Translational genomics to develop a Salmonella enterica serovar Paratyphi A multiplex polymerase chain reaction assay.
The use of pathogen genome sequence data for the control and management of infections remains an ongoing challenge. We describe a broadly applicable, web-enabled approach that can be used to develop bacteria-specific polymerase chain reaction (PCR) assays. Salmonella enterica Paratyphi A has emerged as a major cause of enteric fever in Asia. Culture-based diagnosis is slow and frequently negative in patients with suspected typhoid and paratyphoid fever, potentially compromising patient management and public health. We used the MobilomeFINDER web-server to perform in silico subtractive hybridization, thus identifying 43 protein-coding sequences (CDSs) that were present in two Paratyphi A strains but not in other sequenced Salmonella genomes. After exclusion of 29 CDSs found to be variably present in Paratyphi A strains by microarray hybridization and grouping of remaining CDSs by genomic location, four dispersed targets (stkF, spa2473, spa2539, hsdM) were used to develop a highly discriminatory multiplex PCR assay. All 52 Paratyphi A strains within the diverse panel investigated produced one of two pathognomonic four-band signatures. Given rapid and ongoing expansion of DNA and comparative genomics databases, our universally accessible web-server-supported do-it-yourself approach offers the potential to contribute significantly to the rapid development of species-, serovar-, or pathotype-specific PCR assays targeting pre-existing and emerging bacterial pathogens.
Ou
Hong-Yu
HY
Laboratory of Microbial Metabolism and School of Life Science and Biotechnology, Shanghai Jiaotong University, Shanghai, China.
Ju
Cindy Teh Shuan
CT
Thong
Kwai-Lin
KL
Ahmad
Norazah
N
Deng
Zixin
Z
Barer
Michael R
MR
Rajakumar
Kumar
K
D001483:Base Sequence; D005798:Genes, Bacterial; D023281:Genomics / Q000379:methods*; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000379:methods*; D011110:Polymorphism, Genetic; D012482:Salmonella paratyphi A / Q000235:genetics* / Q000302:isolation & purification*