Introduction to Microbiology PDF

TOPIC 1: INTRODUCTION TO MICROBIOLOGY Types of Cells (Cellular Microbe) Prokaryotic Cells Eukaryotic Cells Scientific study of microorganisms No nucleus Nucleus Micro – too small to be see...

TOPIC 1: INTRODUCTION TO MICROBIOLOGY Types of Cells (Cellular Microbe) Prokaryotic Cells Eukaryotic Cells Scientific study of microorganisms No nucleus Nucleus Micro – too small to be seen with naked eye No organelles Organelles Bio – life Cell wall of If cell wall, cellulose or Logy – study of peptidoglycan chitin Binary Fission Mitosis 1 circular Linera chromosome chromosome Cellular Microorganism: Eukaryotes Fungi ▪ Unicellular form (yeast) – replicate asexually ▪ Filamentous form (mold) – replicate sexually ▪ Dimorphic – exist as yeast or mold o Molds (filaments) – 250C Acellular infectious agents (soil) Prions o Yeasts - 370C (in host ▪ Misfolded protein particles tissue) ▪ Neurodegenerative diseases o Histoplasma capsulatum Virus o Blastomyces dermatidis ▪ DNA or RNA o Paracoccidioides o Not both brasiliensis o Ds or ss o Coccidioides immitis ▪ Enveloped or non-enveloped o Penicillium marneffei o Sporothrix schenkii – (fav ni sir Roel). Also known as rose gardeners’ disease. Parasites ▪ Protozoa o Amoeba – Entamoeba hystolitica (most common amoeba) o Flagellates – hair like structure o Cilia – Balantidium coli (largest protozoan) o Sporozoa – pseudopods ▪ Helminths o Nematodes – Ascaris lumbricoides, Trichiuris trichiuria, Strongyloides (4 hookworms – Necator americanus, Ancylostoma braziliense, Ancylostoma caninum, Ancylostoma duodenale) o Cestodes o Trematodes Cellular Microorganism: Prokaryotes o S. aureus, S. pneumoniae, S. Archaea pyogenes ▪ Archaic – Greek word (Ancient) o Informal designation Bacteria o When referred to as a group ▪ 1-20 MM or larger o Names are neither capitalized nor ▪ Shapes (spheres, rods, spirals) underlined ▪ Spatial (single cells, chains, o staphylococci, streptococci cluster) Identification ▪ Binary fission ▪ Practical use of classification scheme to: ▪ Bacterial cell wall forms: Gram o Isolate and distinguish desirable Positive (foxie) (violet) (thick cell organisms from undesirable ones. wall) and Gram Negative (rod) (red) o Verify the authenticity or special (thin cell wall) properties of a culture in clinical settings Microbial Taxonomy o Isolate and identify the causative ▪ 3 distinct area agent of disease Classification o CONS – coagulase negative ▪ Categorization of organisms into staphylococci taxonomic groups (based on biochemical, Methods physiologic, genetic, and morphologic ▪ Genotypic characteristic (PCR machine) properties) DKPCOFGS ▪ Phenotypic characteristic ▪ Species (abbreviated as sp., singular, or o Macroscopic – visualized by naked spp., plural) eye o Most basic of the taxonomic o Microscopic groups/ collection of bacterial o Staining strains. o Environmental ▪ Subspecies o Resistance profiles o Subgroup within species o Antigenic profiles o Biotype o Subcellular profiles o Serotype o Genotype Other terminologies in Bacterial Taxonomy ▪ Genus (plural, genera) Biogroup – group of microorganisms that o Composed of various species with share the same biochemical properties. common characteristics Serogroup - group of microorganisms ▪ Family having similar antigens. o Composed of similar genera Epithet – proper word for the name of the species. Nomenclature Strain – altered or variant microorganisms ▪ Naming of an organism by international within the same species. rules according to its characteristics Morphovar/Morphotype – prokaryotic ▪ Established guidelines provided by the stain that differs morphologically from international code of Nomenclature of other stains. Bacteria or the Bacteriological Code Polyphasic taxonomy – modern system of o Genus designation – first letter is bacterial classification and identification always capitalized (phylogenetic, phenotypic, and genotypic o Species designation – first letter is characterization). always lower case o Italics or underlines ❖ HISTORY OF MICROBIOLOGY o Staphylococcus aureus, Anton Van Leeuwenhoek (1632-1723) Streptococcus pneumoniae, Considered as the first true Streptococcus pyogenes microbiologist o Abbreviated ▪ Father of Microbiology ▪ Father of Bacteriology  ▪ Father of Protozoology Rudolf Virchow First person to see live bacteria and Challenged the spontaneous generation protozoa theory with the concept of biogenesis. He observed and described microorganism such as bacteria and Louis Pasteur protozoa ads “animalcules” Disproved the Spontaneous generation ▪ Swan-neck flask analysis Proposed the use of heat in killing microorganisms. Developed the vaccine against anthrax and rabies Experiment of Franciso Redi (1668) Fermentation and Demonstrated that maggots could not pasteurization arise spontaneously from decaying meat. John Tyndall Tyndallization ▪ Method of sterilization in 1860 ▪ Bacterial spores can be destroyed ▪ Moist heat for 3 consecutive days ▪ 1000C for 30 minutes Ferdinand Cohn Experiment of John Needham (1745) Discovered that some bacteria could Boiled mutton broth using flask – withstand heating and boiling eventually became cloudy. ▪ Endospores Classification of bacteria into four groups based on shape Robert Koch Germ theory of disease (Koch’s postulate) Show evidence that bacteria can cause diseases Experiment of Lazzaro Spallanzani (1765) Discovered Bacillus anthracis and Improved the previous experiments of Mycobacterium tuberculosis Needham First to cultivate bacteria on boiled ▪ Heating the broth placed in a potatoes, meat extracts, and proteins sealed jar. Developed a culture medium ▪ Spallanzani’s result contradicted the findings of Needham Microbes move through the air and that they could be killed through boiling water. ❖ Collaborators of Koch Walter Hesse Paul Ehrlich Introduced the used of culture media Discovered the treatment of syphilis – Fanny Hesse Arsenic Suggested the use of agar, as ▪ Salvarsan (Arsphenamine) solidifying agent, in the preparation of Syphilis – bacterial infection usually culture media. spread by sexual contact. The disease Julius Richard Petri starts as painless – typically on your Developed the petri dish genitals, rectum or mouth. Martinus Beijerinck and Sergei Syphilis spreads from person to Winogradsky person via skin or mucus membrane Developed the enrichment-culture contact with these sores. technique and the sue of selective media. IMPORTANCE OF MICROBIOLOGY – BACTERIA Edward Jenner Importance of Bacteria in: Introduced the concept of vaccination ▪ Environment ▪ Smallpox vaccine ▪ Plants Coined the word vaccine from Latin ▪ Animals “vacca” for cow ▪ Agriculture ▪ Industry Ignaz Semmelweis ▪ Nitrogen cycle Demonstrated the routine hand ▪ Medicine washing can prevent the spread of CAREERS IN MICROBILOGY disease ▪ Bacteriology – bacteriologist ▪ Protozoology – protozoologist Joseph Lister ▪ Phycology – phycologist Introduced the system of antiseptic ▪ Parasitology – parasitologist surgery in Britain ▪ Mycology – mycologist Use the phenol as antimicrobial agent ▪ Virology – virologist for surgical sound testing DIAGNOSTIC MICROBIOLOGY Selman Waksman Quality of the specimen collection Discovered the streptomycin and from the patient neomycin antibiotics Techniques to demonstrate the Father of Antibiotics microbe in the sample Determine the antimicrobial activity Alexander Flemming Planning treatment Discovered the penicillin ▪ Virulence of the organism ▪ Penicillium chrysogenum ▪ Site of infection Discovered the lysozyme ▪ Patient’s ability to respond to the hospital-parasite Howard Florey interaction. They made the purification process for penicillin and the clinical trials to humans Edward Abraham He was the first to propose the correct biochemical structure of penicillin TOPIC 2: BASIC CELL STRUCTURE AND FUNCTION string test about PROKARYOTE - circular EUKARYOTE - linear 1mm. One circular Paired chromosomes in ▪ Biochemical chromosome. Not in a nuclear membrane. testing – gold membrane standard to No organelles organelles identify bacteria Peptidoglycan cell walls Polysaccharide cell Capsule – for protection walls ▪ Highly organized Binary fission Mitosis tightly attached No histones histones ▪ Gel firmly adherent to cell envelop. ▪ Streptococcus pneumonia – blood agar plate and able to produce a- hemolytic pattern ▪ 3 patterns: o a hemolysis – partial zone o B hemolysis – full zone o Y (gamma) hemolysis – no hemolysis at all ▪ Haemophilus influenzae ▪ Klebsiella 3 LEVELS TO CONSIDER BACTERIAL pneumoniae STRUCTURE b. Flagella (Bacterial Propellers) 1. EXTERNAL STRUCTURES – Appendages ▪ It is the organ of locomotion in (flagella, fimbriae, pili) and glycocalyx bacterial cell and consists of three a. Glycocalyx (Bacterial Surface Coating) parts ▪ Coating of molecules external to 1. Filament – this is long, thin, the cell wall and helical structure. Mostly ▪ Glycoprotein and glycolipid composed of proteins ▪ 2 types: 2. Hook – curved sheath that Slime layer – for protection, looks like mucoid transmit the torque generated ▪ Loosely organized by the basal body to the and attached filament. Responsible why ▪ Gel easily washed bacteria can rotate. off from cell 3. Basal body – stock of bricks, envelope. anchored to bacterial cells. ▪ Best example: Klebsiella pneumoniae – presumptive identification needs to perform  c. Fimbriae ▪ Fine, protein hair-like bristles from the cell surface. ▪ Functions in adhesion to other cells and surfaces. ▪ REMEMBER: use for adhesion to host cells and vital for colonization and infection. d. Pili ▪ It is hairlike structure composed of protein (pili) ▪ Two types (based on functions) Swarming motility – proteus spp. It has a. Common Pili – adherence to burned chocolate smell. cell services Virulence – able to cause infection to human. b. Sex Pili – transferring genetic material called conjugation Flagellar rearrangements 1. Atrichous – bacteria with no flagellum 2. Monotrichous – bacteria with single polar flagellum. (Vibrio cholerae) 3. Lopotrichous – bacteria with bunch of flagella at one pole. (Pseudomonas spp.) 4. Amphitrichous – bacteria with flagella at both poles. 5. Peritrichous – bacteria with flagella all over the surface. (Escherichia coli) Pili is shorter than flagella. Sex pili used for conjugation and flagella is used for attachment. Fimbriae are used as adhesion while pili is used for conjugation process. But fimbriae are shorter compared to pili that is longer but if we compare pili and flagella, flagella are longer. 2. Cell Envelop Cell wall (Boundary Layer of Bacteria) ▪ External covering outside the Endoflagella (Axial filament) cytoplasm ▪ It is the organ of motility found in ▪ Maintains cell integrity periplasmic space of spirochetes. ▪ Composed of Two basic layer ▪ Within the bacterial cell wall only a. Cell wall (inside) ▪ Multilayered structure composed of N-acetyl Muramic acid and N- acetyl Glucosamine back bones cross linked with peptide chain and ▪ Spirochetes w/endoflagella pentaglycine bridge. ▪ Treponema pallidum – syphilis ▪ Peptidoglycan – ▪ Borellia burgdorferin – lyme disease primary component ▪ Leptospira interrogans – Leptospirosis ▪ 2 major types of Cell wall:  o Gram positive o Gram negative Functions of cell wall: 1. Provides a good shape to the bacterium (cocci, rod, spherical, or spiral shape) 2. Gives rigidity to the organism 3. Protects from environment 4. Provides staining characteristics to the bacterium 5. Contains receptor sites for Gram stain: VIAS phages/complements ▪ Crystal violet – attached violet 6. Contains toxic components to host ▪ Iodine – inedible ▪ Alcohol – yes, the crystal violet to the cell wall will be removed in gram negative and turn to colorless ▪ Safranin – this will attach in gram negative cell wall turns color red Components of cell wall of Gram-Positive Bacteria ▪ Peptidoglycan ▪ Teichoic acid ▪ Lipoteichoic acid Gram stain: VIAS ▪ Crystal violet – primary stain ▪ Iodine – mordant ▪ Alcohol – decolorizing agent ▪ Safranin – secondary stain Components of Cell wall of Gram-Negative Bacteria ▪ Peptidoglycan ▪ Lipoprotein Acid Fast Cell Wall ▪ Phospholipids ▪ Gram positive cell wall structure with ▪ Lipopolysaccharide lipid mycolic acid (cord factor) ▪ Pathogenicity and high degree of resistance to certain chemicals and dyes. ▪ Basis for acid fast stain used for diagnosis of infection caused by these microorganisms.  ▪ Mycobacterium and Nocardia Peptidoglycan – prevent osmotic lysis Arabino galactan – provides extra strength, Functions of cell membrane bind glycolic acid and peptidoglycan. Bridge of 1. Regulates the transport of nutrients and waste acid-fast cell wall products into and out of the cell. Endocytosis Acid Fast Staining and exocytosis ▪ Carbol Fuchsin – primary stain 2. Synthesis of cell wall components ▪ Heat/phenol – mordant 3. Assist DNA replication ▪ 0.5% acid alcohol – decolorizer 4. Secretes proteins composed of 0.5% hydrochloric acid 5. Carries on electron transport system in 70% alcohol 6. Provides site for energy reactions, nutrients ▪ Methylene blue – counter stain processing, and synthesis transport into and (secondary stain) out of the cell ▪ Acid fast bacilli – (+ result is red) ▪ Non-acid fast bacilli – (blue) 3. Internal Structures 2 types of Acid-Fast Staining Chromosomes 1. Ziehl Neelsen (heat) ▪ Single, circular, double stranded DNA 2. Kinyoun’s method (cold – tissues) molecule that contains all No Cell Wall Bacteria genetic Pleomorphic information ▪ Filamentous to required by a coccus or doughnut cell. shape ▪ DNA is tightly ▪ Mycoplasma coiled, pneumoniae – fried aggregated in a dense area called egg appearance NUCLEOID. ▪ Blueprint of our body. Plasmids b. Cell membrane ▪ Extra chromosomal genetic material. ▪ It accounts for 30% of ▪ Free or integrated into the the dry weight of chromosome bacterial cell. ▪ Duplicated or passed on too offspring. ▪ Phospholipid bilayer ▪ Confer traits like antibiotic resistance. with embedded Ribosomes proteins – fluid ▪ The ribosome monomer is 70s with mosaic model. two subunits, (30s and 50s) small and ▪ 60% protein, 20-30% large. lipid, 10-20% ▪ Protein synthesis area of the cell carbohydrates  Cytoplasmic granules and Inclusions 3. Spherical – helical or corkscrew-shaped ▪ Intracellular storage bodies vary in bacterium size, numbers, and content Pleomorphic – ability of bacteria to exhibit ▪ Bacterial cell can use them when various shapes and forms. environmental sources are depleted. Morphology - arrangement of bacteria ▪ For survival and can cause virulence ▪ Diplococci factor. ▪ Staphylococci Examples: ▪ Streptococci ▪ Metachromatic granules – phosphate ▪ Coccobacilli reserves ▪ Tetrad ▪ Polysaccharide granules – stores the ▪ Sarcinae energy in the form of CHO ▪ Lipid inclusions – energy storage in lipid form ▪ Sulfur granules – reserves energy from sulfur oxidizing/ containing bacteria ▪ Carboxysomes – enhance carbon fixation ▪ Gas vacuoles – provide buoyancy ▪ Magnetosomes – navigation the earths magnetic field Bacterial Growth and Nutrition Requirements for growth Spores/Endospores ▪ Nutrients (Carbon, Hydrogen, Oxygen, ▪ Bacillus spp. and Clostridium spp. Nitrogen, Sulfur, Phosphate and ions) ▪ A resting structured formed inside ▪ Temperature – 370C some bacteria ▪ pH – acidic or alkaline ▪ Means of survival when their moisture ▪ Aeration – aerobes and anaerobes or nutrient supply is low. ▪ NaCl requirements – salt Nutritional Need ▪ Water ▪ Carbon ▪ Energy ▪ Nitrogen ▪ Minerals Classification of bacteria according to HOW they meet their nutritional needs Autotroph (Lithotroph) ▪ Can cause pathogenicity that’s why it ▪ Produce their own food is infectious. ▪ C02 + H2O + inorganic salts ▪ Grow using CO2 as an only source Bacterial Morphology of carbon atoms 3 basic shapes of bacteria ▪ Obtain energy 1. Coccus – spherical or ovoid bacterium o Photosyntically 2. Bacillus – rod shaped bacterium (phototrophs)  o Oxidation of inorganic Facultative anaerobes - +/- of O2 compounds ▪ Most clinically significant (chemolithotrophs) ▪ Generally, do not need O2 but grow Heterotroph best w/O2 ▪ Requires more complex ▪ Enterobacteriaceae substances for growth Aerotolerant anaerobes – survive in the o Organic source of carbon presence of oxygen but will be unable to o Obtain energy via oxidation perform metabolic processes. or fermentation or organic ▪ Propionibacterium acnes substances – mannitol, Carbon dioxide requirements lactose, sucrose Capnophiles Temperature ▪ Requires increase CO2 Psychrophiles (cryophiles) and ▪ Most anaerobes and facultative Psychrotrophs: grow best at 0-200C anaerobes require only 0.03% CO2 ▪ Listeria monocytogenes – ▪ Haemophilus influenzae – X factor spontaneous abortion, have known as hemin and V facto known as inverted Christmas tree NAD+ appearance ▪ Neisseria gonorrhea – Chocolate Agar ▪ Yersinia enterocolitica – bulls’ eye Plate colony in cultured media ▪ Streptococcus pneumoniae Mesophiles: grow best at 20-450C Other salt and pressure ▪ Most pathogenic bacteria Halophiles Thermophiles: grow best at 50-1250C ▪ Requires increased NaCl ▪ Bacillus stearothermophilus – QC concentration of autoclave, 1210C, 15 psi, 15 ▪ Staphylococcus aureus minutes ▪ Listeria monocytogenes ▪ Thermus aquaticus Barophiles Extremophiles: capable of surviving in ▪ Grow rapidly in high-pressure unfavorable conditions (temperature or environments pressure) above 1250C ▪ Photobacterium, Shewanella, ▪ Bacillus infernus Colwelia pH Bacterial growth Optimal pH: 7.0-7.5 Generation Organism based on pH tolerance: ▪ doubling bacterial cell number ▪ Acidophiles (low pH 0-5.5) - Generation/Doubling time Lactobacillus acidophilus ▪ Time requirement for 1 bacterial cell to ▪ Neutrophiles (equal pH 5.5-8.0) divide into 2 by binary fission ▪ Alkalophiles (high pH 8.0-11.5) – ▪ Varies: Bacillus alkalophilus and Natrono o 20mins (E. coli) bacterium o 24hrs (M. tuberculosis) Aeration (Oxygen requirements) Aerobes (obligate aerobes) – requires 02 for growth. 15-20% of oxygen and 1% of carbon dioxide ▪ Bordetella, brucella, Mycobacterium, Pseudomonas Microaerophiles – 2%-10% for growth ▪ Campylobacter and Treponema pallidum Anaerobes Obligate anaerobes – no O2 for growth ▪ Clostridium and Bacteroides  ▪ Lag phase - metabolically active ▪ Log phase – exponential growth ▪ Stationary phase – plateau ▪ Death phase – exponential decreases in the number of living cells Determination of Cell Numbers Direct microscopic counting ▪ Estimation of bacterial count in a specimen Direct plate count ▪ Number of CFU/ml in broth culture dilutions. ▪ Urine cultures Density measurement ▪ Turbidity of bacterial broth culture in log phase ▪ Preparation of standard inoculum for AST ▪ Effective – susceptible ▪ Not effective – resistance TOPIC 3: BACTERIAL GENETICS AND METABOLISM ▪ Mobile genetic element that can be indicated in larger DNA molecules Bacterial Genome – complete set of DNA ▪ Integrons – type of genetic structure ▪ Single, closed, super coiled, circular that capture the gene cassette and piece of dsDNA able to integrate them into bacterial ▪ Contains all information needed for genome and they provide promoter to growth and replication. drive the expression of the gene cassette. Mobile Genetic Elements ▪ Promoter – they are the initiation site Plasmids to start the process of genes or DNA ▪ Extra chromosomal DNA ▪ Small circular piecers of dsDNA Integrons ▪ Can be gained or lost ▪ Genetic elements that are capable of ▪ Responsible for antimicrobial integrating genes resistance – they able to care the o Does not necessarily include any gene genes that confer resistant antibiotics cassettes ▪ NOT essential for bacterial growth ▪ Via integrons encoded site specific ▪ Considered as virulence factor, able to recombinase cause disease or infection – known as ▪ INT - integrase – responsible for site pathogenicity. Enable to produce specific recombination toxins and able to invade immune ▪ ATT – specific recombination site system. within integron, helps gene cassette to ▪ Able to encode enzymes that allow the integrated as well bacteria to metabolize unusual ▪ PC – promoter located within integron. substances. o Types of Plasmids: MUTATION a. Conjugative – able to transfer themselves ▪ Alteration in the original nucleotide from one bacterium to another bacterium via sequence of the organism’s genome conjugation. ▪ Change in the organism’s genotype b. Non-conjugative – they are not able to ▪ Effect on organism's phenotype may be transfer but they can be transfer if a unnoticeable, noticeable conjugative plasmid is present ▪ Dominant (expressed) and Recessive (not expressed, manifested if 2 recessive trait Transposons is combined) ▪ Jumping genes ▪ Insertion – 1 or more nucleotide is added ▪ Insertion sequence – simplest ▪ Deletion – 1 or more are deleted or o 1000bp removed from DNA sequence ▪ Disrupts and inactivate genes ▪ Frame shift – insertion or deletion that are o Loss of an observable characteristics not in multiple of 3 nucleotides shift the ▪ Usually located in plasmids reading the frame of the codons, o Often carry antibiotic resistance gene potentially altering the entire amino acid ▪ Able to change their position within the sequence downstream. genes that results to inactivation ▪ 3 codons to make protein ▪ For genetic diversity, able to create ▪ AUG – start codon, protein made – mutation methionine ▪ Most common – insertion GENETIC RECOMBINATION Gene cassette ▪ Homologous recombination ▪ Contains a single gene and a ▪ DNA segment of a donor bacterial cell recombination site is inserted to the genome of a recipient o 500-1000bp bacterial cell and is transferred or ▪ May exist with integrons exchanged with a DNA segment of the ▪ Often carry antibiotic resistance genes recipient’s genome.  ▪ Exchange of genetic material between Transformation 2 similar or identical molecules ▪ A recipient cell uptakes free DNA that ▪ Happens during meiosis process is released in the environment from a eukaryotes in DNA ruptured cell. ▪ Crossing over – 2 chromosomes ▪ DNA uptakes – the bacteria take up the overlap each other, and they are able free DNA from the environment to exchange genetic information. 2 through cell membrane. sister chromosomes able to cross ▪ DNA integration – there are internalize each other to transfer genetic DNA integration being processed from information. the outside to become the program of DNA ❖ MECHNISMS OF GENE TRANSFER ▪ Gene expression – expressed the gene Conjugation once enter the system ▪ Transfer of DNA by direct cell-to-cell ▪ Natural transformation – the bacteria contact have evolved mechanism to naturally ▪ A bacterial cell possessing sex pilus take up the DNA from their (donor cell) attaches by means of the surroundings. sex pilus to another bacterial cell ▪ Artificial transformation – made by (recipient cell) scientist to transform bacteria to ▪ Bacteria A – Sex pili used for genetic different level. Heat shock or transfer attached to Bacteria B, able to electroporation to increase transfer genetic information using sex permeability. pili. Transduction ▪ Transfer of genes by a bacteriophage from once cell to another. ▪ Bacteriophage – type of virus that kills off bacteria ▪ Allowing genetic recombination ▪ So, the bacterium they are able to receive the genetic material. This may lack the plasmid or specific genes before conjugation causing because they are able to receive Longman the genetic information. So usually, recipient cell RESTRICTION ENZMES doesn’t have pili. ▪ Endonucleases – proteins that cut DNA at ▪ Pili formation – donor cell extend structure specific sequence called pili that connects recipient cell. If ▪ Found in Bacteria and Archaea where they there’s another bacterium, we have DNA serve as defense mechanism against transfer. Plasmid is transfer from donor to invading foreign DNA such as from viruses. ▪ Mechanisms: ▪ Recognize specific base sequences and 1. Attachment cut the DNA 2. Mating pair formation ▪ Naturally found in bacterial cell 3. DNA transfer ▪ Blunt ends – both strands of DNA are cut 4. Completion – passed genetic information straight across producing fragments with to new bacterial cell from donor cell no overhanging bases. Example: EcoRV ▪ Sticky ends – type of enzymes make staggered cuts. Example: EcoRI  phosphate group to ADP to form a phosphorylated substrate. ▪ This mechanism of phosphorylation occurs only in cytoplasm and mitochondria 2. Oxidative phosphorylation ▪ A process where ATP production that occurs only in mitochondria. ▪ In the mitochondria, the ATP is generated as result of the transfer electron to electron transport chain ▪ ETC and the creation of proton, they’re able to across the inner mitochondrial membrane. FUELING – refers to providing the energy or power in a system, device, or organism Acquisition of nutrients ▪ Passive Transport - does not need energy or ATP ▪ Active Transport – needs energy or ATP ▪ Group Translocation Production of Precursor Metabolites ▪ Embden-Meyerhof- ENERGY UTILIZATION Parnas (EMP) ▪ Biosynthesis of new cell components – ▪ TriCarboxylic acid (TCA) acid where the energy is used to build and ▪ Pentose Phosphate Shunt maintain cellular molecules and structures. ▪ Maintenance of the physical and chemical integrity of the cell – helps the energy in maintaining the cells’ structure and iron balance ▪ Activity of the locomotor organelles – ATP are able to use for the movement of the locomotor like cilia and flagella, etc. ▪ Transport of solutes across membrane – ATP is required for active transport to process inside the cell ▪ Heat production – the energy is expended so we are able to generate energy or heat and manage the thermal regulation. OTHER ▪ Biosynthesis – this is the process of creating complex molecules from simpler ones. ▪ Polymerization – this is specific type of Energy production biosynthesis where the small monomer is 1. Substrate level phosphorylation chemically bonded to long polymers. ▪ A process where the ATP production occurs through the direct transfer of a  ▪ Assembly – type or organization and TYPES OF FERMENTATION arrangement of synthesized molecules Alcoholic fermentation ▪ Turns sugar into RESPIRATION ethanol and Aerobic Respiration – process that needs carbon dioxide oxygen/ oxygen requirement. ▪ Yeasts ▪ Synonymous w/ oxidative (Saccharomyces phosphorylation cerevisiae) – ▪ occurs in obligate aerobes (organism used for to make that needs oxygen) and facultative beer aerobes (either oxygen is present or not) Homolactic fermentation ▪ Energy generating process in which ▪ Pyruvate is reduced to lactate molecular oxygen is the final electron ▪ Streptococcus and Lactobacillus acceptor ▪ Yogurt, cheese, pickled vegetables ▪ Produced 36-38 ATP per glucose. Anaerobic Respiration – no present of oxygen, 2 ATP per glucose ▪ Occurs in anaerobes ▪ Final electron acceptor: inorganic forms or oxygen ▪ Lactic acid, ethanol, carbon dioxide Heterolactic fermentation ▪ Produces substances such as alcohol, carbon dioxide, formic acid, acetic acid ▪ Leukonostoc and Mesenteroides ▪ Sauerkraut and pickles Mixed acid fermentation ▪ Production of ethanol and acids (lactic, acetic, succinic, formic acid) FERMENTATION ▪ Produces strong positive reaction in ▪ Anaerobic process methyl red test ▪ Less efficient in energy generation ▪ Escherichia coli, Salmonella, Shigella ▪ Final electron acceptor is an organic compound. ▪ Occurs in obligate and facultative anaerobes ▪ Used to indicate any type of utilization of CHO o When fermentation occurs – used to identify bacteria o Mixture of the end products accumulates in the medium o Production of an acid pH  Butanediol fermentation ▪ Bromocresol purple – Acidic – ▪ Pyruvate is converted into acetoin then yellow/Neutral – green/Alkaline – blue reduced to 2,3-butanediol with NADH Lactose fermentation – an important step in ▪ Produces small amounts of acids identifying members of the ▪ Presence of acetoin produces positive Enterobacteriaceae family rxn to Vogues-Proskauer (PV) test MacConkey Agar is very useful in identifying ▪ Klebsiella, Enterobacter, Serratia bacteria if they can ferment lactose. The color is reddish/pinkish in color. If colorless – lactose non fermenter Rapid Lactose Fermenters (RLF) – E. coli, Klebsiella, Enterobacter Late Lactose Fermenter (LLF) – Citrobacter, Serratia, Salmonella, Halfnia, Yersinia Butyric acid fermentation ▪ Involves the conversion of pyruvate into butyric acid along with acetic acid, CO2 and H+ IMPORTANCE OF BACTERIAL ▪ Clostridium, Fusobacterium and BIOCHEMISTRY Eubacterium (obligate anaerobes) Metabolic differences are markers of identification. Diagnostic procedure in clinical microbiology laboratory identifies unknown microorganism based: ▪ Utilization of various substrates as a carbon source. ▪ Production of specific end products from various substrates. ▪ Production of an acid or alkaline pH in the test medium. ▪ Citric test- useful test if the bacteria are able to produce carbon as source of energy. CARBOHYDRATE UTILIZATION AND LACTOSE FERMENTATION Utilization of CHO that leads to acid production Detected using pH detectors Acidic – red Neutral – yellow ▪ Neutral red – (6.8-8.0) Acidic – red/Neutral – yellow ▪ Phenol red – (6.8-8.4) Acidic – yellow/ Neutral – red/ Alkaline – pink ▪ Eosin Y – (4.0-6.0) Acidic – yellow to pink/Alkaline – pink ▪ Bromothymol blue – (6.0-7.6) Acidic – yellow/Neutral – green/ Alkaline – blue TOPIC 4: ANTIMICROBIAL AGENTS – ANTIBATERIAL Bacteriostatic Agents that inhibit bacterial growth but Antimicrobial agents generally do not kill the microorganism Agents that can kill and inhibiting the growth of An agent that inhibits the growth or microorganisms. reproduction of bacteria, rather than killing ▪ Antibacterial agents them outright. Bacteriostatic antibiotics allow ▪ Antifungal agents the immune system to clear the infection. ▪ Antiprotozoal agents Examples include tetracyclines and ▪ Antiviral agents sulfonamides. Examples: Characteristics of an Ideal Antimicrobial ▪ Chloramphenicol Agents ▪ Dapsone ▪ Kill or inhibit the growth of pathogens ▪ Erythromycin ▪ Cause no damage to the host (no side ▪ Clindamycin effects) ▪ Isoniazid ▪ Cause no allergic reaction in the host ▪ Rifampicin ▪ Be stable when stored in solid or liquid ▪ Sulfonamides form ▪ Tetracyclines ▪ Remain in specific tissues in the body long enough to be effective Broad-spectrum antibiotic ▪ Kill the pathogens before they mutate and Effective against a wide range of become resistant to it microorganisms An antibiotic that is effective against a Antibiotics wide variety of bacterial species, including ▪ Antimicrobial substances produced by both Gram-positive and Gram-negative microorganisms bacteria. These antibiotics are often used o Bacteria when the specific causative agent of an o Fungi infection is unknown. Examples include ▪ Not Effective for viruses! amoxicillin and ciprofloxacin. All antibiotics are antimicrobial agents, not all Examples: antimicrobial agents are antibiotics. ▪ Chloramphenicol ▪ Ampicillin ❖ TERMINOLOGIES ▪ Tetracycline Bactericidal Agents that kill or destroy microorganisms Narrow-spectrum antibiotic Used for treatment of life-threatening Effective only against specific infections microorganisms An agent that kills bacteria. Bactericidal An antibiotic that is effective against a specific antibiotics work by destroying the bacterial group of bacteria, either Gram-positive or cell wall, inhibiting essential enzymes, or Gram-negative. Narrow-spectrum antibiotics disrupting bacterial DNA. Examples include are typically used when the causative agent is penicillin and vancomycin. known. Examples include penicillin G Examples: (effective against Gram-positive bacteria) and ▪ Aminoglycosides aztreonam (effective against Gram-negative ▪ Beta lactams bacteria). ▪ Bacitracin Examples: ▪ Glycopeptides ▪ Vancomycin – gram + bacteria ▪ Isoniazid ▪ Colistin – gram – bacteria ▪ Quinolones ▪ Nalidixic acid ▪ Metronidazole  Minimal-Inhibitory Concentration (MIC) ▪ Examples include: Lowest concentration of a drug that can still ▪ Morphine (derived from the opium poppy) inhibit bacterial growth ▪ Quinine (extracted from the bark of the The lowest concentration of an antibiotic that cinchona tree) prevents visible growth of a bacterium after a ▪ Aspirin (originally derived from willow specified period. MIC is a critical parameter in bark) determining the efficacy of an antibiotic against a particular bacterial strain. Minimal Lethal Concentration (MLC) Lowest concentration of a drug that can still kill bacterial growth The lowest concentration of an antibiotic that kills a particular bacterium. MLC is typically higher than MIC and is an important measure in determining whether an antibiotic is bactericidal or bacteriostatic. Semi-synthetic drugs Modified natural drugs Therapeutic index Addition of chemical groups Ratio of toxic dose to the therapeutic dose These are drugs that start as natural Higher therapeutic index – more effective substances but are chemically modified in the agent laboratory to enhance their efficacy, reduce side effects, or improve their stability. Semi- synthetic drugs are often designed to overcome limitations of the natural compounds. Examples include: a measure used to assess the safety of a Amoxicillin (a modified form of penicillin) drug. It is the ratio of the toxic dose to the Heroin (a chemically modified derivative of therapeutic dose of a drug. Specifically, it morphine) is calculated as: LSD (initially derived from ergot fungus, then Toxic Dose (TD50): The dose at which the synthesized in the lab) drug causes toxicity in 50% of the Examples: population. ▪ Methicillin Effective Dose (ED50): The dose at which ▪ Ampicillin the drug is therapeutically effective in 50% ▪ Carbenicillin of the population. A high therapeutic index indicates that a Synthetic drugs drug can be used safely with a lower risk of Chemically produced drugs toxicity, as there is a large margin between These are entirely man-made drugs that the effective dose and the toxic dose. are synthesized in laboratories from A low therapeutic index means that the chemical ingredients. They do not effective dose and toxic dose are closer, originate from natural sources. These which requires careful dosing and drugs are designed to mimic or improve monitoring to avoid adverse effects. upon the properties of natural compounds Examples include: ❖ Classification of Antibacterial Drugs Aspirin (modern versions are fully Natural drugs synthetic) ▪ These are medications derived directly Ibuprofen (a synthetic nonsteroidal anti- from natural sources, such as plants, inflammatory drug) animals, or minerals, without significant Diazepam (a synthetic anxiolytic drug) chemical alteration.  Examples: Mechanism of Action: ▪ Sulfonamides ▪ Inhibition of Cell Wall Synthesis: ▪ Trimethoprim Bacitracin works by interfering with the ▪ Chloramphenicol bacterial cell wall synthesis. ▪ Ciprofloxacin ▪ Target: It inhibits the function of the ▪ Isoniazid bactoprenol carrier molecule. ▪ Dapsone ▪ Effect: Bactoprenol is involved in transporting peptidoglycan precursors Antibacterial agents by Mechanism of across the bacterial membrane. By Action blocking this transport, bacitracin prevents the formation of the cell wall, leading to cell lysis and death. 1. Inhibition of Cell wall synthesis The most selective antibiotics with high therapeutic index Beta-lactams Inhibit the activity of transpeptidase enzyme in ▪ Beta-lactams are a large class of which cell growth halts and eventually antibiotics characterized by their beta- followed by cellular death lactam ring structure. This group includes These antibiotics inhibit the synthesis of the several major classes of antibiotics that bacterial cell wall, a vital component for are effective against a wide range of bacterial structure and integrity. The cell wall bacterial infections. provides rigidity and protection to bacteria, Mechanism of Action: particularly against osmotic pressure. When ▪ Inhibition of Cell Wall Synthesis: Beta- cell wall synthesis is inhibited, the bacteria lactams work by interfering with the become vulnerable to lysis. synthesis of the bacterial cell wall. Examples: ▪ Target Enzyme: They inhibit penicillin- Penicillins and Cephalosporins (inhibit binding proteins (PBPs), which are peptidoglycan cross-linking) essential for cross-linking peptidoglycan Vancomycin (blocks peptidoglycan layers in the bacterial cell wall. synthesis) ▪ Effect: By blocking PBPs, beta-lactams Bacitracin prevent the formation of a rigid cell wall, ▪ Inhibits the peptidoglycan precursors leading to bacterial cell lysis and death. synthesis o Inhibit Transpeptidation ▪ Blocks bactoprenol from transporting o Penicillin NAM and NAG sugars across the cell ▪ Penicillin membrane thereby halting further ▪ Amoxicillin synthesis of peptidoglycan. ▪ Ampicillin ▪ Dephosphorylation ▪ Oxacillin ▪ Bacitracin is an antibiotic primarily used o Penicillinase-resistant penicillin for its antibacterial properties against ▪ Methicillin Gram-positive bacteria. It is often used in ▪ Oxacillin topical preparations for minor skin ▪ Nafcillin infections and as part of some o Cephalosporin combination therapies. ▪ Cefazolin  ▪ Cefoxitin (TB). It is a key component in the standard ▪ Cetriaxone regimen for both active TB disease and latent ▪ Cefepime TB infection. ▪ Cephalothin Mechanism of Action: ▪ Cefalexine Inhibition of Mycolic Acid Synthesis: ▪ Cefuroxime Isoniazid targets the synthesis of mycolic o Monobactam acids, essential components of the ▪ Aztreonam mycobacterial cell wall. o Carbapenem Target Enzyme: It inhibits the enzyme ▪ Imipenem InhA, which is involved in the ▪ Meropenem biosynthesis of mycolic acids Major Classes of Beta-Lactams: (Mycobacterium tuberculosis) 1. Penicillins: Effect: By disrupting the cell wall 1. Examples: Penicillin G, Penicillin V, synthesis, isoniazid impairs the Amoxicillin, Ampicillin integrity of the mycobacterial cell wall, 2. Use: Effective against various Gram- leading to bacterial cell death. positive bacteria and some Gram- negative bacteria. Used for infections Vancomycin like strep throat, pneumonia, and ▪ Glycopeptide urinary tract infections. ▪ Inhibits elongation of peptidoglycan 2. Cephalosporins: ▪ Inhibits translocation 1. Examples: Cefazolin, Cefuroxime, ▪ Vancomycin is a powerful antibiotic Ceftriaxone, Cefepime primarily used to treat serious Gram- 2. Use: Broader spectrum compared to positive bacterial infections. It is known for penicillin, covering more Gram- its effectiveness against bacteria that are negative bacteria. Used for infections resistant to other antibiotics. such as pneumonia, skin infections, ▪ Mechanism of Action: and meningitis. ▪ Inhibition of Cell Wall Synthesis: 3. Carbapenems: Vancomycin works by binding to the D- 1. Examples: Imipenem, Meropenem, alanyl-D-alanine terminal of the bacterial Ertapenem cell wall precursor molecules. 2. Use: Very broad-spectrum activity ▪ Binding Site: It binds specifically to this against Gram-positive and Gram- terminal component of the peptidoglycan negative bacteria, including many precursors. resistant strains. Used for severe ▪ Effect: This binding inhibits the cross- infections and polymicrobial linking of peptidoglycan layers in the infections. bacterial cell wall, thereby preventing cell 4. Monobactams: wall synthesis and leading to bacterial cell 1. Examples: Aztreonam lysis and death. 2. Use: Effective mainly against Gram- negative bacteria. Used for patients with penicillin allergies and in combination with other antibiotics for broader coverage. Isoniazid ❖ Stages in Peptidoglycan Synthesis ▪ Acts only on growing cells 1. Synthesis of precursors in the ▪ Bactericidal or Bacteriostatic cytoplasm ▪ Inhibits cord factor synthesis 2. Transport of lipid-bound precursors mycolic acid across the cell membrane ▪ Isoniazid is an antibiotic 3. Insertion of glycan units into the cell primarily used for the treatment wall and prevention of tuberculosis  4. Transpeptidation linking and ▪ Mechanism: These antibiotics target maturation the bacterial ribosome, interfering with the synthesis of proteins, which are essential for bacterial growth and 2. Cell Membrane Function Inhibition function. Because bacterial Polymyxin ribosomes are different from ▪ Disrupt bacterial cell membranes eukaryotic ribosomes, these drugs can ▪ Act as detergent selectively target bacteria. ▪ More effective against gram-negative bacteria ▪ Examples: ▪ Side effects: Neurotoxicity and Nephrotoxicity ▪ Tetracyclines (block the attachment ▪ Polymyxin B and Colistin of tRNA to the ribosome) ▪ Polymyxins are a class of antibiotics primarily ▪ Macrolides (inhibit the translocation used to treat Gram-negative bacterial step in protein synthesis) infections. The most commonly known ▪ Aminoglycosides (cause misreading polymyxins are polymyxin B and colistin (also of mRNA) known as polymyxin E). Mechanism of Action: ▪ Disruption of Cell Membrane: Polymyxins work by binding to the lipopolysaccharides (LPS) in the outer membrane of Gram-negative bacteria. ▪ Binding Site: They interact with the phospholipids and LPS in the bacterial cell membrane. ▪ Effect: This binding disrupts the bacterial cell membrane, leading to increased membrane permeability, leakage of cellular contents, and ultimately bacterial cell death. ❖ Aminoglycoside and Aminocyclitols Inhibit protein synthesis by irreversibly binding to 30S Effective against a variety of aerobic gram (-) bacteria and certain groups of gram (+) bacteria Examples: ▪ Amikacin – treats serious Gram- negative infections, including Pseudomonas aeruginosa and Mycobacterium tuberculosis. Often used for infections resistant to other 3. Inhibition of Protein synthesis aminoglycosides. Binds with 30S subunit ▪ Gentamicin – effective against a range ▪ Misread mRNA; interfere with of Gram-negative bacteria and some aminoacyl-tRNA binding Gram-positive bacteria. Commonly ▪ Examples: Aminoglycosides, used for infections like sepsis, Tetracyclines, Spectinomycin, pneumonia, and urinary tract Minoglycosides infections. Binds with 50S subunit ▪ Kanamycin – used to treat Gram- ▪ Inhibits peptidyl transferase negative infections, particularly in ▪ Inhibits peptide chain elongation tuberculosis therapy. Less commonly ▪ Examples: Macrolide-Lincosamide- used now due to the availability of Streptogramin (MLS) Group, other aminoglycosides. Ketolides Chloramphenicol, ▪ Neomycin – primarily used topically Linezolid for skin infections, eye infections, and  as part of bowel preparation before Minocycline: Used for acne and certain skin surgery. Also used orally for hepatic infections, with good tissue penetration. encephalopathy. ❖ Glycylglycines ▪ Streptomycin – historically used for Semi-synthetic tetracycline derivatives tuberculosis and certain Gram- Binds reversibly to the 30S subunit negative infections. Less commonly Refractory to most common tetracycline used now due to resistance and resistance mechanisms by both gram (+) and availability of other drugs. gram (–) bacteria ▪ Tobramycin – treats Gram-negative Tigecycline infections, particularly Pseudomonas Glycylglycines are a class of antibiotics aeruginosa, and used in cystic fibrosis known for their effectiveness against certain patients for lung infections. Also Gram-negative bacteria, including those available as an inhaled formulation. resistant to other antibiotics. The most prominent drug in this class is tigecycline. ❖ Tetracyclines Mechanism of Action: Broad-spectrum bacteriostatic antibiotics Inhibition of Protein Synthesis: Binds reversibly to the 30S Glycylglycines, like tetracyclines, work by Prevent peptide chain elongation; Binding of binding to the 30S ribosomal subunit of tRNA-amino acid complexes to ribosome bacterial ribosomes. Effective against gram (+) and gram (-) Binding Site: They bind to the 16S rRNA of the organisms, intracellular bacterial pathogens, 30S ribosomal subunit. Neisseria gonorrhea, and some protozoa Effect: This binding prevents the binding of Tetracycline, Demeclocycline, Doxycycline, aminoacyl-tRNA to the ribosomal A-site, Minocycline inhibiting protein synthesis and thereby Tetracyclines are a class of broad-spectrum stopping bacterial growth. antibiotics used to treat a variety of bacterial infections. They are known for their ❖ Macrolide-Lincosamide-Streptogramin effectiveness against both Gram-positive and (MLS) Group Gram-negative bacteria and some atypical Macrolides – most common pathogens. ▪ Binding to the 23sRNA on the bacterial 50S Mechanism of Action: ribosomal subunit Inhibition of Protein Synthesis: Tetracyclines ▪ Disruption of the growing peptide chain work by binding to the 30S ribosomal subunit ▪ Bacteriostatic; Bactericidal if Infective of bacterial ribosomes. Dose is low and used dosage Binding Site: They bind specifically to the 16S ▪ Examples: rRNA of the 30S ribosomal subunit. ▪ Erythromycin – one of the first macrolides, Effect: This binding inhibits the attachment of used for a range of infections including aminoacyl-tRNA to the ribosomal A-site, respiratory and skin infections. preventing the addition of new amino acids to ▪ Azithromycin – known for its long half-life and the growing peptide chain, thereby stopping convenience of once-daily dosing, commonly bacterial protein synthesis and growth. used for respiratory and sexually transmitted Examples of Tetracyclines: infections. Tetracycline: The original tetracycline ▪ Clarithromycin – known effective than antibiotic, used for various infections but less erythromycin against certain bacteria and has commonly used today due to newer options a longer half-life, allowing for less frequent with better safety profiles. dosing. Doxycycline: Commonly used for a range of ▪ Clindamycin infections, including respiratory infections, Mechanism of Action: sexually transmitted infections, and Lyme ▪ Inhibition of Protein Synthesis: Macrolides disease. It has a longer half-life than work by binding to the 50S ribosomal subunit tetracycline. of bacterial ribosomes. ▪ Binding Site: They bind specifically to the 23S rRNA of the 50S ribosomal subunit.  ▪ Effect: This binding inhibits protein synthesis Mechanism of Action: by blocking the exit tunnel through which the ▪ Inhibition of Protein Synthesis: nascent peptide chain exits the ribosome, Streptogramins work by binding to the 50S thus preventing peptide chain elongation and ribosomal subunit of bacterial ribosomes. bacterial growth. ▪ Binding Sites: They bind to different sites on the 23S rRNA of the 50S ribosomal subunit. Lincosamide Quinupristin and dalfopristin, when used ▪ Bind to the 50s ribosomal subunit together, synergistically inhibit protein ▪ Prevent elongation; Interfere peptidyl transfer synthesis. ▪ Can be bacteriostatic or bactericidal ▪ Effect: This binding prevents peptide bond ▪ Effective against gram (+) cocci formation and elongation during translation, ▪ Examples: leading to bacterial growth inhibition and cell ▪ Clindamycin death. ▪ Lincomycin ▪ Lincosamides are a class of antibiotics ❖ Ketolides primarily used to treat Gram-positive bacterial ▪ Chemical derivative of erythromycin A and infections. The most well-known lincosamide other macrolides is clindamycin. ▪ Binds to the 23s rRNA of the 50S ribosomal Mechanism of Action: subunit ▪ Inhibition of Protein Synthesis: ▪ Maintains activity against most macrolide- Lincosamides work by binding to the 50S resistant gram-positive organisms ribosomal subunit of bacterial ribosomes. ▪ Does not induce a common macrolide ▪ Binding Site: They bind specifically to the 23S resistance mechanism rRNA of the 50S ribosomal subunit. ▪ Effective against: Gram (+), some gram (-), ▪ Effect: This binding inhibits protein synthesis Mycoplasma, Mycobacteria, Chlamydia, and by preventing peptide bond formation during Rickettsia spp. and Francisella tularensis translation, which disrupts bacterial growth ▪ Example: Telithromycin and leads to cell death. Mechanism of Action: Clinical Uses: ▪ Inhibition of Protein Synthesis: Ketolides, ▪ Gram-Positive Infections: Effective against like macrolides, work by binding to the 50S infections caused by Gram-positive bacteria, ribosomal subunit of bacterial ribosomes. including: ▪ Binding Site: They bind to the 23S rRNA of the o Staphylococcus aureus (including 50S ribosomal subunit, specifically at the some MRSA strains) peptidyl transferase center. o Streptococcus species ▪ Effect: This binding inhibits protein synthesis by preventing the translocation of peptides Streptogramin during translation, thereby stopping bacterial ▪ Bind irreversibly to the 50s subunit growth. ▪ Interferes with peptide bond formation; Disrupt protein elongation ❖ Oxazolidinones ▪ Effective against gram (+) and some gram (-) ▪ Linezolid bacteria ▪ Synthetic agent ▪ Examples: ▪ Binds to the 23S rRNA of 50S subunit ▪ quinupristin-dalfopristin ▪ Interferes with the binding of tRNA for formyl- ▪ Streptogramins are a class of antibiotics that methionine (inhibits initiation of mRNA are used to treat infections caused by certain translation) Gram-positive bacteria. They are particularly ▪ Effective against most gram (+) bacteria and effective against multi-drug-resistant strains. mycobacteria The most well-known combination of ▪ Not expected to be affected by resistance streptogramins is quinupristin-dalfopristin. mechanism that affect other drug classes Mechanism of Action: ▪ Inhibition of Protein Synthesis: Oxazolidinones work by binding to the 23S  rRNA of the 50S ribosomal subunit, inhibiting Mechanism of Action: the initiation of protein synthesis. ▪ DNA Damage: Metronidazole works by ▪ Binding Site: The binding site is different from entering bacterial and protozoal cells and other antibiotics, which means they are being reduced by the nitro group in the effective against bacteria resistant to other presence of anaerobic conditions. This classes of antibiotics. reduction generates reactive intermediates ▪ Effect: This inhibition prevents the formation that bind to and damage bacterial and of the initiation complex for protein synthesis, protozoal DNA, leading to the disruption of thereby stopping bacterial growth. DNA synthesis and cell death. ❖ Chloramphenicol Rifamycin ▪ Inhibits the addition of amino acids ▪ Semisynthetic ▪ Binds to the 50S subunit; Inhibits ▪ Inhibit RNA synthesis transpeptidation o Bind to the DNA-dependent RNA ▪ Effective against a wide variety of gram (+) and polymerase gram (–) bacteria ▪ Cannot effectively penetrate the outer ▪ Not widely used due to toxicity membrane of gram (-) ▪ Temporary to permanent bone marrow ▪ Commonly used in combination with other suppression, leading to aplastic anemia antimicrobial due to high frequency of Mechanism of Action: resistance ▪ Inhibition of Protein Synthesis: ▪ Used for treating M. tuberculosis infections Chloramphenicol works by binding to the 50S Mechanism of Action: ribosomal subunit of bacterial ribosomes. ▪ Inhibition of DNA-Dependent RNA Specifically, it binds to the 23S rRNA of the 50S Polymerase: Rifamycins work by binding to ribosomal subunit, inhibiting the peptidyl the beta subunit of bacterial DNA-dependent transferase activity. RNA polymerase. This enzyme is essential for ▪ Effect: This inhibition prevents peptide bond the transcription of DNA into RNA. formation during protein synthesis, leading to ▪ Binding Site: Rifamycins bind to the DNA-RNA bacterial growth arrest and cell death. polymerase complex, preventing the enzyme from synthesizing RNA. 4. Inhibition of Nucleic acid synthesis ▪ Effect: This inhibition disrupts RNA synthesis ▪ Inhibition of Nucleic Acid Synthesis: and consequently protein synthesis, leading Mechanism: These antibiotics interfere with the to bacterial cell death. synthesis of DNA or RNA, which are essential for bacterial replication and function. By inhibiting Fluoroquinolones enzymes involved in nucleic acid synthesis, these ▪ AKA quinolones; derivatives of nalidixic acid drugs prevent the bacteria from reproducing. ▪ Bind and interfere with DNA gyrase in gram (-) ▪ Examples: ▪ Newer quinolones also interfere with ▪ Quinolones (inhibit DNA gyrase and topoisomerase IV in gram (+) topoisomerase) ▪ Broad-spectrum bactericidal agents ▪ Rifampin (inhibits RNA polymerase) ▪ Examples: ciprofloxacin, ofloxacin, levofloxacin, gatifloxacin Metronidazole ▪ inhibition of DNA Gyrase and ▪ Contains a nitro group Topoisomerase IV: Fluoroquinolones work by o Reduced by organism’s nitro inhibiting bacterial enzymes called DNA reductase to free radicals gyrase (Topoisomerase II) and ▪ Activation requires low redox potential Topoisomerase IV.DNA Gyrase: Helps in environment relieving the torsional strain of DNA during o Anaerobic conditions replication and transcription. ▪ Effective against anaerobic, gram (-) ▪ Topoisomerase IV: Helps in separating microaerophiles, Trichomonas interlinked DNA strands following DNA ▪ and Giardia spp. and Entamoeba histolytica replication. ▪ By inhibiting these enzymes, fluoroquinolones combination with sulfonamides (as TMP-SMX) prevent DNA replication and transcription, for its synergistic effect. leading to bacterial cell death. Mechanism of Action: ▪ Inhibition of Dihydrofolate Reductase: 5. Inhibition of other metabolic processes Trimethoprim specifically inhibits the enzyme ▪ Inhibition of Other Metabolic Processes: dihydrofolate reductase, which is ▪ Mechanism: These drugs target specific responsible for reducing dihydrofolate to metabolic pathways that are crucial for tetrahydrofolate. Tetrahydrofolate is the active bacterial survival. By disrupting these form of folate required for bacterial nucleic pathways, the bacteria cannot produce the acid and protein synthesis. necessary components for growth and replication. Nitrofurantoin ▪ Examples: ▪ Consists of a nitro group on a heterocyclic ring ▪ Sulfonamides (inhibit the synthesis of folic ▪ Diverse and multifaceted MOA acid, which is necessary for DNA synthesis) ▪ Converted by nitro reductases to reactive ▪ Trimethoprim (also targets folic acid intermediates synthesis but at a different step than ▪ Used to treat uncomplicated UTI sulfonamides) o Has good activity against most gram (+) and gram (-) Sulfonamides ▪ Nitrofurantoin is an antibiotic used primarily ▪ Target and bind to dihydropteroate synthase for the treatment and prevention of urinary ▪ Disrupt folic acid pathway tract infections (UTIs). It is effective against a ▪ Effective against a wide range of gram (+) and range of Gram-positive and Gram-negative gram (-) organisms except P. aeruginosa bacteria that commonly cause UTIs. ▪ Moderately toxic; causes nausea, vomiting ▪ Mechanism of Action: and hypersensitivity reaction ▪ Inhibition of Bacterial Enzymes: ▪ Example: Sulfamethoxazole Nitrofurantoin is a prodrug that, upon entering ▪ Inhibition of Folate Synthesis: Sulfonamides bacterial cells, is reduced to several active inhibit the bacterial enzyme dihydropteroate metabolites. These metabolites interfere with synthase, which is involved in the synthesis of bacterial enzymes involved in several folate (folic acid). processes, including: ▪ Folate is crucial for bacterial DNA, RNA, and o Cell Wall Synthesis protein synthesis. o Protein Synthesis ▪ By blocking this enzyme, sulfonamides o DNA Synthesis prevent the formation of folate, thereby o Aerobic Energy Metabolism inhibiting the growth and replication of ▪ The exact mechanism involves multiple bacteria. actions, but overall, it disrupts bacterial metabolism and inhibits growth. Trimethoprim ▪ Targets folic acid pathway ANTIMICROBIAL