Bacteriophage targeting of gut bacterium attenuates alcoholic liver disease.

Chronic liver disease due to alcohol-use disorder contributes markedly to the global burden of disease and mortality1-3. Alcoholic hepatitis is a severe and life-threatening form of alcohol-associated liver disease. The gut microbiota promotes ethanol-induced liver disease in mice4, but little is known about the microbial factors that are responsible for this process. Here we identify cytolysin-a two-subunit exotoxin that is secreted by Enterococcus faecalis5,6-as a cause of hepatocyte death and liver injury. Compared with non-alcoholic individuals or patients with alcohol-use disorder, patients with alcoholic hepatitis have increased faecal numbers of E. faecalis. The presence of cytolysin-positive (cytolytic) E. faecalis correlated with the severity of liver disease and with mortality in patients with alcoholic hepatitis. Using humanized mice that were colonized with bacteria from the faeces of patients with alcoholic hepatitis, we investigated the therapeutic effects of bacteriophages that target cytolytic E. faecalis. We found that these bacteriophages decrease cytolysin in the liver and abolish ethanol-induced liver disease in humanized mice. Our findings link cytolytic E. faecalis with more severe clinical outcomes and increased mortality in patients with alcoholic hepatitis. We show that bacteriophages can specifically target cytolytic E. faecalis, which provides a method for precisely editing the intestinal microbiota. A clinical trial with a larger cohort is required to validate the relevance of our findings in humans, and to test whether this therapeutic approach is effective for patients with alcoholic hepatitis.

Development and Use of Personalized Bacteriophage-Based Therapeutic Cocktails To Treat a Patient with a Disseminated Resistant Acinetobacter baumannii Infection.

Widespread antibiotic use in clinical medicine and the livestock industry has contributed to the global spread of multidrug-resistant (MDR) bacterial pathogens, including Acinetobacter baumannii We report on a method used to produce a personalized bacteriophage-based therapeutic treatment for a 68-year-old diabetic patient with necrotizing pancreatitis complicated by an MDR A. baumannii infection. Despite multiple antibiotic courses and efforts at percutaneous drainage of a pancreatic pseudocyst, the patient deteriorated over a 4-month period. In the absence of effective antibiotics, two laboratories identified nine different bacteriophages with lytic activity for an A. baumannii isolate from the patient. Administration of these bacteriophages intravenously and percutaneously into the abscess cavities was associated with reversal of the patient's downward clinical trajectory, clearance of the A. baumannii infection, and a return to health. The outcome of this case suggests that the methods described here for the production of bacteriophage therapeutics could be applied to similar cases and that more concerted efforts to investigate the use of therapeutic bacteriophages for MDR bacterial infections are warranted.

The Caulobacter crescentus phage phiCbK: genomics of a canonical phage.

BACKGROUND: The bacterium Caulobacter crescentus is a popular model for the study of cell cycle regulation and senescence. The large prolate siphophage phiCbK has been an important tool in C. crescentus biology, and has been studied in its own right as a model for viral morphogenesis. Although a system of some interest, to date little genomic information is available on phiCbK or its relatives. RESULTS: Five novel phiCbK-like C. crescentus bacteriophages, CcrMagneto, CcrSwift, CcrKarma, CcrRogue and CcrColossus, were isolated from the environment. The genomes of phage phiCbK and these five environmental phage isolates were obtained by 454 pyrosequencing. The phiCbK-like phage genomes range in size from 205 kb encoding 318 proteins (phiCbK) to 280 kb encoding 448 proteins (CcrColossus), and were found to contain nonpermuted terminal redundancies of 10 to 17 kb. A novel method of terminal ligation was developed to map genomic termini, which confirmed termini predicted by coverage analysis. This suggests that sequence coverage discontinuities may be useable as predictors of genomic termini in phage genomes. Genomic modules encoding virion morphogenesis, lysis and DNA replication proteins were identified. The phiCbK-like phages were also found to encode a number of intriguing proteins; all contain a clearly T7-like DNA polymerase, and five of the six encode a possible homolog of the C. crescentus cell cycle regulator GcrA, which may allow the phage to alter the host cell's replicative state. The structural proteome of phage phiCbK was determined, identifying the portal, major and minor capsid proteins, the tail tape measure and possible tail fiber proteins. All six phage genomes are clearly related; phiCbK, CcrMagneto, CcrSwift, CcrKarma and CcrRogue form a group related at the DNA level, while CcrColossus is more diverged but retains significant similarity at the protein level. CONCLUSIONS: Due to their lack of any apparent relationship to other described phages, this group is proposed as the founding cohort of a new phage type, the phiCbK-like phages. This work will serve as a foundation for future studies on morphogenesis, infection and phage-host interactions in C. crescentus.

Solvent Extraction of Klebsiella pneumoniae Bacteriophage Lysates with 1-Dodecanol Results in Endotoxin Reduction with Low Risk of Solvent Contamination.

Antimicrobial resistance in pathogenic bacteria is increasing worldwide. One solution to this crisis is bacteriophage therapy, a treatment that harnesses naturally occurring bacterial viruses to invade and lyse antimicrobial resistant bacterial hosts. In Gram-negative hosts, a by-product of bacteriophage production is bacterial endotoxin, which can cause serious immune reactions in vivo. Purification methods using organic solvent extraction can remove endotoxin in bacteriophage lysates. In this study, we investigate a method for removal of endotoxin from 16 high-titer Klebsiella pneumoniae lysates by extraction with 1-dodecanol, 1-octanol, dodecane, or decane. In these experiments, treatment with either 1-dodecanol or 1-octanol resulted in removal of 104-105 endotoxin units/mL. Recovery of bacteriophage in lysates treated with dodecanol without dialysis was >90%, and residual dodecanol was low (10-1500 ppm). Overall these results suggest that organic solvent extraction using 1-dodecanol is effective at removing bacterial endotoxin, maintaining bacteriophage titer, and reducing solvent contamination in 16 K. pneumoniae bacteriophage lysates.

Complete Genome Sequence of Klebsiella pneumoniae Podophage Pone.

Klebsiella pneumoniae is a Gram-negative pathogen that has become increasingly antibiotic resistant. Phage therapy is potentially a useful approach to controlling this pathogen. Here, we present the genome sequence of the phiKMV-like K. pneumoniae podophage Pone.

Complete Genome Sequence of Klebsiella pneumoniae Jumbo Phage Miami.

Bacteriophage Miami infects Klebsiella pneumoniae, a Gram-negative pathogen that is becoming an increasing threat to public health due to its multidrug resistance. Here, we describe the annotation of the 253,383-bp jumbo phage Miami genome sequence and its similarity to other myophages.

Complete Genome Sequence of Klebsiella pneumoniae Myophage Muenster.

Klebsiella pneumoniae is associated with antibiotic-resistant nosocomial infections. Here, we present the annotated genome sequence of the Klebsiella jumbo phage Muenster. The Muenster genome sequence (346,937 bp) encodes 6 tRNAs and 561 putative protein-coding genes, including 9 tail fibers, suggesting a genetic mechanism to broaden the host range.

Genome-wide screens reveal Escherichia coli genes required for growth of T1-like phage LL5 and V5-like phage LL12.

The host factor requirements of phages and mechanisms of mutational phage insensitivity must be characterized for rational design of phage cocktails. To characterize host dependencies of two novel Escherichia coli phages, the T1-like siphophage LL5 and the V5-like myophage LL12, forward genetic screens were conducted against the Keio collection, a library of single non-essential gene deletions in E. coli str. BW25113. These screens and subsequent experiments identified genes required by phages LL5 and LL12. E. coli mutants deficient in heptose II and the phosphoryl substituent of heptose I of the inner core lipopolysaccharide (LPS) were unable to propagate phage LL5, as were mutants deficient in the outer membrane protein TolC. Mutants lacking glucose I of the LPS outer core failed to propagate LL12. Two additional genes encoding cytoplasmic chaperones, PpiB and SecB, were found to be required for efficient propagation of phage LL5, but not LL12. This screening approach may be useful for identifying host factors dependencies of phages, which would provide valuable information for their potential use as therapeutics and for phage engineering.

Complete Genome Sequence of Klebsiella pneumoniae Myophage Mineola.

Klebsiella pneumoniae is an important human pathogen due to the wide range of infections it can cause and its emerging drug resistance. Isolation and characterization of phage infecting K. pneumoniae could be important for future therapeutic applications. Here, we report the complete genome sequence of the T4-like Klebisella pneumoniae myophage Mineola.

Complete Genome Sequence of Klebsiella pneumoniae Myophage Menlow.

Klebsiella pneumoniae is an opportunistic pathogen that has become an increasing problem in nosocomial infections. Studying phages that infect K. pneumoniae may lead to improvements in phage therapeutics for treating these infections. Here, the full genome sequence of Menlow, a Vi01-like phage, is introduced and described.

Complete Genome Sequence of Enterotoxigenic Escherichia coli Siphophage LL5.

Here, we describe the complete genome sequence of siphophage LL5. LL5 is a T1-like phage isolated against enterotoxigenic Escherichia coli, which causes traveler's diarrhea. LL5 is included as a component phage in the commercial prebiotic product PreforPro.

Complete Genome Sequence of Enterotoxigenic Escherichia coli Myophage LL12.

This work describes the complete genome sequence of the virulent myophage LL12. The 136-kb LL12 genome is related to coliphage V5 and is a component in the prebiotic PreforPro. LL12 was isolated against enterotoxigenic Escherichia coli, which causes traveler's diarrhea.

Complete Genome Sequence of Klebsiella pneumoniae Siphophage Sanco.

Klebsiella pneumoniae is an opportunistic pathogen that is the cause of several hospital-acquired infections. Bacteriophages that target this bacterium could be used therapeutically as novel antimicrobial agents. Here, we present the complete genome sequence of the T1-like K. pneumoniae phage Sanco.

Complete Genome Sequence of Klebsiella pneumoniae Podophage Pylas.

Carbapenemase-producing Klebsiella pneumoniae is an important opportunistic pathogen due to its drug resistance. This study reports on the isolation and characterization of a podophage, named Pylas, infecting this bacterium. The complete genome of phage Pylas is described, and it is distantly related to the well-studied phage N4.

Complete Genome Sequence of Klebsiella pneumoniae Siphophage Seifer.

Carbapenemase-producing Klebsiella pneumoniae poses a significant public health threat due to its resistance to antibiotics. Siphophage Seifer was isolated and characterized as part of an effort to develop phage therapeutics to control this pathogen. This report describes the complete genome sequence of phage Seifer, which is a distant member of the χ-like siphovirus phage cluster.

Complete Genome Sequence of Klebsiella pneumoniae Myophage Mulock.

Klebsiella pneumoniae is an opportunistic pathogen associated with hospital-acquired infections. This report describes the complete genome of the K. pneumoniae myophage Mulock, which appears to be a temperate myophage distantly related to other Klebsiella myophages in morphogenesis genes and is partially syntenic with the canonical Escherichia phage lambda in genes encoding lambda-like functions.

Complete Genome Sequence of Proteus mirabilis Phage Mydo.

Proteus mirabilis is a pathogen that has been linked to nosocomial infections. Studies on phages infecting P. mirabilis may provide therapeutics for infections caused by antibiotic-resistant strains of this pathogen. Here, we announce the complete genome sequence of a P. mirabilis myophage, Mydo, which is distantly related to Escherichia coli phage rv5.

Complete Genome Sequence of Proteus mirabilis Phage Myduc.

Proteus mirabilis as a nosocomial pathogen is often the cause of urinary tract infections. This announcement describes the complete genome sequence of a P. mirabilis myophage named Myduc. Phage Myduc is related to Enterobacteria phage phiEcoM-GJ1, which belongs to a group of myophages with small genome sizes (52,000 to 56,000 bp) possessing a T7-like RNA polymerase.

Complete Genome Sequence of Proteus mirabilis Siphophage Stubb.

Proteus mirabilis, a Gram-negative bacterium belonging to the family Enterobacteriaceae, is a common cause of urinary tract infections. Phages infecting Proteus mirabilis could be used as therapeutics to treat infections caused by this bacterium. This announcement describes the complete genome sequence of the T5-like P. mirabilis phage Stubb.