Drug-resistant Bacterial Pathogens Research Articles

Evaluation of the antifungal and antibacterial action of a new unsaturated 1,2,4-triazole derivative (Nitrotriazon)

The growing clinical importance of drug-resistant fungal and bacterial pathogens has given additional urgency to microbiological research and the development of new antimicrobial compounds. 1,2,4-Triazoles act as important pharmacophores by interacting with high affinity biological receptors due to their dipole character, hydrogen binding capacity, stiffness and solubility. Following the evaluation of the antifungal activity, Nitrotriazon has MIC/MFC values: from 0.07 to 0.19 mM compared to the reference drugs: Ketoconazole, Bifonazole. Nitrotriazone also showed good antibacterial activity with MIC / MBC values from 0.0006 to 0.0013 mM, compared to the reference drugs Ampicillin and Chloramphenicol. The antimycobacterial activity of Nitrotriazone against M. tuberculosis, the reference strain (H37Rw) and the extended resistance of M. tuberculosis (M/XDR) were also studied.

Mechano-Induced Assembly of a Nanocomposite for "Press-N-Go" Coatings with Highly Efficient Surface Disinfection.

Using antimicrobial coatings to control the spread of pathogenic microbes is appreciated in public and healthcare settings, but the performance of most antimicrobial coatings could not fulfill the increasing requirements, particularly the ease of preparation, high durability, rapid response, and high killing efficiency. Herein, we develop a new type of mechano-induced assembly of nanocomposite coating by simple "Press-N-Go" procedures on various substrates such as glassware, gloves, and fabrics, in which the coating shows strong adhesion, high shear stability, and high stiffness, making it durable in daily use to withstand common mechanical deformation and scratches. The coating also shows remarkable disinfection effectiveness over 99.9% to clinically significant multiple drug-resistant bacterial pathogens upon only 6 s near-infrared irradiation, which can be further improved to over 99.9999% upon another 6 s treatment. We envision that the coating can provide convenience and values to control pathogen spread for easily contaminated substrates in high-risk areas.

Effect of Synergistic Action of Bovine Lactoferrin with Antibiotics on Drug Resistant Bacterial Pathogens.

Background and Objectives: The multidrug resistant (MDR) bacterial pathogenic infection is one of the chief worldwide public health threat to humanity. The development of novel antibiotics against MDR Gram negative bacteria has reduced over the last half century. Research is in progress regarding the treatment strategies that could be engaged in combination with antibiotics to extend the duration of these life-saving antibacterial agents. The current study was therefore planned to assess the synergistic effects of bovine lactoferrin (bLF) in combination with different antibiotics that are conventionally used. This synergism would provide a newer therapeutic choice against MDR pathogens. LF is present in mucosal secretions, vastly in milk. LF is considered an important constituent in host defense. In previous reports, LF has been co-administered as a combination antibiotic therapy. Materials and Methods: This study included synergistic (LF + appropriate antibiotic) exposure against 147 locally encountered bacterial pathogens, which were completely characterized strains. The anti-biofilm effects and the outcome of bLF on minimum inhibitory concentrations (MICs) of antibacterials on clinical MDR bacterial pathogens were determined by standard techniques. Results: In our study, synergism of bLF with antibacterial agents were reproducible and found to be significant. LF on its own had an important effect of inhibiting the biofilm production of some significant bacterial pathogens. Conclusion: The results of this study provides useful data on the antibacterial potential of the combination of LF with antibiotics against drug resistant pathogens.

In vitro antioxidant and antibiofilm activities of Microcystis sp. against multidrug-resistant human pathogens

The biofilm of drug-resistant pathogenic bacteria is a serious issue worldwide. The natural growth of blue-green algae Microcystis sp. has the potential to antagonize the bacterial pathogens in a novel approach. In the present study,Microcystis sp. was isolated and its mass production was optimized by Response Surface Methodology (RSM) and also, the invitro antioxidant and antibiofilm potentials against the drug-resistant pathogens were investigated. The experimental design was carried out on mass production of Microcystis sp. with pH, and mineral salts as independent variables.The maximum biomass yield (1.5 g/L) was achieved with NaNO3 (20mM) andK2HPO4 (10mM) at pH 7.5. The Microcystis sp. exhibited remarkable antioxidant (65%) and antibiofilm activities against Staphylococcus aureus and Pseudomonas aeruginosa. The Gas chromatography-mass spectrometry (GC-MS)of Microcystis sp. extract analysis revealed the presence ofsixteen bioactive potential compounds which proved the synergistic activity against the bacterial pathogens. Therefore, the potential bioactive compoundsinMicrocystis sp. could be an effective biocontrol agentagainst the challenging drug-resistant bacterial pathogens.

Electrostatic-Mediated Affinity Tuning of Lysostaphin Accelerates Bacterial Lysis Kinetics and Enhances In Vivo Efficacy

Drug-resistant bacterial pathogens are a serious threat to global health, and antibacterial lysins are at the forefront of innovative treatments for these life-threatening infections. While lysins' general mechanism of action is well understood, the design principles that might enable engineering of performance-enhanced variants are still being formulated. Here, we report a detailed analysis of molecular determinants underlying the in vivo efficacy of lysostaphin, a canonical anti-MRSA (methicillin-resistant Staphylococcus aureus) lysin. Systematic analysis of bacterial binding, growth inhibition, lysis kinetics, and in vivo therapeutic efficacy revealed that binding affinity, and not inherent catalytic firepower, is the dominant driver of lysostaphin efficacy. This insight enabled electrostatic affinity tuning of lysostaphin to produce a single point mutant that manifested dramatically enhanced processivity and lysis kinetics and trended toward improved in vivo efficacy. More generally, these studies provide important insights into the complex relationships between lysin electrostatics, bacterial targeting, cell lysis efficiency, and in vivo efficacy. The lessons learned may enable engineering of other high-performance antibacterial biocatalysts.

Study of bacterial pathogens, their antibiotic susceptibility patterns and multi drug resistance in bacterial pathogens isolated during acute episodes of respiratory infections in chronic obstructive pulmonary disease

Study of bacterial pathogens, their antibiotic susceptibility patterns and multi drug resistance in bacterial pathogens isolated during acute episodes of respiratory infections in chronic obstructive pulmonary disease

Dual stimuli-responsive metal-organic framework-based nanosystem for synergistic photothermal/pharmacological antibacterial therapy

The serious threat of drug-resistant bacterial pathogens has arisen through overuse of antibiotics. Photothermal therapy (PTT) has come to prominence as viable alternative strategy for antibacterial therapy. In this work, we report a NIR/pH dual stimuli-responsive antibacterial formulation based on zeolitic imidazolate frameworks-8 (ZIF-8) with strong antibacterial activity that combines photothermal heating with enhanced antibiotic delivery. ZIF-8 with polydopamine (PDA) surface modification was used to encapsulate the antibiotic vancomycin to construct a dual stimuli-responsive antimicrobial formulation (Van@ZIF-8@PDA). This treatment was tested against Gram-positive Mu50 (a vancomycin-intermediate S. aureus reference strain). Results showed that the PDA coating improved ZIF-8 stability and dispersion, while also conferring a high photothermal conversion efficiency. Hyperthermia activated by near-infrared (NIR) light irradiation, in conjunction with pH-dependent nanoparticle degradation to release vancomycin, enabled tight control of drug delivery that functioned synergistically in the elimination of both planktonic bacteria prior to biofilm formation and established biofilms. We found that this combined formulation compromises cell structure while also degrading bacterial DNA. Moreover, further investigation showed that the Van@ZIF-8@PDA nanoparticles exhibit good biocompatibility, with low toxicity toward host organs and tissues, while also reducing the antibiotic concentration needed for effective bacterial control. Finally, we treated Mu50 in a mouse model of skin abscess and found that Van@ZIF-8@PDA was effective and safe in vivo. Cumulatively, this study shows that this NIR/pH dual stimuli-responsive nanoparticle-based formulation offers a promising potential strategy for clinical application against bacterial infection that circumvents antibiotic resistance.

Antimicrobial activity of polyphenolic compounds from Spirulina against food-borne bacterial pathogens.

Food-borne drug-resistant bacteria have adverse impacts on both food manufacturers and consumers. Disillusionment with the efficacy of current preservatives and antibiotics for controlling food-borne pathogens, especially drug-resistant bacteria, has led to a search for safer alternatives from natural sources. Spirulina have been recognized as a food supplement, natural colorant, and enriched source of bioactive secondary metabolites. The main objectives of this study were to isolate polyphenolic compounds from Spirulina and analyze their antibacterial potential against drug-resistant food-borne bacterial pathogens. We found that fraction B of methanol extract contained a high quantity of polyphenols exhibiting broad spectrum antimicrobial effects against drug-resistant food-borne bacterial pathogens. Potential secondary metabolites, such as benzophenone, dihydro-methyl-phenylacridine, carbanilic acid, dinitrobenzoate, propanediamine, isoquinoline, piperidin, oxazolidin, and pyrrolidine, were identified by gas chromatography and mass spectrophotometry (GCMS). These metabolites are active against both gram-positive and gram-negative pathogens. Our work suggests that phenolic compounds from Spirulina provide a natural and sustainable source of food preservatives for future use.

The roles of tetraspanins in bacterial infections.

Tetraspanins, a wide family composed of 33 transmembrane proteins, are associated with different types of proteins through which they arbitrate important cellular processes such as fusion, adhesion, invasion, tissue differentiation and immunological responses. Tetraspanins share a comparable structural design, which consists of four hydrophobic transmembrane domains with cytoplasmic and extracellular loops. They cooperate with different proteins, including other tetraspanins, receptors or signalling proteins to compose functional complexes at the cell surface, designated tetraspanin-enriched microdomains (TEM). Increasing evidences establish that tetraspanins are exploited by numerous intracellular pathogens as a doorway for entering and replicating within human cells. Although previous surveys focused mainly on viruses and parasites, it is now becoming clear that bacteria interact with tetraspanins, using TEM as a "gateway" to infection. In this review, we examine the biological functions of tetraspanins that are relevant to bacterial infective procedures and consider the available data that reveal how different bacteria benefit from host cell tetraspanins in infection and in the pathogenesis of diseases. We will also emphasise the stimulating potentials of targeting tetraspanins for preventing bacterial infectious diseases, using specific neutralising antibodies or anti-adhesion peptide-based therapies. Such innovative therapeutic opportunities may deliver alternatives for fighting difficult-to-manage and drug-resistant bacterial pathogens.

Antimicrobial Effects of Selected, Cultivated Red Seaweeds and Their Components in Combination with Tetracycline, against Poultry Pathogen Salmonella Enteritidis

Poultry and its products are an economical source of high-quality protein for human consumption. In animal agriculture, antibiotics are used as therapeutic agents to treat disease in livestock, or as prophylactics to prevent disease and in so doing enhance production. However, the extensive use of antibiotics in livestock husbandry has come at the cost of increasingly drug-resistant bacterial pathogens. This highlights an urgent need to find effective alternatives to be used to treat infections, particularly in poultry and especially caused by drug-resistant Salmonella strains. In this study, we describe the combined effect of extracts of the red seaweeds Chondrus crispus (CC) and Sarcodiotheca gaudichaudii (SG) and compounds isolated from these in combinations with industry standard antibiotics (i.e., tetracycline and streptomycin) against Salmonella Enteritidis. Streptomycin exhibited the higher antimicrobial activity against S. Enteritidis, as compared to tetracycline with a MIC25 and MIC50 of 1.00 and 1.63 μg/mL, respectively. The addition of a water extract of CC at a concentration of 200 µg/mL in addition to tetracycline significantly enhanced the antibacterial activity (log CFU/mL 4.7 and 4.5 at MIC25 and MIC50, respectively). SG water extract, at 400 and 800 µg/mL (p = 0.05, n = 9), also in combination with tetracycline, showed complete inhibition of bacterial growth. Combinations of floridoside (a purified red seaweed component) and tetracycline (MIC25 and MIC50) in vitro revealed that only the lower concentration (i.e., 15 μg/mL) of floridoside potentiated the activity of tetracycline. Sub-lethal concentrations of tetracycline (MIC50 and MIC25), in combination with floridoside, exhibited antimicrobial activities that were comparable to full-strength tetracycline (23 μg/mL). Furthermore, the relative transcript levels of efflux-related genes of S. Enteritidis, namely marA, arcB and ramA, were significantly repressed by the combined treatment of floridoside and tetracycline, as compared to control MIC treatments (MIC25 and MIC50). Taken together, these findings demonstrated that the red seaweeds CC and SG and their selected, purified components can be used to increase the lifetime of existing, patented antibiotics and can also help to reduce costly (economic and environmental) therapeutic and prophylactic use of antibiotics in poultry. To our knowledge, this is the first report of antibiotic potentiation of existing industry standard antibiotics using red seaweeds and their selected extracts against S. Enteritidis.

Predicting the impact of infectious diarrhea pathophysiology on drug disposition

In the field of global health, physiologically based pharmacokinetic (PBPK) modeling has improved the confidence in malaria and tuberculosis (TB) drug development programs by assisting with first in human dose predictions, assessing potential drug‐drug interactions, and designing efficacious dosing regimens. PBPK models are composed of a compound file and a virtual physiology/population, and the impact of PBPK modeling in malaria and TB research is largely based on the substantial effort to improve both components. For example, compound files for commonly used therapeutics in malaria and TB have been developed through initiatives led by Medicines for Malaria Venture (MMV) and the Critical Path to TB Drug Regimens (CPTR). In addition, there has been considerable effort to develop virtual populations for these disease areas. Specifically, the focus of physiology and population model development for malaria has been on pediatric and pregnancy models. For TB, a virtual South African population has been generated which captures relevant genetic variants and TB‐related physiological changes that affect drug distribution and metabolism in this population.While the successful application of PBPK modeling in malaria and TB drug development has demonstrated the usefulness of this approach in global health, PBPK modeling has not been extensively applied to the area of drug development for enteric pathogens. In 2015, diarrhea caused > 688 million illnesses in children under 5 and was a leading cause of death among all ages (>1.3 million deaths). Despite advances in both the development of vaccines and small molecule therapeutics, new treatments are urgently needed for the treatment of multi drug resistant (MDR) bacterial pathogens and disease areas without an efficacious vaccine treatment. In symptomatic patients, we expect diarrhea to be associated with a reduction in gastrointestinal (GI) transit times, altered luminal pH, increased luminal volume, and disrupted epithelial integrity. However, the extent of these changes is often not clear. In addition, there has been insufficient clinical data to support how the pathophysiology may impact therapeutic disposition. To initially address this lack of clinical data, an aim of a recently completed clinical study in Malawi (NCT03341767) was to characterize the PK of clofazimine in HIV+ adult populations divided into two groups (diarrhea vs. non‐diarrheal). These clinical data were used to inform PBPK modeling efforts to predict the impact of diarrhea on clofazimine disposition and to relate the clinical study PK and efficacy outcomes to those obtained with pre‐clinical animal models of disease. While this work primarily focused on participants with cryptosporidiosis, our results provide a foundation for future efforts to use PBPK models in the development of therapeutics intended for various study populations with diarrhea.Support or Funding InformationBill and Melinda Gates Foundation Award OPP1172544

Old Drugs, New Synergy: Identifying Novel Synergistic Antibiotic Combinations to Combat MDR Bacterial Pathogens

Dwindling therapeutic options have necessitated a push to identify more efficacious antibiotic regimens for treating drug resistant bacterial pathogens. Acinetobacter baumannii, a member of the ESKAPE pathogens, is of special concern due to its high mortality rate and propensity for broad spectrum antibiotic resistance. We recently described hitherto unexplored azithromycin activity against MDR A. baumannii. To expand on those findings, we sought to compare the activity of azithromycin to minocycline, the current standard of care for MDR A. baumannii infections. We identified disparate media dependent susceptibility profiles for each drug that suggested the potential for an efficacious combination therapy. In combination, the two drugs were found to interact synergistically in standardized laboratory media, human whole blood, and in a murine MDR A. baumannii lung pneumoniae model. A synergistic interaction was a surprising finding as both azithromycin and minocycline are translation inhibitors and would be expected to only have an additive interaction. Synergy was found to be mediated through enhancing the kinetics of translation inhibition, yielding a combination that was faster, more potent, and longer lasting then either drug in monotherapy. The combination was found to have broad applicability as it was also synergistic against MDR strains of Pseudomonas aeruginosa, Klebsiella pneumoniae, and methicillin‐resistant Staphylococcus aureus. Although initially found with azithromycin and minocycline, synergy was also uncovered with other macrolides and tetracyclines and may therefore be a class wide phenomenon. Interestingly, this synergistic interaction does not appear to necessitate bacterial sensitivity to the drugs in monotherapy. Considering both azithromycin and minocycline are already FDA approved, their inclusion in a combination therapy has the potential to have an immediate impact in the clinic for treating high‐priority MDR bacterial pathogen infections in humans.Support or Funding InformationNIAID U01AI124316

Innovation in Antimicrobial Resistance: The CARB-X Perspective.

The spread of drug-resistant bacterial pathogens has been recognized as one of the largest global threats to mankind. In order to continue to benefit from the advancement of modern medicine, new treatments, prevention, and diagnostic products are needed to satisfactorily treat or prevent infections. Combating Antibiotic-Resistant Bacteria Biopharmaceutical Accelerator (CARB-X) is a global nonprofit public-private partnership dedicated to accelerating antibacterial-related research to tackle the rising threat of drug-resistant bacteria. The CARB-X portfolio is the world's largest early development pipeline of new antibiotics, vaccines, rapid diagnostics, and other products to prevent, diagnose, and treat life-threatening bacterial infections. Since inception, three application rounds of funding have been completed with a significant worldwide response.

Development of a Bacteriophage Cocktail to Constrain the Emergence of Phage-Resistant Pseudomonas aeruginosa.

With the emergence of multidrug-resistant and extensively drug-resistant bacterial pathogens, phage therapy and other alternative or additional therapeutic modalities are receiving resurgent attention. One of the major obstacles in developing effective phage therapies is the evolution of phage resistance in the bacterial host. When Pseudomonas aeruginosa was infected with a phage that uses O-antigen as receptor, phage resistances typically achieved through changing or loss of O-antigen structure. In this study, we showed that dsRNA phage phiYY uses core lipopolysaccharide as receptor and therefore efficiently kills the O-antigen deletion mutants. Furthermore, by phage training, we obtained PaoP5-m1, a derivative of dsDNA phage PaoP5, which is able to infect mutants with truncated O-antigen. We then generated a cocktail by mixing phiYY and PaoP5-m1 with additional three wide host range P. aeruginosa phages. The phage cocktail was effective against a diverse selection of clinical isolates of P. aeruginosa, and in the short-term constrained the appearance of the phage-resistant mutants that had beleaguered the effectiveness of single phage. Resistance to the 5-phage cocktail emerges after several days, and requires mutations in both wzy and migA Thus, this study provides an alternative strategy for designing phage cocktail and phage therapy.

Mechanism-of-Action Classification of Antibiotics by Global Transcriptome Profiling.

Antimicrobial resistance (AMR) is an ever-growing public health problem worldwide. The low rate of antibiotic discovery coupled with the rapid spread of drug-resistant bacterial pathogens is causing a global health crisis. To facilitate the drug discovery processes, we present a large-scale study of reference antibiotic challenge bacterial transcriptome profiles, which included 37 antibiotics across 6 mechanisms of actions (MOAs) and provide an economical approach to aid in antimicrobial dereplication in the discovery process. We demonstrate that classical MOAs can be sorted based upon the magnitude of gene expression profiles despite some overlap in the secondary effects of antibiotic exposures across MOAs. Additionally, using gene subsets, we were able to subdivide broad MOA classes into subMOAs. Furthermore, we provide a biomarker gene set that can be used to classify most antimicrobial challenges according to their canonical MOA. We also demonstrate the ability of this rapid MOA diagnostic tool to predict and classify the expression profiles of pure compounds and crude extracts to their expression profile-associated MOA class.

Pastoral community practices, microbial quality and associated health risks of raw milk in the milk value chain of Nakasongola District, Uganda

The informal milk value chain in Uganda controls most of the raw marketed milk, albeit of poor quality. A study was conducted to assess the milk handling practices, the bacterial quality and the associated health concerns of raw milk along the informal milk value in Nakasongola District. The cross-sectional study used both qualitative and quantitative methods to obtain information from key players. The qualitative data was categorized and presented into emerging themes. Bacterial contamination of 200 milk samples was determined by morphological and biochemical tests. The antibiotic susceptibility test was done by the disc diffusion method.Hand milking was carried out mainly by men (92.5%), and most never or rarely practised hand washing (67.5%) during milking. Milk was mainly delivered to mobile milk-collecting centres located under tree shades (50%). The quality of raw milk was affected by poor hygienic, handling and transportation practices. Most participants (75%) were aware of the dangers of drinking raw milk and the requirements to transport milk in metallic cans. Viable bacterial counts above the recommended limit were recorded in 76 samples (38%) that included Staphylococcus aureus (46%), coagulase-negative Staphylococcus (29%), Escherichia coli (12%), Streptococcus agalactiae (8%) and Salmonella spp. (5%). The overall antibiotic resistance was 45% (34/76), which included resistant isolates of S. aureus (17/34), coagulase-negative Staphylococcus (11/22), S. agalactiae (3/6), Salmonella spp. (2/4) and E. coli (2/9).The raw milk contaminated with antibiotic drug-resistant bacterial pathogens is of public health concern. Thus, measures to improve the quality of milk need to be designed for the pastoral community in Nakasongola district.

Synergistic antimicrobial action of phyco-synthesized silver nanoparticles and nano-fungal chitosan composites against drug resistant bacterial pathogens

Here we report the phycosynthesis of silver nanoparticles (Ag-NPs), using Codium capitatum extract, and the synthesis of fungal chitosan nanoparticles (FC-NPs), using extracted chitosan from Aspergillus niger mycelia. Then nanoconjugates from FC/Ag-NPs were produced and evaluated. The synthesized NPs had mean particles’ size diameters of 37.2, 68.4 and 79.6 nm for Ag-NPs, FC-NPs and FC/Ag-NPs, respectively. The FTIR (Fourier-transform infrared spectroscopy) analysis of synthesized NPs indicated their cross-linkage and interaction. The antibacterial activity of each type of NPs was assayed against drug resistant pathogens of Salmonella Typhimurium and Staphylococcus aureus. All NPs had powerful inhibitory effect and FC/Ag nanoconjugates had stronger activity than the other types. Scanning micrographs of FC/Ag-NPs treated S. Typhimurium elucidated vigorous alterations in cell surfaces and lethal damage to bacterial structure after 8 h of treatment. The nanoconjugates form FC-NPs and Ag-NPs had minute particle size with increasing bioactivity as antimicrobial agents to control drug resistant bacterial pathogens, which recommends their further exploration for topical applications in biomedical sectors.

Imipenem/cilastatin sodium/relebactam fixed combination to treat urinary infections and complicated intra-abdominal bacterial infections.

Imipenem/cilastatin sodium/relebactam is a combination of imipenem/cilastatin, a U.S. Food and Drug Administration (FDA)-approved antibiotic, and β-lactamase inhibitor relebactam which has been developed for the treatment of complicated urinary tract infection (cUTI) and complicated intra-abdominal infection (cIAI) due to drug-resistant bacterial pathogens. The combination (Recarbrio) has been designated as a qualified infectious disease product (QIDP) and obtained FDA approval in 2019 for the treatment of cUTI and cIAI caused by susceptible Gram-negative microorganisms in adult patients with limited or no alternative treatment options. The product was also approved by the European Medicines Agency (EMA) in 2020 for the treatment of infections due to aerobic Gram-negative organisms in adults with limited treatment options.

Phytochemical Investigation and Antimicrobial Activity of Root Extracts of <i>Withania Somnifera</i> (L.) Dunal. against Drug- Resistant Bacterial Pathogens

With the emergence of multidrug resistance among clinical bacterial pathogens there is a growing need for the development of herbal alternatives for the effective therapeutic management of bacterial infections. Plants and plants products have long been used in traditional folk medicine worldwide. Hence this study was designed to test the antimicrobial efficacy of Withania somnifera L. Dunal. (root) against drug resistant bacterial pathogens. Extracts of W. somnifera (L.) Dunal (root) was prepared by cold percolation method. Antimicrobial activity of different solvent extracts of W. somnifera (L.) Dunal. (root) was studied against drug-resistant clinical isolates of Gram-positive cocci (vancomycin resistant Enterococcus faecalis and Methicillin resistant Staphylococcus aureus) and Gram-negative bacilli (ESBL Escherichia coli and Pseudomonas aeruginosa). The standard strains of ATCC 25922 E. coli, ATCC 25923 S. aureus, ATCC 27853 P. aeruginosa, and ATCC 29212 E. faecalis were used as controls. Antibacterial activities of extracts were evaluated using agar well diffusion method. Minimum inhibitory concentration was determined by microbroth dilution method. All the extracts (methanol, ethanol, acetone and chloroform) were inhibitory to gram positive bacteria tested. Nevertheless, inhibitory effect waspronounced in the methanol and ethanol extract, against gram negative bacteria (Both ATCC and clinical strains).Further based on the phytochemical and GC-MS analysis, it could be speculated that the antibacterial activity exhibited by the methanol extract of W. somnifera root could be attributed to the presence of phenolic compounds, flavonoids, steroids and terpenes.

Screening of biomarkers of drug resistance or virulence in ESCAPE pathogens by MALDI-TOF mass spectrometry

Rapid identification and characterisation of drug-resistant bacterial pathogens have an important role in diagnostic and antimicrobial stewardship. Response time in the diagnosis of not only the etiological agent but also in antimicrobial susceptibility results is of utmost importance in patient treatment. In this study, matrix-assisted laser desorption ionisation–time of flight (MALDI-TOF) mass spectrometry (MS) was used to screen for biomarkers of ESCAPE (vancomycin-resistant Enterococcus faecium, methicillin-resistant Staphylococcus aureus, hypervirulent NAP1/ribotype 027 Clostridioides [Clostridium] difficile, multidrug resistant Acinetobacter baumannii, multidrug resistant Pseudomonas aeruginosa, and carbapenem-resistant Enterobacteriaceae) pathogens to predict antimicrobial resistance or hypervirulence. Several biomarkers of drug-resistant genotypes in S. aureus, A. baumannii, P. aeruginosa, and K. pneumoniae, as well as hypervirulence in C. difficile, were detected. The fastest possible susceptibility testing with MALDI-TOF MS is simultaneous detection of a characteristic drug-resistant peak and species identification in the same spectra generated in routine processing. According to our approach, resistance or virulence biomarker peaks can be identified while performing routine microbiology analysis, and no additional assays nor prolonged incubation time is needed. Outstanding biomarker peaks detected in our study should be further analysed by additional methods to identify the specific proteins involved.