Microbiology: Introduction 1/18/2017 4:02:00 PM
https://mediakron.bc.edu/microbiology
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Intro- How microbes effect us and why we should care about them
What are microbes?
• Organisms so small that they are not easily visible without a
microscope
• They are diverse- bacteria, archaea, eukaryotes (fungi, protists,
animal parasites), viruses, prions
Pathogens were the first to grab our attention
• = microbes that cause diseases
How they effect us….
• Microbes can spoil or preserve foods
• Tooth Decay
• Needed for vitamin C production—deficiency was an issue for
sailors/ pirates
• C. Difficile infection- fatal infection; resident gut microbiota (without
antibiotic perturbation) often blocks infection
• Chronic wound infection
1/20- history of microbiology
History of Microbiology- to understand how things change and develop
Biology started with the discovery of the microscope discovery of
microorgnaisms
• Antoine Van Leeuwenhoek
• Following Hooke’s 1665 use of microscopes to observe plant cells
• Established that there were forms of life that were not visible to the
naked eye; originally referred to them as animalcules (tiny animals)
• Observed bacteria from his teeth plaque
Sterilization and development of germ theory
• Lazzaro Spallanzani- evidence against “spontaneous generation” of
living creatures
o Boiling broth would sterilize it and kill microorganisms in it
o New microorganisms could only settle in broth if it was
exposed to air
• John Snow- tracing the source of cholera; One of the pioneers of
epidemiology
Broad Street Cholera outbreak- major outbreak in London; many died • Dominant theory was miasma theory- diseases such as cholera and
plague spread by “bad air” from rotten organic matter
• John snow investigates.. skeptical of miasma theory; interviewed
locals and mapped area
o Found that sections where the closest source of water was the
Broad street pump had the most cases of cholera public
water pump was the source of the outbreak
• Anomaly: there were no cases in the Broad Street Brewery—got
beer from boiled water
Vaccination- Edward jenner- Smallpox vaccine= world’s first vaccine “The
father of immunology”
• Observed that milk maids were immune to smallpox Jenner
postulated that contact with cow pox blisters protects them from
small pox
o It was known that infection with cowpox could prevent
against infection with smallpox
o Investigators tested cowpox vaccine with success
• Jenner established immunity
• In 1840, the vaccine was offered to the public by the British
government
• “Saved more lives than the work of any other human”
What are the challenges against vaccination now?
Development of Germ Theory- Louis Pasteur
• Founder of medical microbiology
• Germ Theory 1861= growth and reproduction of microbes within
their host can cause diseases
• Suggested forms of sterilization: filtration, exposure to heat, or
exposure to solution/ chemical solutions
• Developed a method for attenuating a virulent pathogen for
immunization (inspired by Jenner’s success with smallpox
vaccination)
Classification of bacteria- Ferdinand Julius Cohn
• His classification of bacteria was based on shapes- rod, spherical,
spiral, thread
Microbes as causal agents of disease • Robert Koch- he found that the blood of cattle who were infected
with anthrax always had large numbers of Bacillus anthracis
• He could transmit anthrax by injecting a small sample of blood from
an infected animal into a healthy animal
• Founded the scientific method of microbiology
Koch’s postulates; to identify disease-causing agents
• The microorganism or other pathogen must be present in all cases
of the disease
• The pathogen can be isolated from the diseased host and grown in
pure culture
• The pathogen from the pure culture must cause the disease when
inoculated into a healthy, susceptible laboratory animal
• The pathogen must be re- isolated from the new host and shown to
be the same as the originally inoculated pathogen
Discovery of viruses, environmental microbiology
• Martinus Beijerinck- discovery of viruses; development of
enrichment culture techniques
• Sergei Winogradsky- developing the concept of chemoautotrophy;
Nitrogen cycle; Winogradsky columns
Discovery of Archaea- Carl Woese
• Based on differences in Ribosomal RNA
• Wasn’t accepted without resistance at first
Notable discoveries
st
• 1878 Lister- developed the 1 method to isolate a pure bacterial
culture (lactic fermentation of milk)
• 1880 Pasteur- developed a method for attenuating a virulent
pathogen for immunization (inspired by Jenner’s success at
smallpox vaccination)
• 1881 Koch- developed solid media for cultivating cells on the
surface of the plate (simplified the process of isolating microbes)
• 1884 Koch- identified tubercle bacillus as the agent that caused
Tuberculosis and formulated the Koch postulates Noble prize in
1905
• 1889 Beijernick- developed enrichment cultures for isolating a pure
culture if Rhizobium • 1891 Ehrlich- proposed that antibodies are responsible for immunity
against diseases Noble prize in 1908
• 1893 Smith and Kilbourne- established that ticks carry Babesia
microti (which causes babesiosis in animals and humans), providing
a foundation for animal host studies and insect vectors
• 1915/1917 Twort & d=Herrelle- discovery of bacteriophage (virus
that infect bacteria)
• 1928 Griffith- discovered transformtion in bacteria, establishing the
field of molecular gentics
• 1929 Felming- discovered penicillin and its effect on bacteria,
starting the “Antibiotic Era” Nobel prize in 1945
• 1935 Stanley- crystallized tabacco mosaic virus and showed in
remained infectious Nobel prize in 1946
• 1941 Beadle and Tatum- used fungus Neurospora crassa to
establish the link between enzymes and corresponding genes (one
gene- one enzyme) Nobel prixe in 1958
• 1943 Luria and Delbruck- shows that inheritance in bacteria follows
Darwinian principles Nobel prize in 1969
• 1946 Lederberg and Tatum- discovered conjugation in bacteria
• 1949 Enders, Weller, and Robbins- developed technique to grow
viruses in culture Nobel prize in 1954
• 1952 Lederberg and Zinder- transduction: transfer of genetic
information by viruses
o a phage of salmonella can carry DNA from one bacterium to
another
• 1961 Brenner, Jacob and Meselson- used phage-infected bacteria to
show that ribosome’s are the site of protein synthesis
• 1982 Prusiner- discovery of prions Nobel prize in 1997
• 1983 Mullis- used heat-stable enzyme from Thermus aquaticus to
establich PCR Nobel Prize in 1993
• 1993-2013 Mojica, Bolotin, Siksnys, Doudna, Church and Zhang-
Discovery of CRISPR-Cas9 bacterial adaptive immunity machinery,
which enabled flexible gene editing Nobel prize to come soon
Transition to Genomic era
• Genomics has revolutionized every aspect of microbiology, from
tools to conceptual frameworks • Whole genome sequencing has become a common tool to reveal the
genetic potential of organisms
• Application not limited to analysis of genes
• Sequencing to us is what microscope was to van Leeuwenhoek
Tools for modern microbiology
• Genomics: Study of the DNA/ genomes of organisms
• Transcriptomics: Studying RNA molecules in one cell or a potulation
of cells
• Metabolomics: Studying molecules involved in cell metabolism
• Proteomics: Studying the profile, structure, and abundance of
protiens
Form single species studies to communities- many microbial functions
happen within assemblies of interacting species called communities, not as
acts of isolated species
• What microbes do depends on the environment theyre in, and that
environment depends on other organisms
• Requires a shift in studies towards examining microbial activities in
communities
o Examining monocultures might not be enough 1/ 23 Prokaryotes and Eukaryotes 1/18/2017 4:02:00 PM
Bacteria- unicellular organisms
• prokaryotic because they lack a nucleus
• around for more than 3 billion years; evolved into complexity with a
lot of capabilities
• 4 categories by shape= rod, spherical, spiral, curved
• divide asexually; divide by binary fission; may have flagella for
motility
• Gram staining tells differences in structure
• Example= E. Coli; rod shaped bacteria
o Mostly harmless- commonly found in intestines
o Beneficially by producing vitamins (Vit. K) and prevent
colonization by pathogens
o Also pathogenic strains—can cause food poisoning like
symptoms
o Popular- can be grown and cultured easily- most widely
studied prokaryotic organism
• Example= H. pylori- famous because of role in causing ulcers
o Relatively small
o Can remain in our stomach without causing any problem-
more than ½ people have it in upper GI tract and over 80%
of individuals infected will never show symptoms
o To avoid stomach acid, they use flagella to embed themselves
in stomach lining mucus
▪ In some people can cause inflammation ulcers can
become cancerous
Archaea- differ from bacteria in cell wall structure; also prokaryotes with no
clear nucleus
• Avidity to extreme environmental conditions
• Can also produce asexually by binary fission (use flagella)
• Not as well studied- harder to culture/ reproduce and some are
anaerobic
• Example- Korarchaeum cryptofilum- found at high temperatures but
found in low abundances
• Example- Methanococcus maripaludis- found in salt marsh
sediments; obligate anaerobes; produces methane
Fungi- mushrooms, molds and yeasts; eukaryotic cells with a true nucleus • Molds= multi-cellular organisms; form filamentous tubes called
hyphae that help absorb material
• Yeasts- unicellular; may produce sexually or asexually
• Obtain nutrients by absorbing organic material from their
environment; can spread and reproduce by releasing spores
• Example- saccharomyces cerevisiae- oval-shaped yeast; harmless
and beneficial
o Only pathogenic in blood stream and immunosuppressed
patients
o Can be grown and cultured easily for lab; most widely studied
eukaryotic microbe; used to produce alcohols from sugars
Protozoa- unicellular aerobic eukaryotes (nucleus); complex organelles and
obtain nourishment by absorption and ingestion
• Divide based on mode of locomotion
o 1. Flagellates produce their own food and their whip-like
structure to propel forward
o 2. Ciliates- tiny hair to beat to produce movement
o 3. Amoeboid- false feet/ pseudopodia used for feeding and
locomotion
o 4. Sporozoans- non-mobile
• essential in marine environment
• Example= amoeba proteus- eukaryote with phospholipids bilayers
membrane similar to other eukaryotic organisms
Algae- diverse group of photosynthetic organisms which aren’t closely
related
• Poorly defined
• Live in water, soil, rocks and produce oxygen
• Believed that they are the origins of green land plants
• Example= cyanobacteria- photosynthetic; nitrogen fixing bacteria
that live mostly in soil and water
o Chloroplasts found in plants have evolved from cyanobacteria
endosymbiosis
o Harmful- concern for drinking water supplies; can clog filters
and aren’t safe for consumption
o Application in biotechnology because so versatile- agriculture,
generation of renewable energy Helminths- multi-cellular animal parasites; visible to the eye; live and feed
on host, receiving nourishment by disrupting hosts’ nutrient absorption
weakness and disease
• May not all be bad; modulation of immune system may prevent
allergies and autoimmune diseases
o co-evolution of parasites with host aren’t as harmful
• Example= A. lumbricoides- round worm; affect 1 billion worldwide;
eggs live in soil
• Once swallowed, they go in small intestines and become adults
• Female worms produce as many as 200,000 eggs per day for a year
Viruses- non-cellular entities consist of nucleic acid core; different from
everything else; microorganisms, yet not living organisms
• Cannot reproduce or feed themselves/ metabolize without a host
• Infest prokaryotic & eukaryotic cells and exploit the cellular
machinery of the host to replicate
• Example= lambda phage- infects E. coli; typical structure of outer
protein capsid enclosing genetic material; can enter into lytic phase
which it kills and lyses cell to produce offspring
o Used as model organism for studying bacteria-phage
interaction and used as molecular genetics tool for cloning
Prions- infectious agent composed entirely of protein; disease due to
abnormal folding of regular proteins
• Folding can be used as a guide to mis-fold other proteins in the
cell reason they can be pathogenic
• Animal prion diseases- example= mad cow
• Human prion disease are often in NS-
o CJD- can lead to death
Microbes are diverse- Bacteria are more diverse! (picture on graph—top
shows bacteria diversity, bottom is Archaea)
• 1/25 Taxonomy 1/18/2017 4:02:00 PM
Taxonomy- the classification, description, identification and naming of living
organisms
• Carlos Linnaeus- proposed Linnaean taxonomy, a system of
categorizing and naming organisms using a standard format
o Divided natural world into 3 kingdoms: Animals, Plants and
Minerals
• Developed into a hierarchy, based on kingdom, class, order, family,
genus and species
Phylogeny (based on evolutionary relations)= evolutionary history of
organisms
• Arranging organisms by how closely related they are phylogenic
tree (tree of life)
• Initially based on visual classifications
• Linnaeus- developed a new way to categorize animals and plants
Haeckel- proposed the four kingdoms Whittaker- added fungi to
the tree of life
Tree of life- based on similarity to humans
Endosymbiotic theory- history of how mitochondria came about
• an organism originally trapped inside an organism based on
similarity between organelles and bacterial cells- proposed by
Margulis
Classification hierarchy
• Life domain kingdom phylum class order family
genus species
Classification by appearance- most intuitive way of characterizing; helpful for
identification
Classification by traits
• Temperature Response
o Psychrophiles- low temperature; below/ around freezing; can
cause contamination of refrigerated food
o Mesophiles- room/ body temperature; the ones we typically
deal with when we deal with pathogens
o Thermophiles- high temperature (in compost)
o Hyperthermophiles- very high temperature
• Based on oxygen dependency
o Aerobes- (strict/ obligate aerobes) ▪ Need/ use oxygen for growth
▪ Subgroup= Microaerophiles- technically aerobes; use
oxygen for growth but are poisoned by high
concentration of oxygen
o Anaerobes- can grow in the absence of oxygen
▪ Subgroup= aerotolerants- don’t use oxygen but not
harmed by it
o Facultative anaerobes- have both capabilities; can survive in
both aerobic and anaerobic environments
• Classification by Metabolic Capabilities
o Source of energy
▪ Chemotrophs- obtain energy by the oxidation of
compounds
Chemoliphotrophs- inorganic compounds
Cehmoorganotrophs- get oxygen from organic
compounds
▪ Phototrophs- obtain energy from light
o Carbon Dependency
▪ Autotrophs- produce complex organic compounds from
simple substances present in its surrounding
▪ Heterotrophs- cannot fix carbon from inorganic sources
but uses organic carbon for growth
o Metabolic dependency
▪ Prototrophs- capable of synthesizing all compounds
needed for growth
▪ Auxotrophs- cannot synthesize a particular compounds
required for growth; need to obtain it from something
else
• Classification by Life-style
o Association
▪ Endosymbiosis: Symbiont lives within the body or cells
of the host; many instances of endosymbiosis are
obligate
▪ Ectosymbiosis- on the surface of the host, including
internal surfaces such as the lining of the GI tract
• Cell wall staining o Gram-positive or Gram-negative- depending on cell wall
structure
• Growth on specific resources- growth plates are specific to certain
types of organisms
o Can potentially differentiate closely related organisms
Taxonomy in the light of genetics (by genotype)
• Has become the most dominant way of classifying microbes
• American biologist Carl Woese discovered what appeared to be a
living record in the evolution of organisms
• Ribosomal RNA gene: conserved sites (C) and hypervariables sites
(V)
o rRNA gene is one of only a few genes present in all cells
o conserved sites are similar among many organisms
o Hypervariable sites diversity as organisms evolve
o Conserved areas allow universal primers for amplifying this
portion of the genome
o Sequencing of the hypervariable areas allows differentiation
of species
• Ribosomal DNA sequencing for classification
o Target rDNA depends on the branch of organism
▪ Prokaryotes- use 16S subunit
▪ Eukaryotes- use 18S subunit
o Database of rDNA sequences allows identification of microbes
o **Isolation of microbe DNA extraction Amplify RNA gene
using universal primers sequence a portion (16S or 18S) of
the rRNA by PCR Compare the sequence of the Database
• Tools for classifying microbes (mostly bacteria)
o RDP
o NCBI BLAST
• Advantage of Classifying: Access to previous work
o You know what you’re dealing with and certain characteristics
about it
o Staphylococcus Epidermis- the things we can tell about it
from databases; we don’t need to start from scratch even
when we have just isolated the organism
The challenge of defining microbial species • Historically, a species is defined as a group of individuals that
actually or potentially interbreed in nature. In this sense, a species
is the biggest gene pool possible under natural conditions
o BUT
▪ Many organisms form hybrids
▪ Many microbes reproduce mainly asexually
o …The definition of a species isn’t defined for a microbe
▪ we need to change our definition of a species…
• Species identification based on rRNA gene similarity- using 97%
16S similarity as a measure to distinguish species in general works
well
o Limitations still…
▪ Identification limited to those already included in the
database
▪ Early categories based on 527 bp portion of 16S subunit
in bacteria, but later transitioned to full 16S
▪ 97% similarity may not be the best choice
(subjectively; all primates would be grouped together)
▪ based on whole genome sequencing, there are
exceptions and inconsistencies
Pan-genome versus Core-genome (definitions aren’t very set in stone)
• Core-genome= contains gene families shared by all the organisms
within a group***
• Pan-genome contains all the genes families including shared and
strain specific genes
• Picture: showing 4 different strains of E. Coli… out of 4,000 genes
there are about 3,071 that are shared between all strands… AND
there are genes only present in one strand
• Current species definition is mainly based on core genome*** History of Microbial Impact 1/18/2017 4:02:00 PM
Humans have been living with and using microorganisms for much longer
than they have been able to see them
• Example: People were making fermented beverages
• Example: Cheese, yogurt and bread making
• What do we gain by looking into the details of microbial
contribution?
o Safety of the food product
The Iceman- prehistoric humans had a limited grasp on the causes of
disease; still they attempted to treat illnesses and infections
• Otzi the Iceman- a 53000 year old mummy found frozen in ice of
Otzel Alps on Australian-Italian border
• Infected with the eggs of the parasite human whipworm, which may
have caused abdominal pain and anemia
• Also evidence of bacterium that causes Lyme disease
• Trying to treat his infections with the woody fruit of a fungus with
laxative and antibiotic properties, which was discovered tied to his
belongings
• Also had tattoos
Early Notions of Disease and Containment
• Several ancient civilizations appear to have had some
misunderstanding that disease could be transmitted by things they
couldn’t see
o Quarantining leprosy patients
o Miasma theory= transmission of disease via air of rotten
material
o Sewage in Rome- built a channel to carry sewage out from
the city into the river to avoid accumulation of waste, and
prevent disease
What Causes Disease?
• Suspicion of possibly invisible causes for disease
o Hippocrates: Diseases have natural causes from within
patients on their environments, rather than supernatural
causes
o Thucydides- survived the plague in Athens and recorded the
observation that survivors didn’t get re-infected even when
exposed ▪ immunity
o Varro—“certain minute creatures” in air and water were
responsible for contracting disease
Bubonic Plague AKA Black Plague
• AKA black death
• Caused by bacteria Yersenia Pestis
• Killed ¼-1/2 of people in eastern Mediterranean years
• Killed 50-100 million people in the 1300’s—most popular event
1918 Flu pandemic
• another pandemic caused by viruses; unusually deadly; 500 million
people infected around the world and killed 5% of world population
(50-100 million people)
• Contributed to 12 year drop of human’s life expectancy
• Most influenza outbreaks target juvenile, elderly, or already
weakened patients; pandemic killed previously healthy young adults
Great Irish Famine
• The famine in Ireland cause potato crop to fail significant drop in
food resources famine 100 million died and 100 million had to
migrate
• Cause is potato blight, which destroyed potato crops throughout
Europe
• Possibly caused by microbial eukaryote, phytophthora infestans
A look back to the past
• Infections and food spoilage have tainted our impression of
microbes the impression that they can be harmful
• BUT things have been getting better…
o Life expectancy (mostly caused by infections) has increased!
o Infections aren’t killing as many people as they used to
• Increase in life expectancy due to vaccination and antibiotics
Vaccination- active immunity to diseases can be acquired by natural or
intentional exposure to disease through vaccination
• Use weakened/ inactive forms of microbes
o body will produce antibodies to fight them
• more modern techniques involve cloning genes to mass produce the
antigens that are producing the reactions
Antibiotics, the good • Produces by organisms to fight
• Treating a broad range of infections increasing life span
• Lowering the bacterial load prevent infection (during surgery
• Higher yield of livestock in animals, it promotes growth and
prevents disease
Antibiotics, the bad
• AAD- diarrhea
• Colon inflammation (colitis)
o Malabsorption characterized by a celiac- like syndrome
o impaired absorption of medications
o ^ both due to inflammation
• Impaired metabolism and absorption of vitamins disrupted
• Rise of antibiotic resistant organisms
• Susceptibility to other infections
Rise of antibiotic Resistant Strains
• Evolution drives microbes toward developing resistance
How can Antibiotic Resistance Spread?
• We eat it when we use it in livestock resistance
o When its present in our food (meat products)
• Patients use antibiotics to treat things that aren’t a bacterial
infection (such as a cold)/ all infections being treated the same way
o Resistance rises when people who don’t need antibiotics are
treated with them
• ** if we develop high enough resistance, its not effective anymore
The Threat Is Real
• Nevada dies of bacteria that was resistant to several microbiotics
What Can we DO?!
• Responsible administration of antibiotics
o Avoid long-term, unnecessary, sub-lethal doses
• Discovery of new antibiotics
o Not enough corporate support
o Research on previously uncultivated organisms
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