Lyme rashes disease classification using deep feature fusion technique

Automatic classification of Lyme disease rashes on the skin helps clinicians and dermatologists’ probe and investigate Lyme skin rashes effectively. This paper proposes a new in-depth features fusion system to classify Lyme disease rashes. The proposed method consists of two main steps. First, three different deep learning models, Densenet201, InceptionV3, and Exception, were trained independently to extract the deep features from the erythema migrans (EM) images. Second, a deep feature fusion mechanism (meta classifier) is developed to integrate the deep features before the final classification output layer. The meta classifier is a basic deep convolutional neural network trained on original images and features extracted from base level three deep learning models. In the feature fusion mechanism, the last three layers of base models are dropped out and connected to the meta classifier. The proposed deep feature fusion method significantly improved the classification process, where the classification accuracy was 98.97%, which is particularly impressive than the other state-of-the-art models.© 2023 The Authors. Skin Research and Technology published by John Wiley & Sons Ltd. This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited.fi=vertaisarvioitu|en=peerReviewed

Sick of ticks?:Towards a comprehensive understanding of rarely diagnosed<i> Ixodes ricinus</i>-borne diseases

This thesis expands the current knowledge on the impact and risk posed by tick-borne diseases (TBDs) other than LB and TBE in Europe and more specifically in the Netherlands. More importantly, it examines different approaches to do so and how this knowledge could be employed in the control of emerging TBDs. TBDs represent a puzzle of health care, scientific and societal challenges and each chapter of this thesis constitutes the tackling of one of its many pieces. The different research approaches used show the risk posed by TBDs can only be assessed when epidemiological, clinical, experimental and bioinformatical data are combined. These can subsequently be translated into actionable steps for effective monitoring and control of TBDs in the Netherlands and the rest of Europe.Much is left to uncover in the field of TBDs as human populations and their environment mutually reshape each other overtime giving raise to new conditions. Likewise, the tools at our disposal for scientific research, by undergoing constant innovation, expand our ability to solve old problems but also understand new ones prompting further innovation. The task of control and surveillance at the interface between well established and emerging pathogens is thus never ending and comes with a great responsibility that necessitates continuous multidisciplinary collaboration

An appraisal of health datasets to enhance the surveillance of Lyme disease in the United Kingdom

Lyme disease is a tick-borne disease of increasing global public health interest. Clinical presentation is varied, posing challenges for case definition. Currently national incidence figures for the United Kingdom (UK) are derived from two-tier confirmatory laboratory diagnostic results. These figures have the potential to underestimate incidence as clinical cases managed without diagnostic investigation are unrecorded. This thesis aimed to identify and evaluate a variety of datasets for their ability to describe the incidence and sociodemographics of Lyme disease cases in the UK, and to assess whether they could be utilised in future national surveillance programmes. The datasets analysed were: Public Health England (PHE)’s Lyme disease diagnostic laboratory, PHE’s laboratory surveillance system, hospital episode statistics data for England and Wales, an electronic health records database of primary care in the UK, Twitter, and the Small Animals Veterinary Surveillance Network (SAVSNET). A generalised Lyme disease population could be described from these data. This population had a bimodal age distribution, was predominately white, was from rural areas, and increasingly from areas with lower societal deprivation. Geographic distribution of cases could be described for England and Wales and showed the highest incidence of disease in southern central to south western England. These data showed an increasing incidence of Lyme disease. The relative incidence of Lyme disease cases varied between datasets, with the primary care data having the largest incidence of 4.42 per 100,000 person-years (95% CI 4.23-4.67). Multiplication factors were described between the three datasets of routinely collected health care data. The most important being a multiplication factor of 2.35 (95% CI 1.81-2.88) between laboratory-confirmed incidence and primary care incidence in England and Wales. The results from this thesis start to describe the epidemiological picture of Lyme disease in the UK; specifically identified as a research gap by the NICE guidelines. They will provide a platform for the many unanswered questions about the changing landscape of Lyme disease in the UK. It was concluded that a combination of health datasets could be used for future Lyme disease surveillance systems in the UK. Ideally this would include laboratory and primary care data. Until this is in place, the multiplication factor can be used to estimate the national incidence of Lyme disease and the potential burden it places on the National Health Service and the patients it afflicts

MULTI-AFFINITY NANOTRAPS THAT ENHANCE DETECTION OF LOW-ABUNDANT PROTEINS: A NOVEL AND HIGHLY SENSITIVE TEST FOR THE DIAGNOSIS OF LYME DISEASE

In the recent years a lot of emphasis has been placed on the discovery and better detection of clinically relevant biomarkers. Biomarkers are crucial for the early detection of several diseases, and they play an important role in the improvement of current treatments, thus reducing patient mortality rate. Biofluids account for 60% of the human body mass and can be a goldmine of significant biomarkers. Unfortunately, low abundance biomarkers are difficult to detect with mass-spectrometry or immunoassays because of their low concentration in body fluids, their lability, and the presence of high-abundance proteins (i.e. albumin and immunoglobulins). In order to overcome these physiological barriers, we developed nanoparticles made of poly(N-isopropylacrylamide) (NIPAm) and functionalized with affinity reactive baits that one single step capture, concentrate and preserve labile biomarkers in complex body fluids (i.e. urine, blood, sweat, CSF). The design, synthesis and application of the Nanotrap hydrogel particles are described in this thesis. The novelty of the technology relies in the fact that in the past hydrogel nanoparticles have been studied and used as a drug delivery tool, whereas our application focuses on their capturing abilities instead of the releasing of specific drug molecules. Once the functionalized nanoparticles are incubated with a biological fluid, low molecular weight biomarkers are captured by the affinity baits while unwanted high abundance analytes are excluded. The potentially relevant biomarkers are then concentrated into small volumes and analyzed. The concentration factor (up to 10000 fold depending on the initial volume) enhances the effective sensitivity of mass-spectrometry and immunoassays and permits to detect previously invisible proteins thus improving biomarker discovery and diagnostic testing. This thesis discusses the use of hydrogel nanoparticles to develop a urinary antigen test for the detection of Lyme Borreliosis. There is a clinical need to improve the diagnostic specificity of early stage Lyme assays in the period prior to the mounting of a robust serology response and to develop a diagnostic tool to monitor therapy success. Borrelia burgdorferi is the causative agent of Lyme disease. Using our hydrogel particles (Nanotraps) we evaluated the presence of urinary Borrelia Outer surface protein A (OspA) C-terminus peptide in early stage LB before and after treatment, and in patients suspected of late stage disseminated LB. We employed Nanotraps to concentrate urinary OspA and used a highly specific anti-OspA monoclonal antibody (mAb) as a detector of the C-terminus peptides. We mapped the mAb epitope to a narrow specific OspA C-terminal domain OspA236-239 conserved across infectious Borrelia species but with no homology to human proteins and no cross reactivity with relevant viral and non-Borrelia bacterial proteins. 268 urine samples from patients being evaluated for all categories of LB were collected in a LB endemic area. The urinary OspA assay, blinded to outcome, utilized Nanotrap particle pre-processing, western blotting to evaluate the OspA molecular size, and OspA peptide competition for confirmation. OspA test characteristics: sensitivity 1.7 pg/mL (lowest limit of detection), %coefficient of variation (CV)=8%, dynamic range 1.7-30 pg/mL. Pre-treatment, 24/24 newly diagnosed patients with an erythema migrans (EM) rash were positive for urinary OspA while false positives for asymptomatic patients were 0/117 (Chi squared p<10-6). For 10 patients who exhibited persistence of the EM rash during the course of antibiotic therapy, 10/10 were positive for urinary OspA. Urinary OspA of 8/8 patients switched from detectable to undetectable following symptom resolution post-treatment. Specificity of the urinary OspA test for the clinical symptoms was 40/40. Specificity of the urinary OspA antigen test for later serology outcome was 87.5% (21 urinary OspA positive/24 serology positive, Chi squared p=4.072e-15). 41 of 100 patients under surveillance for persistent LB in an endemic area were positive for urinary OspA protein. OspA urinary shedding was strongly linked to concurrent active symptoms (e.g. EM rash and arthritis), while resolution of these symptoms after therapy correlated with urinary conversion to OspA negative. Detection of OspA was performed using Western blot analysis. In order to obtain a quantitative measurement of the antigen, an ELISA was developed. Preliminary results showed a lowest limit of detection of 0.5pg/ml and %coefficient of variation 2%, dynamic range 0.5pg-30 pg/ml. 3/3 of symptomatic patients that resulted positive with the western blot Lyme assay were also found positive when tested on ELISA. Another promising format under development uses Mass Spectrometry Multiple Reaction Monitoring (MRM) for the detection of multiple Borrelia proteins after Nanotrap processing. Developing a diagnostic test against a panel of analytes will improve clinical sensitivity and understanding of staging of disease. MRM is a prime technology that yield multiplex measurement of more than 100 peptides in a single sample. A sensitivity of 5pg/ml and high reproducibility in human urine spiked with OspA was observed. Lastly, partially degradable Nanotraps were employed to produce a prototype of Lateral Flow Immunoassay (LFI) which exploits the use of antigen displaying nanoparticles for a point of care test for Lyme disease. This technology will ensure high accuracy and sensitivity while allowing for rapid testing of Lyme disease antigens in the urine of patients in the doctor office. In summary, this study presents data supporting the successful use of the Nanotrap technology to develop a more accurate and sensitive test for Lyme disease that can diagnose the disease before seroconversion and that can be used to monitor therapy success. Nanotraps increase the effective analytical sensitivity of western blot analysis, ELISA, mass spectrometry MRM and lateral flow immunoassay. This is a concept that can be extended to communicable diseases with different etiologic agents (e.g. Tuberculosis, Chagas disease, Toxoplasmosis, etc.)

The troublesome ticks research protocol: Developing a comprehensive, multidiscipline research plan for investigating human tick-associated disease in Australia

In Australia, there is a paucity of data about the extent and impact of zoonotic tick-related illnesses. Even less is understood about a multifaceted illness referred to as Debilitating Symptom Complexes Attributed to Ticks (DSCATT). Here, we describe a research plan for investigating the aetiology, pathophysiology, and clinical outcomes of human tick-associated disease in Australia. Our approach focuses on the transmission of potential pathogens and the immunological responses of the patient after a tick bite. The protocol is strengthened by prospective data collection, the recruitment of two external matched control groups, and sophisticated integrative data analysis which, collectively, will allow the robust demonstration of associations between a tick bite and the development of clinical and pathological abnormalities. Various laboratory analyses are performed including metagenomics to investigate the potential transmission of bacteria, protozoa and/or viruses during tick bite. In addition, multi-omics technology is applied to investigate links between host immune responses and potential infectious and non-infectious disease causations. Psychometric profiling is also used to investigate whether psychological attributes influence symptom development. This research will fill important knowledge gaps about tick-borne diseases. Ultimately, we hope the results will promote improved diagnostic outcomes, and inform the safe management and treatment of patients bitten by ticks in Australia

Environmental risk factors in infectious diseases: studies in waterborne disease outbreaks, Ebola, and Lyme disease

Thesis (Ph.D.)--Boston UniversityThe resurgence of infectious diseases and global climate change's potential impact on them has refocused public health's attention on the environment's role in infectious disease. The studies in this dissertation utilize the increased availability of satellite image-derived data sets with fine temporal and geographic granularity and the expansion of epidemiologic methods to explore the relationship between the environment and infectious disease in three settings. The first study employed a novel study design and analytic methods to investigate the hypothesis that heavy rainfall is an independent risk factor for waterborne disease outbreaks (WBDOs). We found that a location experiencing a heavy rainfall event had about half the odds of a WBDO two or four weeks later than did a location without a heavy rainfall event. The location-based case-crossover study design utilized in this study may help to expand the research methods available to epidemiologists working in this developing field. The second study employed a location-based case-crossover study design to evaluate standardized differences from historic average of weekly rainfall in locations with a recorded introduction of Ebola into a human. For each 1.0 unit z-score decrease in total rainfall, the odds of an Ebola introduction three weeks later increased by 75%. Given the severity of Ebola outbreaks and the dearth of knowledge about indicators of increased risk, this finding is an important step in advancing our understanding of Ebola ecology. The third study used GIS methods on remote sensing data to estimate the association between peridomestic forest/non-forest interface within 100, 150, 250 meters and Lyme-associated peripheral facial palsy (LAPFP) among pediatric facial palsy patients. After adjustment for sex, age, and socio-economic status, children with the highest level of forest edge in the three radii of analysis had 2.74 (95% CI 1.15, 6.53), 4.58 (1.84, 11.41), and 5.88 (2.11, 16.4) times the odds of LAPFP compared to children with zero forest edge in those radii. This study is the first to examine environmental risk factors for LAPFP. Each of these studies advances the techniques used to investigate environmental risk factors for infectious disease through study design, case definition, data used, or exposure definitions

A Guide to Utilization of the Microbiology Laboratory for Diagnosis of Infectious Diseases: 2013 Recommendations by the Infectious Diseases Society of America (IDSA) and the American Society for Microbiology (ASM)a

The critical role of the microbiology laboratory in infectious disease diagnosis calls for a close, positive working relationship between the physician and the microbiologists who provide enormous value to the health care team. This document, developed by both laboratory and clinical experts, provides information on which tests are valuable and in which contexts, and on tests that add little or no value for diagnostic decisions. Sections are divided into anatomic systems, including Bloodstream Infections and Infections of the Cardiovascular System, Central Nervous System Infections, Ocular Infections, Soft Tissue Infections of the Head and Neck, Upper Respiratory Infections, Lower Respiratory Tract infections, Infections of the Gastrointestinal Tract, Intraabdominal Infections, Bone and Joint Infections, Urinary Tract Infections, Genital Infections, and Skin and Soft Tissue Infections; or into etiologic agent groups, including Tickborne Infections, Viral Syndromes, and Blood and Tissue Parasite Infections. Each section contains introductory concepts, a summary of key points, and detailed tables that list suspected agents; the most reliable tests to order; the samples (and volumes) to collect in order of preference; specimen transport devices, procedures, times, and temperatures; and detailed notes on specific issues regarding the test methods, such as when tests are likely to require a specialized laboratory or have prolonged turnaround times. There is redundancy among the tables and sections, as many agents and assay choices overlap. The document is intended to serve as a reference to guide physicians in choosing tests that will aid them to diagnose infectious diseases in their patients