Microbiology The Science of Microorganisms What is Microbiology

Microbiology: The Science of Microorganisms

What is Microbiology? • Biology is the study of living organisms • Microbiology is the study of very small living organisms • Microorganisms are ubiquitous (they are everywhere!), those living things that are too small to be seen with the naked eye. • Categories of microorganisms include: – Viruses – Bacteria – Archaeans – Protozoa – Some algae and fungi

Type of Organisms Name of Study Characteristics of Organism Microorganisms Bacteria Viruses Simple, single-cell organisms. Bacteriology Grow in many environments. Lack a true nucleus and most organelles Virology Representative Examples Bacteria Composed of nucleic acid (DNA Viruses or RNA) and protein. Can reproduce only within living cells —obligate intracellular parasites Very simple, nongreen, plantlike organisms. Single-cell, globular Fungi forms are yeasts; multicellular filamentous forms are molds Fungi Mycology Protozoa Protozoology Single-cell, animal-like organisms Protozoa

What is Microbiology? • Viruses are technically infectious agents or particles • “Germs” are the microorganisms that cause disease • Disease-causing microorganisms are technically pathogens • Microorganisms that do not cause disease are nonpathogens

Disease • Disease- abnormality of structure or function of a body part, organ or system § May be felt or observed by others. § May have a known or unknown cause. § Severity of effects may be variable.

Where are Microbes? • Soil • Water • Air • Food • Chemicals • Sewage

Why Study Microbiology? • Microorganisms live on and in our bodies (indigenous microflora) • Microorganisms participate in bioremediation and genetic engineering • Some microorganisms are opportunistic pathogens • Microorganism are involved in elemental cycles • Microorganisms are essential for life on this planet • Microorganisms are involved in decomposition of dead organisms and waste (decomposers and saprophytes) • Algae and bacteria serve as food for tiny animals • Microorganisms aid in digestion of food • Microorganisms are used in the food and beverage industry

Why Study Microbiology? • Some microorganisms produce antibiotics used to treat diseases • Microbes are essential in the field of genetic engineering • Microbes are used as “cell models” • Microorganisms cause two categories of disease: – Infectious diseases-a pathogen inhabits the body and subsequently causes disease – Microbial intoxications- an individual ingests a toxin (poisonous substance) that has been produced by a pathogen outside the body

Benefits of Microorganisms • Fertilizing the soil • Aiding plant growth • Returning nitrogen from the air into the soil • Producing yeast (raised breads), cheeses, wine, beer, etc. . • Destruction of waste materials (paper, feces, biodegradable substances) • Purifying waste water.

Relevance in Healthcare • Taking precautions to protect self • Protecting patients and the community by preventing the spread of pathogens • Transferring harmful organisms • Keeping surrounding areas clean

Types of Microbes • Pathogenic-caused by disease causing microbe – virulence refers to the degree of damage caused by a microbe to its host. The pathogenicity of an organism - its ability to cause disease - is determined by its virulence factors • Opportunistic-Microb with the potential to cause disease, but doesn’t under normal circumstances • Normal flora-population of microorganisms that normally grows on body surfaces or cavities. • Saprophytes-organisms that live on dead or decaying matter (mushrooms) • Nitrogen fixing- bacteria capable of converting nitrogen gas to ammonia. Ex Clostridium • Iron-utilizing-bacteria which utilize the oxidation of iron ions as an essential component in their metabolic functioning (brown in color) ex; Bordetella

Pioneers in the Science of Microbiology Anton van Leeuwenhoek (1632 -1723) – “Father of Microbiology” – Not a trained scientist! – Made many simple single-lens microscopes – He was first to describe cells and bacteria ”animalcules” (bacteria and protozoa), seen through his microscopes

Pioneers in the Science of Microbiology Louis Pasteur (1822 -1895) – French chemist who made numerous contributions to microbiology – Investigated different fermentation products – Developed the pasteurization process – Reduces the number of microorganisms in foods such as milk – Discovered life forms that could exist without oxygen – anaerobes – Developed several vaccines, including rabies and anthrax vaccines

Pioneers in the Science of Microbiology Robert Koch (1843 -1910) – German physician who made numerous contributions to microbiology – Significant contribution to germ theory of disease – Discovered that B. anthracis produced spores – Developed method of fixing and staining bacteria – Developed methods to cultivate bacteria by Assisted Julius Petri in creating the Petri dish to grow bacteria for study

Koch’s Postulates 1. A particular microorganism must be found in all cases of the disease and must not be present in healthy animals or humans. 2. The microorganism must be isolated from the diseased animal or human and grown in pure culture in the laboratory. 3. The same disease must be produced when microorganisms from the pure culture are inoculated into healthy susceptible lab animals. 4. The same microorganism must be recovered from the experimentally infected animals and grown again in pure culture.

Koch’s Postulates (continued) • If an organism fulfills Koch’s postulates it is proven to be the cause of that particular infectious disease. • Koch’s Postulates helped to prove the germ theory of disease • Koch gave a tremendous boost to the development of microbiology by stressing lab culture and identification of microorganisms • Circumstances do exist in which Koch’s Postulates cannot be fulfilled.

Exceptions to Koch's Postulates 1. Some microbes are obligate intracellular parasites (like chlamydia or viruses) and are very challenging, or even impossible, to grow on artificial media. 2. Some diseases, such as tetanus, have variable signs and symptoms between patients. 3. Some diseases, such as pneumonia & nephritis, may be caused by a variety of microbes. 4. Some pathogens, such as S. pyogenes, cause several different diseases. 5. Certain pathogens, such as HIV, cause disease in humans only -- it is unethical to purposefully infect a human.

Ignaz Semmelweis/Holmes • Discovered the transmission of disease/microorganisms to patients from cadaver studies • Discovered the importance of handwashing to prevent the spread of infection

John Tyndall • Opposed to spontaneous generation of organisms • Established “Tyndallization” or discontinuous heating with steam as a sterilization technique.

Using the Metric System to Express the Sizes of Microorganisms • Metric units are used to express the sizes of microorganisms. • The basic unit of length in the metric system is the meter (m); it is equivalent to 39. 4 inches. • The sizes of bacteria and protozoa are usually expressed in terms of micrometers (µm). • The sizes of microorganisms are measured using an ocular micrometer.

Microscopes • The human eye, a telescope, a pair of binoculars, a magnifying glass and a microscope are various types of optical instruments. • A microscope is an optical instrument that is used to observe tiny objects; those that cannot be seen with the unaided human eye. • Each optical instrument has a limit as to what can be seen – The limit is referred to as the resolving power or resolution of the instrument.

Simple Microscopes • A simple microscope is one that contains only one magnifying lens. • A magnifying glass could be considered a simple microscope. – With a magnifying glass, images appear 3 -20 times larger than the object’s actual size. • Leeuwenhoek’s simple microscopes had a maximum magnifying power of about 300 X or 300 times.

Compound Microscopes • A compound microscope contains more than one magnifying lens. • Because visible light is the source of illumination, the compound microscope is also referred to as a compound light microscope. • Compound light microscopes usually magnify objects about 1000 times!

Electron Microscopes • Electron microscopes enable us to see extremely small infectious agents such as rabies and smallpox viruses. • Living organisms cannot be seen with an electron microscope – the processing procedures kill the organisms. • An electron beam is used as a source of illumination and magnets are used to focus the beam. • Electron microscopes have a much higher resolving power than the compound light microscope. • There are 2 types of electron microscopes - transmission and scanning.

The Transmission Electron Microscope • Uses an electron gun to fire a beam of electrons through an extremely thin specimen (<1 µm thick). • An image of the specimen is produced on a phosphorcoated screen. • Magnification is 1000 times greater than the compound light microscope.

The Scanning Electron Microscope • Electrons are bounced off the surface of a specimen and the image appears on a monitor. • Used to observe the outer surfaces of specimens. • Resolving power not as high as the transmission electron microscope. • Scanning and transmission electron micrographs are black and white images.

S. aureus and red blood cells as seen by light microscopy S. aureus as seen by transmission electron microscopy (TEM) S. aureus as seen by Scanning electron microscopy

Introduction: Cell Structure and Taxonomy • A cell is the fundamental living unit of any living organism because it exhibits the basic characteristics of life. • Metabolism -the chemical reactions that occur within a cell. • Bacterial cells do not have the complex system of membranes and organelles found in the more advanced cellular organisms.

Introduction (continued) • Bacteria and Archaea are called procaryotes or procaryotic cells because they do not have a true nucleus. • More complex cells that contain a true nucleus and many membrane-bound organelles are called eucaryotes or eucaryotic cells. • Eucaryotes include algae, protozoa, fungi, plants, animals and humans. • Some microorganisms are procaryotic, some are eucaryotic, and some are not cells at all.

Functions of Living Organisms Metabolism ◦ Physical & chemical processes involved in maintaining life Growth Ability to reproduce Irritability (able to react to environment) Motion – gliding, flagella, axial filaments Protection

Acellular and Cellular Microbes

Viruses • Composed of only a few genes protected by a protein coat. • Depend on the energy and metabolic machinery of a host cell in order to reproduce. • They are acellular, that is, they are not composed of cells.

Eucaryotic Cell Structure TEM of Yeast Cell

Eucaryotic Cell Structures • Cell Membrane – Mosaic of large molecules of proteins and phospholipids. – Regulates passage of nutrients, waste products and secretions in and out of the cell. – Has selective permeability.

The Nucleus • The “command center” of the cell. • 3 components: nucleoplasm, chromosomes, and nuclear membrane. • Chromosomes are embedded in the nucleoplasm. • Eucaryotic chromosomes consist of linear DNA molecules and proteins – Genes are located along the DNA molecules. – Each gene contains the information to produce a gene product.

The Nucleus (continued) • Most genes code for proteins but some code for 2 types of ribonucleic acid (RNA) – Ribosomal ribonucleic acid (r. RNA) – Transfer ribonucleic acid (t. RNA) • The organism’s complete collection of genes is the organism’s genotype or genome. • The number and composition of chromosomes and the number of genes on each chromosome are characteristic of the particular species of organism. • Human diploid cells have 46 chromosomes or 23 pairs.

Eucaryotic Cell Structures (continued) • Centrioles – Are near the nucleus, occurring in pairs and involved in the development of spindle fibers in cell division (mitosis) • Cytoplasm • Endoplasmic Reticulum – A highly convoluted system of membranes arranged to form a transport network in the cytoplasm. • Ribosomes – A semi-fluid, gelatinous, nutrient matrix. – Mainly ribosomal RNA and protein. – Contains storage granules and a variety of organelles. – Important in the synthesis of proteins. – Each organelle has a specific function. – The cytoplasm is where most metabolic reactions occur.

Eucaryotic Cell Structures (continued) • Golgi Complex • Lysosomes & Peroxisomes – Also Golgi apparatus or Golgi body. – Originate in the Golgi complex. – Connects or communicates with ER. – Lysosomes contain lysozyme and other digestive enzymes. – Completes the transformation of newly synthesized proteins and packages them for storage or export. – Peroxisomes are membrane bound vesicles where H 2 O 2 is generated and broken down.

Eucaryotic Cell Structures (continued) • Mitochondria – “Power plants, ” “powerhouses, ” or “energy factories” of the eucaryotic cell. – ATP molecules are formed by cellular respiration. – Number of mitochondria varies depending on activities of cell. • Plastids – Membrane-bound structures containing photosynthetic pigments – they are sites of photosynthesis. – Chloroplasts are a type of plastid.

Eucaryotic Cell Structures (continued) • Cytoskeleton – A system of fibers throughout the cytoplasm. – 3 types of fibers: microtubules, microfilaments and intermediate filaments. – Microtubules and microfilaments essential for a variety of activities. • Cell wall – Some eucaryotic cells contain cell walls – an external structure to provide shape, protection and rigidity. – Simpler than procaryotic cell wall. – Chitin found in cell wall of fungi; cellulose in cell wall of algae and plants.

Eucaryotic Cell Structures (continued) • Flagella and Cilia – Some eucaryotic cells (e. g. , spermatozoa) possess long thin structures called flagella. – Organelles of locomotion; may be one or more flagella on a cell. – Cilia are also organelles of locomotion but are shorter, thinner and more numerous; hair-like. – Cilia can be found on some species of protozoa and certain types of cells in our bodies (e. g. , respiratory tract). Cell with numerous cilia

Procaryotic Cell Structure • Procaryotic cells are about 10 times smaller than eucaryotic cells. • Procaryotic cells are simple compared to eucaryotic cells. • Procaryotic cells reproduce by binary fission. • All bacteria are procaryotes as are archaeans. • Unlike eucaryotic cells, the cytoplasm of procaryotic cells is not filled with internal membranes. • Cytoplasm of procaryotic cell surrounded by cell membrane, a cell wall (usually) and sometimes a capsule or slime layer.

Procaryotic Cell

Procaryotic Cell Structure (continued) • Cell membrane – Similar in structure and function to the eucaryotic cell membrane. – Selectively permeable. – Many enzymes are attached to the cell membrane and metabolic reactions take place there. • Chromosome – Procaryotic chromosome usually consists of a single, long, supercoiled, circular DNA molecule – serves as control center of cell. – Plasmids are small circular molecules of DNA that are not part of the chromosome.

Procaryotic Cell Structure (continued) • Cytoplasm – Semi-liquid that consists of water, enzymes, waste products, nutrients, proteins, carbohydrates and lipids – required for metabolic functions of the cell. • Cytoplasmic particles – Most are ribosomes occurring in clusters. – Eucaryotic ribosomes are smaller than procaryotic ribosomes, but function is the same protein synthesis.

Procaryotic Cell Structure (continued) • Bacterial Cell Wall – Rigid exterior that defines the shape of bacterial cells – chemically complex. – Main constituent of most bacterial cell walls is peptidoglycan (only found in bacteria). – Gram-positive bacteria have a thick layer of peptidoglycan; Gram-negative bacteria have a much thinner layer. – Mycoplasma spp. do not have a cell wall.

Gram-negative and Gram-positive Cell Walls Gram-negative bacterium Gram-positive bacterium

Procaryotic Cell Structure (continued) • Glycocalyx (Slime Layers and Capsules) – Some bacteria possess a thick layer of glycocalyx outside their cell wall. – Glycocalyx is a slimy, gelatinous material produced by the cell membrane and secreted outside the cell wall which is loose and allow for bacteria to slide along solid surfaces. – 2 types of glycocalyx – slime layer and capsule. – The capsule is more highly organized and more firmly attached to the cell wall than the slime layer. • Pseudomonas produces a slime layer. • K. pneumoniae, N. meningitidis and S. pneumoniae possess a capsule.

Capsule Stain (Example of a negative staining technique)

Procaryotic Cell Structure (continued) • Flagella – Flagella are threadlike, protein appendages that enable bacteria to move. – Number and arrangement of flagella are characteristic of a particular species: – Peritrichous bacteria – flagella over entire surface – Lophotrichous bacteria – flagella at one end – Amphitrichous bacteria – flagella at both ends – Monotrichous bacteria – single polar flagellum

A Peritrichous Salmonella Cell

Procaryotic Cell Structure (continued) • Pili -Hair-like structures, most often observed on Gramnegative bacteria. – Composed of polymerized protein molecules called pilin. – Pili are thinner than flagella, have a rigid structure and are not associated with motility. – Pili enable bacteria to anchor themselves to surfaces. – Some bacteria possess a sex pilus for conjugation.

Proteus vulgaris cell, showing pili and several flagella Pili Flagella

Procaryotic Cell Structure (continued) Spores (Endospores) • A few genera (e. g. , Bacillus and Clostridium) are capable of forming thick-walled spores as a means of survival. • The process of spore formation is called sporulation – it is not reproduction. • Spores have been shown to survive for many years and are resistant to heat, cold drying and most chemicals. • Usually one spore is produced in a bacterial cell and generates into one vegetative bacterium. • Endospores can be visualized with a spore stain.

Recap of Structural Differences Between Procaryotic and Eucaryotic Cells • Eucaryotic cells contain a true nucleus; procaryotic cells do not. • Eucaryotic cells are divided into plant and animal types – Animal cells do not have a cell wall, plant cells have a simple cell wall. • Eucaryotic cells contain membranous structures and many membrane bound organelles; procaryotic cells possess no membranes other than the cell membrane that encloses the cytoplasm

Reproduction of Organisms and Their Cells • Procaryotic Cell Reproduction – Procaryotic cells reproduce by a process known as binary fission – one cell splits in half to become two daughter cells. • Before a procaryotic cell divides in half, the chromosome must be duplicated.

Taxonomy • Taxonomy is the science of classification of living organisms. • Taxonomy consists of classification, nomenclature and identification. • Classification is the arrangement of organisms into taxonomic groups (known as taxa). • Tool for remembering the sequence of Taxa – “King David Came Over for Good Spaghetti” KDCOFGS, K for Kingdom, D for Division, C for Class, O for Order, F for Family, G for Genus and S for species.

Microbial Classification • The science of taxonomy was established on the binomial system of nomenclature. • In the binomial system, each organism is given 2 names – genus and the specific epithet. Taken together, both names constitute the species. – For example, Escherichia coli; Escherichia is the genus and coli is the specific epithet. – The genus is frequently abbreviated with just a single letter, (e. g. , E for Escherichia). • The abbreviation “sp. ” is used to designate a single species and “spp. ” for more than one species.

Microbial Classification • Organisms are categorized into larger groups based on their similarities and differences. • The Five-Kingdom System of Classification 1. Bacteria and archaeans – Kingdom Procaryotae 2. Algae and protozoa – Kingdom Protista 3. Fungi – Kingdom Fungi 4. Plants – Kingdom Plantae 5. Animals – Kingdom Animalia • Viruses are not included because they are acellular. • Other systems of classification do exist.

Microbial Classification (continued) • The Three-Domain System of Classification 1. Archaea (procaryotic) 2. Bacteria (procaryotic) 3. Eucarya (all eucaryotic organisms)

The Domain Bacteria Characteristics • Bacteria are divided into 3 major phenotypic categories: – Those that are Gram-negative and have a cell wall. – Those that are Gram-positive and have a cell wall. – Those that lack a cell wall. • Characteristics of bacteria used in classification and identification include: cell morphology, staining reactions, motility, colony morphology, atmospheric requirements, nutritional requirements, biochemical and metabolic activities, enzymes that the organism produces, pathogenicity and genetic composition.

The Domain Bacteria Cell Morphology • There are 3 basic shapes of bacteria: – Cocci (round bacteria) – Bacilli (rod-shaped bacteria) – Curved and spiral-shaped bacteria • Cocci may be seen singly or in pairs (diplococci), chains (streptococci), clusters (staphylococci), packets of 4 (tetrads), or packets of 8 (octads). • The average coccus is about 1 µm in diameter. • Some cocci have “coccus” in their name.

The Domain Bacteria Cell Morphology (continued) • Bacilli – Often referred to as rods; may be short or long, thick or thin, and pointed or with curved or blunt ends. – They may occur singly, in pairs (diplobacilli), in chains (streptobacilli), in long filaments, or branched. – An average sized bacillus is 1 x 3 µm. – Examples of medically important bacilli: Escherichia, Klebsiella, and Proteus spp. Pseudomonas, Haemophilus, and Bacillus spp.

The Domain Bacteria Cell Morphology (continued) • Curved and Spiral-Shaped Bacteria – Examples of curved bacteria: • Vibrio spp. • Campylobacter spp. • Helicobacter spp. – Examples of spiral-shaped bacteria: • Treponema spp. • Borrelia spp. Borrelia hermsii in stained blood smear; a cause of relapsing fever.

Factors Influencing Growth of Microorganisms Light ◦ Amount ◦ Type Temperature Moisture Food Availability Atmosphere-Gas-Oxygen supply p. H

Naming Bacteria • Genus name: Capitalized and italic • species name: lowercase and italic • EXAMPLE: Escherichia coli or E. coli

Species Clostridium • Clostridium difficile • Clostridium botulinum • Clostridium sodellii • Clostridium tetani • Clostridium perfringens • These 5 are the most commonly seen but there are many others in this species…….