Proteome partitioning constraints in long-term laboratory evolution.

Adaptive laboratory evolution experiments provide a controlled context in which the dynamics of selection and adaptation can be followed in real-time at the single-nucleotide level. And yet this precision introduces hundreds of degrees-of-freedom as genetic changes accrue in parallel lineages over generations. On short timescales, physiological constraints have been leveraged to provide a coarse-grained view of bacterial gene expression characterized by a small set of phenomenological parameters.

PlyKp104, a Novel Phage Lysin for the Treatment of Klebsiella pneumoniae, Pseudomonas aeruginosa, and Other Gram-Negative ESKAPE Pathogens.

Klebsiella pneumoniae and Pseudomonas aeruginosa are two leading causes of burn and wound infections, pneumonia, urinary tract infections, and more severe invasive diseases, which are often multidrug resistant (MDR) or extensively drug resistant. Due to this, it is critical to discover alternative antimicrobials, such as bacteriophage lysins, against these pathogens. Unfortunately, most lysins that target Gram-negative bacteria require additional modifications or outer membrane permeabilizing agents to be bactericidal.

PaP1, a Broad-Spectrum Lysin-Derived Cationic Peptide to Treat Polymicrobial Skin Infections.

Most skin infections, including those complicating burns, are polymicrobial involving multiple causative bacteria. Add to this the fact that many of these organisms may be antibiotic-resistant, and a simple skin lesion or burn could soon become life-threatening. Membrane-acting cationic peptides from Gram-negative bacteriophage lysins can potentially aid in addressing the urgent need for alternative therapeutics.

On the design principles of metabolic flux sensing.

Metabolism is precisely coordinated, with the goal of balancing fluxes to maintain robust growth. However, coordinating fluxes requires information about rates, which can only be inferred through concentrations. While flux-sensitive metabolites have been reported, the design principles underlying such sensing have not been clearly elucidated.

Prophage integration into CRISPR loci enables evasion of antiviral immunity in Streptococcus pyogenes.

CRISPR loci are composed of short DNA repeats separated by sequences, known as spacers, that match the genomes of invaders such as phages and plasmids. Spacers are transcribed and processed to generate RNA guides used by CRISPR-associated nucleases to recognize and destroy the complementary nucleic acids of invaders. To counteract this defence, phages can produce small proteins that inhibit these nucleases, termed anti-CRISPRs (Acrs).

Macrophage immunomodulation through new polymers that recapitulate functional effects of itaconate as a power house of innate immunity.

Itaconate (ITA) is an emerging powerhouse of innate immunity with therapeutic potential that is limited in its ability to be administered in a soluble form. We developed a library of polyester materials that incorporate ITA into polymer backbones resulting in materials with inherent immunoregulatory behavior. Harnessing hydrolytic degradation release from polyester backbones, ITA polymers resulted in the mechanism specific immunoregulatory properties on macrophage polarization in vitro.

A natural single-guide RNA repurposes Cas9 to autoregulate CRISPR-Cas expression.

CRISPR-Cas systems provide prokaryotes with acquired immunity against viruses and plasmids, but how these systems are regulated to prevent autoimmunity is poorly understood. Here, we show that in the S. pyogenes CRISPR-Cas system, a long-form transactivating CRISPR RNA (tracr-L) folds into a natural single guide that directs Cas9 to transcriptionally repress its own promoter (P).

Evaluation of Topical Lysostaphin as a Novel Treatment for Instrumented Rhesus Macaques () Infected with Methicillin-Resistant .

Lytic enzymes are novel antimicrobial agents that degrade bacterial cell walls, resulting in cell rupture and death. We tested one enzyme, the bacteriocin lysostaphin, for treatment of nonhuman primates () with persistent methicillinresistant (MRSA) infection of their cranial implant margins. The goal of this study was to determine if topical lysostaphin, either alone or as an adjunct therapy, could eliminate MRSA.

Gram-Negative Bacterial Lysins.

Antibiotics have had a profound impact on human society by enabling the eradication of otherwise deadly infections. Unfortunately, antibiotic use and overuse has led to the rapid spread of acquired antibiotic resistance, creating a major threat to public health. Novel therapeutic agents called bacteriophage endolysins (lysins) provide a solution to the worldwide epidemic of antibiotic resistance.

Isolation of Phage Lysins That Effectively Kill Pseudomonas aeruginosa in Mouse Models of Lung and Skin Infection.

Multidrug resistance (MDR) is rapidly increasing in prevalence among isolates of the opportunistic pathogen , leaving few treatment options. Phage lysins are cell wall hydrolases that have a demonstrated therapeutic potential against Gram-positive pathogens; however, the outer membrane of Gram-negative bacteria prevents most lysins from reaching the peptidoglycan, making them less effective as therapeutics. Nevertheless, a few lysins from Gram-negative bacterial phage can penetrate the bacterial outer membrane with the aid of an amphipathic tail found in the molecule's termini.

Lysocins: Bioengineered Antimicrobials That Deliver Lysins across the Outer Membrane of Gram-Negative Bacteria.

The prevalence of multidrug-resistant has stimulated development of alternative therapeutics. Bacteriophage peptidoglycan hydrolases, termed lysins, represent an emerging antimicrobial option for targeting Gram-positive bacteria. However, lysins against Gram-negatives are generally deterred by the outer membrane and their inability to work in serum.

Staphylococcus aureus Infecting and Colonizing Experimental Animals, Macaques, in a Research Animal Facility.

An outbreak of methicillin-resistant Staphylococcus aureus (MRSA) infections on the skin and soft tissues of experimental macaques in the vivarium of The Rockefeller University, New York, triggered this observational and interventional study. We screened 14 macaques in the colony (samples from head, nares, and rectum) and their housing (40 environmental surfaces) four times in 1 year, for S. aureus colonization or contamination, while implementing enhanced decolonization and decontamination procedures.

Strains of bacterial species induce a greatly varied acute adaptive immune response: The contribution of the accessory genome.

A fundamental question in human susceptibility to bacterial infections is to what extent variability is a function of differences in the pathogen species or in individual humans. To focus on the pathogen species, we compared in the same individual the human adaptive T and B cell immune response to multiple strains of two major human pathogens, Staphylococcus aureus and Streptococcus pyogenes. We found wide variability in the acute adaptive immune response induced by various strains of a species, with a unique combination of activation within the two arms of the adaptive response.

Prevalence and Mode of Inheritance of the Dal Blood Group in Dogs in North America.

Background: The Dal blood group system was identified a decade ago by the accidental sensitization of a Dal- Dalmatian with a Dal+ blood transfusion. Similar Dal-related blood incompatibilities have been suspected in other Dalmatians, Doberman Pinschers, and other breeds.

Objectives: To determine the prevalence and mode of inheritance of the Dal antigen expression in dogs.

The Phage Lysin PlySs2 Decolonizes Streptococcus suis from Murine Intranasal Mucosa.

Streptococcus suis infects pigs worldwide and may be zoonotically transmitted to humans with a mortality rate of up to 20%. S. suis has been shown to develop in vitro resistance to the two leading drugs of choice, penicillin and gentamicin.

Survey of Two New (Kai 1 and Kai 2) and Other Blood Groups in Dogs of North America.

Background: Based upon serology, >10 canine blood group systems have been reported.

Objective: We surveyed dogs for dog erythrocyte antigen (DEA) 1 and 2 new blood types (Kai 1 and Kai 2), and some samples also were screened for Dal and DEA 3, 4, and 7.

Methods: Blood samples provided by owners, breeders, animal blood banks, and clinical laboratories were typed for DEA 1 by an immunochromatographic strip technique with a monoclonal antibody and analysis of band intensity.

Effects of BMS-986094, a Guanosine Nucleotide Analogue, on Mitochondrial DNA Synthesis and Function.

BMS-986094, the prodrug of a guanosine nucleotide analogue (2'-C-methylguanosine), was withdrawn from clinical trials due to serious safety issues. Nonclinical investigative studies were conducted as a follow up to evaluate the potential for BMS-986094-related mitochondrial-toxicity. In vitro, BMS-986094 was applied to human hepatoma cells (HepG2 and Huh-7) or cardiomyocytes (hiPSCM) up to 19 days to assess mitochondrial DNA content and specific gene expression.

Xenotransfusion of anemic cats with blood compatibility issues: pre- and posttransfusion laboratory diagnostic and crossmatching studies.

Background: Finding compatible feline blood donors can be challenging. Canine blood has been occasionally used when compatible feline blood was not available in emergency situations.

Objectives: The study goals were to describe the effects of xenotransfusion in 2 anemic cats receiving canine blood because of discordant blood types and acute transfusion reaction, respectively, and to report in vitro heterotyping and crossmatching results between canine and feline blood samples.

Novel Engineered Peptides of a Phage Lysin as Effective Antimicrobials against Multidrug-Resistant Acinetobacter baumannii.

Acinetobacter baumannii is a Gram-negative bacterial pathogen responsible for a range of nosocomial infections. The recent rise and spread of multidrug-resistant A. baumannii clones has fueled a search for alternative therapies, including bacteriophage endolysins with potent antibacterial activities.

Targeted Curing of All Lysogenic Bacteriophage from Streptococcus pyogenes Using a Novel Counter-selection Technique.

Streptococcus pyogenes is a human commensal and a bacterial pathogen responsible for a wide variety of human diseases differing in symptoms, severity, and tissue tropism. The completed genome sequences of >37 strains of S. pyogenes, representing diverse disease-causing serotypes, have been published.

Elimination of Chromosomal Island SpyCIM1 from Streptococcus pyogenes Strain SF370 Reverses the Mutator Phenotype and Alters Global Transcription.

Streptococcus pyogenes chromosomal island M1 (SpyCIM1) integrates by site-specific recombination into the 5' end of DNA mismatch repair (MMR) gene mutL in strain SF370SmR, blocking transcription of it and the downstream operon genes. During exponential growth, SpyCIM1 excises from the chromosome and replicates as an episome, restoring mutL transcription. This process is reversed in stationary phase with SpyCIM1 re-integrating into mutL, returning the cells to a mutator phenotype.

Prediction of Stable Globular Proteins Using Negative Design with Non-native Backbone Ensembles.

Accurate predictions of protein stability have great potential to accelerate progress in computational protein design, yet the correlation of predicted and experimentally determined stabilities remains a significant challenge. To address this problem, we have developed a computational framework based on negative multistate design in which sequence energy is evaluated in the context of both native and non-native backbone ensembles. This framework was validated experimentally with the design of ten variants of streptococcal protein G domain β1 that retained the wild-type fold, and showed a very strong correlation between predicted and experimental stabilities (R(2) = 0.