BOD- Salmonella enterica

2000 Serotypes, 50 in
the US identified i.e..
typhimurium
Causes Salmonellosis
Cells divide inside
vesicles of epithelial
of intestine
Low mortality but load
and age dependant
Microbial Evolution: From rDNA
Gene Sequences to Domains.
Which organism (B,C,D) is most
closely related to A?

A B

D
C
Evolutionary Relationships; how to
determine?
Obviously cannot rely on the shapes of
microorganisms.
Physiology-Look at the Biochemical profile
of the organism. Useful, but is it the whole
story?
Classification as it stood in the late
1970s
Kingdom, Phylum, Class, Order, Family,
Genus, Species
Late 1800s Ernst Haeckel proposed the
Kingdom Monera
Monera included the Eubacteria (true
bacteria), Cyanobacteria (blue-green
algae), and Archeabacteria (ancient
bacteria)
 Classically Determined
Taxonomy of Prokaryotes
Cellular Morphology
Physiological
Characteristics
Biochemical
Properties
Ecological
Characteristics
Genetic
Characteristics
Identification of Microorganisms
CoT Curves
A way to compare the
%GC
DNA absorbs @260nm,
absorption doubles when
DNA melted
Watch results as DNA re-
anneals
Generally more useful to
separate organisms
 No definitive book for
phylogeny
Originally volumes of
Bergys Manual
attempted classification
However it was clear
that Classical Methods
were insufficient for
classification
Moving into the age of Molecular
Research
What else is unique about
each organism, but useful
for comparison?

Their Genes!

Using modern DNA

technologies, DNA
sequencing (Sanger
Method), DNA probes
 Criteria for a target gene
Universally in organisms
Target must remain functionally
homologous
Alignment properties of gene
Target gene mutation rate representative
of phylogeny being compared
 Carl Woese
Chose to compare
organisms using newly
available Molecular
sequence techniques.
The closer two DNA
sequences are from a
given gene the closer
the species are.
Specifically sequenced
16s rRNA
 All cells contain ribosomes
In Bacteria ribosomes are composed of 2
subunits, a large subunit (50S) and a
small subunit (30S) S=Sverberg Unit
Each subunit is composed of several
ribosomal proteins and rRNAs.
The 50S contains a single 23S rRNA and
a 5S rRNA
The 30S subunit contains a single 16S
rRNA
What is 16s rRNA?
Why choose 16S rRNA genes?
16S rRNA genes are ubiquitous in all
microorganisms
Size-they are relatively small
Direct comparisons between sequences
could be used as an Evolutionary
chronometer.
Evolutionary distance in phylogenetic trees
can thus be determined for different species
 Technology
Design two DNA
probes
complementary to
RNA sequence
Using PCR amplify
sequence
DNA product can be
sequenced using
Sanger technique
16s rRNA
Parsimony
 What did Woese find?
Signature 16s rRNA sequences
He discovers discrete sequences of RNA.
Archaeabacteria are as different from
Bacteria as Bacteria are from Eukaryotes
Created the Group Archaea
Woeses analysis led to the proposition
that there are three domains of life.
Woeses Contribution; the Domains
FISH Probes
Fluorescent in-situ
hybridization
Develop labeled probes
for different domains
Can be used for clinical
or environmental tracking
of organisms.
Do not need whole
organisms either; PCR
amplification sufficient
Other Evidence of Domains
Cell wall analysis
Peptidoglycan
Pseudopeptidoglycan
Cellulose/Chiton
Membrane Lipids
Ester vs.. Ether
 Translation
Formyl-methionine vs. methionine
Order of Initiation (IF) factors
Antibiotic sensitivity
Histones, present/absent
Polycistronic vs. Monocistronic
Contain plasmids
 Transcription
TATA box vs.. -10 and -35 region
RNA Polymerases
RNA Product Capped?
Introns?