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•Bacterial Genetics

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• Definitions• Genetics: is the science of heredity and

variation.• Heredity: implies the transmission or

inheritance of constant similar features between generations. (Like father – like son)

• Variation: refers to the fact that similar organisms can be differentiated from each other – parameters for differentiation may be size, coloration, morphology, behaviour, their physiology and biochemistry.

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• Gene: is a hereditary unit, • that occupies a specific location on a

chromosome, • determines a particular characteristic

in an organism by • directing the formation of a specific

protein, and • is capable of replicating itself at each

cell division.

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• A Gene can also be defined as a unit or chemical entity that gives rise to, or controls a trait under a given set of environmental conditions.

• Trait: is any feature about an organism that can be observed.

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• Phenotype: The phenotype of an individual organism is either its total physical appearance and constitution or a specific manifestation of a trait, such as size, eye color, or behavior that varies between individuals. Phenotype is determined to some extent by genotype.

• Genotype: is the genetic constitution of the organism.

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• Interaction between genotype and phenotype in a simple equation:

• genotype + environment → phenotype • Eg• Diabetic genotype environment diabetic phenotype

• Normal diet

• Diabetic genotype environment normal /near normal phenotype • Special diet / • Antidiabetic drugs • • = a phenotype associated with a given genotype

may vary under different environmental conditions.

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•Bacterial genetics-: the study of how the properties of bacteria can be explained by the specific genes that the bacteria contain.

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• medically, we are concerned – with genes that determine basic cellular

processes,•central metabolism •ability to cause disease in man, and to

– resist the action of antibiotics.

contributions to medicine:»rational design of new

antimicrobial compounds, & »the development of highly

effective vaccines.

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variationvariation• bacteria like other organisms breed true and

maintain their characters constant from generation to generation,

• a small proportion of their progeny show variations of particular characters.

• The phenotypes (observed character) of a pure growth are liable to two kinds of variation:

– Temporal variations due to changes in environment. (These changes are usually non-inheritable).

– Permanent variations (mutation) which are inheritable.

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Mutation: Spontaneous and Mutation: Spontaneous and inducedinduced• In a pure culture, one can find

thousands of mutants that will have different genotypes as compared to the parent cell.

• This results from spontaneous mutations that occur in the culture.

• cause of spontaneous mutation is not known.

• mutation can be induced in a culture by exposing it to X’ ray, UV light or other forms of radiation.

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• Eg.penicillin production, mutants of Penicillium chrysogenum with better yields.

• Certain chemical agents (mutagens) can cause induced mutation (eg antibiotic resistance development in chemotherapy).

• Mutation induced by such agents do not become immediately evident but only after several generations when the changed part of the genetic material has become segregated in the newly formed cells and exhibit their controlling influence on their properties and behaviour.

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DNA & Genetic information• Nucleic acids bear genetic

information of organisms (DNA & sometimes RNA)

• Implies that DNA codes for genetic composition and also

• controls the activity/ties of organisms.

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Structure of DNA• Purine bases: adenine, guanine• Pyrimidine bases: thymine, cytosine• Deoxyribose (sugar), PO4• Diagram illustrating a nucleotide, the building blocks of DNA

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• Genetic information is stored in DNA by different sequential arrangement of the purine and pyrimidine nucleotides.

• there are thousands of these compounds in each molecule of DNA

• this makes it possible to store a great deal of information in one DNA molecule.

• One sequential arrangement may control one reaction of the cell.

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• the DNA serves as a template for synthesis of more DNA (which will normally have identical structure)

• any error in the replication of the paired purine – pyrimidine sequence changes the metabolic reaction being controlled by the original DNA.

• this is thought to be the cause of mutation. • In bacteria there is only one large molecule

of DNA (unlike in eukariotic cells = double helix)

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An experiment to illustrate that DNA An experiment to illustrate that DNA bears Genetic Informationbears Genetic Information

• Experiments by Frederick Griffith (1928) found there were two different types of the bacterium Streptococcus pneumoniae:

• An "S" or SMOOTH coat strain, which is lethal to mice.

•  

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  An "R" or rough strain, which will not An "R" or rough strain, which will not hurt the mouse.hurt the mouse.

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Griffith found that he could Griffith found that he could heat inactivate the smooth heat inactivate the smooth

strain.strain.

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However, if he were to take a mixture of the heat-inactivated However, if he were to take a mixture of the heat-inactivated S strain, S strain,

mixed with the R strain, the bacteria would die.  Thus there mixed with the R strain, the bacteria would die.  Thus there was some was some

Material in the heat-killed S strain that was responsible for Material in the heat-killed S strain that was responsible for "transforming" the R strain into a lethal form. "transforming" the R strain into a lethal form.

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• Fred Griffith (and a lab co-worker) was killed in their laboratory in 1940 from a German bomb.  However, their work continued on in the U.S. in 1944, by

• Oswald Avery, C.M. MacLeod, and M. McCarty carefully demonstrated that extracts from the dead cells were responsible for the transformation.

• The chemical substance in the extract was shown to be deoxyribonucleic acid (DNA)–

• The ‘mutagen’ could be inactivated by the specific enzyme - deoxyribonuclease.

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Avery’s exptAvery’s expt• Component of the extract causing the

transformation:– Proteins or DNAR. strain + S. strain extract + Protease =

transformation occurredR. strain + S. strain extract + DNAase = No transformation conclusion: DNA (and not proteins) is the

heredity molecule.

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Alfred Hershy & Martha Chase (1952)Alfred Hershy & Martha Chase (1952)• Another expt to prove DNA as the source

of heredity information:– bacteriophage– Use 2 radioactive labels to track the

movement of – Protein (isotope 35S) & – DNA (isotope 32P) separately.

• Expt: 35S-phage (labeled protein) added to growing culture of E coli.

• Obs: 35S-label remains outside of the bacteria cells

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• Expt: 32P-phage (labeled DNA) added to growing culture of E coli.

• Obs: 32P-label enters interior of the bacteria cells

• Conclusion: DNA was the material that enters the cells & therefore must be the substance that carried the heredity information.

• Progeny bacteriophages: contained 32P and not 35S.

• Shows that DNA passes from one generation to the next.

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Plasmids Plasmids • Plasmids are small extra chromosomal

pieces of genetic materials that can replicate autonomously and maintain themselves in the cytoplasm for generations.

• Plasmids can be transferred from one bacteria to another (even of different spp).

• They are usually non essential to the cell.• Plasmids are commonly found in a

number of gram negative bacilli especially enterobacilli

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• Plasmids can contain genetic information– that determines the ability of bacteria to

mate and ( also to act as the donor during mating).

– that codes for resistance to antibiotics & other chemicals.

– that code for the degradation of various complex organic cpds.

– for toxin production (in pathogenic bacteria).

One bacterial cell can contain more that one plasmid

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GeneGene transfertransfer

The methods by which genes can be transferred in bacteria include

• Phage-mediated transduction– The temperate phage before lysis picks up some

of the genetic material of the lysogenic bacterium.

– Later when the bacterial cell lyses it releases phages some of which contain the genetic material picked from the cell.

– When such phages re-infect other bacterial cells, these take on new characteristics owing to the integration of the genetic material of the phage into the recipient’s genetic make up.

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• Phage conversion– When a susceptible bacterial culture is

infected with a phage every cell that becomes lysogenic acquires new properties imparted by the infecting phage.

– They are retained by the progeny for as long as these are parasitized by the phage in the prophage state and the prophage reproduces in synchrony with the bacterial chromosome.

– If the prophage is lost, the lysogenic stage ends and the acquired property (ties) are lost.

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• Conjugation – requires the presence of a genetic element,

the fertility or F factor. – The cell containing F factor is called the F+. – The female strains (F-) do not contain the F

factor. – a bridge forms between specific pilli (sex pilli)

on the F+ strain and receptor site on the F-. – genetic material passes only from the F+ to

the F- bacterium. – The genetic material of F+ may becomes

integrated into the chromosome of F-.

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– During the transfer across the bridge it is possible for the bridge to break at any time.

The fertility factor is ‘infectious’.

• In F+ males the factor is in the cytoplasm and during conjugation can infect F- bacterium.

• Another type of male bacterium is referred to as “Hfr” (high frequency of recombination),

• in this the F factor is integrated into the bacterial genetic material.

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• Hfr is not infectious,• transfer of chromosomal material

occurs more frequently than in F+ males

• few of the recipients of Hfr become F+ themselves.

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• Episome: a term postulated to explain the mechanisms for hereditary material transfer in transduction and conjugation.

• Episomes are genetic structures containing DNA that may be present or absent in bacteria.

• They can exist in two alternative states: • a) free in the cytoplasm and multiply

independently of cell division or • b) integrate with genetic material and multiply

only when the genetic material replicates. • An integrated episome may become

cytoplasmic and a cytoplasmic episome may become integrated.

• Thus F factor may be present and when present be free (F+) in cytoplasm or bound as in “Hfr”.

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Summary, genetic transfer

• Genetic material can be transferred between bacteria by several means, most often by:

– Conjugation – Transformation, and

– Transduction

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• Conjugation • Is mediated by a particular kind of

circular DNA called F factor (a plasmid), which replicates independently of the chromosome.

• When two cells are in close proximity to each other, a hollow bridge-like structure, forms between two cells.

• This allows a copy of the plasmid, as it is duplicated, to be transferred from one bacterium to the other.

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• Transformation • During this process, genes are

transferred from one bacterium to another as “naked” DNA.

• When cells die and break apart, DNA can be released into the surrounding environment. Other bacteria in close proximity can scavenge this free-floating DNA, and incorporate it into their own DNA. This DNA may contain advantageous genes,

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• Transduction • In this process, bacterial DNA is

transferred from one bacterium to another inside a phage that infects bacteria.

• When a phage infects a bacterium, it essentially takes over the bacteria's genetic processes to produce more phages.

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• In this process of phage infection, bacterial DNA may inadvertently be incorporated into the new phage DNA.

• Upon bacterial death and lysis, these new phages go on to infect other bacteria.

• A phage brings along genes from the previously infected bacterium.

• The imported genes from the phage incorporates into the genome of the new bacterium.

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•Thank you.

•GOOD LUCK!!!