Cronobacter Support
04-27-2009, 10:47 AM
1st International Conference on Cronobacter Poster Abstract 8
Development of Multiple-Locus Variable Number Tandem Repeat Analysis for the molecular subtyping of Enterobacter sakazakii
Enterobacter sakazakii is an emerging opportunistic pathogen that is the cause of rare cases of meningitis, necrotising enterocolitis and bacteremia in infants. The tandem repeat DNA motifs were identified in the genome sequence of E. sakazakii ATCC BAA-894. Four variable number tandem repeat (VNTR) loci selected and amplified individually by PCR with fluorescent labelling. PCR products were electrophoresed on a ABI 377 DNA analyser and size values were assigned to the GeneFlo™ TAMRA labelled ladder. A four-digit allele string based on the number of repeats was assigned to all isolates in the order ESTR-1, ESTR-2, ESTR-3, and ESTR-4. Allele strings were imported into Bionumerics and a minimum-spanning tree (MST) was generated. The genetic relationships of the analysed isolates were deduced by the construction of a MST. Forty-nine unique allele strings were observed among these isolates. A number of branched clusters were visible but no association could be established between isolate type, geographical origin, or biogroup. The discriminatory index was comparable to pulsed-field gel electrophoresis (PFGE). Molecular typing remains an important tool for surveillance, outbreak investigation and tracing of bacteria through the food chain. Although this method has less discriminatory power than PFGE, it could be potentially used to rapidly monitor outbreaks and trace this organism in food production facilities. It is realistic to assume that the inclusion of additional polymorphic loci would further increase the discriminatory power of the method.
Niall Mullane1,2, Mary Ryan2, Carol Iversen1,2, Mary Murphy3, Peadar O'Gaora4, Teresa Quinn1,2, Paul Whyte1,2, Patrick Wall5 and Séamus Fanning1,2
1Centre for Food Safety, 2School of Agriculture, Food Science and Veterinary Medicine,4 Conway Institute of Biomolecular and Biomedical Research,5 School of Public Health and Population Sciences, University College Dublin, Belfield, Dublin 4, Ireland. 3 Veterinary Food Safety Laboratory and Cork County Council, Inniscara, County Cork, Ireland.
Development of Multiple-Locus Variable Number Tandem Repeat Analysis for the molecular subtyping of Enterobacter sakazakii
Enterobacter sakazakii is an emerging opportunistic pathogen that is the cause of rare cases of meningitis, necrotising enterocolitis and bacteremia in infants. The tandem repeat DNA motifs were identified in the genome sequence of E. sakazakii ATCC BAA-894. Four variable number tandem repeat (VNTR) loci selected and amplified individually by PCR with fluorescent labelling. PCR products were electrophoresed on a ABI 377 DNA analyser and size values were assigned to the GeneFlo™ TAMRA labelled ladder. A four-digit allele string based on the number of repeats was assigned to all isolates in the order ESTR-1, ESTR-2, ESTR-3, and ESTR-4. Allele strings were imported into Bionumerics and a minimum-spanning tree (MST) was generated. The genetic relationships of the analysed isolates were deduced by the construction of a MST. Forty-nine unique allele strings were observed among these isolates. A number of branched clusters were visible but no association could be established between isolate type, geographical origin, or biogroup. The discriminatory index was comparable to pulsed-field gel electrophoresis (PFGE). Molecular typing remains an important tool for surveillance, outbreak investigation and tracing of bacteria through the food chain. Although this method has less discriminatory power than PFGE, it could be potentially used to rapidly monitor outbreaks and trace this organism in food production facilities. It is realistic to assume that the inclusion of additional polymorphic loci would further increase the discriminatory power of the method.
Niall Mullane1,2, Mary Ryan2, Carol Iversen1,2, Mary Murphy3, Peadar O'Gaora4, Teresa Quinn1,2, Paul Whyte1,2, Patrick Wall5 and Séamus Fanning1,2
1Centre for Food Safety, 2School of Agriculture, Food Science and Veterinary Medicine,4 Conway Institute of Biomolecular and Biomedical Research,5 School of Public Health and Population Sciences, University College Dublin, Belfield, Dublin 4, Ireland. 3 Veterinary Food Safety Laboratory and Cork County Council, Inniscara, County Cork, Ireland.