Antimicrobial and antibiofilm activity of unionid mussels from the north of Portugal

The populations of freshwater mussels belonging to the family Unionidae have been facing drastic changes in terms of diversity and numbers caused by constant aggressions on their natural habitat. Nevertheless, bivalves are capable of developing strategies of defense to overcome potential aggressors. The present work aimed to assess the potential antibacterial capacity of different species of unionid mussels from the north of Portugal. For this purpose, circulating cells (hemocytes), fluids, and mucus were obtained by nonlethal methods from the species Anodonta anatina (Linnaeus), Anodonta cygnea (Linnaeus), Potomida littoralis (Cuvier), and Unio delphinus (Spengler), and tested against bacterial reference strains and multidrug-resistant isolates. The cellular fraction of A. anatina, A. cygnea, and P. littoralis showed antibacterial activity, detected by the agar disc diffusion method, against Bacillus subtilis ATCC 6683, Pseudomonas aeruginosa ATCC 27853 and Acinetobacter baumannii ATCC 19606. Circulating cells from P. littoralis and A. anatina also inhibited Listeria monocytogenes ATCC 19111 and A. cygnea has also inhibited a multidrug-resistant isolate of Pseudomonas putida. The plasma of all mentioned freshwater mussels, used directly or diluted, showed great ability to hamper or inhibit the biofilm formation of Staphylococcus aureus ATCC 25923, P. aeruginosa ATCC 27853, and Escherichia coli ATCC 25922. Anodonta cygnea hampered the biofilm formation by Enterococcus faecalis ATCC 29212 as well. Overall, these results showed that not only cells play a relevant role in the immune system of these species but also the plasma, which likely contains antibiofilm substances. Anodonta cygnea stood out by presenting the best antibacterial inhibition potential.This work was supported by the Portuguese Foundation for Science and Technology (FCT) with a research grant to M. H. (SFRH/BD/76265/2011). This article was also supported by the project INNOVMAR-Innovation and Sustainability in the Management and Exploitation of Marine Resources (NORTE-01-0145-FEDER-000035, within Research Line NOVELMAR), by North Portugal Regional Operational Programme( NORTE2020), under the PORTUGAL2020 Partnership Agreement, and through the European Regional Development Fund (ERDF).info:eu-repo/semantics/publishedVersio

“Clicking” an ionic liquid to a potent antimicrobial peptide: on the route towards improved stability

A covalent conjugate between an antibacterial ionic liquid and an antimicrobial peptide was produced via “click” chemistry, and found to retain the parent peptide’s activity against multidrug-resistant clinical isolates of Gram-negative bacteria, and antibiofilm action on a resistant clinical isolate of Klebsiella pneumoniae, while exhibiting much improved stability towards tyrosinase-mediated modifications. This unprecedented communication is a prelude for the promise held by ionic liquids -based approaches as tools to improve the action of bioactive peptides.info:eu-repo/semantics/publishedVersio

Turning a Collagenesis-Inducing Peptide Into a Potent Antibacterial and Antibiofilm Agent Against Multidrug-Resistant Gram-Negative Bacteria

Antimicrobial resistance is becoming one the most serious health threats worldwide, as it not only hampers effective treatment of infectious diseases using current antibiotics, but also increases the risks of medical procedures like surgery, transplantation, bone and dental implantation, chemotherapy, or chronic wound management. To date, there are no effective measures to tackle life-threatening nosocomial infections caused by multidrug resistant bacterial species, of which Gram-negative species within the so-called "ESKAPE" pathogens are the most worrisome. Many such bacteria are frequently isolated from severely infected skin lesions such as diabetic foot ulcers (DFU). In this connection, we are pursuing new peptide constructs encompassing antimicrobial and collagenesis-inducing motifs, to tackle skin and soft tissue infections by exerting a dual effect: antimicrobial protection and faster healing of the wound. This produced peptide 3.1-PP4 showed MIC values as low as 1.0 and 2.1 μM against Escherichia coli and Pseudomonas aeruginosa, respectively, and low toxicity to HFF-1 human fibroblasts. Remarkably, the peptide was also potent against multidrug-resistant isolates of Klebsiella pneumoniae, E. coli, and P. aeruginosa (MIC values between 0.5 and 4.1 μM), and hampered the formation of/disaggregated K. pneumoniae biofilms of resistant clinical isolates. Moreover, this notable hybrid peptide retained the collagenesis-inducing behavior of the reference cosmeceutical peptide C16-PP4 ("Matrixyl"). In conclusion, 3.1-PP4 is a highly promising lead toward development of a topical treatment for severely infected skin injuries.info:eu-repo/semantics/publishedVersio

Growth assessment of Helicobacter pylori in liquidmedium – effect of aggregation

Helicobacter pylori is a pathogenic organism associated with gastric diseases. It is described that H. pylori can change morphology when exposed to adverse conditions and H. pylori cells can aggregate in clusters when in liquid culture. Such phenomenon makes it difficult to assess growth using the conventional methods. The development of robust methods to assess growth in a more reliable way is needed. In the present work a method that allows efficient cell disaggregation was developed.Fundação para a Ciência e Tecnologia (FCT) PhD Grant (SFRH/BD/47596/2008

Synergistic and antibiofilm properties of ocellatin peptides against multidrug-resistant Pseudomonas aeruginosa

Aim:To test ocellatin peptides (ocellatins-PT2-PT6) for antibacterial and antibiofilm activities and synergy with antibiotics against Pseudomonas aeruginosa. Materials & methods: Normal- and checkerboard-broth microdilution methods were used. Biofilm studies included microtiter plate-based assays and microscopic analysis by confocal laser scanning microscopy and atomic force microscopy. Results: Ocellatins were more active against multidrug-resistant isolates of P. aeruginosa than against susceptible strains. Ocellatin-PT3 showed synergy with ciprofloxacin and ceftazidime against multidrug-resistant isolates and was capable of preventing the proliferation of 48-h mature biofilms at concentrations ranging from 4 to 8× the MIC. Treated biofilms had low viability and were slightly more disaggregated. Conclusion: Ocellatin-PT3 may be promising as a template for the development of novel antimicrobial peptides against P. aeruginosa.This work received financial support from the European Union (FEDER funds through COMPETE) and National Funds (FCT, Fundação para a Ciência e Tecnologia), under the Partnership Agreement PT2020 through project UID/QUI/50006/2013-POCI/01/0145/FEDER/007265 (LAQV/REQUIMTE) and from Programa Operacional Regional do Norte (ON.2 – O Novo Norte),under the Quadro de Referencia Estrat ˆ egico Nacional (QREN) and funded by Fundo Europeu de Desenvolvimento Regional ´ (Feder) through projects: NORTE-01-0145-FEDER-000024, NORTE-01-0145-FEDER-000011 and NORTE-07-0161-FEDER-000111.A Placido is grateful to FCT for financial support through the grant SFRH ´ /BD/97995/2013, POPH-QREN-Tipologia 4.1-Formação Avançada and Fundo Social Europeu and Ministerio da Ciência, Tecnologia e Ensino Superior. The authors have no other relevant ˆ affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript apart from those disclosed.info:eu-repo/semantics/publishedVersio

Disclosure of a promising lead to tackle complicated skin and skin structure infections: antimicrobial and antibiofilm actions of peptide PP4-3.1

Efficient antibiotics are being exhausted, which compromises the treatment of infections, including complicated skin and skin structure infections (cSSTI) often associated with multidrug resistant (MDR) bacteria, methicillin-resistant S. aureus (MRSA) being the most prevalent. Antimicrobial peptides (AMP) are being increasingly regarded as the new hope for the post-antibiotic era. Thus, future management of cSSTI may include use of peptides that, on the one hand, behave as AMP and, on the other, are able to promote fast and correct skin rebuilding. As such, we combined the well-known cosmeceutical pentapeptide-4 (PP4), devoid of antimicrobial action but possessing collagenesis-boosting properties, with the AMP 3.1, to afford the chimeric peptide PP4-3.1. We further produced its N-methyl imidazole derivative, MeIm-PP4-3.1. Both peptide-based constructs were evaluated in vitro against Gram-negative bacteria, Gram-positive bacteria, and Candida spp. fungi. Additionally, the antibiofilm activity, the toxicity to human keratinocytes, and the activity against S. aureus in simulated wound fluid (SWF) were assessed. The chimeric peptide PP4-3.1 stood out for its potent activity against Gram-positive and Gram-negative bacteria, including against MDR clinical isolates (0.8 ≤ MIC ≤ 5.7 µM), both in planktonic form and in biofilm matrix. The peptide was also active against three clinically relevant species of Candida fungi, with an overall performance superior to that of fluconazole. Altogether, data reveal that PP4-3.1 is as a promising lead for the future development of new topical treatments for severe skin infections.info:eu-repo/semantics/publishedVersio

Neofiscalin A and fiscalin C are potential novel indole alkaloid alternatives for the treatment of multidrug-resistant Gram-positive bacterial infections

Ten indole alkaloids were obtained from the marine sponge-associated fungus Neosartorya siamensis KUFA 0017. We studied the antimicrobial properties of these and of three other compounds previously isolated from the soil fungus N. siamensis KUFC 6349. Only neofiscalin A showed antimicrobial activity against Gram-positive bacteria, including methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant Enterococcus faecalis (VRE); with a minimum inhibitory concentration (MIC) of 8 μg mL(-1) against both strains. Another compound, fiscalin C, presented synergistic activity against MRSA when combined with oxacillin, although alone showed no antibacterial effect. Moreover, neofiscalin A, when present at sub-MICs, hampered the ability of both MRSA and VRE strains to form a biofilm. Additionally, the biofilm inhibitory concentration values of neofiscalin A against the MRSA and VRE isolates were 96 and 80 μg mL(-1), respectively. At a concentration of 200 μg mL(-1), neofiscalin A was able to reduce the metabolic activity of the biofilms by ∼50%. One important fact is that our results also showed that neofiscalin A had no cytotoxicity against a human brain capillary endothelial cell line.info:eu-repo/semantics/publishedVersio

Synthesis of novel sulfide-based cyclic peptidomimetic analogues to solonamides

Eight new sulfide-based cyclic peptidomimetic analogues of solonamides A and B have been synthesized via solid-phase peptide synthesis and SN2’ reaction on a Morita–Baylis–Hillman (MBH) residue introduced at the N-terminal of a tetrapeptide. This last step takes advantage of the electrophilic feature of the MBH residue and represents a new cyclization strategy occurring. The analogues were prepared in moderate overall yields and did not show toxic effects on Staphylococcus aureus growth and were not toxic to human fibroblasts. Two of them inhibited the hemolytic activity of S. aureus, suggesting an interfering action in the bacterial quorum sensing similar to the one already reported for solonamides

Neuroprotective effects on microglia and insights into the structure–activity relationship of an antioxidant peptide isolated from Pelophylax perezi

© 2022 The Authors. Journal of Cellular and Molecular Medicine published by Foundation for Cellular and Molecular Medicine and John Wiley & Sons Ltd. This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly citedTryptophyllins constitute a heterogeneous group of peptides that are one of the first classes of peptides identified from amphibian's skin secretions. Here, we report the structural characterization and antioxidant properties of a novel tryptophyllin-like peptide, named PpT-2, isolated from the Iberian green frog Pelophylax perezi. The skin secretion of P. perezi was obtained by electrical stimulation and fractionated using RP-HPLC. De novo peptide sequencing was conducted using MALDI MS/MS. The primary structure of PpT-2 (FPWLLS-NH2 ) was confirmed by Edman degradation and subsequently investigated using in silico tools. PpT-2 shared physicochemical properties with other well-known antioxidants. To test PpT-2 for antioxidant activity in vitro, the peptide was synthesized by solid phase and assessed in the chemical-based ABTS and DPPH scavenging assays. Then, a flow cytometry experiment was conducted to assess PpT-2 antioxidant activity in oxidatively challenged murine microglial cells. As predicted by the in silico analyses, PpT-2 scavenged free radicals in vitro and suppressed the generation of reactive species in PMA-stimulated BV-2 microglia cells. We further explored possible bioactivities of PpT-2 against prostate cancer cells and bacteria, against which the peptide exerted a moderate antiproliferative effect and negligible antimicrobial activity. The biocompatibility of PpT-2 was evaluated in cytotoxicity assays and in vivo toxicity with Galleria mellonella. No toxicity was detected in cells treated with up to 512 µg/ml and in G. mellonella treated with up to 40 mg/kg PpT-2. This novel peptide, PpT-2, stands as a promising peptide with potential therapeutic and biotechnological applications, mainly for the treatment/prevention of neurodegenerative disorders.This work was financed by FEDER - Fundo Europeu de Desenvolvimento Regional funds through the COMPETE 2020 - Operacional Programme for Competitiveness and Internationalization (POCI), and by Portuguese funds through FCT - Fundação para a Ciência e a Tecnologia in the framework of the project POCI-01-0145-FEDER-031158 – PTDC/BII-BIO/31158/2017. The authors would like to thank the participation and scientific support of the Unit projects UIDB/50006/2020 | UIDP/50006/2020, and the Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Universal Faixa ‘B’ (grant number 32103/2018-0). A.P. is a recipient of a post-doctoral grant from the project PTDC/BII-BIO/31158/2017. The authors would like to thank the researcher Roberto Resendes (CiBio, University of the Azores, Ponta Delgada, São Miguel, Azores, Portugal) for the logistical support in the collection of samples. C.P.A acknowledges FCT-MCTES fellowship PD/BD/136860/2018. A.B.-N. and F.C.D.A.L. acknowledge CNPq (grants 420449/2018-3 and 428211/2018-6) for financial support.info:eu-repo/semantics/publishedVersio