Fernando Teles

Biosensors and rapid diagnostic tests on the frontier between analytical and clinical chemistry for biomolecular diagnosis of dengue disease: a review.

Abstract The past decades have witnessed enormous technological improvements towards the development of simple, cost-effective and accurate rapid diagnostic tests for detection and identification of infectious pathogens. Among them is dengue virus, the etiologic agent of the mosquito-borne dengue disease, one of the most important emerging infectious pathologies of nowadays. Dengue fever may cause potentially deadly hemorrhagic symptoms and is endemic in the tropical and sub-tropical world, being also a serious threat to temperate countries in the developed world. Effective diagnostics for dengue should be able to discriminate among the four antigenically related dengue serotypes and fulfill the requirements for successful decentralized (point-of-care) testing in the harsh environmental conditions found in most tropical regions. The accurate identification of circulating serotypes is crucial for the successful implementation of vector control programs based on reliable epidemiological predictions. This paper briefly summarizes the limitations of the main conventional techniques for biomolecular diagnosis of dengue disease and critically reviews some of the most relevant biosensors and rapid diagnostic tests developed, implemented and reported so far for point-of-care testing of dengue infections. The invaluable contributions of microfluidics and nanotechnology encompass the whole paper, while evaluation concerns of rapid diagnostic tests and foreseen technological improvements in this field are also overviewed for the diagnosis of dengue and other infectious and tropical diseases as well.

Biosensors as rapid diagnostic tests for tropical diseases

Effective diagnosis of infectious pathogens is essential for disease identification and subsequent adequate treatment, to prevent drug resistance and to adopt suitable public health interventions for the prevention and control of epidemic outbreaks. Particular situations under which medical diagnostics operate in tropical environments make the use of new easy-to-use diagnostic tools the preferred (or even unique) option. These diagnostic tests and devices, usually based on biosensing methods, are being increasingly exploited as promising alternatives to classical, “heavy” lab instrumentation for clinical diagnosis, allowing simple, inexpensive and point-of-care testing. However, in many developing countries the lack of accessibility and affordability for many commercial diagnostic tests remains a major cause of high disease burden in such regions. We present a comprehensive overview about the problems of conventional medical diagnosis of infectious pathologies in tropical regions, while pointing out new m...Effective diagnosis of infectious pathogens is essential for disease identification and subsequent adequate treatment, to prevent drug resistance and to adopt suitable public health interventions for the prevention and control of epidemic outbreaks. Particular situations under which medical diagnostics operate in tropical environments make the use of new easy-to-use diagnostic tools the preferred (or even unique) option. These diagnostic tests and devices, usually based on biosensing methods, are being increasingly exploited as promising alternatives to classical, “heavy” lab instrumentation for clinical diagnosis, allowing simple, inexpensive and point-of-care testing. However, in many developing countries the lack of accessibility and affordability for many commercial diagnostic tests remains a major cause of high disease burden in such regions. We present a comprehensive overview about the problems of conventional medical diagnosis of infectious pathologies in tropical regions, while pointing out new methods and analytical tools for in-the-field and decentralized diagnosis of current major infectious tropical diseases. The review includes not only biosensor-based rapid diagnostic tests approved by regulatory entities and already commercialized, but also those at the early stages of research.

Genotypic diversity of environmental Cryptococcus neoformans isolates from Northern Portugal

The Cryptococcus neoformans/C. gattii species complex members are the main agents of systemic cryptococcosis. This disease is believed to be acquired from the environment via fungal cell inhalation. Often, isolates recovered from environmental and clinical sources have proven to be genotypically similar. We assessed the occurrence of C. neoformans and C. gattii in environmental substrates collected in a Portuguese region. Twenty‐eight isolates were identified as C. neoformans – five from decaying Eucalyptus leaves and 23 from domestic pigeon droppings. The isolates were genotyped using a URA5‐RFLP approach. The C. neoformans VNIV (53.6%, n = 15) and VNI (32.1%, n = 9) genotypes were abundantly present among environmental isolates. The hybrid VNIII (14.3%, n = 4) genotype was underrepresented and the VNII was not found. Cryptococcus gattii was also not found although some isolates yielded a positive canavanine–glycine–bromothymol blue test.

Nucleic-Acid Testing, New Platforms and Nanotechnology for Point-of-Decision Diagnosis of Animal Pathogens

Accurate disease diagnosis in animals is crucial for animal well-being but also for preventing zoonosis transmission to humans. In particular, livestock diseases may constitute severe threats to humans due to the particularly high physical contact and exposure and, also, be the cause of important economic losses, even in non-endemic countries, where they often arise in the form of rapid and devastating epidemics. Rapid diagnostic tests have been used for a long time in field situations, particularly during outbreaks. However, they mostly rely on serological approaches, which may confirm the exposure to a particular pathogen but may be inappropriate for point-of-decision (point-of-care) settings when emergency responses supported on early and accurate diagnosis are required. Moreover, they often exhibit modest sensitivity and hence significantly depend on later result confirmation in central or reference laboratories. The impressive advances observed in recent years in materials sciences and in nanotechnology, as well as in nucleic-acid synthesis and engineering, have led to an outburst of new in-the-bench and prototype tests for nucleic-acid testing towards point-of-care diagnosis of genetic and infectious diseases. Manufacturing, commercial, regulatory, and technical nature issues for field applicability more likely have hindered their wider entrance into veterinary medicine and practice than have fundamental science gaps. This chapter begins by outlining the current situation, requirements, difficulties, and perspectives of point-of-care tests for diagnosing diseases of veterinary interest. Nucleic-acid testing, particularly for the point of care, is addressed subsequently. A range of valuable signal transduction mechanisms commonly employed in proof-of-concept schemes and techniques born on the analytical chemistry laboratories are also described. As the essential core of this chapter, sections dedicated to the principles and applications of microfluidics, lab-on-a-chip, and nanotechnology for the development of point-of-care tests are presented. Microdevices already applied or under development for application in field diagnosis of animal diseases are reviewed.

The future of novel diagnostics in medical mycology.

Several fungal diseases have become serious threats to human health and life, especially upon the advent of human immunodeficiency virus/AIDS epidemics and of other typical immunosuppressive conditions of modern life. Accordingly, the burden posed by these diseases and, concurrently, by intensive therapeutic regimens against these diseases has increased worldwide. Existing and available rapid tests for point-of-care diagnosis of important fungal diseases could enable the limitations of current laboratory methods for detection and identification of medically important fungi to be surpassed, both in low-income countries and for first-line diagnosis (screening) in richer countries. As with conventional diagnostic methods and devices, former immunodiagnostics have been challenged by molecular biology-based platforms, as a way to enhance the sensitivity and shorten the assay time, thus enabling early and more accurate diagnosis. Most of these tests have been developed in-house, without adequate validation and standardization. Another challenge has been the DNA extraction step, which is especially critical when dealing with fungi. In this paper, we have identified three major research trends in this field: (1) the application of newer biorecognition techniques, often applied in analytical chemistry; (2) the development of new materials with improved physico-chemical properties; and (3) novel bioanalytical platforms, allowing fully automated testing. Keeping up to date with the fast technological advances registered in this field, primarily at the proof-of-concept level, is essential for wise assessment of those that are likely to be more cost effective and, as already observed for bacterial and viral pathogens, may provide leverage to the current tepid developmental status of novel and improved diagnostics for medical mycology.

Nanotechnology for the Diagnosis of Parasitic Infections

Infectious diseases can trigger a large range of clinical symptoms in humans and animals. Most of them are nonspecific signs, especially in their earlier stages, usually rendering differential diagnosis difficult. Many infections also result in cutaneous manifestations, which may present as isolated symptoms as a result of a more complex, or even serious, systemic disorder. In some cases, cutaneous signs may allow for the generation of an accurate diagnosis of disease by facile and direct examination. However, in many cases, the need for additional diagnostic tools and techniques are required. Repeated histopathological analysis of multiple skin samples in cases of equivocal diagnosis leads to significant cost and discomfort to the patient, despite the frequent reliance on clinical findings for diagnosis in dermatology [1]. As such, rapid and specific diagnosis of skin and soft tissues infections require improved diagnostic methods and tools. Ideally, these should be able to provide simple, cost-effective, rapid, specific and sensitive detection, identification and quantification of the etiologic agent. A rapid and accurate diagnosis of infection not only allows prompt initiation of therapy, but also, when resistant strains are detected, allows for a change of therapeutic regimen tailored to the patient’s pathogen profile. In addition, quantitative methods of detection, able to accurately determine the effective burden of a microorganism in host tissues, may constitute an important means to predict disease progression and prognosis. Aesthetic concerns associated with current diagnostic methods must also be taken into account then managing infections. Some diagnostic techniques such as punch and excisional biopsies may result in considerable disfigurement and scarring. New diagnostic approaches based on biosensor technology, especially involving nanotechnological structures, have been developed in all stages of the research cycle from proof-of-concept, to prototype, to early clinical trials. Apart from technical and commercial constraints in developed countries, shifting from lab-based molecular analysis to point-of-care testing faces many ethical concerns and obstacles. Although this is less so for the diagnosis of infectious than genetic diseases [2]. Nanotechnology represents a promising approach to develop and utilize novel and improved tests to diagnose dermatological infections.