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Dr. Wasan Sami Shukur
Genetics
Genetics is the study of inheritance. Except for RNA viruses all
hereditary characteristics are encoded in DNA. The unit of heredity is the
gene, a segment of DNA that carries in its nucleotide sequence
information for a specific biochemical or physiologic property. The
chromosomal DNA plays a pivotal role in the maintenance of genetic
stability with in the organism and the species by providing genetic
information to the progeny possessed by the parent.
Bacterial genetic material
A] Chromosomes
Bacteria have one unique chromosome (haploid) that can encode up to
4000 separate genes necessary for bacterial maintenance and propagation.
The chromosome of bacteria are circular, naked, double strand DNA
molecules, the DNA is usually found attached to the cell membrane at
some point or points. Although bacteria do not possess a nucleus, the
DNA is localized in a distinct area with in the cell called the nucleoid
region. There is no membrane around the nucleoid region and lies free in
the cytoplasm of bacteria.
DNA molecule is composed of two strands of complementary
polynucleotide chains wound together in the form of a double helix.
Each strand has a backbone of deoxyribose & phosphate groups. One of
the four nitrogenous bases, the purines ( adenine "A" & guanine "G") &
the pyrimidines (thymine "T" & cytosine "C") is attached to each
deoxyribose. The two strands are held together by hydrogen bonding
between the bases on the opposite strands in such a specific manner that
hydrogen bonds can only be formed between adenine & thymine (A-T) &
between guanine & cytosine (G-C). Adenine & thymine form one
complementary base pair (bp), & guanine & cytosine form another bp.
Thus, when the arrangement of bases along one strand is G-C-C…. The
arrangement of bases in the other strand will be C-G-G…..
In a molecule of DNA there are as many units of adenine as thymine, &
of guanine as cytosine. The ratio of each pair of bases (A+T) to (G+C) is
constant for each species, & varies considerably with different species of
bacteria. A segment of chromosomal DNA that specifies the production
of a particular polypeptide chain is called a gene and the total
complement of genes in a cell is known as the cells genome. Genetic code
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or information is contained in the sequence of purine & pyrimidine bases
of the nucleotides. One codon (unit of code) consist of three bases,i.e. the
code is triplet. Each triplet codon codes for a single aminoacid (aa) e.g.,
ACG codes for threonine. There are a total number of 64 codons, 61 of
which code for 20 essential aminoacids and three codons (UAA, UAG,
UGA) do not code for any aa & the latter are called "nonsense codons". It
should be noted that there are more than one codon for the same aa, such
as, ACU, ACC & ACA all code for threonine. The function of nonsense
codons is to terminate the elongation of polypeptide chains, hence known
as stop codons.
Bacterial genome may include plasmid & prophage genes as well as the
bacterial chromosome.
B] Plasmids
Plasmids are extrachromosomal DNA molecules & consist of double
stranded, circular DNA molecules that are capable of replicating
independently of the bacterial chromosome. Although plasmids are
usually extrachromosomal, they can be integrated into the bacterial
chromosome.
Plasmids are not essential for normal function of host bacterium but their
presence in bacteria confers properties of drug resistance, toxigenecity,
conjugative plasmid & others. Some plasmids are self transmissible to
other bacteria of the same & also of different species. Transfer occurs
usually by conjugation plasmids occur in both G-ve & G+ve bacteria , &
several different types of plasmids can exist in one cell.
Plasmid types
Plasmids are typed based on their function:
1-R-plasmids: R-plasmids (R-factor) consist of two components one
component is resistance transfer factor (RTF) which carries the genes that
govern the process of inter-cellular transfer & the other component is
called resistant determinant (R-determinant) carrying the resistant genes
for each of the several drugs sometimes the RTF may dissociate from the
R-determinant & may exist separately, then the drug resistance is not
transferable. Determinants other than those for drug resistance may be
attached to RTF, such as, genes responsible for enterotoxin & haemolysin
production in some enteropathogenic E.coli & are able to transfer to other
bacteria.
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2- Col factors: The colicinogenic (col) factors are found in several
species of coliforms which produce extracellular colicins. These
bacterial substances are lethal toxins for other strains of the same or
closely related species of bacteria. Since similar antibacterial substances
are also produced by bacteria other than coliforms, this group of
substances has been named as bacteriocins. Colicins are produced by
E.coli, pyocin by pseudomonas aeroginosa, marscesins by serretia
marcescens & diphthericin by C.diphtheriae. Composition of bacteriocin
varies from proteins to LPS complex. Bacterial strains producing
bacteriocin are resistant to their own bacteriocin which helps interspecies
typing of organisms.
3- F or Fertility factor : The fertility plasmid, F-factor, is a transfer
factor, that contains the basic genetic information necessary for extrachromosomal existence, self transfer & for the synthesis of sex pilus, but
is devoid of other identifiable genetic marker, such as, drug resistance
genes.
Cell carrying F factor (F+cell) possess sex pilus & extrude an
extracellular protein that attaches donor cell (F+) to recipient cell lacking
fertility factor (F-). The sex pilus of male bacteria (F+) is probably
responsible for forming conjugation-tube with female (F-) bacteria,
through which genes are transferred from F+ to F- cells. In a small
proportion of F+ cells (1 per 105 cells at each generation), F becomes
integrated into the loci of bacterial chromosome. When the integration is
relatively stable, the cell in which it has occurred, gives rise to a clone of
cells which are known as Hfr strains (high frequency recombination
strains). The entire chromosome behaves like an enormous F-plasmid &
hence certain chromosomal genes can be transferred into F-cells with
high frequency. The rate of chromosomal transfer from Hfr cell
conjugates with F- cell, the latter receives chromosome from the donor
but rarely becomes F+.
The integration of F factor with chromosomal DNA of bacteria is a
reversible process & detachment of F takes place 1 per 10 5 cells at each
generation. When F factor reverts from integrated to the free state, some
chromosomal genes may be carried by F factor from near to its site of
attachment. Such type of F factor that incorporates some chromosomal
genes is called F prime (F`) factor. When such F` cell mates with a
recipient, the F factor along with incorporated host genes are transferred.
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Molecular classes of plasmids
1-Small (non conjugative) plasmids, common in G+ve cocci as well
as in some G-ve orgs.(e.g. H.influenzae, N.gonorroeae) are usually
small (1-10 million Daltons), they are frequently present in many (1060) copies per cell.
Small plasmids can also be transferred from cell to cell when the same
bacterium carries both conjugative & non conjugative plasmids. Once
conjugation is established, the donor can transfer non conjugative
plasmids.
2- Large (conjugative plasmids) are transferred from bacterium to
bacterium ( usually members of the same species or of very closely
related species) through conjugation. These plasmids are common in
G-ve bacilli and are relatively large (25-150 million Daltons). Large
plasmids are usually present at 1-3 copies per cell.
Cell properties encoded by plasmids
1)
2)
3)
4)
5)
6)
Drug resistance
Virulence-toxigenicity & invasiveness
Antimicrobial agents & bacteriocins production
Genes determining metabolic pathways
Specialized recombination system
Conjugation functions ( transfer genes)
C] Transposons
Transposons are pieces of DNA that move readily from one site to
another; either within or between the DNAs of bacteria, plasmids, &
bacteriophages. They can code for drug resistance enzymes, toxins or a
variety of metabolic enzymes, & they can either cause mutations in the
gene into which they insert or alter the expression of nearby genes.
The transfer of a transposons to a plasmid & the subsequent transfer of
the plasmid to another bacterium by conjugation contributes significantly
to the spread of antibiotic resistance.
Transposons typically have four identifiable domains. One each end is a
short DNA sequence of inverted repeats, which are involved in the
integration of the transposon into the recipient DNA. The second domain
is the gene for the transposase, which is the enzyme that mediates the
excision & integration processes . The third region is the gene for the
repressor that regulates the synthesis both of the transposase & of the
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gene product of the fourth domain, which, in many cases, is an enzyme
mediating antibiotic resistance. In contrast to plasmids or bacterial,
viruses, transposons are not capable of independent replication; they
replicate as part of recipient DNA. More than one transposon can be
located in the DNA; ex, aplasmid can contain several transposons carring
drug resistance genes. Insertion sequences are a type of transposon that
has fewer bases (800-1500 bp). They can cause mutations at their site of
integration & can be found in multiple copies at the ends of larger
transposons units.
Transposons genes
Transfer of DNA between bacterial cells
Transfer of genetic information from one cell to another can occur by
three methods:
1-Conjugation is the mating of two bacterial cells during which DNA is
transferred from the donor to the recipient cell. The mating process is
controlled by an F(fertility) plasmids, which carries the genes for the
proteins required for conjugation.
2-Transduction is the transfer of cell DNA by means of a bacterial virus
(bacteriophage, phage) During the growth of the virus within the cell, a
piece of bacterial DNA is incorporated into the virus particle and is
carried into the recipient cell at the time of infection. Within the recipient
cell, the phage DNA can integrate into the cell DNA and the cell can
acquire a new trait, a process called lysogenic conversion. This process
can change a nonpathogenic org into a pathogenic one. Diphtheria toxin,
botulinum toxin & cholera toxin are encoded by bacteriophages & can be
transferred by transduction.
There are two types of transduction, generalized & specialized. The
generalized type occurs when the virus carries a segment from any part of
the bacterial chromosome. This occurs because the cell DNA is
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fragmented after phage infection & pieces of cell DNA the same size as
the viral DNA are incorporated into the virus particle at a frequency of
about 1 in every 1000 virus particles. The specialized type occurs when
the bacterial virus DNA that has integrated into the cell DNA is excised
& carries with it an adjacent part of the cell DNA. Since most lysogenic
phages integrate at specific sites in the bacterial DNA, the adjacent
cellular genes that are transduced are usually specific to that virus.
3) Transformation is the transfer of DNA itself from one cell to another.
This occurs by either of the two following methods. In nature, dying
bacteria may release their DNA, which may be taken up by recipient
cells. There is little evidence that this natural process plays a significant
role in disease. In the laboratory, an investigator may extract DNA from
one type of bacteria & introduce it into genetically different bacteria.
When purified DNA is injected into the nucleus of a eukaryotic cell, the
process is called transfection. Transfection is frequently used in genetic
engineering procedures.
The experimental use of transformation has revealed important
information about DNA. In 1944, it was shown that DNA extracted from
encapsulated smooth pneumococci could transform non encapsulated
rough pneumococci into encapsulated smooth orgs. This demonstration
that the transforming principle was DNA marked the first evidence that
DNA was the genetic material.
Comparison of conjugation, transduction, and transformation.
Transfer
procedure
Conjugation
Process
Type of cells Nature of DNA
involved
transferred
DNA transferred Prokaryotic
Chromosomal or
from
one
plasmid
bacterium
to
another
DNA transferred Prokaryotic
Any gene in
Transduction
by a virus from
generalized
one
cell
to
transduction
another
Transformation Purified DNA Prokaryotic or Any DNA
taken up by a eukaryotic
cell
(eg, human)
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Mutations
Mutation: any detectable & heritable change in the genetic material not
caused by genetic recombination
Origins
1-Spontaneous mutations: result from replication errors, genetic
mispairing, or DNA changes that cause replication errors & mispairing.
Spontaneous mutations may cause genetic variation that is advantageous
or disadvantageous.
2-Experimentally induce mutations
* Chemical mutagenesis : Some chemicals significantly increase the
mutation rate to rates as high as one mutation per 103-104 cells, ex: nitrous
acid .
*Radiation mutagenesis: is caused by exposure to radiation,ex.,
ultraviolet light, ionizing radiation, X-ray…..etc.
Types of mutations
There are two types of mutations:
a-Point M: also called microlesion, is in general reversible & is of two
classes
a.a-Base pair substitution: involve the replacement of a nucleotide in
the coding sequence & can be subdivided into:
1. Transition: This may happen by replacement of one pyrimidine by
another pyrimidine, & one purine by another purine,i.e. AT bp
replaced by GC or TA by CG. Transition is the most frequent type of
mutation.
2.Transversion: In a bp substitution, when a pyrimidine is replaced
by purine &vice versa, it is called transversion, e.g. GC changes to
CG. Transversions is less frequently observed.
a.b- Frame shift mutations: Sometimes, during replication, one or a
few adjacent bps have been inserted to or deleted from the DNA.
b- Multisite mutations
It is also known as macrolesions where there are alterations of DNA
involving large numbers of bps. Change of genetic material in
macrolesions are of 4 types such as, loss (deletion), again (addition),
duplications or inversions.
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