General feautures Sporulation cycle and germination Genetic elements of bacteria- chromosomes, plasmids, transposable elements, integrons and gene cassettes

1. Darshan
ENDOSPORES AND BACTERIAL
NUCLEIC ACIDS

2. Contents
• Introduction (endospores)
• Endospore structure
• Sporulation cycle
• Germination
• Introduction (bacterial nucleic acids)
• Chromosomes
• Plasmids
• Transposons
• Integrons and antibiotic resistance cassettes
• Conclusion
• Reference

3. Introduction (endospores)
• Extreme survival strategy employed by certain low G+C gram
positive bacteria (Bacillus, Clostridium and Sporosarcina)
• Initiated in nutrient deprivation
• Bacteria produces a dormant and highly resistant cell to
preserve genetic material from environmental stresses
• First studied by Ferdinand Cohn (1876)
• Remain viable for longer period of time (e.g. Thioactinomyces
vulgaris’ 7500 years endospores)
• Dangerous pathogens and are practically important in the fields
of food, industrial and medical microbiology

4. Endospore structure
• Visible by light microscopy as strongly
refractive structures
• Basic stains and malachite green
• Electron micrographs show core surrounded by
several layers
• Cortex- made of less cross-linked
peptidoglycan
• Core wall- contains peptidoglycan that will
form cell wall on germination
• Coat- composed of highly cross-linked proteins

5. Contd.
• Exosporium- a thin delicate covering
• Various membranes contribute to resistance of spore
Spore coat- protects chemicals and lytic enzymes (e.g. lysozymes)
Inner membrane- impermeable to various chemicals
Core has very low water content and high amounts of
dipicolinic acid complexed with Ca ions
• DNA is stabilized by Ca-DPA complexes (inserted between
nitrogenous bases) and small acid-soluble DNA-binding
proteins (SASPs)

7. Sporulation Cycle
A complex process
that occurs in a
highly organized
fashion for several
hours

8. Germination
• Transformation of dormant spores into
active vegetative cells
Activation- prepares spores for
germination and can result from
treatments such as heating
Germination- the breaking down of the
spores dormant state
Begins with germinant receptors which
detect sugars and amino acids
Release of Ca-DPA complexes, breakdown
of peptidoglycan in cortex and water
uptake
Outgrowth- emergence of spores from
spore coat and development into an active
vegetative cell
Clostridium pectinovorum emerging from
the spore during germination

9. Introduction (bacterial nucleic acids)
• Genetic information in cells is embedded
in the nucleic acids DNA or RNA
• DNA carries genetic blueprint for the cell
• RNA is an intermediate molecule that
converts blueprints into defined amino
acid sequences in proteins
• Genetic information resides in the
sequence of monomers in nucleic acids

10. Contd.
• Nucleotides, the monomers of nucleic acids (pentose sugar+
nitrogenous base + phosphate)
• Nucleosides, a nitrogenous base attatched to its sugar, but
lacking phosphate
• Bacteria are prokaryotes
• Nucleoid, an irregular shaped region in bacterial cell that contains
cell’s chromosomes and numerous proteins
• Extra-chromosomal genetic elements, the plasmids

11. Chromosomes
• Chromosomes of most bacteria are single circle
of dsDNA
• Exceptions- Linear chromosomes found on
most species of Steptomyces and presence of
more than one chromosomes in bacteria such
as Vibrio cholerae and Boriella burgdorferi
• Longer than the length of the bacterial cell. E.g.
E. coli’s circular chromosome measure 1400μm
and are 230-700 times longer than cell. Thus
are supercoiled that decreases overall size

13. Plasmids
• Extra chromosomal DNA molecules
• Borelia burgdorferi has 12 linear plasmids and 9 circular plsmids
• Small dsDNA molecules that can exist and replicate independently
of the chromosome
• Have relatively few genes (<30), which are not essential to the
bacterium
• Carry genes that confer a selective advantage to the bacterium in
certain environments

14. Phenotypes
• Antibiotic resistance
• Antibiotic production
• Degradation of aromatic compounds
• Haemolysin production
• Sugar fermentation
• Enterotoxin production
• Heavy metal resistance
• Hydrogen sulphide production

15. Episomes
Plasmids integrated to bacterial chromosomes
Replicate along with bacterial chromosomes
Integrative plasmids
Eg: Ti-plasmid of Agrobacterium tumaeficiens
• Inherited stably during cell division, but are not always equally
appointed equally to daughter cells and sometimes lost
• Curing- loss of plasmid

16. Classification based on phenotype
• R-plasmids: confer resistance against antibacterial chemical agents Eg: RP4
plasmid found in Pseudomonas
• F-plasmids: conjugative plasmids found in F+ bacterium with high
frequency of conjugation Eg: F plasmid of E. Coli
• Col plasmids: have genes encoding colicins, proteins that kill other bacteria
Eg: ColE1 of E. coli
• Degradative plasmids: that allow host bacterium to metabolize unusual
molecules such as toulene and salicylic acid Eg: TOL of Pseudomonas putida
• Virulence plasmids: confer pathogenicity on host bacterium Eg: Ti
plasmids of Agrobacterium tumefacians
• Cryptic plasmids: do not have any apparent effect on phenotype of
bacterium. They just code for enzymes

17. Transposable elements
• Stretches of DNA that can move from one site to another
by transposition
• Found inserted to plasmids or chromosomes of bacteria
• Do not possess their own origin of replication
• Replicate during host DNA replication
• Insertion sequences and transposons
• Both carry genes encoding transposases and have
inverted terminal repeats at their ends

18. Insertion sequences
• Short DNA segments, about 1000 nucleotides long, 10-50bp
• Only encodes transposases

19. Transposons
• Larger than IS elements
• Transposases recognize inverted repeats and transpose the
segment of DNA flanked by them from one site to another

20. Mechanism of transposition

21. Integrons and antibiotic resistance cassettes
• Genetic entities that are
able to capture small
mobile elements known
as gene cassettes
• Gene cassettes are small,
descrete mobile elements
comprising a single gene
and a downstream 59-be

22. Chromosomal(Cis) and mobile integrons(MIs)
• CIs are located on chromosomes of of many bacterial species
• Often found in bacteria from marine and terrestrial ecosystems
• E.g. Vibrio and Xanthomonas spp.
• Also called as super integrons as they carry upto 200 cassettes
that encodes proteins
• MIs are located on mobile genetic elements such as transposons
and plasmids
• Contain limited number of gene cassettes (<10 GCs)
• GCs are responsible for antibiotic resistance and are called as
antibiotic resistance cassettes

23. Antibiotic resistance cassettes
• Enclosed within attI and attC sites
• Dependent on integrons for expression
Production of antibiotic resistance enzymes
Formation of efflux pump
Modification of antibiotic binding target

24. Conclusion
Endospores
• Dormant cells produced by certain bacteria in nutrient difficient
conditions that are resistant to various environmental stresses
• Once favourable conditions return they germinate to get back to
normal lifecycle
Nucleic acids
• Single circular DNA as chromosomes (some exceptions are there)
which are compactly packed into nucleoid-associated
• Bacteria have extracellular genetic elements, plasmids
• Transposable elements are present in bacteria which can act as
recombination agents
• Integrons capture gene cassettes and help them in their replication
• Antibiotic resistance cassettes- degradation enzymes, efflux pumps and
modification of antibiotic binding site

25. Reference
• Sherwood, and Woolverton Willey (2014), Prescott, Harley, and
Klein's Microbiology (9th International Edition), McGraw-Hill
• Michael T. Madigan, David P. Clark, David Stahl, John M.
Martinko, 2012, Brock Biology of Microorganisms 13th Edition,
Benjamin Cummings
• Jeffrey C. Pommerville (2010) Alcamo’s Fundamentals of
Microbiology, 9th Revised edition, Jones and Bartlett Publishers,
Inc
• Sydney Brenner and Jeffery H. Miller, Encyclopedia of genetics,
volume 1, Academia press

26. Thank you