A requirement of TolC and MDR efflux pumps for acid adaptation and GadAB induction in Escherichia coli.

BACKGROUND: The TolC outer membrane channel is a key component of several multidrug resistance (MDR) efflux pumps driven by H(+) transport in Escherichia coli. While tolC expression is under the regulation of the EvgA-Gad acid resistance regulon, the role of TolC in growth at low pH and extreme-acid survival is unknown. METHODS AND PRINCIPAL FINDINGS: TolC was required for extreme-acid survival (pH 2) of strain W3110 grown aerobically to stationary phase. A tolC deletion decreased extreme-acid survival (acid resistance) of aerated pH 7.0-grown cells by 10(5)-fold and of pH 5.5-grown cells by 10-fold. The requirement was specific for acid resistance since a tolC defect had no effect on aerobic survival in extreme base (pH 10). TolC was required for expression of glutamate decarboxylase (GadA, GadB), a key component of glutamate-dependent acid resistance (Gad). TolC was also required for maximal exponential growth of E. coli K-12 W3110, in LBK medium buffered at pH 4.5-6.0, but not at pH 6.5-8.5. The TolC growth requirement in moderate acid was independent of Gad. TolC-associated pump components EmrB and MdtB contributed to survival in extreme acid (pH 2), but were not required for growth at pH 5. A mutant lacking the known TolC-associated efflux pumps (acrB, acrD, emrB, emrY, macB, mdtC, mdtF, acrEF) showed no growth defect at acidic pH and a relatively small decrease in extreme-acid survival when pre-grown at pH 5.5. CONCLUSIONS: TolC and proton-driven MDR efflux pump components EmrB and MdtB contribute to E. coli survival in extreme acid and TolC is required for maximal growth rates below pH 6.5. The TolC enhancement of extreme-acid survival includes Gad induction, but TolC-dependent growth rates below pH 6.5 do not involve Gad. That MDR resistance can enhance growth and survival in acid is an important consideration for enteric organisms passing through the acidic stomach.

Rapid and cost-effective SNP detection method: application of SmartAmp2 to pharmacogenomics research.

Pharmacogenomics data can facilitate our understanding of the sources of variability in drug response, which can potentially lead to improved safety and efficacy of drug therapy for individual patients. A key requirement for the development of individualized medicine or personalized therapy is the ability to rapidly and conveniently test patients for genetic polymorphisms and/or mutations. However, in today's world, genotyping technology remains a bottleneck in clinical applications because of its slow speed and high cost. Therefore, we have recently developed a rapid and cost-effective method for SNP detection, named Smart Amplification Process 2 (SmartAmp2), which enables us to detect genetic polymorphisms or mutations in 30-45 min under isothermal conditions without DNA isolation and PCR amplification. This article presents the SNP detection method and its underlying molecular mechanism as well as clinical research applications.