Michelle Rogers

Transfer of antibiotic resistance by transformation with eDNA within oral biofilms

We demonstrate that live donor Veillonella dispar cells can transfer the conjugative transposon Tn916 to four different Streptococcus spp. recipients in a multispecies oral consortium growing as a biofilm in a constant depth film fermentor. Additionally, we demonstrate that purified V. dispar DNA can transform Streptococcus mitis to tetracycline resistance in this experimental system. These data show that transfer of conjugative transposon-encoded antibiotic resistance can occur by transformation in addition to conjugation in biofilms.

Continuous online microdialysis using microfluidic sensors: dynamic neurometabolic changes during spreading depolarization

Microfluidic glucose biosensors and potassium ion selective electrodes were used in an in vivo study to measure the neurochemical effects of spreading depolarizations (SD), which have been shown to be detrimental to the injured human brain. A microdialysis probe implanted in the cortex of rats was connected to a microfluidic PDMS chip containing the sensors. The dialysate was also analyzed using our gold standard, rapid sampling microdialysis (rsMD). The glucose biosensor performance was validated against rsMD with excellent results. The glucose biosensors successfully monitored concentration changes, in response to SD wave induction, in the range of 10–400 μM with a second time-resolution. The data show that during a SD wave, there is a time delay of 62 ± 24.8 s (n = 4) between the onset of the increase in potassium and the decrease in glucose. This delay can be for the first time demonstrated, thanks to the high-temporal resolution of the microfluidic sensors sampling from a single tissue site (the microdialysis probe), and it indicates that the decrease in glucose is due to the high demand of energy required for repolarization.

ATP microelectrode biosensor for stable long-term in vitro monitoring from gastrointestinal tissue.

We have developed a stable and selective ATP biosensor for long-term in vitro tissue monitoring. The electrode was fabricated by entrapping glucose oxidase (GOx) and hexokinase (HEX) in a poly-phenol film on a Pt microelectrode. The biosensor was stable to a fixed concentration of glucose for over 20 min and had a limit of detection of 9.9 ± 3.2 nM, with a sensitivity of 45.8 ± 1.22 pA μM(-1). Most significantly of all, the response on the ATP biosensor did not alter in the presence of 1mM ascorbic acid, 5 μM dopamine, 5 μM serotonin, 5 μM ADP and 5 μM AMP. The ATP biosensor was also shown to have excellent stability over 7 days, and showed only a 23.92 ± 3.55% loss in sensitivity. The ATP biosensor was utilised for the in vitro detection of ATP from gastrointestinal tissue. The ATP biosensor response was stable for 5h during in vitro recordings from ileum tissue. ATP release was shown to be greater from the mucosal surface in the ileum compared to the colon.

Optimisation of a microfluidic analysis chamber for the placement of microelectrodes

The behaviour of droplets entering a microfluidic chamber designed to house microelectrode detectors for real time analysis of clinical microdialysate is described. We have designed an analysis chamber to collect the droplets produced by multiphase flows of oil and artificial cerebral spinal fluid. The coalescence chamber creates a constant aqueous environment ideal for the placement of microelectrodes avoiding the contamination of the microelectrode surface by oil. A stream of alternating light and dark coloured droplets were filmed as they passed through the chamber using a high speed camera. Image analysis of these videos shows the colour change evolution at each point along the chamber length. The flow in the chamber was simulated using the general solution for Poiseuille flow in a rectangular chamber. It is shown that on the centre line the velocity profile is very close to parabolic, and an expression is presented for the ratio between this centre line velocity and the mean flow velocity as a function of channel aspect ratio. If this aspect ratio of width/height is 2, the ratio of flow velocities closely matches that of Poiseuille flow in a circular tube, with implications for connections between microfluidic channels and connection tubing. The droplets are well mixed as the surface tension at the interface with the oil dominates the viscous forces. However once the droplet coalesces with the solution held in the chamber, the no-slip condition at the walls allows Poiseuille flow to take over. The meniscus at the back of the droplet continues to mix the droplet and acts as a piston until the meniscus stops moving. We have found that the no-slip conditions at the walls of the chamber, create a banding effect which records the history of previous drops. The optimal position for sensors is to be placed at the plane of droplet coalescence ideally at the centre of the channel, where there is an abrupt concentration change leading to a response time ≪16 ms, the compressed frame rate of the video. Further away from this point the response time and sensitivity decrease due to convective dispersion.

High-Performance Bioinstrumentation for Real-Time Neuroelectrochemical Traumatic Brain Injury Monitoring

Traumatic brain injury (TBI) has been identified as an important cause of death and severe disability in all age groups and particularly in children and young adults. Central to TBIs devastation is a delayed secondary injury that occurs in 30–40% of TBI patients each year, while they are in the hospital Intensive Care Unit (ICU). Secondary injuries reduce survival rate after TBI and usually occur within 7 days post-injury. State-of-art monitoring of secondary brain injuries benefits from the acquisition of high-quality and time-aligned electrical data i.e., ElectroCorticoGraphy (ECoG) recorded by means of strip electrodes placed on the brains surface, and neurochemical data obtained via rapid sampling microdialysis and microfluidics-based biosensors measuring brain tissue levels of glucose, lactate and potassium. This article progresses the field of multi-modal monitoring of the injured human brain by presenting the design and realization of a new, compact, medical-grade amperometry, potentiometry and ECoG recording bioinstrumentation. Our combined TBI instrument enables the high-precision, real-time neuroelectrochemical monitoring of TBI patients, who have undergone craniotomy neurosurgery and are treated sedated in the ICU. Electrical and neurochemical test measurements are presented, confirming the high-performance of the reported TBI bioinstrumentation.

Real-Time Clinical Monitoring of Biomolecules

Continuous monitoring of clinical biomarkers offers the exciting possibility of new therapies that use biomarker levels to guide treatment in real time. This review explores recent progress toward this goal. We initially consider measurements in body fluids by a range of analytical methods. We then discuss direct tissue measurements performed by implanted sensors; sampling techniques, including microdialysis and ultrafiltration; and noninvasive methods. A future directions section considers analytical methods at the cusp of clinical use.

Simultaneous monitoring of potassium, glucose and lactate during spreading depolarization in the injured human brain – Proof of principle of a novel real-time neurochemical analysis system, continuous online microdialysis:

Spreading depolarizations occur spontaneously and frequently in injured human brain. They propagate slowly through injured tissue often cycling around a local area of damage. Tissue recovery after an spreading depolarization requires greatly augmented energy utilisation to normalise ionic gradients from a virtually complete loss of membrane potential. In the injured brain, this is difficult because local blood flow is often low and unreactive. In this study, we use a new variant of microdialysis, continuous on-line microdialysis, to observe the effects of spreading depolarizations on brain metabolism. The neurochemical changes are dynamic and take place on the timescale of the passage of an spreading depolarization past the microdialysis probe. Dialysate potassium levels provide an ionic correlate of cellular depolarization and show a clear transient increase. Dialysate glucose levels reflect a balance between local tissue glucose supply and utilisation. These show a clear transient decrease of variable magnitude and duration. Dialysate lactate levels indicate non-oxidative metabolism of glucose and show a transient increase. Preliminary data suggest that the transient changes recover more slowly after the passage of a sequence of multiple spreading depolarizations giving rise to a decrease in basal dialysate glucose and an increase in basal dialysate potassium and lactate levels.

A High‐Performance Application Specific Integrated Circuit for Electrical and Neurochemical Traumatic Brain Injury Monitoring

Abstract This paper presents the first application specific integrated chip (ASIC) for the monitoring of patients who have suffered a Traumatic Brain Injury (TBI). By monitoring the neurophysiological (ECoG) and neurochemical (glucose, lactate and potassium) signals of the injured human brain tissue, it is possible to detect spreading depolarisations, which have been shown to be associated with poor TBI patient outcome. This paper describes the testing of a new 7.5 mm2 ASIC fabricated in the commercially available AMS 0.35 μm CMOS technology. The ASIC has been designed to meet the demands of processing the injured brain tissues ECoG signals, recorded by means of depth or brain surface electrodes, and neurochemical signals, recorded using microdialysis coupled to microfluidics‐based electrochemical biosensors. The potentiostats use switchedcapacitor charge integration to record currents with 100 fA resolution, and allow automatic gain changing to track the falling sensitivity of a biosensor. This work supports the idea of a “behind the ear” wireless microplatform modality, which could enable the monitoring of currently non‐monitored mobile TBI patients for the onset of secondary brain injury.

Chemical Monitoring in Clinical Settings: Recent Developments toward Real-Time Chemical Monitoring of Patients

This review covers references found in the recent literature (from 2015) describing clinically relevant chemical monitoring that either gives continuous chemical information in real time or has the near prospect of doing so.