Gram-Positive Pathogens

Vincent A. Fischetti, Richard P. Novick, Joseph J. Ferretti, Daniel A. Portnoy, Mirian Braunstein, Julian I. Rood, Vincent A. Fischetti, Richard P. Novick, Joseph J. Ferretti, Daniel A. Portnoy, Mirian Braunstein, Julian I. Rood

• Publisher: ASM Press
• Year: 2019
• ISBN:
  9781683672876
• Edition: 3
• Topic: Biological Sciences
• Subtopic: Microbiology
• Index: Biological Sciences

Gram-positive bacteria, lacking an outer membrane and related secretory systems and having a thick peptidoglycan, have developed novel approaches to pathogenesis by acquiring (among others) a unique family of surface proteins, toxins, enzymes, and prophages. For the new edition, the editors have enhanced this fully researched compendium of Gram-positive bacterial pathogens by including new data generated using genomic sequencing as well as the latest knowledge on Gram-positive structure and mechanisms of antibiotic resistance and theories on the mechanisms of Gram-positive bacterial pathogenicity. This edition emphasizes streptococci, staphylococci, listeria, and spore-forming pathogens, with chapters written by many of the leading researchers in these areas. The chapters systematically dissect these organisms biologically, genetically, and immunologically, in an attempt to understand the strategies used by these bacteria to cause human disease.

"This textbook comprises a superb collection of scientific knowledge making it a must-read for any graduate student, medical doctor, or investigator studying these gram-positive bacteria and inspiring future imaginations of biological knowledge." - William R. Jacobs, Jr., PhD, Professor Microbiology & Immunology, Albert Einstein College of Medicine

The Gram-Positive Bacterial Cell Wall

Manfred Rohde

1

HISTORICAL BACKGROUND

In 1884, the Danish bacteriologist Hans Christian Gram developed a staining procedure to view stained bacteriaunder the light microscope (1). His staining method, nowadays simply called Gram staining, discriminated between a Gram-positive and Gram-negative bacterial cell wall. He introduced a dye, gentian violet, which penetrates the cell wall and cytoplasmic membrane, thus staining the cytoplasm of the heat-fixed bacteria. After addition of iodine, an insoluble complex is formed which is retained by the Gram-positive bacterial cell wall upon addition of a decolorizer such as ethanol. Therefore, Gram-positive bacteria appear almost purple, while Gram-negative bacteria retain the dye to a lesser extent or not at all and have to be counterstained with a second dye, safranin or fuchsine, appearing pink or reddish. It is noteworthy that some mycobacteria showed an indifferent staining behavior when Gram stained, suggesting that the cell wall of mycobacteria might be somehow different from the other two types. In the following decades, it became obvious that cell walls/cell envelopes are more diverse and that Gram staining alone often could lead to misinterpretations of the cell wall composition.

Before the early 1950s, when the chemical composition of bacterial cell walls was not known, it was speculated that chitin or cellulose, polymers recognized as providing rigid structures to other organisms, might also represent the building material of the bacterial cell wall. In 1951, experiments with a phenol-insoluble material from Corynebacterium diphtheria (2) revealed glucosamine and diaminopimelic acid as components of the bacterial cell wall which are associated with polysaccharides. Chemical examination of streptococcal cell wall layers highlighted the presence of amino acids and hexosamines in the cell wall extract, as well as rhamnose as a main component in Gram-positive bacteria (3, 4). Systematic analyses of a number of Gram-positive bacteria identified the hexosamines glucosamine and muramic acid as major components together with three prevalent amino acids, namely, d-alanine, lysine or diaminopimelic acid, and glutamic acid. By then, a typical basic basal unit in Gram-positive cell walls was also recognized in which glucosamine and muramic acid are linked with three amino acids via a peptide bond (5, 6). Gram-negative bacteria express the identical basal unit. Numerous analyses of other bacteria revealed that each bacterial genus or even species is often characterized by a distinctive pattern of amino acids, amino sugars, and sugars connected to the basic basal unit. It was believed that these differences should provide a valuable pattern to discriminate between bacterial genera/species (7, 8). Over the following years other compounds of the Gram-positive cell wall were recognized, such as teichoic acids (TAs), which are polyribitol phosphates (9), and lipoteichoic acid (LTA). Furthermore, numerous proteins were found to be linked to the cell wall.

Methods of staining bacteria for light microscopic examinations have limitations since the resolution is not high enough to reveal structural details. With the advent of transmission electron microscopes (TEMs) in the 1930s and the parallel development of preparation methods for biological samples, electron microscopic imaging of ultrathin sections of embedded bacteria became the method of choice to study bacterial cell walls in detail at high resolutions (10–13). With this methodology, it was possible for the first time to discriminate between the structures of Gram-positive and Gram-negative bacteria based on morphological differences in an image. First, electron microscopic preparation protocols developed for eukaryotic cells or tissues were also applied for bacteria. The most fruitful era started when embedding protocols were customized for bacteria and new kinds of embedding resins became available, for example, the Lowicryl resins for low-temperature embedding, which allowed the introduction of the progressive lowering of temperature method (14–16). This development was paralleled by technical inventions, especially cryo-methods in which bacteria are physically instead of chemically fixed, and it opened up a new horizon in understanding bacterial cell walls. It should be mentioned that even today new methodologies are arising and pushing morphological studies toward vitrified and unstained bacteria in a fully hydrated status and therefore in a close-to-nature condition. It is noteworthy that major developments in electron microscopic methodology required a long period of invention and testing before the technique was introduced to the market. For example, three-dimensional (3D) electron microscopy was developed about 30 years after the invention of the TEM. Invention and precommercial development of cryo-electron tomography (CET) occurred another 30 years later. Due to the rapid development of computer performance and the progress in specialized software nowadays, one can estimate that new imaging techniques are introduced faster. For example, the introduction of cryo-focused ion beam (cryo-FIB) combined with a scanning electron microscope (cryo-FIB-SEM) as a new close-to-nature approach was sold a few years after the first advent of FIB-SEMs for conventional resin-embedded biological samples.

THE BACTERIAL CELL WALL

Bacteria are mostly unicellular organisms which can be found in a wide variety of environments. Therefore, bacterial cell walls deserve special attention because they (i) provide the essential structure for bacterial...