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More on Microbes Notes

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MICB 202
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More on Microbes • Are everywhere • A strain=pop of bacteria descened from a single cell • Virulence=measure of pathogenicity of microbe (quantitative eg. Cell number needed to cause pathogenic response) • Hygiene hypothesis: because we are too clean, immune system not stimulated early on, end up being hyper-reactive afterwards • Bubble boy: had no T, B cells that acted normallydied • Host benefits o Prevents bad bacteria from growing by changing pH, O2, producing inhibitory substances, hogging nutrients, occupying space o Gives VK, B12, o Steroid metabolism to break down bile acids o Produce organic acids o Breaks down sugar o Stimulate + enhance host defenses  Need normal flora to develop normal immune system • Colonization at birth o From environment: dirt, food, fluids, inhalation, contact w/ people and animals o Stable micro-flora established rapidly o Mostly first from contact with mother  Pre-mastication of food  Mammary  Cutaneous  Vaginal (passage thru) • Bacteria in GI tract o Anaerobes o 3 main phyla in gut, 500 dif species (gram positive, proteobacteria, bacteroide) • During infectious diarrhea o Shift in species in gut, decrease in total bacteria • Fecal microbiota transplantation to treat irritable bowel disease, c. difficile infection • Normal skin flora (mostly associated with sweat glands and hair follicles) o Epidermis is impregnable o Need brekas in skin: wounds, bites, sweat glands o Skin is dry, high salt, acidic, antimicrobials in sweat • Flora in oral cavity o Saliva not good (lysozyme, peroxidase) o Teeth at first aerotolerantanaerobes o Bacteria grow in layers=plaques o Caries: bacteria destroy enamel by making lactic acid o Periodontal disease: gingivitis and inflammation • Flora in resp. tract o Nose: s. aureus, s. epidermis o Sinuses: few bacteria, can cause sinusitis o Nasopharynx: many dif types o Lower resp. tract: few bacteria w/ mucociliary system clearance  Secondary infection can follow bronchitis or pneumonia • Normal flora of Urogenital o Urine + urogenital tract usually sterile o Vagina, distal urethra have bacteria  Lactobacillus in vagina • Protects against pathogens by lowering pH with lactic acid production • Without them, get UTIs o Bacteria in GI tract all anaerobes • Flora establish by its ligands bind to host cell receptors eg. Capsules, cell wall components • If animals isolated w/o bacteria o Underdeveloped immune system, very susceptible to pathogens, low Ab titer, thin intestinal walls, abnormal features BIOFILMS • Are everywhere, where bacteria form communities o 2/3 of all bacterial infections=caused by biofilms  Eg. On medical equipment and heart valve impants, catheters, hemodialysis equipment o Form on surfaces, edges, bottoms of aqueous environments • Examples o Slippery rocks in stream o Slippery roof o Ship hull coatings causing corrosion o Harmful biofilms in diseases  Lungs: cystic fibrosis  Lungs: TB  UTI  Upper resp tract (ear infection, tonsillitis) • Biofilms: bacteria in terrestrial environment within communities that susbtain enviro ecosystems • Benign biofilms provide surface for pathogens (eg. Bacillus anthracis) o Biofilms can be found in hospitals and air conditioning and drinking water. • Why form biofilms: o Stability in growth environment: physical structure to protect and anchor o Dependence on interactive metabolism o Protection from enviro: uv, metals, acid, phagocytosis, Antibiotics o Similar bacteria work together to undertake a big job) o Diverse bacteria work together to occupy diverse niches • What is it? o Communities of bacteria adhered to surface on matrix of extracellular polymeric substances (EPS)  Slimy, film-like, produced by bacteria: polysaccharides, proteins, nucleic acids • Can be thick with different bacteria in different layers • Layout of matrix particles is not homogenous  Formation occurs with coordinated chemical signaling between cells=quorum sensing • Uses autoinducers=signaling molecules produced by bacteria that can bind to bacteria receptors • When enough receptors bound, gene expression changes to encourage community development o How community formed  1. Relocate/swim--flagella  2. Attach to surface (triggered by enviro signals such as T, osmolartiy, pH, iron, oxygen, nutrient availability)  3. Spread on surface • Movement along solid surface mediated by pili  4. Biofilm in 3D with EPS • EPS synthesis triggered by quorum sensing chemicals=homoserine lactones (HSL or AHL or autoregulators)  5. Dispersal (propagate new communities etiher single cells or chunks come off) o Immunity  Planktonic cells (free-living) eradicated by antibodies and antibacterials  Biofilm bacteria provoke same inflammation and immune response but resist death, generate chronic inflammation and tissue necrosis • Also more resistant to antibiotics o EPS=barrier to drugs and immune cells, nutrient barrier so bacteria grow slowly, resistant to antibiotics can develop,  Has persister cells=small subpop that remains inactive and resistant to antibiotics o Altnerate therapies  Block EPS synthesis by coating surfaces or electric charges  Inhibit adherence of biofilms—coat with Teflon or antibiotics  Block cell-cell communication: INHIBIT AUTOINDUCER formation. How? Bacteria and Immune Response • Innate: always present, fast, effective • Cells have TLRs and Nods (intracellular) o Recognize LPS, flagella, peptidoglycan, CpG DNA o Initiates inflammation (chemotaxis, phagocytosis, cytokines, activate complements) • Acute inflammation o Limit bacteria spread and to activate complement system  Release histamine and vasodilation o Recruit phagocytes, activate phagocytes, create clot • Chronic inflammation o Chemotaxins produced by lymphocytes attract more neutrophils/monocytes o Activated macrophages release reactive oxygen species that destroy local cells and tissues Defence against Bacteria • Disinfectants: chemicals that kill microorganisms on non-living surfaces • Antiseptics: chemicals used on living tissues to kill microorganisms • Chemotherapeutic agents: chemicals used in body to kill bacteria, antibiotics used to kill bacteria • Most antibiotics we have=from streptomyces=soil bacteria o Sulfonamides, penicillins o Act by inhibiting bacterial processes  Cell wall synthesis, translation, transcription, replication, membrane permeability, metabolism o Can be bactericidal or bacteriostatic o Can be broad spectrum or narrow spectrum • A good antibiotic should be o Well tolerated, low allergic o Low MIC (effective dose) o Good penetration to reach pathogen in host o Low frequency of resistance appearing o Retained in body—long half life o Broad spectrum • Development of drug resistance o Target modifications: mutations at target site, have enzymes that modify target site o Have enzymes that degrade or modify antibiotics o Efflux pumps to remove antibiotics from inside o How?  Spontaneous mutations • Selection for genes having alternative functions (natural selection) • Then spreads into other bacteria by horizontal transmission=conjugation, or via transformation (get genetics from surroundings), or transduction (bacteriophage injects) • Today, we have more and more superbugs and large pharma don’t make new antibiotics since resistance develops too quickly o Due to patient, physician misuse and agricultural use • Potentially invest in cmpds that inhibit bacterial resistance o Eg. Beta lactamase inhibitors o Understand mechanisms of disease for new drug targets • Vaccines: o Don’t directly kill pathogen but stimulates body to make Ig  Immunize with modified pathogen molecule (toxoid), pieces of pathogen, live attenuated pathogen, or dead pathogen o Active vs passive immunization Classifying Microbes • Why classify: o Choose right antibiotic o Identify source and prevent spread in nosocomial infections • How to classify o Sample the infection site, preserve sample, analyze with colony growth or gram stain • Microscopy o Shape: spherical? Rods? Filamentous? Groups? o Gram positive or negative? o Problems  Can’t see viruses  Complex samples eg. Stool  Many different microbes present • Take sample, streak onto complete media o Can use enrichment media for difficult bacteria or o Selective media when only want to grow 1 thing o Use differential media to visualize colonies with various indicators: dyes, or blood agar to see hemolysis o Use a media to lmeasure metabolic requirements (aerobic, anaerobic, use of different C, N sources, specific nutrients) o Does it produce gas? • Zones of inhibition show susceptibility to antibiotics • Or classify by sequencing genes with qPCR • Or growth-independent ways to identify bacteria o Attach antibody to bacteria to fluorescent marker, if fluorescence detected=infection o Assay based on pathogen-specific enzymnes eg. S. aureus  Coagulase, heat stable DNAase • Epidemiology: when, where, why disease occur, how spread locally, around world • Sporadic=occur occasionally • Endemic: low, continually occurring • Epidemic: localized outbreak, spreading rapidly • Pandemic: spreads over countries/continents • TB o Invade lung, fatal in 10% o Remains in body dormant (30% of world) o No vaccine o Difficult to treat: broad spectrum abtibiotics ineffective o Current standard treamtne=3 drug combo x 6 months, but emerging completely resistant strains • Modes of transmission o Human contact, food, water, blood, drugs, airborne, vectors o Microbial virulence measured by lethal dose of pathogen • Infectious dose=# of bacteria required to cause disease • Lethal dose=number of bacteria needed to kill o Depends on route of entry • Nosocomial infections very important since o Patients often have compromised immunity o These strains are usually antibiotic resistant since so many antibiotics used o Can be endogenous (caused by own flora) or exogenous(transmitted from another person) or from medical procedures (eg. Surgery, catheters, implants) • Incidence=fraction of pop that contracts disease expressed as new cases in time period per 100,000 people • Prevalence: fraction of population with symptoms in a specific time o Includes new and established cases •
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