Introduction to Food & Industrial Microbiology MB1001 PDF

ntroduction to Food & Industrial Microbiology, Module MB1001 Module coordinator Prof Douwe van Sinderen, School of Microbiology ([email protected]) General introduction All organisms can be divided into 1) Macroorganisms (we can see them) 2) Microorganisms (we can’t see th...

ntroduction to Food & Industrial Microbiology, Module MB1001 Module coordinator Prof Douwe van Sinderen, School of Microbiology ([email protected]) General introduction All organisms can be divided into 1) Macroorganisms (we can see them) 2) Microorganisms (we can’t see them) Microbiology is the study of microorganisms. Microorganisms are ubiquitous and include viruses, bacteria, algae, fungi and protozoa.  Various subdisciplines of Microbiology: Food Microbiology, Industrial Microbiology, Medical Microbiology, Environmental Microbiology Impacts on various fundamental sciences: Genetics, Molecular Biology, Biochemistry, Immunology, Cell Physiolo Some Aspects of microbiology are: Living cells and how they work Function and effect of microorganisms Microbial diversity and evolution  Tree of Life t es Euka r yo ryote ka s Pro Archaea Euryarchaeota Eucarya G reen Slim e Anim als nonsulfur Entam oebae m olds Bacteria G ram bacteria Fungi +ve Cren- H alophiles Plants bacteria archaeota Ciliates Purple bacteria Therm oproteus Cyanobacteria Pyrodictium Flagelattes Flavobacteria Trichom onads Therm otogales M icrosporodia Diplom onads Viruses are not (evolutionary) related to prokaryotes and eukaryotes  How big is a…? Virus Bacterium 0.05-0.1 microns 0.5-1.5 microns Red blood cell 5 microns 10 microns (0.01 mm) Micron = micrometer = m  So what, if they’re that small why bother? Obvious reason: They are the major causes of diseases – COVID-19, AIDS, TB and malaria Not so obvious reason: The ecosystem depends on them - they are the major contributors to the Carbon cycle Nitrogen cycle Phosphorous cycle Decomposition cycles And other important geochemical cycles  The Cell The cell is the basic unit of all living systems. The vast majority of living organisms are unicellular (microorganisms), while those of higher organisms (animals and plants) are multicellular (e.g. a human body is composed of ~37 trillion cells). We can classify cells into two main types: Eucaryotic and Procaryotic (Bacteria & Archaea) Eu-(true) caryos (nucleus) pro-(before) caryos (nucleus) All animal and plant cells belong to the Eucaryotes (eukaryotes), but there are also many unicellular eucaryotes, such as yeast. B Viruses areAs notaconsidered general rule, eucaryotic living cells are entities/obligate (they are biological larger than parasite procaryotes (prokaryotes), and are much more complex. Basic morphologies (shapes) of prokaryotes Diplococci Staphylococci Rod-shaped Vibrios Micrococci Streptococci Sarcina Spirilla There are square bacteria – like living in very high salt concentrations This also makes it very difficult to identify bacteria by simply looking at them Bifidobacterium Morphologies of some eukaryotic cells Nerve cell Rods Egg cell Cones Retinal cells  Examples of where microbiology matters Agriculture Food N2 fixation Food preservation Nutrient cycling Fermented Food Animal husbandry Food additives Disease/medicine Identifying (new) diseases Treatment and cure Disease prevention Energy/Environment Biotechnology Biofuels (methane and ethanol) Genetically modified organisms Bioremediation Production of pharmaceuticals Microbial mining Gene therapy for certain diseases  1900 Influenza & pneumonia Tuberculosis Gastroenteritis Heart disease Stroke Kidney disease Accidents Cancer Infant disease Diphtheria 0 100 200 Deaths per 100,000 population  1990s Heart disease 310 Cancer Stroke Accidents Influenza & pneumonia Diabetes Cirrhosis of the liver Suicide Arteriosclerosis Homicide 0 100 200 Deaths per 100,000 population Sources: CDC, WORLDOMETER IMPORTANT: COVID-19 data is the number of ACTUAL US deaths since March 15th, 2020 as reported on Worldometer against the backdrop of the EXTRAPOLATED AVERAGE DAILY number of deaths for top 15 causes of death in the US based on the latest (2018) data from the CDC. This chart is not meant to represent statistical analysis of any kind, it is meant for visual purposes only to help raise public awareness of the exponentially increasing COVID-19 deaths in the US. The three major diseases of the 21st century ~1.2 million die of Tuberculosis – Bacterial. Between 2000 and 2025, it is estimated that: Over one billion people will be newly infected with TB. >200 million people will become sick from TB. TB will claim at least 35 million lives. ~0.6 million people, mostly in Africa, die of Malaria – Protozoal.  Infectious Disease – historical impact Plague – 25 million died Smallpox/TB - 8 million people in Caribbean post 1492 US Civil War (1861-1865) – 90% died of disease, 10% died in battle Spanish Flu 1918 – more killed than in entire WW I (525 million infected, 21 million died) 1900 USA – 1/3rd to ½ all babies died before age of 5  Infectious Disease – historical impact Microbes have markedly affected the course of human history: Decline of Rome (Justinian) was hastened by smallpox (virus) & the bubonic plague (Yersinia pestis - bacterium) (A.D. 565). In 1346 the Black Death struck (Yersinia pestis). The population of Europe, North Africa and the Middle East totalled 100 million. After the plague 25% had died. Typhus, pneumonia and dysentery played a role in Napoleon’s departure by decimating his army during their retreat from Moscow in 1812 and contributed to him losing at Waterloo in 1815.  Food borne disease (FBD) - Costs FBD refers to those diseases which result from ingesting contaminated food. Not all due to microorganisms (toxins/allergies/ etc). For example in 2011 in the US alone 48 million get sick due to FBD, 128,000 require hospitalization, 3000 die Food poisoning in the UK affects 1 in 5 people per annum  How We Study Microbial Cells The discovery and early study of cells progressed with the invention and improvement of microscopes in the 17th century. In a light microscope (LMs) visible light passes through the specimen and then through glass lenses. – The lenses refract light such that the image is magnified into the eye or a video screen. Bacteria on a pin head  Resolution of a light microscope is about 2 microns, the size of a small bacterium Light microscopes can magnify effectively to about 1,000 times the size of the actual specimen. – At higher magnifications, the image blurs. We can also study microbes by isolating them as pure cultures in the laboratory. Use selective media to isolate different types of microbes. Individual colonies (each colony consists of billions of bacteria) Atoms are the building blocks of all matter Each element consists of atoms (e.g. H2O or O2). An atom is the smallest unit of matter that still retains the properties of an element. Atoms are composed of even smaller parts: Neutrons and protons which are packed together to form a dense core, the atomic nucleus, at the center of an atom, and electrons that form a cloud around the nucleus  Each electron has one unit of negative charge. Each proton has one unit of positive charge. Neutrons are electrically neutral. Fig 2.5 In Campbell. Models of the Helium atom Different elements have different numbers of electrons  Atoms combine by chemical bonding to form molecules Atoms can interact with other atoms and share their electrons These interactions typically result in the atoms remaining close together, held by attractions called chemical bonds. Chemical bonds can form between atoms of the same element or atoms of different elements. While both types are molecules, the latter are also compounds. Water, H2O, is a compound in which two hydrogen atoms form single covalent bonds with an oxygen atom. THE STRUCTURE AND FUNCTION OF MACROMOLECULES Three of the four classes of macromolecules form chain-like molecules called polymers. Polymers consist of many similar or identical building blocks linked by covalent bonds. The repeated units are small molecules called monomers. The chemical mechanisms that cells use to make and break polymers are similar for all classes of macromolecules. Monomers are connected by covalent bonds via a condensation reaction or dehydration reaction. When cells need to break down polymers they use the opposite reaction. The covalent bonds connecting monomers in a polymer are disassembled by hydrolysis.  Polymers are like beads (monomers) connected to make a string Monomers are the same Monomers are different, e.g. DNA/RNA 4 different monomers Protein 20 different monomers ings (polymers) can be linear, but biological polymers can also contain branches  An immense variety of polymers can be built from a small set of monomers Each cell has thousands of different macromolecules. This diversity comes from various combinations of the 40- 50 common monomers and other rarer ones. These monomers can be connected in various combinations like the 26 letters in the alphabet can be used to create a great diversity of words. Biological molecules are even more diverse.  Bacterial cell composition There is no typical cell or species Water constitutes 70% or more of a typical bacterial cell Remaining 30 % of the material will be composed of the elements C - carbon H - hydrogen O - oxygen N – nitrogen With smaller amounts of K - potassium, P - phosphorous, S - sulphur, Ca - calcium, Mg - magnesium, Na - sodium, Fe – iron and other trace elements e.g. Mn – manganese; Mo - Molybdenum Proteins are instrumental in about everything that an organism does. These functions include structural support, storage, transport of other substances, intercellular signaling, movement, and defense against foreign substances. Many proteins act as enzymes in a cell and regulate metabolism by selectively accelerating chemical reactions. Proteins are the most structurally complex molecules known. All protein polymers are constructed from the same set of 20 monomers, called amino acids. Polymers of amino acids are called polypeptides. A protein consists of one or more polypeptides folded and coiled into a specific conformation. They can constitute 50-55% of the dry weight of a cell. A polypeptide is a polymer of amino acids connected in a specific sequence. Amino acids consist of four components attached to a central carbon, the alpha carbon. These components include a hydrogen atom, a carboxyl group, an amino group, and a variable R group (or side chain). Differences in R groups produce the 20 different amino acids. H Amino H O Carboxyl group N C C group H OH R The twenty different R groups may be as simple as a hydrogen atom (as in the amino acid glycine) to a carbon skeleton with various functional groups attached. The physical and chemical characteristics of the R group determine the unique characteristics of a particular amino acid. One group of amino acids has hydrophobic (or non-polar) R groups. Another group of amino acids has polar R groups, making them hydrophilic. The last group of amino acids includes those with functional groups that are charged (ionized) at cellular pH. Amino acids are joined together when a condensation reaction removes a hydroxyl group from the carboxyl end of one amino acid and a hydrogen from the amino group of another. The resulting covalent bond is called a peptide bond. Repeating the process over and over creates a long polypeptide chain. At one end is an amino acid with a free amino group (the N- terminus) and at the other is an amino acid with a free carboxyl group the (the C-terminus). Attached to the polypeptide backbone are the various R groups. Polypeptides range in size from a few monomers to thousands. A protein’s function depends on its specific conformation A functional protein consists of one or more polypeptides that have been precisely twisted, folded, and coiled into a unique shape. It is the order of amino acids that determines what the 3-D conformation will be. The folding of a protein from a chain of amino acids occurs more or less spontaneously. Three levels of protein structure: primary, secondary, and tertiary structure, are used to organize the folding within a single polypeptide. Quaternary structure arises when two or more polypeptides join to form a protein.  Lysozyme 129 aa The primary structure of a Attacks protein is its unique Bacteria sequence of amino acids. The precise primary structure of a protein is determined by inherited genetic information. Even a slight change in primary structure can affect a protein’s conformation and ability to function The secondary structure of a protein results from hydrogen bonds at regular intervals along the polypeptide backbone  Helix  Pleated sheet Tertiary structure is determined by a variety of interactions among R groups and between R groups and the polypeptide backbone. Quaternary structure results from the aggregation of two or more polypeptide subunits In spite of the knowledge of the three-dimensional shapes of over 10,000 proteins, it is still difficult to predict the conformation of a protein from its primary structure alone. Environment (high temp, extremes in pH) can disrupt sec/tert/quat structure: denaturation of protein, which inactivates the biological activity of proteins Traditionally, scientists used X-ray crystallography to determine protein conformation, although in recent years electron microscopes have become powerful enough to obtain 3D images of (macro)molecules  How many ways can 20 amino acids be combined in a 200 amino acid protein? 1.6 x 10260 160000000000000000000000000000000000000 000000000000000000000000000000000000000 000000000000000000000000000000000000000 000000000000000000000000000000000000000 000000000000000000000000000000000000000 000000000000000000000000000000000000000 0000000000000000000000000000  Bacterial cell composition There is no typical cell or species Macromolecules: Nucleic Acids There are two types of nucleic acids: ribonucleic acid (RNA) and deoxyribonucleic acid (DNA). Nucleic acids are polymers of monomers called nucleotides. Each nucleotide consists of three parts: a nitrogen base, a pentose sugar, and a phosphate group. Nucleotide structure Phosphate group Nitrogen base Pentose sugar The nitrogen bases, rings of carbon and nitrogen, come in two types: purines and pyrimidines. Pyrimidines have a single six-membered ring. Three types: cytosine (C), thymine (T), and uracil (U). They differ in the atoms attached to the ring. NB  Purines have a six-membered ring joined to a five-membered ring. The two purines are adenine (A) and guanine (G). DNA: GATC RNA: GAUC The pentose joined to the nitrogen base is ribose in nucleotides of RNA and deoxyribose in DNA. The only difference between the sugars is the lack of an oxygen atom on carbon two in deoxyribose. The combination of a pentose and nitrogen base is a nucleoside. The addition of a phosphate group creates nucleotide. Polynucleotides are synthesized by connecting the sugars of one nucleotide to the phosphate of the next with a phosphodiester link. This creates a repeating backbone of sugar- phosphate units with the nitrogen bases as appendages The linear sequence of nucleotides in a nucleic acid chain is commonly abbreviated by a one-letter code, for example A—G—C—T—T—A—C—A ; this is referred to as the DNA sequence. Furthermore two polynucleotides will combine with each other to form a double helix – the bases interact with other bases on the second chain. Base pairing in DNA Most DNA molecules have thousands to millions of base pairs.  https://i.pinimg.com/originals/d8/e5/4 b/d8e54b944370214729f9ff718f7ed3 71.gif Discovered by Watson & Crick in 1953 Rosalind Franklin generated X-ray data of DNA What is the main function of nucleic acid? Nucleic acids store and transmit hereditary information. They contain the blueprint for life. The basic unit of hereditary information is the gene. The flow of genetic information is from DNA  RNA  protein. The sequence of nitrogen bases along a DNA polymer is unique for each gene. Genes are normally hundreds to thousands of nucleotides long. The number of possible combinations of the four DNA bases is massive. Q: How many different ways can 4 bases be combined, in any order, in a piece of DNA 1000 bases long? A: 10602 different ways. (10130 atoms in the universe). The linear order of bases in a gene specifies the order of amino acids - the primary structure of a protein. The primary structure in turn determines three-dimensional conformation and function. Amino acids Nucleus mRNA Ribosome Protein DNA (messenger) Site of protein synthesis While DNA has the information for all the cell’s activities, it is not directly involved in the day to day operations of the cell. Each gene along a DNA molecule directs the synthesis of a specific type of RNA molecule - messenger RNA (mRNA). The mRNA interacts with the protein-synthesizing machinery to direct the ordering of amino acids in a polypeptide.  Nucleotides have many other functions They carry chemical energy ATP They combine with other groups to form coenzymes. Coenzyme A They are used as specific signaling molecules in the cell. cAMP  Bacterial cell composition There is no typical cell or species  Macromolecules: Carbohydrates (sugars)  monosaccharides Sucrose disaccharides Lactose raffinose stachyose amylose Fig. 5.3 Monosaccharides, particularly glucose, are a major energy source for cellular work. They also function as the raw material for the synthesis of other monomers, including those of amino acids and fatty acids. Fig. 5.4 Two monosaccharides can join with a glycosidic linkage to form a disaccharide via dehydration. Maltose, malt sugar, is formed by joining two glucose molecules. Fig. 5.5a Sucrose, table sugar, is formed by joining glucose and fructose and is the major transport form of sugars in plants. Fig. 5.5 Polysaccharides are polymers of hundreds to thousands of monosaccharides joined by glycosidic linkages. One function of polysaccharides is as an energy storage macromolecule that is hydrolyzed as needed. Other polysaccharides serve as building materials for the cell or whole organism. Starch is a storage polysaccharide composed entirely of glucose monomers. Fig. 5.6a Animals also store glucose in a polysaccharide called glycogen. Glycogen is highly branched. Humans and other vertebrates store glycogen in the liver and muscles. Insert Fig. 5.6b - glycogen Fig. 5.6b Structural polysaccharides form strong building materials. Cellulose is a major component of the tough wall of plant cells. Cellulose is also a polymer of glucose monomers, but using beta linkages. Fig. 5.7c Fig. 5.8 The enzymes that digest starch cannot hydrolyze the (beta) linkages in cellulose. Some microbes can digest cellulose to its glucose monomers through the use of cellulase enzymes. Many eukaryotic herbivores, like cows and termites, have symbiotic relationships with cellulolytic microbes, allowing them access to this rich source of energy. Macromolecules: Lipids/fats Lipids are an exception among macromolecules because they do not form polymers. The unifying feature of lipids is that they all are insoluble in water. This is because their structures are dominated by non-polar covalent bonds. Lipids are highly diverse in form and function. Fats store large amounts of energy Although fats are not strictly polymers, they are large molecules assembled from smaller molecules by dehydration reactions. A fat is constructed from two kinds of smaller molecules, glycerol and fatty acids. Glycerol consists of a three carbon skeleton with a hydroxyl group attached to each. A fatty acid consists of a carboxyl group attached to a long carbon skeleton, often 16 to 18 carbons long. Fig. 5.10a The many nonpolar C-H bonds in the long hydrocarbon skeleton make fats hydrophobic. In a fat, three fatty acids are joined to glycerol by an ester linkage, creating a triacylglycerol. Fig. 5.10b The three fatty acids in a fat can be the same or different. Fatty acids may vary in length (number of carbons) and in the number and locations of double bonds. If there are no carbon-carbon double bonds, then the molecule is a saturated fatty acid - a hydrogen at every possible position. Fig. 5.11a If there are one or more carbon-carbon double bonds, then the molecule is an unsaturated fatty acid - formed by the removal of hydrogen atoms from the carbon skeleton. Saturated fatty acids are straight chains, but unsaturated fatty acids have a kink wherever there is a double bond. Fig. 5.11b Fats containing fatty acids without double bonds are saturated fats. Most animal fats are saturated. Saturated fat are solid at room temperature. Fats with unsaturated fatty acids are unsaturated fats. Plant and fish fats, known as oils, are liquid are room temperature. The kinks provided by the double bonds prevent the molecules from packing tightly together. The major function of fats is energy storage. A gram of fat stores more than twice as much energy as a gram of a polysaccharide. Plants use starch for energy storage when mobility is not a concern but use oils when dispersal and packing is important, as in seeds. Humans and other mammals store fats as long-term energy reserves in adipose cells. A layer of fats can also function as insulation.  Phospholipids are major components of cell membranes Phospholipids have two fatty acids attached to glycerol and a phosphate group at the third position. The phosphate group carries a negative charge. Additional smaller groups may be attached to the phosphate group. At the surface of a cell phospholipids are arranged as a bilayer: the hydrophilic heads are on the outside in contact with the aqueous solution and the hydrophobic tails from the core. The phospholipid bilayer forms a barrier between the cell and the external environment. They are the major component of membranes Fig. 5.12b  Plasma membrane (cytoplasmic membrane) Selectively permeable barrier: allows nutrients in and waste products out Phospholipid: Outside of cell Hydrophilic H 2O region Fatty acids Hydrophobic region Phospholipid Bilayer Glycerol Phosphate H 2O Inside of cell - cytosol A pure phospholipid bilayer would be relatively impermeable. So how can substances and signals traverse the membrane? Some molecules (water,small fatty acids) can traverse the membrane without the need for any assistance. Charged molecules (H+) and larger molecules must be transported by CARRIERS (proteins) in the membrane.  Steroids Structurally different from lipids, but have in common that they are insoluble in water. They represent complex organic molecules with four interlocking ring structures (e.g. cholesterol, testosterone, estrogen, cortisone) A historical view of microbiology  Microbiology before 1650 Human diseases prior to the 17th century were thought to be caused by Supreme beings Miasmas – unwholesome atmosphere Spontaneous generation of life Magic Chemical instabilities  Spontaneous generation Accepted idea, for centuries, started with Aristotle (4th Century BC) The idea that organisms originate directly from non-living matter – mud to frogs. "life from non-life" abiogenesis - (a-not bio-life genesis- origin) disproved by Francesco Redi’s and Louis Pasteur’s experiments.  Microbiology before 1650 However, the scientific community was not easily persuaded that such processes as food decay were due to biological entities. Chemical instabilities and spontaneous generation were accepted as the reason for diseases and life. One example of this school of thought was Jan Baptista van Helmont (c. 1580-1644). Who published a recipe for producing mice from soiled underwear and wheat. This was considered “state of the art” science.  Van Helmont’s “experiment” on spontaneous generation Wheat 21 Days Sweaty underwear Open-mouthed jar  Microbiology 1650 to 1850 During this period the greatest advancement in microbiology came from the invention and application of the microscope. This instrument lead to the realization that living matter was composed of individual cells. Many pioneers in the area of microscopy (e.g. Robert Hooke) but for microbiology the main one was Antonie van Leeuwenhoek (c. 1632- 1723) Founder of microbiology  Microbiology 1650 to 1850 Van Leeuwenhoek, Antonie (1676): First to describe microorganisms (animalcules) Born in Delft Holland – tradesman/draper  Microbiology 1650 to 1850 Using his microscope he described a variety of microbes or “animalcules” from environments as diverse as seawater and the mouth. Drawing made by A. van Leeuwenhoek. Probable identification: A: Bacillus B: motile Selenomonas E: Micrococcus F: Leptothrix G: a spirocete Microbiology 1650 to 1850 Edward Jenner (c. 1749-1823) 1798: was the first to use vaccination in order to protect against a disease I selected a healthy boy, about eight years old. The matter was taken from the [cowpox] sore on the hand of Sarah Nelmes and it was inserted on 14 May 1796 into the boy by two cuts each about half an inch long. On the seventh day he complained of uneasiness, on the ninth he became a little chilly, lost his appetite and had a slight headache and spent the night with some degree of restlessness, but on the following day he was perfectly well. In order to ascertain that the boy was secure from the contagion of the Empirically smallpox, he was inoculated with smallpox matter, but no disease followed. Several months later he was again inoculated with smallpox showed the matter but again no disease followed." efficacy of the vaccine  Microbiology after 1850: The beginning of modern microbiology The Pasteur School Louis Pasteur (1822 - 1895) Disproved spontaneous generation Supported germ theory of disease Developed vaccines (anthrax and rabies)  Microbiology after 1850: Pasteur’s disproves spontaneous generation Pasteur delivered the fatal blow to the idea of spontaneous generation, after Rudolf Virchow (1858) put forward the idea of biogenesis as the reason for life.  Pasteur and spontaneous generation Steam Dust escapes Air moves in Infusion from air From open and out of Is heated settles end of flask flask in bend Months Infusion sits Infusion No microbes appear remains sterile indefinitely Flask is tilted Sterile infusion Sterile infusion Microbes appear sites in flask comes into in infusion contact with dust  Microbiology after 1850: At the same time, Pasteur discovered the existence of life without oxygen: anaerobic life. This discovery paved the way for the study of germs that cause septicaemia and gangrene, among other infections. It also lead to an understanding of the process of fermentation – the degradation of sugars in the absence of oxygen. This lead to the development of pasteurisation.  Microbiology after 1850: The Koch School Robert Koch (1843-1910) This school of thought concentrated on the isolation of pure cultures of pathogens. Confirmed germ theory Proposed a system to prove an organism was a disease causing agent – Koch’s postulates Developed pure culture methods  Microbiology after 1850: Koch/Petri/Hesse Koch and his assistant (Richard Petri) discovered agar & Petri dishes (plates) for isolation of pure cultures. In 1876 plated samples from diseased animals and obtained pure cultures of Bacillus anthracis – cause of anthrax. In 1882 discovered organism responsible for tuberculosis.  Microbiology after 1850: Koch’s Postulates 1. The bacterium must be present in every case of the disease. 2. The bacterium must be isolated from the diseased host & grown in pure culture. 3. The specific disease must be reproduced when a pure culture of the bacterium is inoculated into a healthy susceptible host. 4. The bacterium must be recoverable from the experimentally infected host.  Diseased animal Healthy animal 1 Red blood cells Observe blood/ tissue under the Suspecte microscope d pathoge n 2 Steak agar Colonies plate with of sample from suspecte either animal d Inoculate healthy Colonies animal with cells 4 of of suspected Pure culture suspecte pathogens must be same d organism as pathogen before Observe blood/ tissue under the microscope Laboratory culture 3 Suspecte Diseased animal d pathoge  Microbiology after 1850: Koch’s Postulates - Limitations A bacterium that is usually part of the normal flora may become a pathogen in certain situations: - opportunistic Not all of those infected will develop disease – asymptomatic. Some bacteria are not culturable in the lab (e.g. Mycobacterium leprae) or there may be no suitable animal model of infection. Different groups of microbes What different types of microbes do we know? NB. Many microbes live as single cells or cell clusters; some multicellular, but not as complex as animals, plants. Eukaryotes: Algae – Unicellular as well multicellular. Use light to generate energy – photosynthesis. Can be found in aquatic environments. Some are motile. Protozoa - Usually motile unicellular organisms, derive energy from degradation of organic materials. Many are pathogenic: malaria, African sleeping sicknes. Fungi – Consist of yeasts (unicellular) and moulds (filamentous, can be macroscopic). Example: Saccharomyces cerevisiae (Yeast). No photosynthesis, can grow in drier and more acidic environments than other microorganisms. Reproduce sexually and asexually, usually through spore formation.  What different types of microbes do we know (cont’d)? Prokaryotes - Bacteria and Archaea Very diverse group Unicellular Classification based on physiology, biochemistry and genetics Reproduce through binary fission Daughter cells are identical clones DNA Asexual reproduction One generation  What different types of microbes do we know (cont’d)? Viruses Strictly speaking a virus would not be considered a living organism (acellular). They are parasites that can only reproduce by using a suitable host. They can be seen with an electron microscope and may contain DNA or RNA as their genetic material  Viruses and bacteria compared Typical Viruse bacteria s Intracellular parasite No Yes Plasma membrane Yes No Binary fission Yes No Pass through bacterial No Yes filters Possess both DNA and Yes No RNA ATP-generating Yes No metabolism Ribosomes Yes No Sensitive to antibiotics Yes No  The microbial cell – size and shape The size of a microbial cell or virus is measured in µm (micrometer – 10-6 meter) or nm (nanometer – 10-9 meter). Due to their size we need to use a microscope to see them. Typical bacterial cell measures 1-5 m. Two types. 1. Light microscope – uses visible light to illuminate the subject Four variations - Bright field, Phase contrast, Dark- field and Fluorescence 2) Electron microscope – This has a much higher resolution than a light microscope. Can study cell structure. Scanning electron Transmission electron microscopy microscopy Microscopic image of a drop of lake water  Staining of micro organisms Purpose: to increase contrast between microbial cells and background, but can also be used to distinguish cells from each other Positive staining: stains cells Negative staining: stains background Gram stain is used to distinguish bacteria into Gram-negative and Gram-positive. Cell wall is stained with crystal violet (purple), with also the use of a counter stain (safranin). Hans Christian Gram 1853 - 1938  Nomenclature of Bacteria Strict convention exists regarding the way in which the names of bacteria are written. The genus (plural: genera) name is written first with a capital letter. It is always written in italics (or underlined). Bacillus – B. Salmonella – S. Lactobacillus – Lb. or L. The genus may be abbreviated to the first (or first and a second) letter. Carl Linnaeus 1707 –1778 Binomial nomenclature The species name comes second and is never abbreviated. Bacillus cereus or B. cereus Bacillus subtilis or B. subtilis Lactobacillus bulgaricus or Lb. bulgaricus To further distinguish between species we can use strain names. These are not in italics. Lactobacillus bulgaricus CHO or Lb. bulgaricus CHO To write this on paper we show the italics by underlining the name. Lactobacillus bulgaricus CHO = Lactobacillus bulgaricus CHO  Prokaryotic and Eukaryotic cell types There are two fundamental cell types. Prokaryotic (before nucleus) – they have no membrane enclosed nucleus. Bacteria and Archaea (or Archaebacteria). Eukaryotic (true nucleus) – have a membrane-enclosed nucleus. erences/similarities between prokaryotes and eukaryotes Structure Function Procaryote Eucaryote Cell membrane Semi-permeable barrier Yes Yes Cytoplasm Semi fluid medium Yes Yes Peptidoglycan cell wall Gives rigidity and shape Yes No Cytoskeleton Gives rigidity and shape No Yes Ribosomes Sites of protein synthesis Yes Yes Chromosome(s) Repository of genetic information Yes (usually one) Yes Nucleus Membrane bound location of DNA No (nucleoid) Yes Organelles (mitochondria, Chloroplast, Endoplasmic Various roles in cell metabolism No Yes reticulum) Flagellum (plural: flagella) Responsible for cell motility Some Some Cell structures  Common Features of All Cells All cells, whether they are prokaryotic or eukaryotic, have some common features.  Procaryotes Small and simple (1/10 to 1/1000 human cell) Possess a plasma membrane (as do eukaryotes) Can possess internal structures (but no internal membrane) Have a peptidoglycan cell wall (unique) May have structures outside cell wall (flagella, pili) Some bacteria can form endospores (internal to cell)  Prokaryotic architecture The simplest prokaryotic cells are spherical (cocci) or rod-shaped (bacilli), but many different shapes are possible (conferred by the cell wall),including spiral,filamentous and square cells. om ) es NA Re cle ne e Pla all sule som gi oid Nu bra (D me sma la w el os on Ca p ag m Cell Ribo es Fl i Pil M A typical cell has the following structure  Pili: attachment structures Flagella: not all bacteria:locomotion Capsule: not all bacteria:slippery layer Cell wall: confers rigidity and shape Cell Cell membrane: semi-permeable barrier envelop Nucleoid: DNA e Ribosomes: sites of protein synthesis Mesosomes: invagination of membrane will first consider the cell envelope and associated structure  Bacterial cell envelope: There is considerable pressure on the cell membrane of a bacterium, due to the concentration of dissolved solutes inside relative to outside (~2 atmospheres = pressure of a car tyre). Therefore, bacteria require cell walls to resist this pressure, and also to confer shape and rigidity to the cell. Bacteria have chosen two distinct routes to building a cell wall. This choice divides the bacteria into two groups – Gram positive and Gram negative. Named after the Danish bacteriologist in 1884, Hans Christian Gram (1853 - 1938)  Peptidoglycan N-acetylmuramic acid (NAM) N-acetylglucosamine Peptide side chains on NAM Back bone is twisted Individual sugar chains can Peptide align as interbridges sheets. Sheets can stack up to give a porous, strong S-layer Lipoteichoic acid Polysaccharide Teichoic acid Peptidoglyc an Plasma Membra ne Gram positive Structure of Staphylococcus aureus peptidoglycan Penicillin kills bacteria by interfering with the cross-linking and weakening the cell wall-cells eventually lyse. Penicillin structure mimics amino acid bonds in peptide side chain. The enzyme (transpeptidase) which normally links individual peptidoglycan chains to one another binds irreversibly to penicillin. Typical animal cell Typical plant cell The endoplasmic reticulum (ER): is a system of membrane bound tubes and sacs called cisternae. cisternae It is divided into two types smooth and rough. rough Rough ER – appears rough due the bound ribosomes which are attached to the cytoplasmic surface of the membrane Ribosomes Rough ER Smooth ER The rough ER has two main functions 1. Synthesis of secretory proteins – for example insulin made by pancreatic cells 2. Membrane factory. Smooth ER – is so named because its membrane lacks ribosomes. Included in its functions are synthesis of lipids, metabolism of carbohydrates and detoxification of drugs and toxins. The Golgi apparatus – finishes, sorts and exports cell products. Many products made in the ER are transported to the Golgi apparatus via vesicles. It is a collection of flattened membranous sacs - which look like a stack or pitta breads. It also has a distinct polarity i.e. distinct sides. Relationship between ER and Golgi apparatus Rough ER Fig 7.16 Golgi apparatus Other membranous organelles Mitochondria (Mitochondrion) – they are found in nearly all cells – 1 to 1000s. It is enclosed in an envelop of two membranes each made of phospholipids. It is the site of cellular respiration where they extract energy from sugars, fats and other fuels with the help of oxygen to generate ATP. Contain their own DNA and ribosomes. Chloroplasts – Only found in plants these organelles are the site of photosynthesis – where light energy is converted into chemical energy used to synthesis organic compounds from CO2 and H20. They also contain their own DNA and ribosomes.