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At least 343 records · Page 19Linked to original sources
PubMed1984

Intraphagocytic bactericidal activity of bacterial DNA gyrase inhibitors against Serratia marcescens.

Addition of phenylbutazone (2 mg/ml) to 55 vol% of fresh defibrinated human blood permitted leukocytic ingestion of Serratia marcescens bacteria, but blocked phagocytic killing activity. The group A (phage tail) bacteriocin bA+ 16 served to kill extraphagocytic test bacteria. Five bacterial DNA gyrase inhibitors (ciprofloxacin, enoxacin, norfloxacin, nalidixic acid and pipemidic acid) revealed intraphagocytic bactericidal activity against ingested S. marcescens bacteria of three test strains; the extent of intraphagocytic activity of these drugs approximated that of rifampin. Following combination with unmodified defibrinated human blood (55 vol%), ciprofloxacin, enoxacin, norfloxacin and pipemidic acid yielded additive effects against 3 assay strains of S. marcescens and Escherichia coli strain ATCC 25922; nalidixic acid was inferior in this regard. These data might guide chemotherapy of parenchymatous urinary tract infections.

Blood Bactericidal Activity↗
PubMed1989

Tracking bacterial DNA replication forks in vivo by pulsed field gel electrophoresis.

The location of chromosomal DNA replication forks was identified in synchronously replicating E. coli cultures by pulse labeling DNA at specific times with 14C-thymidine and following incorporation of radionucleotide into genomic Not I restriction fragments. This technique could be used to characterize chromosomal DNA replication, to characterize mutations which affect this process, to identify the location of DNA replication origins and termini as well as aid in the construction of macrorestriction maps. Here, we further characterize the DNA replication mutations divE and dnaK and preliminary characterize the genomic organization of E. coli isolate 15.

Chromosome Mapping↗
PubMed1993

Cloning of the large subunit of activator 1 (replication factor C) reveals homology with bacterial DNA ligases.

We have cloned a gene encoding a DNA-binding protein by Southwestern screening of a murine cDNA library with a double-stranded oligonucleotide containing the sequence from the bidirectional promoter of the alpha 1 and alpha 2 collagen IV genes. The middle portion of this 1131-amino acid protein has a region homologous to bacterial DNA ligases, and the more carboxyl portion contains several domains homologous to p40, p38, p37, and p36.5 subunits of activator 1 (A1, also called replication factor C), a human replication protein complex. Western blotting revealed that antiserum generated against part of the recombinant protein reacted specifically with the 145-kDa component of the purified human A1 complex, indicating that it is the murine counterpart of the A1 p145. Characterization of the DNA-binding activity of the recombinant fusion protein by gel mobility-shift assay revealed that it had a preference for a run of pyrimidines on one strand. Deletion analysis using recombinant proteins revealed that the DNA ligase-like domain was required for DNA-binding activity. The finding that the region required for the binding of murine A1 p145 to DNA has similarity to a domain found in DNA ligases suggests that this region may be utilized by both proteins in recognizing DNA.

Amino Acid Sequence↗
PubMed2003

Isolation and structure elucidation of Sch 642305, a novel bacterial DNA primase inhibitor produced by Penicillium verrucosum.

A novel primase inhibitor, Sch 642305 (1), was isolated from the fermentation broth of the fungal culture Penicillium verrucosum. The structure of 1 was elucidated on the basis of MS and NMR spectroscopic data as a new and unusual bicyclic 10-membered macrolide. The absolute configuration of the asymmetric centers was determined by X-ray crystallographic analysis of the p-bromobenzoate derivative (3). Compound 1 exhibited inhibitory activity against bacterial DNA primase enzyme with an EC(50) of 70 microM.

Arizona↗
PubMed2003

Real-time quantitative PCR assay on bacterial DNA: In a model soil system and environmental samples.

Real-time quantitative PCR (RTQ-PCR) was used to quantify the bacterial target DNA extracted by three commonly used DNA extraction protocols (bead mill homogenization, grinding in presence of liquid nitrogen and hot detergent SDS based enzymatic lysis). For the purpose of our study, pure culture of Bacillus cereus (model organism), sterilized soil seeded with a known amount of B. cereus (model soil system) and samples from woodland and grassland (environmental samples) were chosen to extract DNA by three different protocols. The extracted DNA was then quantified by RTQ-PCR using 16S rDNA specific universal bacterial primers. The standard curve used for the quantification by RTQ-PCR was linear and revealed a strong linear relationship (r(2)=0.9968) with a higher amplification efficiency, e5=1.02. High resolution gel electrophoresis was also carried out to observe the effect of these extraction methods on diversity analysis. For the model soil system, the liquid nitrogen method showed the highest target DNA copy number (1.3 x 10(9) copies/microl). However, for both the environmental samples, the bead beating method was found to be suitable on the basis of the high target DNA copy numbers (5.38 x 10(9) and 4.01 x 10(8) copies/ml for woodland and grassland respectively), high yield (6.4 microg/g and 1.76 microg/g of soil for woodland and grassland respectively) and different band patterns on high resolution gel electrophoresis suggesting an overall high extraction efficiency. This difference in the extraction efficiency between the model soil system and environmental samples may be attributed to different affinity of seeded and native DNA to soil particles.

Bacillus cereus↗
PubMed2002

Meningococcal bacterial DNA load at presentation correlates with disease severity.

AIMS: To determine bacterial loads in meningococcal disease (MCD), their relation with disease severity, and the factors which determine bacterial load. METHODS: Meningococcal DNA quantification was performed by the Taqman PCR method on admission and sequential blood samples from patients with MCD. Disease severity was assessed using the Glasgow Septicaemia Prognostic Score (GMSPS, range 0-15, severe disease > or =8). RESULTS: Median admission bacterial load was 1.6 x 10(6) DNA copies/ml of blood (range 2.2 x 10(4) to 1.6 x 10(8)). Bacterial load was significantly higher in patients with severe (8.4 x 10(6)) compared to milder disease (1.1 x 10(6), p = 0.018). This difference was greater in septicaemic patients (median 1.6 x 10(7) versus 9.2 x 10(5), p < 0.001). Bacterial loads were significantly higher in patients that died (p = 0.017). Admission bacterial load was independent of the duration of clinical symptoms prior to admission, with no difference between the duration of symptoms in mild or severe cases (median, 10.5 and 11 hours respectively). Bacterial loads were independent of DNA elimination rates following treatment. CONCLUSION: Patients with MCD have higher bacterial loads than previously determined with quantitative culture methods. Admission bacterial load is significantly higher in patients with severe disease (GMSPS > or =8) and maximum load is highest in those who die. Bacterial load is independent of the duration of clinical symptoms or the decline in DNA load.

Bacteremia↗
PubMed1999

The kinetoplast structure-specific endonuclease I is related to the 5' exo/endonuclease domain of bacterial DNA polymerase I and colocalizes with the kinetoplast topoisomerase II and DNA polymerase beta during replication.

The mitochondrial DNA (kinetoplast DNA) of the trypanosomatid Crithidia fasciculata has an unusual structure composed of minicircles and maxicircles topologically interlocked into a single network and organized in a disc-shaped structure at the base of the flagellum. We previously purified a structure-specific endonuclease (SSE1), based on its RNase H activity, that is enriched in isolated kinetoplasts. The endonuclease gene has now been cloned, sequenced, and found to be closely related to the 5' exonuclease domain of bacterial DNA polymerase I proteins. Although the protein does not contain a typical mitochondrial leader sequence, the enzyme is shown to colocalize with a type II DNA topoisomerase and a DNA polymerase beta at antipodal sites flanking the kinetoplast disc. Cell synchronization studies with an epitope-tagged construct show that the localization of the endonuclease to the antipodal sites varies in a cell cycle-dependent manner similar to that of the DNA polymerase beta [Johnson, C. E. & Englund, P. T. (1998) J. Cell Biol. 143, 911-919]. Immunofluorescent localization of SSE1 to the antipodal sites is only observed during kinetoplast replication. Together, these results suggest a point of control for kinetoplast DNA replication through the regulation of the availability of DNA replication proteins and a possible role for the antipodal sites in removal of RNA primers and the repair of gaps in newly replicated minicircles.

Amino Acid Sequence↗
PubMed1995

Cloning of the murine cDNA encoding VDJP, a protein homologous to the large subunit of replication factor C and bacterial DNA ligases.

A putative full-length 1.7-kb cDNA, encoding a murine protein that specifically binds to the nonamer portion of the V(D)J recombinational signal sequence (RSS) element, has been cloned. By its sequence analysis, this cDNA is identical to a portion of the 4.5-kb murine replication factor C large-subunit-encoding cDNA. By Northern blot analysis, the 1.7-kb mRNA species is observed in murine immature B cells but not in non-lymphoid cells and tissues, while the 4.5-kb replication factor C-encoding cDNA is expressed in all cell types. The deduced VDJP amino-acid sequence includes a region of homology with bacterial DNA ligases at the C terminus of each of the proteins. VDJP has been synthesized as a fusion protein in bacteria, and the purified protein has been previously shown to mediate the joining of DNA fragments in a V(D)J RSS-dependent fashion (Guilliams et al., Biochem. Biophys. Res. Commun. 202 (1994) 1134-1141).

Amino Acid Sequence↗
PubMed1996

Absence of bacterial DNA in the bladder of patients with interstitial cystitis.

PURPOSE: Although bacterial infection has been long considered a possible cause of interstitial cystitis (IC), no definitive proof for or against this hypothesis has been presented so far. We have used 16S rDNA bacterial polymerase chain reaction to study bladder biopsies and sterile urine samples from patients suffering from IC. This method is sensitive and detects all known eubacteria. MATERIALS AND METHODS: Bladder biopsies and sterile urine samples obtained by transabdominal puncture were studied from 11 patients with IC. As controls we studied 4 patients with other urological problems leading to partly similar symptoms and 5 healthy individuals. RESULTS: All samples from the IC patients were negative. One positive sample was obtained from a woman with a history of urinary tract infections who suffered from nonIC ulcerative cystitis. Her sterile urine sample yielded Lactobacillus acidophilus. CONCLUSION: These results indicate that an ongoing bacterial infection is not the cause of interstitial cystitis.

Cystitis, Interstitial↗
PubMed2001

CpG motifs in bacterial DNA and their immune effects.

Unmethylated CpG motifs are prevalent in bacterial but not vertebrate genomic DNAs. Oligodeoxynucleotides (ODN) containing CpG motifs activate host defense mechanisms leading to innate and acquired immune responses. The recognition of CpG motifs requires Toll-like receptor (TLR) 9, which triggers alterations in cellular redox balance and the induction of cell signaling pathways including the mitogen activated protein kinases (MAPKs) and NF kappa B. Cells that express TLR-9, which include plasmacytoid dendritic cells (PDCs) and B cells, produce Th1-like proinflammatory cytokines, interferons, and chemokines. Certain CpG motifs (CpG-A) are especially potent at activating NK cells and inducing IFN-alpha production by PDCs, while other motifs (CpG-B) are especially potent B cell activators. CpG-induced activation of innate immunity protects against lethal challenge with a wide variety of pathogens, and has therapeutic activity in murine models of cancer and allergy. CpG ODN also enhance the development of acquired immune responses for prophylactic and therapeutic vaccination.

Animals↗
PubMed1996

Rapid preparation of bacterial DNA for pulsed-field gel electrophoresis.

A disadvantage of genotyping bacterial strains by pulsed-field gel electrophoresis is that the procedure requires up to 6 days to complete. We modified a standard pulsed-field gel electrophoresis method (B.E. Murray, K.V. Singh, J.D. Health, B.R. Sharma, and G.M. Weinstock, J.Clin. Microbiol. 28:2059-2063, 1990) so that it could be completed in less than 3 days. We successfully applied this method to the analysis of a variety of gram-positive and gram-negative bacteria.

DNA, Bacterial↗
PubMed1996

Histone-like protein HU and bacterial DNA topology: suppression of an HU deficiency by gyrase mutations.

The abundant bacterial protein called HU has the ability to wrap and bend DNA in vitro, and thus it has long been thought to play a role in DNA supercoiling. In the absence of HU, Escherichia coli formed tiny colonies on agar, rapidly accumulated suppressor mutations, and was hypersensitive to novobiocin. Three types of evidence implicated gyrase in the suppression of an HU deficiency. First, spontaneous suppressors that restored normal growth and reduced sensitivity to novobiocin mapped in gyrB, one of the genes encoding DNA gyrase. Second, a pair of known gyrB mutations (gyrB-203 Ts gyrB-221 NovR) allowed normal growth at permissive (30 degrees C) but not at intermediate (37 degrees C) conditions. Third, introduction of a gyrB-expressing plasmid restored normal colony size. DNA supercoiling comparisons showed that chromosomal supercoiling decreased in the absence of HU and increased toward wild-type levels in the presence of a spontaneous gyrB suppressor. Taken together, these data establish that HU has a physiological role in chromosomal DNA topology, probably by facilitating the action of gyrase.

Alleles↗
PubMed1987

Bacterial DNA reactivity of the novel antitumor antibiotics PD 114,759 and PD 115,028 (veractamycins A and B).

The novel antitumor antibiotics PD 114,759 and PD 115,028 were evaluated for their ability to cause repairable DNA damage and the induction of SOS functions in bacterial systems. PD 114,759 and PD 115,028 were preferentially toxic to DNA repair-defective Escherichia coli WP100 uvrA recA in comparison to wild-type E. coli WP2 at concentrations of 10 approximately 30 micrograms/ml in agar diffusion assays. Both compounds were inducers of cell filamentation and prophage lambda (two E. coli SOS functions) at concentrations of 0.1 approximately 1 microgram/ml. In addition, the ability of PD 114,759 and PD 115,028 to retain their filamentation-inducing effects under both aerobic conditions and anaerobic conditions suggests that a bioreductive, rather than an oxygen-requiring, mechanism is involved in the DNA-reactive effects of these agents.

Antibiotics, Antineoplastic↗