Microbiology exam 1 learning objectives

Explain how the structure of the bacterial cell wall relates to its Gram-stain status.

-Gram-positive bacteria retain the crystal violet stain because of their thicker cell wall., thick peptidoglycan layer, lack outer membrane, techoic acid Gram-negative bacteria do not, thin peptidoglycan layer in between 2 cell walls, periplasmic space between outer membrane and peptidogycan, porins, lipopolysaccharides, cell wall retains e coli/salmonella is gram - staph/strep (baccilus) is gram +

Identify bacterial morphologies (e.g., bacilli, cocci, and spirochetes). ESSAY

Cocci are spherical or oval cell (strep or staph and are gram + purple) Bacilli (from baculus meaning rod) are rod shaped cells (salmonella, e.coli, TYPICALLY GRAM -, ) 3. Vibrios are comma shaped curved rods and derive their name from their characteristics vibratory motility. 4. Spirilla are rigid spiral forms. 5. Spirochetes (from speira meaning coil and chaite meaning hair) are flexuous spiral forms(boriella, lyme disease)

Define essential nutrients, macronutrients, and micronutrients.

Essential nutrients are those that must be supplied from the environment §Macronutrients •Major elements in cell macromolecules -C, O, H, N, P, S •Cations necessary for protein function -Mg2+, Ca2+, Fe2+, K+ §Micronutrients •Trace elements necessary for enzyme function -Co, Cu, Mn, Zn

Explain the roles of the replisome, cell expansion, and septation in bacterial cell division.

replisome: At each fork, DNA is synthesized by DNA polymerase, with the help of accessory proteins. This protein complex is called the replisome cell expansion: cells elongate which septation occurs septation:The septum grows inward from the sides of the cell, at last constricting and sealing off the two daughter cells. FtsZ subunit assembly circles around the septum in a "treadmilling" pattern, stepwise around the cell, that directs septal growth. Septation requires rapid biosynthesis of all envelope components, including membranes and cell wall. Coordinated with DNA replication

Compare the cell envelopes of firmicutes, proteobacteria, and mycobacteria.

Gram-positive bacteria (thick cell wall) -Example: the phylum Firmicutes §Has multiple layers of peptidoglycan •Threaded by teichoic acids (shown here) §The capsule •Made of polysaccharide and glycoprotein •Protects cells from phagocytosis •Found in some Gram-positive (and some Gram-negative) cells •Gram-negative bacteria (thin cell wall) -Example: the phylum Proteobacteria •Covered by an outer membrane, which confers defensive abilities and toxigenic properties on many pathogens -Inward-facing leaflet including lipoprotein -Outward-facing leaflet containing lipopolysaccharides -Contains transmembrane proteins called porins •Mycobacteria (complex, multilayered cell wall) -Example: the phylum Actinobacteria •Include unusual membrane lipids (mycolic acids) and unusual sugars (arabinogalactans) §The mycobacterial envelope may be 103-fold less permeable to nutrients and toxins than that of E. coli. •Thus, mycobacteria must grow slowly—but they effectively resist host defenses.

Recall the structures and functions of thylakoids, carboxysomes, storage granules, pili, stalks, nanotubes, and flagella.

thylakoids: extensively folded intracellular membrane in photosynthetic bacteria carboxysomes: polyhedral bodiees packed with enzyme, rubisco for CO2 fixation storage granules:•Glycogen •Other polymers, such as polyhydroxybutyrate (PHB) and poly-3-hydroxyalkanoate (PHA) -For energy •Sulfur globules -Either inside or outside the cell -For oxidation pills: Pili (also called fimbriae) are straight filaments of pilin protein. •Used in attachment stalks:Stalks are membrane-embedded extensions of the cytoplasm. nanotubes: membrane extensions that merge directly with the membranes of neighboring organisms. flagella: Each flagellum is a spiral filament of protein monomers called flagellin. •The filament is rotated by a motor driven by the proton motive force. -The motor possesses an axle and rotary parts, all composed of specific proteins. oMotB - Forms part of the proton channel oFliG - Forms part of device that generates torque Note: flagella rotate either clockwise (CW) or counterclockwise (CCW) relative to the cell

Describe the stages of biofilm development.

twitching motility->quorem sensing->microcolony->exopolysaccharide formation

list examples of microbes

viruses, bacteria, archaea, eukarya; prochlorococcus, e.coli,spirogyra, pelomxya, halococcus

Recall the forms in which cells store energy.

§A membrane potential is generated when chemical energy is used to pump protons outside of the cell. §The H+ gradient plus the charge difference form an electrochemical potential called the proton motive force. §The potential energy stored can be used to transport nutrients, drive flagellar rotation, and make ATP by the F1FO ATP synthase.

Classify stains as simple or differential.

§A simple stain adds dark color specifically to cells, but not to the external medium or surrounding tissue. •Methylene blue is the most commonly used stain. §A differential stain stains one kind of cell but not another. •The most famous differential stain is the Gram stain. Acid-fast stain: carbolfuchsin used to stain Mycobacterium species §Mycolic acids in cell walls unique to this clade take the stain §Spore stain: malachite green used to detect spores of Bacillus and Clostridium §Stain is specific to components of the endospore coat §Negative stain: colors the background, which makes capsules more visible

State the structure and function of cyanobacterial heterocysts.

§Anabaena differentiates into specialized cells called heterocysts. §Allows it to fix nitrogen anaerobically while maintaining oxygenic photosynthesis They fix nitrogen from dinitrogen (N₂) in the air using the enzyme nitrogenase, in order to provide the cells in the filament with nitrogen for biosynthesis.

Explain how chemoreceptors and magnetosomes direct flagellar motility.

§Attractants cause CCW rotation. •Flagella bundle together •Push cell forward •"Run" §Repellents (or absence of attractants) cause CW rotation. •Flagellar bundle falls apart. •"Tumble" -Bacterium briefly stops, then changes direction. Chemoreceptors receive signals and transmit them to the flagellar motor monotrichous-cells single flagella lophotrichous-cells have flagella at ends peritrichous-flagella randomly around

Explain how lysis and ultracentrifugation contribute to cell fractionation.

§Cell fractionation is a method used to separate cellular components such as membranes, ribosomes, and flagella. •This requires techniques that lyse (break open) the cell. -The lysis method must generate enough force to separate the membrane lipids but not enough to disintegrate complexes of protein and RNA. §A key tool of subcellular fractionation is the ultracentrifuge. •The high rotation rate produces centrifugal forces strong enough to separate particles by size. •Particles are collected in fractions of sample from the tube— and are then observed by electron microscopy

Differentiate among complex, synthetic, selective, and defined media ESSAY

§Complex media are nutrient rich but poorly defined (gram - and positive, staph and e coli). §Minimal defined media contain only those nutrients that are essential for growth of a given microbe. (gram -, e coli) §Enriched media are complex media to which specific blood components are added. §Selective media favor the growth of one organism over another. §Differential media exploit differences between two species that grow equally well.

Categorize different types of active transport.

§Coupled transport systems are those in which energy released by a driving ion moving down its gradient is used to move a solute up its gradient. •In symport, the two molecules travel in the same direction. In antiport, the actively transported molecule moves in the direction opposite to the driving ion §The largest family of energy-driven transport systems is the ATP-binding cassette superfamily, or ABC transporters. •They are found in all three domains of life. §Two main types: •Uptake ABC transporters are critical for transporting nutrients. •Efflux ABC transporters are generally used as multidrug efflux pumps.

Compare and contrast batch culture and continuous culture.

§In a continuous culture, all cells in a population achieve a steady state, which allows detailed study of bacterial physiology.The complex relationships among dilution rate, cell mass, and generation time in a chemostat are illustrated here (parabolic shape) §Exponential growth never lasts indefinitely. §The simplest way to model the effects of a changing environment is to culture bacteria in a batch culture.

Define biofilm.

§In nature, many bacteria form specialized, surface-attached communities called biofilms. •These can be constructed by one or multiple species and can form on a range of organic or inorganic surfaces.

Explain why some microbes require growth factors for culture and why some cannot be grown in pure culture.

§Microbes can evolve to require specific growth factors depending on the nutrient richness of their natural ecological niche. §Growth factors are specific nutrients not required by other species. •A microbe needs them in order to be able to grow in laboratory media. •e.g., Streptococcus pyogenes requires glutamic acid and alanine because it can no longer synthesize them. •Some of these uncultured organisms depend on factors provided by other species that cohabit their niche.-codependent cannot grow alone Rickettsia prowazekii, the cause of epidemic typhus fever, has adapted to grow within the cytoplasm of eukaryotic cells and nowhere else.

Compare and contrast different types of direct and indirect microbial counts.

§Microorganisms can be counted directly by placing dilutions on a special microscope slide called a Petroff-Hausser counting chamber. §Living cells may be distinguished from dead cells by fluorescence microscopy using fluorescent chemical dyes. •Dead bacterial cells fluoresce orange or yellow because propidium (red) can enter the cells and intercalate the base pairs of DNA. •Live cells fluoresce green because Syto-9 (green) enters the cell. §The instrument is called a fluorescence-activated cell sorter (FACS) or flow cytometer. •Viable cells can be counted via the pour plate method. §Microorganisms can be counted indirectly via biochemical assays of cell mass, protein content, or metabolic rate. •Also, by measuring optical density

. State the components of the bacterial cell envelope.

§Most bacteria share fundamental traits: •Thick, complex outer envelope •Compact genome •Tightly coordinated cell functions §Archaea, like bacteria, are prokaryotes: •Have unique membrane and envelope structures ell wall, cell membrane and an outer membrane. The cell envelope of gram-positive bacteria comprises cell wall, cytoplasmic membrane and capsule. The cell envelope of gram-negative bacteria comprises cell wall, cytoplasmic membrane, outer membrane, periplasmic space and capsule. archaea have ether-terpenoid chain bacteria (saccule, mesh like structure) and eukarya have esther bonds for bacteria- NAM and NAG is how cross bridging occurs to create mesh like structure in peptidoglycan

Recall how microbes obtain iron.

§Siderophores are specialized molecules secreted to bind ferric ion (Fe3+) and transport it into the cell. •The iron is released into the cytoplasm and reduced to the more useful ferrous (Fe2+) form. §Note: Neisseria gonorrhoeae employs receptors on its surface that bind human iron complexes and wrest the iron from them.

outline the structure and function of bacterial intercellular nanotubes

§Some bacteria and archaea can form membrane extensions that merge directly with the membranes of neighboring organisms. These nanotubes allow bacteria to directly share proteins and mRNA useful under hostile conditions, such as when exposed to antibiotics ex:baccilus subtilis, just a membrane

Compute the generation time, growth rate constant, or number of cells present given the appropriate starting information.

§The growth rate, or rate of increase in cell numbers or biomass, is proportional to the population size at a given time. •Such a growth rate is called "exponential" because it generates an exponential curve, a curve whose slope increases continually. •If a cell divides by binary fission, the number of cells is proportional to 2^n. -Where n = number of generations §Generation time is the time it takes for a population to double. §For cells undergoing binary fission, Nt = N0 x 2n where Nt is the final cell number, N0 is the original cell number, and n is the number of generations.

definition of a microbe

a microbe is a living organism that requires a microscope to be seen. All three domains of life contain some microbial organisms viruses do not adhere to this definition (non-living), but are considered microbes Each microbe contains in its genome the capacity to reproduce its own kind.

Explain why the microscope is an important tool in the field of microbiology.

allows us to see microbes: robert hooke- first microscope to view mold, termed cell; leuwenhook-single lense magnifiers, first to see microbes (animacules);

Explain how limiting nutrient supplies limits bacterial growth

bacteria need certain nutrients to grow, All of Earth's life-forms are based on carbon; •Phototrophs obtain energy from chemical reactions triggered by light. •Chemotrophs obtain energy from oxidation-reduction reactions. •Lithotrophs use inorganic molecules as a source of electrons. Organotrophs use organic molecules

State some uses of microbes in medicine and industry.

experimental: medical: epidemiology: immunology: food: industrial: production of drugs environemntal:

Recall cell membrane structure.

hoponoid, phsospholipid, ATP synthase, protein transporter, cytoplasm §A phospholipid consists of glycerol with ester links to two fatty acids and a phosphoryl head group. The two layers of phospholipids in the bilayer are called leaflets §Membrane proteins often contain both hydrophilic and hydrophobic regions, allowing them to be inserted into the cell membrane. •In eukaryotic membranes, the reinforcing agents are sterols, such as cholesterol. •In bacteria, the same function is served by hopanoids, or hopanes. §Archaea have the most extreme variations in phospholipid side-chain structures. •Ether links between glycerol and fatty acids •Diethers or tetraethers •Hydrocarbon chains are branched terpenoids. •Can cyclize to form cyclopentane The most common structural support is the cell wall

Compare the roles of immunization, antiseptics, and antibiotics in human disease treatment and prevention.

immunization:the stimulation of an immune response by deliberate inoculation with an attenuated pathogen. antiseptic: chlorine, carbolic acid to treat wounds and clean surgical instruments. antibiotic: Penicillin quickly became the first commercial antibiotic used to save human lives.Alexander Fleming discovered that Penicillium mold generated a substance that kills bacteria.

List some functions of biofilms.

increases the cells' survival through improved defense against antibiotics, increased availability of nutrients, food and protection

List and describe the four stages of bacterial growth in batch culture.

lag (adjusting to new environment before cell growth)->log(growth of bacterial pops)->stationary(not growing anymore, plateau)->death

Recall the advantages and disadvantages of a wet mount.

-Advantages: oObservation of cells in natural state -Disadvantages: oLittle contrast between cell and background oSample may dry out quickly.

Describe spore formation.

1. septum forms and dna replicates to axial filament 2.septum separates forespore 3. mother engulfs forespore 4.chromosomes of mother cell disinegrate 5.forespore develops cortex layer of peptioglycan 6.dipliconic acid synthesized and calcium in spore coat 7. mother cell releases spore

Describe the different methods of obtaining single bacterial colonies.

1.Liquid or broth -Useful for studying the growth characteristics of a pure culture 1.Dilution streaking -A loop is dragged across the surface of an agar plate. 2.Spread plate -Tenfold serial dilutions are performed on a liquid culture. A small amount of each dilution is then plated

List Koch's postulates.

1.Suspected microbe is always present in diseased hosts. -absent in healthy hosts 2.Suspected microbe is grown in pure culture outside hosts. -no other microbes present in culture 3.Cultured microbe is introduced into healthy hosts. -Individuals become sick with same disease as original hosts. 4. Same microbial suspect is re-isolated from sick individuals

State the importance of DNA sequencing in classification.

16s rRNA helped show linkage between bacteria, archea, and eukarya, archaea and eukarya are more closely related

Recall the role of the ultracentrifuge in advancing our knowledge of cell structure and function.

Enabled separation of subcellular parts

Explain the functions of the components of a compound microscope.

Eyepiece: The lens the viewer looks through to see the specimen. The eyepiece usually contains a 10X or 15X power lens. Diopter Adjustment: Useful as a means to change focus on one eyepiece so as to correct for any difference in vision between your two eyes. Body tube (Head): The body tube connects the eyepiece to the objective lenses. Arm: The arm connects the body tube to the base of the microscope. Coarse adjustment: Brings the specimen into general focus. Fine adjustment: Fine tunes the focus and increases the detail of the specimen. Nosepiece: A rotating turret that houses the objective lenses. The viewer spins the nosepiece to select different objective lenses. Objective lenses: One of the most important parts of a compound microscope, as they are the lenses closest to the specimen. A standard microscope has three, four, or five objective lenses that range in power from 4X to 100X. When focusing the microscope, be careful that the objective lens doesn't touch the slide, as it could break the slide and destroy the specimen. Specimen or slide: The specimen is the object being examined. Most specimens are mounted on slides, flat rectangles of thin glass. The specimen is placed on the glass and a cover slip is placed over the specimen. This allows the slide to be easily inserted or removed from the microscope. It also allows the specimen to be labeled, transported, and stored without damage. Stage: The flat platform where the slide is placed. Stage clips: Metal clips that hold the slide in place. Stage height adjustment (Stage Control): These knobs move the stage left and right or up and down. Aperture: The hole in the middle of the stage that allows light from the illuminator to reach the specimen. On/off switch: This switch on the base of the microscope turns the illuminator off and on. Illumination: The light source for a microscope. Older microscopes used mirrors to reflect light from an external source up through the bottom of the stage; however, most microscopes now use a low-voltage bulb. Iris diaphragm: Adjusts the amount of light that reaches the specimen. Condenser: Gathers and focuses light from the illuminator onto the specimen being viewed.

Explain the role of microbes in geochemical cycling, especially that of nitrogen.

Geochemical cycling is the global conversion of inorganic and organic forms of nitrogen, sulfur, and phosphorus, and other minerals. Nitrifiers turn ammonia NH4 to nitrite NO2. Nitrogen fixation is taking N2 and reducing it to ammonia NH4.

Recall the contributions of various individuals to the discovery and implementation of vaccination.

Lady Mary Montagu introduced the practice of smallpox inoculation to Europe in 1717. Edward Jenner (1749-1823) deliberately infected patients with material he collected from cowpox lesions. •The practice of cowpox inoculation was called vaccination (from Latin vacca for "cow"). Louis Pasteur developed the first vaccines based on attenuated (weakened) strains of microbes. •Fowl cholera •Rabies

Appraise endosymbiosis as an explanation for mitochondria and chloroplasts.

Lynn Margulis (1938-2011). •She proposed that eukaryotic organelles, such as mitochondria and chloroplasts, evolved by endosymbiosis from prokaryotic cells engulfed by pre-eukaryotes. The endosymbiosis theory was highly controversial. •It implied a polyphyletic ancestry of living species, instead of the long-held assumption that species evolve only by divergence from a common ancestor (monophyletic ancestry).

Compare the roles of animal endosymbionts and plant endosymbionts.

Plants have endosymbionts to form special nodules to fix nitrogen into biomass in their roots which is shared with plant cells. Animal endosymbionts have biofilms of bacteria to help the digestive tract.

Compare and contrast Spallanzani's, Pasteur's, and Tyndall's experiments that tested spontaneous generation.

Spallanzani: Showed that microbes arise from preexisting microbes and demonstrated that heat sterilization can prevent microbial growthPasteur: Providing oxygen does not enable spontaneous generation Tyndall: Showed the repeated cycles of heat were necessary to eliminate spores formed by certain kinds of bacteria

Recall the properties of spores.

Starvation initiates an elaborate 8-hour genetic program that involves: •An asymmetrical cell division process that produces a forespore and ultimately an endospore §Sporulation can be divided into discrete stages based primarily on morphological appearance.

Outline how microbial classification has changed over time.

The famous classifier of species, Swedish botanist Carolus Linnaeus (1707-1778), called the microbial world "chaos." See the Ring of Life hypothesis 2004 by James LakeAnalysis of archea 16S rRNA revealed that these prokaryotes were a distinct form of life. §Woese's discovery replaced the classification scheme of five kingdoms with three equally distinct groups called domains. •Bacteria •Archaea •Eukarya In the three-domain model, the bacterial ancestor of mitochondria derives from ancient proteobacteria, whereas chloroplasts derive from ancient cyanobacteria. issues: Resolution of the light microscope was too low Microbial species are hard to define

Explain how studies of microbes fostered our knowledge of DNA function and enhanced DNA technology.

microbes are simplized oraganisms so its easy to see thei cell structures §In 1928, Frederick Griffith discovered transformation in bacteria. §In 1944, Oswald Avery and colleagues showed that the transforming substance is DNA. §In 1953, Rosalind Franklin used X-ray crystallography to determine that DNA is a double helix. §Later that year, James Watson and Francis Crick discovered the complementary bases and antiparallel nature of DNA. §The promise of DNA was first fulfilled in bacteria and bacteriophages. •Bacteria readily recombine DNA from unrelated organisms. -Recombinant DNA ultimately enabled us to transfer genes. •A heat-stable bacterial DNA polymerase was used for amplifying DNA via the polymerase chain reaction (PCR). •Gene regulation discovered in bacteria provided models for animals and plants.

identify the contributions of the following individuals: Nightingale, Hooke, van Leeuwenhoek, Pasteur, and Tyndall.

nightinggale: •Founder of modern nursing •She founded the science of medical statistics. •To show the deaths of soldiers due to various causes, she devised the "polar area chart." Hooke:•Built the first compound microscope •Used it to observe mold •Published Micrographia, the first manuscript that illustrated objects observed with a microscope •Coined the term "cell" Leewenhoek:•Built single-lens magnifiers, complete with sample holder and focus adjustment •First to observe single-celled microbes -He called them "animalcules" / "small animals." pasteur: •Began his scientific career as an organic chemist •Discovered the microbial basis of fermentation •Produced data that refuted spontaneous generation -Invented the "swan-neck" flask -Showed that a broth boiled in a swan-neck flask remained free of microbial growth, despite being exposed to air Tyndall: Showed the repeated cycles of heat were necessary to eliminate spores formed by certain kinds of bacteria

Compare and contrast autotrophs, heterotrophs, chemotrophs, and phototrophs.

•Autotrophs -Photoautotrophs -Chemolithoautotrophs (or lithotrophs) •Heterotrophs -Photoheterotrophs -Chemoheterotrophs (or organotrophs) •Phototrophs obtain energy from chemical reactions triggered by light. •Chemotrophs obtain energy from oxidation-reduction reactions. •Lithotrophs use inorganic molecules as a source of electrons. •Organotrophs use organic molecules.

Recall the structure and functions of the bacterial cell wall.

•Nevertheless, a few prokaryotes, such as the mycoplasmas, have a cell membrane with no outer layers. §The cell wall confers shape and rigidity to the cell and helps it withstand turgor pressure. § §The bacterial cell wall, or the sacculus, consists of a single interlinked molecule. •Like a flexible mesh bag or scaffold Most bacterial cell walls are made up of peptidoglycan •Long polymers of repeating disaccharides called N-acetylglucosamine and N-acetylmuramic acid bound to a peptide of four to six amino acids The peptides can form cross-bridges connecting the parallel glycan strands §Envelope composition defines: •Gram-positive bacteria (thick cell wall) -Example: the phylum Firmicutes •Gram-negative bacteria (thin cell wall) -Example: the phylum Proteobacteria •Mycobacteria (complex, multilayered cell wall) -Example: the phylum Actinobacteria

Compare and contrast prokaryotes and eukaryotes with respect to DNA replication, transcription, and translation.

•Unlike eukaryotes, prokaryotes synthesize RNA and proteins continually while the cell's DNA undergoes replication. Bacteria do not undergo mitosis or meiosis §Bacteria DNA synthesis terminates, the cell divides by a process called septation, the formation of the septum. eukaryotes have nucleus, In contrast, prokaryotic cells have a nucleoid region that extends throughout the cytoplasm and is not enclosed by a membrane. §In prokaryotes, translation is tightly coupled to transcription. The ribosomes bind to mRNA and begin translation even before the transcription of the mRNA strand is complete with replisome in prokaryotes, a circular chromosome begins to replicate at its origin, or ori site.