Microbiology Lec 1 & 2 PDF

Lec. 1 Structure & Characteristics of Prokaryotic & Eukaryotic Cells Lec. 2 Microbial Growth Microbiology: study of organisms that are too small to see with the Naked eyes which need machine such as light microscope & E.M or stain or culture in media to see them such as bacteria viruses, fungi. TW...

Lec. 1 Structure & Characteristics of Prokaryotic & Eukaryotic Cells Lec. 2 Microbial Growth Microbiology: study of organisms that are too small to see with the Naked eyes which need machine such as light microscope & E.M or stain or culture in media to see them such as bacteria viruses, fungi. TWO GENERAL TYPES OF CELLS A. Prokaryotic ("before nucleus") - a cell lacking a membrane-bound nucleus & membrane-bound organelles (ex. bacteria); these cells do have some organelles, but they are not membrane-bound; all prokaryotic cells have a cell wall, its primary component being peptidoglycan; prokaryotic cells are much smaller than eukaryotic cells (about 10 times smaller); their small size allows them to grow faster & multiply more rapidly than eukaryotic cells (they have a higher surface area to volume ratio than larger cells; thus, because they are small, they can easily meet their modest nutritional needs and grow rapidly). This group includes all bacteria. B. Eukaryotic ("true nucleus") - a cell having a membrane-bound nucleus & membrane-bound organelles (“little organs” – specialized structures that perform specific functions within the cell); evolved about 2 million years after the prokaryotes; cell walls are sometimes present, but they are composed of cellulose or chitin; organisms with eukaryotic cells include fungi, algae, protozoa, plants, & animals. It is important to know the differences between prokaryotic and eukaryotic cells; allows us to control disease-causing bacteria without harming our own cells. Eukaryotic Cell Bacteria 1. Bacteria are relatively simple in structure. 2. They are procaryotic organism. 3. Simple unicellular organism with no nuclear membrane, Mitochondria, G. bodies, endoplasmic reticulum. 4. Bacteria reproduce a sexually by Binary fission or sexually by conjugation. 5. The nucleus consists of single chromosome which is circular double strand DNA (helix). 6. Bacteria have cell membrane which surrounds the cytoplasm and all metabolic activity found in it. 7. There is extra circular chromosomal DNA in the cytoplasm called plasmid. 8. All Bacteria have cell membrane, which is differentially permeable and very important for protection, the cell membrane contain mesosomes which support the cell by energy. 9. 99% of Bacteria have cell wall which is rigid & rough in structure but some do not have this wall permeable for water solution and nutrient the 10. The chemical structure of the cell wall is peptidoglycan which accessory specific material + Amino group & techoic acid. 11. The cell wall give protection for bacteria and give its shape, the shapes of Bacteria differ from each other. The Bacterial arrange as single cell such as micrococcus or double cell united pneumococcus or as chain such as streptococcus, or clusters staphylococcus or spiral as spirillum such as Nocardia. Classification of bacteria is mainly based on the following: 1. Shape 2. Composition of cell wall 3. Mode of respiration, nutrition 4. Arrangement Classification based on shape: Cell Shape: one function of the cell wall is to confer shape on the bacterium; most bacteria fall into one of these general groups. 1. Coccus (spherical) or round e.g Streptococcus pneumoniae 2. Bacilli. Rod shaped e.g Escherichia coli (E-coli) 3. Vibrio curved Vibrio cholerae or (comma shaped) e.g Vibrio cholerae 4. Spirilla (spiral)-rigid & short int length contain flagella in end e.g Nacardia. 5. Spirochetes - which is helix-Filament similar to corkscrew in which cell body is wrapped around a central fiber called the axial filament with internal flagella or end flagellum. e.g Spirillum volutans. * Some bacterial groups lack typical cell wall structure i.e. Mycobacterium and Nocardia Also classified into two major subgroups according to the composition and staining characteristic of the cell walls. 1. Peptidoglycan Gram+ve bacteria (purple) 2. Lipopoly Saccharide Gram -ve bacteria (pink) Some uncommon bacteria: 1. Mycoplasma: have no cell wall. 2. Actinomycetes filaments bacteria. Classification of bacteria based on the mode of nutrition: 1. Autotrophic bacterial Cyanobacteria 2. Heterotrophic bacteria e.g all diseases causing bacteria Classification of bacteria based on the mode of respiration 1. Anaerobic bacteria e.g Actinomycetes 2. Aerobic bacteria e.g Mycobacterium tuberculosis Bacterial arrangements: Bacterial arrange themselves according to the plane of successive cell division as single cells, pairs (diplococci), tetrads or chains streptococci, grape-like (clusters) e.g staphylococci, or as angled pairs. External Structures of the Bacterial Cell  Appendages  Glycocalyx – surface coating Two major groups of appendages used for: 1. Motility – flagella and axial filaments (periplasmic flagella) 2. Attachment or channels – fimbriae and pili  3 parts of Flagella (structure of Flagella) Filament, long, thin, helical structure (spiral) composed of protein flagellin.  Hook, curved sheath. Basal body, stack of rings firmly anchored in cell wall. Functions in motility of cell through environment.  Flagellar Arrangements: Monotrichous – single flagellum at one end. Lophotrichous – small bunches arising from one end of cell. Amphitrichous – flagella at both ends of cell. Peritrichous – flagella dispersed over surface of cell.  Flagellar Function: Guide bacteria in a direction in response to external stimulus: Chemical stimuli, chemotaxis; positive and negative. Light stimuli, phototaxis.  Many bacilli (rods) have flagella, but most cocci do not and are therefore non-motile. Spirochaetes move by using a flagellum-like structure called the axial filament, which wraps around the cell to produce an undulating motion.  Fimbriae Fine, proteinaceous, hairlike bristles from the cell surface. Function in adhesion to other cells and surfaces, attach bacteria to plant or animal cells to maintain themselves in a favorable environment; if fimbriae have been lost (maybe due to a mutation) in disease-causing bacteria, the bacteria will not be able to establish an infection.  Pili Rigid tubular structure made of pilin protein, they are usually short; all Gram negative bacteria have pili; function is to attach bacteria to other bacteria, other cells, or other surfaces. Sex pili allow one bacterial cell to adhere to another (cells can actually exchange genetic material through the pili to get sexual reproduction); called conjugation.  Glycocalyx: Coating of molecules external to the cell wall, made of sugars and/or proteins, Two types: Slime layer - loosely organized and attached. Capsule - highly organized, tightly attached, it is composed of polysaccharide, and sometimes protein (e.g., anthrax bacillus). The sugar components of the polysaccharide vary in different bacterial species and frequently determine the serological type within a species.   Functions: Protect cells from dehydration and nutrient loss. Inhibit killing by white blood cells by phagocytosis contributing to pathogenicity. Attachment - formation of biofilms this is especially true in the case of Streptococcus mutans, a major cariogenic organism, which has the ability to produce vast quantities of extracellular polysaccharide in the presence of dietary sugars such as sucrose.  Cell Envelope: External covering outside the cytoplasm Composed of two basic layers: cell wall and cell membrane Maintains cell integrity Two generally different groups of bacteria demonstrated by Gram stain: Gram-positive bacteria: thick cell wall composed primarily of peptidoglycan and cell membrane Gram-negative bacteria: outer cell membrane, thin peptidoglycan layer, and cell membrane. Structure of Cell Walls: Determines cell shape, Protect the cell, prevents lysis (bursting) or collapsing due to changing osmotic pressures. Peptidoglycan is primary component: unique macromolecule composed of a repeating long chains of polysaccharides (glycan) cross-linked by short proteins (peptide) fragments. When linked together, these chains create the single rigid mesh (resembles a chain link fence). Gram-positive Cell Wall Thick, homogeneous sheath of peptidoglycan, 20-80 nm thick. Function in cell wall maintenance and enlargement during cell division, stimulate a specific immune response. Gram-negative Cell Wall Composed of an outer membrane and a thin peptidoglycan layer Outer membrane is similar to cell membrane bilayer structure it has 3 layers. Outermost layer contains lipopolysaccharides (LPS). LPS compounds are endotoxins and are only released when the bacteria die and their cell walls are broken down. Endotoxin that may become toxic when released during infections may function as receptors and blocking immune response contains porin proteins in upper layer – regulate molecules entering and leaving cell. Endotoxins cause fever and dilate blood vessels (drop in blood pressure results). Killing the bacteria may increase the concentrations of this toxin. Bottom layer composed of phospholipids and lipoproteins So the cell wall peptidoglycan layer is: Single, thin sheet of peptidoglycan Protective structure while providing some flexibility and sensitivity to lysis  Periplasmic space surrounds peptidoglycan The Gram Stain: Differential stain that distinguishes cells with a Gram-positive cell wall from those with a Gram- negative cell wall. Gram-positive - retain crystal violet and stain purple. Gram-negative - lose crystal violet and stain red from safranin counterstain.  A group of bacteria that lack a cell wall; they avoid lysis from pressure by maintaining a nearly equal pressure between their cytoplasm and their external environment by actively pumping sodium ions out of the cell; additionally, their cell membranes are strengthened because they contain cholesterol, a lipid found in eukaryotic cell membranes.  Some bacterial groups lack typical cell wall structure i.e. Mycobacterium and Nocardia  Some have no cell wall i.e. Mycoplasma Plasma or Cell Membrane: membrane that encloses cytoplasm of any cell; major function is to contain cytoplasm and to transport and regulate what comes in and what goes out of the cell. Many prokaryotic cell membranes are similar to eukaryotic cell membranes. Its structure is referred to as the Fluid Mosaic Model, because the structure behaves more like a fluid than a solid. Contains:  Membrane Lipids: (composed primarily of phospholipid molecules), it is a phospholipid bilayer (hydrophobic fatty acid tails & hydrophilic phosphate heads). Cell Membrane Invaginations: the cell membrane sometimes invaginates or folds back on itself, forming structures that extend into the cytoplasm; since prokaryotic cells lack organelles, these invaginations are called Mesosome, they provide increased surface area for peripheral proteins (enzymes) to catalyze chemical reactions, functions as the origin of the transverse septum that divides the cell in half during cell division. It is also the binding site of the DNA that will become the genetic material of each daughter cell. Bacterial Internal Structures Cell cytoplasm: Dense gelatinous solution of sugars, amino acids, and salts, 90% water. Serves as solvent for materials used in all cell functions. It has the following structures: Nucleoid: or nuclear region is a mass of DNA; well defined, although it is not surrounded by a membrane; most of a bacterium's DNA is arranged in a single circular supercoiled double-stranded DNA molecule called a chromosome; some bacteria also contains smaller circular DNA molecules called plasmids which passed on to offspring, not essential to bacterial growth and metabolism, may encode antibiotic resistance, tolerance to toxic metals, enzymes and toxins, used in genetic engineering- readily manipulated and transferred from cell to cell. There are two types of plasmids, F- plasmid or (F factor) or (fertility) play a major role in conjugation of bacteria, R. Plasmid (resistance plasmid) to antibiotic & other factors.  Ribosomes: Site of protein synthesis; prokaryotic ribosomes are smaller than eukaryotic ribosomes. Antibiotics such as tetracycline, erythromycin, and streptomycin can specifically target bacterial ribosomes & not harm the host's eukaryotic ribosomes. They are made of 60% ribosomal RNA and 40% protein, consist of two subunits: large and small.  Cytoplasmic Inclusions and granules: intracellular storage bodies, vary in size, number and content. Bacterial cell can use them when environmental sources are depleted. Examples: glycogen, gas vesicles for floating, sulfur and phosphate granules.  Endospores: extremely hardy, resting (non-growing) structures that some bacteria, principally G(+), produce through the process of sporulation when nutrients are exhausted, they are able to withstand harsh environmental conditions (heat, drying, freezing, radiation and chemicals) because they contain so little water and high concentrations of calcium and dipicolinic acid; when favorable conditions return, the spore germinates into a new vegetative cell, which grow & reproduce. Some of endospore-producing bacteria are pathogenic to humans. Ex. Clostridium tetani causes tetanus (other species of this genus cause botulism & gas gangrene). Bacillus is another genus of bacteria that forms spores. Spores are called either terminal or subterminal, depending on their position. Useful bacteria Not all bacteria are harmful to humans. There are some bacteria which are beneficial to in different way: 1. Convert milk into curd, Lactobacillus or Lactic acid bacteria. 2. Ferment food products, Streptococcus & Bacillus 3. Help in digestion & improving the body's immunity system, Actinobacteria - Firmicutes proteobacteria. 4. Production of antibiotic, which is used for treatment prevention of bacterial infections. 5. Soil bacteria - Nitrogen fixation. Rhizobium Harmful bacteria: Those bacteria cause illness & responsible for infectious disease like Pneumonia, Tuberculosis, Viridians. Lec.2 Microbial Growth The term microbial growth refers to the growth of a population (or an increase in the number of cells), not to an increase in the size of the individual cell. Cell division leads to the growth of cells in the population. Two Types of Asexual Reproduction in Microbes: 1. Binary Fission - Bacterial reproduction occurs through fission, a primitive form of cell division that does not employ a spindle fiber apparatus. [A spindle fiber apparatus made of protein filaments is responsible for moving the chromosomes around during cell division (mitosis & meiosis) in most eukaryotic cells. Bacteria do not have these structures.] The bacterial cell doubles in size and replicates its chromosome. Following DNA replication, the two chromosomes attach to separate sites on the plasma membrane, and the cell wall is laid down between them, producing two daughter cells. 2. Budding - A few bacteria and some eukaryotes (including yeasts) may also replicate by budding, forming a bubble-like growth that enlarges and separates from the parent cell. Phases of Growth - A microbial lab culture typically passes through 4 distinct, sequential phases of growth that form the standard bacterial growth curve: (Not all growth phases occur in all cultures). 1. Lag Phase: In the lag phase, the number of cells doesn't increase. However, considerable metabolic activity is occurring as the cells prepare to grow. (This phase may not occur, if the cells used to inoculate a new culture are in the log phase & provided conditions are the same). 2. Log Phase (logarithmic or exponential phase): cell numbers increase exponentially; during each generation time, the number of cells in the population increases by a factor of two. The number of microbes in an exponentially increasing population increases slowly at first, then extremely rapidly. Organisms in a tube of culture medium can maintain log growth for only a limited time, as nutrients are used up, metabolic wastes accumulate, and microbes suffer from oxygen depletion. 3. Stationary Phase: The number of cells doesn't increase, but changes in cells occur: cell become smaller and synthesizes components to help them survive longer periods without growing (some may even produce endospores); the signal to enter this phase may have to do with overcrowding (accumulation of metabolic byproducts, depletion of nutrients, etc.). 4. Death Phase: In this phase, cells begin to die out. Death occurs exponentially, but at a low rate. Death occurs because cells have depleted intracellular ATP reserves. Not all cells necessarily die during this phase. Growth Factors: Microbes can exist in a great many environments because they are small, easily dispersed, need only small quantities of nutrients, are diverse in their nutritional requirements. A. Physical Factors 1. pH – bacteria can classified as: acidophiles (acid-loving): grow best at a pH of 1 to 5.4; Ex. Lactobacilllus (ferments milk) neutrophiles: exist from pH to 5.4 to 8.5; most bacteria that cause human disease are in this category. alkaliphiles (base loving): exist from pH to 7.0 to 11.5; ex. Vibrio cholerae (causes cholera) 2. Temperature – bacteria can be classified as: psychrophiles: (cold-loving) 15oC to 20oC; some can grow at 0oC. mesophiles: grow best between 25oC and 40oC; human body temp is 37oC. thermophiles: (heat-loving) 50oC to 60oC; found in compost heaps and in boiling hot springs. 3. Moisture – only the spores of sport-forming bacteria can exist in a dormant state in a dry environment. 4. Hydrostatic pressure – pressure exerted by standing water (ex. lakes, oceans, etc.); some bacteria can only survive in high hydrostatic pressure environments (ex. ocean valleys in excess of 7000 meters); the high pressure is necessary to keep their enzymes in the proper 3-D shape; without it, the enzymes lose their shape and denature and the cell dies. 5. Tonicity (hypotonic, hypertonic, isotonic) – The use of salt as a preservative in curing meats and the use of sugar in making jellies is based on the fact that a hypertonic environment kills or inhibits microbial growth. Halophiles (salt lovers) inhabit the oceans. 6. Radiation – UV rays and gamma rays can cause mutations in DNA and even kill microorganisms. Some bacteria have enzyme systems that can repair some mutations. B. Oxygen Requirements strict or obligate anaerobes – oxygen kills the bacteria; ex. Clostridium tetani strict or obligate aerobes – lack of oxygen kills the bacteria; ex. Pseudomonas facultative anaerobes – can shift their metabolism (anaerobic if oxygen is absent or aerobic if oxygen is present); ex. Escherichia coli, Staphylococcus aerotolerant – the bacteria don’t use oxygen, but oxygen doesn’t harm them; ex. Lactobacillus microaerophiles – like low oxygen concentrations and higher carbon dioxide concentrations; ex. Campylobacter C. Nutritional (Biochemical) Factors – Nutrients needed by microorganisms include: Carbon: carbon containing compounds are needed as an energy source (ex. glucose) and for building blocks. Nitrogen: needed for amino acids and nucleotides; some can synthesize all 20 amino acids; others have to have some provided in their medium. Sulfur: needed for amino acids, coenzymes, Phosphorus: needed for ATP, phospholipids, and nucleotides  Vitamins: a vitamin is an organic substance that an organism requires in small amounts and that is typically used as a coenzyme; many bacteria make their own, but some are required in the medium; microbes living in the human intestine manufacture vitamin K, needed for blood clotting, and some of the B vitamins, thus benefiting their host. Certain trace elements: ex. copper, iron, zinc, sodium, chloride, potassium, calcium, etc.; often serve as cofactors in enzymatic reactions.