scholarly journals Enzyme-catalyzed biodegradation of penicillin fermentation residues by β-lactamase OtLac from Ochrobactrum tritici

2021 ◽  
Vol 20 (1) ◽  
Author(s):  
Peng Wang ◽  
Chen Shen ◽  
Qinqin Cong ◽  
Kaili Xu ◽  
Jialin Lu
Keyword(s):  
Large Scale ◽  
In Vitro Study ◽  
Scale Removal ◽  
E Coli ◽  
Penicillin V ◽  
Highly Active ◽  
Steady State Kinetics ◽  
Biodegradation Process ◽  
Km Value

Abstract Background Biodegradation of antibiotics is a promising method for the large-scale removal of antibiotic residues in the environment. However, the enzyme that is involved in the biodegradation process is the key information to be revealed. Results In this study, the beta-lactamase from Ochrobactrumtritici that mediates the biodegradation of penicillin V was identified and characterized. When searching the proteins of Ochrobactrumtritici, the β-lactamase (OtLac) was identified. OtLac consists of 347 amino acids, and predicted isoelectric point is 7.0. It is a class C β-lactamase according to BLAST analysis. The coding gene of OtLac was amplified from the genomic DNA of Ochrobactrumtritici. The OtLac was overexpressed in E. coli BL21 (DE3) and purified with Ni2+ column affinity chromatography. The biodegradation ability of penicillin V by OtLac was identified in an in vitro study and analyzed by HPLC. The optimal temperature for OtLac is 32 ℃ and the optimal pH is 7.0. Steady-state kinetics showed that OtLac was highly active against penicillin V with a Km value of 17.86 μM and a kcat value of 25.28 s−1 respectively. Conclusions OtLac demonstrated biodegradation activity towards penicillin V potassium, indicating that OtLac is expected to degrade penicillin V in the future.

Acetate Kinase (AcK) is Essential for Microbial Growth and Betel-derived Compounds Potentially Target AcK, PhoP and MDR Proteins in M. tuberculosis, V. cholerae and Pathogenic E. coli: An in silico and in vitro Study

2019 ◽  
Vol 18 (31) ◽  
pp. 2731-2740 ◽  
Author(s):  
Sandeep Tiwari ◽  
Debmalya Barh ◽  
M. Imchen ◽  
Eswar Rao ◽  
Ranjith K. Kumavath ◽  
...  
Keyword(s):  
Broad Spectrum ◽  
In Silico ◽  
Pathogenic Bacteria ◽  
In Vitro Study ◽  
Docking Studies ◽  
Acetate Kinase ◽  
E Coli ◽  
Pathogenic Escherichia Coli ◽  
Novel Target

Background: Mycobacterium tuberculosis, Vibrio cholerae, and pathogenic Escherichia coli are global concerns for public health. The emergence of multi-drug resistant (MDR) strains of these pathogens is creating additional challenges in controlling infections caused by these deadly bacteria. Recently, we reported that Acetate kinase (AcK) could be a broad-spectrum novel target in several bacteria including these pathogens. Methods: Here, using in silico and in vitro approaches we show that (i) AcK is an essential protein in pathogenic bacteria; (ii) natural compounds Chlorogenic acid and Pinoresinol from Piper betel and Piperidine derivative compound 6-oxopiperidine-3-carboxylic acid inhibit the growth of pathogenic E. coli and M. tuberculosis by targeting AcK with equal or higher efficacy than the currently used antibiotics; (iii) molecular modeling and docking studies show interactions between inhibitors and AcK that correlate with the experimental results; (iv) these compounds are highly effective even on MDR strains of these pathogens; (v) further, the compounds may also target bacterial two-component system proteins that help bacteria in expressing the genes related to drug resistance and virulence; and (vi) finally, all the tested compounds are predicted to have drug-like properties. Results and Conclusion: Suggesting that, these Piper betel derived compounds may be further tested for developing a novel class of broad-spectrum drugs against various common and MDR pathogens.

Effect of Ethanol/Trisodium Citrate Lock on Microorganisms Causing Hemodialysis Catheter-Related Infections

2007 ◽  
Vol 8 (4) ◽  
pp. 262-267 ◽  
Author(s):  
T.A. Takla ◽  
S.A. Zelenitsky ◽  
L.M. Vercaigne
Keyword(s):  
In Vitro Study ◽  
Trisodium Citrate ◽  
Methicillin Resistant ◽  
Hemodialysis Catheter ◽  
E Coli ◽  
Lock Solution ◽  
Mueller Hinton ◽  
Mueller Hinton Broth ◽  
Made In

Purpose This in vitro study tested the effectiveness of a novel 30% ethanol/4% trisodium citrate (TSC) lock solution against the most common pathogens causing hemodialysis catheter-related infections. Methods Clinical isolates of methicillin-resistant Staphylococcus aureus (MRSA) (n=4), methicillin-sensitive S. aureus (MSSA) (n=8), methicillin-resistant Staphylococcus epidermidis (MRSE) (n=8), Pseudomonas aeruginosa (n=4) and Escherichia coli (n=4) were tested in duplicate. Bacterial suspensions of each isolate were made in a control solution of normal saline and Mueller-Hinton broth (MHB), and in a lock solution of ethanol 30%, TSC 4% and MHB. Suspensions were incubated at 37 °C for 48 h. Colony counts were determined from samples collected at t=0 h (before exposure to the ethanol/TSC lock), t=1 h (one hour after exposure to the ethanol/TSC lock), t=24 h and t=48 h. To confirm the absence of viable organisms in the lock solution, the remaining volume at 48 h was filtered through a 0.45 μm filter. The filter was rinsed with 15 mL sterile water and plated on tryptic soy agar (TSA). Results All controls demonstrated significant growth over 48 h. In the lock solutions, initial inocula were reduced to 0 viable colonies by t=1 h (6-log kill), and there was no growth at t=24 and 48 h. Filtering of lock solutions also showed no growth. These results were consistent among duplicates of all isolates. Conclusions The 30% ethanol/4% TSC lock solution consistently eradicated MRSA, MSSA, MRSE, P. aeruginosa and E. coli within 1 h of exposure. Experiments are currently underway to test this novel lock solution on preventing biofilm production by these pathogens.

scholarly journals The Production of the Recombinant Protein of Anthrax Bacteriolysine PlyPH and In Vitro Study of the Lytic Properties of the Protein Coded for them Against Bacillus anhracis Microbial Cells

2019 ◽  
pp. 5-22
Author(s):  
Onuchina N.V., Soybanov V.D.
Keyword(s):  
In Vitro Study ◽  
Coli Strain ◽  
Vitro Study ◽  
Chromosomal Dna ◽  
Lytic Enzymes ◽  
Microbial Cells ◽  
Phage Gene ◽  
E Coli ◽  
Species Specific

The causative agent of anthrax - Bacillus anthracis, due to the prevalence of its natural foci in Russia, high virulence for humans and most mammals, the unique resistance of spore forms to environmental factors and repeated use in terrorist acts, is an extremely dangerous biological agent. Therefore, the search for new effective drugs for the diagnosis and treatment of anthrax, including diseases caused by antibiotic-resistant strains of B. anthracis is necessary. The use of lytic enzymes of species-specific bacteriophages is a new trend in the diagnosis, prevention and treatment of infectious diseases. The goal of this work is the cloning of the anthrax bacteriolysin PlyPH gene as part of the pTrcHis2C vector in Escherichia coli and the in vitro study of the lytic properties of the protein encoded by it against B. anthracis microbial cells. According to the complete sequencing of the B. anthracis genomes of the Ames, Stern 34F2 and JB17 strains, a prophage was found in their chromosomal DNA, which lost part of the structural genes necessary for its replication, but retained a gene with a high degree of homology with the bacteriolysin γ phage gene. For amplification and subsequent cloning of the PlyPH gene, we developed primers containing EcoRI and BamHI restriction enzyme recognition sites. Amplification of the PlyPH gene in a polymerase chain reaction (PCR) with a developed pair of primers was performed using the Stern 34F2 strain of the anthrax microbe as a template. Based on the obtained amplification products and the pTrcHis2C vector, we constructed a recombinant plasmid containing the bacteriolysin synthesis PlyPH gene and stably functioning in the cells of the recombinant E. coli strain. In the course of research, it has been established that microbial cells of the E. coli recombinant TOP10 strain provide for the production of the bacteriolysin of the anthrax prophage, PlyPH , which has the ability to in vitro lyse the vegetative cells of the STI-1 vaccine strain of B. anthracis

scholarly journals Antimicrobial efficacy of a new tri-antibiotic combination against resistant endodontic pathogens: An in-vitro study

2020 ◽  
Vol 23 (4) ◽  
pp. 8p ◽  
Author(s):  
Prasanna T. Dahake ◽  
Sudhindra M Baliga
Keyword(s):  
Root Canal ◽  
Pathogenic Bacteria ◽  
Culture Media ◽  
In Vitro Study ◽  
Vitro Study ◽  
New Combination ◽  
Antimicrobial Efficacy ◽  
E Coli ◽  
Background Removal

Background: Removal of all the pathogenic bacteria from the root canal system is of prime importance for the success of endodontic therapy. Objective: The study aimed to determine the antimicrobial efficacy of three antibiotics and their new combination against selected endodontic pathogens. Methods: In this in-vitro study, we used bacterial strains associated with the refractory endodontic condition and determined MIC and MBC of Clindamycin (C), Metronidazole (M), Doxycycline (D) as well as their combination CMD. We cultured Candida Albicans, Pseudomonas Aeruginosa, Escherichia Coli, Enterococcus Faecalis, Streptococcus Mutans, Bacillus Subtilis subsp. spizizenii, Actinomyces Actinomycetemcomitans on selective culture media. We analyzed the data using paired 't' test, one-way ANOVA, and Tuckey's HSD post hoc test. Results: Clindamycin inhibited the growth of C. Albicans (90%) and S. Mutans (90%) significantly and P. Aeruginosa, E. Coli, E. Faecalis, B. Subtilis, and A. Actinomycetemcomitans were resistant to it. Metronidazole did not inhibit any of the bacteria. Doxycycline inhibited C. Albicans (90%), P. Aeruginosa (90%), and S. Mutans (90%) significantly while E. Coli, E. Faecalis, B. Subtilis, and A. Actinomycetemcomitans were resistant to it. The combination of CMD inhibited all the microbes significantly. However, at bactericidal concentrations of CMD, E. Faecalis (p = 0.024), B. Subtilis (p = 0.021) and A. Actinomycetemcomitans (p = 0.041) were eliminated significantly, while C. Albicans (p = 0.164), P. Aeruginosa (p = 0.489), E. Coli (p = 0.106) and S. Mutans (p = 0.121) showed resistance. Conclusion: Combination CMD can be used against resistant endodontic pathogens to achieve predictable endodontic results.KEYWORDSAntimicrobial agents; Clindamycin; Doxycycline; Metronidazole; Root canal therapy.    

scholarly journals Volatile Oils ofNepeta tenuifolia(Jing Jie) as an Alternative Medicine against Multidrug-Resistant Pathogenic Microbes

2018 ◽  
Vol 2018 ◽  
pp. 1-8 ◽  
Author(s):  
Yi-Hsuan Lee ◽  
Chao-Min Wang ◽  
Po-Yu Liu ◽  
Ching-Chang Cheng ◽  
Zong-Yen Wu ◽  
...  
Keyword(s):  
Essential Oils ◽  
In Vitro Study ◽  
Multidrug Resistant ◽  
Clinical Isolates ◽  
Gram Negative ◽  
E Coli ◽  
Wide Range ◽  
Main Component ◽  
Reference Strains

Essential oils from the dried spikes ofNepeta tenuifolia(Benth) are obtained by steam distillation. Pulegone was identified as the main component in the spikes ofN. tenuifoliathrough analysis, with greater than 85% purity obtained in this study. The essential oils are extremely active against all Gram-positive and some Gram-negative reference bacteria, particularlySalmonella enterica,Citrobacter freundii, andEscherichia coli. The minimum inhibitory concentration was found to be between 0.08 and 0.78% (againstS. enterica), 0.39 and 0.78% (againstC. freundii), and 0.097 and 0.39% (againstE. coli), whereas the minimum bactericidal concentration varied in range from 0.097% to 1.04%. In general, the essential oils show a strong inhibitory action against all tested reference strains and clinical isolates. However, the antibacterial activity of EOs against bothPseudomonas aeruginosareference strains and clinical isolates was relatively lower than other Gram-negative pathogens. The essential oils ofN. tenuifoliaalso displayed bactericidal activities (MBC/MIC < 4) in this study. These findings reflect the bactericidal activity of the essential oils against a wide range of multidrug-resistant clinical pathogens in an in vitro study. In addition, we propose the fragmentation pathways of pulegone and its derivatives by LC-ESI-MS/MS in this study.

scholarly journals 2214. Comparison of Cefepime–Zidebactam (WCK 5222), Ceftazidime–Avibactam, and Ceftolozane–Tazobactam Tested Against Gram-Negative Organisms Causing Pneumonia in United States Hospitals in 2018

2019 ◽  
Vol 6 (Supplement_2) ◽  
pp. S755-S755 ◽  
Author(s):  
Helio S Sader ◽  
Cecilia G Carvalhaes ◽  
Rodrigo E Mendes ◽  
Mariana Castanheira ◽  
Robert K Flamm
Keyword(s):  
Therapeutic Option ◽  
High Dose ◽  
Gram Negative ◽  
E Coli ◽  
Highly Active ◽  
Medical Centers ◽  
Carbapenem Resistant ◽  
Gram Negative Organisms ◽  
Dual Mechanism

Abstract Background Zidebactam (ZID) is a bicyclo-acyl hydrazide antibiotic with a dual mechanism of action: selective Gram-negative PBP2 binding and β-lactamase inhibition. We evaluated the frequency and antimicrobial susceptibility (S) of Gram-negative bacilli (GNB) isolated from patients with pneumonia in US hospitals. Methods All 3,086 clinical isolates were consecutively collected from patients hospitalized with pneumonia (1/patient) in 29 US medical centers in 2018, and the GNB (n = 2,171) were S tested against cefepime (FEP)-ZID (1:1 ratio) and comparators by reference broth microdilution methods. The FEP S breakpoint of ≤8 mg/L (CLSI, high dose) was applied to FEP-ZID for comparison purposes. An FEP-ZID S breakpoint of ≤64 mg/L has been proposed for non-fermentative GNB based on pharmacokinetic/pharmacodynamic target attainment and was applied. Enterobacterales (ENT) isolateswere screened for β-lactamase genes by whole-genome sequencing. Results GNB represented 70.3% of the collection, and the most common GNB were P. aeruginosa (PSA; 34.9% of GNB), K. pneumoniae (10.9%), E. coli (9.7%), S. marcescens (7.7%), and S. maltophilia (XM; 6.4%). FEP-ZID was highly active against PSA (MIC50/90, 2/8 mg/L; 98.8% and 99.9% inhibited at ≤8 and ≤16 mg/L, respectively; highest MIC, 32 mg/L), including resistant subsets (table). Among comparators, colistin (99.6%S), ceftazidime–avibactam (CAZ-AVI; 95.2%S), and ceftolozane–tazobactam (C-T; 94.5%S) were the most active compounds against PSA. FEP-ZID inhibited all ENT at ≤4 mg/L, including ESBL-producers (MIC90, 0.25 mg/L) and carbapenem-resistant ENT (MIC90, 4 mg/L). The most active comparators against ENT were CAZ-AVI (99.9%S), amikacin (98.5%S), and meropenem (MEM; 98.3%S). FEP-ZID inhibited 75.0% and 97.9% of XM isolates at ≤8 and ≤16 mg/L, respectively (highest MIC, 64 mg/L). The only other compounds active against XM were co-trimoxazole (MIC50/90, ≤0.12/2 mg/L; 95.7%S) and levofloxacin (MIC50/90, 1/2 mg/L; 70.7%S). FEP-ZID inhibited 71.0% and 98.9% of A. baumannii isolates at ≤8 and ≤64 mg/L,, respectively. Conclusion FEP-ZID showed potent in vitro activity against GNB causing pneumonia in US hospitals and may represent a valuable therapeutic option for these difficult-to-treat infections Disclosures All authors: No reported disclosures.

Functional and structural differences between photosynthetic and heterotrophic Rhodospirillum rubrum ribosomes and S-100 fractions

1976 ◽  
Vol 22 (10) ◽  
pp. 1522-1539 ◽  
Author(s):  
C. T. Chow
Keyword(s):  
Rhodospirillum Rubrum ◽  
Photosynthetic Pigment ◽  
Free Protein ◽  
E Coli ◽  
Highly Active ◽  
Structural Differences ◽  
S 100 ◽  
Cell Free Protein Synthesis ◽  
Fraction Addition

Cell-free, protein-synthesizing activity has been tested by using various combinations of the S-100 and ribosome fractions prepared from photosynthetic and heterotrophic Rhodospirillum rubrum. The photosynthetic ribosomes are highly active when combined with either the photosynthetic or the heterotrophic S-100 fractions, whereas the heterotrophic ribosomes are active only when combined with the photosynthetic S-100 fraction. Addition of a photosynthetic pigment-containing fraction to the homologous heterotrophic system is, however, able to stimulate its activity. An inhibitor and an activator involved in cell-free protein synthesis have been isolated from the stationary heterotrophic cells. The inhibitor is a very small, dialyzable compound which inhibits not only the R. rubrum but also the E. coli protein-synthesizing activity in vitro, whereas the activator is a non-dialyzable, small RNA molecule capable of stimulating only the R. rubrum activity. Differences exist between the photosynthetic and the heterotrophic systems in their response to various chemical compounds and to light as well as in their structure.

scholarly journals Comparative antimicrobial efficacy study of different commercially available toothpaste in India: An in vitro study

2021 ◽  
pp. 122-131
Keyword(s):  
In Vitro Study ◽  
Chemotherapeutic Agents ◽  
Oral Bacteria ◽  
Oral Microbiome ◽  
Antimicrobial Efficacy ◽  
Bacteria Growth ◽  
E Coli ◽  
Test Organisms ◽  
Oral Pathogens

Antimicrobial chemotherapeutic agents have been recommended for lowering oral bacteria growth. The main purpose of this study is to examine the efficacy of different toothpaste formulations in providing complete oral cavity protection against oral pathogens. By using a modified well agar diffusion assay, twenty kinds of toothpaste were examined for antimicrobial efficacy against two oral pathogens: Streptococcus aureus and Escherichia coli. The examination indicated that the majority of the non-herbal dentifrices and combinations of herbal and chemical-based dentifrices chosen for the investigation were viable against both microbial strains, however, to differing degrees. TP1 and TP17 were found the best against E. coli and S. aureus, respectively, with 21.553 mm and 23.443 mm as the zone of inhibition. From the herbal dentifrices, TP15 was found to have significant effect on E. coli, followed by TP19 for S. aureus. Nevertheless, toothpaste TP15 and TP19 were not effective against S. aureus and E. coli, respectively. In correlation, the inhibition zones of every single other dentifrice were found to be less. Antimicrobial activity against test organisms was stronger in a sodium lauryl sulphate-based dental formulations, when combined with fluoride. A formulation including TP15 exhibited substantial activity against the tested bacterium E. coli among herbal dentifrices. Statistical analysis demonstrated that the effectiveness against Gram-negative bacteria was greater than against Gram-positive bacteria. Furthermore, herbal toothpaste can be incorporated with chemotherapeutic agents to enhance its effectiveness against pathogens present in the oral microbiome. This comparison aids in the identification of the toothpaste’s shortcomings and benefits over other formulations, widening the scope of more potent toothpaste products.

scholarly journals Ebolavirus Entry: From Molecular Characterization to Drug Discovery

2019 ◽  
Author(s):  
Cristiano Salata ◽  
Arianna Calistri ◽  
Gualtiero Alvisi ◽  
Michele Celestino ◽  
Cristina Parolin ◽  
...  
Keyword(s):  
Life Cycle ◽  
Viral Entry ◽  
Large Scale ◽  
Ebola Virus ◽  
Virus Disease ◽  
Drug Repurposing ◽  
In Vitro Study ◽  
Rational Drug Design ◽  
Global Public Health

Ebola Virus Disease (EVD) is one of the most lethal transmissible infections characterized by a high fatality rate, and caused by members of the Filoviridae family. The recent large outbreak of EVD in West Africa (2013-2016), highlighted the worldwide danger of this disease and its impact on global public health and economy. The development of highly needed anti-Filoviridae antivirals has been so far hampered by the shortage of tools to study their life cycle in vitro, and therefore screen for potential active compounds outside a biosafety level-4 (BSL-4) containment. Importantly, the development of surrogate models to in vitro study of Filoviridae entry in a BSL-2 setting, such as viral pseudotypes and Ebola virus like particles, tremendously boosted both our knowledge on viral life cycle and the identification of promising anti-Filoviridae compounds interfering with viral entry. In this context, the combination of such surrogate systems with large-scale small molecule compounds and haploid genetic screenings, as well as rational drug design and drug repurposing approaches will prove priceless in our quest for the development of a treatment for EVD.

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