Lysins as a powerful alternative to combat Bacillus anthracis.

This review gathers all, to the best of our current knowledge, known lysins, mainly bacteriophage-derived, that have demonstrated activity against Bacillus anthracis strains. B. anthracis is a spore-forming, toxin-producing bacteria, naturally dwelling in soil. It is best known as a potential biowarfare threat, an etiological agent of anthrax, and a severe zoonotic disease. Anthrax can be treated with antibiotics (ciprofloxacin, penicillin, doxycycline); however, their administration may take up even to 60 days, and different factors can compromise their effectiveness. Bacterial viruses, bacteriophages (phages), are natural enemies of bacteria and use their lytic enzymes, endolysins (lysins), to specifically kill bacterial cells. Harnessing the potential of lysins to combat bacterial infections holds promise for diminishing antibiotic usage and, consequently, addressing the escalating antibiotic resistance in bacteria. In this context, we list the lysins with the activity against B. anthracis, providing a summary of their lytic properties in vitro and the outcomes observed in animal models. Bacillus cereus strain ATCC 4342/RSVF1, a surrogate for B. anthracis, was also included as a target bacteria. KEY POINTS: • More than a dozen different B. anthracis lysins have been identified and studied. • They fall into three blocks regarding their amino acid sequence similarity and most of them are amidases. • Lysins could be used in treating B. anthracis infections.

New bacteriophage-derived lysins, LysJ and LysF, with the potential to control Bacillus anthracis.

Bacillus anthracis is an etiological agent of anthrax, a severe zoonotic disease that can be transmitted to people and cause high mortalities. Bacteriophages and their lytic enzymes, endolysins, have potential therapeutic value in treating infections caused by this bacterium as alternatives or complements to antibiotic therapy. They can also be used to identify and detect B. anthracis. Endolysins of two B. anthracis Wbetavirus phages, J5a and F16Ba which were described by us recently, differ significantly from the best-known B. anthracis phage endolysin PlyG from Wbetavirus genus bacteriophage Gamma and a few other Wbetavirus genus phages. They are larger than PlyG (351 vs. 233 amino acid residues), contain a signal peptide at their N-termini, and, by prediction, have a different fold of cell binding domain suggesting different structural basis of cell epitope recognition. We purified in a soluble form the modified versions of these endolysins, designated by us LysJ and LysF, respectively, and depleted of signal peptides. Both modified endolysins could lyse the B. anthracis cell wall in zymogram assays. Their activity against the living cells of B. anthracis and other species of Bacillus genus was tested by spotting on the layers of bacteria in soft agar and by assessing the reduction of optical density of bacterial suspensions. Both methods proved the effectiveness of LysJ and LysF in killing the anthrax bacilli, although the results obtained by each method differed. Additionally, the lytic efficiency of both proteins was different, which apparently correlates with differences in their amino acid sequence. KEY POINTS: • LysJ and LysF are B. anthracis-targeting lysins differing from lysins studied so far • LysJ and LysF could be overproduced in E. coli in soluble and active forms • LysJ and LysF are active in killing cells of B. anthracis virulent strains.

Electrochemical, optical and mass-based immunosensors: A comprehensive review of Bacillus anthracis detection methods.

A biosensor is an analytical device whose main components include transducer and bioreceptor segments. The combination of biological recognition with the ligand is followed by transformation into physical or chemical signals. Many publications describe biological sensors as user-friendly, easy, portable, and less time-consuming than conventional methods. Among major categories of methods for the detection of Bacillus anthracis, such as culture-based microbiological method, polymerase chain reaction (PCR) and enzyme-linked immunosorbent assay (ELISA), microarray-based techniques sensors with bioreceptors have been highlighted which particular emphasis is placed on herein. There are several types of biosensors based on various chemical or physical transducers (e.g., electrochemical, optical, piezoelectric, thermal or magnetic electrodes) and the type of biological materials used (e.g., enzymes, nucleic acids, antibodies, cells, phages or tissues). In recent decades, antibody-based sensors have increasingly gained popularity due to their reliability, sensitivity and rapidness. The fundamental principle of antibody-based sensors is mainly based on the molecular recognition between antigens and antibodies. Therefore, immunosensors that detect B. anthracis surface antigens can provide a rapid tool for detecting anthrax bacilli and spores, especially in situ. This review provides a comprehensive summary of immunosensor-based methods using electrochemical, optical, and mass-based transducers to detect B. anthracis.

Three Novel Bacteriophages, J5a, F16Ba, and z1a, Specific for Bacillus anthracis, Define a New Clade of Historical Wbeta Phage Relatives.

Bacillus anthracis is a potent biowarfare agent, able to be highly lethal. The bacteria dwell in the soil of certain regions, as natural flora. Bacteriophages or their lytic enzymes, endolysins, may be an alternative for antibiotics and other antibacterials to fight this pathogen in infections and to minimize environmental contamination with anthrax endospores. Upon screening environmental samples from various regions in Poland, we isolated three new siphophages, J5a, F16Ba, and z1a, specific for B. anthracis. They represent new species related to historical anthrax phages Gamma, Cherry, and Fah, and to phage Wbeta of Wbetavirus genus. We show that the new phages and their closest relatives, phages Tavor_SA, Negev_SA, and Carmel_SA, form a separate clade of the Wbetavirus genus, designated as J5a clade. The most distinctive feature of J5a clade phages is their cell lysis module. While in the historical phages it encodes a canonical endolysin and a class III holin, in J5a clade phages it encodes an endolysin with a signal peptide and two putative holins. We present the basic characteristic of the isolated phages. Their comparative genomic analysis indicates that they encode two receptor-binding proteins, of which one may bind a sugar moiety of B. anthracis cell surface.

Bacteriophages, phage endolysins and antimicrobial peptides - the possibilities for their common use to combat infections and in the design of new drugs.

The antibiotic resistance in many pathogenic bacteria has become a major clinical problem, therefore, the necessity arises to search for new therapeutic strategies. The most promising solution lies in bacteriophages, phage endolysins and antimicrobial peptides. The aim of this study is to review the possibilities for the common use of bacteriophages, phage endolysins and antimicrobial peptides, both in the form of combined therapies and new strategies for the production of peptide drugs. Bacteriophages are viruses that specifically infect and destroy pathogenic bacteria by penetration into bacterial cells, causing metabolism disorders and, consequently, cell lysis. Phage-encoded endolysins are bacteriolytic proteins produced at the end of the phage lytic cycle that destroy elements of bacterial cell wall and enable the release of phage progeny from host cells. Antimicrobial peptides (AMPs) constitute an element of the innate immunity of living organisms and are characterized by the activity against a broad spectrum of bacteria. In the literature, there are only a few reports on the direct interaction of bacteriophages, phage endolysins and antimicrobial peptides against pathogenic bacteria. In each of them, a synergistic effect was observed, and Phage-encoded antimicrobial peptides as a specific group of AMPs have were also discussed. Phage-display technique was also reviewed in terms of its applications to produce and deliver biologically active peptides. The literature data also suggest that bacteriophages, phage endolysins and antimicrobial peptides can be used in combined therapy, thus negating many of the limitations resulting from their specificity as a single antimicrobial agent.