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In vitro destruction of Streptococcus pneumoniae biofilms with bacterial and phage peptidoglycan hydrolases.

ABSTRACT: Host- and phage-coded cell wall hydrolases have been used to fight Streptococcus pneumoniae growing as planktonic cells in vitro as well as in animal models. Until now, however, the usefulness of these enzymes in biofilm-grown pneumococci has gone untested. The antipneumococcal activity of different cell wall hydrolases produced by S. pneumoniae and a number of its phages was examined in an in vitro biofilm model. The major pneumococcal autolysin LytA, an N-acetylmuramoyl-l-alanine amidase, showed the greatest efficiency in disintegrating S. pneumoniae biofilms. The phage-encoded lysozymes Cpl-1 and Cpl-7 were also very efficient. Biofilms formed by the close pneumococcal relatives Streptococcus pseudopneumoniae and Streptococcus oralis were also destroyed by the phage endolysins but not by the S. pneumoniae autolysin LytA. A cooperative effect of LytA and Cpl-1 in the disintegration of S. pneumoniae biofilms was recorded.

SUBMITTER: Domenech M

PROVIDER: S-EPMC3165314 | biostudies-literature | 2011 Sep

REPOSITORIES: biostudies-literature

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In vitro destruction of Streptococcus pneumoniae biofilms with bacterial and phage peptidoglycan hydrolases.

Domenech Mirian M García Ernesto E Moscoso Miriam M

Antimicrobial agents and chemotherapy 20110711 9

Host- and phage-coded cell wall hydrolases have been used to fight Streptococcus pneumoniae growing as planktonic cells in vitro as well as in animal models. Until now, however, the usefulness of these enzymes in biofilm-grown pneumococci has gone untested. The antipneumococcal activity of different cell wall hydrolases produced by S. pneumoniae and a number of its phages was examined in an in vitro biofilm model. The major pneumococcal autolysin LytA, an N-acetylmuramoyl-l-alanine amidase, show ...[more]

PMID: 21746941

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Project description:Streptococcus pneumoniae is a Gram positive bacterium that causes severe invasive infection such as pneumonia, septicemia, meningitis and otitis media especially in children, the elderly and immune-compromised patients. Pneumococcal colonization and disease is often associated with biofilm formation. Bacteria in biofilms exhibit elevated resistance both to antibiotics and to host defense systems, which often results in persistent and difficult-to-treat infections. Therefore, the ongoing treat to human health posed by pneumococcal biofilms has prompted extensive research aimed to identify alternative targets and new antimicrobial agents that are effective against bacteria biofilms. The effective anti-biofilm strategies should include inhibition of microbial adhesion to the surface and of colonization, interference with the signal molecules modulating biofilm development and the disaggregation of the biofilm matrix. In this study, we examine the effect of DAM inhibitor small molecule pyrimidine-diones on streptococcus pneumoniae D-39 strain growth (planktonic and biofilm) and evaluate the changes in global gene expression using c-DNA microarray. The microarray analysis was performed on total RNA extracted from biofilms grown in 24-well microtiter plate with 7µm/ml pyrimidine-diones small molecule and control biofilms (biofilms grown without pyrimidine-diones small molecule). To validate the results of microarray, real-time RT-PCR was performed on 12 differentially expressed genes from six different functional groups. cDNA-microarray analysis detected a total of 259 genes that were significantly differentially expressed in biofilm growth with pyrimidine-diones small molecule. 204 genes were significantly down expressed and 55 genes were significantly up expressed in biofilms grown with 7µm/ml pyrimidine-diones small molecule. Among the 204 down expressed genes, 45 were hypothetical protein encoding gene and 159 were functional protein encoding genes. Of 55 up-regulated genes 21 were hypothetical genes and 34 were functional protein encoding genes. The functional annotation showed that gene involve in fatty acid metabolism, cell division, cell cycles, DNA metabolism, cell assembly were significantly down regulated and galactose metabolism related gene were up-expressed in biofilm grown with pyrimidine-diones small molecule.

Project description:UnlabelledMost bacterial cells are enclosed in a single macromolecule of the cell wall polymer, peptidoglycan, which is required for shape determination and maintenance of viability, while peptidoglycan biosynthesis is an important antibiotic target. It is hypothesized that cellular enlargement requires regional expansion of the cell wall through coordinated insertion and hydrolysis of peptidoglycan. Here, a group of (apparent glucosaminidase) peptidoglycan hydrolases are identified that are together required for cell enlargement and correct cellular morphology of Staphylococcus aureus, demonstrating the overall importance of this enzyme activity. These are Atl, SagA, ScaH, and SagB. The major advance here is the explanation of the observed morphological defects in terms of the mechanical and biochemical properties of peptidoglycan. It was shown that cells lacking groups of these hydrolases have increased surface stiffness and, in the absence of SagB, substantially increased glycan chain length. This indicates that, beyond their established roles (for example in cell separation), some hydrolases enable cellular enlargement by making peptidoglycan easier to stretch, providing the first direct evidence demonstrating that cellular enlargement occurs via modulation of the mechanical properties of peptidoglycan.ImportanceUnderstanding bacterial growth and division is a fundamental problem, and knowledge in this area underlies the treatment of many infectious diseases. Almost all bacteria are surrounded by a macromolecule of peptidoglycan that encloses the cell and maintains shape, and bacterial cells must increase the size of this molecule in order to enlarge themselves. This requires not only the insertion of new peptidoglycan monomers, a process targeted by antibiotics, including penicillin, but also breakage of existing bonds, a potentially hazardous activity for the cell. Using Staphylococcus aureus, we have identified a set of enzymes that are critical for cellular enlargement. We show that these enzymes are required for normal growth and define the mechanism through which cellular enlargement is accomplished, i.e., by breaking bonds in the peptidoglycan, which reduces the stiffness of the cell wall, enabling it to stretch and expand, a process that is likely to be fundamental to many bacteria.

Dataset Information

In vitro destruction of Streptococcus pneumoniae biofilms with bacterial and phage peptidoglycan hydrolases.

Publications

In vitro destruction of Streptococcus pneumoniae biofilms with bacterial and phage peptidoglycan hydrolases.

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