Bacterial Genetics Thymine Uracil

T:A U:A
Genetics - the study of genes, genetic variation and C:G C:G
heredity in living organisms

Genes – functional unit of heredity Strand Double stranded Single

(stable) stranded
(unstable)
Gene Expression

Exon – portion of the gene that codes for amino acids

DNA – RNA – Protein
Intron – portion of the gene that is non-coding of amino
1. DNA Replication – duplication of DNA to DNA.
acids
Responsible enzyme: DNA polymerase
Components of Nucleotides 2. Transcription - DNA to mRNA. Responsible
enzyme: RNA polymerase occurs in nucleus
• Nitrogenous base
3. Translation - tRNA to protein. Responsible
• Five-carbon sugar (ribose or deoxyribose) enzyme: Reverse Transcriptase converting RNA
• Phosphate group to DNA occurs in cytoplasm

Human Immunodeficiency virus – Example of virus with

reverse transcriptase

Polymerase Chain Reaction – simply DNA replication in

vitro (outside the body); amplification. SARS-CoV-2 not
will process this

Reverse Transcriptase Polymerase Chain Reaction –

convert RNA genetic material of SARS-CoV-2 into DNA

Amino Acids - made up of proteins

Mutation – change that occurs in the DNA sequence

and result of exposure to physical agent (e.g. UV) that
can lead to cancer

• Deletion
• Duplication
Deoxyribonucleic Ribonucleic • Inversion
Acid (DNA) Acid (RNA) • Translocation

Recombination – genes are transferred or exchange

Sugar Deoxyribose Ribose sugar
sugar between homologous region of two DNA molecules

Nitrogenous Guanine Guanine

bases Cytosine Cytosine
Adenine Adenine

Mechanism of Gene Transfer
Plasmid – are extra-chromosomal DNA containing the
coding of the virulence factor (e.g. antimicrobial
susceptibility, capsule formation)
1. Transformation
- Principle: When the bacteria dies, there will be
a liberation of the bacterial genetic makeup
that will take up by the other living bacteria.
The gene material will be incorporated into the
main gene material of the host
- Uptake and incorporation of free or naked DNA
into bacterial cell
2. Transduction
- Principle: uses bacteriophage or phage - a virus
that feeds on and infects the bacteria
- Bacteriophage will attach to the donor
bacterium and it will inject the genetic material
once attachment happened
- Protein and Genetic material (major parts of the
virus)
- Inside the donor bacterium, there will be the
development of the virus forming new virus
using the genetic material of that organism
- as a result, some of the bacteriophage will get
the genetic material of the donor bacteria
instead of getting the genetic material of the
previous bacteriophage
- Bacteriophage containing the gene material of
the donor bacteria and incorporated in the
genetic material of the receiving host
- Viruses: will only infect a living organism
- Summarized: Transfer of bacterial gene by a
bacteriophage or a virus from one cell (donor
bacteria) to another (receiving host)
3. Conjugation
- Principle: The transfer of gene material will be
on bacteria to bacteria through the use of sex
pili (transfer gene material)
- Transfer of genetic material from bacterial
strain to a recipient strain
Culture and Culture Media
Culture - growth of microorganism on a culure medium
Culture medium – is a liquid, semi-solid or solid
medium utilized to observe growth patterns of
microorganism as well as for transport and storage
Mucoid colonies
Type of culture facultative
anaerobes
1. Pure culture – only one organism (one shape,
color, consistency) Examples Tryptic Soy Sulfide Indole Triple Sugar
2. Mixed culture – made up of more than one Broth (TSB) Motility Iron (TSI)
organism (different morphology, appearance, Thioglycolate Medium (can Agar,
Brain heart determine the MacConkey,
color) infusion (BHI) motility of the Blood Agar
- Never use mixed colonies: may cause erroneous organism) Plate (BAP),
results and hard for identifying organisms Chocolate
- Perform subculture when dealing with mixed agar
colonies. Getting one colony and subculture to Growth Formation of Haziness
indicator Turbidity and within the tube
another medium and repeat.
Sediments (overall the
3. Stock culture – composed of several species medium)
contained in a separate culture medium (one means motile
species/culture medium) organism
- commonly used in academic and industrial
Growth in stub
purposes
area means
organism is
non-motile
Others blackening of
the medium
indicated
formation of
hydrogen
sulfide
Pure Culture Mixed Culture Used in Identification
of
Enterobacteria
• Agar
- Solidifying agent
- Made up of: Sulfated polymer made up of D-
galactose, 3-6-anhydro-L-galactose, and D-
glucoronic acid
Stock Culture - Derived from red algae
- Karenia brevis – causative agent of red tide
Classification of Culture Medium
- Agar will melt if the temperature reaches 80 to
A. According to consistency 90 C. It will produce moisture thus
inappropriate in culturing
Liquid Semi-solid Sodium
- Solidify at 40 to 50 C
medium medium medium
Agar 0% agar 0.5 to 1% agar 2-3% agar - Cooling temp for distribution of agar into
Definition they are fluid does not have plates: 55-60 C – RECOMMENDED
no solidifying the same TEMPERATURE
agent (agar) consistency in
the solid agar Liquid Medium
nor in liquid
medium

Other Broth
term
Function Allows the
growth of:
aerobes
anaerobes
Semi-solid Medium

• Obligate intracellular bacteria – requires a living

host to survive like cells, Chlamydia and
rickettsia. Without a host/cell, they will not
grow and survive.
• At the bottom of the medium, it can see cancer
cells. Purpose: cancer cells are the culture
medium.
• Uses Inverted microscope to observe the
sample (light source at the top, lens at the
bottom because cells are found in the bottom
layer)

Solid Medium

C. According to How the Medium is

Dispensed/Distributed

Plated Medium Tube Medium

B. According to composition Orientation Placed in petri Placed in a tube
dish
Synthetic Non- Tissue
or defined synthetic culture Types Butt
medium or Complex medium Slanted
Medium Slant with butt
Definition Contains a Composed Used for
known of some cultivation
component unknown of obligate
to the user substances intracellular
(peptones, bacteria
meat, yeast
extract)
Other term Defined Complex
medium medium
Uses Research Isolation of Obligate Plated medium Slant with butt
purposes bacteria intracellular
bacteria
Common
use in the
laboratory
Composition Fluid
Example Nutrient W138,
medium broth, HeLa 229
Tryptic Soy cells,
Broth (TSB), Mc Coy cells
MacConkey
Agar
D. According to use:
1. Simple media/ General purpose media/
Supportive media
- routinely used in the laboratory
- without added supplement (non-fastidious
bacteria)
- Composition: Meat and soybean extracts
- Examples:
o Nutrient agar (NA)
o Broth
o Trypticase Soy broth (TSB) 4. Differential media
- Media allow the visualization of the metabolic
differences between groups of bacteria
- Differentiation based on the pH indicators
- Example:
o MacConkey Agar – can differentiate a
lactose to a non-lactose fermenter)
Lactose fermenter: pink colony (+)
2. Enrichment broth Non-lactose fermenter: colorless
- Used to propagate the growth of certain group colony (-)
of organisms o Blood Agar Plate – differential medium
- Contains specific nutrients: liquid type media and enriched medium. It can
- Examples: differentiate medium using its
o Alkaline Peptone Water - culture hemolytic properties/pattern. No
medium for Vibrio spp. indicator used.
o Selenite F – culture medium for Beta hemolysis is when the organism
salmonella can completely lyse the blood = clear
o Thioglycolate – culture medium for zone of hemolysis
aerobes Alpha hemolysis can give partial
o Tetrathionate - culture medium for hemolysis = greenish zone of inhibition
salmonella and Proteus Gamma hemolysis = no hemolysis (or
o Gram-Negative broth - culture medium no hemolytic pattern)
for Salmonella and Shigella
o Lim broth - culture medium for group B
streptococci
3. Enriched media/non selective media
- With added supplements
- Supplements: blood, vitamins and yeast extract
(fastidious bacteria)
- These are solid-type media
- Examples:
o Blood Agar Plate - media with blood of
sheep or horse.
*Horse blood used for the cultivation of
haemophilus
o Chocolate Agar Plate - resembles of
chocolate color. Same procedure with
BAP procedure except that it has
heating)
 6. Special media
- Used to isolate bacteria with specific growth
5. Selective media
requirements
- Media incorporated with antibiotics, dyes or
- Examples:
chemicals (inhibitors) to inhibit the growth of
o Löwenstein-Jensen Medium –
other organism while promoting the growth of
cultivation for Mycobacterium
the desired organism
tuberculosis
Examples:
o Thiosulfate-citrate-bile salts (TCBS) -
o MacConkey Agar – both a selective and
cultivation of Vibrio cholerae
differential media. It promotes the
growth of gram-positive organism
o Xylose Lysine Desoxycholate agar (XLD)
- promotes the growth of gram-
negative organism
o Bismuth Sulfate Agar - promotes the
growth of gram-negative organism
o Mannitol Salt agar (7.5% NaCl) –
contains high-salt concentration and
thus, not suitable for halophiles pH Acid Alkaline Example
organisms Phenol Red TSI, XLD,
MSA
Others: Bromthymol HEA, SCA,
blue TCBS
o Hektoen Enteric Agar – contains bile salt and
Neutral red MAC, SSa
dyes (acid fuchsin) for non-enteric pathogens; it
Bromcresol LIA
inhibits the normal flora of the lower GIT; purple
selective media
o XLD agar – contains 0.25 % sodium
desoxycholate salt and sodium thiosulfate to
differentiate salmonella (red with black centers)
from shigella (red or clear colonies). Both a
differential and selective media.

Inhibitory substances in the Selective Media

Gram-positive Gram-negative Swarming

bacteria bacteria bacteria
Crystal/gentian Potassium Alcohol,
violet, tellurite, Choral hydrate
Basic Fuchsin Sodium azide
Bile salt
NOTES TO REMEMBER:

1. Selection of media is based on the type of

specimen (mahon table 6.4 p 115)
2. If several plates will be inoculated for a given
specimen, the media should be arranged
beginning with the MOST ENRICHED medium
and the progressing to the most selective