recA Expression Research Articles

Comparison of gene expression among normally divided cells, elongated cells, spheroplasts at the beginning of growth, and enlarged spheroplasts at 43 h of growth in Lelliottia amnigena

We compared the expression levels of cell division related genes ftsA and ftsZ, DNA maintenance related genes hupA, recA, and topA, stress response related genes clpB, pspA and spy, and outer membrane synthesis related genes ompA and pbp1b, among normally divided cells, elongated (filamentous) cells, spheroplasts at the beginning of growth, and enlarged spheroplasts at 43h of growth in Lelliottia amnigena, by using quantitative RT-PCR. Although the expression level of ftsA was similar among the four samples, the expression level of ftsZ was higher in spheroplasts, and that in enlarged spheroplasts was the highest among the four samples. Although the hupA and recA expression levels in spheroplasts at 0h and 43h of growth were higher than those in divided cells, their topA expression was lower. This suggests that HupA and RecA may play a role in DNA replication during spheroplast enlargement. The stress response genes clpB, pspA, and spy were upregulated in spheroplasts. However, the expression levels of pspA and spy in enlarged spheroplasts at 43h of growth were lower than those in spheroplasts at the beginning of growth, suggesting that enlarged spheroplasts at 43h may be adapted to stress. The expression level of ompA in spheroplasts at 0h of growth was approximately 3 times higher than that in divided cells, but the ompA expression in spheroplasts at 43h of growth was approximately half of that in divided cells. This suggests that OmpA may be dispensable for spheroplast enlargement. Our findings thus show that the gene expression in spheroplast enlargement is quite different from cell elongation.

Threshold effect of growth rate on population variability of Escherichia coli cell lengths.

A long-standing question in biology is the effect of growth on cell size. Here, we estimate the effect of Escherichia coli growth rate (r) on population cell size distributions by estimating the coefficient of variation of cell lengths (CVL) from image analysis of fixed cells in DIC microscopy. We find that the CVL is constant at growth rates less than one division per hour, whereas above this threshold, CVL increases with an increase in the growth rate. We hypothesize that stochastic inhibition of cell division owing to replication stalling by a RecA-dependent mechanism, combined with the growth rate threshold of multi-fork replication (according to Cooper and Helmstetter), could form the basis of such a threshold effect. We proceed to test our hypothesis by increasing the frequency of stochastic stalling of replication forks with hydroxyurea (HU) treatment and find that cell length variability increases only when the growth rate exceeds this threshold. The population effect is also reproduced in single-cell studies using agar-pad cultures and ‘mother machine’-based experiments to achieve synchrony. To test the role of RecA, critical for the repair of stalled replication forks, we examine the CVL of E. coli ΔrecA cells. We find cell length variability in the mutant to be greater than wild-type, a phenotype that is rescued by plasmid-based RecA expression. Additionally, we find that RecA-GFP protein recruitment to nucleoids is more frequent at growth rates exceeding the growth rate threshold and is further enhanced on HU treatment. Thus, we find growth rates greater than a threshold result in increased E. coli cell lengths in the population, and this effect is, at least in part, mediated by RecA recruitment to the nucleoid and stochastic inhibition of division.

A Bacillus sp. strain with antagonistic activity against Fusarium graminearum kills Microcystis aeruginosa selectively

Cyanobacterial harmful algal blooms (CyanoHABs) cause severe environmental problems, economic losses and threaten human health seriously. In the present study, a Bacillus sp. strain, designated as AF-1, with strong antagonistic activity against plant pathogenic fungus Fusarium graminearum was isolated from purple soil. Bacillus sp. AF-1 selectively killed Microcystis aeruginosa at low cell density (1.6×103cfu/mL), and showed the strongest bactericidal activity against M. aeruginosa NIES-843 (Ae=93%, t=6d). The algicidal substances originated from strain AF-1 were stable in the temperature range of 35-100°C, and pH range of 3-11. Cell-free filtrate of AF-1 culture caused excessive accumulation of intracellular reactive oxygen species (ROS), cell death and the efflux of intracellular components of M. aeruginosa NIES-843 cells. The expression of genes recA, psbA1, psbD1, rbcL and mcyB, involved in DNA repair, photosynthesis and microcystin synthesis of NIES 843, were significantly influenced by the cell-free filtrate of AF-1 culture. Bacillus sp. AF-1 has the potential to be developed as a bifunctional biocontrol agent to control CyanoHABs and F. graminearum caused plant disease.

ITRAQ-based proteome analysis of fluoroquinolone-resistant Staphylococcus aureus.

The aim of this study was to compare global protein expression changes during fluoroquinolone (FQ) exposure of Staphylococcus aureus. Total protein extracts of wild-type S. aureus ATCC 29213 and six multidrug-resistant (MDR) strains derived from the wild-type under different FQ exposures were analysed using the 8-plex isobaric tag for relative and absolute quantitation (iTRAQ) method combined with LC-MS/MS analysis. Differentially expressed proteins were searched for their Gene Ontology (GO) annotation (UniProt database) and protein-protein interaction network (STRING v.10.0). recA expression was determined by real-time quantitative reverse transcription PCR (qRT-PCR) analysis. Overall, 582 unique proteins were identified at a confidence level of >95% (unused cut-off >1.3). After strict filtering for proteins with significant expression changes in comparison with the wild-type S. aureus ATCC 29213, 147 unique proteins were identified. GO searching showed that development of FQ resistance was associated with altered expression of various proteins involved in the SOS response (RecA), antibiotic resistance (MgrA), pathogenesis (uncharacterised leukocidin-like proteins 1 and 2, immunoglobulin-binding protein Sbi, triosephosphate isomerase, enolase, EsxA, SaeR, SarA, MgrA) and the stress response (alkyl hydroperoxide reductase subunit C, ClpB, ClpC, ClpL, ClpX, HslU, l-lactate dehydrogenase 1 and 2, SAV1710). Network analysis of antibiotic resistance-related proteins identified three major protein clusters involved in metabolic pathways, aminoacyl-tRNA biosynthesis and ribosome structure. qRT-PCR results were consistent with the proteomics data. Development of resistance to multiple drugs, including FQs, under drug exposure mostly involves upregulation of SOS and stress response proteins.

Antimicrobial Cluster Bombs: Silver Nanoclusters Packed with Daptomycin

Integration of two distinctive bactericides into one entity is a promising platform to improve the efficiency of antimicrobial agents. We report an efficient antimicrobial hybrid formed through conjugating silver nanoclusters (AgNCs) with daptomycin. The as-designed antimicrobial hybrid (D-AgNCs) inherits intrinsic properties of both bactericides with an enhanced synergistic performance. In particular, the chemically integrated D-AgNCs showed improved bacterial killing efficiency over the physically mixed daptomycin and AgNCs (D+AgNCs). More interestingly, the as-designed D-AgNCs could effectively damage the bacterial membrane. Propidium iodide (PI) stain showed bacterial membrane damage in about 85% of the bacteria population after treatment with D-AgNCs through creation of larger pores on the membrane as compared to D+AgNCs, largely due to the localization of daptomycin within the hybrid structure. These larger pores facilitated the entry of the D-AgNCs into the cell and led to more severe DNA damage of the bacterial DNA as compared to D+AgNCs in genomic DNA PAGE analysis. TUNEL assay further depicted more bacterial DNA breaks induced by D-AgNCs. The RecA gene expression level was upregulated, suggestive of DNA repair activation. The strong induced DNA damage benefited from the localization of AgNCs in the core of the antimicrobial hybrid structure, which could generate localized high ROS concentration and work as a critical ROS reservoir to continually generate ROS within the bacterium. The continual bombardments by these ROS generators restrict the ability of the bacteria to now develop resistance against this.

Biodetection of potential genotoxic pollutants entering the human food chain through ashes used in livestock diets

Ash derived from energy generation is used as a source of minerals in livestock feeds. The microbial biosensor recApr–Luc2 was built to detect genotoxic hazard in recycled ash. Escherichia coli SOS gene (recA, lexA, dinI and umuC) expression in response to cisplatin-induced DNA damage led to the selection of the recA promoter. The biosensor required functional RecA expression to respond to genotoxic heavy metals (Cr>Cd≈Pb), and polluted ash induced a strong recApr–Luc2 response. In human liver and intestinal cells, heavy metals induced acute toxicity (Cr>Cd>Pb) at concentrations sufficient to activate recApr–Luc2. Cytostatic effects, including genotoxicity, were cell- and metal-dependent, apart from Cr. In agreement with the recApr–Luc2 bioassay, Cr had the strongest effect in all cells. In conclusion, recApr–Luc2 could be useful for evaluating the genotoxic risk of pollutants present in ash that might be concentrated in animal products and, thus, entering the human food chain.

Overexpression of SOS genes in ciprofloxacin resistant Escherichia coli mutants

IntroductionFluoroquinolones are important antibiotics for the treatment of urinary tract infections caused by Escherichia coli. Mutational studies have shown that ciprofloxacin, a member of fluoroquinolones induces SOS response and mutagenesis in pathogenic bacteria which in turn develop antibiotic resistance. However, inhibition of SOS response can increase recombination activity which in turn leads to genetic variation. ObjectiveThe aim of this study was to measure 5 SOS genes expressions in nine E. coli mutants with different MICs for ciprofloxacin following exposure to ciprofloxacin. MethodsGene expression was assessed by quantitative real time PCR. Gene alteration assessment was conducted by PCR amplification and DNA sequencing. ResultsResults showed that the expression of recA was increased in 5 mutants. This overexpression is not related to gene alteration, and enhances the expression of polB and umuCD genes encoding nonmutagenic and mutagenic polymerases, respectively. The direct relationship between the level of SOS expression and the level of resistance to ciprofloxacin was also indicated. ConclusionIt was concluded that novel therapeutic strategy that inhibits RecA activity would enhance the efficiency of common antibiotics against pathogenic bacteria.

Evaluation of Potential Effects of NaCl and Sorbic Acid on Staphylococcal Enterotoxin A Formation.

The prophage-encoded staphylococcal enterotoxin A (SEA) is recognized as the main cause of staphylococcal food poisoning (SFP), a common foodborne intoxication disease, caused by Staphylococcus aureus. Studies on the production of SEA suggest that activation of the SOS response and subsequent prophage induction affect the regulation of the sea gene and the SEA produced, increasing the risk for SFP. The present study aims to evaluate the effect of NaCl and sorbic acid, in concentrations relevant to food production, on SOS response activation, prophage induction and SEA production. The impact of stress was initially evaluated on steady state cells for a homogenous cell response. NaCl 2% was found to activate the SOS response, i.e., recA expression, and trigger prophage induction, in a similar way as the phage-inducer mitomycin C. In contrast, sorbic acid decreased the pH of the culture to a level where prophage induction was probably suppressed, even when combined with NaCl stress. The impact of previous physiological state of the bacteria was also addressed on cells pre-exposed to NaCl, and was found to potentially affect cell response upon exposure to further stress. The results obtained highlight the possible SFP-related risks arising from the use of preservatives during food processing.

Evaluation of the photodynamic therapy as a potential technique for blood decontamination

Evaluation of the photodynamic therapy as a potential technique for blood decontamination

Fine-tuning recA expression in Staphylococcus aureus for antimicrobial photoinactivation: importance of photo-induced DNA damage in the photoinactivation mechanism.

Bacterial cell envelope is generally accepted as the primary target for a photo-induced oxidative stress. It is plausible that DNA damage occurs during the antimicrobial photoinactivation. Here we investigate the correlation between DNA damage and photoinactivation by evaluating the level of RecA-based DNA repair system in Staphylococcus aureus. By using exogenous photosensitizers (new methylene blue (NMB), toluidine blue O (TBO), 5,10,15,20-tetrakis(1-methyl-4-pyridinio)porphyrin tetra(p-toluenesulfonate) (TMPyP), zinc phthalocyanine (ZnPc), Rose Bengal (RB)) and ALA-induced endogenous porphyrin-dependent blue light (405 nm), several outcomes were observed: (i) an increase of DNA damage (from gel electrophoresis in DNA damage assay), (ii) an increase of recA expression (luminescence assay in recA-lux strain), and (iii) an increase of RecA protein level (Western blotting). When recA expression was repressed by novobiocin, or abolished by deleting the gene, S. aureus susceptibility towards photoinactivation was increased at approximately a hundred-fold. The absence of RecA increases DNA damage to yield bactericidal effect. In novobiocin-resistant mutant (gyrB), as opposed to wild type, neither RecA protein level nor cell’s susceptibility was affected by photoinactivation (when novobiocin is present). This is to suggest that GyrB-dependent inhibition mediated recA repression. Therefore, we have established the role of RecA in DNA damage during photoinactivation. With the use of rifampicin mutation frequency and Ames tests, we demonstrated that photoinactivation did not increase S. aureus mutagenesis and potentially is not mutagenic toward eukaryotic cells. The results suggest that the treatment is considered safe. In conclusion, we provide an evidence that recA inhibitor may serve as therapeutic adjuvant for antimicrobial photoinactivation. Clinical relevance of our findings warrants further investigations.

Development of a stress-induced mutagenesis module for autonomous adaptive evolution of Escherichia coli to improve its stress tolerance.

BackgroundMicrobial tolerance to different environmental stresses is of importance for efficient production of biofuels and biochemical. Such traits are often improved by evolutionary engineering approaches including mutagen-induced mutagenesis and successive passage. In contrast to these approaches which generate mutations in rapidly growing cells, recent research showed that mutations could be generated in non-dividing cells under stressful but non-lethal conditions, leading to the birth of the theory of stress-induced mutagenesis (SIM). A molecular mechanism of SIM has been elucidated to be mutagenic repair of DNA breaks. This inspired us to develop a synthetic SIM module to simulate the mutagenic cellular response so as to accelerate microbial adaptive evolution for an improved stress tolerance.ResultsA controllable SIM evolution module was devised based on a genetic toggle switch in Escherichia coli. The synthetic module enables expression and repression of the genes related to up- and down-regulation responses during SIM in a bistable way. Upon addition of different inducers, the module can be turned on or off, triggering transition to a mutagenic or a high-fidelity state and thus allowing periodic adaptive evolution. Six genes (recA, dinB, umuD, ropS, ropE, and nusA) in the up-regulation responses were evaluated for their potentials to enhance the SIM rate. Expression of recA, dinB, or ropS alone increased the SIM rate by 4.5- to 13.7-fold, whereas their combined expression improved the rate by 31.9-fold. Besides, deletion of mutL increased the SIM rate by 82-fold. Assembly of these genes into the SIM module in the mutL-deletion E. coli strain elevated the SIM rate by nearly 3000-fold. Accelerated adaptive evolution of E. coli equipped with this synthetic SIM module was demonstrated under n-butanol stress, with the minimal inhibitory concentration of n-butanol increasing by 56 % within 2.5 months.ConclusionsA synthetic SIM module was constructed to simulate cellular mutagenic responses during SIM. Based on this, a novel evolutionary engineering approach—SIM-based adaptive evolution—was developed to improve the n-butanol tolerance of E. coli.Electronic supplementary materialThe online version of this article (doi:10.1186/s13068-015-0276-1) contains supplementary material, which is available to authorized users.

Effect of SOS-response Inhibition on Enrofloxacin Resistance in <italic>Escherichia coli</italic>

To explore the effect of SOS response on enrofloxacin resistance in E. coli , we deleted the recA gene, which encodes the inducer of SOS response, and evaluated the combined inhibitory effect of RecA inhibitor and enrofloxacin on E. coli in vitro . By Red recombination, a deficient strain E. coli ATCC 25922/Δ recA was obtained. The expression of recA and umuC under the pressure of enrofloxacin was measured by real-time PCR. The minimum inhibitory concentration (MIC) of E. coli ATCC 25922 and E. coli ATCC 25922/Δ recA to many kinds of antibiotics, including enrofloxacin, were detected by broth dilution method. And the effect of recA deficiency on resistance variant to enrofloxacin was investigated by gradient plate method. N6-(1-naphthyl)-ADP, the inhibitor of SOS response, was synthesized and its influence combined with enrofloxacin on bacterial growth and resistance was evaluated. As results, E. coli ATCC 25922/Δ recA was established successfully and its MIC was decreased to an eighth of original strain. Moreover, the recA deficient strain was less prone to develop resistance than E. coli ATCC 25922 when exposed to enrofloxacin. The results of real-time PCR showed that SOS response was inhibited in recA -deleted strain or with the action of RecA inhibitor. The recA -deleted E. coli ATCC 25922 was more susceptive to enrofloxacin and the rate of resistant variation was lower than that in wild-type strain, and the inhibitor of RecA can inhibit SOS response to a certain degree and assist enrofloxacin to inhibit bacteria growth.

An inducible recA expression Bacillus subtilis genome vector for stable manipulation of large DNA fragments.

BackgroundThe Bacillus subtilis genome (BGM) vector is a novel cloning system based on the natural competence that enables B. subtilis to import extracellular DNA fragments into the cell and incorporate the recombinogenic DNA into the genome vector by homologous recombination. The BGM vector system has several attractive properties, such as a megabase cloning capacity, stable propagation of cloned DNA inserts, and various modification strategies using RecA-mediated homologous recombination. However, the endogenous RecA activity may cause undesirable recombination, as has been observed in yeast artificial chromosome systems. In this study, we developed a novel BGM vector system of an inducible recA expression BGM vector (iREX), in which the expression of recA can be controlled by xylose in the medium.ResultsWe constructed the iREX system by introducing the xylose-inducible recA expression cassette followed by the targeted deletion of the endogenous recA. Western blot analysis showed that the expression of recA was strictly controlled by xylose in the medium. In the absence of xylose, recA was not expressed in the iREX, and the RecA-mediated recombination reactions were greatly suppressed. By contrast, the addition of xylose successfully induced RecA expression, which enabled the iREX to exploit the same capacities of transformation and gene modifications observed with the conventional BGM vector. In addition, an evaluation of the stability of the cloned DNA insert demonstrated that the DNA fragments containing homologous sequences were more stably maintained in the iREX by suppressing undesirable homologous recombination.ConclusionsWe developed a novel BGM vector with inducible recA expression system, iREX, which enables us to manipulate large DNA fragments more stably than the conventional BGM vector by suppressing undesirable recombination. In addition, we demonstrate that the iREX can be applied to handling the DNA, which has several homologous sequences, such as multiple-reporter expression cassettes. Thus, the iREX expands the utility of the BGM vector as a platform for engineering large DNA fragments.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-015-1425-4) contains supplementary material, which is available to authorized users.

Recombinase and translesion DNA polymerase decrease the speed of replication fork progression during the DNA damage response in Escherichia coli cells

The SOS response is a DNA damage response pathway that serves as a general safeguard of genome integrity in bacteria. Extensive studies of the SOS response in Escherichia coli have contributed to establishing the key concepts of cellular responses to DNA damage. However, how the SOS response impacts on the dynamics of DNA replication fork movement remains unknown. We found that inducing the SOS response decreases the mean speed of individual replication forks by 30–50% in E. coli cells, leading to a 20–30% reduction in overall DNA synthesis. dinB and recA belong to a group of genes that are upregulated during the SOS response, and encode the highly conserved proteins DinB (also known as DNA polymerase IV) and RecA, which, respectively, specializes in translesion DNA synthesis and functions as the central recombination protein. Both genes were independently responsible for the SOS-dependent slowdown of replication fork progression. Furthermore, fork speed was reduced when each gene was ectopically expressed in SOS-uninduced cells to the levels at which they are expressed in SOS-induced cells. These results clearly indicate that the increased expression of dinB and recA performs a novel role in restraining the progression of an unperturbed replication fork during the SOS response.

Cellular and transcriptional responses in Microcystis aeruginosa exposed to two antibiotic contaminants.

The responses of Microcystis aeruginosa under exposure to spiramycin and amoxicillin were investigated on both cellular and genetic levels through a 7-day exposure test. Algal growth was inhibited by spiramycin while promoted by amoxicillin at test concentrations of 0.6-1.8 μg L(-1), indicating a higher toxicity of spiramycin than amoxicillin. During the whole exposure period, the chlorophyll a content and expression levels of psbA, psaB, and rbcL were significantly inhibited by spiramycin at test concentrations of 1.2 and 1.8 μg L(-1) (p < 0.05) and stimulated by 0.6-1.8 μg L(-1) of amoxicillin (p < 0.05), with respective decreases of up to 26, 75, 72, and 82% compared to the control and respective increases of 20, 70, 135, and 55%. During the 4 to 7 days of exposure, the microcystin-LR content and expression levels of mcyB and grpE were reduced by up to 66, 47, and 72% in spiramycin-treated algal cells, respectively, and stimulated by up to 1.3-, 1.4-, and 1.5-folds in amoxicillin-treated algal cells, respectively. Elevated recA expression was only observed in 1.2 and 1.8 μg L(-1) of spiramycin-treated algal cells, indicating severe DNA damage due to the high toxicity. Target antibiotics were suspected to regulate the growth and microcystin-production in M. aeruginosa via the photosynthesis system.

Lycopene-induced hydroxyl radical causes oxidative DNA damage in Escherichia coli.

Lycopene, which is a well-known red carotenoid pigment, has been drawing scientific interest because of its potential biological functions. The current study reports that lycopene acts as a bactericidal agent by inducing reactive oxygen species (ROS)-mediated DNA damage in Escherichia coli. Lycopene treatment elevated the level of ROS-in particular, hydroxyl radicals ((•)OH) -which can damage DNA in E. coli. Lycopene-induced DNA damage in bacteria was confirmed and we also observed cell filamentation caused by cell division arrest, an indirect marker of the DNA damage repair system, in lycopene-treated E. coli. Increased RecA expression was observed, indicating activation of the DNA repair system (SOS response). To summarize, lycopene exerts its antibacterial effects by inducing (•)OH -mediated DNA damage that cannot be ameliorated by the SOS response. Lycopene may be a clinically useful adjuvant for current antimicrobial therapies.

MG428 is a novel positive regulator of recombination that triggersmgpBandmgpCgene variation inMycoplasma genitalium

The human pathogen Mycoplasma genitalium employs homologous recombination to generate antigenic diversity in the immunodominant MgpB and MgpC proteins. Only recently, some of the molecular factors involved in this process have been characterized, but nothing is known about its regulation. Here, we show that M. genitalium expresses N-terminally truncated RecA isoforms via alternative translation initiation, but only the full-length protein is essential for gene variation. We also demonstrate that overexpression of MG428 positively regulates the expression of recombination genes, including recA, ruvA, ruvB and ORF2, a gene of unknown function co-transcribed with ruvAB. The co-ordinated induction of these genes correlated with an increase of mgpBC gene variation. In contrast, cells lacking MG428 were unable to generate variants despite expressing normal levels of RecA. Similarly, deletion analyses of the recA upstream region defined sequences required for gene variation without abolishing RecA expression. The requirement of these sequences is consistent with the presence of promoter elements associated with MG428-dependent recA induction. Sequences upstream of recA also influence the relative abundance of RecA isoforms, possibly through translational regulation. Overall, these results suggest that MG428 is a positive regulator of recombination and that precise control of recA expression is required to initiate mgpBC variation.

Kinetics of recA and recX induction in drug-susceptible and MDR clinical strains of Mycobacterium tuberculosis.

To investigate and compare the expression of recA and recX, components of the SOS pathway, following rifampicin treatment in drug-susceptible and MDR clinical strains of Mycobacterium tuberculosis. Strains (M. tuberculosis and Mycobacterium smegmatis) were subjected to rifampicin- and mitomycin-induced stress for 36 h followed by RNA extraction. recA and recX in the RNA extract were estimated using qRT-PCR. The MDR clinical strain induced faster (24 h) and higher (7-fold) levels of recA as compared with the drug-susceptible strain (36 h) in response to rifampicin. recX levels were found to rise with an increase in levels of recA; however, the levels were relatively higher than recA. Drug-susceptible and MDR strains have different kinetics of induction of DNA repair.

A novel mechanism for the antibacterial effect of silver nanoparticles on Escherichia coli.

Silver nanoparticles are known to have antimicrobial properties and have been used extensively in medicine, although the mechanism(s) of action have not yet been clearly established. In the present study, the findings suggest a novel mechanism for the antibacterial effect of silver nanoparticles on Escherichia coli, namely, the induction of a bacterial apoptosis-like response. We propose a possible mechanism for the bacterial apoptosis-like response that includes the following: accumulation of reactive oxygen species (ROS) (detected with H2DCFDA staining), increased intracellular calcium levels (detected with Fura-2 AM), phosphatidylserine exposure in the outer membrane (detected with Annexin V) which is the hallmarks of early apoptosis, disruption of the membrane potential [detected with DiBAC4(3)], activation of a bacterial caspase-like protein (detected by FITC-VAD-FMK staining) and DNA degradation (detected with TUNEL assay) which is the hallmarks of late apoptosis in bacterial cells treated with silver nanoparticles. We also performed RecA expression assay with western blotting and observed activation of SOS response to repair the damaged DNA. To summarize, silver nanoparticles are involved in the apoptosis-like response in E. coli and the novel mechanisms which were identified in this study, suggest that silver nanoparticles may be an effective antimicrobial agent with far lower propensity for inducing microbial resistance than antibiotics.

Magainin 2 induces bacterial cell death showing apoptotic properties.

Magainin 2 is pore-forming antimicrobial peptide on lipid matrix of bacterial membrane, secreted from the skin of the African clawed frog Xenopus laevis. The aim of this study was to investigate a new concept for antibacterial mechanisms called bacterial apoptosis-like cell death. We examined the morphological changes induced by magainin 2 in Escherichia coli, regarding apoptosis. Specifically, phosphatidylserine externalization from the inner to outer membrane surface was detected by Annexin V staining, and DNA fragmentation and chromatin condensation was detected by TUNEL and DAPI assay. We also found much mechanistic evidence to support the hypothesis that magainin 2 induces bacterial apoptosis-like death-including disturbance of membrane detected by DiBAC4(3), caspase activation observed by FITC-VAD-FMK staining, and analyzing the role of RecA in bacterial apoptosis-like death through the RecA expression assay by Western blot-in E. Coli when treated with magainin 2. On the basis of these results, magainin 2 exerts antibacterial activity with a new mechanism which is bacterial apoptosis-like death. Searching antimicrobial agents with novel mechanisms of action can be an effective strategy to coping with the emergence of new resistance mechanisms. Magainin 2 deserves further research as a potential antimicrobial therapeutic agent.