Chapter 2: Bacterial genetics, metabolism, and structure

Bacterial metabolism

1. Aerobic respiration includes: -Glycolysis (aka Embden-Meyerhof-Parnas pathway) -Krebs cycle -Electron transport chain with O2 electron acceptor 2. Anaerobic respiration includes: -Glycolysis -Krebs cycle -Electron transport chain without O2 electron acceptor 3. Fermentation -Just includes glycolysis

Microbial genetics

1. Bacteria: -Single, circular, ds DNA chromosome -DNA located in the nucleoid region (as opposed to inside a nucleus 2. Fungi and parasites -Linear ds DNA chromosome -Chromosome found inside the nucleus 3. Viruses -DNA or RNA found inside a protein coat (capsid) 4. Extrachromosomal DNA -Plasmids -Transposons

Acid-fast (AF) cell envelope

1. Cell wall with inconsistent gram stain results -Basic structure more closely akin to GP organisms (thick layer of peptidoglycan) 2. 60% of the wall is composed of a waxy layer of glycolipids and fatty acids (specifically mycolic acid) on the outside of the cell wall 3. Seen in Mycobacterium and Nocardia spp.

Bacterial cell components

1. Cytosol 2. Polysome (mRNA complexed with 50s ribosome) 3. Inclusions -Glycogen and phosphate reserves 4. Nucleoid region (containing the chromosome) 5. Plasmids 6. Endospores (produced under adverse conditions)

Transposons

1. Extrachromosomal DNA 2. Pieces of DNA that may be transferred from one location to another within the chromosomes or between chromosomes (?) 3. May be simple or composit

Plasmids

1. Extrachromosomal piece of DNA (usually circular) 2. Contains several genes that control its replication, and may code for specific protein products

Acridine orange

1. Fluorochrome die used to evaluate levels of both living and dead bacteria -Used to identify low levels of bacteria, such as in the blood or CSF 2. Stains both GP and GN organisms, whether living or dead (the gram stain only stains living bacteria) 3. The dye binds to the nucleic acids in the cell and fluoresces bright orange

Calcofluor white

1. Fluorochrome dye 2. Binds to chitin in fungal cell walls 3. Fluoresces bright green or blue-white

Cell exterior and appendages

1. Glycocalyx -Includes the capsule and slime layers 2. Biofilms -Bacterial colonies in an extracelular polymeric substance -May have one or many bacterial species 3. Appendages: -Fimbriae (adhesins) -Pili (used for adhesion or conjugation -Flagella (motility)

Morphological characteristics

1. Gram stain -Based on differences in the cell wall -May be gram positive (purple) or negative (pink) 2. Shapes: -Cocci -Coccobacilli (oval shaped) -Bacillus -Fusiform (pointed end) -Curved bacilli -Spiral 3. Arrangements -Single -Pairs -Chains -Clusters -Other (like tetrads)

Gram negative cell envelope

1. Inner and outer membranes -Porins in the membranes allow transport of nutrients inside the cell wall and may also be involved in antimicrobial resistance 2. Thin peptidoglycan layer between the membranes -Periplasmic space between the cell membrane and outer layer is important to antibiotic resistance (antibiotics may be broken down in this area?) 3. The outer membrane contains proteins, phospholipids, and lipopolysaccharides (LPS) -LPS has three regions -Antigenic region (has O-specific polysaccharides?) -Inner lipid A (acts as endotoxin)

Kinyoun stain

1. Method used for the acid-fast stain 2. Uses a detergent to treat the cell wall, allowing the stain to penetrate

Ziehl-Neelsen stain

1. Method used for the acid-fast stain 2. Uses heat to treat the cell wall, allowing the stain to penetrate

India ink

1. Negative stain (stains the background instead of the microorganism 2. Used to visualize the capsules of certain yeasts like Cryptococcus neoformans -Dark background with a halo around the yeast

Peptidoglycan

1. Polysaccharide found in bacterial cell walls 2. Formation of this polysaccharide is targeted by antibiotics like penicillin 3. Penicillin binding proteins confer resistance by preventing penicillin from blocking polysaccharide formation -Includes transpeptidase and D alanine carboxypeptidase

Gram stain

1. Positive stain purple, negative stain red/pink 2. Staining process: -Crystal violet (primary stain, 1 minute) -Iodine (mordant/fixative, 1 minute) -Alcohol (decolorizer; 6-7 drops, rinse immediately) -Safranin (secondary stain/counterstain; 2 minutes)

No cell envelope

1. Some species of bacteria have no cell wall -Includes *Mycoplasma and Ureaplasma spp* -Have sterols in the cell membrane -Shape of cells vary under the microscope 2. Some bacteria (both GP and GN) may lose their cell envelopes -This is referred to as the L-form -May grow on media supplemented with serum or sugar (?) -Helps the bacteria to escape antibiotics attacking the cell walls

Methylene blue

1. Stain used for bacteria 2. May be used as the counterstain for the acid fast stain 3. Or, traditionally used to stain Corynebacterium diphtheriae -Shows the metachromatic granules (gives the bacteria a "beaded look")

Endospore stain

1. Stains these structures that are only found in gram positive bacilli 2. Staining process -Malachite green (primary stain) -Water (decolorizer) -Safranin (counterstain) 3. Results: -These structures are green while bacterial cells are pink/red -Location of these structures within the bacteria can be used to identify the bacteria

Acid-Fast stain

1. The cell wall must first be treated so that the stain can actually penetrate -Ziehl-Neelsen method → heat -Kinyoun method → detergent 2. Staining process -Carbolfuchsin (primary stain, red) -Acidified alcohol (decolorized) -Methylene blue (secondary stain, blue 3. Positive stains red, negative stains blue

Inner lipid A

1. The portion of the LPSs on the GN cell envelope that acts as an endotoxin 2. Results in fever and shock, which may result to DIC in the case of septicemia

Gram positive cell envelope

1. Thick layer of peptidoglycan 2. Also contains teichoic acids and lipoteichoic acid (act as antigens and help give the cell its polarity)

Conjugation

1. Type of bacteria genetic exchange 2. DNA exchanged between living cells 3. Requires F factor -The powerpoint implies that the sex pilus is what's doing the transferring, but I'm pretty certain Dr. B said that it's just used for attachment. I can't find that anywhere else, though.

Transduction

1. Type of bacteria genetic exchange 2. DNA from one bacteria gets transported to another bacteria by a bacteriophage

Transformation

1. Type of bacteria genetic exchange 2. Free DNA fragments in the environment are taken up by competent cells

String test

1. Used to confirm GN organisms 2. Process: -Add 3% KOH -GN cells lyse, which results in the release of DNA, making the solution stringy and viscous

Recombinant DNA (in bacteria)

DNA from a donor bacterium gets inserted into a recipient bacterium

Cell envelope

May contain: 1. Outer membrane (in GN bacteria) 2. Cell wall (aka murein layer) -In GP bacteria, thick layer of peptidoglycan (the murein layer) -In GN bacteria, thin layer of peptidoglycan -In acid fact (AF) bacteria, contains mycolic acids (waxy substances) 3. Periplasmic space (GN bacteria) -Contains gel-like substance to capture nutrients and contains enzymes 4. Cell membrane -Osmotic barrier -Contains proteins and enzymes for metabolism

Lactophenol cotton blue

Stain used for medically important fungi