Mutagenicity Testing and In Vitro Tests

A variety of mutagenicity tests has been developed. They can be categorized as tests that detect gene mutations, chromosome effects, DNA repair and recombination, or others. The others include tests for transformation of mammalian cells into malignant cancer cells in vitro.

Gene mutation tests detect mutations directly. They may be conducted in vitro or in vivo, and with prokaryotic or eukaryotic organisms. One of the best known is the Ames test. This test uses a strain of Salmonella typhimurium that has been mutated artificially so that it cannot synthesize the amino acid histidine. In the test, this organism is cultured in a histidine-deficient medium with the chemical being tested. Mutagens cause a reverse mutation, again allowing the organism to produce its own histidine. The presence of growth in the histidine-deficient medium thus serves to indicate the occurrence of a mutation. Since many mutagens require bioactivation, microsomes extracted from animal livers are added to the medium. These supply active cytochrome enzyme systems. Similar tests have been developed using eukaryotic yeast cell cultures and mammalian cell cultures.

(The developer of this test, Bruce Ames, is an outspoken critic of the way that tests are used to identify carcinogens. He states: "The effort to eliminate synthetic pesticides because of unsubstantiated fears about residues in food will make fruits and vegetables more expensive, decrease consumption, and thus increase cancer rates. The levels of synthetic pesticide residues are trivial in comparison to natural chemicals, and thus their potential for cancer causation is extremely low.'')

A type of in vivo gene mutation test is the host-mediated assay. In this test, cells that are susceptible to reverse mutation (whether bacterial or eukaryotic) are injected with the toxicant into a live animal, such as a mouse. After several hours the animal is sacrificed and the injected cells are cultured to detect mutations. This has the advantage of using all the biotransformation mechanisms of the living animal.

Mutations can also be detected directly in test animals. The most popular is the fruit fly Drosphila melanogaster, but tests have also been developed to detect mutations in mice.

Chromosomal effects include structural aberrations such as deletion or duplication of chromosomes, or translocation, involving switching of sections between different chromosomes. These are conducted either in vitro using mammalian cell cultures or in vitro with Drosophila sp. or mammals. In either case the effect is detected by microscopic examination a period of time after dosing the system with the toxicant.

The third type of mutagenicity test involve the DNA repair mechanisms to detect mutations. Some strains of Escherichia coli and Bacillus subtilis are deficient in repair enzymes. Their growth will be inhibited relative to normal strains when mutations occur, and this inhibition can be detected by an assay. In another test of this type, DNA synthesis stimulated by mutations can be detected in human cell culture by the uptake of radioactive thymidine.

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