The New SH3b_T Domain Increases the Structural and Functional Variability Among SH3b-Like CBDs from Staphylococcal Phage Endolysins.

Endolysins, proteins encoded by phages to lyse their hosts and release their progeny, have evolved to adapt to the structural features of each host. The endolysins from Staphylococcus-infecting phages typically feature complex architectures with two enzymatically active domains (EADs) and one cell wall-binding domain (CBD) belonging to the bacterial SH3 (SH3b) superfamily. This study focuses on three SH3b-like CBDs from representative staphylococcal phage endolysins (LysRODI, LysC1C and LysIPLA5) that were structurally and functionally characterized.

You get what you test for: The killing effect of phage lysins is highly dependent on buffer tonicity and ionic strength.

The phage lysin field has done nothing but grow in the last decades. As a result, many different research groups around the world are contributing to the field, often with certain methodological differences that pose a challenge to the interpretation and comparison of results. In this work, we present the case study of three Acinetobacter baumannii-targeting phage lysins (wild-type endolysin LysMK34 plus engineered lysins eLysMK34 and 1D10) plus one lysin with broad activity against Gram-positive bacteria (PlySs2) to provide exemplary evidence on the risks of generalization when using one of the most common lysin evaluation assays: the killing assay with resting cells.

Distinct mode of action of a highly stable, engineered phage lysin killing Gram-negative bacteria.

Engineered lysins are considered as highly promising alternatives for antibiotics. Our previous screening study using VersaTile technology identified 1D10 as a possible lead compound with activity against strains under elevated human serum concentrations. In this manuscript, we reveal an unexpected mode of action and exceptional thermoresistance for lysin 1D10.

BRCA1 frameshift variants leading to extended incorrect protein C termini.

Carriers of BRCA1 germline pathogenic variants are at substantially higher risk of developing breast and ovarian cancer than the general population. Accurate identification of at-risk individuals is crucial for risk stratification and the implementation of targeted preventive and therapeutic interventions. Despite significant progress in variant classification efforts, a sizable portion of reported BRCA1 variants remain as variants of uncertain clinical significance (VUSs).

A Bioluminescence-Based Ex Vivo Burn Wound Model for Real-Time Assessment of Novel Phage-Inspired Enzybiotics.

The silent pandemic of antibiotic resistance is thriving, prompting the urgent need for the development of new antibacterial drugs. However, within the preclinical pipeline, in vitro screening conditions can differ significantly from the final in vivo settings. To bridge the gap between in vitro and in vivo assays, we developed a pig-skin-based bioluminescent ex vivo burn wound infection model, enabling real-time assessment of antibacterials in a longitudinal, non-destructive manner.

Screening method for Enzyme-based liquefaction of corn stover pellets at high solids.

Liquefaction of high solid loadings of unpretreated corn stover pellets has been demonstrated with rheology of the resulting slurries enabling mixing and movement within biorefinery bioreactors. However, some forms of pelleted stover do not readily liquefy, so it is important to screen out lots of unsuitable pellets before processing is initiated. This work reports a laboratory assay that rapidly assesses whether pellets have the potential for enzyme-based liquefaction at high solids loadings.

The Specific Capsule Depolymerase of Phage PMK34 Sensitizes to Serum Killing.

The rising antimicrobial resistance is particularly alarming for , calling for the discovery and evaluation of alternatives to treat infections. Some bacteriophages produce a structural protein that depolymerizes capsular exopolysaccharide. Such purified depolymerases are considered as novel antivirulence compounds.

Essential Topics for the Regulatory Consideration of Phages as Clinically Valuable Therapeutic Agents: A Perspective from Spain.

Antibiotic resistance is one of the major challenges that humankind shall face in the short term. (Bacterio)phage therapy is a valuable therapeutic alternative to antibiotics and, although the concept is almost as old as the discovery of phages, its wide application was hindered in the West by the discovery and development of antibiotics in the mid-twentieth century. However, research on phage therapy is currently experiencing a renaissance due to the antimicrobial resistance problem.

A new model for the formation of an Enterococcus faecalis endodontic biofilm with nutritional restriction.

An in vitro model for the formation of an Enterococcus faecalis endodontic biofilm under nutritional restriction was established, simulating clinical conditions for the evaluation of antimicrobial substances. Biofilm formation in dentin was standardized using root quarters incubated with E. faecalis ATCC 29212 at 37°C without nutritional changes.

Engineering a Lysin with Intrinsic Antibacterial Activity (LysMK34) by Cecropin A Fusion Enhances Its Antibacterial Properties against Acinetobacter baumannii.

Bacteriophage-encoded lysins are increasingly reported as alternatives to combat Acinetobacter baumannii infections, for which limited therapeutic options are available. Some lysins, such as LysMK34, have a C-terminal amphipathic helix allowing them to penetrate the otherwise-impermeable outer membrane barrier. Another approach to kill Gram-negative pathogens with lysins relies on fusion of a peptide with outer membrane-permeabilizing properties to the lysin.

Design and Selection of Engineered Lytic Proteins With Decolonizing Activity.

causes various infections in humans and animals, the skin being the principal reservoir of this pathogen. The widespread occurrence of methicillin-resistant (MRSA) limits the elimination and treatment of this pathogen. Phage lytic proteins have been proven as efficient antimicrobials against .

New strategy for liquefying corn stover pellets.

The movement of solid material into and between unit operations within a biorefinery is a bottleneck in reaching design capacity, with formation of biomass slurries needed to introduce feedstock. Corn stover slurries have been achieved from dilute acid, pretreated materials resulting in slurry concentrations of up to about 150 g/L, above which flowability is compromised. We report a new strategy to liquefy corn stover at higher solids concentration (300 g/L) by initially cooking it with the enzyme mimetic maleic acid at 40 mM and 150 °C.

Poppers-Induced Methemoglobinemia: A Curious Case of the Blues.

Methemoglobinemia (MetHb) can be a deadly condition at certain levels, presenting in a fulminant form of cyanosis or disguising itself with vague symptoms. Methemoglobinemia is an altered state of the body's hemoglobin, which can be congenital or acquired. We report a case of a 62-year-old male who presented with altered mental status and hypoxia after consuming "Jungle Juice", raising concern for methemoglobinemia.

Synergistic action of phage phiIPLA-RODI and lytic protein CHAPSH3b: a combination strategy to target Staphylococcus aureus biofilms.

Staphylococcus aureus is considered a priority pathogen due to its increasing acquisition of antibiotic resistance determinants. Additionally, this microbe has the ability to form recalcitrant biofilms on different biotic and inert surfaces. In this context, bacteriophages and their derived lytic proteins may be a forward-looking strategy to help combat staphylococcal biofilms.

Cellular strategies against metal exposure and metal localization patterns linked to phosphorus pathways in Ochrobactrum anthropi DE2010.

Cytotoxic, chemical, biochemical, compositional, and morphometric responses were analyzed against heavy metal exposure in Ochrobactrum anthropi DE2010, an heterotrophic bacterium isolated from Ebro Delta microbial mats (Tarragona, NE Spain). Several parameters of effect and exposure were evaluated to determine tolerance to a range of cadmium (Cd), lead (Pb(II)), copper (Cu(II)), chromium (Cr(III)), and zinc (Zn) concentrations. Additionally, removal efficiency, polyphosphate production and metal localization patterns were also analyzed.

Lysins breaking down the walls of Gram-negative bacteria, no longer a no-go.

Phage lysins are one of the most advanced classes of antibacterials under clinical evaluation and have a new mode of action based on peptidoglycan degradation. Lysins were initially excluded from use against Gram-negative pathogens because of their impermeable outer membrane, but are now increasingly developed as effective antibacterials against these critical priority pathogens. Generally, three routes of investigation have been recently explored and advanced to different extents, including the use of lysins that possess intrinsic activity due to a positively charged C-terminus that destabilizes the outer membrane, the use of physical or chemical means to disrupt the outer membrane integrity and protein engineering to equip the lysin with the necessary tools to overcome the outer membrane.

Environmental pH is a key modulator of Staphylococcus aureus biofilm development under predation by the virulent phage phiIPLA-RODI.

Previous work had shown that, in some Staphylococcus aureus strains, low concentrations of the virulent phage vB_SauM_phiIPLA-RODI (phiIPLA-RODI) promoted the formation of DNA-rich biofilms, whose cells exhibited significant transcriptional differences compared to an uninfected control. This study aimed to dissect the sequence of events leading to these changes. Analysis of phage propagation throughout biofilm development revealed that the number of phage particles increased steadily up to a certain point and then declined.

Advanced engineering of third-generation lysins and formulation strategies for clinical applications.

One of the possible solutions for the current antibiotic resistance crisis may be found in (often bacteriophage-derived) peptidoglycan hydrolases. The first clinical trials of these natural enzymes, coined here as first-generation lysins, are currently ongoing. Moving beyond natural endolysins with protein engineering established the second generation of lysins.

Characterization of Clinical MRSA Isolates from Northern Spain and Assessment of Their Susceptibility to Phage-Derived Antimicrobials.

Methicillin-resistant (MRSA) is a prevalent nosocomial pathogen, causing a wide range of diseases. The increased frequency of MRSA isolates in hospitals and the emergence of vancomycin resistance have sparked the search for new control strategies. This study aimed to characterize sixty-seven MRSA isolates collected from both infected patients and asymptomatic carriers in a Spanish hospital.

Lysin LysMK34 of Bacteriophage PMK34 Has a Turgor Pressure-Dependent Intrinsic Antibacterial Activity and Reverts Colistin Resistance.

The prevalence of extensively and pandrug-resistant strains of leaves little or no therapeutic options for treatment for this bacterial pathogen. Bacteriophages and their lysins represent attractive alternative antibacterial strategies in this regard. We used the extensively drug-resistant strain MK34 to isolate the bacteriophage PMK34 (vB_AbaP_PMK34).

Encapsulation of the Antistaphylococcal Endolysin LysRODI in pH-Sensitive Liposomes.

Phage lysins are promising new therapeutics against multidrug-resistant bacteria. These so-called enzybiotics offer, amongst their most notable advantages, high target specificity and low resistance development. Moreover, there are numerous recent and ongoing studies aimed at demonstrating the efficacy and safety of endolysins in animal models or even in clinical trials.

Phage Lytic Protein LysRODI Prevents Staphylococcal Mastitis in Mice.

Phage lytic proteins are promising antimicrobials that could complement conventional antibiotics and help to combat multi-drug resistant bacteria that cause important human and animal infections. Here, we report the characterization of endolysin LysRODI (encoded by staphylophage phiIPLA-RODI) and its application as a prophylactic mastitis treatment. The main properties of LysRODI were compared with those of endolysin LysA72 (encoded by staphylophage phiIPLA35) and the chimeric protein CHAPSH3b (derived from the virion-associated peptidoglycan hydrolase HydH5 and lysostaphin).

Methicillin-Resistant Staphylococcus aureus in Hospitals: Latest Trends and Treatments Based on Bacteriophages.

Even though antibiotic resistance in bacteria is a natural phenomenon, the alarming increase in pathogenic bacteria refractory to a wide range of antimicrobials is attracting attention worldwide. Indeed, the World Health Organization (WHO) has recently published a list of priority pathogens for which new antimicrobial alternatives are urgently needed. Among these pathogens, methicillin-resistant (MRSA) strains are perhaps the best known by the general public.