Generation Of Antimicrobials Research Articles

Combatting Antibiotic Resistance: Regulatory Exclusivities for New Antibiotic Development in the United States and Europe

Abstract Antimicrobial resistance is a global concern that has led to millions of deaths worldwide. Genetic changes occur naturally in pathogens such as viruses, bacteria, parasites, and fungi. To overcome these concerns, there is a need to develop a new generation of antimicrobials. In an effort to encourage innovation, both the United States (US) and the European Union have introduced exclusivity programs. The US offered 5 years of additional nonpatent exclusivity for Qualified Infectious Disease Products under the Generating Antibiotic Incentives Now Act. In parallel, the European Commission offers a patent extension of 12 months across Europe (EU) countries or the option to sell the vouchers to other pharmaceutical companies through transferable exclusivity extension vouchers. This review focuses on the most important innovative strategies of antibiotic development the drug approvals, their mode of action, spectra of activity, and targeting of microorganisms, in past 5 years were mentioned, and discusses how to overcome the challenges of drug approval under antibiotic exclusivity in the US and EU, which will benefit the companies to develop novel drugs in several nations.

Inhibition of DXR in the MEP pathway with lipophilic N-alkoxyaryl FR900098 analogs.

In Mycobacterium tuberculosis (Mtb) and Plasmodium falciparum (Pf), the methylerythritol phosphate (MEP) pathway is responsible for isoprene synthesis. This pathway and its products are vital to bacterial/parasitic metabolism and survival, and represent an attractive set of drug targets due to their essentiality in these pathogens but absence in humans. The second step in the MEP pathway is the conversion of 1-deoxy-d-xylulose-5-phosphate (DXP) to MEP and is catalyzed by 1-deoxy-d-xylulose-5-phosphate reductoisomerase (DXR). Natural products fosmidomycin and FR900098 inhibit DXR, but are too polar to reach the desired target inside some cells, such as Mtb. Synthesized FR900098 analogs with lipophilic substitution in the position α to the phosphorous atom showed promise, resulting in increased activity against Mtb and Pf. Here, an α substitution, consisting of a 3,4-dichlorophenyl substituent, in combination with various O-linked alkylaryl substituents on the hydroxamate moiety is utilized in the synthesis of a novel series of FR900098 analogs. The purpose of the O-linked alkylaryl substituents is to further enhance DXR inhibition by extending the structure into the adjacent NADPH binding pocket, blocking the binding of both DXP and NADPH. Of the initial O-linked alkylaryl substituted analogs, compound 6e showed most potent activity against Pf parasites at 3.60 μM. Additional compounds varying the phenyl ring of 6e were synthesized. The most potent phosphonic acids, 6l and 6n, display nM activity against PfDXR and low μM activity against Pf parasites. Prodrugs of these compounds were less effective against Pf parasites but showed modest activity against Mtb cells. Data from this series of compounds suggests that this combination of substituents can be advantageous in designing a new generation of antimicrobials.

Bacillus Strains Isolated from Korean Fermented Food Products with Antimicrobial Activity Against Staphylococci, an Alternative for Control of Antibiotic-Resistant Pathogens.

Multidrug resistance in foodborne and clinical pathogens is a worldwide health problem. The urgent need for new alternatives to the existing antibiotics is emerging. Bacteriocin-like inhibitory substances can be considered part of the new generation of antimicrobials, which can be potentially applied in the food industry and health care practices. This study aimed to select Bacillus strains with antimicrobial activity against Staphylococcus spp. with future application in the formulation of pharmaceutical antimicrobial preparations. Putative antimicrobial agent-producing strains, previously isolated and preidentified as Bacillus spp. were profiled by repetitive element sequence-based polymerase chain reaction (rep-PCR) and 16s rRNA sequencing identified the strains as Bacillus tequilensis ST1962CD with 99.47% identity confidence and as Bacillus subtilis subsp. stercoris ST2056CD with 98.45% identity confidence. Both the selected Bacillus strains were evaluated via biomolecular and physiological approaches related to their safety and virulence, beneficial properties, enzyme production profile, and presence of corresponding genes for the production of antimicrobials and virulence. Both strains were confirmed to harbor srfa and sbo genes and be free of hemolysin binding component (B) and two lytic components (L1 and L2) [BL] and nonhemolytic enterotoxin-associated genes. Produced antimicrobial agents by strains ST1962CD and ST2056CD were partially purified through the combination of ammonium sulfate precipitation and hydrophobic-based chromatography on SepPakC18 and evaluated regarding their cytotoxicity. The dynamics of bacterial growth, pH change, accumulation of produced antimicrobials, and the mode of action were evaluated. Obtained results were pointing to the potential application of safe B. tequilensis ST1962CD and B. subtilis subsp. stercoris ST2056CD strains as functional beneficial microbial cultures that are putative producers of surfactin and/or subtilosin, as potent antimicrobials, for the treatment of some staphylococcal-associated infections. Expressed antimicrobials were shown to be not cytotoxic, and appropriate biotechnological approaches need to be developed for cost-effective production, isolation, and purification of expressed antimicrobials by studied strains.

Inhibitory-like Substances Produced by Yeasts Isolated from Andean Blueberries: Prospective Food Antimicrobials.

Natural agents from microorganisms have emerged as suitable options to replace chemical preservatives in foods. In this study, the antibacterial activity of cell-free supernatant (CFS) from five native yeasts (Saccharomyces cerevisiae Lev6 and Lev30, C. pseudointermedia Lev8, Candida intermedia Lev9, C. parapsilosis Lev15) and the reference S. boulardi SSB, was evaluated against some indicator food pathogens. The generation of antimicrobials was reliant on strain-, and sugar-supplemented media, which supported yeast growth established at 30 °C and 200 rpm for 48 h. Treatment with proteinase K and catalase was unable to completely abolish the inhibitory effect, indicating that the active components are likely complex combinations of acids, proteins, hydrogen peroxide, and other metabolites. Although there was no impact on Listeria monocytogenes, exposure to CFS and extracellular fractions obtained through precipitation with methanol (PPm) at 120 °C for 60 min significantly (p < 0.05) increased the inhibitory activity against Escherichia coli, Salmonella enterica, Kosakonia cowanii, and Staphylococcus aureus, indicating that the inhibitory activity was stimulated by heat. Likewise, a synergistic inhibitory action against Listeria was obtained following the pretreatment of PPm with EDTA (ethylenediaminetetraacetic acid). These activities were yeast strain-dependent, with Lev6, Lev8, and Lev30 showing the highest activity. In addition, a heat-stable low-molecular-mass molecule under 5 kDa was detected in Lev30. Further research is required to evaluate the mode of action and characterize the composition of the released molecules in the CFS in order to develop a novel biocontrol agent based on yeasts.

Changing Trend in the Antibiotic Resistance Pattern of Klebsiella Pneumonia Isolated From Endotracheal Aspirate Samples of ICU Patients of a Tertiary Care Hospital in North India.

Introduction Klebsiella pneumonia is one of the most prevalent bacteria that cause nosocomial infections, particularly in critically ill patients in the intensive care unit (ICU). Multi-drug-resistant Klebsiella pneumoniae (MDRKP) has become an urgent risk to public health as its prevalence has sharply surged around the globe in recent decades. Therefore, this research was conducted to evaluate shifts over a four-year period in drug susceptibility patterns among Klebsiella pneumoniae isolates from mechanically ventilated intensive care unit patients. Materials and methods This is a retrospective observational study conducted in a tertiary care multi-specialty hospital and teaching institute in North India and was approved by the institutional ethics committee. The research comprised Klebsiella pneumoniae isolates from endotracheal aspirates (ETA) of patients on mechanical ventilation admitted to the general intensive care unit (ICU) of our tertiary care facility. The data from January to June 2018 and January to June 2022 were collected. According to the antimicrobial resistance profile of the strains, they were categorized as susceptible, resistant to one or two antimicrobial categories, multidrug-resistant (MDR), extensively drug-resistant (XDR), or pan-drug-resistant (PDR). The criteria for MDR, XDR, and PDR were proposed by the European Centre for Disease Prevention and Control (ECDC). IBM Statistical Package for the Social Sciences (SPSS) for Windows, Version 24.0, Armonk, NY, IBM Corp., was used for data input and analysis. Results A total of 82 cases of Klebsiella pneumonia were included in the study. Of these 82 isolates, 40 were isolated over a period of six months from January to June 2018, and the remaining 42 were isolated from January to June 2022. Among the 2018 group, five strains (12.5%) were classified as susceptible, three (7.5%) as resistant, seven (17.5%) as MDR, and 25 (62.5%) as XDR. The highest percentages of antimicrobial resistance in the 2018 group were observed with amoxicillin/clavulanic acid (90%), ciprofloxacin (100%), piperacillin/tazobactam (92.5%), and cefoperazone/sulbactam (95%). In comparison, the 2022 group showed no strain as susceptible; nine strains (21.4%) were classified as resistant; three strains (7%) as MDR; and 30 strains (93%) were classified as XDR. There was a significant increase in resistance to amoxicillin, from 10% in 2018 to nil in 2022. Overall, the rate of resistant Klebsiella pneumonia (K. pneumonia) increased from 7.5% (3/40) in 2018 to 21.4% (9/42) in 2022, while XDR Klebsiella pneumonia among the mechanically ventilated ICU patients significantly increased from 62.5% (25/40) in 2018 to 71% (30/42) in 2022. Conclusion K. pneumoniae antibiotic resistance is a real threat in Asia and requires close monitoring to be controlled. More careful attempts should be made to create a new generation of antimicrobials since the prevalence of resistance to existingmedications is rising. Antibiotic resistance should be monitored and reported by healthcare institutions regularly.

The antibacterial activity of LL-37 peptide against multidrug-resistant Pseudomonas aeruginosa isolated from burn infections

Multidrug-resistant Pseudomonas aeruginosa has emerged as a significant problem worldwide, posing a severe hazard to burn-infected patients. Antimicrobial peptides produced from humans or animals and synthetic peptides have received interest as antibiotic options for treating resistant bacteria, particularly those obtained from burn patients. The current work evaluated the role of antimicrobial peptide LL-37 as an antibacterial agent against multidrug P. aeruginosa isolates from burn infections. The study samples were collected between November 2021 and the end of February 2022 and included 157 clinical specimens as burn swabs from patients with burn infections admitted to four Baghdad hospitals in Baghdad, Iraq. The results of selective media, biochemical tests, and the ITEK2 system identified 39 isolates (24.8%) as p. aeruginosa from all collected bacterial cultures. The findings of the antimicrobial susceptibility test by disc diffusion method for the isolates under investigation revealed that P. aeruginosa clinical isolates were moderately resistant to antibiotics tested. Most P. aeruginosa isolates were highly resistant to Tetracycline (89.7%), Azithromycin (71.7%), and Amikacin, Cefepime, and Gentamycin. Also, the highest sensitivity was recorded for Ciprofloxacin, Piperacillin/tazobactam, C ceftazidime and Levofloxacin. The results of minimum inhibitory concentrations (MICs) of LL-37 against (8) multidrug-resistant P. aeruginosa isolates revealed that the concentration range of LL-37 was (15.6-1000 µg/ml), indicating that LL-37 has a significant effect on P. aeruginosa growth at low concentrations. In conclusion, t using the antimicrobial peptides LL-37 in treating life-threaded infections could lead to developing a new generation of antimicrobials that can overcome bacterial resistance mechanisms. Keywords: Antibacterial, Burns, LL-37, Pseudomonas aeruginosa

Synergistic effects of short peptides and antibiotics against bacterial and fungal strains.

There is a rising tide of concern about the antibiotic resistance issue. To reduce the possibility of antibiotic-resistant infections, a new generation of antimicrobials must be developed. Antimicrobial peptides are potential alternatives to antibiotics that can be used alone or together with conventional antibiotics to combat antimicrobial resistance. In this work, lead compounds LP-23, DP-23, SA4, and SPO from previously published studies were synthesized by solid-phase peptide synthesis and their antimicrobial evaluation was carried out against various bacterial and fungal strains. Peptide combinations with antibiotics were evaluated by using the checkerboard method and their minimal inhibitory concentration (MIC) in combination was calculated by using the fractional inhibitory concentration (FIC) index. Cytotoxicity evaluations of these peptides further confirmed their selectivity toward microbial cells. Based on the FIC values, LP-23, DP-23, and SPO demonstrated synergy in combination with gentamicin against a gentamicin-resistant clinical isolate of Escherichia coli. For Staphylococcus aureus, Escherichia coli, and Salmonella typhimurium, seven combinations exhibited synergistic effects between peptide/peptoids and the tested antibiotics. Additionally, almost all the combinations of peptides/peptoids with amphotericin B and fluconazole also showed effective synergy against Aspergillus niger and Aspergillus flavus. The synergy found between LP-23, DP-23, SA4, and SPO with the selected antibiotics may have the potential to be used as a combination therapy against various microbial infections.

Essential Oils: A Novel Approach for Anti-Microbial Therapy

Abstract: Overexploitation of antibiotics has led to significant challenges with antimicrobial resistance. The gravity of this trend has resulted in the rapid emergence of multi-resistant pathogens. Increased frequency to treat infections and the unfeasibility to reverse this resistance have burdened researchers in developing novel mechanisms to counteract and obstruct antimicrobial resistance. An effective medical strategy to control resistance was to develop new and potent antibiotics although, a sobering reality is that the product pipeline towards new antibiotics is inadequate. To ensure continued effective treatment of bacterial infections, there is an urgent need to control as well as conserve existing antibiotics through novel approaches. Utilization of Essential Oils (EO) in a multi-target anti-infective therapy addresses this need by targeting the mechanism of bacterial resistance and discovering synergism between EO’s antimicrobial properties to anti-infectives. When introduced into a pharmaceutical formulation, this novel and rational approach will open the door towards the development of a new generation of antimicrobials. Today, several essential oils have the ability to act as resistant modifying agents and enhance the antimicrobial activity of anti-infectives. This review article intends to focus on the effectiveness of essential oils on drug-resistant pathogens, mechanisms to counteract antimicrobial resistance, approaches to enhance efficacy, and explore potential applications.

Biological Dark Matter Exploration using Data Mining for the Discovery of Antimicrobial Natural Products.

The discovery of novel antimicrobials has significantly slowed down over the last three decades. At the same time, humans rely increasingly on antimicrobials because of the progressive antimicrobial resistance in medical practices, human communities, and the environment. Data mining is currently considered a promising option in the discovery of new antibiotics. Some of the advantages of data mining are the ability to predict chemical structures from sequence data, anticipation of the presence of novel metabolites, the understanding of gene evolution, and the corroboration of data from multiple omics technologies. This review analyzes the state-of-the-art for data mining in the fields of bacteria, fungi, and plant genomic data, as well as metabologenomics. It also summarizes some of the most recent research accomplishments in the field, all pinpointing to innovation through uncovering and implementing the next generation of antimicrobials.

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.

Antimicrobial Peptides from Black Soldier Fly (Hermetia illucens) as Potential Antimicrobial Factors Representing an Alternative to Antibiotics in Livestock Farming.

Simple SummaryMicrobial resistance to antibiotics is a constant threat to livestock farming, and unreasonable use of antibiotics has increased the prevalence of infectious diseases in humans and animals. Antimicrobial peptides derived from black soldier fly, Hermetia illucens (Diptera: Stratiomyidae), have great potential as alternatives to antibiotics for prophylaxis and treatment of diseases in animals because they have extensive antimicrobial properties and a lower tendency to induce resistance. Additionally, several studies have shown that Hermetia illucens larvae can participate in a circular economy by digesting organic waste alone and then promoting the growth performance of domestic animals fed the larvae. Therefore, antimicrobial peptides from Hermetia illucens are promising candidate for replacement of antibiotics in livestock farming.Functional antimicrobial peptides (AMPs) are an important class of effector molecules of innate host immune defense against pathogen invasion. Inability of microorganisms to develop resistance against the majority of AMPs has made them alternatives to antibiotics, contributing to the development of a new generation of antimicrobials. Due to extensive biodiversity, insects are one of the most abundant sources of novel AMPs. Notably, black soldier fly insect (BSF; Hermetia illucens (Diptera: Stratiomyidae)) feeds on decaying substrates and displays a supernormal capacity to survive under adverse conditions in the presence of abundant microorganisms, therefore, BSF is one of the most promising sources for identification of AMPs. However, discovery, functional investigation, and drug development to replace antibiotics with AMPs from Hermetia illucens remain in a preliminary stage. In this review, we provide general information on currently verified AMPs of Hermetia illucens, describe their potential medical value, discuss the mechanism of their synthesis and interactions, and consider the development of bacterial resistance to AMPs in comparison with antibiotics, aiming to provide a candidate for substitution of antibiotics in livestock farming or, to some extent, for blocking the horizontal transfer of resistance genes in the environment, which is beneficial to human and animal welfare.

Spotlight on the Selected New Antimicrobial Innate Immune Peptides Discovered During 2015-2019

Antibiotic resistance is a global issue and new anti-microbials are required. Anti-microbial peptides are important players of host innate immune systems that prevent infections. Due to their ability to eliminate drug-resistant pathogens, AMPs are promising candidates for developing the next generation of anti-microbials. The anti-microbial peptide database provides a useful tool for searching, predicting, and designing new AMPs. In the period from 2015-2019, ~500 new natural peptides have been registered. This article highlights a selected set of new AMP members with interesting properties. Teixobactin is a cell wall inhibiting peptide antibiotic, while darobactin inhibits a chaperone and translocator for outer membrane proteins. Remarkably, cOB1, a sex pheromone from commensal enterococci, restricts the growth of multidrug-resistant Enterococcus faecalis in the gut at a picomolar concentration. A novel proline-rich AMP has been found in the plant Brassica napus. A shrimp peptide MjPen- II comprises three different sequence domains: serine-rich, proline-rich, and cysteine-rich regions. Surprisingly, an amphibian peptide urumin specifically inhibits H1 hemagglutinin-bearing influenza A virus. Defensins are abundant and typically consist of three pairs of intramolecular disulfide bonds. However, rat rattusin dimerizes via forming five pairs of intermolecular disulfide bonds. While human LL-37 can be induced by vitamin D, vitamin A induces the expression of resistin-like molecule alpha (RELMα) in mice. The isolation and characterization of an alternative human cathelicidin peptide, TLN-58, substantiates the concept of one gene multiple peptides. The involvement of a fly AMP nemuri in sleep induction may promote the research on the relationship between sleep and infection control. The functional roles of AMPs continue to grow and the general term "innate immune peptides" becomes useful. These discoveries widen our view on the anti-microbial peptides and may open new opportunities for developing novel peptide therapeutics for different applications.

Sisters in structure but different in character, some benzaldehyde and cinnamaldehyde derivatives differentially tune Aspergillus flavus secondary metabolism

Great are the expectations for a new generation of antimicrobials, and strenuous are the research efforts towards the exploration of diverse molecular scaffolds—possibly of natural origin – aimed at the synthesis of new compounds against the spread of hazardous fungi. Also high but winding are the paths leading to the definition of biological targets specifically fitting the drug’s structural characteristics. The present study is addressed to inspect differential biological behaviours of cinnamaldehyde and benzaldehyde thiosemicarbazone scaffolds, exploiting the secondary metabolism of the mycotoxigenic phytopathogen Aspergillus flavus. Interestingly, owing to modifications on the parent chemical scaffold, some thiosemicarbazones displayed an increased specificity against one or more developmental processes (conidia germination, aflatoxin biosynthesis, sclerotia production) of A. flavus biology. Through the comparative analysis of results, the ligand-based screening strategy here described has allowed us to delineate which modifications are more promising for distinct purposes: from the control of mycotoxins contamination in food and feed commodities, to the environmental management of microbial pathogens, to the investigation of specific structure–activity features for new generation drug discovery.

Bacteriocins as a new generation of antimicrobials: toxicity aspects and regulations.

ABSTRACTIn recent decades, bacteriocins have received substantial attention as antimicrobial compounds. Although bacteriocins have been predominantly exploited as food preservatives, they are now receiving increased attention as potential clinical antimicrobials and as possible immune-modulating agents. Infections caused by antibiotic-resistant bacteria have been declared as a global threat to public health. Bacteriocins represent a potential solution to this worldwide threat due to their broad- or narrow-spectrum activity against antibiotic-resistant bacteria. Notably, despite their role in food safety as natural alternatives to chemical preservatives, nisin remains the only bacteriocin legally approved by regulatory agencies as a food preservative. Moreover, insufficient data on the safety and toxicity of bacteriocins represent a barrier against the more widespread use of bacteriocins by the food and medical industry. Here, we focus on the most recent trends relating to the application of bacteriocins, their toxicity and impacts.

In the Age of Synthetic Biology, Will Antimicrobial Peptides be the Next Generation of Antibiotics?

Antibiotics have changed human health and revolutionised medical practice since the Second World War. Today, the use of antibiotics is increasingly limited by the rise of antimicrobial-resistant strains. Additionally, broad-spectrum antibiotic activity is not adapted to maintaining a balanced microbiome essential for human health. Targeted antimicrobials could overcome these two drawbacks. Although the rational design of targeted antimicrobial molecules presents a formidable challenge, in nature, targeted genetically encoded killing molecules are used by microbes in their natural ecosystems. The use of a synthetic biology approach allows the harnessing of these natural functions. In this commentary article we illustrate the potential of applying synthetic biology towards bacteriocins to design a new generation of antimicrobials.

Developing high through put screening strategies to identify the next generation of antimicrobials

Area of expertise: My research interests focus on identifying novel compounds that can disrupt key phenotypes associated with antibiotic resistance such as biofilm formation and motility in the pathogens Acinetobacter baumannii and Pseudomonas aeruginosa. To this end we have developed a range of screens and assays to facilitate compound identification and mechanism of action profiling in these pathogens. These include: Antibiotic Resistance Inhibitor Screens, Biofilm Inhibition Screens, Biofilm Disruption Screens, Signalling Pathway Specific Biosensor Screens , Mechanism of Action Assays, Resistance Development Screens, Antibiotic Resistance Inhibitor Screens, In vivo biofilm and pathogenicity models to validate lead compounds and perform toxicity profiling. Future goals: Future goals include being part of a collaborative multidisciplinary drug discovery network that has access to diverse and bespoke extract/compound collections and the expertise to identify the bioactive leads, characterise their mechanism of action and ultimately take them forward to clinical trials. Further to this the goal is to target national and international funding opportunities that will facilitate taking active compounds right through the drug discovery pipeline from the bench to the bedside. I am hoping to identify collaborators across all aspects of drugs discovery, particularly collaborators with: Access to compound or extract collections, Expertise in compound identification ,Clinical trial expertise.

Synergy Between Two Chimeric Lysins to Kill Streptococcus pneumoniae.

Phage lysins constitute a new generation of antimicrobials that are becoming a promising alternative and complementation to current antibiotic therapies, which are nowadays called into question by the increasing numbers of multiresistant bacteria. Streptococcus pneumoniae is a leading human pathogen causing serious infectious diseases in children and adults. Within the host-parasite interplay system of pneumococcus and its phages, several antipneumococcal lysins have been described and, among them, chimeric lysins Cpl-711 and PL3 stand out for their potent bactericidal activities. Here, evidence is presented on the synergistic cooperation of the catalytically diverse lysins Cpl-711 and PL3 in different assays, like purified cell wall enzymatic degradation, in vitro bacterial cell growth inhibition, and killing of both planktonic and biofilm grown cells. Synergy between Cpl-711 and PL3 has been shown to reduce the amount of enzyme necessary to inhibit growth in checkerboard assays with a sum of fractional inhibitory concentrations ≤0.5 for all pneumococcal strains tested, while also significatively increasing bactericidal effect by ≥2 logs with respect to the sum of activities of Cpl-711 and PL3 individual treatments. Moreover, the combination of these two lysins showed synergy in an adult zebrafish model of pneumococcal infection. This study consolidates the possibility of formulating highly efficient and synergistic antibacterial enzymes that could improve our ability to fight multiresistant bacterial infections.

DPD-Inspired Discovery of Novel LsrK Kinase Inhibitors: An Opportunity To Fight Antimicrobial Resistance.

Antibiotic resistance is posing a continuous threat to global public health and represents a huge burden for society as a whole. In the past decade, the interference with bacterial quorum sensing (QS) (i.e., cell-cell communication) mechanisms has extensively been investigated as a valid therapeutic approach in the pursuit of a next generation of antimicrobials. ( S)-4,5-Dihydroxy-2,3-pentanedione, commonly known as ( S)-DPD, a small signaling molecule that modulates QS in both Gram-negative and Gram-positive bacteria, is phosphorylated by LsrK, and the resulting phospho-DPD activates QS. We designed and prepared a small library of DPD derivatives, characterized by five different scaffolds, and evaluated their LsrK inhibition in the context of QS interference. SAR studies highlighted the pyrazole moiety as an essential structural element for LsrK inhibition. Particularly, four compounds were found to be micromolar LsrK inhibitors (IC50 ranging between 100 μM and 500 μM) encouraging further exploration of novel analogues as potential new antimicrobials.

Phage Lysins for Fighting Bacterial Respiratory Infections: A New Generation of Antimicrobials.

Lower respiratory tract infections and tuberculosis are responsible for the death of about 4.5 million people each year and are the main causes of mortality in children under 5 years of age. Streptococcus pneumoniae is the most common bacterial pathogen associated with severe pneumonia, although other Gram-positive and Gram-negative bacteria are involved in respiratory infections as well. The ability of these pathogens to persist and produce infection under the appropriate conditions is also associated with their capacity to form biofilms in the respiratory mucous membranes. Adding to the difficulty of treating biofilm-forming bacteria with antibiotics, many of these strains are becoming multidrug resistant, and thus the alternative therapeutics available for combating this kind of infections are rapidly depleting. Given these concerns, it is urgent to consider other unconventional strategies and, in this regard, phage lysins represent an attractive resource to circumvent some of the current issues in infection treatment. When added exogenously, lysins break specific bonds of the peptidoglycan and have potent bactericidal effects against susceptible bacteria. These enzymes possess interesting features, including that they do not trigger an adverse immune response and raise of resistance is very unlikely. Although Gram-negative bacteria had been considered refractory to these compounds, strategies to overcome this drawback have been developed recently. In this review we describe the most relevant in vitro and in vivo results obtained to date with lysins against bacterial respiratory pathogens.

A Versatile Strategy for the Synthesis of 4,5-Dihydroxy-2,3-Pentanedione (DPD) and Related Compounds as Potential Modulators of Bacterial Quorum Sensing.

Resistance to antibiotics is an increasingly serious threat to global public health and its management translates to significant health care costs. The validation of new Gram-negative antibacterial targets as sources for potential new antibiotics remains a challenge for all the scientists working in this field. The interference with bacterial Quorum Sensing (QS) mechanisms represents a potentially interesting approach to control bacterial growth and pursue the next generation of antimicrobials. In this context, our research is focused on the discovery of novel compounds structurally related to (S)-4,5-dihydroxy-2,3-pentanedione, commonly known as (S)-DPD, a small signaling molecule able to modulate bacterial QS in both Gram-negative and Gram-positive bacteria. In this study, a practical and versatile synthesis of racemic DPD is presented. Compared to previously reported syntheses, the proposed strategy is short and robust: it requires only one purification step and avoids the use of expensive or hazardous starting materials as well as the use of specific equipment. It is therefore well suited to the synthesis of derivatives for pharmaceutical research, as demonstrated by four series of novel DPD-related compounds described herein.