Method for the estimation of antibacterial activity of the polyfunctional protein from transferrin family in the experimental model of the kinetics of Staphylococcus aureus development

Introduction. Lactoferrin is a cationic monomeric glycoprotein produced by acinar cells and glands, present in different places of the mucous membrane in different concentrations. In connection with the development of various variants of hygienic and medicinal products for the treatment of inflammatory diseases of the oral cavity based on lactoferrin, there was a need for an objective assessment of its antibacterial and antibiofilms properties, followed by an analysis of the preservation of activity in various variants of the isolation of this protein from the substrate and storage.Aim - to improve the effectiveness of evaluating the antibacterial activity of lactoferrin and the duration of its preservation in various biological substrates containing the active substance and individual experimental batches of the manufactured drug using automatic cultivation.Materials and methods. As part of the experiment, a microbiological diagnostic technique employing a system for the automatic cultivation of microbial populations was used. A pre-prepared bacterial suspension was inoculated into the nutrient broth and the studied lactoferrin samples were added, followed by cultivation and analysis of the possible antibacterial effects of transferrin protein. To determine the sensitivity of the isolated strains, we used our own modification of the serial dilution method developed at the Department of microbiology, virology, immunology of the A.I. Yevdokimov Moscow State University of Medicine and Dentistry. The experiment was based on the programmed automatic cultivation using the RTS-1 bioreactor. The interpretation of the results was carried out by changing the optical density at a wavelength of λ = 850 nm. The study of the growth dynamics of microorganisms was carried out in several repetitions, which was reflected in the graphs of the development of bacterial populations. The assessment of the growth control of the corresponding bacterial species was reflected in the change in the optical density values, on the basis of which the curve was built. Results and discussion. According to the results of an experimental study of the growth curves of bacterial populations, statistically significant differences in the number of viable cells in different phases of the growth curves were noted, when using different lactoferrin samples. Higher activity of human recombinant lactoferrin samples was established. An analysis of growth dynamics revealed differences in the onset of the maximum reproduction and its inhibition under the influence of various aggravating factors during cultivation. The bacteriostatic effect of lactoferrin is realized through the binding of iron ions, depriving the bacteria of this microelement, causing inhibition of their development. Along with this, lactoferrin is active against certain virulence factors of microorganisms, splitting them like serine proteases, and thus prevents their penetration into human cells.Conclusion. The method used for automatic cultivation of microorganisms in the bioreactor used allows one to obtain reproducible results, is available for wide use, and can be recommended for obtaining objective, comparable, reliable information about the antimicrobial properties of various samples of the bactericidal protein lactoferrin produced by the domestic pharmaceutical industry. The studied substrate containing recombinant human lactoferrin of Russian production is characterized by high antibacterial activity that persists for 3 years as minimum.

Download Full-text

Stochasticity in Colonial Growth Dynamics of Individual Bacterial Cells

Applied and Environmental Microbiology ◽

10.1128/aem.03629-12 ◽

2013 ◽

Vol 79 (7) ◽

pp. 2294-2301 ◽

Cited By ~ 64

Author(s):

Konstantinos P. Koutsoumanis ◽

Alexandra Lianou

Keyword(s):

Kinetic Parameters ◽

Single Cells ◽

Growth Curves ◽

Growth Dynamics ◽

Time Lapse ◽

Initial Population ◽

Bacterial Cells ◽

Stochastic Growth ◽

Bacterial Populations ◽

Content Type

ABSTRACTConventional bacterial growth studies rely on large bacterial populations without considering the individual cells. Individual cells, however, can exhibit marked behavioral heterogeneity. Here, we present experimental observations on the colonial growth of 220 individual cells ofSalmonella entericaserotype Typhimurium using time-lapse microscopy videos. We found a highly heterogeneous behavior. Some cells did not grow, showing filamentation or lysis before division. Cells that were able to grow and form microcolonies showed highly diverse growth dynamics. The quality of the videos allowed for counting the cells over time and estimating the kinetic parameters lag time (λ) and maximum specific growth rate (μmax) for each microcolony originating from a single cell. To interpret the observations, the variability of the kinetic parameters was characterized using appropriate probability distributions and introduced to a stochastic model that allows for taking into account heterogeneity using Monte Carlo simulation. The model provides stochastic growth curves demonstrating that growth of single cells or small microbial populations is a pool of events each one of which has its own probability to occur. Simulations of the model illustrated how the apparent variability in population growth gradually decreases with increasing initial population size (N0). For bacterial populations withN0of >100 cells, the variability is almost eliminated and the system seems to behave deterministically, even though the underlying law is stochastic. We also used the model to demonstrate the effect of the presence and extent of a nongrowing population fraction on the stochastic growth of bacterial populations.

Download Full-text

Evaluation of Antibacterial Activity of Some Traditionally Used Medicinal Plants against Human Pathogenic Bacteria

BioMed Research International ◽

10.1155/2015/265425 ◽

2015 ◽

Vol 2015 ◽

pp. 1-6 ◽

Cited By ~ 30

Author(s):

Bishnu P. Marasini ◽

Pankaj Baral ◽

Pratibha Aryal ◽

Kashi R. Ghimire ◽

Sanjiv Neupane ◽

...

Keyword(s):

Antibacterial Activity ◽

Medicinal Plants ◽

Inhibitory Action ◽

Pathogenic Bacteria ◽

Cynodon Dactylon ◽

Bacterial Species ◽

Dilution Method ◽

Chloroform Fraction ◽

Ethanolic Extracts

The worldwide increase of multidrug resistance in both community- and health-care associated bacterial infections has impaired the current antimicrobial therapy, warranting the search for other alternatives. We aimed to find thein vitroantibacterial activity of ethanolic extracts of 16 different traditionally used medicinal plants of Nepal against 13 clinical and 2 reference bacterial species using microbroth dilution method. The evaluated plants species were found to exert a range ofin vitrogrowth inhibitory action against the tested bacterial species, andCynodon dactylonwas found to exhibit moderate inhibitory action against 13 bacterial species including methicillin-resistantStaphylococcus aureus, imipenem-resistantPseudomonas aeruginosa, multidrug-resistantSalmonella typhi, andS. typhimurium. The minimum inhibitory concentration (MIC) values of tested ethanolic extracts were found from 31 to >25,000 μg/mL. Notably, ethanolic extracts ofCinnamomum camphora, Curculigo orchioides, andCurcuma longaexhibited the highest antibacterial activity againstS. pyogeneswith a MIC of 49, 49, and 195 μg/mL, respectively; whereas chloroform fraction ofCynodon dactylonexhibited best antibacterial activity againstS. aureuswith a MIC of 31 μg/mL. Among all,C. dactylon, C. camphora, C. orchioides, andC. longaplant extracts displayed a potential antibacterial activity of MIC < 100 μg/mL.

Download Full-text

In-vitro evaluation of cisplatin nanoparticles encompass natural polymer

International Journal of Pharmacometrics and Integrated Biosciences ◽

10.26452/ijpib.v3i1.1204 ◽

2020 ◽

Vol 3 (1) ◽

pp. 1-6

Author(s):

Bhavani J ◽

Sunil Kumar Prajapati ◽

Ravichandran S

Keyword(s):

Antibacterial Activity ◽

Bacterial Infections ◽

Pathogenic Bacteria ◽

Bacterial Species ◽

Research Work ◽

Dilution Method ◽

Bacterial Strains ◽

Active Components ◽

Synthetic Drugs

Opportunistic bacterial infections are common in the various parts of human body. In recent years bacterial species have shown resistance against a number of synthetic drugs. This study measured the antibacterial activity of bacterial strains against five common pathogenic bacteria related strains. Cup plate method and two fold serial dilution method were used to evaluated by antibacterial activity by the help of different bacterial related strains. The results revealed that Cisplatin (CIP) using natural as a polymer showed a minimum inhibitory concentration (MIC) at 250 mg/ml to 500 mg/ml of the broth against all bacterial strains. CIP using natural as a polymer was prepared different doses1000 μg/ml and 2000 μg/ ml and measured zone of inhibition dose dementedly reduced when compared to standard. The CIP using natural as a polymer exhibited strong anti-bacterial activity against five different species of bacteria and this may be attributed to various active components. Our research work has been indicated Nanoparticles containing CIP using natural as a polymer formulated for the enhanced anti-cancer activity through antimicrobial mechanism.

Download Full-text

Antibacterial Activity and Mechanism of Litsea cubeba L. Essential Oil Against Acinetobacter baumannii

Natural Product Communications ◽

10.1177/1934578x21999146 ◽

2021 ◽

Vol 16 (3) ◽

pp. 1934578X2199914

Author(s):

Kaiyuan Hao ◽

Baichang Xu ◽

Geyin Zhang ◽

Feifei Lv ◽

Yuhan Wang ◽

...

Keyword(s):

Antibacterial Activity ◽

Essential Oil ◽

Cell Membrane ◽

Acinetobacter Baumannii ◽

Growth Curves ◽

Antibacterial Activities ◽

Dilution Method ◽

Obvious Effect ◽

Strong Activity ◽

Litsea Cubeba

The main objective of this study was to investigate the antibacterial activity and mechanism of Litsea cubeba essential oil (LCEO) against Acinetobacter baumannii. The antibacterial activity was examined by a serial dilution method and growth curves. The essential oil showed strong activity against A. baumannii. The mechanism of the antibacterial action was evaluated by the integrity and permeability of the membrane, scanning electron microscopy ( SEM) and SDS-PAGE. The change in permeability of the cell membrane and leakage of cell intracellular biomacromolecules verified that LCEO has an obvious effect on the cell membrane. SEM showed the damaging effect of the essential oil on cells since the morphology of the treated bacteria was significantly changed. Different expression of proteins indicated the effect of the essential oil on protein synthesis. It is suggested that LCEO with both emulsifying and antibacterial activities, has a potential to serve as an inhibitor against A. baumannii.

Download Full-text

Novel co-culture plate enables growth dynamic-based assessment of contact-independent microbial interactions

10.1101/145615 ◽

2017 ◽

Author(s):

Thomas J. Moutinho ◽

John C. Panagides ◽

Matthew B. Biggs ◽

Gregory L. Medlock ◽

Glynis L. Kolling ◽

...

Keyword(s):

Real Time ◽

Optical Density ◽

Low Cost ◽

Bacterial Species ◽

Effective Interaction ◽

Microbial Interactions ◽

Burkholderia Cenocepacia ◽

Bacterial Populations ◽

Culture Plate ◽

Mediated Contact

AbstractInteractions between microbes are central to the dynamics of microbial communities. Understanding these interactions is essential for the characterization of communities, yet challenging to accomplish in practice. There are limited available tools for characterizing diffusion-mediated, contact-independent microbial interactions. A practical and widely implemented technique in such characterization involves the simultaneous co-culture of distinct bacterial species and subsequent analysis of relative abundance in the total population. However, distinguishing between species can be logistically challenging. In this paper, we present a low-cost, vertical membrane, co-culture plate to quantify contact-independent interactions between distinct bacterial populations in co-culture via real-time optical density measurements. These measurements can be used to facilitate the analysis of the interaction between microbes that are physically separated by a semipermeable membrane yet able to exchange diffusible molecules. We show that diffusion across the membrane occurs at a sufficient rate to enable effective interaction between physically separate cultures. Two bacterial species commonly found in the cystic fibrotic lung,Pseudomonas aeruginosaandBurkholderia cenocepacia, were co-cultured to demonstrate how this plate may be implemented to study microbial interactions. We have demonstrated that this novel co-culture device is able to reliably generate real-time measurements of optical density data that can be used to characterize interactions between microbial species.

Download Full-text

Potential Antibacterial Activity of Yemeni Sidr Honey Against Pseudomonas aeruginosa and Streptococcus pyogenes

Anti-Infective Agents ◽

10.2174/2211352519666210319100204 ◽

2021 ◽

Vol 19 ◽

Author(s):

Mohammad A. Al-kafaween ◽

Abu Bakar Mohd Hilmi ◽

Hamid A. Nagi Al-Jamal ◽

Rania M. Al-Groom ◽

Nour A. Elsahoryi ◽

...

Keyword(s):

Antibacterial Activity ◽

Pathogenic Bacteria ◽

Growth Curves ◽

Cell Number ◽

Sem Analysis ◽

Dilution Method ◽

Microbial Infection ◽

Log Reduction ◽

Kill Curve ◽

Time Kill

Background: Sidr honey has been reported to exhibit antimicrobial activity against numerous pathogenic bacteria making this honey a promising functional food for the treatment of wounds or stomach ulcers. Objective: The purpose of this study was to investigate the effect of Sidr honey against P. aeruginosa and S. pyogenes. Methods: Minimum inhibitory concentration (MIC) and Minimum bactericidal concentration (MBC) for Sidr honey were determined by the broth dilution method. The growth curve of both bacteria with MIC, half-MIC and quarter-MIC were monitored by optical density (at OD570). The time-kill curve was used to determine the bacteriostatic and bactericidal activity of Sidr honey on both bacteria by plotting colony forming unit (CFUs) versus time. The effect of Sidr honey on the ultrastructure of the P. aeruginosa and S. pyogenes was investigated using scanning electron microscopy (SEM). The effect of Sidr honey on the expression of virulence genes in both bacteria was determined using quantitative reverse transcription polymerase chain reaction (RT-qPCR). Results: The results showed that Sidr honey possessed the lowest MIC value against P. aeruginosa and S. pyogenes with 12.5 % (w/v) and 20 % (w/v) respectively. In addition, the MBC value for Sidr honey was found to be 20% (w/v) and 25% (w/v) against P. aeruginosa and S. pyogenes respectively. Growth curves conducted with MIC Sidr honey resulted in no growth of P. aeruginosa and S. pyogenes. Growth curves with half-MIC Sidr honey resulted in a reduced growth rate and reduction in overall cell number in both bacteria over a period of 24 h, compared with cells grown without honey. In time-kill curve, treatment of P. aeruginosa and S. pyogenes with Sidr honey for 8 hours resulted in decreases of 4-log reduction (P < 0.05) in total viable counts (TVCs). SEM analysis revealed that there were marked changes in the bacterial cell morphology for both bacteria following treatment with Sidr honey. These changes included the appearance of irregular shapes, incomplete cell division, and swelling cells. The RT-qPCR results showed that the expression of algD, oprF, fleN, fleQ, fleR, fliA, and fliC in P. aeruginosa decreased 0.43-fold, 0.38-fold, 0.41-fold, 0.51-fold, 0.40-fold, 0.61-fold, and 0.39-fold respectively after exposure to Sidr honey. Meanwhile the expression of sof, sfbl, and scpA in S. pyogenes decreased 0.18-fold, 0.21-fold, and 0.28-fold respectively after treated with Sidr honey. Conclusion: Using varying methods to evaluate the planktonic integrity, this study demonstrated that Sidr honey has antibacterial activity against both bacteria and has potential as a therapeutic agent for microbial infection particularly against these two organisms. To our knowledge, this study is the first to indicate that Sidr honey is effective at inducing cell lysis and identify targets genes, at the genetic level, that might be involved in this process.

Download Full-text

Isolasi, Identifikasi Senyawa Alkaloid Dan Uji Efektivitas Penghambatan Dari Ekstrak Daun Sirih (Piper betle L.) Terhadap Bakteri Staphylococcus epidermidis

Jurnal e-Biomedik ◽

10.35790/ebm.v8i2.28999 ◽

2020 ◽

Vol 8 (2) ◽

Author(s):

Gwendolyn L. Kapondo ◽

. Fatimawali ◽

Meilani Jayanti

Keyword(s):

Antibacterial Activity ◽

Optical Density ◽

Staphylococcus Epidermidis ◽

Leaf Extract ◽

Ethanol Extract ◽

Dilution Method ◽

Piper Betle ◽

Betel Leaf ◽

Before And After ◽

Density Values

Abstract: Betel leaf (Piper betle L) contains alkaloid compounds and has antibacterial and antiseptic functions. The purpose of this research was to isolated alkaloid compounds also to tested the effectiveness of inhibition of the betel leaf extract against the Staphylococcus epidermidis bacterial growth. Simplicia betel leaf was extracted used 96% of ethanol solvent. Antibacterial activity tested of betel leaf extract with a concentration variation of 25%, 20%, 15%, 10% and 5% using the liquid dilution method. The results of identification of alkaloid isolates used UV-Vis Spectrophotometry method can be known alkaloid compounds contained in betel leaves include alkaloids with indole base framework that was absorption at 262 nm and 274 nm wavelengths. The results of the effectiveness of betel leaf on Bacteria Staphylococcus epidermidis used the liquid dilution method showed that betel leaf extract with Optical density values at concentrations of 25%, 20%, 15% and 10% before and after incubation decreased respectively by -0.347, -0.304, -0.192 and -0.104, while at a concentration of 5% there was increase in Optical density values of 0.162. From the results of the research, it can be concluded that the ethanol extract of betel leaf contains an alkaloid with an indole base framework and was inhibitory activity against Staphylococcus epidermidis bacteria with a MIC at a concentration of 10%.Keywords: betel leaf, Staphylococcus epidermidis, alkaloid compound, liquid dilution. Abstrak: Daun Sirih (Piper betle L) mengandung senyawa alkaloid dan memiliki fungsi sebagai antibakteri dan antiseptik. Penelitian ini bertujuan untuk mengisolasi senyawa alkaloid dan menguji efektivitas penghambatan dari ekstrak daun sirih hijau terhadap pertumbuhan bakteri Staphylococcus epidermidis. Simplisia daun sirih hijau diekstraksi dengan menggunakan pelarut etanol 96%. Uji aktivitas antibakteri ekstrak daun sirih dengan variasi konsentrasi 25%, 20%, 15%, 10% dan 5% menggunakan metode dilusi cair. Hasil identifikasi terhadap isolat alkaloid menggunakan metode Spektrofotometri UV-Vis dapat diketahui senyawa alkaloid yang terkandung dalam daun sirih termasuk alkaloid dengan kerangka dasar Indol yang mempunyai serapan pada panjang gelombang 262 nm dan 274 nm. Hasil uji efektivitas daun sirih terhadap bakteri Staphylococcus epidermidis dengan metode dilusi cair menunjukan bahwa ekstrak daun sirih dengan nilai densitas optik pada konsentrasi 25%, 20%, 15% dan 10% sebelum dan sesudah inkubasi mengalami penurunan berturut-turut sebesar -0.347, -0.304, -0.192 dan -0.104, sedangkan pada konsentrasi 5% mengalami kenaikan nilai densitas optik sebesar 0.162. Dari hasil penelitian yang diperoleh, dapat disimpulkan bahwa ekstrak etanol daun sirih mengandung alkaloid dengan kerangka dasar indol dan memiliki aktivitas penghambatan terhadap bakteri Staphylococcus epidermidis dengan KHM pada konsentrasi 10%.Kata Kunci: daun sirih, Staphylococcus epidermidis, senyawa alkaloid, dilusi cair.

Download Full-text

Acalypha Indica mediated MgO NPs: A novel approach in greener route with its antibacterial activity against pathogens

Nanoscale Reports ◽

10.26524/nr.3.12 ◽

2020 ◽

Vol 3 (2) ◽

Author(s):

Kavipriya K C ◽

Sudha A P ◽

Sujatha K ◽

Sowmya Lakshmi K

Keyword(s):

Antibacterial Activity ◽

Metal Oxides ◽

Bacterial Species ◽

Magnesium Nitrate ◽

Precipitation Method ◽

Average Crystalline Size ◽

Face Centered Cubic ◽

X Ray ◽

Novel Approach ◽

Acalypha Indica

The interest in miniaturization of particles revealed the hidden applications of metal oxides. The potential applications of the particles may vary when the size of the particle is reduced. One of the alternative routes to the conventional approach is the use of plant extract for the synthesis of metal oxides NPs. In the framework of this study, the ecofriendly MgO nanoparticles were synthesized using Acalypha Indica leaf extract,functioning as reducing and capping agent by co-precipitation method. The predecessor taken here was Magnesium Nitrate. The biologically synthesized MgO NPs were characterized by various techniques like X ray diffraction(XRD), Fourier Transform infrared spectroscopy(FTIR), Scanning electron microscope (SEM) with Energy Dispersive X-ray spectroscopy(EDX) profile and its antibacterial activity is evaluated against causative organisms. XRD studies confirmed the face centered cubic crystalline structure of MgO NPs and the average crystalline size of MgO NPs calculated using Scherer’s formula was found to be 13 nm. FTIR spectrum shows a significant Mg-O vibrational band. Purity, surface morphology and chemical composition of elements were confirmed by SEM with EDX. The SEM result shows the fine spherical morphology with the grain size range between 43nm to 62nm. Antimicrobial assay of MgO NPs was examined against gram positive and negative bacteria. Appreciated activity was observed on the Staphylococcus aureus bacterial species. In general, the renewed attempt of this facile approach gave the optimum results of multifunctional MgO NPs.

Download Full-text

Microbiological substantiation of application of phytopreparations in treatment of inflammatory periodontal diseases

Periodontology ◽

10.33925/1683-3759-2019-24-3-196-202 ◽

2019 ◽

Vol 24 (3) ◽

pp. 196-202 ◽

Cited By ~ 1

Author(s):

S. A. Abdurakhmanova ◽

G. S. Runova ◽

M. S. Podporin ◽

E. V. Tsareva ◽

E. V. Ippolitov ◽

...

Keyword(s):

Tooth Loss ◽

Periodontal Diseases ◽

Adult Population ◽

Experimental Studies ◽

Herbal Medicines ◽

Growth Dynamics ◽

Integrated Approach ◽

Bacterial Populations ◽

Streptococcus Constellatus ◽

Urgent Task

Relevance: Inﬂammatory-destructive periodontal diseases are the most complicated and became the main cause of tooth loss in adult population. Herbal medicines have a variety of pharmacological properties, so the development and introduction of new forms for the treatment of inﬂammatory periodontal diseases is an urgent task today.Purpose – experimental evaluation of eﬀectiveness of the use of herbal medicines “Tonzinal” and “CM-1” in relation to the priority periodontal pathogenes.Materials and methods: in experimental studies, the basis for the experiment was the system for the cultivation of microorganisms in real time – the Revers-Spinner RTS-1 bioreactor. With the priority strains of periodontitis pathogens, the study of the growth dynamics of the culture was carried out in several parallels.Results: herbal medicines “CM-1” and “Tonsinal” has a multilateral therapeutic eﬀect, exerting a diverse inﬂuence on the key stages of development of such bacterial populations as Aggregatibacter actinomycetemcomitans, Streptococcus constellatus, Candida albicans.Conclusion: tan integrated approach in the treatment of patients with inﬂammatory periodontal diseases is promising and will contribute to a more prolonged remission and increase the eﬀectiveness of treatment.

Download Full-text

Investigation on the antibacterial activity of electronic cigarette liquids (ECLs): a proof of concept study

Current Pharmaceutical Biotechnology ◽

10.2174/1389201021666200903121624 ◽

2020 ◽

Vol 21 ◽

Cited By ~ 1

Author(s):

Virginia Fuochi ◽

Massimo Caruso ◽

Rosalia Emma ◽

Aldo Stivala ◽

Riccardo Polosa ◽

...

Keyword(s):

Antibacterial Activity ◽

Bacterial Species ◽

A549 Cells ◽

Bactericidal Effect ◽

Minimal Bactericidal Concentration ◽

Viability Assay ◽

Dependent Inhibition ◽

Dose Dependent Inhibition ◽

Dose Dependent

Background: The key ingredients of e-cigarettes liquid are commonly propane-1,2-diol (also called propylene glycol) and propane-1,2,3-triol (vegetal glycerol) and their antimicrobial effects are already established. The nicotine and flavors which are often present in e-liquids can interfere with the growth of some microorganisms. Objective: The effect of the combining these elements in e-liquids is unknown. The aim of the study was to investigate the possible effects of these liquids on bacterial growth in the presence or absence of nicotine and flavors. Methods: Susceptibilities of pathogenic strains (Klebsiella pneumoniae, Staphylococcus aureus, Pseudomonas aeruginosa, Acinetobacter baumannii, Escherichia coli, Enterococcus faecalis and Sarcina lutea) were studied by means of a multidisciplinary approach. Cell viability and antioxidant assays were also evaluated. Results: All e-liquids investigated showed antibacterial activity against at least one pathogenic strain. A higher activity was correlated to the presence of flavors and nicotine. Discussion: In most cases the value of minimal bactericidal concentration is equal to the value of minimal inhibitory concentration showing that these substances have a bactericidal effect. This effect was observed in concentrations up to 6.25% v/v. Antioxidant activity was also correlated to presence of flavors. Over time, the viability assay in human epithelial lung A549 cells showed a dose-dependent inhibition of cell growth. Conclusion: Our results have shown that flavors considerably enhance the antibacterial activity of propane-1,2-diol and propane-1,2,3-triol. This study provides important evidence that should be taken into consideration in further investigative approaches, to clarify the different sensitivity of the various bacterial species to e-liquids, including the respiratory microbiota, to highlight the possible role of flavors and nicotine.

Download Full-text