Unit 1.2 - Antibiotic Treatment Antibiotic Resistance The mass use of antibiotics has resulted in new strains of bacteria that are resistant to antibiotics Over the last decade, almost every type of bacteria known to cause disease has become stronger and less responsive to antibiotic treatment Increasing number of antibiotic resistant bacteria: Tuberculosis, gonorrhea, malaria, skin infections, pneumonia, & ear infections Antibiotic resistance has been called one of the world’s most pressing public health problems and is one of the Centers for Disease Control’s top concerns.

Unit 1.2 - Antibiotic Treatment Mechanism of Action Specific antibiotics are effective at preventing the growth of certain strains of bacteria The effectiveness of antibiotics is dependent on the mechanism of action of the drug and the structure of the bacteria

Unit 1.2 - Antibiotic Treatment KEY TERMS Crossword Puzzle

Unit 1.2 - Antibiotic Treatment

Unit 1.2 - Antibiotic Treatment Activity 1.2.3 Introduction The purpose of this activity is for you to review and build upon your knowledge of bacterial cells. In the last lesson, you were introduced to 18-year-old Sue Smith, who was diagnosed with bacterial meningitis. In this activity, you will learn about the structure of a bacterial cell in order to understand how different classes of antibiotics work. Your goal is to put all the information together to determine the best type of antibiotic to use to treat Sue Smith.    

Unit 1.2 - Antibiotic Treatment What is a bacterium? What are the types of bacteria? How do antibiotics kill bacteria?

Unit 1.2 - Antibiotic Treatment What is a bacterium? Bacteria are a large group of unicellular, prokaryote, microorganisms Typically a few micrometers in length A wide range of shapes (ranging from spheres to rods and spirals) 40 million bacterial cells in a gram of soil A million bacterial cells in a milliliter of fresh water Approximately five nonillion (5×1030) bacteria on Earth

Unit 1.2 - Antibiotic Treatment A Typical Bacterium

Unit 1.2 - Antibiotic Treatment Bacterial DNA Chromosomal: bacteria possess a single chromosome composed of double‐stranded DNA in a closed loop. The DNA is located in the nucleoid of the cell and is not associated with protein. Transfer of chromosomal DNA is accomplished through replication then division to daughter cells. Plasmid: A small circular double-stranded DNA molecule that carries accessory genes separate from those of the bacterial chromosome. They carry a small number of genes. Easily transferred between bacteria in cellular contact.

Unit 1.2 - Antibiotic Treatment Bacterial Gene Transfer Three primary types of gene transfer between bacterial cells Bacterial conjugation is the transfer of genetic material (plasmid) between bacterial cells by direct cell-to-cell contact or by a bridge-like connection between two cells Transduction is the process by which DNA is transferred from one bacterium to another by a virus. It also refers to the process whereby foreign DNA is introduced into another cell via a viral vector. Transformation is the genetic alteration of a cell resulting from the direct uptake and incorporation of exogenous genetic material (exogenous DNA) from its surroundings and taken up through the cell membrane(s).

Unit 1.2 - Antibiotic Treatment Gram Staining Gram-negative bacteria Thin cell wall consisting of a few layers of peptidoglycan surrounded by a second lipid membrane containing lipopolysaccharides and lipoproteins Do not retain crystal violet dye in the Gram staining protocol due to their lipopolysaccharide outer layer In a Gram stain test, a counterstain (commonly safranin) is added after the crystal violet, coloring all Gram-negative bacteria with a red or pink color. Gram-positive bacteria Do not have an outer membrane BUT they have a thickened peptidoglycan layer Will retain the crystal violet dye when washed in a decolorizing solution

Unit 1.2 - Antibiotic Treatment Gram Staining The higher lipid content of the gram-negative bacteria cell walls allows the alcohol destain to wash the purple colored stain out of the cells. Gram positive bacteria will retain the purple color because the alcohol is not able to destain the purple color from the cells. Gram negative bacteria stain reddish-pink Gram positive bacteria stain purple “Positively Purple”

Unit 1.2 Antibiotic Treatment Gram Staining Gram Negative Gram Positive

Unit 1.2 Antibiotic Treatment Gram Staining

Unit 1.2 - Antibiotic Treatment Mechanism of Action Specific antibiotics are effective at preventing the growth of certain strains of bacteria The effectiveness of antibiotics is dependent on the mechanism of action of the drug and the structure of the bacteria

Unit 1.2 - Antibiotic Treatment How do antibiotics work? A number of bacterial processes, including the synthesis of bacterial cell walls, proteins, metabolic pathways, and the integrity of the cytoplasmic membrane, are the targets of most antibacterial drugs. 4 Main Classes of Antibiotics: β-Lactam Antibiotics Tetracyclines Fluoroquinolones Sulfonamides

Unit 1.2 - Antibiotic Treatment β-Lactam Antibiotics “Blocks cell wall synthesis” Irreversibly inhibit enzymes involved in the final steps of cell wall synthesis. These drugs vary in their spectrum of activity; some are more active against Gram positive bacteria; whereas, others are more active against Gram negative bacteria. Tetracyclines “Blocks protein synthesis” Reversibly bind to the 30S ribosomal subunit, blocking the attachment of tRNA to the ribosome and preventing the continuation of protein synthesis. They are effective against certain Gram positive and Gram negative bacteria.

Unit 1.2 - Antibiotic Treatment Fluoroquinolones “Disrupts bacterial DNA organization” Inhibit one or more of a group of enzymes called topoisomerases, which maintain the supercoiling of the chromosomal DNA within the bacterial cells. The inhibition of these enzymes prevents essential cell processes. The fluoroquinolones are active against a wide variety of bacteria, including both Gram positive and Gram negative. Sulfonamides “Disrupts metabolic pathways” Inhibit the growth of many Gram positive and Gram negative bacteria. They are structurally similar to paraminobenzoic acid (PABA), a substrate in the pathway for folic acid biosynthesis. Because of this similarity, the enzyme that normally binds with PABA preferentially binds with the sulfonamide drugs, resulting in its competitive inhibition. Human cells are not affected by these drugs because they lack this enzyme.

Unit 1.2 Antibiotic Treatment Antibiotic Resistance It may take the form of a spontaneous or induced genetic mutation, or the acquisition of resistance genes from other bacterial species by horizontal gene transfer via conjugation, transduction, or transformation. Many antibiotic resistance genes reside on transmissible plasmids, facilitating their transfer via conjugation.

Unit 1.2 Antibiotic Treatment Unit 1.2.4.A – When Antibiotics Fail Exposure to an antibiotic naturally selects for the survival of the organisms with the genes for resistance. In this way, a gene for antibiotic resistance may readily spread through an ecosystem of bacteria. Antibiotic-resistance plasmids frequently contain genes conferring resistance to several different antibiotics. What does it mean by select? After completing the conclusion questions please address this in the form of a paragraph in your notebook (diagrams welcomed)

Unit 1.2.4.A – When Antibiotics Fail Begin Unit 1.2.4.A – When Antibiotics Fail

Unit 1.2 - Antibiotic Treatment Mechanism of Action Specific antibiotics are effective at preventing the growth of certain strains of bacteria. The effectiveness of antibiotics is dependent on the mechanism of action of the drug and the structure of the bacteria. In the last lesson, students were introduced to 18-year-old Sue Smith, who was diagnosed with bacterial meningitis. In this lesson, students will review bacterial structure, investigate various types of antibiotics and their mode of action against the bacteria they target, and suggest an antibiotic treatment for Sue Smith. Students will then investigate the mechanisms by which DNA from one bacterial cell is transferred to another bacterial cell and explore antibiotic resistance.

Review Unit 1.2 –Antibiotics

Quiz Review The one-way transfer of DNA between bacteria in cellular contact (1 pt) Transformation Replication Conjugation Transduction The gel-like region within the cytoplasm containing the single, circular, double-stranded DNA molecule? (1pt) Nucleoid Pili Plasma Membrane Organelle

Quiz Review True or False: Gram negative bacteria will stain purple? (1 pt) Match the correct term to the structure shown below. (3 pts)   _______ Cell Wall _______ Flagellum _______ Cell membrane _______ Nucleoid _______ Pilus _______ Ribosome Positively Purple!!! B E A D F C

Quiz Review For Project 1.2.3.P ("Superbugs") we have been working with 2 strains of bacteria. The first strain, E. coli Strain I, contains a gene found on the chromosomal DNA coding for streptomycin resistance. The second strain, E. coli Strain II, contains a gene found on the plasmid DNA coding for ampicillin resistance. On the back of this page, answer the following question based on the what you have learned about gene transfer and bacterial resistance. If your results indicate that you did create a superbug with both streptomycin and ampicillin resistance, was the streptomycin resistant gene transferred from Strain I to Strain II, or was the ampicillin resistant gene transferred from Strain II to Strain I? You must explain in detail your reasoning for full credit. (4 pts) The ampicillin resistant gene on the plasmid DNA is transferred from Strain II to Strain I. The mechanism leading to this gene transfer is conjugation.

Quiz Review b d c a β-Lactam Antibiotics ______ Fluoroquinolones ______ Sulfonamides _______ Tetracyclines _______ d c a “Blocks protein synthesis” “Blocks cell wall synthesis” “Disrupts metabolic pathways” “Disrupts bacterial DNA organization”

Unit 1.2 - Antibiotic Treatment Understandings Antibiotics disrupt the pathways that bacteria use to survive. Bacterial cells use multiple pathways to gain resistance to antibiotics. Overuse and misuse of antibiotics promotes the selection of resistant bacteria. Knowledge and Skills It is expected that you will: Label the structures of a bacterial cell. Explain the method of action for different classes of antibiotics. Describe the pathways through which bacterial cells transfer genes. Explain the importance of taking antibiotics as prescribed. Use proper laboratory techniques to “mate” a streptomycin resistant strain of E. coli with an ampicillin resistant strain of E. coli. Simulate the effects of antibiotics on a bacterial population during an infection. Simulate the effect of a missed dose of antibiotics on a bacterial population during an infection.

Unit 1.2 - Antibiotic Treatment Unit 1.2 - Essential Questions How do antibiotics work to fight bacterial infections? What methods do bacteria use to share antibiotic resistant genes? What actions are humans taking that are contributing to bacteria becoming resistant to commonly used antibiotics?