Your browser doesn't support javascript.
loading
Show: 20 | 50 | 100
Results 1 - 2 de 2
Filter
Add more filters










Database
Language
Publication year range
1.
J Microbiol Methods ; 146: 16-21, 2018 03.
Article in English | MEDLINE | ID: mdl-29360487

ABSTRACT

The Burkholderia cepacia complex (Bcc) consists of at least 20 phenotypically similar but genotypically distinct Gram-negative bacteria that are ubiquitous in nature, are capable of promoting plant growth and biodegradation of pollutants, but that also are highly antibiotic resistant and produce damaging effects towards plants, fungi, and humans. To study these genetically recalcitrant bacteria in detail, molecular tools are required that work efficiently with the many strains and species of the Bcc. One mutagenesis strategy that has been used effectively to analyze the genes of Burkholderia cenocepacia is based upon the activity of the Sce-I restriction enzyme. Unfortunately, this system is limited in its applicability to many members of the Bcc. Therefore, we undertook the expansion of this system to create an Sce-I mutagenesis system that could be used with many different species and strains of the Bcc, including members of the B. cenocepacia IIIB Midwest clones. We demonstrated the use of this system by clean-deleting the lipo-oligosaccharide (LOS) inner core biosynthesis gene waaC, to create a B. cenocepacia PC184 strain variant with truncated LOS. This enhanced mutagenesis system can be used to analyze a wide range of Burkholderia and other Gram-negative bacteria.


Subject(s)
Bacterial Typing Techniques/methods , Burkholderia cepacia complex/genetics , Mutagenesis , DNA, Bacterial , Genes, Bacterial/genetics , Genotype , Lipopolysaccharides/genetics , Plasmids/genetics , Transformation, Bacterial
2.
BMC Genomics ; 14: 574, 2013 Aug 27.
Article in English | MEDLINE | ID: mdl-23978260

ABSTRACT

BACKGROUND: As is true for many other antibiotic-resistant Gram-negative pathogens, members of the Burkholderia cepacia complex (BCC) are currently being assessed for their susceptibility to phage therapy as an antimicrobial treatment. The objective of this study was to perform genomic and limited functional characterization of the novel BCC phage JG068 (vB_BceP_JG068). RESULTS: JG068 is a podovirus that forms large, clear plaques on Burkholderia cenocepacia K56-2. Host range analysis indicates that this phage can infect environmental, clinical, and epidemic isolates of Burkholderia multivorans, B. cenocepacia, Burkholderia stabilis, and Burkholderia dolosa, likely through interaction with the host lipopolysaccharide as a receptor. The JG068 chromosome is 41,604 base pairs (bp) in length and is flanked by 216 bp short direct terminal repeats. Gene expression originates from both host and phage promoters and is in the forward direction for all 49 open reading frames. The genome sequence shows similarity to Ralstonia phage ϕRSB1, Caulobacter phage Cd1, and uncharacterized genetic loci of blood disease bacterium R229 and Burkholderia pseudomallei 1710b. CoreGenesUniqueGenes analysis indicates that JG068 belongs to the Autographivirinae subfamily and ϕKMV-like phages genus. Modules within the genome encode proteins involved in DNA-binding, morphogenesis, and lysis, but none associated with pathogenicity or lysogeny. Similar to the signal-arrest-release (SAR) endolysin of ϕKMV, inducible expression of the JG068 SAR endolysin causes lysis of Escherichia coli that is dependent on the presence of an N-terminal signal sequence. In an in vivo assay using the Galleria mellonella infection model, treatment of B. cenocepacia K56-2-infected larvae with JG068 results in a significant increase in larval survival. CONCLUSIONS: As JG068 has a broad host range, does not encode virulence factors, is obligately lytic, and has activity against an epidemic B. cenocepacia strain in vivo, this phage is a highly promising candidate for BCC phage therapy development.


Subject(s)
Burkholderia cenocepacia/virology , Genome, Viral , Podoviridae/genetics , Base Sequence , Host Specificity , Molecular Sequence Annotation , Molecular Sequence Data , Podoviridae/isolation & purification , Podoviridae/ultrastructure , Promoter Regions, Genetic , Sequence Analysis, DNA , Sequence Homology, Nucleic Acid , Sewage/virology , Soil Microbiology , Terminator Regions, Genetic , Transcription Initiation Site , Viral Proteins/genetics , Virion/genetics , Virion/isolation & purification , Virion/ultrastructure , Virulence
SELECTION OF CITATIONS
SEARCH DETAIL
...