Dependence between acylation degree and specific lysis actitivy of the Pseudomonas aeruginosa M6 bacteriophage

Abstract

Introduction

The purpose of this work is to study the influence of various degrees of protein’s acylation on lytic activity and other biological properties of Pseudomonas M6 bacteriophage. Materials and methods. The subject of the study were samples of the Pseudomonas O 12 bacteriophage, hereinafter named the M6 phage. Pure phage lines were obtained from individual standard-morphology phage plaques (plaque-forming units, or PFUs) after ten passes over the indicator strain. In subsequent experiments, M6 phage samples were selected that had a sufficient level of purity (homogeneous in the morphology of virions that inactivated the homologic antiphage serum by no less than 99.0% and the heterologic antibacterial serum by no more than 1%), and a titer no lower than 10 PFU per ml (PFU/ml). Titration of the phages was conducted using the standard bilayer agar. The crude protein content in the phage suspensions was determined using the spectroscopic method (at 280 and 260 nm). Succinylation of the phage samples was conducted according to the method developed by T.J. Molenaar. The phage’s lytic activity spectrum was determined through spot tests on 224 strains of the Pseudomonas genus and 106 strains of other genera (correspondingly, of strains Enterobacter -40, Escherichia -10, Citrobacter -8, Hafnia -7, Serratia -6, Shigella -10, Salmonella -8, Proteus -7) through the use of a phage suspension containing 10 ⁵ - 10 ⁶ PFU/ml. (This concentration of infectious phage corpuscles corresponds with the critical distribution of the M6 phage). The results of the research were subjected to statistical processing using single-factor dispersion analysis. Results and discussion. One hundred percent phage protein acylation leads to a loss of adsorption ability on sensitive cells and is accompanied by a significant (more than 10³ times) decrease in the level of specific lytic activity in the phage preparations. The succinylation type being studied does not cause changes to the morphology or antigen structure of phage virions and does not substantially affect the lytic activity spectrum or the average yield of infectious phage corpuscles on a single sensitive bacterial cell. Considering the relative uniqueness of any biological object (including the M6 phage being studied), the authors recognize the advisability of verifying the established patterns of the effect of various acylation levels on the biological properties of other types and morphological groups of viruses. It was proven that at phage sample acylation levels of 5%, 10%, and 20%, the specific lytic activity increased by a factor of ninety, twelve, and eight respectively. The adsorption speed constant increased by a factor of nine, eight, and eight respectively. And the duration of the latent period fell by 20% in all cases with the development of a productive infection caused by the acylated samples of the phages. The use of this chemical modification method does not cause changes to the morphology or antigen structure of phage virions and does not substantially affect the average yield of infectious phage corpuscles on a sensitive bacterial cell.