One important concept in microbiology is attenuation. Attenuation is a form of adaptation which indicates a loss in the disease ability of a specific organism. When an organism whose virulence is decreased, it is said to be attenuated. A highly pathogenic organism may be rendered temporarily or permanently nonpathogenic if repeatedly subcultured on artificial laboratory media. The concept of attenuation can be further studied under the microscope For instance, by cultivation of a strain of bovine tubercle bacilli on media containing bile until the strain had lost its ability to cause disease, a suitable preparation (BCG vaccine) was developed for vaccination against tuberculosis. Although artificially eliminated, virulence may often be restored by animal passage, through the serial injection of microorganism into and their recovery from susceptible animals. This procedure is best observed with the aid of the microscope.
Genetic factors
Some internal factors operative in bacterial variations pertain to changes in the genetic apparatus of bacteria.
MUTATIONS
Mutations occur in higher forms of life. It is also possible in bacteria. Since the specific intracellular enzyme (protein) is regulated biochemically by a specific gene positioned on the chromosome of the bacterial cell, the related structure and function of the cell must depend on the integrity of that gene. There is a significant component within the gene which is the sequence of the nucleotide bases, with any change in this pattern projecting an effect on the cell. The sudden changes often constitute a mutation. Mutations are inheritable. The mutant is the individual bacterial cell exhibiting the mutation. The appearance of the mutant cell can be observed and examined well using the microscope.
The rearrangement of the nucleotide sequence of a gene is the outcome from an error in replication or from a breakage of the sugar-phosphate backbone of the DNA molecule. Mutations occur spontaneously. They are induced by certain mutagenic agents that can alter the nucleotide bases in such a way as to promote replication errors. The most effective mutagens are certain alkylating agents and forms of radiant energy.
INTERMICROBIAL TRANSFER
As observed under the microscope variations resulting from the passage of nuclear DNA from one bacterium to another are easily recognized in alterations of specific properties. If two bacterial strains which are variants of a single species are incubated together under special circumstances such a transfer is borne out by the appearance in the culture of new organisms displaying qualities of both original strains. The best classic example of the antimicrobial transfer of genetic material is the experiment performed by the famous British pathologist named Frederick Griffith in 1928. In his experiment Griffith injected a mixed culture of nonencapsulated or avirulent and heatkilled, encapsulated or virulent before heat-killed pneumococci into mice. The mice are laboratory animals highly susceptible to pneumococci. The following day, Griffith examined the blood of the sick mice, under the microscope. Griffith discovered a fully encapsulated type of pneumococcus in the blood. Since the encapsulated bacteria used for injection had been killed, the origin of the encapsulated forms could only be explained by assuming that some genetic material from the killed organisms was able to enter the living nonencapsulated cells and change or transform them into living and encapsulated ones. It was discovered later that the substance transferring the genetic capability from one bacterium to another resides in DNA alone.
When genetic material known as nuclear DNA is transferred from one cell to another, the receiving cell usually does not gain the full complement of chromosome but only a portion. This has been observed well under the microscope. Immediately after the event of transfer, this portion must be matched to the corresponding segments of the chromosome in the cell, and genetic material is exchanged and eliminated. Newly formed chromosome emerges from this arrangement, containing DNA from two bacterial cells. This is the known as the recombinant chromosome It is shaped by the process of recombination. Recombination can be defined therefore as the acquisition by the chromosome, as a genetic linkage group, of information from another such unit.
Transformation is defined as a process of direct transfer of nucleic acid from one bacterial cell which is the donor to another recipient. Certain species of bacteria release DNA into the medium in which they are growing, such as certain species of Neisseria that release it in their extracellular slime. A small number of bacteria in a given population can pick up this DNA as observed keenly under the microscope. If DNA is extracted from a donor chromosome, it fragments into several hundred pieces. Of these, only some 10 can be taken up by the recipient, about 2% to 5% of the total amount. Only double-stranded DNA participates in the process of transformation, but within the recipient bacterium, one strand is degraded. The other strand is permanently integrated with that cell’s chromosome, the net effect of which is the replacement of a short region of the chromosome with a new portion of genetic material. The large molecules of DNA can enter a cell only during certain transient periods of growth during which time the recipient cells can be changed. This period is indicated as competence, and the bacteria in a given population receptive to change as competent or transformable.
Transduction on the other hand is a special form of indirect transfer of genetic material. A bacterial virus or bacteriophage carries a small fragment of the chromosome from the bacterial cell in which it was produced to the bacterium it invades. Transduction occurs in coliforms, enteric pathogens, and staphylococci. These microorganisms can best be viewed using the microscope.
Conjugation is known to be the process effecting passage of genetic material from one bacterial cell to another on a transient physical contact. As seen under the electron micrograph two individual bacteria are seen to unite by means of the sex pilus, a cytoplasmic bridge between them. Even male and female mating types have been defined for the closely related groups in the family Enterobacteriaceae that have been studied. The difference in the appearance of the male and female can be observed carefully with the aid of the microscope. Conjugation can occur between any two members of the family, not just within a given genus. The process is not as its designation suggests a method of reproduction but a provision for the oneway passage of genetic material. No fusion of cells or new cell results as a consequence. Progeny are produced in the usual way by binary fission of the parent cell. Probably the least common method of genetic exchange in bacteria, conjugation differs from both transformation and transduction in that, unlike them, it results in the transfer of very large segments, sometimes even the entire chromosome.
Extrachromosomal factors such as plasmids and episomes, are widely seen among bacteria and not essential to viability. Circular molecules of DNA 1% to 3% of the weight of the bacterial chromosome they determine bacterial traits crucial to adaptation and may carry their own genes for replication and transfer. Plasmids are independent in the bacterial cell cytoplasm. Episomes may be independent or at times be integrated into the bacterial chromosome. The shapes form and appearance of the plasmids and episomes can be described well with the aid of the microscope.
Drug-fastness
A microbial variation with considerable therapeutic importance is one that arises through a mutation or other genetic mechanism giving the organism an increased tolerance for an antimicrobial drug. This tolerance for the drug is termed drug resistance or drug fastness. An. organism that becomes resistant to a drug used in the treatment of disease is said to be drug-fast. Strains of bacteria are always emerging resistant to one or more antibiotics. This resistance usually acquired when clinical infectious disease has been inadequately treated with the given antibiotic. Bacteria may become tolerant of more than one antimicrobial drug. This cross-resistance is noted especially in the case of closely related antibiotics. Under the microscope, one can compare clearly the difference between the drug resistant virus and from one which is not.