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Sample records for aureus pc221 plasmid

  1. Single-stranded plasmid DNA in Bacillus subtilis and Staphylococcus aureus.

    OpenAIRE

    te Riele, H; Michel, B.; Ehrlich, S D

    1986-01-01

    Plasmid pC194 was found to exist in a double-stranded and a single-stranded DNA form in Bacillus subtilis and Staphylococcus aureus. This single-stranded DNA was found as a circular molecule of the same size as the parental monomer and corresponded to only one of the two DNA strands. It represented one-third of plasmid copies. Single- and double-stranded DNA copies in similar proportions to the above were detected for five other S. aureus plasmids (pC221, pC223, pE194, pT127, and pT181) and o...

  2. Comparative analysis of conjugative plasmids mediating gentamicin resistance in Staphylococcus aureus.

    OpenAIRE

    Goering, R. V.; Ruff, E A

    1983-01-01

    Five gentamicin-resistant clinical isolates of Staphylococcus aureus were found to contain self-transmissible plasmids of 32 to 37 megadaltons in size. Restriction endonuclease digests of the plasmids were markedly similar to those of reference plasmids of unrelated geographical origin, thus suggesting the significant contribution of common conjugal plasmids to the emergence of gentamicin resistance in S. aureus populations.

  3. Common R-plasmids in Staphylococcus aureus and Staphylococcus epidermidis during a nosocomial Staphylococcus aureus outbreak.

    OpenAIRE

    Cohen, M. L.; Wong, E. S.; Falkow, S

    1982-01-01

    During a 7-month period in 1978 to 1979, 31 patients and personnel at a Kentucky hospital were colonized or infected with a Staphylococcus aureus strain resistant to clindamycin, erythromycin, gentamicin, methicillin, penicillin, and tetracycline. S. epidermidis with similar antibiotic resistance patterns had been isolated in this hospital in the year before the S. aureus outbreak. A 32-megadalton R-plasmid, pUW3626, mediating resistance to penicillin and gentamicin, was present in these isol...

  4. Plasmid profiles and antibiotic susceptibility patterns of Staphylococcus aureus isolates from Nigeria.

    Science.gov (United States)

    Olukoya, D K; Asielue, J O; Olasupo, N A; Ikea, J K

    1995-06-01

    In an investigation into the problems of infections due to Staphylococcus aureus in Nigeria, 100 strains were isolated from various hospitals in Lagos. The strains were screened for the presence of plasmids and for susceptibility to antimicrobial agents. Plasmids were extracted by modification of the method of Takahashi and Nagono[1]. The plasmids were diverse in nature. The strains were found to be highly resistant to commonly prescribed antibiotics. PMID:8669391

  5. A stable luciferase reporter plasmid for in vivo imaging in murine models of Staphylococcus aureus infections.

    Science.gov (United States)

    Bacconi, Marta; Haag, Andreas F; Torre, Antonina; Castagnetti, Andrea; Chiarot, Emiliano; Delany, Isabel; Bensi, Giuliano

    2016-04-01

    In vivo imaging of bioluminescent bacteria permits their visualization in infected mice, allowing spatial and temporal evaluation of infection progression. Most available bioluminescent strains were obtained by integration of the luciferase genes into the bacterial chromosome, a challenging and time-consuming approach. Recently, episomal plasmids were used, which were introduced in bacteria and expressed all genes required for bioluminescence emission. However, the plasmid was progressively lost in vitro and in vivo, if bacteria were not maintained under antibiotic selective pressure. Increased stability could be obtained inserting into the plasmid backbone sequences that assured plasmid partition between daughter bacterial cells, or caused death of bacteria that had lost the plasmid. So far, no detailed analysis was performed of either plasmid stability in vivo or contribution of different stabilizing sequence types. Here we report the construction of a plasmid, which includes the Photorhabdus luminescens lux cassette expressed under the control of a Staphylococcus aureus specific gene promoter, and toxin/antitoxin (T/A) and partition sequences (Par) conferring stability and transmissibility of the plasmid. Following infection of mice with S. aureus carrying this plasmid, we demonstrated that the promoter-lux fusion was functional in vivo, that the plasmid was retained by 70-100% of bacterial cells 7 days post-infection, and that both stabilizing sequence types were required to maximize plasmid retention. These data suggest that the plasmid can be a valuable tool to study gene expression and bacterial spread in small laboratory animals infected with S. aureus or possibly other Gram-positive human pathogens. PMID:26685857

  6. Expansion of a Plasmid Classification System for Gram-Positive Bacteria and Determination of the Diversity of Plasmids in Staphylococcus aureus Strains of Human, Animal, and Food Origins

    OpenAIRE

    Lozano, Carmen; García-Migura, Lourdes; Aspiroz, Carmen; Zarazaga, Myriam; Torres, Carmen; Aarestrup, Frank Møller

    2012-01-01

    An expansion of a previously described plasmid classification was performed and used to reveal the plasmid content of a collection of 92 Staphylococcus aureus strains of different origins. rep genes of other genera were detected in Staphylococcus. S1 pulsed-field gel electrophoresis (PFGE) hybridizations were performed with 18 representative S. aureus strains, and a high number of plasmids of different sizes and organizations were detected.

  7. Efficient non-enzymatic cleavage of Staphylococcus aureus plasmid DNAs mediated by neodymium ions.

    Science.gov (United States)

    Zovčáková, Monika; Španová, Alena; Pantůček, Roman; Doškař, Jiří; Rittich, Bohuslav

    2016-08-15

    Staphylococcus aureus plasmids are the main factor in the spreading of antibacterial resistance among bacterial strains that has emerged on a worldwide scale. Plasmids recovered from 12 clinical and food isolates of S. aureus were treated with 10 mM free lanthanide Nd(3+) ions (non-enzymatic cleavage agent) in Hepes buffer (pH 7.5) at 70 °C. Topological forms of plasmids-closed circular (ccc), open circular (oc), and linear (lin)-produced by cleavage at different times were separated using pulsed-field agarose gel electrophoresis. The method is proposed to detect and differentiate several plasmids in the same bacterial strain according to their size. PMID:27237372

  8. Exfoliative toxin plasmids of bacteriophage group 2 Staphylococcus aureus: sequence homology.

    OpenAIRE

    Warren, R. L.

    1980-01-01

    The plasmid contents of seven exfoliative toxin-producing strains of phage group 2 Staphylococcus aureus were analyzed by agarose gel electrophoresis and deoxyribonucleic acid-deoxyribonucleic acid hybridization. All strains were found to contain a large plasmid with a molecular weight of 27 X 10(6) except for strain RW1005. A comparison of the restriction endonuclease cleavage products by agarose gel electrophoresis showed that the number and size distribution of the fragments of all these T...

  9. Expansion of a plasmid classification system for Gram-positive bacteria and determination of the diversity of plasmids in Staphylococcus aureus strains of human, animal, and food origins

    DEFF Research Database (Denmark)

    Lozano, C.; Garcia-Migura, L.; Aspiroz, C.;

    2012-01-01

    An expansion of a previously described plasmid classification was performed and used to reveal the plasmid content of a collection of 92 Staphylococcus aureus strains of different origins. rep genes of other genera were detected in Staphylococcus. S1 pulsed-field gel electrophoresis (PFGE) hybrid...

  10. Penicillinase plasmid-linked genetic determinants for enterotoxins B and C1 production in Staphylococcus aureus.

    OpenAIRE

    Altboum, Z; Hertman, I; Sarid, S

    1985-01-01

    The genes encoding for beta-lactamase (bla+) and resistance to metallic ions (cadmium, mercury, lead, arsenate, and arsenite) were located in a 56.2-kilobase plasmid, pZA10, isolated from a clinical strain, Staphylococcus aureus 6344. This strain produced enterotoxin B and enterotoxin C1. Elimination of pZA10 by either sodium dodecyl sulfate or heat treatment (43 degrees C) resulted in the loss of the capability of the bacteria to produce both enterotoxin B and enterotoxin C1. A physical map ...

  11. Nucleotide sequence and expression of the mercurial-resistance operon from Staphylococcus aureus plasmid pI258

    International Nuclear Information System (INIS)

    The mercurial-resistance determinant from Staphylococcus aureus plasmid pI258 is located on a 6.4-kilobase-pair Bgl II fragment. The determinant was cloned into both Bacillus subtilis and Escherichia coli. Mercury resistance was found only in B. subtilis. The 6404-base-pair DNA sequence of the Bgl II fragment was determined. The mer DNA sequence includes seven open reading frames, two of which have been identified by homology with the merA (mercuric reductase) and merB (organomercurial lyase) genes from the mercurial-resistance determinants of Gram-negative bacteria. Whereas 40% of the amino acid residues overall were identical between the pI258 merA polypeptide product and mercuric reductases from Gram-negative bacteria, the percentage identity in the active-site positions and those thought to be involved in NADPH and FAD contacts was above 90%. The 216 amino acid organomercurial lyase sequence was 39% identical with that from a Serratia plasmid, with higher conservation in the middle of the sequences and lower homologies at the amino and carboxyl termini. The remaining five open reading frames in the pI258 mer sequence have no significant homologies with the genes from previously sequenced Gram-negative mer operons

  12. Plasmid-borne cadmium resistance genes in Listeria monocytogenes are similar to cadA and cadC of Staphylococcus aureus and are induced by cadmium.

    OpenAIRE

    Lebrun, M; AUDURIER, A.; Cossart, P

    1994-01-01

    pLm74 is the smallest known plasmid in Listeria monocytogenes. It confers resistance to the toxic divalent cation cadmium. It contains a 3.1-kb EcoRI fragment which hybridizes with the cadAC genes of plasmid pI258 of Staphylococcus aureus. When introduced into cadmium-sensitive L. monocytogenes or Bacillus subtilis strains, this fragment conferred cadmium resistance. The DNA sequence of the 3.1-kb EcoRI fragment contains two open reading frames, cadA and cadC. The deduced amino acid sequences...

  13. Cadmium resistance from Staphylococcus aureus plasmid pI258 cadA gene results from a cadmium-efflux ATPase.

    OpenAIRE

    Nucifora, G; Chu, L; Misra, T K; Silver, S

    1989-01-01

    Cadmium resistance specified by the cadA determinant of Staphylococcus aureus plasmid pI258 results from the functioning of a cadmium-efflux system. In the nucleotide sequence of the DNA fragment containing the cadA determinant, two open reading frames were identified. The larger one, corresponding to a predicted polypeptide of 727 amino acid residues, is necessary and sufficient for expression of cadmium resistance. Comparison of the CadA amino acid sequence with known protein sequences sugg...

  14. Introduction of plasmid DNA into an ST398 livestock-associated methicillin-resistant Staphylococcus aureus strain

    Science.gov (United States)

    MRS926 is a livestock-associated methicillin-resistant Staphylococcus aureus (MRSA) strain of sequence type (ST) 398. In order to facilitate in vitro and in vivo studies of this strain, we sought to tag it with a fluorescent marker. We cloned a codon-optimized gene for TurboGFP into a shuttle vector...

  15. Analysis of the beta-lactamase plasmid of borderline methicillin-susceptible Staphylococcus aureus: focus on bla complex genes and cadmium resistance determinants cadD and cadX.

    Science.gov (United States)

    Massidda, Orietta; Mingoia, Marina; Fadda, Daniela; Whalen, Michael B; Montanari, Maria Pia; Varaldo, Pietro E

    2006-03-01

    Borderline methicillin-susceptible Staphylococcus aureus strains are a rather homogeneous group, characterized by MICs of penicillinase-resistant penicillins (PRPs) at or just below the susceptibility breakpoint. Other features unique to this group include the presence of a pBW15-like beta-lactamase plasmid, the association with phage complex 94/96, and the production of a PRP-hydrolyzing beta-lactamase activity in addition to the classical penicillinase activity. The four HindIII fragments of pBORa53, a pBW15-like plasmid from the well-studied borderline S. aureus strain a53, were cloned in Escherichia coli, sequenced and analyzed. The plasmid (17,334 bp in size) contains 14 open reading frames (ORFs) and a complete copy of transposon Tn552, which harbors the three genes of the bla complex (blaZ, blaR1, and blaI) necessary for penicillinase production. Among the other 11 ORFs identified, two were homologous to cadmium resistance determinants of Staphylococcus lugdunensis and to the cadD and cadX genes recently detected in S. aureus. Consistent with this, strain a53 was found to be cadmium resistant. From a collection of 30 S. aureus isolates with borderline PRP MIC levels, 27 matched strain a53 in the positive amplification reactions with all of the four primer pairs targeting the cadD-cadX region, the presence of the 17.3-kb plasmid, and the level of cadmium resistance. The well-established S. aureus laboratory strain ATCC 29213 was also found to express cadD-cadX-mediated cadmium resistance. pBORa53 could be re-isolated from transformants obtained by transferring it into a PRP-susceptible recipient. However, while the transformants demonstrated levels of cadmium and penicillin resistance similar to those of strain a53, they remained fully susceptible to PRPs. PMID:16229889

  16. Methicillin-resistant S. aureus (MRSA), extended-spectrum (ESBL)- and plasmid-mediated AmpC ß-lactamase -producing Gram-negative bacteria associated with skin and soft tissue infections in hospital and community settings

    OpenAIRE

    Selma Uzunović; Branka Bedenić; Ana Budimir; Amir Ibrahimagić; Farah Kamberović; Zlatko Fiolić; Michelle I. A. Rijnders; Stobberingh, Ellen E

    2015-01-01

    Aim To investigate the characteristics of meticillin-resistant S. aureus (MRSA), extended-spectrum (ESBL), and plasmid-mediated AmpC beta-lactamase producing Gram-negative bacteria causing skin and soft tissue infections (SSTIs) in hospital and outpatient settings of Zenica-Doboj Canton, Bosnia and Herzegovina. Methods Antibiotic susceptibility was determined by disc-diffusion and broth microdillution methods according to CLSI guidelines. MecA gene was detected by PCR, and genetic charact...

  17. Plasmid-mediated transformation in Bacillus megaterium.

    OpenAIRE

    Brown, B. J.; Carlton, B C

    1980-01-01

    A transformation system was developed for Bacillus megaterium by using antibiotic resistance plasmid deoxyribonucleic acid molecules derived from Staphylococcus aureus and Bacillus cereus. Lysozyme-generated protoplasts of B. megaterium allowed uptake of plasmid deoxyribonucleic acid in the presence of polyethylene glycol. Transformants expressed the antibiotic resistance determinants present on the plasmid deoxyribonucleic acid, and reisolated plasmid deoxyribonucleic acid yielded restrictio...

  18. Methicillin-resistant S. aureus (MRSA, extended-spectrum (ESBL- and plasmid-mediated AmpC ß-lactamase -producing Gram-negative bacteria associated with skin and soft tissue infections in hospital and community settings

    Directory of Open Access Journals (Sweden)

    Selma Uzunović

    2015-08-01

    Full Text Available Aim To investigate the characteristics of meticillin-resistant S. aureus (MRSA, extended-spectrum (ESBL, and plasmid-mediated AmpC beta-lactamase producing Gram-negative bacteria causing skin and soft tissue infections (SSTIs in hospital and outpatient settings of Zenica-Doboj Canton, Bosnia and Herzegovina. Methods Antibiotic susceptibility was determined by disc-diffusion and broth microdillution methods according to CLSI guidelines. MecA gene was detected by PCR, and genetic characterization of MRSA was performed using spa-typing and the algorithm based upon repeat patterns (BURP. Double-disk-synergy test was used to screen for ESBLs. PCR was used to detect blaESBL alleles. Genetic relatedness of the strains was tested by PFGE. Results Seventeen in-patients with MRSA, 13 with ESBL-producing Gram-negative bacteria and three patients co-infected with both, were detected. Five MRSA and 16 ESBL-producing Gramnegative bacteria were found in outpatient samples. Klebsiella spp. was isolated in 11 in- and seven outpatients. MLST CC152 was the most prevalent MRSA. Seven (38.9% Klebsiella spp. yielded amplicons with primers specific for SHV, TEM-1 and CTXM group 1 β-lactamases. Eight K. pneumonia (44.4% and 16 (64% MRSA (including the in- and outpatient strains were clonally related. Conclusion The presence of MRSA and ESBL-producing organisms causing SSTIs in the community poses a substantial concern, due to the high morbidity and mortality associated with possible consequent hospital infections.

  19. Detection and characterization of mupirocin resistance in Staphylococcus aureus.

    OpenAIRE

    Janssen, D A; Zarins, L T; Schaberg, D R; Bradley, S. F.; Terpenning, M S; Kauffman, C A

    1993-01-01

    Fourteen mupirocin-resistant Staphylococcus aureus strains were isolated over 18 months; 12 exhibited low-level resistance, while two showed high-level resistance. Highly mupirocin-resistant strains contained a large plasmid which transferred mupirocin resistance to other S. aureus strains and to Staphylococcus epidermidis. This plasmid and pAM899-1, a self-transferable gentamicin resistance plasmid, have molecular and biologic similarities.

  20. Transformation of Bacillus thuringiensis protoplasts by plasmid deoxyribonucleic acid.

    OpenAIRE

    Martin, P A; Lohr, J. R.; Dean, D H

    1981-01-01

    A method has been developed to transform plasmid deoxyribonucleic acid into protoplasts of the insect pathogen Bacillus thuringiensis. Protoplasts were formed by treatment of cells with lysozyme. The efficiency of formation of protoplasts was affected by the strain, the media, and the cell density. Deoxyribonucleic acid uptake was induced by polyethylene glycol. Deoxyribonucleic acid from the Staphylococcus aureus plasmid pC194 was used for transformation. Although this plasmid could not be i...

  1. Expression and inducibility in Staphylococcus aureus of the mecA gene, which encodes a methicillin-resistant S. aureus-specific penicillin-binding protein.

    OpenAIRE

    Ubukata, K; Nonoguchi, R; Matsuhashi, M; Konno, M

    1989-01-01

    A beta-lactam-sensitive strain of Staphylococcus aureus could be converted to methicillin resistance by the introduction of a plasmid carrying the 4.3-kilobase HindIII chromosomal DNA fragment which encoded the mecA gene from a methicillin-resistant S. aureus. Transformant cells produced methicillin-resistant S. aureus-specific penicillin-binding protein constitutively, and additional insertion of an inducible penicillinase plasmid caused production of the pencillin-binding protein to become ...

  2. Rapid procedure for isolation of plasmid DNA and application to epidemiological analysis.

    OpenAIRE

    Takahashi, S; Nagano, Y

    1984-01-01

    A rapid and simple plasmid isolation procedure was developed for the epidemiological analysis of plasmid-mediated antimicrobial resistance. By this method, plasmid DNAs ranging in molecular weight between 2.0 and 122 X 10(6) could be detected. Various bacteria, such as strains of the family Enterobacteriaceae, Pseudomonas aeruginosa, Haemophilus influenzae, and Staphylococcus aureus, could be analyzed. The plasmid DNA obtained could be directly used for restriction endonuclease analysis witho...

  3. Transmissible mupirocin resistance in Staphylococcus aureus.

    OpenAIRE

    Rahman, M.; Noble, W. C.; Cookson, B

    1989-01-01

    The spread of two strains of Staphylococcus aureus with high level resistance to mupirocin is described. The resistance proved to be easily transferred to other S. aureus strains by filter mating experiments and on the skin of mice. No plasmid band corresponding to the resistance could be demonstrated by agarose gel electrophoresis or by caesium chloride gradient centrifugation but cleavage of 'chromosomal' DNA from resistant recipients showed bright bands of DNA absent from sensitive controls.

  4. An updated view of plasmid conjugation and mobilization in Staphylococcus.

    Science.gov (United States)

    Ramsay, Joshua P; Kwong, Stephen M; Murphy, Riley J T; Yui Eto, Karina; Price, Karina J; Nguyen, Quang T; O'Brien, Frances G; Grubb, Warren B; Coombs, Geoffrey W; Firth, Neville

    2016-01-01

    The horizontal gene transfer facilitated by mobile genetic elements impacts almost all areas of bacterial evolution, including the accretion and dissemination of antimicrobial-resistance genes in the human and animal pathogen Staphylococcus aureus. Genome surveys of staphylococcal plasmids have revealed an unexpected paucity of conjugation and mobilization loci, perhaps suggesting that conjugation plays only a minor role in the evolution of this genus. In this letter we present the DNA sequences of historically documented staphylococcal conjugative plasmids and highlight that at least 3 distinct and widely distributed families of conjugative plasmids currently contribute to the dissemination of antimicrobial resistance in Staphylococcus. We also review the recently documented "relaxase-in trans" mechanism of conjugative mobilization facilitated by conjugative plasmids pWBG749 and pSK41, and discuss how this may facilitate the horizontal transmission of around 90% of plasmids that were previously considered non-mobilizable. Finally, we enumerate unique sequenced S. aureus plasmids with a potential mechanism of mobilization and predict that at least 80% of all non-conjugative S. aureus plasmids are mobilizable by at least one mechanism. We suggest that a greater research focus on the molecular biology of conjugation is essential if we are to recognize gene-transfer mechanisms from our increasingly in silico analyses. PMID:27583185

  5. cmp, a cis-acting plasmid locus that increases interaction between replication origin and initiator protein.

    OpenAIRE

    Gennaro, M L; Novick, R P

    1986-01-01

    pT181, a 4.4-kilobase multicopy plasmid of Staphylococcus aureus, encodes a trans-acting initiator protein, RepC, which was rate limiting for replication. Deletions in a 500-base-pair region of the plasmid external to the minimal replicon decreased the ability of the plasmid to compete with a coexisting incompatible plasmid. These deletions, which define a region called cmp (for competition), appeared to affect the interaction of RepC and the plasmid origin of replication. However, in the hom...

  6. Spread of Staphylococcus aureus resistant to penicillin and tetracycline within and between dairy herds

    DEFF Research Database (Denmark)

    Waage, S.; Bjorland, J.; Caugant, D. A.;

    2002-01-01

    One hundred and seven bovine isolates of penicillin and tetracycline resistant Staphylococcus aureus, recovered from 25 different dairy herds in various parts of Norway, were characterized using antimicrobial susceptibility testing, multilocus enzyme electrophoresis, ribotyping, plasmid analysis...... different counties, were assigned to 6 different strains. Seven out of these 8 isolates had the same plasmid restriction profile. In conclusion, penicillin and tetracycline resistant S. aureus occurring in dairy herds in Norway mainly seems to represent one particular strain that has achieved widespread...

  7. Plasmid segregation mechanisms

    DEFF Research Database (Denmark)

    Ebersbach, Gitte; Gerdes, Kenn; Charbon, Gitte Ebersbach

    2005-01-01

    Bacterial plasmids encode partitioning (par) loci that ensure ordered plasmid segregation prior to cell division. par loci come in two types: those that encode actin-like ATPases and those that encode deviant Walker-type ATPases. ParM, the actin-like ATPase of plasmid R1, forms dynamic filaments...... that segregate plasmids paired at mid-cell to daughter cells. Like microtubules, ParM filaments exhibit dynamic instability (i.e., catastrophic decay) whose regulation is an important component of the DNA segregation process. The Walker box ParA ATPases are related to MinD and form highly dynamic, oscillating...... filaments that are required for the subcellular movement and positioning of plasmids. The role of the observed ATPase oscillation is not yet understood. However, we propose a simple model that couples plasmid segregation to ParA oscillation. The model is consistent with the observed movement...

  8. SPP1-mediated plasmid transduction.

    OpenAIRE

    Canosi, U; Lüder, G; Trautner, T A

    1982-01-01

    The virulent Bacillus subtilis phage SPP1 transduces plasmid DNA. Plasmid-transducing phages contain only plasmid DNA. Such DNA represents a concatemer of monomeric plasmid molecules with the molecular weight of mature SPP1 DNA. Biological parameters of plasmid transduction are described.

  9. Transformation of microorganisms with the plasmid vector with the replicon from pAC1 from Acetobacter pasteurianus.

    Science.gov (United States)

    Grones, J; Turna, J

    1995-01-26

    A number of gram-negative and gram-positive bacteria species was screened for the expression of the gram-negative plasmid pACK5 and pACT72 with replicon of pAC1 plasmid from Acetobacter pasteurianus. As was described previously, both plasmids were expressed in Escherichia coli, Acetobacter pasteurianus, Acetobacter aceti, Shigella spp. and Citrobacter spp. Expressions of plasmids were successful in twelve species tested, Comamonas terrigena, Salmonella typhimurium, Serratia marcescens, Bacillus cereus, Bacillus megatericum, Bacillus subtilis, Lactobacillus helveticus, Micrococcus luteus, Sarcina lutea, Staphylococcus aureus, Staphylococcus epidermidis, Streptoccocus feacalis, and the stability of plasmid DNA was tested after cultivation in non-selective conditions. PMID:7832808

  10. Plasmids encoding therapeutic agents

    Science.gov (United States)

    Keener, William K.

    2007-08-07

    Plasmids encoding anti-HIV and anti-anthrax therapeutic agents are disclosed. Plasmid pWKK-500 encodes a fusion protein containing DP178 as a targeting moiety, the ricin A chain, an HIV protease cleavable linker, and a truncated ricin B chain. N-terminal extensions of the fusion protein include the maltose binding protein and a Factor Xa protease site. C-terminal extensions include a hydrophobic linker, an L domain motif peptide, a KDEL ER retention signal, another Factor Xa protease site, an out-of-frame buforin II coding sequence, the lacZ.alpha. peptide, and a polyhistidine tag. More than twenty derivatives of plasmid pWKK-500 are described. Plasmids pWKK-700 and pWKK-800 are similar to pWKK-500 wherein the DP178-encoding sequence is substituted by RANTES- and SDF-1-encoding sequences, respectively. Plasmid pWKK-900 is similar to pWKK-500 wherein the HIV protease cleavable linker is substituted by a lethal factor (LF) peptide-cleavable linker.

  11. Staphylococcus aureus and Pregnancy

    Science.gov (United States)

    Staphylococcus aureus and Pregnancy In every pregnancy, a woman starts out with a 3-5% chance of ... risk. This sheet talks about whether exposure to staphylococcus aureus may increase the risk for birth defects ...

  12. Staphylococcus aureus toxins

    OpenAIRE

    Otto, Michael

    2013-01-01

    Staphylococcus aureus is a dangerous pathogen that causes a variety of severe diseases. The virulence of S. aureus is defined by a large repertoire of virulence factors, among which secreted toxins play a preeminent role. Many S. aureus toxins damage biological membranes, leading to cell death. In particular, S. aureus produces potent hemolysins and leukotoxins. Among the latter, some were recently identified to lyse neutrophils after ingestion, representing an especially powerful weapon agai...

  13. Toxin plasmids of Clostridium perfringens.

    Science.gov (United States)

    Li, Jihong; Adams, Vicki; Bannam, Trudi L; Miyamoto, Kazuaki; Garcia, Jorge P; Uzal, Francisco A; Rood, Julian I; McClane, Bruce A

    2013-06-01

    In both humans and animals, Clostridium perfringens is an important cause of histotoxic infections and diseases originating in the intestines, such as enteritis and enterotoxemia. The virulence of this Gram-positive, anaerobic bacterium is heavily dependent upon its prolific toxin-producing ability. Many of the ∼16 toxins produced by C. perfringens are encoded by large plasmids that range in size from ∼45 kb to ∼140 kb. These plasmid-encoded toxins are often closely associated with mobile elements. A C. perfringens strain can carry up to three different toxin plasmids, with a single plasmid carrying up to three distinct toxin genes. Molecular Koch's postulate analyses have established the importance of several plasmid-encoded toxins when C. perfringens disease strains cause enteritis or enterotoxemias. Many toxin plasmids are closely related, suggesting a common evolutionary origin. In particular, most toxin plasmids and some antibiotic resistance plasmids of C. perfringens share an ∼35-kb region containing a Tn916-related conjugation locus named tcp (transfer of clostridial plasmids). This tcp locus can mediate highly efficient conjugative transfer of these toxin or resistance plasmids. For example, conjugative transfer of a toxin plasmid from an infecting strain to C. perfringens normal intestinal flora strains may help to amplify and prolong an infection. Therefore, the presence of toxin genes on conjugative plasmids, particularly in association with insertion sequences that may mobilize these toxin genes, likely provides C. perfringens with considerable virulence plasticity and adaptability when it causes diseases originating in the gastrointestinal tract. PMID:23699255

  14. Determination of aminoglycoside resistance in Staphylococcus aureus by DNA hybridization.

    OpenAIRE

    Dickgiesser, N; Kreiswirth, B N

    1986-01-01

    A method is described for identification of the genes conferring aminoglycoside resistance in Staphylococcus aureus by dot-blot and Southern blot techniques. As radioactive probes, fragments of plasmids pAT48, pUBH2, and pH13, carrying the genes for an aminocyclitol-3'-phosphotransferase, an aminocyclitol-4'-adenylyltransferase, and an aminocyclitol-2''-phosphotransferase-aminocyclitol-6'-acetyltransferase, respectively, were used.

  15. Persistence Mechanisms of Conjugative Plasmids

    DEFF Research Database (Denmark)

    Bahl, Martin Iain; Hansen, Lars H.; Sørensen, Søren Johannes

    2009-01-01

    maintenance in the host cell. These importantly include the ability to self-mobilize in a process termed conjugative transfer, which may occur across species barriers. Other plasmid stabilizing mechanisms include the multimer resolution system, active partitioning, and post-segregational-killing of plasmid......Are plasmids selfish parasitic DNA molecules or an integrated part of the bacterial genome? This chapter reviews the current understanding of the persistence mechanisms of conjugative plasmids harbored by bacterial cells and populations. The diversity and intricacy of mechanisms affecting the...... successful propagation and long-term continued existence of these extra-chromosomal elements is extensive. Apart from the accessory genetic elements that may provide plasmid-harboring cells a selective advantage, special focus is placed on the mechanisms conjugative plasmids employ to ensure their stable...

  16. Determining of antibiotic resistance profile inStaphylococcus aureus isolates

    Institute of Scientific and Technical Information of China (English)

    Hossein Motamedi; Hadis Mirzabeigi; Tahere Shirali

    2010-01-01

    Objective:To determine the pattern of antibiotic resistance amongStaphylococcus aureus (S. aureus) isolates from clinical specimens and to identify community-acquired methicillin-resistantStaphylococcus aureus(CA-MRSA)in specimens that have been collected from patients referring to one of the hospitals of Ahvaz.Methods:S. aureus isolates from a hospital in Ahvaz were screened for resistance to various antibiotics including methicillin. The susceptibility of the isolates was determined by Kirby-Bauer disc diffusion method. TheMRSA was also treated with ethidium bromide to find the origin of resistance.Results: Among the bacterial isolates, all of 11S. aureus were resistant to methicillin and cefixime,2 were resistant to ciprofloxacine,6 were resistant to tetracycline and the reminder were sensitive or intermediate to other antibiotics. The treated isolates were reminded resistant to methicillin and this suggested that the plasmid was not the origin of resistance in these isolates.Conclusions: These results showed that infection due toMRSA is widespread in Ahvaz and with respect to the spread of vancomycin resistance among MRSA and appearance of overwhelming infections. It is necessary to identify continuously the profile of antibiotic resistance amongS. aureus isolates in other regions and finding appropriate antibiotic for infection control and eradication.

  17. In Silico Detection and Typing of Plasmids using PlasmidFinder and Plasmid Multilocus Sequence Typing

    DEFF Research Database (Denmark)

    Carattoli, Alessandra; Zankari, Ea; García-Fernández, Aurora; Larsen, Mette Voldby; Lund, Ole; Villa, Laura; Aarestrup, Frank Møller; Hasman, Henrik

    2014-01-01

    genomes of multidrug-resistant Enterobacteriaceae species by the rapid detection of known plasmid types. Replicon sequences from 559 fully sequenced plasmids associated with the family Enterobacteriaceae in the NCBI nucleotide database were collected to build a consensus database for integration into a...... plasmid sequence types (STs) and new alleles and ST variants. In conclusion, testing of the two Web tools using both fully assembled plasmid sequences and WGS-generated draft genomes showed them to be able to detect a broad variety of plasmids that are often associated with antimicrobial resistance in......In the work presented here, we designed and developed two easy-to-use Web tools for in silico detection and characterization of whole-genome sequence (WGS) and whole-plasmid sequence data from members of the family Enterobacteriaceae. These tools will facilitate bacterial typing based on draft...

  18. Conjugative plasmids of Neisseria gonorrhoeae.

    Directory of Open Access Journals (Sweden)

    Emilia Pachulec

    Full Text Available Many clinical isolates of the human pathogen Neisseria gonorrhoeae contain conjugative plasmids. The host range of these plasmids is limited to Neisseria species, but presence of a tetracycline (tetM determinant inserted in several of these plasmids is an important cause of the rapid spread of tetracycline resistance. Previously plasmids with different backbones (Dutch and American type backbones and with and without different tetM determinants (Dutch and American type tetM determinants have been identified. Within the isolates tested, all plasmids with American or Dutch type tetM determinants contained a Dutch type plasmid backbone. This demonstrated that tetM determinants should not be used to differentiate between conjugal plasmid backbones. The nucleotide sequences of conjugative plasmids with Dutch type plasmid backbones either not containing the tetM determinant (pEP5233 or containing Dutch (pEP5289 or American (pEP5050 type tetM determinants were determined. Analysis of the backbone sequences showed that they belong to a novel IncP1 subfamily divergent from the IncP1alpha, beta, gamma, delta and epsilon subfamilies. The tetM determinants were inserted in a genetic load region found in all these plasmids. Insertion was accompanied by the insertion of a gene with an unknown function, and rearrangement of a toxin/antitoxin gene cluster. The genetic load region contains two toxin/antitoxins of the Zeta/Epsilon toxin/antitoxin family previously only found in Gram positive organisms and the virulence associated protein D of the VapD/VapX toxin/antitoxin family. Remarkably, presence of VapX of pJD1, a small cryptic neisserial plasmid, in the acceptor strain strongly increased the conjugation efficiency, suggesting that it functions as an antitoxin for the conjugative plasmid. The presence of the toxin and antitoxin on different plasmids might explain why the host range of this IncP1 plasmid is limited to Neisseria species. The isolated plasmids

  19. Plasmid profile of bacteria isolated from tears of HIV/AIDS patients

    Directory of Open Access Journals (Sweden)

    O B Ajayi

    2009-01-01

    Full Text Available Objective: The purpose of this study is to determine the presence and transfer of plasmids in bacteria isolated from tears of HIV/AIDS patients, their sensitivity and resistance to commercially available antibiotics. Design: This was a cross sectional experimental study. Materials and methods:One hundred tears samples from HIV/ AIDS patients and fifty tears samples from HIV/AIDS negative patients were screened for resistance to 14 commercially available antibiotics using disc diffusion method. Result: Three multiple antibiotics resistant strains of staphylococcus aureus and four multiple antibiotics resistance strains of Pseudomonas aeruginosa were identified. staphylococcus aureus strains showed 100% resistance to Ampiclox and erythromycin, 66.6% to Perfloxacin, amoxicillin and septrin, 33.33% to ciprofloxacin. Pseudomonas aeruginosa strains showed 100% resistance to streptomycin, amoxicillin, septrin and chloramphenicol. Only I strain of staphylococcus aureus showed presence of plasmid which was not transferable to Escherichia coli because of presence of disulphide cross--linked cell wall. Other strains of both staphylococcus aureus and Pseudomonas aeruginosa remained resistant after curing . Conclusion: Further studies are needed in this area to show if antibiotic resistance in HIV/AIDS positive patients could be as a result of plasmid as well as other factors.

  20. A plasmid in the archaebacterium Sulfolobus acidocaldarius

    OpenAIRE

    Yeats, Siobhán; McWilliam, Peter; Zillig, Wolfram

    1982-01-01

    A plasmid of mol. wt. ∼9 × 106 has been isolated from the archaebacterium Sulfolobus acidocaldarius strain B12. Plasmid production is induced by u.v. radiation. A copy of the plasmid is probably carried by the chromosome, integrated at a specific site. The entire plasmid, and also restriction fragments of it, has been cloned into Escherichia coli plasmid vectors, and the cleavage sites on the plasmid DNA of three restriction endonucleases have been mapped.

  1. Plasmid acquisition in microgravity

    Science.gov (United States)

    Juergensmeyer, Margaret A.; Juergensmeyer, Elizabeth A.; Guikema, James A.

    1995-01-01

    In microgravity, bacteria often show an increased resistance to antibiotics. Bacteria can develop resistance to an antibiotic after transformation, the acquisition of DNA, usually in the form of a plasmid containing a gene for resistance to one or more antibiotics. In order to study the capacity of bacteria to become resistant to antibiotics in microgravity, we have modified the standard protocol for transformation of Escherichia coli for use in the NASA-flight-certified hardware package, The Fluid Processing Apparatus (FPA). Here we report on the ability of E. coli to remain competent for long periods of time at temperatures that are readily available on the Space Shuttle, and present some preliminary flight results.

  2. Evolved plasmid-host interactions reduce plasmid interference cost.

    Science.gov (United States)

    Yano, Hirokazu; Wegrzyn, Katarznya; Loftie-Eaton, Wesley; Johnson, Jenny; Deckert, Gail E; Rogers, Linda M; Konieczny, Igor; Top, Eva M

    2016-09-01

    Antibiotic selection drives adaptation of antibiotic resistance plasmids to new bacterial hosts, but the molecular mechanisms are still poorly understood. We previously showed that a broad-host-range plasmid was poorly maintained in Shewanella oneidensis, but rapidly adapted through mutations in the replication initiation gene trfA1. Here we examined if these mutations reduced the fitness cost of TrfA1, and whether this was due to changes in interaction with the host's DNA helicase DnaB. The strains expressing evolved TrfA1 variants showed a higher growth rate than those expressing ancestral TrfA1. The evolved TrfA1 variants showed a lower affinity to the helicase than ancestral TrfA1 and were no longer able to activate the helicase at the oriV without host DnaA. Moreover, persistence of the ancestral plasmid was increased upon overexpression of DnaB. Finally, the evolved TrfA1 variants generated higher plasmid copy numbers than ancestral TrfA1. The findings suggest that ancestral plasmid instability can at least partly be explained by titration of DnaB by TrfA1. Thus under antibiotic selection resistance plasmids can adapt to a novel bacterial host through partial loss of function mutations that simultaneously increase plasmid copy number and decrease unfavorably high affinity to one of the hosts' essential proteins. PMID:27121483

  3. Cloning, expression, and mutagenesis of phosphatidylinositol-specific phospholipase C from Staphylococcus aureus: a potential staphylococcal virulence factor.

    OpenAIRE

    Daugherty, S; Low, M G

    1993-01-01

    Staphylococcus aureus secretes a phosphatidylinositol (PI)-specific phospholipase C (PI-PLC) which is able to hydrolyze the membrane lipid PI and membrane protein anchors containing glycosyl-PI. The gene for PI-PLC (plc) was cloned from S. aureus into Escherichia coli. Oligonucleotide probes based on partial protein sequence and polyclonal antibodies raised against the purified protein were used to identify positive clones. E. coli transformed with a plasmid containing the plc gene expressed ...

  4. Molecular Characterization of Resistance to Mupirocin in Methicillin-Susceptible and -Resistant Isolates of Staphylococcus aureus from Nasal Samples

    OpenAIRE

    Chaves, Fernando; García-Martínez, Jesus; Miguel, Sonia; Otero, Joaquín R.

    2004-01-01

    A total of 15 of 101 (14.8%) nasal methicillin-resistant Staphylococcus aureus (MRSA) isolates exhibited mupirocin resistance (Mupr) compared with 1 of 154 (0.6%) methicillin-susceptible Staphylococcus aureus isolates. A total of 14 (93%) isolates exhibiting high-level Mupr belonged to a single clone. Horizontal plasmid transfer and transmission of Mupr strains contribute to a high incidence of Mupr MRSA at our institution.

  5. Evaluation of phenotypic and genotypic methods for epidemiological typing of Staphylococcus aureus isolates from bovine mastitis in Denmark

    DEFF Research Database (Denmark)

    Aarestrup, Frank Møller; Wegener, H. C.; Rosdahl, V. T.

    1995-01-01

    The value of five different typing methods (antibiogram typing, biotyping, phage typing, plasmid profiling and restriction fragment length polymorphism of the gene encoding 16S and 23S ribosomal RNA (ribotyping)), in discriminating 105 Staphylococcus aureus strains from bovine milk samples obtained...... combination of phage, bio- or ribotyping or all three methods in combination are considered to be an efficient combination of typing methods for epidemiological investigation of S. aureus mastitis....

  6. Staphylococcus aureus Transcriptome Architecture

    DEFF Research Database (Denmark)

    Mäder, Ulrike; Nicolas, Pierre; Depke, Maren;

    2016-01-01

    Staphylococcus aureus is a major pathogen that colonizes about 20% of the human population. Intriguingly, this Gram-positive bacterium can survive and thrive under a wide range of different conditions, both inside and outside the human body. Here, we investigated the transcriptional adaptation of S...... to their dependence on the RNA polymerase sigma factors SigA or SigB, and allow identification of new potential targets for several known transcription factors. In particular, this study revealed a relatively low abundance of antisense RNAs in S. aureus, where they overlap only 6% of the coding genes, and only 19...... antisense RNAs not co-transcribed with other genes were found. Promoter analysis and comparison with Bacillus subtilis links the small number of antisense RNAs to a less profound impact of alternative sigma factors in S. aureus. Furthermore, we revealed that Rho-dependent transcription termination...

  7. Staphylococcus aureus CC398

    DEFF Research Database (Denmark)

    Price, Lance B.; Stegger, Marc; Hasman, Henrik;

    2012-01-01

    Since its discovery in the early 2000s, methicillin-resistant Staphylococcus aureus (MRSA) clonal complex 398 (CC398) has become a rapidly emerging cause of human infections, most often associated with livestock exposure. We applied whole-genome sequence typing to characterize a diverse collection...... of CC398 isolates (n = 89), including MRSA and methicillin-susceptible S. aureus (MSSA) from animals and humans spanning 19 countries and four continents. We identified 4,238 single nucleotide polymorphisms (SNPs) among the 89 core genomes. Minimal homoplasy (consistency index = 0.9591) was detected...

  8. Interactions between octopine and nopaline plasmids in Agrobacterium tumefaciens.

    OpenAIRE

    Hooykaas, P J; Den Dulk-Ras, H; Ooms, G.; Schilperoort, R.A.

    1980-01-01

    Transfer of octopine Ti plasmids to strains already carrying an octopine Ti plasmid was found to occur at the same (high) frequency as transfer to Ti plasmid lacking recipients, showing that resident Ti plasmids do not exhibit entry exclusion towards incoming Ti plasmids. The resident octopine Ti plasmid was lost by the recipient after the entrance of the incoming Ti plasmid, which is indicative of the incompatibility between the Ti plasmids. Octopine Ti plasmids were found to become establis...

  9. Effects of genes exerting growth inhibition and plasmid stability on plasmid maintenance.

    OpenAIRE

    Boe, L; Gerdes, K; Molin, S

    1987-01-01

    Plasmid stabilization mediated by the parA+ and parB+ genes of the R1 plasmid and the ccd+ and sop+ genes of the F plasmid was tested on a mini-R1 plasmid and a pBR322 plasmid derivative. The mini-R1 plasmid is thought to be unstably inherited owing to a low copy number and to random segregation of the plasmid at cell division, whereas cells harboring the pBR322 derivative used in this work are lost through competition with plasmid-free cells, mainly as a result of the shorter generation time...

  10. Recurrent Methicillin-Resistant Staphylococcus aureus Cutaneous Abscesses and Selection of Reduced Chlorhexidine Susceptibility during Chlorhexidine Use

    OpenAIRE

    Johnson, Ryan C.; Schlett, Carey D.; Crawford, Katrina; Lanier, Jeffrey B.; Merrell, D. Scott; Ellis, Michael W.

    2015-01-01

    We describe the selection of reduced chlorhexidine susceptibility during chlorhexidine use in a patient with two episodes of cutaneous USA300 methicillin-resistant Staphylococcus aureus abscess. The second clinical isolate harbors a novel plasmid that encodes the QacA efflux pump. Greater use of chlorhexidine for disease prevention warrants surveillance for resistance.

  11. Antiparallel plasmid-plasmid pairing may control P1 plasmid replication.

    OpenAIRE

    Abeles, A L; Austin, S J

    1991-01-01

    The copy number of the P1 plasmid replicon is stringently controlled, giving only one or two copies per newborn cell. Control is achieved by the action of the copy-control locus incA, which contains nine repeats of the 19-basepair binding site for the plasmid-encoded initiator protein RepA. A set of five similar repeats are present in the replication origin where RepA acts to trigger initiation. Using an in vitro replication system consisting of an Escherichia coli extract, the P1 origin as a...

  12. A plasmid in Legionella pneumophila.

    OpenAIRE

    Knudson, G B; Mikesell, P

    1980-01-01

    Sixteen strains from the six serogroups of Legionella pneumophila were examined for the presence of extrachromosomal genetic elements by a modified cleared lysate procedure, dye-buoyant centrifugation, and agarose gel electrophoresis. Two strains, Atlanta-1 and Atlanta-2 from serogroup II, each contained a plasmid of cryptic function with a molecular weight of ca. 30 megadaltons.

  13. Bacterial Plasmids in Antarctic Natural Microbial Assemblages

    OpenAIRE

    Kobori, Hiromi; Sullivan, Cornelius W.; Shizuya, Hiroaki

    1984-01-01

    Samples of psychrophilic and psychrotrophic bacteria were collected from sea ice, seawater, sediments, and benthic or ice-associated animals in McMurdo Sound, Antarctica. A total of 155 strains were isolated and tested for the presence of plasmids by DNA agarose gel electrophoresis. Thirty-one percent of the isolates carried at least one kind of plasmid. Bacterial isolates taken from sediments showed the highest plasmid incidence (42%), and isolates from seawater showed the lowest plasmid inc...

  14. Plasmid maintenance functions encoded on Dictyostelium discoideum nuclear plasmid Ddp1.

    OpenAIRE

    Hughes, J E; H. Kiyosawa; Welker, D L

    1994-01-01

    All of the plasmid-carried genes expressed during vegetative growth are essential for long-term maintenance of plasmid Ddp1 in the nucleus of Dictyostelium discoideum. Deletion of Ddp1 genes expressed only during development had no detectable effect on plasmid maintenance. Deletion of vegetatively expressed genes, either singly or in pairs, resulted in (i) a rapid loss of plasmid from cells grown in the absence of selection for plasmid retention, (ii) variation in the proportion of monomer to...

  15. Characterization of plasmids in Erwinia stewartii.

    OpenAIRE

    Coplin, D. L.; Rowan, R G; Chisholm, D A; Whitmoyer, R E

    1981-01-01

    Plasmids in 39 strains of Erwinia stewartii were examined by agarose gel electrophoresis. Most virulent strains had from 11 to 13 plasmids ranging in molecular mass from 2.8 to 210 megadaltons and contained plasmids of 210, 70, 49, 43, 29.5, 16.8, 8.8, and 2.8 megadaltons. Plasmids in strains SW2 and SS104 were characterized by both electron microscopy and agarose gel electrophoresis and may be useful as convenient references for sizing plasmids by electrophoresis. Specific size classes of pl...

  16. Regulation of protein A synthesis by the sar and agr loci of Staphylococcus aureus.

    OpenAIRE

    Cheung, A L; Eberhardt, K.; Heinrichs, J H

    1997-01-01

    The synthesis of protein A in Staphylococcus aureus is regulated by global regulatory loci such as sar and agr. Phenotypic data indicate that both sar and agr suppress protein A synthesis; like agr, sar also regulates protein A production at the transcriptional level. To determine the genetic requirement of sar in protein A suppression, we transformed shuttle plasmids containing various sar fragments into a sar mutant. Our results indicated that the 560-bp sarA transcript, or, more probably, ...

  17. Virulence Plasmids of Spore-Forming Bacteria.

    Science.gov (United States)

    Adams, Vicki; Li, Jihong; Wisniewski, Jessica A; Uzal, Francisco A; Moore, Robert J; McClane, Bruce A; Rood, Julian I

    2014-12-01

    Plasmid-encoded virulence factors are important in the pathogenesis of diseases caused by spore-forming bacteria. Unlike many other bacteria, the most common virulence factors encoded by plasmids in Clostridium and Bacillus species are protein toxins. Clostridium perfringens causes several histotoxic and enterotoxin diseases in both humans and animals and produces a broad range of toxins, including many pore-forming toxins such as C. perfringens enterotoxin, epsilon-toxin, beta-toxin, and NetB. Genetic studies have led to the determination of the role of these toxins in disease pathogenesis. The genes for these toxins are generally carried on large conjugative plasmids that have common core replication, maintenance, and conjugation regions. There is considerable functional information available about the unique tcp conjugation locus carried by these plasmids, but less is known about plasmid maintenance. The latter is intriguing because many C. perfringens isolates stably maintain up to four different, but closely related, toxin plasmids. Toxin genes may also be plasmid-encoded in the neurotoxic clostridia. The tetanus toxin gene is located on a plasmid in Clostridium tetani, but the botulinum toxin genes may be chromosomal, plasmid-determined, or located on bacteriophages in Clostridium botulinum. In Bacillus anthracis it is well established that virulence is plasmid determined, with anthrax toxin genes located on pXO1 and capsule genes on a separate plasmid, pXO2. Orthologs of these plasmids are also found in other members of the Bacillus cereus group such as B. cereus and Bacillus thuringiensis. In B. thuringiensis these plasmids may carry genes encoding one or more insecticidal toxins. PMID:26104459

  18. Homemade Site Directed Mutagenesis of Whole Plasmids

    Science.gov (United States)

    Laible, Mark; Boonrod, Kajohn

    2009-01-01

    Site directed mutagenesis of whole plasmids is a simple way to create slightly different variations of an original plasmid. With this method the cloned target gene can be altered by substitution, deletion or insertion of a few bases directly into a plasmid. It works by simply amplifying the whole plasmid, in a non PCR-based thermocycling reaction. During the reaction mutagenic primers, carrying the desired mutation, are integrated into the newly synthesized plasmid. In this video tutorial we demonstrate an easy and cost effective way to introduce base substitutions into a plasmid. The protocol works with standard reagents and is independent from commercial kits, which often are very expensive. Applying this protocol can reduce the total cost of a reaction to an eighth of what it costs using some of the commercial kits. In this video we also comment on critical steps during the process and give detailed instructions on how to design the mutagenic primers. PMID:19488024

  19. Plasmid and chromosome partitioning: surprises from phylogeny

    DEFF Research Database (Denmark)

    Gerdes, Kenn; Møller-Jensen, Jakob; Bugge Jensen, Rasmus

    2000-01-01

    Plasmids encode partitioning genes (par) that are required for faithful plasmid segregation at cell division. Initially, par loci were identified on plasmids, but more recently they were also found on bacterial chromosomes. We present here a phylogenetic analysis of par loci from plasmids...... and chromosomes from prokaryotic organisms. All known plasmid-encoded par loci specify three components: a cis-acting centromere-like site and two trans-acting proteins that form a nucleoprotein complex at the centromere (i.e. the partition complex). The proteins are encoded by two genes in an operon...... that is autoregulated by the par-encoded proteins. In all cases, the upstream gene encodes an ATPase that is essential for partitioning. Recent cytological analyses indicate that the ATPases function as adaptors between a host-encoded component and the partition complex and thereby tether plasmids and chromosomal...

  20. Uptake of plasmid deoxyribonucleic acid by Haemophilus.

    OpenAIRE

    Gromkova, R; Goodgal, S

    1981-01-01

    The uptake of circular and linear plasmid RSF0885 deoxyribonucleic acids, (DNAs) obtained from Haemophilus parainfluenzae 14, in both homologous and heterologous recipients was studied and compared with that of chromosomal DNA. High concentrations of divalent cations stimulated the uptake of either circular or linear plasmid DNA in H. parainfluenzae 14 competent cells but did not affect the uptake of chromosomal DNA. The biological activity of linear plasmid DNA was similar to that of circula...

  1. Construction of a Multiplex Promoter Reporter Platform to Monitor Staphylococcus aureus Virulence Gene Expression and the Identification of Usnic Acid as a Potent Suppressor of psm Gene Expression.

    Science.gov (United States)

    Gao, Peng; Wang, Yanli; Villanueva, Iván; Ho, Pak Leung; Davies, Julian; Kao, Richard Yi Tsun

    2016-01-01

    As antibiotic resistance becomes phenomenal, alternative therapeutic strategies for bacterial infections such as anti-virulence treatments have been advocated. We have constructed a total of 20 gfp-luxABCDE dual-reporter plasmids with selected promoters from S. aureus virulence-associated genes. The plasmids were introduced into various S. aureus strains to establish a gfp-lux based multiplex promoter reporter platform for monitoring S. aureus virulence gene expressions in real time to identify factors or compounds that may perturb virulence of S. aureus. The gene expression profiles monitored by luminescence correlated well with qRT-PCR results and extrinsic factors including carbon dioxide and some antibiotics were shown to suppress or induce the expression of virulence factors in this platform. Using this platform, sub-inhibitory ampicillin was shown to be a potent inducer for the expression of many virulence factors in S. aureus. Bacterial adherence and invasion assays using mammalian cells were employed to measure S. aureus virulence induced by ampicillin. The platform was used for screening of natural extracts that perturb the virulence of S. aureus and usnic acid was identified to be a potent repressor for the expression of psm. PMID:27625639

  2. pLS010 plasmid vector

    Science.gov (United States)

    Lacks, Sanford A.; Balganesh, Tanjore S.

    1988-01-01

    Disclosed is recombinant plasmid pLS101, consisting essentially of a 2.0 Kb malM gene fragment ligated to a 4.4 Kb T.sub.c r DNA fragment, which is particularly useful for transforming Gram-positive bacteria. This plasmid contains at least four restriction sites suitable for inserting exogeneous gene sequences. Also disclosed is a method for plasmid isolation by penicillin selection, as well as processes for enrichment of recombinant plasmids in Gram-positive bacterial systems.

  3. Chromate resistance plasmid in Pseudomonas fluorescens.

    OpenAIRE

    Bopp, L H; Chakrabarty, A M; Ehrlich, H. L.

    1983-01-01

    Chromate resistance of Pseudomonas fluorescens LB300, isolated from chromium-contaminated sediment in the upper Hudson River, was found to be plasmid specified. Loss of the plasmid (pLHB1) by spontaneous segregation or mitomycin C curing resulted in a simultaneous loss of chromate resistance. Subsequent transformation of such strains with purified pLHB1 plasmid DNA resulted in a simultaneous re-acquisition of the chromate resistance phenotype and the plasmid. When pLHB1 was transferred by con...

  4. ANALYSIS OF A MODEL OF PLASMID-BEARING, PLASMID-FREE COMPETITION IN A PULSED CHEMOSTAT

    OpenAIRE

    XIANGYUN SHI; XINYU SONG; XUEYONG ZHOU

    2006-01-01

    We introduce and study a chemostat model with plasmid-bearing, plasmid-free competition and impulsive effect. According to the stability analysis of the boundary periodic solution, we obtain the invasion threshold of the plasmid-free organism and plasmid-bearing organism. Furthermore, by using standard techniques of bifurcation theory, we prove the system has a positive τ-periodic solution, which shows that the impulsive effect destroys the equilibria of the unforced continuous system and ini...

  5. Plasmid ColVBtrp maintenance in Erwinia carotovora.

    OpenAIRE

    Schukin, N N

    1981-01-01

    Plasmid ColVBtrp maintenance in Erwinia carotovora cells was followed by measuring kinetics of elimination of plasmid genetic markers and loss of plasmid deoxyribonucleic acid. An E. carotovora mutant stably carrying plasmid ColVBtrp was isolated. Besides stable plasmid maintenance, the mutant showed altered sensitivity to male-specific phage MS2, sensitivity to drugs, and colony morphology.

  6. Linezolid resistant Staphylococcus aureus

    Directory of Open Access Journals (Sweden)

    Pavani Gandham

    2014-08-01

    Full Text Available Linezolid is the only antibiotic available as an oral formulation for resistant staphylococcal infections. It is effective in skin and soft tissue infections, nosocomial pneumonias including VAP, infective endocarditis and MRSA meningitis. It is also effective in the eradication of both nasal and throat colonization of MRSA. Its high bioavailability and post antibiotic effect, ease of switching to oral therapy during its use and the fact that it can be used in patients of all ages, also in patients with liver disease and poor kidney function and its increased effectiveness over glycopeptides makes this drug a precious drug in the treatment of resistant staphylococcal infections. Linezolid resistance in staphylococcus is defined as a linezolid MIC of and #8805;8 mg/L. Reported Linezolid resistance in India and elsewhere is 2-20%. There is clonal dissemination of Linezolid Resistant Staphylococcus aureus (LRSA within or across health care settings which demands continuous surveillance to determine the emergent risk of resistance strains and to establish guidelines for appropriate use. Clinical laboratories should confirm any LRSA preferably by a second method, prior to using linezolid for serious infections. Effective surveillance, more judicious use of this antibiotic, avoiding linezolid usage for empiric therapy in hospital acquired staphylococcus infections, optimization of the pharmacological parameters of the antibiotics in specific clinical situation, decreasing bacterial load by timely surgical debridement or drainage of collections, use of combination therapies would prevent the emergence of resistance to linezolid in staphylococcus aureus. [Int J Res Med Sci 2014; 2(4.000: 1253-1256

  7. The expression of a plasmid-specified exported protein causes structural plasmid instability in Bacillus subtilis

    NARCIS (Netherlands)

    Cordes, C.; Meima, R; Twiest, B; Kazemier, B; Venema, G; vanDijl, JM; Bron, S

    1996-01-01

    The rolling-circle plasmid pGP1 was used to study the effects of the expression of a plasmid-specified exported protein on structural plasmid stability in Bacillus subtilis. pGP1 contains a fusion between the Bacillus licheniformis penP gene, encoding a C-terminally truncated penicillinase, and the

  8. Identification, characterization and preliminary X-ray diffraction analysis of the rolling-circle replication initiator protein from plasmid pSTK1

    International Nuclear Information System (INIS)

    A proteolytically stable fragment of a plasmid replication initiation protein from the thermophile G. stearothermophilus has been biochemically characterized, crystallized and diffraction data collected to a resolution of 2.5 Å. Antibiotic resistance in bacterial pathogens poses an ever-increasing risk to human health. In antibiotic-resistant strains of Staphylococcus aureus this resistance often resides in extra-chromosomal plasmids, such as those of the pT181 family, which replicate via a rolling-circle mechanism mediated by a plasmid-encoded replication initiation protein. Currently, there is no structural information available for the pT181-family Rep proteins. Here, the crystallization of a catalytically active fragment of a homologous replication initiation protein from the thermophile Geobacillus stearothermophilus responsible for the replication of plasmid pSTK1 is reported. Crystals of the RepSTK1 fragment diffracted to a resolution of 2.5 Å and belonged to space group P212121

  9. Genomic organization of a vancomycin-resistant staphylococcus aureus

    International Nuclear Information System (INIS)

    Objective: To study the genomic organization of vancomycin resistance in a local isolate of vancomycin resistant Staphylococcus aureus (VRSA). Study Design: Experimental study. Place and Duration of Study: Department of Microbiology, University of Karachi, January 2008 through December 2010. Methodology: A vancomycin-resistant Staphylococcus aureus (VRSA-CP2) isolate (MIC 16 mu g/ml) was isolated from a local hospital of Karachi. Species identification was confirmed by Gram staining, standard biochemical tests and PCR amplification of the nuc gene. The vancomycin MIC was re-confirmed by E-test. For the genetic determination of vancomycin resistance, in-vitro amplification of vanA cassette was performed by using plasmid DNA of CP2, CP2's transformant as template on MWG Thermo-Cycler. Amplified products of vanR, vanS, vanH, vanA, vanY, orf2, orf1D, orf2E, orf-Rev and IS element genes were subjected to Sanger's electrophoresis based sequence determination using specific primers. The Basic Local Alignment Search Tool (BLAST) algorithm was used to identify sequences in GenBank with similarities to the vanA cassette genes. Results: The vancomycin-resistant isolate CP2 was found to be resistant to oxacillin, chloramphenicol, erythromycin, rifampicin, gentamicin, tetracycline and ciprofloxacin, as well. The isolate CP2 revealed four bands: one of large molecular size approx 56.4 kb and three of small size approx 6.5 kb, approx 6.1 kb and approx 1.5 kb by agarose gel electrophoresis indicating the presence of 3 plasmids. The plasmid DNA of isolate CP2 was analyzed by PCR for the presence of the van cassettes with each of the vanA , vanB and vanC specific primers. It carried vanA cassette, which comprises of vanR, vanS, vanH, vanA, vanY, and orf2. The vanA cassette of isolate CP2 also carried an insertion element (IS). However, it did not show the PCR product for orf1. Vancomycin resistance was successfully transferred from the donor CP2 to a vancomycin-sensitive recipient S

  10. Isolation of Plasmid DNA from Butyrivibrio fibrisolvens†

    OpenAIRE

    Teather, Ronald M.

    1982-01-01

    A procedure based on successive precipitation of cell lysates with sodium dodecyl sulfate-NaCl and polyethylene glycol 6000 was developed which allows the isolation of plasmid DNA from Butyrivibrio fibrisolvens. A survey of B. fibrisolvens strains isolated from the bovine rumen showed that plasmids are a common feature of this species.

  11. Plasmid deoxyribonucleic acid replication in Streptomyces griseus.

    OpenAIRE

    Xue, Y.; Zhuang, Z.; Zhu, Y.; Xu, Y.; Dong, K.

    1981-01-01

    A series of electron micrographs showing the presence of different molecular forms representing various replication stages of plasmid deoxyribonucleic acid from Streptomyces griseus was obtained. Based upon an analysis of these electron micrographs, a tentative model for plasmid deoxyribonucleic acid replication in S. griseus is proposed.

  12. Staphylococcus aureus triggered reactive arthritis.

    OpenAIRE

    Siam, A R; M. Hammoudeh

    1995-01-01

    OBJECTIVES--To report two patients who developed reactive arthritis in association with Staphylococcus aureus infection. METHODS--A review of the case notes of two patients. RESULTS--Two adult female patients have developed sterile arthritis in association with Staph aureus infection. The first patient has had two episodes of arthritis; the first followed olecranon bursitis, the second followed infection of a central venous catheter used for dialysis. The second patient developed sterile arth...

  13. Staphylococcus aureus and sore nipples.

    OpenAIRE

    Livingstone, V. H.; Willis, C. E.; Berkowitz, J

    1996-01-01

    OBJECTIVE: To correlate clinical symptoms and signs of sore nipples with the presence of Staphylococcus aureus and to determine the probability of mothers having S aureus-infected nipples when these local symptoms and signs are found. DESIGN: Two cohorts of consecutive patients were enrolled regardless of presenting complaint. A questionnaire was administered to determine the presence and severity of sore nipples. Objective findings on breast examination were documented. A nipple swab was tak...

  14. Plasmid-mediated horizontal gene transfer is a coevolutionary process

    OpenAIRE

    Harrison, Ellie; Brockhurst, Michael A

    2012-01-01

    Conjugative plasmids are key agents of horizontal gene transfer (HGT) that accelerate bacterial adaptation by vectoring ecologically important traits between strains and species. However, although many conjugative plasmids carry beneficial traits, all plasmids exert physiological costs-of-carriage on bacteria. The existence of conjugative plasmids, therefore, presents a paradox because non-beneficial plasmids should be lost to purifying selection, whereas beneficial genes carried on plasmids ...

  15. Bifurcation Analysis of a Chemostat Model of Plasmid-Bearing and Plasmid-Free Competition with Pulsed Input

    OpenAIRE

    Zhong Zhao; Baozhen Wang; Liuyong Pang; Ying Chen

    2014-01-01

    A chemostat model of plasmid-bearing and plasmid-free competition with pulsed input is proposed. The invasion threshold of the plasmid-bearing and plasmid-free organisms is obtained according to the stability of the boundary periodic solution. By use of standard techniques of bifurcation theory, the periodic oscillations in substrate, plasmid-bearing, and plasmid-free organisms are shown when some conditions are satisfied. Our results can be applied to control bioreactor aimed at producing co...

  16. Absence in Bacillus subtilis and Staphylococcus aureus of the sequence-specific deoxyribonucleic acid methylation that is conferred in Escherichia coli K-12 by the dam and dcm enzymes.

    OpenAIRE

    Dreiseikelmann, Brigitte; Wackernagel, Wilfried

    1981-01-01

    Restriction analysis of plasmid pHV14 deoxyribonucleic acid isolated from Escherichia coli K-12, Bacillus subtilis, and Staphylococcus aureus with restriction endonucleases MboI, Sau3AI, and EcoRII was used to study the methylation of those nucleotide sequences which in E. coli contain the major portions of N6-methyladenine and 5-methylcytosine. The results showed that neither B. subtilis nor S. aureus methylates deoxyribonucleic acid at the same sites and nucleotides which are recognized and...

  17. Nucleotide sequence and phylogeny of the tet (L) tetracycline resistance determinant encoded by the plasmid pSTE1 from Staphylococcus hyicus

    DEFF Research Database (Denmark)

    Schwarz, S.; Cardoso, M.; Wegener, Henrik Caspar

    1992-01-01

    amino acid sequences of the pSTE1-encoded Tet from S. hyicus and the previously sequenced Tet K variants from Staphylococcus aureus, Tet L variants from Bacillus cereus, Bacillus stearothermophilus, and Bacillus subtilis, Tet M variants from Steptococcus faecalis and Staphylococcus aureus as well as Tet...... variants on one hand and the Tet K and Tet L variants on the other hand. The pSTE1-encoded Tet proved to be closely related to the Tet L proteins originally found on small Bacillus plasmids. The observed extensive similarities in the nucleotide sequences of the tet genes and in the deduced Tet amino acid...

  18. PcrA function in plasmid replication

    OpenAIRE

    Chisty, L. T.

    2014-01-01

    PcrA is a DNA helicase involved in unwinding plasmi ds as a part of a complex in asymmetric rolling - circle replication of certain plasmids carrying antibiotic resistance genes. PcrA translocates on single stranded DNA by coupling ATP hydrolysis to movement on DNA. Initiator protein, RepD is required to nick supercoiled plasmid site - specifically and open an ssDNA stretch that PcrA can bind. The presence of RepD is needed throughout plasmid unwinding to maintain processivity. Using fluo...

  19. Plasmids as Tools for Containment.

    Science.gov (United States)

    García, José L; Díaz, Eduardo

    2014-10-01

    Active containment systems are a major tool for reducing the uncertainty associated with the introduction of monocultures, genetically engineered or not, into target habitats for a large number of biotechnological applications (e.g., bioremediation, bioleaching, biopesticides, biofuels, biotransformations, live vaccines, etc.). While biological containment reduces the survival of the introduced organism outside the target habitat and/or upon completion of the projected task, gene containment strategies reduce the lateral spread of the key genetic determinants to indigenous microorganisms. In fundamental research, suicide circuits become relevant tools to address the role of gene transfer, mainly plasmid transfer, in evolution and how this transfer contributes to genome plasticity and to the rapid adaptation of microbial communities to environmental changes. Many lethal functions and regulatory circuits have been used and combined to design efficient containment systems. As many new genomes are being sequenced, novel lethal genes and regulatory elements are available, e.g., new toxin-antitoxin modules, and they could be used to increase further the current containment efficiencies and to expand containment to other organisms. Although the current containment systems can increase the predictability of genetically modified organisms in the environment, containment will never be absolute, due to the existence of mutations that lead to the appearance of surviving subpopulations. In this sense, orthogonal systems (xenobiology) appear to be the solution for setting a functional genetic firewall that will allow absolute containment of recombinant organisms. PMID:26104372

  20. Plasmid DNA fermentation strategies: influence on plasmid stability and cell physiology.

    Science.gov (United States)

    Silva, Filomena; Queiroz, João A; Domingues, Fernanda C

    2012-03-01

    In order to provide sufficient pharmaceutical-grade plasmid DNA material, it is essential to gain a comprehensive knowledge of the bioprocesses involved; so, the development of protocols and techniques that allow a fast monitoring of process performance is a valuable tool for bioprocess design. Regarding plasmid DNA production, the metabolic stress of the host strain as well as plasmid stability have been identified as two of the key parameters that greatly influence plasmid DNA yields. The present work describes the impact of batch and fed-batch fermentations using different C/N ratios and different feeding profiles on cell physiology and plasmid stability, investigating the potential of these two monitoring techniques as valuable tools for bioprocess development and design. The results obtained in batch fermentations showed that plasmid copy number values suffered a pronounced increase at the end of almost all fermentation conditions tested. Regarding fed-batch fermentations, the strategies with exponential feeding profiles, in contrast with those with constant feeding, showed higher biomass and plasmid yields, the maximum values obtained for these two parameters being 95.64 OD(600) and 344.3 mg plasmid DNA (pDNA)/L, respectively, when using an exponential feed rate of 0.2 h(-1). Despite the results obtained, cell physiology and plasmid stability monitoring revealed that, although higher pDNA overall yields were obtained, this fermentation exhibited lower plasmid stability and percentage of viable cells. In conclusion, this study allowed clarifying the bioprocess performance based on cell physiology and plasmid stability assessment, allowing improvement of the overall process and not only plasmid DNA yield and cell growth. PMID:22089386

  1. Enhancement of plasmid-mediated gene therapy for muscular dystrophy by directed plasmid integration

    OpenAIRE

    Bertoni, Carmen; Jarrahian, Sohail; Wheeler, Thurman M.; LI, YINING; Olivares, Eric C.; Michele P Calos; Rando, Thomas A.

    2005-01-01

    Plasmid-mediated gene therapy can restore dystrophin expression in skeletal muscle in the mdx mouse, a model of Duchenne muscular dystrophy. However, sufficient long-term expression and distribution of dystrophin remain a hurdle for translating this technology into a viable treatment for Duchenne muscular dystrophy. To improve plasmid-mediated gene therapy for muscle diseases, we studied the effects of targeted plasmid integration using a phage integrase (φC31) that can mediate the integratio...

  2. Collagen binding to Staphylococcus aureus

    International Nuclear Information System (INIS)

    Staphylococcus aureus can bind soluble collagen in a specific, saturable manner. We have previously shown that some variability exists in the degree of collagen binding between different strains of heat-killed, formaldehyde-fixed S. aureus which are commercially available as immunologic reagents. The present study demonstrates that live S. aureus of the Cowan 1 strain binds amounts of collagen per organism equivalent to those demonstrated previously in heat-killed, formaldehyde-fixed bacteria but has an affinity over 100 times greater, with Kd values of 9.7 X 10(-11) M and 4.3 X 10(-8) M for live and heat-killed organisms, respectively. Studies were also carried out with S. aureus killed by ionizing radiation, since this method of killing the organism seemed less likely to alter the binding moieties on the surface than did heat killing. Bacteria killed by exposure to gamma radiation bound collagen in a manner essentially indistinguishable from that of live organisms. Binding of collagen to irradiated cells of the Cowan 1 strain was rapid, with equilibrium reached by 30 min at 22 degrees C, and was fully reversible. The binding was not inhibited by fibronectin, fibrinogen, C1q, or immunoglobulin G, suggesting a binding site for collagen distinct from those for these proteins. Collagen binding was virtually eliminated in trypsin-treated organisms, indicating that the binding site has a protein component. Of four strains examined, Cowan 1 and S. aureus ATCC 25923 showed saturable, specific binding, while strains Woods and S4 showed a complete lack of binding. These results suggest that some strains of S. aureus contain high-affinity binding sites for collagen. While the number of binding sites per bacterium varied sixfold in the two collagen-binding strains, the apparent affinity was similar

  3. Elimination of multicopy plasmid R6K by bleomycin.

    OpenAIRE

    Attfield, P V; Pinney, R. J.

    1985-01-01

    Bleomycin eliminated multicopy plasmid R6K from growing cells of Escherichia coli AB1157 but failed to cure either of the low-copy plasmids R1 or R46. Measurements of R6K-encoded beta-lactamase and of covalently closed plasmid DNA indicated that the drug causes a progressive reduction in plasmid copy number.

  4. Behavior of IncQ Plasmids in Agrobacterium tumefaciens

    NARCIS (Netherlands)

    Hille, Jacques; Schilperoort, Rob

    1981-01-01

    Inc-Q plasmids were introduced into Agrobacterium tumefuciens, by mobilization from Escherichia coli with an Inc-P plasmid, or by transformation with purified plasmid DNA. It was found that they were stably maintained. The presence of an Inc-Q plasmid did not influence tumorigenicity. These results

  5. Curing of plasmid pXO1 from Bacillus anthracis using plasmid incompatibility.

    Directory of Open Access Journals (Sweden)

    Xiankai Liu

    Full Text Available The large plasmid pXO1 encoding the anthrax toxin is important for the virulence of Bacillus anthracis. It is essential to cure pXO1 from B. anthracis to evaluate its role in the pathogenesis of anthrax infection. Because conventional methods for curing plasmids (e.g., curing agents or growth at elevated temperatures can induce mutations in the host chromosomal DNA, we developed a specific and reliable method to eliminate pXO1 from B. anthracis using plasmid incompatibility. Three putative replication origins of pXO1 were inserted into a temperature-sensitive plasmid to generate three incompatible plasmids. One of the three plasmids successfully eliminated the large plasmid pXO1 from B. anthracis vaccine strain A16R and wild type strain A16. These findings provided additional information about the replication/partitioning of pXO1 and demonstrated that introducing a small incompatible plasmid can generate plasmid-cured strains of B. anthracis without inducing spontaneous mutations in the host chromosome.

  6. Positive epistasis between co-infecting plasmids promotes plasmid survival in bacterial populations

    OpenAIRE

    San Millan, Alvaro; Heilbron, Karl; MacLean, R. Craig

    2014-01-01

    Plasmids have a key role in the horizontal transfer of genes among bacteria. Although plasmids are catalysts for bacterial evolution, it is challenging to understand how they can persist in bacterial populations over the long term because of the burden they impose on their hosts (the ‘plasmid paradox'). This paradox is especially perplexing in the case of ‘small' plasmids, which are unable to self-transfer by conjugation. Here, for the first time, we investigate how interactions between co-in...

  7. Mini-F plasmid genes that couple host cell division to plasmid proliferation.

    OpenAIRE

    Ogura, T; Hiraga, S

    1983-01-01

    A mechanism for stable maintenance of plasmids, besides the replication and partition mechanisms, has been found to be specified by genes of a mini-F plasmid. An oriC plasmid carrying both a mini-F segment necessary for partition [coordinates 46.4-49.4 kilobase pairs (kb) on the F map] and another segment (42.9-43.6 kb), designated ccd (coupled cell division), is more stably maintained than are oriC plasmids carrying only the partition segment; the stability is comparable to that of the paren...

  8. Adaptive Plasmid Evolution Results in Host-Range Expansion of a Broad-Host-Range Plasmid

    OpenAIRE

    De Gelder, Leen; Williams, Julia J.; Ponciano, José M; Sota, Masahiro; Eva M. Top

    2008-01-01

    Little is known about the range of hosts in which broad-host-range (BHR) plasmids can persist in the absence of selection for plasmid-encoded traits, and whether this “long-term host range” can evolve over time. Previously, the BHR multidrug resistance plasmid pB10 was shown to be highly unstable in Stenotrophomonas maltophilia P21 and Pseudomonas putida H2. To investigate whether this plasmid can adapt to such unfavorable hosts, we performed evolution experiments wherein pB10 was maintained ...

  9. Host-adaptive evolution of Staphylococcus aureus

    OpenAIRE

    Lowder, Bethan Victoria

    2011-01-01

    Staphylococcus aureus is a notorious human pathogen associated with severe nosocomial and community-acquired infections. In addition, S. aureus is a major cause of animal diseases including skeletal infections of poultry and bovine and ovine mastitis, which are a large economic burden on the broiler chicken and dairy farming industries. The population structure of S. aureus associated with humans has been well studied. However, despite the prevalence of S. aureus infections in ...

  10. Characterization of a Haemophilus ducreyi mobilizing plasmid.

    OpenAIRE

    McNicol, P J; Albritton, W L; Ronald, A R

    1986-01-01

    The OriV site of Haemophilus ducreyi mobilizing plasmid pHD147, determined by replication in Escherichia coli polA, is located close to the OriT site. The OriT site, located by recombination-proficient and -deficient cells, and the OriV site map in a region of pHD147 homologous to the beta-lactamase-specifying plasmids of H. ducreyi and Neisseria gonorrhoeae.

  11. Plasmid and chromosome segregation in prokaryotes

    DEFF Research Database (Denmark)

    Møller-Jensen, Jakob; Bugge Jensen, Rasmus; Gerdes, Kenn

    2000-01-01

    Recent major advances in the understanding of prokaryotic DNA segregation have been achieved by using fluorescence microscopy to visualize the localization of cellular components. Plasmids and bacterial chromosomes are partitioned in a highly dynamic fashion, suggesting the presence of a mitotic......-like apparatus in prokaryotes. The identification of chromosomal homologues of the well-characterized plasmid partitioning genes indicates that there could be a general mechanism of bacterial DNA partitioning. Udgivelsesdato: July 1...

  12. Plasmid Evolution and Interaction between the Plasmid Addiction Stability Systems of Two Related Broad-Host-Range IncQ-Like Plasmids

    OpenAIRE

    Deane, Shelly M.; Rawlings, Douglas E

    2004-01-01

    Plasmid pTC-F14 contains a plasmid stability system called pas (plasmid addiction system), which consists of two proteins, a PasA antitoxin and a PasB toxin. This system is closely related to the pas of plasmid pTF-FC2 (81 and 72% amino acid identity for PasA and PasB, respectively) except that the pas of pTF-FC2 contains a third protein, PasC. As both pTC-F14 and pTF-FC2 are highly promiscuous broad-host-range plasmids isolated from bacteria that share a similar ecological niche, the plasmid...

  13. Genome Sequence of the Clinical Isolate Staphylococcus aureus subsp. aureus Strain UAMS-1

    OpenAIRE

    Sassi, Mohamed; Sharma, Deepak; Brinsmade, Shaun ,; Felden, Brice; Augagneur, Yoann

    2015-01-01

    We report here the draft genome sequence of Staphylococcus aureus subsp. aureus strain UAMS-1. UAMS-1 is a virulent oxacillin-susceptible clinical isolate. Its genome is composed of 2,763,963 bp and will be useful for further gene expression analysis using RNA sequencing (RNA-seq) technology. S taphylococcus aureus is an opportunistic human bacterial pathogen responsible for nosocomial and community-associated infections. S. aureus subsp. aureus strain UAMS-1 was originally isolated from the ...

  14. Relationship between plasmid content and auxotype in Neisseria gonorrhoeae isolates.

    OpenAIRE

    Dillon, J R; Pauzé, M

    1981-01-01

    One hundred and forty strains of Neisseria gonorrhoeae, representing 12 different auxotype groups, were examined for differences in plasmid content. Most auxotype groups harbored a phenotypically cryptic 2,6-megadalton plasmid; a few groups also carried a 24.5-megadalton plasmid which has been previously characterized as a transfer plasmid. However, isolates of the proline-, citrulline-, and uracil-requiring (PCU-) auxotype were consistently free of plasmids. The correlation between auxotype ...

  15. Plasmid stability and maintenance of copy number using natural marker

    OpenAIRE

    Hamzah Basil Mohammed; Sudhakar Malla

    2015-01-01

    Present study was conducted to study the plasmid stability with the help of natural plasmid isolated from the bacteria which lodges the ink gland of the sea squid and emits bioluminescence. Isolated bacterial strain was identified by using 16srRNA sequencing and its plasmid DNA was used for the experimental studies. The plasmid is found to be responsible for the bioluminescence. The stability of this plasmid was studied in shake flask method using the different sugar sources (Gluc...

  16. Plasmids spread very fast in heterogeneous bacterial communities.

    OpenAIRE

    Dionisio, Francisco; Matic, Ivan; Radman, Miroslav; Rodrigues, Olivia R; Taddei, François

    2002-01-01

    Conjugative plasmids can mediate gene transfer between bacterial taxa in diverse environments. The ability to donate the F-type conjugative plasmid R1 greatly varies among enteric bacteria due to the interaction of the system that represses sex-pili formations (products of finOP) of plasmids already harbored by a bacterial strain with those of the R1 plasmid. The presence of efficient donors in heterogeneous bacterial populations can accelerate plasmid transfer and can spread by several order...

  17. Plasmid Segregation: Spatial Awareness at the Molecular Level

    DEFF Research Database (Denmark)

    Møller-Jensen, Jakob; Gerdes, Kenn

    2007-01-01

    In bacteria, low-copy number plasmids ensure their stable inheritance by partition loci (par), which actively distribute plasmid replicates to each side of the cell division plane. Using time-lapse fluorescence microscopic tracking of segregating plasmid molecules, a new study provides novel...... insight into the workings of the par system from Escherichia coli plasmid R1. Despite its relative simplicity, the plasmid partition spindle shares characteristics with the mitotic machinery of eukaryotic cells....

  18. Mobilization of Thiobacillus ferrooxidans plasmids among Escherichia coli strains.

    OpenAIRE

    Rawlings, D. E.; Woods, D R

    1985-01-01

    Nonconjugative Thiobacillus ferrooxidans plasmids were mobilized at high frequencies among Escherichia coli strains by the IncP plasmid RP4 and at low frequencies by the IncN plasmid R46, but not by the IncW plasmid pSa. The mobilization region of a nonconjugative T. ferrooxidans plasmid was located on a 5.3-kilobase T. ferrooxidans DNA fragment.

  19. Inducible Escherichia coli fermentation for increased plasmid DNA production.

    Science.gov (United States)

    Carnes, Aaron E; Hodgson, Clague P; Williams, James A

    2006-11-01

    Bacterial plasmids are the vectors of choice for DNA vaccines and short-term gene therapeutics. Growing plasmid DNA by microbial (Escherichia coli) fermentation is usually combined with alkaline lysis/chromatography methods of purification. To date, typical plasmid fermentation media and processes result in yields of 100-250 mg of plasmid DNA/l of culture medium, using standard high-copy pUC origin-containing plasmids. In order to address this initial and yield-limiting upstream step, we identified novel fermentation control parameters for fed-batch fermentation. The resulting fermentation strategies significantly increased specific plasmid yield with respect to cell mass while enhancing plasmid integrity and maintaining supercoiled DNA content. Fed-batch fermentation yield exceeding 1000 mg of plasmid DNA/l was obtained after reduction of plasmid-mediated metabolic burden during growth, and yields up to 1500 mg of plasmid DNA/l have been achieved with optimized plasmid backbones. Interestingly, by inducing high plasmid levels after sufficient biomass accumulation at low temperature and restricted growth, cells were able to tolerate significantly higher plasmid quantities than cells grown by conventional processes. This 5-10-fold increase in plasmid yield dramatically decreases plasmid manufacturing costs and improves the effectiveness of downstream purification by reducing the fraction of impurities. PMID:16819941

  20. Conservation of Salmonella typhimurium virulence plasmid maintenance regions among Salmonella serovars as a basis for plasmid curing.

    OpenAIRE

    Tinge, S A; Curtiss, R

    1990-01-01

    The association of large plasmids with virulence in invasive Salmonella serovars has led to a number of studies designed to uncover the role of these plasmids in virulence. This study addresses two aspects of virulence-associated plasmids. The first is the distribution of the replication and maintenance regions among the plasmids of different Salmonella serovars, and the second is the use of the conserved virulence plasmid par region to provide a rapid method for eliminating the virulence pla...

  1. Staphylococcus aureus and hand eczema severity

    DEFF Research Database (Denmark)

    Haslund, P; Bangsgaard, N; Jarløv, J O;

    2009-01-01

    BACKGROUND: The role of bacterial infections in hand eczema (HE) remains to be assessed. OBJECTIVES: To determine the prevalence of Staphylococcus aureus in patients with HE compared with controls, and to relate presence of S. aureus, subtypes and toxin production to severity of HE. METHODS......: Bacterial swabs were taken at three different visits from the hand and nose in 50 patients with HE and 50 controls. Staphylococcus aureus was subtyped by spa typing and assigned to clonal complexes (CCs), and isolates were tested for exotoxin-producing S. aureus strains. The Hand Eczema Severity Index...... was used for severity assessment. RESULTS: Staphylococcus aureus was found on the hands in 24 patients with HE and four controls (P aureus was found to be related to increased severity of the eczema (P aureus types on the hands...

  2. Phenotypic and molecular characterization of Staphylococcus aureus isolates expressing low- and high-level mupirocin resistance in Nigeria and South Africa

    Directory of Open Access Journals (Sweden)

    Udo Edet E

    2009-01-01

    Full Text Available Abstract Background Mupirocin is a topical antimicrobial agent which is used for the treatment of skin and postoperative wound infections, and the prevention of nasal carriage of methicillin-resistant Staphylococcus aureus (MRSA. However, the prevalence of mupirocin resistance in S. aureus, particularly in MRSA, has increased with the extensive and widespread use of this agent in hospital settings. This study characterized low- and high-level mupirocin-resistant S. aureus isolates obtained from Nigeria and South Africa. Methods A total of 17 mupirocin-resistant S. aureus isolates obtained from two previous studies in Nigeria and South Africa, were characterized by antibiogram, PCR-RFLP of the coagulase gene and PFGE. High-level mupirocin resistant isolates were confirmed by PCR detection of the mupA gene. The genetic location of the resistance determinants was established by curing and transfer experiments. Results All the low-level mupirocin resistant isolates were MRSA and resistant to gentamicin, tetracycline and trimethoprim. PFGE identified a major clone in two health care institutions located in Durban and a health care facility in Pietermaritzburg, Greytown and Empangeni. Curing and transfer experiments indicated that high-level mupirocin resistance was located on a 41.1 kb plasmid in the South African strain (A15. Furthermore, the transfer of high-level mupirocin resistance was demonstrated by the conjugative transfer of the 41.1 kb plasmid alone or with the co-transfer of a plasmid encoding resistance to cadmium. The size of the mupirocin-resistance encoding plasmid in the Nigerian strain (35 IBA was approximately 35 kb. Conclusion The emergence of mupirocin-resistant S. aureus isolates in Nigeria and South Africa should be of great concern to medical personnel in these countries. It is recommended that methicillin-susceptible S. aureus (MSSA and MRSA should be routinely tested for mupirocin resistance even in facilities where the agent

  3. Genome sequence of type strain of Staphylococcus aureus subsp. aureus

    OpenAIRE

    Kim, Bong-Soo; Yi, Hana; Chun, Jongsik; Cha, Chang-Jun

    2014-01-01

    Background Staphylococcus aureus is a pathogen that causes food poisoning and community-associated infection with antibiotic resistance. This species is an indigenous intestinal microbe found in infants and not found in adult intestine. The relatively small genome size and rapid evolution of antibiotic resistance genes in the species have been drawing an increasing attention in public health. To extend our understanding of the species and use the genome data for comparative genomic studies, w...

  4. BioShuttle-mediated Plasmid Transfer

    Directory of Open Access Journals (Sweden)

    Klaus Braun, Leonie von Brasch, Ruediger Pipkorn, Volker Ehemann, Juergen Jenne, Herbert Spring, Juergen Debus, Bernd Didinger, Werner Rittgen, Waldemar Waldeck

    2007-01-01

    Full Text Available An efficient gene transfer into target tissues and cells is needed for safe and effective treatment of genetic diseases like cancer. In this paper, we describe the development of a transport system and show its ability for transporting plasmids. This non-viral peptide-based BioShuttle-mediated transfer system consists of a nuclear localization address sequence realizing the delivery of the plasmid phNIS-IRES-EGFP coding for two independent reporter genes into nuclei of HeLa cells. The quantification of the transfer efficiency was achieved by measurements of the sodium iodide symporter activity. EGFP gene expression was measured with Confocal Laser Scanning Microscopy and quantified with biostatistical methods by analysis of the frequency of the amplitude distribution in the CLSM images. The results demonstrate that the “BioShuttle”-Technology is an appropriate tool for an effective transfer of genetic material carried by a plasmid.

  5. BioShuttle-mediated Plasmid Transfer

    Science.gov (United States)

    Braun, Klaus; von Brasch, Leonie; Pipkorn, Ruediger; Ehemann, Volker; Jenne, Juergen; Spring, Herbert; Debus, Juergen; Didinger, Bernd; Rittgen, Werner; Waldeck, Waldemar

    2007-01-01

    An efficient gene transfer into target tissues and cells is needed for safe and effective treatment of genetic diseases like cancer. In this paper, we describe the development of a transport system and show its ability for transporting plasmids. This non-viral peptide-based BioShuttle-mediated transfer system consists of a nuclear localization address sequence realizing the delivery of the plasmid phNIS-IRES-EGFP coding for two independent reporter genes into nuclei of HeLa cells. The quantification of the transfer efficiency was achieved by measurements of the sodium iodide symporter activity. EGFP gene expression was measured with Confocal Laser Scanning Microscopy and quantified with biostatistical methods by analysis of the frequency of the amplitude distribution in the CLSM images. The results demonstrate that the “BioShuttle”-Technology is an appropriate tool for an effective transfer of genetic material carried by a plasmid. PMID:18026568

  6. Electrotransfer of Plasmid Vector DNA into Muscle

    Science.gov (United States)

    Miyazaki, Satsuki; Miyazaki, Jun-Ichi

    Wolff et al. (1990) first reported that plasmid DNA injected into skeletal muscle is taken up by muscle cells and the genes in the plasmid are expressed for more than two months thereafter, although the transfected DNA does not usually undergo chromosomal integration (Wolff et al., 1991, 1992). However, the relatively low expression levels attained by this method have hampered its applications for uses other than as a DNA vaccine (Davis et al., 1995). There are a number of reports analyzing the conditions that affect the efficiency of gene transfer by intramuscular DNA injection and assessing the fine structures of expression plasmid vectors that may affect expression levels (Davis et al., 1993; Liang et al., 1996; Norman et al., 1997). Furthermore, various attempts were done to improve the efficiency of gene transfer by intramus cular DNA injection. Consequently, regenerating muscle was shown to produce 80-fold or more protein than did normal muscle, following injection of an expression plas-mid. Muscle regeneration was induced by treatment with cardiotoxin or bupivacaine (Wells, 1993; Vitadello et al., 1994). We previously demonstrated that by combining a strong promoter and bupivacaine pretreatment intramuscular injection of an IL-5 expression plasmid results in IL-5 production in muscle at a level sufficient to induce marked proliferation of eosinophils in the bone marrow and eosinophil infiltration of various organs (Tokui et al., 1997). It was also reported that a single intramuscular injection of an erythropoietin expression plasmid produced physiologically significant elevations in serum erythropoietin levels and increased hematocrits in adult mice (Tripathy et al., 1996). Hematocrits in these animals remained elevated at >60% for at least 90 days after a single injection. However, improvements to this method have not been sufficient to extend its applications including clinical use.

  7. BioShuttle-mediated Plasmid Transfer

    OpenAIRE

    Braun, Klaus; von Brasch, Leonie; Pipkorn, Ruediger; Ehemann, Volker; Jenne, Juergen; Spring, Herbert; Debus, Juergen; Didinger, Bernd; Rittgen, Werner; Waldeck, Waldemar

    2007-01-01

    An efficient gene transfer into target tissues and cells is needed for safe and effective treatment of genetic diseases like cancer. In this paper, we describe the development of a transport system and show its ability for transporting plasmids. This non-viral peptide-based BioShuttle-mediated transfer system consists of a nuclear localization address sequence realizing the delivery of the plasmid phNIS-IRES-EGFP coding for two independent reporter genes into nuclei of HeLa cells. The quantif...

  8. Competition between Plasmid-Bearing and Plasmid-Free Organisms in a Chemostat with Pulsed Input and Washout

    Directory of Open Access Journals (Sweden)

    Sanling Yuan

    2009-01-01

    Full Text Available We consider a model of competition between plasmid-bearing and plasmid-free organisms in the chemostat with pulsed input and washout. We investigate the subsystem with nutrient and plasmid-free organism and study the stability of the boundary periodic solutions, which are the boundary periodic solutions of the system. The stability analysis of the boundary periodic solution yields the invasion threshold of the plasmid-bearing organism. By using the standard techniques of bifurcation theory, we prove that above this threshold there are periodic oscillations in substrate, plasmid-free, and plasmid-bearing organisms. Numerical simulations are carried out to illustrate our results.

  9. Plasmid maintenance functions of the large virulence plasmid of Shigella flexneri.

    OpenAIRE

    Radnedge, L; Davis, M. A.; Youngren, B; Austin, S. J.

    1997-01-01

    The large virulence plasmid pMYSH6000 of Shigella flexneri contains a replicon and a plasmid maintenance stability determinant (Stb) on adjacent SalI fragments. The presence of a RepFIIA replicon on the SalI C fragment was confirmed, and the complete sequence of the adjacent SalI O fragment was determined. It shows homology to part of the transfer (tra) operon of the F plasmid. Stb stabilizes a partition-defective P1 miniplasmid in Escherichia coli. A 1.1-kb region containing a homolog of the...

  10. Plasmid transfer and plasmid-mediated genetic exchange in Brucella abortus.

    OpenAIRE

    Rigby, C E; Fraser, A.D.

    1989-01-01

    Naturally-occurring plasmids and gene transfer mechanisms have not yet been reported in brucellae. Here we show that Brucella abortus is capable of maintaining and transferring the broad-host-range plasmids pTH10 (IncP), pSa (IncW) and R751 (IncP), and describe pTH10-mediated transfer of B. abortus chromosomal genes to Escherichia coli. All three plasmids transferred by conjugation from E. coli to B. abortus S19, and from B. abortus S19 to B. abortus 292 (biovar 4). They were stably maintaine...

  11. Plasmid transduction by Bacillus subtilis bacteriophage SPP1: effects of DNA homology between plasmid and bacteriophage.

    OpenAIRE

    Deichelbohrer, I; Alonso, J.C.; Lüder, G; Trautner, T A

    1985-01-01

    Any SPP1 DNA restriction fragment cloned into Bacillus subtilis plasmid pC194 or pUB110 increased the transduction frequency of the plasmid by SPP1 100- to 1,000-fold over the transduction level of the plasmid alone. This increment was observed irrespective of whether a fragment contained the SPP1 packaging origin (pac). Furthermore, an SPP1 derivative into whose genome pC194 DNA had been integrated transduced pC194 DNA with a greatly enhanced frequency. Transduction enhancement mediated by D...

  12. Evasion of Neutrophil Killing by Staphylococcus aureus

    Directory of Open Access Journals (Sweden)

    Will A. McGuinness

    2016-03-01

    Full Text Available Staphylococcus aureus causes many types of infections, ranging from self-resolving skin infections to severe or fatal pneumonia. Human innate immune cells, called polymorphonuclear leukocytes (PMNs or neutrophils, are essential for defense against S. aureus infections. Neutrophils are the most prominent cell type of the innate immune system and are capable of producing non-specific antimicrobial molecules that are effective at eliminating bacteria. Although significant progress has been made over the past few decades, our knowledge of S. aureus-host innate immune system interactions is incomplete. Most notably, S. aureus has the capacity to produce numerous molecules that are directed to protect the bacterium from neutrophils. Here we review in brief the role played by neutrophils in defense against S. aureus infection, and correspondingly, highlight selected S. aureus molecules that target key neutrophil functions.

  13. Plasmid maintenance and protein overproduction in selective recycle bioreactors.

    Science.gov (United States)

    Ogden, K L; Davis, R H

    1991-02-20

    A new plasmid construct has been used in conjunction with selective recycle to successfully maintain otherwise unstable plasmid-bearing E. coli cells in a continuous bioreactor and to produce significant amounts of the plasmid-encoded protein beta-lactamase. The plasmid is constructed so that pilin expression, which leads to bacterial flocculation, is under control of the tac operon. The plasmid-bearing cells are induced to flocculate in the separator, whereas cell growth and product synthesis occur in the main fermentation vessel without the inhibiting effects of pilin production. Selective recycle allows for the maintenance of the plasmid-bearing cells by separating flocculent, plasmid-bearing cells from nonflocculent, segregant cells in an inclined settler, and recycling only the plasmid-bearing cells to the reactor. As a result, product expression levels are maintained that are more than ten times the level achieved without selective recycle. All experimental data agree well with theoretical predictions. PMID:18597374

  14. Simple method for identification of plasmid-coded proteins

    International Nuclear Information System (INIS)

    Proteins encoded by plasmid DNA are specifically labeled in uv-irradiated cells of Escherichia coli carrying recA and uvrA mutations because extensive degradation of the chromosome DNA occurs concurrently with amplification of plasmid DNA

  15. Population structure of Staphylococcus aureus in China

    OpenAIRE

    Yan, Xiaomei

    2015-01-01

    The present PhD research was aimed at analysing the population structure of Staphylococcus aureus in China. Between 2000 and 2005 we found that patients from a single Chinese hospital showed increasing trends in antimicrobial resistance. Among methicillin-resistant S. aureus (MRSA), resistance against rifampicin doubled to 68%. Staphylococcal food poisoning (SFP) is frequent in China. Two predominant S. aureus lineages, ST6 and ST943, were identified causing outbreaks of SFP in Southern China...

  16. Immunomodulation and Disease Tolerance to Staphylococcus aureus

    OpenAIRE

    Zhigang Li; Peres, Adam G.; Andreea C. Damian; Joaquín Madrenas

    2015-01-01

    The Gram-positive bacterium Staphylococcus aureus is one of the most frequent pathogens that causes severe morbidity and mortality throughout the world. S. aureus can infect skin and soft tissues or become invasive leading to diseases such as pneumonia, endocarditis, sepsis or toxic shock syndrome. In contrast, S. aureus is also a common commensal microbe and is often part of the human nasal microbiome without causing any apparent disease. In this review, we explore the immunomodulation and d...

  17. DISTRIBUTION OF PLASMIDS IN GROUNDWATER BACTERIA

    Science.gov (United States)

    Bacteria isolated from groundwater aquifer core materials of pristine aquifers at Lula and Pickett, Oklahoma, and from a site with a history of aromatic hydrocarbon contamination and natural renovation located at Conroe, Texas, were screened for the presence of plasmid Deoxyribon...

  18. Plasmid mediated quinolone resistance in Enterobacteriaceae

    NARCIS (Netherlands)

    Veldman, K.T.

    2014-01-01

    This thesis describes the occurrence of Plasmid Mediated Quinolone Resistance (PMQR) in Salmonella and E. coli from The Netherlands and other European countries. Furthermore, the genetic background of these genes was characterized. Fluoroquinolones are widely used antibiotics in both human and veter

  19. The influence of biofilms in the biology of plasmids

    OpenAIRE

    Cook, Laura C.C.; Dunny, Gary M.

    2014-01-01

    The field of plasmid biology has historically focused on bacteria growing in liquid culture. Surface attached communities of bacterial biofilms have recently been understood to be the normal environment of bacteria in the natural world. Thus, studies examining plasmid replication, maintenance, and transfer in biofilms are essential for a true understanding of bacterial plasmid biology. This chapter reviews the current knowledge of the interplay between bacterial biofilms and plasmids, focusin...

  20. The 2 micron plasmid purloins the yeast cohesin complex

    OpenAIRE

    Mehta, Shwetal; Yang, Xian Mei; Chan, Clarence S.; Dobson, Melanie J.; Jayaram, Makkuni; Velmurugan, Soundarapandian

    2002-01-01

    The yeast 2 micron plasmid achieves high fidelity segregation by coupling its partitioning pathway to that of the chromosomes. Mutations affecting distinct steps of chromosome segregation cause the plasmid to missegregate in tandem with the chromosomes. In the absence of the plasmid stability system, consisting of the Rep1 and Rep2 proteins and the STB DNA, plasmid and chromosome segregations are uncoupled. The Rep proteins, acting in concert, recruit the yeast cohesin complex to the STB locu...

  1. Linear and Circular Plasmid Content in Borrelia burgdorferi Clinical Isolates

    OpenAIRE

    Iyer, Radha; Kalu, Ogori; Purser, Joye; Norris, Steven; Stevenson, Brian; Schwartz, Ira

    2003-01-01

    The genome of Borrelia burgdorferi, the etiologic agent of Lyme disease, is composed of a linear chromosome and more than 20 linear and circular plasmids. Typically, plasmid content analysis has been carried out by pulsed-field gel electrophoresis and confirmed by Southern hybridization. However, multiple plasmids of virtually identical sizes (e.g., lp28 and cp32) complicate the interpretation of such data. The present study was undertaken to investigate the complete plasmid complements of B....

  2. Plasmid DNA entry into postmitotic nuclei of primary rat myotubes.

    OpenAIRE

    Dowty, M E; Williams, P; Zhang, G.; Hagstrom, J E; Wolff, J A

    1995-01-01

    These studies were initiated to elucidate the mechanism of DNA nuclear transport in mammalian cells. Biotin- or gold-labeled plasmid and plasmid DNA expression vectors for Escherichia coli beta-galactosidase or firefly luciferase were microinjected into the cytoplasm of primary rat myotubes in culture. Plasmid DNA was expressed in up to 70% of the injected myotubes, which indicates that it entered intact, postmitotic nuclei. The nuclear transport of plasmid DNA occurred through the nuclear po...

  3. Mastite com lesões sistêmicas por Staphylococus aureus subesp. aureus em coelhos Mastitis with systemic lesions due to Staphylococus aureus subesp. aureus in rabbits

    OpenAIRE

    Sandra Davi Traverso; Leonardo da Cunha; Joaquim César Teixeira Fernandes; Alexandre Paulino Loretti; Adriana Rhoden; Elsio Wunder Jr.; David Driemeier

    2003-01-01

    Em uma criação composta por 1800 coelhos, 33% das matrizes apresentaram mastite e lesões cutâneas crostosas e purulentas. Estes animais apresentavam-se entre 10 a- 12 meses de idade e em segunda parição. Quinze coelhos afetados foram sacrificados e necropsiados. Na necropsia, além das lesões cutâneas haviam microabscessos em diversos órgãos. Das amostras coletadas isolou-se Staphylococcus aureus subesp. aureus. S. aureus subesp. aureus também foi isolado de "swab" nasal coletado do tratador e...

  4. Immunomodulation and Disease Tolerance to Staphylococcus aureus

    Directory of Open Access Journals (Sweden)

    Zhigang Li

    2015-11-01

    Full Text Available The Gram-positive bacterium Staphylococcus aureus is one of the most frequent pathogens that causes severe morbidity and mortality throughout the world. S. aureus can infect skin and soft tissues or become invasive leading to diseases such as pneumonia, endocarditis, sepsis or toxic shock syndrome. In contrast, S. aureus is also a common commensal microbe and is often part of the human nasal microbiome without causing any apparent disease. In this review, we explore the immunomodulation and disease tolerance mechanisms that promote commensalism to S. aureus.

  5. Methicillin-Resistant Staphylococcus aureus (MRSA) Treatment

    Science.gov (United States)

    ... Laboratory of Bacteriology Network on Antimicrobial Resistance in Staphylococcus aureus (NARSA) Antibacterial Resistance Leadership Group (ARLG) NIAID Antimicrobial Resistance Funding Information ...

  6. Methicillin-Resistant Staphylococcus aureus (MRSA) Diagnosis

    Science.gov (United States)

    ... Laboratory of Bacteriology Network on Antimicrobial Resistance in Staphylococcus aureus (NARSA) Antibacterial Resistance Leadership Group (ARLG) NIAID Antimicrobial Resistance Funding Information ...

  7. Postsegregational killing does not increase plasmid stability but acts to mediate the exclusion of competing plasmids

    OpenAIRE

    Cooper, Tim F; Heinemann, Jack A.

    2000-01-01

    Postsegregational killing (PSK) systems consist of a tightly linked toxin–antitoxin pair. Antitoxin must be continually produced to prevent the longer lived toxin from killing the cell. PSK systems on plasmids are widely believed to benefit the plasmid by ensuring its stable vertical inheritance. However, experimental tests of this “stability” hypothesis were not consistent with its predictions. We suggest an alternative hypothesis to explain the evolution of PSK: ...

  8. Plasmid Capture by the Bacillus thuringiensis Conjugative Plasmid pXO16▿

    OpenAIRE

    Timmery, Sophie; Modrie, Pauline; Minet, Olivier; Mahillon, Jacques

    2009-01-01

    Conjugation, mobilization, and retromobilization are three related mechanisms of horizontal gene transfer in bacteria. They have been extensively studied in gram-negative species, where retromobilization, the capture of DNA from a recipient by a donor cell, was shown to result from two successive steps: the transfer of the conjugative plasmid from the donor to the recipient followed by the retrotransfer of the mobilizable plasmid to the donor. This successive model was established for gram-ne...

  9. Conjugative plasmids: Vessels of the communal gene pool

    DEFF Research Database (Denmark)

    Norman, Anders; Hansen, Lars H.; Sørensen, Søren Johannes

    2009-01-01

    over large taxonomic distances. These plasmids are collections of discrete regions of genes that function as ‘backbone modules' to undertake different aspects of overall plasmid maintenance and propagation. Conjugative plasmids often carry suites of ‘accessory elements' that contribute adaptive traits...

  10. Compositional discordance between prokaryotic plasmids and host chromosomes

    Directory of Open Access Journals (Sweden)

    van Kampen Antoine HC

    2006-02-01

    Full Text Available Abstract Background Most plasmids depend on the host replication machinery and possess partitioning genes. These properties confine plasmids to a limited range of hosts, yielding a close and presumably stable relationship between plasmid and host. Hence, it is anticipated that due to amelioration the dinucleotide composition of plasmids is similar to that of the genome of their hosts. However, plasmids are also thought to play a major role in horizontal gene transfer and thus are frequently exchanged between hosts, suggesting dinucleotide composition dissimilarity between plasmid and host genome. We compared the dinucleotide composition of a large collection of plasmids with that of their host genomes to shed more light on this enigma. Results The dinucleotide frequency, coined the genome signature, facilitates the identification of putative horizontally transferred DNA in complete genome sequences, since it was found to be typical for a certain genome, and similar between related species. By comparison of the genome signature of 230 plasmid sequences with that of the genome of each respective host, we found that in general the genome signature of plasmids is dissimilar from that of their host genome. Conclusion Our results show that the genome signature of plasmids does not resemble that of their host genome. This indicates either absence of amelioration or a less stable relationship between plasmids and their host. We propose an indiscriminate lifestyle for plasmids preserving the genome signature discordance between these episomes and host chromosomes.

  11. Competition between Plasmid-Bearing and Plasmid-Free Organisms in a Chemostat with Pulsed Input and Washout

    OpenAIRE

    Sanling Yuan; Yu Zhao; Anfeng Xiao

    2009-01-01

    We consider a model of competition between plasmid-bearing and plasmid-free organisms in the chemostat with pulsed input and washout. We investigate the subsystem with nutrient and plasmid-free organism and study the stability of the boundary periodic solutions, which are the boundary periodic solutions of the system. The stability analysis of the boundary periodic solution yields the invasion threshold of the plasmid-bearing organism. By using the standard techniques of bifurcation theory, w...

  12. Co-segregation of yeast plasmid sisters under monopolin-directed mitosis suggests association of plasmid sisters with sister chromatids

    OpenAIRE

    Liu, Yen-Ting; Ma, Chien-Hui; Jayaram, Makkuni

    2013-01-01

    The 2-micron plasmid, a high copy extrachromosomal element in Saccharomyces cerevisiae, propagates itself with nearly the same stability as the chromosomes of its host. Plasmid stability is conferred by a partitioning system consisting of the plasmid-coded proteins Rep1 and Rep2 and a cis-acting locus STB. Circumstantial evidence suggests that the partitioning system couples plasmid segregation to chromosome segregation during mitosis. However, the coupling mechanism has not been elucidated. ...

  13. Staphylococcus aureus small colony variants in diabetic foot infections

    OpenAIRE

    Cervante-García, Estrella; García-Gonzalez, Rafael; Reyes-Torres, Angélica; Resendiz-Albor, Aldo Arturo; Salazar-Schettino, Paz María

    2015-01-01

    Background: Staphylococcus aureus (S. aureus) is one of the major pathogens causing chronic infections. The ability of S. aureus to acquire resistance to a diverse range of antimicrobial compounds results in limited treatment options, particularly in methicillin-resistant S. aureus (MRSA). A mechanism by which S. aureus develops reduced susceptibility to antimicrobials is through the formation of small colony variants (SCVs). Infections by SCVs of S. aureus are an upcoming problem due to diff...

  14. Evaluation of Two New Chromogenic Media, CHROMagar MRSA and S. aureus ID, for Identifying Staphylococcus aureus and Screening Methicillin-Resistant S. aureus

    OpenAIRE

    Hedin, Göran; Fang, Hong

    2005-01-01

    Thirty-nine methicillin-resistant Staphylococcus aureus (MRSA) isolates with diverse genetic backgrounds and two reference strains were correctly identified as S. aureus on CHROMagar MRSA and S. aureus ID media. Growth inhibition on CHROMagar MRSA was noted. A combination of cefoxitin disk and S. aureus ID was found suitable for rapid MRSA screening.

  15. THE INTEGRATED STATE OF THE ROLLING-CIRCLE PLASMID PTB913 IN THE COMPOSITE BACILLUS PLASMID PTB19

    NARCIS (Netherlands)

    OSKAM, L; HILLENGA, DJ; VENEMA, G; BRON, S

    1992-01-01

    pTB19, a 27 kb plasmid originating from a thermophilic Bacillus species, contains integrated copies of two rolling-circle type plasmids on a 10.6 kb DNA fragment. In the present study we analysed the part of pTB19 that contains the rolling-circle plasmid pTB913 and the region in between the two roll

  16. Stable inheritance of plasmid R1 requires two different loci.

    OpenAIRE

    Gerdes, K; Larsen, J E; Molin, S

    1985-01-01

    The largest EcoRI fragment from plasmid R1 mediates a stability phenotype which is required to ensure the stable inheritance of this low-copy-number plasmid. When covalently linked to small, unstable R1 derivatives, this fragment makes the plasmids as stable as the wild-type R1 plasmid. A genetic analysis showed that two independently acting stabilization functions are encoded by this EcoRI fragment, both of which have the potential of partial stabilization of mini-R1 plasmids. The two loci a...

  17. DNA restriction-modification systems mediate plasmid maintenance.

    OpenAIRE

    Kulakauskas, S; Lubys, A; Ehrlich, S. D.

    1995-01-01

    Two plasmid-carried restriction-modification (R-M) systems, EcoRI (from pMB1 of Escherichia coli) and Bsp6I (from pXH13 of Bacillus sp. strain RFL6), enhance plasmid segregational stability in E. coli and Bacillus subtilis, respectively. Inactivation of the endonuclease or the presence of the methylase in trans abolish the stabilizing activity of the R-M systems. We propose that R-M systems mediate plasmid segregational stability by postsegregational killing of plasmid-free cells. Plasmid-enc...

  18. The cell surface proteome of Staphylococcus aureus

    NARCIS (Netherlands)

    Dreisbach, Annette; van Dijl, Jan Maarten; Buist, Girbe

    2011-01-01

    The Gram-positive bacterium Staphylococcus aureus is a wide spread opportunistic pathogen that can cause a range of life-threatening diseases. To obtain a better understanding of the global mechanisms for pathogenesis and to identify novel targets for therapeutic interventions, the S. aureus proteom

  19. Population structure of Staphylococcus aureus in China

    NARCIS (Netherlands)

    Yan, Xiaomei

    2015-01-01

    The present PhD research was aimed at analysing the population structure of Staphylococcus aureus in China. Between 2000 and 2005 we found that patients from a single Chinese hospital showed increasing trends in antimicrobial resistance. Among methicillin-resistant S. aureus (MRSA), resistance again

  20. Role of Plasmid in Production of Acetobacter Xylinum Biofilms

    Directory of Open Access Journals (Sweden)

    Abbas Rezaee

    2005-01-01

    Full Text Available Acetobacter xylinum has the ability to produce cellulotic biofilms. Bacterial cellulose is expected to be used in many industrial or biomedical materials for its unique characteristics. A. xylinum contains a complex system of plasmid DNA molecules. A 44 kilobases (kb plasmid was isolated in wild type of A. xylinum. To improve the cellulose producing ability of A. xylinum, role of the plasmid in production of cellulose was studied. The comparisons between wild type and cured cells of A. xylinum showed that there is considerably difference in cellulose production. In order to study the relationship between plasmid and the rate of cellulose production, bacteria were screened for plasmid profile by a modified method for preparation of plasmid. This method yields high levels of pure plasmid DNA that can be used for common molecular techniques, such as digestion and transformation, with high efficiency.

  1. Deficient sumoylation of yeast 2-micron plasmid proteins Rep1 and Rep2 associated with their loss from the plasmid-partitioning locus and impaired plasmid inheritance.

    Directory of Open Access Journals (Sweden)

    Jordan B Pinder

    Full Text Available The 2-micron plasmid of the budding yeast Saccharomyces cerevisiae encodes copy-number amplification and partitioning systems that enable the plasmid to persist despite conferring no advantage to its host. Plasmid partitioning requires interaction of the plasmid Rep1 and Rep2 proteins with each other and with the plasmid-partitioning locus STB. Here we demonstrate that Rep1 stability is reduced in the absence of Rep2, and that both Rep proteins are sumoylated. Lysine-to-arginine substitutions in Rep1 and Rep2 that inhibited their sumoylation perturbed plasmid inheritance without affecting Rep protein stability or two-hybrid interaction between Rep1 and Rep2. One-hybrid and chromatin immunoprecipitation assays revealed that Rep1 was required for efficient retention of Rep2 at STB and that sumoylation-deficient mutants of Rep1 and Rep2 were impaired for association with STB. The normal co-localization of both Rep proteins with the punctate nuclear plasmid foci was also lost when Rep1 was sumoylation-deficient. The correlation of Rep protein sumoylation status with plasmid-partitioning locus association suggests a theme common to eukaryotic chromosome segregation proteins, sumoylated forms of which are found enriched at centromeres, and between the yeast 2-micron plasmid and viral episomes that depend on sumoylation of their maintenance proteins for persistence in their hosts.

  2. Prevalence of plasmid mediated pesticide resistant bacterial assemblages in crop fields.

    Science.gov (United States)

    Umamaheswari, S; Murali, M

    2010-11-01

    Three crop fields namely paddy sugarcane and tomato exposed to bavistin [Methyl (1H-benzimidazol-2-yl) carbomate], monocrotophos[Dimethyl(E)-1-methyl-2-(methyl-carbamoyl) vinyl phosphate] and kinado plus [(EZ)-2-chloro-3-dimethoxyphosphinoyloxy-X1, X1-diethylbut-2-enamide], respectively were chosen for the present investigation to know the bacterial population and degradation of pesticides. The chemical nature of the soil and water samples from the pesticide contaminated fields was analysed along with counting of the total heterotrophic bacteria (THB), Staphylococci and Enterococcci population. Mean calcium, phosphate and biological oxygen demand were maximum in tomato field water Field water recorded maximum phophate and silicate content, whereas, sugarcane field water elicited maximum dissolved oxygen content. On the other hand, available phosphate and exchangeable potassium were maximum is sugarcane field soil. Significant variations in the bacterial population were evident between the treatments in sugarcane field soil and tomato field water exposed to monocrotophos and kinado plus, respectively In addition, significant variations between THB, Staphlyococci and Enterococci population were also evinced in both the sugarcane andtomato fields. The dominant pesticide resistant bacteria, Staphylococcus aureus, Enterococcus faecalis and Pseudomonas aeuroginosa harboured plasmids and the resistant trait observed were found to be plasmid borne. PMID:21506482

  3. Antibacterial effect of cationic porphyrazines and anionic phthalocyanine and their interaction with plasmid DNA

    Science.gov (United States)

    Hassani, Leila; Hakimian, Fatemeh; Safaei, Elham; Fazeli, Zahra

    2013-11-01

    Resistance to antibiotics is a public health issue and identification of new antibacterial agents is one of the most important goals of pharmacological research. Among the novel developed antibacterial agents, porphyrin complexes and their derivatives are ideal candidates for use in medical applications. Phthalocyanines differ from porphyrins by having nitrogen atoms link the individual pyrrol units. The aza analogues of the phthalocyanines (azaPcs) such as tetramethylmetalloporphyrazines are heterocyclic Pc analogues. In this investigation, interaction of an anionic phthalocyanine (Cu(PcTs)) and two cationic tetrapyridinoporphyrazines including [Cu(2,3-tmtppa)]4+ and [Cu(3,4-tmtppa)]4+ complexes with plasmid DNA was studied using spectroscopic and gel electrophoresis methods. In addition, antibacterial effect of the complexes against Gram-positive (Staphylococcus aureus) and Gram-negative (Escherichia coli) bacteria was investigated using dilution test method. The results indicated that both porphyrazines have significant antibacterial properties, but Cu(PcTs) has weak antibacterial effect. Compairing the binding of the phthalocyanine and the porphyrazines to DNA demonstrated that the interaction of cationic porphyrazines is stronger than the anionic phthalocyanine remarkably. The extent of hypochromicity and red shift of absorption spectra indicated preferential intercalation of the two porphyrazine into the base pairs of DNA helix. Gel electrophoresis result implied Cu(2,3-tmtppa) and Cu(3,4-tmtppa) are able to perform cleavage of the plasmid DNA. Consequently, DNA binding and cleavage might be one of the antibacterial mechanisms of the complexes.

  4. Modeling sRNA-regulated Plasmid Maintenance

    CERN Document Server

    Gong, Chen Chris

    2016-01-01

    We study a theoretical model for the toxin-antitoxin (hok/sok) mechanism for plasmid maintenance in bacteria. Toxin-antitoxin systems enforce the maintenance of a plasmid through post-segregational killing of cells that have lost the plasmid. Key to their function is the tight regulation of expression of a protein toxin by an sRNA antitoxin. Here, we focus on the nonlinear nature of the regulatory circuit dynamics of the toxin-antitoxin mechanism. The mechanism relies on a transient increase in protein concentration rather than on the steady state of the genetic circuit. Through a systematic analysis of the parameter dependence of this transient increase, we confirm some known design features of this system and identify new ones: for an efficient toxin-antitoxin mechanism, the synthesis rate of the toxin's mRNA template should be lower that of the sRNA antitoxin, the mRNA template should be more stable than the sRNA antitoxin, and the mRNA-sRNA complex should be more stable than the sRNA antitoxin. Moreover, ...

  5. Determination of Plasmid Segregational Stability in a Growing Bacterial Population.

    Science.gov (United States)

    Kramer, M Gabriela

    2016-01-01

    Bacterial plasmids are extensively used as cloning vectors for a number of genes for academic and commercial purposes. Moreover, attenuated bacteria carrying recombinant plasmids expressing genes with anti-tumor activity have shown promising therapeutic results in animal models of cancer. Equitable plasmid distribution between daughter cells during cell division, i.e., plasmid segregational stability, depends on many factors, including the plasmid copy number, its replication mechanism, the levels of recombinant gene expression, the type of bacterial host, and the metabolic burden associated with all these factors. Plasmid vectors usually code for antibiotic-resistant functions, and, in order to enrich the culture with bacteria containing plasmids, antibiotic selective pressure is commonly used to eliminate plasmid-free segregants from the growing population. However, administration of antibiotics can be inconvenient for many industrial and therapeutic applications. Extensive ongoing research is being carried out to develop stably-inherited plasmid vectors. Here, I present an easy and precise method for determining the kinetics of plasmid loss or maintenance for every ten generations of bacterial growth in culture. PMID:26846807

  6. Plasmid transfer between bacteria in soil microcosms and the field

    Directory of Open Access Journals (Sweden)

    Eric Smit

    1997-01-01

    Full Text Available In ibis review factors influencing conjugal plasmid transfer between bacteria and the possible role of naturally occurring selftransmissible plasmide for the dissemination of recombinant DNA in soil will be discussed. In microcosm studies, plasmid transfer between various species of introduced bacteria has been detected. Moreover, plamid transfer to indigenous soil micoorganisms was observed. Soil is an oligotrophic environment and plasmid transfer occurred mainly under conditions which were nutritionally favourable for bacteria, such as in the plant rhizosphere and in the presence of clay minerais or added nutrients. Mobilizable plasmids, lacking the ability to transfer themselves, have been reported to be transferred in the presence of selftransmissible plasmids. A study comparing conjugal transfer in microcosme with those in the field revealed that the transfer rates found in microcosme and in the field were similar. Transfer of chromosomal DNA by plasmid RP4 could only be shown on filters and was not observed in soil. Transfer of plasmids carrying biodegradative genes appeared to be favoured in the presence of the compound that can be degraded. Evidence was found for the presence of naturally-occurring selftransmissible plasmids in bacteria in the rhizosphere which could mobilize recombinant plasmids.

  7. Characterization of toxin plasmids in Clostridium perfringens type C isolates.

    Science.gov (United States)

    Gurjar, Abhijit; Li, Jihong; McClane, Bruce A

    2010-11-01

    Clostridium perfringens type C isolates cause enteritis necroticans in humans or necrotizing enteritis and enterotoxemia in domestic animals. Type C isolates always produce alpha toxin and beta toxin but often produce additional toxins, e.g., beta2 toxin or enterotoxin. Since plasmid carriage of toxin-encoding genes has not been systematically investigated for type C isolates, the current study used Southern blot hybridization of pulsed-field gels to test whether several toxin genes are plasmid borne among a collection of type C isolates. Those analyses revealed that the surveyed type C isolates carry their beta toxin-encoding gene (cpb) on plasmids ranging in size from ∼65 to ∼110 kb. When present in these type C isolates, the beta2 toxin gene localized to plasmids distinct from the cpb plasmid. However, some enterotoxin-positive type C isolates appeared to carry their enterotoxin-encoding cpe gene on a cpb plasmid. The tpeL gene encoding the large clostridial cytotoxin was localized to the cpb plasmids of some cpe-negative type C isolates. The cpb plasmids in most surveyed isolates were found to carry both IS1151 sequences and the tcp genes, which can mediate conjugative C. perfringens plasmid transfer. A dcm gene, which is often present near C. perfringens plasmid-borne toxin genes, was identified upstream of the cpb gene in many type C isolates. Overlapping PCR analyses suggested that the toxin-encoding plasmids of the surveyed type C isolates differ from the cpe plasmids of type A isolates. These findings provide new insight into plasmids of proven or potential importance for type C virulence. PMID:20823204

  8. Genomic Analysis of Companion Rabbit Staphylococcus aureus

    Science.gov (United States)

    Holmes, Mark A.; Harrison, Ewan M.; Fisher, Elizabeth A.; Graham, Elizabeth M.; Parkhill, Julian; Foster, Geoffrey; Paterson, Gavin K.

    2016-01-01

    In addition to being an important human pathogen, Staphylococcus aureus is able to cause a variety of infections in numerous other host species. While the S. aureus strains causing infection in several of these hosts have been well characterised, this is not the case for companion rabbits (Oryctolagus cuniculus), where little data are available on S. aureus strains from this host. To address this deficiency we have performed antimicrobial susceptibility testing and genome sequencing on a collection of S. aureus isolates from companion rabbits. The findings show a diverse S. aureus population is able to cause infection in this host, and while antimicrobial resistance was uncommon, the isolates possess a range of known and putative virulence factors consistent with a diverse clinical presentation in companion rabbits including severe abscesses. We additionally show that companion rabbit isolates carry polymorphisms within dltB as described as underlying host-adaption of S. aureus to farmed rabbits. The availability of S. aureus genome sequences from companion rabbits provides an important aid to understanding the pathogenesis of disease in this host and in the clinical management and surveillance of these infections. PMID:26963381

  9. [Staphylococcus aureus and antibiotic resistance].

    Science.gov (United States)

    Sancak, Banu

    2011-07-01

    After the report of first case of methicillin-resistant Staphylococcus aureus (MRSA) in 1961, MRSA become a major problem worldwide. Over the last decade MRSA strains have emerged as serious pathogens in nosocomial and community settings. Glycopeptides (vancomycin and teicoplanin) are still the current mainstay of therapy for infections caused by MRSA. In the last decade dramatic changes have occurred in the epidemiology of MRSA infections. The isolates with reduced susceptibility and in vitro resistance to vancomycin have emerged. Recently, therapeutic alternatives such as quinupristin/dalfopristin, linezolid, tigecycline and daptomycin have been introduced into clinical practice for treating MRSA infections. Nevertheless, these drugs are only approved for certain indication and resistance has already been reported. In this review, the new information on novel drugs for treating MRSA infections and the resistance mechanisms of these drugs were discussed. PMID:21935792

  10. Exfoliative Toxins of Staphylococcus aureus

    Directory of Open Access Journals (Sweden)

    Michal Bukowski

    2010-05-01

    Full Text Available Staphylococcus aureus is an important pathogen of humans and livestock. It causes a diverse array of diseases, ranging from relatively harmless localized skin infections to life-threatening systemic conditions. Among multiple virulence factors, staphylococci secrete several exotoxins directly associated with particular disease symptoms. These include toxic shock syndrome toxin 1 (TSST-1, enterotoxins, and exfoliative toxins (ETs. The latter are particularly interesting as the sole agents responsible for staphylococcal scalded skin syndrome (SSSS, a disease predominantly affecting infants and characterized by the loss of superficial skin layers, dehydration, and secondary infections. The molecular basis of the clinical symptoms of SSSS is well understood. ETs are serine proteases with high substrate specificity, which selectively recognize and hydrolyze desmosomal proteins in the skin. The fascinating road leading to the discovery of ETs as the agents responsible for SSSS and the characterization of the molecular mechanism of their action, including recent advances in the field, are reviewed in this article.

  11. Unique type of plasmid maintenance function: postsegregational killing of plasmid-free cells.

    OpenAIRE

    Gerdes, K; Rasmussen, P. B.; Molin, S

    1986-01-01

    The stability locus parB+ of plasmid R1 has been found to specify a unique type of plasmid maintenance function. Two genes, hok (host killing) and sok (suppressor of killing), are required for the stabilizing activity. The hok gene encodes a highly toxic gene product, whose overexpression causes a rapid killing and a concomitant dramatic change in morphology of the host cell. The other gene, sok, was found to encode a product that counteracts the hok gene-mediated killing. The parB+ region wa...

  12. Molecular characterization of "plasmid-free" antibiotic-resistant Haemophilus influenzae.

    OpenAIRE

    Roberts, M C; Smith, A. L.

    1980-01-01

    We examined 14 multiresistant and 8 ampicillin- or tetracycline-resistant Haemophilus influenzae isolates and 4 ampicillin-resistant H. parainfluenzae isolates for plasmid deoxyribonucleic acid. Sixteen strains carried plasmids. Both "plasmid-free" and plasmid-carrying isolates transferred the antibiotic resistance by conjugation. All transconjugants carried plasmid deoxyribonucleic acid, suggesting that the apparent plasmid-free strains contained R plasmids encoding for antibiotic resistance.

  13. Phospholipase C-related catalytically inactive protein participates in the autophagic elimination of Staphylococcus aureus infecting mouse embryonic fibroblasts.

    Directory of Open Access Journals (Sweden)

    Kae Harada-Hada

    Full Text Available Autophagy is an intrinsic host defense system that recognizes and eliminates invading bacterial pathogens. We have identified microtubule-associated protein 1 light chain 3 (LC3, a hallmark of autophagy, as a binding partner of phospholipase C-related catalytically inactive protein (PRIP that was originally identified as an inositol trisphosphate-binding protein. Here, we investigated the involvement of PRIP in the autophagic elimination of Staphylococcus aureus in infected mouse embryonic fibroblasts (MEFs. We observed significantly more LC3-positive autophagosome-like vacuoles enclosing an increased number of S. aureus cells in PRIP-deficient MEFs than control MEFs, 3 h and 4.5 h post infection, suggesting that S. aureus proliferates in LC3-positive autophagosome-like vacuoles in PRIP-deficient MEFs. We performed autophagic flux analysis using an mRFP-GFP-tagged LC3 plasmid and found that autophagosome maturation is significantly inhibited in PRIP-deficient MEFs. Furthermore, acidification of autophagosomes was significantly inhibited in PRIP-deficient MEFs compared to the wild-type MEFs, as determined by LysoTracker staining and time-lapse image analysis performed using mRFP-GFP-tagged LC3. Taken together, our data show that PRIP is required for the fusion of S. aureus-containing autophagosome-like vacuoles with lysosomes, indicating that PRIP is a novel modulator in the regulation of the innate immune system in non-professional phagocytic host cells.

  14. Construction of scFv that bind both fibronectin-binding protein A and clumping factor A of Stapylococcus aureus.

    Science.gov (United States)

    Wang, Man; Zhang, Yan; Li, Benqiang; Zhu, Jianguo

    2015-06-01

    Bovine mastitis (BM) causes significant losses to the dairy industry. Vaccines against the causative agent of BM, Staphylococcus aureus, do not confer adequate protection. Because passive immunization with antibodies permits disease prevention, we constructed a recombinant single-chain antibody (scFv) against fibronectin-binding protein A (FnBPA) and clumping factor A (ClfA), two important virulence factors in S. aureus infection. The DNA coding sequences of the variable heavy (VH) and variable light (VL) domains of antibodies produced in the peripheral blood lymphocytes of cows with S. aureus-induced mastitis were obtained using reverse transcription and polymerase chain reaction, and the VH and VL cDNAs were assembled in-tandem using a DNA sequence encoding a (Gly4Ser)3 peptide linker. The scFv cDNAs were cloned into the pOPE101 plasmid for the expression of soluble scFv protein in Escherichia coli. The binding of the scFvs to both FnBPA and ClfA was confirmed using an indirect ELISA and Western blotting. The DNA sequences of the framework regions of the VH and VL domains were highly conserved, and the complementarity-determining regions displayed significant diversity, especially in CDR3 of the VH domain. These novel bovine antibody fragments may be useful as a therapeutic candidate for the prevention and treatment of S. aureus-induced bovine mastitis. PMID:25910693

  15. The Heme Sensor System of Staphylococcus aureus

    OpenAIRE

    Stauff, Devin L; Skaar, Eric P.

    2009-01-01

    The important human pathogen Staphylococcus aureus is able to satisfy its nutrient iron requirement by acquiring heme from host hemoglobin in the context of infection. However, heme acquisition exposes S. aureus to heme toxicity. In order to detect the presence of toxic levels of exogenous heme, S. aureus is able to sense heme through the heme sensing system (HssRS) two-component system. Upon sensing heme, HssRS directly regulates the expression of the heme-regulated ABC transporter HrtAB, wh...

  16. Staphylococcus aureus infections in psittacine birds.

    Science.gov (United States)

    Hermans, K; Devriese, L A; De Herdt, P; Godard, C; Haesebrouck, F

    2000-10-01

    Staphylococcus aureus was isolated from internal organs of 13 different psittacine birds submitted for necropsy over a period of 6 years. The birds all had lesions consistent with septicaemia. S. aureus isolates included three different phage types. In seven of the 13 birds, concurrent infections with Chlamydophila species, Enterococcus hirae, Candida species, unidentified streptococci and coagulasenegative staphylococci were detected. One bird also had lesions of lymphoid leucosis. Few indications were found that staphylococcosis associated problems may spread epidemically. The present studies suggest that S. aureus is pathogenic for psittacine birds, although it does not seem to be a frequent cause of disease. PMID:19184832

  17. Laboratory Maintenance of Methicillin-Resistant Staphylococcus aureus (MRSA)

    OpenAIRE

    Nicholas P Vitko; Richardson, Anthony R.

    2013-01-01

    Staphylococcus aureus is an important bacterial pathogen in the hospital and community settings, especially Staphylococcus aureus clones that exhibit methicillin-resistance (MRSA). Many strains of S. aureus are utilized in the laboratory, underscoring the genetic differences inherent in clinical isolates. S. aureus grows quickly at 37°C with aeration in rich media (e.g. BHI) and exhibits a preference for glycolytic carbon sources. Furthermore, S. aureus has a gold pigmentation, exhibits β-hem...

  18. Safety and efficacy of DNA vaccines: Plasmids vs. minicircles

    OpenAIRE

    Stenler, Sofia; Blomberg, Pontus; Smith, Ci Edvard

    2014-01-01

    While DNA vaccination using plasmid vectors is highly attractive, there is a need for further vector optimization regarding safety, stability, and efficiency. In this commentary, we review the minicircle vector (MC), which is an entity devoid of plasmid bacterial sequences, as an alternative to the traditional plasmid construct. The commentary highlights the recent discovery by Stenler et al. (2014) that the small size of an MC enables improved resistance to the shearing forces associated wit...

  19. Pathogenomics of the Virulence Plasmids of Escherichia coli

    OpenAIRE

    Johnson, Timothy J.; Lisa K. Nolan

    2009-01-01

    Summary: Bacterial plasmids are self-replicating, extrachromosomal elements that are key agents of change in microbial populations. They promote the dissemination of a variety of traits, including virulence, enhanced fitness, resistance to antimicrobial agents, and metabolism of rare substances. Escherichia coli, perhaps the most studied of microorganisms, has been found to possess a variety of plasmid types. Included among these are plasmids associated with virulence. Several types of E. col...

  20. Identification of plasmid partition function in coryneform bacteria.

    OpenAIRE

    Kurusu, Y; Satoh, Y.; Inui, M.; Kohama, K; Kobayashi, M.; Terasawa, M.; Yukawa, H

    1991-01-01

    We have identified and characterized a partition function that is required for stable maintenance of plasmids in the coryneform bacteria Brevibacterium flavum MJ233 and Corynebacterium glutamicum ATCC 31831. This function is localized to a HindIII-NspV fragment (673 bp) adjacent to the replication region of the plasmid, named pBY503, from Brevibacterium stationis IFO 12144. The function was independent of copy number control and was not associated directly with plasmid replication functions. ...

  1. Streamlined Purification of Plasmid DNA From Prokaryotic Cultures

    OpenAIRE

    Pueschel, Laura; Li, Hongshan; Hymes, Matthew

    2011-01-01

    We describe the complete process of AcroPrep Advance Filter Plates for 96 plasmid preparations, starting from prokaryotic culture and ending with high purity DNA. Based on multi-well filtration for bacterial lysate clearance and DNA purification, this method creates a streamlined process for plasmid preparation. Filter plates containing silica-based media can easily be processed by vacuum filtration or centrifuge to yield appreciable quantities of plasmid DNA. Quantitative analyses determine ...

  2. Comparative genetic organization of incompatibility group P degradative plasmids.

    OpenAIRE

    Burlage, R S; Bemis, L A; Layton, A C; Sayler, G. S.; Larimer, F

    1990-01-01

    Plasmids that encode genes for the degradation of recalcitrant compounds are often examined only for characteristics of the degradative pathways and ignore regions that are necessary for plasmid replication, incompatibility, and conjugation. If these characteristics were known, then the mobility of the catabolic genes between species could be predicted and different catabolic pathways might be combined to alter substrate range. Two catabolic plasmids, pSS50 and pSS60, isolated from chlorobiph...

  3. The Heme Sensor System of Staphylococcus aureus

    Science.gov (United States)

    Stauff, Devin L.; Skaar, Eric P.

    2016-01-01

    The important human pathogen Staphylococcus aureus is able to satisfy its nutrient iron requirement by acquiring heme from host hemoglobin in the context of infection. However, heme acquisition exposes S. aureus to heme toxicity. In order to detect the presence of toxic levels of exogenous heme, S. aureus is able to sense heme through the heme sensing system (HssRS) two-component system. Upon sensing heme, HssRS directly regulates the expression of the heme-regulated ABC transporter HrtAB, which alleviates heme toxicity. Importantly, the inability to sense or respond to heme alters the virulence of S. aureus, highlighting the importance of heme sensing and detoxification to staphylococcal pathogenesis. Furthermore, potential orthologues of the Hss and Hrt systems are found in many species of Gram-positive bacteria, a possible indication that heme stress is a challenge faced by bacteria whose habitats include host tissues rich in heme. PMID:19494582

  4. Construction of a eukaryotic expression plasmid of Humanin*

    OpenAIRE

    Luo, Ben-yan; Chen, Xiang-ming; Tang, Min; Chen, Feng; Chen, Zhi

    2004-01-01

    Objective: To construct a eukaryotic expression plasmid pcDNA3.1(-)-Humanin. Methods: The recombinant plasmid pGEMEX-1-Humanin was digested with restriction endonucleases BamH I and Hind III and the Humanin gene fragments, about 100 bp length, were obtained. Then the Humanin gene fragments were inserted into eukaryotic expression vector pcDNA3.1(-) and the recombinant plasmids pcDNA3.1(-)-Humanin were identified by sequencing. Results: Recombinant plasmid DNA successfully produced a band whic...

  5. Characterization of a plasmid from moderately halophilic eubacteria

    OpenAIRE

    Fernández Castillo, Rosario; Vargas, C.; Nieto Gutiérrez, Joaquín José; Ventosa Ucero, Antonio; Ruiz Berraquero, Francisco

    1992-01-01

    A plasmid has been isolated for the first time from moderately halophilic eubacteria. Halomonas elongata, Halomonrrs halmphila, Deleya halophila and Vibrio costkola were found to harbour an 11.5 kbp plasmid (pMH1). The plasmid was isolated and characterized after transformation into Escherichia coh'JM101 cells. A restriction map was constructed, and unique restriction sites for EcoRI, EcoRV and ClaI were detected. The occurrence of such a plasmid in the original halophilic strains was confirm...

  6. Characterization of Chlamydia trachomatis Plasmid-Encoded Open Reading Frames

    OpenAIRE

    Gong, Siqi; Yang, Zhangsheng; Lei, Lei; Shen, Li; Zhong, Guangming

    2013-01-01

    The recent success in transformation of Chlamydia trachomatis represents a major advancement in Chlamydia research. Plasmid-free C. trachomatis serovar L2 organisms can be transformed with chlamydial plasmid-based shuttle vectors pGFP::SW2 and pBRCT. Deletion of plasmid genes coding for Pgp1 to Pgp8 in pBRCT led to the identification of Pgp1, -2, -6, and -8 as plasmid maintenance factors; Pgp4 as a transcriptional regulator of chlamydial virulence-associated gene expression; and Pgp3, -5, and...

  7. Plasmid P1 replication: negative control by repeated DNA sequences.

    OpenAIRE

    Chattoraj, D; Cordes, K.; Abeles, A

    1984-01-01

    The incompatibility locus, incA, of the unit-copy plasmid P1 is contained within a fragment that is essentially a set of nine 19-base-pair repeats. One or more copies of the fragment destabilizes the plasmid when present in trans. Here we show that extra copies of incA interfere with plasmid DNA replication and that a deletion of most of incA increases plasmid copy number. Thus, incA is not essential for replication but is required for its control. When cloned in a high-copy-number vector, pi...

  8. Identification of plasmid partition function in coryneform bacteria

    Energy Technology Data Exchange (ETDEWEB)

    Kurusu, Yasurou; Satoh, Yukie; Inui, Masayuki; Kohama, Keiko; Kobayashi, Miki; Terasawa, Masato; Yukawa, Hideaki (Mitsubishi Petrochemical Co., Ltd., Ibaraki (Japan))

    1991-03-01

    The authors have identified and characterized a partition function that is required for stable maintenance of plasmids in the coryneform bacteria Brevibacterium flavum MJ233 and Corynebacterium glutamicum ATCC 31831. This function is localized to a HindIII-NspV fragment (673 bp) adjacent to the replication region of the plasmid, named pBY503, from Brevibacterium stationis IFO 12144. The function was independent of copy number control and was not associated directly with plasmid replication functions. This fragment was able to stabilize the unstable plasmids in cis but not in trans.

  9. Degradative Plasmid and Heavy Metal Resistance Plasmid Naturally Coexist in Phenol and Cyanide Assimilating Bacteria

    OpenAIRE

    Bahig E.  Deeb; Abdullah D. Altalhi

    2009-01-01

    Problem statement: Heavy metals are known to be powerful inhibitors of xenobiotics biodegradation activities. Alleviation the inhibitory effect of these metals on the phenol biodegradation activities in presence of heavy metals resistant plasmid was investigated. Approach: Combination of genetic systems of degradation of xenobiotic compound and heavy metal resistance was one of the approaches to the creation of polyfunctional strains for bioremediation of s...

  10. Photonic plasmid stability of transformed Salmonella Typhimurium: A comparison of three unique plasmids

    Directory of Open Access Journals (Sweden)

    Lay Donald

    2009-07-01

    Full Text Available Abstract Background Acquiring a highly stable photonic plasmid in transformed Salmonella Typhimurium for use in biophotonic studies of bacterial tracking in vivo is critical to experimental paradigm development. The objective of this study was to determine stability of transformed Salmonella Typhimurium (S. typh-lux using three different plasmids and characterize their respective photonic properties. Results In presence of ampicillin (AMP, S. typh-lux with pCGLS-1, pAK1-lux and pXEN-1 plasmids exhibited 100% photon-emitting colonies over a 10-d study period. Photon emitters of S. typh-lux with pCGLS-1, pAK1-lux and pXEN-1 without AMP selection decreased over time (P 7 to 1 × 109 CFU, P 0.05; although photonic emissions across a range of bacterial concentrations were not different (1 × 104 to 1 × 106 CFU, P > 0.05. For very low density bacterial concentrations imaged in 96 well plates photonic emissions were positively correlated with bacterial concentration (P 3 to 1 × 105 CFU low to high were different in the 96-well plate format (P Conclusion These data characterize photon stability properties for S. typh-lux transformed with three different photon generating plasmids that may facilitate real-time Salmonella tracking using in vivo or in situ biophotonic paradigms.

  11. Photonic Plasmid Stability of Transformed Salmonella Typhimurium: A Comparison of Three Unique Plasmids

    Science.gov (United States)

    Background: Acquiring a highly stable photonic plasmid in transformed Salmonella Typhimurium for use in biophotonic studies of bacterial tracking in vivo is critical to experimental paradigm development. The objective of this study was to determine stability of transformed Salmonella Typhimurium (S....

  12. Photonic plasmid stability of transformed Salmonella typhimurium: A comparison of three unique plasmids

    Science.gov (United States)

    Acquiring a highly stable photonic plasmid in transformed Salmonella typhimurium for use in biophotonic studies of bacterial tracking in vivo is critical to experimental paradigm development. The objective of this study was to determine stability of transformed Salmonella typhimurium (S. typh-lux) u...

  13. Bacterial Mitosis: ParM of Plasmid R1 Moves Plasmid DNA by an Actin-like Insertional Polymerization Mechanism

    DEFF Research Database (Denmark)

    Møller-Jensen, Jakob; Borch, Jonas; Dam, Mette;

    2003-01-01

    Bacterial DNA segregation takes place in an active and ordered fashion. In the case of Escherichia coli plasmid R1, the partitioning system (par) separates paired plasmid copies and moves them to opposite cell poles. Here we address the mechanism by which the three components of the R1 par system...... act together to generate the force required for plasmid movement during segregation. ParR protein binds cooperatively to the centromeric parC DNA region, thereby forming a complex that interacts with the filament-forming actin-like ParM protein in an ATP-dependent manner, suggesting that plasmid...... movement is powered by insertional polymerization of ParM. Consistently, we find that segregating plasmids are positioned at the ends of extending ParM filaments. Thus, the process of R1 plasmid segregation in E. coli appears to be mechanistically analogous to the actin-based motility operating...

  14. Binding of heparan sulfate to Staphylococcus aureus.

    OpenAIRE

    Liang, O D; Ascencio, F; Fransson, L A; Wadström, T

    1992-01-01

    Heparan sulfate binds to proteins present on the surface of Staphylococcus aureus cells. Binding of 125I-heparan sulfate to S. aureus was time dependent, saturable, and influenced by pH and ionic strength, and cell-bound 125I-heparan sulfate was displaced by unlabelled heparan sulfate or heparin. Other glycosaminoglycans of comparable size (chondroitin sulfate and dermatan sulfate), highly glycosylated glycoprotein (hog gastric mucin), and some anionic polysaccharides (dextran sulfate and RNA...

  15. The Staphylococcus aureus “superbug”

    OpenAIRE

    FOSTER, TIMOTHY JAMES

    2004-01-01

    PUBLISHED There has been some debate about the disease-invoking potential of Staphylococcus aureus strains and whether invasive disease is associated with particularly virulent genotypes, or "superbugs." A study in this issue of the JCI describes the genotyping of a large collection of nonclinical, commensal S. aureus strains from healthy individuals in a Dutch population. Extensive study of their genetic relatedness by amplified restriction fragment typing and comparison with strains that...

  16. Triclosan Promotes Staphylococcus aureus Nasal Colonization

    OpenAIRE

    Syed, Adnan K.; Ghosh, Sudeshna; Love, Nancy G.; Boles, Blaise R.

    2014-01-01

    ABSTRACT The biocide triclosan is used in many personal care products, including toothpastes, soaps, clothing, and medical equipment. Consequently, it is present as a contaminant in the environment and has been detected in some human fluids, including serum, urine, and milk. Staphylococcus aureus is an opportunistic pathogen that colonizes the noses and throats of approximately 30% of the population. Colonization with S. aureus is known to be a risk factor for several types of infection. Here...

  17. Degradative Plasmid and Heavy Metal Resistance Plasmid Naturally Coexist in Phenol and Cyanide Assimilating Bacteria

    Directory of Open Access Journals (Sweden)

    Bahig E.  Deeb

    2009-01-01

    Full Text Available Problem statement: Heavy metals are known to be powerful inhibitors of xenobiotics biodegradation activities. Alleviation the inhibitory effect of these metals on the phenol biodegradation activities in presence of heavy metals resistant plasmid was investigated. Approach: Combination of genetic systems of degradation of xenobiotic compound and heavy metal resistance was one of the approaches to the creation of polyfunctional strains for bioremediation of soil after co-contamination with organic pollutants and heavy metals. Results: A bacterial strain Pseudomonas putida PhCN (pPhCN1, pPhCN2 had been obtained. This bacterium contained two plasmids, a 120 Kb catabolic plasmid that encode for breakdown of phenol (pPhCN1 and pPhCN2 plasmid (100 Kb that code for cadmium and copper resistant. Cyanide assimilation by this bacterium was encoded by chromosomal genes. The inhibitory effect of cadmium (Cd2+ or copper (Cu2+ on the degradation of phenol and cyanide by P. putida strains PhCN and PhCN1 (contained pPhCN1 were investigated. The resistant strain PhCN showed high ability to degrade phenol and cyanide in presence of Cd2+ or Cu2+ comparing with the sensitive strain PhCN1. In addition, Cd2+ or Cu2+ was also found to exert a strong inhibitory effect on the C23O dioxygenase enzyme activity in the presence of cyanide as a nitrogen source. Conclusion: The presence of heavy metal resistance plasmid alleviated the inhibitory effect of metals on the phenol and cyanide assimilation by resistant strain.

  18. Induced mutagenesis of plasmids and chromosomal genes inserted into plasmid DNA 1. Mutagenic effects of irradiations

    International Nuclear Information System (INIS)

    Effect of two physical agents: UV- and γ-radiation has been considered in comparison. DNA of RSF2124 plasmid, determining colcine synthesis and ampicillin resistance, was used as a model. Mutagenous effect is taken into account according to the appearance of Col--mutants, which are not capable of colicine synthesis. Lethal effect is determined according to ampicillin marker inactivation. After reisolation of plasmid DNA from mutant transformant, new traits and antibiotic resistance are preserved during subsequent transformations and reseedings of transformed colonies, which proves mutational nature of the transformations. Under short-wave UV irradiation (lambda=254 nm) of RSF2124 DNA a clear mutagenous effect is detected: relative amount of Col--mutants at the optimum for mutagenesis doses increased by a factor of 10. Under conditions of W-reactivation (additional UV-irradiation of recipient cells of wild C600 type) of lethal injuries an increase in mutagenous effect was observed, which is reliable for 95%. A distinct increase in mutagenesis (approximately by a factor of 4) is observed during UV-irradiation in small doses of only one recipient cell (a so-called indirect UV-mutagenesis). Thus, according to its ability to W- and indirect UV-mutagenesis plasmid DNA behaves as DNA of moderate phages, which can testify to their evolution relationship. Treatment of plasmid DNA with acridine orange before UV-irradiation protected only from lethal injuries. γ-irradiation of 60Co at inactivation approximately 10-2 increased by an order the yield of Col--mutants. The presence of the plasmid in a cell did not affect its UV-resistance

  19. Mutations in an antisense RNA, involved in the replication control of a repABC plasmid, that disrupt plasmid incompatibility and mediate plasmid speciation.

    Science.gov (United States)

    Rivera-Urbalejo, América; Pérez-Oseguera, Ángeles; Carreón-Rodríguez, Ofelia E; Cevallos, Miguel A

    2015-03-01

    The maintenance of large plasmid in a wide variety of alpha-proteobacteria depends on the repABC replication/segregation unit. The intergenic repB-repC region of these plasmids encodes a countertranscribed RNA (ctRNA) that modulates the transcription/translation rate of RepC, the initiator protein. The ctRNA acts as a strong incompatibility factor when expressed in trans. We followed a site directed mutagenesis approach to map those sequences of the ctRNA that are required for plasmid incompatibility and for plasmid replication control. We found that the first three nucleotides of the 5'-end of the ctRNA are essential for interactions with its target RNA. We also found that stretches of 4-5 nucleotides of non-complementarity within the first 10 nucleotides of the left arm of the ctRNA and the target RNA are sufficient to avoid plasmid incompatibility. Additionally, miniplasmid derivatives expressing ctRNAs with mutations in the 5' end or small deletions in the ctRNA are capable of controlling their own replication and coexisting with the parental plasmid. We suggest that a mechanism that could have a crucial role in the speciation process of repABC plasmids is to accumulate enough changes in this small region of the ctRNA gene to disrupt heteroduplex formation between the target RNA of one plasmid and the ctRNA of the other. Plasmids carrying these changes will not have defects in their maintenance. PMID:25644116

  20. Construction and Application of R Prime Plasmids, Carrying Different Segments of an Octopine Ti Plasmid from Agrobacterium tumefaciens, for Complementation of vir Genes

    NARCIS (Netherlands)

    Hille, Jacques; Klasen, Ina; Schilperoort, Rob

    1982-01-01

    Several R prime plasmids have been obtained with high efficiency, by enclosing the R plasmid replicator, in an R::Ti cointegrate plasmid, between two copies of the transposon Tn1831, in the same orientation. These R primes carry different segments of an octopine Ti plasmid, and are compatible with T

  1. Production and pharmaceutical formulation of plasmid DNA vaccines

    NARCIS (Netherlands)

    van der Heijden, I.

    2013-01-01

    Research leading to the thesis ‘Production and pharmaceutical formulation of plasmid DNA vaccines‘ can be divided into two parts. The first part describes the development of a Good Manufacturing Practice (GMP) compliant plasmid DNA production process of pDNA vaccines for the treatment of Human papil

  2. Homology of plasmids in strains of unicellular cyanobacteria

    NARCIS (Netherlands)

    Hondel, C.A.M.J.J. van den; Keegstra, W.; Borrias, W.E.; Arkel, G.A. van

    1979-01-01

    Six strains of unicellular cyanobacteria were examined for the presence of plasmids. Analysis of lysates of these strains by CsCl-ethidium bromide density centrifugation yielded a major chromosomal DNA band and a minor band containing covalently closed circular plasmid DNA, as shown by electron micr

  3. Plasmid-encoded copper resistance and precipitation by Mycobacterium scrofulaceum.

    OpenAIRE

    Erardi, F X; Failla, M L; Falkinham, J. O.

    1987-01-01

    A copper-tolerant Mycobacterium scrofulaceum strain was able to remove copper from culture medium by sulfate-dependent precipitation as copper sulfide. Such precipitation of copper sulfide was not observed in a derivative that lacks a 173-kilobase plasmid. In addition, the plasmid-carrying strain has a sulfate-independent copper resistance mechanism.

  4. Two novel conjugative plasmids from a single strain of Sulfolobus

    NARCIS (Netherlands)

    Erauso, G.; Stedman, K.M.; Werken, van de H.J.G.; Zillig, W.; Oost, van der J.

    2006-01-01

    Two conjugative plasmids (CPs) were isolated and characterized from the same 'Sulfolobus islandicus' strain, SOG2/4, The plasmids were separated from each other and transferred into Sulfolobus soltataricus. One has a high copy number and is not stable (pSOG1) whereas the other has a low copy number

  5. Inc A/C Plasmids in Multidrug resistant Salmonella enterica

    Science.gov (United States)

    Bacteria plasmids are fragments of extra-chromosomal double stranded deoxyribonucleic acid (DNA) that can contain a variety of genes beneficial to the survival of the host bacteria. Classification and tracking of plasmids is beneficial because they are potentially a medium of horizontal gene transf...

  6. Transformation of Bacillus subtilis by single-stranded plasmid DNA.

    OpenAIRE

    Rudolph, C F; Schmidt, B J; Saunders, C W

    1986-01-01

    The single-stranded form of a pE194-based plasmid transformed Bacillus subtilis protoplasts at least as efficiently as did the double-stranded plasmid, but the single-stranded form did not detectably transform B. subtilis competent cells.

  7. Curing of the 2 mu DNA plasmid from Saccharomyces cerevisiae.

    OpenAIRE

    Toh-e, A; Wickner, R B

    1981-01-01

    The 2 mu DNA plasmid is often eliminated from yeast cells when they are transformed with the 2 mu DNA-LEU2-pMB9 composite plasmid pJDB219. Since pJDB219 is subsequently lost with high frequency, derivatives lacking all 2 mu DNA can be prepared from any strain.

  8. Purification of large plasmids with methacrylate monolithic columns.

    Science.gov (United States)

    Krajnc, Nika Lendero; Smrekar, Franci; Cerne, Jasmina; Raspor, Peter; Modic, Martina; Krgovic, Danijela; Strancar, Ales; Podgornik, Ales

    2009-08-01

    The rapid evolution of gene therapy and DNA vaccines results in an increasing interest in producing large quantities of pharmaceutical grade plasmid DNA. Most current clinical trials involve plasmids of 10 kb or smaller in size, however, future requirements for multigene vectors including extensive control regions may require the production of larger plasmids, e. g., 20 kb and bigger. The objective of this study was to examine certain process conditions for purification of large plasmids with the size of up to 93 kb. Since there is a lack of knowledge about production and purification of bigger plasmid DNA, cell lysis and storage conditions were investigated. The impact of chromatographic system and methacrylate monolithic column on the degradation of plasmid molecules under nonbinding conditions at different flow rates was studied. Furthermore, capacity measurements varying salt concentration in loading buffer were performed and the capacities up to 13 mg of plasmid per mL of the monolithic column were obtained. The capacity flow independence in the range from 130 to 370 cm/h was observed. Using high resolution monolithic column the separation of linear and supercoiled isoforms of large plasmids was obtained. Last but not least, since the baseline separation of RNA and pDNA was achieved, the one step purification on larger CIM DEAE 8 mL tube monolithic column was performed and the fractions were analyzed by CIM analytical monolithic columns. PMID:19598166

  9. Potassium Uptake Modulates Staphylococcus aureus Metabolism.

    Science.gov (United States)

    Gries, Casey M; Sadykov, Marat R; Bulock, Logan L; Chaudhari, Sujata S; Thomas, Vinai C; Bose, Jeffrey L; Bayles, Kenneth W

    2016-01-01

    As a leading cause of community-associated and nosocomial infections, Staphylococcus aureus requires sophisticated mechanisms that function to maintain cellular homeostasis in response to its exposure to changing environmental conditions. The adaptation to stress and maintenance of homeostasis depend largely on membrane activity, including supporting electrochemical gradients and synthesis of ATP. This is largely achieved through potassium (K(+)) transport, which plays an essential role in maintaining chemiosmotic homeostasis, affects antimicrobial resistance, and contributes to fitness in vivo. Here, we report that S. aureus Ktr-mediated K(+) uptake is necessary for maintaining cytoplasmic pH and the establishment of a proton motive force. Metabolite analyses revealed that K(+) deficiency affects both metabolic and energy states of S. aureus by impairing oxidative phosphorylation and directing carbon flux toward substrate-level phosphorylation. Taken together, these results underline the importance of K(+) uptake in maintaining essential components of S. aureus metabolism. IMPORTANCE Previous studies describing mechanisms for K(+) uptake in S. aureus revealed that the Ktr-mediated K(+) transport system was required for normal growth under alkaline conditions but not under neutral or acidic conditions. This work focuses on the effect of K(+) uptake on S. aureus metabolism, including intracellular pH and carbon flux, and is the first to utilize a pH-dependent green fluorescent protein (GFP) to measure S. aureus cytoplasmic pH. These studies highlight the role of K(+) uptake in supporting proton efflux under alkaline conditions and uncover a critical role for K(+) uptake in establishing efficient carbon utilization. PMID:27340697

  10. Plasmid DNA induces dodecyl triethyl ammonium bromide to aggregate into vesicle

    Institute of Scientific and Technical Information of China (English)

    Xiang Mei Ran; Xia Guo; Jia Tong Ding

    2012-01-01

    Single-chained cationic surfactant dodecyl triethyl ammonium bromide and plasmid DNA together can form vesicles once the concentration of plasmid DNA reaches a critical value (Ccvc).Bigger the size of plasmid DNA,higher the value of Ccvc.

  11. Construction of a eukaryotic expression plasmid of Humanin

    Institute of Scientific and Technical Information of China (English)

    LUO Ben-yan; CHEN Xiang-ming; TANG Min; CHEN Feng; CHEN Zhi

    2005-01-01

    Objective: To construct a eukaryotic expression plasmid pcDNA3.1 (-)-Humanin. Methods: The recombinant plasm pGEMEX- 1-Humanin was digested with restriction endonucleases BamH I and Hind Ⅲ and the Humanin gene fragments, abo 100 bp length, were obtained. Then the Humanin gene fragments were inserted into eukaryotic expression vector pcDNA3.1 (-) and the recombinant plasmids pcDNA3. l(-)-Humanin were identified by sequencing. Results: Recombinant plasmid DNA succesfully produced a band which had the same size as that of the Humanin positive control. The sequence of recombinant plasmids accorded with the Humnain gene sequence. Conclusions: A eukaryotic expression plasmid of Humanin was successfully constructed.

  12. Permissiveness of soil microbial communities towards broad host range plasmids

    DEFF Research Database (Denmark)

    Klümper, Uli

    of fluorescently tagged conjugative plasmids into a soil microbial community in solid-surface filter matings under maximized cell-to-cell contact, followed by quantification of transfer events through advanced fluorescent microscopy, isolation of transconjugants through triple-gated fluorescent activated cell...... using model BHR plasmid pKJK5 introduced through the γ-proteobacterial donor E. coli. I found that the permissiveness towards plasmids was modified through stress on a taxon specific basis and cannot be generally predicted for the whole community. The response of the phylogenetic group was specific...... between phylogenetically distant groups. Finally, I extended the high-throughput method to quantify the potential of a microbial community to actively mobilize and transfer exogenous mobilizable plasmids to its indigenous members. I evaluated the transfer frequency of model plasmid RSF1010 by comparing...

  13. Radiosensitivity of plasmid DNA: role of topology and concentration

    International Nuclear Information System (INIS)

    Using the plasmid relaxation assay, the induction of single strand breaks (SSB) by ionizing radiation was investigated in two plasmids of different length, pBS and pSP189. The dose-response was linear for both plasmids but pSP189 exhibited a three times higher sensitivity than mBS. This disparity may be explained by a reduced accessibility to hydroxyl radicals due to a different topology of each plasmid, i.e. degree of compaction, as observed with electron microscopy. pBS plasmid was also exposed at various DNA concentrations to γ rays. The yield of SSB decreased with increasing concentration, suggesting a diminution in the amount of hydroxyl radicals efficient for radiolytic attack. This effect of concentration was also observed with densely ionizing radiation. In conclusion, the accessibility of DNA is a key-parameter in the formation of damage in vitro and in vivo as well. (authors)

  14. Positive selection and compensatory adaptation interact to stabilize non-transmissible plasmids

    OpenAIRE

    Millan, A. San; Peña-Miller, R.; Toll-Riera, M.; Halbert, Z. V.; McLean, A R; Cooper, B. S.; Maclean, R. C.

    2014-01-01

    Plasmids are important drivers of bacterial evolution, but it is challenging to understand how plasmids persist over the long term because plasmid carriage is costly. Classical models predict that horizontal transfer is necessary for plasmid persistence, but recent work shows that almost half of plasmids are non-transmissible. Here we use a combination of mathematical modelling and experimental evolution to investigate how a costly, non-transmissible plasmid, pNUK73, can be maintained in popu...

  15. Requirements for the formation of plasmid-transducing particles of Bacillus subtilis bacteriophage SPP1.

    OpenAIRE

    Alonso, J.C.; Lüder, G; Trautner, T A

    1986-01-01

    We had previously proposed that the production of concatemeric plasmid DNA in plasmid-transducing SPP1 particles is a consequence of phage-directed rolling-circle-type replication of plasmid DNA. The production of such DNA was greatly enhanced when DNA/DNA homology was provided between phage and plasmid DNAs (facilitation of transduction). Here we present evidence that synthesis of concatemeric plasmid DNA can proceed after phage infection under conditions non-permissive for plasmid replicati...

  16. Construction of Streptococcus lactis subsp. lactis Strains with a Single Plasmid Associated with Mucoid Phenotype

    OpenAIRE

    von Wright, Atte; Tynkkynen, Soile

    1987-01-01

    Lactose-fermenting mucoid (Lac+ Muc+) variants of plasmid-free Streptococcus lactis subsp. lactis MG1614 were obtained by protoplast transformation with total plasmid DNA from Muc+S. lactis subsp. cremoris ARH87. By using plasmid DNA from these variants for further transformations followed by novobiocininduced plasmid curing, Lac− Muc+ MG1614 strains containing only a single 30-megadalton plasmid could be constructed. This plasmid, designated pVS5, appeared to be associated with the Muc+ phen...

  17. Plasmid stability in immobilized and free recombinant Escherichia coli JM105(pKK223-200): importance of oxygen diffusion, growth rate, and plasmid copy number.

    OpenAIRE

    de Taxis du Poët, P; Arcand, Y; Bernier, R.; Barbotin, J N; Thomas, D.

    1987-01-01

    Stability of the plasmid pKK223-200 in Escherichia coli JM105 was studied for both free and immobilized cells during continuous culture. The relationship between plasmid copy number, xylanase activity, which was coded for by the plasmid, and growth rate and culture conditions involved complex interactions which determined the plasmid stability. Generally, the plasmid stability was enhanced in cultured immobilized cells compared with free-cell cultures. This stability was associated with modif...

  18. Bifurcation Analysis of a Chemostat Model of Plasmid-Bearing and Plasmid-Free Competition with Pulsed Input

    Directory of Open Access Journals (Sweden)

    Zhong Zhao

    2014-01-01

    to the stability of the boundary periodic solution. By use of standard techniques of bifurcation theory, the periodic oscillations in substrate, plasmid-bearing, and plasmid-free organisms are shown when some conditions are satisfied. Our results can be applied to control bioreactor aimed at producing commercial producers through genetically altered organisms.

  19. Intra-cellular Staphylococcus aureus alone causes infection in vivo

    Directory of Open Access Journals (Sweden)

    T Hamza

    2013-07-01

    Full Text Available Chronic and recurrent bone infections occur frequently but have not been explained. Staphylococcus aureus (S. aureus is often found among chronic and recurrent infections and may be responsible for such infections. One possible reason is that S. aureus can internalize and survive within host cells and by doing so, S. aureus can evade both host defense mechanisms and most conventional antibiotic treatments. In this study, we hypothesized that intra-cellular S. aureus could induce infections in vivo. Osteoblasts were infected with S. aureus and, after eliminating extra-cellular S. aureus, inoculated into an open fracture rat model. Bacterial cultures and radiographic observations at post-operative day 21 confirmed local bone infections in animals inoculated with intra-cellular S. aureus within osteoblasts alone. We present direct in vivo evidence that intra-cellular S. aureus could be sufficient to induce bone infection in animals; we found that intra-cellular S. aureus inoculation of as low as 102 colony forming units could induce severe bone infections. Our data may suggest that intra-cellular S. aureus can “hide” in host cells during symptom-free periods and, under certain conditions, they may escape and lead to infection recurrence. Intra-cellular S. aureus therefore could play an important role in the pathogenesis of S. aureus infections, especially those chronic and recurrent infections in which disease episodes may be separated by weeks, months, or even years.

  20. Antimicrobial (Drug) Resistance: Methicillin-Resistant Staphylococcus aureus (MRSA)

    Science.gov (United States)

    ... Marketing Share this: Main Content Area Methicillin-Resistant Staphylococcus aureus (MRSA) During the past four decades, methicillin-resistant Staphylococcus aureus , or MRSA, has evolved from a controllable ...

  1. Evolution of methicillin-resistant Staphylococcus aureus towards increasing resistance

    DEFF Research Database (Denmark)

    Strommenger, Birgit; Bartels, Mette Damkjær; Kurt, Kevin;

    2014-01-01

    To elucidate the evolutionary history of Staphylococcus aureus clonal complex (CC) 8, which encompasses several globally distributed epidemic lineages, including hospital-associated methicillin-resistant S. aureus (MRSA) and the highly prevalent community-associated MRSA clone USA300....

  2. Classification of Healthcare-Associated Staphylococcus aureus Bacteremia

    DEFF Research Database (Denmark)

    Smit, Jesper; Søgaard, Mette; Schønheyder, Henrik Carl; Nielsen, Henrik; Thomsen, Reimar Wernich

    2016-01-01

    We investigated whether different definitions of healthcare-associated infection influenced the prevalence, characteristics, and mortality of patients with Staphylococcus aureus bacteremia. With different definitions, the proportion of patients classified as having healthcare-associated S. aureus...

  3. Comparative Efficacy of Ceftaroline with Linezolid against Staphylococcus Aureus and Methicillin Resistant Staphylococcus Aureus

    International Nuclear Information System (INIS)

    Objective:To compare the in vitro antimicrobial efficacy of ceftaroline with linezolid against Staphylococcus aureus and methicillin resistant Staphylococcus aureus. Study Design: Quasi-experimental study. Place and Duration of Study: Microbiology Department, Army Medical College, Rawalpindi, from January to December 2013. Methodology: Clinical samples from respiratory tract, blood, pus and various catheter tips routinely received in the Department of Microbiology, Army Medical College, Rawalpindi were innoculated on blood and MacConkey agar. Staphylococcus aureus was identified by colony morphology, Gram reaction, catalase test and coagulase test. Methicillin resistant Staphylococcus aureus detection was done by modified Kirby Bauer disc diffusion method using cefoxitin disc (30g) and the isolates were considered methicillin resistant if the zone of inhibition around cefoxitin disc was /sup 2/ 21 mm. Bacterial suspensions of 56 Staphylococcus aureus isolates and 50 MRSA isolates were prepared, which were standardized equal to 0.5 McFarland's turbidity standard and inoculated on Mueller-Hinton agar plates followed by application of ceftaroline and linezolid disc (Oxoid, UK), according to manufacturer's instructions. The plates were then incubated at 37 Degree C aerobically for 18 - 24 hours. Diameters of inhibition zone were measured and interpretated as per Clinical and Laboratory Standards Institute (CLSI) guidelines. Results: Out of 106 isolates all of the 56 Staphylococcus aureus (100%) were sensitive to ceftaroline and linezolid. However, out of 50 methicillin resistant Staphylococcus aureus, 48 (96%) were sensitive to ceftaroline whereas, 49 (98%) were sensitive to linezolid. Conclusion: Ceftaroline is equally effective as linezolid against Staphylococcus aureus and methicillin resistant Staphylococcus aureus. (author)

  4. Plasmid Copy Number Determination by Quantitative Polymerase Chain Reaction.

    Science.gov (United States)

    Anindyajati; Artarini, A Anita; Riani, Catur; Retnoningrum, Debbie S

    2016-01-01

    Recombinant therapeutic proteins are biopharmaceutical products that develop rapidly for years. Recombinant protein production in certain hosts requires vector expression harboring the gene encoding the corresponding protein. Escherichia coli is the prokaryote organism mostly used in recombinant protein production, commonly using a plasmid as the expression vector. Recombinant protein production is affected by plasmid copy number harboring the encoded gene, hence the determination of plasmid copy number also plays an important role in establishing a recombinant protein production system. On the industrial scale, a low copy number of plasmids are more suitable due to their better stability. In the previous study we constructed pCAD, a plasmid derived from the low copy number pBR322 plasmid. This study was aimed to confirm pCAD's copy number by quantitative polymerase chain reaction (qPCR). Plasmid copy number was determined by comparing the quantification signal from the plasmid to those from the chromosome. Copy number was then calculated by using a known copy number plasmid as a standard. Two pairs of primers, called tdk and ori, were designed for targeting a single gene tdk in the chromosome and a conserved domain in the plasmid's ori, respectively. Primer quality was analyzed in silico using PrimerSelect DNASTAR and PraTo software prior to in vitro evaluation on primer specificity and efficiency as well as optimization of qPCR conditions. Plasmid copy number determination was conducted on E. coli lysates harboring each plasmid, with the number of cells ranging from 10(2)-10(5) cells/μL. Cells were lysed by incubation at 95ºC for 10 minutes, followed by immediate freezing at -4°C. pBR322 plasmid with the copy number of ~19 copies/cell was used as the standard, while pJExpress414-sod plasmid possessing the high copy number pUC ori was also determined to test the method being used. In silico analysis based on primer-primer and primer-template interactions showed

  5. Plasmid-associated sensitivity of Bacillus thuringiensis to UV light

    International Nuclear Information System (INIS)

    Spores and vegetative cells of Bacillus thuringiensis were more sensitive to UV light than were spores or cells of plasmid-cured B. thuringiensis strains or of the closely related Bacillus cereus. Introduction of B. thuringiensis plasmids into B. cereus by cell mating increased the UV sensitivity of the cells and spores. Protoxins encoded by one or more B. thuringiensis plasmids were not involved in spore sensitivity, since a B. thuringiensis strain conditional for protoxin accumulation was equally sensitive at the permissive and nonpermissive temperatures. In addition, introduction of either a cloned protoxin gene, the cloning vector, or another plasmid not containing a protoxin gene into a plasmid-cured strain of B. thuringiensis all increased the UV sensitivity of the spores. Although the variety of small, acid-soluble proteins was the same in the spores of all strains examined, the quantity of dipicolinic acid was about twice as high in the plasmid-containing strains, and this may account for the differences in UV sensitivity of the spores. The cells of some strains harboring only B. thuringiensis plasmids were much more sensitive than cells of any of the other strains, and the differences were much greater than observed with spores

  6. Investigation of plasmid-induced growth defect in Pseudomonas putida.

    Science.gov (United States)

    Mi, Jia; Sydow, Anne; Schempp, Florence; Becher, Daniela; Schewe, Hendrik; Schrader, Jens; Buchhaupt, Markus

    2016-08-10

    Genetic engineering in bacteria mainly relies on the use of plasmids. But despite their pervasive use for physiological studies as well as for the design and optimization of industrially used production strains, only limited information about plasmid induced growth defects is available for different replicons and organisms. Here, we present the identification and characterization of such a phenomenon for Pseudomonas putida transformants carrying the pBBR1-derived plasmid pMiS1. We identified the kanamycin resistance gene and the transcription factor encoding rhaR gene to be causal for the growth defect in P. putida. In contrast, this effect was not observed in Escherichia coli. The plasmid-induced growth defect was eliminated after introduction of a mutation in the plasmid-encoded rep gene, thus enabling construction of the non-toxic variant pMiS4. GFP reporters construct analyses and qPCR experiments revealed a distinctly lowered plasmid copy number for pMiS4, which is probably the reason for alleviation of the growth defect by this mutation. Our work expands the knowledge about plasmid-induced growth defects and provides a useful low-copy pBBR1 replicon variant. PMID:27287537

  7. Sociobiological control of plasmid copy number in bacteria.

    Directory of Open Access Journals (Sweden)

    Mukta M Watve

    Full Text Available All genes critical for plasmid replication regulation are located on the plasmid rather than on the host chromosome. It is possible therefore that there can be copy-up "cheater" mutants. In spite of this possibility, low copy number plasmids appear to exist stably in host populations. We examined this paradox using a multilevel selection model. Simulations showed that, a slightly higher copy number mutant could out-compete the wild type. Consequently, another mutant with still higher copy number could invade the first invader. However, the realized benefit of increasing intra-host fitness was saturating whereas that of inter-host fitness was exponential. As a result, above a threshold, intra-host selection was overcompensated by inter-host selection and the low copy number wild type plasmid could back invade a very high copy number plasmid. This led to a rock-paper-scissor (RPS like situation that allowed the coexistence of plasmids with varied copy numbers. Furthermore, another type of cheater that had lost the genes required for conjugation but could hitchhike on a conjugal plasmid, could further reduce the advantage of copy-up mutants. These sociobiological interactions may compliment molecular mechanisms of replication regulation in stabilizing the copy numbers.

  8. A single copy integration vector that integrates at an engineered site on the Staphylococcus aureus chromosome

    Directory of Open Access Journals (Sweden)

    Lei Mei G

    2012-01-01

    Full Text Available Abstract Background Single-copy integration vectors based upon the site-specific recombination systems of bacteriophage are invaluable tools in the study of bacterial pathogenesis. The utility of such vectors is often limited, however, by the fact that integration often results in the inactivation of bacterial genes or has undesirable effects on gene transcription. The aim of this study is to develop an integration vector that does not have a detectable effect on gene transcription upon integration. Findings We have developed a single-copy integration system that enables the cloning vector to integrate at a specific engineered site, within an untranscribed intergenic region, in the chromosome of Staphylococcus aureus. This system is based on the lysogenic phage L54a site-specific recombination system in which the L54a phage (attP and chromosome (attB attachment sites, which share an 18-bp identical core sequence, were modified with identical mutations. The integration vector, pLL102, was constructed to contain the modified L54a attP site (attP2 that was altered at 5 nucleotide positions within the core sequence. In the recipient strain, the similarly modified attB site (attB2 was inserted in an intergenic region devoid of detectable transcription read-through. Integration of the vector, which is unable to replicate in S. aureus extrachromosomally, was achieved by providing the L54a integrase gene in a plasmid in the recipient. We showed that pLL102 integrated specifically at the engineered site rather than at the native L54a attB site and that integration did not have a significant effect on transcription of genes immediately upstream or downstream of the integration site. Conclusions In this work, we describe an E. coli-S. aureus shuttle vector that can be used to introduce any cloned gene into the S. aureus chromosome at a select site without affecting gene expression. The vector should be useful for genetic manipulation of S. aureus and for

  9. Characterization of a Mouse-Adapted Staphylococcus aureus Strain

    OpenAIRE

    Holtfreter, Silva; Fiona J Radcliff; Grumann, Dorothee; Read, Hannah; Johnson, Sarah; Monecke, Stefan; Ritchie, Stephen; Clow, Fiona; Goerke, Christiane; Bröker, Barbara M.; Fraser, John D.; Wiles, Siouxsie

    2013-01-01

    More effective antibiotics and a protective vaccine are desperately needed to combat the ‘superbug’ Staphylococcus aureus. While in vivo pathogenicity studies routinely involve infection of mice with human S. aureus isolates, recent genetic studies have demonstrated that S. aureus lineages are largely host-specific. The use of such animal-adapted S. aureus strains may therefore be a promising approach for developing more clinically relevant animal infection models. We have isolated a mouse-ad...

  10. Meticillin-resistant Staphylococcus aureus (MRSA)

    DEFF Research Database (Denmark)

    Stefani, Stefania; Chung, Doo Ryeon; Lindsay, Jodi A;

    2012-01-01

    This article reviews recent findings on the global epidemiology of healthcare-acquired/associated (HA), community-acquired/associated (CA) and livestock-associated (LA) meticillin-resistant Staphylococcus aureus (MRSA) and aims to reach a consensus regarding the harmonisation of typing methods...... health. Continuous efforts to understand the changing epidemiology of S. aureus infection in humans and animals are therefore necessary, not only for appropriate antimicrobial treatment and effective infection control but also to monitor the evolution of the species. The group made several consensus...

  11. A sensitive assay for Staphylococcus aureus nucleases

    International Nuclear Information System (INIS)

    A sensitive assay for staphylococcal nuclease involving incubation of the enzyme sample with heat-denatured [3H] thymidine labelled DNA from E.coli, precipitation with trichloroacetic acid and measurement of the radioactivity of acid-soluble nucleotides released has been developed. The assay is sensitive enough to be used for comparing the levels of nucleases elaborated by different strains of S. aureus as well as for determining the extent of contamination of S. aureus in food and water samples even at levels at which the conventional spectrophotometric and toluidine blue-DNA methods are totally inadequate. (author). 26 refs., 3 figs ., 3 tabs

  12. Staphylococcus aureus bacteremia in hemodialysis patients.

    Science.gov (United States)

    Latos, D L; Stone, W J; Alford, R H

    1977-01-01

    Fifteen male hemodialysis patients developed 21 episodes of S. aureus bacteremia. Infections involving vascular access were responsible for 65% of initial bacteremias. The arteriovenous fistula was the most prevalent type of access used, and thus was responsible for the majority of these illnesses. Phage typing indicated that recurrent episodes were due to reinfection rather than relapse. Complications included endocarditis, osteomyelitis, septic embolism, and pericarditis. One patient died of infectious complications. It is recommended that hemodialysis patients developing bacteremia due to S. aureus receive at least 6 weeks of beta lactamase-resistant antimicrobial therapy. PMID:608860

  13. Key genetic elements and regulation systems in methicillin-resistant Staphylococcus aureus.

    Science.gov (United States)

    Hao, Haihong; Dai, Menghong; Wang, Yulian; Huang, Lingli; Yuan, Zonghui

    2012-11-01

    Methicillin-resistant Staphylococcus aureus (MRSA), popularly known as a type of superbug, has been a serious challenge for animal and human health. S. aureus has developed methicillin resistance mainly by expression of β-lactamase and PBP2a, which is regulated by the blaZ-blaI-blaR1 and mecA-mecI-mecRI systems. Other genetic elements, including murE and femA, also participate in expression of methicillin resistance, but the mechanism remains unclear. The evolution of the staphylococcal cassette chromosome mec determines the epidemiological risk of MRSA. The plasmid-located gene cfr might contribute to multiresistance and transmission of MRSA. Some virulence factors, including Panton-Valentine leukocidin, phenol-soluble modulin, arginine catabolic mobile element and other toxin elements enhance the pathogenesis and fitness of MRSA. Two-component regulation systems (agr, saeRS and vraRS) are closely associated with pathogenesis and drug resistance of MRSA. The systematic exploration of key genetic elements and regulation systems involved in multidrug resistance/pathogenesis/transmission of MRSA is conclusively integrated into this review, providing fundamental information for the development of new antimicrobial agents and the establishment of reasonable antibiotic stewardship to reduce the risk of this superbug. PMID:23075449

  14. 21 CFR 866.3700 - Staphylococcus aureus serological reagents.

    Science.gov (United States)

    2010-04-01

    ... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Staphylococcus aureus serological reagents. 866... Staphylococcus aureus serological reagents. (a) Identification. Staphylococcus aureus serological reagents are... diagnosis of disease caused by this bacterium belonging to the genus Staphylococcus and...

  15. 9 CFR 113.115 - Staphylococcus Aureus Bacterin-Toxoid.

    Science.gov (United States)

    2010-01-01

    ... 9 Animals and Animal Products 1 2010-01-01 2010-01-01 false Staphylococcus Aureus Bacterin-Toxoid... REQUIREMENTS Inactivated Bacterial Products § 113.115 Staphylococcus Aureus Bacterin-Toxoid. Staphylococcus... Staphylococcus aureus which has been inactivated and is nontoxic. Each serial of biological product...

  16. A pig model of acute Staphylococcus aureus induced pyemia

    DEFF Research Database (Denmark)

    Nielsen, O. L.; Iburg, T.; Aalbæk, B.;

    2009-01-01

    Background: Sepsis caused by Staphylococcus aureus constitutes an important cause of morbidity and mortality in humans, and the incidence of this disease-entity is increasing. In this paper we describe the initial microbial dynamics and lesions in pigs experimentally infected with S. aureus....... aureus isolated from man and an extension of the timeframe aiming at inducing sepsis, severe sepsis and septic shock....

  17. Multidrug-Resistant Staphylococcus aureus in US Meat and Poultry

    OpenAIRE

    Waters, Andrew E.; Contente-Cuomo, Tania; Buchhagen, Jordan; Liu, Cindy M.; Watson, Lindsey; Pearce, Kimberly; Foster, Jeffrey T.; Bowers, Jolene; Driebe, Elizabeth M; Engelthaler, David M.; Keim, Paul S; Lance B Price

    2011-01-01

    We characterized the prevalence, antibiotic susceptibility profiles, and genotypes of Staphylococcus aureus among US meat and poultry samples (n = 136). S. aureus contaminated 47% of samples, and multidrug resistance was common among isolates (52%). S. aureus genotypes and resistance profiles differed significantly among sample types, suggesting food animal–specific contamination.

  18. Separation of plasmid DNA topoisomers by multimodal chromatography.

    Science.gov (United States)

    Silva-Santos, A Rita; Alves, Cláudia P A; Prazeres, Duarte Miguel F; Azevedo, Ana M

    2016-06-15

    The ability to analyze the distribution of topoisomers in a plasmid DNA sample is important when evaluating the quality of preparations intended for gene therapy and DNA vaccination or when performing biochemical studies on the action of topoisomerases and gyrases. Here, we describe the separation of supercoiled (sc) and open circular (oc) topoisomers by multimodal chromatography. A medium modified with the ligand N-benzyl-N-methyl ethanolamine and an elution scheme with increasing NaCl concentration are used to accomplish the baseline separation of sc and oc plasmid. The utility of the method is demonstrated by quantitating topoisomers in a purified plasmid sample. PMID:27033004

  19. Linearization of donor DNA during plasmid transformation in Neisseria gonorrhoeae.

    OpenAIRE

    Biswas, G D; Burnstein, K. L.; Sparling, P F

    1986-01-01

    We examined the fate of plasmid DNA after uptake during transformation in Neisseria gonorrhoeae. An 11.5-kilobase plasmid, pFA10, was processed to linear double-stranded DNA during uptake by competent cells, but cleavage of pFA10 was not site specific. A minority of pFA10 entered as open circles. A 42-kilobase plasmid, pFA14, was degraded into small fragments during uptake; no intracellular circular forms of pFA14 were evident. Since pFA10 DNA linearized by a restriction enzyme was not furthe...

  20. 金黄色葡萄球菌FnBP配体结合区基因的克隆及其原核表达%Cloning and Prokaryotic Expression of FnBP Ligand Binding Gene of Staphylococcus aureus

    Institute of Scientific and Technical Information of China (English)

    尹荣兰; 杨正涛; 张艳晶; 刘辉; 刘珊; 杨琦; 曹永国; 张乃生

    2008-01-01

    [Objective] The study aimed to clone the FnBP ligand binding gene of Staphylococcus aureus and run prokaryotic expression by constructing a prokaryotic expression vector. [Method] The gene encoding FnBP ligand binding gene was amplified from S.aureus chromosomal DNA by PCR technique. After T-A cloning, plasmid pMD18- FnBP was constructed. pMD18- FnBP and pET28a(+)were digested by BamH Ⅰ and EcoR Ⅰ double enzymes, then the purified FnBP ligand binding gene was subcloned into the expression vector pET28a(+), and the prokaryotic expression vector pET28a-FnBP was thus constructed. The constructed plasmid pET28a-FnBP was transformed into Escherichia coli BL21(DE3) competent cells. The bacterium was induced by IPTG and the expressed products were analyzed by SDS-PAGE and Western blot. [Result] The gene fragment with the length of 370 bp was amplified by PCR approach. One approximately 30 kD exogenous protein was observed in SDS-PAGE analysis. Western blot analysis indicates the protein has antigenicity of S.aureus. [Conclusion] The FnBP ligand binding gene of S.aureus was successfully cloned and expressed in prokaryotic cells.

  1. Action of cholecalciferol and alpha-tocopherol on Staphylococcus aureus efflux pumps.

    Science.gov (United States)

    Tintino, Saulo R; Morais-Tintino, Cícera D; Campina, Fábia F; Pereira, Raimundo L; Costa, Maria do S; Braga, Maria Flaviana B M; Limaverde, Paulo W; Andrade, Jacqueline C; Siqueira-Junior, José P; Coutinho, Henrique Douglas Melo; Balbino, Valdir Q; Leal-Balbino, Tereza C; Ribeiro-Filho, Jaime; Quintans-Júnior, Lucindo J

    2016-01-01

    Alpha-tocopherol is one the most abundant and biologically active isoforms of vitamin E. This compound is a potent antioxidant and one of most studied isoforms of vitamin E. Vitamin D3 (cholecalciferol) is an important nutrient for calcium homeostasis and bone health, that has also been recognized as a potent modulator of the immune response. Methicillin-resistant Staphylococcus aureus (MRSA) is the most important causative agent of both nosocomial and community-acquired infections. The aim of this study was to evaluate the inhibitory effect of alpha-tocopherol and cholecalciferol on both S. aureus and multidrug resistant S. aureus efflux pumps. The RN4220 strain has the plasmid pUL5054 that is the carrier of gene that encodes the macrolide resistance protein (an efflux pump) MsrA; the IS-58 strain possesses the TetK tetracycline efflux protein in its genome and the 1199B strain resists to hydrophilic fluoroquinolones via a NorA-mediated mechanism. The antibacterial activity was evaluated by determining the Minimal Inhibitory Concentration (MIC) and a possible inhibition of efflux pumps was associated to a reduction of the MIC. In this work we observed that in the presence of the treatments there was a decrease in the MIC for the RN4220 and IS-58 strains, suggesting that the substances presented an inhibitory effect on the efflux pumps of these strains. Significant efforts have been done to identify efflux pump inhibitors (EPIs) from natural sources and, therefore, the antibacterial properties of cholecalciferol and alpha-tocopherol might be attributed to a direct effect on the bacterial cell depending on their amphipathic structure. PMID:27298617

  2. Risk factors for Staphylococcus aureus and methicillin-resistant S aureus colonization among health care workers in pediatrics departments.

    Science.gov (United States)

    Gomes, Ivete Martins; Marlow, Mariel Asbury; Pinheiro, Marcos Gabriel; de Freitas, Maria de Fátima Nogueira; Fonseca, Fernanda Fernandes; Cardoso, Claudete Aparecida Araújo; Aguiar-Alves, Fábio

    2014-08-01

    Risk factors for Staphylococcus aureus and methicillin-resistant S aureus (MRSA) were evaluated for 178 health care workers from a public hospital pediatrics department in Brazil. Colonization rates were 33.1% for S aureus and 5.1% for MRSA. Risk factors for S aureus colonization differed from those for MRSA. Results suggest nurses with prolonged pediatric patient contact in inpatient units are at higher risk for MRSA colonization. PMID:25087145

  3. Plasmid pGA1 from Corynebacterium glutamicum codes for a gene product that positively influences plasmid copy number.

    OpenAIRE

    Nesvera, J; Pátek, M; Hochmannová, J; Abrhámová, Z; Becvárová, V; Jelínkova, M; Vohradský, J

    1997-01-01

    The complete nucleotide sequence (4,826 bp) of the cryptic plasmid pGA1 from Corynebacterium glutamicum was determined. DNA sequence analysis revealed four putative coding regions (open reading frame A [ORFA], ORFA2, ORFB, and ORFC). ORFC was identified as a rep gene coding for an initiator of plasmid replication (Rep) according to the high level of homology of its deduced amino acid sequence with the Rep proteins of plasmids pSR1 (from C. glutamicum) and pNG2 (from Corynebacterium diphtheria...

  4. Cloning and Analysis of a Large Plasmid pBMB165 from Bacillus thuringiensis Revealed a Novel Plasmid Organization

    OpenAIRE

    Yueying Wang; Donghai Peng; Zhaoxia Dong; Lei Zhu; Suxia Guo; Ming Sun

    2013-01-01

    In this study, we report a rapid cloning strategy for large native plasmids via a contig linkage map by BAC libraries. Using this method, we cloned a large plasmid pBMB165 from Bacillus thuringiensis serovar tenebrionis strain YBT-1765. Complete sequencing showed that pBMB165 is 77,627 bp long with a GC-content of 35.36%, and contains 103 open reading frames (ORFs). Sequence analysis and comparison reveals that pBMB165 represents a novel plasmid organization: it mainly consists of a pXO2-like...

  5. Staphylococcus aureus resistente a la meticilina (SARM)

    Centers for Disease Control (CDC) Podcasts

    2007-10-22

    Datos importantes sobre las infecciones por SARM en Estados Unidos, en las escuelas y los entornos médicos. (Title: Methicillin-resistant Staphylococcus aureus (MRSA)Created: 10/2007).  Created: 10/22/2007 by National Center for Preparedness, Detection, and Control of Infectious Diseases.   Date Released: 11/9/2007.

  6. Staphylococcus aureus enterotoxins A- and B

    DEFF Research Database (Denmark)

    Danielsen, E Michael; Hansen, Gert H; Karlsdóttir, Edda

    2013-01-01

    Enterotoxins of Staphylococcus aureus are among the most common causes of food poisoning. Acting as superantigens they intoxicate the organism by causing a massive uncontrolled T cell activation that ultimately may lead to toxic shock and death. In contrast to our detailed knowledge regarding...

  7. Staphylococcus aureus vaccines: Deviating from the carol.

    Science.gov (United States)

    Missiakas, Dominique; Schneewind, Olaf

    2016-08-22

    Staphylococcus aureus, a commensal of the human nasopharynx and skin, also causes invasive disease, most frequently skin and soft tissue infections. Invasive disease caused by drug-resistant strains, designated MRSA (methicillin-resistant S. aureus), is associated with failure of antibiotic therapy and elevated mortality. Here we review polysaccharide-conjugate and subunit vaccines that were designed to prevent S. aureus infection in patients at risk of bacteremia or surgical wound infection but failed to reach their clinical endpoints. We also discuss vaccines with ongoing trials for combinations of polysaccharide-conjugates and subunits. S. aureus colonization and invasive disease are not associated with the development of protective immune responses, which is attributable to a large spectrum of immune evasion factors. Two evasive strategies, assembly of protective fibrin shields via coagulases and protein A-mediated B cell superantigen activity, are discussed as possible vaccine targets. Although correlates for protective immunity are not yet known, opsonophagocytic killing of staphylococci by phagocytic cells offers opportunities to establish such criteria. PMID:27526714

  8. Carriage of Staphylococcus aureus in the elderly

    NARCIS (Netherlands)

    R. M. Parnaby; G. O'Dwyer; H. A. Monsey; M. S. Shafi

    1996-01-01

    textabstractThe point prevalence and incidence of Staphylococcus aureus (methicillin-sensitive and -resistant) carriage by inpatients on acute elderly care wards was estimated. The relationship to body site and to previous admissions to hospital or other institutions was determined. Fifty-five patie

  9. Increasing resistance of Staphylococcus aureus to ciprofloxacin.

    OpenAIRE

    Daum, T E; Schaberg, D R; Terpenning, M S; Sottile, W S; Kauffman, C A

    1990-01-01

    We demonstrated the marked emergence of resistance to ciprofloxacin among Staphylococcus arueus strains isolated at the Ann Arbor Veterans Administration Medical Center. All S. aureus isolates tested from 1984 to 1985 were susceptible, whereas 55.1% of methicillin-resistant and 2.5% of methicillin-susceptible strains from 1989 had high-level resistance to ciprofloxacin.

  10. Staphylococcus aureus spa type t437

    DEFF Research Database (Denmark)

    Glasner, C; Pluister, G; Westh, H;

    2015-01-01

    large epidemiological studies in Europe. Nevertheless, the overall numbers identified in some Northern European reference laboratories have increased during the past decade. To determine whether the S. aureus t437 clone is present in other European countries, and to assess its genetic diversity across...

  11. A series of template plasmids for Escherichia coli genome engineering.

    Science.gov (United States)

    Deb, Shalini S; Reshamwala, Shamlan M S; Lali, Arvind M

    2016-06-01

    Metabolic engineering strategies often employ multi-copy episomal vectors to overexpress genes. However, chromosome-based overexpression is preferred as it avoids the use of selective pressure and reduces metabolic burden on the cell. We have constructed a series of template plasmids for λ Red-mediated Escherichia coli genome engineering. The template plasmids allow construction of genome integrating cassettes that can be used to integrate single copies of DNA sequences at predetermined sites or replace promoter regions. The constructed cassettes provide flexibility in terms of expression levels achieved and antibiotics used for selection, as well as allowing construction of marker-free strains. The modular design of the template plasmids allows replacement of genetic parts to construct new templates. Gene integration and promoter replacement using the template plasmids are illustrated. PMID:27071533

  12. Construction and Identification of Plasmid pTA-TUB2

    Institute of Scientific and Technical Information of China (English)

    2003-01-01

    An about 1.40 Kb target gene fragment was yielded by PCR amplification with the plasmid pRB 129,which was identified by restriction enzyme digestion that the PCR product was TU B2 gene.The gene was digested by the restriction enzyme and was linked with pTA plasmid to construct pTA-TU B2 plasmid.The plasmid was transformed into Chaetomium spp.by PEG method and the transformation rate was 27/(2×105) and it is nine times higher than that of pRB 129.The transformants can grow on the PDA containing 1 000 μg*mL-1 carbendazim,which is 1 000 times higher than the original Chaetomium spp.The resistance was stable after 10 times transfer on non-selective medium.

  13. Characterization of Nocardia Plasmid pXT107

    Institute of Scientific and Technical Information of China (English)

    Hai-Yang XIA; Yong-Qiang TIAN; Ran ZHANG; Kai-Chun LIN; Zhong-Jun QIN

    2006-01-01

    Nocardia, Rhodococcus and Streptornyces, all members of the actinomycetes family, are Gram-positive eubacteria with high G+C content and able to form mycelium. We report here a newly identified plasmid pXT107 of Nocardia sp. 107, one of the smallest circular plasmids found in Nocardia.The complete nucleotide sequence of pXT107 consisted of 4335 bp with 65% G+C content, and encoded one replication extragenic palindromic (Rep) and six hypothetical proteins. The Rep, double-strand origin and single-strand origin of pXT107 resembled those of typical rolling-circle-replication plasmids, such as pNI100 of Nocardia, pRE8424 of Rhodococcus and plJ101 of Streptomyces. The Escherichia coli-Nocardia shuttle plasmid pHAQ22, containing thc rep gene of pXT107, is able to propagate in Nocardia but not in Streptomyces.

  14. Plasmid DNA labelled with 14C or 3H

    International Nuclear Information System (INIS)

    Plasmid DNA labelled with 14C or 3H in thymine was isolated from the thymine-dependent strain of Escherichia coli 15 SPT bacteria. The specific activity of the plasmid DNA preparations lay in the range from 0.5 to 20 MBq/mg, their relative molecular weight was 1.7 x 106 dalton. Molecular weight, preparation purity, and the degree of damage of the plasmid DNA molecules were examined by UV absorption spectroscopy, by gel electrophoresis, and by electron micrography. The quality of the [thymine-2-14C] plasmid DNA was verified in a diagnostic test for the determination of the anti-dsDNA bonding activity in human serum. (author). 1 tab., 5 figs., 30 refs

  15. Characterization of the Double-Partitioning Modules of R27: Correlating Plasmid Stability with Plasmid Localization

    OpenAIRE

    Trevor D Lawley; Taylor, Diane E.

    2003-01-01

    Plasmid R27 contains two independent partitioning modules, designated Par1 and Par2, within transfer region 2. Par1 is member of the type I partitioning family (Walker-type ATPase), and Par2 is a member of the type II partitioning family (actin-type ATPase). Stability tests of cloned Par1 and Par2 and insertional disruptions of Par1 and Par2 within R27 demonstrated that Par1 is the major stability determinant whereas Par2 is the minor stability determinant. Creation of double-partitioning mut...

  16. The Salmonella typhimurium virulence plasmid encodes a positive regulator of a plasmid-encoded virulence gene.

    OpenAIRE

    Caldwell, A L; Gulig, P A

    1991-01-01

    The 90-kb virulence plasmid of Salmonella typhimurium is necessary for invasion beyond the Peyer's patches to the mesenteric lymph nodes and spleens of orally inoculated mice. Two Tn5 insertions located on the left side of a previously identified 14-kb virulence region (P. A. Gulig and R. Curtiss III, Infect. Immun. 58:3262-3271, 1988) and mapping 272 bp from each other exhibited opposite effects on splenic infection of mice after oral inoculation. spvR23::Tn5 decreased splenic infection by 1...

  17. Detection of plasmid deoxyribonucleic acid in an isolate of Lactobacillus acidophilus.

    OpenAIRE

    Klaenhammer, T R; Sutherland, S M

    1980-01-01

    Eight strains of Lactobacillus acidophilus were examined for the presence of plasmid deoxyribonucleic acid, and one, a pig intestinal isolate, showed the presence of a 13.7- and a 6.3-megadalton plasmid. This is the first reported evidence for plasmid deoxyribonucleic acid in Lactobacillus acidophilus. The functions of these plasmids are presently unknown.

  18. Production of Fibronectin Binding Protein A at the surface of Lactococcus lactis increases plasmid transfer in vitro and in vivo.

    Directory of Open Access Journals (Sweden)

    Daniela Pontes

    Full Text Available Lactococci are noninvasive lactic acid bacteria frequently used as protein delivery vectors and, more recently, as DNA delivery vehicles. We previously showed that Lactococcus lactis (LL expressing the Fibronectin-Binding Protein A of Staphylococcus aureus (LL-FnBPA+ showed higher internalization rates in vitro in Caco-2 cells than the native (wt lactococci and were able to deliver a eukaryotic Green Fluorescent Protein (GFP expression plasmid in 1% of human Caco-2 cells. Here, using the bovine beta-lactoglobulin (BLG, one of the major cow's milk allergen, and GFP we characterized the potential of LL-FnBPA+ as an in vivo DNA vaccine delivery vehicle. We first showed that the invasive strain LL-FnBPA+ carrying the plasmid pValac:BLG (LL-FnBPA+ BLG was more invasive than LL-BLG and showed the same invasivity as LL-FnBPA+. Then we demonstrated that the Caco-2 cells, co-incubated with LL-FnBPA+ BLG produced up to 30 times more BLG than the Caco-2 cells co-incubated with the non invasive LL-BLG. Using two different gene reporters, BLG and GFP, and two different methods of detection, EIA and fluorescence microscopy, we showed in vivo that: i in order to be effective, LL-FnBPA+ required a pre-coating with Fetal Calf Serum before oral administration; ii plasmid transfer occurred in enterocytes without regard to the strains used (invasive or not; iii the use of LL-FnBPA+ increased the number of mice producing BLG, but not the level of BLG produced. We thus confirmed the good potential of invasive recombinant lactic acid bacteria as DNA delivery vector in vivo.

  19. Pharmaceutical development of the plasmid DNA vaccine pDERMATT

    OpenAIRE

    Quaak, S.G.L.

    2009-01-01

    The discovery of tumor specific antigens and self tolerance mechanisms against these antigens led to the assumption that antigens circulating at sufficient concentration levels could break this self tolerance mechanism and evoke immunological antitumor effects. pDERMATT (plasmid DNA encoding recombinant MART-1 and tetanus toxin fragment-c) is a plasmid that encodes for MART-1, a melanoma associated antigen that is expressed in a large fraction of melanomas. In animal models administration of ...

  20. A novel chromatographic procedure for purification of bacterial plasmids.

    Science.gov (United States)

    Bywater, M; Bywater, R; Hellman, L

    1983-07-01

    A new, rapid procedure for purifying bacterial plasmids with high recovery is described. The sequence of operations consists essentially of treatment with alkali, ribonuclease, and proteinase K, followed by chisam extraction and gel filtration on Sephacryl S-1000, and finally a precipitation step using isopropanol at room temperature. The method gives rather good yields of plasmid DNA of high purity, and lends itself to scaling up. PMID:6312836

  1. Isolation and properties of plasmids from Deinococcus radiodurans Sark

    International Nuclear Information System (INIS)

    Radioresistant bacterium, Deinococcus radiodurans, can repair completely almost all of DNA damages including double strand breaks induced by gamma-rays up to about 5 kGy. In order to reveal the repair mechanism, it is necessary to develop a cloning vector available for the genetic analysis. We tried to isolate plasmids from D.radiodurans Sark strain. In the present paper the isolation and properties of plasmids were described. (author)

  2. Construction of a bioluminescence reporter plasmid for Francisella tularensis

    OpenAIRE

    Bina, Xiaowen R.; Miller, Mark A.; James E Bina

    2010-01-01

    A Francisella tularensis shuttle vector that constitutively expresses the Photorhabdus luminescens lux operon in type A and type B strains of F. tularensis was constructed. The bioluminescence reporter plasmid was introduced into the live vaccine strain of F. tularensis and used to follow F. tularensis growth in a murine intranasal challenge model in real time by bioluminescence imaging. The results show that the new bioluminescence reporter plasmid represents a useful tool for tularemia rese...

  3. Conjugative plasmid transfer from Enterococcus faecalis to Escherichia coli.

    OpenAIRE

    Trieu-Cuot, P; Carlier, C; Courvalin, P

    1988-01-01

    The possibility of transfer of genetic information by conjugation from gram-positive to gram-negative bacteria was investigated with a pBR322-pAM beta 1 chimeric plasmid, designated pAT191. This shuttle vector, which possesses the tra functions of the streptococcal plasmid pAM beta 1, was conjugatively transferred from Enterococcus faecalis to Escherichia coli with an average frequency of 5 x 10(-9) per donor colony formed after mating.

  4. Intragenic variation by site-specific recombination in the cryptic plasmid of Neisseria gonorrhoeae.

    OpenAIRE

    Hagblom, P; Korch, C; Jonsson, A B; Normark, S

    1986-01-01

    Cryptic plasmid DNA of Neisseria gonorrhoeae was found integrated into the gonococcal chromosome in both plasmid-bearing strains and plasmid-free strains. At several chromosomal locations only segments of the plasmid were found. However, in at least two strains an intact copy of the plasmid seemed to be present with the joints between the plasmid and the chromosomal DNA being located within the cppB gene of the cryptic plasmid. The cppB gene was shown to undergo a sequence-specific intragenic...

  5. Plasmid vector with temperature-controlled gene expression

    International Nuclear Information System (INIS)

    In plasmid pBR327, a fragment 169 b.p. long including promotor p3 of the bla gene has been deleted. The deletional derivative so obtained (pSP2) has been used to construct a recombinant plasmid bearing a fragment of phage λ DNA with the p/sub R/ promotor and the gene of the temperature-sensitive repressor cI. It has been shown that the plasmid vector so constructed (pCE119) with promotor cR performs repressor-cI-controlled transcription of the bla gene, as a result of which induction for an hour at 420C leads to an almost 100-fold increase in the amount of product of the bla gene as compared with that at 320C. The possibility of the use of plasmid cPE119 for the expression of other genes has been demonstrated for the case of the semisynthetic β-galactosidase gene of E. coli. In this case, on induction of the cells with recombinant plasmid pCEZ12 for 3 hours at 420C, a 300-fold increase in the amount of active β-galactosidase, as compared with that at 320C, was observed. It is important to point out that under these conditions (at 420C), at least 99% of the cells containing the plasmid retain the phenotype lacZ+, which indicates the stability of the proposed vector system

  6. Transfer of conjugative plasmids among bacteria under environmentally relevant conditions

    DEFF Research Database (Denmark)

    Musovic, Sanin

    det oprindelige bakteriesamfund der tager andel i plasmid overførsel blev udviklet. Dyrknings-minimal metode i kombination med reporter gen teknologi og moderne mikroskopi viste en meget høj forekomst af RP4:gfp plasmid overførsel til oprindelige jord bakterier af et bredt værtskab. Der blev også vist......Mobile genetiske elementer (f.eks. plasmider), der ofte bærer ekstra funktioner såsom antibiotikaresistens, eller kataboliske- og xenobiotiske nedbrydnings gener, antages at have en meget vigtigt evolutionær rolle for bakterier. I denne PhD afhandling undersøgte jeg størrelsen af plasmid overførsel...... under de miljørelevante substrat-begrænsede forhold, den del og diversitet af bakteriel samfund der er involveret i overførslen, og effekten af plasmid donor cellens fysiologiske status og de miljørelevante faktorer (selektive tryk) på plasmid spredning. En ny metode til at kvantificere den fraktion af...

  7. The characteristics of micrococcus (deinococcus) radiodurans sark plasmids

    International Nuclear Information System (INIS)

    The characterization of micrococcus (deinococcus) radiodurans sark plasmids. This bacterium has been classified as a new genus deinococcus radiodurans which is resistant to gamma-rays. It can repair itself completely almost all of DNA damages including double strand breaks induced by gamma-rays up to about 5 KGy. To reveal the repair mechanism, several investigations had been done to develop a cloning vector available for the genetic analysis. For this purpose D. radiodurans Sark are to be prepared as a vector by studying the characteristics of its plasmid. Plasmids were isolated by electrophoresis using 0.6% low-melting-temperature agarose in TAE and run for 5.5 hours, followed by the identification. An antibiotic marker was also carried out in this experiment to identify its location in the genetic materials of the cell, beside making a restriction map of the plasmid. Results have shown that D. radiodurans Sark has 4 plasmids (P1, P2, P3, and P4) and the refampicin resistant genes were not found in the plasmid. (authors). 14 refs; 4 figs

  8. Plasmid-free T7-based Escherichia coli expression systems.

    Science.gov (United States)

    Striedner, Gerald; Pfaffenzeller, Irene; Markus, Luchner; Nemecek, Sabine; Grabherr, Reingard; Bayer, Karl

    2010-03-01

    In order to release host cells from plasmid-mediated increases in metabolic load and high gene dosages, we developed a plasmid-free, T7-based E. coli expression system in which the target gene is site-specifically integrated into the genome of the host. With this system, plasmid-loss, a source of instability for conventional expression systems, was eliminated. At the same time, system leakiness, a challenging problem with recombinant systems, was minimized. The efficiency of the T7 RNA polymerase compensates for low gene dosage and provides high rates of recombinant gene expression without fatal consequences to host metabolism. Relative to conventional pET systems, this system permits improved process stability and increases the host cell's capacity for recombinant gene expression, resulting in higher product yields. The stability of the plasmid-free system was proven in chemostat cultivation for 40 generations in a non-induced and for 10 generations in a fully induced state. For this reason plasmid-free systems benefit the development of continuous production processes with E. coli. However, time and effort of the more complex cloning procedure have to be considered in relation to the advantages of plasmid-free systems in upstream-processing. PMID:19891007

  9. Relationship between Vancomycin-Resistant Staphylococcus aureus, Vancomycin-Intermediate S. aureus, High Vancomycin MIC, and Outcome in Serious S. aureus Infections

    OpenAIRE

    Holmes, Natasha E.; Johnson, Paul D. R.; Howden, Benjamin P.

    2012-01-01

    Vancomycin has been used successfully for over 50 years for the treatment of Staphylococcus aureus infections, particularly those involving methicillin-resistant S. aureus. It has proven remarkably reliable, but its efficacy is now being questioned with the emergence of strains of S. aureus that display heteroresistance, intermediate resistance, and, occasionally, complete vancomycin resistance. More recently, an association has been established between poor outcome and infections with strain...

  10. Partition-associated incompatibility caused by random assortment of pure plasmid clusters

    DEFF Research Database (Denmark)

    Ebersbach, Gitte; Sherratt, David J; Gerdes, Kenn;

    2005-01-01

    Summary Bacterial plasmids and chromosomes encode centromere-like partition loci that actively segregate DNA before cell division. The molecular mechanism behind DNA segregation in bacteria is largely unknown. Here we analyse the mechanism of partition-associated incompatibility for plasmid pB171......, a phenotype associated with all known plasmid-encoded centromere loci. An R1 plasmid carrying par2 from plasmid pB171 was destabilized by the presence of an F plasmid carrying parC1, parC2 or the entire par2 locus of pB171. Strikingly, cytological double-labelling experiments revealed no evidence of long......-lived pairing of plasmids. Instead, pure R1 and F foci were positioned along the length of the cell, and in a random order. Thus, our results raise the possibility that partition-mediated plasmid incompatibility is not caused by pairing of heterologous plasmids but instead by random positioning of pure plasmid...

  11. Tyrosine Partners Coordinate DNA Nicking by the Salmonella typhimurium Plasmid pCU1 Relaxase Enzyme

    OpenAIRE

    Nash, Rebekah P.; Niblock, Franklin C.; Redinbo, Matthew R.

    2011-01-01

    Conjugative plasmid transfer results in the spread of antibiotic resistance genes and virulence factors between bacterial cells. Plasmid transfer is dependent upon the DNA nicking activity of a plasmid-encoded relaxase enzyme. Tyrosine residues within the relaxase cleave the DNA plasmid nic site in a highly sequence-specific manner. The conjugative resistance plasmid pCU1 encodes a relaxase with four tyrosine residues surrounding its active site (Y18,19,26,27). We use activity assays to demon...

  12. Linear Plasmid SLP2 Is Maintained by Partitioning, Intrahyphal Spread, and Conjugal Transfer in Streptomyces▿

    OpenAIRE

    Hsu, Chin-Chen; Chen, Carton W.

    2009-01-01

    Low-copy-number plasmids generally encode a partitioning system to ensure proper segregation after replication. Little is known about partitioning of linear plasmids in Streptomyces. SLP2 is a 50-kb low-copy-number linear plasmid in Streptomyces lividans, which contains a typical parAB partitioning operon. In S. lividans and Streptomyces coelicolor, a parAB deletion resulted in moderate plasmid loss and growth retardation of colonies. The latter was caused by conjugal transfer from plasmid-co...

  13. Transfer of chimeric plasmids among Salmonella typhimurium strains by P22 transduction.

    OpenAIRE

    Orbach, M J; Jackson, E N

    1982-01-01

    Salmonella typhimurium bacteriophage P22 transduced plasmids having P22 sequences inserted in the vector pBR322 with high frequency. Analysis of the structure of the transducing particle DNA and the transduced plasmids indicates that this plasmid transduction involves two homologous recombination events. In the donor cell, a single recombination between the phage and the homologous sequences on the plasmid inserted the plasmid into the phage chromosome, which was then packaged by headfuls int...

  14. Exposing Plasmids as the Achilles’ Heel of Drug-Resistant Bacteria

    OpenAIRE

    Williams, Julia J.; Hergenrother, Paul J.

    2008-01-01

    Many multi-drug resistant bacterial pathogens harbor large plasmids that encode proteins conferring resistance to antibiotics. While the acquisition of these plasmids often enables bacteria to survive in the presence of antibiotics, it is possible that plasmids also represent a vulnerability that can be exploited in tailored antibacterial therapy. This review highlights three recently described strategies designed to specifically combat bacteria harboring such plasmids: Inhibition of plasmid ...

  15. Sequence-nonspecific replication of transfected plasmid DNA in poxvirus-infected cells.

    OpenAIRE

    DeLange, A. M.; McFadden, G

    1986-01-01

    A system in which transfected plasmid DNA replicates in the cytoplasm of poxvirus-infected cells is described. A variety of recombinant plasmids was introduced into poxvirus-infected cells by transfection, and replication of input plasmid DNA was monitored by (i) digestion with restriction enzymes that discriminate between input methylated plasmid DNA and unmethylated DNA produced by replication in mammalian cells; (ii) amplification of intracellular plasmid DNA; and (iii) density shift analy...

  16. Characterization of plasmid burden and copy number in Saccharomyces cerevisiae for optimization of metabolic engineering applications

    OpenAIRE

    Karim, Ashty S.; Curran, Kathleen A.; Alper, Hal S.

    2012-01-01

    Many metabolic engineering and genetic engineering applications in yeast rely on the use of plasmids. Despite their pervasive use and the diverse collections available, there is a fundamental lack of understanding of how commonly used DNA plasmids affect the cell’s ability to grow and how the choice of plasmid components can influence plasmid load and burden. In this study, we characterized the major attributes of the 2μ and centromeric plasmids typically used in yeast by examining the impact...

  17. Hofmeister series salts enhance purification of plasmid DNA by non-ionic detergents

    OpenAIRE

    Lezin, George; Kuehn, Michael R.; Brunelli, Luca

    2011-01-01

    Ion-exchange chromatography is the standard technique used for plasmid DNA purification, an essential molecular biology procedure. Non-ionic detergents (NIDs) have been used for plasmid DNA purification, but it is unclear whether Hofmeister series salts (HSS) change the solubility and phase separation properties of specific NIDs, enhancing plasmid DNA purification. After scaling-up NID-mediated plasmid DNA isolation, we established that NIDs in HSS solutions minimize plasmid DNA contamination...

  18. Comparison of Ti plasmids from three different biotypes of Agrobacterium tumefaciens isolated from grapevines.

    OpenAIRE

    Knauf, V C; Panagopoulos, C G; Nester, E. W.

    1983-01-01

    Twenty-six plasmids from grapevine isolates of Agrobacterium tumefaciens were analyzed by SmaI fingerprinting and by hybridization of nick-translated DNA to DNA of another plasmid. These experiments established that octopine Ti plasmids are not highly conserved, although octopine Ti plasmids from biotype 1 A. tumefaciens strains appeared to be very similar. Octopine Ti plasmids from biotype 3 strains are more variable in terms of host range and SmaI fingerprints, but share extensive DNA homol...

  19. Proteome data from a host-pathogen interaction study with Staphylococcus aureus and human lung epithelial cells

    Directory of Open Access Journals (Sweden)

    Kristin Surmann

    2016-06-01

    Full Text Available To simultaneously obtain proteome data of host and pathogen from an internalization experiment, human alveolar epithelial A549 cells were infected with Staphylococcus aureus HG001 which carried a plasmid (pMV158GFP encoding a continuously expressed green fluorescent protein (GFP. Samples were taken hourly between 1.5 h and 6.5 h post infection. By fluorescence activated cell sorting GFP-expressing bacteria could be enriched from host cell debris, but also infected host cells could be separated from those which did not carry bacteria after contact (exposed. Additionally, proteome data of A549 cells which were not exposed to S. aureus but underwent the same sample processing steps are provided as a control. Time-resolved changes in bacterial protein abundance were quantified in a label-free approach. Proteome adaptations of host cells were monitored by comparative analysis to a stable isotope labeled cell culture (SILAC standard. Proteins were extracted from the cells, digested proteolytically, measured by nanoLC–MS/MS, and subsequently identified by database search and then quantified. The data presented here are related to a previously published research article describing the interplay of S. aureus HG001 and human epithelial cells (Surmann et al., 2015 [1]. They have been deposited to the ProteomeXchange platform with the identifiers PRIDE: http://www.ebi.ac.uk/pride/archive/projects/PXD002384 for the S. aureus HG001 proteome dataset and PRIDE: http://www.ebi.ac.uk/pride/archive/projects/PXD002388 for the A549 proteome dataset.

  20. Staphylococcus aureus methicillin-resistance mechanisms

    Directory of Open Access Journals (Sweden)

    Petrović-Jeremić Ljiljana

    2008-01-01

    Full Text Available Background/Aim. In many hospitals in the world and in our country, the spread of methicillin-resistant Staphylococcus aureus (MRSA is so wide that nowdays vancomycin is recommended for empiric treatment of staphylococcal life threatening infections (sepsis, pneumonia instead of beta-lactam antibiotics. The aim of this study was to determine the production of beta-lactamases in hospital and community isolates of staphyloococus aureus, i. e. hospital associated MRSA (HA-MRSA and community associated MRSA (CA-MRSA, the presence of homogeneous and heterogeneous type of methicillin resistance, and border-line resistance in Staphylococcus aureus (BORSA. The aim of this study was also to determine if there was a statistically significant difference between mechanisms of resistance in HA-MRSA and CA-MRSA. Methods. A total 216 clinical Staphylococcus aureus isolates from the General Hospital in the town of Cuprija and 186 ambulance Staphylococcus aureus isolates from the community were examined for the presence of methicillin-resistance using disk-diffusion test with penicillin disk (10 ij, oxacillin disk (1 μg and cefoxitin disk (30 μg. Betalactamases production was detected by nitrocefin disk and betalactamase tablets. Determination of oxacillin minimum inhibitory concentracion (MIC was done by agar-dilution method. Results. The prevalence of HA-MRSA was 57.4%, and CA-MRSA was 17.7% (p < 0.05. There was a higher rate of heterogeneous type of resistance among clinical MRSA isolates (11.1% compared with ambulance ones (3.8% (p < 0.05. The rates of beta-lactamases production were similar among hospital associated isolates (97.5%, as well as in the community associated isolates (95.5% (p > 0.05. There were 4.6 % of BORSA hospital isolates and 3.3 % of BORSA ambulance isolates (p > 0.05. Conclusion. The frequency of MRSA isolates in hospital was significantly higher than in community, as well as the heterogeneous type of resistance. The frequency of BORSA

  1. Conjugative plasmids isolated from bacteria in marine environments show various degrees of homology to each other and are not closely related to well-characterized plasmids.

    OpenAIRE

    Dahlberg, C; Linberg, C; Torsvik, V L; Hermansson, M

    1997-01-01

    Mercury resistance plasmids were exogenously isolated, i.e., recovered after transfer to a model recipient bacterium, from marine air-water interface, bulk water, and biofilm communities during incubation in artificial seawater without added nutrients. Ninety-five plasmids from different environments were classified by restriction endonuclease digestion, and 12 different structural plasmid groups were revealed. The plasmid types isolated from different habitats and from different sampling occ...

  2. Replication of a low-copy-number plasmid by a plasmid DNA-membrane complex extracted from minicells of Escherichia coli.

    OpenAIRE

    Firshein, W; Strumph, P; Benjamin, P; Burnstein, K; Kornacki, J

    1982-01-01

    A DNA-membrane complex was extracted from minicells of an Escherichia coli mutant harboring a "miniplasmid" derivative (11.2 kilobases) of the low-copynumber plasmid RK2 (56 kilobases). The complex contained various species of supercoiled and intermediate forms of plasmid DNA, of which approximately 20% was bound firmly to the membrane after centrifugation in a CsCl density gradient. The plasmid DNA-membrane complex synthesized new plasmid DNA without the addition of exogenous template, enzym...

  3. Plasmids and rickettsial evolution: insight from Rickettsia felis.

    Directory of Open Access Journals (Sweden)

    Joseph J Gillespie

    Full Text Available BACKGROUND: The genome sequence of Rickettsia felis revealed a number of rickettsial genetic anomalies that likely contribute not only to a large genome size relative to other rickettsiae, but also to phenotypic oddities that have confounded the categorization of R. felis as either typhus group (TG or spotted fever group (SFG rickettsiae. Most intriguing was the first report from rickettsiae of a conjugative plasmid (pRF that contains 68 putative open reading frames, several of which are predicted to encode proteins with high similarity to conjugative machinery in other plasmid-containing bacteria. METHODOLOGY/PRINCIPAL FINDINGS: Using phylogeny estimation, we determined the mode of inheritance of pRF genes relative to conserved rickettsial chromosomal genes. Phylogenies of chromosomal genes were in agreement with other published rickettsial trees. However, phylogenies including pRF genes yielded different topologies and suggest a close relationship between pRF and ancestral group (AG rickettsiae, including the recently completed genome of R. bellii str. RML369-C. This relatedness is further supported by the distribution of pRF genes across other rickettsiae, as 10 pRF genes (or inactive derivatives also occur in AG (but not SFG rickettsiae, with five of these genes characteristic of typical plasmids. Detailed characterization of pRF genes resulted in two novel findings: the identification of oriV and replication termination regions, and the likelihood that a second proposed plasmid, pRFdelta, is an artifact of the original genome assembly. CONCLUSION/SIGNIFICANCE: Altogether, we propose a new rickettsial classification scheme with the addition of a fourth lineage, transitional group (TRG rickettsiae, that is unique from TG and SFG rickettsiae and harbors genes from possible exchanges with AG rickettsiae via conjugation. We offer insight into the evolution of a plastic plasmid system in rickettsiae, including the role plasmids may have played in

  4. Methicillin-resistant Staphylococcus aureus: the superbug.

    Science.gov (United States)

    Ippolito, Giuseppe; Leone, Sebastiano; Lauria, Francesco N; Nicastri, Emanuele; Wenzel, Richard P

    2010-10-01

    Over the last decade, methicillin-resistant Staphylococcus aureus (MRSA) strains have emerged as serious pathogens in the nosocomial and community setting. Hospitalization costs associated with MRSA infections are substantially greater than those associated with methicillin-sensitive S. aureus (MSSA) infections, and MRSA has wider economic effects that involve indirect costs to the patient and to society. In addition, there is some evidence suggesting that MRSA infections increase morbidity and the risk of mortality. Glycopeptides are the backbone antibiotics for the treatment of MRSA infections. However, several recent reports have highlighted the limitations of vancomycin, and its role in the management of serious infections is now being reconsidered. Several new antimicrobials demonstrate in vitro activity against MRSA and other Gram-positive bacteria. Data from large surveys indicate that linezolid, daptomycin, and tigecycline are almost universally active against MRSA. This review will briefly discuss the epidemiology, costs, outcome, and therapeutic options for the management of MRSA infections. PMID:20851011

  5. Curcumin Reverse Methicillin Resistance in Staphylococcus aureus

    OpenAIRE

    Su-Hyun Mun; Sung-Bae Kim; Ryong Kong; Jang-Gi Choi; Youn-Chul Kim; Dong-Won Shin; Ok-Hwa Kang; Dong-Yeul Kwon

    2014-01-01

    Curcumin, a natural polyphenolic flavonoid extracted from the rhizome of Curcuma longa L., was shown to possess superior potency to resensitize methicillin-resistant Staphylococcus aureus (MRSA) to antibiotics. Previous studies have shown the synergistic activity of curcumin with β-lactam and quinolone antibiotics. Further, to understand the anti-MRSA mechanism of curcumin, we investigated the potentiated effect of curcumin by its interaction in diverse conditions. The mechanism of anti-MRSA ...

  6. Xanthgranulomatous pyelonephritis associated with Staphylococcus aureus.

    Science.gov (United States)

    Al-Hwiesh, Abdulla K

    2007-11-01

    A 44-year old man with xanthogranulomatous pyelonephritis presented with abdominal distention, left lumber pain, fever, loss of appetite, and loss of weight. He had been known to have diabetes mellitus type II for 20 years, and he was diagnosed to have a left renal stone three months prior to this presentation. The patient's urine and the left psous abscess grew staphylococcus aureus. PMID:17951953

  7. Xanthgranulomatous pyelonephritis associated with staphylococcus aureus

    International Nuclear Information System (INIS)

    A 44-year-old man with xanthgranulomatous pyelonephritis presented with abdominal distention, left lumber pain, fever, loss of appetite and loss of weight. He had been known to have diabetes mellitus type II for 20 years and he was diagnosed to have a left renal stone three months prior to this presentation. The patient's urine and the left psous abscess grew staphylococcus aureus. (author)

  8. Aspartate inhibits Staphylococcus aureus biofilm formation.

    Science.gov (United States)

    Yang, Hang; Wang, Mengyue; Yu, Junping; Wei, Hongping

    2015-04-01

    Biofilm formation renders Staphylococcus aureus highly resistant to conventional antibiotics and host defenses. Four D-amino acids (D-Leu, D-Met, D-Trp and D-Tyr) have been reported to be able to inhibit biofilm formation and disassemble established S. aureus biofilms. We report here for the first time that both D- and L-isoforms of aspartate (Asp) inhibited S. aureus biofilm formation on tissue culture plates. Similar biofilm inhibition effects were also observed against other staphylococcal strains, including S. saprophyticus, S. equorum, S. chromogenes and S. haemolyticus. It was found that Asp at high concentrations (>10 mM) inhibited the growth of planktonic N315 cells, but at subinhibitory concentrations decreased the cellular metabolic activity without influencing cell growth. The decreased cellular metabolic activity might be the reason for the production of less protein and DNA in the matrix of the biofilms formed in the presence of Asp. However, varied inhibition efficacies of Asp were observed for biofilms formed by clinical staphylococcal isolates. There might be mechanisms other than decreasing the metabolic activity, e.g. the biofilm phenotypes, affecting biofilm formation in the presence of Asp. PMID:25687923

  9. Whole-Genome Sequence of Staphylococcus aureus Strain LCT-SA112

    OpenAIRE

    Wang, Junfeng; Liu, Yanhong; Wan, Daiwei; Fang, Xiangqun; Li, Tianzhi; Guo, Yinghua; Chang, De; Su, Longxiang; Wang, Yajuan; Zhao, Jiao; Liu, Changting

    2012-01-01

    Staphylococcus aureus is a facultative anaerobic Gram-positive coccal bacterium. S. aureus is the most common species of Staphylococcus to cause staphylococcal infections, which are very common in clinical medicine. Here we report the genome sequence of S. aureus strain LCT-SA112, which was isolated from S. aureus subsp. aureus CGMCC 1.230.

  10. Plasma-activated air mediates plasmid DNA delivery in vivo.

    Science.gov (United States)

    Edelblute, Chelsea M; Heller, Loree C; Malik, Muhammad A; Bulysheva, Anna; Heller, Richard

    2016-01-01

    Plasma-activated air (PAA) provides a noncontact DNA transfer platform. In the current study, PAA was used for the delivery of plasmid DNA in a 3D human skin model, as well as in vivo. Delivery of plasmid DNA encoding luciferase to recellularized dermal constructs was enhanced, resulting in a fourfold increase in luciferase expression over 120 hours compared to injection only (P plasmid DNA encoding green fluorescent protein (GFP) was confirmed in the epidermal layers of the construct. In vivo experiments were performed in BALB/c mice, with skin as the delivery target. PAA exposure significantly enhanced luciferase expression levels 460-fold in exposed sites compared to levels obtained from the injection of plasmid DNA alone (P plasmid DNA encoding GFP to mouse skin was confirmed by immunostaining, where a 3-minute exposure at a 10 mm distance displayed delivery distribution deep within the dermal layers compared to an exposure at 3 mm where GFP expression was localized within the epidermis. Our findings suggest PAA-mediated delivery warrants further exploration as an alternative approach for DNA transfer for skin targets. PMID:27110584

  11. Replicase-based plasmid DNA shows anti-tumor activity

    Directory of Open Access Journals (Sweden)

    Weiss Richard

    2011-03-01

    Full Text Available Abstract Background Double stranded RNA (dsRNA has multiple anti-tumor mechanisms. Over the past several decades, there have been numerous attempts to utilize synthetic dsRNA to control tumor growth in animal models and clinical trials. Recently, it became clear that intracellular dsRNA is more effective than extracellular dsRNA on promoting apoptosis and orchestrating adaptive immune responses. To overcome the difficulty in delivering a large dose of synthetic dsRNA into tumors, we propose to deliver a RNA replicase-based plasmid DNA, hypothesizing that the dsRNA generated by the replicase-based plasmid in tumor cells will inhibit tumor growth. Methods The anti-tumor activity of a plasmid (pSIN-β that encodes the sindbis RNA replicase genes (nsp1-4 was evaluated in mice with model tumors (TC-1 lung cancer cells or B16 melanoma cells and compared to a traditional pCMV-β plasmid. Results In cell culture, transfection of tumor cells with pSIN-β generated dsRNA. In mice with model tumors, pSIN-β more effectively delayed tumor growth than pCMV-β, and in some cases, eradicated the tumors. Conclusion RNA replicase-based plasmid may be exploited to generate intracellular dsRNA to control tumor growth.

  12. Replicase-based plasmid DNA shows anti-tumor activity

    International Nuclear Information System (INIS)

    Double stranded RNA (dsRNA) has multiple anti-tumor mechanisms. Over the past several decades, there have been numerous attempts to utilize synthetic dsRNA to control tumor growth in animal models and clinical trials. Recently, it became clear that intracellular dsRNA is more effective than extracellular dsRNA on promoting apoptosis and orchestrating adaptive immune responses. To overcome the difficulty in delivering a large dose of synthetic dsRNA into tumors, we propose to deliver a RNA replicase-based plasmid DNA, hypothesizing that the dsRNA generated by the replicase-based plasmid in tumor cells will inhibit tumor growth. The anti-tumor activity of a plasmid (pSIN-β) that encodes the sindbis RNA replicase genes (nsp1-4) was evaluated in mice with model tumors (TC-1 lung cancer cells or B16 melanoma cells) and compared to a traditional pCMV-β plasmid. In cell culture, transfection of tumor cells with pSIN-β generated dsRNA. In mice with model tumors, pSIN-β more effectively delayed tumor growth than pCMV-β, and in some cases, eradicated the tumors. RNA replicase-based plasmid may be exploited to generate intracellular dsRNA to control tumor growth

  13. Functional amyloids as inhibitors of plasmid DNA replication.

    Science.gov (United States)

    Molina-García, Laura; Gasset-Rosa, Fátima; Moreno-Del Álamo, María; Fernández-Tresguerres, M Elena; Moreno-Díaz de la Espina, Susana; Lurz, Rudi; Giraldo, Rafael

    2016-01-01

    DNA replication is tightly regulated to constrain the genetic material within strict spatiotemporal boundaries and copy numbers. Bacterial plasmids are autonomously replicating DNA molecules of much clinical, environmental and biotechnological interest. A mechanism used by plasmids to prevent over-replication is 'handcuffing', i.e. inactivating the replication origins in two DNA molecules by holding them together through a bridge built by a plasmid-encoded initiator protein (Rep). Besides being involved in handcuffing, the WH1 domain in the RepA protein assembles as amyloid fibres upon binding to DNA in vitro. The amyloid state in proteins is linked to specific human diseases, but determines selectable and epigenetically transmissible phenotypes in microorganisms. Here we have explored the connection between handcuffing and amyloidogenesis of full-length RepA. Using a monoclonal antibody specific for an amyloidogenic conformation of RepA-WH1, we have found that the handcuffed RepA assemblies, either reconstructed in vitro or in plasmids clustering at the bacterial nucleoid, are amyloidogenic. The replication-inhibitory RepA handcuff assembly is, to our knowledge, the first protein amyloid directly dealing with DNA. Built on an amyloid scaffold, bacterial plasmid handcuffs can bring a novel molecular solution to the universal problem of keeping control on DNA replication initiation. PMID:27147472

  14. Dcm methylation is detrimental to plasmid transformation in Clostridium thermocellum

    Energy Technology Data Exchange (ETDEWEB)

    Guss, Adam M [ORNL; Olson, Daniel G. [Thayer School of Engineering at Dartmouth; Caiazza, Nicky [Mascoma Corporation; Lynd, Lee R [Thayer School of Engineering at Dartmouth

    2012-01-01

    BACKGROUND: Industrial production of biofuels and other products by cellulolytic microorganisms is of interest but hindered by the nascent state of genetic tools. Although a genetic system for Clostridium thermocellum DSM1313 has recently been developed, available methods achieve relatively low efficiency and similar plasmids can transform C. thermocellum at dramatically different efficiencies. RESULTS: We report an increase in transformation efficiency of C. thermocellum for a variety of plasmids by using DNA that has been methylated by Escherichia coli Dam but not Dcm methylases. When isolated from a dam+ dcm+ E. coli strain, pAMG206 transforms C. thermocellum 100-fold better than the similar plasmid pAMG205, which contains an additional Dcm methylation site in the pyrF gene. Upon removal of Dcm methylation, transformation with pAMG206 showed a four- to seven-fold increase in efficiency; however, transformation efficiency of pAMG205 increased 500-fold. Removal of the Dcm methylation site from the pAM205 pyrF gene via silent mutation resulted in increased transformation efficiencies equivalent to that of pAMG206. Upon proper methylation, transformation efficiency of plasmids bearing the pMK3 and pB6A origins of replication increased ca. three orders of magnitude. CONCLUSION: E. coli Dcm methylation decreases transformation efficiency in C. thermocellum DSM1313. The use of properly methylated plasmid DNA should facilitate genetic manipulation of this industrially relevant bacterium.

  15. [Raw milk-associated Staphylococcus aureus intoxication in children].

    Science.gov (United States)

    Giezendanner, N; Meyer, B; Gort, M; Müller, P; Zweifel, C

    2009-07-01

    Four hours after the consumption of raw goat milk, three Swiss children came down with emesis and diarrhea in July 2008. First investigations showed that the milk originated from a goat suffering from clinical mastitis (Staphylococcus aureus). In the milk sample from the untreated left udder, Staphylococcus aureus counts reached 5.0 x 10(7) CFU ml(-1). By PCR, the gene for the staphylococcal enterotoxin D was found in isolated strains. The consumption of raw milk is rarely associated with Staphylococcus aureus intoxications. Due to the flora naturally present in raw milk, Staphylococcus aureus normally cannot multiply sufficiently. However, in the present case, high Staphylococcus aureus counts were already present in the milk due to the mastitis of the goat. This amount sufficed to cause a Staphylococcus aureus intoxication in the children. PMID:19565455

  16. Mitochondrial pAL2-1 plasmid homologs are senescence factors in Podospora anserina independent of intrinsic senescence

    NARCIS (Netherlands)

    van Diepeningen, Anne D; Debets, Alfons J M; Slakhorst, S Marijke; Hoekstra, Rolf F

    2008-01-01

    Since the first description of a linear mitochondrial plasmid in Podospora anserina, pAL2-1, and homologous plasmids have gone from being considered beneficial longevity plasmids, via neutral genetic elements, toward mutator plasmids causing senescence. The plasmid has an invertron structure, with t

  17. Antibody responses in patients with invasive Staphylococcus aureus infections

    OpenAIRE

    Jacobsson, G; Colque-Navarro, P.; Gustafsson, E.; Andersson, R.; Möllby, R

    2010-01-01

    Abstract Correlation between antibody response and clinical outcome in Staphylococcus aureus bacteremia has yielded conflicting results. Immunization schedules have failed in clinical trials. Is the humoral response toward S. aureus of protective nature? A prospective study was performed in patients with invasive S. aureus (ISA) infections during the period 2003?2005. The antibody levels were determined at the beginning and at the end of treatment and one month later (n?=?96, n?=?7...

  18. Vitamin D sufficiency and Staphylococcus aureus infection in children.

    Science.gov (United States)

    Wang, Jeffrey W; Hogan, Patrick G; Hunstad, David A; Fritz, Stephanie A

    2015-05-01

    Vitamin D promotes epithelial immunity by upregulating antimicrobial peptides, including LL-37, which have bactericidal activity against Staphylococcus aureus. We found that children with vitamin D deficiency or insufficiency [25-hydroxyvitamin D recurrent, rather than primary, S. aureus skin or soft tissue infection. Vitamin D sufficiency may be one of a myriad of host and environmental factors that can be directly impacted to reduce the frequency of S. aureus skin and soft tissue infection. PMID:25860535

  19. Mapping the Distribution of Invasive Staphylococcus aureus across Europe

    OpenAIRE

    Grundmann, Hajo; Aanensen, David M.; van den Wijngaard, Cees C.; Brian G Spratt; Harmsen, Dag; Friedrich, Alexander W.; ,

    2010-01-01

    Editors' Summary Background The bacterium Staphylococcus aureus lives on the skin and in the nose of about a third of healthy people. Although S. aureus usually coexists peacefully with its human carriers, it is also an important disease-causing organism or pathogen. If it enters the body through a cut or during a surgical procedure, S. aureus can cause minor infections such as pimples and boils or more serious, life-threatening infections such as blood poisoning and pneumonia. Minor S. aureu...

  20. Infection control of Staphylococcus aureus : spa typing to elucidate transmission

    OpenAIRE

    Mernelius, Sara

    2015-01-01

    Staphylococcus aureus is a commensal of the human flora, primarily colonizing the anterior nares and throat, but it may also cause infections ranging from mild skin and soft tissue infections to severe diseases such as endocarditis and septicemia. S. aureus is also a major nosocomial problem increasing with the worldwide dissemination of methicillin-resistant S. aureus (MRSA). The main vector for bacterial cross-transmission in healthcare settings is the hands of healthcare workers (HCWs). No...

  1. Staphylococcus aureus atsparumas antibiotikams ir fagotipų paplitimas

    OpenAIRE

    Kareivienė, Violeta; Pavilonis, Alvydas; Sinkutė, Gintarė; Liegiūtė, Sigutė; Gailienė, Greta

    2006-01-01

    Objective. The aim of this study was to identify the phage groups of Staphylococcus aureus strains, their prevalence, and resistance of different phage groups to antibiotics. Materials and methods. A total of 294 Staphylococcus aureus strains in Kaunas hospitals were obtained; they were phage typed and their resistance to antibiotics was determined. We used the method of routine dilution to test 17 antibiotics against the isolates. Susceptibility of Staphylococcus aureus to studied antibio...

  2. Threat of drug resistant Staphylococcus aureus to health in Nepal

    OpenAIRE

    Ansari, Shamshul; Nepal, Hari Prasad; Gautam, Rajendra; Rayamajhi, Nabin; Shrestha, Sony; Upadhyay, Goma; Acharya, Anju; Chapagain, Moti Lal

    2014-01-01

    Background Staphylococcus aureus is the most commonly isolated organism from the different clinical samples in hospital. The emergence and dissemination of methicillin resistant Staphylococcus aureus (MRSA) and growing resistance to non-beta-lactam antibiotics is making treatment of infections due to this organism increasingly difficult. Methods This study was conducted to determine the frequency of Staphylococcus aureus isolated from different clinical samples, rates of MRSA and full antibio...

  3. Epicutaneous Model of Community-Acquired Staphylococcus aureus Skin Infections

    OpenAIRE

    Prabhakara, Ranjani; Foreman, Oded; De Pascalis, Roberto; Lee, Gloria M.; Plaut, Roger D.; Kim, Stanley Y.; Stibitz, Scott; Elkins, Karen L.; Merkel, Tod J.

    2013-01-01

    Staphylococcus aureus is one of the most common etiological agents of community-acquired skin and soft tissue infection (SSTI). Although the majority of S. aureus community-acquired SSTIs are uncomplicated and self-clearing in nature, some percentage of these cases progress into life-threatening invasive infections. Current animal models of S. aureus SSTI suffer from two drawbacks: these models are a better representation of hospital-acquired SSTI than community-acquired SSTI, and they involv...

  4. Evolution of genes on the Salmonella Virulence plasmid phylogeny revealed from sequencing of the virulence plasmids of S. enterica serotype Dublin and comparative analysis.

    Science.gov (United States)

    Chu, Chishih; Feng, Ye; Chien, An-Chi; Hu, Songnian; Chu, Chi-Hong; Chiu, Cheng-Hsun

    2008-11-01

    Salmonella enterica serotype Dublin harbors an approximately 80-kb virulence plasmid (pSDV), which mediates systemic infection in cattle. There are two types of pSDV: one is pSDVu (pOU1113) in strain OU7025 and the other pSDVr (pOU1115) in OU7409 (SD Lane) and many clinical isolates. Sequence analysis showed that pSDVr was a recombinant plasmid (co-integrate) of pSDVu and a plasmid similar to a 35-kb indigenous plasmid (pOU1114) of S. Dublin. Most of the F-transfer region in pSDVu was replaced by a DNA segment from the pOU1114-like plasmid containing an extra replicon and a pilX operon encoding for a type IV secretion system to form pSDVr. We reconstructed the particular evolutionary history of the seven virulence plasmids of Salmonella by comparative sequence analysis. The whole evolutionary process might begin with two different F-like plasmids (IncFI and IncFII), which then incorporated the spv operon and fimbriae operon from the chromosome to form the primitive virulence plasmids. Subsequently, these plasmids descended by deletion from a relatively large plasmid to smaller ones, with some recombination events occurring over time. Our results suggest that the phylogeny of virulence plasmids as a result of frequent recombination provides the opportunity for rapid evolution of Salmonella in response to the environmental cues. PMID:18718522

  5. Recombinogenic engineering of conjugative plasmids with fluorescent marker cassettes

    DEFF Research Database (Denmark)

    Reisner, A.; Molin, Søren; Zechner, E.L.

    2002-01-01

    An efficient approach for the insertion of fluorescent marker genes with sequence specificity into conjugative plasmids in Escherichia coli is described. For this purpose, homologous recombination of linear double-stranded targeting DNA was mediated by the bacteriophage lambda recombination...... functions using very short regions of homology. Initial manipulation of the IncFII target plasmids R1 and R1drd19 indicated that the linear targeting DNA should be devoid of all extraneous homologies to. the target molecule for optimal insertion specificity. Indeed, a simple recombination assay proved...... resistance genes and fluorescent markers. The choice of 5' non-homologous extensions in primer pairs used for amplifying the marker cassettes determines the site specificity of the targeting DNA. This methodology is applicable to the modification of all plasmids that replicate in E coli and is not restricted...

  6. A novel plasmid pEA68 of Erwinia amylovora and the description of a new family of plasmids.

    Science.gov (United States)

    Ismail, Emadeldeen; Blom, Jochen; Bultreys, Alain; Ivanović, Milan; Obradović, Aleksa; van Doorn, Joop; Bergsma-Vlami, Maria; Maes, Martine; Willems, Anne; Duffy, Brion; Stockwell, Virginia O; Smits, Theo H M; Puławska, Joanna

    2014-12-01

    Recent genome analysis of Erwinia amylovora, the causal agent of fire blight disease on Rosaceae, has shown that the chromosome is highly conserved among strains and that plasmids are the principal source of genomic diversity. A new circular plasmid, pEA68, was found in E. amylovora strain 692 (LMG 28361), isolated in Poland from Sorbus (mountain ash) with fire blight symptoms. Annotation of the 68,763-bp IncFIIa-type plasmid revealed that it contains 79 predicted CDS, among which two operons (tra, pil) are associated with mobility. The plasmid is maintained stably in E. amylovora and does not possess genes associated with antibiotic resistance or known virulence genes. Curing E. amylovora strain 692 of pEA68 did not influence its virulence in apple shoots nor amylovoran synthesis. Of 488 strains of E. amylovora from seventeen countries, pEA68 was only found in two additional strains from Belgium. Although the spread of pEA68 is currently limited to Europe, pEA68 comprises, together with pEA72 and pEA78 both found in North America, a new plasmid family that spans two continents. PMID:25178659

  7. The development of plasmid-free strains of Agrobacterium tumefaciens by using incompatibility with a Rhizobium meliloti plasmid to eliminate pAtC58.

    Science.gov (United States)

    Hynes, M F; Simon, R; Pühler, A

    1985-03-01

    Agrobacterium tumefaciens strains LBA275 and LBA290 were cured of their cryptic plasmid pAtC58 by the introduction of the Rhizobium meliloti plasmid pRme41a, which is incompatible with pAtC58. pRme41a and pTiC58, the resident Ti plasmid of LBA275, were subsequently eliminated by growth at supraoptimal temperature (40 degrees C). The resulting plasmid-free Agrobacterium strains, UBAPF1 and UBAPF2, have proved extremely useful for the study of Rhizobium plasmids. The loss of the cryptic plasmid pAtC58 has no effect on the tumor-forming ability of the Agrobacterium strains; when the Ti plasmid is present, normal tumors are formed on Kalanchoe daigremontiana. PMID:4001194

  8. Characterization of atypical Aeromonas salmonicida isolates by ribotyping and plasmid profiling

    DEFF Research Database (Denmark)

    Pedersen, Karl; Dalsgaard, Inger; Larsen, J.L.

    1996-01-01

    ) and plasmid profiles. Most epidemiologically unrelated strains had different ribotypes, whereas isolates from the same outbreak were identical. All strains, except one, carried one or more large plasmids (>55 kbp) and all strains, except two, additionally carried one or more smaller plasmids. Many strains...... isolated from the same outbreak showed different plasmid profiles although some plasmids were identical. The results suggest the existence of several atypical Aer, salmonicida. It also seems that ribotypes are stable properties for these bacteria while the plasmids are more labile....

  9. Anion exchange purification of plasmid DNA using expanded bed adsorption.

    Science.gov (United States)

    Ferreira, G N; Cabral, J M; Prazeres, D M

    2000-01-01

    Recent developments in gene therapy with non-viral vectors and DNA vaccination have increased the demand for large amounts of pharmaceutical-grade plasmid DNA. The high viscosity of process streams is of major concern in the purification of plasmids, since it can cause high back pressures in column operations, thus limiting the throughput. In order to avoid these high back pressures, expanded bed anion exchange chromatography was evaluated as an alternative to fixed bed chromatography. A Streamline 25 column filled with 100 ml of Streamline QXL media, was equilibrated with 0.5 M NaCl in TE (10 mM Tris, 1 mM EDTA, pH = 8.0) buffer at an upward flow of 300 cmh-1, E. coli lysates (obtained from up to 3 liters of fermentation broth) were injected in the column. After washing out the unbound material, the media was allowed to sediment and the plasmid was eluted with 1 M NaCl in TE buffer at a downward flow of 120 cmh-1. Purification factors of 36 +/- 1 fold, 26 +/- 0.4 plasmid purity, and close to 100% yields were obtained when less than one settled column volume of plasmid feed was injected. However, both recovery yield and purity abruptly decreased when larger amounts were processed-values of 35 +/- 2 and 5 +/- 0.7 were obtained for the recovery yield and purity, respectively, when 250 ml of feedstock were processed. In these cases, gel clogging and expansion collapse were observed. The processing of larger volumes, thus larger plasmid quantities, was only possible by performing an isopropanol precipitation step prior to the chromatographic step. This step led to an enhancement of the purification step. PMID:10840595

  10. Evaluation of phenotypic and genotypic methods for epidemiological typing of Staphylococcus aureus isolates from bovine mastitis in Denmark

    DEFF Research Database (Denmark)

    Aarestrup, Frank Møller; Wegener, H. C.; Rosdahl, V. T.

    The value of five different typing methods (antibiogram typing, biotyping, phage typing, plasmid profiling and restriction fragment length polymorphism of the gene encoding 16S and 23S ribosomal RNA (ribotyping)), in discriminating 105 Staphylococcus aureus strains from bovine milk samples obtained...... from 105 different Danish dairy herds was investigated. A total of 85 strains (81%) proved susceptible to all of the 11 antibiotics tested, and the remaining 20 strains could be divided into 5 different antibiogram patterns. The predominant resistance pattern, penicillin resistance, was observed in 15...... (75%) of the 20 antibiotic resistant strains. Biotyping assigned the strains to 14 different types, with the most common type accounting for 25.7% of the strains. Ninety eight (93.3%) strains could be typed by phages, assigning them to 19 different phage types. The predominant phage type accounted for...

  11. Sustained plasmid DNA release from dissolving mineral coatings

    OpenAIRE

    Choi, Siyoung; Murphy, William L.

    2010-01-01

    Calcium phosphate (Ca-P) minerals such as hydroxyapatite are able to bind a diverse range of biological molecules due to the presence of anions and cations in their crystal structure. The well-characterized ability of Ca-P minerals to bind and release plasmid DNA, coupled with the ability of biodegradable Ca-P coatings to form on the surface of common biomaterials, provides a potential mechanism for controlled release of plasmid DNA from various biomaterials. In this study we hypothesized tha...

  12. Restriction Fragment Length Polymorphisms of Virulence Plasmids in Rhodococcus equi

    OpenAIRE

    Takai, Shinji; Shoda, Masato; Sasaki, Yukako; Tsubaki, Shiro; Fortier, Guillaume; Pronost, Stephane; Rahal, Karim; Becu, Teotimo; Begg, Angela; Browning, Glenn; Nicholson, Vivian M.; Prescott, John F.

    1999-01-01

    Virulent Rhodococcus equi, which is a well-known cause of pyogranulomatous pneumonia in foals, possesses a large plasmid encoding virulence-associated 15- to 17-kDa antigens. Foal and soil isolates from five countries—Argentina, Australia, Canada, France, and Japan—were investigated for the presence of 15- to 17-kDa antigens by colony blotting, using the monoclonal antibody 10G5, and the gene coding for 15- to 17-kDa antigens by PCR. Plasmid DNAs extracted from positive isolates were digested...

  13. VISA/VRSA (Vancomycin-Intermediate/Resistant Staphylococcus aureus) in Healthcare Settings

    Science.gov (United States)

    ... to vancomycin and other antimicrobial agents. What is Staphylococcus aureus? Staphylococcus aureus is a bacterium commonly found on the ... control personnel. Investigation and Control of Vancomycin-Resistant Staphylococcus aureus (VRSA) [PDF - 300 KB] - This document is ...

  14. DNA sequence analysis of plasmids from multidrug resistant Salmonella enterica serotype Heidelberg isolates.

    Directory of Open Access Journals (Sweden)

    Jing Han

    Full Text Available Salmonella enterica serovar Heidelberg is among the most detected serovars in swine and poultry, ranks among the top five serotypes associated with human salmonellosis and is disproportionately associated with invasive infections and mortality in humans. Salmonella are known to carry plasmids associated with antimicrobial resistance and virulence. To identify plasmid-associated genes in multidrug resistant S. enterica serovar Heidelberg, antimicrobial resistance plasmids from five isolates were sequenced using the 454 LifeSciences pyrosequencing technology. Four of the isolates contained incompatibility group (Inc A/C multidrug resistance plasmids harboring at least eight antimicrobial resistance genes. Each of these strains also carried a second resistance plasmid including two IncFIB, an IncHI2 and a plasmid lacking an identified Inc group. The fifth isolate contained an IncI1 plasmid, encoding resistance to gentamicin, streptomycin and sulfonamides. Some of the IncA/C plasmids lacked the full concert of transfer genes and yet were able to be conjugally transferred, likely due to the transfer genes carried on the companion plasmids in the strains. Several non-IncA/C resistance plasmids also carried putative virulence genes. When the sequences were compared to previously sequenced plasmids, it was found that while all plasmids demonstrated some similarity to other plasmids, they were unique, often due to differences in mobile genetic elements in the plasmids. Our study suggests that Salmonella Heidelberg isolates harbor plasmids that co-select for antimicrobial resistance and virulence, along with genes that can mediate the transfer of plasmids within and among other bacterial isolates. Prevalence of such plasmids can complicate efforts to control the spread of S. enterica serovar Heidelberg in food animal and human populations.

  15. Genetic Characterization of ExPEC-Like Virulence Plasmids among a Subset of NMEC.

    Science.gov (United States)

    Nicholson, Bryon A; West, Aaron C; Mangiamele, Paul; Barbieri, Nicolle; Wannemuehler, Yvonne; Nolan, Lisa K; Logue, Catherine M; Li, Ganwu

    2016-01-01

    Neonatal Meningitis Escherichia coli (NMEC) is one of the most common causes of neonatal bacterial meningitis in the US and elsewhere resulting in mortality or neurologic deficits in survivors. Large plasmids have been shown experimentally to increase the virulence of NMEC in the rat model of neonatal meningitis. Here, 9 ExPEC-like plasmids were isolated from NMEC and sequenced to identify the core and accessory plasmid genes of ExPEC-like virulence plasmids in NMEC and create an expanded plasmid phylogeny. Results showed sequenced virulence plasmids carry a strongly conserved core of genes with predicted functions in five distinct categories including: virulence, metabolism, plasmid stability, mobile elements, and unknown genes. The major functions of virulence-associated and plasmid core genes serve to increase in vivo fitness by adding multiple iron uptake systems to the genetic repertoire to facilitate NMEC's survival in the host's low iron environment, and systems to enhance bacterial resistance to host innate immunity. Phylogenetic analysis based on these core plasmid genes showed that at least two lineages of ExPEC-like plasmids could be discerned. Further, virulence plasmids from Avian Pathogenic E. coli and NMEC plasmids could not be differentiated based solely on the genes of the core plasmid genome. PMID:26800268

  16. Genetic Characterization of ExPEC-Like Virulence Plasmids among a Subset of NMEC.

    Directory of Open Access Journals (Sweden)

    Bryon A Nicholson

    Full Text Available Neonatal Meningitis Escherichia coli (NMEC is one of the most common causes of neonatal bacterial meningitis in the US and elsewhere resulting in mortality or neurologic deficits in survivors. Large plasmids have been shown experimentally to increase the virulence of NMEC in the rat model of neonatal meningitis. Here, 9 ExPEC-like plasmids were isolated from NMEC and sequenced to identify the core and accessory plasmid genes of ExPEC-like virulence plasmids in NMEC and create an expanded plasmid phylogeny. Results showed sequenced virulence plasmids carry a strongly conserved core of genes with predicted functions in five distinct categories including: virulence, metabolism, plasmid stability, mobile elements, and unknown genes. The major functions of virulence-associated and plasmid core genes serve to increase in vivo fitness by adding multiple iron uptake systems to the genetic repertoire to facilitate NMEC's survival in the host's low iron environment, and systems to enhance bacterial resistance to host innate immunity. Phylogenetic analysis based on these core plasmid genes showed that at least two lineages of ExPEC-like plasmids could be discerned. Further, virulence plasmids from Avian Pathogenic E. coli and NMEC plasmids could not be differentiated based solely on the genes of the core plasmid genome.

  17. Staphylococcus aureus: resistance pattern and risk factors

    Directory of Open Access Journals (Sweden)

    Mohammad Naghavi-Behzad

    2015-03-01

    Full Text Available Introduction: Methicillin resistant Staphylococcus aureus (MRSA has emerged as a nosocomial pathogen of major worldwide importance and is an increasingly frequent cause of community-acquired infections. In this study, different risk factors and MRSA resistance pattern were investigated. Methods: In a 24 months period, all of the patients who were confined to bed in the surgery ward were included in the study. Then they were assessed to find out as if they had MRSA infection when hospitalized and once when they were discharged. Almost 48 h after admission, when patients were discharged, social and medical histories were acquired. Acquired samples were examined. Results: During the present study of 475 patients, 108 patients (22.8% had S. aureus. About frequency of antibiotic resistance among collected S. aureus colonies, erythromycin resistance, was the most frequent antibiotic resistance, also resistance to vancomycin was 0.4% that was the least. Only hospitalization duration had statistically significant correlation with antibiotic resistance, also resistance to erythromycin had statistically significant relation with history of surgery and alcohol consumption. Of all 34 MRSA species, 22 (64.7% samples were resistant to erythromycin, 17 (50.0% resistant to cefoxitin, 5 (14.7% resistant to mupirocin, 1 (2.9% resistant to vancomycin and 1 (2.9% resistant to linezolid. Conclusion: The results of the current study show that among hospitalized patients, there is resistance against methicillin. Since based on results of the study there is resistance against oxacillin and erythromycin in most cases, administering appropriate antibiotics have an important role in minimizing the resistance burden among bacterial species.

  18. [Methicillin-resistant Staphylococcus aureus (MRSA) in veterinary medicine: a "new emerging pathogen"?].

    Science.gov (United States)

    Walther, Birgit; Friedrich, Alexander W; Brunnberg, Leo; Wieler, Lothar H; Lübke-Becker, Antina

    2006-01-01

    The problem of nosocomial infections is of increasing importance in veterinary medicine. As an example, this review summarizes current knowledge regarding methicillin-resistant Staphylococcus aureus (MRSA) as a typical example, as these pathogens are the most important agents of nosocomial infections in human medicine worldwide and are being increasingly reported in veterinary medicine. MRSA are classified by their ability to be resistant against oxacillin/methicillin, this feature being confered by mecA, a gene which was acquired by horizontal gene transfer of the staphylococcal gene cassette (SCCmec). It is this genetic information that enables MRSA to be resistant against all penicillins, cehalosporins and carbapenems. In addition, MRSA are often resistant against a variety of other antiinfectives, i.e. aminoglycosides, macrolides, lincosamide, streptomycins, tetracyclin, chloramphenicol, but also against fluorquinolones and rifampicin. Presumably, these highly adapted strains are particularly able to acquire resistance genes located on plasmids or transposons. They are also able to develop point mutations, further leading to resistant phenotypes. If these pathogens are leading to infectious diseases, veterinarians may be confronted with a worst-case scenario, being left without any antiinfective therapeutic. As Staphylococcus aureus is highly tenacid, professional hygiene management is of utmost importance. The increasing number of published sporadic MRSA infections, MRSA-infectious diseases as well as MRSA outbreaks in veterinary medicine justifies their recognition as a "New Emerging Pathogen". So far, horses and dogs are mostly affected by MRSA. Although transmission between humans and animals has been reported occasionally, the sources, routes of transmission or the epidemiological relevance of MRSA infections in animals are far from being understood. Therefore, epidemiological investigations utilizing molecular typing tools are mandatory. Typing tools like

  19. Construction and Use of Flow Cytometry Optimized Plasmid-Sensor Strains

    DEFF Research Database (Denmark)

    Bahl, Martin Iain; Oregaard, Gunnar; Sørensen, Søren Johannes;

    2009-01-01

    Determining the stability of plasmids in bacterial populations is traditionally performed by isolating a large number of clones followed by screening for the presence of plasmids by replica transfer to plasmid-selective agar plates. This is often a laborious task, especially when the intrinsic...... stability of the plasmid is high. The method presented here relies on a phenotypic (green fluorescence protein) marker, which is switched on if the host bacteria loses the residing plasmid. The incorporation of flow cytometry for single-cell detection and discrimination between plasmid-free and plasmid......-harboring cells in a bacterial population facilitates a very high throughput of cells and thus provides excellent sensitivity and statistics toward detecting even very low levels of plasmid instability....

  20. Survival and evolution of a large multidrug resistance plasmid in new clinical bacterial hosts

    DEFF Research Database (Denmark)

    Porse, Andreas; Schønning, Kristian; Munck, Christian;

    2016-01-01

    and population sequencing to show that the long-term persistence and molecular integrity of the plasmid is highly influenced by multiple factors within a 25 kb plasmid region constituting a host-dependent burden. In the E. coli hosts investigated here, improved plasmid stability readily evolves via IS......Large conjugative plasmids are important drivers of bacterial evolution and contribute significantly to the dissemination of antibiotic resistance. Although plasmid borne multidrug resistance is recognized as one of the main challenges in modern medicine, the adaptive forces shaping the evolution...... of these plasmids within pathogenic hosts are poorly understood. Here we study plasmid-host adaptations following transfer of a 73 kb conjugative multidrug resistance plasmid to naïve clinical isolates of Klebsiella pneumoniae and Escherichia coli We use experimental evolution, mathematical modelling...

  1. Survival and evolution of a large multidrug resistance plasmid in new clinical bacterial hosts

    DEFF Research Database (Denmark)

    Porse, Andreas; Schønning, Kristian; Munck, Christian;

    2016-01-01

    Large conjugative plasmids are important drivers of bacterial evolution and contribute significantly to the dissemination of antibiotic resistance. Although plasmid borne multidrug resistance is recognized as one of the main challenges in modern medicine, the adaptive forces shaping the evolution...... of these plasmids within pathogenic hosts are poorly understood. Here we study plasmid-host adaptations following transfer of a 73 kb conjugative multidrug resistance plasmid to naïve clinical isolates of Klebsiella pneumoniae and Escherichia coli We use experimental evolution, mathematical modelling and population...... sequencing to show that the long-term persistence and molecular integrity of the plasmid is highly influenced by multiple factors within a 25 kb plasmid region constituting a host-dependent burden. In the E. coli hosts investigated here, improved plasmid stability readily evolves via IS26 mediated deletions...

  2. Conservation of plasmids among plant-pathogenic Pseudomonas syringae isolates of diverse origins.

    Science.gov (United States)

    von Bodman, S B; Shaw, P D

    1987-05-01

    Thirty isolates of Pseudomonas syringae pv. tabaci, pv. angulata (pathogens on tobacco), pv. coronafaciens, and pv. striafaciens (pathogens on oats) were examined for plasmid DNAs. The strains were obtained from plants throughout the world, some over 50 years ago. Of the 22 tobacco pathogens, 16 contain predominantly one type of plasmid, the pJP27.00 type. The remaining six tobacco-specific strains do not harbor detectable plasmids. The oat pathogens contain one, two, or three plasmids. DNA homology studies indicate that the plasmid DNAs are highly conserved. More importantly, the plasmids harbored by strains isolated from one host plant are conserved most stringently; e.g., the plasmids from the tobacco pathogens are, with one exception, indistinguishable by restriction endonuclease digestion and Southern hybridization. There is also extensive homology among plasmids indigenous to the oat-specific P. syringae pv. coronafaciens and pv. striafaciens strains. PMID:3628554

  3. Possible involvement of a plasmid in arginine auxotrophic mutation of Streptomyces kasugaensis.

    OpenAIRE

    Nakano, M M; Ozawa, K; Ogawara, H

    1980-01-01

    Streptomyces kasugaensis gave arginine auxotrophic mutants at high frequency, The coupled loss and reappearance of plasmid deoxyribonucleic acid with arginine auxotrophy suggested that the insertion of the plasmid into chromosomal deoxyribonucleic acid caused the arginine auxotrophy.

  4. A Time-Efficient and User-Friendly Method for Plasmid DNA Restriction Analysis.

    Science.gov (United States)

    LaBanca, Frank; Berg, Claire M.

    1998-01-01

    Describes an experiment in which plasmid DNA is digested with restriction enzymes that cleave the plasmid either once or twice. The DNA is stained, loaded on a gel, electrophoresed, and viewed under normal laboratory conditions during electrophoresis. (DDR)

  5. Transcription of ColE1Ap mbeC induced by conjugative plasmids from twelve different incompatibility groups.

    OpenAIRE

    Selvaratnam, S; Gealt, M A

    1993-01-01

    Although nonconjugative mobilizable plasmids require helping functions of conjugative plasmids in order to be mobilized into recipients, at least some genes from the nonconjugative plasmids may be induced to assist in the DNA transfer process. Conjugative plasmids from 12 different incompatibility groups mobilized the nonconjugative plasmid ColE1Ap between Escherichia coli strains. Introduction of any of the conjugative plasmids into the ColE1Ap-containing strain resulted in an induction of m...

  6. Partition locus-based classification of selected plasmids in Klebsiella pneumoniae, Escherichia coli and Salmonella enterica spp.: An additional tool

    OpenAIRE

    Bousquet, A.; Henquet, S.; Compain, F.; Genel, N.; Arlet, G.; Decré, D

    2015-01-01

    The dissemination of antimicrobial resistance genes in Enterobacteriaceae has been largely attributed to plasmids, circular DNA molecules capable of autonomous replication. Whereas high-copy-number plasmids primarily rely on passive diffusion for plasmid maintenance, low-copy-number plasmids utilize so-called partition (par) systems. Plasmid partition relies on three structures, i.e. a centromere like DNA site, a centromere-binding protein and an ATPase or a GTPase motor protein for plasmid p...

  7. New tetracycline resistance determinant on R plasmids from Vibrio anguillarum.

    OpenAIRE

    Aoki, T.; Satoh, T.; Kitao, T.

    1987-01-01

    Two classes of tetracycline resistance determinants on R plasmids were detected in Vibrio anguillarum strains isolated from ayu (sweat fish; Plecoglossus altivelis) farms in Japan. Tetracycline resistance genes categorized as class B were prevalent from 1973 to 1977; however, a new tetracycline resistance gene, which was not classified into tetracycline resistance determinant class A, B, C, or D, has been prevalent since 1981.

  8. Pharmaceutical development of the plasmid DNA vaccine pDERMATT

    NARCIS (Netherlands)

    Quaak, S.G.L.

    2009-01-01

    The discovery of tumor specific antigens and self tolerance mechanisms against these antigens led to the assumption that antigens circulating at sufficient concentration levels could break this self tolerance mechanism and evoke immunological antitumor effects. pDERMATT (plasmid DNA encoding recombi

  9. Stability of Integrated Plasmids in the Chromosome of Lactococcus lactis

    NARCIS (Netherlands)

    Leenhouts, Kees J.; Kok, Jan; Venema, Gerhardus

    1990-01-01

    Derivatives of plasmids pBR322, pUB110, pSC101, and pTB19, all containing an identical fragment of lactococcal chromosomal DNA, were integrated via a Campbell-like mechanism into the same chromosomal site of Lactococcus lactis MG1363, and the transformants were analyzed for the stability of the inte

  10. Plasmid-determined copper resistance in Pseudomonas syringae from impatiens

    Energy Technology Data Exchange (ETDEWEB)

    Cooksey, D.A. (Univ. of California, Riverside (USA))

    1990-01-01

    A strain of Pseudomonas syringae was recently identified as the cause of a new foliar blight of impatiens. The bacterium was resistant to copper compounds, which are used on a variety of crops for bacterial and fungal disease control. The bacterium contained a single 47-kilobase plasmid (pPSI1) that showed homology to a copper resistance operon previously cloned and characterized from P. syringae pv. tomato plasmid pPT23D (D. Cooksey, Appl. Environ. Microbiol. 53:454-456, 1987). pPSI1 was transformed by electroporation into a copper-sensitive P. syringae strain, and the resulting transformants were copper resistant. A physical map of pPSI1 was constructed, and the extent of homology to pPT23D outside the copper resistance operon was determined in Southern hybridizations. The two plasmids shared approximately 20 kilobases of homologous DNA, with the remainder of each plasmid showing no detectable homology. The homologous regions hybridized strongly, but there was little or no conservation of restriction enzyme recognition sites.

  11. Studying plasmid horizontal transfer in situ: a critical review

    DEFF Research Database (Denmark)

    Sørensen, Søren Johannes; Bailey, Mark; Hansen, Lars Hestbjerg;

    2005-01-01

    This review deals with the prospective, experimental documentation of horizontal gene transfer (HGT) and its role in real-time, local adaptation. We have focused on plasmids and their function as an accessory and/or adaptive gene pool. Studies of the extent of HGT in natural environments have...

  12. Use of plasmid DNA for induction of protective immunity

    DEFF Research Database (Denmark)

    Lorenzen, Niels

    2004-01-01

    Vaccines based on plasmid DNA have been tested for a number of fish pathogens but so far it is only in case of the rhabdoviruses, where the technology has been a real break through in vaccine research. Aspects of dose, time-course and mechanisms of protection, as well as practical use are discussed....

  13. Multilocus sequence typing of IncN plasmids

    DEFF Research Database (Denmark)

    García-Fernández, Aurora; Villa, Laura; Moodley, Arshnee; Hasman, Henrik; Miriagou, Vivi; Guardabassi, Luca; Carattoli, Alessandra

    2011-01-01

    OBJECTIVES: Incompatibility group N (IncN) plasmids have been associated with the dissemination of antimicrobial resistance and are a major vehicle for the spread of blaVIM-1 in humans and blaCTX-M-1 in animals. A plasmid multilocus sequence typing (pMLST) scheme was developed for rapid......, Greece, Denmark, UK and The Netherlands) were classified by DNA sequencing of the amplicons obtained for the repA, traJ and korA loci. RESULTS: Eleven sequence types (STs) were defined on the basis of allele sequences of the three selected loci. Most plasmids carrying blaCTX-M-1 (24/27) isolated in...... spread and persistence of this particular IncN-carrying blaVIM-1 lineage in Greece. CONCLUSIONS: This study proposes the use of pMLST as a suitable and rapid method for identification of IncN epidemic plasmid lineages. The recent spread of blaCTX-M-1 among humans and animals seems to be associated with...

  14. [Transfer of plasmid beta-lactamases in enterobacteria].

    Science.gov (United States)

    Umaran, A; Garaizar, J; Gallego, L; Colom, K; Cisterna, R

    1989-04-01

    The aim of the present study was to determine which types of beta-lactamases codified by plasmids are transferred by conjugation from several species of enterobacteria. To this end, 352 strains of ampicillin-resistant enterobacteria from clinical samples from the Hospital Civil of Bilbao were evaluated. Their beta-lactamase activity and their capacity to transfer this capacity by conjugation were evaluated. The several types of plasmidic beta-lactamases in the strains that conjugated and in their respective transconjugants were characterized by analytic isoelectric approach, and also the sensitivity of these stains to 20 beta-lactamic antibiotics and the size of their plasmids. Twenty different types were detected, with a clear predominance of TEM 1. Type TEM 2 was found in 19% of the strains which conjugated, and much less commonly the types SHV 1, HMS 1 and a beta-lactamase of an approximate pl of 4.9 were found. The transfer of these beta-lactamases is mediated by a great variety of plasmids and is associated with variable levels of resistance to penicillins and unstable cephalosporins. The presence of betalactamases with activity on the more stable cephalosporins has not been detected. PMID:2490696

  15. Plasmid profiles of clinical and environmental isolates of Legionella pneumophila serogroup 1.

    OpenAIRE

    Maher, W E; Plouffe, J F; Para, M F

    1983-01-01

    Clinical and environmental Legionella pneumophila serogroup 1 isolates from a single water source in Columbus, Ohio, exhibited five different plasmid profiles. The multiplicity of plasmid profiles observed within a single geographic area and a skewed distribution of isolates bearing these plasmid profiles within this area suggest that plasmid analyses will be useful in the study of the epidemiology of Legionnaires disease and the environmental distribution of legionellae.

  16. Plasmid marker rescue transformation proceeds by breakage-reunion in Bacillus subtilis.

    OpenAIRE

    Weinrauch, Y; Dubnau, D

    1987-01-01

    Bacillus subtilis carrying a plasmid which replicates with a copy number of about 1 was transformed with linearized homologous plasmid DNA labeled with the heavy isotopes 2H and 15N, in the presence of 32Pi and 6-(p-hydroxyphenylazo)-uracil to inhibit DNA replication. Plasmid DNA was isolated from the transformed culture and fractionated in cesium chloride density gradients. The distribution of total and donor plasmid DNA was examined, using specific hybridization probes. The synthesis of new...

  17. Incidence of Plasmids in Thermus spp. Isolated in Yellowstone National Park

    OpenAIRE

    Munster, Michael J.; Munster, Ann P.; Sharp, Richard J.

    1985-01-01

    Forty-eight strains of Thermus spp. were isolated from thermal sites in Yellowstone National Park, Wyo., and 62.5% showed evidence of plasmid DNA. Attempts to assign function to the plasmid DNA were unsuccessful, and the presence of plasmid DNA could not be correlated with antibiotic or heavy metal resistance. A number of these cryptic plasmids are now being investigated for their potential as vectors for molecular cloning in Thermus spp.

  18. A DNA polymerase mutation that suppresses the segregation bias of an ARS plasmid in Saccharomyces cerevisiae.

    OpenAIRE

    Houtteman, S W; Elder, R T

    1993-01-01

    Yeast autonomously replicating sequence (ARS) plasmids exhibit an unusual segregation pattern during mitosis. While the nucleus divides equally into mother and daughter cells, all copies of the ARS plasmid will often remain in the mother cell. A screen was designed to isolate mutations that suppress this segregation bias. A plasmid with a weak ARS (wARS) that displayed an extremely high segregation bias was constructed. When cells were grown under selection for the wARS plasmid, the resulting...

  19. Modulation of ColE1-like Plasmid Replication for Recombinant Gene Expression

    OpenAIRE

    Camps, Manel

    2010-01-01

    ColE1-like plasmids constitute the most popular vectors for recombinant protein expression. ColE1 plasmid replication is tightly controlled by an antisense RNA mechanism that is highly dynamic, tuning plasmid metabolic burden to the physiological state of the host. Plasmid homeostasis is upset upon induction of recombinant protein expression because of non-physiological levels of expression and because of the frequently biased amino acid composition of recombinant proteins. Disregulation of p...

  20. Participation of the lytic replicon in bacteriophage P1 plasmid maintenance.

    OpenAIRE

    Yarmolinsky, M B; Hansen, E B; Jafri, S; Chattoraj, D K

    1989-01-01

    P1 bacteriophage carries at least two replicons: a plasmid replicon and a viral lytic replicon. Since the isolated plasmid replicon can maintain itself stably at the low copy number characteristic of intact P1 prophage, it has been assumed that this replicon is responsible for driving prophage replication. We provide evidence that when replication from the plasmid replicon is prevented, prophage replication continues, albeit at a reduced rate. The residual plasmid replication is due to incomp...

  1. Stability in Escherichia coli of an antibiotic resistance plasmid from Bacteroides fragilis.

    OpenAIRE

    Rashtchian, A; Booth, S J

    1981-01-01

    A Bacteroides fragilis strain resistant to penicillin G, tetracycline, and clindamycin was screened for the presence of plasmid deoxyribonucleic acid (DNA). Agarose gel electrophoresis of ethanol-precipitated DNA from cleared lysates of this strain revealed two plasmid DNA bands. The molecular weights of the plasmids were estimated by their relative mobility in agarose gel and compared with standard plasmids with known molecular weights. The molecular weights were 3.40 +/- 0.20 x 10(6) and 1....

  2. Properties of R1162, a broad-host-range, high-copy-number plasmid.

    OpenAIRE

    R. MEYER; Hinds, M; Brasch, M.

    1982-01-01

    Regions of plasmid DNA encoding characteristic properties of the IncQ (P-4) group plasmid R1162 were identified by mutagenesis and in vitro cloning. Coding sequences sufficient for expression of incompatibility and efficient conjugal mobilization by plasmid R751 were found to be linked to the origin of DNA replication. In contrast, there was a region remote from the origin, and active in trans, that was required for plasmid maintenance. A derivative that was temperature sensitive for stabilit...

  3. Characterization of different plasmid-borne dihydropteroate synthases mediating bacterial resistance to sulfonamides.

    OpenAIRE

    Swedberg, G; Sköld, O

    1980-01-01

    Plasmid-borne resistance to sulfonamides was studied in both newly isolated and earlier characterized R plasmids. Two different classes of drug-resistant dihydropteroate synthases were found to be responsible for most cases of plasmid-mediated sulfonamide resistance. The plasmid-coded enzymes could be completely separated from their chromosomal counterpart and also showed differences in heat stability and molecular size. The resistant and chromosomal enzymes could bind the normal substrate, p...

  4. Use of agarose gel electrophoresis of plasmid deoxyribonucleic acid to fingerprint gram-negative bacilli.

    OpenAIRE

    Schaberg, D.R.; Tompkins, L S; Falkow, S

    1981-01-01

    Agarose gel electrophoresis of the plasmid deoxyribonucleic acids from 60 gram-negative bacilli recovered during investigations of nosocomial epidemics was used to fingerprint the strains. This method was as specific at differentiating bacterial strains as more conventional phenotyping methods. In all cases, plasmid band fingerprints of epidermic strains isolates were identical whereas coisolate plasmid deoxyribonucleic acid patterns were different. Agarose gel electrophoresis of plasmid deox...

  5. Identification of tetracycline-resistant R-plasmids in Streptococcus agalactiae (group B).

    OpenAIRE

    Burdett, V

    1980-01-01

    In this report, 30 tetracycline-resistant clinical isolates of group B Streptococcus were examined to assess the extent to which tetracycline resistance is plasmid mediated. Of these, 27 showed no physical or genetic evidence of plasmid-mediated resistance; however, one conjugative and two small (3.5 X 10(6)-dalton) multicopy non-self-transmissible tetracycline resistance plasmids were identified. The conjugative plasmid was transmissible to Streptococcus faecalis as well as to Streptococcus ...

  6. The Salmonella virulence plasmid enhances Salmonella-induced lysis of macrophages and influences inflammatory responses.

    OpenAIRE

    Guilloteau, L. A.; Wallis, T S; A.V. Gautier; Macintyre, S.; Platt, D. J.; Lax, A J

    1996-01-01

    The Salmonella dublin virulence plasmid mediates systemic infection in mice and cattle. Here, we analyze the interaction between wild-type and plasmid-cured Salmonella strains with phagocytes in vitro and in vivo. The intracellular recovery of S. dublin from murine peritoneal and bovine alveolar macrophages cultured in the presence of gentamicin in vitro was not related to virulence plasmid carriage. However, the virulence plasmid increased the lytic activity of S. dublin, Salmonella typhimur...

  7. Complete DNA Sequence and Analysis of the Large Virulence Plasmid of Shigella flexneri

    OpenAIRE

    Venkatesan, Malabi M.; Goldberg, Marcia B.; Rose, Debra J.; Grotbeck, Erik J.; Burland, Valerie; Blattner, Frederick R.

    2001-01-01

    The complete sequence analysis of the 210-kb Shigella flexneri 5a virulence plasmid was determined. Shigella spp. cause dysentery and diarrhea by invasion and spread through the colonic mucosa. Most of the known Shigella virulence determinants are encoded on a large plasmid that is unique to virulent strains of Shigella and enteroinvasive Escherichia coli; these known genes account for approximately 30 to 35% of the virulence plasmid. In the complete sequence of the virulence plasmid, 286 ope...

  8. The critical role of the linear plasmid lp36 in the infectious cycle of Borrelia burgdorferi

    OpenAIRE

    Mollie W Jewett; Lawrence, Kevin; Bestor, Aaron C; Tilly, Kit; Grimm, Dorothee; Shaw, Pamela; VanRaden, Mark; Gherardini, Frank; Rosa, Patricia A.

    2007-01-01

    Borrelia burgdorferi, the aetiological agent of Lyme disease, follows a life cycle that involves passage between the tick vector and the mammalian host. To investigate the role of the 36 kb linear plasmid, lp36 (also designated the B. burgdorferi K plasmid), in the infectious cycle of B. burgdorferi, we examined a clone lacking this plasmid, but containing all other plasmids known to be required for infectivity. Our results indicated that lp36 was not required for spirochete survival in the t...

  9. Conservation of Plasmid-Encoded Traits among Bean-Nodulating Rhizobium Species

    OpenAIRE

    Brom, Susana; Girard, Lourdes; García-de los Santos, Alejandro; Sanjuan-Pinilla, Julio M.; Olivares, José; Sanjuan, Juan

    2002-01-01

    Rhizobium etli type strain CFN42 contains six plasmids. We analyzed the distribution of genetic markers from some of these plasmids in bean-nodulating strains belonging to different species (Rhizobium etli, Rhizobium gallicum, Rhizobium giardinii, Rhizobium leguminosarum, and Sinorhizobium fredii). Our results indicate that independent of geographic origin, R. etli strains usually share not only the pSym plasmid but also other plasmids containing symbiosis-related genes, with a similar organi...

  10. Tn5-mediated transposition of plasmid DNA after transduction to Myxococcus xanthus.

    OpenAIRE

    Downard, J S

    1988-01-01

    After coliphage P1-mediated transfer of Tn5-containing plasmid DNA from Escherichia coli to Myxococcus xanthus, transductants were identified which contained plasmid sequences integrated at many sites on the bacterial chromosome. The unaltered plasmid DNA sequences in these transductants were apparently flanked by intact Tn5 or IS50 sequences. These results suggest that Tn5-mediated transposition has occurred and provide a method for integrating plasmid DNA into the M. xanthus chromosome with...

  11. Microneedle-mediated transcutaneous immunization with plasmid DNA coated on cationic PLGA nanoparticles

    OpenAIRE

    Kumar, Amit; Wonganan, Piyanuch; Sandoval, Michael A.; Li, Xinran; Zhu, Saijie; Cui, Zhengrong

    2012-01-01

    Previously, it was shown that microneedle-mediated transcutaneous immunization with plasmid DNA can potentially induce a stronger immune response than intramuscular injection of the same plasmid DNA. In the present study, we showed that the immune responses induced by transcutaneous immunization by applying plasmid DNA onto a skin area pretreated with solid microneedles were significantly enhanced by coating the plasmid DNA on the surface of cationic nanoparticles. In addition, the net surfac...

  12. Characterization of the OCT plasmid encoding alkane oxidation and mercury resistance in Pseudomonas putida.

    OpenAIRE

    Harder, P A; Kunz, D A

    1986-01-01

    Transformation of Pseudomonas putida and analysis for plasmid DNA revealed that both n-alkane oxidation and mercury resistance are encoded on a single 220-megadalton OCT plasmid molecule. Derivatives of OCT having lost the mercury resistance function could be readily isolated and contained a smaller plasmid estimated to be 170 megadaltons. The results show that segregation of the mercury resistance property occurs not by loss of a separate MER plasmid as previously thought but by a deletion i...

  13. Enhanced expressions and histological characteristics of intravenously administered plasmid DNA in rat lung.

    OpenAIRE

    Rha, S. J.; Y. P. Wang

    2001-01-01

    Cationic liposome-mediated gene transfection is a promising method for gene therapy. In this study, the transfection efficiency and histological patterns were evaluated in rat lung after intravenous administration via femoral vein of naked plasmid DNA, naked plasmid DNA with pretreatment of DOTAP, and DOTAP-cholesterol-plasmid DNA complex. Plasmid DNA encoding bacterial LacZ gene was used. For quantification of LacZ gene expression, beta-galactosidase assay was performed. For histologic exami...

  14. Comparison of electrically mediated and liposome-complexed plasmid DNA delivery to the skin

    OpenAIRE

    Heller, Loree C.; Jaroszeski, Mark J; Coppola, Domenico; Heller, Richard

    2008-01-01

    Background Electroporation is an established technique for enhancing plasmid delivery to many tissues in vivo, including the skin. We have previously demonstrated efficient delivery of plasmid DNA to the skin utilizing a custom-built four-plate electrode. The experiments described here further evaluate cutaneous plasmid delivery using in vivo electroporation. Plasmid expression levels are compared to those after liposome mediated delivery. Methods Enhanced electrically-mediated delivery, and ...

  15. Staphylococcus aureus bacteriuria as a prognosticator for outcome of Staphylococcus aureus bacteremia: a case-control study

    Directory of Open Access Journals (Sweden)

    Weinstein Robert A

    2010-07-01

    Full Text Available Abstract Background When Staphylococcus aureus is isolated in urine, it is thought to usually represent hematogenous spread. Because such spread might have special clinical significance, we evaluated predictors and outcomes of S. aureus bacteriuria among patients with S. aureus bacteremia. Methods A case-control study was performed at John H. Stroger Jr. Hospital of Cook County among adult inpatients during January 2002-December 2006. Cases and controls had positive and negative urine cultures, respectively, for S. aureus, within 72 hours of positive blood culture for S. aureus. Controls were sampled randomly in a 1:4 ratio. Univariate and multivariable logistic regression analyses were done. Results Overall, 59% of patients were African-American, 12% died, 56% of infections had community-onset infections, and 58% were infected with methicillin-susceptible S. aureus (MSSA. Among 61 cases and 247 controls, predictors of S. aureus bacteriuria on multivariate analysis were urological surgery (OR = 3.4, p = 0.06 and genitourinary infection (OR = 9.2, p = 0.002. Among patients who died, there were significantly more patients with bacteriuria than among patients who survived (39% vs. 17%; p = 0.002. In multiple Cox regression analysis, death risks in bacteremic patients were bacteriuria (hazard ratio 2.9, CI 1.4-5.9, p = 0.004, bladder catheter use (2.0, 1.0-4.0, p = 0.06, and Charlson score (1.1, 1.1-1.3, p = 0.02. Neither length of stay nor methicillin-resistant Staphylococcus aureus (MRSA infection was a predictor of S. aureus bacteriuria or death. Conclusions Among patients with S. aureus bacteremia, those with S. aureus bacteriuria had 3-fold higher mortality than those without bacteriuria, even after adjustment for comorbidities. Bacteriuria may identify patients with more severe bacteremia, who are at risk of worse outcomes.

  16. Intracellular proliferation of S. aureus in osteoblasts and effects of rifampicin and gentamicin on S. aureus intracellular proliferation and survival

    Directory of Open Access Journals (Sweden)

    W Mohamed

    2014-10-01

    Full Text Available Staphylococcus aureus is the most clinically relevant pathogen regarding implant-associated bone infection and its capability to invade osteoblasts is well known. The aim of this study was to investigate firstly whether S. aureus is not only able to invade but also to proliferate within osteoblasts, secondly to delineate the mechanism of invasion and thirdly to clarify whether rifampicin or gentamicin can inhibit intracellular proliferation and survival of S. aureus. The SAOS-2 osteoblast-like cell line and human primary osteoblasts were infected with S. aureus EDCC5055 and S. aureus Rosenbach 1884. Both S. aureus strains were able to invade efficiently and to proliferate within human osteoblasts. Immunofluorescence microscopy showed intracellular invasion of S. aureus and transmission electron microscopy images could demonstrate bacterial division as a sign of intracellular proliferation as well as cytosolic bacterial persistence. Cytochalasin D, the major actin depolymerisation agent, was able to significantly reduce S. aureus invasion, suggesting that invasion was enabled by promoting actin rearrangement at the cell surface. 7.5 μg/mL of rifampicin was able to inhibit bacterial survival in SAOS-2 cells with almost complete elimination of bacteria after 4 h. Gentamicin could also kill intracellular S. aureus in a dose-dependent manner, an effect that was significantly lower than that observed using rifampicin. In conclusion, S. aureus is not only able to invade but also to proliferate in osteoblasts. Invasion seems to be associated with actin rearrangement at the cell surface. Rifampicin is effective in intracellular eradication of S. aureus whereas gentamicin only poorly eliminates intracellularly replicating bacteria.

  17. Duplex Identification of Staphylococcus aureus by Aptamer and Gold Nanoparticles.

    Science.gov (United States)

    Chang, Tianjun; Wang, Libo; Zhao, Kexu; Ge, Yu; He, Meng; Li, Gang

    2016-06-01

    Staphylococcus aureus is the top common pathogen causing infections and food poisoning. Identification of S. aureus is crucial for the disease diagnosis and regulation of food hygiene. Herein, we report an aptamer-AuNPs based method for duplex identification of S. aureus. Using AuNPs as an indicator, SA23, an aptamer against S. aureus, can well identify its target from Escherichia coli, Listeria monocytogenes and Pseudomonas aeruginosa. Furthermore, we find citrate-coated AuNPs can strongly bind to S. aureus, but not bind to Salmonella enterica and Proteus mirabilis, which leads to different color changes in salt solution. This colorimetric response is capable of distinguishing S. aureus from S. enteritidis and P. mirabilis. Thus, using the aptasensor and AuNPs together, S. aureus can be accurately identified from the common pathogens. This duplex identification system is a promising platform for simple visual identification of S. aureus. Additionally, in the aptasensing process, bacteria are incubated with aptamers and then be removed before the aptamers adding to AuNPs, which may avoid the interactions between bacteria and AuNPs. This strategy can be potentially applied in principle to detect other cells by AuNPs-based aptasensors. PMID:27427591

  18. Lysostaphin in treatment of neonatal Staphylococcus aureus infection.

    Science.gov (United States)

    Oluola, Okunola; Kong, Lingkun; Fein, Mindy; Weisman, Leonard E

    2007-06-01

    This study describes lysostaphin's effect against methicillin-sensitive Staphylococcus aureus in suckling rats. Standard techniques determined minimal inhibitory and bactericidal concentrations, pharmacokinetics, and efficacy. The numbers of surviving rats after vancomycin, oxacillin, and lysostaphin treatment were comparable and were different from that of controls (P < 0.00001). Lysostaphin appears effective in the treatment of neonatal S. aureus infection. PMID:17420212

  19. Susceptibility of methicillin-resistant Staphylococcus aureus to lysostaphin.

    OpenAIRE

    Huber, M M; Huber, T. W.

    1989-01-01

    One hundred and eleven isolates of methicillin-resistant Staphylococcus aureus recovered from patients at the Olin E. Teague Veterans Center from March 1983 to April 1987 were as susceptible to lysis by lysostaphin as methicillin-susceptible S. aureus controls were.

  20. Mechanism of resistance to some cephalosporins in Staphylococcus aureus.

    OpenAIRE

    Kono, M; Sasatsu, M; O'Hara, K; Shiomi, Y.; HAYASAKA, T.

    1983-01-01

    The mechanism of resistance to some cephalosporins in Staphylococcus aureus strains was investigated with high-pressure liquid chromatography and nuclear magnetic resonance spectrometry. Drug inactivation by penicillinase was found to be the main mechanism of resistance to cefazolin, cephaloridine, and cephalothin in S. aureus.

  1. Activity of and Resistance to Moxifloxacin in Staphylococcus aureus

    OpenAIRE

    Ince, Dilek; Zhang, Xiamei; Hooper, David C.

    2003-01-01

    Moxifloxacin has enhanced potency against Staphylococcus aureus, lower propensity to select for resistant mutants, and higher bactericidal activity against highly resistant strains than ciprofloxacin. Despite similar activity against purified S. aureus topoisomerase IV and DNA gyrase, it selects for topoisomerase IV mutants, making topoisomerase IV the preferred target in vivo.

  2. Changing Trends in Resistance Pattern of Methicillin Resistant Staphylococcus aureus

    OpenAIRE

    Kali, Arunava; Stephen, Selvaraj; Umadevi, Sivaraman; Kumar, Shailesh; Joseph, Noyal Mariya; Srirangaraj, Sreenivasan

    2013-01-01

    Background: Methicillin resistance in Staphylococcus aureus is associated with multidrug resistance, an aggressive course, increased mortality and morbidity in both community and health care facilities. Monitoring of newly emerging and prevalent Methicillin Resistant Staphylococcus aureus (MRSA) strains for their resistance patterns to conventional as well as novel drugs, are essential for infection control.

  3. Effect of Mupirocin on Nasal Carriage of Staphylococcus Aureus

    NARCIS (Netherlands)

    M. Bulanda; M. Gruszka; B. Heczko

    1989-01-01

    textabstractMupirocin eliminates nasal carriage of staphylococcal aureus among medical and surgical personnel for periode varying from several weeks upto one year. In persons recolonized after therapy densites of S. aureus population in nares were much lower than in the same persons before therapy.

  4. Plasmid profiling of bacterial isolates from confined environments

    Science.gov (United States)

    van Houdt, Rob; Provoost, Ann; Coninx, Ilse; Leys, Natalie; Mergeay, Max

    Plasmid profiling of bacterial isolates from confined environments R. Van Houdt, I. Coninx, A. Provoost, N. Leys, and M. Mergeay Expertise group for Molecular and Cellular Biology, Institute for Environment, Health and Safety, Belgian Nuclear Research Centre (SCK•CEN), Boeretang 200, B-2400 Mol, Belgium. Human exploration of extreme and isolated hostile environments such as space requires special confined small volume habitats to protect and house the crew. However, human confinement in such small volume habitats has restrictions on waste disposal and personal hygiene and inevitably generates a particular community of microorganisms within the habitat. These microorganisms are mainly originating from the crew (skin, mucous membranes, upper respiratory tract, mouth, and gastrointestinal tract) but also include the residing environmental microorganisms. Earth-based confined habitats such as the Antarctic Research Station Concordia are used as test beds for long-duration spaceflights to study the physiologic and psychological adaptation to isolated environments. The dynamics of the environmental microbial population in such a test bed could render additional insights in assessing the potential health risks in long-duration space missions. Not only total bacterial contamination levels are important, but it is essential to identify also the predominant microbial taxa and their mobile genetic elements (MGE). These MGEs could be exchanged between bacteria by horizontal gene transfer and may alter the pathogenic potential since they often carry antibiotic resistance or more in general adaptation-enhancing traits. In this study several bacterial strains isolated in the Concordia research station were examined for their plasmid content. An optimized protocol for extraction of large plasmids showed the present of at least one plasmid in 50% of the strains. For all strains the minimal inhibitory concentration of a range of antibiotics was determined indicating resistance to

  5. GeneGuard: A modular plasmid system designed for biosafety.

    Science.gov (United States)

    Wright, Oliver; Delmans, Mihails; Stan, Guy-Bart; Ellis, Tom

    2015-03-20

    Synthetic biology applications in biosensing, bioremediation, and biomining envision the use of engineered microbes beyond a contained laboratory. Deployment of such microbes in the environment raises concerns of unchecked cellular proliferation or unwanted spread of synthetic genes. While antibiotic-resistant plasmids are the most utilized vectors for introducing synthetic genes into bacteria, they are also inherently insecure, acting naturally to propagate DNA from one cell to another. To introduce security into bacterial synthetic biology, we here took on the task of completely reformatting plasmids to be dependent on their intended host strain and inherently disadvantageous for others. Using conditional origins of replication, rich-media compatible auxotrophies, and toxin-antitoxin pairs we constructed a mutually dependent host-plasmid platform, called GeneGuard. In this, replication initiators for the R6K or ColE2-P9 origins are provided in trans by a specified host, whose essential thyA or dapA gene is translocated from a genomic to a plasmid location. This reciprocal arrangement is stable for at least 100 generations without antibiotic selection and is compatible for use in LB medium and soil. Toxin genes ζ or Kid are also employed in an auxiliary manner to make the vector disadvantageous for strains not expressing their antitoxins. These devices, in isolation and in concert, severely reduce unintentional plasmid propagation in E. coli and B. subtilis and do not disrupt the intended E. coli host's growth dynamics. Our GeneGuard system comprises several versions of modular cargo-ready vectors, along with their requisite genomic integration cassettes, and is demonstrated here as an efficient vector for heavy-metal biosensors. PMID:24847673

  6. Novel plasmid conferring kanamycin and tetracycline resistance in turkey-derived Campylobacter jejuni strain 11601MD

    Science.gov (United States)

    In Campylobacter spp., resistance to the antibiotics kanamycin and tetracycline is frequently associated with plasmid-borne genes. However, relatively few plasmids of Campylobacter jejuni have been fully characterized to date. A novel plasmid (p11601MD; 44,095 bp.) harboring tet(O) was identified in...

  7. Usefulness of plasmid profiles for differentiation of Shigella isolates in Bangladesh.

    OpenAIRE

    Tacket, C O; Shahid, N.; Huq, M I; Alim, A R; Cohen, M L

    1984-01-01

    We studied the plasmid profiles of 136 Shigella isolates in Bangladesh to determine whether plasmid profiles could be used for differentiation of strains for epidemiological studies. Many different plasmid patterns were observed within each species, indicating that many genetically different strains of Shigella are responsible for illness in Bangladesh.

  8. Mobilization of a Sym plasmid from a fast-growing cowpea Rhizobium strain.

    OpenAIRE

    Morrison, N.A.; Cen, Y H; Chen, H.C.; Plazinski, J; Ridge, R; Rolfe, B G

    1984-01-01

    A large Sym plasmid from a fast-growing cowpea Rhizobium species was made mobilizable by cointegration with plasmid pSUP1011, which carries the oriT region of RP4. This mobilizable Sym plasmid was transferred to a number of Rhizobium strains, in which nodulation and nitrogen fixation functions for symbiosis with plants of the cowpea group were expressed.

  9. A porcine model of haematogenous brain infectionwith staphylococcus aureus

    DEFF Research Database (Denmark)

    Astrup, Lærke Boye; Agerholm, Jørgen Steen; Nielsen, Ole Lerberg;

    2012-01-01

    A PORCINE MODEL OF HAEMATOGENOUS BRAIN INFECTION WITH STAPHYLOCOCCUS AUREUS Astrup Lærke1, Agerholm Jørgen1, Nielsen Ole1, Jensen Henrik1, Leifsson Páll1, Iburg Tine2. 1: Faculty of Health and Medical Sciences, University of Copenhagen, Denmark boye@life.ku.dk 2: National Veterinary Institute......, Uppsala, Sweden Introduction Staphylococcus aureus (S.aureus) is a common cause of sepsis and brain abscesses in man and a frequent cause of porcine pyaemia. Here we present a porcine model of haematogenous S. aureus-induced brain infection. Materials and Methods Four pigs had two intravenous catheters...... inserted surgically, one in a. carotis communis and one in v. jugularis externa. All pigs received 106 CFU/kg body weight S. aureus through the arterial catheter. Bacteria were either suspended in isotonic saline infused at constant flow for 60 minutes (two pigs) or given as a bolus injection of autologoue...

  10. Antimicrobial drug resistance ofStaphylococcus aureus in dairy products

    Institute of Scientific and Technical Information of China (English)

    Sasidharan S; Prema B; Yoga Latha L

    2011-01-01

    Objective:To evaluate the prevalence of multidrug resistantStaphylococcus aureus(S. aureus) in dairy products.Methods:Isolation and identification ofS. aureus were performed in3 dairy-based food products. The isolates were tested for their susceptibility to5 different common antimicrobial drugs.Results:Of50 samples examined,5 (10%) were contaminated with S. aureus. Subsequently, the5 isolates were subjected to antimicrobial resistance pattern using five antibiotic discs (methicillin, vancomycin, kanamycin, chloramphenicol and tetracycline). Sample 29 showed resistance to methicillin and vancomycin. Sample18 showed intermediate response to tetracycline. The other samples were susceptible to all the antibiotics tested.Conclusions:The results provide preliminary data on sources of food contamination which may act as vehicles for the transmission of antimicrobial-resistantStaphylococcus.Therefore, it enables us to develop preventive strategies to avoid the emergence of new strains of resistantS. aureus.

  11. Characterization of methicillin-resistant Staphylococcus aureus Sequence Type 398

    DEFF Research Database (Denmark)

    Christiansen, Mette Theilgaard

    Staphylococcus aureus is an opportunistic pathogen that colonizes the nares and skin surfaces of several animal species, including man. S. aureus can cause a wide variety of infections ranging from superficial soft tissue and skin infections to severe and deadly systemic infections. Traditionally S...... for LA-MRSA ST398 survival on porcine skin and nasal epithelium ex vivo were identified. These genes could represent targets for de-colonization, which could help prevent further spread and adaption of LA-MRSA ST398. Manuscript III describes the construction of the S. aureus VirulenceFinder database....... The database can be applied for identification of virulence genes in S. aureus using whole genome 5 sequence data. The S. aureus VirulenceFinder will be part of the tool package generated for the Centre for Genomic Epidemiology (CGE) (www.genomicepidemiology.org)....

  12. Where does a Staphylococcus aureus vaccine stand?

    Science.gov (United States)

    Fowler, V G; Proctor, R A

    2014-05-01

    In this review, we examine the current status of Staphylococcus aureus vaccine development and the prospects for future vaccines. Examination of the clinical trials to date show that murine models have not predicted success in humans for active or passive immunization. A key factor in the failure to develop a vaccine to prevent S. aureus infections comes from our relatively limited knowledge of human protective immunity. More recent reports on the elements of the human immune response to staphylococci are analysed. In addition, there is some controversy concerning the role of antibodies for protecting humans, and these data are reviewed. From a review of the current state of understanding of staphylococcal immunity, a working model is proposed. Some new work has provided some initial candidate biomarker(s) to predict outcomes of invasive infections and to predict the efficacy of antibiotic therapy in humans. We conclude by looking to the future through the perspective of lessons gleaned from the clinical vaccine trials. PMID:24476315

  13. Plasmid vectors for Xylella fastidiosa utilizing a toxin-antitoxin system for plasmid stability in the absence of antibiotic selection

    Science.gov (United States)

    The phytopathogen Xylella fastidiosa causes disease in a variety of important crop and landscape plants. Functional genetic studies have led to a broader understanding of virulence mechanisms used by this pathogen in the grapevine host. Plasmid shuttle vectors are important tools in studies of bacte...

  14. Efektivitas Ekstrak Daun Jambu Biji Buah Putih Terhadap Pertumbuhan Staphylococcus aureus Dari Abses Dan Staphylococcus aureus (ATCC® 29213™)

    OpenAIRE

    Sinurat, Jojor

    2016-01-01

    Daun jambu biji mengandung senyawa aktif seperti tanin, triterpenoid, flavonoid, saponin yang mempunyai efek antibakteri. Mekanisme tanin sebagai antibakteri dengan mengkerutkan dinding sel dan membran sel, inaktivasi enzim, inaktivasi fungsi materi genetik bakteri. Flavonoid merusak sel bakteri, denaturasi protein, inaktivasi enzim dan menyebabkan lisis. Triterpenoid dan saponin menghambat pertumbuhan Staphylococcus aureus dengan cara merusak struktur membran sel. Staphylococcus aureus adala...

  15. Factors contributing to the biofilm-deficient phenotype of Staphylococcus aureus sarA mutants.

    Directory of Open Access Journals (Sweden)

    Laura H Tsang

    Full Text Available Mutation of sarA in Staphylococcus aureus results in a reduced capacity to form a biofilm, but the mechanistic basis for this remains unknown. Previous transcriptional profiling experiments identified a number of genes that are differentially expressed both in a biofilm and in a sarA mutant. This included genes involved in acid tolerance and the production of nucleolytic and proteolytic exoenzymes. Based on this we generated mutations in alsSD, nuc and sspA in the S. aureus clinical isolate UAMS-1 and its isogenic sarA mutant and assessed the impact on biofilm formation. Because expression of alsSD was increased in a biofilm but decreased in a sarA mutant, we also generated a plasmid construct that allowed expression of alsSD in a sarA mutant. Mutation of alsSD limited biofilm formation, but not to the degree observed with the corresponding sarA mutant, and restoration of alsSD expression did not restore the ability to form a biofilm. In contrast, concomitant mutation of sarA and nuc significantly enhanced biofilm formation by comparison to the sarA mutant. Although mutation of sspA had no significant impact on the ability of a sarA mutant to form a biofilm, a combination of protease inhibitors (E-64, 1-10-phenanthroline, and dichloroisocoumarin that was shown to inhibit the production of multiple extracellular proteases without inhibiting growth was also shown to enhance the ability of a sarA mutant to form a biofilm. This effect was evident only when all three inhibitors were used concurrently. This suggests that the reduced capacity of a sarA mutant to form a biofilm involves extracellular proteases of all three classes (serine, cysteine and metalloproteases. Inclusion of protease inhibitors also enhanced biofilm formation in a sarA/nuc mutant, with the combined effect of mutating nuc and adding protease inhibitors resulting in a level of biofilm formation with the sarA mutant that approached that of the UAMS-1 parent strain. These results

  16. Role of the 85-Kilobase Plasmid and Plasmid-Encoded Virulence-Associated Protein A in Intracellular Survival and Virulence of Rhodococcus equi

    OpenAIRE

    Giguère, Steeve; Hondalus, Mary K.; Yager, Julie A.; Darrah, Patricia; Mosser, David M.; Prescott, John F.

    1999-01-01

    Rhodococcus equi is a facultative intracellular pathogen of macrophages and a cause of pneumonia in young horses (foals) and immunocompromised people. Isolates of R. equi from pneumonic foals typically contain large, 85- or 90-kb plasmids encoding a highly immunogenic virulence-associated protein (VapA). The objective of this study was to determine the role of the 85-kb plasmid and VapA in the intracellular survival and virulence of R. equi. Clinical isolates containing the plasmid and expres...

  17. The stb operon balances the requirements for vegetative stability and conjugative transfer of plasmid R388.

    Directory of Open Access Journals (Sweden)

    Catherine Guynet

    2011-05-01

    Full Text Available The conjugative plasmid R388 and a number of other plasmids carry an operon, stbABC, adjacent to the origin of conjugative transfer. We investigated the role of the stbA, stbB, and stbC genes. Deletion of stbA affected both conjugation and stability. It led to a 50-fold increase in R388 transfer frequency, as well as to high plasmid loss. In contrast, deletion of stbB abolished conjugation but provoked no change in plasmid stability. Deletion of stbC showed no effect, neither in conjugation nor in stability. Deletion of the entire stb operon had no effect on conjugation, which remained as in the wild-type plasmid, but led to a plasmid loss phenotype similar to that of the R388ΔstbA mutant. We concluded that StbA is required for plasmid stability and that StbA and StbB control conjugation. We next observed the intracellular positioning of R388 DNA molecules and showed that they localize as discrete foci evenly distributed in live Escherichia coli cells. Plasmid instability of the R388ΔΔstbA mutant correlated with aberrant localization of the plasmid DNA molecules as clusters, either at one cell pole, at both poles, or at the cell center. In contrast, plasmid molecules in the R388ΔΔstbB mutant were mostly excluded from the cell poles. Thus, results indicate that defects in both plasmid maintenance and transfer are a consequence of variations in the intracellular positioning of plasmid DNA. We propose that StbA and StbB constitute an atypical plasmid stabilization system that reconciles two modes of plasmid R388 physiology: a maintenance mode (replication and segregation and a propagation mode (conjugation. The consequences of this novel concept in plasmid physiology will be discussed.

  18. Plasmid-Chromosome Recombination of Irradiated Shuttle Vector DNA in African Green Monkey Kidney Cells.

    Science.gov (United States)

    Mudgett, John Stuart

    1987-09-01

    An autonomously replicating shuttle vector was used to investigate the enhancement of plasmid-chromosome recombination in mammalian host cells by ultraviolet light and gamma radiation. Sequences homologous to the shuttle vector were stably inserted into the genome of African Green Monkey kidney cells to act as the target substrate for these recombination events. The SV40- and pBR322-derived plasmid DNA was irradiated with various doses of radiation before transfection into the transformed mammalian host cells. The successful homologous transfer of the bacterial ampicillin resistance (amp^{rm r}) gene from the inserted sequences to replace a mutant amp^->=ne on the shuttle vector was identified by plasmid extraction and transformation into E. coli host cells. Ultraviolet light (UV) was found not to induce homologous plasmid-chromosome recombination, while gamma radiation increased the frequency of recombinant plasmids detected. The introduction of specific double -strand breaks in the plasmid or prolonging the time of plasmid residence in the mammalian host cells also enhanced plasmid-chromosome recombination. In contrast, plasmid mutagenesis was found to be increased by plasmid UV irradiation, but not to change with time. Plasmid survival, recombination, and mutagenesis were not affected by treating the mammalian host cells with UV light prior to plasmid transfection. The amp^{rm r} recombinant plasmid molecules analyzed were found to be mostly the result of nonconservative exchanges which appeared to involve both homologous and possibly nonhomologous interactions with the host chromosome. The observation that these recombinant structures were obtained from all of the plasmid alterations investigated suggests a common mechanistic origin for plasmid -chromosome recombination in these mammalian cells.

  19. Coupling between the Basic Replicon and the Kis-Kid Maintenance System of Plasmid R1: Modulation by Kis Antitoxin Levels and Involvement in Control of Plasmid Replication

    OpenAIRE

    Juan López-Villarejo; Damián Lobato-Márquez; Ramón Díaz-Orejas

    2015-01-01

    kis-kid, the auxiliary maintenance system of plasmid R1 and copB, the auxiliary copy number control gene of this plasmid, contribute to increase plasmid replication efficiency in cells with lower than average copy number. It is thought that Kis antitoxin levels decrease in these cells and that this acts as the switch that activates the Kid toxin; activated Kid toxin reduces copB-mRNA levels and this increases RepA levels that increases plasmid copy number. In support of this model we now repo...

  20. Coupling between the Basic Replicon and the Kis-Kid Maintenance System of Plasmid R1: Modulation by Kis Antitoxin Levels and Involvement in Control of Plasmid Replication

    Directory of Open Access Journals (Sweden)

    Juan López-Villarejo

    2015-02-01

    Full Text Available kis-kid, the auxiliary maintenance system of plasmid R1 and copB, the auxiliary copy number control gene of this plasmid, contribute to increase plasmid replication efficiency in cells with lower than average copy number. It is thought that Kis antitoxin levels decrease in these cells and that this acts as the switch that activates the Kid toxin; activated Kid toxin reduces copB-mRNA levels and this increases RepA levels that increases plasmid copy number. In support of this model we now report that: (i the Kis antitoxin levels do decrease in cells containing a mini-R1 plasmid carrying a repA mutation that reduces plasmid copy number; (ii kid-dependent replication rescue is abolished in cells in which the Kis antitoxin levels or the CopB levels are increased. Unexpectedly we found that this coordination significantly increases both the copy number of the repA mutant and of the wt mini-R1 plasmid. This indicates that the coordination between plasmid replication functions and kis-kid system contributes significantly to control plasmid R1 replication.

  1. Studies on the plasmid stability, plasmid copy number and endo(1, 3)(1, 4) b-glucanase production by free and alginate immobilised recombinant saccharomyces cerevisiae cells

    OpenAIRE

    Canavan, Peter D.

    1994-01-01

    A recombinant yeast strain, Saccharomyces cerevisiae DBY746, containing the plasmid pJG317, was grown in a variety of fermentation modes including batch, serial batch and chemostat culture incorporating a wide range of media types Plasmid pJG317 consists of a 2^-denved yeast episomal plasmid containing the gene which encodes for the bacterial enzyme endo (1,3)(1,4) P-glucanase. The concentration of enzyme produced appears to be proportional to the number of plasmid copies per cell. Specific e...

  2. Escherichia coli dnaT gene function is required for pBR322 plasmid replication but not for R1 plasmid replication.

    OpenAIRE

    Masai, H.; Arai, K.

    1989-01-01

    Plasmid pBR322 was unable to replicate in a temperature-sensitive dnaT1 strain at a nonpermissive temperature, whereas a pBR322-derived plasmid carrying the wild-type dnaT+ gene was able to replicate under the same conditions. In contrast to pBR322, plasmid R1 could replicate in the dnaT1 strain at a nonpermissive temperature. In keeping with this finding, in vitro replication of plasmid R1 did not require DnaT protein.

  3. Clone and construction of prokaryotic expression plasmid of MRSA-PBP2a%耐甲氧西林葡萄球菌PBP2a的质粒克隆与构建

    Institute of Scientific and Technical Information of China (English)

    聂红; 王云英; 陈维贤

    2011-01-01

    Objective To clone and construct the prokaryotic expression plasmid of penicillin binding protein 2a (PBP2a) of methicillin resistant Staphylococcus aureus (MRSA).Method According to the sequence of mecA gene published in GenBank, the target gene fragments were amplified by PCR, and then inserted into PET-32a plasmid.The recombined plasmids were identified by enzyme digestion and the inserted fragments were further confirmed by DNA sequencing.Result The corresponding prokaryotic expression plasmids of PBP2a had been successfully constructed.Conclusion The full length of PBP2a and its transpeptidase domain expressing plasmid are successfully constructed, which establishes the foundation for its purification, identification and application.%目的 克隆并构建耐甲氧西林金黄色葡萄球菌(MRSA)青霉素结合蛋白2a(PBP2a)全长及转肽酶区的原核表达质粒.方法 登录基因文库查找获得mecA基因的编码序列,应用PCR技术扩增获得DNA片段,将此基因片段插入PET-32a载体,同时酶切鉴定阳性克隆,DNA序列测定验证序列正确性.结果 PCR扩增获得了mecA基因全长及转肽酶区DNA片段,成功插入到原核表达载体PET32a,双酶切鉴定及DNA序列测定证实插入片段正确.结论 成功构建了PBP2a全长及转肽酶区片段表达质粒,为该蛋白的纯化表达和疫苗研究奠定了基础.

  4. Enterococcus faecalis hemolysin-bacteriocin plasmids belong to the same incompatibility group.

    OpenAIRE

    Colmar, I; Horaud, T

    1987-01-01

    Plasmid pair coexistence was studied both among nine Enterococcus faecalis hemolysin-bacteriocin (Hly-Bcn) plasmids, including pJH2, pAD1, pAM gamma 1, and pIP964, and between pIP964 and five R plasmids. Some of the Hly-Bcn plasmids used were derivatives encoding resistance to erythromycin or tetracycline. The Hly-Bcn plasmids were incompatible with each other; 40 to 100% displacement was observed bilaterally for eight pairs and unilaterally for one pair. In contrast, pIP964 stably coexisted ...

  5. Structural and genetic analyses of a par locus that regulates plasmid partition in Bacillus subtilis.

    OpenAIRE

    Chang, S.; Chang, S Y; Gray, O

    1987-01-01

    The Bacillus plasmid pLS11 partitions faithfully during cell division. Using a partition-deficient plasmid vector, we randomly cloned DNA fragments of plasmid pLS11 and identified the locus that regulates plasmid partition (par) by cis complementation in Bacillus subtilis. The cloned par gene conferred upon the vector plasmid a high degree of segregational stability. The par locus was mapped to a 167-base-pair segment on pLS11, and its nucleotide sequence was determined. The cloned par fragme...

  6. Remarkable stability of an instability-prone lentiviral vector plasmid in Escherichia coli Stbl3

    OpenAIRE

    Al-Allaf, Faisal A.; Tolmachov, Oleg E.; Zambetti, Lia Paola; Tchetchelnitski, Viktoria; Mehmet, Huseyin

    2012-01-01

    Large-scale production of plasmid DNA to prepare therapeutic gene vectors or DNA-based vaccines requires a suitable bacterial host, which can stably maintain the plasmid DNA during industrial cultivation. Plasmid loss during bacterial cell divisions and structural changes in the plasmid DNA can dramatically reduce the yield of the desired recombinant plasmid DNA. While generating an HIV-based gene vector containing a bicistronic expression cassette 5′-Olig2cDNA-IRES-dsRed2-3′, we encountered ...

  7. The incC Sequence Is Required for R27 Plasmid Stability

    OpenAIRE

    Tassinari, Eleonora; Aznar, Sonia; Urcola, Imanol; Prieto, Alejandro; Hüttener, Mário; Juárez, Antonio

    2016-01-01

    IncHI plasmids account for multiple antimicrobial resistance in Salmonella and other enterobacterial genera. These plasmids are generally very stable in their bacterial hosts. R27 is the archetype of IncHI1 plasmids. A high percentage of the R27-encoded open reading frames (ORFs) (66.7%) do not show similarity to any known ORFs. We performed a deletion analysis of all non-essential R27 DNA sequences to search for hitherto non-identified plasmid functions that might be required for plasmid sta...

  8. [Plasmids of streptomycetes strains isolated from soils of Ukraine with different anthropogenic loading].

    Science.gov (United States)

    Luk'ianchuk, V V; Polishchuk, L V; Matseliukh, B P

    2010-01-01

    Screening of plasmid DNA was carried out among 94 streptomycetes cultures which were isolated from the samples of Ukrainian soils with different anthropogenic contamination. Seventeen streptomycetes strains containing plasmid DNA were found. It is established that some cultures contain more than one plasmid (Streptomyces sp.M15, S.sp.T8, S.sp.T19). Depending on a molecular sizes the found plasmids were divided in 2 groups: 3 kb-15 kb, and 30 kb-70 kb. Research of a few morphological and physiological properties of plasmid strains of streptomycetes was carried out. The paper is presented in Ukrainian. PMID:21117293

  9. recE4-Independent Recombination Between Homologous Deoxyribonucleic Acid Segments of Bacillus subtilis Plasmids

    OpenAIRE

    Tanaka, T

    1980-01-01

    A plasmid (pLS104) carrying a tandem repetition of the leu region of the Bacillus subtilis chromosome arose spontaneously from pLS103, which carried a single copy of the leu region. Plasmid preparations from strains harboring pLS104 also contained the original plasmid, pLS103, and, in some preparations, plasmids carrying three or four repetitions of the leu region. These plasmids were shown to be generated by recombination between homologous deoxyribonucleic acid (DNA) segments in the tandeml...

  10. An Improved Method for Including Upper Size Range Plasmids in Metamobilomes

    DEFF Research Database (Denmark)

    Norman, Anders; Riber, Leise; Luo, Wenting; Li, Li Li; Hansen, Lars Hestbjerg; Sørensen, Søren Johannes

    2014-01-01

    Two recently developed isolation methods have shown promise when recovering pure community plasmid DNA (metamobilomes/plasmidomes), which is useful in conducting culture-independent investigations into plasmid ecology. However, both methods employ multiple displacement amplification (MDA) to ensure...... suitable quantities of plasmid DNA for high-throughput sequencing. This study demonstrates that MDA greatly favors smaller circular DNA elements (<10 Kbp), which, in turn, leads to stark underrepresentation of upper size range plasmids (>10 Kbp). Throughout the study, we used two model plasmids, a 4.4 Kbp...

  11. Tumor induction by Agrobacterium rhizogenes involves the transfer of plasmid DNA to the plant genome

    OpenAIRE

    White, Frank F; Ghidossi, Gina; Gordon, Milton P.; Nester, Eugene W.

    1982-01-01

    The DNA from tumors of Nicotiana glauca initiated by strains of Agrobacterium rhizogenes was shown to contain sequences that are homologous to the root-inducing (Ri) plasmid of the bacterium. Two independently established tumor lines contained a similar portion of the Ri-plasmid. The Ri-plasmid also hybridized to DNA fragments from uninfected N. glauca. A cosmid clone of the Ri-plasmid encompassing the region containing the Ri-plasmid sequences that are stably transferred to the plant also hy...

  12. The molecular changing mechanism of Ampicillin-Sulbactam resistant Staphylococcus aureus towards Methicillin resistant Staphylococcus aureus

    Directory of Open Access Journals (Sweden)

    Mieke Hemiawati Satari

    2005-12-01

    Full Text Available The aim of this study was to determine the molecular changing of S.aureus, which is resistant to Ampicillin-Sulbactam and then become resistant to Methicillin as a result of improper dosage. The study was conducted by isolating Ampicillin-Sulbactam resistant and Methicillin Resistant S.aureus (MRSA, afterwards an amplification process was performed by PCR (Polymerase Chain Reaction. to isolate the betalactamase enzyme regulator and PBP 2a genes. The result of this research showed that there were a deletion of few amino acids from the regulator gene, and a suspicion that the DNA sequence had been substituted from PBP 2 gene into PBP 2a (gen mec. This process had formed MRSA.

  13. Indole and 7-benzyloxyindole attenuate the virulence of Staphylococcus aureus.

    Science.gov (United States)

    Lee, Jin-Hyung; Cho, Hyun Seob; Kim, Younghoon; Kim, Jung-Ae; Banskota, Suhrid; Cho, Moo Hwan; Lee, Jintae

    2013-05-01

    Human pathogens can readily develop drug resistance due to the long-term use of antibiotics that mostly inhibit bacterial growth. Unlike antibiotics, antivirulence compounds diminish bacterial virulence without affecting cell viability and thus, may not lead to drug resistance. Staphylococcus aureus is a major agent of nosocomial infections and produces diverse virulence factors, such as the yellow carotenoid staphyloxanthin, which promotes resistance to reactive oxygen species (ROS) and the host immune system. To identify novel antivirulence compounds, bacterial signal indole present in animal gut and diverse indole derivatives were investigated with respect to reducing staphyloxanthin production and the hemolytic activity of S. aureus. Treatment with indole or its derivative 7-benzyloxyindole (7BOI) caused S. aureus to become colorless and inhibited its hemolytic ability without affecting bacterial growth. As a result, S. aureus was more easily killed by hydrogen peroxide (H₂O₂) and by human whole blood in the presence of indole or 7BOI. In addition, 7BOI attenuated S. aureus virulence in an in vivo model of nematode Caenorhabditis elegans, which is readily infected and killed by S. aureus. Transcriptional analyses showed that both indole and 7BOI repressed the expressions of several virulence genes such as α-hemolysin gene hla, enterotoxin seb, and the protease genes splA and sspA and modulated the expressions of the important regulatory genes agrA and sarA. These findings show that indole derivatives are potential candidates for use in antivirulence strategies against persistent S. aureus infection. PMID:23318836

  14. The changing epidemiology of Staphylococcus aureus bloodstream infection

    DEFF Research Database (Denmark)

    Galbraith, J.C.; Valiquette, G.; Kennedy, K.J.;

    2013-01-01

    Clin Microbiol Infect ABSTRACT: Although the epidemiology of Staphylococcus aureus bloodstream infection (BSI) has been changing, international comparisons are lacking. We sought to determine the incidence of S. aureus BSI and assess trends over time and by region. Population-based surveillance...... episodes of S. aureus BSI were identified. The overall annual incidence rate for S. aureus BSI was 26.1 per 100 000 population, and those for methicillin-sensitive S. aureus (MSSA) and methicillin-resistant S. aureus (MRSA) were 24.2 and 1.9 per 100 000, respectively. Although the overall incidence...... of community-onset MSSA BSI (15.0 per 100 000) was relatively similar across regions, the incidence rates of hospital-onset MSSA (9.2 per 100 000), community-onset MRSA (1.0 per 100 000) and hospital-onset MRSA (0.8 per 100 000) BSI varied substantially. Whereas the overall incidence of S. aureus BSI did...

  15. STABILITY OF PLASMIDS IN 5 STRAINS OF SALMONELLA MAINTAINED IN STAB CULTURE AT DIFFERENT TEMPERATURES

    DEFF Research Database (Denmark)

    Olsen, J. E.; Brown, D. J.; Baggesen, Dorte Lau;

    1994-01-01

    Four strains of Salmonella berta and one of Salm. enteritidis were stored as stab cultures in sugar-free agar at 5 degrees, 22 degrees and 30 degrees C and in 15% glycerol at -80 degrees C. The stability of the plasmid profiles in each of the strains was monitored over a period of 2.5 years....... Plasmid profiles were stable in all strains stored at -80 degrees C, and only six of 450 colonies examined from strains kept in sugar-free agar at 5 degrees C had lost plasmid molecules. Seventy of 440 colonies from stab cultures that were kept at 22 degrees C, and 71 of 440 colonies at 30 degrees C...... showed changed plasmid profiles. The total number of plasmids lost increased with time, and occasionally, more than one plasmid molecule was lost in the same strain. The virulence associated plasmid of Salm. enteritidis was remarkably stable as it was maintained in all colonies examined at all...

  16. Differences in the stability of the plasmids of Yersinia pestis cultures in vitro: impact on virulence

    Directory of Open Access Journals (Sweden)

    TC Leal-Balbino

    2004-11-01

    Full Text Available Plasmid and chromosomal genes encode determinants of virulence for Yersinia pestis, the causative agent of plague. However, in vitro, Y. pestis genome is very plastic and several changes have been described. To evaluate the alterations in the plasmid content of the cultures in vitro and the impact of the alterations to their pathogenicity, three Y. pestis isolates were submitted to serial subculture, analysis of the plasmid content, and testing for the presence of characteristic genes in each plasmid of colonies selected after subculture. Different results were obtained with each strain. The plasmid content of one of them was shown to be stable; no apparent alteration was produced through 32 subcultures. In the other two strains, several alterations were observed. LD50 in mice of the parental strains and the derived cultures with different plasmid content were compared. No changes in the virulence plasmid content could be specifically correlated with changes in the LD50.

  17. Novel assay to measure the plasmid mobilizing potential of mixed microbial communities

    DEFF Research Database (Denmark)

    Klümper, Uli; Droumpali, Ariadni; Dechesne, Arnaud;

    2014-01-01

    Mobilizable plasmids lack necessary genes for complete conjugation and are therefore non-self-transmissible. Instead, they rely on the conjugation system of conjugal plasmids to be horizontally transferred to new recipients. While community permissiveness, the fraction of a mixed microbial...... community that can receive self-transmissible conjugal plasmids, has been studied, the intrinsic ability of a community to mobilize plasmids that lack conjugation systems is unexplored. Here, we present a novel framework and experimental method to estimate the mobilization potential of mixed communities. We...... compare the transfer frequency of a mobilizable plasmid to that of a mobilizing and conjugal plasmid measured for a model strain and for the assayed community. With Pseudomonas putida carrying the gfp-tagged mobilizable RSF1010 plasmid as donor strain, we conducted solid surface mating experiments...

  18. Molecular epidemiological study on tetracycline resistance R plasmids in enterohaemorrhagic Escherichia coli O157:H7.

    Science.gov (United States)

    Makino, S; Asakura, H; Obayashi, T; Shirahata, T; Ikeda, T; Takeshi, K

    1999-08-01

    Restriction patterns obtained with EcoRI and Southern hybridization were used for the differentiation of tetracycline-resistant (Tet(r)) R plasmids in enterobaemorrhagic Escherichia coli (EHEC) O157:H7 isolates from a mass outbreak at a kindergarten in Obihiro-City, Hokkaido, Japan, 1996. Two kinds of Tet(r) R plasmids of 50 and 95 kb were detected. The 50-kb plasmids were identical to each other, while the 93-kb plasmids were of three types that were very similar to each other. The tet genes of both 50- and 95-kb R plasmids were 100% identical to the tet gene of pSC101 and all plasmids hybridized to a probe for tet. Because food-origin O157 strains were sensitive to tetracycline, we concluded that such Tet(r) R-plasmids might transfer to drug-sensitive O157 strains in the infected individuals. PMID:10487638

  19. Molecular epidemiological study on tetracycline resistance R plasmids in enterohaemorrhagic Escherichia coli O157:H7.

    Science.gov (United States)

    Makino, S.; Asakura, H.; Obayashi, T.; Shirahata, T.; Ikeda, T.; Takeshi, K.

    1999-01-01

    Restriction patterns obtained with EcoRI and Southern hybridization were used for the differentiation of tetracycline-resistant (Tet(r)) R plasmids in enterobaemorrhagic Escherichia coli (EHEC) O157:H7 isolates from a mass outbreak at a kindergarten in Obihiro-City, Hokkaido, Japan, 1996. Two kinds of Tet(r) R plasmids of 50 and 95 kb were detected. The 50-kb plasmids were identical to each other, while the 93-kb plasmids were of three types that were very similar to each other. The tet genes of both 50- and 95-kb R plasmids were 100% identical to the tet gene of pSC101 and all plasmids hybridized to a probe for tet. Because food-origin O157 strains were sensitive to tetracycline, we concluded that such Tet(r) R-plasmids might transfer to drug-sensitive O157 strains in the infected individuals. PMID:10487638

  20. Staphylococcus aureus: portadores entre manipuladores de alimentos Staphylococcus aureus: food handler carriers

    Directory of Open Access Journals (Sweden)

    Maria Stella Gonçalves Raddi

    1988-02-01

    Full Text Available Foram colhidas amostras de mãos e fossas nasais de 48 manipuladores de alimentos das principais casas comerciais da cidade de Araraquara, Estado de São Paulo (Brasil, e de 20 estudantes universitários. Dentre os indivíduos foram encontrados 44,1% e 34,8% que portavam Staphylococcus aureus em fossas nasais e mãos, respectivamente. Observou-se predomínio de fagotipos dos grupos I e III. Dos 12 portadores do microrganismo, concomitantemente em mãos e fossas nasais, 75,0% apresentaram cepas com vínculo epidemiológico. Os achados mostram o risco potencial representado pelas mãos nas intoxicações alimentares.Material was collected from the hands and nasal passages of forty-eight food handlers and twenty college students of Araraquara (S. Paulo State, Brazil and analized in order to evaluate the carrier function with regard to Staphylococcus aureus. The organism discovered in both samples of nine out of the twelve volunteers were of the same S. aureus phage types. The incidence of carriage on the hands was much greater in the handlers' group. These findings demonstrate the potential risk represented by hands in the transmission of food poisoning.

  1. Methicillin-resistant Staphylococcus aureus transmission

    DEFF Research Database (Denmark)

    Andersen, Leif Percival; Nielsen, Xiaohui

    2015-01-01

    increase the risk of contaminating hands, arms and the front of the uniform. Hand hygiene is therefore essential, but the use of protection gowns with long sleeves is also important in order to prevent transmission of MRSA. After culture of MRSA and implementation of specific precautions to prevent......INTRODUCTION: Even though methicillin-resistant Staphylococcus aureus (MRSA) is a common cause of nosocomial infections, it may often be difficult to evaluate the exact route of transmission. METHODS: In this study, we describe four cases of nosocomial transmission of MRSA in a hospital with a low...... transmission of MRSA, no further transmissions were observed. FUNDING: not relevant. TRIAL REGISTRATION: The data in this study are included in the routine surveillance of MRSA at Rigshospitalet and do not form part of a trial....

  2. Replisome Assembly at Bacterial Chromosomes and Iteron Plasmids.

    Science.gov (United States)

    Wegrzyn, Katarzyna E; Gross, Marta; Uciechowska, Urszula; Konieczny, Igor

    2016-01-01

    The proper initiation and occurrence of DNA synthesis depends on the formation and rearrangements of nucleoprotein complexes within the origin of DNA replication. In this review article, we present the current knowledge on the molecular mechanism of replication complex assembly at the origin of bacterial chromosome and plasmid replicon containing direct repeats (iterons) within the origin sequence. We describe recent findings on chromosomal and plasmid replication initiators, DnaA and Rep proteins, respectively, and their sequence-specific interactions with double- and single-stranded DNA. Also, we discuss the current understanding of the activities of DnaA and Rep proteins required for replisome assembly that is fundamental to the duplication and stability of genetic information in bacterial cells. PMID:27563644

  3. Polymerase chain reaction-based gene removal from plasmids

    Directory of Open Access Journals (Sweden)

    Vishnu Vardhan Krishnamurthy

    2015-09-01

    Full Text Available This data article contains supplementary figures and methods to the research article entitled, “Multiplex gene removal by two-step polymerase chain reactions” (Krishnamurthy et al., Anal. Biochem., 2015, doi:http://dx.doi.org/10.1016/j.ab.2015.03.033, which presents a restriction-enzyme free method to remove multiple DNA segments from plasmids. Restriction-free cloning methods have dramatically improved the flexibility and speed of genetic manipulation compared to conventional assays based on restriction enzyme digestion (Lale and Valla, 2014. DNA Cloning and Assembly Methods, vol. 1116. Here, we show the basic scheme and characterize the success rate for single and multiplex gene removal from plasmids. In addition, we optimize experimental conditions, including the amount of template, multiple primers mixing, and buffers for DpnI treatment, used in the one-pot reaction for multiplex gene removal.

  4. Plasmid DNA damage induced by helium atmospheric pressure plasma jet

    Science.gov (United States)

    Han, Xu; Cantrell, William A.; Escobar, Erika E.; Ptasinska, Sylwia

    2014-03-01

    A helium atmospheric pressure plasma jet (APPJ) is applied to induce damage to aqueous plasmid DNA. The resulting fractions of the DNA conformers, which indicate intact molecules or DNA with single- or double-strand breaks, are determined using agarose gel electrophoresis. The DNA strand breaks increase with a decrease in the distance between the APPJ and DNA samples under two working conditions of the plasma source with different parameters of applied electric pulses. The damage level induced in the plasmid DNA is also enhanced with increased plasma irradiation time. The reactive species generated in the APPJ are characterized by optical emission spectra, and their roles in possible DNA damage processes occurring in an aqueous environment are also discussed.

  5. Transfer of the lambdadv plasmid to new bacterial hosts

    International Nuclear Information System (INIS)

    Lambda dv, which was derived from bacteriophage lambda, replicates autonomously as a plasmid in Escherichia coli and consists of only the immunity region (imm/sup lambda/) and DNA replication genes (O, P) of the ancestral phage. Addition phages (lambda imm21--lambda dv) carry the lambda dv fragment inserted as a tandem duplication in their genome (sequence A imm21 O P imm/sup lambda/ O P R) are formed as recombinants after lambda imm21 infection of strains carrying lambda dv. Addition phages were used to transfer lambda dv to new bacterial hosts. Lambda dv transfer by excision of the lambda dv segment from the addition phage genome requires a bacterial Rec or a phage Red recombination system. Successful transfer is stimulated by uv irradiation of the addition phage before infection. Some properties of the newly transferred lambda dv plasmids are described. (U.S.)

  6. Plasmids and packaging cell lines for use in phage display

    Science.gov (United States)

    Bradbury, Andrew M.

    2012-07-24

    The invention relates to a novel phagemid display system for packaging phagemid DNA into phagemid particles which completely avoids the use of helper phage. The system of the invention incorporates the use of bacterial packaging cell lines which have been transformed with helper plasmids containing all required phage proteins but not the packaging signals. The absence of packaging signals in these helper plasmids prevents their DNA from being packaged in the bacterial cell, which provides a number of significant advantages over the use of both standard and modified helper phage. Packaged phagemids expressing a protein or peptide of interest, in fusion with a phage coat protein such as g3p, are generated simply by transfecting phagemid into the packaging cell line.

  7. Bovine Staphylococcus aureus: Subtyping, evolution, and zoonotic transfer.

    Science.gov (United States)

    Boss, R; Cosandey, A; Luini, M; Artursson, K; Bardiau, M; Breitenwieser, F; Hehenberger, E; Lam, Th; Mansfeld, M; Michel, A; Mösslacher, G; Naskova, J; Nelson, S; Podpečan, O; Raemy, A; Ryan, E; Salat, O; Zangerl, P; Steiner, A; Graber, H U

    2016-01-01

    Staphylococcus aureus is globally one of the most important pathogens causing contagious mastitis in cattle. Previous studies using ribosomal spacer (RS)-PCR, however, demonstrated in Swiss cows that Staph. aureus isolated from bovine intramammary infections are genetically heterogeneous, with Staph. aureus genotype B (GTB) and GTC being the most prominent genotypes. Furthermore, Staph. aureus GTB was found to be contagious, whereas Staph. aureus GTC and all the remaining genotypes were involved in individual cow disease. In addition to RS-PCR, other methods for subtyping Staph. aureus are known, including spa typing and multilocus sequence typing (MLST). They are based on sequencing the spa and various housekeeping genes, respectively. The aim of the present study was to compare the 3 analytic methods using 456 strains of Staph. aureus isolated from milk of bovine intramammary infections and bulk tanks obtained from 12 European countries. Furthermore, the phylogeny of animal Staph. aureus was inferred and the zoonotic transfer of Staph. aureus between cattle and humans was studied. The analyzed strains could be grouped into 6 genotypic clusters, with CLB, CLC, and CLR being the most prominent ones. Comparing the 3 subtyping methods, RS-PCR showed the highest resolution, followed by spa typing and MLST. We found associations among the methods but in many cases they were unsatisfactory except for CLB and CLC. Cluster CLB was positive for clonal complex (CC)8 in 99% of the cases and typically positive for t2953; it is the cattle-adapted form of CC8. Cluster CLC was always positive for tbl 2645 and typically positive for CC705. For CLR and the remaining subtypes, links among the 3 methods were generally poor. Bovine Staph. aureus is highly clonal and a few clones predominate. Animal Staph. aureus always evolve from human strains, such that every human strain may be the ancestor of a novel animal-adapted strain. The zoonotic transfer of IMI- and milk-associated strains

  8. Identification of the ClpX Regulon in Staphylococcus aureus

    DEFF Research Database (Denmark)

    Jelsbak, Lotte; Thomsen, Line Elnif; Ingmer, Hanne;

    Staphyloccous aureus is a major human pathogen capable of causing a wide spectrum of infections ranging from superficial wound infections to life-threatening endocarditis and toxic shock syndrome. Essential for S. aureus virulence is a large number of cell-surface-associated proteins and secreted...... we show here that almost 400 genes (15%) are influenced by the clpX deletion. Furthermore, ClpX not only regulates many virulence factors, but rather serves as a global regulator of central functions for S. aureus lifestyle and pathogenicity....

  9. Proton-induced direct and indirect damage of plasmid DNA

    Czech Academy of Sciences Publication Activity Database

    Vyšín, Luděk; Pachnerová Brabcová, Kateřina; Štěpán, V.; Moretto-Capelle, P.; Bugler, B.; Legube, G.; Cafarelli, P.; Casta, R.; Champeaux, J. P.; Sence, M.; Vlk, M.; Wagner, Richard; Štursa, Jan; Zach, Václav; Incerti, S.; Juha, Libor; Davídková, Marie

    2015-01-01

    Roč. 54, č. 3 (2015), s. 343-352. ISSN 0301-634X R&D Projects: GA ČR GA13-28721S; GA MŠk LD12008; GA MŠk LM2011019 Institutional support: RVO:68378271 ; RVO:61389005 Keywords : proton radiation * DNA plasmid * direct and indirect effects * clustered damage * repair enzymes Subject RIV: BO - Biophysics Impact factor: 1.528, year: 2014

  10. Recombination between short direct repeats in Streptomyces lavendulae plasmid DNA.

    OpenAIRE

    Nakano, M M; Ogawara, H; Sekiya, T

    1984-01-01

    Streptomyces lavendulae S985 carried two plasmids, pSL1 and pSL2. pSL2 contained all of the pSL1 sequences plus a tandem duplication of 900 base pairs from a region of pSL1. Sequence analysis of the duplication junction suggested that the duplication occurred by recombination between short direct repeats of as little as 5 base pairs.

  11. Characterization of Toxin Plasmids in Clostridium perfringens Type C Isolates▿

    OpenAIRE

    Gurjar, Abhijit; Li, Jihong; McClane, Bruce A.

    2010-01-01

    Clostridium perfringens type C isolates cause enteritis necroticans in humans or necrotizing enteritis and enterotoxemia in domestic animals. Type C isolates always produce alpha toxin and beta toxin but often produce additional toxins, e.g., beta2 toxin or enterotoxin. Since plasmid carriage of toxin-encoding genes has not been systematically investigated for type C isolates, the current study used Southern blot hybridization of pulsed-field gels to test whether several toxin genes are plasm...

  12. Plasmid addiction systems: perspectives and applications in biotechnology

    OpenAIRE

    Kroll, Jens; Klinter, Stefan; Schneider,Cornelia; Voß, Isabella; Steinbüchel, Alexander

    2010-01-01

    Summary Biotechnical production processes often operate with plasmid‐based expression systems in well‐established prokaryotic and eukaryotic hosts such as Escherichia coli or Saccharomyces cerevisiae, respectively. Genetically engineered organisms produce important chemicals, biopolymers, biofuels and high‐value proteins like insulin. In those bioprocesses plasmids in recombinant hosts have an essential impact on productivity. Plasmid‐free cells lead to losses in the entire product recovery a...

  13. Deep sequencing reveals complex spurious transcription from transiently transfected plasmids

    Czech Academy of Sciences Publication Activity Database

    Nejepínská, Jana; Malík, Radek; Moravec, Martin

    2012-01-01

    Roč. 7, č. 8 (2012), e43283. E-ISSN 1932-6203 R&D Projects: GA ČR GA204/09/0085 Grant ostatní: EMBO(XE) 0001488 Institutional research plan: CEZ:AV0Z50520514 Institutional support: RVO:68378050 Keywords : transient plasmid transfection * deep sequencing Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 3.730, year: 2012

  14. Conjugative Plasmid Transfer in Gram-Positive Bacteria

    OpenAIRE

    Grohmann, Elisabeth; Muth, Günther; Espinosa, Manuel

    2003-01-01

    Conjugative transfer of bacterial plasmids is the most efficient way of horizontal gene spread, and it is therefore considered one of the major reasons for the increase in the number of bacteria exhibiting multiple-antibiotic resistance. Thus, conjugation and spread of antibiotic resistance represents a severe problem in antibiotic treatment, especially of immunosuppressed patients and in intensive care units. While conjugation in gram-negative bacteria has been studied in great detail over t...

  15. Plasmid Mediated Antibiotic Resistance in Isolated Bacteria From Burned Patients

    OpenAIRE

    Beige, Fahimeh; Baseri Salehi, Majid; Bahador, Nima; Mobasherzadeh, Sina

    2014-01-01

    Background: Nowadays, the treatment of burned patients is difficult because of the high frequency of infection with antibiotic resistance bacteria. Objectives: This study was conducted to evaluate the level of antibiotic resistance in Gram-negative bacteria and its relation with the existence of plasmid. Materials and Methods: The samples were collected from two hundred twenty hospitalized burned patients in Isfahan burn hospital during a three-month period (March 2012 to June 2012). The samp...

  16. Efficient transformation of Bacillus thuringiensis requires nonmethylated plasmid DNA.

    OpenAIRE

    Macaluso, A; Mettus, A M

    1991-01-01

    The transformation efficiency of Bacillus thuringiensis depends upon the source of plasmid DNA. DNA isolated from B. thuringiensis, Bacillus megaterium, or a Dam- Dcm- Escherichia coli strain efficiently transformed several B. thuringiensis strains, B. thuringiensis strains were grouped according to which B. thuringiensis backgrounds were suitable sources of DNA for transformation of other B. thuringiensis strains, suggesting that B. thuringiensis strains differ in DNA modification and restri...

  17. Reconstruction of the yeast 2 micron plasmid partitioning mechanism.

    OpenAIRE

    Dobson, M J; Yull, F E; Molina, M.; Kingsman, S M; Kingsman, A J

    1988-01-01

    The effect of the yeast 2 micron circle encoded REP1 and REP2 gene products on plasmid partitioning and copy number control was analyzed by removing the open reading frames from their normal sequence context and transcriptional control regions and directing their expression using heterologous promoters in [cir0] host strains. Both the REP1 and REP2 gene products are directly required at appropriate levels of expression to reconstitute the 2 microns circle partitioning system in conjunction wi...

  18. Enhancer-activated plasmid transcription complexes contain constrained supercoiling.

    OpenAIRE

    Bonilla, P J; Freytag, S O; Lutter, L C

    1991-01-01

    It has been proposed that transcriptionally active chromatin contains totally unconstrained supercoiling. The results of recent studies have raised the possibility that this topological state is the property of highly transcribed genes. Since the transcription rate of RNA polymerase II genes can be dramatically increased by the presence of an enhancer, we have determined if the transcription complex of an enhancer-activated plasmid contains totally unconstrained supercoils. Following transfec...

  19. Spaceflight Effects on Genetics and Plasmids of Streptomycetes

    Science.gov (United States)

    Voeikova, T. A.; Emelyanova, L. K.; Tyaglov, B. V.; Novikova, L. M.; Goins, T. L.; Pyle, B. H.

    2008-06-01

    In 2007, experiments with streptomycetes were conducted during a 12-day flight of the Russian Foton-M3 spacecraft. The flight (F), synchronous control (SC) and laboratory control (LC) specimens were kept at 30°C. The objective of the experiments was to study spaceflight effects on the streptomycetes growth, differentiation, pigmentation, enzyme formation, genetic stability of plasmid and crossing between strains. It was found that the frequency of strain Streptomyces lividans segregation, the enzyme synthesis, pigmentation, and the level of sporulation were higher in F than in SC organisms. The study of pIJ702 plasmid inheritance in S. lividans showed that the frequency of plasmid loss in F and LC was similar and lower than that in SC specimens. The study of melanin synthesis in S. lividans (pIJ702) strain demonstrated decreased melanin specific yield and increased biomass accumulation in F microorganisms. HPTLC analysis of melanin showed that the number, molecular mass and the percentage of fractions were similar in SC and LC but different in F organisms. The study of spaceflight effects on genetic recombination in crosses between Streptomyces coelicolor A3(2) auxotrophic mutants showed that the frequency of various recombinant classes in F specimens differed from that in SC and LC. The frequency of a distal donor marker entry to the recipient in F was higher than in SC and LC.

  20. Two domains at the origin are required for replication and maintenance of broad-host-range plasmid R1162.

    OpenAIRE

    Kim, Y.J.; Lin, L. S.; Meyer, R. J.

    1987-01-01

    Two domains at the replicative origin of broad-host-range plasmid R1162 are required in cis for plasmid maintenance in Escherichia coli and for plasmid DNA replication in cell extracts. Increasing the distance between the domains reduces replication in vitro, without substantially changing plasmid DNA content or stability in vivo.

  1. Comparative Physical and Genetic Maps of the Virulence Plasmids of Salmonella enterica Serovars Typhimurium, Enteritidis, Choleraesuis, and Dublin

    OpenAIRE

    Chu, Chishih; Hong, Seau-Feng; Tsai, Chingju; Lin, Wen-Shiun; Liu, Tsui-Ping; Ou, Jonathan T.

    1999-01-01

    Using fragment profiling, PCR, and Southern hybridization, we found that Salmonella enterica serovar Choleraesuis harbored virulence plasmids of various sizes, whereas serovars Typhimurium, Enteritidis, and Dublin carried a plasmid of a unique size. Also, the virulence plasmid of Typhimurium contained genes in the same order detected in the other three plasmids, all of which contained deletions.

  2. A Site-Specific Integrative Plasmid Found in Pseudomonas aeruginosa Clinical Isolate HS87 along with A Plasmid Carrying an Aminoglycoside-Resistant Gene.

    Directory of Open Access Journals (Sweden)

    Dexi Bi

    Full Text Available Plasmids play critical roles in bacterial fitness and evolution of Pseudomonas aeruginosa. Here two plasmids found in a drug-resistant P. aeruginosa clinical isolate HS87 were completely sequenced. The pHS87b plasmid (11.2 kb carries phage-related genes and function-unknown genes. Notably, pHS87b encodes an integrase and has an adjacent tRNAThr-associated attachment site. A corresponding integrated form of pHS87b at the tRNAThr locus was identified on the chromosome of P. aeruginosa, showing that pHS87b is able to site-specifically integrate into the 3'-end of the tRNAThr gene. The pHS87a plasmid (26.8 kb displays a plastic structure containing a putative replication module, stability factors and a variable region. The RepA of pHS87a shows significant similarity to the replication proteins of pPT23A-family plasmids. pHS87a carries a transposon Tn6049, a truncated insertion sequence ΔIS1071 and a Tn402-like class 1 integron which contains an aacA4 cassette that may confer aminoglycoside resistance. Thus, pHS87b is a site-specific integrative plasmid whereas pHS87a is a plastic antibiotic resistance plasmid. The two native plasmids may promote the fitness and evolution of P. aeruginosa.

  3. A Site-Specific Integrative Plasmid Found in Pseudomonas aeruginosa Clinical Isolate HS87 along with A Plasmid Carrying an Aminoglycoside-Resistant Gene.

    Science.gov (United States)

    Bi, Dexi; Xie, Yingzhou; Tai, Cui; Jiang, Xiaofei; Zhang, Jie; Harrison, Ewan M; Jia, Shiru; Deng, Zixin; Rajakumar, Kumar; Ou, Hong-Yu

    2016-01-01

    Plasmids play critical roles in bacterial fitness and evolution of Pseudomonas aeruginosa. Here two plasmids found in a drug-resistant P. aeruginosa clinical isolate HS87 were completely sequenced. The pHS87b plasmid (11.2 kb) carries phage-related genes and function-unknown genes. Notably, pHS87b encodes an integrase and has an adjacent tRNAThr-associated attachment site. A corresponding integrated form of pHS87b at the tRNAThr locus was identified on the chromosome of P. aeruginosa, showing that pHS87b is able to site-specifically integrate into the 3'-end of the tRNAThr gene. The pHS87a plasmid (26.8 kb) displays a plastic structure containing a putative replication module, stability factors and a variable region. The RepA of pHS87a shows significant similarity to the replication proteins of pPT23A-family plasmids. pHS87a carries a transposon Tn6049, a truncated insertion sequence ΔIS1071 and a Tn402-like class 1 integron which contains an aacA4 cassette that may confer aminoglycoside resistance. Thus, pHS87b is a site-specific integrative plasmid whereas pHS87a is a plastic antibiotic resistance plasmid. The two native plasmids may promote the fitness and evolution of P. aeruginosa. PMID:26841043

  4. Expansion of the IncX plasmid family for improved identification and typing of novel plasmids in drug-resistant Enterobacteriaceae

    DEFF Research Database (Denmark)

    Johnson, Timothy J.; Bielak, Eliza Maria; Fortini, Daniela;

    2012-01-01

    IncX plasmids are narrow host range plasmids of Enterobactericeae that have been isolated for over 50years. They are known to encode type IV fimbriae enabling their own conjugative transfer, and to provide accessory functions to their host bacteria such as resistance towards antimicrobial agents ...

  5. Bacillus anthracis Virulent Plasmid pX02 Genes Found in Large Plasmids of Two Other Bacillus Species

    OpenAIRE

    Luna, Vicki A.; King, Debra S.; Peak, K. Kealy; Reeves, Frank; Heberlein-Larson, Lea; Veguilla, William; Heller, L.; Duncan, Kathleen E; Cannons, Andrew C.; Amuso, Philip; Cattani, Jacqueline

    2006-01-01

    In order to cause the disease anthrax, Bacillus anthracis requires two plasmids, pX01 and pX02, which carry toxin and capsule genes, respectively, that are used as genetic targets in the laboratory detection of the bacterium. Clinical, forensic, and environmental samples that test positive by PCR protocols established by the Centers for Disease Control and Prevention for B. anthracis are considered to be potentially B. anthracis until confirmed by culture and a secondary battery of tests. We ...

  6. Genetic transformation of a clinical (genital tract, plasmid-free isolate of Chlamydia trachomatis: engineering the plasmid as a cloning vector.

    Directory of Open Access Journals (Sweden)

    Yibing Wang

    Full Text Available Our study had three objectives: to extend the plasmid-based transformation protocol to a clinical isolate of C. trachomatis belonging to the trachoma biovar, to provide "proof of principle" that it is possible to "knock out" selected plasmid genes (retaining a replication competent plasmid and to investigate the plasticity of the plasmid. A recently developed, plasmid-based transformation protocol for LGV isolates of C. trachomatis was modified and a plasmid-free, genital tract C. trachomatis isolate from Sweden (SWFP- was genetically transformed. Transformation of this non-LGV C. trachomatis host required a centrifugation step, but the absence of the natural plasmid removed the need for plaque purification of transformants. Transformants expressed GFP, were penicillin resistant and iodine stain positive for accumulated glycogen. The transforming plasmid did not recombine with the host chromosome. A derivative of pGFP::SW2 carrying a deletion of the plasmid CDS5 gene was engineered. CDS5 encodes pgp3, a protein secreted from the inclusion into the cell cytoplasm. This plasmid (pCDS5KO was used to transform C. trachomatis SWFP-, and established that pgp3 is dispensable for plasmid function. The work shows it is possible to selectively delete segments of the chlamydial plasmid, and this is the first step towards a detailed molecular dissection of the role of the plasmid. The 3.6 kb β-galactosidase cassette was inserted into the deletion site of CDS5 to produce plasmid placZ-CDS5KO. Transformants were penicillin resistant, expressed GFP and stained for glycogen. In addition, they expressed β-galactosidase showing that the lacZ cassette was functional in C. trachomatis. An assay was developed that allowed the visualisation of individual inclusions by X-gal staining. The ability to express active β-galactosidase within chlamydial inclusions is an important advance as it allows simple, rapid assays to measure directly chlamydial infectivity without

  7. Separation of Staphylococcus aureus causing serious infections by electrophoretic techniques

    Czech Academy of Sciences Publication Activity Database

    Tesařová, Marie; Horká, Marie; Moravcová, Dana; Šťavíková, Lenka; Růžička, F.

    2014. s. 237-238. [Chemtech /14./. 22.10.2014-25.10.2014, Istambul] R&D Projects: GA MV VG20112015021 Institutional support: RVO:68081715 Keywords : Staphylococcus aureus * electrophoretic techniques Subject RIV: CB - Analytical Chemistry, Separation

  8. Improving Diagnosis and Treatment of Staphylococcus aureus Infections : Experimental Studies

    NARCIS (Netherlands)

    S. van den Berg (Sanne)

    2015-01-01

    markdownabstract__Abstract__ Staphylococcus aureus is an opportunistic pathogen that causes a variety of infections, ranging from mild skin infections like furuncles and impetigo, to severe, lifethreatening infections including endocarditis, osteomyelitis and pneumonia. Invasive infections are freq

  9. A polymerization-based method to construct a plasmid containing clustered DNA damage and a mismatch.

    Science.gov (United States)

    Takahashi, Momoko; Akamatsu, Ken; Shikazono, Naoya

    2016-10-01

    Exposure of biological materials to ionizing radiation often induces clustered DNA damage. The mutagenicity of clustered DNA damage can be analyzed with plasmids carrying a clustered DNA damage site, in which the strand bias of a replicating plasmid (i.e., the degree to which each of the two strands of the plasmid are used as the template for replication of the plasmid) can help to clarify how clustered DNA damage enhances the mutagenic potential of comprising lesions. Placement of a mismatch near a clustered DNA damage site can help to determine the strand bias, but present plasmid-based methods do not allow insertion of a mismatch at a given site in the plasmid. Here, we describe a polymerization-based method for constructing a plasmid containing clustered DNA lesions and a mismatch. The presence of a DNA lesion and a mismatch in the plasmid was verified by enzymatic treatment and by determining the relative abundance of the progeny plasmids derived from each of the two strands of the plasmid. PMID:27449134

  10. Rapid Mini-Prep Isolation of High-Quality Plasmid DNA from Lactococcus and Lactobacillus spp. †

    OpenAIRE

    O'Sullivan, Daniel J.; Klaenhammer, Todd R.

    1993-01-01

    A simple, rapid plasmid mini-prep procedure for lactococci and lactobacilli which gives high yields and can be performed on overnight broth cultures is presented. Large plasmids were isolated from both lactococci and lactobacilli, including a 70-kb plasmid from Lactobacillus acidophilus C7. The purity of the resulting plasmid DNA makes it suitable for subsequent molecular manipulations. The convenience of the technique makes this rapid mini-prep procedure suitable for routine plasmid isolatio...

  11. Effect of excessive cadmium chloride on the plasmids of E. coli HB 101 in vivo

    Institute of Scientific and Technical Information of China (English)

    1999-01-01

    After Escherichia coli HB101 with plasmid pWH58, pWH98, or pTBa5 were cultered respectively in amp LB broth which contained 50 mg/L CdCl2 constantly for 24 h, these plasmids were isolated from E. coli, and the effect of excessive CdCl2 on the E. coli HB101 and plasmid DNA was studied by surveying the growth of E. coli HB101 and plasmid, argarose gel electrophoresis and analysis of restriction fragment length polymorphism (RFLP) of plasmids, and plasmid transformation. The results showed that 50 mg/L CdCl2 treatment lagged the growth of E. coli HB101 for at least 4h, but after grown for 24h there were not significant differences in the growths of E. coli HB101s and the productions of plasmids between the treatment and control. These results implified that E. coli HB101 have induced adaptability to cadmium stress and excessive CdCl2 did not inhibit the replication and amp+ gene's expression of plasmid DNA in vivo of E. coli significantly. 50 mg/L CdCl2 treatment for 24 hours might cause the sequence's change of plasmid DNA, but could not lead to the random breakage of plasmid DNA strands. Moreover, after 50 mg/L of CdCl2 treatment in vivo the transformation activities of plasmid did not altered, implied excessive CdCl2 could not affect the superhelical structure of plasmid and also not break the loop of plasmid DNA evidently.

  12. Staphylococcus aureus Peptidoglycan Tertiary Structure from Carbon-13 Spin Diffusion

    OpenAIRE

    Sharif, Shasad; Singh, Manmilan; Kim, Sung Joon; Schaefer,Jacob

    2009-01-01

    The cell-wall peptidoglycan of Staphylococcus aureus is a heterogeneous, highly cross-linked polymer of unknown tertiary structure. We have partially characterized this structure by measuring spin diffusion from 13C labels in pentaglycyl cross-linking segments to natural-abundance 13C in the surrounding intact cell walls. The measurements were performed using a version of centerband-only detection of exchange (CODEX). The cell walls were isolated from S. aureus grown in media containing [1-13...

  13. S. aureus bacteria : a new target of serum calcification activity

    OpenAIRE

    Dy, Diane Jazmin

    2009-01-01

    Staphylococcus aureus are gram- positive bacteria that cause skin and soft tissue infections. The continual incidence of infection is of great concern especially with the advent of methicillin resistant S. aureus (MRSA). Continued investigation on mechanisms our body uses to fight bacterial infection is vital. Our study suggests that the body takes advantage of a mechanism that mineralizes type-I collagen of bone and tendon to also mineralize bacteria. Serum driven bacterial mineralization ma...

  14. Comparative Pharmacodynamics of Gentamicin against Staphylococcus aureus and Pseudomonas aeruginosa†

    OpenAIRE

    Tam, Vincent H.; Kabbara, Samer; Vo, Giao; Schilling, Amy N.; Coyle, Elizabeth A.

    2006-01-01

    Aminoglycosides are often used to treat severe infections with gram-positive organisms. Previous studies have shown concentration-dependent killing by aminoglycosides of gram-negative bacteria, but limited data are available for gram-positive bacteria. We compared the in vitro pharmacodynamics of gentamicin against Staphylococcus aureus and Pseudomonas aeruginosa. Five S. aureus strains were examined (ATCC 29213 and four clinical isolates). Time-kill studies (TKS) in duplicate (baseline inocu...

  15. Role of Monocytes in Experimental Staphylococcus aureus Endocarditis

    OpenAIRE

    Veltrop, Marcel H. A. M.; Bancsi, Maurice J. L. M. F.; Bertina, Rogier M.; Thompson, Jan

    2000-01-01

    In the pathogenesis of bacterial endocarditis (BE), the clotting system plays a cardinal role in the formation and maintenance of the endocardial vegetations. The extrinsic pathway is involved in the activation of the coagulation pathway with tissue factor (TF) as the key protein. Staphylococcus aureus is a frequently isolated bacterium from patients with BE. We therefore investigated whether S. aureus can induce TF activity (TFA) on fibrin-adherent monocytes, used as an in vitro model of BE....

  16. Vancomycin Resistance Pattern of Staphylococcus Aureus among Clinical Samples

    OpenAIRE

    S Saadat; K Solhjoo; A. Kazemi; Erfanian, S. (MSc); Ashrafian, F. (MSc)

    2014-01-01

    Background and Objective: Vancomycin is used for treatment of methicillin-resistant S. Aureus (MRSA) infections; therefore, resistance to this antibiotic is increasing. We aimed to determine the antibiotic resistance pattern and frequency of vancomycin resistant S. Areas (VRSA) strains isolated from clinical samples. Material and Methods: In this cross-sectional study, 100 S. Aureus isolates collected from hospitals in Shiraz during six months, 2012, were identified by biochemical, microbiolo...

  17. AKTIVITAS ANTIBAKTERI KITOSAN TERHADAP BAKTERI S.aureus

    OpenAIRE

    Mardiyah Kurniasih; Dwi Kartika

    2009-01-01

    Chitosan is the N-deacetylated derivative of chitin. Chitosan is biodegradable, biocompatible and non-toxic. Chitosan is polycationic in acidic media and give antibacterial activity. In this paper, antibacterial activity of chitosan have been studied. Chitosan had been isolated from white shrimp. Antibacterial activity of chitosan solutions was examined against S. aureus The result showed that antimicrobial effect on S. aureus was strengthened as the choitosan concentrate decreased.

  18. Susceptibility of Staphylococcus aureus biofilms to reactive discharge gases

    OpenAIRE

    Traba, Christian; Liang, Jun F.

    2011-01-01

    Formation of bacterial biofilms at solid-liquid interfaces creates numerous problems in both industrial and biomedical sciences. In this study, the susceptibility of Staphylococcus aureus biofilms to discharge gas generated from plasma was tested. It was found that despite distinct chemical/physical properties, discharge gases from oxygen, nitrogen, and argon demonstrated very potent and almost the same anti-biofilm activity. The bacterial cells in S. aureus biofilms were killed (>99.9%) by d...

  19. Resistance to Antimicrobials Mediated by Efflux Pumps in Staphylococcus aureus

    OpenAIRE

    Isabel Couto; Leonard Amaral; José Melo-Cristino; Miguel Viveiros; Cláudia Palma; Elisabete Junqueira; Costa, Sofia S.

    2013-01-01

    Resistance mediated by efflux has been recognized in Staphylococcus aureus in the last few decades, although its clinical relevance has only been recognized recently. The existence of only a few studies on the individual and overall contribution of efflux to resistance phenotypes associated with the need of well-established methods to assess efflux activity in clinical isolates contributes greatly to the lack of solid knowledge of this mechanism in S. aureus. This study aims to provide inform...

  20. Methicillin resistance & inducible clindamycin resistance in Staphylococcus aureus

    OpenAIRE

    Soumyadeep Ghosh; Mandira Banerjee

    2016-01-01

    Background & objectives: Methicillin resistant Staphylococcus aureus (MRSA) isolates with inducible clindamycin resistance (iCR) are resistant to erythromycin and sensitive to clindamycin on routine testing and inducible clindamycin resistance can only be identified by D-test. This study was aimed to detect methicillin resistance and iCR among S. aureus isolates, effectiveness of some commonly used antibiotics and correlation between methicillin resistance and iCR. Methods: The present cro...

  1. Methicillin-Resistant Staphylococcus aureus Ocular Infection in Taiwan

    OpenAIRE

    Kang, Yu-Chuan; Hsiao, Ching-Hsi; Yeh, Lung-Kun; Ma, David H. K.; Chen, Phil Y. F.; Lin, Hsin-Chiung; Tan, Hsin-Yuan; Chen, Hung-Chi; Chen, Shin-Yi; Huang, Yhu-Chering

    2015-01-01

    Abstract Methicillin-resistant Staphylococcus aureus (MRSA) infection is an important public health issue. This observational study aimed to characterize clinical features, antibiotic susceptibility, and genotypes of ocular infections caused by MRSA based on the clinical and molecular definitions of community-associated (CA) and healthcare-associated (HA) strains. Fifty-nine patients with culture-proven S aureus ocular infection were enrolled from January 1, 2010 to December 31, 2011 at Chang...

  2. Resistance to Antimicrobials Mediated by Efflux Pumps in Staphylococcus aureus

    Directory of Open Access Journals (Sweden)

    Isabel Couto

    2013-03-01

    Full Text Available Resistance mediated by efflux has been recognized in Staphylococcus aureus in the last few decades, although its clinical relevance has only been recognized recently. The existence of only a few studies on the individual and overall contribution of efflux to resistance phenotypes associated with the need of well-established methods to assess efflux activity in clinical isolates contributes greatly to the lack of solid knowledge of this mechanism in S. aureus. This study aims to provide information on approaches useful to the assessment and characterization of efflux activity, as well as contributing to our understanding of the role of efflux to phenotypes of antibiotic resistance and biocide tolerance in S. aureus clinical isolates. The results described show that efflux is an important contributor to fluoroquinolone resistance in S. aureus and suggest it as a major mechanism in the early stages of resistance development. We also show that efflux plays an important role on the reduced susceptibility to biocides in S. aureus, strengthening the importance of this long neglected resistance mechanism to the persistence and proliferation of antibiotic/biocide-resistant S. aureus in the hospital environment.

  3. Photoreactivation of ultraviolet-irradiation damage in Staphylococcus aureus

    International Nuclear Information System (INIS)

    This study reports the capacity of Staphylococcus aureus strain 7 - 8 to undergo photoenzymatic repair of UV-irradiation induced damage and compares it to the photoreactivation (PR) response of Escherichia coli strain B. Staphylococcus aureaus showed greater inhibition by UV irradiation than E. coli, consistent with its higher adenine and thymine content of DNA. Staphylococcus aureus showed an enhanced rate of photoreactivation with no lag in initiation of the PR response at low PR doses compared to E. coli. Maximum PR capacity of both cultures was about equal and occurred in cultures incubated at 23 - 250. The PR responses at 11 - 12 and 35 - 370 for S. aureus and E. coli differed although both were capable of PR at each of these temperatures. The PR response of E. coli was directly related to the dosage of PR light (J/m2); however, the photoenzymatic capacity of S. aureus was not directly responsive to continued decrease in light intensity. The capacity of S. aureus to undergo liquid holding recovery (LHR) occurred at 23 - 250 (not at 11 - 120 or 35 - 370), whereas E. coli underwent LHR at 11 - 120 and 23 - 250 but not at 35 - 370. The LHR response of S. aureus was somewhat more effective than E. coli and did not show the direct response to increased liquid-holding period as did E. coli. (author)

  4. Staphylococcus aureus – antimicrobial resistance and the immunocompromised child

    Directory of Open Access Journals (Sweden)

    McNeil JC

    2014-05-01

    Full Text Available J Chase McNeilDepartment of Pediatrics, Section of Infectious Diseases, Baylor College of Medicine, Houston, TX, USAAbstract: Children with immunocompromising conditions represent a unique group for the acquisition of antimicrobial resistant infections due to their frequent encounters with the health care system, need for empiric antimicrobials, and immune dysfunction. These infections are further complicated in that there is a relative paucity of literature on the clinical features and management of Staphylococcus aureus infections in immunocompromised children. The available literature on the clinical features, antimicrobial susceptibility, and management of S. aureus infections in immunocompromised children is reviewed. S. aureus infections in children with human immunodeficiency virus (HIV are associated with higher HIV viral loads and a greater degree of CD4 T-cell suppression. In addition, staphylococcal infections in children with HIV often exhibit a multidrug resistant phenotype. Children with cancer have a high rate of S. aureus bacteremia and associated complications. Increased tolerance to antiseptics among staphylococcal isolates from pediatric oncology patients is an emerging area of research. The incidence of S. aureus infections among pediatric solid organ transplant recipients varies considerably by the organ transplanted; in general however, staphylococci figure prominently among infections in the early posttransplant period. Staphylococcal infections are also prominent pathogens among children with a number of immunodeficiencies, notably chronic granulomatous disease. Significant gaps in knowledge exist regarding the epidemiology and management of S. aureus infection in these vulnerable children.Keywords: pediatric, HIV, cancer, transplant

  5. Multiple drug resistant carbapenemases producing Acinetobacter baumannii isolates harbours multiple R-plasmids

    Directory of Open Access Journals (Sweden)

    Rajagopalan Saranathan

    2014-01-01

    Full Text Available Background & objectives: The nosocomial human pathogen Acinetobacter baumannii has high propensity to develop resistance to antimicrobials and to become multidrug resistant (MDR, consequently complicating the treatment. This study was carried out to investigate the presence of resistant plasmids (R-plasmids among the clinical isolates of A. baumannii. In addition, the study was performed to check the presence of common β-lactamases encoding genes on these plasmids. Methods: A total of 55 clinical isolates of A. baumannii were included in the study and all were subjected to plasmid DNA isolation, followed by PCR to check the presence of resistance gene determinants such as blaOXA-23 , blaOXA-51, blaOXA-58 and blaIMP-1 on these plasmids that encode for oxacillinase (OXA and metallo-β-lactamase (MBL type of carbapenemases. Plasmid curing experiments were carried out on selected isolates using ethidium bromide and acridine orange as curing agents and the antibiotic resistance profiles were evaluated before and after curing. Results: All the isolates were identified as A. baumannii by 16SrDNA amplification and sequencing. Plasmid DNA isolated from these isolates showed the occurrence of multiple plasmids with size ranging from 500bp to ≥ 25 kb. The percentage of blaOXA-51 and blaOXA-23 on plasmids were found to be 78 and 42 per cent, respectively and 20 isolates (36% carried blaIMP-1 gene on plasmids. Significant difference was observed in the antibiograms of plasmid cured isolates when compared to their parental ones. The clinical isolates became susceptible to more than two antibiotic classes after curing of plasmids indicating plasmid borne resistance. Interpretation & conclusions: Our study determined the plasmid mediated resistance mechanisms and occurrence of different resistance genes on various plasmids isolated from MDR A. baumannii. The present findings showed the evidence for antibiotic resistance mediated through multiple plasmids in

  6. Plasmid purification by phenol extraction from guanidinium thiocyanate solution: development of an automated protocol.

    Science.gov (United States)

    Fisher, J A; Favreau, M B

    1991-05-01

    We have developed a novel plasmid isolation procedure and have adapted it for use on an automated nucleic acid extraction instrument. The protocol is based on the finding that phenol extraction of a 1 M guanidinium thiocyanate solution at pH 4.5 efficiently removes genomic DNA from the aqueous phase, while supercoiled plasmid DNA is retained in the aqueous phase. S1 nuclease digestion of the removed genomic DNA shows that it has been denatured, which presumably confers solubility in the organic phase. The complete automated protocol for plasmid isolation involves pretreatment of bacterial cells successively with lysozyme, RNase A, and proteinase K. Following these digestions, the solution is extracted twice with a phenol/chloroform/water mixture and once with chloroform. Purified plasmid is then collected by isopropanol precipitation. The purified plasmid is essentially free of genomic DNA, RNA, and protein and is a suitable substrate for DNA sequencing and other applications requiring highly pure supercoiled plasmid. PMID:1713749

  7. Growth dependence of conjugation explains limited plasmid invasion in biofilms: an individual‐based modelling study

    DEFF Research Database (Denmark)

    Merkey, Brian; Lardon, Laurent; Seoane, Jose Miguel;

    2011-01-01

    Plasmid invasion in biofilms is often surprisingly limited in spite of the close contact of cells in a biofilm. We hypothesized that this poor plasmid spread into deeper biofilm layers is caused by a dependence of conjugation on the growth rate (relative to the maximum growth rate) of the donor....... By extending an individual‐based model of microbial growth and interactions to include the dynamics of plasmid carriage and transfer by individual cells, we were able to conduct in silico tests of this and other hypotheses on the dynamics of conjugal plasmid transfer in biofilms. For a generic model plasmid......, we find that invasion of a resident biofilm is indeed limited when plasmid transfer depends on growth, but not so in the absence of growth dependence. Using sensitivity analysis we also find that parameters related to timing (i.e. a lag before the transconjugant can transfer, transfer proficiency...

  8. Conjugation is necessary for a bacterial plasmid to survive under protozoan predation.

    Science.gov (United States)

    Cairns, Johannes; Jalasvuori, Matti; Ojala, Ville; Brockhurst, Michael; Hiltunen, Teppo

    2016-02-01

    Horizontal gene transfer by conjugative plasmids plays a critical role in the evolution of antibiotic resistance. Interactions between bacteria and other organisms can affect the persistence and spread of conjugative plasmids. Here we show that protozoan predation increased the persistence and spread of the antibiotic resistance plasmid RP4 in populations of the opportunist bacterial pathogen Serratia marcescens. A conjugation-defective mutant plasmid was unable to survive under predation, suggesting that conjugative transfer is required for plasmid persistence under the realistic condition of predation. These results indicate that multi-trophic interactions can affect the maintenance of conjugative plasmids with implications for bacterial evolution and the spread of antibiotic resistance genes. PMID:26843557

  9. Plasmid DNA lesions induced by α particle exposure observed with the atomic force microscopy

    International Nuclear Information System (INIS)

    Objective: To examine the lesions of plasmid DNA induced by α particle exposure. Methods: The α-particle irradiated plasmid pGEM-Tl DNA was deposited on mica and was observed with Nanoscope III a atomic force microscopy. The irradiated DNA was concurrently analyzed by electrophoresis. Results: The three types of plasmid DNA molecules, i.e, open circle (OC), super coiled (SC) and linear were seen clearly under atomic force microscopy. The proportion of OC plasmid DNA was obviously increased at as less as 2 Gy of α particle exposure as compared with 10 Gy of γ-ray exposure. Conclusions: Clear image of plasmid DNA molecules was obtained using atomic force microscopy. The lesions of plasmid DNA induced by α particles were more serious than those by γ-ray with the same dose of exposure in vitro

  10. Characterization of a Cryptic and Intriguing Low Molecular Weight Plasmid.

    Science.gov (United States)

    Carneiro, Lilian C; Mendes, Paulo Vinicius C; Silva, Silvana P; Souza, Guilherme R L; Bataus, Luiz Artur M

    2016-03-01

    The complete nucleotide sequence of cryptic plasmid pVCM04 isolated from Salmonella enterica serovar Enteritidis was determined and analyzed. pVCM04 contains 3853 bp with 53.6 % GC content and has twelve ORFs with more than 50 amino acids. Five of these sequences showed homology with replication and mobilization proteins. ORF1 and ORF2 showed homology with replication proteins, while ORFs 3-5 showed homology with mobilization proteins. The pVCM04 possesses a region associated with the theta-type replication mechanism. BLASTn search analysis revealed unexpectedly no similarity with sequences deposited in GenBank. The nucleotide sequence of pVCM04 can be divided into two arms: the region between nucleotides 552-1774 (encoding RepA and RepB) and the region between nucleotides 1775-3853 (encoding MobA, MobB and MobC). Codon bias pattern is distinct between mobA and repA, so the program Modeltest was used to select the best evolutionary model to study these genes. The result of ModelTest (model GTR+G for mobA and model HKY+G for repA) suggests that these genes would be subject to different selective pressures. Considering the differences in the codon usage, the selection of two different evolutionary models, and the absence of plasmids with homology to pVCM04 in GenBank, we believe that pVCM04 is a chimeric molecule and represents a new plasmid lineage. PMID:26670037

  11. Plasmid DNA damage caused by stibine and trimethylstibine

    International Nuclear Information System (INIS)

    Antimony is classified as 'possibly carcinogenic to humans' and there is also sufficient evidence for antimony carcinogenicity in experimental animals. Stibine is a volatile inorganic antimony compound to which humans can be exposed in occupational settings (e.g., lead-acid battery charging). Because it is highly toxic, stibine is considered a significant health risk; however, its genotoxicity has received little attention. For the work reported here, stibine was generated by sodium borohydride reduction of potassium antimony tartrate. Trimethylstibine is a volatile organometallic antimony compound found commonly in landfill and sewage fermentation gases at concentrations ranging between 0.1 and 100 μg/m3. Trimethylstibine is generally considered to pose little environmental or health risk. In the work reported here, trimethylstibine was generated by reduction of trimethylantimony dichloride using either sodium borohydride or the thiol compounds, dithioerythritol (DTE), L-cysteine, and glutathione. Here we report the evaluation of the in vitro genotoxicities of five antimony compounds--potassium antimony tartrate, stibine, potassium hexahydroxyantimonate, trimethylantimony dichloride, and trimethylstibine--using a plasmid DNA-nicking assay. Of these five antimony compounds, only stibine and trimethylstibine were genotoxic (significant nicking to pBR 322 plasmid DNA). We found stibine and trimethylstibine to be about equipotent with trimethylarsine using this plasmid DNA-nicking assay. Reaction of trimethylantimony dichloride with either glutathione or L-cysteine to produce DNA-damaging trimethylstibine was observed with a trimethylantimony dichloride concentration as low as 50 μM and L-cysteine or glutathione concentrations as low as 500 and 200 μM, respectively, for a 24 h incubation

  12. Physical characterization of plasmids determining synthesis of a microcin which inhibits methionine synthesis in Escherichia coli.

    OpenAIRE

    Perez-Diaz, J C; Clowes, R C

    1980-01-01

    Plasmid deoxyribonucleic acid (DNA) isolated from each of three antibiotic-resistant clinical strains of Escherichia coli producing the same microcin showed multiple bands upon agarose gel electrophoresis. Transformants selected either for microcin resistance or ampicillin resistance yielded plasmid DNA corresponding in size to only one of the multiple bands. Plasmids, isolated from all three hosts, which determined microcin resistance and microcin production measured about 4 megadaltons by s...

  13. Redundant transfer of F' plasmids occurs between Escherichia coli cells during nonlethal selections.

    OpenAIRE

    Peters, J E; Benson, S A

    1995-01-01

    Surface exclusion is the mechanism by which F plasmids prevent the redundant entry of additional F plasmids into the host cell during exponential growth. This mechanism is relaxed in cells that are in stationary phase. Using genetically marked F' plasmids and host strains, we extend this finding to Escherichia coli populations during extended nonlethal selection in bacterial lawns. We show that a high level of redundant transfer occurs between these nongrowing cells during the selection. This...

  14. Parallel Compensatory Evolution Stabilizes Plasmids across the Parasitism-Mutualism Continuum

    OpenAIRE

    Harrison, Ellie; Guymer, David; Spiers, Andrew J.; Paterson, Steve; Brockhurst, Michael A.

    2015-01-01

    Plasmids drive genomic diversity in bacteria via horizontal gene transfer [1, 2]; nevertheless, explaining their survival in bacterial populations is challenging [3]. Theory predicts that irrespective of their net fitness effects, plasmids should be lost: when parasitic (costs outweigh benefits), plasmids should decline due to purifying selection [4-6], yet under mutualism (benefits outweigh costs), selection favors the capture of beneficial accessory genes by the chromosome and loss of the c...

  15. Stability of plasmid content in Salmonella wien in late phases of the epidemic history.

    OpenAIRE

    Casalino, M.; Comanducci, A; Nicoletti, M; Maimone, F

    1984-01-01

    Prevalence, genetic characteristics, and EcoRI cleavage analysis of plasmids identified in clinical strains of Salmonella wien isolated in recent years showed that the plasmid content in this serotype has remained uniform and stable over more than a decade and also late in the epidemic history. No correlation between decrease in S. wien isolations and naturally occurring systematic changes in the DNA of its most common FIme plasmid was structurally detectable.

  16. Differences in the stability of the plasmids of Yersinia pestis cultures in vitro: impact on virulence

    OpenAIRE

    TC Leal-Balbino; NC Leal; CV Lopes; AMP de Almeida

    2004-01-01

    Plasmid and chromosomal genes encode determinants of virulence for Yersinia pestis, the causative agent of plague. However, in vitro, Y. pestis genome is very plastic and several changes have been described. To evaluate the alterations in the plasmid content of the cultures in vitro and the impact of the alterations to their pathogenicity, three Y. pestis isolates were submitted to serial subculture, analysis of the plasmid content, and testing for the presence of characteristic genes in each...

  17. Adaptation of model genetically engineered microorganisms to lake water: growth rate enhancements and plasmid loss.

    OpenAIRE

    Sobecky, P A; Schell, M A; Moran, M. A.; Hodson, R. E.

    1992-01-01

    When a genetically engineered microorganism (GEM) is released into a natural ecosystem, its survival, and hence its potential environmental impact, depends on its genetic stability and potential for growth under highly oligotrophic conditions. In this study, we compared plasmid stability and potential for growth on low concentrations of organic nutrients of strains of Pseudomonas putida serving as model GEMs. Plasmid-free and plasmid-bearing (NAH7) prototrophic isogenic strains and two amino-...

  18. Cloning in Streptococcus lactis of plasmid-mediated UV resistance and effect on prophage stability.

    OpenAIRE

    Chopin, M C; Chopin, A; Rouault, A.; Simon, D.

    1986-01-01

    Plasmid pIL7 (33 kilobases) from Streptococcus lactis enhances UV resistance and prophage stability. A 5.4-kilobase pIL7 fragment carrying genes coding for both characters was cloned into S. lactis, using plasmid pHV1301 as the cloning vector. The recombinant plasmid was subsequently transferred to three other S. lactis strains by transformation or protoplast fusion. Cloned genes were expressed in all tested strains.

  19. Replication and segregational stability of Bacillus plasmid pBAA1.

    OpenAIRE

    DEVINE, KEVIN; MC CONNELL, DAVID JOHN

    1989-01-01

    PUBLISHED A cryptic plasmid, pBAA1, was identified in an industrial Bacillus strain. The plasmid is 6.8 kilobases in size and is present in cells at a copy number of approximately 5 per chromosome equivalent. The plasmid has been maintained under industrial fermentation conditions without apparent selective pressure and so is assumed to be partition proficient. The minimal replicon was localized to a 1.4-kilobase fragment which also contains the functions required for copy number control. ...

  20. A cell engineering strategy to enhance supercoiled plasmid DNA production for gene therapy

    OpenAIRE

    Hassan, S.; Keshavarz-Moore, E.; J. Ward

    2016-01-01

    With the recent revival of the promise of plasmid DNA vectors in gene therapy, a novel synthetic biology approach was used to enhance the quantity, (yield), and quality of the plasmid DNA. Quality was measured by percentage supercoiling and supercoiling density, as well as improving segregational stability in fermentation. We examined the hypothesis that adding a Strong Gyrase binding Site (SGS) would increase DNA gyrase-mediated plasmid supercoiling. SGS from 3 different replicons, (the Mu b...

  1. Characterization and molecular cloning in Escherichia coli of a plasmid from the mollicute Spiroplasma citri.

    OpenAIRE

    Mouches, C; Barroso, G.; Bové, J M

    1983-01-01

    Two plasmids, pMH1 with 7 kilobase pairs and pM41 with 8 kilobase pairs, were purified from the plant pathogen Spiroplasma citri and characterized by restriction mapping. Upon in vitro DNA recombination with plasmid pBR328 as a vector, we have cloned pMH1 in Escherichia coli. A radioactive probe obtained upon nick translation of the recombinant plasmid was used to further characterize and compare pMH1 and pM41.

  2. Cloning in Streptococcus lactis of plasmid-mediated UV resistance and effect on prophage stability

    International Nuclear Information System (INIS)

    Plasmid pIL7 (33 kilobases) from Streptococcus lactis enhances UV resistance and prophage stability. A 5.4-kilobase pIL7 fragment carrying genes coding for both characters was cloned into S. lactis, using plasmid pHV1301 as the cloning vector. The recombinant plasmid was subsequently transferred to three other S. lactis strains by transformation or protoplast fusion. Cloned genes were expressed in all tested strains

  3. Transferable plasmid-mediated resistance to streptomycin in a clinical isolate of Yersinia pestis.

    OpenAIRE

    Guiyoule, A; Gerbaud, G; Buchrieser, C.; Galimand, M.; Rahalison, L.; Chanteau, S.; Courvalin, P; Carniel, E

    2001-01-01

    Plasmid-mediated high-level resistance to multiple antibiotics was reported in a clinical isolate of Yersinia pestis in Madagascar in 1997. We describe a second Y. pestis strain with high-level resistance to streptomycin, isolated from a human case of bubonic plague in Madagascar. The resistance determinants were carried by a self-transferable plasmid that could conjugate at high frequencies to other Y. pestis isolates. The plasmid and the host bacterium were different from those previously a...

  4. A New Extant Respirometric Assay to Estimate Intrinsic Growth Parameters Applied to Study Plasmid Metabolic Burden

    DEFF Research Database (Denmark)

    Seoane, Jose Miguel; Sin, Gürkan; Lardon, Laurent;

    2010-01-01

    burden caused by the carriage of a pWW0 TOL plasmid in the model organism Pseudomonas putida KT2440; The metabolic,burden associated was manifested as a reduction in the yield and the specific growth rate of the host, with both plasmid maintenance and the over-expression of recombinant proteins from...... the plasmid contributing equally to the overall effect. Biotechnol. Bioeng. 2010;105: 141-149. (C) 2009 Wiley Periodicals, Inc....

  5. General Mutagenesis of F Plasmid TraI Reveals Its Role in Conjugative Regulation

    OpenAIRE

    Haft, Rembrandt J F; Palacios, Gilberto; Nguyen, Tran; Mally, Manuela; Gachelet, Eliora G.; Zechner, Ellen L.; Traxler, Beth

    2006-01-01

    Bacteria commonly exchange genetic information by the horizontal transfer of conjugative plasmids. In gram-negative conjugation, a relaxase enzyme is absolutely required to prepare plasmid DNA for transit into the recipient via a type IV secretion system. Here we report a mutagenesis of the F plasmid relaxase gene traI using in-frame, 31-codon insertions. Phenotypic analysis of our mutant library revealed that several mutant proteins are functional in conjugation, highlighting regions of TraI...

  6. Transfer of plasmid RSF1010 by conjugation from Escherichia coli to Streptomyces lividans and Mycobacterium smegmatis.

    OpenAIRE

    Gormley, E P; Davies, J.

    1991-01-01

    The plasmid RSF1010 belongs to a class of plasmids (IncQ) that replicate in a range of bacterial hosts. Although non-self-transmissible, it can be mobilized at high frequency between different gram-negative bacterial species if transfer functions are supplied in trans. We report the transfer of RSF1010 by conjugation from Escherichia coli to the gram-positive actinomycetes Streptomyces lividans and Mycobacterium smegmatis. In its new hosts, the plasmid was stable with respect to structure and...

  7. Selective Conditions for a Multidrug Resistance Plasmid Depend on the Sociality of Antibiotic Resistance

    OpenAIRE

    Bottery, Michael; Wood, A. Jamie; Brockhurst, Michael

    2016-01-01

    Multidrug resistance (MDR) plasmids frequently carry antibiotic resistance genes conferring qualitatively different mechanisms of resistance. We show here that the antibiotic concentrations selecting for the RK2 plasmid in Escherichia coli depend upon the sociality of the drug resistance: the selection for selfish drug resistance (efflux pump) occurred at very low drug concentrations, just 1.3% of the MIC of the plasmid-free antibiotic-sensitive strain, whereas selection for cooperative drug ...

  8. Plasmid transfer between strains of Bacillus thuringiensis infecting Galleria mellonella and Spodoptera littoralis.

    OpenAIRE

    Jarrett, P.; Stephenson, M

    1990-01-01

    To determine the possibility of plasmid transfer occurring between strains of Bacillus thuringiensis in infected lepidopterous larvae, Galleria mellonella and Spodoptera littoralis were infected with two or more strains of B. thuringiensis and the resulting bacteria from the dead insects were examined for plasmid transfer. Transfer rates of plasmids coding for crystal production and tetracycline resistance were high, reaching levels similar to those obtained in laboratory broth cultures. Tran...

  9. Reporter gene expression in dendritic cells after gene gun administration of plasmid DNA

    OpenAIRE

    Watkins, Craig; Hopkins, John; Harkiss, Gordon

    2005-01-01

    Dendritic cells (DC) play an integral role in plasmid DNA vaccination. However, the interaction between plasmid DNA and DC in vivo is incompletely understood. In this report, we utilise the sheep pseudoafferent cannulation model to examine the interaction between plasmid DNA encoding enhanced green fluorescent protein (pEGFP) and afferent lymph DC (ALDC) following gene gun administration. The results show that peaks of fluorescent ALDC tended to appear around days 1-4 and 9-13, then erratical...

  10. Quantification Bias Caused by Plasmid DNA Conformation in Quantitative Real-Time PCR Assay

    OpenAIRE

    Lin, Chih-Hui; Chen, Yu-Chieh; Pan, Tzu-Ming

    2011-01-01

    Quantitative real-time PCR (qPCR) is the gold standard for the quantification of specific nucleic acid sequences. However, a serious concern has been revealed in a recent report: supercoiled plasmid standards cause significant over-estimation in qPCR quantification. In this study, we investigated the effect of plasmid DNA conformation on the quantification of DNA and the efficiency of qPCR. Our results suggest that plasmid DNA conformation has significant impact on the accuracy of absolute qu...

  11. Use of locked nucleic acid oligonucleotides to add functionality to plasmid DNA

    OpenAIRE

    Hertoghs, Kirsten M. L.; Ellis, Jonathan H.; Catchpole, Ian R.

    2003-01-01

    The available reagents for the attachment of functional moieties to plasmid DNA are limiting. Most reagents bind plasmid DNA in a non-sequence- specific manner, with undefined stoichiometry, and affect DNA charge and delivery properties or involve chemical modifications that abolish gene expression. The design and ability of oligonucleotides (ODNs) containing locked nucleic acids (LNAs) to bind supercoiled, double-stranded plasmid DNA in a sequence-specific manner are described for the first ...

  12. Natural Transformation of Acinetobacter calcoaceticus by Plasmid DNA Adsorbed on Sand and Groundwater Aquifer Material

    OpenAIRE

    Chamier, Bärbel; Lorenz, Michael G.; Wackernagel, Wilfried

    1993-01-01

    It is known that plasmid DNA and linear duplex DNA molecules adsorb to chemically purified mineral grains of sand and to particles of several clay fractions. It seemed desirable to examine whether plasmid DNA would also adsorb to nonpurified mineral materials taken from the environment and, particularly, whether adsorbed plasmid DNA would be available for natural transformation of bacteria. Therefore, microcosms consisting of chemically pure sea sand plus buffered CaCl2 solution were compared...

  13. Using Mahalanobis distance to compare genomic signatures between bacterial plasmids and chromosomes

    OpenAIRE

    Suzuki, Haruo; Sota, Masahiro; Brown, Celeste J.; Top, Eva M.

    2008-01-01

    Plasmids are ubiquitous mobile elements that serve as a pool of many host beneficial traits such as antibiotic resistance in bacterial communities. To understand the importance of plasmids in horizontal gene transfer, we need to gain insight into the ‘evolutionary history’ of these plasmids, i.e. the range of hosts in which they have evolved. Since extensive data support the proposal that foreign DNA acquires the host's nucleotide composition during long-term residence, comparison of nucleoti...

  14. Plasmid-associated transfer of tetracycline resistance in Bacteroides ruminicola.

    OpenAIRE

    Flint, H J; Thomson, A. M.; Bisset, J

    1988-01-01

    Tetracycline resistance was transferred at frequencies between 10(-7) and 10(-6) per recipient cell in anaerobic matings between two strains of the strictly anaerobic rumen bacterium Bacteroides ruminicola. The donor strain, 223/M2/7, was a multiple-plasmid-bearing tetracycline-resistant strain from the ovine rumen, and the recipient, F101, was a rifampin-resistant mutant of B14, a bovine strain belonging to B. ruminicola subsp. brevis. Resistance transfer could occur in the presence of DNase...

  15. [Plasmid characteristics of naphthalene and salicylate biodegradation in Pseudomonas putida].

    Science.gov (United States)

    Zakharian, R A; Bakunin, K A; Gasparian, N S; Kocharian, Sh M; Arakelov, G M

    1980-01-01

    The object of this work was to study the physico-chemical and biological properties of DNAs of the biodegradation plasmids NAH and SAL. A comparative analysis of the physico-chemical parameters for these DNAs made it possible to detect a number of identical properties in them: the same sedimentation profile for covalently-closed circular DNA forms, 68--70 S; the molecular weight of ca. 50 MD; a roughly equal number of fragments (up to 23) was found when the DNAs of NAH and SAL were restricted by EcoRI endonuclease. The transformation of the plasmidless strain PpGI was done. PMID:6259498

  16. Cold atmospheric pressure plasma jet interactions with plasmid DNA

    International Nuclear Information System (INIS)

    The effect of a cold (<40 deg. C) radio frequency-driven atmospheric pressure plasma jet on plasmid DNA has been investigated. Gel electrophoresis was used to analyze the DNA forms post-treatment. The experimental data are fitted to a rate equation model that allows for quantitative determination of the rates of single and double strand break formation. The formation of double strand breaks correlates well with the atomic oxygen density. Taken with other measurements, this indicates that neutral components in the jet are effective in inducing double strand breaks.

  17. Peaceful coexistence amongst Borrelia plasmids: getting by with a little help from their friends?

    OpenAIRE

    Chaconas, George; Norris, Steven J

    2013-01-01

    Borrelia species comprise a unique genus of bacterial pathogens. These organisms contain a segmented genome with up to two dozen plasmids ranging in size from 5kb up to about 200 kb. The plasmids have also been referred to as mini-chromosomes or essential genetic elements, as some of them carry information important for infection of vertebrates or for survival in the tick vector. Most of the plasmids are linear with covalently closed hairpin telomeres and these linear plasmids are in a consta...

  18. A procedure for large-scale plasmid isolation without using ultracentrifugation.

    Science.gov (United States)

    Chakrabarti, A; Sitaric, S; Ohi, S

    1992-10-01

    An expedient procedure for large-scale plasmid isolation from Escherichia coli strains without using ultracentrifugation or special setups or reagents is described. The protocol, which utilizes a modified alkaline extraction procedure as well as differential precipitations by isopropanol and lithium chloride, is simple and rapid and yet produces plasmid DNA with a yield of about 2 mg/liter culture. The isolated plasmids consisted of mostly monomeric and dimeric covalently closed circular DNA. The plasmids could be digested by various restriction endonucleases and were compatible with gene cloning, transfection-gene expression, and viral production. PMID:1333773

  19. Plasmid marker rescue transformation proceeds by breakage-reunion in Bacillus subtilis

    International Nuclear Information System (INIS)

    Bacillus subtilis carrying a plasmid which replicates with a copy number of about 1 was transformed with linearized homologous plasmid DNA labeled with the heavy isotopes 2H and 15N, in the presence of 32Pi and 6-(p-hydroxyphenylazo)-uracil to inhibit DNA replication. Plasmid DNA was isolated from the transformed culture and fractionated in cesium chloride density gradients. The distribution of total and donor plasmid DNA was examined, using specific hybridization probes. The synthesis of new DNA, associated with the integration of donor moiety, was also monitored. Donor-specific sequences were present at a density intermediate between that of light and hybrid DNA. This recombinant DNA represented 1.4% of total plasmid DNA. The latter value corresponded well with the transforming activity (1.7%) obtained for the donor marker. Newly synthesized material associated with plasmid DNA at the recombinant density amounted to a minor portion of the recombinant plasmid DNA. These data suggest that, like chromosomal transformation, plasmid marker rescue transformation does not require replication for the integration of donor markers and, also like chromosomal transformation, proceeds by a breakage-reunion mechanism. The extent of donor DNA replacement of recipient DNA per plasmid molecule of 54 kilobases (27 kilobase pairs) was estimated as 16 kilobases

  20. Simian virus 40 promoters direct expression of the tetracycline gene in plasmid pACYC184.

    OpenAIRE

    Jenkins, F J; Howett, M K; Rapp, F

    1983-01-01

    Insertion of HindIII DNA fragments into the HindIII site of plasmid pACYC184 destroys the promoter of the plasmid tetracycline resistance gene and causes Escherichia coli cells harboring recombinant plasmids to be tetracycline sensitive and chloramphenicol resistant. The HindIII-C DNA fragment of simian virus 40 contains the two virus promoters and the virus origin of replication. We report the isolation of recombinant plasmids that contained the simian virus 40 HindIII-C DNA fragment at the ...

  1. Plasmid profile in oral Fusobacterium nucleatum from humans and Cebus apella monkeys

    Directory of Open Access Journals (Sweden)

    Paula Marcia O.

    2003-01-01

    Full Text Available Fusobacterium nucleatum is a strict anaerobe and is indigenous of the human oral cavity. This organism is commonly recovered from different monomicrobial and mixed infections in humans and animals. In this study, the plasmid profile, the plasmid stability and the penicillin-resistance association in oral F. nucleatum isolated from periodontal patients, healthy subjects and Cebus apella monkeys were evaluated. Forty-five F. nucleatum strains from patients, 38 from healthy subjects and seven from C. apella were identified and analyzed. Plasmid extraction was performed in all the isolated strains. These elements were found in 26.7% strains from patients and one strain from C. apella. Strains from healthy subjects did not show any plasmid. Most of strains showed two plasmid bands ranging from 4 to 16 Kb, but digestions with endonucleases showed that they belonged to a single plasmid. The plasmid profile was similar and stable in human and monkey strains. Also, plasmids were classified into three groups according to size. Two strains were positive to beta-lactamase production and no plasmid DNA-hybridization with a beta-lactamase gene probe was observed, suggesting a chromosomal resistance.

  2. The stb operon balances the requirements for vegetative stability and conjugative transfer of plasmid R388

    OpenAIRE

    Catherine Guynet; Ana Cuevas; Gabriel Moncalián; Fernando de la Cruz

    2011-01-01

    The conjugative plasmid R388 and a number of other plasmids carry an operon, stbABC, adjacent to the origin of conjugative transfer. We investigated the role of the stbA, stbB, and stbC genes. Deletion of stbA affected both conjugation and stability. It led to a 50-fold increase in R388 transfer frequency, as well as to high plasmid loss. In contrast, deletion of stbB abolished conjugation but provoked no change in plasmid stability. Deletion of stbC showed no effect, neither in conjugation n...

  3. Autoregulation of the stability operon of IncFII plasmid NR1.

    OpenAIRE

    Tabuchi, A; Min, Y N; Womble, D D; Rownd, R H

    1992-01-01

    The stb locus of IncFII plasmid NR1, which mediates stable inheritance of the plasmid, is composed of an essential cis-acting DNA site located upstream from two tandem genes that encode essential stability proteins. The two tandem genes, stbA and stbB, are transcribed as an operon from promoter PAB. Using PAB-lacZ gene fusions, it was found that the stb operon is autoregulated. A low-copy-number stb+ plasmid introduced into the same cell with the PAB-lacZ fusion plasmid repressed beta-galacto...

  4. Genomics of microbial plasmids: classification and identification based on replication and transfer systems and host taxonomy.

    Science.gov (United States)

    Shintani, Masaki; Sanchez, Zoe K; Kimbara, Kazuhide

    2015-01-01

    Plasmids are important "vehicles" for the communication of genetic information between bacteria. The exchange of plasmids transmits pathogenically and environmentally relevant traits to the host bacteria, promoting their rapid evolution and adaptation to various environments. Over the past six decades, a large number of plasmids have been identified and isolated from different microbes. With the revolution of sequencing technology, more than 4600 complete sequences of plasmids found in bacteria, archaea, and eukaryotes have been determined. The classification of a wide variety of plasmids is not only important to understand their features, host ranges, and microbial evolution but is also necessary to effectively use them as genetic tools for microbial engineering. This review summarizes the current situation of the classification of fully sequenced plasmids based on their host taxonomy and their features of replication and conjugative transfer. The majority of the fully sequenced plasmids are found in bacteria in the Proteobacteria, Firmicutes, Spirochaetes, Actinobacteria, Cyanobacteria and Euryarcheota phyla, and key features of each phylum are included. Recent advances in the identification of novel types of plasmids and plasmid transfer by culture-independent methods using samples from natural environments are also discussed. PMID:25873913

  5. Replication of the R6K plasmid during the Escherichia coli cell cycle.

    OpenAIRE

    Keasling, J.D.; Palsson, B O; Cooper, S.

    1992-01-01

    The cell-cycle replication pattern of the R6K plasmid has been investigated by using the membrane-elution technique to produce cells labelled at different times during the division cycle and scintillation counting for quantitative analysis of radioactive plasmid DNA. The high-copy plasmid R6K replicates exponentially in a cell-cycle-independent manner. A mini-R6K plasmid deleted for the ori alpha origin of replication also replicates, exponentially in a cell-cycle-independent manner.

  6. Construction of Recombinant Plasmid Containing S. Mutans F-ATPase β Subunit Gene

    Institute of Scientific and Technical Information of China (English)

    YU Dan-ni; JIANG Li

    2005-01-01

    objective: construct a homologous recombinant plasmid which was expected to be transformed into S. mutans Methods: a region at the 5' terminus of the S. mutans F-ATPase β subunit gene was amplified by PCR, the PCR product was inserted into vector pVA891, yielding recombinant plasmid. Results: the DNA sequence of the recombinant plasmid was identified correct in whole by restriction endonuclease and DNA sequence techniques. Conclusion: the recombinant plasmid of S. mutans DNA was cloned in effect ,it may assist in construction of homologues recombinant mutant.

  7. Effects of medium composition on the production of plasmid DNA vector potentially for human gene therapy

    Institute of Scientific and Technical Information of China (English)

    XU Zhi-nan; SHEN Wen-he; CHEN Hao; CEN Pei-lin

    2005-01-01

    Plasmid vector is increasingly applied to gene therapy or gene vaccine. The production of plasmid pCMV-AP3 for cancer gene therapy was conducted in a modified MBL medium using a recombinant E. coli BL21 system. The effects of different MMBL components on plasmid yield, cell mass and specific plasmid DNA productivity were evaluated on shake-flask scale. The results showed that glucose was the optimal carbon source. High plasmid yield (58.3 mg/L) was obtained when 5.0 g/L glucose was added to MMBL. Glycerol could be chosen as a complementary carbon source because of the highest specific plasmid productivity (37.9 mg DNA/g DCW). After tests of different levels of nitrogen source and inorganic phosphate, a modified MMBL medium was formulated for optimal plasmid production. Further study showed that the initial acetate addition (less than 4.0 g/L) in MMBL improved plasmid production significantly, although it inhibited cell growth. The results will be useful for large-scale plasmid production using recombinant E. coli system.

  8. The 2-micron plasmid as a nonselectable, stable, high copy number yeast vector

    Science.gov (United States)

    Ludwig, D. L.; Bruschi, C. V.

    1991-01-01

    The endogenous 2-microns plasmid of Saccharomyces cerevisiae has been used extensively for the construction of yeast cloning and expression plasmids because it is a native yeast plasmid that is able to be maintained stably in cells at high copy number. Almost invariably, these plasmid constructs, containing some or all 2-microns sequences, exhibit copy number levels lower than 2-microns and are maintained stably only under selective conditions. We were interested in determining if there was a means by which 2-microns could be utilized for vector construction, without forfeiting either copy number or nonselective stability. We identified sites in the 2-microns plasmid that could be used for the insertion of genetic sequences without disrupting 2-microns coding elements and then assessed subsequent plasmid constructs for stability and copy number in vivo. We demonstrate the utility of a previously described 2-microns recombination chimera, pBH-2L, for the manipulation and transformation of 2-microns as a pure yeast plasmid vector. We show that the HpaI site near the STB element in the 2-microns plasmid can be utilized to clone yeast DNA of at least 3.9 kb with no loss of plasmid stability. Additionally, the copy number of these constructs is as high as levels reported for the endogenous 2-microns.

  9. Conjugal transfer and characterization of bacteriocin plasmids in group N (lactic acid) streptococci.

    Science.gov (United States)

    Neve, H; Geis, A; Teuber, M

    1984-01-01

    Thirteen bacteriocin-producing strains of group N (lactic acid) streptococci were screened for their potential to transfer this property by conjugation to Streptococcus lactis subsp. diacetylactis Bu2-60. Bacteriocin production in three strains was plasmid encoded as shown by conjugal transfer and by analysis of cured, bacteriocin-negative derivatives of the donor strains and the transconjugants. With Streptococcus cremoris strains 9B4 and 4G6 and S. lactis subsp. diacetylactis 6F7 as donors, bacteriocin-producing transconjugants were isolated with frequencies ranging from ca. 2 X 10(-2) to 2 X 10(-1) per recipient cell. Bacteriocin-producing transconjugants had acquired a 39.6-megadalton plasmid from the donor strains 9B4 and 4G6, and a 75-megadalton plasmid from the donor strain 6F7. As shown by restriction endonuclease analysis, the plasmids from strains 9B4 and 4G6 were almost identical. The plasmid from strain 6F7 yielded some additional fragments not present in the two other plasmids. In hybridization experiments any of the three plasmids strongly hybridized with each other and with some other bacteriocin but nontransmissible plasmids from other S. cremoris strains. Homology was also detected to a variety of cryptic plasmids in lactic acid streptococci. Images PMID:6321437

  10. Low-dose plasmid DNA treatment increases plasma vasopressin and regulates blood pressure in experimental endotoxemia

    OpenAIRE

    Malardo Thiago; Batalhão Marcelo E; Panunto-Castelo Ademilson; Almeida Luciana P; Padilha Everton; Fontoura Isabela C; Silva Célio L; Carnio Evelin C; Coelho-Castelo Arlete AM

    2012-01-01

    Abstract Background Although plasmid DNA encoding an antigen from pathogens or tumor cells has been widely studied as vaccine, the use of plasmid vector (without insert) as therapeutic agent requires further investigation. Results Here, we showed that plasmid DNA (pcDNA3) at low doses inhibits the production of IL-6 and TNF-α by lipopolysaccharide (LPS)-stimulated macrophage cell line J774. These findings led us to evaluate whether plasmid DNA could act as an anti-inflammatory agent in a Wist...

  11. High-frequency transformation of Brevibacterium lactofermentum protoplasts by plasmid DNA.

    OpenAIRE

    Santamaria, R I; Gil, J.A.; Martin, J. F.

    1985-01-01

    An efficient polyethylene glycol-assisted method for transformation of Brevibacterium lactofermentum protoplasts that uses plasmid vectors has been developed. Two small plasmids, pUL330 (5.2 kilobases) and pUL340 (5.8 kilobases), both containing the kanamycin resistance gene from transposon Tn5 and the replication origin of the natural plasmid pBL1 of B. lactofermentum, were selected as vectors. Supercoiled forms of the plasmids yielded a 100-fold higher transformation frequency than did line...

  12. Plasmids which make their host bacteria mutable as well as resistant to ultraviolet irradiation

    International Nuclear Information System (INIS)

    Some of the naturally occurring Iα, I zeta, M, N, O and T group plasmids increase both the mutability and UV resistance of their host bacteria, while group H and S plasmids only increase mutability. This suggests that these two plasmid-mediated repair functions are separable. The two functions have no direct relation to their restriction-modification systems and nitrofuran resistant functions. In addition, the close linking between the restriction-modification genes and these repair function genes was suggested in group N plasmids. (author)

  13. Relationship between the limited and wide host range octopine-type Ti plasmids of Agrobacterium tumefaciens.

    OpenAIRE

    Thomashow, M F; Knauf, V C; Nester, E W

    1981-01-01

    The relationship between the limited host range octopine Ti plasmids and the wide host range octopine Ti plasmids pTiB6806 and pTiA6 was studied. The limited host range Ti plasmids shared extensive deoxyribonucleic acid homology; pTiAg63 and pTiAg162 were essentially completely homologous with pTiAg158 while pTiAg57 shared approximately 64% homology with pTiAg158. In contrast, the limited host range octopine Ti plasmids only shared 6 to 15% homology with the wide host range octopine Ti plasmi...

  14. Stable maintenance of plasmid in continuous culture of yeast under non-selective conditions.

    Science.gov (United States)

    Gupta, J C; Mukherjee, K J

    2001-01-01

    A recombinant yeast plasmid containing the gene for beta-galactosidase was tested for stability in a host auxotrophic for leucine. Plasmid loss was studied at different dilution rates in continuous culture under selective as well as non-selective conditions. It was observed that the instability of the culture was higher at low dilution rates in selective medium, while the pattern was reversed when complex non-selective medium was used, with plasmid-containing cells competing effectively with plasmid-free cells at low dilution rates. This was attributed to a low residual yeast extract concentration in the medium at low dilution rates. Since yeast extract was the sole source of leucine, this limited the growth of plasmid-free cells, which were auxotrophic for leucine. Growth rate studies also indicated a competitive advantage of the plasmid-containing cells over the plasmid-free cells at low yeast extract concentrations in semi-defined medium. Using the above data, a modified continuous culture was run using non-selective medium at a low dilution rate of 0.05 h(-1). This resulted in stable coexistence of plasmid-containing and plasmid-free cells and hence sustained expression of beta-galactosidase at approximately 330 OD420l(-1) h(-1) throughout the period of cultivation (134 h). PMID:16233104

  15. Determination of plasmid DNA concentration maintained by nonculturable Escherichia coli in marine microcosms.

    OpenAIRE

    Byrd, J J; Leahy, J G; Colwell, R. R.

    1992-01-01

    The concentration of plasmid pBR322 DNA in nonculturable Escherichia coli JM83 was measured to determine whether the plasmid concentration changed during survival of E. coli in marine and estuarine water. E. coli JM83 containing the plasmid pBR322 was placed in both sterile seawater and sterile estuarine water and analyzed for survival (i.e., culturability) and plasmid maintenance. The concentration of pBR322 DNA remained stable in E. coli JM83 for 28 days in an artificial seawater microcosm,...

  16. Repair of UV-irradiated plasmid DNA in excision repair deficient mutants of Saccharomyces cerevisiae

    International Nuclear Information System (INIS)

    The repair of UV-irradiated DNA of plasmid YEp13 was studied in the incision defective strains by measurement of cell transformation frequency. In Saccharomyces cerevisiae, rad1,2,3 and 4 mutants could repair UV-damaged plasmid DNA. In Escherichia coli, uvrA mutant was unable to repair UV-damaged plasmid DNA; however, pretreatment of the plasmid with Micrococcus luteus endonuclease increased repair. It was concluded that all the mutations of yeast were probably limited only to the nuclear DNA. (author)

  17. Staphylococcus aureus and methicillin-resistant S. aureus (MRSA) at ambient freshwater beaches

    Science.gov (United States)

    Fogarty, Lisa R.; Haack, Sheridan K.; Johnson, Heather E.; Brennan, Angela K.; Isaacs, Natasha M.; Spencer, Chelsea

    2015-01-01

    Methicillin-resistant Staphylococcus aureus (MRSA) are a threat to human health worldwide, and although detected at marine beaches, they have been largely unstudied at freshwater beaches. Genes indicating S. aureus (SA; femA) and methicillin resistance (mecA) were detected at 11 and 12 of 13 US Great Lakes beaches and in 18% or 27% of 287 recreational water samples, respectively. Eight beaches had mecA + femA (potential MRSA) detections. During an intensive study, higher bather numbers, staphylococci concentrations, and femA detections were found in samples collected after noon than before noon. Local population density, beach cloud cover, and beach wave height were significantly correlated with SA or MRSA detection frequency. The Panton-Valentine leukocidin gene, associated with community-acquired MRSA, was detected in 12 out of 27 potential MRSA samples. The femA gene was detected less frequently at beaches that met US enterococci criteria or EU enterococci ‘excellent’ recreational water quality, but was not related to Escherichia coli-defined criteria. Escherichia coli is often the only indicator used to determine water quality at US beaches, given the economic and healthcare burden that can be associated with infections caused by SA and MRSA, monitoring of recreational waters for non-fecal bacteria such as staphylococci and/or SA may be warranted.

  18. Staphylococcus aureus small colony variants in diabetic foot infections.

    Science.gov (United States)

    Cervantes-García, Estrella; García-Gonzalez, Rafael; Reyes-Torres, Angélica; Resendiz-Albor, Aldo Arturo; Salazar-Schettino, Paz María

    2015-01-01

    Background : Staphylococcus aureus (S. aureus) is one of the major pathogens causing chronic infections. The ability of S. aureus to acquire resistance to a diverse range of antimicrobial compounds results in limited treatment options, particularly in methicillin-resistant S. aureus (MRSA). A mechanism by which S. aureus develops reduced susceptibility to antimicrobials is through the formation of small colony variants (SCVs). Infections by SCVs of S. aureus are an upcoming problem due to difficulties in laboratory diagnosis and resistance to antimicrobial therapy. Methods : A prospective study was performed on 120 patients diagnosed with both type 2 diabetes mellitus and infected diabetic foot ulcers. The study was carried out from July 2012 to December 2013 in Hospital General de Mexico. The samples were cultured in blood agar, mannitol salt agar, and MacConkey agar media, and incubated at 37°C in aerobic conditions. Results : We describe the first known cases of diabetic foot infections caused by MRSA-SCVs in patients diagnosed with type 2 diabetes mellitus and infected diabetic foot ulcers. In all of our cases, the patients had not received any form of gentamicin therapy. Conclusions : The antibiotic therapy commonly used in diabetic patients with infected diabetic foot ulcers fails in the case of MRSA-SCVs because the intracellular location protects S. aureus-SCVs from the host's defenses and also helps them resist antibiotics. The cases studied in this article add to the spectrum of persistent and relapsing infections attributed to MRSA-SCVs and emphasizes that these variants may also play a relevant role in diabetic foot infections. PMID:25787018

  19. Staphylococcus aureus small colony variants in diabetic foot infections

    Directory of Open Access Journals (Sweden)

    Estrella Cervantes-García

    2015-03-01

    Full Text Available Background: Staphylococcus aureus (S. aureus is one of the major pathogens causing chronic infections. The ability of S. aureus to acquire resistance to a diverse range of antimicrobial compounds results in limited treatment options, particularly in methicillin-resistant S. aureus (MRSA. A mechanism by which S. aureus develops reduced susceptibility to antimicrobials is through the formation of small colony variants (SCVs. Infections by SCVs of S. aureus are an upcoming problem due to difficulties in laboratory diagnosis and resistance to antimicrobial therapy. Methods: A prospective study was performed on 120 patients diagnosed with both type 2 diabetes mellitus and infected diabetic foot ulcers. The study was carried out from July 2012 to December 2013 in Hospital General de Mexico. The samples were cultured in blood agar, mannitol salt agar, and MacConkey agar media, and incubated at 37°C in aerobic conditions. Results: We describe the first known cases of diabetic foot infections caused by MRSA-SCVs in patients diagnosed with type 2 diabetes mellitus and infected diabetic foot ulcers. In all of our cases, the patients had not received any form of gentamicin therapy. Conclusions: The antibiotic therapy commonly used in diabetic patients with infected diabetic foot ulcers fails in the case of MRSA-SCVs because the intracellular location protects S. aureus-SCVs from the host's defenses and also helps them resist antibiotics. The cases studied in this article add to the spectrum of persistent and relapsing infections attributed to MRSA-SCVs and emphasizes that these variants may also play a relevant role in diabetic foot infections.

  20. Mutations in a Partitioning Protein and Altered Chromatin Structure at the Partitioning Locus Prevent Cohesin Recruitment by the Saccharomyces cerevisiae Plasmid and Cause Plasmid Missegregation

    OpenAIRE

    Yang, Xian-Mei; Mehta, Shwetal; Uzri, Dina; Jayaram, Makkuni; Velmurugan, Soundarapandian

    2004-01-01

    The 2μm circle is a highly persistent “selfish” DNA element resident in the Saccharomyces cerevisiae nucleus whose stability approaches that of the chromosomes. The plasmid partitioning system, consisting of two plasmid-encoded proteins, Rep1p and Rep2p, and a cis-acting locus, STB, apparently feeds into the chromosome segregation pathway. The Rep proteins assist the recruitment of the yeast cohesin complex to STB during the S phase, presumably to apportion the replicated plasmid molecules eq...

  1. Complete Nucleotide Sequence of TOL Plasmid pDK1 Provides Evidence for Evolutionary History of IncP-7 Catabolic Plasmids

    OpenAIRE

    Yano, Hirokazu; Miyakoshi, Masatoshi; Ohshima, Kenshiro; Tabata, Michiro; Nagata, Yuji; Hattori, Masahira; Tsuda, Masataka

    2010-01-01

    To understand the mechanisms for structural diversification of Pseudomonas-derived toluene-catabolic (TOL) plasmids, the complete sequence of a self-transmissible plasmid pDK1 with a size of 128,921 bp from Pseudomonas putida HS1 was determined. Comparative analysis revealed that (i) pDK1 consisted of a 75.6-kb IncP-7 plasmid backbone and 53.2-kb accessory gene segments that were bounded by transposon-associated regions, (ii) the genes for conjugative transfer of pDK1 were highly similar to t...

  2. A cohort study of the Copenhagen CF Centre eradication strategy against Staphylococcus aureus in patients with CF

    DEFF Research Database (Denmark)

    Dalbøge, Christina Schjellerup; Pressler, Tacjana; Høiby, Niels;

    2013-01-01

    Staphylococcus aureus is an important pathogen in CF. Centre prevalence of intermittent colonization and chronic S. aureus infections and the effectiveness of an anti-S. aureus eradication strategy was assessed....

  3. Regulation of the pT181 encoded tetracycline resistance gene in Straphylococcus aureus

    International Nuclear Information System (INIS)

    pT181 is a naturally-occurring 4437 basepair (bp) plasmid isolated from Staphylococcus aureus which encodes inducible resistance to tetracycline (Tc). The DNA sequence data has identified three open reading frames (ORFs). The largest ORF B, has been found to be responsible for the Tc resistance phenotype of pT181. Since most Tc resistance systems appear to be regulated by an effector protein and a repressor protein, several Bal 31 deletion mutants of pT181 were constructed and analyzed in an effort to identify the elements involved in Tc resistance. Two transcomplementing groups of mutants were identified within the tet gene. The mechanism of Tc resistance was studied by assaying the accumulation of [7-3H] Tc by Tc sensitive cells, and uninduced and induced pT181-containing cells. A sharp decrease in accumulation of the drug after an initial increase was observed in Tc induced pT181-containing cells. In vivo labeling of Bacillus subtilis minicells containing pT181 was performed with 35S-methionine to identify the polypeptide product of the tet gene. A Tc-inducible protein having a molecular weight of approximately 50,000 daltons was detected only in B. subtilis minicells carrying pT181. Cell fractionation studies of S. aureus cells with and without pT181 showed that an approximately 28,000 daltons Tc-inducible protein was present in membranes of pT181 containing cells. The amount of TET protein in Tc induced minicells was about fifteen-fold higher than that in uninduced minicells. RNA prepared from stationary phase cells analyzed by Northern blot hybridization showed that the steady-state level of the tet mRNA in induced pT181-containing cells was bout four-fold higher than that in uninduced pT181-containing cells. When RNA synthesis was blocked with rifampicin, tet mRAN was found to be much more stable in Tc induced cells as compared to that in uninduced cells over a 30 min period

  4. Analysis of plasmid DNA synthesis by double tracer method

    International Nuclear Information System (INIS)

    An Escherichia coli strain, CR34, harboring both pSC101 and ColEl-amp plasmids was exposed to media containing rifampicin (100 μg/ml) and/or chloramphenicol (180 μg/ml) and the cells were labeled for 20 min with 3H-thymine at 3,25 and 50 min after exposure to drug(s). The plasmid DNA synthesis was assayed by DNA-DNA hybridization with 14C-labeled pSC134 DNA as internal marker. In the presence of rifampicin, the replication of pSC 101 was from 57 to 104% that in its absence, and that of ColEl-amp was from 17 to 26%. The DNA replication of pSC101 after addition of chloramphenicol was reduced to 35 to 75%, and that of ColEl-amp was reduced to 39% and then restored to 92%. This restoration was not observed in the presence of rifampicin. (author)

  5. Rapid and inexpensive method for isolating plasmid DNA

    International Nuclear Information System (INIS)

    A small-scale and economical method for isolating plasmid DNA from bacteria is described. The method provides DNA of suitable quality for most DNA manipulation techniques. This DNA can be used for restriction endonuclease digestion, southern blot hybridization, nick translation and end labeling of DNA probes, Polymerase Chain Reaction (PCR) -based techniques, transformation, DNA cycle-sequencing, and Chain-termination method for DNA sequencing. The entire procedure is adapted to 1.5 ml microfuge tubes and takes approximately 30 mins. The DNA isolated by this method has the same purity produced by CTAB and cesium chloride precipitation and purification procedures respectively. The two previous methods require many hours to obtain the final product and require the use of very expensive equipment as ultracentrifuge. This method is well suited for the isolation of plasmid DNA from a large number of bacterial samples and in a very short time and low cost in laboratories where chemicals, expensive equipment and finance are limited factors in conducting molecular research. (authors). 11refs. 11refs

  6. Germline Competent Pluripotent Mouse Stem Cells Generated by Plasmid Vectors.

    Science.gov (United States)

    Chen, Chien-Hong; Su, Yu-Hsiu; Lee, Kun-Hsiung; Chuang, Chin-Kai

    2016-07-01

    We developed nonintegrated methods to reprogram mouse embryonic fibroblast (MEF) cells into induced pluripotent stem cells (iPSCs) using pig pOct4, pSox2, and pc-Myc as well as human hKLF4, hAID, and hTDG that were carried by plasmid vectors. The 4F method employed pOct4, pSox2, pc-Myc, and hKLF4 to derive iPSC clones with naive embryonic stem cell (ESC)-like morphology. These 4F clones expressed endogenous mouse Nanog protein and could generate chimeras. In addition to the four conventional reprogramming factors used in the 4F method, hAID and hTDG were utilized in a 6F method to increase the conversion efficiency of reprogramming by approximately five-fold. One of the 6F plasmid derived iPSC (piPSC) clones was shown to be germline transmission competent. PMID:26980563

  7. Radioimmunoassays for protein A of Staphylococcus aureus

    International Nuclear Information System (INIS)

    Radioimmunoassays have been developed that can detect nanogram amounts of protein A (SpA), a product generated by Staphylococcus aureus that binds selectively to the Fc region of IgG from most mammalian species. Competition assays for fluid phase SpA utilize antibodies produced in chickens, 125I-labeled SpA as the tracer molecule, and either F(ab')2 fragments of rabbit IgG anti-chicken IgG or 40% ammonium sulfate as the precipitating agent to separate antigen-antibody complexes from free antigen. The double antibody assay could be carried out in serum from species that form only soluble complexes with SpA (e.g., rabbit), that react poorly with SpA (e.g., rat) or under appropriate conditions in serum from species (e.g., dog) that show high reactivity with SpA and form precipitating complexes. Chicken antibodies prepared by affinity chromatography on SpA-Sepharose and labeled with 125I were used in a direct binding assay for SpA present either on the cell wall of Cowan strain I or Wood 46 bacteria, in insoluble complexes prepared from SpA and whole serum or purified IgG, or in C1q binding complexes that were formed by passage of serum from normal or tumor bearing humans or dogs over SpA-collodion charcoal. Since both types of assays could detect SpA even in the presence of serum or IgG, they offer advantages over other techniques in which the SpA-Fc interaction may interfere. (Auth.)

  8. A systematic review of animal models for Staphylococcus aureus osteomyelitis

    Directory of Open Access Journals (Sweden)

    W Reizner

    2014-03-01

    Full Text Available Staphylococcus aureus (S. aureus osteomyelitis is a significant complication for orthopaedic patients undergoing surgery, particularly with fracture fixation and arthroplasty. Given the difficulty in studying S. aureus infections in human subjects, animal models serve an integral role in exploring the pathogenesis of osteomyelitis, and aid in determining the efficacy of prophylactic and therapeutic treatments. Animal models should mimic the clinical scenarios seen in patients as closely as possible to permit the experimental results to be translated to the corresponding clinical care. To help understand existing animal models of S. aureus, we conducted a systematic search of PubMed and Ovid MEDLINE to identify in vivo animal experiments that have investigated the management of S. aureus osteomyelitis in the context of fractures and metallic implants. In this review, experimental studies are categorised by animal species and are further classified by the setting of the infection. Study methods are summarised and the relevant advantages and disadvantages of each species and model are discussed. While no ideal animal model exists, the understanding of a model’s strengths and limitations should assist clinicians and researchers to appropriately select an animal model to translate the conclusions to the clinical setting.

  9. Methicillin-resistant Staphylococcus aureus in central Iowa wildlife.

    Science.gov (United States)

    Wardyn, Shylo E; Kauffman, Lin K; Smith, Tara C

    2012-10-01

    Livestock and pets have been identified as carriers of Staphylococcus aureus; however, the role of wild animals as a reservoir of S. aureus strains has not yet been examined. We conducted a pilot study to determine the prevalence of methicillin-sensitive S. aureus (MSSA) and methicillin-resistant S. aureus (MRSA) in 37 species of wild animals rehabilitated at a university clinic. Nasal, wing, wound, and cloacal swabs were collected. Of 114 animals, seven (6.1%) were MSSA-positive and three (2.6%) were MRSA-positive. The MRSA isolates were obtained from two eastern cottontail rabbits (Sylvilagus floridanus) and a Lesser Yellowlegs (Tringa flavipes), a migratory shorebird. Antibiotic resistance testing of the MRSA isolates revealed that two were additionally resistant to tetracycline and erythromycin, and the third isolate was also resistant to erythromycin, clindamycin, and levofloxacin. All three isolates were positive for the Panton-Valentine leukocidin (PVL) gene. Sequence typing of the staphylococcal protein A (spa) region revealed one MRSA isolate to be t002, whereas the other two MRSA isolates were found to be t008. Our results suggest that S. aureus, including MRSA, is being carried by wild animals, although at a low prevalence with the limited number of animals tested. Additional studies are needed to determine how this may impact human health. PMID:23060511

  10. Crystal Violet and XTT Assays on Staphylococcus aureus Biofilm Quantification.

    Science.gov (United States)

    Xu, Zhenbo; Liang, Yanrui; Lin, Shiqi; Chen, Dingqiang; Li, Bing; Li, Lin; Deng, Yang

    2016-10-01

    Staphylococcus aureus (S. Aureus) is a common food-borne pathogenic microorganism. Biofilm formation remains the major obstruction for bacterial elimination. The study aims at providing a basis for determining S. aureus biofilm formation. 257 clinical samples of S. aureus isolates were identified by routine analysis and multiplex PCR detection and found to contain 227 MRSA, 16 MSSA, 11 MRCNS, and 3 MSCNS strains. Two assays for quantification of S. aureus biofilm formation, the crystal violet (CV) assay and the XTT (tetrazolium salt reduction) assay, were optimized, evaluated, and further compared. In CV assay, most isolates formed weak biofilm 74.3 %), while the rest formed moderate biofilm (23.3 %) or strong biofilm (2.3 %). However, most isolates in XTT assay showed weak metabolic activity (77.0 %), while the rest showed moderate metabolic activity (17.9 %) or high metabolic activity (5.1 %). In this study, we found a distinct strain-to-strain dissimilarity in terms of both biomass formation and metabolic activity, and it was concluded from this study that two assays were mutual complementation rather than being comparison. PMID:27324342

  11. Enterotoxin production by Staphylococcus aureus isolated from mastitic cows.

    Science.gov (United States)

    Cenci-Goga, B T; Karama, M; Rossitto, P V; Morgante, R A; Cullor, J S

    2003-09-01

    Staphylococcus aureus is an important cause of mastitis in cows. The ability of S. aureus strains to produce one or more enterotoxins in milk and dairy products is linked to staphylococcal food poisoning. To determine whether staphylococci causing bovine mastitis could cause human foodborne intoxication, the production of staphylococcal enterotoxins A through D (SEA, SEB, SEC, and SED) by 160 S. aureus isolates was evaluated with the use of a reverse passive latex agglutination enterotoxin kit. All S. aureus strains were isolated over a 9-month period from 2,343 routine submissions of a composite quarter collection of individual mastitic cows at 18 dairy farms in the San Joaquin Valley in California. Prior to enterotoxin detection, isolates were grown by a method that enhances the in vitro synthesis of enterotoxin. Twenty-two of 160 S. aureus isolates produced enterotoxin. Seven produced SEC, 12 produced SED, and 3 produced both SEC and SED. None of the isolates produced SEA or SEB. PMID:14503727

  12. Prevalence of Salmonella and Staphylococcus aureus in chorizo and longaniza

    Directory of Open Access Journals (Sweden)

    Refugio Torres-Vitela

    2011-12-01

    Full Text Available Epidemiological research in developed and developing countries, had found meat products as the principal cause for foodbourne diseases. In addition, Salmonella and Staphyococcus aureus are well known pathogens for their mayor impact in public health. The objective for the present study consisted on determinate the sanitary quality from chorizo and longaniza samples from several butcheries in Guadalajara, Jalisco, Mexico. Samples of chorizo (50 and longaniza (50 were obtained from different points in Guadalajara metropolis. Presence of Salmonella and recounts for S. aureus were tested in 25 g samples. Procedure was followed according Mexican NOM 145-SSA1-1995 methods. In chorizo, 18 samples were positive to Salmonella. The count of S. aureus showed a mean of 24,600 UFC/g. On the other hand, 24 samples of longaniza were positive to Salmonella spp. In this case, the mean of S. aureus was 7,800 UFC/g. The serotypes of Salmonella spp were: Derby (30%, Adelaile (17%, Azteca (15%, Infantis (15%, Muenster(10% y Anatum (13 %. The high positivity of Salmonella spp. and S. aureus is a potential hazard to consumers.

  13. Novel assay to measure the plasmid mobilizing potential of mixed microbial communities

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    Uli eKlümper

    2014-12-01

    Full Text Available Mobilizable plasmids lack necessary genes for complete conjugation and are therefore non-self-transmissible. Instead, they rely on the conjugation system of conjugal plasmids to be horizontally transferred to new recipients. While community permissiveness, the fraction of a mixed microbial community that can receive self-transmissible conjugal plasmids, has been studied, the intrinsic ability of a community to mobilize plasmids that lack conjugation systems is unexplored. Here, we present a novel framework and experimental method to estimate the mobilization potential of mixed communities. We compare the transfer frequency of a mobilizable plasmid to that of a mobilizing and conjugal plasmid measured for a model strain and for the assayed community. With Pseudomonas putida carrying the gfp-tagged mobilizable RSF1010 plasmid as donor strain, we conducted solid surface mating experiments with either a P. putida strain carrying the mobilizing plasmid RP4 or a model bacterial community that was extracted from the inner walls of a domestic shower conduit. Additionally, we estimated the permissiveness of the same community for RP4 using P. putida as donor strain. The permissiveness of the model community for RP4 (at 1.16x10-4 transconjugants per recipient (T/R was similar to that previously measured for soil microbial communities. RSF1010 was mobilized by the model community at a frequency of 1.16x10-5 T/R, only one order of magnitude lower than its permissiveness to RP4. This mobilization frequency is unexpectedly high considering that (i mobilization requires the presence of mobilizing conjugal plasmids within the permissive fraction of the recipients; (ii in pure culture experiments with P. putida retromobilization of RSF1010 through RP4 only took place in approximately half of the donors receiving the conjugal plasmid in the first step. Further work is needed to establish how plasmid mobilization potential varies within and across microbial

  14. Structural and replicative diversity of large plasmids from sphingomonads that degrade polycyclic aromatic compounds and xenobiotics.

    Science.gov (United States)

    Basta, Tamara; Buerger, Sibylle; Stolz, Andreas

    2005-06-01

    The plasmids from 16 sphingomonads which degrade various xenobiotics and polycyclic aromatic compounds were compared with the previously sequenced plasmid pNL1 from Sphingomonas aromaticivorans F199. The replicase genes repAaAb from plasmid pNL1 were amplified by PCR and used as a gene probe for the identification of plasmids belonging to the same incompatibility group as plasmid pNL1. Plasmids were prepared from various sphingomonads and hybridized with the repA gene probe. Positive hybridization signals were obtained with plasmids of approximately 160-195 kb from Sphingomonas subterranea and S. aromaticivorans B0695, which had been isolated from the same subsurface location as S. aromaticivorans F199. The repA probe also hybridized with plasmids from Sphingomonas xenophaga BN6, Sphingomonas sp. HH69 and Sphingomonas macrogoltabidus, which had been isolated from different continents and which utilize different organic compounds than S. aromaticivorans F199 and the other subsurface strains. The results of the hybridization experiments were confirmed by PCR experiments using primers deduced from the repAaAb region of plasmid pNL1. Nucleotide sequence comparisons suggested that three gene clusters were conserved between plasmid pNL1 and plasmid pBN6 from the naphthalenesulfonate- degrading strain S. xenophaga BN6. From these sequence comparisons, PCR primers were derived in order to detect the respective gene clusters in the other strains and to deduce their position relative to each other. These experiments demonstrated that all analysed subsurface strains harboured the same three gene clusters, but that the position and distance from each other of the clusters varied considerably among the different strains. PMID:15942009

  15. Molecular and population analyses of a recombination event in the catabolic plasmid pJP4.

    Science.gov (United States)

    Larraín-Linton, Juanita; De la Iglesia, Rodrigo; Melo, Francisco; González, Bernardo

    2006-10-01

    Cupriavidus necator JMP134(pJP4) harbors a catabolic plasmid, pJP4, which confers the ability to grow on chloroaromatic compounds. Repeated growth on 3-chlorobenzoate (3-CB) results in selection of a recombinant strain, which degrades 3-CB better but no longer grows on 2,4-dichlorophenoxyacetate (2,4-D). We have previously proposed that this phenotype is due to a double homologous recombination event between inverted repeats of the multicopies of this plasmid within the cell. One recombinant form of this plasmid (pJP4-F3) explains this phenotype, since it harbors two copies of the chlorocatechol degradation tfd gene clusters, which are essential to grow on 3-CB, but has lost the tfdA gene, encoding the first step in degradation of 2,4-D. The other recombinant plasmid (pJP4-FM) should harbor two copies of the tfdA gene but no copies of the tfd gene clusters. A molecular analysis using a multiplex PCR approach to distinguish the wild-type plasmid pJP4 from its two recombinant forms, was carried out. Expected PCR products confirming this recombination model were found and sequenced. Few recombinant plasmid forms in cultures grown in several carbon sources were detected. Kinetic studies indicated that cells containing the recombinant plasmid pJP4-FM were not selectable by sole carbon source growth pressure, whereas those cells harboring recombinant plasmid pJP4-F3 were selected upon growth on 3-CB. After 12 days of repeated growth on 3-CB, the complete plasmid population in C. necator JMP134 apparently corresponds to this form. However, wild-type plasmid forms could be recovered after growing this culture on 2,4-D, indicating that different plasmid forms can be found in C. necator JMP134 at the population level. PMID:16980481

  16. Induced mutagenesis of plasmid and chromosomal genes inserted into the plasmid DNA. II. Mutagenic action of chemical factors

    International Nuclear Information System (INIS)

    Following the study of the mutagenic action of UV and γ-radiation on plasmid DNA in vitro, they investigated the induction of mutations under the influence of chemical mutagens on the same DNA of plasmid RSF2124, determining the synthesis of colicine E1 and resistance to ampicillin. The inactivating action of the mutagen was assessed from the yield of transformants resistant to the antibiotic and the mutagenic effect from the loss by colonies of transformants that were capable of releasing colicine into the external medium. In these experiments they mainly used chemical compounds whose mutagenic effect if well known in other systems (transforming and transfecting DNA, microbial viruses). As a result all mutagens tested for their activity were divided into four groups: first group, those exceeding the level of mutagenesis by more than 100-fold above the spontaneous background (hydroxylamine, O-methylhydroxylamine); second group, those exceeding it by a factor of 10 (UV radiation (λ = 254 nm), W-mutagenesis, ionizing radiation, nitrous acid, mitomycin C); third group, those exceeding it by a factor of <10 (indirect UV mutagenesis, nitrous acid, β-chloroethyldiethylamine hydrochloride, nitrosoguanidine); fourth group, no mutagenic effect (acridine orange, ethyl methane sulfonate, sodium azide, 0-β-diethylaminoethylhydroxylamine)

  17. Introduction of temperature-sensitive helper and donor plasmids into Bac-to-Bac baculovirus expression systems

    Institute of Scientific and Technical Information of China (English)

    Zhihong; Huang; Ao; Li; Mengjia; Pan; Wenbi; Wu; Meijin; Yuan; Kai; Yang

    2015-01-01

    In the baculovirus shuttle vector(bacmid) system, a helper plasmid and a donor plasmid are employed to insert heterologous genes into a cloned baculovirus genome via Tn7 transposition in Escherichia coli. The helper and donor plasmids are usually cotransfected with constructed bacmids into insect cells, which will lead to integration of these plasmids into the viral genome,and hence to the production of defective virions. In this study, to facilitate the preparation of plasmid-free recombinant bacmids, we modified a set of helper and donor plasmids by replacing their replication origins with that of a temperature-sensitive(ts) plasmid, p SIM6. Using the resulting ts helper plasmid p MON7124 ts and the ts donor plasmid p FB1ts-PH-GFP, a recombinant bacmid,b Ac WT-PG(-), was constructed, and the transposition efficiency was found to be 33.1%. The plasmids were then removed by culturing at 37 °C. For b Ac WT-PG(-), the infectious progeny virus titer and the protein expression level under the control of the polyhedrin promoter were similar to those of a bacmid constructed with unmodified helper and donor plasmids. These ts plasmids will be useful for obtaining plasmid-free bacmids for both heterologous protein production and fundamental studies of baculovirus biology.

  18. Occurrence of Multidrug Resistant Staphylococcus aureus in horses in Malaysia

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    Z. Zunita

    2008-12-01

    Full Text Available A total of 22 Staphylococcus aureus were isolated from 50 samples from 8 stable horses. They are positive in the catalase and coagulase tests. Upon testing the cultures with SLIDEX test kit all formed agglutination within a few seconds, confirming they are of S. aureus. When cultured onto MSA, all isolates formed yellow colonies. However, none of the isolates produced blue colonies on ORSAB indicating that there were no MRSA among the S. aureus. There were 13 isolates which were multiresistant. Eleven are resistant to eight out of ten antibiotics tested. All these isolates were found to originate from stable G. One isolate is resistant to 5 antibiotics while another one isolate is resistant to 3 antibiotics. The rest of the isolates are not multiresistant to the antibiotics tested. [Veterinary World 2008; 1(6.000: 165-167

  19. Risk factors for Staphylococcus aureus nasal colonization in Danish middle-aged and elderly twins

    DEFF Research Database (Denmark)

    Andersen, P S; Larsen, Lisbeth Aagaard; Fowler, V G;

    2013-01-01

    Staphylococcus aureus is a human commensal bacterium found in the nasal cavity and other body sites. Identifying risk factors for S. aureus nasal carriage is of interest, as nasal carriage is a risk factor for subsequent invasive infection. We recently investigated the influence of host genetics...... on S. aureus carriage in Danish middle-aged and elderly twins, which indicated no significant heritability that could account for the observed S. aureus carriage. In the present study, we performed a questionnaire-based study of S. aureus colonization on the same cohort of 2,196 Danish middle......-aged and elderly twins to identify specific risk factors for S. aureus nasal colonization, including analyzing the paired twins (n = 478) that were discordant for S. aureus colonization. We found associations between risk factors and S. aureus nasal colonization among middle-aged and elderly twins, including age...

  20. Draft Genome Sequence of Methicillin-Sensitive Staphylococcus aureus ATCC 29213

    OpenAIRE

    Soni, Isha; Chakrapani, Harinath; Chopra, Sidharth

    2015-01-01

    Staphylococcus aureus subsp. aureus ATCC 29213 is one of the most commonly used strains in drug discovery research and for quality control. We report the completed draft genome sequence for the strain.