Irradiated Bacteria Research Articles

Gamma Irradiated Pasteurella multocida Vaccine induces strong humoral immunity and protects rabbits from disease

Pasteurella multocida is affecting a multitude of animals and severely affects livestock production. Existing vaccines are mostly chemically inactivated and do not lead to wide protection. Irradiated vaccines are enjoying a renaissance and the concept of “replication defficient but metabolically active” vaccines was recently evaluated in several vaccine trials. P. multocida was isolated from the nasal swab, blood, and lung swab samples from infected rabbits. Gamma irradiation of P. multocida for inhibition of replication was evaluated at an optimized irradiation dose of 10 Kgy established. Four groups of rabbits were (mock) vaccinated with a commercial P. multocida vaccine and three irradiated formulations as liquid, lyophilized formulations with added Trehalose and lyophilized-Trehalose with an “activation” culturing the irradiated bacteria for 24 in broth. Evaluation of humoral immune response by ELISA showed that all three irradiated vaccines produced an effective, protective, and continued IgG serum level after vaccination and bacterial challenge. The IFN-γ expression is maintained at a normal level, within each individual group however, the lyophilized trehalose irradiated vaccine showed peak mean of IFN-γ titer at one week after booster dose (day 21) which was statistically significant. Cumulatively, the results of this study show that gamma-irradiated P. multocida vaccines are safe and protect rabbits against disease. Moreover, Rabbits’ immunization with the three irradiated formulations avoided adverse side effects as compared to commercial polyvalent vaccine, the body weight gain for the irradiated vaccine groups indicates less stress compared to the commercial polyvalent vaccine.

TMT-based quantitative proteomic and scanning electron microscopy reveals biological and morphological changes of Staphylococcus aureus irradiated by electron beam

Electron beam irradiation is an efficient cold sterilization technique that has been widely utilized in food preservation. To understand the biological and morphological changes of microorganisms by electron beam irradiation, the D10 value, growth curve, and morphological characterization by scanning electron microscope (SEM) of Staphylococcus aureus were studied. Furthermore, Tandem mass tag (TMT)-based quantitative proteomic was performed to investigate the responses of Staphylococcus aureus to electron beam. The results showed that electron beam prolonged the lag phase of bacterial growth, and irradiated bacteria were more sensitive to irradiation than unirradiated bacteria. Compared to unirradiated cells, there were 53, 96, 1055, and 431 differentially expressed proteins (DEPs) detected in 1-kGy and 2-kGy irradiated cells after 0 h and 24 h incubation, respectively. Most of the DEPs involved in DNA repair were found to be up-regulated, whereas those involved in biofilm formation and staphylococcal enterotoxin production were down-regulated. No noticeable morphological changes were observed in irradiated S. aureus. These findings provided valuable information in the responses of S. aureus to electron beam irradiation, leading to a better understanding of the effects of electron beam irradiation on food pathogens.

Evaluation of room temperature (30°C to 35°C) lyophilized vaccine with radio inactivated Mannheimia haemolytica whole cells isolated from infected sheep.

Background and Aim:Vaccines are one of the important tools for fighting diseases and limiting their spread. The development of vaccines with high efficacy against diseases is essential. Ionizing radiation is the method used for the preparation of the irradiated gamma Mannheimia haemolytica vaccine. The study aimed to measure the metabolic activity and electron microscopic examination of the irradiated bacterial cells and immunological efficiency of different preparations of the irradiated M. haemolytica vaccine.Materials and Methods:The irradiated vaccines were prepared in three forms at a dose of 2×109 colony-forming unit (CFU) (irradiated M. haemolytica, trehalose irradiated M. haemolytica, and trehalose lyophilized irradiated M. haemolytica). The formalin-killed vaccine was prepared at a dose of 2×109 CFU. Scanning electron microscopy was used to determine the difference between the non-irradiated bacterial cells and the bacterial cells exposed to gamma radiation. The metabolic activity of the irradiated bacterial cells was measured using the Alamar blue technique. Rabbits were divided into five groups (control, vaccinated groups with the formalin-killed vaccine, irradiated bacterial cells without trehalose, trehalose irradiated bacteria, and trehalose lyophilized irradiated bacterial cells). The rabbits were subcutaneously inoculated twice in 2-week intervals. Enzyme-linked immunosorbent assay, interferon-gamma (IFNγ), and interleukin 4 (IL4) assays were used to evaluate the vaccines’ immunological efficiency in rabbits.Results:The metabolic activity tests showed that the bacterial cells exposed to gamma radiation at the lowest lethal dose have metabolic activity. The difference in the metabolic activity between preparations of the irradiated bacterial cells varied according to the cell concentration and incubation time. The highest level of metabolic activity was 8 h after incubation in the nutrient broth medium compared with 4 and 18 h. The scanning electron microscopy of irradiated bacterial cells showed a cavity at the bacterial cell center without rupture of the surrounding cell membrane compared to the non-irradiated bacterial cells. The antibody level in the groups vaccinated with the different preparations of the irradiated bacterial cells was high compared with the control and formalin-killed vaccine groups. The level of the IFNγ showed an increase after the second dose in the group vaccinated with irradiated bacterial cells without trehalose compared with the other groups. The IL4 level in the vaccinated groups with the irradiated bacterial cells without trehalose, irradiated bacterial cells with trehalose, and trehalose lyophilized irradiated bacterial cells were at a high level when compared with the formalin-killed vaccinated group and control group after the second inoculation.Conclusion:The irradiated M. haemolytica vaccine provides a wide range of humoral and cellular immunity. This study showed high immunological efficiency in rabbits inoculated with the irradiated M. haemolytica vaccine that was shown in the high levels of antibodies (IFNγ and IL4) compared with the group treated with the formalin-killed vaccine. The second dose of irradiated M. haemolytica vaccine is an immune booster that gives the irradiated vaccine a long-acting immunological efficiency.

Short-Term Biodegradation of Crude Petroleum Oil in Water by Photostimulated Janibacter terrae Strain S1N1.

Biodegradation is a sustainable green strategy that gives the opportunity for remediation of water contaminated with petroleum products. In this study, 12 bacterial isolates were recovered from River Nile, Egypt and screened for their potential to degrade a mixture of paraffinic petroleum crude oil. The most promising isolate was identified according to 16S rRNA sequencing as Janibacter terrae strain S1N1 (GenBank accession No. KX570955.1). In order to boost the biodegradation efficiency, the bacterial suspension was photostimulated by exposure to different irradiation doses using a low-power helium–neon (He–Ne) laser (λ = 632.8 nm). Maximum biodegradation was achieved after 4 min of exposure (134.07 J cm–2) at optimized pH value (6) and temperature (35 °C). The gas chromatography–mass spectrometry (GC-MS) analysis revealed the biodegradation of 96.5% of the substrate after only 48 h of incubation. The n-C17/Pr and n-C18/Ph ratios indicated a preferential biodegradation of iso-paraffines over normal ones. Meanwhile, pristane/phytane (Pr/Ph) ratios were indicative of selective biodegradation for pristane. The carbon preference index (CPI) was nearly around unity indicating the ability of Janibacter terrae to attack the odd and even n-alkanes simultaneously. These results support the superiority of irradiated bacteria in optimizing the biodegradation efficiency and shortening the time of treatment, thus proposing an eco-friendly technique in water bioremediation programs.

Min Oscillations as Real-time Reporter of Sublethal Effects in Photodynamic Treatment of Bacteria.

The Min protein system is a cell division regulator in Escherichia coli. Under normal growth conditions, MinD is associated with the membrane and undergoes pole-to-pole oscillations. The period of these oscillations has been previously proposed as a reporter for the bacterial physiological state at the single-cell level and has been used to monitor the response to sublethal challenges from antibiotics, temperature, or mechanical fatigue. Using real-time single-cell fluorescence imaging, we explore here the effect of photodynamic treatment on MinD oscillations. Irradiation of bacteria in the presence of the photosensitizer methylene blue disrupts the MinD oscillation pattern depending on its concentration. In contrast to antibiotics, which slow down the oscillation, photodynamic treatment results in an abrupt interruption, reflecting divergent physiological mechanisms leading to bacterial death. We show that MinD oscillations are sensitive to mild photodynamic effects that are overlooked by traditional methods, expanding the toolbox for mechanistic studies in antimicrobial photodynamic therapy.

E. coli aggregation and impaired cell division after terahertz irradiation

In this study we demonstrated that exposure of Escherichia coli (E. coli) to terahertz (THz) radiation resulted in a change in the activities of the tdcABCDEFGR and matA–F genes (signs of cell aggregation), gene yjjQ (signs of suppression of cell motility), dicABCF, FtsZ, and minCDE genes (signs of suppression of cell division), sfmACDHF genes (signs of adhesin synthesis), yjbEFGH and gfcA genes (signs of cell envelope stabilization). Moreover, THz radiation induced E. coli csg operon genes of amyloid biosynthesis. Electron microscopy revealed that the irradiated bacteria underwent increased aggregation; 20% of them formed bundle-like structures consisting of two to four pili clumped together. This could be the result of changes in the adhesive properties of the pili. We also found aberrations in cell wall structure in the middle part of the bacterial cell; these aberrations impaired the cell at the initial stages of division and resulted in accumulation of long rod-like cells. Overall, THz radiation was shown to have adverse effects on bacterial populations resulting in cells with abnormal morphology.

Bacterial cell growth is arrested by violet and blue, but not yellow light excitation during fluorescence microscopy

BackgroundFluorescence microscopy is a powerful tool in cell biology, especially for the study of dynamic processes. Intensive irradiation of bacteria with UV, blue and violet light has been shown to be able to kill cells, but very little information is available on the effect of blue or violet light during live-cell imaging.ResultsWe show here that in the model bacterium Bacillus subtilis chromosome segregation and cell growth are rapidly halted by standard violet (405 nm) and blue light (CFP) (445–457 nm) excitation, whereas they are largely unaffected by green light (YFP). The stress sigma factor σB and the blue-light receptor YtvA are not involved in growth arrest. Using synchronized B. subtilis cells, we show that the use of blue light for fluorescence microscopy likely induces non-specific toxic effects, rather than a specific cell cycle arrest. Escherichia coli and Caulobacter crescentus cells also stop to grow after 15 one-second exposures to blue light (CFP), but continue growth when imaged under similar conditions in the YFP channel. In the case of E. coli, YFP excitation slows growth relative to white light excitation, whereas CFP excitation leads to cell death in a majority of cells. Thus, even mild violet/blue light excitation interferes with bacterial growth. Analyzing the dose-dependent effects of violet light in B. subtilis, we show that short exposures to low-intensity violet light allow for continued cell growth, while longer exposures do not.ConclusionsOur experiments show that care must be taken in the design of live-cell imaging experiments in that violet or blue excitation effects must be closely controlled during and after imaging. Violet excitation during sptPALM or other imaging studies involving photoactivation has a threshold, below which little effects can be seen, but above which a sharp transition into cell death occurs. YFP imaging proves to be better suited for time-lapse studies, especially when cell cycle or cell growth parameters are to be examined.

Effect of gamma irradiated Lactobacillus bacteria as an edible coating on enhancing the storage of tomato under cold storage conditions

This study was carried out during the two successive seasons of 2017 and 2018 to study the effect of both unirradiated and irradiated Lactobacillus paracasei cell-free supernatant in combination wi...

Natural outer membrane permeabilizers boost antibiotic action against irradiated resistant bacteria

BackgroundThis study sought to develop new strategies for reverting the resistance of pathogenic Gram-negative bacilli by a combination of conventional antibiotics, potent permeabilizers and natural beta lactamase inhibitors enhancing the activity of various antibiotics.MethodsThe antibiotic susceptibility in the presence of natural non-antibacterial tested concentrations of phytochemicals (permeabilizers and natural beta lactamase inhibitors) was performed by disk diffusion and susceptibility assays. Thymol and gallic acid were the most potent permeabilizers and facilitated the passage of the antibiotics through the outer membrane, as evidenced by their ability to cause LPS release, sensitize bacteria to SDS and Triton X-100.ResultsThe combination of permeabilizers and natural beta lactamase inhibitors (quercetin and epigallocatechin gallate) with antibiotics induced greater susceptibility of resistant isolates compared to antibiotic treatment with beta lactamase inhibitors alone. Pronounced effects were detected with 24.4 Gy in vitro gamma irradiation on permeability barrier, beta lactamase activity, and outer membrane protein profiles of the tested isolates.ConclusionsThe synergistic effects of the studied natural phytochemicals and antibiotics leads to new clinical choices via outer membrane destabilization (permeabilizers) and/or inactivation of the beta lactamase enzyme, which enables the use of older, more cost-effective antibiotics against resistant strains.

Antistress and antimicrobial studies of biphenyl chalcone derivatives

This work reports the synthesis, characterization of new (2E)-1-(biphenyl-4-yl)-3-aryl)-prop-2-en-1-one derivatives ( C1-C10 ). The chalcone derivatives were evaluated for in vitro antibacterial, antifungal, antioxidant activities. All the compounds exhibited moderate to good antimicrobial activity at MIC 10-40 µg/mL. The compounds were docked at the active site of the methionyl-tRNA synthetase (metRS) (PDB ID: 1A8H) to predict their putative interactions. The compounds further screened for antioxidant property, amid C5, C7 and C8 exhibited excellent DPPH radical scavenging activity. Compound C5 was selected for antistress studies against gamma radiation induced oxidative stress markers in E .coli K 12. The C5 pretreatment and irradiated bacteria sample showed modulatory action of stress enzymes SOD and CAT to near basal level and significant demulating effect on the level of TBARS.

Er:YAG Laser Irradiation Induces Behavioral Changes in V. harveyi.

Laser technologies have many different applications in medical, agricultural, and industrial fields. Studies have shown several effects of laser energy on different bacterial species, in a wide variety of settings. Recent reports have found that one of the unique features of bacteria is their ability to communicate among themselves (quorum sensing). We sought to investigate whether low-energy laser irradiation affects bacterial behavior, which is regulated by quorum sensing. Laser irradiations were performed using Er:YAG laser (2940 nm wavelength) at output powers of 0.5, 1.5, 2.5, and 4 W on wild-type Vibrio harveyi. Bioluminescence, motility, and biofilm forming capability were assessed on the bacteria after irradiation. After irradiation of bacteria, positive dose/output power dependencies were found in the bioluminescence omitted from tested experimental groups. Motility of colonies on semi-solid media was inhibited as irradiation output power was increased. However, after irradiation, biomass analysis of biofilm samples showed negligible differences between the irradiated samples and controls. Results indicate the impact of low-energy laser irradiation on bacterial behavior such as quorum sensing and motility, without affecting bacterial growth patterns.

Biophysical Investigations on the Interaction between Antimicrobial Peptides and Bacteria Killed by Cs-137 Irradiation

Two major problems arise while investigating the interaction between antimicrobial peptides (AMPs) and bacteria using biophysical techniques: (i) working with living pathogenic bacteria cause safety issues and (ii) some effects might depend on the metabolic activity of the bacteria. To distinguish between AMP-activities depending either on processes in active living bacteria or on interactions between AMPs and passive bacterial components, e.g. membrane permeabilization or DNA binding, one have to kill the bacteria before addition of the AMPs. Most techniques to kill bacteria are not suitable because they pre-damage or modify the bacterial envelope. Therefore, we investigated the usability of Cs-137 irradiation to kill the bacteria. Morphological studies using AFM and TEM showed no or only minor damage using a minimal bactericidal dose (MBD). At a higher dose, the bacterial integrity is damaged. The Zeta-potential of the bacteria is not changed by the irradiation and the ability of the bacteria to stimulate immune cells is not influenced. Binding experiments between AMPs and living bacteria as well as heat killed bacteria using ITC result in complex data because of metabolic activities as well as binding to intracellular components being exposed after heat killing. In contrast, experiments using bacteria killed by irradiation results in typical binding curves. The permeabilization assay based on the binding of Sytox Green to the bacterial DNA demonstrated that the envelopes of most of the irradiated bacteria are still intact. Surprisingly, the two AMPs LL32 (a derivative of the human cathelicidin) and polymyxin B cannot permeabilize the bacterial membrane of the killed bacteria anymore. In summary, biophysical investigations on bacteria killed by Cs-137 irradiation can be used as a tool for a better understanding of the interaction between whole bacteria and AMPs.

Microfluidic growth chambers with optical tweezers for full spatial single-cell control and analysis of evolving microbes

Single-cell analysis in microfluidic systems has opened up new possibilities in biotechnological research enabling us to deal with large eukaryotic cells and even small bacteria. In particular, transient investigations in laminar flow or diffusive environments can be performed to unravel single cell behaviour. Up to now, most systems have been limited with respect to precise cell inoculation and sampling methods. Individual cell selection and manipulations have now been made possible by combining laser tweezers with microfluidic cell cultivation environments specifically tailored for micrometre-sized bacteria. Single cells were optically seeded into various micrometre-sized growth sites arranged in parallel. During cultivation, single-cell elongation, morphology and growth rates were derived from single cells and microcolonies of up to 500 cells. Growth of irradiated bacteria was not impaired by minimizing the exposed laser dosage as confirmed by exceptional growth rates. In fact, Escherichia coli exhibited doubling times of less than 20min. For the first time, a filamentous Escherichia coli WT (MG1655) was safely relocated from its growing microcolony by laser manipulations. The cell was transferred to an empty cultivation spot allowing single-cell growth and morphology investigations. Contrary to previous discussions, the filamentous E. coli exhibited normal cell morphology and division after a few generations. This combination of optical tweezers and single-cell analysis in microfluidics adds a new degree of freedom to microbial single-cell analysis.

Comparison of the efficiency of titanium(IV) and iron(III) oxide nanoparticles as mediators in suppression of bacterial growth by radiation of a blue (405 nm) light-emitting diode

The effect of blue (405 nm) radiation of a light-emitting diode in combination with titanium(IV) and iron(III) oxide nanoparticles on S. aureus 209 P, S. simulans, and D. hominis bacteria is studied. It is shown that, upon irradiation of bacteria by blue (405 nm) light, Fe2O3 nanoparticles have a stronger (by 5–30%) antibacterial effect than TiO2 nanoparticles.

Influence of the photothermal effect of a gold nanorod cluster on biofilm disinfection

We evaluate a method for biofilm disinfection by raising biofilm temperature using the photothermal effect of a gold nanorod cluster. Gold nanorods (GNRs) are capable of generating enough heat to lyse bacteria by heating biofilm via laser irradiation. To test this, GNRs are synthesized using wet chemistry and a single GNR cluster is fabricated using photo-lithography technique. The GNR cluster is directly applied to the biofilm and its effects on bacteria are measured before and after laser irradiation. The photothermal effect of GNRs on the biofilm structure results in a considerable reduction of cell viability and biofilm thickness. Several quantitative measurements of bacterial mortality and biofilm destruction show an increase in efficacy with increasing durations of laser irradiation. Scanning electron microscopy images of the irradiated bacteria show obvious morphological damage such as rupture or collapse of the bacterial cell membrane in the biofilm. These results indicate that GNRs are useful and a potential material for use in photothermal treatments, particularly biofilm disinfection.

Pathogen germs response to low-dose radiation — medical approach

The side effects of radiation therapy in the case of microbial loading of irradiated organs was considered as phenomenological basis of the experiment carried out on Staphylococcus aureus (ATCC germ) exposed to low X-ray doses. The inoculum was prepared in a liquid culture medium with standard composition, the volumes of 3 ml identical samples (in sterile glass tubes) being irradiated in hospital conditions. Five experimental variants were developed corresponding to irradiation time durations between 25 and 100 minutes. The spectro-colorimetric assay was accomplished at 560 nm and 420 nm, the resulting average values (for three repetitions) being analyzed from the viewpoint of cell density in the irradiated variants compared to control ones. The resistance to antibiotics of the irradiated bacteria was tested on agarized cultures against five antibiotic molecules (ampicillin, cloramphenicol, tetracycline, tobramicin and ofloxacin) by assessing the diameter of inhibition growth areas in each case. The increase of the inhibition area diameter with up to 15% (in the case of tetracycline) was noticed for the lowest irradiation time for all five antibiotics, which is suggesting a weakening of the bacteria resistance to the pharmaceutical agents following the X-ray treatment. This was concordant with the results of the spectro-colorimetric assay of the cell density within the directly irradiated bacteria cultures. The main issue of this study is concerning the optimization of the radiotherapy protocol in patients with potential microbial loading.

Novel DNA photocleavage properties of [Cr(NN)3]3+ complexes

Abstract The aim of this study was to investigate the ability of chromium(III) tris-diimine complexes, [Cr(NN)3]3+, to induce DNA photodamage and to examine its capability to impair the survival of irradiated bacteria with the purpose of using these compounds as photocleavage reagents. [Cr(NN)3]3+ complexes, where NN stands for the ligands: phen (1,10-phenanthroline), 5-Mephen, 4,7-diMephen, 3,4,7,8-tetraMephen, 5-Phphen and 5-Clphen were used for this purpose. Their properties of DNA photocleavage were investigated by electrophoretic studies and assays of the photosensitization of transformed bacteria. The results clearly indicate that [Cr(NN)3]3+ complexes promote the photocleavage of plasmid DNA with varied degrees of efficiency after 12 h of irradiation. The combination of DNA, [Cr(NN)3]3+ and light proved to be required to induce the breakdown of DNA. Irradiated plasmid DNA-[Cr(NN)3]3+ association is also capable of impairing the transforming capacity of bacteria. These results provide evidence which confirms the responsibility and essential role of the excited state of [Cr(NN)3]3+ for inducing damage. Moreover, assays of the photosensitization of transformed bacterial suspensions suggest that Escherichia coli may be photoinactivated by irradiation in the presence of [Cr(NN)3]3+ complexes.

Inactivation of bacteria by high-power 940nm laser irradiation

Summary Background and objective Diode lasers are used in dentistry for oral surgery or disinfection of root canals in endodontic treatment. The aim of the study was to elucidate the basic mechanism of action in laser-induced root canal disinfection, which could be a photothermal and/or photochemical effect. Materials and methods To find out whether high-power NIR laser bacterial killing is caused by the light itself (photochemical effect or direct bacterial heating by selective absorption) or by a general thermal process, bacterial suspensions of a non-pathogenic strain of Escherichia coli were heated in a water bath and by diode laser (940 nm) each with the same temporal temperature course. Furthermore, bacterial suspensions were irradiated during which the temperature was fixed by iced water. Efficacy of bacterial inactivation was determined using double fluorescence labeling to distinguish vital from dead bacteria. Results Comparison of bacteria-killing efficacy between laser and water bath-induced heating showed no significant differences. The most important parameter was the maximum temperature achieved during heating. Laser irradiation of bacteria at low temperatures (elimination of temperature effect) did not show any bactericidal effect. Conclusion The experiments show that the inactivation of bacteria, at least for E. coli , using high-power 940 nm laser irradiation is solely based on a thermal mechanism. This should be taken into consideration during laser-assisted endodontic disinfection, where efficient bacterial inactivation must be achieved whilst preserving temperature-sensitive neighboring tissue.

Alterations of Outer Membrane Proteins and Virulence Genes Expression in Gamma-IrradiatedVibrio parahaemolyticusandVibrio alginolyticus

Gamma-irradiation technology sterilizes microorganisms and thereby prevents decay and improves the safety and shelf stability of food products. In this study we treated the foodborne pathogens Vibrio parahaemolyticus and Vibrio alginolyticus with gamma-irradiation (0.5 kGy) to evaluate their adaptative response. Outer membrane protein patterns of irradiated bacteria were found altered when analyzed by sodium dodecyl sulphate polyacrylamide gel electrophoresis. These modifications were manifested by the appearance and/or disappearance of bands as well as in the expression level of certain proteins. In addition, we searched for the presence of eight Vibrio cholerae virulence genes, toxR, toxS, toxRS, ctxA, zot, ace, toxT, and virulence pathogenicity island (VPI), in the genome of investigated strains. The expression of toxR, toxS, VPI, and ace genes in gamma-irradiated bacteria, studied by reverse transcriptase polymerase chain reaction, was altered. These variations were manifested by an increase and/or a decrease in the expression level of tested virulence genes.

A new perspective on radiation resistance based on Deinococcus radiodurans

In classical models of radiation toxicity, DNA is the molecule that is most affected by ionizing radiation (IR). However, recent data show that the amount of protein damage caused during irradiation of bacteria is better related to survival than to DNA damage. In this Opinion article, a new model is presented in which proteins are the most important target in the hierarchy of macromolecules affected by IR. A first line of defence against IR in extremely radiation-resistant bacteria might be the accumulation of manganese complexes, which can prevent the production of iron-dependent reactive oxygen species. This would allow an irradiated cell to protect sufficient enzymatic activity needed to repair DNA and survive.