Resistance-modifying Agents Research Articles

Deciphering the impact of Acinetobacter baumannii on human health, and exploration of natural compounds as efflux pump inhibitors to treat multidrug resistance.

Acinetobacter baumannii is an ESKAPE pathogen and threatens human health by generating infections with high fatality rates. A. baumannii leads to a spectrum of infections such as skin and wound infections, endocarditis, meningitis pneumonia, septicaemia and urinary tract infections. Recently, strains of A. baumannii have emerged as multidrug-resistant (MDR), meaning they are resistant to at least three different classes of antibiotics. MDR development is primarily intensified by widespread antibiotic misuse and inadequate stewardship. The World Health Organization (WHO) declared A. baumannii a precarious MDR species. A. baumannii maintains the MDR phenotype via a diverse array of antimicrobial metabolite-hydrolysing enzymes, efflux of antibiotics, impermeability and antibiotic target modification, thereby complicating treatment. Hence, a deeper understanding of the resistance mechanisms employed by MDR A. baumannii can give possible approaches to treat antimicrobial resistance. Resistance-nodulation-cell division (RND) efflux pumps have been identified as the key contributors to MDR determinants, owing to their capacity to force a broad spectrum of chemical substances out of the bacterial cell. Though synthetic inhibitors have been reported previously, their efficacy and safety are of debate. As resistance-modifying agents, phytochemicals are ideal choices. These natural compounds could eliminate the bacteria or interact with pathogenicity events and reduce the bacteria's ability to evolve resistance. This review aims to highlight the mechanism behind the multidrug resistance in A. baumannii and elucidate the utility of natural compounds as efflux pump inhibitors to deal with the infections caused by A. baumannii.

Polyphenols and their nanoformulations as potential antibiofilm agents against multidrug-resistant pathogens.

The emergence of multidrug-resistant (MDR) pathogens is a major problem in the therapeutic management of infectious diseases. Among the bacterial resistance mechanisms is the development of an enveloped protein and polysaccharide-hydrated matrix called a biofilm. Polyphenolics have demonstrated beneficial antibacterial effects. Phenolic compounds mediate their antibiofilm effects via disruption of the bacterial membrane, deprivation of substrate, protein binding, binding to adhesion complex, viral fusion blockage and interactions with eukaryotic DNA. However, these compounds have limitations of chemical instability, low bioavailability, poor water solubility and short half-lives. Nanoformulations offer a promising solution to overcome these challenges by enhancing their antibacterial potential. This review summarizes the antibiofilm role of polyphenolics, their underlying mechanisms and their potential role as resistance-modifying agents.

Antibiotic adjuvants: synergistic tool to combat multi-drug resistant pathogens.

The rise of multi-drug resistant (MDR) pathogens poses a significant challenge to the field of infectious disease treatment. To overcome this problem, novel strategies are being explored to enhance the effectiveness of antibiotics. Antibiotic adjuvants have emerged as a promising approach to combat MDR pathogens by acting synergistically with antibiotics. This review focuses on the role of antibiotic adjuvants as a synergistic tool in the fight against MDR pathogens. Adjuvants refer to compounds or agents that enhance the activity of antibiotics, either by potentiating their effects or by targeting the mechanisms of antibiotic resistance. The utilization of antibiotic adjuvants offers several advantages. Firstly, they can restore the effectiveness of existing antibiotics against resistant strains. Adjuvants can inhibit the mechanisms that confer resistance, making the pathogens susceptible to the action of antibiotics. Secondly, adjuvants can enhance the activity of antibiotics by improving their penetration into bacterial cells, increasing their stability, or inhibiting efflux pumps that expel antibiotics from bacterial cells. Various types of antibiotic adjuvants have been investigated, including efflux pump inhibitors, resistance-modifying agents, and compounds that disrupt bacterial biofilms. These adjuvants can act synergistically with antibiotics, resulting in increased antibacterial activity and overcoming resistance mechanisms. In conclusion, antibiotic adjuvants have the potential to revolutionize the treatment of MDR pathogens. By enhancing the efficacy of antibiotics, adjuvants offer a promising strategy to combat the growing threat of antibiotic resistance. Further research and development in this field are crucial to harness the full potential of antibiotic adjuvants and bring them closer to clinical application.

Prenylated isoflavonoids from Fabaceae against the NorA efflux pump in Staphylococcus aureus

Overexpression of NorA efflux pumps plays a pivotal role in the multidrug-resistance mechanism in S. aureus. Here, we investigated the activities of prenylated isoflavonoids, present in the legume plant family (Fabaceae), as natural efflux pump inhibitors (EPIs) in fluoroquinolone-resistant S. aureus. We found that four prenylated isoflavonoids, namely neobavaisoflavone, glabrene, glyceollin I, and glyceollin III, showed efflux pump inhibition in the norA overexpressing S. aureus. At sub-inhibitory concentrations, neobavaisoflavone (6.25 µg/mL, 19 µM) and glabrene (12.5 µg/mL, 39 µM), showed up to 6 times more Eth accumulation in norA overexpressing S. aureus than in the control. In addition, these two compounds boosted the MIC of fluoroquinolones up to eightfold. No fluoroquinolone potentiation was observed with these isoflavonoids in the norA knockout strain, indicating NorA as the main target of these potential EPIs. In comparison to the reported NorA EPI reserpine, neobavaisoflavone showed similar potentiation of fluoroquinolone activity at 10 µM, higher Eth accumulation, and less cytotoxicity. Neobavaisoflavone and glabrene did not exhibit membrane permeabilization effects or cytotoxicity on Caco-2 cells. In conclusion, our findings suggest that the prenylated isoflavonoids neobavaisoflavone and glabrene are promising phytochemicals that could be developed as antimicrobials and resistance-modifying agents to treat fluoroquinolone-resistant S. aureus strains.

A standardized nomenclature for resistance-modifying agents in the Comprehensive Antibiotic Resistance Database.

While increasing rates of antimicrobial resistance undermine our current arsenal of antibiotics, resistance-modifying agents (RMAs) hold promise to extend the lifetime of these important molecules. We here provide a standardized nomenclature for RMAs within the Comprehensive Antibiotic Resistance Database in aid of RMA discovery, data curation, and genome mining.

Hydrocinnamic Acid and Perillyl Alcohol Potentiate the Action of Antibiotics against Escherichia coli.

The treatment of bacterial infections has been troubled by the increased resistance to antibiotics, instigating the search for new antimicrobial therapies. Phytochemicals have demonstrated broad-spectrum and effective antibacterial effects as well as antibiotic resistance-modifying activity. In this study, perillyl alcohol and hydrocinnamic acid were characterized for their antimicrobial action against Escherichia coli. Furthermore, dual and triple combinations of these molecules with the antibiotics chloramphenicol and amoxicillin were investigated for the first time. Perillyl alcohol had a minimum inhibitory concentration (MIC) of 256 µg/mL and a minimum bactericidal concentration (MBC) of 512 µg/mL. Hydrocinnamic acid had a MIC of 2048 µg/mL and an MBC > 2048 µg/mL. Checkerboard and time-kill assays demonstrated synergism or additive effects for the dual combinations chloramphenicol/perillyl alcohol, chloramphenicol/hydrocinnamic acid, and amoxicillin/hydrocinnamic acid at low concentrations of both molecules. Combenefit analysis showed synergism for various concentrations of amoxicillin with each phytochemical. Combinations of chloramphenicol with perillyl alcohol and hydrocinnamic acid revealed synergism mainly at low concentrations of antibiotics (up to 2 μg/mL of chloramphenicol with perillyl alcohol; 0.5 μg/mL of chloramphenicol with hydrocinnamic acid). The results highlight the potential of combinatorial therapies for microbial growth control, where phytochemicals can play an important role as potentiators or resistance-modifying agents.

CARD 2023: expanded curation, support for machine learning,and resistome prediction at the Comprehensive Antibiotic Resistance Database.

The Comprehensive Antibiotic Resistance Database (CARD; card.mcmaster.ca) combines the Antibiotic Resistance Ontology (ARO) with curated AMR gene (ARG) sequences and resistance-conferring mutations to provide an informatics framework for annotation and interpretation of resistomes. As of version 3.2.4, CARD encompasses 6627 ontology terms, 5010 reference sequences, 1933 mutations, 3004 publications, and 5057 AMR detection models that can be used by the accompanying Resistance Gene Identifier (RGI) software to annotate genomic or metagenomic sequences. Focused curation enhancements since 2020 include expanded β-lactamase curation, incorporation of likelihood-based AMR mutations for Mycobacterium tuberculosis, addition of disinfectants and antiseptics plus their associated ARGs, and systematic curation of resistance-modifying agents. This expanded curation includes 180 new AMR gene families, 15 new drug classes, 1 new resistance mechanism, and two new ontological relationships: evolutionary_variant_of and is_small_molecule_inhibitor. In silico prediction of resistomes and prevalence statistics of ARGs has been expanded to 377 pathogens, 21,079 chromosomes, 2,662 genomic islands, 41,828 plasmidsand 155,606 whole-genome shotgun assemblies, resulting in collation of 322,710 unique ARG allele sequences. New features include the CARD:Live collection of community submitted isolate resistome data and the introduction of standardized 15 character CARD Short Names for ARGs to support machine learning efforts.

Structure–activity relationship studies of [1,2,5]oxadiazolo[3,4-b]pyrazine-containing polymyxin-selective resistance-modifying agents

Multidrug-resistant (MDR) Gram-negative bacteria are an urgent and rapidly spreading threat to human health with limited treatment options. Previously, we discovered a novel [1,2,5]oxadiazolo[3,4-b]pyrazine-containing compound (1) that selectively re-sensitized a variety of MDR Gram-negative bacteria to colistin, one of the last-resort antibiotic. Herein, we report the structure–activity relationship studies of compound 1 that led to the discovery of several more potent and/or less toxic resistance-modifying agents (RMAs). Further evaluation of these RMAs showed that they were effective in a wide range of MDR bacteria. These results demonstrated these compounds as a novel class of RMAs and may be further developed as therapeutic agents.

Synergistic Role of Plant Extracts and Essential Oils against Multidrug Resistance and Gram-Negative Bacterial Strains Producing Extended-Spectrum β-Lactamases.

Plants, being the significant and natural source of medication for humankind against several ailments with characteristic substances hidden on them, have been recognized for many centuries. Accessibility of various methodologies for the revelation of therapeutically characteristic items has opened new avenues to redefine plants as the best reservoirs of new structural types. The role of plant metabolites to hinder the development and movement of pathogenic microbes is cherished. Production of extended-spectrum β-lactamases is an amazing tolerance mechanism that hinders the antibacterial treatment of infections caused by Gram-negative bacteria and is a serious problem for the current antimicrobial compounds. The exploration of the invention from sources of plant metabolites gives sustenance against the concern of the development of resistant pathogens. Essential oils are volatile, natural, complex compounds described by a solid odor and are framed by aromatic plants as secondary metabolites. The bioactive properties of essential oils are commonly controlled by the characteristic compounds present in them. They have been commonly utilized for bactericidal, virucidal, fungicidal, antiparasitic, insecticidal, medicinal, and antioxidant applications. Alkaloids are plant secondary metabolites that have appeared to have strong pharmacological properties. The impact of alkaloids from Callistemon citrinus and Vernonia adoensis leaves on bacterial development and efflux pump activity was assessed on Pseudomonas aeruginosa. Plant-derived chemicals may have direct antibacterial activity and/or indirect antibacterial activity as antibiotic resistance modifying agents, increasing the efficiency of antibiotics when used in combination. The thorough screening of plant-derived bioactive chemicals as resistance-modifying agents, including those that can act synergistically with antibiotics, is a viable method to overcome bacterial resistance. The synergistic assessment studies with the plant extract/essential oil and the antibiotic compounds is essential with a target for achieving a redesigned model with sustainable effects which are appreciably noticeable in specific sites of the plants compared to the entirety of their individual parts.

Flavonoids as Inhibitors of Bacterial Efflux Pumps.

Flavonoids are widely occurring secondary plant constituents, and are abundant in vegetable and fruit diets as well as herbal medicines. Therapeutic treatment options for bacterial infections are limited due to the spread of antimicrobial resistances. Hence, in a number of studies during the last few years, different classes of plant secondary metabolites as resistance-modifying agents have been carried out. In this review, we present the role of flavonoids as inhibitors of bacterial efflux pumps. Active compounds could be identified in the subclasses of chalcones, flavan-3-ols, flavanones, flavones, flavonols, flavonolignans and isoflavones; by far the majority of compounds were aglycones, although some glycosides like kaempferol glycosides with p-coumaroyl acylation showed remarkable results. Staphylococcus aureus NorA pump was the focus of many studies, followed by mycobacteria, whereas Gram-negative bacteria are still under-investigated.

EVALUATION OF CITRIC ACID AND CLOVE OIL AS SAFE ANTI-VIRULENCE AND RESISTANCE-MODIFYING AGENTS AGAINST PROBLEMATIC KLEBSIELLA CLINICAL ISOLATES IN EGYPT

Klebsiella species are regarded as terrifying pathogens that threaten different healthcare systems. These pathogens possess a variety of virulence factors which markedly account for their pathogenicity. Three virulence factors (capsule formation, hypermucoviscosity and biofilm formation) were screened, phenotypically and genotypically, among selected Egyptian Klebsiella clinical isolates. Two safe edible agents, clove oil and citric acid, were tested for their effect on bacterial capsules using the Anthony capsular staining method and the transmission electron microscopy. The antibiotic-resistance modifying activity of both agents was studied against selected troublesome isolates showing resistance to amikacin, meropenem and cefotaxime. Molecular docking of citric acid and eugenol with NDM-1, as well as the docking of citric acid with AAC(6′)-Ib and CTX-M-15 were carried out to explain the resistance modifying activity of the tested agents. Capsule formation was the most prominent virulence factor that was detected in all isolates. Both citric acid and clove oil could effectively reduce the size of the formed bacterial capsules. Citric acid showed promising synergistic effects when combined with the studied antibiotics against all the tested isolates. However, clove oil showed a synergistic effect only against 50% of the tested isolates when combined with meropenem. Relying on their binding efficiency mode obtained from molecular docking studies, the tested agents were recognized as promising inhibitors of the studied enzymes. In conclusion, both citric acid and clove oil are auspicious anti-virulence and resistance-modifying agents that can be combined with conventional antibiotics to combat virulent and multidrug resistant Klebsiella isolates.

Critical discovery and synthesis of novel antibacterial and resistance-modifying agents inspired by plant phytochemical defense mechanisms

Antimicrobial resistance is at increasing risk worldwide since it is threatening the ability to control common infectious diseases, resulting in prolonged illness, disability, and death. Herein, we inspired by the effective plant phytochemical mechanisms evolved to overcome microbial pathogenesis and evolved resistance. Cuminaldehyde is previously reported as the main antibacterial component in Calligonum comosum essential oil. The toxicity of cuminaldehyde limits its medical application for human use. On the other hand, compared to cuminaldehyde, the plant total extract showed similar antibacterial activities, while maintained lower toxicity, although it contains 22 times less cuminaldehyde. Thus, we assumed that other components in the plant extracts specifically affect bacteria but not mammalian cells. Bioassay-guided fractionations combined with comparative metabolomics analysis of different plant extracts were employed. The results revealed the presence of bacterial species-specific phytochemicals. Cinnamyl linoleate and linoleic acid enhanced the antibacterial activities of cuminaldehyde and ampicillin against S. aureus including MRSA, while decanal and cinnamyl linoleate enhanced the activities against E. coli. Computational modeling and enzyme inhibition assays indicated that cinnamyl linoleate selectively bind to bacterial ribosomal RNA methyltransferase, an important enzyme involved in the virulence and resistance of multidrug resistant bacteria. The results obtained can be employed for the future preparation of pharmaceutical formula containing cinnamyl linoleate in order to overcome evolved multidrug resistance behaviors by microbes.

Antifungal resistance-modifying multiplexing action of Momordica charantia protein and phosphorylated derivatives on the basis of growth-dependent gene coregulation in Candida albicans.

A new platform to modify resistance is fungal growth-dependent gene coregulation. MAP30 and phosphorylated derivatives are candidate resistance-modifying agents. Low-dose stepwise treatment absolutely modifies azole resistance in model fungus.

Antimicrobial Efficiency of Moringa oleifera Leave Extracts against Some Multidrug Resistant Pathogen

THE MEDICINAL plants have vital sources of natural antimicrobial substances for human health avoiding side effects precipitated by using synthetic chemical. The decreasing effectiveness of traditional antibiotics towards multi drug resistant microorganism is a global public health. It was found that the mixtures between different plant extract and commercial antibiotics increased the effectiveness of antibiotics multi drug resistant bacteria. The combinations between different plant extract and commercial antibiotics increased the effectiveness of antibiotics against multi drug resistant strains. Extracts of Moringa oleifera leaves showed remarkable antimicrobial activity against the growth of the gram positive and negative bacteria drug resistant clinical strains (Escherichia coli ATCC 10536, Pseudomonas aeruginosa ATCC 9027, Staphylococcus aureus ATCC 29737 and Staph epidermidis ATCC 12228, Bacillus pumilis ATCC 14884, Micrococcus luteus ATCC 10240, Bordetella sp, Bacillus subtilis and Klebsiella sp . Antimicrobial activity of methanol extract of Moringa oleifera leaves (MML) using well diffusion method was higher (30mm inhibition zone) than ethanolic and aqueous extract. The minimum inhibition concentration (MIC) of the selected pathogenic bacteria ranged between 23 -800μg/ml. The interactions between different extract of Moringa oleifera leaves and different antibiotic groups of Ampicillin (Am10), Amoxicillin (AMC30), Amikacin (AK30), Gentamycin (CN10), Tetracycline (TE30), Erythromycin (E15), Bacitracin (B10) were conducted using agar diffusion method against the selected pathogenic bacteria. Methanol extract of Moringa oleifera leaves could be used as a source for resistance-modifying agents against infectious multi-drug resistant bacteria.

A Clerodane Diterpene from Callicarpa americana Resensitizes Methicillin-Resistant Staphylococcus aureus to β-Lactam Antibiotics.

The rise of antibiotic resistance presents a significant healthcare challenge and precludes the use of many otherwise valuable antibiotics. One potential solution to this problem is the use of antibiotics in combination with resistance-modifying agents, compounds that act synergistically with existing antibiotics to resensitize previously resistant bacteria. In this study, 12(S),16ξ-dihydroxycleroda-3,13-dien-15,16-olide, a clerodane diterpene isolated from the medicinal plant Callicarpa americana, was found to synergize with oxacillin against methicillin-resistant Staphylococcus aureus. This synergy was confirmed by checkerboard (fractional inhibitory concentration index (FICI) = 0.125) and time-kill assays, with a subinhibitory dose of 12(S),16ξ-dihydroxycleroda-3,13-dien-15,16-olide causing the effective concentration of oxacillin to fall below the susceptibility breakpoint for S.aureus, a >32-fold decrease in both cases.

Jatrorrhizine suppresses the antimicrobial resistance of methicillin-resistant Staphylococcus aureus.

Bacterial resistance to antimicrobial agents, including multidrug resistance, is an increasing problem in the treatment of infectious diseases. The development of resistance-modifying agents represents a potential strategy to alleviate the spread of bacterial resistance to antibiotics. A checkerboard microdilution assay was used to determine the synergy of jatrorrhizine and the antibiotic, norfloxacin (NFX). A bacterial ethidium bromide efflux assay, reverse transcription semi-quantitative polymerase chain reaction analysis and molecular docking study were performed. The three-dimensional structure of NorA multidrug efflux pump (NorA) was generated using a multiple threading approach. A murine thigh infection model was used to evaluate the in vivo synergistic effect. As a natural product, jatrorrhizine exhibited little antibacterial activity against methicillin-resistant Staphylococcus aureus (MRSA) SA1199B with a minimum inhibitory concentration (MIC) of 64 mg/l. According to the investigations of the mechanism, jatrorrhizine significantly inhibited bacterial drug efflux and the expression of NorA in the mRNA level as it can bind to NorA by hydrogen-bonds, hydrophobic and electrostatic interactions. The in vivo synergistical bactericidal activity of jatrorrhizine and NFX against MRSA was confirmed in a murine thigh infection model. As a novel resistance-modifying agent, jatrorrhizine exhibited in vitro and in vivo synergistic activities against MRSA, and inhibited bacterial drug efflux. The effects were mediated by the suppression of NorA mRNA expression and/or interactions with NorA efflux pump. These data support the hypothesis that jatrorrhizine is a potential agent for therapeutic use in infections caused by MRSA.

Tryptoline-based benzothiazoles re-sensitize MRSA to β-lactam antibiotics

Resistance-modifying agents (RMAs) offer a promising solution to combat bacterial antibiotic resistance. Here we report the discovery and structure–activity relationships of a new class of RMAs with a novel tryptoline-based benzothiazole scaffold. Our most potent compound in this series (4ad) re-sensitizes multiple MRSA strains to cephalosporins at low concentrations (2 μg/mL) and has low mammalian cytotoxicity with a half growth inhibitory concentration (GI50) > 100 μg/mL in human cervical carcinoma (HeLa) cells. In addition, the same core scaffold with different substitutions also gives good antibacterial activity against MRSA.

Synergistic Antimicrobial Effects of Cefabronchin®.

Antibiotic resistance of Streptococcus pneumoniae has risen to worrying levels in the past few decades worldwide, and subsequently, effective treatment of respiratory tract infections has become even more challenging. While the need to develop new strategies to combat bacterial infections is urgent, novel antibiotic compounds are no longer a priority of the pharmaceutical industry. However, resistance-modifying agents can alleviate the spread of antibiotic resistance and render existing antibiotics effective again. In the present study, we aimed to determine the combinatory antimicrobial effects of the commercial herbal product Cefabronchin® and antibiotic compounds, such as amoxicillin and clarithromycin, on 6 clinical isolates of S. pneumoniae. Therefore, the minimal inhibitory concentration (MIC) of each agent before and after adding Cefabronchin® at different concentrations was determined by applying the checkerboard method. Sub-inhibitory concentrations of the added Cefabronchin® were found to reduce the MIC down to between 3.4% and 29.2% of the amoxicillin MIC and down to between 10.4% and 45.8% of the clarithromycin MIC in all 6 strains. In conclusion, this study provides evidence for the improved antimicrobial effects of commonly used antibiotics in combination with Cefabronchin® in order to combat infections with antibiotic-resistant S. pneumoniae strains.

Novel approach to combat antibiotic resistance: evaluation of some Armenian herb crude extracts for their antibiotic modulatory and antiviral properties.

One of the strategies to combat antibiotic resistance can be the use of plant materials in combination with antibiotics, taking into account that phytochemicals can act as antibiotic resistance-modifying agents. This can give a second life to the traditional antibiotics. The aim was to evaluate antibiotic modulatory effect of crude extracts from Agrimonia eupatoria, Hypericum alpestre, Rumex obtusifolius and Sanguisorba officinalis herbs towards several commercial antibiotics using some Gram-positive and Gram-negative bacteria. The antibiotic modulatory activity was tested by determining MICs of antibiotics in the absence and presence of plant crude extracts at subinhibitory concentrations. Antiviral potential of different extracts of tested plant materials was also explored by double overlay plaque assay. The tested plant crude extracts exhibited high modulatory activity towards used antibiotics. Particularly, high modulatory activity was observed with extracts of H. alpestre and R. obtusifolius. Many plant-antibiotic combinations induced the decrease in MICs of antibiotics up to ~fourfold indicating synergy. Moreover, the similar change was observed at both subinhibitory concentrations (MIC/2 and MIC/4) of the same plant crude extract. High anti-phage activity of plants with the exception of Lilium armenum against T4 phage of Escherichia coli C-T4 was also shown. Plant crude extract or commercial antibiotic combinations significantly increased the efficiency of antibiotics. Tested plant materials with exception of L. armenum have antiviral property. For the first time, antibiotic modulatory activity of tested herb extracts was shown, which could have potential in practical applications. Tested plant materials with exception of L. armenum could have prospective, as a source of new antiviral compounds.

Antimicrobial Activity of Polyphenols and Alkaloids in Middle Eastern Plants.

Antibiotic-resistant microorganisms have been an ever-growing concern over the past years. This has led researchers to direct their attention onto plants to be able to discover new possible antimicrobial compounds. The Middle East encompasses a wide spectrum of plant diversity with over 20,000 different species in habitats ranging from deserts to snow-capped mountains. Several plant secondary metabolites and their derivatives have been identified as possible antimicrobial agents. Among the secondary metabolites studied, alkaloids and polyphenols have shown strong antimicrobial activity. Polyphenols are one of the most numerous and diverse group of secondary metabolites; their antioxidant properties provide the basis for antimicrobial effects. Alkaloids provided the underlying structure for the development of several antibiotics with a diverse range of action. The ability of some plant secondary metabolites to act as resistance-modifying agents is a promising field in mitigating the spread of bacterial resistance.