A review on impedimetric immunosensors for pathogen and biomarker detection.

Since the discovery of antibiotics in the first quarter of the twentieth century, their use has been the principal approach to treat bacterial infection. Modernized medicine such as cancer therapy, organ transplantation or advanced major surgeries require effective antibiotics to manage bacterial infections. However, the irresponsible use of antibiotics along with the lack of development has led to the emergence of antimicrobial resistance which is considered a serious global threat due to the rise of multidrug-resistant bacteria (Wang et al. in Antibiotic resistance: a rundown of a global crisis, pp. 1645-1658, 2018). Currently employed diagnostics techniques are microscopy, colony counting, ELISA, PCR, RT-PCR, surface-enhanced Raman scattering and others. These techniques provide satisfactory selectivity and sensitivity (Joung et al. in Sens Actuators B Chem 161:824-831, 2012). Nevertheless, they demand specialized personnel and expensive and sophisticated machinery which can be labour-intensive and time-consuming, (Malvano et al. in Sensors (Switzerland) 18:1-11, 2018; Mantzila et al. in Anal Chem 80:1169-1175, 2008). To get around these problems, new technologies such as biosensing and lab-on-a-chip devices have emerged in the last two decades. Impedimetric immunosensors function by applying electrochemical impedance spectroscopy to a biosensor platform using antibodies or other affinity proteins such as Affimers (Tiede et al. in Elife 6(c):1-35, 2017) or other binding proteins (Weiss et al. in Electrochim Acta 50:4248-4256, 2005) as bioreceptors, which provide excellent sensitivity and selectivity. Pre-enrichment steps are not required and this allows miniaturization and low-cost. In this review different types of impedimetric immunosensors are reported according to the type of electrode and their base layer materials, either self-assembled monolayers or polymeric layers, composition and functionalization for different types of bacteria, viruses, fungi and disease biomarkers. Additionally, novel protein scaffolds, both antibody derived and non-antibody derived, used to specifically target the analyte are considered.

Biosensor Regeneration: A Review of Common Techniques and Outcomes.

Biosensors are ideally portable, low-cost tools for the rapid detection of pathogens, proteins, and other analytes. The global biosensor market is currently worth over 10 billion dollars annually and is a burgeoning field of interdisciplinary research that is hailed as a potential revolution in consumer, healthcare, and industrial testing. A key barrier to the widespread adoption of biosensors, however, is their cost. Although many systems have been validated in the laboratory setting and biosensors for a range of analytes are proven at the concept level, many have yet to make a strong commercial case for their acceptance. Though it is true with the development of cheaper electrodes, circuits, and components that there is a downward pressure on costs, there is also an emerging trend toward the development of multianalyte biosensors that is pushing in the other direction. One way to reduce the cost that is suitable for certain systems is to enable their reuse, thus reducing the cost per test. Regenerating biosensors is a technique that can often be used in conjunction with existing systems in order to reduce costs and accelerate the commercialization process. This article discusses the merits and drawbacks of regeneration schemes that have been proven in various biosensor systems and indicates parameters for successful regeneration based on a systematic review of the literature. It also outlines some of the difficulties encountered when considering the role of regeneration at the point of use. A brief meta-analysis has been included in this review to develop a working definition for biosensor regeneration, and using this analysis only ∼60% of the reported studies analyzed were deemed a success. This highlights the variation within the field and the need to normalize regeneration as a standard process across the field by establishing a consensus term.

Systematic review of methods to predict and detect anastomotic leakage in colorectal surgery.

AIM: Anastomotic leakage is a serious complication of gastrointestinal surgery resulting in increased morbidity and mortality, poor function and predisposing to cancer recurrence. Earlier diagnosis and intervention can minimize systemic complications but is hindered by current diagnostic methods that are non-specific and often uninformative. The purpose of this paper is to review current developments in the field and to identify strategies for early detection and treatment of anastomotic leakage. METHOD: A systematic literature search was performed using the MEDLINE, Embase, PubMed and Cochrane Library databases. Search terms included 'anastomosis' and 'leak' and 'diagnosis' or 'detection' and 'gastrointestinal' or 'colorectal'. Papers concentrating on the diagnosis of gastrointestinal anastomotic leak were identified and further searches were performed by cross-referencing. RESULTS: Computerized tomography CT scanning and water-soluble contrast studies are the current preferred techniques for diagnosing anastomotic leakage but suffer from variable sensitivity and specificity, have logistical constraints and may delay timely intervention. Intra-operative endoscopy and imaging may offer certain advantages, but the ability to predict anastomotic leakage is unproven. Newer techniques involve measurement of biomarkers for anastomotic leakage and have the potential advantage of providing cheap real-time monitoring for postoperative complications. CONCLUSION: Current diagnostic tests often fail to diagnose anastomotic leak at an early stage that enables timely intervention and minimizes serious morbidity and mortality. Emerging technologies, based on detection of local biomarkers, have achieved proof of concept status but require further evaluation to determine whether they translate into improved patient outcomes. Further research is needed to address this important, yet relatively unrecognized, area of unmet clinical need.

Intra-operative tumour detection and staging in colorectal cancer surgery.

AIM: Surgical resection for colorectal cancer involves segmental resection and regional lymphadenectomy. The appropriateness of this 'one-size-fits-all' strategy is questioned as bowel cancer screening programmes result in a shift to earlier stage disease. Currently, the nodal status of a colorectal cancer can only be reliably determined by histopathological examination of the resected specimen. New methods of intra-operative staging are required to allow surgical resection to be tailored to the stage of the disease. METHOD: A literature search was performed of PubMed and Embase databases using the terms 'colon' OR 'colorectal' AND 'intra-operative detection' OR 'intra-operative staging' OR 'intra-operative detection' OR 'radioimmunoguided surgery'. Articles published between January 1980 and January 2012 were included. Technologies that have the potential to allow intra-operative staging and treatment stratification were identified and further searches performed. RESULTS: Established techniques such as sentinel lymph node mapping and radioimmunoguided surgery have benefited from combination with other technologies to allow real-time intra-operative staging. Intra-operative fluorescence, using naturally fluorescent biomarkers or fluorescent tumour probes, probably offers the most practical means of intra-operative lymph node staging and may be facilitated using nanotechnology. Optical coherence tomography and real-time elastography have the potential to provide an in vivo'virtual biopsy'. CONCLUSION: Technological advances may allow accurate intra-operative lymph node staging to facilitate tailored surgical resection. This may become the next paradigm shift in colorectal cancer surgery.

The auxin signal.

Recent work on the auxin signal has yielded clear answers to some questions and produced puzzling new data to explain. It is now established that the auxin-binding protein functions extracellularly, but it is unclear how it reaches that location. Important clues on the mechanism(s) by which auxin achieves its genetic and cell biological effects are emerging.

Estimated cumulative X-ray exposure and additional cancer risk during the evaluation and treatment of scoliosis in children and young people requiring surgery.

INTRODUCTION: Clinicians and patients must weigh the benefits of radiological imaging against the risks of radiation exposure in the diagnosis and treatment of scoliosis. This report aims to estimate the cumulative absorbed and equivalent dose of radiation in patients undergoing surgical treatment for scoliosis, and to present this as an estimated risk of cancer compared to background radiation levels. METHODS: Retrospective review of estimated absorbed dose on the Computerised Radiology Information System (CRIS®). Patients undergoing surgical correction of scoliosis (age ≤ 25) from August 2010 to August 2015 investigated. Estimated absorbed dose [milligrays (mGy)] recorded. Pedicle screws inserted using image intensification. Equivalent dose [millisieverts (mSv)] and additional cancer risk calculated from the National Research Council document 'Health risks from exposure to low levels of ionising radiation' (2006). RESULTS: 271 patients identified. Mean age 15 (range 2-25). Mean total absorbed dose 2136 mGy [standard deviation (SD) 1700 mGy]. Mean number of plain spine radiographs was 8 (SD 3) with total 1884 mGy exposure (SD 1609 mGy). Additional dose provided by CT (mean 0.17 episodes), plain chest and abdominal radiographs and image intensification. Mean number of image intensification episodes was 1.1 with mean estimated exposure 180 mGy (SD 238 mGy). Image intensification accounted for 8% of the estimated absorbed dose during treatment. Estimated mean effective dose delivered was 20.952 mSv equating to an additional cancer risk of 0.27-0.45%. CONCLUSION: Additional cancer risk from cumulative imaging is small and equivalent to approximately 8 years of natural background radiation. Use of image intensification for pedicle screw insertion is a minor contribution (8%) to the total patient dose.

Arabidopsis AUX1 gene: a permease-like regulator of root gravitropism.

The plant hormone auxin regulates various developmental processes including root formation, vascular development, and gravitropism. Mutations within the AUX1 gene confer an auxin-resistant root growth phenotype and abolish root gravitropic curvature. Polypeptide sequence similarity to amino acid permeases suggests that AUX1 mediates the transport of an amino acid-like signaling molecule. Indole-3-acetic acid, the major form of auxin in higher plants, is structurally similar to tryptophan and is a likely substrate for the AUX1 gene product. The cloned AUX1 gene can restore the auxin-responsiveness of transgenic aux1 roots. Spatially, AUX1 is expressed in root apical tissues that regulate root gravitropic curvature.

Current concepts in the surgical management of adolescent idiopathic scoliosis.

Adolescent idiopathic scoliosis (AIS) is a complex 3D deformity of the spine. Its prevalence is between 2% and 3% in the general population, with almost 10% of patients requiring some form of treatment and up to 0.1% undergoing surgery. The cosmetic aspect of the deformity is the biggest concern to the patient and is often accompanied by psychosocial distress. In addition, severe curves can cause cardiopulmonary distress. With proven benefits from surgery, the aims of treatment are to improve the cosmetic and functional outcomes. Obtaining correction in the coronal plane is not the only important endpoint anymore. With better understanding of spinal biomechanics and the long-term effects of multiplanar imbalance, we now know that sagittal balance is equally, if not more, important. Better correction of deformities has also been facilitated by an improvement in the design of implants and a better understanding of metallurgy. Understanding the unique character of each deformity is important. In addition, using the most appropriate implant and applying all the principles of correction in a bespoke manner is important to achieve optimum correction. In this article, we review the current concepts in AIS surgery. Cite this article: Bone Joint J 2018;100-B:415-24.

Acetylecholinesterase-based biosensor electrodes for organophosphate pesticide detection. II. Immobilization and stabilization of acetylecholinesterase.

The dry and wet stability of Drosophila acetylcholinesterase non-covalently immobilized onto polyethyleneimine modified screen-printed carbon electrodes was improved when compared to non-immobilized acetylcholinesterase, and acetylcholinesterase covalently immobilized onto dialdehyde and polyethyleneimine modified electrodes. Stabilizer mixtures were characterized for additional stabilization of acetylcholinesterase during storage in the dry state, with dextran-sulphate/sucrose and polygalacturonic acid/sucrose mixtures proving highly effective for long-term storage of biosensor electrodes.

ADP-ribosylation of thylakoid membrane polypeptides by cholera toxin is correlated with inhibition of thylakoid GTPase activity and protein phosphorylation.

Incubation of pea thylakoid membranes with [32P]-NAD+ in the presence of cholera toxin resulted in the [32P]-ADP-ribosylation of a 60 kDa thylakoid membrane polypeptide. When ATP was included in the incubation mixture, a 29 kDa polypeptide was also labelled. In the absence of electron transfer cofactors or inhibitors, the extent of labelling depended on whether the membranes were preincubated in the light or dark and also on the developmental stage of the leaves used for thylakoid isolation. Irrespective of the latter, the strongest labelling was observed when DCMU was present in the light. After pretreatment of the thylakoid membranes with cholera toxin plus NAD+ under the same conditions, light-stimulated GTPase activity and protein phosphorylation were inhibited. The extent of inhibition for both processes appeared to be correlated with the amount of [32P]-ADP-ribosylation found when [32P]-NAD+ was included in the pretreatment mixture. The data presented are fully consistent with the 60 and 29 kDa polypeptides functioning as thylakoid membrane associated guanine nucleotide binding regulatory proteins.

Personalised image-based templates for intra-operative guidance.

The high rate of unplanned perforation, poor fixation, and nerve injury with freehand pedicle screw insertion has led to the use of image-guided navigation systems. Although these improve accuracy, they have several drawbacks that could be overcome by using image-based drilling guide templates. The accuracy of such templates was tested in a cadaveric study of screw placement in the lumbar, thoracic, and cervical regions of the spine. The dimensional stability with autoclaving of duraform polyamide, to be used for manufacturing the guides, was first determined using test specimens. Computed tomography (CT) images were acquired of 4 cadaveric spines, and placement of 4 cervical, 32 thoracic, and 14 lumbar screws was planned. Eighteen personalized drilling guide templates, in four different designs, were built. Orthopaedic surgeons experienced in the freehand techniques used the templates. CT images were acquired to assess placement position with respect to the pedicle. Duraform polyamide was found to be unaffected by sterilization. Two of the template designs facilitated the placement of 20/20 screws without error. Templates can lead to successful screw placement, even in small pedicles, providing their design is optimized for the application area, e.g. with enhanced rotational stabilization.

Isolation of a putative receptor from Zea mays microsomal membranes that interacts with the G-protein, GP alpha 1.

The C-terminal region of a heterotrimeric G-protein alpha-subunit is known to be one of the principal determinants governing its interaction with its cognate receptor. Use of an oligopeptide corresponding to the fifteen C-terminal residues of the Arabidopsis G alpha-subunit (GP alpha 1), as an affinity ligand, led to the resolution of a tightly binding 37 kDa membrane polypeptide from detergent solubilised Zea microsomal fraction membranes. An identical polypeptide bound tightly to an affinity matrix containing recombinant GP alpha 1 protein as ligand: binding and release of this 37 kDa protein was dependent on the activation state of GP alpha 1 which was regulated by inclusion or omission of the G-protein activator AlF-4. Finally, the isolated 37 kDa protein was labelled with the lectin concanavalin A, indicating it to be glycosylated. These data are consistent with the identity of the 37 kDa membrane polypeptide as a receptor that interacts with the Zea homologue of GP alpha 1.

Investigation of the substrate specificity of thylakoid protein kinase using synthetic peptides.

Synthetic peptide analogues of the N-terminal region of the light harvesting chlorophyll a/b binding polypeptide of photosystem II (LHC II) were used to probe the effect of charged groups on the protein kinase activity of pea (Pisum sativum) thylakoid membranes. The effectiveness of the synthetic peptides as substrates for protein kinase activity or as inhibitors of LHC II phosphorylation was correlated with their net positive charge, which ranged between +2 and +5. The effects of the synthetic peptides on phosphorylation of other, non-LHC II, thyakoid polypeptides are also discussed.

Heterotrimeric G-proteins in plant cell signaling.

Heterotrimeric G-proteins, which couple cell surface receptors with internal effectors, are evident in all eukaryotes. Their operation involves receptor activation, GTP/GDP exchange and modulation of effector activity; deactivation occurs by an intrinsic GTPase activity. Structurally, G-proteins comprise three dissimilar subunits; Gα, Gß and Gγ. The Gα subunit consists of an α-helical and a GTPase domain, the latter is responsible for interaction with Gßγ, receptor and effector. Gß and Gγ form a tightly associated heterodimer which can also modulate effector activity when released by the activated Gα. Genome sequence and other data suggest that, in plants, there are several (~8-10?) Gα, one or two Gß and one Gγ. These proteins are expressed throughout the plant, mainly in the plasma membrane and endoplasmic reticulum. In vivo, there is strong evidence for G-protein control of ion channels, particularly K+ , in the response pathways to fungal and bacterial pathogens as well as in some aspects of gibberellin, abscisic acid and auxin signaling pathways. Finally, future prospects for understanding plant G-protein linked signaling will rely on new and emerging technologies; these include antisense suppression, gene knockouts, yeast two-hybrid and phage display molecular approaches, intracellular immunization using recombinant single chain antibodies and expression of peptide encoding minigenes.

Acetylcholinesterase-based biosensor electrodes for organophosphate pesticide detection. I. Modification of carbon surface for immobilization of acetylcholinesterase.

Screen-printed carbon electrodes modified with the dialdehydes, glutaraldehyde and terephthaldicarboxaldehyde, and then polyethyleneimine have been utilized for production of pesticide biosensors based on acetylcholinesterase. To improve the extent of dialdehyde modification, the electrodes were NH2-derivatized, initially by electrochemical reduction of 4-nitrobenzenediazonium to a nitroaryl radical permitting attachment to the carbon surface. Subsequent reduction of the 4-nitrobenzene yields a 4-aminobenzene modified carbon surface. Drosophila melanogaster acetylcholinesterase was immobilized either covalently onto dialdehyde modified electrodes or non-covalently onto polyethyleneimine modified electrodes. Internal diffusion limitations due to the dialdehyde and polyethyleneimine modifications increased the apparent Km of the immobilized enzyme. The thiocholine sensitivity was about 90% for dialdehyde modified electrodes and about 10% for polyethyleneimine modified electrodes as compared with non-modified carbon electrodes. The detection limit of the biosensors produced by non-covalent immobilization of acetylcholinesterase onto polyethyleneimine modified carbon electrodes was found to be about 10(-10) M for the organophosphate pesticide dichlorvos.

Late-onset idiopathic scoliosis in children six to fourteen years old. A cross-sectional prevalence study.

We performed a point-prevalence survey of 15,799 children, six to fourteen years old, who formed part of a prospective longitudinal study. Our purpose was to detect the prevalence of scoliosis and to investigate associated factors. On the basis of the initial screening, 934 children (5.9 per cent) were referred for additional clinical and radiographic examinations; 896 children returned for this second evaluation. A lateral spinal curve with a Cobb angle of more than 5 degrees was seen in 431 children (2.7 per cent of the 15,799 children). Only seventy-six children (0.5 per cent) had a curve that met our definition of idiopathic scoliosis (a curve of more than 10 degrees with concordant apical rotation). The point-prevalence rate was higher in girls, and it increased with age. The rate was 0.1 per cent (four of 5246) in the age-group of six to eight years, 0.3 per cent (sixteen of 5831) in the age-group of nine to eleven years, and 1.2 per cent (fifty-six of 4722) in the age-group of twelve to fourteen years old. With allowance for the fact that different definitions of idiopathic scoliosis have been used in earlier studies, our results suggest that the natural history of idiopathic scoliosis may be becoming more benign spontaneously.

Indications of disordered eating behaviour in adolescent patients with idiopathic scoliosis.

We have investigated whether patients with adolescent-onset idiopathic scoliosis (AIS) are more likely to have a low body-weight. Measurements of weight, height and body mass index (BMI) were made in 44 young women with AIS and compared with age- and gender-adjusted normative data. The body mass criteria of the International Classification of Diseases for eating disorders was used to determine how many patients were within the range considered to be 'eating disordered'. Compared with the normative data, the AIS group did not differ significantly in terms of height, (p = 0.646), but they were significantly lighter (p < 0.001) and had significantly lower BMI scores (p < 0.001); 25% of the series had BMI scores which were within the range considered to be anorexic. The relationship between a diagnosis of AIS and low body-weight may indicate disordered eating and is thus a cause for concern, particularly in the light of the well-established relationship between eating psychopathology and osteoporosis. Aspects of organic health may need to be considered in addition to the cosmetic deformity.

Primary bone tumours of the hand and wrist.

Primary bone tumours are rare in the hand and wrist. This 34-year survey of the Leeds Regional Bone Tumour Registry reveals 80 such tumours, representing 3.9% of the total number of bone tumours in the registry. The large majority of these tumours were benign (86%), and were found predominantly in younger patients. Malignant lesions were found more frequently in older patients. 61% of tumours were found in the metacarpals and proximal phalanges. Two-thirds of patients (67.5%) presented with features of swelling, either with or without pain. We conclude that the information gathered from Bone Tumour Registries is of value in describing tumour characteristics, where such information could not be gathered by personal experience alone.

The removal of indicator microorganisms from primary treated wastewater in subsurface reed beds using different substrates.

Subsurface, horizontal flow, experimental reed beds, were designed and built based on a combination of two design methodologies, that of the WRc and Severn Trent Water Plc and that of the USA, EPA. Four different growing media were used with a combination of top soil, gravel, river sand and mature sewage sludge compost, aiming to determine the best substrate for enteric pathogen removal. Eight units were constructed, two for each material. One bed for each pair was planted with Typha latifolia plants commonly known as cattails. Primary treated domestic wastewater, was continuously fed in to the bed for more than six months. All beds achieved a high reduction of Escherichia coli and faecal coliforms with the best results recorded in the gravel reed beds with an average removal above 3.3 log for E. coli and 3.0 for faecal coliforms. There was no significance difference in the performance of planted and unplanted reed beds.