metabolite produced by Escherichia coli 1-deoxy-ᴅ-xylulose 5-phosphate synthase (DXS), the first enzyme of the MEP pathway, structurally resembles (S)-4,5-dihydroxy-2,3-pentanedione, a signal molecule implied in bacterial cell communication. In this review, we shed light on the diversity of potential uses of the MEP pathway in antibacterial therapies, starting with an overview of the antibacterials developed for each of its enzymes. Then, we provide insight into HMBPP, its synthetic analogs, and their prodrugs. Finally, we discuss the potential contribution of the MEP pathway to quorum sensing mechanisms. The MEP pathway, providing simultaneously antibacterial drug targets and potent immunostimulants, coupled with its potential role in bacterial cell-cell communication, opens new therapeutic perspectives.

Targeting multidrug resistant Staphylococcus infections with bacterial histidine kinase inhibitors.

The emergence of drug-resistant bacteria, such as methicillin-resistant Staphylococcus aureus (MRSA), which are not susceptible to current antibiotics has necessitated the development of novel approaches and targets to tackle this growing challenge. Bacterial two-component systems (TCSs) play a central role in the adaptative response of bacteria to their ever-changing environment. They are linked to antibiotic resistance and bacterial virulence making the proteins of the TCSs, histidine kinases and response regulators, attractive for the development of novel antibacterial drugs. Here, we developed a suite of maleimide-based compounds that we evaluated against a model histidine kinase, HK853, in vitro and in silico. The most potent leads were then assessed for their ability to decrease the pathogenicity and virulence of MRSA, resulting in the identification of a molecule that decreased the lesion size caused by a methicillin-resistant S. aureus skin infection by 65% in a murine model.

Antibacterial Properties and Potential Mechanism of Serum from Chinese Alligator.

The Chinese alligator (Alligator sinensis) is an ancient reptile with strong immunity that lives in wetland environments. This study tested the antibacterial ability of Chinese alligator serum (CAS) against Klebsiella pneumoniae, Escherichia coli, Staphylococcus aureus, and Pseudomonas aeruginosa and analyzed the potential underlying mechanisms. Results showed that the CAS had a marked antibacterial effect on K. pneumoniae, E. coli, and P. aeruginosa, while S. aureus was only mildly affected. However, these effects disappeared when Protease K was added to the serum. The serum proteome analysis revealed that the antibacterial ability of CAS was produced by interactions among various proteins and that the complement proteins played a major antibacterial role. Therefore, we made relevant predictions about the structure and function of complement component 3. In addition, sequence alignment and phylogenetic analysis of complement component 3d (C3d) in four mammalian species and two alligator species showed that the amino acids that make up the acid pocket on the concave surface of alligator C3d are not identical to those in mammals. This study provided evidence that CAS elicits significant antibacterial effects against some pathogens and provides the basis for further development of novel antibacterial drugs.

GCN-Based Structure-Activity Relationship and DFT Studies of Staphylococcus aureus FabI Inhibitors

The enoyl-[acyl-carrier-protein] reductase (FabI) is an important enzyme in the fatty acid metabolism of Gram-positive bacteria, such as Staphylococcus aureus. FabI is also a potential target for the development of novel antibacterials. Several machine learning-driven studies were reported to develop FabI inhibitors, describing robust and predictive models. Herein, the authors applied the kGCN, a graph convolutional network framework, to generate classification models to select potential S. aureus FabI inhibitors. The most predictive model showed robustness for both active and inactive class prediction, according to statistical validation. Finally, the chemical interpretation of the model was consistent with prior experimental and theoretical works. The SAR analysis highlighted the importance of the occupation of hydrophobic pockets and polar interactions with Tyr-156 and NADPH cofactor present in the FabI catalytic site by potential inhibitors. A density functional theory study endorsed the SAR, where the electrostatic surfaces were consistent with the expected interactions with the pocket.

Redox Impact on Bacterial Macromolecule: A Promising Avenue for Discovery and Development of Novel Antibacterials.

Antibiotic resistance in bacteria has remained a serious public health concern, resulting in substantial deaths and morbidity each year. Factors such as mutation and abuse of currently available antibiotics have contributed to the bulk of the menace. Hence, the introduction and implementation of new therapeutic strategies are imperative. Of these strategies, data supporting the role of reactive oxygen species (ROS) in bacterial lethality are intriguing, with several antimicrobials, including antibiotics such as fluoroquinolones, β-lactams, and aminoglycosides, as well as natural plant compounds, being remarkably implicated. Following treatment with ROS-inducing antimicrobials, ROS such as O2•-, •OH, and H2O2 generated in bacteria, which the organism is unable to detoxify, damage cellular macromolecules such as proteins, lipids, and nucleic acids and results in cell death. Despite the unique mechanism of action of ROS-inducing antibacterials and significant studies on ROS-mediated means of bacterial killing, the field remains a topical one, with contradicting viewpoints that require frequent review. Here, we appraised the antibacterial agents (antibiotics, natural and synthetic compounds) implicated in ROS generation and the safety concerns associated with their usage. Further, background information on the sources and types of ROS in bacteria, the mechanism of bacterial lethality via oxidative stress, as well as viewpoints on the ROS hypothesis undermining and solidifying this concept are discussed.

In Vitro Study of the Effect of Inhibition of Quorum Sensing by Brominated Furanone on Peritoneal Dialysis-Associated Peritonitis Associated with Escherichia Coli Infection.

In recent years, the occurrence of peritoneal dialysis (PD)-associated peritonitis (PDAP) with Escherichia coli infection has gradually increased. The presence of quorum sensing (QS) among bacteria facilitates the expansion of antibiotic resistance. Brominated furanone (BMF), a halogenated furanone compound isolated from macroalgae, is a new type of quorum-sensing inhibitor that can inhibit bacterial quorum sensing and reduce bacterial resistance. In this study, we established an in vitro peritoneal dialysis-associated peritonitis biofilm model. After intervention with BMF, the biofilm was destroyed, as shown by scanning electron microscopy, and the number of viable bacteria was reduced. Crystal violet semiquantitative determination showed that biofilm absorption significantly decreased, and RT-PCR showed that luxS expression was downregulated after drug intervention. Therefore, we propose that BMF can effectively inhibit E. coli QS by disrupting the bacterial biofilm and downregulating QS gene expression to reduce the bacterial resistance, providing a direction for the development of novel antibacterial drugs.

Novel metronidazole-derived three-component hybrids as promising broad-spectrum agents to combat oppressive bacterial resistance

The dreadful bacterial resistance to clinical drugs calls for the development of novel antibacterials. This work developed a class of unique metronidazole-derived three-component hybrids as promising antibacterial therapeutic alternatives. Bioactive assay discovered that p-chlorophenylhydrazone derivative 6b possessed excellent ability to suppress the growth of drug-resistant E. coli (MIC = 0.5 µg/mL), being 16 folds more potent than norfloxacin (MIC = 8 µg/mL). The active molecule 6b with imperceptible hemolysis could effectively retard the development of bacterial drug resistance within 30 passages. Moreover, compound 6b displayed a favorable inhibitory effect on E. coli biofilms and could act rapidly in bactericidal efficacy. Subsequent exploration of mechanism revealed that 6b could destruct the bacterial cytoplasmic membrane, leading to the leakage of intracellular protein. The inactivation of lactate dehydrogenase, metabolic stagnation and the accumulation of reactive oxygen species caused by 6b were observed. Furthermore, molecule 6b could form a supramolecular complex with DNA to obstruct DNA replication. These results demonstrated that metronidazole-derived three-component hybrids provided a large potential for deep development as prospective antibacterial agents.

The diversity of heme sensor systems - heme-responsive transcriptional regulation mediated by transient heme protein interactions.

Heme is a versatile molecule that is vital for nearly all cellular life by serving as prosthetic group for various enzymes or as nutritional iron source for diverse microbial species. However, elevated levels of heme is toxic to cells. The complexity of this stimulus has shaped the evolution of diverse heme sensor systems, which are involved in heme-dependent transcriptional regulation in eukaryotes and prokaryotes. The functions of these systems are manifold-ranging from the specific control of heme detoxification or uptake systems to the global integration of heme and iron homeostasis. This review focuses on heme sensor systems, regulating heme homeostasis by transient heme protein interaction. We provide an overview of known heme-binding motifs in prokaryotic and eukaryotic transcription factors. Besides the central ligands, the surrounding amino acid environment was shown to play a pivotal role in heme binding. The diversity of heme-regulatory systems, therefore, illustrates that prediction based on pure sequence information is hardly possible and requires careful experimental validation. Comprehensive understanding of heme-regulated processes is not only important for our understanding of cellular physiology, but also provides a basis for the development of novel antibacterial drugs and metabolic engineering strategies.

Antimicrobial activities and mechanisms of extract and components of herbs in East Asia.

Antibacterial drugs face increasing challenges due to drug resistance and adverse reactions, which has created a pressing need for the discovery and development of novel antibacterial drugs. Herbs have played an important role in the treatment of infectious diseases. This review aims to summarize, analyze and evaluate the antibacterial activities and mechanisms of components from popular herbs in East Asia. In this review, we have searched and summarized the scientific papers published during the past twenty-year period from electronic databases such as PubMed, ScienceDirect, and Web of Science. These herbs and their components, including alkaloids, flavonoids, essential oils, terpenes, organic acids, coumarins and lignans, display potential antimicrobial effects. Herbal medicine formulas (HMFs) usually show stronger antibacterial activity than single herbs. Herbs and HMFs bring forth antibacterial activities by damaging cell membranes and walls, inhibiting nucleic acid and protein synthesis, and increasing intracellular osmotic pressure. These herbs and their components can be developed as potential and promising novel antibacterial herbal products.

Synthesis and biological evaluation of indole-based peptidomimetics as antibacterial agents against Gram-positive bacteria

The emergence of bacterial multidrug resistance and the lack of new antimicrobial agents urgently demand the discovery and development of novel antibacterials that avoid bacterial resistance. Antimicrobial peptidomimetics represent a promising approach for overcoming antibiotic resistance. Herein we report the synthesis and evaluation of indole-based amphiphilic antimicrobial peptidomimetics, bearing hydrophobic side chains and hydrophilic cationic moieties. Among these derivatives, compound 28 demonstrated potent antimicrobial activity against Gram-positive bacteria, low hemolytic activity and low toxicity towards mammalian cells, as well as good stability in salt conditions. Moreover, compound 28 showed the rapid killing of bacteria via membrane-targeting action without developing bacterial resistance. More importantly, compound 28 displayed high antimicrobial potency against Gram-positive bacteria in a murine model of bacterial keratitis, and was found to be more efficient than vancomycin. Thus, compound 28 had great potential as a promising lead compound for the treatment of Gram-positive bacterial infection.

Bacteriophage Cocktails Protect Dairy Cows Against Mastitis Caused By Drug Resistant Escherichia coli Infection.

Mastitis caused by Escherichia coli (E. coli) remains a threat to dairy animals and impacts animal welfare and causes great economic loss. Furthermore, antibiotic resistance and the lagged development of novel antibacterial drugs greatly challenge the livestock industry. Phage therapy has regained attention. In this study, three lytic phages, termed vB_EcoM_SYGD1 (SYGD1), vB_EcoP_SYGE1 (SYGE1), and vB_EcoM_SYGMH1 (SYGMH1), were isolated from sewage of dairy farm. The three phages showed a broad host range and high bacteriolytic efficiency against E. coli from different sources. Genome sequence and transmission electron microscope analysis revealed that SYGD1 and SYGMH1 belong to the Myoviridae, and SYGE1 belong to the Autographiviridae of the order Caudovirales. All three phages remained stable under a wide range of temperatures or pH and were almost unaffected in chloroform. Specially, a mastitis infected cow model, which challenged by a drug resistant E. coli, was used to evaluate the efficacy of phages. The results showed that the cocktails consists of three phages significantly reduced the number of bacteria, somatic cells, and inflammatory factors, alleviated the symptoms of mastitis in cattle, and achieved the same effect as antibiotic treatment. Overall, our study demonstrated that phage cocktail may be a promising alternative therapy against mastitis caused by drug resistant E. coli.

Antibacterial Properties of Antimicrobial peptides (AMP) from Skin Extracts of Catla catla against Wound Pathogens

Wound infection is a major complication in healing of chronic wounds. Diabetic wounds are highly prone to wound infection and development of drug resistance among pathogens has created a necessity for development of new antimicrobials for treatment of such wound infections. Antimicrobial peptides (AMP) are found to be potent source showing promising antimicrobial action. Therefore the present study focuses on extraction of AMP from Catla catla skin using two different solvents and determining antibacterial activity against wound pathogens. Specimens from wound were collected using sterile swabs and streaked on nutrient agar. Then the colonies were subjected to gram staining and series of biochemical tests for identification. Live fish of Catla catla weighing 470gm was collected, homogenized in two different solvents (Ethanol and water). Antibacterial activity of the crude extracts of AMP was evaluated by using well diffusion method. Based on the colony morphology and biochemical tests the pathogens were found to be Staphylococcus aureus and Pseudomonas aeruginosa. The wound pathogens isolated were also found to be a biofilm producer. Antibacterial analysis showed ethanolic extract had higher inhibition than aqueous extracts. Therefore the extracted AMP can be used for development of novel antibacterial drugs for treatment of drug resistant bacterial infections.

Immunopeptidomics for next-generation bacterial vaccine development.

Antimicrobial resistance is an increasing global threat and alternative treatments substituting failing antibiotics are urgently needed. Vaccines are recognized as highly effective tools to mitigate antimicrobial resistance; however, the selection of bacterial antigens as vaccine candidates remains challenging. In recent years, advances in mass spectrometry-based proteomics have led to the development of so-called immunopeptidomics approaches that allow the untargeted discovery of bacterial epitopes that are presented on the surface of infected cells. Especially for intracellular bacterial pathogens, immunopeptidomics holds great promise to uncover antigens that can be encoded in viral vector- or nucleic acid-based vaccines. This review provides an overview of immunopeptidomics studies on intracellular bacterial pathogens and considers future directions and challenges in advancing towards next-generation vaccines.

Antibacterial Properties of Lichens Parmelia sulcata and Parmelia vagans Extracts

The objective of this study is to analyze the antibacterial properties of secondary metabolites extracted from lichens Parmelia sulcata and Parmelia vagans collected from the semi-deserts of Russia. Identifying these natural compounds that can inhibit proliferation of bacterial cells can enhance the development of antibacterial therapeutics. Acetone extracts of two lichens (P. sulcata and P. vagans) were prepared by using a Soxhlet apparatus. After drying the extracted solution, we added a known volume of acetone and rocked the flask for 10 minutes to dissolve the dry extract. We then transferred the dissolved solution to a 50-ml tube to get Fraction #1 and repeated this step to get multiple fractions. We first confirmed the antibacterial activity against Staphylococcus aureus by using a disk diffusion test from different fractions of the lichens. We assessed the antibacterial activity against S. aureus by using Resazurin Assay to measure the viability of the bacterial cells. Our experimental data from a disc diffusion test showed different sizes of inhibition rings around the pellets with lichen extracts from different fractions. The inhibition was also confirmed by Resazurin Assay which demonstrates that bacterial growth is inhibited differently by different fractions of lichen extracts. Our findings indicate that these lichen species produce at least two secondary metabolites that slow bacterial growth and proliferation. Future experiments will be focused on isolating and characterizing the antibacterial compounds from P. sulcata and P. vagans crude extracts. In summary, the lichen's secondary metabolites may be important for the development of novel antibacterial drugs.

Crystal structures of UDP-N-acetylmuramic acid L-alanine ligase (MurC) from Mycobacterium bovis with and without UDP-N-acetylglucosamine.

Peptidoglycan comprises repeating units of N-acetylmuramic acid, N-acetylglucosamine and short cross-linking peptides. After the conversion of UDP-N-acetylglucosamine (UNAG) to UDP-N-acetylmuramic acid (UNAM) by the MurA and MurB enzymes, an amino acid is added to UNAM by UDP-N-acetylmuramic acid L-alanine ligase (MurC). As peptidoglycan is an essential component of the bacterial cell wall, the enzymes involved in its biosynthesis represent promising targets for the development of novel antibacterial drugs. Here, the crystal structure of Mycobacterium bovis MurC (MbMurC) is reported, which exhibits a three-domain architecture for the binding of UNAM, ATP and an amino acid as substrates, with a nickel ion at the domain interface. The ATP-binding loop adopts a conformation that is not seen in other MurCs. In the UNAG-bound structure of MbMurC, the substrate mimic interacts with the UDP-binding domain of MbMurC, which does not invoke rearrangement of the three domains. Interestingly, the glycine-rich loop of the UDP-binding domain of MbMurC interacts through hydrogen bonds with the glucose moiety of the ligand, but not with the pyrophosphate moiety. These findings suggest that UNAG analogs might serve as potential candidates for neutralizing the catalytic activity of bacterial MurC.

DeepT3_4: A Hybrid Deep Neural Network Model for the Distinction Between Bacterial Type III and IV Secreted Effectors.

Gram-negative bacteria can deliver secreted proteins (also known as secreted effectors) directly into host cells through type III secretion system (T3SS), type IV secretion system (T4SS), and type VI secretion system (T6SS) and cause various diseases. These secreted effectors are heavily involved in the interactions between bacteria and host cells, so their identification is crucial for the discovery and development of novel anti-bacterial drugs. It is currently challenging to accurately distinguish type III secreted effectors (T3SEs) and type IV secreted effectors (T4SEs) because neither T3SEs nor T4SEs contain N-terminal signal peptides, and some of these effectors have similar evolutionary conserved profiles and sequence motifs. To address this challenge, we develop a deep learning (DL) approach called DeepT3_4 to correctly classify T3SEs and T4SEs. We generate amino-acid character dictionary and sequence-based features extracted from effector proteins and subsequently implement these features into a hybrid model that integrates recurrent neural networks (RNNs) and deep neural networks (DNNs). After training the model, the hybrid neural network classifies secreted effectors into two different classes with an accuracy, F-value, and recall of over 80.0%. Our approach stands for the first DL approach for the classification of T3SEs and T4SEs, providing a promising supplementary tool for further secretome studies.