Влияние in vitro изолированного и сочетанного с антибактериальными средствами применения бовгиалуронидазы азоксимер на целостность бактериальной биопленки и жизнеспосоность микроорганизмов

2020 ◽  
Vol 83 (2) ◽  
pp. 38-44
Author(s):  
Е. Ю. Тризна ◽  
Д. Р. Байдамшина ◽  
Александр А. Виницкий ◽  
А. Р. Каюмов
Keyword(s):  
E Coli

Исследована способность лиофилизата бовгиалуронидазы азоксимера («Лонгидаза») разрушать бактериальные биопленки S. aureus, E. faecalis, E. coli, а также сочетанное действие препарата с антибактериальными средствами. Показано, что 2 ч инкубации бовгиалуронидазы азоксимер в концентрации 750 – 1500 МЕ/мл вызывает двукратное снижение биомассы матрикса зрелых биопленок E. faecalis и E. coli, и на 60 % — S. aureus. Данный ферментный препарат не влияет на образование бактериальных биопленок. При сочетанном применении с антибактериальными средствами препарат повышает их эффективность в отношении бактерий в составе биопленок. Так, концентрация ципро-флоксацина и амоксициллина, необходимая для снижения количества КОЕ на 3 порядка в биопленке E. faecalis, в присутствии бовгиалуронидазы азоксимера снижается в 16 раз (p < 0,05). В присутствии фермента в 16 раз меньшие концентрации цефуроксима, фосфомицина, ципрофлоксацина и амикацина достаточны для снижения количества КОЕ на 3 порядка в биопленке E. coli (p < 0,05), и в значительно меньшей концентрации цефуроксим оказывает бактерицидное действие на клетки в биопленке S. aureus (p < 0,05). Вероятно, бовгиалуронидаза азоксимер увеличивает проникновение антибактериальных средств к клеткам бактерий в биопленке, что обеспечивает потенцирование их антибактериального эффекта. Такое действие ферментного препарата позволяет снизить дозу и повысить безопасность антибактериальных средств при сохранении их эффективности.

Rapid, Refined, and Robust Method for Expression, Purification, and Characterization of Recombinant Human Amyloid-beta M1-42

2019 ◽  
Author(s):  
Priya Prakash ◽  
Travis Lantz ◽  
Krupal P. Jethava ◽  
Gaurav Chopra
Keyword(s):  
Amyloid Beta ◽  
Western Blots ◽  
Force Microscopy ◽  
E Coli ◽  
Atomic Force ◽  
Set Up ◽  
Short Time

Amyloid plaques found in the brains of Alzheimer’s disease (AD) patients primarily consists of amyloid beta 1-42 (Ab42). Commercially, Ab42 is synthetized using peptide synthesizers. We describe a robust methodology for expression of recombinant human Ab(M1-42) in Rosetta(DE3)pLysS and BL21(DE3)pLysS competent E. coli with refined and rapid analytical purification techniques. The peptide is isolated and purified from the transformed cells using an optimized set-up for reverse-phase HPLC protocol, using commonly available C18 columns, yielding high amounts of peptide (~15-20 mg per 1 L culture) in a short time. The recombinant Ab(M1-42) forms characteristic aggregates similar to synthetic Ab42 aggregates as verified by western blots and atomic force microscopy to warrant future biological use. Our rapid, refined, and robust technique to purify human Ab(M1-42) can be used to synthesize chemical probes for several downstream in vitro and in vivo assays to facilitate AD research.

Антимикробная активность лиофилизированного водного извлечения Caragana jubata (Pall.) Poir.

2020 ◽  
Vol 54 (3) ◽  
pp. 42-44
Author(s):  
Павел Алексеевич Какорин ◽  
Татьяна Владимировна Фатеева ◽  
Ольга Ивановна Терешкина ◽  
Ирина Борисовна Перова ◽  
Галина Владиславовна Раменская ◽  
...  
Keyword(s):  
E Coli

На основании ранее проведенных исследований установлен профиль флавоноидов лиофилизированного водного извлечения, полученного из побегов C. jubata. В связи с тем, что, согласно данным литературы, флавоноиды являются потенциальными ингибиторами микроорганизмов, проведено изучение антимикробной активности лиофилизата в опытах in vitro с использованием скринигового метода определения антимикробной активности для препаратов растительного происхождения. При изучении бактериостатической и фунгистатической активности в опытах in vitro использовали метод двукратного серийного разведения препаратов в жидких питательных средах. В результате исследования лиофилизированного водного извлечения караганы гривастой установлено наличие умеренной антимикробной активности в отношении всех изученных штаммов патогенных микроорганизмов: грамположительных и грамотрицательных бактерий (S. aureus, E. coli, P. vulgaris, P. aeruginosa), дрожжеподобных и мицелиальных грибов (C. albicans, M. canis). Полученные данные позволяют рекомендовать лиофилизированное водное извлечение караганы гривастой для создания на его основе лекарственных форм наружного применения для лечения заболеваний кожи и слизистых оболочек, связанных с бактериальным воспалительным процессом.

Properties and Biotechnological Application of mutant Derivatives of Mini-intein PRP8 from Penicillium chrysogenum with Improved Control of C-terminal Processing

2019 ◽  
Vol 35 (6) ◽  
pp. 91-101
Author(s):  
F.A. Klebanov ◽  
S.E. Cheperegin ◽  
D.G. Kozlov
Keyword(s):  
Penicillium Chrysogenum ◽  
Protein Denaturation ◽  
Biologically Active ◽  
Cultivation Temperature ◽  
Target Proteins ◽  
E Coli ◽  
Complete Inactivation ◽  
Target Molecules ◽  
Proteins And Peptides

Mutant variants of mini-intein PRP8 from Penicillium chrysogenum (Int4b) with improved control of C-terminal processing were characterized. The presented variants can serve as a basis for self-removed polypeptide tags capable of carrying an affine label and allowing to optimize the process of obtaining target proteins and peptides in E. coli cells. They allow to synthesize target molecules in the composition of soluble and insoluble hybrid proteins (fusions), provide their afnne purification, autocatalytic processing and obtaining mature target products. The presented variants have a number of features in comparison with the known prototypes. In particular the mutant mini-intein Int4bPRO, containing the L93P mutation, has temperature-dependent properties. At cultivation temperature below 30 °C it allows the production of target molecules as part of soluble fusions, but after increasing of cultivation temperature to 37 °C it directs the most of synthesized fusions into insoluble intracellular aggregates. The transition of Int4bPRO into insoluble form is accompanied by complete inactivation of C-terminal processing. Further application of standard protein denaturation-renaturation procedures enable efficiently reactivate Int4bPRO and to carry out processing of its fusions in vitro. Two other variants, Int4b56 and Int4b36, containing a point mutation T62N or combination of mutations D144N and L146T respectively, have a reduced rate of C-terminal processing. Their use in E. coli cells allows to optimize the biosynthesis of biologically active target proteins and peptides in the composition of soluble fusions, suitable for afnne purification and subsequent intein-dependent processing without the use of protein denaturation-renaturation procedures. intein, fusion, processing, processing rate, gelonin The work was supported within the framework of the State Assignment no. 595-00003-19 PR.

In Vitro Holdase Activity of E. coli Small Heat-Shock Proteins IbpA, IbpB and IbpAB: A Biophysical Study with Some Unconventional Techniques

2014 ◽  
Vol 21 (6) ◽  
pp. 564-571 ◽  
Author(s):  
Sourav Roy ◽  
Monobesh Patra ◽  
Suman Nandy ◽  
Milon Banik ◽  
Rakhi Dasgupta ◽  
...  
Keyword(s):  
Heat Shock ◽  
Heat Shock Proteins ◽  
Small Heat Shock Proteins ◽  
E Coli ◽  
Biophysical Study ◽  
Shock Proteins

Substituted 4-Formyl-2H-chromen-2-ones: Their Reaction with N-(2,3,4,6-Tetra-Oacetyl- β-D-galactopyranosyl)thiosemicarbazide, Antibacterial and Antifungal Activity of Their Thiosemicarbazone Products

2020 ◽  
Vol 24 (19) ◽  
pp. 2272-2282
Author(s):  
Vu Ngoc Toan ◽  
Nguyen Minh Tri ◽  
Nguyen Dinh Thanh
Keyword(s):  
Escherichia Coli ◽  
Staphylococcus Aureus ◽  
Candida Albicans ◽  
Selenium Dioxide ◽  
Antibacterial And Antifungal Activity ◽  
E Coli ◽  
Antibacterial And Antifungal Activities ◽  
Alkyl Ethers ◽  
Antibacterial And Antifungal

Several 6- and 7-alkoxy-2-oxo-2H-chromene-4-carbaldehydes were prepared from corresponding alkyl ethers of 6- and 7-hydroxy-4-methyl-2-oxo-2H-chromen-2-ones by oxidation using selenium dioxide. 6- and 7-Alkoxy-4-methyl-2H-chromenes were obtained with yields of 57-85%. Corresponding 4-carbaldehyde derivatives were prepared with yields of 41-67%. Thiosemicarbazones of these aldehydes with D-galactose moiety were synthesized by reaction of these aldehydes with N-(2,3,4,6-tetra-O-acetyl-β-Dgalactopyranosyl) thiosemicarbazide with yields of 62-74%. These thiosemicarbazones were screened for their antibacterial and antifungal activities in vitro against bacteria, such as Staphylococcus aureus, Escherichia coli, and fungi, such as Aspergillus niger, Candida albicans. Several compounds exhibited strong inhibitory activity with MIC values of 0.78- 1.56 μM, including 8a (against S. aureus, E. coli, and C. albicans), 8d (against E. coli and A. niger), 9a (against S. aureus), and 9c (against S. aureus and C. albicans).

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.

In Vitro Antibacterial activity of ethanolic extract of Myrsine africana against laboratory Strains of Pathogenic Organisms

2016 ◽  
Vol 5 (04) ◽  
pp. 4512
Author(s):  
Jackie K. Obey ◽  
Anthoney Swamy T* ◽  
Lasiti Timothy ◽  
Makani Rachel
Keyword(s):  
Antibacterial Activity ◽  
Minimum Inhibitory Concentration ◽  
Inhibitory Concentration ◽  
Ethanolic Extract ◽  
Ethanol Extract ◽  
E Coli ◽  
Zones Of Inhibition ◽  
In Vitro Antibacterial Activity ◽  
Myrsine Africana

The determination of the antibacterial activity (zone of inhibition) and minimum inhibitory concentration of medicinal plants a crucial step in drug development. In this study, the antibacterial activity and minimum inhibitory concentration of the ethanol extract of Myrsine africana were determined for Escherichia coli, Bacillus cereus, Staphylococcus epidermidis and Streptococcus pneumoniae. The zones of inhibition (mm±S.E) of 500mg/ml of M. africana ethanol extract were 22.00± 0.00 for E. coli,20.33 ±0.33 for B. cereus,25.00± 0.00 for S. epidermidis and 18. 17±0.17 for S. pneumoniae. The minimum inhibitory concentration(MIC) is the minimum dose required to inhibit growth a microorganism. Upon further double dilution of the 500mg/ml of M. africana extract, MIC was obtained for each organism. The MIC for E. coli, B. cereus, S. epidermidis and S. pneumoniae were 7.81mg/ml, 7.81mg/ml, 15.63mg/ml and 15.63mg/ml respectively. Crude extracts are considered active when they inhibit microorganisms with zones of inhibition of 8mm and above. Therefore, this study has shown that the ethanol extract of M. africana can control the growth of the four organisms tested.

scholarly journals Functional identification of ygiP as a positive regulator of the ttdA-ttdB-ygjE operon

Microbiology ◽  
2006 ◽  
Vol 152 (7) ◽  
pp. 2129-2135 ◽  
Author(s):  
Taku Oshima ◽  
Francis Biville
Keyword(s):  
Functional Characterization ◽  
Anaerobic Growth ◽  
Functional Identification ◽  
New Members ◽  
E Coli ◽  
Inner Membrane Protein ◽  
Tartrate Dehydratase

Functional characterization of unknown genes is currently a major task in biology. The search for gene function involves a combination of various in silico, in vitro and in vivo approaches. Available knowledge from the study of more than 21 LysR-type regulators in Escherichia coli has facilitated the classification of new members of the family. From sequence similarities and its location on the E. coli chromosome, it is suggested that ygiP encodes a lysR regulator controlling the expression of a neighbouring operon; this operon encodes the two subunits of tartrate dehydratase (TtdA, TtdB) and YgiE, an integral inner-membrane protein possibly involved in tartrate uptake. Expression of tartrate dehydratase, which converts tartrate to oxaloacetate, is required for anaerobic growth on glycerol as carbon source in the presence of tartrate. Here, it has been demonstrated that disruption of ygiP, ttdA or ygjE abolishes tartrate-dependent anaerobic growth on glycerol. It has also been shown that tartrate-dependent induction of the ttdA-ttdB-ygjE operon requires a functional YgiP.

scholarly journals 1443. Activity of Ceftazidime-Avibactam against Carbapemenase-negative Carbapenem-resistant Enterobacterales (CRE) Isolates from US Hospitals

2020 ◽  
Vol 7 (Supplement_1) ◽  
pp. S725-S725
Author(s):  
Mariana Castanheira ◽  
Timothy B Doyle ◽  
Cory Hubler ◽  
Rodrigo E Mendes ◽  
Helio S Sader
Keyword(s):  
Resistance Mechanisms ◽  
Chemical Diversity ◽  
Services Research ◽  
New Agents ◽  
E Coli ◽  
Department Of Health ◽  
Carbapenem Resistant ◽  
Center Research ◽  
Research Grant

Abstract Background Most CRE isolates in US hospitals produce KPC enzymes, but some do not carry carbapenemases. We investigated the prevalence, resistance mechanisms and activity of ceftazidime-avibactam and comparator agents against CRE that did not carry carbapenemase genes from US hospitals. Additionally, meropenem-resistant isolates were tested for meropenem-vaborbactam. Methods A total of 28,904 Enterobacterales isolates were collected in 70 US hospitals during 2016-2018, and susceptibility tested by reference broth microdilution. Meropenem-vaborbactam was tested using lyophilized panels following the manufacturer’s instructions. CRE isolates were submitted to whole genome sequencing for the screening of b-lactamase genes, multilocus sequence typing, changes in outer membrane protein (OMP) genes and AmpC expression levels. Results A total of 304 (1.1%) CREs were observed in the study period and 45 (14.8%) isolates did not carry carbapenemases. These isolates were mainly Klebsiella aerogenes, Enterobacter cloacae and Klebsiella pneumoniae (11, 11 and 10 isolates, respectively), but also included 5 other species. Acquired b-lactamase genes were detected among 17 isolates and blaCTX-M-15 was the most common (13 isolates). All K. aerogenes and 10 E. cloacae did not carry acquired b-lactamase genes. Ceftazidime-avibactam (100% susceptible) inhibited all isolates at the current breakpoint, followed by tigecycline and amikacin (&gt; 80% susceptible). Other comparators were not active against non-carbapenemase-producing CRE. Nine of 35 meropenem-resistant isolates displayed meropenem-vaborbactam MIC values of ≥ 8 mg/L (nonsusceptible). Further analysis showed that 23 isolates had disruption of OmpC/OmpK36, 4 had disrupted OmpF/OmpK35 and 13 had both OMP genes disrupted. Additionally, 7 isolates had elevated AmpC expression among 17 isolates tested. Among 7 E. coli, 4 were ST131 and only 2 of 10 K. pneumoniae were clonal complex 11. Conclusion Therapy options for treatment of infections caused by CRE were very limited until recent approval of new agents with activity against these isolates. Ceftazidime-avibactam demonstrated full in vitro activity against all carbapenemase-negative CRE carrying multiple resistance mechanisms. Disclosures Mariana Castanheira, PhD, 1928 Diagnostics (Research Grant or Support)A. Menarini Industrie Farmaceutiche Riunite S.R.L. (Research Grant or Support)Allergan (Research Grant or Support)Allergan (Research Grant or Support)Amplyx Pharmaceuticals (Research Grant or Support)Cidara Therapeutics (Research Grant or Support)Cidara Therapeutics (Research Grant or Support)Cipla Ltd. (Research Grant or Support)Cipla Ltd. (Research Grant or Support)Fox Chase Chemical Diversity Center (Research Grant or Support)GlaxoSmithKline (Research Grant or Support)Melinta Therapeutics, Inc. (Research Grant or Support)Melinta Therapeutics, Inc. (Research Grant or Support)Melinta Therapeutics, Inc. (Research Grant or Support)Merck (Research Grant or Support)Merck (Research Grant or Support)Merck & Co, Inc. (Research Grant or Support)Merck & Co, Inc. (Research Grant or Support)Paratek Pharma, LLC (Research Grant or Support)Pfizer (Research Grant or Support)Qpex Biopharma (Research Grant or Support) Timothy B. Doyle, Allergan (Research Grant or Support)Allergan (Research Grant or Support)Cipla Ltd. (Research Grant or Support)Melinta Therapeutics, Inc. (Research Grant or Support)Pfizer (Research Grant or Support)Qpex Biopharma (Research Grant or Support) Cory Hubler, Allergan (Research Grant or Support) Rodrigo E. Mendes, PhD, A. Menarini Industrie Farmaceutiche Riunite S.R.L. (Research Grant or Support)Allergan (Research Grant or Support)Allergan (Research Grant or Support)Basilea Pharmaceutica International, Ltd (Research Grant or Support)Cipla Ltd. (Research Grant or Support)Department of Health and Human Services (Research Grant or Support)GlaxoSmithKline (Research Grant or Support)Melinta Therapeutics, Inc. (Research Grant or Support)Merck (Research Grant or Support)Merck (Research Grant or Support)Pfizer (Research Grant or Support) Helio S. Sader, MD, PhD, A. Menarini Industrie Farmaceutiche Riunite S.R.L. (Research Grant or Support)Allergan (Research Grant or Support)Allergan (Research Grant or Support)Allergan (Research Grant or Support)Cipla Ltd. (Research Grant or Support)Cipla Ltd. (Research Grant or Support)Melinta (Research Grant or Support)Merck (Research Grant or Support)Merck (Research Grant or Support)Paratek Pharma, LLC (Research Grant or Support)Pfizer (Research Grant or Support)

scholarly journals Influence of nutrient availability on in vitro growth of major bovine mastitis pathogens

2021 ◽  
Vol 88 (1) ◽  
pp. 80-88
Author(s):  
Remo Stürmlin ◽  
Josef J. Gross ◽  
Olga Wellnitz ◽  
Lea A. Wagner ◽  
Camille Monney ◽  
...  
Keyword(s):  
Bacterial Growth ◽  
Nutrient Availability ◽  
Bovine Mastitis ◽  
Milk Composition ◽  
B Vitamins ◽  
E Coli ◽  
Streptococcus Uberis ◽  
Whole Milk ◽  
Mastitis Pathogens

AbstractThe aim of the present study was to investigate the effects of milk composition changes on the in vitro growth of bovine mastitis pathogens. Nutritional requirements of three major bovine mastitis pathogens Escherichia coli (E. coli), Staphylococcus aureus (S. aureus), and Streptococcus uberis (S. uberis) were investigated in vitro. We used ultra-high temperature (UHT) treated milk with different contents of fat, protein, and carbohydrates to test the influence of the availability of various milk constituents on pathogen growth characteristics. Additionally, the bacterial growth was investigated under experimentally modified nutrient availability by dilution and subsequent supplementation with individual nutrients (carbohydrates, different nitrogen sources, minerals, and different types of B vitamins) either to milk or to a conventional medium (thioglycolate broth, TB). Varying contents of fat, protein or lactose did not affect bacterial growth with the exception of growth of S. uberis being promoted in protein-enriched milk. The addition of nutrients to diluted whole milk and TB partly revealed different effects, indicating that there are media-specific growth limiting factors after dilution. Supplementation of minerals to diluted milk did not affect growth rates of all studied bacteria. Bacterial growth in diluted whole milk was decreased by the addition of high concentrations of amino acids in S. aureus, and by urea and additional B vitamins in E. coli and S. aureus. The growth rate of S. uberis was increased by the addition of B vitamins to diluted whole milk. The present results demonstrate that growth-limiting nutrients differ among pathogen types. Because reduced bacterial growth was only shown in diluted milk or TB, it is unlikely that alterations in nutrient availability occurring as a consequence of physiological changes of milk composition in the cow's udder would directly affect the susceptibility or course of bovine mastitis.

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