DNA analysis of a radiotolerant bacterium Pantoea agglomerans by FT-IR spectroscopy

A radiation tolerance strain, Pantoea agglomerans was isolated from γ-irradiated carrot samples (Daucus carota). D₁₀ determination showed that the radioresistance of this bacterium is five-fold higher than Escherichia coli, both belonging to the family of Enterobacteriaceae. DNA isolated from untreated and irradiated bacterial cells was analyzed by FT-IR spectroscopy to investigate the radiotolerance of this bacterium. At doses <5 kGy, an alteration of the interbase hydrogen networks was observed and characterized mainly by an increase of bands assigned to the carbonyl non-pairing and the free amine groups. Moderate breakage of the DNA backbone and damage of the osidic structure were also observed. Similar spectral profiles were noticed at doses ≥5 kGy, but additional increase of the band intensity of CC and CN suggests damages of nucleobases. High number of asymmetric PO₂⁻ and upper shift of symmetric PO₂⁻ are indicative of DNA strand breaks. Osidic damages were evidenced by decrease of the absorption bands ascribed to deoxyribosyl moieties and by appearance of C–OH band. DNA degradation at high irradiation doses was also noticed by electrophoresis using agarose gel. It appeared that DNA underwent covalent cross-linking, as revealed by agglomeration of DNA in the wells of agarose gel.     

PROTECTION OF ESCHERICHIA COLI CELLS AGAINST THE LETHAL EFFECTS OF ULTRAVIOLET AND X IRRADIATION BY PRIOR X IRRADIATION: A GENETIC AND PHYSIOLOGICAL STUDY

Abstract— When log phase cells of wild-type E. coli K-12 were maintained in growth medium after X irradiation, they became progressively more resistant to a subsequent exposure to UV or X radiation. The time to achieve maximum resistance was about 60 min. The uvrB, uvrD, polA and certain exrA strains (W3110 background) also demonstrated this X ray-induced resistance to subsequent UV or X irradiation but recA, recB, lex (AB1157 or W3110 backgrounds) and other exrA strains (AB1157 background) did not. The resistance induced in wild-type, uvrB and uvrD cells was characterized by the production or enhancement of a shoulder on the survival curves obtained for the second irradiation, while the resistance induced in the W3110 exrA strains was expressed only as a change in slope. The induction of resistance in the W3110 exrA strain was not inhibited by the presence of chloramphenicol, but that in the wild-type cells appeared to be. The production or enhancement of a shoulder on the survival curves of the rec ⁺ lex ⁺ exr ⁺ cells is consistent with the concept of the radiation induction of repair enzymes. Alternative explanations, however, are discussed.     

Induction of phr gene expression in E. coli strain KY706/pPL-1 by He-Ne laser (632.8 nm) irradiation

We have observed that He–Ne laser irradiation of E. coli strain KY706/pPL-1 leads to induction of photolyase gene, phr. The magnitude of induction was found to depend on the He–Ne laser fluence, fluence rate and post-irradiation incubation period in the nutrient medium. The optimum values for fluence and fluence rate were 7×10³ J/m² and 100 W/m², respectively, and the induction of phr gene was observed to saturate beyond an incubation period of 2 h. Experiments carried out with singlet oxygen quenchers and with D₂O suggest that the effect is mediated via singlet oxygen. Photoreactivation studies carried out after UVC exposure of both the He–Ne laser-exposed as well as unexposed cells showed a larger surviving fraction in the He–Ne laser pre-irradiated cells. This can be attributed to He–Ne laser irradiation-induced induction of phr expression. However, since even without photoreactivating light He–Ne laser pre-irradiated cells show higher survival against UVC radiation it appears that He–Ne laser irradiation induces both light-dependent as well as dark DNA repair processes.     

NON-NUCLEAR DAMAGE AND CELL LYSIS ARE INDUCED BY UVA, BUT NOT UVB OR UVC, RADIATION IN THREE STRAINS OF L5178Y CELLS

The potential to induce non-nuclear changes in mammalian cells has been examined for (1) UVA1 radiation (340–400 nm, UVASUN 2000 lamp), (2) UVA + UVB (peak at 313 nm) radiation (FS20 lamp), and (3) UVC (254 nm) radiation (GI5T8 lamp). The effects of irradiation were monitored in vitro using three strains of L5178Y (LY) mouse lymphoma cells that markedly differ in sensitivity to UV radiation. Comparisons were made for the effects of approximately equitoxic fluences that reduced cell survival to 1–15%. Depending on the cell strain, the fluences ranged from 830 to 1600 kJ/m² for the UVASUN lamp, 75 to 390 J/m² for the FS20 lamp and 3.8 to 17.2 J/m² for the G15T8 lamp. At the exposure level used in this study, irradiation with the UVASUN, but not the FS20 or G15T8, lamp induced a variety of non-nuclear changes including damage to cytoplasmic organelles and increased plasma membrane permeability and cell lysis. Cell lysis and membrane permeabilization were induced by the UVA1 emission of the UVASUN lamp, but not by its visible + IR components (>400 nm). The results show that the plasma membrane and other organelles of LY cells are highly sensitive to UVA1 but not to UVB or UVC radiation. Also UVA1, but not UVB or UVC radiation, causes rapid and extensive lysis of LY cells. In conclusion, non-nuclear damage contributes substantially to UVA cytotoxicity in all three strains of LY cells.     

Effect of dose rate on inactivation of microorganisms in spices by electron-beams and gamma-rays irradiation

Total aerobic bacteria in spices used in this study were determined to be 1 × 10⁶ to 6 × 10⁷ per gram. A study on the inactivation of microorganisms in spices showed that doses of 6–9kGy of EB (electron-beams) or γ-irradiation were required to reduce the total aerobic bacteria in many However, a little increase of resistance was observed on the inactivation of total aerobic bacteria in many spices in case of EB irradiation. These difference of radiation sensitivities between EB and γ-rays was explained by dose rate effect on oxidation damage to microorganisms from the results of radiation sensitivities of Bacillus pumilus and B. megaterium spores at dry conditions. On the other hand, these high dose rate of EB irradiation suppressed the increase of peroxide values in spices at high dose irradiation up to 80 kGy. However, components of essential oils in spices were not changed even irradiated up to 50 kGy with EB and γ-rays.     

OXYGEN EFFECT IN SURVIVAL OF Dictyostelium discoideum TREATED WITH NEAR ULTRAVIOLET RADIATION

Abstract— Fluence-response survival curves have been measured for the cellular slime mold Dictyostelium discoideum exposed to near ultraviolet radiation. Data were obtained for a wild type strain and three UV-sensitive mutant strains in exponential growth phase. Fluences for 10% survival (F₁₀) are about 1 MJ m⁻² for cells irradiated in saline solution saturated with nitrogen. When air is bubbled through the saline, the Fm values are only one third as large. Strain HPS50, which is the strain most sensitive to gamma radiation and to 254 nm UV, also exhibits the greatest sensitivity to near UV. However, the difference in sensitivity to near UV between wild type and mutant strains is small compared to other physical and chemical agents known to damage DNA.     

POSTREPLICATION REPAIR IN uvrA AND uvrB STRAINS OF ESCHERICHIA COLI K-12 IS INHIBITED BY RICH GROWTH MEDIUM

Abstract Escherichia coli K-12 uvrA or uvrB strains grown to logarithmic phase in minimal medium showed higher survival after ultraviolet (UV) irradiation (254 nm) if plated on minimal medium (MM) instead of rich medium. This'minimal medium recovery'(MMR) was largely blocked by additional recA56 (92% inhibition) or lexA101 (77%) mutations, was partially blocked by additional recB21 (54%), uvrD3 (31%) or recF143 (22%) mutations, but additional polA1 or polA5 mutations had no effect on MMR. When incubated in MM after UV irradiation, the uvrB5 and uvrB5 uvrD3 strains showed essentially complete repair of DNA daughter-strand gaps (DSG) produced after UV radiation fluences up to ∼ 6 J/m² and ∼1 J/m², respectively, and then they accumulated unrepaired DSG as a linear function of UV radiation fluence. However, when they were incubated in rich growth medium after UV irradiation, they did not show the complete repair of DSG and unrepaired DSG accumulated as a linear function of UV radiation fluence. The fluence-dependent correlation observed for the uvrB and uvrB uvrD cells between UV radiation-induced killing and the accumulation of unrepaired DSG, indicates that the molecular basis of MMR is the partial inhibition of postreplication repair by rich growth medium. Rich growth medium can be just MM plus Casamino Acids or the 13 pure amino acids therein in order to have an adverse effect on survival, regardless of whether the cells were grown in rich medium or not before UV irradiation.     

Stretched exponential survival statistics for microorganisms in radiation field

The so-called concave or tailed survival curves are reported both for multi and single species bacterial populations. Taking as an example Bacillus pumilus, frequently encountered in decontamination studies, it is shown that the tailed survival curves are adequately described by stretched (0<<1) exponential SF=exp[−(D/D₀)], where SF denotes the fraction of species surviving the irradiation dose D, D₀ is the effective dose, and is the dispersion parameter interpreted phenomenologically in terms of radiation sensivity distribution for single species population under the given experimental conditions.     

Structure and synthesis of mexolide: a new antibiotic dicoumarin from murraya exotica linn. [syn. m. paniculata (l) jack.]

In continuation of our studies on Murraya exotica Linn. (Syn. M. paniculata) from which we reported mexoticin¹ (1), hibiscetin heptamethyl ether,² carbazole alkaloids, murrayazoline, mahanimbine and girinimbine,³ we now report the structure of a new antibiotic dimeric coumarin named mexolide (2) having an unusual cyclohexene system similar to that in cyclobissuberodiene⁴ [thamnosin⁵ (3)].     

EFFECTS OF 8-METHOXYPSORALEN AND NEAR ULTRAVIOLET RADIATION ON THE SURVIVAL OF THE LOWER EUKARYOTE D. Discoideum

Abstract— Seven axenic wild-type and repair-deficient mutant strains of the cellular slime mold Dictyostelium discoideum have been treated with the furocoumarin 8-methoxypsoralen (8-MOP) up to 50 μg/mζ and then exposed to near ultraviolet light (UVA 320-400 nm) up to 21 kJ/m². Fluence-response survival curves exhibit shoulders at lower fluences and an exponential lethal response at higher fluences. Neither the psoralen alone nor the irradiation alone produced any measurable lethal effect. Wild-type strains, which show resistance to 254 nm UV and gamma radiation, also show resistance to psoralen plus UVA. The moderate sensitivity of a rad D repair-deficient mutant strain and the extreme sensitivity of a rad B mutant strain to 8-MOP plus UVA parallel their responses to UV and gamma radiation. However a rad C mutant which is sensitive to UV, exhibits wild-type response to photoactivated psoralen.     

Comparison of the effects of visible femtosecond laser pulses and continuous wave laser radiation of low average intensity on the clonogenicity of Escherichia coli.

To evaluate the contribution of local pulsed heating of light-absorbing microregions to biochemical activity, irradiation of Escherichia coli was carried out using femtosecond laser pulses (λ = 620 nm, τ_p=3 × 10⁻¹³ s, f_p = 0.5 Hz, E_p = 1.1 × 10⁻³J cm⁻², I_av = 5.5 × 10⁻⁴ W cm⁻², I_p = 10⁹ W cm⁻²) and continuous wave (CW) laser radiation (λ = 632.8 nm, I = 1.3 W cm⁻²). The irradiation dose required to produce a similar biological effect (a 160%–190% increase in the clonogenic activity of the irradiated cells compared with the non-irradiated controls) is a factor of about 10³ lower for pulsed radiation than for CW radiation (3.3 × 10⁻¹ and 7.8 × 10² J cm⁻² respectively). The minimum size of the microregions transiently heated on irradiation with femtosecond laser pulses is estimated to be about 10 Å, which corresponds to the size of the chromophores of hypothetical primary photoacceptors—respiratory chain components.     

REC A +-DEPENDENT SYNERGISM BETWEEN 365 NM AND IONIZING RADIATION IN LOG-PHASE ESCHERICHIA COLI: A MODEL FOR OXYGEN-DEPENDENT NEAR-UV INACTIVATION BY DISRUPTION OF DNA REPAIR

Abstract —The oxygen dependence of 365 nm inactivation of colony-forming ability of Escherichia coli has been investigated in two series of DNA repair-deficient K12 mutants grown to mid-exponential phase. All strains except a uvr A rec A double mutant are more sensitive to inactivation under O₂ and show a lower threshold dose. The inactivation of photoreactivating enzyme in a crude cell extract and DNA repair disruption are both reduced when irradiation is carried out under nitrogen. The rec A gene-dependent synergism between 365 nm and ionising radiation is reversible if cells are incubated in full growth medium before ionising radiation treatment. In a wildtype strain, incubation for 2.5 h in full growth medium after 10⁶ J m^-2 365 nm radiation changes a sensitised response to a protection from ionising radiation. Protection is not seen at 1.5 times 10⁶ J m^-2. A tentative model for near UV lethality in logarithmic phase cells is suggested which proposes two classes of lesions. One requires oxygen for it's induction, is rapidly fixed as a lethal event as a result of repair disruption, and is primarily responsible for cell death after aerobic 365 nm irradiation. The other lesion, possibly pyrimidine dimers, may lead to cell death under anaerobic conditions.     

Targeted Synthesis of Anthranilic Diamides Insecticides Containing Trifluoroethoxyl Phenylpyrazole

Aseries of novel anthranilic diamides analogues(9a-9t) containing trifluoroethoxyl pyrazole moiety was designed and synthesized and their insecticidal bioactivities against Mythimna separata(Walker, M. separata) and Plutellaxylostella (P. xylostella) were evaluated. The structures of the title compounds were confirmed by ¹H NMR, ¹³C NMR and HRMS. Preliminary insecticidal activities showed that some of the title compounds possessed good to excellent bioactivities towards M. separata and P. xylostella. Compounds 9c and 9t exhibited 100% mortality rate against M. separate at 0.2 mg/L. For the P. xylostella, the synthesized compounds(9c-9e, 9i and 9o) showed 70%, 80%, 75%, 65% and 60% insecticidal activities at 1×10^-6 mg/L, respectively, higher than that of chlorantraniliprole(0). Based on excellent insecticidal activities, the mode of the action was tested by the calcium-imaging technique, the results of which demonstrated that the novel compounds shared the same target with chlorantraniliprole. The binding pose of the most active compound 9t in RyRs of P. xylostella was predicted by molecular docking, which showed that compound 9t interacted with the residues Glu140(A) and His147(A) via hydrogen bond.     

EFFECTS OF DOSE FRACTIONATION ON ULTRAVIOLET SURVIVAL OF ESCHERICHIA COLI

Abstract— Exposure of E. coli B/r and B at low average dose rates of u.v. radiation (2537 Å), produced either by fractionated doses or by continuous irradiation at a very low dose rate (80 ergs/mm²/hr), results in much increased survival compared to single exposure at high dose rate. This increase is attributed to repair taking place during the irradiation period. The effect is small in the repair-deficient strains E. coli B_8-1_, and C syn- , and is absent in phage T1 and T4, which cannot undergo repair in the extracellular state. However, the prolonged time available for repair in these experiments accounts for only a very minor part of the increase in survival. The principal factor apparently is that the number of lesions present at any time remains relatively low. Presumably complete repair, not only the excision step, can occur in buffer during the irradiation period. This interpretation is supported by experiments in which cells were exposed to combinations of highly fractionated irradiation and single-dose irradiation. We therefore propose that mutual interference in repair, possibly by overlapping of repair regions in complementary DNA strands, reduces considerably the repair efficiency if many lesions are present. This hypothesis explains the 'shouldered' survival curves of B/r and possibly other E. coli strains as due to decreasing repair efficiency with increasing u.v. dose     

Effects of solar and artificial UV irradiation on pigments and assimilation of N ammonium and N nitrate by macroalgae

Three macroalgal species, Fucus vesiculosus, Laminaria saccharina and Ulva lactuca were exposed to natural daylight in UV-transparent and UV-non-transparent Plexiglas vessels at different water depths. Uptake of ¹⁵NH₄⁺ and ¹⁵NO₃⁻ was measured at different light intensities. In general, uptake rates in August surpassed those in May 1987 and Ulva assimilated significantly more nitrogen than Fucus and Laminaria. A negative influence of ambient solar UV radiation on nitrogen uptake was dound in Fucus and Ulva. Additionally, the impact of UV of different wave bands, using cut-off filters (WG 295, WG 305 and WG 320) and special UV-A and UV-B lamps, on pigments and ¹⁵N ammonium assimilation by several macroalgae was studied under controlled laboratory conditions. UV-B irradiation of shorter wave bands led to a reduction in the main pigments of Fucus except antheraxanthin and those of Ulva except lutein and violaxanthin. Uptake of ¹⁵N-ammonium by Fucus and Ulva was reduced after UV-B radiation whereas an increase was observed after UV-A irradiance in Ulva. Ammonia utilization by Halidrys was more damaged by UV-A than UV-B. The impact of UV radiation on the labelling of free amino acids and the pattern of pool sizes varied with the species and the UV wave bands. A marked reduction in ¹⁵N incorporation into the amino acids of Plocamium was found after UV-B exposure. The ¹⁵N labelling of the amino acids of Halidrys and Rhodomela was less affected. Results are discussed with reference to photoinhibitory effects on the enzymes of the nitrogen metabolism.     

Effect of γ irradiation on optical properties of Ce-doped phosphate and silicate scintillating glasses

A set of Ce³⁺ activated silicate- and phosphate-based scintillating glasses were submitted to γ irradiation in the ⁶⁰Co radioisotope source “Calliope” (ENEA-Casaccia in Rome, Italy) in the dose range between 1 and 250 Gy (3.7 Gy/h). The effect of ionising radiation was probed by transmission measurements performed before and after each irradiation on all analysed samples. From these data, the radiation-induced absorption coefficient was calculated, proportional to the density of colour centres induced by irradiation in the solid matrix. Results are discussed by taking into account the possible dependence of radiation hardness on the composition of glass matrix.     

Nitrite formation in the radiolysis of aerated aqueous solutions of ammonia

Gamma irradiation of aerated aqueous solutions of ammonia leads to the formation of nitrite as a radiolytic product. Its yield increases with increasing concentration of NH₃ as well as O₂. OH radicals react with NH₃ to give NH₂ radicals, which in the presence of O₂ form NH₂O₂ radicals. These radicals finally lead to the formation of nitrite. G(NO₂⁻) decreases with increasing radiation dose due to secondary reactions. Its initial yield, however, is more than 1/2G_OH, while hydrogen peroxide yield is less than the expected value viz. G(H₂O₂) + 1/2G(e_aq⁻), indicating its participation in reactions with radiation produced free radicals. G(H₂) is 0.35 in aerated aqueous solutions of 10⁻³ mol dm⁻³ NH₃ and 0.23 in the absence of oxygen. Implications of these results to the use of NH₃ in primary coolant water of pressurized water nuclear power reactors of the VVER type are discussed.     

Optical microscopy and deuterium N.M.R. of non-ionic amphiphiles which exhibit thermotropic and aqueous lyotropic mesophases

Tetrabenzocyclododecatetraene (I) substituted with eight methoxy-mono-, di-, and triethyleneoxide side chains exhibit the same highly ordered thermotropic mesophase, M. In the methoxydiethyleneoxide (I-2), and methoxytriethyleneoxide (I-3) derivatives this mesophase is also lyotropic and can sustain (at room temperature) up to 40 wt per cent water. The homologue (I-2) exhibits at higher water content another lyomesophase, M_F, which is more fluid and nematic-like. Optical microscopy, differential scanning calorimetry and deuterium N.M.R. of both labelled (I-2) and water as well as of the water ¹⁷O were used to construct the (I-2)-water phase diagram and to characterize the various mesophases. It is found that both M and M_F are uniaxial with negative birefringence and negative anisotropic magnetic susceptibility. The M_F phase readily aligns in a magnetic field and is characterized by fast (∼ 10⁷ s^-1) reorientation of the mesogen molecules about the director. The M phase is apparently columnar, highly ordered and is not readily aligned by a magnetic field. The rate of molecular reorientation in this mesophase is much slower (∼ 10³ s^-1) compared with M_F.     

Biological effects of helium-neon (HeNe) laser irradiation on acrosome reaction in bull sperm cells

The purpose of this study was to determine the effect of He---Ne laser irradiation (632.8 nm, 10 mW) on the induction of acrosome reaction and mortality in bull sperm cells in comparison with two important capacitation agents; calcium and heparin. Frozen-thawed bull sperm cells were washed in percoll gradient and suspended at a concentration of 1 × 10⁶ ml⁻¹ in sp-TALP medium, capacitated in the presence of 2 mM CaCl₂, 10 μg ml⁻¹ heparin, or irradiated at fluences from 2 to 16 J cm⁻², and incubated for 0, 30, 60 and 90 minutes. At the end of the incubation period, the percentage of sperm that were acrosome-reacted and dead was determined. The results obtained indicated that laser irradiation at all fluences produced a significant increase (p < 0.001) in the percentage of sperm cells that were acrosome reacted, and a significant decrease (p < 0.001) in the percentage of dead sperm at 90 minutes of incubation in comparison to other capacitation agents and the control group. The percentage of sperm cells with acrosome reaction was increased with increasing fluences of laser irradiation and time of incubation. It is conclude that the application of He---Ne laser irradiation at fluences from 2 to 16 J cm⁻² induced the acrosome reaction and decreased the sperm mortality percentage in vitro of bull sperm cells.