Three unlinked gene clusters are involved in clavam metabolite biosynthesis in Streptomyces clavuligerus

2004 ◽  
Vol 50 (10) ◽  
pp. 803-810 ◽  
Author(s):  
Kapil Tahlan ◽  
Hyeon Ung Park ◽  
Susan E Jensen
Keyword(s):  
Clavulanic Acid ◽  
Gel Electrophoresis ◽  
Genomic Dna ◽  
Biosynthetic Pathway ◽  
Gene Clusters ◽  
Streptomyces Clavuligerus ◽  
Pulsed Field Gel Electrophoresis ◽  
Single Family ◽  
Chromosome Walking ◽  
Restriction Fragments

In Streptomyces clavuligerus, three groups of genes are known to be involved in the biosynthesis of the clavam metabolites. Since antibiotic biosynthetic genes are invariably clustered on the chromosome in prokaryotes, chromosome walking was undertaken in an attempt to show that the three groups of clavam genes would resolve into a single super-cluster when analyzed at larger scale. However, no evidence of linkage between the three groups was obtained. Furthermore, Southern analysis of macro-restriction fragments of genomic DNA separated by pulsed-field gel electrophoresis also indicated that the three groups of genes are not linked. Despite the structural and biosynthetic relatedness of the clavam metabolites, our results suggest that the genes involved in their production lie in three unlinked gene clusters. We believe that this represents the first instance in bacteria of genes involved in the biosynthesis of a single family of antibiotics sharing a common biosynthetic pathway and yet residing in three separate locations on the chromosome.Key words: Streptomyces, clavulanic acid, clavams, paralogues, gene clusters.

scholarly journals Physical Map of the Chromosome of the Apple Proliferation Phytoplasma

2000 ◽  
Vol 182 (5) ◽  
pp. 1415-1418 ◽  
Author(s):  
Ulrich Lauer ◽  
Erich Seemüller
Keyword(s):  
Gel Electrophoresis ◽  
Genomic Dna ◽  
Southern Blot Analysis ◽  
Physical Map ◽  
Pulsed Field Gel Electrophoresis ◽  
Apple Proliferation ◽  
Circular Chromosome ◽  
Tobacco Plants ◽  
Restriction Fragments ◽  
Partial Digestion

ABSTRACT A physical map of the apple proliferation phytoplasma strain AT chromosome was constructed from genomic DNA extracted from diseased tobacco plants. The map was generated with single and double digestions of the chromosome with BssHII, SmaI,MluI, and ApaI restriction endonucleases and resolving the fragments by pulsed-field gel electrophoresis. Partial digestion and Southern blot analysis were used to assist in the arrangement of the 14 contiguous restriction fragments obtained. From the restriction fragments generated by double digestions, the size of the circular chromosome was calculated to be approximately 645 kb. Locations of the two rRNA operons, the operon including thefus and tuf genes, and three other genes were placed on the map. Genome sizes and BssHII restriction profiles of apple proliferation strain AP15 and the pear decline and European stone fruit yellows phytoplasmas were different from that of strain AT.

scholarly journals Use of pulsed field gel electrophoresis and transposon mutagenesis to estimate the minimal number of genes required for motility in Caulobacter crescentus.

Genetics ◽  
1989 ◽  
Vol 123 (4) ◽  
pp. 649-654 ◽  
Author(s):  
B Ely ◽  
T W Ely
Keyword(s):  
Gel Electrophoresis ◽  
Caulobacter Crescentus ◽  
Pulsed Field Gel Electrophoresis ◽  
Restriction Fragments ◽  
Pulsed Field ◽  
Transposon Insertion ◽  
Number Of Genes ◽  
Physical And Genetic Map ◽  
Insertion Mutations ◽  
Flagellar Genes

Abstract To facilitate the mapping of transposon insertion mutations in Caulobacter crescentus, we have used pulsed field gel electrophoresis to construct a detailed physical and genetic map of the C. crescentus genome. Restriction fragments were generated by DraI, AseI, or SpeI which cleave the C. crescentus 40, 13, and 26 times, respectively, and Tn5 insertions were used to align the restriction fragments generated by each of the enzymes. The utility of the resulting map was demonstrated by determining the chromosomal locations of a collection of flagellar mutations. As a result of this study, we were able to identify ten new flagellar genes at various locations on the chromosome. Thus, at least 48 genes are required for the assembly of a functional flagellum in C. crescentus.

scholarly journals Comparison of Genomic Methods for Differentiating Strains of Enterococcus faecium: Assessment Using Clinical Epidemiologic Data

1998 ◽  
Vol 36 (11) ◽  
pp. 3327-3331 ◽  
Author(s):  
Connie Savor ◽  
Michael A. Pfaller ◽  
Julie A. Kruszynski ◽  
Richard J. Hollis ◽  
Gary A. Noskin ◽  
...  
Keyword(s):  
Enterococcus Faecium ◽  
Gel Electrophoresis ◽  
Genomic Dna ◽  
Restriction Endonuclease Analysis ◽  
Pulsed Field Gel Electrophoresis ◽  
Agarose Gel Electrophoresis ◽  
Subspecies Level ◽  
Vancomycin Resistant ◽  
Ideal Method ◽  
Endonuclease Analysis

Genomic DNA extracted from 45 vancomycin-resistantEnterococcus faecium (VRE) isolates was cleaved withHindIII and HaeIII and subjected to agarose gel electrophoresis. The ability of this method (restriction endonuclease analysis [REA]) to distinguish strains at the subspecies level was compared with results previously determined by pulsed-field gel electrophoresis (PFGE). Chart reviews were performed to provide a clinical correlation of possible epidemiologic relatedness. A likely clinical association was found for 29 patients as part of two outbreaks. REA found 21 of 21 isolates were the same type in the first outbreak, with PFGE calling 19 strains the same type. In the second outbreak with eight patient isolates, HindIII found six were the same type and two were unique types. HaeIII found three strains were the same type, two strains were a separate type, and three more strains were unique types, while PFGE found three were the same type and five were unique types. No single “ideal” method can be used without clinical epidemiologic investigation, but any of these techniques is helpful in providing focus to infection control practitioners assessing possible outbreaks of nosocomial infection.

scholarly journals Genotypic Characterization of Clarithromycin-Resistant and -Susceptible Helicobacter pylori Strains from the Same Patient Demonstrates Existence of Two Unrelated Isolates

1998 ◽  
Vol 36 (9) ◽  
pp. 2730-2731 ◽  
Author(s):  
Ge Wang ◽  
Qin Jiang ◽  
Diane E. Taylor
Keyword(s):  
Helicobacter Pylori ◽  
Gel Electrophoresis ◽  
Genomic Dna ◽  
Pulsed Field Gel Electrophoresis ◽  
23S Rrna ◽  
Rrna Gene ◽  
Clarithromycin Resistance ◽  
23S Rrna Gene ◽  
Mode Of Development

Clarithromycin-susceptible and clarithromycin-resistantHelicobacter pylori isolates from the same patient were investigated for the mode of development and mechanism of clarithromycin resistance. The clarithromycin-resistant strain UA1182 harbors homozygous A-to-G mutations at position 2143 in both copies of the 23S rRNA gene and has a phenotype of resistance to clarithromycin and clindamycin but no significant resistance to streptogramin B. Pulsed-field gel electrophoresis patterns of NruI- andNotI-digested genomic DNA from the Clas and Clar isolates demonstrated that they are genetically distinct, suggesting that the development of clarithromycin resistance is not from the mutation of the existing Clas strain but from a completely new strain.

scholarly journals Plasmid-Borne Type E Neurotoxin Gene Clusters in Clostridium botulinum Strains

2013 ◽  
Vol 79 (12) ◽  
pp. 3856-3859 ◽  
Author(s):  
Zhen Zhang ◽  
Hannamari Hintsa ◽  
Ying Chen ◽  
Hannu Korkeala ◽  
Miia Lindström
Keyword(s):  
Gene Cluster ◽  
Gel Electrophoresis ◽  
Clostridium Botulinum ◽  
Southern Hybridization ◽  
Gene Clusters ◽  
Pulsed Field Gel Electrophoresis ◽  
Content Type ◽  
Pulsed Field ◽  
Large Plasmids

ABSTRACTA collection of 36Clostridium botulinumtype E strains was examined by pulsed-field gel electrophoresis (PFGE) and Southern hybridization with probes targeted tobotEandorfX1in the neurotoxin gene cluster. Three strains were found to contain neurotoxin subtype E1 gene clusters in large plasmids of about 146 kb in size.

scholarly journals Epidemiological and genetic characterization of Clostridium butyricum cultured from neonatal cases of necrotizing enterocolitis in China

2020 ◽  
Vol 41 (8) ◽  
pp. 900-907
Author(s):  
Yinping Dong ◽  
Ying Li ◽  
Di Zhang ◽  
Scott Nguyen ◽  
Nikunj Maheshwari ◽  
...  
Keyword(s):  
Necrotizing Enterocolitis ◽  
Gel Electrophoresis ◽  
Genomic Dna ◽  
Genetic Characterization ◽  
Pulsed Field Gel Electrophoresis ◽  
Species Level ◽  
Clostridium Butyricum ◽  
Neonatal Necrotizing Enterocolitis ◽  
Dna Profiles

AbstractObjective:Laboratory-based characterization and traceback of Clostridium butyricum isolates linked to outbreak cases of neonatal necrotizing enterocolitis (NEC) in a hospital in China.Methods:In total, 37 samples were collected during the NEC outbreak. Classical bacteriological methods were applied to isolate and identify Clostridium spp. Meanwhile, 24 samples collected after an outbreak were similarly tested. All Clostridium isolates were identified to species level as either C. butyricum or C. sporogenes. These isolates were subsequently subtyped using pulsed-field gel electrophoresis (PFGE). Genomic DNA was purified from 2 representative C. butyricum isolates and sequenced to completion.Results:Of 37 samples collected during the NEC outbreak, 17 (45.95%) were positive for Clostridium spp. One species, C. butyricum, was cultured from 10 samples. Another species cultured from 2 other samples was identified as C. sporogenes. Both of these species were cocultured from 5 samples. Pulsotyping showed that the 15 C. butyricum and the 7 C. sporogenes isolates produced indistinguishable DNA profiles. No NEC cases were reported after disinfection following the outbreak, and all samples collected after the outbreak were negative for Clostridium spp. Whole-genome sequencing (WGS) indicated that sialidase, hemolysin, and enterotoxin virulence factors were located on the chromosomes of 2 C. butyricum isolates.Conclusions:The outbreak of NEC was epidemiologically linked to C. butyricum contamination within the hospital. This is the first report of an NEC outbreak associated with C. butyricum infection in China.

scholarly journals Evaluation of pulsed-field gel electrophoresis of genomic restriction fragments in the discrimination of Yersinia enterocolitica O[ratio ]3

1998 ◽  
Vol 121 (3) ◽  
pp. 579-586 ◽  
Author(s):  
K. ASPLUND ◽  
T. JOHANSSON ◽  
A. SIITONEN
Keyword(s):  
Yersinia Enterocolitica ◽  
Gel Electrophoresis ◽  
Pulsed Field Gel Electrophoresis ◽  
Restriction Fragments ◽  
Pulsed Field ◽  
Different Types

One hundred and six Yersinia enterocolitica serogroup O[ratio ]3, biotype 4 isolated from human and porcine samples in 1984 and in the years 1993–5 were examined by pulsed-field gel electrophoresis (PFGE). The genomic profiles produced by the enzymes NotI and XbaI were studied. Sixteen (A–P) and 8 (1·8) different pulsotypes were obtained, respectively. By combining the pulsotypes produced by both NotI and XbaI 24 different types were distinguished. The two major types, designated as A1 and B1, comprised 36% of all strains tested. The proportions of pulsotypes A1 and B1 were, 35·9 and 25·6%, respectively, among strains isolated in 1984. The corresponding figures among the strains isolated in 1993–5 were 35·8 and 41·8%. Nine pulsotypes were found only in 1984 and nine only in 1993–5. The proportions of the major pulsotypes, A1 and B1, in human isolates were 42·9 and 35·7% and in porcine isolates 22·2 and 36·1% respectively. Six types were found among both human and porcine isolates, 8 only among human strains and 10 only among porcine strains.

Characterization of Antimicrobial-Resistant Phenotypes and Genotypes among Salmonella enterica Recovered from Pigs on Farms, from Transport Trucks, and from Pigs after Slaughter

2004 ◽  
Vol 67 (4) ◽  
pp. 698-705 ◽  
Author(s):  
WONDWOSSEN A. GEBREYES ◽  
PETER R. DAVIES ◽  
PAA-KOBINA TURKSON ◽  
W. E. MORGAN MORROW ◽  
JULIE A. FUNK ◽  
...  
Keyword(s):  
Antimicrobial Resistance ◽  
Salmonella Typhimurium ◽  
Clavulanic Acid ◽  
Gel Electrophoresis ◽  
Rank Correlation ◽  
Pulsed Field Gel Electrophoresis ◽  
Mesenteric Lymph Nodes ◽  
Chloramphenicol Resistance ◽  
Pulsed Field ◽  
Amoxicillin Clavulanic Acid

The main objectives of this study were to determine antimicrobial resistance patterns among Salmonella serotypes and to evaluate the role of transport trucks in dissemination of antimicrobial-resistant strains of Salmonella. Salmonella from groups of nursery and finishing pigs on farms, from trucks, and from pigs after slaughter were compared using serotyping, patterns of antimicrobial resistance, and pulsed-field gel electrophoresis patterns. The five farms included in the study yielded 858 isolates representing 27 Salmonella serovars. The most common resistance observed (80% of all isolates) was to tetra-cycline; resistance to ampicillin (42%), chloramphenicol (31%), amoxicillin/clavulanic acid (30%), and piperacillin (31%) also were common. We found a correlation between serovar and antimicrobial resistance. High correlation was found between Salmonella Typhimurium var. Copenhagen and chloramphenicol resistance (Spearman rank correlation, ρ = 0.7). Multidrug resistance was observed primarily in Salmonella Typhimurium var. Copenhagen (94%) and Salmonella Typhimurium (93%) and was much less common in the other common serovars, including Salmonella Derby (7%) and Salmonella Heidelberg (8%). Of the 225 isolates exhibiting the most common pentaresistance pattern in this study, amoxicillin/clavulanic acid–ampicillin–chloramphenicol–piperacillin–tetracycline, 220 (98%) were Salmonella Typhimurium var. Copenhagen, and 86% of the isolates of this serovar had this pattern. Isolates from the trucks were similar, based on pulsed-field gel electrophoresis patterns, to those from the cecum and mesenteric lymph nodes of pigs on two of the farms, suggesting the probable infection of pigs during transport. Class I integrons were also common among various serovars.

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