Towards a smartphone-connected point-of-care test for HIV

The devastation caused by HIV is driving the development of new point-of-care diagnostics. The work presented in this thesis aims to help develop a new generation of smartphone- connected HIV tests designed to address the very high levels of undiagnosed HIV-infected individuals, by widening access to HIV testing to doctors surgeries, pharmacies and developing countries. The biosensor is based on mass manufacturable surface acoustic wave (SAW) devices, and uses piezoelectricity to transduce the binding of biomarkers on the surface of the device into a measurable electric signal, making the test low cost, easy to use and reliable. In addition, the SAW biosensor presented here has the ability to wirelessly and securely transmit results to healthcare providers to potentially offer follow-up appointments at local clinics, or virtually. This thesis begins with the theory behind SAW biosensors. A more focussed characterisation of the specific device developed is then presented, followed by the details of the work done to optimise the biosensor in order to make it a good candidate for a point-of-care test for HIV. Key results include the proof of concept detection of different biomarkers of HIV infection, as well as a demonstration of the ability of the SAW biosensor to deliver a fast response. Different pilot studies are then presented, demonstrating the performance of the device as a diagnostic test, highlighting 100% sensitivity and 100% specificity. These were conducted with more than 30 confirmed HIV positive patient samples and more than 100 healthy volunteers. The following chapter then examines the fundamental mechanisms underpinning the SAW biosensor output and an empirical method to ultimately design more sensitive devices in future antigen detection. This thesis concludes with a summary of the main results and future work, including the potential for larger clinical studies, and field trials in developing countries

Soil biodiversity: functions, threats and tools for policy makers

Human societies rely on the vast diversity of benefits provided by nature, such as food, fibres, construction materials, clean water, clean air and climate regulation. All the elements required for these ecosystem services depend on soil, and soil biodiversity is the driving force behind their regulation. With 2010 being the international year of biodiversity and with the growing attention in Europe on the importance of soils to remain healthy and capable of supporting human activities sustainably, now is the perfect time to raise awareness on preserving soil biodiversity. The objective of this report is to review the state of knowledge of soil biodiversity, its functions, its contribution to ecosystem services and its relevance for the sustainability of human society. In line with the definition of biodiversity given in the 1992 Rio de Janeiro Convention, soil biodiversity can be defined as the variation in soil life, from genes to communities, and the variation in soil habitats, from micro-aggregates to entire landscapes. Bio Intelligence Service, IRD, and NIOO, Report for European Commission (DG Environment

Harnessing recombinase polymerase amplification for rapid multi-gene detection of SARS-CoV-2 in resource-limited settings

The COVID-19 pandemic is challenging diagnostic testing capacity worldwide. The mass testing needed to limit the spread of the virus requires new molecular diagnostic tests to dramatically widen access at the point-of-care in resource-limited settings. Isothermal molecular assays have emerged as a promising technology, given the faster turn-around time and minimal equipment compared to gold standard laboratory PCR methods. However, unlike PCR, they do not typically target multiple SARS-CoV-2 genes, risking sensitivity and specificity. Moreover, they often require multiple steps thus adding complexity and delays. Here we develop a multiplexed, 1-2 step, fast (20-30 minutes) SARS-CoV-2 molecular test using reverse transcription recombinase polymerase amplification to simultaneously detect two conserved targets - the E and RdRP genes. The agile multi-gene platform offers two complementary detection methods: real-time fluorescence or dipstick. The analytical sensitivity of the fluorescence test was 9.5 (95% CI: 7.0-18) RNA copies per reaction for the E gene and 17 (95% CI: 11-93) RNA copies per reaction for the RdRP gene. The analytical sensitivity for the dipstick was 130 (95% CI: 82-500) RNA copies per reaction. High specificity was found against common seasonal coronaviruses, SARS-CoV and MERS-CoV model samples. The dipstick readout demonstrated potential for point-of-care testing in decentralised settings, with minimal or equipment-free incubation methods and a user-friendly prototype smartphone application. This rapid, simple, ultrasensitive and multiplexed molecular test offers valuable advantages over gold standard tests and in future could be configurated to detect emerging variants of concern

Beyond protocols:improving the reliability of expert-based risk analysis underpinning invasive species policies

Risk assessment tools for listing invasive alien species need to incorporate all available evidence and expertise. Beyond the wealth of protocols developed to date, we argue that the current way of performing risk analysis has several shortcomings. In particular, lack of data on ecological impacts, transparency and repeatability of assessments as well as the incorporation of uncertainty should all be explicitly considered. We recommend improved quality control of risk assessments through formalized peer review with clear feedback between assessors and reviewers. Alternatively, a consensus building process can be applied to better capture opinions of different experts, thereby maximizing the evidential basis. Elaborating on manageability of invasive species is further needed to fully answer all risk analysis requirements. Tackling the issue of invasive species urges better handling of the acquired information on risk and the exploration of improved methods for decision making on biodiversity management. This is crucial for efficient conservation resource allocation and uptake by stakeholders and the public

Quantifying Biomolecular Binding Constants Using Video Paper Analytical Devices

A novel ultra‐low‐cost biochemical analysis platform to quantify protein dissociation binding constants and kinetics using paper microfluidics is reported. This approach marries video imaging with one of humankind's oldest materials: paper, requiring no large, expensive laboratory equipment, complex microfluidics or external power. Temporal measurements of nanoparticle–antibody conjugates binding on paper is found to follow the Langmuir Adsorption Model. This is exploited to measure a series of antibody–antigen dissociation constants on paper, showing excellent agreement with a gold‐standard benchtop interferometer. The concept is demonstrated with a camera and low‐end smartphone, 500‐fold cheaper than the reference method, and can be multiplexed to measure ten reactions in parallel. These findings will help to widen access to quantitative analytical biochemistry, for diverse applications spanning disease diagnostics, drug discovery, and environmental analysis in resource‐limited settings

Deep learning of HIV field-based rapid tests

Although deep learning algorithms show increasing promise for disease diagnosis, their use with rapid diagnostic tests performed in the field has not been extensively tested. Here we use deep learning to classify images of rapid human immunodeficiency virus (HIV) tests acquired in rural South Africa. Using newly developed image capture protocols with the Samsung SM-P585 tablet, 60 fieldworkers routinely collected images of HIV lateral flow tests. From a library of 11,374 images, deep learning algorithms were trained to classify tests as positive or negative. A pilot field study of the algorithms deployed as a mobile application demonstrated high levels of sensitivity (97.8%) and specificity (100%) compared with traditional visual interpretation by humans-experienced nurses and newly trained community health worker staff-and reduced the number of false positives and false negatives. Our findings lay the foundations for a new paradigm of deep learning-enabled diagnostics in low- and middle-income countries, termed REASSURED diagnostics1, an acronym for real-time connectivity, ease of specimen collection, affordable, sensitive, specific, user-friendly, rapid, equipment-free and deliverable. Such diagnostics have the potential to provide a platform for workforce training, quality assurance, decision support and mobile connectivity to inform disease control strategies, strengthen healthcare system efficiency and improve patient outcomes and outbreak management in emerging infections

Assessing the assessments: evaluation of four impact assessment protocols for invasive alien species

Aim: Effective policy and management responses to the multiple threats posed by invasive alien species (IAS) rely on the ability to assess their impacts before conclusive empirical evidence is available. A plethora of different IAS risk and/or impact assessment protocols have been proposed, but it remains unclear whether, how and why the outcomes of such assessment protocols may differ. Location: Europe. Methods: Here, we present an in-depth evaluation and informed assessment of the consistency of four prominent protocols for assessing IAS impacts (EICAT, GISS, Harmonia and NNRA), using two non-native parrots in Europe: the widespread ring-necked parakeet (Psittacula krameri) and the rapidly spreading monk parakeet (Myiopsitta monachus). Results: Our findings show that the procedures used to assess impacts may influence assessment outcomes. We find that robust IAS prioritization can be obtained by assessing species based on their most severe documented impacts, as all protocols yield consistent outcomes across impact categories. Additive impact scoring offers complementary, more subtle information that may be especially relevant for guiding management decisions regarding already established invasive alien species. Such management decisions will also strongly benefit from consensus approaches that reduce disagreement between experts, fostering the uptake of scientific advice into policy-making decisions. Main conclusions: Invasive alien species assessments should take advantage of the capacity of consensus assessments to consolidate discussion and agreement between experts. Our results suggest that decision-makers could use the assessment protocol most fit for their purpose, on the condition they apply a precautionary approach by considering the most severe impacts only. We also recommend that screening for high-impact IAS should be performed on a more robust basis than current ad hoc practices, at least using the easiest assessment protocols and reporting confidence scores.This study is a joint effort from a workshop organized under COST European Cooperation in Science and Technology Actions Parrotnet (ES1304) and Alien Challenge (TD1209). We would also like to thank two anonymous referees for helpful comments on an earlier version of this manuscript. D.S is currently funded by a Marie Skłodowska‐Curie Action under the Horizon 2020 call (H2020‐MSCA‐IF‐2015, grant number 706318) and acknowledges the Danish National Research Foundation for support to the Center for Macroecology, Evolution and Climate (grant number DNRF96). J.C.S. was supported by funds from the Ministry of Economy and Competitivity, Spanish Research Council (CGL‐2016‐79568‐C3‐3‐P)