Effect of Microwaves on Escherichia coli and Bacillus subtilis

Suspensions of Escherichia coli and Bacillus subtilis spores were exposed to conventional thermal and microwave energy at 2,450 MHz. The degrees of inactivation by the different energy sources were compared quantitatively. During the transient heating period by microwave energy, approximately a 6 log cycle reduction in viability was encountered for E. coli. This reduction was nearly identical to what is expected for the same time-temperature exposure to conventional heating. Heating of B. subtilis spores by conventional and microwave energy was also carried out at 100 C, in ice and for transient heating. The degree of inactivation by microwave energy was again identical to that by conventional heating. In conclusion, inactivation of E. coli and B. subtilis by exposure to microwaves is solely due to the thermal energy, and there is no per se effect of microwaves.     

Cephalexin-Induced Morphological Alterations in the Surface Structures of Staphylococcus aureus and Escherichia coli Demonstrated by Scanning Electron Microscopy

The scanning electron microscope was used to investigate the alterations in surface morphology of Staphylococcus aureus 209P and Escherichia coli NIH induced by the action of cephalexin known to interfere with cell-wall synthesis. Exposure to cephalexin produced a series of changes on the surface morphology in proportion to the concentrations of cephalexin added. Untreated S. aureus cells had smooth contours. Exposure to 1 μg/ml of cephalexin during the logarithmic phase of growth in S. aureus did not produce any detectable changes. Upon exposure of S. aureus to 5 μ/ml or 10 μg/ml, some cells were larger than normal and showed abnormal cell division-like structures in part. When S. aureus was exposed to 50 μg/ml, cell division was completely inhibited, and no formation of grape-like clusters was observed. Untreated E. coli cells appeared to have smooth and regular contours. E. coli propagated almost normally upon exposure of the organisms to 1 μg/ml of cephalexin. Filamentous structures were observed with the exposure of E. coli to 12.5 μg/ml or 25 μg/ml, but spheroplast-like structures were not observed. Exposure to 100 μg/ml of cephalexin resulted in the formation of marked filamentous cells and spheroplast-like structures having multiple small saccular outpouchings. Scanning electron microscope demonstrated more completely the morphological abnormalities induced by cephalexin.     

Nonthermal killing effect of microwave irradiation

Summary Suspensions of E. coli were exposed to microwave irradiation at constant temperatures. No cell death was observed at 35°C, whereas, at 45, 47, and 50°C, the death rates of E. coli exposed to microwave irradiation were higher than those obtained in conventional heat sterilization at the same temperatures.     

Thermal and Nonthermal Effects of Discontinuous Microwave Exposure (2.45 Gigahertz) on the Cell Membrane of Escherichia coli

The aim of this study was to investigate the effects on the cell membranes of Escherichia coli of 2.45-GHz microwave (MW) treatment under various conditions with an average temperature of the cell suspension maintained at 37°C in order to examine the possible thermal versus nonthermal effects of short-duration MW exposure. To this purpose, microwave irradiation of bacteria was performed under carefully defined and controlled parameters, resulting in a discontinuous MW exposure in order to maintain the average temperature of the bacterial cell suspensions at 37°C. Escherichia coli cells were exposed to 200- to 2,000-W discontinuous microwave (DW) treatments for different periods of time. For each experiment, conventional heating (CH) in a water bath at 37°C was performed as a control. The effects of DW exposure on cell membranes was investigated using flow cytometry (FCM), after propidium iodide (PI) staining of cells, in addition to the assessment of intracellular protein release in bacterial suspensions. No effect was detected when bacteria were exposed to conventional heating or 200 W, whereas cell membrane integrity was slightly altered when cell suspensions were subjected to powers ranging from 400 to 2,000 W. Thermal characterization suggested that the temperature reached by the microwave-exposed samples for the contact time studied was not high enough to explain the measured modifications of cell membrane integrity. Because the results indicated that the cell response is power dependent, the hypothesis of a specific electromagnetic threshold effect, probably related to the temperature increase, can be advanced.     

Dose‐dependent fate of GFP‐expressing Escherichia coli in the alimentary canal of adult house flies

The adult house fly Musca domestica (L.) (Diptera: Muscidae) can disseminate bacteria from microbe‐rich substrates to areas in which humans and domesticated animals reside. Because bacterial abundance fluctuates widely across substrates, flies encounter and ingest varying amounts of bacteria. This study investigated the dose‐dependent survival of bacteria in house flies. Flies were fed four different ‘doses’ of green fluorescent protein (GFP)‐expressing Escherichia coli (GFP E. coli) (very low, low, medium, high) and survival was determined at 1, 4, 10 and 22 h post‐ingestion by culture and epifluorescent microscopy. Over 22 h, the decline in GFP E. coli was significant in all treatments (P < 0.04) except the very low dose treatment (P = 0.235). Change in survival (ΔS) did not differ between flies fed low and very low doses of bacteria across all time‐points, although ΔS in both treatments differed from that in flies fed high and medium doses of bacteria at several time‐points. At 4, 10 and 22 h, GFP E. coli ΔS significantly differed between medium and high dose‐fed flies. A threshold dose, above which bacteria are detected and destroyed by house flies, may exist and is likely to be immune‐mediated. Understanding dose‐dependent bacterial survival in flies can help in predicting bacteria transmission potential.     

Pathogenic bacteria and TNF do not induce production of macrophage migration inhibitory factor (MIF) by human monocytes

Elevated serum macrophage migration inhibitory factor (MIF) is associated with severe sepsis, but it is not clear whether bacteria stimulate synthesis of MIF by blood leukocytes directly or via induction of TNF. Here we assess production of MIF mRNA and protein by blood leukocytes from healthy human subjects (n = 28) following exposure to bacteria commonly associated with sepsis (Escherichia coli and Streptococcus pneumoniae). Bacteria did not increase levels of MIF mRNA or secreted protein. CD14⁺ monocytes were the main cell type producing MIF before and after stimulation. Exposure of leukocytes to TNF did not induce MIF. Hence elevated levels of serum MIF observed in sepsis may not reflect MIF produced by blood leukocytes stimulated directly by bacteria or TNF.     

Effect of Gamma-Ray upon Food Microorganisms

The author has carried out a study concerning a simplified method for the measurement of gamma-ray resistivity of E. coli. This report deals with comparison of each survival curve of 11 species of bacteria and the influence of addition of chemicals on a medium containing E. coli by gamma-ray irradiation by application of this method.     

Survival of Shewanella oneidensis MR-1 after UV Radiation Exposure

We systematically investigated the physiological response as well as DNA damage repair and damage tolerance in Shewanella oneidensis MR-1 following UVC, UVB, UVA, and solar light exposure. MR-1 showed the highest UVC sensitivity among Shewanella strains examined, with D₃₇ and D₁₀ values of 5.6 and 16.5% of Escherichia coli K-12 values. Stationary cells did not show an increased UVA resistance compared to exponential-phase cells; instead, they were more sensitive at high UVA dose. UVA-irradiated MR-1 survived better on tryptic soy agar than Luria-Bertani plates regardless of the growth stage. A 20% survival rate of MR-1 was observed following doses of 3.3 J of UVC m⁻², 568 J of UVB m⁻², 25 kJ of UVA m⁻², and 558 J of solar UVB m⁻², respectively. Photoreactivation conferred an increased survival rate to MR-1 of as much as 177- to 365-fold, 11- to 23-fold, and 3- to 10-fold following UVC, UVB, and solar light irradiation, respectively. A significant UV mutability to rifampin resistance was detected in both UVC- and UVB-treated samples, with the mutation frequency in the range of 10⁻⁵ to 10⁻⁶. Unlike in E. coli, the expression levels of the nucleotide excision repair (NER) component genes uvrA, uvrB, and uvrD were not damage inducible in MR-1. Complementation of Pseudomonas aeruginosa UA11079 (uvrA deficient) with uvrA of MR-1 increased the UVC survival of this strain by more than 3 orders of magnitude. Loss of damage inducibility of the NER system appears to contribute to the high sensitivity of this bacterium to UVR as well as to other DNA-damaging agents.     

Synthesis of a new class of antimicrobial agents incorporating the indolin-2-one moiety

New furanone derivatives incorporating the indolin-2-one moiety 3 were prepared via the Perkin reaction of isatins 1 with aroylpropionic acids 2 under conventional conditions or microwave irradiation. A series of functionally heterocyclic derivatives (e.g., pyridazines, pyrroles, and sulfonamides) incorporating the indolin-2-one moiety was achieved via reaction of 3 with different reagents under microwave irradiation conditions. The newly synthesized compounds were characterized on the basis of FTIR, ¹H, ¹³C NMR and mass spectral studies. Some of the new synthesized compounds were screened for antibacterial activity against Gram-positive bacteria (Staphylococcus aureus and Bacillus cereus), Gram-negative bacteria (Escherichia coli and Shigilla flexneri) and antifungal activity against Aspergillus flavus and Candida albicans. Compound 8 j was equipotent to chloramphenicol in inhibiting the growth of E. coli minimum inhibitory concentration (MIC 2.5 μg/mL). Compound 8j may possibly be used as a lead compound for developing a new antibacterial agents. The antibacterial activity is expressed as the corresponding MIC (μg/mL) values.     

Competition for growth substrates in river water between Escherichia coli and indigenous bacteria illustrated by high-resolution mass spectrometry

Escherichia coli normally cannot grow in the environment. One environmental stress that prevents E. coli growth may be the competition for growth substrates with co-existing micro-organisms. In this study, the growth substrates of E. coli were screened by high-resolution mass spectrometry and compared with those of indigenous bacteria in river water. In an incubation experiment, E. coli multiplied in sterilized river water, but did not multiply when indigenous micro-organisms were present in the water. By analysing dissolved organic matter in the river water before and after E. coli growth, 35 compounds were identified as putative growth substrates of E. coli. Among them, 33 compounds were also identified as putative growth substrates of indigenous bacteria. These results indicate that E. coli and indigenous bacteria compete for organic substrates in river water, which could suppress the growth of E. coli.     

Rapid and sensitive detection of Pseudomonas aeruginosa in chlorinated water and aerosols targeting gyrB gene using real-time PCR

Aims: This study aimed to assess the contamination risk of Escherichia coli in commercial lettuce grown under three different irrigation systems (overhead sprinkler, subsurface drip and surface furrow). Methods and Results: Three replicated field trials were conducted. In an initial trial, we consistently observed higher mesophilic bacteria counts under sprinkler irrigation but visual quality was found to be dependent on the water potential of leaves at harvest. Further, in the other two trials, E. coli K‐12 strains LMM1010 and ATCC 25253, was injected into the water stream of the different irrigation systems to determine survival in the field. Results showed that product samples were positive for E. coli up to 7 days when using sprinkler irrigation, whereas only one product sample was found positive for E. coli when using other irrigation methods. Survival of bacteria in soil persisted longer in furrow‐irrigated areas, ranging from an estimated 17 days in winter months to 5 days during the warmer summer periods. This finding combined with results from a parallel 3‐year survey of canal waters indicate that while highest risk of finding E. coli in irrigation water is in warmer months, the survival in soil is lower during the same time period. Conclusions: Our results in a study set under common commercial conditions confirmed the enhanced risk of E. coli contamination when using sprinkle irrigation. Furthermore, E. coli persistence in furrow‐irrigated soil validates the importance of an early irrigation termination for both sprinkler and furrow methods. Significance and Impact of the Study: Stringent monitoring and in‐field food safety controls should be emphasized during the last few days before harvest.     

Lysosomal involvement in apoptosis

Abstract

The involvement of reactive oxygen species (ROS) in the induction of DNA damage to Escherichia coli cells caused by UVC (254 nm) irradiation was studied. We verified the expression of the soxS gene induced by UVC (254 nm) and its inhibition by sodium azide, a singlet oxygen (¹O₂) scavenger. Additional results showed that a water-soluble carotenoid (norbixin) protects against the lethal effects of UVC. These results suggest that UVC radiation can also cause ROS-mediated lethality.     

Effect of copper hydroxide sprays for citrus canker control on wild-type Escherichia coli

Aims: To show that application of copper hydroxide citrus sprays mixed with field source water (possibly contaminated) will not support Escherichia coli on plant surfaces. Environmental stresses of transient phyllosphere bacteria and presence of copper will eradicate these bacteria before harvest. Methods and Results: Studies were performed in vitro with bacteria grown in broth and then subjected to field spray copper hydroxide concentrations in the broth and on citrus leaves. Escherichia coli exposed to copper hydroxide in vitro were eradicated from the broth within 6–8 h depending on the broth pH. Even with near neutral pH (7·2), cells began to die immediately after exposure to copper. No E. coli survived on leaf surfaces sprayed with copper. Conclusions: Copper field sprays mixed with water that may contain E. coli can help eliminate E. coli from plant surfaces. Significance and Impact of the Study: HACCP mandates are becoming more restrictive because of the increased illness resulting from food pathogens on fresh produce. Use of potable water in fields, a proposed mandate, is not feasible for large grove owners. These data show that copper sprays aimed at reducing citrus canker also affect E. coli and may help to ease water quality mandates.     

Selection of Escherichia coli-inhibiting strains of Lactobacillus paracasei subsp. paracasei

The aim of this study was to select Escherichia coli-inhibiting strains among lactic acid bacteria. On the basis of phenotypical and technological characteristics, 20 strains of lactic acid bacteria were screened from a total of 225 isolates, obtained from nine samples of artisanal Caprino d'Aspromonte cheese, made from raw goats' milk. The antagonistic activity of these 20 strains was detected in plates against three different strains of E. coli. Two strains of Lactobacillus paracasei subsp. paracasei showed a marked anti-E. coli activity against all three strains tested; the other lactic acid bacteria did not exhibit inhibiting activity. The E. coli inhibition can be ascribed to production of bacteriocin-like compounds. The use of L. paracasei subsp. paracasei strains to increase the safety of the cheeses made from raw milk is recommended because these cultures strongly inhibit E. coli, without foreseeable adverse sensory changes. Received 27 December 2001/ Accepted in revised form 28 June 2002     

Activities involved in base excision repair of bacteriophage T4 and lambda DNA in vivo

Summary The in vivo excision repair functions of Escherichia coli exonuclease III and 3-methyladenine DNA glycosylase I, and bacteriophage T4 pyrimidine dimer-DNA glycosylase were investigated. Following exposure of bacteriophage T4 or lambda to methyl methanesulfonate or ultraviolet irradiation, survival was determined by plating on E. coli have various genetic backgrounds. Although exonuclease III was shown to participate in base excision repair initiated by 3-methyladenine DNA glcosylase I, it had no detectable role in base excision repair initiated by the T4 pyrimidine dimer-DNA glycosylase. Despite its 3 apurinic/apyrimidinic endonuclease activity in vitro, T4 pyrimidine dimer-DNA glycosylase, even in large quantities, did not complement mutants defective in exonuclease III in the repair of apurinic sites generated by 3-methyladenine DNA glycosylase I in vivo.     

UV light killing efficacy of fluorescent protein‐expressing cancer cells in vitro and in vivo

We investigated the cell‐killing efficacy of UV light on cancer cells expressing GFP in the nucleus and RFP in the cytoplasm (dual‐color cells). After exposure to various doses of UVA, UVB, or UVC, apoptotic and viable cells were quantitated under fluorescence microscopy using dual‐color 143B human osteosarcoma cells, HT‐1080 human fibrosarcoma cells, Lewis lung carcinoma (LLC), and XPA‐1 human pancreatic cancer cells in vitro. UV‐induced cancer cell death was wave‐length and dose dependent, as well as cell‐line dependent. After UVA exposure, most cells were viable even when the UV dose was increased up to 200 J/m². With UVB irradiation, cell death was observed with irradiation at 50 J/m². For UVC, as little as 25 J/m² UVC irradiation killed approximately 70% of the 143B dual‐color cells. This dose of UVB or UVA had almost no effect on the cancer cells. UV‐induced cancer cell death varied among the cell lines. Cell death began about 4 h after irradiation and continued until 10 h after irradiation. UVC exposure also suppressed cancer cell growth in nude mice in a model of minimal residual cancer (MRC). No apparent side effects of UVC exposure were observed. This study opens up the possibility of UVC treatment for MRC after surgical resection. J. Cell. Biochem. 110: 1439–1446, 2010. © 2010 Wiley‐Liss, Inc.     

The influence of colicinogenic plasmids ColIb-P9, ColIa-CA53 and ColV-K30 on the repair, mutagenesis and induction of colicin E1 synthesis

Summary The presence of colicinogenic plasmids ColIb-P9 and ColIa-CA53 in E. coli K-12 cells, wild-type with respect to repair, enhanced the survival of cells after UV irradiation and increased the frequency of UV-induced argE3 and his-4 reversions, while the presence of ColV-K30 negatively affected repair and mutagenesis. The plasmid ColIb-P9 showed a UV-protective effect in E. coli cells carrying mutations in genes uvrA, uvrB, uvrC, polA, recB, recF, though in none of the mutants did cell survival reach the wild-type level. The effect of ColIb-P9 on mutagenesis did not depend on the uvrA or recB genes. The plasmids' protective effect and the enhancement of mutagenesis depended on the recA ⁺ lexA⁺ genotype. The frequency of 2-aminopurine-induced mutations was not affected by ColIb-P9 or ColV-K30. The presence of ColIb-P9 decreased the ability of ColEl-carrying cells to induce colicin E1 synthesis caused by DNA-damaging agents: UV, MNNG, mitomycin C, whereas ColV-K30 increased the percentage of colicin E1-producing cells. These plasmid effects on the level of induction of colicin E1 synthesis were not observed in the case of induction caused by chloramphenicol which did not depend on the products of recA and lexA genes.Abbreviations AP 2-aminopurine - MNNG N-methyl-N-nitro-N-nitrosoguanidine - ICS induction of colicin synthesis - CM chlorampheniol - MC mitomycin C     

Examination of indigenous microbiota and survival of Escherichia coli 0157 and Salmonella in a paper milling environment

Aims: A public beach was frequently cited for health advisories because of high Escherichia coli levels, the source suspected to be a paper mill located upstream. This investigation sought to confirm whether or not the paper mill was the pollution source, and to characterize the risk to recreational bathers imposed by the source. Methods and Results: Quantification of E. coli in river water collected at incremental distances showed that paper mill effluent caused elevated E. coli levels in beach samples. Samples collected throughout the mill were variably positive for heterotrophic bacteria, total coliforms and E. coli, but negative for pathogenic E. coli O157 and Salmonella. Escherichia coli O157 or Salmonella spiked into mill samples (4·2 log₁₀ or 5·6 log₁₀ CFU per 100 ml, respectively) fell below detection levels within 14–24 h in raw (unaltered) samples, while in heat‐sterilized replicates, the counts remained at initial levels or increased over 36 h. Conclusions: Pathogenic E. coli O157 and Salmonella were not isolated from paper mill samples. The absence of native bacteria allowed the survival of pathogens, while their presence accelerated pathogen decline. Significance and Impact of the Study: The co‐existence of paper mill and swimming beach may be reasonable for now in spite of the limitations of an E. coli‐based assay for beach water.     

Escherichia coli heme oxygenase modulates host innate immune responses

Induction of mammalian heme oxygenase (HO)‐1 and exposure of animals to carbon monoxide (CO) ameliorates experimental colitis. When enteric bacteria, including Escherichia coli, are exposed to low iron conditions, they express an HO‐like enzyme, chuS, and metabolize heme into iron, biliverdin and CO. Given the abundance of enteric bacteria residing in the intestinal lumen, our postulate was that commensal intestinal bacteria may be a significant source of CO and those that express chuS and other Ho‐like molecules suppress inflammatory immune responses through release of CO. According to real‐time PCR, exposure of mice to CO results in changes in enteric bacterial composition and increases E. coli 16S and chuS DNA. Moreover, the severity of experimental colitis correlates positively with E. coli chuS expression in IL‐10 deficient mice. To explore functional roles, E. coli were genetically modified to overexpress chuS or the chuS gene was deleted. Co‐culture of chuS‐overexpressing E. coli with bone marrow‐derived macrophages resulted in less IL‐12p40 and greater IL‐10 secretion than in wild‐type or chuS‐deficient E. coli. Mice infected with chuS‐overexpressing E. coli have more hepatic CO and less serum IL‐12 p40 than mice infected with chuS‐deficient E. coli. Thus, CO alters the composition of the commensal intestinal microbiota and expands populations of E. coli that harbor the chuS gene. These bacteria are capable of attenuating innate immune responses through expression of chuS. Bacterial HO‐like molecules and bacteria‐derived CO may represent novel targets for therapeutic intervention in inflammatory conditions.