Phages shape the structure of natural bacterial communities and can be effective therapeutic agents. Bacterial resistance to phage infection, however, limits the usefulness of phage therapies and ...could destabilise community structures, especially if individual resistance mutations provide cross-resistance against multiple phages. We currently understand very little about the evolution of cross-resistance in bacteria-phage interactions. Here we show that the network structure of cross-resistance among spontaneous resistance mutants of Pseudomonas aeruginosa evolved against each of 27 phages is highly modular. The cross-resistance network contained both symmetric (reciprocal) and asymmetric (nonreciprocal) cross-resistance, forming two cross-resistance modules defined by high within- but low between-module cross-resistance. Mutations conferring cross-resistance within modules targeted either lipopolysaccharide or type IV pilus biosynthesis, suggesting that the modularity of cross-resistance was structured by distinct phage receptors. In contrast, between-module cross-resistance was provided by mutations affecting the alternative sigma factor, RpoN, which controls many lifestyle-associated functions, including motility, biofilm formation, and quorum sensing. Broader cross-resistance range was not associated with higher fitness costs or weaker resistance against the focal phage used to select resistance. However, mutations in rpoN, providing between-module cross-resistance, were associated with higher fitness costs than mutations associated with within-module cross-resistance, i.e., in genes encoding either lipopolysaccharide or type IV pilus biosynthesis. The observed structure of cross-resistance predicted both the frequency of resistance mutations and the ability of phage combinations to suppress bacterial growth. These findings suggest that the evolution of cross-resistance is common, is likely to play an important role in the dynamic structure of bacteria-phage communities, and could inform the design principles for phage therapy treatments.
This study has reviewed the many roles of lumican as a biomarker of tissue pathology in health and disease. Lumican is a structure regulatory proteoglycan of collagen-rich tissues, with cell ...instructive properties through interactions with a number of cell surface receptors in tissue repair, thereby regulating cell proliferation, differentiation, inflammation and the innate and humoral immune systems to combat infection. The exponential increase in publications in the last decade dealing with lumican testify to its role as a pleiotropic biomarker regulatory protein. Recent findings show lumican has novel roles as a biomarker of the hypercoagulative state that occurs in SARS CoV-2 infections; thus, it may also prove useful in the delineation of the complex tissue changes that characterize COVID-19 disease. Lumican may be useful as a prognostic and diagnostic biomarker of long COVID disease and its sequelae.
New applications for phage integrases Fogg, Paul C M; Colloms, Sean; Rosser, Susan ...
Journal of molecular biology,
07/2014, Volume:
426, Issue:
15
Journal Article
Peer reviewed
Open access
Within the last 25 years, bacteriophage integrases have rapidly risen to prominence as genetic tools for a wide range of applications from basic cloning to genome engineering. Serine integrases such ...as that from ϕC31 and its relatives have found an especially wide range of applications within diverse micro-organisms right through to multi-cellular eukaryotes. Here, we review the mechanisms of the two major families of integrases, the tyrosine and serine integrases, and the advantages and disadvantages of each type as they are applied in genome engineering and synthetic biology. In particular, we focus on the new areas of metabolic pathway construction and optimization, biocomputing, heterologous expression and multiplexed assembly techniques. Integrases are versatile and efficient tools that can be used in conjunction with the various extant molecular biology tools to streamline the synthetic biology production line.
Phage therapy is a promising alternative to chemotherapeutic antibiotics for the treatment of bacterial infections. However, despite recent clinical uses of combinations of phages to treat ...multidrug-resistant infections, a mechanistic understanding of how bacteria evolve resistance against multiple phages is lacking, limiting our ability to deploy phage combinations optimally. Here, we show, using
and pairs of phages targeting shared or distinct surface receptors, that the timing and order of phage exposure determine the strength, cost, and mutational basis of resistance. Whereas sequential exposure allowed bacteria to acquire multiple resistance mutations effective against both phages, this evolutionary trajectory was prevented by simultaneous exposure, resulting in quantitatively weaker resistance. The order of phage exposure determined the fitness costs of sequential resistance, such that certain sequential orders imposed much higher fitness costs than the same phage pair in the reverse order. Together, these data suggest that phage combinations can be optimized to limit the strength of evolved resistances while maximizing their associated fitness costs to promote the long-term efficacy of phage therapy.
Globally rising rates of antibiotic resistance have renewed interest in phage therapy where combinations of phages have been successfully used to treat multidrug-resistant infections. To optimize phage therapy, we first need to understand how bacteria evolve resistance against combinations of multiple phages. Here, we use simple laboratory experiments and genome sequencing to show that the timing and order of phage exposure determine the strength, cost, and mutational basis of resistance evolution in the opportunistic pathogen
These findings suggest that phage combinations can be optimized to limit the emergence and persistence of resistance, thereby promoting the long-term usefulness of phage therapy.
Physician burnout is increasing, but navigating its prevention involves a complex intersection of physical, psychological, social, and organizational strategies.
Heterocycle‐containing cyclic peptides are promising scaffolds for the pharmaceutical industry but their chemical synthesis is very challenging. A new universal method has been devised to prepare ...these compounds by using a set of engineered marine‐derived enzymes and substrates obtained from a family of ribosomally produced and post‐translationally modified peptides called the cyanobactins. The substrate precursor peptide is engineered to have a non‐native protease cleavage site that can be rapidly cleaved. The other enzymes used are heterocyclases that convert Cys or Cys/Ser/Thr into their corresponding azolines. A macrocycle is formed using a macrocyclase enzyme, followed by oxidation of the azolines to azoles with a specific oxidase. The work is exemplified by the production of 17 macrocycles containing 6–9 residues representing 11 out of the 20 canonical amino acids.
Heterocycle‐containing cyclic peptides are promising scaffolds for the pharmaceutical industry, but their chemical synthesis is very challenging. A new universal method has been devised to prepare these compounds by using a set of engineered marine‐derived enzymes and substrates.
Background:
Minimally invasive surgery is being used increasingly, including for hallux valgus surgery. Despite the growing interest in minimally invasive procedures, there have been few publications ...on percutaneous chevron/akin (PECA) procedures, and no studies have been published comparing PECA to open scarf/akin osteotomies (SA).
Methods:
This was a prospective, randomized study of 50 patients undergoing operative correction of hallux valgus using one of 2 techniques (PECA vs open SA). Data were collected preoperatively and on 1 day, 2 weeks, 6 weeks, and 6 months postoperatively. Outcome measures include the American Orthopaedic Foot & Ankle Society Hallux-Metatarsophalangeal-Interphalangeal (AOFAS-HMI) Score, visual analog pain score, hallux valgus angle (HVA), and 1-2 intermetatarsal angle (IMA). Twenty-five patients underwent PECA procedures and 25 patients received SA procedures.
Results:
Both groups showed significantly improved AOFAS-HMI scores after surgery (PECA group: 61.8 to 88.9, SA group: 57.3 to 84.1, P = .560) with comparable final scores. HVA and IMA also presented similar outcomes at final follow-up (P = .520 and P = .270, respectively). However, the PECA group showed significantly lower pain level (VAS) in the early postoperative phase (postoperative day 1 to postoperative week 6, P < .001 and P = .004, respectively). No serious complications were observed in either group.
Conclusion:
Both groups showed comparable good to excellent clinical and radiologic outcomes at final follow-up. However, the PECA group had significantly less pain in the first 6 weeks following surgery.
Level of Evidence
Level II, prospective comparative study.
Summary
The serine integrase, Int, from the Streptomyces phage φC31 mediates the integration and excision of the phage genome into and out of the host chromosome. Integrases usually require a ...recombination directionality factor (RDF) or Xis to control integration and excision and, as φC31 Int only mediates integration in the absence of other phage proteins, we sought to identify a φC31 RDF. Here we report that the φC31 early protein, gp3 activated attL x attR recombination and inhibited attP x attB recombination. Gp3 binds to Int in solution and when Int is bound to the attachment sites. Kinetic analysis of the excision reaction suggested that gp3 modifies the interactions between Int and the substrates to form an active recombinase. In the presence of gp3, Int assembles an excision synaptic complex and the accumulation of the integration complex is inhibited. The structure of the excision synaptic complex, like that of the hyperactive mutant of Int, IntE449K, appeared to be biased towards one that favours the production of correctly joined products. The functional properties of φC31 gp3 resemble those of the evolutionarily unrelated RDF from phage Bxb1, suggesting that these two RDFs have arisen through convergent evolution.
Peptide macrocycles are found in many biologically active natural products. Their versatility, resistance to proteolysis and ability to traverse membranes has made them desirable molecules. Although ...technologies exist to synthesize such compounds, the full extent of diversity found among natural macrocycles has yet to be achieved synthetically. Cyanobactins are ribosomal peptide macrocycles encompassing an extraordinarily diverse range of ring sizes, amino acids and chemical modifications. We report the structure, biochemical characterization and initial engineering of the PatG macrocyclase domain of Prochloron sp. from the patellamide pathway that catalyzes the macrocyclization of linear peptides. The enzyme contains insertions in the subtilisin fold to allow it to recognize a three-residue signature, bind substrate in a preorganized and unusual conformation, shield an acyl-enzyme intermediate from water and catalyze peptide bond formation. The ability to macrocyclize a broad range of nonactivated substrates has wide biotechnology applications.
•Genome sequence of the 4-androstene-3,17-dione (AD) producer strain ‘M. neoaurum’ NRRL B-3805 (formerly Mycobacterium sp. NRRL B-3805).•Sequence of B-3805 was nearly identical to that of VKM ...Ac-1815D, but the detected differences determine its higher AD and lower ADD production.•These genome data permit the species-level identification of the strain using three marker sequences as Mycobacterium neoaurum.
Microbial bioconversion of sterols into high value steroid precursors, such as 4-androstene-3,17-dione (AD), is an industrial challenge. Genes and enzymes involved in sterol degradation have been proposed, although the complete pathway is not yet known. The genome sequencing of the AD producer strain ‘Mycobacterium neoaurum’ NRRL B-3805 (formerly Mycobacterium sp. NRRL B-3805) will serve to elucidate the critical steps for industrial processes and will provide the basis for further genetic engineering. The genome comprises a circular chromosome (5 421 338bp), is devoid of plasmids and contains 4844 protein-coding genes.
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