ID-UCLA

Electrochemical Sensor Research

Emergence of drug resistant pathogens is a global healthcare crisis that is forcing physicians to treat common infectious diseases with ever more potent antibiotics.  New strategies are urgently needed for rapid identification of drug-resistant healthcare-associated infections and to provide clinicians with real-time information to guide antibiotic selection. The goal of this research project is to develop a biosensor for rapid, high-throughput bacterial pathogen identification and antibiotic susceptibility determination. 

Electrochemical sensors are molecular sensing devices that intimately couple a biological recognition element to an electrode transducer.  Such devices produce an accurate, sensitive and yet simple, inexpensive, compact and low-power platform for point-of-care diagnosis. The best examples are the widely used blood glucose sensors that use glucose-specific oxidases on a sensor strip to link the blood glucose concentration to current flow . Another successful portable electrochemical device is the hand-held i-STAT Abbot system that performs eight common clinical tests simultaneously in about 90 seconds .

Rapid Bacterial Identification.  In collaboration with GeneFluidics, we developed an electrochemical sensor assay for microbial identification involving “sandwich” hybridization of the abundant target rRNA (~25,000 copies per bacterial cell) to a panel of capture and detector probes for rapid (30 min), species-specific identification of a broad array of bacterial pathogens in clinical urine specimens (1, 2).  Together with Joseph Wang’s laboratory at UCSD, the sensor assay’s limit of detection has recently been lowered to 250 bacteria/milliliter (3) and was expanded to direct bioassays for target molecules in raw body fluids (4). 

Rapid Detection of Antibiotic Resistance.  We are developing the electrochemical sensor assay as a rapid antimicrobial susceptibility assay for bacteria in clinical specimens.  Step One of the assay involves identification and quantitation with a 7-probe sensor.  Step Two involves specimen inoculation and incubation in growth medium with and without antibiotics.  Step Three involves bacterial quantitation for measurement of the phenotypic response to antibiotics.  The rapid antimicrobial susceptibility assay is being performed using an automated, robotic high-throughput system developed by GeneFluidics.

The electrochemical sensor assay for 16s rRNA can detect aminoglycoside resistance within 30 minutes.  Detection of quinolone resistance can be determined within 15 minutes by measurement of precursor-rRNA (5).  Recently we discovered that antibiotic resistance testing for a broad range of beta-lactam antibiotics can be dramatically accelerated and determined within 30-45 minutes by addition of the penicillin-binding protein 2 inhibitor, amdinocillin (6).

References

1. Liao JC, Mastali M, Gau V, Suchard MA, Moller AK, Bruckner DA, et al. Use of electrochemical DNA biosensors for rapid molecular identification of uropathogens in clinical urine specimens. J Clin Microbiol. 2006;44(2):561-70.

2. Liao JC, Mastali M, Li Y, Gau V, Suchard M, Babbitt JT, et al. Development of an advanced electrochemical DNA biosensor for bacterial pathogen detection. J Mol Diagn. 2007;9:158-68.

3. Wu J, Campuzano S, Halford C, Haake DA, Wang J. Ternary Surface Monolayers for Ultrasensitive (Zeptomole) Amperometric Detection of Nucleic Acid Hybridization without Signal Amplification. Anal Chem. 2010;82:8830-7.

4. Campuzano S, Kuralay F, Jesús Lobo-Castañón J, Bartošík M, Vyavahare K, Paleček E, et al. Ternary monolayers as DNA recognition interfaces for direct and sensitive electrochemical detection in untreated clinical samples. Biosens Bioelectron. 2011;26:3577-84.

5. Halford C, Gonzalez R, Campuzano S, Hu B, Babbit J T, Liu J, Wang J, Churchill B M, Haake D A.  Rapid antimicrobial susceptibility testing by sensitive detection of precursor rRNA using a novel electrochemical biosensing platform.  Antimicrob Agents Chemother. 2013; 57:936-43.

6. Halford C, Humphries R, Churchill B M, Haake D A.  Rapid electrochemical detection of antimicrobial resistance to beta-lactam antibiotics using the penicillin-binding protein 2 inhibitor, amdinocillin. Manuscript in Preparation.