Methods of Detection and Identification of Bacteria (1977)

B. M. Mitruka

• Publisher: CRC Press
• Year: 2017
• Print ISBN: 9781138550568
• eBook ISBN: 9781351360081
• Topic: Sciences biologiques
• Subtopic: Biologie

The objective of this book is to present a critical review and evaluation of the so-called conventional methods currently being used for bacterial identification, as well as to discuss the new approaches for the detection and identification of bacteria. Morphological, biochemical, and serological methods of detection and identification of bacteria in clinical specimens are emphasised, and current methods of characterization and enumeration of bacteria in air, water, milk, and other food materials are also described.

Chapter 1
INTRODUCTION

Mary J. Bonner and Brij M. Mitruka

DIFFERENTIATION OF PROCARYOTIC AND EUCARYOTIC CELLS

Concepts proffered in Darwin’s Origin of the Species¹ provided the framework for the description of a third kingdom to serve as the transition form between the plant and animal kingdoms. Haeckel² termed this kingdom the Protista. Chatton³ recognized two general patterns of cellular organization in Protista, the eucaryotes (Greek, true nucleus), which include protozoa, fungi, and most algae, and the procaryotes, which include all bacteria and the small group of blue-green algae (Cyanophyceae). While the original division of Protista was based on the simple organization of the procaryotic bacteria, the electron microscope has revealed a fundamental division based on complexity of organization.

Bacteria form a heterogeneous group of procaryotic cells anatomically, physiologically, and biochemically. In general, the main distinguishing features of the procaryotic cells (bacteria and related organisms such as rickettsiae, chlamydiae, and mycoplasmas) are as follows.

1. Their nucleus is a simple homogeneous body without a nuclear membrane separating it from the cytoplasm. They lack nucleolus, a spindle, and a number of separate nonidentical chromosomes.

2. Procaryotes lack the internal membranes isolating the respiratory and photosynthetic enzyme systems in specific organelles comparable to the membrane-bound mitochondria and the chloroplasts of eucaryotic cells. Thus, the respiratory enzymes in bacteria are located mainly in the peripheral cytoplasmic membrane, and their effective functioning is dependent on the integrity of the cell protoplast as a whole.

3. Procaryotes have a rigid cell wall structure containing a specific mucopeptide not found in eucaryotic cells.

4. Morphologically, bacteria are characterized by their small cell size (usually between 0.4 and 1.5 μm in short diameter); characteristic shape, which may be spherical (coccus), rod shaped (bacillus), comma shaped (vibrio), spiral (spirillum and spirochete), or filamentous (actinomyces); and arrangement, such as clusters, chains, rods, filaments, or mycelia.

5. Although unicellular bacteria may grow attached to one another so as to appear multicellular (in clusters, chains, rods, etc.), each cell is physiologically self-sufficient and, if isolated artificially, able to nourish itself, grow, and reproduce the species by binary fission.

6. Another distinctive feature of the procaryotic cell is that its ribosomes are small⁴,⁵ (10 to 20 nm) with a sedimentation constant of 70S as compared to the larger 80S ribosome of eucaryotes. The 70S ribosome is composed of 30S and 50S subunits, whereas the 80S ribosome of the eucaryotic cell is composed of 40S and 60S subunits.

7. The basic chemical composition of all microorganisms is essentially similar, i.e., made up of compounds of lower molecular weight which are about 10% of dry weight in procaryotes and 15% in eucaryotes. Protein, nucleic acid, and lipid contents are also slightly lower in procaryotic cells as compared to eucaryotic cells.

The procaryotic bacteria show a considerably narrow range of structural and biochemical variations as compared to those in eucaryotic cells. Thus, evolutionary specialization among bacteria is expressed in metabolic rather than structural terms. However, there is great metabolic variation among bacteria; for example, representatives of all four primary nutritional categories (photoauto-trophs, photoheterotrophs, chemoautotrophs, and chemoheterotrophs) occur in bacteria. Almost every type of metabolic activity present in eucaryotic cells can be found in bacteria. The physiological diversity of bacteria, which explains both their numerical abundance and their ubiquity throughout the biosphere, is reflected in the role they play as agents in the cycles of carbon, nitrogen, sulfur, oxygen, and phosphorus.

Comparative cytology using the light microscope and classical staining methods, extension of comparative cytology to the ultrastructural level with the electron microscope (EM), and advances in biochemical techniques have established the uniqueness of bacteria.⁶,⁷ By using these three basic experimental approaches, it was recognized that bacteria have a wide range of anaerobic energy-yielding reactions, synthesize unique cell wall polymers (except for the “L”-form Mycoplasma and those without walls and exceptional bacteria such as Halobacterium), fix nitrogen, and accumulate poly-0-hydroxybutyrate as a reserve material. Such properties are virtually or completely absent from eucaryotes.

The cytological recognition of procaryotic organization supported by biochemical and physiological data provided the foundation for a coherent, systematic view of the nature of bacteria such as that described in Bergey’s Manual of Determinative Bacteriology.¹⁴

TAXONOMY OF BACTERIA

Bacteria are usually classified according to the Linnean binomial scheme of genus and species, although many kinds of systematic compilations are possible. Detailed classification of bacteria utilizes results of cytological, biochemical, and physiological tests as well as specialized techniques of strain identification (such as serotyping, bio-typing, bacteriophage typing, and genetic analyses including measurement of guanine and cytosine (GC) contents and DNA-DNA homology analysis).⁷¹³

Phylogenetic Scheme

The most widely used system of classification and nomenclature in the United States is Bergey’s Manual of Determinative Bacteriology.¹⁴ The first edition appeared in 1923 and the eighth edition in 1974. Bacteria are placed into the following 19 parts:

• Kingdom – Procaryotae
  • Division I – Cyanobacteria
• Division II – Bacteria
  • Part 1 – Phototrophic bacteria
  • Part 2 – Gliding bacteria
  • Part 3 – Sheathed bacteria
  • Part 4 – Budding and/or appendaged bacteria
  • Part 5 – Spirochetes
  • Part 6 – Spiral and curved bacteria
  • Part 7 – Gram-negative aerobic rods and cocci
  • Part 8 – Gram-negative facultatively anaerobic rods
  • Part 9 – Gram-negative anaerobic bacteria
  • Part 10 – Gram-negative cocci and coccobacilli
  • Part 11 – Gram-negative anaerobic cocci
  • Part 12 – Gram-negative chemolitho-trophic bacteria
  • Part 13 – Methane-producing bacteria
  • Part 14 – Gram-positive cocci
  • Part 15 – Endospore-forming rods and cocci
  • Part 16 – Gram-positive, asporo-genous, rod-shaped bacteria
  • Part 17 – Actinomycetes and related organisms
  • Part 18 – Rickettsias
  • Part 19 – Mycoplasmas

Each part is also divided, where appropriate, into classes, orders, families, tribes, genera, and species.

In the older phylogenetic scheme of classification, the class Schizomycetaceae included all true bacteria, filamentous bacteria (including mycobacteria), spirochetes, and mycoplasmas. For example, in this scheme Escherichia coli was placed as follows:

• Kingdom — Monera
  • Class (-aceae) — Schizomycetaceae
  • Order (-ales) — Eubacteriales
  • Family (-aceae) — Entero-bacteriaceae
  • Genus - Escherichia
  • Species - coli

However, since the publication of Bergey’s eighth edition, Escherichia coli is placed as follows:

• Kingdom – Procaryotae

• Division II – Bacteria

• Part 8 – Gram-negative facultatively

• anaerobic rods

• Family I – Enterobacteriaceae

• Genus I – Escherichia

• Species – coli

As specified in the International Code of Nomenclature of Bacteria,¹⁵ scientific...

Table of contents

• Cover Page
• Half Title
• Title Page
• Copyright Page
• Preface
• The Author
• Dedication
• Acknowledgment
• Table of Contents
• Chapter 1 Introduction
• Chapter 2 Review of Current Methods used in Bacteriology
• Chapter 3 New Methods of Detection and Identification of Bacteria
• Chapter 4 Detection and Identification of Bacteria by Gas Chromatography
• Chapter 5 Automated Methods in Bacteriology
• Chapter 6 Computer Identification of Bacteria