Top PDF Action of antimicrobial peptides against bacterial biofilms

Action of antimicrobial peptides against bacterial biofilms

Action of antimicrobial peptides against bacterial biofilms

Based on their secondary structure, AMPs are generally categorized into four groups (1) α-helical AMPs; (2) β-sheet AMPs; (3) extended AMPs; and (4) cationic loop AMPs [ 53 ]. Alpha-helical peptides are the largest group of AMPs representing 30–50% of all AMPs of known secondary structure [ 54 – 56 ]. These peptides commonly consist of 12–40 amino acids and contain an abundance of helix stabilizing amino acids such as alanine, leucine, and lysine [ 56 ]. Beta-sheet AMPs usually consist of two to ten cysteine residues that from one to five inter-chain disulfide bonds that help the peptides to form the beta-sheet [ 57 ]. Beta-sheet antimicrobial peptides include the defensin family of peptides [ 58 , 59 ]. Defensins consist of two to three antiparallel beta-sheets however, in some cases alpha-helical or unstructured segments can be found at their N- or C-termini [ 60 ]. Compared with α-helical antimicrobial peptides, the defensins adopt a globular structure in aqueous solutions [ 60 , 61 ]. Despite extensive variations in length, amino acid composition and net positive charge, β-strands are observed in all α- and β-defensins [ 62 , 63 ]. Extended AMPs are not folded into α-helix or β-sheet structures. These AMPs often contain high numbers of arginine, tryptophan, proline or cystine residues [ 64 ]. Some of these AMPs can fold into defined amphipathic molecules in bacterial membranes, but often these are not membrane active [ 65 ]. The proline-rich insect-derived pyrrhocoricin, drosocin and apidaecin peptides penetrate membranes and exert their antimicrobial activities by interacting with intracellular proteins such as the heat-shock protein DnaK and GroEL to inhibit the DnaK ATPase and chaperone-assisted protein folding related activities, respectively [ 66 , 67 ]. Cationic loop AMPs are proline-arginine rich peptides, and because of their high numbers of proline residues, they rarely form amphipathic characteristics and tend to adopt polyproline helical type-II structures [ 68 ].
Show more

15 Read more

Host Antimicrobial Peptides: the promise of new treatment strategies against Tuberculosis

Host Antimicrobial Peptides: the promise of new treatment strategies against Tuberculosis

1 Faculty of Biosciences, Department of Biochemistry and Molecular Biology, Universitat Autònoma de Barcelona, Cerdanyola del Vallès, Spain, 2 Mycobacteria Research Laboratory, Department of Biological Sciences, Institute of Structural and Molecular Biology, Birkbeck University of London, London, United Kingdom Tuberculosis (TB) continues to be a devastating infectious disease and remerges as a global health emergency due to an alarming rise of antimicrobial resistance to its treat- ment. Despite of the serious effort that has been applied to develop effective antitubercular chemotherapies, the potential of antimicrobial peptides (AMPs) remains underexploited. A large amount of literature is now accessible on the AMP mechanisms of action against a diversity of pathogens; nevertheless, research on their activity on mycobacteria is still scarce. In particular, there is an urgent need to integrate all available interdisciplinary strategies to eradicate extensively drug-resistant Mycobacterium tuberculosis strains. In this context, we should not underestimate our endogenous antimicrobial proteins and peptides as ancient players of the human host defense system. We are confident that novel antibiotics based on human AMPs displaying a rapid and multifaceted mechanism, with reduced toxicity, should significantly contribute to reverse the tide of antimycobac- terial drug resistance. In this review, we have provided an up to date perspective of the current research on AMPs to be applied in the fight against TB. A better understanding on the mechanisms of action of human endogenous peptides should ensure the basis for the best guided design of novel antitubercular chemotherapeutics.
Show more

17 Read more

Biosynthesis and mode of action of ribosomally synthesized and post-translationally modified antimicrobial peptides

Biosynthesis and mode of action of ribosomally synthesized and post-translationally modified antimicrobial peptides

12 environmental stresses and enabling the bacteria to withstand high internal osmotic pressures. Although the cell walls of different bacterial strains contain several structural differences, the composition of the peptidoglycan polymer is remarkably similar and the pathway to peptidoglycan biosynthesis is highly conserved. The cell wall in Gram-positive bacteria is approximately 20 layers thick, with each layer containing numerous peptidoglycan subunits. By contrast, a relatively small amount of bactoprenyl-phosphate molecules are present at around 200,000 molecules per cell, 52,53 so the amount of lipid II synthesized and available for incorporation is limited. As a result, it is a requirement that the lipid II cycle is an extremely dynamic and efficient process, with each lipid II molecule in the cell having a high turnover rate. 54 The recycling process of lipid phosphate carrier molecule must be tightly controlled and efficient, and is thus regarded as the crux of bacterial cell-wall synthesis. Because an intact cell wall is absolutely essential for bacterial cell survival, peptidoglycan biosynthesis is a key target of many important classes of antibiotics, including penicillins, carbapenems, cephalosporins, the glycopeptides vancomycin 55 and ramoplanin, 56-59 host defense peptides, 46,47 and an increasing number of lantibiotics described in further detail below.
Show more

291 Read more

Nanochitosan antimicrobial activity against Streptococcus mutans and Candida albicans dual species biofilms

Nanochitosan antimicrobial activity against Streptococcus mutans and Candida albicans dual species biofilms

Nanochitosan inhibition capability was evaluated by measuring: cell viability, remaining biofilm mass, and bio- film mass reduction in dual-species biofilms after treating with nanochitosan at various concentrations. It has been previously studied [31, 32] that nanochitosan can inhibit the viability of biofilm cells with the effectivity of more than 90%. Accordingly, the novelty of this study stands on the effect of nanochitosan on dual-species biofilms. Dual- species biofilms are more resistant to antimicrobial agent because the interactions that occur can affect develop- ment, function, and structure of the biofilms formed; dif- ferent from those in the single-species biofilm. One of the reasons is because dual-species biofilms produce more extracellular matrix than single-species biofilms, which causes limited diffusion of antimicrobial substances to reach microbial cells. As a nanoparticle, chitosan has higher penetration rate rather than other antimicro- bial agents (micro size). After penetrating extracellular matrix of biofilms, nanochitosan as a cationic molecule will interact with anionic particles on the cell surface of microorganisms. Modes of action of nanochitosan as cationic biocide are adsorption on microorganism cells, diffusion through the cell wall, adsorption and destruc- tion of the plasma membrane, cytoplasmic component leakage and cell death [33, 34]. Hence, we observed the S. mutans and C. albicans cell viability reduction compar- ing the treatment groups (15%, 30%, 45% nanochitosan concentration) with the control group (0% nanochitosan) (Fig. 3a, b).
Show more

7 Read more

Host Antimicrobial Peptides: the promise of new treatment strategies against Tuberculosis

Host Antimicrobial Peptides: the promise of new treatment strategies against Tuberculosis

secondary granules of eosinophils (162) and secondarily in neu- trophils (163). Complementarily, the signal peptide of the ECP (ECPsp) was found to promote the migration of macrophages via pro-inflammatory molecules to sites of infection and inflam- mation (164). Interestingly, ECP is secreted, together with α-defensin, in response to M. bovis BCG infection (79). Although the recruitment of eosinophils in the respiratory tract during Mtb infection was first regarded as a mere response to inflammation (165), further work has shown that this cell type together with neutrophils can directly participate in the removal of the infec- tion focus (166). Eosinophils are activated via TLR2 induction by the specific mycobacterial wall component, the lipomannan (79). Eosinophils, together with neutrophils, would then release the content of their granules into the granuloma macrophages (84, 159). To note, the eosinophil peroxidase, another eosinophil protein stored in the secondary granules, is also endowed with antimycobacterial activity (119). On the other hand, macrophages express upon bacterial infection two additional RNases, RNase6 and RNase7 (114). In addition, human RNase7, also called the skin derived RNase, is also secreted by keratinocytes and exerts a protective role against a variety of pathogens at the skin bar- rier (39, 115). Interestingly, RNase7, together with RNase3, can eradicate mycobacteria in  vitro (117). Moreover, very recent results indicate that human RNases 3, 6, and 7 can also inhibit the growth of mycobacteria in a macrophage infection model (167). Considering that RNase6 and RNase 7 expression is induced in macrophages upon bacterial infection (114), one might hypothesize that these antimicrobial proteins can also play a physiological role against intracellular dwelling mycobacteria. Eventually, we cannot disregard a complementary contribution of the RNases reported immunomodulatory properties, such as the induction of pro-inflammatory cytokines and the dendritic cell chemoattraction (168, 169).
Show more

18 Read more

Antimicrobial peptides from echinoderms as antibiofilm agents: a natural strategy to combat bacterial infections

Antimicrobial peptides from echinoderms as antibiofilm agents: a natural strategy to combat bacterial infections

Recent research has shown that AMPs also have a high potential for inhibiting the formation of or destroying bio film. In fact, they can act at several stages of bio film formation and with different mechanisms of action: they may minimize the initial adhesion of microbial cells to abiotic surfaces by altering the adhesive features of abiotic surfaces, reducing flagellum-dependent swimming motility, stimulating twitching motility, a type of surface moti- lity that promotes the disassembly of bio film struc- tures or by binding to microbial surfaces via electrostatic interactions; they may prevent bio film maturation by killing the early surface colonizers, or by inhibiting quorum sensing (QS) – that is, the communication system used by many bacteria to build a bio film. QS is a system composed of small molecules that control collective behaviors, such as bioluminescence, virulence factor production and bio film formation (Spoering & Gilmore 2006 ; Horswill et al. 2007 ; Picioreanu et al. 2007 ; Huang et al. 2010 ; Brogden & Brogden 2011 ; de la Fuente- Nunez et al. 2012 ).
Show more

11 Read more

PLANT ANTIMICROBIAL PEPTIDES: A NOVEL APPROACH AGAINST DRUG RESISTANT MICROORGANISMS

PLANT ANTIMICROBIAL PEPTIDES: A NOVEL APPROACH AGAINST DRUG RESISTANT MICROORGANISMS

NaD1 defensin from Nicotiana alata showed antifungal activity against Candida albicans 62 . NaD1 interact with fungal cell wall and cause membrane permeabilization. When NaD1 reaches cytoplasm, it leads to hyper-production of Reactive Oxygen Species (ROS) that induce oxidation damage. High osmolarity glycerol (HOG) pathway was identified which play an important role in protection of fungal cell against NaD1. This indicated that HOG pathway could be a suitable target site for increasing the effectiveness of the antimicrobial peptides against Candida albicans. Putative defensin gene was isolated from Vigna uniguiculata (cowpea). Sequence, amino acid arrangement, splicing analysis, secondary and tertiary structures of mature putative defensin peptide showed similarity towards a typical defensin peptide. Response of biotic stimuli on pathogen treated and untreated plants were examined by the expression level of defensin gene, using RT-PCR 63 . A list of defensin peptides, their origin and action against specific microorganisms is listed in Table 2.
Show more

15 Read more

The effect of chlorhexidine and other antimicrobial agents on the formation and viability of oral bacterial biofilms

The effect of chlorhexidine and other antimicrobial agents on the formation and viability of oral bacterial biofilms

ability to exert an antibacterial effect in the microenvironment (pH, redox potential etc) within the biofilm. The molar concentrations of the three agents used were very similar (0.0013, 0.0014 and 0.001 for CH, CPC and T respectively) but little is known concerning their ability to penetrate biofilms or their ability to kill S. sanguis under the conditions existing within biofilms of the organism. The effectiveness of the three agents in terms of minimum inhibitory concentrations (MIC) shows that CPC is the most effective having an MIC of 0.00024 % compared with 0.001 % for triclosan and 0.0078 % for chlorhexidine (Lim, 1982). But much of this data is, of course, relevant only to planktonic bacteria. Work by Bonesvoll and Gjermo (1978) showed that quaternary ammonium compounds such as CPC are rapidly cleared from the oral cavity, the model could be mimicking this action and cause the CPC to act as in vivo. If the MIC data were used as a guide we would expect triclosan to be more effective as an antibacterial agent than chlorhexidine when used at the same concentrations. Both T and CH achieved greater kills in the biofilms grown on enamel, at least after short-term (1 min.) exposure, due possibly to the uptake of the agent by the biofilm and subsequent retention on it’s surface. In a study of the effects of CH and CPC on biofilms of S. sanguis grown on PTFE (Wilson et a!., 1996), similar kills after 5 mins were found although CPC appeared to be more effective than CH. Differences in the findings of the two studies may be attributable to a number of factors including the nature of the substratum, the presence of other ingredients in the mouthwashes used in the present study and the differing concentrations of the CH used.
Show more

215 Read more

Epithelial antimicrobial peptides in host defense against infection

Epithelial antimicrobial peptides in host defense against infection

Most antimicrobial peptides are cationic (polar) molecules with spatially separated hydrophobic and charged regions. These structural hallmarks are important for the proposed mechanisms of action of peptide antibiotics (see below). Beside these characteristics, antimicrobial peptides of various families differ in size, amino acid sequence and certain structural motifs (Fig. 2). Antimicrobial peptides are gene-encoded, meaning that one gene codes for one peptide. Families of antimicrobial peptide genes are located in clustered arrangements in the genome and map to syntenic chromosomal regions in different mammalian species, providing clues about the evolutionary develop- ment of this host defense system. The primary translational product is a prepropeptide consisting of an N-terminal signal sequence for targeting of the endoplasmic reticu- lum, a pro segment, and a C-terminal cationic peptide that has antimicrobial activity after cleavage (Fig. 3). The pro segment is often anionic in charge and might have several biological functions including the correct folding of the C-terminus, intracellular trafficking, or the inhibition of the Figure 1
Show more

10 Read more

ISOLATION AND CHARACTERIZATION OF ANTIMICROBIAL PEPTIDES FROM DATURA INOXIA LEAVES HAVING ANTIMICROBIAL ACTIVITY AGAINST SELECTED BACTERIA

ISOLATION AND CHARACTERIZATION OF ANTIMICROBIAL PEPTIDES FROM DATURA INOXIA LEAVES HAVING ANTIMICROBIAL ACTIVITY AGAINST SELECTED BACTERIA

well diffusion method against Gram-positive bacteria namely Bacillus subtilis and Staphylococcus aureus, Gram-negative bacteria Escherichia coli and fungi Candida albicans. The clear zone of inhibition was observed and the diameter of the zone was measured in millimeter (mm). Antimicrobial activity was performed with the positive and negative control sample. For positive control samples, Ampicillin (60µg/ml) against Escherichia coli and Staphylococcus aureus, Chloramphenicol (50µg/ml) against Bacillus subtilis and Fluconazole (70µg/ml) against Candida albicans were used (Table 3 and Fig. 2). PBS buffer was used as negative control. The antimicrobial activity of different protein pellets and supernatant indicated that Datura inoxia leaf extract had antimicrobial activity against selected gram- positive bacteria i.e. Staphylococcus aureus and Bacillus subtilis whereas no antimicrobial activity was observed against selected gram-negative bacteria (Escherichia coli) and fungi (Candida albicans) (Table 4 and Fig. 3). The negative result obtained against gram-negative bacteria and fungi might be due to the absence of action against these microbes or due to a low concentration of an active compound or due to loss of its activity in the presence of the microorganism. [11] In the previous reports, PAMPs isolated from leaves with antimicrobial activity the pathogens used were different from that of ours except E. coli. [11,19] Small peptides having antimicrobial properties of less then 10kDa were reported from germinated and non-germinated 50 different types of seeds. [20] Four novel cyclotides (macrocyclic knotted proteins) was isolated from Viola hederaceae (vhl). vhl- 1 was leaf-specific cyclotides with 31-residue. EC50 for vhl-1 was found to be 0.87M against HIV- virus. [21]
Show more

12 Read more

Circular Dichroism studies on the interactions of antimicrobial peptides with bacterial cells

Circular Dichroism studies on the interactions of antimicrobial peptides with bacterial cells

T he emergence of bacterial strains resistant to common antibiotics has strongly encouraged studies on antimicrobial peptides (AMPs) from natural sources and on their mechanism of action 1,2 . It has been demonstrated that antimicrobial peptides impair cell viability by mechanisms which likely depend on their sequence and structure 3 . Biophysical studies carried out using molecules which mimic the outer leaflet of bacterial cells, as lipid mixtures or purified lipopolysaccharides (LPS), leaded to hypothesize three different mechanisms by which peptides kill bacterial cells, namely the barrel stave, toroidal pore and carpet mechanism 4–7 . According to those mechanisms, antimicrobial peptides interact with bacterial outer membranes, perturb their integrity causing their disgregation. Nuclear Magnetic Resonance (NMR) and Circular Dichroism (CD) studies carried out in the presence of either detergents, as mimic of the bacterial outer leaflet, or LPS, which is the main component of the Gram negative bacteria outer membrane, allowed the determination of the three-dimensional and secondary structure of antimicrobial peptides in cell-like environments 8–12 . A different perspective is actually offered by solid state NMR studies, which give interesting insights on the changes occurring to the membranes after interaction with the antimicrobial peptides, other than on the membrane bound structure of peptides 13–15 .
Show more

7 Read more

Exploring the Antimicrobial Action of Quaternary Amines against Acinetobacter baumannii

Exploring the Antimicrobial Action of Quaternary Amines against Acinetobacter baumannii

Protein aggregate isolation. Cellular protein aggregate isolation and analysis were performed as previously described (37). Briefly, 50-ml LB cultures of exponentially growing bacteria were treated with the indicated concentration of BZK or otilonium bromide. Following isolation, aggregates were normal- ized and separated by SDS-PAGE and Coomassie stained for visualization. Aggregates were quantified using the area density feature in VisionWorks LS software (UVP, Inc.) on an image of the aggregate gel. Aggregate proteomics. Protein identification was provided by the Proteomics Facility at the University of Texas at Austin following previously published procedures (61). Scaffold (version Scaf- fold_4.8.2; Proteome Software Inc., Portland, OR) was used to validate tandem mass spectrometry (MS/MS)-based peptide and protein identifications. Tandem and Sequest were set up to search acinetobacter_07-15.fasta (unknown version; 3,798 entries), assuming the presence of the digestion enzyme trypsin. Peptide identifications were accepted if they could be established at greater than 89.0% probability to achieve an FDR of less than 1.0%. Peptide probabilities evaluated using Sequest software were assigned by the Scaffold Local FDR algorithm. Peptide probabilities evaluated using X! Tandem software were assigned by the Peptide Prophet algorithm (62) with Scaffold delta-mass correction. Protein identifications were accepted if they could be established to achieve an FDR of less than 5.0% at greater than 99.0% probability and contained at least 2 identified peptides. Protein probabilities were assigned by the Protein Prophet algorithm (63). Proteins that contained similar peptides and that could not be differentiated based on MS/MS analysis alone were grouped to satisfy the principles of parsimony. Proteins were annotated with Gene Ontology (GO) terms from gene_association.goa_uniprot (down- loaded 14 January 2015) (64). Relative abundances were quantified by the use of normalized exponen- tially modified protein abundance index (emPAI) values, with a minimal value setting of 0.
Show more

13 Read more

On the Physiology and Pathophysiology of Antimicrobial Peptides

On the Physiology and Pathophysiology of Antimicrobial Peptides

An interesting approach has been de- scribed in which gene therapy methods were used to deliver the LL-37 gene into cystic fibrosis xenografts (127). In an- other gene therapy study, adenovirus- mediated gene transfer of the anti- microbial peptide elafin increased the antimicrobial activity of mouse lung cells against S. aureus in vitro and in vivo (128). In exploring potential treatment ap- proaches, the use of bacterial gene ther- apy strategies (alternative gene therapy and bactofection) is interesting, because of higher selectivity of bacteria for air- way cells compared with other gene therapy vectors (129). In general, applica- tion of antimicrobial peptides by using gene therapy can be more effective than direct peptide use. In a recent gene ther- apy study, the cutaneous adenoviral delivery of human cathelicidin was sig- nificantly more effective than the admin- istration of synthetic host defense pep- tides in the treatment of burn wound infections (130).
Show more

9 Read more

DESIGN OF SYNTHETIC ANTIMICROBIAL PEPTIDES AGAINST RESISTANT ACINETOBACTER BAUMANNII USING COMPUTATIONAL APPROACH

DESIGN OF SYNTHETIC ANTIMICROBIAL PEPTIDES AGAINST RESISTANT ACINETOBACTER BAUMANNII USING COMPUTATIONAL APPROACH

Their mechanism of antibacterial action is largely due to electrostatic interactions with anionic moieties of the microbial cell membranes leading to disintegration/ perturbation of cell membrane resulting in cell death 6, 8 , they are less likely to induce resistance in microbes 9 . Three model of actions were suggested, barrel-stave, carpet model and toroidal-pore model in addition to other mechanisms 10 , their mechanisms are completely different from the action of common antibiotics 4 . Despite their great potentials, AMPs have several drawbacks that limits their clinical utility, including hemolytic activity 11 , rapid turnover in human body
Show more

7 Read more

Synergy Pattern of Short Cationic Antimicrobial Peptides Against Multidrug-Resistant Pseudomonas aeruginosa

Synergy Pattern of Short Cationic Antimicrobial Peptides Against Multidrug-Resistant Pseudomonas aeruginosa

The 13mer peptide indolicidin (ILPWKWPWWPWRR- CONH 2 ) is, like bactenecin, a bovine peptide and also belongs to the cathelicidin family. It is present in the cytoplasmic granules of neutrophils ( Selsted et al., 1992 ). The NMR structure reveals that indolicidin forms an extended boat-like structure ( Rozek et al., 2000 ). The peptide is modestly active against various Gram-positive and Gram-negative bacteria, but exhibits high hemolytic activity and cytotoxicity ( Selsted et al., 1992 ; Ahmad et al., 1995 ). In Gram-negative bacteria, indolicidin interacts with surface-exposed lipopolysaccharides (LPS), resulting in a self-promoted uptake across the outer membrane, followed by channel formation in the cytoplasmic membrane, leading to cell death ( Falla et al., 1996 ). Indolicidin has rather weak membrane permeabilization characteristics ( Wu et al., 1999 ). Similarly to Bac2A, indolicidin seems to have multiple modes of action; it interacts with ATP and can inhibit ATP dependent enzymes, inhibits DNA/RNA synthesis, and inhibits protein synthesis ( Subbalakshmi and Sitaram, 1998 ; Hilpert et al., 2010 ). In this study, optimized indolicidin variants are used in addition to indolicidin itself. Variants Indopt1-6 show a similar pattern compared to indolicidin itself. Indopt 2 and 3 also show synergy with MER; Indopt 4 shows additional synergy to KAN and CIP. Indopt 5 shows only two synergistic interactions, again showing that small changes in the sequence can have a strong effect on the synergy. Indopt 7-12 show only very few synergistic interactions. The FICs between combinations of antibiotics and peptides show several synergistic effects if antibiotics are used in tandem with short AMPs. In general, the beta-lactams and beta-lactam- like antibiotics show the lowest amount of synergy, along with ciprofloxacin, a gyrase inhibitor, which also showed a rather low amount of synergy. Cefepime and piperacillin showed no detectable synergy at all. However, antibiotics acting on the ribosome show a higher amount of synergy, with the highest proportion of synergy observed in the use of polymyxin B, which acts on the cell wall and cell membrane. The majority of the results were confirmed by three independent measurements of the selected combinations. Some combinations, however, showed larger FIC values than determined in the first screen, thus proving the importance of verifying FIC data ( Hsieh et al., 1993 ).
Show more

15 Read more

ANTIMICROBIAL ACTIVITY OF PHYLLANTHUS NIRURI AGAINST DIFFERENT HUMAN PATHOGENIC BACTERIAL STRAINS

ANTIMICROBIAL ACTIVITY OF PHYLLANTHUS NIRURI AGAINST DIFFERENT HUMAN PATHOGENIC BACTERIAL STRAINS

P. niruri has several benefits as a herbal medicine. The plant has been found to have hepatoprotective, antilithic, pain-relieving, anti-fungal, diuretic, antispasmodic, hypoglycemic, antiviral and anti-bacterial actions (Paithankar, 2011). The therapeutic action has been investigated with respond to following diseases: diarrhea, dysentery, dropsy, mouth and throat infection, veneral diseases, pimples, eczemas, gangrene, malaria, syphilis, ulcer, urethral secretion, hepatic diseases and gastro-intestinal disorders (Tona et al; 2004).

6 Read more

Antimicrobial and Antibiofilm Activity of Ag and Ni Nanoparticles Against Some Bacterial Pathogens

Antimicrobial and Antibiofilm Activity of Ag and Ni Nanoparticles Against Some Bacterial Pathogens

bacteria. Fortunately, antibiotic resistance mechanisms are irrelevant to NPs. This is because NPs mode of action targets to the cell wall directly with no need to bacterial cell penetration. Accordingly, Attention was focused on materials based on nanoparticles with antibacterial effect [1]. Moreover, NPs act also as a carrier of antibiotics. They increase antibiotics serum levels and inhibit bacterial resistance [3]. Therefore, nanoparticles are promising and can replace conventional materials in many applications due to their ultra- small size and high surface to volume ratio [4]. Antimicrobial activities of metal NPs like Ag, Ni, Co, and Cu have been previously reported [5]. Silver is an antimicrobial metal that is widely used for sterilization purposes including medical devices and water sanitization [6]. Ag NPs have concentration dependent antibacterial activity against strong pathogens such as Escherichia coli and Pseudomonas aeruginosa. [1]. Accordingly, Ag NPs are widely used in antibacterial coating of implantable devices, bone cement, dental materials, wound dressing and other applications [7-10]. Moreover, Ag NPs prevent biofilm formation by inhibition of expression of some bacterial genes [11]. Accordingly, they have an effect on both the developing and matured biofilms [12].
Show more

9 Read more

Mode of Action Studies on Synthetic Antimicrobial Peptides

Mode of Action Studies on Synthetic Antimicrobial Peptides

38 FLOW CYTOMETRY Flowcytometry experiments was included, to investigate the peptide effect on both bacterial DNA content and change in cell size after exposure with peptides. In order to be able to follow the changes over time, samples were extracted at 0, 20, 40, 80, 120 and 180 minutes. ON culture of E.coli ATCC 25922 was diluted 1:50 in fresh MH broth and allowed to grow until an OD 600 of 0,1 had been reached. The suspension was then diluted 1:10 in fresh MH broth and regrown to an OD 600 of 0,1 to ensure a uniform bacterial population. 90 µl of bacterial suspension was loaded into a flat bottomed 96-well Greiner plate for peptide and antibiotic samples and 100 µl for control samples. After extraction of the zero sample, 10 µl of peptide solution corresponding to 1x and 4x the MIC concentration was added for a total volume of 100µl. Two wells were loaded for each treatment to ensure enough bacteria for the flow cytometry analysis. Immediately after extraction, the contents of two wells were transferred to a single E-tube and put on ice. The plate was placed at 37°C between extractions. The samples were, whenever appropriate during the experiment, centrifuged at 10.000x g for 5 min at 4°C. The supernatant was then carefully removed and the cells were resuspended in 100 µl 10mM Tris HCL pH 7,4 and 1000 µl 77% Ethanol was added before storing the samples at 4° C for later analysis. The Rifampicin/Cephalexin samples were prepared in the same way as the other samples with the exception that the 200 µl from the wells were transferred to an E-tube with 45 µl mixture of Rifampicin (300µg/ml) and Cephalexin (36µg/ml) and placed in a 37°C water bath for 2½ hours. Simultaneously, the OD 600 was measured approximately every 25 minutes in a multi- detection microplate reader Synergy HT to monitor bacterial growth. The following day, the samples were centrifuged at 10.000x g for 5 min before gently removing the supernatant and adding 140 µl staining solution (90µg/ml Mitramycin and 20µg/ml Ethidium Bromide in 10 mM Tris pH 7,4, 10mM MgCl 2 ). The samples where run on an A10 Bryte Flow Cytometer and when possible, 20.000 events were included in the analysis.
Show more

90 Read more

Biofilms: Novel strategies based on antimicrobial peptides

Biofilms: Novel strategies based on antimicrobial peptides

* Correspondence: annarita.falanga@unina.it; Tel.: +39-0812534503 Received: 21 May 2019; Accepted: 6 July 2019; Published: 10 July 2019 !"#!$%&'( !"#$%&' ! Abstract: The problem of drug resistance is very worrying and ever increasing. Resistance is due not only to the reckless use of antibiotics but also to the fact that pathogens are able to adapt to di↵erent conditions and develop self-defense mechanisms such as living in biofilms; altogether these issues make the search for alternative drugs a real challenge. Antimicrobial peptides appear as promising alternatives but they have disadvantages that do not make them easily applicable in the medical field; thus many researches look for solutions to overcome the disadvantages and ensure that the advantages can be exploited. This review describes the biofilm characteristics and identifies the key features that antimicrobial peptides should have. Recalcitrant bacterial infections caused by the most obstinate bacterial species should be treated with a strategy to combine conventional peptides functionalized with nano-tools. This approach could e↵ectively disrupt high density infections caused by biofilms. Moreover, the importance of using in vivo non mammalian models for biofilm studies is described. In particular, here we analyze the use of amphibians as a model to substitute the rodent model.
Show more

15 Read more

3D Bioprinting of mature bacterial biofilms for antimicrobial resistance drug testing

3D Bioprinting of mature bacterial biofilms for antimicrobial resistance drug testing

The minimum inhibitory concentration ( MIC ) of antimicrobial agents ( de fi ned as the lowest concentra- tion of an antimicrobial agent at which visible bacterial growth is inhibited after overnight incubation ) is fre- quently calculated during AST to assess antimicrobial ef fi cacy and bacterial resistance [ 10 ] . Methods to determine the MIC based on two-dimensional ( 2D ) planktonic cultures of bacteria are well established [ 11 ] . However, determining the minimal bio fi lm era- dicating concentration ( MBEC ) in bio fi lm infections is much more challenging. This is primarily because in vivo bio fi lm formation is 3D in architecture, which differs to most currently available laboratory models that tend to involve 2D biofilm culture [12–14]. AST of planktonic bacteria therefore tends to give misleading results that do not reflect the increased resistance of bacteria living in a 3D biofilm [15, 16]. This has sig- nificant clinical implications; for example, anti- microbial agents are usually chosen on the basis of their efficacy against 2D planktonic cultures which are more sensitive to treatment than 3D biofilms. Clini- cally this is well demonstrated by cystic fibrosis patients, where treatment of P.aeruginosa infection with antibiotics originally developed against plank- tonic cultures often becomes ineffective once biofilm formation occurs [15]. To develop novel anti- microbials capable of disrupting biofilm formation and resistance in future, 3D in vitro biofilm models more representative of clinical infection are required.
Show more

12 Read more

Show all 10000 documents...