BIOL341/BIOL982: Introduction to Microbiology Part 1 PDF

BIOL341/BIOL982: Week 1 Introduction to Microbiology Part 1 Dr Emma-Jayne Proctor 1 [email protected] Defending Ourselves Against Microbes  Microbes are found everywhere  1030 microorganisms distributed amongst 1 trillion species on Earth  A complex system of def...

BIOL341/BIOL982: Week 1 Introduction to Microbiology Part 1 Dr Emma-Jayne Proctor 1 [email protected] Defending Ourselves Against Microbes  Microbes are found everywhere  1030 microorganisms distributed amongst 1 trillion species on Earth  A complex system of defenses protect us from pathogenic microbes  Microbes have evolved complex, elegant mechanisms to protect themselves from host defenses 2 Introduction to Microbiology Lecture 1 Outline:  Brief history of Microbiology  Koch’s postulates  Microbiology today  Biofilms  Emerging and reemerging disease  Antibiotic resistance  Microbiome 3 Learning Outcomes  Awareness of the history of microbiology  Describe Koch's postulates and their limitations  Describe Koch's molecular postulates and their limitations  Provide examples of how both sets of postulates would be fulfilled experimentally. 4  A Brief History of Microbiology 1676: Antony van Leeuwenhoek was able to identify “invisible creatures” with the help of his primitive microscope calling them “animalcules” 1850s: Semmelweis and handwashing, determined through observation and experimentation 1820-1910: Florence Nightingale founded the science of medical statistics – demonstrating the impact of infectious disease on overcrowded populations Late 1800’s: Major advances in understanding what causes disease and how to identify it 1900’s: Key discoveries in vaccine and antibiotic development Maresso, A.W. (2019). A Short History of Microbiology. In: Bacterial Virulence. Springer, Cham. https://doi.org/10.1007/978-3-030-20464-8_1 5 Pasteur: Spontaneous Generation  Louis Pasteur demonstrated that microorganisms do not spontaneously generate  Pasteur suggested such germs could also cause human illness and developed the “Germ Theory”  Tried to isolate the pathogen responsible for Cholera - mixed broth cultures 6 Koch: Cause of Disease  Bacillus anthracis as the cause of anthrax  Pure culture technique using a solid media  Developed bacterial staining methods  Isolated Vibrio cholerae and Tuberculosis bacillus  Koch’s postulates first published  Criteria for judging whether a given bacteria is the cause of a given disease  Koch's criteria brought some much-needed scientific clarity to what was then a very confused field 7 The Importance of Jam  Koch used gelatin to set beef broth for bacterial culture  Many bacteria produce gelatinase  Fanny Hesse - Agar, derived from seaweed, to set Jam  Recommended to her husband, Koch’s lab assistant 8 9 Koch’s Postulates 1. The bacteria must be present in every case of the disease. 2. The bacteria must be isolated from the host with the disease and grown in pure culture. 3. The specific disease must be reproduced when a pure culture of the bacteria is inoculated into a healthy susceptible host. 4. The bacteria must be recoverable from the experimentally infected host. 10 Bacteria were being discovered as the agents of some human diseases BUT Koch’s postulates couldn’t confirm bacterial origin for diseases such as measles, mumps, smallpox and yellow fever – WHY? 11 Discovery of Viruses 1892 Dimitri Ivanovsky Soluble toxin 1898 Martinus Beijerinck Contagious living fluid 12 Koch’s Postulates – limitations “Failure to fulfil the Koch postulates does not eliminate a putative microorganism from playing a causative role in a disease. It did not at the time of Koch's presentation in 1890, and it certainly does not today” – Stanley Falkow http://ctl.unbc.ca/outloud/docs/marko/  Microorganisms (such as the bacterium that causes leprosy, Mycobacterium leprae) that cannot be "grown in pure culture" in the laboratory.  No animal model of infection for that particular microorganism e.g. leprosy. 13 Kochs’s Postulates – limitations  Additionally, a harmless microorganism may cause disease if;  It has acquired extra virulence factors making it pathogenic.  It gains access to deep tissues via trauma, surgery, an IV line, etc.  It infects an immuno-compromised patient.  Not all people infected by a bacteria may develop disease-subclinical infection is usually more common than clinically obvious infection.  Still a useful benchmark in judging whether there is a cause-and- effect relationship between a microorganism and a clinical disease 14 Kochs’s Postulates – limitations  Koch's postulates have been modified over the years to encompass:  Viruses  Obligate parasites  Slow viruses (viruses that cause symptoms in an infected host long after the original infection and which progress slowly)  The microbial causation of cancer.  More recently sequence-based identification of bacterial pathogens, to resolve outbreaks of infectious disease and even to define the causation of certain non-infectious diseases. 15 Koch’s Molecular Postulates  Put forward by Stanley Falkow (1988);  Koch’s Postulates  Criteria for determining whether a specific bacterial strain causes a disease.  Koch’s Molecular Postulates  Criteria for determining whether a specific bacterial virulence factor has a role in pathogenesis. 16 Kochs’s Molecular Postulates  Identify gene (or gene product) responsible for virulence determinant  Show gene present in strains of bacteria that cause the disease  Not present in avirulent strains  Disrupting the gene reduces virulence  Complementation with the gene restores virulence  Introduction of the cloned gene into avirulent strain restores virulence  The gene is expressed in vivo  Specific immune response to gene product is protective 17  Experimental demonstration – Koch’s Molecular Postulates  Prp is a multifunctional plasminogen binding M protein. Alanine replacement of 2 amino acids abolishes plasminogen binding activity. Cm XX NS88.2 prp NS88.2prp 18 FASEB J. 2008 22: 2715-22.  Experimental demonstration – Koch’s Molecular Postulates  Complementation of the prp mutant was achieved by reverse complementation. Cm XX NS88.2prp prp NS88.2prpRC Reverse complementation results in stable complemented strains 19 for in vivo studies or where gene copy number is an issue. FASEB J. 2008 22: 2715-22.  Experimental demonstration – Koch’s Molecular Postulates  Plasminogen binding was reduced in the prp mutant and restored in the reverse complemented mutant. 100 Plasminogen binding (%) 75 50 25 * 0 NS88.2 NS88.2prp NS88.2prpRC 20 FASEB J. 2008 22: 2715-22.  Experimental demonstration – Koch’s Molecular Postulates  Virulence was reduced in the prp mutant and restored in the reverse complemented mutant. 100 NS88.2prp Percent survival 75 50 25 NS88.2 NS88.2prpRC 0 0.0 2.5 5.0 7.5 10.0 Time (days) 21 FASEB J. 2008 22: 2715-22. FASEB J. 2008 22: 2715-22. Molecular Postulates  “The molecular Koch's postulates were not intended to be anything more than a means to provide a basis of dialogue among interested investigators…. the dialogue among investigators now takes on less of a phenotypic description based on only a few, often observational, criteria. The dialogue about bacterial virulence genes now centers increasingly on better defined biochemical mechanisms that are less equivocal.” - Stanley Falkow 22 What do you think about the road to microbiology as the discipline we know it today? 23 Microbiology Today  Infectious disease still preoccupies much of microbiology  Polymicrobial diseases and biofilm  Emergence of antibiotic resistance (MRSA)  Emergence of more virulent infectious agents (SARS)  Microbiome  Microbial ecology and evolution, environmental microbiology  Only 1% of all microbes on earth have been studied  Microbiology today is more focused on the relationships among microorganisms and with their environment rather than specific microbes Global Disease Burden – 2019 vs 2021 Infectious Disease Infectious Disease combined 2019 – 15.85% combined 2021 – 26.2% 25 https://ourworldindata.org/burden-of-disease Biofilms  Majority of microbes survive in complex communities rather than individually  One or more types microorganisms physically linked together and to the underlying surface by substances they secrete  Provide increased resistance to antibiotics and the immune system 26 Emerging and Reemerging Infectious Disease Emerging: recently surfaced in the human population for the first time  Eg. HIV, SARS, Lyme disease, Bird Flu and Zika virus all of which have no cure Reemerging: has existed in the past but has shown resurgance in resistant forms and expansion in geographic location  Eg. Drug-resistant tuberculosis, cholera, dengue fever 27 And of course.. COVID! https://www.health.gov.au/topics/covid-19/reporting https://ourworldindata.org/covid-deaths 28  Group A Streptococcus (Strep A) Scarlet Fever Necrotising Fasciitis Streptococcal Toxic Shock Syndrome Global outbreaks since 2019 attributed to new lineage called 'M1UK’ (STSS) Invasive infections are associated with 50% mortality rates Reported increase in antibiotic resistance and clinical treatment failures Urgent need for novel therapeutic strategies Key features of the strains causing these infections is bacterial toxins Antibiotic Resistance  Increasing inability to fight infections because so many pathogens becoming resistant to one or more antibiotics  Rate of antimicrobial resistance developing faster than rate of discovery of new antibiotics  Taking longer to develop a drug from discovery to market  Emergence of superbugs!  Major health threat  One of the most important challenges facing microbiology today https://www.who.int/publications/i/item/9789240093461 Microbiome  Definition 1: The entire collection of genes found in all of the microbes associated with a particular host.  Definition 2: The ecosystem made up of microbes within and on the human body — that is, the collection of microbes that live in the human ”habitat.”  Emerging as an important factor in inflammation, immunity and general health. Changes in Microbiome are associated with many diseases https://www.nature.com/articles/s4139 2-022-00974-4  The microbiome in disease treatment  As research in field of microbiome increases, so has the potential to modulate the microbiome therapeutically  Since the human gut is involved in a wide range of physiologic functions, its modulation is expected to prevent or treat the corresponding diseases  Therefore, many number of clinical trials are ongoing to investigate this possibility https://www.nature.com/articles/s41392-022-00974-4 https://www.nature.com/articles/s41392-022-00974-4  Limitations to microbiome treatment studies  While these response rate are promising, it should be noted:  The trials are mainly pilot studies with small sample size  The underlying mechanism for complete response required further investigation to optimise the experimental design and to personalise the treatment https://www.nature.com/articles/s41392-022-00974-4 Summary  We are surrounded by microbes  The history of microbiology is built on observation and experimentation  Definitive experiments by Koch and Pasteur  Microbiology is still relevant in 2024 36 37  BIOL341/BIOL982: Week 1 Introduction to Microbiology Part 2 Dr Emma-Jayne Proctor [email protected] Introduction to Microbiology Lecture 2 Outline (Textbook, Pommerville Fundamentals of Microbiology chapters 4 and 15): Microorganisms: diversity Pathogenic Microorganisms Structural features of Bacteria Structural features of Viruses Assumed Knowledge: BIOL103, BIOL215, BIOL340 2 Learning Outcomes  Be aware of the techniques used to study microbial diversity  Knowledge of basic bacterial structure  Knowledge of basic viral structure  Understand some techniques used to classify bacteria and viruses What do you know about Microorganisms?  Definition?  What are the 5 types of microorganisms?  What proportion of microorganisms affecting humans are pathogenic?  What are the 10 most dangerous microorganisms to human health? Microorganisms  Colonise every habitat or environment on and in the earth = ubiquitous  Profound influence on all aspects of life  Bacteria, fungi, algae, protozoa and viruses  Vast numbers  Very diverse  Not always harmful but pathogens driving force for much microbiology development 5 Microbial Diversity  Metagenomics is the study of the total number of genomes in a sample  Estimates are conducted by PCR amplification of 16S or 23S rRNA genes because many bacterial Genera are non-culturable by known techniques….  Oceans may support over 2 million bacterial Genera  Half a ton of soil may contain over 4 million bacterial Genera  Human skin houses approximately 2 x 109 bacteria  Human gut content contains approximately 1 x 1014 bacteria Microbial Diversity  Whole genome sequencing becoming more standard  Challenges of computational power, bioinformatics tools and mix of biological and mathematical skillsets. Microbial Diversity  The vast majority of microorganisms are benign and do not cause disease. In fact, most microorganisms are beneficial;  Niche occupation, preventing pathogens gaining entry into gut, lungs, skin etc.  Performing digestion functions in the gut  Turning over nitrogen and carbon compounds in the soil  Photosynthesis  Biotechnology Pathogenic organisms are present in all 5 kingdoms e.g. helminths- worms e.g. red algae e.g. Trichophyton BACTERIA Ring worm VIRUSES e.g. protozoa e.g. bacteria 9 Prokaryotes  Hereditary material – DNA  Complex biochemical processes  Reproduce to produce new generations  Evolutionary adaptation in response to other organisms and the environment  Complex and regulated responses to stimuli Often described as “simple single-cell” organisms, but are they really? Bacterial Cells Have an Organised Structure Figure 4.3: Bacterial cell * structure. * * * *not in all bacteria * 11 The Bacterial Cell Wall  Covers entire cell surface  Serves as exoskeleton  Provides structural integrity  Anchors cellular appendages  Protects the cell from injury  Maintains water balance and prevents cell Figure 4.9: Cell rupture (lysis). rupture The Bacterial Cell Wall Structure  Bacteria are classified into two major groups: Gram-positive or Gram- negative following staining  Gram stain results reflect the thickness of the peptidoglycan layer in the bacterial cell wall.  Gram-positive bacteria stain purple/blue whilst Gram-negative bacteria stain pink. Staphylococcus aureus Escherichia coli The cell wall governs the shape of the bacterium : cocci, rod or spiral shaped. 13 www.cat.cc.md.us Gram-Positive and Gram-Negative Bacterial Cells  Gram-positive bacteria:  Thick peptidoglycan cell walls containing teichoic acid. Figure 4.10B1: Gram- positive cell wall.  Gram-negative bacteria:  Two-dimensional peptidoglycan layer and no teichoic acid.  Outer membrane containing porins separated from the cell membrane by the periplasmic space. Figure 4.10C1: Gram-14 negative cell wall. Bacterial Cell Wall Structure The structure of the Gram-positive cell wall: Peptidoglycan chains - alternating units of 2 amino containing sugars; N acetyl glucosamine (NAG) and N acetyl muramic acid (NAM) 15 www.cat.cc.md.us Bacterial Cell Wall Structure The structure of the Gram-negative cell wall: 16 www.cat.cc.md.us  Courtesy of Elliot Juni, Department of Microbiology The Glycocalyx Serves and Immunology, The University of Michigan. Several Functions  The glycocalyx is a sticky layer of polysaccharides secreted externally to the cell wall. Figure 4.8A: The bacterial  Capsule: Thick, firmly bound layer. glycocalyx in Acinetobacter cells.  Slime layer: Diffuse, water-soluble layer. © George Musil/Visuals Unlimited.  It protects cells from the environment and allows them to attach to surfaces.  Also helps pathogens evade immune system. Figure 4.8B: The17 glycocalyx of E. coli. Cell-Surface Structures Interact with the Environment  Pili are short protein fibers extending from the surface of many gram-negative bacteria.  Type I: Contain adhesins that attach cells to surfaces. Figure 4.4A: Bacterial pili on E. coli.  Actas virulence factor in pathogenic bacteria.  Type IV: Provide “twitching motility” to cells.  Conjugation pili: Used to transfer genetic material between cells. 18 Neisseria gonorrhoeae http://textbookofbacteriology.net Flagella Provide Motility  Prokaryotic flagella are very long corkscrew appendages extending from the cell surface at one or more points.  Bacterial flagella are used for locomotion and chemotaxis (moving up or down chemical gradients). Courtesy of Dr. Jeffrey Pommerville.  Flagella differences are used to classify bacterial strains Figure 4.5A: Bacterial flagella on Proteus vulgaris. 19 Bacterial Spores  Bacterial spores survive desiccation and heating. Spores “germinate” to form vegetative cells eg. Bacillus anthracis.  Vegetative cells usually are destroyed at 60oC while spores require heating to 121oC for 15 min. 20 http://www.wired.com Viruses Have a Simple Structural Organisation  Viruses are tiny infectious agents that are obligate intracellular parasites.  They lack the machinery for generating energy/large molecules and need a host cell to replicate. Figure 15.3: Size relationships Figure 15.4: The viral among cells and viruses. replication cycle. 21 Viruses  Viruses have either a DNA or RNA genome  single- or double-stranded  Viruses are typically less than 1 um (1/1000 mm) in size HIV virus  Viruses lack metabolic enzymes, DNA replication enzymes, ribosomes etc…  Viruses infect all classes and types of living organisms (animals, plants, fungi, bacteria) 22 Viral Structure  The major components of viruses include:  Capsid (or core) which is the protein shell of the virus.  The genome which can be either single- or double-stranded DNA or RNA.  Many animal viruses contain an envelope, which is made up of host cell membrane and viral proteins. Figure 15.5: The components of viruses. 23 Viral Structure  Viruses can be grouped by their shape:  Those with a rod or filament shape have helical symmetry.  Viruses with a capsid that has 20 triangular sides have isocahedral symmetry.  Other viruses are classified as having complex symmetry. 24 Figure 15.6: Viral shapes. Bacterial Viruses  Bacteriophage (or coliphage) T4 infects E. coli.  Many bacteriophage contain a tail structure, which is used like a syringe to inject the bacteriophage genome into the host cell. 25 biologyonline.us Dr Graham Beards via wikimedia.commons Eukaryotic viruses  Eukaryote viruses are usually rod-shaped, oblong or round  These viruses do not possess a “tail” structure Influenza virus Rotavirus 26 http://web.uct.ac.za  Viruses Can Be Classified by Their Genome Figure 15.7: A classification for the medically relevant 27 human viruses. Figure 15.9: Diversity of viruses. Diversity of Viruses 28 Pie charts modified from Small Things Considered (January 2016). Viral Abundance and Diversity. Detection of Viruses  Detection of viruses is critical to disease identification: © Dennis Kunkel Microscopy/Science Source.  Some diseases have specific symptoms, such as mumps or measles.  Light microscopes look for cytopathic effects (e.g., syncytia or giant cells in RSV).  Electron microscopes examine cellular Figure 15.17: Cells and viruses. components.  Serology looks for antibodies.  PCR 29  Cultivation of Viruses  In a primary cell culture, cells form a monolayer in a culture dish and cytopathic effects are noted.  Viruses can also be identified by the formation of plaques—clear zones within the monolayer—and phage typing of the plaques, as specific strains create characteristic plaques. © James King-Holmes/Science Source. Courtesy of Giles Scientific Inc., CA, Courtesy of Greg Knobloch/CDC. www.biomic.com. Figure 15.18: Infection of cells in embryonated eggs and cell culture. 30 Today’s Tools to Study Microbiology  Advanced microscopes  High throughput technologies  Automation  Advanced computer analysis systems, data storage, data retrieval etc… (bioinformatics)  Model systems  The “OMICS” - 31 Summary  Basic bacterial and viral structures and their function  Host immune response differs between microbes  Modern tools for the study of microbiology 32  integers deaths 251 ally pre job BIOL341/981: Infection & Immunity Microbial Diseases and Prevention Martina Sanderson-Smith Lecture 3, Week 2 [email protected] 31st July 2024 1 Page 1  Lecture 3 Outline 1. Development and progression of disease 2. Viral infection: SARS as an example 3. Bacterial infection: H. pylori 4. Host susceptibility and resistance 2 Page 2  Lecture 3 Outline 1. Development and progression of disease 2. Viral infection: SARS as an example 3. Bacterial infection: H. pylori 4. Host susceptibility and resistance 3 Page 3  Agents of Infectious Disease Viruses Bacteria Protozoa (protists) Fungi Helminths (worms) 4 Page 4  b if The Progression and Outcome of Infection and Disease i S Eration my station relo ifeng.dtens fm.ir 5 inflationcontinually Youldreopers Page 5  Stages of Disease Progression met Incubation period: time between entryexposure v8path 1. of the microbe and onset of symptoms. 2. Prodromal: mild signs or symptoms. i think I'm getting sick 3. Acute: symptoms are most intense and disease-specific. Also called 4. the climax. most disease specificsymptoms Decline: A period of decline occurs as signs and symptoms subside. Might 5. Convalescence: Return to normal health Figure 20.10: The course of an infectious disease. JE 6 rigging ayy.my yggggyyy stops proliferating does Imagine system has restructured body a lot medication tissue cells used of energy to allow this phase endary infection occur fail to do so Page 6  guitarists Sequence of events leading to an infectious disease Eisai_want Invasiveness: Portal of entry: Linked to virulence determinants midriff The route by which an exogenous pathogen enters the host Figure 20.5 Infectious Dose: The number of microorganisms or viral particles needed to initiate infection 7 eg typhoid 100 cells few million cells cholera dose infectious according to risk variys Page 7 gffidsPortals of exit from the body Portal of Exit: Many portal of exit are the same sites as portal of entry. Pathogens often leave the body via bodily secretions and excretions produced at those sites. Figure 20.9 Figure 20.13 8 Page 8  Modes of Infectious Disease e 9 Transmission S Direct: Indirect Idiots EE o Person-person contact o Inanimate objects o Exchange of bodily fluids o Vector transmission (arthropods) o Droplet/aerosols Biological Vector o o Animal contact During labour or delivery www.Ii.int Mechanical Vector o Vehicle Transmission Water Food See Figure 20.12 in Pommerville Page 9  Examples & Comparison wifsiftin Mode of transmission Bacteria away Virus Parasite Person to person contact Eg. Neisseria gonorrhoeae - Eg. HIV - acquired immune Eg. scabies mites, hair gonorrhoea in mucus deficiency syndrome from lice secretions semen/vaginal fluid; Ebola virus in blood Aerosols Eg. Bordetella pertussis - Eg. Influenza virus Rarely... whooping cough Food or water contamination Eg. Samonella typhi - Eg. Rota virus - diarrhoea Eg. Giardia lamblia – Salmonellosis (diarrhoea, Giardiasis fever) (gastroenteritis) Insect bite Eg. Yersinia pestis – Bubonic Eg. Ross River virus - Ross Eg. Plasmodium spp. - plague (via flea) River fever (via mosquito) malaria (via mosquito); Trypanosoma - African sleeping sickness (via Tsetse fly) Animal carrier Eg. Bacillus anthracis – Eg. Lyssavirus – includes Eg. Echinococcus Anthrax (via domesticated or rabies and other diseases granulosus - hydatid wild animals) (via bats) disease (via dogs) 10 Page 10  Lecture 3 Outline 1. Development and progression of disease 2. Viral infection: SARS as an example 3. Bacterial infection: H. pylori 4. Host susceptibility and resistance 11 Page 11  Viral Infectious Disease Viruses cause many different types of disease: o Colds (Rhinovirus, Coronavirus, Coxsackie virus and Echovirus) o Flu (Influenza virus type A, B and C) o Diarrhoea (Rotavirus) o Polio o Hepatitis (Hepatitis virus type A, B and C) o Smallpox o SARS - severe acute respiratory syndrome (Coronavirus) 12 Page 12  Viral Infectious Disease © Gary Gaugler/Medical Images RM. SARS is an emerging infectious disease caused by the coronavirus SARS CoV. 2000s early o SARS spreads through close person- to-person contact. Figure 16.7: Coronaviruses. o Many patients are asymptomatic. o May result in only dry cough and labored breathing, but severe infections cause pneumonia and may require mechanical ventilation due to lack of oxygen reaching the blood. Middle East respiratory syndrome (MERS) is similar to but more dangerous than SARS. to not who announced Spread of SARS. named based on region Just L Emine 13 elitetenographic Page 13  Viral Infectious Disease SARS - severe acute respiratory syndrome (Coronavirus) SARS emerged in China in November 2002. The outbreak became a global epidemic after a doctor from Guangdon infected several people at a Hong Kong hotel. From Hong Kong, the disease spread rapidly to more than 24 countries in North and South America, Europe and Asia. SARS spread so rapidly because symptoms were delayed 2-7 days, via air travel. WHO reports that 8,000 people were infected, with a death rate of 10%. 14 Page 14 regiviredpeknetsenogntae oa.it Epidemiology of SARS Outbreak Primary Index Case Individual persons infected by primary Secondary cases and index case in Honk contacts who spread Kong Hotel the infection 15 Page 15  Viral Infectious Disease of SARS - symptoms and spread: system portal entry SARS symptoms include resp o high fever o diarrhoea o headache and body ache o dry cough after 2-7 days o atypical pneumonia SARS is spread via air droplets and www.cdc.gov close contact and can survive for 6+ hours on tissue or as an aerosol 16 Page 16  Viral Infectious Disease SARS – pathogenesis: o Enters the body through the respiratory tract o Infects the epithelial cells of the respiratory tract then infects infiltrating and circulating immune cells. o Circulating immune cells carry the virus to other organs (spleen, lymphoid tissue etc) o Weakened immune system leads to rapid deterioration and pneumonia the vulnerable elderly ghildren immunocompromised Gu et al. 2005 202 (3) p415–424 17 Page 17  Viral Infectious Disease SARS - genome sequence: The genome sequence of the SARS coronavirus was published in May 2003. Coronavirus belong to Group IV single-stranded (+) sense RNA. 5 Kb 10 Kb 15 Kb 20 Kb 25 Kb 30 Kb i.info is Replicase 1A Replicase 1B Structural proteins Replicase is a polymerase enzyme that catalyses self replication of single stranded RNA and made up a significant portion of its 30Kb genome. 18 Science 300: 1399‐1404 Tiny genome, relative to bacterial (~ 5 million bp) or human (3 billion). SARS-COV-2 similar size. Page 18  Viral Infectious Disease greastfated IBV BCoV SARS - phylogeny:  Molecular phylogeny based on the MHV TGEV gene sequence of the Replicase 1A gene suggests SARS virus is not HCoV-229E SARS closely related to other coronaviruses. SARS - origin:  Epidemiological (PCR and sequence analysis) studies have shown that bats are the natural reservoir of SARS virus. 19 against wave wous Etf circuit see.is atem Page 19  Lecture 3 Outline 1. Development and progression of disease 2. Viral infection: SARS as an example 3. Bacterial infection: H. pylori 4. Host susceptibility and resistance 20 Page 20  Bacterial Infectious Disease Bacteria cause many different types of disease: o Sore throat and flesh eating disease (Streptococcus pyogenes) o Anthrax (Bacillus anthracis) o Tetanus (Clostridium tetani) o Tuberculosis (Mycobacterium tuberculosis) o Gonorrhoea (Neisseria gonorrhoeae) o Plague (Yersinia pestis) o Stomach ulcers (Helicobacter pylori) 21 Page 21  Bacterial Infectious Disease Helicobacter pylori - the causative agent of gastritis and EGstomach ulcers o Helicobacter pylori first described by Barry Marshall and Robin Warren in 1983 in Perth - awarded the 2005 Nobel prize o Stomach and duodenal ulcers originally thought to be caused by stress and diet not elective by postulates o Treatment by antacids originally but now antibiotics o Approximately 2/3 of the human population is infected with H. pylori, yet the majority do not develop disease whitemicrobiome weather.fmtffgfatsa https://www.mja.com.au/journal/2005/183/11/2005-nobel-prize-physiology-or-medicine#0_i1091639 22 Page 22  Bacterial Infectious Disease Helicobacter pylori - symptoms and spread: H. pylori symptoms include: o burning stomach pain o nausea o vomiting o inflammation and bleeding www.gicare.com The transmission method of H. pylori is not completely understood, but it is more common with increased age and in less developed countries. Faecal-oral, oral-oral transmission? Contaminated food or water sources? 23 Page 23  Bacterial infectious disease Helicobacter pylori – infection: H. pylori resides in the mucosal layers of the stomach. The helical jÉÉ shape is thought to facilitate movement through the mucosal layer. to H. pylori urease breaks down urea in the stomach into ammonia. The production of an "ammonia cloud" around the bacterium mine protects it from the low pH of the stomach, where few other organisms are able to survive. urease C=O(NH2)2 + 2H2O CO2 + 2NH3 urea 24 Page 24  is.ms iii Bacterial Infectious Disease Helicobacter pylori - infection: I expose Tsified 25 Page 25  Genomic 1 may changesthe way 26 host cells behave Page 26  Bacterial Infectious Disease Helicobacter pylori - gastric cancer: prisoner  Epidemiological studies have shown an association between H. pylori and the development of stomach cancer. H. pylori infection is considered a major risk factor, though not necessarily a direct cause. www.gastrolab.net 27 Page 27  Fungal, protozoan, and parasitic infectious disease Eukaryote micro-organisms also cause many different types of disease: o Thrush (Candida albicans) o AIDS-related infections (Pneumocystis carinii) o Malaria (Plasmodium spp.) o African sleeping sickness (Trypanosoma brucei) o Intestinal worm infections (Ascaris spp.) o Hydatid disease (Echinococcus granulosus) my deportation POEX factors host 28 AND Page 28  Lecture 3 Outline 1. Development and progression of disease 2. Viral infection: SARS as an example 3. Bacterial infection: H. pylori 4. Host susceptibility and resistance 29 Page 29  Severity of Infectious Disease pod said Infectious disease severity is governed by; Virulence determinants of pathogens o Attachment systems o Toxins/enzymes o Self-destruction o Changing antigens o Camouflage Host factors o Human susceptibility o Resistance genes 30 Page 30  Severity of Infectious Disease Virulence determinants include: Virulence determinant Role during infectious disease ECM adhesins Colonisation Fimbriae/Flagella Colonisation Siderophores Iron acquisition Lactoferrin binding proteins Iron acquisition Proteases/Enzymes Disruption of immune system, nutrient acquisition, cell lysis, ECM/tissue damage Capsule Camouflage - anti-phagocytosis Toxins Disrupt host cells, host function, immunity Urease Protects against acid 31 Page 31  Severity of Infectious Disease Human susceptibility and resistance genes include: we Gene Resistance/ susceptibility Infectious agent malaria if Hbs (sickle cell gene) R Plasmodium falciparum Duffy blood group glycoprotein S Plasmodium vivax CCR5 receptor polymorphisms R HIV Vitamin D receptor polymorphisms R/S Mycobacterium tuberculosis Human leukocyte antigen (HLA) R/S Streptococcus pyogenes 32 Page 32  Host Resistance Genes 33 mid feed Progressing CCR5 polymorphism and HIV CD4+ T cells (Th cells) are the primary target of HIV infection forget CCR5 and CXCR4 are chemokine receptors present on immune cells HIV uses CCR5 as a secondary receptor to gain entry into macrophages phagocytes cell Mutations in CCR5 (e.g. CCR5-wt/Δ32) have been associated with a slower progression towards AIDS delayprogrammised CD4+ T cells = T-helper cells. They are the primary target of HIV infection. Their destruction leads to weakening immune system (basis for AIDS). Binding of gp120 to CD4 receptor causes conformational change and then binding to CCR5, and/or CXCR4. Page 33  Host Resistance Genes whataddthis.int dictates HIV tropism HIV strains display tropism for the different chemokine cCR5 receptors Some strains, known as X4- tropic or dual-tropic strains, can engage CXCR4 This strain can emerge as a result of mutation within the individual Kerina Duri (2012). Coreceptor Usage in HIV Infection, Can overcome host resistance Immunodeficiency, Prof. Krassimir Metodiev (Ed.), ISBN: 978-953-51-0791-0, InTech, DOI: 10.5772/52060 conferred by CCR5 mutation low 34 would evolve to CCRs infected it eg target CXCR4 rapid mechanisms wave negate come over Page 34  Host Susceptibility and Resistance Genes flesh eaffase Group A streptococcal invasive diseases include necrotising fasciitis I and toxic shock syndrome Severity is associated with human leukocyte antigen (HLA) class II polymorphism Patients with necrotising fasciitis, caused by S. pyogenes. Limb may require amputation for the survival of the patient. www.nnff.org www.nnff.org 35 Page 35  Host Susceptibility and Resistance Genes cells Human leukocyte antigen (HLA) site on surfimture HLA is the name given to the human form of the expressed Major Histocompatibility Complex (MHC) MHC is a large locus in all vertebrates that encodes for surface proteins essential in immune function. There are two classes of MHC HLA class II (also called MHC class II) are expressed primarily on antigen-presenting cells of the immune system, such as macrophages Importantly, HLA class II is highly polymorphic (variable) 36 Page 36  Host Susceptibility and Resistance Genes Usually, class II HLA binds to T cell N receptor specifically and reversibly This is a crucial process of the adaptive immune response APC Group A streptococci produce super- antigens that non-specifically cross- link HLA class II proteins and TCR This causes massive T cell over- f proliferation and inflammatory TCR cytokine production Excessive, systemic inflammatory response can lead to organ damage superantigen N and death TH cell Some HLA haplotypes are less vulnerable (more resistant) to superantigen-mediated cross-linking Nature Medicine 8: 1398-1404 37 Page 37  Lecture 3 Outline 1. Development and progression of disease 2. Viral infection: SARS as an example 3. Bacterial infection: H. pylori 4. Host susceptibility and resistance 38 severity reliant SARS Dyson PG affecting te piscate VIRAL Bacterial Page 38  BIOL341/981: Infection & Immunity Microbial Diseases and Prevention Martina Sanderson-Smith Lecture 4, Week 2 [email protected] 31st July 2024 1 1 Page 1  Lecture Outline 1. Public health measures 2. Chemotherapeutics A. Penicillin B. Vancomycin 3. Immunotherapeutics A. Interferon alpha 4. Action and efficacy of antimicrobials 5. Antimicrobial resistance 6. Bacteriophages 2 2 Page 2  Lecture Outline 1. Public health measures 2. Chemotherapeutics A. Penicillin B. Vancomycin 3. Immunotherapeutics A. Interferon alpha 4. Action and efficacy of antimicrobials 5. Antimicrobial resistance 6. Bacteriophages 3 3 Page 3  Public Health Measures Following the discovery of microorganisms by Pasteur, Koch and others, measures were taken to reduce the incidence and transmission of microorganisms. Such measures include; o clean water o sewerage systems o reduction in overcrowding o better food handling and nutrition o sterilisation methods o use of disinfectants and antiseptics o education on microbial transmission Public health measures are comparatively cheap, cost effective, and save millions of lives every year. 4 4 Page 4  f miiini.IE Public Health Measures England/Wales death rates Measles Scarlet fever Typhoid Whooping cough Diptheria Deaths per 100,000 Penicillin discovered in 1928, but only widely available in 1940s.... year 5 5 Page 5  Lecture Outline 1. Public health measures 2. Chemotherapeutics A. Penicillin B. Vancomycin 3. Immunotherapeutics A. Interferon alpha 4. Action and efficacy of antimicrobials 5. Antimicrobial resistance 6. Bacteriophages 6 6 Page 6  Chemotherapeutics Chemotherapeutic agents are natural or synthetic substances which are either toxic or growth inhibitory to microorganisms (or cancer) and are used internally in humans. Relenza (flu - influenza virus type A, B and C) Antibiotics (antibacterials - eg. penicillin, streptomycin, vancomycin) Quinine (malaria) Artemisinin (malaria) Metronidazole (systemic amoebiasis) 7 7 Page 7  we E've Natural antibiotics: Produced by Microbes Secondary metabolites, or chemical compounds not directly used by the producing organism for growth and reproduction. – Often produced in the stationary phase of growth to help compete for limited resources. – They might help the producer kill competitors that are sensitive to the antibiotic. – They give the producer a selective advantage for nutrients and space. Possible roles of antibiotics. 8 firitimer Page 8  AB is antibiotic Eat Mode of Action 5 major targets for antibacterial agents inhibits Cell wall Cell membrane Ribosomes (protein synthesis) Nucleic acid synthesis (RNA and DNA) Metabolic reactions 9 9 Page 9  Beta-Lactam Antibiotics Target Bacterial Cell Wall Synthesis – Pencillin was the first beta-lactam discovered – Discovered by Alexander Fleming (1928), studied in greater detail by Howard Florey (1938-1941) – Active against many gram-positives and some gram-negatives – They interfere with cell wall synthesis in rapidly growing cells, – Specifically interfere with cross-linking of peptidoglycan layers – Exhibits greatest effect on rapidly growing populations of bacteria – More dormant communities may not be as affected slow – Allergic reaction and evolution of resistance are problematic 5 The Action of Beta-Lactamase on Sodium Penicillin G. 10 metamate L p viii finger peptides Page 10  Other Antibiotics Target Metabolism Glycopeptide antibiotics inhibit bacterial cell wall synthesis. gram ve bacteria – Vancomycin: o Effective against gram-positive bacteria such as staphylococci. o Side effects include damage to the ears and kidneys. o Used as a drug of last resort, especially to treat MRSA. 11 Page 11  Chemotherapeutics was Vancomycin: antibiotic of "last resort" probe Vancomycin is a 1.5 kDa glycoprotein used to kill bacteria (not viruses or parasites etc). This antibiotic is used to kill multiple resistant bacteria such as methicillin resistant Staphylococcus aureus (MRSA). Vancomycin is produced by the soil bacterium Amycolatopis orientalis. Vancomycin was first deployed in 1956, while the first resistant Enterococcus spp. were detected in 1988. www-che.syr.edu 12 12 Page 12  Chemotherapeutics Vancomycin: mode of action Vancomycin kills bacteria by interfering with cell wall synthesis Binds to D-ala terminus, which interferes with transpeptidase activity Peptidoglycan layers cannot be cross-linked, resulting in weakness of fall EE peptidoglycan Vancomycin binding site www-che.syr.edu 13 13 In WHEN Page 13  Chemotherapeutics Vancomycin: origin of resistance Antibiotics have been used as "growth promoters" in intensive agriculture (chickens/pigs etc) since the 1960s. o Avoparcin extensively used, structurally similar to vancomycin. o Avoparcin resistant Enterococci isolated from animals are also vancomycin-resistant. o Europe banned Avoparcin in 1997. o A set of resistance genes (van genes) encodes resistance. 14 14 Page 14  Chemotherapeutics Vancomycin: mechanism of resistance Van genes encode proteins that modify peptidoglycan structure, resulting in vancomycin losing the capacity to bind, and hence to block peptidoglycan synthesis. wait D-lactate we peptidoglycan Vancomycin binding site D-lactate P aka http://www.sumanasinc.com/scienceinfocus/sif_antibiotics.html 15 www-che.syr.edu 15 afore diffide Page 15  Polypeptide Antibiotics Affect the Cell Membrane Topical use only - unacceptable therapeutic window for internal use - cause renal failure and respiratory paralysis Bacitracin interferes with transport of cell wall precursors through the membrane. – skin infections by gram-positive and gram-negative bacteria. – Combined with neomycin and polymyxin B, it is sold as Neosporin. Polymyxins increase membrane permeability of gram- negative rods, leading to cell death. – Polymyxin E is used against many superbugs. – Polymyxin B and bacitracin are combined to make Polysporin. 16 Page 16  Many Antibiotics Affect Translation Aminoglycosides attach to bacterial ribosomes, blocking transcription. – Streptomycin was discovered in 1943 by Waksman. Antibiotics and their affect on protein synthesis. 17 Page 17  Lecture Outline 1. Public health measures 2. Chemotherapeutics A. Penicillin B. Vancomycin 3. Immunotherapeutics A. Interferon alpha 4. Action and efficacy of antimicrobials 5. Antimicrobial resistance 6. Bacteriophages 18 18 Page 18  Immunotherapeutics Immunotherapeutic agents are molecules of the human immune system that are used to "boost" the immune response against microorganisms (or cancer)  Interferon alpha (viruses)  Humanised antibodies (all infectious agents)  Interlukin-2 and interlukin-12 (viruses)  Antibody-toxin conjugates (antimicrobial and anti-cancer) 19 19 Page 19  Immunotherapeutics Interferon alpha: role in immune system. Interferon alpha is produced by many cell types, including B cells, T cells and macrophages and exerts important antiviral role Recombinant interferon alpha is used in the treatment of Hepatitis C virus. en.wikipedia.org 20 20 Page 20  nature boost Immunotherapeutics infection Interferon alpha: mode of action. Interferon alpha is part of the innate antiviral response Induces expression of other host antiviral proteins (protein kinase, 2'-5' oligoadenylate synthetase) which block viral production Although best known for antiviral activity, IFNs have recently been shown to be induced by, and active against – rickettsia, mycobacteria and several protozoa 21 www.med.howard.edu 21 Page 21  Lecture Outline 1. Public health measures 2. Chemotherapeutics A. Penicillin B. Vancomycin signature on warning 3. Immunotherapeutics A. Interferon alpha give eventsThan to m 4. Action and efficacy of antimicrobials 5. Antimicrobial resistance 6. Bacteriophages 22 22 Page 22  ethical abets wilt Characteristics of Antimicrobial Drugs Readily Available Inexpensive Chemically stable (so that it can be transported easily and stored for long periods of time) Easily administered Non-toxic and non-allergenic Selectively toxic against a wide range of pathogens 23 23 Page 23  Clinical Considerations in Prescribing Antimicrobial Drugs Spectrum of Action – The number of different bacterial pathogens a drug acts against o Narrow spectrum (eg. Penicillin) o Broad spectrum (eg. Tetracycline) Efficacy (versus toxicity) - ascertained by: o Diffusion susceptibility test o Minimum inhibitory concentration test o Minimum bactericidal concentration test 24 24 Page 24  Therapeutic Window Selective toxicity says a drug should harm the pathogen but not the host. – The toxic dose of a drug is the concentration causing harm to the host. – The therapeutic dose is the concentration eliminating pathogens in the host. – Together, the toxic and therapeutic doses are used to formulate the therapeutic window. A Representation of the Therapeutic Window. 25 Page 25  Antimicrobial Spectrum Drugs have a range of pathogens on which they will work, which is known as the antimicrobial spectrum. nisheyaifof.rs – Broad-spectrum drugs affect many taxonomic groups. – Narrow-spectrum drugs affect only a few pathogens. broad narrow The Antimicrobial Spectrum of Activity. 26 Page 26  Antibiotic Susceptibility Assays Tube dilution method: decreasing concentrations to determine the lowest concentration of the antibiotic that is effective. mineralisition MIC: The smallest amount of the drug that will inhibit the growth and reproduction of the pathogen 27 Page 27  published abide determine MC MIC – result interpretation Plot data – easy identification of break-point Compared to known MIC values in break-point tables MIC 28 28 Page 28  Colonies acone Amniotes widthMinimum bactericidal concentration test (MBC) MBC: Determines the amount of drug required to kill the microbe rather than just the amount to inhibit it. The lowest concentration of drug for which no growth occurs in the subculture is the MBC 29 29 Lust dont E netflix.ie iiiide our Page 29  Plate based Methods Etest: Determines MIC using an antibiotic-infused strip placed on an agar plate. iii Disk diffusion method: antibiotics Courtesy of bioMerieux, Inc. diffusing from paper disks on a bacterial confluent growth. - Both Etest and disk diffusion method involve measuring the zone of inhibition of bacterial growth. Figure 10.14A: Figure 10.14B: Kirby- Etest Method. Bauer Method. 30 30 Page 30  Zone of inhibition in a diffusion susceptibility test Susceptible Resistant ELI 31 31 Page 31  Zone of inhibition – result interpretation Compare inhibition diameter to known data in break-point tables (published by manufacturer) Species-specific values for each drug diff MC none fewanee Abbreviation Zone Resistant Intermed Sensitive Sensitive/ Antibiotic on disc Diameter Diameter Diameter Diameter Resistant Streptomycin S10 15 mm Ampicillin AM 15 mm Gentamicin GM 15 mm Erythromycin E 23 mm Chloramphenicol C 18 mm Tetracycline Te 19 mm 32 32 Page 32  Lecture Outline 1. Public health measures 2. Chemotherapeutics A. Penicillin B. Vancomycin 3. Immunotherapeutics A. Interferon alpha 4. Action and efficacy of antimicrobials 5. Antimicrobial resistance 6. Bacteriophages 33 33 Page 33  Figure 10.18: Timeline for antibiotic introduction and appearance of antibiotic resistance. 34 Page 34  resisted preadfari snowing Resistance to Antimicrobial Drugs 35 35 Page 35  Resistance to Antimicrobial Drugs The Development of Resistance in Populations — Some pathogens are naturally resistant — Resistance can be acquired by bacteria in two ways L  New mutations of chromosomal genes  Acquisition of R-plasmids via transformation, transduction, and it war conjugation Darwinian theory of evolution = survival of the fittest 36 36 Page 36  Multiple strategies of antibiotic resistance _splitethyk to sina.ttcell or Antibiotic hydrolysis. streptomycinfosphate group Antibiotic modification. Membrane modification/efflux. Target modification. Metabolic pathway alteration. no overcome side effects the Figure 10.20: Antibiotic Resistance Tactics. 37 Page 37  Resistance to Antimicrobial Drugs Improper or excessive use of antibiotics causes antibiotic resistance. – If resistant strains spread to other patients, a superinfection occurs. Three factors have contributed: – Prescription abuse. – Prescription misuse by individuals. – Prescription misuse by healthcare organizations. Figure 10.21: How antibiotic resistance develops. 38 Page 38  Resistance to Antimicrobial Drugs Limiting Resistance Maintain high concentration of drug in patient for sufficient time o Kills all sensitive cells and inhibits others so immune system can destroy (PK/PD modelling) Use antimicrobial agents in combination (synergy) Use specific antimicrobials and only when necessary Develop new variations of existing drugs a variety o Second/Third-generation drugs to overcommen Search for new antibiotics, semi-synthetics, and synthetics o Bacteriocins e.g. antimicrobial peptides (BLIS; http://blis.co.nz/) o Identify new bacterial proteins to target 39 39 Page 39  Lecture Outline 1. Public health measures 2. Chemotherapeutics A. Penicillin B. Vancomycin 3. Immunotherapeutics A. Interferon alpha 4. Action and efficacy of antimicrobials 5. Antimicrobial resistance 6. Bacteriophages 40 40 Page 40  Bacteriophage Therapeutics Bacteriophages: viruses that infect bacteria o Bacteriophages cannot reproduce and survive on their own, take over host cell o Bacteriophage have 2 distinct life cycles waited lytic (T4) lysogenic (lambda) Lytic phage may perhaps be weaponised against pathogenic bacteria. 41 41 Page 41  Feet detention 42 42 Page 42  Bacteriophage: Lytic life cycle 1. Adsorption to the host cell and penetration Phages then inject DNA into the cell 2. Synthesis of phage nucleic acids and proteins Assembly of phage particles 3. Release of phage particles (300 in 22 min) Many phages lyse their host by damaging the cell membrane and cell wall Holin – enzyme which destabilizes the host cell membrane (pokes holes) Lysin – phage enzyme which breaks host cell wall (lyses host bacteria) Utilisation of either complete phage or holin and lysin proteins for therapy? 43 43 Page 43  Bacteriophage Therapeutics Bacteriophages: medical applications Need for alternative therapies for treating bacterial infections o resistance exists to every antibiotic we have Phages are potent anti-bacterials Self-replicating (smart drugs?) Narrow specificity so don’t damage the normal flora Resistance not as significant Resurgent interest in the application of phages to agriculture and human health Used for years in Eastern Europe and Russia Reassessment of Medicinal Phage Spurs Companies to Study Therapeutic Uses. ASM News 64:620-623, 1998. Phages eyed as agents to protect against harmful E. coli. ASM News 65:666-667, 1999. 44 44 Page 44  Bacteriophage Therapeutics Bacteriophages: medical applications Clearance from circulatory system (not a problem if topical application) Immune response against phage upon repeated use Access to site of infection http://ehp.niehs.nih.gov/ 45 45 Page 45  Bacteriophage Therapeutics Bacteriophages: medical applications https://www.sciencemag.org/news/2019/05/viruses-genetically-engineered-kill-bacteria- rescue-girl-antibiotic-resistant-infection 46 46 Page 46  commandments seventh Other Antimicrobial Drugs Target Viruses, Fungi, and Parasites Antiviral drugs interfere with viral replication. – Virus attachment/penetration: Several classes of antifungal drugs cause membrane damage The goal of antiprotozoal agents is to eradicate the parasite Antihelminthic agents target non-dividing Types of Human Infectious Disease. Helminths 47 Page 47  Lecture Outline 1. Public health measures 2. Chemotherapeutics A. Penicillin B. Vancomycin 3. Immunotherapeutics A. Interferon alpha 4. Action and efficacy of antimicrobials 5. Antimicrobial resistance 6. Bacteriophages 48 48 Page 48  A BAD DAY AT THE OFFICE A work lunch that almost resulted in death! Started with acute vomiting, headache and fainting followed by severe diarrhea and delirium ICU for a week – multiple organ shut-down within 24 hours of onset 5 week recovery in hospital and months at home 1  BIOL341/982 Infection and Immunity Interactions of Microbes with the Host: Colonisation and Survival Mechanisms Prof. Martina Sanderson- Smith [email protected] 2 A BALANCING ACT Microbe Factors Microbe burden 3  A BAD DAY AT THE OFFICE: Shigella dysenteriae Produces shiga toxin = one of the most toxic and lethal toxins known 4 MICROBIAL COLONISATION Lecture 5 outline Colonisation, attachment and invasion survival 5  ENTRY AND EXIT § Pathogens enter and exit the host through portals § Portal of entry: Refers to the site at which the pathogen enters the host §Varies considerably between organisms, and is a key factor in the establishment of disease § Portal of exit: Refers to the site from which the pathogen is able to leave the body after the end of its pathogenicity cycle § Easy transmission enables organism to continue its pathogenic existence § Shed in large numbers in secretions and excretions § Present in the blood for uptake – e.g. blood sucking arthropods 6  COLONISATION § Colonisation is the establishment of a stable microbial population in the host § Attachment to host cells plays a major role in colonisation § First step in development of an infectious disease § Primary sites of colonisation include: § Respiratory tract § Intestinal tract § Reproductive tract § Urinary tract § Skin, mucosa 7  COLONISATION § Host organisms have developed many ways to try and prevent microbial colonisation: Type of Host Defences Microbial Evasion Mechanism Examples Infection Respiratory tract Mucociliary clearance Adhere to epithelial cells, interfere with ciliary Influenza virus action Alveolar macrophage Replicate in alveolar macrophage Legionella, tuberculosis Intestinal tract Mucus, peristalsis, acidic Adhere to epithelial cells, Resist acid, bile Rotavirus, environment, bile, gut Salmonella, flora Helicobacter pylori Reproductive Flushing action of urine Adhere to urethral/vaginal epithelial cells Gonococcus, tract and secretions, mucosal Chlamydia defences Skin, mucosa Layers of constantly Invade skin/mucosa Measles shed cells (mucosa), Penetrate intact skin Staphylococci Dead keratinised skin layers Streptococci 8  POST-COLONISATION § Some pathogens remain at the site of infection, whereas others invade host tissues Bacterial pathogens restricted to epithelial surfaces include: Respiratory tract Urogenital tract Skin Intestinal tract Mycoplasma pneumoniae Neisseria gonorrhoeae Staphylococcus aureus Salmonella enterica (Atypical pneumonia) (Gonorrhea) (Skin infections) (Food poisoning) Bordetella pertussis Proteus mirabilis Streptococcus pyogenes Shigella spp. (Whooping cough) (Urinary tract infection) (Impetigo) (food poisoning) Corynebacterium Escherichia coli Campylobacter jejuni diptheriae (Diphtheria) (Urinary tract infection) (Food poisoning) Streptococcus pyogenes Vibrio cholerae (Pharyngitis) (Cholera) Escherichia coli (Food poisoning) 9  POST-COLONISATION § Post-colonisation, some pathogens remain at the site of infection, whereas others invade host tissues Bacterial pathogens that cross epithelial surfaces include: Respiratory tract Urogenital tract Skin Intestinal tract Mycobacterium Treponema pallidum Bacillus anthracis Salmonella typhi tuberculosis (TB) (syphilis) (anthrax) (Typhoid fever) Yersinia pestis Streptococcus pyogenes (plague) (Necrotising fasciitis) Legionella pneumophila (Legionnaire’s) 10  VIRAL ATTACHMENT TO HOST CELLS AND TISSUES Viruses gain access to host cells by binding to specific receptors expressed on the surface of host cells. 11 VIRUSES HAVE A HOST RANGE AND TISSUE SPECIFICITY § A host range refers to what organisms the virus can infect and depends on capsid or envelope structure. § Many viruses infect certain cell or tissue types within the host (tissue tropism). § The virus needs a specific receptor to invade the host cell. 12 VIRAL ATTACHMENT TO HOST CELLS AND TISSUES: Adenovirus § Adenoviruses are a double-stranded DNA virus and a major cause of human respiratory disease § 7 human species and over 50 serotypes § Found in mammals, birds and amphibians § Attach via a Penton fibre: § Projects from each apex § Consist of a slender shaft § Globular head § Different adenovirus subtypes attach to different receptors 13 14 BACTERIAL ATTACHMENT TO HOST CELLS AND TISSUES § Bacterial adhesins: surface structures that bind to specific host receptors § Fimbrial/pilus adhesins § Afimbrial adhesins (capsule/protein adhesins) § “Lock and key" mechanism § Overcome electrostatic repulsion (bacterial and host cells are both negatively charged) fimbrial/pilus adhesins afimbrial adhesins 15  BACTERIAL ATTACHMENT TO HOST CELLS AND TISSUES: Fimbrial Adhesins § Bacterial fimbriae/pili: § Hair-like structures 5-7nm diameter with varied morphology § Structurally diverse and different fimbriae recognise different host receptors § Comprised primarily of protein subunits called pilin “Bald” E. coli Bundle-forming pili CS1-piliated (non-pathogenic) enteropathogenic E. coli enterotoxigenic E. coli (ETEC) (EPEC) 16  BACTERIAL ATTACHMENT TO HOST CELLS AND TISSUES: Fimbrial Adhesins § Shaft - repeating protein subunits (pilins) assembled into a helical array § Adhesive subunit(s) § Specialised tip structure (Pap pili – UPEC) § Recognise specific cells e.g. bladder and kidney cells § No specialised tip structure (N. meningitidis). § Adhesive subunits along length of shaft § Interact with cells via multiple anchorage points Pap (pyelonephritis associated pili) associated with uropathogenic E. coli (UPEC) which causes UTIs (bladder, kidney infections)....... 17  BACTERIAL ATTACHMENT TO HOST CELLS AND TISSUES: Fimbrial Adhesins Type 1 pilus-mediated UPEC attachment to bladder epithelium UPEC Bladder epithelium Mulvey M A et al. PNAS 2000;97:8829-8835 18  BACTERIAL ATTACHMENT TO HOST CELLS AND TISSUES: Fimbrial Adhesins Host and tissue types specified by some E. coli fimbriae: Fimbriae/Pili Host/Tissue Disease Enterotoxigenic E. coli (ETEC) K88 Pig / intestine Diarrhoea K99 Cow, sheep, pig / intestine Diarrhoea CS1 Human/ intestine Diarrhoea Enteropathogenic E. coli (EPEC) Bundle-forming pili (Bfp) Human/ intestine Severe diarrhoea (children) Uropathogenic E. coli (UPEC) Pap Human/Urinary tract Urinary tract infections Type1 non-specific Urinary tract infections Extraintestinal pathogenic E. coli (ExPEC) ExPEC pili Human infant brain, kidney, Meningitis, septicaemia epithelium 19 BACTERIAL ATTACHMENT TO HOST CELLS AND TISSUES: Afimbrial Adhesins Afimbrial adhesins: contribute to tighter binding after pilus anchorage Dehio et al, 1998 Trends Microbiol. 6: 489-495 20  INTRACELLULAR PATHOGENESIS Why intracellular? § Immune evasion § Nutrient rich (in the cytoplasm) § Transport around the body Intracellular survival (olpaimages.nsf.gov/admin/images/listerias.jpg) § Subverting intracellular antimicrobial mechanisms such as phagosome- lysosome fusion, modulating phagosomal pH, damaging phagosomal membranes, and/or quenching microbicidal oxidative bursts § Escape from the phagosome to the cytoplasm § Replicate 21  BACTERIAL TISSUE INVASION § Invasiveness is the ability of a pathogen to invade tissues § Invasiveness encompasses: § Mechanisms for colonisation (adherence and initial multiplication) § Production of extracellular substances ("invasins"), that promote the immediate invasion of tissues – “spreading factor” § Ability to bypass or overcome host defence mechanisms which facilitate the actual invasive process 22 SUCCESSFUL INFECTION OFTEN DEPENDS ON A VARIETY OF ENZYME VIRULENCE FACTORS § Pathogens have to adapt to a new environment when they enter a host. § Enzymes can help pathogens resist body defenses.. 23  BACTERIAL TISSUE INVASION: Staphylococcus Clot formation contributes to staphylococcal skin boil 24 Pommerville