ABSTRACT
We envisioned that the rumen of Kitasato Yakumo beef cattle would contain unique microorganisms which produce bioactive compounds as their defense response to the external environment. The variety of microorganisms were collected from the feces of Kitasato Yakumo beef cattle. We evaluated the biological activity of the culture broth of the isolated strains, proving the utility of our approach.
ABSTRACT
In this paper, we describe our discovery of burnettiene A (1) as an anti-malarial compound from the culture broth of Lecanicillium primulinum (Current name: Flavocillium primulinum) FKI-6715 strain utilizing our original multidrug-sensitive yeast system. This polyene-decalin polyketide natural product was originally isolated as an anti-fungal active compound from Aspergillus burnettii. However, the anti-fungal activity of 1 has been revealed in only one fungal species for and the mechanism of action of 1 remains unknown. After the validation of mitochondrial function inhibitory of 1, we envisioned a new anti-malarial drug discovery platform based on mitochondrial function inhibitory activity. We evaluated anti-malarial activity and 1 showed anti-malarial activity against Plasmodium falciparum FCR3 (chloroquine sensitive) and K1 strain (chloroquine resistant). Our study revealed the utility of our original screening system based on a multidrug-sensitive yeast and mitochondrial function inhibitory activity for the discovery of new anti-malarial drug candidates.
ABSTRACT
KSP-1007 is a novel bicyclic boronate-based broad-spectrum ß-lactamase inhibitor and is being developed in combination with meropenem (MEM) for the treatment of infections caused by carbapenem-resistant Gram-negative bacteria, a global health concern, and here, we describe its characteristics. KSP-1007 exhibited low apparent inhibition constant (Ki app) values against all classes of ß-lactamase, including imipenemase types and oxacillinase types from Acinetobacter baumannii. Against 207 Enterobacterales and 55 A. baumannii, including carbapenemase producers, KSP-1007 at fixed concentrations of 4, 8, and 16 µg/mL dose-dependently potentiated the in vitro activity of MEM in broth microdilution MIC testing. The MIC90 of MEM/KSP-1007 at 8 µg/mL against Enterobacterales was lower than those of MEM/vaborbactam, ceftazidime/avibactam, imipenem/relebactam, and colistin and similar to those of aztreonam/avibactam, cefiderocol, and tigecycline. The in vitro activity of MEM/KSP-1007 at ≥4 µg/mL against Enterobacterales harboring metallo-ß-lactamase was superior to that of cefepime/taniborbactam. MEM/KSP-1007 showed excellent activity against Escherichia coli with PBP3 mutations and New Delhi metallo-ß-lactamase compared to aztreonam/avibactam, cefepime/taniborbactam, and cefiderocol. MEM/KSP-1007 at 8 µg/mL showed greater efficacy against A. baumannii than these comparators except for cefiderocol, tigecycline, and colistin. A 2-fold reduction in MEM MIC against 96 Pseudomonas aeruginosa was observed in combination with KSP-1007. MEM/KSP-1007 demonstrated bactericidal activity against carbapenemase-producing Enterobacterales, A. baumannii, and P. aeruginosa based on minimum bactericidal concentration/MIC ratios of ≤4. KSP-1007 enhanced the in vivo activity of MEM against carbapenemase-producing Enterobacterales, A. baumannii, and P. aeruginosa in murine systemic, complicated urinary tract, and thigh infection models. Collectively, MEM/KSP-1007 has a good profile for treating carbapenem-resistant Gram-negative bacterial infections.
ABSTRACT
We found that the culture broth of fungi showed anti-fungal activity against multidrug-sensitive budding yeast. However, we could not identify the anti-fungal compound due to the small quantity. Therefore, we attempted to increase the productivity of the target compound by the introduction of a global secondary metabolism regulator, laeA to the strain, which led to the successful isolation of 10-folds greater amount of MS-347a (1) than Aspergillus sp. FKI-5362. Compound 1 was not effective against Candida albicans and the detailed anti-fungal activity of 1 remains unverified. After our anti-fungal activity screening, 1 was found to inhibit the growth of broad plant pathogenic fungal species belonging to the Ascomycota. It is noteworthy that 1 showed little insecticidal activity against silkworms, suggesting its selective biological activity against plant pathogenic fungi. Our study implies that the combination strategy of multidrug-sensitive yeast and the introduction of laeA is useful for new anti-fungal drug discovery.
Subject(s)
Drug Discovery , Saccharomyces cerevisiae , Saccharomyces cerevisiae/drug effects , Saccharomyces cerevisiae/genetics , Saccharomyces cerevisiae/metabolism , Drug Discovery/methods , Candida albicans/drug effects , Secondary Metabolism , Fungicides, Industrial/pharmacology , Antifungal Agents/pharmacology , Antifungal Agents/chemistry , Microbial Sensitivity Tests , Ascomycota/drug effects , Ascomycota/genetics , Aspergillus/drug effects , Aspergillus/genetics , Aspergillus/metabolism , Drug Evaluation, Preclinical/methods , Saccharomyces cerevisiae Proteins/genetics , Saccharomyces cerevisiae Proteins/metabolism , Fungal Proteins/genetics , Fungal Proteins/metabolismABSTRACT
Utilizing a binding mode-based physicochemical screening method using d-Ala-d-Ala silica gel, two new macrolactams, named banglactams A (1) and B (2), were discovered from the culture broth of Nonomuraea bangladeshensis K18-0086. In the course of our investigation, we found that d-Ala-d-Ala silica gel precisely differentiated the chemical structures of banglactams and separated them. However, we were not able to obtain enough of 1 to elucidate the structure due to its instability and insolubility. To overcome this challenge, we chemically modified 1 to improve solubility, enabling us to obtain a sufficient material supply for the indirect determination of the structure. Antibacterial activity evaluation of banglactams revealed that 1 binding to d-Ala-d-Ala silica gel exhibited antibacterial activity against Staphylococcus aureus; however, this was not the case with 2. This research indicates the utility of our original binding mode-based PC screening method, and the combination strategy of PC and chemical modifications led us to discover novel antibacterial compounds.
ABSTRACT
Anti-microbial resistance (AMR) is one of the greatest threats to global health. The continual battle between the emergence of AMR and the development of drugs will be extremely difficult to stop as long as traditional anti-biotic approaches are taken. In order to overcome this impasse, we here focused on the type III secretion system (T3SS), which is highly conserved in many Gram-negative pathogenic bacteria. The T3SS is known to be indispensable in establishing disease processes but not essential for pathogen survival. Therefore, T3SS inhibitors may be innovative anti-infective agents that could dramatically reduce the evolutionary selective pressure on strains resistant to treatment. Based on this concept, we previously identified a polyketide natural product, aurodox (AD), as a specific T3SS inhibitor using our original screening system. However, despite its promise as a unique anti-infective drug of AD, the molecular target of AD has remained unclear. In this paper, using an innovative chemistry and genetic biology-based approach, we show that AD binds to adenylosuccinate synthase (PurA), which suppresses the production of the secreted proteins from T3SS, resulting in the expression of bacterial virulence both in vitro and in vivo experiments. Our findings illuminate the potential of PurA as a target of anti-infective drugs and vaccination and could open a avenue for application of PurA in the regulation of T3SS.
Subject(s)
Aurodox , Type III Secretion Systems , Type III Secretion Systems/metabolism , Aurodox/pharmacology , Anti-Bacterial Agents/pharmacology , Anti-Bacterial Agents/chemistry , Gram-Negative Bacteria/metabolism , Bacterial Proteins/metabolismABSTRACT
The emergence and spread of antimicrobial resistance are global threats. Pseudomonas aeruginosa (P. aeruginosa) is responsible for a substantial proportion of this global health issue because of its intrinsic resistance to many antibiotics due to the impermeability of its outer membrane and its multidrug efflux pump systems. Therefore, therapeutic drugs are limited, and the development of new drugs is extremely challenging. As an alternative approach, we focused on a combinational treatment strategy and found that 5-O-mycaminosyltylonolide (OMT) showed potent antibacterial activity against P. aeruginosa in the presence of an efflux pump inhibitor, phenylalanine-arginine beta-naphthylamide (PAßN). In this report, we prepared a PAßN derivative and compared the potentiation activity of OMT by PAßNs against multidrug-resistant P. aeruginosa clinical isolates.
Subject(s)
Anti-Bacterial Agents , Dipeptides , Drug Resistance, Multiple, Bacterial , Microbial Sensitivity Tests , Pseudomonas aeruginosa , Tylosin/analogs & derivatives , Pseudomonas aeruginosa/drug effects , Anti-Bacterial Agents/pharmacology , Anti-Bacterial Agents/chemistry , Drug Resistance, Multiple, Bacterial/drug effects , Dipeptides/pharmacology , Dipeptides/chemistry , Drug Synergism , HumansABSTRACT
Fusaramin (1) was isolated as a mitochondrial inhibitor. However, the fungal producer stops producing 1, which necessitates us to supply 1 by total synthesis. We proposed the complete stereochemical structure based on the biosynthetic pathway of sambutoxin. We have established concise and robust total synthesis of 1, enabling us to determine the complete stereochemical structure and to elucidate the structure-activity relationship, and uncover the hidden antiplant pathogenic fungal activity.
Subject(s)
Anti-Infective Agents , Fungi , Anti-Infective Agents/chemistry , Structure-Activity Relationship , Mycotoxins/chemistryABSTRACT
We discovered a new tetronomycin analog, C-32-OH tetronomycin (2) from the Streptomyces sp. K20-0247 strain, which produces tetronomycin (1). After NMR analysis of 2, we determined the planar structure. Futhermore, the absolute stereochemistry of 2 was deduced based on the biosynthetic pathway of 1 in the K20-0247 strain and a comparison of experimental electronic circular dichroism (ECD) results of 1 with 2. While 2 exihibits potent antibacterial activity aganist Gram-positive baceria including vancomycin-intermediate Staphylococcus aureus (VISA) strains and vancomycin-resistant Enterococci (VRE), the antibacterial activity of 2 shows 16-32-folds weaker than that of 1 suggesting that the C-34 methyl group in 1 is one of the very important functinal group. Moreover, we evaluated the ionophore activity of 1 and 2 and neither compound shows ionophore activity at reasonable concetrations. Our research suggests that 1 and 2 would have different target(s) from an ionophore mechanism in the antibacterial activity and tetronomycins are promising natural products for broad-spectrum antibiotics.
Subject(s)
Anti-Bacterial Agents , Ethers , Anti-Bacterial Agents/pharmacology , Gram-Positive Bacteria , Ionophores , Microbial Sensitivity TestsABSTRACT
Antimicrobial resistance (AMR) causes a global health threat and enormous damage for humans. Among them, Methicillin-resistant Staphylococcus aureus (MRSA) resistant to first-line therapeutic ß-lactam drugs such as meropenem (MEPM) is problematic. Therefore, we focus on combination drug therapy and have been seeking new potentiators of MEPM to combat MRSA. In this paper, we report the isolation of phomoidrides A-D and its new analog, phomoidride H along with a polyketide compound, oxasetin from the culture broth of Neovaginatispora clematidis FKI-8547 strain as potentiators of MEPM against MRSA.
Subject(s)
Methicillin-Resistant Staphylococcus aureus , Pyrroles , Humans , Anti-Bacterial Agents/pharmacology , beta-Lactams/pharmacology , Naphthalenes , Meropenem/pharmacology , Microbial Sensitivity TestsABSTRACT
Six aromatic secondary metabolites, pestalone (1), emodin (2), phomopsilactone (3), pestalachlorides B (4), C (5), and D (6), were isolated from Pestalotiopsis sp. FKR-0115, a filamentous fungus collected from white moulds growing on dead branches in Minami Daito Island. The efficacy of these secondary metabolites against methicillin-resistant Staphylococcus aureus (MRSA) with and without meropenem (ß-lactam antibiotic) was evaluated using the paper disc method and broth microdilution method. The chemical structures of the isolated compounds (1-6) were characterised using spectroscopic methods, including nuclear magnetic resonance and mass spectrometry. All six isolated compounds exhibited synergistic activity with meropenem against MRSA. Among the six secondary metabolites, pestalone (1) overcame bacterial resistance in MRSA to the greatest extent.
Subject(s)
Benzophenones , Methicillin-Resistant Staphylococcus aureus , Methicillin-Resistant Staphylococcus aureus/metabolism , Anti-Bacterial Agents/pharmacology , Meropenem/metabolism , Meropenem/pharmacology , Pestalotiopsis , beta-Lactams/pharmacology , beta-Lactams/metabolism , beta-Lactam Resistance , Microbial Sensitivity TestsABSTRACT
Vancomycin is a potent and broad-spectrum antibiotic that binds to the d-Ala-d-Ala moiety of the growing bacterial cell wall and kills bacteria. This fascinating binding model prompted us to design and synthesize d-Ala-d-Ala silica gels for the establishment of a new physicochemical (PC) screening method. In this report, we confirmed that vancomycin binds to d-Ala-d-Ala silica gel and can be eluted with MeOH containing 50 mM TFA. Finally, d-Ala-d-Ala silica gel enables to purify vancomycin from the culture broth of a vancomycin-producing strain, Amycolatopsis orientalis.
ABSTRACT
The filamentous fungus Synnemellisia sp. strain FKR-0921 was obtained from soil collected on Kume Island, Okinawa. The MeOH extract of FKR-0921 cultured on a solid rice medium yielded a new aromatic compound, synnemellisitriol A (1). The structure, including the absolute configuration, was elucidated by spectroscopic analysis (FT-IR, NMR, and HR-ESI-MS), and the absolute configuration at C-9 of 1 was determined using the modified Mosher's method. Additionally, 1 was evaluated for its biological activities, including metallo-ß-lactamase inhibitory activity, type III secretion system inhibitory activity, antimicrobial activity, antimalarial activity, and cytotoxicity.
Subject(s)
Hypocreales , Phenols , Hypocreales/chemistry , Magnetic Resonance Spectroscopy , Molecular Structure , Spectroscopy, Fourier Transform Infrared , Phenols/chemistry , Phenols/pharmacology , beta-Lactamase Inhibitors/chemistry , beta-Lactamase Inhibitors/pharmacology , Anti-Infective Agents/chemistry , Anti-Infective Agents/pharmacologyABSTRACT
Luminamicin (1) isolated in 1985, is a macrodiolide compound exhibiting selective antibacterial activity against anaerobes. However, the antibacterial activity of 1 was not fully examined. In this research, re-evaluation of the antibacterial activity of 1 revealed that 1 is a narrow spectrum and potent antibiotic againstClostridioides difficile(C. difficile) and effective against fidaxomicin resistantC. difficilestrain. This prompted us to obtain luminamicin resistantC. difficilestrains for the determination of the molecular target of 1 inC. difficile. Sequence analysis of 1-resistantC. difficileindicated that the mode of action of 1 differs from that of fidaxomicin. This is because no mutation was observed in RNA polymerase and mutations were observed in a hypothetical protein and cell wall protein. Furthermore, we synthesized derivatives from 1 to study the structure-activity relationship. This research indicated that the maleic anhydride and the enol ether moieties seem to be pivotal functional groups to maintain the antibacterial activity againstC. difficileand the 14-membered lactone may contribute to taking an appropriate molecular conformation.
ABSTRACT
The emergence and spread of antimicrobial resistant pathogens continue to threaten our ability to combat several infections. Among them, Pseudomonas aeruginosa (P. aeruginosa) poses a major threat to human health. P. aeruginosa has intrinsic resistance to many antibiotics due to the impermeability of its outer membrane and a resistance-nodulation-cell division type multidrug efflux pump system. Therefore, only limited therapeutic drugs are effective against the pathogen. To address this problem, we have recently discovered an overlooked anti- P. aeruginosa compound, 5-O-mycaminosyltylonolide (OMT) from the Omura Natural Compound library using an efflux pump deletion P. aeruginosa mutant strain, YM64. In this report, we aim to demonstrate the promising potential of OMT for as a novel anti- P. aeruginosa compound and performed combination assays of OMT with polymyxin B nonapeptide, an example of a permeabilizing agent, against multi-drug resistant P. aeruginosa clinical isolates.
Subject(s)
Macrolides , Pseudomonas aeruginosa , Humans , Anti-Bacterial Agents/pharmacology , Drug Resistance, Multiple, Bacterial , Macrolides/pharmacology , Microbial Sensitivity Tests , Polymyxin B/pharmacologyABSTRACT
Tetronomycin (1), first isolated from a cultured broth of Streptomyces sp. by Juslen et al. in 1974, is a polycyclic polyether compound. However, the biological activity of 1 has not been thoroughly examined. In this study, we found that 1 exhibits more potent antibacterial activity than two well-known antibacterial drugs (vancomycin and linezolid) and is effective against several drug-resistant clinical isolates including methicillin-resistant Staphylococcus aureus and vancomycin-resistant Enterococci. Furthermore, we reassigned the 13C NMR spectra of 1 and performed a preliminary structure-activity relationship study of 1 to synthesize a chemical probe for target identification, which implied different targets based on its ionophore activity.
ABSTRACT
Screening for bioactivity related to anti-infective, anti-methicillin-resistant Staphylococcus aureus (MRSA) and anti-viral activity, led us to identify active compounds from a methanol extract of Litsea japonica (Thub.) Juss. and the hot water extract of bark of Cinnamomum sieboldii Meisn (also known as Karaki or Okinawa cinnamon). The two main components in these extracts were identified as the catechin trimers (+)-cinnamtannin B1 and pavetannin B5. Moreover, these extracts exhibited anti-severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) activity. The structures of these catechin trimers were previously determined by chemical and spectroscopic methods. Pavetanin B5 has never been reported to be isolated as a pure form and has been obtained as a mixture with another component. Although other groups have reported the putative structure of pavetannin B5, preparation of the methylated derivative of pavetannin B5 in this study allowed us to obtain the pure form for the first time as the undecamethyl derivative and confirm its exact structure. Commercially available (+)-cinnamtannin B1 and aesculitannin B (C2'-epimer of cinnamtannin B1) both of which contained pavetannin B5 as a minor component, and C. sieboldii bark extract (approx. 5/2 mixture of (+)-cinnamtannin B1/pavetannin B5) were assessed for anti-SARS-CoV-2 activity. Both C. sieboldii bark extract and commercially available aesculitannin B showed viral growth inhibitory activity.
Subject(s)
COVID-19 , Catechin , Cinnamomum , Methicillin-Resistant Staphylococcus aureus , Catechin/pharmacology , Plant Bark/chemistry , SARS-CoV-2 , Plant Extracts/chemistryABSTRACT
BACKGROUND: Although crowds are considered to be a risk factor for SARS-CoV-2 transmission, little is known about the changes in environmental surface contamination with the virus when a large number of people attend an event. In this study, we evaluated the changes in environmental surface contamination with SARS-CoV-2. METHODS: Environmental samples were collected from concert halls and banquet rooms before and after events in February to April 2022 when the 7-day moving average of new COVID-19 cases in Tokyo was reported to be 5000-18000 cases per day. In total, 632 samples were examined for SARS-CoV-2 by quantitative reverse transcription polymerase chain reaction (RT-qPCR) tests, and RT-qPCR-positive samples were subjected to a plaque assay. RESULTS: The SARS-CoV-2 RNA detection rate before and after the events ranged from 0% to 2.6% versus 0%-5.0% in environmental surface samples, respectively. However, no viable viruses were isolated from all RT-qPCR-positive samples by the plaque assay. There was no significant increase in the environmental surface contamination with SARS-CoV-2 after these events. CONCLUSIONS: These findings revealed that indirect contact transmission from environmental fomite does not seem to be of great magnitude in a community setting.
Subject(s)
COVID-19 , SARS-CoV-2 , Humans , SARS-CoV-2/genetics , COVID-19/epidemiology , RNA, Viral/genetics , Japan/epidemiology , Risk FactorsABSTRACT
Antimicrobial resistance is a serious, worldwide problem. Pseudomonas aeruginosa (P. aeruginosa) is the pathogen that poses a major threat to human health. However, resistance-nodulation-cell division type multidrug efflux pump systems defend P. aeruginosa from many antibiotics. Therefore, only limited therapeutic drugs are available. In this regard, we screened overlooked anti- P. aeruginosa compounds from the Omura Natural Compound library using an efflux pump deletion P. aeruginosa mutant strain, YM64, which led us to find a semisynthetic macrolide, 5-O-mycaminosyltylonolide, whose anti- P. aeruginosa activity against a standard laboratory adapted strain, PAO1, was enhanced by an efflux pump inhibitor, phenylalanine-arginine beta-naphthylamide.
Subject(s)
Macrolides , Pseudomonas aeruginosa , Humans , Pseudomonas aeruginosa/genetics , Macrolides/pharmacology , Membrane Transport Proteins , Anti-Bacterial Agents/pharmacology , Microbial Sensitivity Tests , Bacterial Proteins/genetics , Bacterial Proteins/metabolismABSTRACT
In this report, we disclose our discovery of a new antifungal natural product, sakurafusariene (1), from an in-house fractionated library of the culture broth of Fusarium sp. FKI-7550 strain by using a combination strategy of multidrug-sensitive yeast and chemical modification. Throughout our investigation, we encountered challenges in the isolation of natural product 1. A chemical modification strategy via alkylation of 1 allowed for removal of the impurities enabling us to elucidate the structure of 1. Furthermore, we synthesized ester derivatives using a method inspired by the isolation study of 1, which gave us valuable information to understand a preliminary structure-activity relationship against Pyricularia oryzae growth inhibitory activity.