Detection on surfaces and in Caco-2 cells of Campylobacter jejuni cells transformed with new gfp, yfp, and cfp marker plasmids.
We have developed two sets of Campylobacter shuttle vectors containing either the gfp (green fluorescent protein), yfp (yellow fluorescent protein), or cfp (cyan fluorescent protein) reporter gene. In one set, the reporter gene is fused to a consensus Campylobacter promoter sequence (P(c)). The other set contains a pUC18 multicloning site upstream of the reporter gene, allowing the construction of transcriptional fusions using known promoters or random genomic fragments. C. jejuni cells transformed with the P(c) fusion plasmids are strongly fluorescent and easily visualized on chicken skin, on plant tissue, and within infected Caco-2 cells. In each C. jejuni strain tested, these plasmids were maintained over several passages in the absence of antibiotic selection. Also, in many C. jejuni strains, >91% of the cells transformed with the P(c) fusion plasmids remained fluorescent after several days. Experiments with yellow fluorescent and cyan fluorescent C. jejuni transformants suggest that aggregates containing two or more strains of C. jejuni may be present in an enrichment broth culture. Colonies arising from these aggregates would be heterologous in nature; therefore, isolation of a pure culture of C. jejuni, by selecting single colonies, from an environmental sample may not always yield a single strain.
Miller
W G
WG
Food Safety and Health Research Unit, Agricultural Research Service, U.S. Department of Agriculture, Albany, California 94710, USA. bmiller@pw.usda.gov
Bates
A H
AH
Horn
S T
ST
Brandl
M T
MT
Wachtel
M R
MR
Mandrell
R E
RE
D000818:Animals; D001422:Bacterial Adhesion; D001426:Bacterial Proteins / Q000235:genetics; D001483:Base Sequence; D018938:Caco-2 Cells; D016123:Campylobacter jejuni / Q000235:genetics* / Q000302:isolation & purification / Q000472:pathogenicity; D017931:DNA Primers / Q000235:genetics; D005516:Food Microbiology; D017930:Genes, Reporter; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins; D006801:Humans; D008164:Luminescent Proteins / Q000235:genetics; D018613:Microscopy, Confocal; D008969:Molecular Sequence Data; D010957:Plasmids / Q000235:genetics*; D014170:Transformation, Genetic
D001426:Bacterial Proteins; D017931:DNA Primers; D008164:Luminescent Proteins; C054562:yellow fluorescent protein, Bacteria; D049452:Green Fluorescent Proteins