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Necrotic fasciitis (NF) is a particularly aggressive and serious infection of the fascia that can penetrate into the musculature and internal organs, resulting in death if not treated promptly. In this work, an integrated microfluidic system composed of micropumps, microvalves, and micromixers was used to automate the detection of pathogens associated with NF. The entire molecular diagnostic process, including bacteria isolation, lysis, nucleic acid amplification and optical detection steps, was enacted on this developed system. Mannose binding lectin coated magnetic beads were first used as probes to isolate all bacteria in a sample. In this work, polymerase chain reaction assays featuring primers specific to genes from each of four NF-causing bacteria (Vibrio vulnificus, Aeromonas hydrophila, and methicillin-sensitive and resistant Staphylococcus aureus) were used to rapidly and exclusively verify the presence of the respective bacterial strains, and the limits of detection were experimentally found to be 11, 1960, 14, and 11 400 colony forming units/reaction, respectively; all values reflect improvement over ones reported in literature. This integrated microfluidic chip may then be valuable in expediting diagnosis and optimizing treatment options for those with NF; such diagnostic improvements could ideally diminish the need for amputation and even reduce the morality rate associated with this life-threatening illness. [ABSTRACT FROM AUTHOR]

Cardiovascular diseases (CVDs) cause more than 17×106 deaths worldwide on a yearly basis. Early diagnosis of CVDs is therefore of great need. The C-reactive protein (CRP) is an important biomarker for analyzing the risks of CVDs. In this work, CRP-specific aptamers with high sensitivity and specificity and field effect-transistor (FET) devices were used to recognize and detect CRP by using an integrated microfluidic system automatically while consuming less volumes of reagents and samples (about 5 μm). In order to package the FET device into the microfluidic chip, a new method to prevent liquid leakage was proposed. Sensitive detection of CRP has been demonstrated on the developed microfluidic system. It is the first time that aptamer-FET assays could be realized on an integrated microfluidic system. Experimental results showed that the aptamer-FET assay was capable of detecting CRP with concentrations ranging from 0.625 mg/l to 10.000 mg/l, which may be promising for early diagnosis of CVDs. [ABSTRACT FROM AUTHOR]

Vancomycin-resistant Enterococcus (VRE) is a kind of enterococci, which shows resistance toward antibiotics. It may last for a long period of time and meanwhile transmit the vancomycin-resistant gene (vanA) to other bacteria. In the United States alone, the resistant rate of Enterococcus to vancomycin increased from a mere 0.3% to a whopping 40% in the past two decades. Therefore, timely diagnosis and control of VRE is of great need so that clinicians can prevent patients from becoming infected. Nowadays, VRE is diagnosed by antibiotic susceptibility test or molecular diagnosis assays such as matrix-assisted laser desorption ionization/time-of-flight mass spectrometry and polymerase chain reaction. However, the existing diagnostic methods have some drawbacks, for example, time-consumption, no genetic information, or high false-positive rate. This study reports an integrated microfluidic system, which can automatically identify the vancomycin resistant gene (vanA) from live bacteria in clinical samples. A new approach using ethidium monoazide, nucleic acid specific probes, low temperature chemical lysis, and loop-mediated isothermal amplification (LAMP) has been presented. The experimental results showed that the developed system can detect the vanA gene from live Enterococcus in joint fluid samples with detection limit as low as 10 colony formation units/reaction within 1 h. This is the first time that an integrated microfluidic system has been demonstrated to detect vanA gene from live bacteria by using the LAMP approach. With its high sensitivity and accuracy, the proposed system may be useful to monitor antibiotic resistance genes from live bacteria in clinical samples in the near future. [ABSTRACT FROM AUTHOR]