Infectious Disease Diagnostics Research Articles

Fighting Antibiotic Resistance in Hospital-Acquired Infections: Current State and Emerging Technologies in Disease Prevention, Diagnostics and Therapy.

Rising antibiotic resistance is a global threat that is projected to cause more deaths than all cancers combined by 2050. In this review, we set to summarize the current state of antibiotic resistance, and to give an overview of the emerging technologies aimed to escape the pre-antibiotic era recurrence. We conducted a comprehensive literature survey of >150 original research and review articles indexed in the Web of Science using “antimicrobial resistance,” “diagnostics,” “therapeutics,” “disinfection,” “nosocomial infections,” “ESKAPE pathogens” as key words. We discuss the impact of nosocomial infections on the spread of multi-drug resistant bacteria, give an overview over existing and developing strategies for faster diagnostics of infectious diseases, review current and novel approaches in therapy of infectious diseases, and finally discuss strategies for hospital disinfection to prevent MDR bacteria spread.

Portable Pathogen Diagnostics Using Microfluidic Cartridges Made from Continuous Liquid Interface Production Additive Manufacturing.

Biomedical diagnostics based on microfluidic devices have the potential to significantly benefit human health; however, the manufacturing of microfluidic devices is a key limitation to their widespread adoption. Outbreaks of infectious disease continue to demonstrate the need for simple, sensitive, and translatable tests for point-of-care use. Additive manufacturing (AM) is an attractive alternative to conventional approaches for microfluidic device manufacturing based on injection molding; however, there is a need for development and validation of new AM process capabilities and materials that are compatible with microfluidic diagnostics. In this paper, we demonstrate the development and characterization of AM cartridges using continuous liquid interface production (CLIP) and investigate process characteristics and capabilities of the AM microfluidic device manufacturing. We find that CLIP accurately produces microfluidic channels as small as 400 μm and that it is possible to routinely produce fluid channels as small as 100 μm with high repeatability. We also developed a loop-mediated isothermal amplification (LAMP) assay for detection of E. coli from whole blood directly on the CLIP-based AM microfluidic cartridges, with a 50 cfu/μL limit of detection, validating the use of CLIP processes and materials for pathogen detection. The portable diagnostic platform presented in this paper could be used to investigate and validate other AM processes for microfluidic diagnostics and could be an important component of scaling up the diagnostics for current and future infectious diseases and pandemics.

Powering Diagnostics for Infectious Disease: The COVID-19 pandemic saw a rush of companies developing diagnostics for the infectious disease that should benefit the detection of other diseases

Clinical OMICsVol. 8, No. 4 NewsPowering Diagnostics for Infectious Disease: The COVID-19 pandemic saw a rush of companies developing diagnostics for the infectious disease that should benefit the detection of other diseasesMalorye Allison BrancaContributing EditorMalorye Allison BrancaContributing EditorSearch for more papers by this authorPublished Online:26 Aug 2021https://doi.org/10.1089/clinomi.08.04.24AboutSectionsView articleView Full TextPDF/EPUB Permissions & CitationsPermissionsDownload CitationsTrack CitationsAdd to favorites Back To Publication ShareShare onFacebookTwitterLinked InRedditEmail View articleFiguresReferencesRelatedDetails Volume 8Issue 4Jul 2021 InformationCopyright © GEN PublishingTo cite this article:Malorye Allison BrancaContributing Editor.Powering Diagnostics for Infectious Disease: The COVID-19 pandemic saw a rush of companies developing diagnostics for the infectious disease that should benefit the detection of other diseases.Clinical OMICs.Jul 2021.36-39.http://doi.org/10.1089/clinomi.08.04.24Published in Volume: 8 Issue 4: August 26, 2021PDF download

EpitopeVec: linear epitope prediction using deep protein sequence embeddings.

MotivationB-cell epitopes (BCEs) play a pivotal role in the development of peptide vaccines, immuno-diagnostic reagents and antibody production, and thus in infectious disease prevention and diagnostics in general. Experimental methods used to determine BCEs are costly and time-consuming. Therefore, it is essential to develop computational methods for the rapid identification of BCEs. Although several computational methods have been developed for this task, generalizability is still a major concern, where cross-testing of the classifiers trained and tested on different datasets has revealed accuracies of 51–53%.ResultsWe describe a new method called EpitopeVec, which uses a combination of residue properties, modified antigenicity scales, and protein language model-based representations (protein vectors) as features of peptides for linear BCE predictions. Extensive benchmarking of EpitopeVec and other state-of-the-art methods for linear BCE prediction on several large and small datasets, as well as cross-testing, demonstrated an improvement in the performance of EpitopeVec over other methods in terms of accuracy and area under the curve. As the predictive performance depended on the species origin of the respective antigens (viral, bacterial and eukaryotic), we also trained our method on a large viral dataset to create a dedicated linear viral BCE predictor with improved cross-testing performance.Availability and implementationThe software is available at https://github.com/hzi-bifo/epitope-prediction.Supplementary information Supplementary data are available at Bioinformatics online.

The importance of molecular diagnostics for infectious diseases in low-resource settings.

In settings with limited resources and a wide range of possible etiologies, molecular technologies offer an effective solution for infectious disease diagnostics, because they are agile, fast and flexible. Health systems that routinely use molecular diagnostics will achieve economies of scale, maximize limited expertize and rapidly respond to new threats.

Genome editing of a hybridoma cell line via the CRISPR/Cas9 system: A new approach for constitutive high-level expression of heterologous proteins in eukaryotic system

The power of the CRISPR/Cas9 system has revolutionized genome editing in many fields of biology. These applications have expanded exponentially over recent years, including those regarding protein expression technologies. The CRISPR/Cas9 system avoids random integration of the gene of interest and due to this characteristic can be exploited to obtain a stable cell line for the high-yield expression of recombinant proteins. Here we propose a method to edit a hybridoma cell line for the constitutive expression of proteins of interest using the CRISPR/Cas9 system. First, with the scope of optimizing the method, we replaced part of the light chain of immunoglobulin with the Green Fluorescent Protein (GFP) gene, obtaining a precise knock-in in the hybridoma genome. We confirmed the expression and secretion of GFP into the culture medium via fluorimetric analysis, as well as correct genome editing by RNA sequencing. Then, using the same approach, we included the gene encoding a protein of diagnostic interest, the Bovine Herpesvirus 1 glycoprotein E, in the donor DNA. We obtained a stable clone able to secrete gE protein in fusion with GFP into the culture medium. This result was confirmed by ELISA and Western Blot analysis. This study confirms the suitability of this cell line for the production of proteins of diagnostic interest by stable gene expression in a mammalian system. These experiments will enable the technique to be developed from its proof of concept to more specific applications in the field of infectious disease diagnostics.

Fast Antibiotic Susceptibility Testing via Raman Microspectrometry on Single Bacteria: An MRSA Case Study.

Despite recent advances in molecular diagnostics, ultrafast determination of the antibiotic susceptibility phenotype of pathogenic microorganisms is still a major challenge of in vitro diagnostics (IVD) of infectious diseases. Raman microspectroscopy has been proposed as a means to achieve this goal. Previous studies have shown that susceptibility phenotyping could be done through Raman analysis of microbial cells, either in large clusters or down to the single-cell level in the case of Gram-negative rods. Gram-positive cocci such as Staphylococcus aureus pose several challenges due to their size and their different metabolic and chemical characteristics. Using a tailored automated single-cell Raman spectrometer and a previously proposed sample preparation protocol, we acquired and analyzed 9429 S. aureus single cells belonging to three cefoxitin-resistant strains and two susceptible strains during their incubation in the presence of various concentrations of cefoxitin. We observed an effect on S. aureus spectra that is weaker than what was detected on previous bacteria/drug combinations, with a higher cell-to-cell response variability and an important impact of incubation conditions on the phenotypic resistance of a given strain. Overall, the proposed protocol was able to correlate strains’ phenotype with a specific modification of the spectra using majority votes. We, hence, confirm that our previous results on single-cell Raman antibiotic susceptibility testing can be extended to the S. aureus case and further clarify potential limitations and development requirements of this approach in the move toward industrial applications.

Biological and clinical significance of quorum sensing alkylquinolones: current analytical and bioanalytical methods for their quantification.

Quorum sensing (QS) is a sophisticated bacterial communication system which plays a key role in the virulence and biofilm formation of many pathogens. The Pseudomonas aeruginosa QS network consists of four sets of connected systems (las, rlh, pqs and iqs) hierarchically organized. The pqs system involves characteristic autoinducers (AI), most of them sharing an alkylquinolone (AQ) structure, and is able to carry out several relevant biological functions besides its main signalling activity. Their role in bacterial physiology and pathogenicity has been widely studied. Indeed, the presence of these metabolites in several body fluids and infected tissues has pointed to their potential value as biomarkers of infection. In this review, we summarize the most recent findings about the biological implications and the clinical significance of the main P. aeruginosa AQs. These findings have encouraged the development of analytical and bioanalytical techniques addressed to assess the role of these metabolites in bacterial growth and survival, during pathogenesis or as biomarkers of infections. The availability of highly sensitive reliable analytical methods suitable for clinical analysis would allow getting knowledge about pathogenesis and disease prognosis or progression, supporting clinicians on the decision-making process for the management of these infections and guiding them on the application of more effective and appropriate treatments. The benefits from the implementation of the point-of-care (PoC)-type testing in infectious disease diagnostics, which are seen to improve patient outcomes by promoting earlier therapeutic interventions, are also discussed.

An integrated E-Tube cap for sample preparation, isothermal amplification and label-free electrochemical detection of DNA

A simple, disposable, and integrated electronic-tube cap (E-tube cap) for DNA detection at the point-of-care was designed, fabricated, and tested. The E-tube cap contains a 3D printed electrode substrate for DNA extraction and label-free pH sensing detection. One Flinders Technology Associates (Whatman FTA) membrane was incorporated into the 3D printed electrode substrate for the isolation, concentration, and purification of DNA. The E-tube cap with captured DNA by the membrane was inserted directly into a reaction tube for loop-mediated isothermal amplification (LAMP). The isothermal amplification process was monitored in real-time by a 3D printed electrochemical electrode coated with pH-sensitive material (carbon/iridium oxide layer). The pH sensing electrode showed an excellent linear response within the pH range of 6–9 with a slope of −31.32 ± 0.5 mV/pH at room temperature. The utility of the integrated E-tube cap was demonstrated by detecting the presence of lambda DNA spiked in saliva samples with a sensitivity of 100 copies per mL sample within 30 min. Such a simple, rapid, and affordable diagnostic device is particularly suitable for point-of-care molecular diagnostics of infectious diseases.

RNA-extraction-free nano-amplified colorimetric test for point-of-care clinical diagnosis of COVID-19.

The global pandemic of coronavirus disease 2019 (COVID-19) highlights the shortcomings of the current testing paradigm for viral disease diagnostics. Here, we report a stepwise protocol for an RNA-extraction-free nano-amplified colorimetric test for rapid and naked-eye molecular diagnosis of COVID-19. The test employs a unique dual-prong approach that integrates nucleic acid (NA) amplification and plasmonic sensing for point-of-care detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), with a sample-to-assay response time of <1 h. The RNA-extraction-free nano-amplified colorimetric test utilizes plasmonic gold nanoparticles capped with antisense oligonucleotides (ASOs) as a colorimetric reporter to detect the amplified nucleic acid from the COVID-19 causative virus, SARS-CoV-2. The ASOs are specific for the SARS-CoV-2 N-gene, and binding of the ASOs to their target sequence results in the aggregation of the plasmonic gold nanoparticles. This highly specific agglomeration step leads to a change in the plasmonic response of the nanoparticles. Furthermore, when tested using clinical samples, the accuracy, sensitivity and specificity of the test were found to be >98.4%, >96.6% and 100%, respectively, with a detection limit of 10 copies/μL. The test can easily be adapted to diagnose other viral infections with a simple modification of the ASOs and primer sequences. It also provides a low-cost, rapid approach requiring minimal instrumentation that can be used as a screening tool for the diagnosis of COVID-19 at point-of-care settings in resource-poor situations. The colorimetric readout of the test can even be monitored using a handheld optical reader to obtain a quantitative response. Therefore, we anticipate that this protocol will be widely useful for the development of biosensors for the molecular diagnostics of COVID-19 and other infectious diseases.

A practitioner's guide to understanding infectious disease diagnostics in the United Kingdom. Part 2: Serological diagnostic testing methods and diagnostic test result interpretation

SummaryThis article is the second in a review series about infectious disease diagnostics and summarises serological testing techniques, diagnostic test validation and result interpretation. It highlights the requirement for results to be interpreted with testing limitations in mind and in conjunction with the case's clinical presentation. With these foundations, the treating clinician should always be well placed to deal with equine infectious diseases.

Challenges in the morphological diagnostics of infectious lung diseases among HIV-infected patients: a case-series study

HIV-infection is a long-term infectious disease, characterized by deterioration of the immune system and it is often complicated by the development of opportunistic diseases, mainly of infectious origin. HIV-associated infections become generalized in the late stages of the disease and often may combine, that makes challenges in the diagnosis of these diseases. Differential morphological diagnosis of HIV-associated lung infections is quite complex. Most frequently, there is the need to differentiate between tuberculosis, bacterial or septic pneumonia, cytomegalovirus lung lesions, and deep mycoses. The aim. To demonstrate the expediency and necessity to apply special complex morphological methods in HIV-infected patients with secondary lung lesions using the series of clinical cases as an example. Materials and methods. The case series consisted of 7 cases with discordance between the clinical-radiologic, laboratory data and histological conclusions. Of these, there were 5 cases of life-time and 2 autopsy diagnoses. All the clinical and laboratory data of these cases were retrospectively analyzed, and a number of additional special histological examinations were performed. The Ziehl–Neelsen and fluorescent methods, Gomori–Grocot and alcian blue-PAS staining as well as immunohistochemical examinations were used for the detection of mycobacterial antigens with different antibodies to M. tuberculosis. Results. The case series of initial morphological misdiagnosis or indeterminate histological diagnosis was revealed among hospitalized HIV-infected patients. Clinical data did not provide an accurate diagnosis of pulmonary pathology. According to the traditional histological examination, a tuberculous process was diagnosed in 3 cases, a chronic granulomatous process – in 2 cases, but without specifying the etiology. In two cases of autopsy, pneumocystis pneumonia was suspected in combination with other infections. Additional morphological examination with using 2–6 special methods made it possible to determine the correct diagnosis. In 2 cases of provisional diagnosis of tuberculosis, invasive cryptococcosis and “pneumonia in a drug user” were revealed. Cryptococcosis and pneumocystis pneumonia were detected in 2 cases with an unspecified process. In the autopsy cases, the diagnosis of pneumocystis pneumonia was ruled out, while viral-bacterial pneumonia and tuberculosis associated with cytomegalovirus infection were detected. Conclusions. The presented factual material demonstrates the principal importance of using a comprehensive morphological examination in cases of lung inflammatory processes in HIV-infected patients. Clinical patient data, namely HIV-status at the time of diagnostic procedures as well as radiological and microbiological findings serve as a substantial aid in the diagnosis.

A practitioner's guide to understanding equine infectious disease diagnostics in the United Kingdom. Part 1: How to optimise sampling approaches and a guide to agent detection testing methods

SummaryDiagnostic testing is routinely performed by the equine clinician when dealing with suspected infectious disease cases and outbreaks. Optimal sample timing, choice and handling are fundamental to attain an accurate diagnosis, and a good understanding of laboratory‐based sample analysis techniques, and their validation is necessary for effective diagnostic test result interpretation. This two‐part series highlights the importance of interpreting results bearing testing limitations and specific clinical findings in mind, and on these foundations, the treating clinician should always be well placed to deal with equine infectious diseases. Part 1 in this series will provide a treating clinician with an overview of the importance of testing horses in infectious disease outbreaks and how this is achieved. The different laboratory testing options available for agent detection and their methods will also be discussed. Part 2 will summarise serological (antibody) testing techniques, sample processing (including how tests are performed and validated) and result interpretation.

THE ROLE OF “CLINICAL PRACTICE” DISCIPLINE IN PREPARATION OF FUTURE SPECIALISTS OF HIGHER MEDICAL EDUCATIONAL INSTITUTIONS

Резюме. Клінічна практика студентів-медиків одна із найважливіших складових професійної підготовки будь-якого спеціаліста і є центральним елементом викладання і вивчення навчальної програми. Клінічна практика з терапії є процесом закріплення теоретичних знань, практичних умінь і навичок, отриманих в процесі навчання студента, і формування універсальних та професійних компетенцій майбутнього лікаря-терапевта, сімейного лікаря, лікаря загальної практики, тобто набуття досвіду у вирішенні реальних професійних завдань.&#x0D; На кафедрі внутрішньої медицини №2 та медсестринства із 2020-2021 навчального року введена вибіркова дисципліна «Клінічна практика в терапії» для студентів 5 курсу медичного факультету (спеціальність «лікувальна справа», «педіатрія»). Клінічна практика в терапії складається із трьох змістових модулів: «Основні методи обстеження та діагностики в кардіології», «Особливості обстеження та діагностики в ревматології, гастроентерології, нефрології. Діагностика основних інфекційних хворіб», «Клінічна практика в пульмонології, ендокринології, гематології».&#x0D; На практичних заняттях велика увага приділяється виконанню практичних навичок і умінню розв’язувати ситуаційні задачі. Студенти вдосконалюють свої навички з фізикального обстеження всіх органів і систем та встановлюють ступінь їх порушення, навчаються реєстрації електрокардіограми, проводити серцево-легеневу реанімацію, пікфлуометрію, аспірацію кісткового мозку, інтерпретувати результати лабораторних та інструментальних методів дослідження, обґрунтовувати та формулювати попередній діагноз, призначити лікування і вибрати подальший маршрут пацієнта залежно від діагностованої патології.

Molecular genetic techniques in current biomedical research. Part II: PCR applications in diagnostics of human infectious diseases

Polymerase chain reaction (PCR)-based diagnostics is currently established as a gold standard for the detection of microorganisms. The features of PCR include rapid amplification of DNA and RNA as well as high sensitivity and specificity. In contrast to diagnostic microbiology, PCR diagnostics does not require preliminary culture of the microorganisms for their identification, reducing both time and costs of the diagnostic procedure. The lecture discusses the molecular basis behind the modern technical solutions for the PCR diagnostics of human infectious diseases including multiplex and reverse transcription PCR. We describe the principles of qualitative and quantitative PCR-based detection of pathogens in biological samples and provide the examples of PCR application for solving specific diagnostic scenarios. The lecture is primarily designed for students of biomedical specialties and healthcare professionals using molecular genetic techniques in their practice.

Engineering the interface between electroactive bacteria and electrodes

Chelsea Catania is a Postdoctoral Associate in the Department of Mechanical Engineering at the Massachusetts Institute of Technology. She received a BS in Materials Science and Engineering from the University of Florida. She was an NSF graduate research fellow and obtained her PhD in Materials Science from the University of California, Santa Barbara. Her research interests are in the modification of biotic-abiotic interfaces for microbial electrochemical technologies and human health. Amruta A. Karbelkar is a Postdoctoral Associate in the Department of Chemical Engineering at the Massachusetts Institute of Technology. She obtained her MS in Chemistry from the Institute of Chemical Technology, Mumbai, India. She obtained her PhD in Chemistry from the University of Southern California, Los Angeles. Her research focuses on developing electrochemical platforms to study microbial electron transfer and diagnostics for infectious diseases. Ariel L. Furst is an Assistant Professor of Chemical Engineering at the Massachusetts Institute of Technology. She obtained her BS in Chemistry from the University of Chicago and her PhD in Chemistry from the California Institute of Technology. She was then an Arnold O. Beckman Postdoctoral Fellow at the University of California, Berkeley. Her lab combines electrochemical methods with biomolecular and materials engineering to address challenges in human health and clean energy.

Application of fluorescein copolymer to improve the efficiency of counter-immunoelectrophoresis for diagnostics of animal infectious diseases

Application of fluorescein copolymer to improve the efficiency of counter-immunoelectrophoresis for diagnostics of animal infectious diseases

Less is more… in infectious diseases

The increasing development of antimicrobial resistance has been identified as one of the greatest threats to public health and is caused to arelevant extent by falsely indicated antibiotic treatment. The main aim of this article is to identify areas in infectious disease diagnostics and treatment where overuse occurs and to provide recommendations on how to avoid it. The authors identified current and relevant studies on the topic of medical overuse in infectious diseases via a literature search. In particular, contributions from international "less is more" initiatives were included. The focus was on areas in which areduction of diagnostic and therapeutic measures leads to an optimization of patient outcomes. In many cases overuse in the context of diagnostics and treatment of infectious diseases not only leads to an unnecessary financial burden on the healthcare system and is not beneficial but can also increase the risk of development of antimicrobial resistance and have adverse consequences for patients. Correct indications as well as focused selection and adequate application of antimicrobial agents is crucial to provide the best possible medical care. Diagnostic and antibiotic stewardship measures, which should be implemented in collaboration with infectious disease specialists, can help to identify and reduce areas of overuse and misuse.

Point-of-care diagnostics for infectious diseases: From methods to devices.

The current widespread of COVID-19 all over the world, which is caused by SARS-CoV-2 virus, has again emphasized the importance of development of point-of-care (POC) diagnostics for timely prevention and control of the pandemic. Compared with labor- and time-consuming traditional diagnostic methods, POC diagnostics exhibit several advantages such as faster diagnostic speed, better sensitivity and specificity, lower cost, higher efficiency and ability of on-site detection. To achieve POC diagnostics, developing POC detection methods and correlated POC devices is the key and should be given top priority. The fast development of microfluidics, micro electro-mechanical systems (MEMS) technology, nanotechnology and materials science, have benefited the production of a series of portable, miniaturized, low cost and highly integrated POC devices for POC diagnostics of various infectious diseases. In this review, various POC detection methods for the diagnosis of infectious diseases, including electrochemical biosensors, fluorescence biosensors, surface-enhanced Raman scattering (SERS)-based biosensors, colorimetric biosensors, chemiluminiscence biosensors, surface plasmon resonance (SPR)-based biosensors, and magnetic biosensors, were first summarized. Then, recent progresses in the development of POC devices including lab-on-a-chip (LOC) devices, lab-on-a-disc (LOAD) devices, microfluidic paper-based analytical devices (μPADs), lateral flow devices, miniaturized PCR devices, and isothermal nucleic acid amplification (INAA) devices, were systematically discussed. Finally, the challenges and future perspectives for the design and development of POC detection methods and correlated devices were presented. The ultimate goal of this review is to provide new insights and directions for the future development of POC diagnostics for the management of infectious diseases and contribute to the prevention and control of infectious pandemics like COVID-19.

The role of cardioenzymes in the diagnostics of infectious heart diseases

Purpose: to evaluate the importance of increasing cardioenzymes in the diagnosis of infectious heart lesions.Object and methods: Under observation were 71 children aged 3 to 17 years (average age – 6.6 ± 0.4 years) who received treatment at the clinic of Pediatric Research and Clinical Center for Infectious Diseases. All children in the acute period of illness and 49 patients with follow-up history were identified with levels of creatine phosphokinase (CPK), MВ fractions of creatine phosphokinase (CPK-MВ), lactate dehydrogenase (LDH), C-reactive protein (CRP), aspartate aminotransferase (AST). Follow-up observation and examination of patients was performed after 1 month and after 1 year. All children were examined for a wide range of pathogens of acute respiratory viral infections, active herpes virus infections (EBV, CMV, HHV-6 type), B19 parvovirus, Chlamydophila pneumoniae, Mycoplasma pneumonia, enteroviruses and bacterial respiratory pathogens. All examined patients underwent electrocardiography and echocardiography.The results of the study. In the examined patients with an increased level of “cardioenzymes” (CPK-MВ, LDH, AST) against the background of acute respiratory infection (ARI), a wide range of diseases was established with damage to both the upper respiratory tract and lower (bronchitis, bronchiolitis, pneumonia), and also other infectious nosologies, including respiratory syndrome (infectious mononucleosis, enterovirus and parvovirus infection). The etiological structure was dominated by a group of herpesvirus (53%) and bacterial infections (25%), as well as their combinations. An analysis of the dynamics of the main biochemical and hematological parameters characterizing the severity of the systemic inflammation syndrome (leukocytes, SRE, CRP, CPK), as well as reflecting myocardial injuries and used in cardiology practice (CPK- MВ, LDH, AST), revealed a long-term (within 1 year observations) the preservation of a moderately elevated level of CPK-MВ, unlike other indicators that returned to normal within 1 month. An increase in CPK-MВ was recorded in 79% of patients with drip infections, while the average level of CPK-MВ in the total sample exceeded the normal values by 1.8-2.4 times.Conclusion. It was found that elevated levels of CPK-MВ and LDH can be used as criteria for the formation of dynamic observation groups and early rehabilitation.