Jayasri Basak

DNA damage and prophage induction and toxicity of nitrofurantoin in Escherichia coli and Vibrio cholerae cells

Repair-deficient and repair-proficient strains of E. coli K12 were sensitive to nitrofurantoin treatment to varying degrees with the double mutant strain (uvrA 6, recA 13) being most sensitive. Ultraviolet absorption data and thermal chromatography through a hydroxyapatite column revealed that nitrofurantoin treatment of V. cholerae strain OGAWA 154 produced a maximal amount of 55% reversibly bihelical DNA at a nitrofurantoin dose of 120 micrograms/ml/h, which indicated the formation of inter-strand cross-links in DNA. Nitrofurantoin also produced prophage-lambda induction in E. coli K12 strain GY 5027: envA, uvrB, ampA 1, strA (lambda), in a dose-dependent manner, the maximum induction being highly significant (P less than 0.001). Previously published mutation data coupled with the prophage induction data presented here suggest that the genotoxic properties of nitrofurantoin are mediated through the SOS pathway.

Inter-strand cross-linking of Vibrio cholerae DNA induced by furazolidone: a quantitative assay by four simple methods

Four simple methods, i.e., (i) UV absorption spectrophotometry, (ii) hydroxyapatite chromatography, (iii) fluorescence analysis of ethidium bromide bound to DNA and (iv) assay of S1 endonuclease action, were used in parallel for the estimation of furazolidone-induced inter-strand cross-links in Vibrio cholerae DNA. The data produced by the four methods were in reasonable agreement with each other and provided similar linear dose-response relations, the correlation (between dose and response) coefficient being in any case numerically greater than 0.98. When the data obtained by four independent methods were plotted in a single graph, the resulting dose-response relation could be described by the equation log NR = 1.41 - 0.54 log D, where NR is the % non-reversible DNA remaining in the cells treated by furazolidone at dose D micrograms/ml x h. The correlation coefficient in this plot was -0.98 and significant to a level better than 0.1%. This study thus brings out that any one of these four methods can be used with reasonable confidence for the diagnosis and assay of inter-strand cross-links in DNA.

Characterization of the adaptive response to ionizing radiation induced by low doses of X-rays to Vibrio cholerae cells

Pretreatment with sublethal doses of X-rays induced an adaptive response in Vibrio cholerae cells as indicated by their greater resistance to the subsequent challenging doses of X-irradiation. The adaptive response was maximum following a pre-exposure dose of 1.7 Gy X-rays and an optimum incubation period of 40 min at 37 degrees C. Pre-exposure to a sublethal dose of 1.7 Gy X-rays made the Vibrio cholerae cells 3.38-fold more resistant to the subsequent challenge by X-rays. Pretreatment with a sublethal dose of hydrogen peroxide offered a similar degree of protection to the bacterial cells against subsequent treatment with challenging doses of X-ray radiation. However, exposure of Vibrio cholerae cells to mild heat (42 degrees C for 10 min) before X-ray irradiation decreased their survival following X-irradiation.

Induction of adaptive response by nitrofurantoin against oxidative DNA damage in some bacterial cells

Pretreatment with a sublethal dose of nitrofurantoin did not give any protection to Vibrio cholerae OGAWA 154 (wild-type) cells against subsequent treatment with challenging doses of MNNG and vice versa. However, pretreatment with a sublethal dose of nitrofurantoin offered significant protection to the bacterial cells against subsequent treatment with challenging doses of H2O2 and vice versa. Further, sublethal doses of nitrofurantoin or H2O2 produced almost the same degree of protection against challenges by H2O2 or nitrofurantoin. Both the alkylating agent MNNG and the oxidative agent H2O2 induced adaptive responses in Vibrio cholerae OGAWA 154 cells against subsequent challenge by the respective agents. The experiments presented in this communication revealed that nitrofurantoin produced an adaptive response in bacterial cells against oxidative and not alkylating DNA damage.

Furazolidone-induced interstrand cross-links in Vibrio cholerae DNA. Study of conformational change by circular dichroism

Vibrio cholerae DNA bearing furazolidone-induced interstand cross-links show a change in the characteristic circular dichroism spectra of the DNA itself in dilute buffer. The change in c.d. spectra was characterized by a shift of the positive band around 272 nm to lower wavelength and a loss of ellipticity of the negative band around 242 nm, and is similar to that exhibited by mitomycin linked Vibrio cholerae DNA under identical conditions and is suggestive of a conformational change of DNA bearing such cross-links. Both furazolidone-induced and mitomycin-induced cross-linking of Vibrio cholerae DNA inhibited the salt-induced conformation change, i.e. increase in winding angle of DNA, the percentage inhibition being greater for mitomycin-linked DNA.