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Molecular diagnosis of biofilm-related genes (BRGs) in common bacteria that cause periprosthetic joint infections may provide crucial information for clinicians. In this study, several BRGs, including ica, fnbA, and fnbB, were rapidly detected (within 1 h) with a new integrated microfluidic system. Mannose-binding lectin (MBL)-coated magnetic beads were used to isolate these bacteria, and on-chip nucleic acid amplification (polymerase chain reaction, PCR) was then performed to detect BRGs. Both eukaryotic and prokaryotic MBLs were able to isolate common bacterial strains, regardless of their antibiotic resistance, and limits of detection were as low as 3 and 9 CFU for methicillin-resistant Staphylococcus aureus and Escherichia coli, respectively, when using a universal 16S rRNA PCR assay for bacterial identification. It is worth noting that the entire process including bacteria isolation by using MBL-coated beads for sample pre-treatment, on-chip PCR, and fluorescent signal detection could be completed on an integrated microfluidic system within 1 h. This is the first time that an integrated microfluidic system capable of detecting BRGs by using MBL as a universal capturing probe was reported. This integrated microfluidic system might therefore prove useful for monitoring profiles of BRGs and give clinicians more clues for their clinical judgments in the near future.

this study presents a simple microfluidic device which can perform sample pretreatment automatically for rapid bacteria detection. Herein, a new approach to rapidly isolate bacteria incorporated with vancomycin-conjugated magnetic beads by using a new membrane-type microfluidic component with a floating block was proposed and its performance was tested and verified. The experimental results showed the proposed device can automatically isolate methicillin-resistant Staphylococcus aureus from 4 µL of samples within 90 seconds including bacteria isolation and washing processes. Not only does the new membrane-type microfluidic device perform rigid mixing, but it is also capable of uniform sample transportation. Furthermore, it has been successfully demonstrated that it could isolate bacteria in an automated format. With its great mixing efficiency and robust performance, the proposed device might be a promising approach for bacteria diagnosis.

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.

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.