TY - JOUR
T1 - Subspecies characterization of urease-positive thermophilic Campylobacter (UPTC) isolated from shellfish employing modified flagellin (flaA) restriction fragment length polymorphism (RFLP) typing
AU - Moore, John E.
AU - Matsuda, Motoo
AU - Sekizuka, Tsuyoshi
AU - Fanning, Seamus
AU - Murayama, Ohoshi
AU - Yokoi, Taeko
AU - Kagawa, Shizuko
AU - Nagano, Kentaro
AU - Shimura, Hiromi
AU - Watabe, Miyuki
AU - Nagano, Yuriko
AU - Usui, Kaori
AU - Imamaki, Rie
AU - Aritomi, Takao
AU - Harada, Chisato
AU - Iida, Haruna
AU - Mitsuhashi, Naomi
AU - Miyatake, Takeshi
AU - Shigematsu, Makoto
AU - Rao, Juluri R.
AU - Lowery, Colm
AU - Millar, B. Cherie
AU - Dooley, James
AU - Rooney, Paul J.
PY - 2006/8
Y1 - 2006/8
N2 - Shellfish including oysters (Crassostrea gigas), cockles (Cerastoderma edule) and mussels (Mytilus edulis), have previously been described as an important source of thermophilic campylobacters, with the potential of causing acute bacterial gastroenteritis in humans. Previous genotyping studies employing the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) typing, based on the flagellin (flaA) gene have been unable to generate an amplicon for the urease-positive thermophilic Campylobacter (UPTC), which are the predominant taxa associated with shellfish, largely caused by sequence diversity between the UPTC group and C. jejuni. Hence the aim of this study was to develop a modified PCR-RFLP genotyping assay, employing polymorphisms within the flagellin (flaA) gene of UPTC organisms, which would now allow the successful amplification and typing of previously nontypable UPTC isolates obtained from natural marine environments. A novel primer pair (UPTC flaF/UPTC flaR) was designed based on conserved regions within the flaA gene locus of UPTC organisms to generate a 1,358 bp amplicon for all UPTC organisms tested. RFLP analysis with DdeI in combination with computational analysis of genetic relatedness using BioNumerics software demonstrated the presence of four distinct flaA genotypes, among the seven UPTC isolates. In conclusion, this study describes a PCR-RFLP method, based on modified primers from UPTC flaA gene sequences that may be successfully applied to examine subspecies relatedness of UPTC organisms from natural environments, including shellfish.
AB - Shellfish including oysters (Crassostrea gigas), cockles (Cerastoderma edule) and mussels (Mytilus edulis), have previously been described as an important source of thermophilic campylobacters, with the potential of causing acute bacterial gastroenteritis in humans. Previous genotyping studies employing the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) typing, based on the flagellin (flaA) gene have been unable to generate an amplicon for the urease-positive thermophilic Campylobacter (UPTC), which are the predominant taxa associated with shellfish, largely caused by sequence diversity between the UPTC group and C. jejuni. Hence the aim of this study was to develop a modified PCR-RFLP genotyping assay, employing polymorphisms within the flagellin (flaA) gene of UPTC organisms, which would now allow the successful amplification and typing of previously nontypable UPTC isolates obtained from natural marine environments. A novel primer pair (UPTC flaF/UPTC flaR) was designed based on conserved regions within the flaA gene locus of UPTC organisms to generate a 1,358 bp amplicon for all UPTC organisms tested. RFLP analysis with DdeI in combination with computational analysis of genetic relatedness using BioNumerics software demonstrated the presence of four distinct flaA genotypes, among the seven UPTC isolates. In conclusion, this study describes a PCR-RFLP method, based on modified primers from UPTC flaA gene sequences that may be successfully applied to examine subspecies relatedness of UPTC organisms from natural environments, including shellfish.
M3 - Article
VL - 25
SP - 625
EP - 629
JO - Journal of Shellfish Research
JF - Journal of Shellfish Research
IS - 2
ER -
