MICROBIAL GENETIC

BY
Dr Ola-Bello Olafoyekemi Ibiwunmi
Consultant Microbial Pathologist
UNIMED/UNIMEDTH
 Aims and Objectives
• An overview of microbial genetic content
• Basic definitions
• Understand the methods of gene expression
• Understand the basis of microbial gene
replication
• Explore the mechanisms of genetic transfer
• Relevance of microbial genetics in modern
medicine
 Introduction
• Microbial genetics involves both discovering the
regulatory genes and sites that control individual gene
expression and determining which genes are co-
regulated and likely to participate in the same process.

• Genetics is the study of genes including the structure of

genetic materials, what information is stored in the
genes, how the genes are expressed and how the
genetic information is transferred.

• The central dogma of molecular biology is that DNA

carries all genetic information.
 Basic definitions
• A DNA sequence that encodes for a specific
product (RNA or protein) is defined as a gene.
• GENOME: The total complement of genes in a
cell or in a chromosome
• The genome is organized into discrete
elements known as chromosomes.
 Basic definitions
• Chromosomes are chains of double stranded
DNA, where the cell's genome is stored
• DNA is the genetic material of bacteria. It
carries all genetic information.
– The two essential functions of genetic material
are replication and expression
• Genes are sequences of nucleotides within
DNA that code for functional proteins.
 STRUCTURE OF DNA
The DNA molecule is composed of two
chains of nucleotides wound around
each other in the form of double helix.
Each nucleotide contains one
phosphate group, one deoxyribose
sugar, and one purine or pyrimidine
base.
In DNA the sugar is deoxyribose; in
RNA the sugar is ribose.
The double helix is stabilized by
hydrogen bonds between purine and
pyrimidine bases on the opposite
strands
 Introd contd
Nucleotides DNA RNA

Purines Adenine Adenine

Guanine Guanine

Pyrimidines Thymine Uracil

Cytosine Cytosine
 Basic definition
• RNA rarely exists as a double-stranded
molecule.
• The three major types of RNA :
 messenger RNA [mRNA],
 transfer RNA[tRNA], and
 ribosomal RNA [rRNA])
• All play key roles in gene expression.
 Bacterial genome
• Bacteria are classified as prokaryotes
• Nearly all prokaryotes are unicellular.
• Prokaryotes contain no membrane-bound
organelles; their only membrane is the
membrane that separates the cell form the
outside world.
• Prokaryote genetic material is located in the
cytoplasm (although sometimes confined to a
particular region called a “nucleoid”).
• The Bacterial Genome
– Is in a single, giant, circular loop of
DNA called a Bacterial chromosome.
– lacks a nuclear membrane
– lies naked in cytoplasm
– Is made up of about 3000-6000 genes
 Replication and expression of
genetic information
• Bacteria multiply by cell division, resulting in the
production of two daughter cells from one
parent cell.
• The genome replicate to produce two daughter
cells with an identical copy of functional DNA.
• Gene expression is the processing of information
encoded in genetic elements which results in
the production of biochemical molecules,
including RNA molecules and proteins.
 Flow of genetic information
• Replication of DNA to make more DNA,
• Transcription of the DNA into mRNA
• Translation of mRNA into proteins
 Gene expression
Gene expression is the process by which information
from a gene is used in the synthesis of a functional
gene product.
Composed of;
– Transcription: synthesis of RNA from a DNA
template.
– Translation : formation of a protein (amino acid
sequence) from RNA sequence.
– Folding.
– Transport.
 Gene expression
• Transcription proceeds in a 5’ to 3’ direction.
• DNA helicase – unfold the parental DNA strand
that is used as a template.
• Other enzymes involved include DNA
polymerase, DNA ligase.
• Results in complimentary DNA or RNA
production.
• Transcription process produces mRNA, tRNA
and rRNA.
 Gene expression
• Translation involves protein synthesis.
• Through this process the genetic code in mRNA
molecules is translated into specific amino acid
sequences that are responsible for protein
structure and function
• Three parts:
Initiation - start codon (AUG)
Elongation - ribosome moves along mRNA
Termination - stop codon reached/polypeptide
released and new protein forms.
 Gene expression
• NOTE!
 Each set of three bases is known as a codon
– AAC, GCT, TAG etc.
 Each codon codes for a specific amino acid.
e.g., leucine, valine
 There are over 64 different triplet sequences
and 20 amino acids
• These then undergo processing in form of
folding and conformation to result in functional
proteins.
Regulation and Control of Gene Expression

• Regulation occurs at one of three levels of

information transfer from :
• the gene expression level
• protein synthesis pathway:
 transcriptional regulation; Genetic control
 Translational;
• Posttranslational level.
 Genetic variation
• Rate of reproduction is so rapid and bacterial
populations so vast that changes in properties
(variations) are easily detected.
• These are of two types-
– Phenotypic
– Genotypic
 Genetic variation
Genotype is the arrangement (order of
DNA bases) of genes within an organism
A change in the sequence of template DNA
is called mutation
Phenotype is the physical characteristics
of an organism (based on its genotype)
and the interaction with its environment
 Genotypic Variation
– This is an inheritable variation due to
changes in the genetic constitution. This
may be a consequence of:
• mutation
• genetic transfer- acquisition of new heritable
properties from other organisms
 Phenotypic Variation:.

• This is a non-heritable variation, which

represents a temporary adjustment to
changes in the environment
• It normally involves all the cells of a culture,
• Respond physiologically within the range of
potential of the genotype
 Phenotypic Variation (cont)

• May manifest as changes in

– Morphology (size, shape, staining reactions)
– metabolism and chemical reactions,
– induction of an enzyme, e.g., b-lactamase by the presence
of penicillin
– repression of enzymes due to the presence of the end
products in the system.
• They revert back to type when the inducing
circumstance is removed.
 Mutation
• This is an important aspect of Bacterial evolution.
Spontaneous mutation is common. Occurs at a rate
between 1 in 107 and 1 in 1010
• The environment is important only in that it may
favour or select for the variant, which may then grow
and replace the wild type
• Mutations are heritable changes in genotype that can
occur spontaneously or be induced by chemical or
physical treatments
 Mutation (cont)

– Example - Smooth-Rough (S-R) variation in which the

bacterium loses a surface component and therefore
becomes less virulent.
– Best seen in Pneumococci, Salmonellae and Shigellae and
occurs when bacteria are grown for a long time on artificial
media.
 Significance of mutation
• Mutations can result in change in phenotype
such as
– appearance of novel surface antigen,
– alternation in physiological properties,
– change in colony morphology, nutritional
requirements, biochemical reactions,
growth characteristics, virulence and host
range
 Significance of mutation
• Discovery of a mutation in a gene can help in
identifying the function of that gene.
• • Mutations can be induced at a desired region to
create a suitable mutant, especially to produce
vaccines.
• • Spontaneous mutations can result in emergence of
antibiotic resistance in bacteria.
• The process of mutation is called mutagenesis and
the agent inducing mutation is called mutagen.
 Mutagens
Mutagens can be chemicals

 nitrous acid, which alters adenine to pair with

cytosine instead of thymine.
acridine dyes, nucleoside analogs that are similar
in structure to nitrogenous bases,
Radiation can also be a cause of DNA mutations. High
energy light waves such as X-rays, gamma rays, and
ultraviolet light have been shown to damage DNA.
These change the physical shape of the DNA
preventing transcription and replication
 Types of Mutation
• Point mutations
– Base-Pair substitution
– Insertions or deletion
• Conditional Lethal Mutations
 Base- Pair Substitution

• E.g., GAA = leucine to

GTA = histidine.
• This is called a Missence Mutation
• Results in the substitution of one amino-
acid for another.
• It may or may not affect the function of
the polypeptide.
 Insertion or deletion
• The insertion or deletion of a base will lead to
a frame shift mutation.
• The deletion of a base unless it is
compensated for, by the insertion of a new
base very close to the deleted base, will lead
to a new polypeptide being formed.
 Insertion and deletion
• DNA = AAC GAA CGC TGA
• RNA = UUG CUU GCG ACU…. =
leucine, histidine, alanine, threonine

• If A is deleted there will be a left shift such that

the above DNA code will read starting from the
second A as:

• DNA = ACG AAC GCT GA…..

RNA =UGC UUG CUU CU……
cystine, leucine, arginine, leucine
 Conditional Lethal Mutations
• Mutations that are lethal under one set
of circumstances but not under another.
• “Permissive” conditions e.g.,
temperature-sensitive mutants
 Genetic Exchange
• Genetic transfer - results in genetic variation.
• Genetic variation - needed for evolution.
• Genes are then transferred by
• Vertical method
– Parent to offspring
• Horizontal methods
Mechanism of Gene Exchange
• Conjugation
– Occur between two living cells
– Direct transfer of DNA from one organism to another through a
modified fimbriae (pilus) called a sex pilus
 Mechanism of Gene Exchange
• Transformation
– gene transfer resulting from the uptake by a recipient cell of
naked DNA from a donor cell (another bacterial cell)-dies, lysis
• Such bacteria are said to be competent- Haemophilus,
Streptococcus, and Neisseria
• Transduction
– mechanism by which DNA from two bacteria may come
together in one cell
– Transfer of foreign genes by bacteriophages (bacterial
virus)
 Transferable genetic materials
Foreign DNA being transferred may be
– Plasmids
– Viral DNA
– Parts of host chromosome
– Transposable Elements
• Transposons
• Insertional Sequences
 Plasmids
• Plasmids are extra-chromosomal elements found
inside a bacterium. They are not essential for the
survival of the bacterium but confer certain extra
advantages to the cell.

• They are closed circular molecules of double

stranded DNA that range in size from 2-3000 kilo
bases (100,000-150,000 base pairs) but typically
contain the size of the chromosome.
 Plasmids
• Plasmids multiply independently of the
chromosome and are inherited regularly by
the daughter cells.
• Types of plasmids
 R factor, Col factor, RTF and F factor
• No and sizes
– A bacterium can have no plasmids at all or have
many plasmids (20-30) or multiple copies of a
plasmid.
 Significance of plasmids
• Code for resistance to several antibiotics especially
in Gram-negative bacteria
• Code for the production of bacteriocins.
• Code for the production of toxins (such as
Enterotoxins by Escherichia coli, Vibrio cholerae,
exfoliative toxin by Staphylococcus aureus and
neurotoxin of Clostridium tetani).
 Types of plasmids
F factor: (fertility factor or sex factor)
Most plasmids are unable to mediate their own transfer to
other cells.
F factor is a plasmid that codes for sex pili for its own
transfer to other cells.
Those bacteria that possess transfer factor are called F+,
(male, donor) such bacteria have sex pili on their surface.
Those cells lacking this factor are designated F- (female,
recipient)
This plasmid is transferred to other cells through
conjugation.
An F- cell will become F+ when it receives the fertility factor
from an F+ cell.
 Types of plasmids
R factor: a plasmid that codes for the transmissible
drug resistance
These plasmids contain genes that code for
resistance to many antibiotics.
Bacteria possessing such plasmids are resistant to
many antibiotics and this drug resistance is
transferred to closely related species through
conjugation
 They are usually transferred to related species along
with RTF.
 Transposable elements
• These are of two types -
1. Insertion sequences simply contain genes for
insertion and replication in host
chromosome.
2. Transposons is the process by which these
genetic elements excise from one genomic
location and insert into another.
 Transposable elements
• Transposons do not exist independently in the cell.
• must be incorporated into the chromosome or
plasmids or both
• They are often referred to as “jumping genes”
because of their ability to change location within and
even between the genomes of bacterial cells.
• Transposons carry genes that have products that help
mediate the transposition process, in addition to
genes that encode for other accessory characteristics,
such as antimicrobial resistance.
 Benefits of understanding and using
Bacterial genetics
Recombinant- DNA techniques
Recombinant DNA techniques, i.e., the
transfer of a gene (length of DNA) from one
bacterium, virus or animal cell into the
genetic apparatus of a living bacterium
where it will replicate and instruct the cell.
 Recombinant- DNA techniques -(cont)

Usually done between closely related organisms.

The new bacteria are known as recombinant
organisms and the technique is known as
recombinant – DNA technique or cloning.
 Recombinant- DNA techniques (Applications)

In medicine, novel strains of E.coli have been

created which are capable of producing
– Insulin,
– Growth hormone,
– Components for vaccines (hepatitis B, H. influenzae
and foot and mouth disease.),
– Interferon,
– Blood clotting factor 8,
– DNA probes for the detection of specific sequences of
particular pathogens.
Genetic mechanisms of drug resistance
• Mutations
• Transferable drug resistance:
The physiological mechanisms of antibiotic
resistance include
• Inactivation of the antibiotic by enzymes produced
by the bacteria
• Alteration of target proteins such that the antibiotic
doesn’t bind or binds with decreased affinity
• Alteration of the membrane which decreases the
permeability of the antibiotic
• Active efflux of the antibiotic
• Development of alternate metabolic pathway to
bypass the action of antibiotic
Thank you for the attention