Response of Bacillus cereus ATCC 14579 to challenges with sublethal concentrations of enterocin AS-48. - Related Documents




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622901.0000Response of Bacillus cereus ATCC 14579 to challenges with sublethal concentrations of enterocin AS-48. BACKGROUND: Enterocin AS-48 is produced by Enterococcus faecalis S48 to compete with other bacteria in their environment. Due to its activity against various Gram positive and some Gram negative bacteria it has clear potential for use as a food preservative. Here, we studied the effect of enterocin AS-48 challenges on vegetative cells of Bacillus cereus ATCC 14579 by use of transcriptome analysis. RESULTS: Of the 5200 genes analysed, expression of 24 genes was found to change significantly after a 30 min treatment with a subinhibitory bacteriocin concentration of 0.5 microg/ml. Most of up-regulated genes encode membrane-associated or secreted proteins with putative transmembrane segments or signal sequences, respectively. One operon involved in arginine metabolism was significantly downregulated. The BC4206-BC4207 operon was found to be the most upregulated target in our experiments. BC4206 codes for a PadR type transcriptional regulator, while BC4207 codes for a hypothetical membrane protein. The operon structure and genes are conserved in B. cereus and B. thuringiensis species, but are not present in B. anthracis and B. subtilis. Using real-time qPCR, we show that these genes are upregulated when we treated the cells with AS-48, but not upon nisin treatment. Upon overexpression of BC4207 in B. cereus, we observed an increased resistance against AS-48. Expression of BC4207 in B. subtilis 168, which lacks this operon also showed increased resistance against AS-48. CONCLUSION: BC4207 membrane protein is involved in the resistance mechanism of B. cereus cells against AS-48.200919863785
621710.9995Identification of the sigmaB regulon of Bacillus cereus and conservation of sigmaB-regulated genes in low-GC-content gram-positive bacteria. The alternative sigma factor sigma(B) has an important role in the acquisition of stress resistance in many gram-positive bacteria, including the food-borne pathogen Bacillus cereus. Here, we describe the identification of the set of sigma(B)-regulated genes in B. cereus by DNA microarray analysis of the transcriptome upon a mild heat shock. Twenty-four genes could be identified as being sigma(B) dependent as witnessed by (i) significantly lower expression levels of these genes in mutants with a deletion of sigB and rsbY (which encode the alternative sigma factor sigma(B) and a crucial positive regulator of sigma(B) activity, respectively) than in the parental strain B. cereus ATCC 14579 and (ii) increased expression of these genes upon a heat shock. Newly identified sigma(B)-dependent genes in B. cereus include a histidine kinase and two genes that have predicted functions in spore germination. This study shows that the sigma(B) regulon of B. cereus is considerably smaller than that of other gram-positive bacteria. This appears to be in line with phylogenetic analyses where sigma(B) of the B. cereus group was placed close to the ancestral form of sigma(B) in gram-positive bacteria. The data described in this study and previous studies in which the complete sigma(B) regulon of the gram-positive bacteria Bacillus subtilis, Listeria monocytogenes, and Staphylococcus aureus were determined enabled a comparison of the sets of sigma(B)-regulated genes in the different gram-positive bacteria. This showed that only three genes (rsbV, rsbW, and sigB) are conserved in their sigma(B) dependency in all four bacteria, suggesting that the sigma(B) regulon of the different gram-positive bacteria has evolved to perform niche-specific functions.200717416654
620520.9993Characterization of Caulobacter crescentus response to low temperature and identification of genes involved in freezing resistance. Free-living bacteria must respond to a wide range of temperature changes, and have developed specific mechanisms to survive in extreme environments. In this work we describe a remarkable resistance of mesophilic bacterium Caulobacter crescentus to several cycles of freezing at -80 degrees C, which was able to grow at low temperatures. Exponentially growing cells and late stationary-phase cells presented higher freezing resistance at both -20 and -80 degrees C than early stationary-phase cells. Cryotolerance was observed when log-phase cultures grown at 30 degrees C were preincubated at 5, 15 or 20 degrees C before freezing at -20 degrees C. A transposon library was screened to identify mutants sensitive to freezing at -80 degrees C and three strains presenting <10% survival were isolated. Identification of genes disrupted in each mutant showed that they encoded an AddA family DNA helicase, a DEAD/DEAH box RNA helicase and a putative RND (resistance, nodulation, cell division) efflux system component. These strains showed longer generation times than wild-type cells when growing at 15 degrees C, with the RNA helicase mutant presenting a severe growth defect. These analyses suggest that the singular intrinsic resistance to freezing of C. crescentus is in fact a consequence of several independent traits, especially the maintenance of a proper degree of supercoiling of nucleic acids.200818801049
66230.9993Gene expression and physiological role of Pseudomonas aeruginosa methionine sulfoxide reductases during oxidative stress. Pseudomonas aeruginosa PAO1 has two differentially expressed methionine sulfoxide reductase genes: msrA (PA5018) and msrB (PA2827). The msrA gene is expressed constitutively at a high level throughout all growth phases, whereas msrB expression is highly induced by oxidative stress, such as sodium hypochlorite (NaOCl) treatment. Inactivation of either msrA or msrB or both genes (msrA msrB mutant) rendered the mutants less resistant than the parental PAO1 strain to oxidants such as NaOCl and H2O2. Unexpectedly, msr mutants have disparate resistance patterns when exposed to paraquat, a superoxide generator. The msrA mutant had a higher paraquat resistance level than the msrB mutant, which had a lower paraquat resistance level than the PAO1 strain. The expression levels of msrA showed an inverse correlation with the paraquat resistance level, and this atypical paraquat resistance pattern was not observed with msrB. Virulence testing using a Drosophila melanogaster model revealed that the msrA, msrB, and, to a greater extent, msrA msrB double mutants had an attenuated virulence phenotype. The data indicate that msrA and msrB are essential genes for oxidative stress protection and bacterial virulence. The pattern of expression and mutant phenotypes of P. aeruginosa msrA and msrB differ from previously characterized msr genes from other bacteria. Thus, as highly conserved genes, the msrA and msrB have diverse expression patterns and physiological roles that depend on the environmental niche where the bacteria thrive.201323687271
628840.9993Regulation of ofloxacin resistance in Escherichia coli strains causing calf diarrhea by quorum-sensing acyl-homoserine lactone signaling molecules. Escherichia coli is a major pathogen responsible for calf diarrhea. However, it has developed resistance to many antimicrobial drugs for their inappropriate usage. The bacterial quorum sensing system transmits information between bacteria, it's important in regulating bacterial virulence, drug and acid resistance and so on. This system can found in Gram-negative bacteria and operates through acyl-homoserine lactone (AHL) signaling molecules. In this study, a type I quorum sensing AHL, N-Octanoyl-L-Homoserine lactone (C8), was added to E. coli growth medium to investigate its regulatory functions in drug resistance. After screening out the strains of E. coli that showed an obvious regulatory effect to the drug ofloxacin (OFX), transcriptomic sequencing was performed on the E. coli strains from the sub-inhibitory concentration group that concentration plus C8 group, and the control group. It shows that C8 significantly influenced resistance to OFX and the minimum inhibitory concentration of OFX in the tested strain was significantly increased. To Analyze transcriptome sequencing results identified 415 differentially expressed genes between the control and sub-inhibitory concentration groups, of which 201 were up-regulated and 214 were down. There were 125 differentially expressed genes between bacteria treated with a sub-inhibitory concentration of OFX and those treated with C8, of which 102 were up-regulated and 23 were down. Finally, It found that to adding the C8 significantly increased the resistance of tested bacteria to OFX. Data from transcriptome sequencing on differently expressed genes helps to explain how the type I quorum sensing system controls drug resistance in E. coli.202539974163
629450.9993Comparison of Gene Expression Profiles of Uropathogenic Escherichia Coli CFT073 after Prolonged Exposure to Subinhibitory Concentrations of Different Biocides. Biocides are chemical compounds widely used for sterilization and disinfection. The aim of this study was to examine whether exposure to subinhibitory biocide concentrations influenced transcriptional expression of genes that could improve a pathogen's drug resistance or fitness. We used DNA microarrays to investigate the transcriptome of the uropathogenic Escherichia coli strain CFT073 in response to prolonged exposure to subinhibitory concentrations of four biocides: benzalkonium chloride, chlorhexidine, hydrogen peroxide and triclosan. Transcription of a gene involved in polymyxin resistance, arnT, was increased after treatment with benzalkonium chloride. However, pretreatment of the bacteria with this biocide did not result in cross-resistance to polymyxin in vitro. Genes encoding products related to transport formed the functional group that was most affected by biocides, as 110 out of 884 genes in this category displayed altered transcription. Transcripts of genes involved in cysteine uptake, sulfate assimilation, dipeptide transport, as well as cryptic phage genes were also more abundant in response to several biocides. Additionally, we identified groups of genes with transcription changes unique to single biocides that might include potential targets for the biocides. The biocides did not increase the resistance potential of the pathogen to other antimicrobials.201931569631
15860.9992Homology- and cross-resistance of Lactobacillus plantarum to acid and osmotic stress and the influence of induction conditions on its proliferation by RNA-Seq. In this study, homology- and cross-resistance of Lactobacillus plantarum L1 and Lactobacillus plantarum L2 to acid and osmotic stress were investigated. Meanwhile, its proliferation mechanism was demonstrated by transcriptomic analysis using RNA sequencing. We found that the homologous-resistance and cross-resistance of L. plantarum L1 and L. plantarum L2 increased after acid and osmotic induction treatment by lactic acid and sodium lactate solution in advance, and the survival rate of live bacteria was improved. In addition, the count of viable bacteria of L. plantarum L2 significantly increased cultivated at a pH 5.0 with a 15% sodium lactate sublethal treatment, compared with the control group. Further study revealed that genes related to membrane transport, amino acid metabolism, nucleotide metabolism, and cell growth were significantly upregulated. These findings will contribute to promote high-density cell culture of starter cultures production in the fermented food industry.202133945164
623070.9992dpr and sod in Streptococcus mutans are involved in coexistence with S. sanguinis, and PerR is associated with resistance to H2O2. Large numbers of bacteria coexist in the oral cavity. Streptococcus sanguinis, one of the major bacteria in dental plaque, produces hydrogen peroxide (H(2)O(2)), which interferes with the growth of other bacteria. Streptococcus mutans, a cariogenic bacterium, can coexist with S. sanguinis in dental plaque, but to do so, it needs a means of detoxifying the H(2)O(2) produced by S. sanguinis. In this study, we investigated the association of three oxidative stress factors, Dpr, superoxide dismutase (SOD), and AhpCF, with the resistance of S. sanguinis to H(2)O(2). The knockout of dpr and sod significantly increased susceptibility to H(2)O(2), while the knockout of ahpCF had no apparent effect on susceptibility. In particular, dpr inactivation resulted in hypersensitivity to H(2)O(2). Next, we sought to identify the factor(s) involved in the regulation of these oxidative stress genes and found that PerR negatively regulated dpr expression. The knockout of perR caused increased dpr expression levels, resulting in low-level susceptibility to H(2)O(2) compared with the wild type. Furthermore, we evaluated the roles of perR, dpr, and sod when S. mutans was cocultured with S. sanguinis. Culturing of the dpr or sod mutant with S. sanguinis showed a significant decrease in the S. mutans population ratio compared with the wild type, while the perR mutant increased the ratio. Our results suggest that dpr and sod in S. mutans are involved in coexistence with S. sanguinis, and PerR is associated with resistance to H(2)O(2) in regulating the expression of Dpr.201323263955
68380.9992Integrative Multiomics Analysis of the Heat Stress Response of Enterococcus faecium. A continuous heat-adaptation test was conducted for one Enterococcus faecium (E. faecium) strain wild-type (WT) RS047 to obtain a high-temperature-resistant strain. After domestication, the strain was screened with a significantly higher ability of heat resistance. which is named RS047-wl. Then a multi-omics analysis of transcriptomics and metabolomics was used to analyze the mechanism of the heat resistance of the mutant. A total of 98 differentially expressed genes (DEGs) and 115 differential metabolites covering multiple metabolic processes were detected in the mutant, which indicated that the tolerance of heat resistance was regulated by multiple mechanisms. The changes in AgrB, AgrC, and AgrA gene expressions were involved in quorum-sensing (QS) system pathways, which regulate biofilm formation. Second, highly soluble osmotic substances such as putrescine, spermidine, glycine betaine (GB), and trehalose-6P were accumulated for the membrane transport system. Third, organic acids metabolism and purine metabolism were down-regulated. The findings can provide target genes for subsequent genetic modification of E. faecium, and provide indications for screening heat-resistant bacteria, so as to improve the heat-resistant ability of E. faecium for production.202336979372
894390.9992Effects of indole on drug resistance and virulence of Salmonella enterica serovar Typhimurium revealed by genome-wide analyses. BACKGROUND: Many Gram-positive and Gram-negative bacteria produce large quantities of indole as an intercellular signal in microbial communities. Indole demonstrated to affect gene expression in Escherichia coli as an intra-species signaling molecule. In contrast to E. coli, Salmonella does not produce indole because it does not harbor tnaA, which encodes the enzyme responsible for tryptophan metabolism. Our previous study demonstrated that E. coli-conditioned medium and indole induce expression of the AcrAB multidrug efflux pump in Salmonella enterica serovar Typhimurium for inter-species communication; however, the global effect of indole on genes in Salmonella remains unknown. RESULTS: To understand the complete picture of genes regulated by indole, we performed DNA microarray analysis of genes in the S. enterica serovar Typhimurium strain ATCC 14028s affected by indole. Predicted Salmonella phenotypes affected by indole based on the microarray data were also examined in this study. Indole induced expression of genes related to efflux-mediated multidrug resistance, including ramA and acrAB, and repressed those related to host cell invasion encoded in the Salmonella pathogenicity island 1, and flagella production. Reduction of invasive activity and motility of Salmonella by indole was also observed phenotypically. CONCLUSION: Our results suggest that indole is an important signaling molecule for inter-species communication to control drug resistance and virulence of S. enterica.201222632036
8945100.9992Adaptation of a fluoroquinolone-sensitive Shigella sonnei to norfloxacin exposure. Shigella causes shigellosis that requires antibiotic treatment in severe cases. Sublethal antibiotic concentrations can promote resistance, but their effect on antibiotic-sensitive bacteria before resistance development is unclear. This study investigated the effects of sublethal norfloxacin (NOR) challenges on a NOR-sensitive strain, Shigella sonnei UKMCC1015. Firstly, the whole genome of S. sonnei UKMCC1015 was assembled, and 45 antimicrobial resistance (AMR) genes were identified. Interestingly, transcriptomic analysis showed that low NOR levels do not change either the expression of the AMR genes or NOR targets such as gyrA. Instead, multiple ribosomal protein genes were downregulated, which could be attributed to decreased ribosomal protein promoter activity, modulated by elevated guanosine pentaphosphate and tetraphosphate (ppGpp) levels. This alarmone is involved in the bacterial stringent response during environmental stress, and it is mainly produced from the ppGpp synthetase (relA). Additionally, we observed that a relA overexpression (prolonged period of elevated ppGpp levels) may negatively affect the NOR tolerance of the bacteria. In conclusion, this study revealed that a NOR-sensitive strain responds differently to sublethal NOR than commonly reported in resistant strains.202439100177
6327110.9992The Response of Enterococcus faecalis V583 to Chloramphenicol Treatment. Many Enterococcus faecalis strains display tolerance or resistance to many antibiotics, but genes that contribute to the resistance cannot be specified. The multiresistant E. faecalis V583, for which the complete genome sequence is available, survives and grows in media containing relatively high levels of chloramphenicol. No specific genes coding for chloramphenicol resistance has been recognized in V583. We used microarrays to identify genes and mechanisms behind the tolerance to chloramphenicol in V583, by comparison of cells treated with subinhibitory concentrations of chloramphenicol and untreated V583 cells. During a time course experiment, more than 600 genes were significantly differentially transcribed. Since chloramphenicol affects protein synthesis in bacteria, many genes involved in protein synthesis, for example, genes for ribosomal proteins, were induced. Genes involved in amino acid biosynthesis, for example, genes for tRNA synthetases and energy metabolism were downregulated, mainly. Among the upregulated genes were EF1732 and EF1733, which code for potential chloramphenicol transporters. Efflux of drug out of the cells may be one mechanism used by V583 to overcome the effect of chloramphenicol.201020628561
6292120.9992Genome-Wide Screening and Characterization of Genes Involved in Response to High Dose of Ciprofloxacin in Escherichia coli. The global emergence of antibiotic resistance, especially in Gram-negative bacteria, is an urgent threat to public health. Inevitably, considering its extensive use and misuse, resistance toward ciprofloxacin has increased in almost all clinically relevant bacteria. This study aimed to investigate the transcriptome changes at a high concentration of ciprofloxacin in Escherichia coli. In brief, 1,418 differentially expressed genes (DEGs) were identified, from which 773 genes were upregulated by ciprofloxacin, whereas 651 genes were downregulated. Enriched biological pathways reflected the upregulation of biological processes such as DNA damage and repair system, toxin/antitoxin systems, formaldehyde detoxification system. With kyoto encyclopedia of genes and genomes pathway analysis, higher expressed DEGs were associated with "LPS biosynthesis," "streptomycin biosynthesis," and "polyketide sugar unit biosynthesis." Lower expressed DEGs were associated with "biosynthesis of amino acids" and "flagellar assembly" pathways. After treatment of ciprofloxacin, lipopolysaccharide (LPS) release was increased by two times, and the gene expression level of LPS synthesis was elevated (p < 0.05) in both reference and clinical strains. Our results demonstrated that transient exposure to high-dose ciprofloxacin is a double-edged sword. Cautions should be taken when administering high-dose antibiotic treatment for infectious diseases.202235512736
187130.9992Functional coexistence of twin arsenic resistance systems in Pseudomonas putida KT2440. The genome of the soil bacterium Pseudomonas putida KT2440 bears two virtually identical arsRBCH operons putatively encoding resistance to inorganic arsenic species. Single and double chromosomal deletions in each of these ars clusters of this bacterium were tested for arsenic sensitivity and found that the contribution of each operon to the resistance to the metalloid was not additive, as either cluster sufficed to endow cells with high-level resistance. However, otherwise identical traits linked to each of the ars sites diverged when temperature was decreased. Growth of the various mutants at 15°C (instead of the standard 30°C for P. putida) uncovered that ars2 affords a much higher resistance to As (III) than the ars1 counterpart. Reverse transcription polymerase chain reaction of arsB1 and arsB2 genes as well as lacZ fusions to the Pars1 and Pars2 promoters traced the difference to variations in transcription of the corresponding gene sets at each temperature. Functional redundancy may thus be selected as a stable condition - rather than just as transient state - if it affords one key activity to be expressed under a wider range of physicochemical settings. This seems to provide a straightforward solution to regulatory problems in environmental bacteria that thrive under changing scenarios.201524673935
6291140.9992Adaptive Resistance of Staphylococcus aureus to Cefquinome Sulfate in an In Vitro Pharmacokinetic Model with Transcriptomic Insights. Cefquinome sulfate has a strong killing effect against Staphylococcus aureus (S. aureus), but bacterial resistance has become increasingly widespread. Experiments were conducted to investigate the pattern of adaptive resistance of S. aureus to cefquinome sulfate under different dosage regimens by using pharmacokinetic-pharmacodynamics (PK-PD) modeling, and the adaptive-resistant bacteria in different states were screened and subjected to transcriptomic sequencing. The results showed that the minimum inhibitory concentration of Staphylococcus aureus under the action of cefquinome sulfate was 0.5 μg/mL, the anti-mutation concentration was 1.6 μg/mL, and the mutation selection window range was 0.5~1.6 μg/mL. In the in vitro pharmacokinetic model to simulate different dosing regimens in the animal body, there are certain rules for the emergence of adaptive drug-resistant bacteria: the intensity of bacterial resistance gradually increased with culture time, and the order of emergence was tolerant bacteria (TO) followed by persistent bacteria (PE) and finally resistant bacteria (RE). The sequence reflected the evolution of adaptive drug resistance. Transcriptome Gene Ontology (GO) analysis revealed that differentially expressed genes were involved in cellular respiration, energy derivation by oxidation of organic compounds, and oxidation-reduction processes. The differentially expressed genes identified functioned in the synthesis of cell membranes, cytoplasm, and intracellular parts. A Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis found that 65 genes were differentially expressed after cefquinome sulfate treatment, of which 35 genes were significantly upregulated and 30 genes were significantly downregulated. Five genes, sdhB, sdhA, pdhA, lpdA, and sucC, may be involved in network regulation. This study revealed the cross-regulation of multiple metabolic pathway networks and the targets of network regulation of S. aureus to produce adaptive drug resistance. The results will provide guidance for clinical drug use in animals infected with S. aureus.202540005696
6171150.9992Host response to infection with a temperature-sensitive mutant of Salmonella typhimurium in a susceptible and a resistant strain of mice. The inoculation of a temperature-sensitive mutant of Salmonella typhimurium induced a long-lasting infection in susceptible (C57BL/6) and resistant (A/J) mice. During week 1 of infection, the number of bacteria in the spleens was similar in both mouse strains. Then, the decrease of bacteria was more rapid in the resistant strain. Splenomegaly and granulomatous hepatitis were more severe in the susceptible strain. The immune response induced by this infection was studied. In both mouse strains delayed-type hypersensitivity to Salmonella antigens was present, and resistance to reinfection with a virulent strain of S. typhimurium or with Listeria monocytogenes appeared with the same kinetics. Thus, it does not seem that the gene(s) controlling natural resistance to S. typhimurium act(s) on acquired immunity.19853897053
8941160.9992Salicylate reduces the antimicrobial activity of ciprofloxacin against extracellular Salmonella enterica serovar Typhimurium, but not against Salmonella in macrophages. OBJECTIVES: Salicylate, a potent inducer of the MarA activator in Salmonella enterica, is the principal metabolite of aspirin, which is often consumed for medicinal and cosmetic uses. Our research was aimed at testing if salicylate activates the mar regulon in macrophage-associated Salmonella (intracellular bacteria), and investigating its effects on bacterial susceptibility to ciprofloxacin extracellularly and intracellularly. METHODS: J774 macrophages were infected with S. enterica serovar Typhimurium (wild-type and marA null mutant), treated with ciprofloxacin with and without pre-exposure to salicylate, and the surviving bacteria were counted. Similar experiments were conducted with bacteria in broth (extracellular bacteria). Phe-Arg-beta-naphthylamide (PAbetaN) was added to investigate the role of efflux pumps in resistance. The transcriptional regulation of marRAB, acrAB and micF in extracellular and intracellular Salmonella Typhimurium with and without salicylate and ciprofloxacin was investigated using green fluorescent protein as a marker protein and quantitative real time PCR. RESULTS: Pre-exposure of Salmonella to salicylate increased the resistance of extracellular but not intracellular bacteria to ciprofloxacin, although salicylate stimulated the expression of mar genes in intracellular and extracellular bacteria. Using marA mutants and the inhibitor PAbetaN, we showed that the improved resistance in extracellular bacteria is derived from the induction of acrAB by salicylate, which is mediated by MarA. CONCLUSIONS: In intracellular bacteria, the expression of acrAB is already higher when compared with extracellular cells; therefore, salicylate does not result in significant acrAB induction intracellularly and subsequent resistance enhancement. Results show that conclusions raised from extracellular studies cannot be applied to intracellular bacteria, although the systems have similar functions.201020237076
251170.9992Deep sequencing analysis of the Kineococcus radiotolerans transcriptome in response to ionizing radiation. Kineococcus radiotolerans is a gram-positive, radiation-resistant bacterium that was isolated from a radioactive environment. The synergy of several groups of genes is thought to contribute to the radio-resistance of this species of bacteria. Sequencing of the transcriptome, RNA sequencing (RNA-seq), using deep sequencing technology can reveal the genes that are differentially expressed in response to radiation in this bacterial strain. In this study, the transcriptomes of two samples (with and without irradiation treatment) were sequencing by deep sequencing technology. After the bioinformatics process, 143 genes were screened out by the differential expression (DE) analysis. In all 143 differentially expressed genes, 20 genes were annotated to be related to the radio-resistance based on the cluster analysis by the cluster of orthologous groups of proteins (COG) annotation which were validated by the quantitative RT-PCR. The pathway analysis revealed that these 20 validated genes were related to DNA damage repair, including recA, ruvA and ruvB, which were considered to be the key genes in DNA damage repair. This study provides the foundation to investigate the regulatory mechanism of these genes.201525467197
189180.9992Arsenate detoxification in a Pseudomonad hypertolerant to arsenic. Pseudomonas sp. strain As-1, obtained from an electroplating industrial effluent, was capable of growing aerobically in growth medium supplemented with up to 65 mM arsenate (As (V)), significantly higher concentrations than those tolerated by other reference arsenic resistant bacteria. The majority of the arsenic was detected in culture supernatants as arsenite (As (III)) and X-ray absorbance spectroscopy suggested that 30% of this cell-bound arsenic was As (V), 65% As (III) and 5% of arsenic was associated with sulphur. PCR analysis using primers designed against arsenic resistance genes of other Gram-negative bacteria confirmed the presence of an arsenic resistance operon comprising of three genes, arsR, arsB and arsC in order of predicted transcription, and consistent with a role in intracellular reduction of As (V) and efflux of As (III). In addition to this classical arsenic resistance mechanism, other biochemical responses to arsenic were implicated. Novel arsenic-binding proteins were purified from cellular fractions, while proteomic analysis of arsenic-induced cultures identified the upregulation of additional proteins not normally associated with the metabolism of arsenic. Cross-talk with a network of proteins involved in phosphate metabolism was suggested by these studies, consistent with the similarity between the phosphate and arsenate anions.200717160678
8456190.9992Identification of genes required by Bacillus thuringiensis for survival in soil by transposon-directed insertion site sequencing. Transposon-directed insertion site sequencing was used to identify genes required by Bacillus thuringiensis to survive in non-axenic plant/soil microcosms. A total of 516 genetic loci fulfilled the criteria as conferring survival characteristics. Of these, 127 (24.6 %) were associated with uptake and transport systems; 227 loci (44.0 %) coded for enzymatic properties; 49 (9.5 %) were gene regulation or sensory loci; 40 (7.8 %) were structural proteins found in the cell envelope or had enzymatic activities related to it and 24 (4.7 %) were involved in the production of antibiotics or resistance to them. Eighty-three (16.1 %) encoded hypothetical proteins or those of unknown function. The ability to form spores was a key survival characteristic in the microcosms: bacteria, inoculated in either spore or vegetative form, were able to multiply and colonise the soil, whereas a sporulation-deficient mutant was not. The presence of grass seedlings was critical to colonisation. Bacteria labelled with green fluorescent protein were observed to adhere to plant roots. The sporulation-specific promoter of spo0A, the key regulator of sporulation, was strongly activated in the rhizosphere. In contrast, the vegetative-specific promoters of spo0A and PlcR, a pleiotropic regulator of genes with diverse activities, were only very weakly activated.201424310935