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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