Effect of tandem repeats of antimicrobial peptide CC34 on production of target proteins and activity of Pichia pastoris.

Antimicrobial peptides (AMPs) are attracting attention in the fields of medicine, food, and agriculture because of their broad-spectrum antibacterial properties, low resistance, and low-residue in the body. However, the low yield and instability of the prepared AMP drugs limit their application. In this study, we designed a tetramer of the AMP CC34, constructed and transfected two recombinant expression vectors with pGAPZαA containing a haploid CC34 and tetraploid CC34 (CC34-4js) into Pichia pastoris to explore the effect of biosynthesized peptides. The results showed that CC34 and CC34-4js expression levels were 648.2 and 1105.3 mg/L, respectively, in the fermentation supernatant of P. pastoris. The CC34-4js tetramer showed no antibacterial activity, could be cleaved to the monomer using formic acid, and the hemolytic rate of the polyploid was slightly lower than that of monomeric CC34. The average daily gain, average daily feed intake, feed conversion ratio and immune organ index of rats fed CC34 and CC34-4js showed no differences. In conclusion, CC34-4js exhibited a higher yield and lower hemolysis in P. pastoris than those of CC34. Finally, CC34 and CC34-4js enterokinase lysates showed similar antibacterial activity and both expressed peptides potentially improved the growth performance and organ indices of rats.

Synthetic mucus biomaterials for antimicrobial peptide delivery.

Despite the promise of antimicrobial peptides (AMPs) as treatments for antibiotic-resistant infections, their therapeutic efficacy is limited due to the rapid degradation and low bioavailability of AMPs. To address this, we have developed and characterized a synthetic mucus (SM) biomaterial capable of delivering LL37 AMPs and enhancing their therapeutic effect. LL37 is an AMP that exhibits a wide range of antimicrobial activity against bacteria, including Pseudomonas aeruginosa. LL37 loaded SM hydrogels demonstrated controlled release with 70%-95% of loaded LL37 over 8 h due to charge-mediated interactions between mucins and LL37 AMPs. Compared to treatment with LL37 alone where antimicrobial activity was reduced after 3 h, LL37-SM hydrogels inhibited P. aeruginosa (PAO1) growth over 12 h. LL37-SM hydrogel treatment reduced PAO1 viability over 6 h whereas a rebound in bacterial growth was observed when treated with LL37 only. These data demonstrate LL37-SM hydrogels enhance antimicrobial activity by preserving LL37 AMP activity and bioavailability. Overall, this work establishes SM biomaterials as a platform for enhanced AMP delivery for antimicrobial applications.

Expression, purification and investigation of antibacterial activity of a novel hybrid peptide LL37/hBD-129 by applied comprehensive computational and experimental approaches.

Antibiotic-resistant pathogens have become a great universal health concern. Antimicrobial peptides (AMPs) are small amphipathic and cationic polypeptides with high therapeutic potential against various microorganisms containing drug-resistant strains. Two major groups of these peptides, which have antibacterial activity against Gram-positive and Gram-negative bacteria, antiviral activity, and even antifungal activity, are defensins and cathelicidins. Hybridization of various AMPs is an appropriate approach to achieving new fusion AMPs with high antibacterial activity but low cellular toxicity. In the current research, the amino-acid sequence of human cathelicidin LL-37 (2-31) and Human beta-defensin (hBD)-129 were combined, and the fusion protein was evaluated by bioinformatics tool. The designed AMP gene sequence was commercially synthesized and cloned in the pET-28a expression vector. The LL-37/hBD-129 fusion protein was expressed in E.coli BL21-gold (DE3). The expression of the recombinant protein was evaluated using the SDS-PAGE method. The LL37/hBD-129 was successfully expressed as a recombinant hybrid AMP in E.coli BL21-gold (DE3) strain. Purification of the expressed AMP was performed by Ni-NTA column affinity chromatography, and the purified AMP was validated using the Western blot technic. Finally, the antimicrobial activity of the fusion AMP against Staphylococcus aureus and Escherichia coli bacteria was assessed. Based on the in silico analysis and experimental evaluations, the fusion AMP showed a significant antimicrobial effect on E. coli and Staphylococcus aureus bacteria.

Advancements, challenges and future perspectives on peptide-based drugs: Focus on antimicrobial peptides.

Among other health related issues, the rising concerns on drug resistance led to look for alternative pharmaceutical drugs that are effective both against infectious and noninfectious diseases. Antimicrobial peptides (AMPs) emerged as potential therapeutic molecule with wide range of applications. With their limitations, AMPs have gained reputable attentions in research as well as in the pharmaceutical industry. This review highlighted the historical background, research trends, technological advancements, challenges, and future perspectives in the development and applications of peptide drugs. Some vital questions related with the need for pharmaceutical production, factors for the slow and steady journey, the importance of oral bioavailability, and the drug resistance possibilities of AMPs were raised and addressed accordingly. Therefore, the current study is believed to provide a profound understanding in the past and current scenarios and future directions on the therapeutic impacts of peptide drugs.

In Silico and In Vitro Analyses Reveal Promising Antimicrobial Peptides from Myxobacteria.

Antimicrobial resistance (AMR) is a global concern, and as soon as new antibiotics are introduced, resistance to those agents emerges. Therefore, there is an increased appetite for alternative antimicrobial agents to traditional antibiotics. Here, we used in silico methods to investigate potential antimicrobial peptides (AMPs) from predatory myxobacteria. Six hundred seventy-two potential AMP sequences were extracted from eight complete myxobacterial genomes. Most putative AMPs were predicted to be active against Klebsiella pneumoniae with least activity being predicted against Staphylococcus aureus. One hundred seventeen AMPs (defined here as 'potent putative AMPs') were predicted to have very good activity against more than two bacterial pathogens, and these were characterized further in silico. All potent putative AMPs were predicted to have anti-inflammatory and antifungal properties, but none was predicted to be active against viruses. Twenty six (22%) of them were predicted to be hemolytic to human erythrocytes, five were predicted to have anticancer properties, and 56 (47%) were predicted to be biofilm active. In vitro assays using four synthesized AMPs showed high MIC values (e.g. So_ce_56_913 250 µg/ml and Coral_AMP411 125 µg/ml against E. coli). However, antibiofilm assays showed a substantial reduction in numbers (e.g. Coral_AMP411 and Myxo_mac104 showed a 69% and 73% reduction, respectively, at the lowest concentration against E. coli) compared to traditional antibiotics. Fourteen putative AMPs had high sequence similarity to proteins which were functionally associated with proteins of known function. The myxobacterial genomes also possessed a variety of biosynthetic gene clusters (BGCs) that can encode antimicrobial secondary metabolites, but their numbers did not correlate with those of the AMPs. We suggest that AMPs from myxobacteria are a promising source of novel antimicrobial agents with a plethora of biological properties.

Feedback-AVPGAN: Feedback-guided generative adversarial network for generating antiviral peptides.

In this study, we propose Feedback-AVPGAN, a system that aims to computationally generate novel antiviral peptides (AVPs). This system relies on the key premise of the Generative Adversarial Network (GAN) model and the Feedback method. GAN, a generative modeling approach that uses deep learning methods, comprises a generator and a discriminator. The generator is used to generate peptides; the generated proteins are fed to the discriminator to distinguish between the AVPs and non-AVPs. The original GAN design uses actual data to train the discriminator. However, not many AVPs have been experimentally obtained. To solve this problem, we used the Feedback method to allow the discriminator to learn from the existing as well as generated synthetic data. We implemented this method using a classifier module that classifies each peptide sequence generated by the GAN generator as AVP or non-AVP. The classifier uses the transformer network and achieves high classification accuracy. This mechanism enables the efficient generation of peptides with a high probability of exhibiting antiviral activity. Using the Feedback method, we evaluated various algorithms and their performance. Moreover, we modeled the structure of the generated peptides using AlphaFold2 and determined the peptides having similar physicochemical properties and structures to those of known AVPs, although with different sequences.

Efficacy of natural antimicrobial peptides versus peptidomimetic analogues: a systematic review.

Aims: This systematic review was carried out to determine whether synthetic peptidomimetics exhibit significant advantages over antimicrobial peptides in terms of in vitro potency. Structural features - molecular weight, charge and length - were examined for correlations with activity. Methods: Original research articles reporting minimum inhibitory concentration  values against Escherichia coli, indexed until 31 December 2020, were searched in PubMed/ScienceDirect/Google Scholar and evaluated using mixed-effects models. Results: In vitro antimicrobial activity of peptidomimetics resembled that of antimicrobial peptides. Net charge significantly affected minimum inhibitory concentration values (p < 0.001) with a trend of 4.6% decrease for increments in charge by +1. Conclusion: AMPs and antibacterial peptidomimetics exhibit similar potencies, providing an opportunity to exploit the advantageous stability and bioavailability typically associated with peptidomimetics.

Highly selective performance of rationally designed antimicrobial peptides based on ponericin-W1.

Antimicrobial peptides (AMPs) or host-defence peptides act by penetrating and disrupting the bacterial membranes and are therefore less prone to antimicrobial resistance (AMR) compared to conventional antibiotics. However, there are still many challenges in the clinical application of the naturally occurring AMPs which necessitates further studies to establish the relationship between the chemical structure of AMPs and their antimicrobial activity and selectivity. Herein, we report a study on the relationship between the chemical structure and the biological activity of a series of rationally designed AMPs derived from Ponericin-W1, a naturally occurring AMP from ants. The peptides were designed by modification of the hydrophobic and hydrophilic regions of the lead peptide sequence in a systematic way. Their antibacterial and hemolytic activities were determined in vitro. The antibacterial activity of a representative peptide, At5 was also tested in a mouse model of skin wound infection. Furthermore, the relationship between the physicochemical properties of the peptides and their antibacterial activity was investigated. Replacing the cationic amino acids in the hydrophobic region of the peptides with hydrophobic amino acids enhanced their antibacterial activity and increasing the number of cationic amino acids in the hydrophilic region reduced their toxicity to human red blood cells and thus improved their selectivity for bacteria. Four of the designed peptides, coded as At3, At5, At8, and At10, displayed considerable antibacterial activity and high selectivity for bacteria. At5 also accelerated the wound healing in mice indicating high in vivo efficiency of this peptide. The peptides were more effective against Gram-negative bacteria and no AMR was developed against them in the bacteria even after 25 generations. The results from this study can provide a better understanding of the structural features required for strong antibacterial activity and selectivity, and serve as a guide for the future rational design of AMPs.

Stable isotope analysis confirms substantial changes in the fatty acid composition of bacteria treated with antimicrobial random peptide mixtures (RPMs).

Resistance of plant-pathogenic bacteria to classic antibiotics has prompted the search for suitable alternative antimicrobial substances. One promising strategy could be the use of purposely synthesized random peptide mixtures (RPMs). Six plant-pathogenic bacteria were cultivated and treated with two RPMs previously found to show antimicrobial activity mainly by bacterial membrane disruption. Here, we show that bacteria treated with RPMs showed partly remarkable changes in the fatty acid pattern while those unaffected did not. Quantitative changes could be verified by compound specific isotope analysis of δ13C values (‰). This technique was employed due to the characteristic feature of stronger bonds between heavier isotopes in (bio)chemical reactions. As a proof of concept, the increase in abundance of a fatty acid group after RPM treatment was accompanied with a decrease in the 13C content and vice versa. We propose that our findings will help designing and synthesizing more selective antimicrobial peptides.

Identification of Bacterial Membrane Selectivity of Romo1-Derived Antimicrobial Peptide AMPR-22 via Molecular Dynamics.

The abuse or misuse of antibiotics has caused the emergence of extensively drug-resistant (XDR) bacteria, rendering most antibiotics ineffective and increasing the mortality rate of patients with bacteremia or sepsis. Antimicrobial peptides (AMPs) are proposed to overcome this problem; however, many AMPs have attenuated antimicrobial activities with hemolytic toxicity in blood. Recently, AMPR-11 and its optimized derivative, AMPR-22, were reported to be potential candidates for the treatment of sepsis with a broad spectrum of antimicrobial activity and low hemolytic toxicity. Here, we performed molecular dynamics (MD) simulations to clarify the mechanism of lower hemolytic toxicity and higher efficacy of AMPR-22 at an atomic level. We found four polar residues in AMPR-11 bound to a model mimicking the bacterial inner/outer membranes preferentially over eukaryotic plasma membrane. AMPR-22 whose polar residues were replaced by lysine showed a 2-fold enhanced binding affinity to the bacterial membrane by interacting with bacterial specific lipids (lipid A or cardiolipin) via hydrogen bonds. The MD simulations were confirmed experimentally in models that partially mimic bacteremia conditions in vitro and ex vivo. The present study demonstrates why AMPR-22 showed low hemolytic toxicity and this approach using an MD simulation would be helpful in the development of AMPs.

Characterization of a Novel Antimicrobial Peptide Isolated from Moringa oleifera Seed Protein Hydrolysates and Its Membrane Damaging Effects on Staphylococcus aureus.

The present study sought to identify and characterize a novel antimicrobial peptide, named MOp2 from Moringa oleifera seed protein hydrolysates, and elucidate its potential antimicrobial effects on Staphylococcus aureus. MOp2, with the amino acid sequence of His-Val-Leu-Asp-Thr-Pro-Leu-Leu (HVLDTPLL), was characterized as a hydrophobic anionic AMP of the ß-sheet structure. MOp2 exhibited negligible hemolytic activity at 2.0× MIC, suggesting its inhibitory effect on the growth of S. aureus (MIC: 2.204 mM). It maintained more than 90% of antimicrobial activity under 5% salt and about 78% of antimicrobial activity at a high temperature of 115 °C for 30 min. Protease, especially acid protease, reduced its antimicrobial activity to different extents. Moreover, MOp2 caused irreversible membrane damage to S. aureus cells by increasing the membrane permeability, resulting in the release of intracellular nucleotide pools. Additionally, molecular docking revealed that MOp2 could inhibit S. aureus growth by interacting with dihydrofolate reductase and DNA gyrase through hydrogen bonding and hydrophobic interactions. Overall, MOp2 could be a potential novel antimicrobial agent against S. aureus in food processing.

Machine Learning Prediction of Antimicrobial Peptides.

Antibiotic resistance constitutes a global threat and could lead to a future pandemic. One strategy is to develop a new generation of antimicrobials. Naturally occurring antimicrobial peptides (AMPs) are recognized templates and some are already in clinical use. To accelerate the discovery of new antibiotics, it is useful to predict novel AMPs from the sequenced genomes of various organisms. The antimicrobial peptide database (APD) provided the first empirical peptide prediction program. It also facilitated the testing of the first machine-learning algorithms. This chapter provides an overview of machine-learning predictions of AMPs. Most of the predictors, such as AntiBP, CAMP, and iAMPpred, involve a single-label prediction of antimicrobial activity. This type of prediction has been expanded to antifungal, antiviral, antibiofilm, anti-TB, hemolytic, and anti-inflammatory peptides. The multiple functional roles of AMPs annotated in the APD also enabled multi-label predictions (iAMP-2L, MLAMP, and AMAP), which include antibacterial, antiviral, antifungal, antiparasitic, antibiofilm, anticancer, anti-HIV, antimalarial, insecticidal, antioxidant, chemotactic, spermicidal activities, and protease inhibiting activities. Also considered in predictions are peptide posttranslational modification, 3D structure, and microbial species-specific information. We compare important amino acids of AMPs implied from machine learning with the frequently occurring residues of the major classes of natural peptides. Finally, we discuss advances, limitations, and future directions of machine-learning predictions of antimicrobial peptides. Ultimately, we may assemble a pipeline of such predictions beyond antimicrobial activity to accelerate the discovery of novel AMP-based antimicrobials.

Mechanism of Action and Therapeutic Potential of the ß-Hairpin Antimicrobial Peptide Capitellacin from the Marine Polychaeta Capitella teleta.

Among the most potent and proteolytically resistant antimicrobial peptides (AMPs) of animal origin are molecules forming a ß-hairpin structure stabilized by disulfide bonds. In this study, we investigated the mechanism of action and therapeutic potential of the ß-hairpin AMP from the marine polychaeta Capitella teleta, named capitellacin. The peptide exhibits a low cytotoxicity toward mammalian cells and a pronounced activity against a wide range of bacterial pathogens including multi-resistant bacteria, but the mechanism of its antibacterial action is still obscure. In view of this, we obtained analogs of capitellacin and tachyplesin-inspired chimeric variants to identify amino acid residues important for biological activities. A low hydrophobicity of the ß-turn region in capitellacin determines its modest membranotropic activity and slow membrane permeabilization. Electrochemical measurements in planar lipid bilayers mimicking the E. coli membrane were consistent with the detergent-like mechanism of action rather than with binding to a specific molecular target in the cell. The peptide did not induce bacterial resistance after a 21-day selection experiment, which also pointed at a membranotropic mechanism of action. We also found that capitellacin can both prevent E. coli biofilm formation and destroy preformed mature biofilms. The marked antibacterial and antibiofilm activity of capitellacin along with its moderate adverse effects on mammalian cells make this peptide a promising scaffold for the development of drugs for the treatment of chronic E. coli infections, in particular those caused by the formation of biofilms.

Molecular mechanism underlying the TLR4 antagonistic and antiseptic activities of papiliocin, an insect innate immune response molecule.

SignificanceSimilar to mammalian TLR4/MD-2, the Toll9/MD-2-like protein complex in the silkworm, Bombyx mori, acts as an innate pattern-recognition receptor that recognizes lipopolysaccharide (LPS) and induces LPS-stimulated expression of antimicrobial peptides such as cecropins. Here, we report that papiliocin, a cecropin-like insect antimicrobial peptide from the swallowtail butterfly, competitively inhibits the LPS-TLR4/MD-2 interaction by directly binding to human TLR4/MD-2. Structural elements in papiliocin, which are important in inhibiting TLR4 signaling via direct binding, are highly conserved among insect cecropins, indicating that its TLR4-antagonistic activity may be related to insect Toll9-mediated immune response against microbial infection. This study highlights the potential of papiliocin as a potent TLR4 antagonist and safe peptide antibiotic for treating gram-negative sepsis.

Comparative study of antibacterial activity and stability of D-enantiomeric and L-enantiomeric bovine NK-lysin peptide NK2A.

L-enantiomers of antimicrobial peptides (AMPs) are sensitive to proteolytic degradation; however, D-enantiomers of AMPs are expected to provide improved proteolytic resistance. The present study aimed to comparatively investigate the in vitro antibacterial activity, trypsin and serum stability, toxicity, and in vivo antibacterial activity of L-enantiomeric bovine NK2A (L-NK2A) and its D-enantiomeric NK2A (D-NK2A). Circular dichroism spectroscopy of D-NK2A and L-NK2A in anionic liposomes showed α-helical structures and the α-helical conformation of D-NK2A was a mirror image of L-NK2A. Both D-NK2A and L-NK2A displayed minimal in vitro and in vivo toxicities. RP-HPLC and mass spectrometry analyses revealed that D-NK2A, but not L-NK2A, was resistant to trypsin digestion. D-NK2A and L-NK2A showed similar in vitro bacterial killing activities against Histophilus somni. Slightly reduced antibacterial activity was observed when D-NK2A and L-NK2A were pre-incubated with serum. Confocal and transmission electron microscopic findings confirmed that both peptides induced disruption of bacterial inner- and outer-membranes. Improved survivals with D-NK2A treatment were observed when compared to L-NK2A in a murine model of acute H. somni septicemia. We conclude that antibacterial activity and mode of action of NK2A are not chiral specific. With further optimization, D-NK2A may be a viable AMP candidate to combat bacterial infections.

[Molecular design and biological activity analysis of antimicrobial peptide RIKL].

Natural antimicrobial peptides have strong bactericidal activities. An obstacle of the development of antimicrobial peptides resides in the difficulty of developing peptides with high biocompatibility. In this study, molecular dynamics analysis was employed to assess the structural characteristics and biological activities of peptides. A (RXKY)2(YRY)2 structure was used as a template to design an antimicrobial peptide RIKL of high-efficiency and low-toxicity, where X represents Ile and Y represents Leu. The secondary structure of the antimicrobial peptide was detected by circular dichroism (CD), and the structures of RIKL in water and in POPC/POPG membrane environment were measured using molecular dynamics. The biological activity of RIKL was further studied by assessing its antimicrobial activity, hemolytic activity, eukaryotic cytotoxicity, and salt ion stability. CD results showed that RIKL presented an α-helical structure in a simulated bacterial membrane environment. Molecular dynamics simulation predicted that the secondary structure of RIKL could be partly retained in water and POPG environment, while this secondary structure was weakened in the POPC environment. Antimicrobial test suggested that RIKL had high antimicrobial activities, and the geometric mean of the Minimum Inhibitory Concentration (MIC) was 3.1 µmol/L. The hemolysis indicated that RIKL had no hemolytic activity within the detection range, and cytotoxicity test suggested the cytotoxicity of RIKL was low. Stability test showed that RIKL maintained antimicrobial activities under different pH, serum concentrations and salt environments. Based on the above results, RIKL has high cell selectivity and has the potential as a highly effective antibacterial drug.

Accelerating the discovery of antifungal peptides using deep temporal convolutional networks.

The application of machine intelligence in biological sciences has led to the development of several automated tools, thus enabling rapid drug discovery. Adding to this development is the ongoing COVID-19 pandemic, due to which researchers working in the field of artificial intelligence have acquired an active interest in finding machine learning-guided solutions for diseases like mucormycosis, which has emerged as an important post-COVID-19 fungal complication, especially in immunocompromised patients. On these lines, we have proposed a temporal convolutional network-based binary classification approach to discover new antifungal molecules in the proteome of plants and animals to accelerate the development of antifungal medications. Although these biomolecules, known as antifungal peptides (AFPs), are part of an organism's intrinsic host defense mechanism, their identification and discovery by traditional biochemical procedures is arduous. Also, the absence of a large dataset on AFPs is also a considerable impediment in building a robust automated classifier. To this end, we have employed the transfer learning technique to pre-train our model on antibacterial peptides. Subsequently, we have built a classifier that predicts AFPs with accuracy and precision of 94%. Our classifier outperforms several state-of-the-art models by a considerable margin. The results of its performance were proven as statistically significant using the Kruskal-Wallis H test, followed by a post hoc analysis performed using the Tukey honestly significant difference (HSD) test. Furthermore, we identified potent AFPs in representative animal (Histatin) and plant (Snakin) proteins using our model. We also built and deployed a web app that is freely available at https://tcn-afppred.anvil.app/ for the identification of AFPs in protein sequences.

Theta-Defensins to Counter COVID-19 as Furin Inhibitors: In Silico Efficiency Prediction and Novel Compound Design.

Coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), was characterized as a pandemic by the World Health Organization (WHO) in Dec. 2019. SARS-CoV-2 binds to the cell membrane through spike proteins on its surface and infects the cell. Furin, a host-cell enzyme, possesses a binding site for the spike protein. Thus, molecules that block furin could potentially be a therapeutic solution. Defensins are antimicrobial peptides that can hypothetically inhibit furin because of their arginine-rich structure. Theta-defensins, a subclass of defensins, have attracted attention as drug candidates due to their small size, unique structure, and involvement in several defense mechanisms. Theta-defensins could be a potential treatment for COVID-19 through furin inhibition and an anti-inflammatory mechanism. Note that inflammatory events are a significant and deadly condition that could happen at the later stages of COVID-19 infection. Here, the potential of theta-defensins against SARS-CoV-2 infection was investigated through in silico approaches. Based on docking analysis results, theta-defensins can function as furin inhibitors. Additionally, a novel candidate peptide against COVID-19 with optimal properties regarding antigenicity, stability, electrostatic potential, and binding strength was proposed. Further in vitro/in vivo investigations could verify the efficiency of the designed novel peptide.

Selective Detection of Legionella pneumophila Serogroup 1 and 5 with a Digital Photocorrosion Biosensor Using Antimicrobial Peptide-Antibody Sandwich Strategy.

Rapid detection of Legionella pneumophila (L. pneumophila) is important for monitoring the presence of these bacteria in water sources and preventing the transmission of the Legionnaires' disease. We report improved biosensing of L. pneumophila with a digital photocorrosion (DIP) biosensor functionalized with an innovative structure of cysteine-modified warnericin antimicrobial peptides for capturing bacteria that are subsequently decorated with anti-L. pneumophila polyclonal antibodies (pAbs). The application of peptides for the operation of a biosensing device was enabled by the higher bacterial-capture efficiency of peptides compared to other traditional ligands, such as those based on antibodies or aptamers. At the same time, the significantly stronger affinity of pAbs decorating the L. pneumophila serogroup-1 (SG-1) compared to serogroup-5 (SG-5) allowed for the selective detection of L. pneumophila SG-1 at 50 CFU/mL. The results suggest that the attractive sensitivity of the investigated sandwich method is related to the flow of an extra electric charge between the pAb and a charge-sensing DIP biosensor. The method has the potential to offer highly specific and sensitive detection of L. pneumophila as well as other pathogenic bacteria and viruses.

An antimicrobial peptide specifically active against Listeria monocytogenes is secreted by Bacillus pumilus SF214.

BACKGROUND: Members of the Bacillus genus produce a large variety of antimicrobial peptides including linear or cyclic lipopeptides and thiopeptides, that often have a broad spectrum of action against Gram-positive and Gram-negative bacteria. We have recently reported that SF214, a marine isolated strain of Bacillus pumilus, produces two different antimicrobials specifically active against either Staphylococcus aureus or Listeria monocytogenes. The anti-Staphylococcus molecule has been previously characterized as a pumilacidin, a nonribosomally synthesized lipopetide composed of a mixture of cyclic heptapeptides linked to fatty acids of variable length. RESULTS: Our analysis on the anti-Listeria molecule of B. pumilus SF214 indicated that it is a peptide slightly smaller than 10 kDa, produced during the exponential phase of growth, stable at a wide range of pH conditions and resistant to various chemical treatments. The peptide showed a lytic activity against growing but not resting cells of Listeria monocytogenes and appeared extremely specific being inactive also against L. innocua, a close relative of L. monocytogenes. CONCLUSIONS: These findings indicate that the B. pumilus peptide is unusual with respect to other antimicrobials both for its time of synthesis and secretion and for its strict specificity against L. monocytogenes. Such specificity, together with its stability, propose this new antimicrobial as a tool for potential biotechnological applications in the fight against the dangerous food-borne pathogen L. monocytogenes.