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VIRAL and BACTERIAL GENETICS
M.Tevfik Dorak
Historical landmarks in viral and bacterial genetics
1944 Avery's pneumococcal transformation experiment shows that DNA is the hereditary material
1946 Lederberg & Tatum describes bacterial conjugation using biochemical mutants
1950 Barbara McClintock finds transposable elements in Maize
1952 Hersey & Chase shows that the hereditary material of the bacteriophage is DNA; Zinder & Lederberg achieves phage-mediated
transduction in Salmonella
1953 Cavalli-Sforza et al show the F factor in bacteria (Cavalli-Sforza LL, Lederberg J, Lederberg M. J Gen Microbiol 1953;8:89)
1961 Jacob & Monad describes the operon structure
1977 Sanger sequences the phage FX174 as the first viral genome (identification of overlapping genes)
1995 First bacterial genome (H. influenza) is sequenced (Fleischmann et al, Science 1995)
Definitions
A virus is a disease causing agent consisting of a nucleic acid molecule and protein coat. Viruses are incapable of autonomous replication and have to use a host cell's translational system. A bacterium is a prokaryotic cell with its own circular DNA. A bacteriophage is a virus that infects bacteria only. Viruses would appear to be the simplest infectious particle. The discovery of viroids, nucleic acid without a protein capsule and prions, infectious proteins, subtracts another level of complexity. Both viroids and prions can cause diseases.
Properties of Viruses
They can be morphologically variable and even complex. Their morphology is one way of classifying viruses. They contain DNA or RNA but never both. Although they have a protein coat around their hereditary material, they lack properties of cells such as membranes, ribosomes, enzymes and ATP synthesis ability. Thus, they can only proliferate by using a host cell's translational machinery. In other words, they are obligate intracellular parasites. Their replication cycle consists of attachment and entry into the cell; replication of viral nucleic acid; synthesis of viral proteins; and finally, assembly of viral components and escape from host cells. Since a virus is not a living cell, they are neither prokaryotes nor eukaryotes. Viruses are widely used as vectors in gene therapy.
Properties of Bacteriophages
They have a simple structure, which consists of their double-stranded DNA and protein coat. Only the DNA enters into the bacteria. Bacterial RNA polymerase is composed of five individual polypeptide subunits (a2, b, b', s). The s (sigma) factor is responsible for initiating transcription by recognizing bacterial promoter DNA sequences. Some phages supply their own s factors to instruct the bacteria to transcribe phage genes preferentially. They are virtually viruses but can only infect bacteria. Bacteriophages usually infect only one species of bacteria but there are some who can infect several species even in different genera. Their life cycle may be either lytic (virulent phage) or lysogenic (temperate phage). Their DNA may be integrated into the host chromosome and remain as a prophage. Integration is achieved by recombination between a 15 bp sequence called att (for attachment) in the host chromosome and an identical sequence in the phage chromosome. This recombination requires an integrase (Int) enzyme encoded by the phage. Bacteriophages are used for DNA cloning in molecular biology. DNA fragments can be inserted into a phage and following transfection of a competent bacteria, many copies of the desired DNA fragment can be obtained.
Properties of Bacteria
A bacterium has four types of genetic material: its single (haploid), covalently closed, circular dsDNA chromosome (in a supercoiled state); a plasmid(s); a bacteriophage or prophage; and a transposon. Genetic exchange between bacteria can occur by transfection, transduction or conjugation. Conjugation involves F+ male bacterium and F- female bacterium. Bacteria are haploid, but following a gene transfer (such as conjugation), they can be partially diploid (merozygote). This may result in a double cross-over event between the circular DNA and the linear newly introduced DNA if the two copies of the DNA are related. Sexual reproduction and meiosis do not occur in bacteria but genetic recombination to increase diversity is still possible by horizontal gene transfer (see below).
While bacteria are haploid organisms, plasmids can be considered as additional mini-chromosomes. Plasmids can be 1 to 300 kb long and may exist as multiple, free copies in a bacterium. As a rule, small plasmids occur in multiple copies per cell (high copy number), and large plasmids have a low copy number. Plasmids cannot replicate outside a bacterium. More than one types of plasmids can co-inhabit the same bacterium. Up to 10 kb (on average 3 kb) long DNA fragments can be inserted into a plasmid. They can enter the cells in two ways: vertical (via cell division - binary fission) or horizontal transmission (bacterial gene swapping). Some plasmids may contain genes that confer an evolutionary advantage to their hosts. These can be anti-bacterial toxins, catabolic enzymes (to use unusual carbon sources), virulence factors (pathogenic toxins), enzymes to degrade toxic compounds (like polychlorinated biphenyls, pesticides) and most importantly, antibiotic resistance (conferred by R plasmids). Sometimes, they may confer resistance up to five antibiotics at the same time. Plasmids can be exchanged between unrelated bacteria. This is the reason for speedy spread of antibiotic resistance among them.
Sometimes, a bacterium also contains a prophage as an inserted DNA fragment into its chromosome and this additional genetic material may be beneficial for it. For example, a prophage of the bacterium Corynobacterium diphtheriae carries a gene that encodes the diphtheria toxin causing the disease. Temperate phages may exist in a bacteria in a non-replicating, latent state (prophage). Naturally, every time a bacteria divides (every 15 to 20 minutes), the prophage will also be replicated.
Transposable elements cannot exist as free particles in bacteria. They are integrated in the bacterial genome or into the genetic material of a plasmid or a prophage. They have the ability to move between these sites using an enzyme called transposase. Transposons may also encode proteins that are useful for bacteria (such as antibiotic or heavy metal resistance factors).
Transformation involves the uptake of DNA from the environment. Cells that are able to take up DNA are called competent. While some bacteria (H. influenzae, B. subtilis) are naturally competent owing to some surface proteins they possess, others can be made competent by various treatments (calcium chloride treatment or electroporation). This kind of transformation is an important method used in genetic engineering. Bacteria can take up DNA from other bacteria in nature (potentially from genetically modified bacteria) but the fate of such DNA is usually degradation. Historically, the principal application of transformation experiments was genetic mapping studies on naturally competent bacteria (co-transformation frequencies are inversely related to map distances).
Transduction involves the transfer of bacterial DNA by means of a phage particle. Here, a desired piece of DNA is packaged into the bacteriophage head. The bacterial DNA (which can be an entire plasmid) can be transferred to a new cell when it is infected by the phage particle. In specialized transduction, the genome of a temperate phage (such as l) integrates as a prophage into a bacterium's chromosome usually at a specific site. In generalized transduction, it is not a specific DNA segment but whatever DNA has been loaded into the phage is transferred. For example, the m and P1 phages of E.coli can achieve generalized transduction. Transduction can also be used to establish gene order and for mapping purposes (only closely spaced genes will show co-transduction). In nature, a phage may transfer parts of bacterial DNA from one bacterium to another.
In conjugation, a direct contact between a male (carrying a fertility factor, or F+) and a female (F-) bacteria results in a one-way genetic material transfer (from male-to-female). Gram-negative bacteria (like E.coli) use a physical bridge called (sex) pilus (encoded by a conjugative plasmid) for gene transfer in conjugation, whereas, gram-positive bacteria (like pneumococcus) use a protein called clumping factor to get together. Some phage use the pili as receptors to attach to the bacteria. During conjugation in E.coli, the F factor (which is a conjugative plasmid) is not lost from the donor as it is only one of the strands of the plasmid that has been transferred. Subsequent replications of the bacteria restore the double-stranded state of the plasmid. When a conjugative plasmid initiates conjugation, other plasmids can be transferred (this is called mobilization). Conjugation is the exception to the rule that bacteria reproduce asexually. Although conjugation resembles sexual reproduction, an important difference is that conjugation is a one-way process. The F factor may exist as a free plasmid or may be inserted into the bacterial genome. Some conjugative plasmids (like the F factor of E.coli) can achieve transfer of chromosomal genes. An E.coli strain that has this property is called Hfr strain (for high frequency recombination). It is important to know that chromosomal genes are transferred before the plasmid itself. If the bridge is broken during transfer, the recipient will remain F-. Controlled conjugation experiments can be used for gene mapping. Indeed, this approach was used to show the circularity of the E. coli chromosome and to determine the location of 1900 of its genes. Transformation, transduction and conjugation are means of horizontal gene transfer in nature (see Bacterial Gene Swapping in Nature by RV Miller. Scientific American 1998 (January), pp.47-51).
SEE ALSO:
Microbial Genetics (Ch. 9 of Brock Biology of Microorganisms, Prentice-Hall, Inc.)
Online Microbiology Book: Chapter 9-Genetic Regulatory Mechanisms in Bacteria
The Biology Project: Molecular Genetics of Prokaryotes
Scitable: Simple Viral and Bacterial Genomes
M.Tevfik Dorak, M.D., Ph.D.
Last updated on Aug 16, 2003
Last edited on July 14, 2013
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