bacterial-genetics-variation-new (1).pptx

Bacterial genetics

1. Bacterial genetics &disease By Prof. Rasheed Ajani Bakare, M.B.B.S (Ib), FMCPath, FWACP

2. Bacterial Genetics And Variations

3. The Bacterial Genome Is in a single, giant, circular loop of DNA called a Bacterial chromosome. lacks a nuclear membrane lies naked in cytoplasm located in a region of the cell known as the nucleoid. Is made up of about 3000-6000 genes

4. What is a Gene? Genes are distinct DNA sequences specifying the sequences of amino acids in a polypeptide chain

5. Characteristics of a gene Each gene determines a particular type of amino acid assembly. The chain of nucleotides that constitute a gene are composed of groups of bases called Purines and Pyrimidines Gene function can be altered or the entire gene deleted by genetic mutation

6. Characteristics of a gene (2) DNA RNA Purines Adenine Adenine Guanine, Guanine Pyrimidines Thymine Uracil Cytosine Cytosine

7. Characteristics of a bacterial genome (2) Bacteria are haploid. Have unpaired chromosomes so no heterozygotes occur Do not show dominance Each bacterium can be regarded as a clone (genetically identical to daughter cells)

8. Characteristics of a bacterial genome (3) 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

9. Characteristics of a bacterial genome (4) Undirected spontaneous mutation occurs Associated with rate of growth Accentuated by generation time

10. Control of proteins The synthesis of proteins is therefore controlled by DNA. The design of each protein is transmitted from DNA to mRNA (transcription), which then instructs the cellular machinery (tRNA and rRNA etc) (translation) to assemble a protein

11. Control of proteins (2) Proteins determine structural and metabolic function structural Enzymes Total genetic potential of DNA is the GENOTYPE Part that is manifest or discernable is the PHENOTYPE .

12. GENETIC VARIATION. Variations easily detected because Rapid rate of reproduction Large bacterial populations) are easily detected. Two types- Phenotypic Genotypic

13. Phenotypic Variation a non-heritable variation, Temporary adjustment to changes in the environment Normally involves all the cells of a culture Respond physiologically within the range of potential of the genotype

14. Phenotypic Variation (2) May manifest as changes in Morphology (size, shape, staining reactions) metabolism and chemical reactions, induction of an enzyme, e.g.,  -lactamase by the presence of penicillin repression of enzymes due to the presence of the end products in the system. Revert back to type when the inducing circumstance is removed.

15. Genotypic Variation Heritable variation Result of genetic changes mutation genetic transfer- acquisition of new heritable properties from other organisms

16. Mutation Important part of Bacterial evolution. Spontaneous mutation is common rate between 1 in 10 7 and 1 in 10 10 Environment not really important may favour or select for the variant, which may then grow and replace the wild type

17. Effect of Mutation Morphology size of a cell, colonial appearance Ability to form spores, flagella or capsules), Nutritional requirements, Antigenic properties, Ability to produce toxins, Drug sensitivity/resistance Loss of Capsule Smooth-Rough (S-R) variation Becomes less virulent. Best seen in Pneumococci, Salmonellae and Shigellae Occurs when bacteria are grown for a long time on artificial media

18. Types of Mutation Base-Pair substitution Insertions or deletion

19. Base- Pair Substitution E.g., G A A = leucine to G T A = histidine . 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 .

20. Insertion or deletion The insertion or deletion of a base will lead to a frame shift mutation 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.

21. Insertion and deletion - 2 DNA = A AC G AA 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

22. Gene exchange (genetic transfer) In eukaryotes occurs by sexual reproduction haploid gametes fuse to become a diploid zygote In Bacteria, NO DIPLOID zygotes are formed processes exist that allow for the acquisition of foreign DNA

23. Methods of Gene exchange (genetic transfer) Transformation gene transfer resulting from the uptake by a recipient cell of naked DNA from a donor cell. Transduction Transfer of foreign genes by bacteriophages Conjugation Direct transfer of DNA from one organism to another through a modified fimbriae (pilus) called a sex pilus

24. Transferable genetic materials Foreign DNA being transferred may be Plasmids Viral DNA Parts of host chromosome Transposable Elements Transposons Insertional Sequences

25. Plasmids Closed circular molecules Double stranded DNA Size from 2-3000 kilobases (100,000-150,000 base pairs) Can replicate independently of the bacterial chromosome. Code for various characteristics Fertility (F), drug resistance transfer (R), toxin production

26. Plasmids (2) Two categories Transmissible by conjugation (usually Large plasmids) Transmissible by transducing phages May be lost during growth Some cell characteristics they confer are unstable e.g., erythrogenic toxin in some Group A  -haemolytic Streptococci. Information is often in the form of transposable genetic elements called Transposons

27. Transposons- (Jumping genes) Segments of DNA Usually containing several gene s Easily transferable into the DNA of bacterial chromosomes, plasmids and infecting phages. incapable of autonomous replication. Have special sequences at the end of their DNA called terminal sequences about 15-40 base pairs are inverted repeats Sticky

28. Insertion sequences (IS elements) Can also be transferred from plasmids to chromosomes Short segments of DNA usually about 100-1000 bases (usually less than 1,500) Contain no instructions for protein synthesis Act as gene inactivators when inserted in the middle of a gene structure Presence is noted only when interference with function of the chromosomal gene occurs.

29. Implication Some toxin-, drug-inactivating enzyme can be on different plasmids or phages. E.g.,  -lactamase of E. coli, N.gonorrhoea and H. influenzae are on different plasmids which all have the same transposon. heat stable enterotoxin of E.coli is on a transposon that is also found in Yersinia enterolitica

30. Transformation- Initially regarded as a laboratory phenomenon Artificial transformation has been achieved in members of the Enterobacteriaceae (e.g. Escherichia coli, Klebsiella spp.) Certain bacteria ( e.g. Bacillus, Haemophilus, Neisseria, Pneumococcus) can take up DNA from the environment DNA that is taken up can be incorporated into the recipient's chromosome

31. Transformation First detected in the pneumococcus in 1928 by Griffith R (rough, non-virulent) strains converted to the corresponding parental S (smooth, virulent) type Also transformed to a different S-type by heat-killed cells of that type.

32. Factors affecting transformation DNA size state Double stranded DNA of at least 5 X 10 5 daltons works best. Competence of the recipient – Some bacteria are able to take up DNA naturally. By producing a specific protein called a competence factor . Other bacteria are not able to take up DNA naturally. competence can be induced in vitro by treatment with chemicals ( e.g. CaCl 2 )

33. Steps in transformation Uptake of DNA Uptake of DNA by Gram+ and Gram- bacteria differs. Gram Positive bacteria Take up single stranded DNA Complementary strand is made in the recipient. Gram Negative bacteria Take up double stranded DNA .

34. Significance of transformation Transformation occurs in nature and it can lead to increased virulence. Drug resistance Widely used in recombinant DNA technology

35. Transduction-1 Transfer of genes by infection with a non-lethal (temperate) phage. Involves transfer of DNA from the host bacteria to another bacterium. First discovered in salmonellae by Lederberg and Zinder Not all phages can mediate transduction.

36. Transduction-2 Gene transfer - usually between members of the same bacterial species. Range of species determined by Phage host range. if a particular phage has a wide host range then transfer between species can occur. Ability of a phage to mediate transduction is related to the life cycle of the phage

37. Types of Transduction Generalized Transduction Potentially any bacterial gene from the donor can be transferred to the recipient. Specialized transduction only certain donor genes can be transferred to the recipient.

38. Significance Lysogenic (phage) conversion occurs in nature and is the source of virulent strains of bacteria Responsible for penicillin-resistant strains of Staphylococcus aureus

39. Conjugation-1 [ A-B-] + [C-D-] = [ A+B+ C+D+] a 1 a 2 a 3 First reported in 1946, Lederberg while investigating genetic recombination in E.coli K12, Mated two doubly auxotrophic strains, (a 1 & a 2 ), each different in its requirements for two essential nutrients. Got a prototroph that did not require any of these nutrients

40. Conjugation -2 It was discovered that transfer of DNA was by direct contact Process called conjugation. Conjugation is a quasisexual introduction of donor DNA into a recipient through a modified pilus (fimbriae) called a sex pilus .

41. Conjugation-4 In bacteria two mating types donor (male ) recipient (female) Direction of transfer of genetic material is one way from a donor to a recipient

42. Mating Types Donor Has a plasmid called the F factor or fertility factor or sex factor.(F+) Recipient Lacks the F factor. (F-)

43. F-Factor Plasmid Has genes needed for its replication and for its ability to transfer DNA to a recipient. codes for the ability to produce a sex pilus (F pilus)

44. Conjugation E.coli undergoing conjugation. Note that the strain on the left has fimbriae and the specialised sex pilus, i.e., has an F-factor and is therefore an F+ F+ F--

45. Effect of Gene transfer Never leads to the formation of a zygote as in eukaryotic cell. At best, a merozygote is formed part of a donor bacterium’s genome (exogenote is transferred to an intact DNA of a recipient Recombination then takes place replacement of resident genes by exogenote genes or the addition of exogenote genes to the resident pool. exogenote genes may lie separate from the chromosome or may be integrated into it.

46. Recombinant- DNA Techniques Or Cloning Transfer of a gene from one bacterium,virus or animal cell into DNA of a living bacterium where it will replicate and instruct the cell .

47. Recombinant- DNA techniques Usually done between closely related organisms. New bacteria are known as recombinant organisms. Applications From novel strains of E.coli Insulin, Growth hormone, Components for vaccines (hepatitis B, H.influenzae and foot and mouth disease.), Interferon, Bloodclotting factor 8, DNA probes for the detection of specific sequences of particular pathogens