Download Microbiology: Bacterial Structure and Physiology I pg. 1 Jenny

Microbiology Transcriber: Jenny Seibert
09/10/2008
49:16
Bacterial Structure and Physiology I
Slide 2:
I am going to give you an intro to bacteriology. For those who have taken a course like this
before, this may be refresher. For those who haven't had this before, this will be a broad overview and
we'll get into the details later. Some things I tell you today will be very brief compared to what you'll get
later on, but this is just to set the stage for the rest of the course. Listed here are some relevant
chapters out of the book. I don't talk about everything in the chapters. They are merely for reference or
a backup for what I talk about in class. I will talk about important points that I want you to know for the
exam. For detail and clarity, you can refer to your book.
Slide 4:
So I'm going to start off with bacterial nomenclature, just to give you an idea of how bacteria are
named and how we refer to them. Bacteria is part of a kingdom called Eubacteria. That kingdom has
been further subdivided into many different divisions, classes, and so forth. As you go down this line
you get more specific, referring to a specific bacterium. When we get to order and family, you will often
hear bacteria referred that way, but it's still a fairly general grouping, such that the bacteria family
Spirochaetaceae will be referred to as “Spirochetes.” If we go down further, we will talk about the
specific bacteria and this case I have listed the genus Borrelia and the species burgdorferi. This is how
most bacteria are referred to, as a genus and a species. You can also subdivide the species. Once you go
beyond this, you begin to talk about really specific strains. Some of those are subdivided as well.
When you write out the name of the bacteria, you write it italicized. If you can't italicize, then
you underline. When you're writing a formal report, you would initially refer to this bacteria as Borrelia
burgdoferi and later in the paper it should be mentioned as B. burgdoferi. This bacterium causes Lyme
disease.
Slide 3:
So bacteria you study are the Eubacteria and if we put them in the context of all the life on
Earth we know, then there are three kingdoms or domains in which life is divided. Eukaryotes
encompass the plants, animals, protists, and the fungi. The Eubacteria are the ones we will be studying
most. The Archaebacteria are a group of bacteria that are evolutionarily distinct from the Eubacteria.
Slide 5:
So if we divide these three groups a little bit further, what distinguishes them is whether or not
their DNA is encompassed by a nuclear membrane. Eukaryotes have a true nuclear membrane. In
Prokaryotic organisms there is no membrane. The Eukaryotic bacteria are considered true bacteria.
Microbiology: Bacterial Structure and Physiology I
Jenny Seibert
pg. 2
The Archaebacteria are primitive and separated evolutionarily. They can grow in extreme
situations, in high salt, extreme pH, high heat. They can produce methane. What distinguishes them
from Eubacteria is their cell wall. Archaebacteria cell wall lacks peptidoglycan (which is a major
component of Eubacteria cell wall). Their membranes have ether rather than ester-linked lipids. Their
ribosome components and metabolism are very different from Eubacteria. In some aspects, they share
features with Eukaryotic organisms. They have introns and histones, things that normally do not occur in
the prokaryotic world.
Slide 6:
What are the distinguishing features of Prokaryotic and Eukaryotic cells? From now on, when we
talk about bacterial cells, we are talking strictly about Eubacteria, not Archaebacteria. (So for distinctive
features, she mostly reads off the chart, but these are the things that are not on the chart:
Eukaryotes have linear chromosomes and Prokayotes usually don't. The exception is Borrelia, which is a
Eubacteria that has a linear shromosome.
In Eubacteria, because the membrane is the only interface with the outside world, it has many functions.
Mycoplasma is a type of bacteria that causes walking pneumonia.
Eukaryotes can have cell walls, just not with peptidoglycan.
Mycoplasma take sterols from the host, i.e. they don't synthesize them.
Ribosomes different in terms of their structure. They are of a different size as quantified by their
Svedberg coefficients (how it sediments in a particular centrifugal gradient). Each ribosome has two
main subunits, each having their own Svedberg coefficient listed in parentheses by the first number.
Not only are these features important for distinguishing these organisms, but they are
important differences we capitalize on to use antibacterials against prokaryotes, so that they kill the
prokaryotic organism, but they don't harm the eukaryotic organism. So many antibacterials act against
ribosomes or cell wall components, in particular the peptidoglycan and other things within the bacterial
cell that are unique. So because in a eukaryotic cell there is no peptidoglycan and because the
ribosomes are different, we are able to target drugs against those inherent properties without harming
the host that we are trying to treat.)
Slide 7:
Bacteria are single-celled organisms that reproduce by simple division, i.e. binary fission in which they
just divide in half. If you'll look a few slides over, you'll see this in process.
Slide 10:
Microbiology: Bacterial Structure and Physiology I
Jenny Seibert
pg. 3
This is a Streptococcus chain of bacteria that's growing and it's been labeled with an antibody to
the surface that had a fluorescent label. If you wash out the remaining label and let the bacteria
continue to grow in the absence of the label, what occurs is cell division down the middle of the
bacteria. The bacteria replicate over time. The new cell wall or cell membrane is not labeled and you
can see down the chain parts where there is label and parts where there is no label. This demonstrates
that cell division is occurring in the middle because that is where new cell wall, cell membrane is being
added.
Slide 7:
The size of bacteria can vary greatly from as little as 0.2 microns in size to 10 microns in size
(some Bacilli) with an average of one micron. When you are doing filter sterilization procedures in the
laboratory, filtering out bacteria in solution, you would use a pore size on your filter of about 0.4
microns. The reason for that is most bacteria are bigger than 0.4 microns, so they'll be trapped on the
filter and the solution that goes through will then be sterilized from those bacteria.
Some bacteria are smaller than that. When you do cell cultures, things like Mycobacterium
(which are smaller than 0.4 microns) can contaminate your culture. In this case a filter with a pore size
smaller than 0.4 microns is necessary.
Most bacteria are free-living which means they do not need a eukaryotic cell to survive. A few.
However, are obligate parasites such as rickettsiae and chlamydiae. These cells need to reside inside a
eukaryotic cell to survive.
Slide 8:
Bacteria come in various shapes. The first is streptococcus which is just a rounded cell. Then you
have rods which are things like bacilli. Then you have spirochetes which are spiral shaped. And Cholera
which are vibrio shaped.
Slide 9:
The bacteria can be associated in different ways. Some grow in groups of four, or bigger clusters,
like grapes. Some can grow in long chains. Streptococcus likes to grow this way.
Slide 11:
Let's talk about the composition of the bacterial cell. (She reads the slide word for word...If you
want it again:
50% Protein, 20%nuclei acids (10X more RNA than DNA), 10% polysaccharides, 10% lipids)
Microbiology: Bacterial Structure and Physiology I
Jenny Seibert
pg. 4
Slide 12:
Bacteria have single, circular, double-stranded DNA with the exception of borrelia. They are
haploid meaning they have only one chromosome. Under certain conditions, there may be more than
one copy of that chromosome. It is the same chromosome with multiple copies as compared to diploid
organisms that have two distinct chromosomes that have different alleles. The reason why there may
be more than one copy is that if the cell is rapidly dividing the cell has to ensure that each daughter cell
will get a copy of the DNA.
The size of the chromosome varies much like the size of the cell does. It can range anywhere
from 600 to 5000 kb in size. The smaller the genome, the more dependent it is on the host. For example
mycoplasmas have a very small chromosome whereas things like E. coli have a larger chromosome.
Mycoplasmas need their host. E.coli and bacillus are ok in many different environments. They have
more genes to synthesize what they need.
One kb on average is equal to one gene. The average genome is 4500 kb so this turns out to be
about 1 mm in length. How does this fit into a 1 micron sized cell? It is done by a process of supercoiling.
It's like the twisting of a rubber band.
Slide 13:
Bacteria DNA is not just free-floating in the cytoplasm. It's actually contained within a nucleoid.
A nucleoid is not membrane bound. The chromosome does not have histones, but it does have proteins
involved in determining the shape and structure of the chromosome, maintaining its composition. They
also have polyamines that neutralize the negative charges on phosphates. This is very important in
neutralizing viral DNA. The DNA chromosome can associate with the cell membrane, but it is not
membrane bound.
Slide 14:
Bacteria can have extrachromosomal DNA. One form can be in plasmids. These are DNA that
replicate inside the cytoplasm that are independent of the chromosome. They are circular with the
exception of borrelia. They can range in size from a few to several hundred kb. They can be conjugative
or nonconjugative and can code a number of different functions. Conjugative plasmids are ones that can
confer antibiotic resistance, metabolic differences or virulence from one bacteria to another. Particularly
with gram-negative bacteria, antibiotic resistance can be conferred through conjugation via sex pili.
Transfer of DNA can occur through other mechanisms with other types of bacteria.
Another way extrachromosomal DNA can occur is through bacteriophages (phages). Phages are
viruses that infect bacterial cells. There are specific viruses that infect specific bacterium. They can
replicate in the cytoplasm or integrate into the chromosome. Many virulence factors associated with the
bacteria are actually encoded by the virus DNA carried by the bacterial DNA.
Slide 15:
Microbiology: Bacterial Structure and Physiology I
Jenny Seibert
pg. 5
These are examples of bacteriophage. In the head is where the DNA material is kept. The spikes
are actually what land on the bacterial cell and the DNA is injected into the bacterial cell.
Slide 16:
Bacteria are basically surrounded by a typical lipid bilayer. It forms a permeability barrier for the
cell. It involves transport and factors that are important for photosynthesis in those bacteria that are
capable of photosynthesis.
It also is affected by antibacterials. Cytoplasmic membranes are affected by detergents. When
you wash your hands, the part of the bacteria you are affecting is the cytoplasmic membrane. They put
extra antibacterials in soap which is not necessary because soap, inherently, is antibacterial. Polymyxins
and ionophores both damage the cytoplasmic membrane or bacteria in different ways. Polymyxins
damage PE-containing membranes and ionophores disrupt the membrane potential.
Slide 17:
The cell wall is very important to the bacteria. It gives and maintains their characteristic shapes.
If you removed it, all bacteria would look the same. It's a barrier as well and serves as osmotic
resistance. The cell wall is composed of highly cross-linked peptidogylcan. This linking can be affected
by antibacterials. Things like B-lactam and lysozyme affect the cross-linking of peptidoglycan. It also
serves the basis of the gram stain. Characterizing bacteria based on evolutionarily similar characteristics
started with gram stain.
Slide 18:
Peptidoglycan itself is the basis for the cell wall. It has a backbone of two sugars N-acetyl
glucosamine (GlcNAc) and N-acetyl muramic acid (MurNAc). These two sugars are cross-linked by
peptide bridges at MurNAc. The peptide side chain of one MurNAc is linked to another MurNAc.
Slide 19:
The cross-linking of many MurNAc allows the cell wall to remain on the surface (not sure if this is
exactly what she said; audio is muffled).
Slide 20:
Looking a little closer at the peptidoglycan. We have the two sugars here GlcNAc and MurNAc
and the multiple peptide side chains linked to the MurNAc. And the side chains are cross-linked. This
particular peptidoglycan structure is from a gram-positive organism. Peptidoglycans from gram+ and
gram- are very similar but they are not identical. Gram+ has a cross-bridge composed of many glycines
while Gram- has a direct cross-link, nothing to bridge the peptide side chains. (Here is a better picture
from online and the link to the website, which has good info on it, is:
http://web.virginia.edu/Heidi/chapter9/chp9.htm).
Microbiology: Bacterial Structure and Physiology I
Jenny Seibert
pg. 6
Figure 9.22  (a) The cross-link
in Gram-positive cell walls is a
pentaglycine bridge. (b) In
Gram-negative cell walls, the
linkage between the tetrapeptides
of adjacent carbohydrate chains
in peptidoglycan involves a
direct amide bond between the
lysine side chain of one
tetrapeptide and d -alanine of the
other.
Peptidoglycans can also differ in the amino acids they have in their side chains.
The sugars are linked together via transglycosylated links. Transpeptidases link the peptide side
chains together. Transpeptidases are also called penicillin binding proteins. We will come back to why
they are called this. Hydrolases can cleave the backbone. Lysosyme, a type of hydrolase found in
secretions, can cleave this backbone and it is used as a first line of defense against bacteria. Amidases
can cleave the peptide structure. Hydrolases, such as lysosyme, recognize bacteria by the acetyl group
off of one of the sugars in the backbone. One thing bacteria do to avoid being recognized by the
lysosyme, is hide that acetyl group. Some bacteria have enzymes that cleave off the acetyl group after
the peptidoglycan has been made. This makes lysosyme ineffective.
Amidase cleaves peptide bonds. It is mostly created by the cell itself for its own needs.
Slide 21:
B-lactams are antibacterials that simulate transpeptidase substrates. They work by blocking the
cross-link of the growing chain. Normally the transpeptidase recognizes the specific D-ala to D-ala
structure. The B-lactam ring so closely resembles D-ala that transpeptidase recognizes it, binds it and
prevents is from recognizing another D-ala to complete the link. This is only effective for cells that are
actively growing. The B-lactam cannot break an already formed peptidoglycan chain. In contrast to that,
lysosyme can cleave a quiescent bacteria.
Slide 22:
Microbiology: Bacterial Structure and Physiology I
Jenny Seibert
pg. 7
Gram stain was one of the first ways people could appreciate a difference in the types of
bacteria. First you put your bacteria on a microscope slide, then you pass it over a flame. This kills the
bacteria and causes them to stick to the slide, hence the name for this procedure heat-fixing. This allows
you to add dye and wash it off without washing the bacteria off. The first dye you add is Gram's crystal
violet, a purple stain. Then you add potassium-iodide. Then you add ethanol which pulls the water out of
the cell wall. Then you wash the cell. What you are forming is a crystal iodide complex. When you shrink
the cell wall with ethanol, this CV-I becomes trapped in the thick cell wall. When you have a thick cell
wall, you end up trapping a lot of these complexes and so the cell is purple. If you have a bacteria that
has a thin cell wall, you wouldn't have any dye trapped and the bacteria will have no color. But if you
come back and add a second dye, safranin, will turn the bacteria red.
So what distinguishes gram+ from gram- is the thickness of their cell walls. And there are
similarities between the bacteria in each classification. Gram+ bacteria resemble each other while all
gram- bacteria resemble each other.
Slide 23:
There are exceptions to the gram-staining rule. Mycoplasmas have no cell wall, no
peptidoglycan, therefore nothing to trap the stain. Mycobacteria, the bacteria that causes TB, does have
a cell wall but around it is a lot of lipids and these lipids block the ability to absorb the stain. It can be
stained with an acid fast stain. Based on genetic analysis, both of these organisms are related to gram+
organisms.
Slide 24:
If we look at gram+ organisms, they have a thick cell wall. The two things to remember are they
have lipoteichoic acid in the membrane and teichoic acid further out in the cell wall.
Slide 25:
Wall teichoic acids are in the cell wall of gram+ organisms. They have repeating units of
phophosiester-linked glycerol or ribitol backbone plus side chains. These are covalently linked to
peptidoglycan. The cell wall itself is not just peptidoglycan. The cell wall contains peptidoglycan and
covalently linked wall teichoic acids. There is an envelope that is the outside part of the cell that gram+
have that also make lipoteichoic acid. Structurally is may be the same or it may be different from the
wall teichoic acid present on that bacterium. The main difference here is that the lipoteichoic acid is
anchored into the cell membrane while the WTA is covalently linked to the PG.
Slide 26:
Both LTA and WTA are important for maintaining the ion binding and charge of the membrane
as well as the membrane integrity. Because LTA is anchored in the membrane, it might have a bigger
role in structure. These acids are also important allowing bacteria to adhere to their eukaryotic host.
And they also have the function of anchoring proteins. Cell walls have parts that can induce
inflammation in the host. This occurs in meningitis when the brain swells due to bacterial infection.
Microbiology: Bacterial Structure and Physiology I
Jenny Seibert
pg. 8
Microbiology: Bacterial Structure and Physiology I
Jenny Seibert
pg. 9