Molecular Diagnostic Technologies Research Articles

A Novel High-Throughput Sample-in-Result-Out Device for the Rapid Detection of Viral Nucleic Acids

Clustered regularly interspaced short palindromic repeats (CRISPR) molecular diagnostic technology is one of the most reliable diagnostic tools for infectious diseases due to its short reaction time, high sensitivity, and excellent specificity. However, compared with fluorescent polymerase chain reaction (PCR) technology, CRISPR molecular diagnostic technology lacks high-throughput automated instrumentation and standardized detection reagents for high sensitivity, limiting its large-scale clinical application. In this study, a high-throughput automated device was developed by combining reagent lyophilization, extraction-free technology, and a one-pot consumable system. This innovative approach enabled the rapid sample-in-result-out detection of 48 samples in 25 min and demonstrated high sensitivity and specificity for the qualitative analysis of clinical samples. The obtained results show that the detection limit of the designed system for African swine fever virus (ASFV) is 0.5 copies/μL. As a proof concept, a single-tube dual-target nucleic acid detection method was developed, achieving a detection limit of 5 copies/μL for the ORF1ab and N genes of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) within 45 min. The method is highly specific, reliable, and stable, providing a feasible solution for the clinical application of CRISPR nucleic acid detection technology.

At the mercy of the AI algorithm - discrepant PCR results.

As pathogen molecular diagnostics technology develops, the software analysis of polymerase chain reaction (PCR) sigmoidal curves, crossing threshold (Ct) values and the limit of nucleic acid detection for that assay, is not always available when reporting test results. These are critical components in interpreting results and can affect the management of patients. Our view is that all commercial assay providers must ensure users, as they can with laboratory developed tests, are able to see the curve and Ct value for all samples. Some examples demonstrating this principle are included. Regulatory bodies have a key role to play.

Companion Diagnostics (CDx) Based on Molecular Biology Techniques

Molecular profiling based on genomic mutations provides clinically important diagnostic and prognostic information. Companion diagnostic (CDx) testing, which is based on targeted drug therapy, is being applied to a variety of molecular diagnostic techniques (e.g., fluorescent in situ hybridization—FISH; polymerase chain reaction—PCR; and next-generation sequencing—NGS) to diagnose complex etiologies using a minimal number of specimens, replacing immunohistochemical analysis, which may show bias at certain stages. The safety and effectiveness of CDx testing using molecular diagnostic technology in precision medicine is an important factor in determining the treatment outcome and prognosis of patients. Meeting minimum safety and effectiveness performance standards is essential for CDx testing, and a thorough understanding of regulatory considerations is necessary to plan and design the optimal product. In this review, we focus on the diagnostic field of precision medicine and discuss the safety and effectiveness that each molecular diagnostic technology must meet according to CDx testing diversity.

Expert consensus for clinical application of molecular diagnosis on hepatobiliary tumors (2024 edition)

Molecular diagnosis is of enormous significance for the clinical diagnosis and treatment of hepatobiliary tumors. The expert group first released the "Expert Consensus for Clinical Application of Molecular Diagnosis on Hepatobiliary Tumors" in 2020. The introduction of this consensus has promoted the standardized application of molecular diagnostic technology in the clinical work of hepatobiliary tumors and improved the diagnosis and treatment capabilities of clinical physicians for hepatobiliary tumors. Research on hepatobiliary tumors has gradually been carried out in recent years, with the continuous publication of new research results. Hence, the expert group updated this consensus on the basis of the latest research progress, the original consensus, with a focus on the detection of molecular markers for targeted and immunotherapy of hepatobiliary tumors, in order to provide a reference for clinical physicians in treatment decisions and maximize the benefits for patients.

Research advances in tumor diagnosis and early detection

This review explores recent advances in tumor diagnosis and early detection, focusing on cutting-edge developments in molecular diagnostic technologies, imaging techniques, and the integration of multi-omics data. Current tumor diagnostic methods have limitations in terms of sensitivity and specificity, particularly for early tumor detection. However, with continuous progress in research and emerging technologies, especially the advent of liquid biopsy, which enables the detection of circulating tumor DNA (ctDNA), exosomes, and tumor-educated platelets (TEPs), the sensitivity and accuracy of early cancer detection have significantly improved. Moreover, the combined application of artificial intelligence and high-resolution imaging technology has enhanced the precision of diagnosis. Despite these advances, challenges, such as the high cost of technology and difficulties in data integration, continue to impede widespread clinical adoption. Therefore, I believe that future research should prioritize the innovation of these technologies to improve their applicability for early detection across various cancer types, ultimately contributing to the advancement of personalized therapy.

Shape-Specific Gold Nanoparticles for Multiplex Biosensing Applications.

The biosensing field faces a significant challenge in efficiently detecting multiple analytes in a single diagnostic sample in order to compete with other established multiplex molecular diagnostic technologies such as PCR and ELISA. In response, we have developed a colorimetric nanobiosensor based on multiple morphological forms of functionalized gold nanoparticles (AuNPs) for the simultaneous detection of the influenza virus and SARS-CoV-2 virus. Gold nanospheres (GNSp) were modified with oligonucleotides specific for the influenza A virus, while gold nanoshells (GNSh) were modified with oligonucleotides specific for the SARS-CoV-2 virus. In the presence of their respective targets, AuNPs remain stable due to DNA-DNA interactions; conversely, in the absence of targets, AuNPs aggregate. Consequently, the hybrid system exhibits an indigo color with the SARS-CoV-2 target, a blue color with the Influenza A target, and a purple color with both targets, visible to the naked eye. Analytical sensitivity was 100 nM, and no cross-reactivity was observed with potentially confounding pathogens. This approach holds great promise for the simultaneous identification of multiple pathogens in a rapid manner without the need for equipment or trained personnel.

Successful development of molecular diagnostic technology combining mini-barcoding and high-resolution melting for traditional Chinese medicine agarwood species based on single-nucleotide polymorphism in the chloroplast genome.

Agarwood is a valuable traditional medicine and fragrance. The production process is a typical injury-induced defense response. Currently, there are approximately 22 known species in the genus Aquilaria Lam., all of which can produce agarwood, whereas there are only two legal species of traditional Chinese medicinal agarwood, Aquilaria sinensis (Lour.) Spreng. and Aquilaria agallocha (Lour.) Roxb. The Taiwan herbal Pharmacopoeia of China stipulates that the medicinal agarwood species are A. sinensis and its relatives in the same genus. Moreover, there are five species of agarwood available for clinical medicinal use in Japan, including A. agallocha and A. sinensis, which are often confused with each other or used in a mixed way in the trade process. Therefore, accurate identification of traditional Chinese medicinal agarwood species is important to ensure the authenticity of traditional medicines and to guide the safety of clinical medication. In this study, 59 specific single-nucleotide polymorphism loci were screened and obtained from the chloroplast genomes of 12 species of the genus Aquilaria Lam. We established an identification method for traditional Chinese medicinal agarwood using mini-barcoding combined with high-resolution melting (HRM) and designed and validated 10 pairs of primers from the psbM-trnD, psbA, rps16, petN, ndhE-psaC, rps4, atpE, ycf1, rps15-trnN, and matK regions. The amplification products were all less than 200 bp, with a high success rate of amplification. The method was applied to successfully identify traditional Chinese medicinal agarwood species from commercial agarwood samples. Overall, the sensitivity of this method was sufficient to detect 1% of adulterants in medicinal agarwood products, proving that mini-barcoding HRM is a powerful and flexible tool. This method can be used as a fast and effective high-throughput method for authenticity testing of traditional Chinese medicinal agarwood and its raw materials containing agarwood-containing proprietary Chinese medicines and is recommended for industrial applications.

Multiplex PCR in septic arthritis and periprosthetic joint infections microorganism identification: Results from the application of a new molecular testing diagnostic algorithm.

Pathogen identification is key in the treatment of septic arthritis (SA) and periprosthetic joint infections (PJI). This study evaluates the outcome of the application of a new, score-based SA and PJI diagnostic algorithm, which includes the execution of molecular testing on synovial fluid. A score-based diagnostic algorithm, which includes serologic and synovial fluid markers determination using multiplex PCR (mPCR) and Next Generation Sequencing (NGS) molecular testing, has been applied to a consecutive series of patients with clinically suspected SA or PJI. Patients with a score ≥6 underwent synovial fluid molecular testing, together with traditional culture, to identify the pathogen and its genetically determined antibiotic resistance. One hundred and seventeen joints in 117 patients (62.5% women; average age 73 years) met the criteria for possible SA/PJI. The affected joint was the knee in 87.5% (joint replacement 66.5%; native joint 21%) and the hip in 12.5% (all replaced joints). 43/117 patients (36.7%) were ultimately diagnosed with SA/PJI. Among the various testing technologies applied, mPCR was the main determinant for pathogen identification in 63%, standard culture in 26%, and mNGS in 11%.Staphylococcus aureusand Enterococcus faecalis were the top two microorganisms identified by mPCR, whileStaphylococcus epidermidiswas the prevalent organism identified by NGS. mPCR detected the presence/absence of the genetically determined antibiotic resistance of all identified microorganisms. The average timeframe for pathogen identification was 3.13 h for mPCR, 4.5 days for culture, and 3.2 days for NGS. Molecular diagnostic technologies represent an innovative screening for fast microorganism identification when a joint infection is clinically suspected. Level IV, case series.

The clinical application and laboratory management of molecular genetic diagnosis in children's hospital

AbstractBackgroundMolecular diagnostic technology is the foundation of precision medicine, which has the advantages of good specificity, high sensitivity, strong targeting, rapiddiagnosis, etc. It has a wide range of applications in the field of pediatrics. However, molecular diagnostic technology is characterized by complicated experimental operation, high difficulty in data analysis and interpretation, in consistent standards among laboratories, and difficult standardization of technical processes. Enhancing the application value of molecular diagnostic technology in pediatrics and promoting the high‐quality development of the discipline requires careful consideration by relevant management and professionals.MethodsThis study firstly outlines the development history of molecular diagnostic technology. Then, it analyzes the application of molecular diagnostic technology in the field of pediatrics. Finally, it explores the countermeasures for the management of molecular diagnostic laboratories.ResultsThis study highlights the importance of molecular diagnostic technology in providing information and decision‐making basis for disease prevention, prediction,diagnosis, treatment and regression. It has a wide range of applications in the molecular diagnosis of pediatric hereditary diseases, malignant tumors and infectious diseases. In addition, the countermeasures for the management of molecular diagnostic laboratories are proposed from the five aspects of laboratory, personnel team construction, standardized management,multidisciplinary cross‐discipline, research and translation, safety managementand ethical supervision, and management upgrading and modernization.ConclusionsMolecular diagnostic technology, as the basis of precision medicine, has become one of the important frontier fields in the development of contemporary pediatric medicine. Enhanced laboratory capacity in molecular diagnostic techniques can improve outcomes in the prevention, prediction, diagnosis, treatment, prognosis and research of pediatricdiseases, and lay the groundwork for child healthcare.

Simple, sensitive, and visual detection of 12 respiratory pathogens with one-pot-RPA-CRISPR/Cas12a assay.

Respiratory infections pose a serious threat to global public health, underscoring the urgent need for rapid, accurate, and large-scale diagnostic tools. In recent years, the CRISPR/Cas (clustered regularly interspaced short palindromic repeats/CRISPR-associated) system, combined with isothermal amplification methods, has seen widespread application in nucleic acid testing (NAT). However, achieving a single-tube reaction system containing all necessary components is challenging due to the competitive effects between recombinase polymerase amplification (RPA) and CRISPR/Cas reagents. Furthermore, to enable precision medicine, distinguishing between bacterial and viral infections is essential. Here, we have developed a novel NAT method, termed one-pot-RPA-CRISPR/Cas12a, which combines RPA with CRISPR molecular diagnostic technology, enabling simultaneous detection of 12 common respiratory pathogens, including sixbacteria and sixviruses. RPA and CRISPR/Cas12a reactions are separated by paraffin, providing an independent platform for RPA reactions to generate sufficient target products before being mixed with the CRISPR/Cas12a system. Results can be visually observed under LED blue light. The sensitivity of the one-pot-RPA-CRISPR/Cas12a method is 2.5 × 100copies/μL plasmids, with no cross-reaction with other bacteria or viruses. Additionally, the clinical utility was evaluated by testing clinical isolates of bacteria and virus throat swab samples, demonstrating favorable performance. Thus, our one-pot-RPA-CRISPR/Cas12a method shows immense potential for accurate and large-scale detection of 12 common respiratory pathogens in point-of-care testing.

Abstract 6158: Tumor genomic heterogeneity in non-small cell lung cancer (NSCLC) patients from Latin America

Abstract Motivation: Life prospects of non-small cell lung cancer (NSCLC) patients have greatly improved over the last decades with the adoption of tumor genomic profiling. A better understanding of the mutational landscape has allowed for a more precise molecular characterization and the development of more effective treatment alternatives. However, tumors present a widespread heterogeneity, and many challenges remain to fully comprehend the influencing factors. Of particular importance is the Latin America population which i) is commonly underrepresented in clinical trials and large-scale epidemiological studies, ii) presents higher incidence and mortality rates in comparison to the developed world, and iii) is constituted by a genetically admixed population. Our research attempts to close this gap by investigating the interplay between sociodemographic, clinical, and lifestyle factors, and the effect these exert on the mutational profiles of Latin-American NSCLC patients. Methodology: The study population is derived from the NIRVANA study, a cross-sectional multicenter observational study aiming to characterize and validate molecular diagnostic technologies for NSCLC patients in Chile, Brazil, and Peru. Between July 2015 to October 2018, 5030 NSCLC patients were recruited from 37 centers, meeting inclusion criteria of age ≥18, histologically or cytological proven NSCLC diagnostic, and naïve treatment. Exclusion criteria included prior chemotherapy treatment and refusal to give informed consent. Primary or metastatic NSCLC samples were collected during standard of care biopsy and processed for the DNA and RNA isolation. Next Generation Sequencing (NSG) genomics profiles were obtained using the Oncomine Focus Assay (OFA). Covariates of interest, including sociodemographic, clinical, and lifestyle factors, were assessed at enrollment. QC-approved genomic profiles were obtained for 1864 participants. Results and Conclusions: The prevalence of mutations and fusions in the eight most relevant NSCLC genes (EGFR, KRAS, ALK, MET, RET, BRAF, ROS1 and ERBB2) varies based on sociodemographic, clinical and lifestyle characteristics. Clear distinctions emerged in the prevalence of EGFR, KRAS and ERBB2 mutations among the three countries (EGFR: 20.9%, 17.6%, 35.3%; KRAS: 21.8%, 15.6%, 10.3%; ERBB2: 2.8%, 3.3%, 4.4% for Brazil, Chile, and Peru, respectively). Furthermore, distinct association patterns were identified between the prevalence of genetic alterations and the studied factors, with attributes such as sex, tobacco use and ethnicity mostly influencing the occurrence of EGFR, ALK and ROS1 alterations. These findings provide novel insights into NSCLC studies among Latin-American patients, potentially serving as guidelines for more tailored strategies in the overall management of this underrepresented population. Citation Format: Javiera Garrido, Evelin González, Gonzalo Sepúlveda-Hermosilla, Matias Freire, Alejandro Blanco, Solange Rivas, Katherine Marcelain, Gareth I. Owen, Carolina Ibañez, Alejandro Corvalan, Marcelo Garrido, Rodrigo Assar, Rodrigo Lizana, Javier Cáceres-Molina, Diego Ampuero, Liliana Ramos, Paola Pérez, Osvaldo Aren, Sara Chernilo, Cristina Fernández, María Loreto Spencer, Jacqueline Flores, Giuliano Bernal, Mónica Ahumada, Germán Rasse, Carolina Sánchez, Maria Galli de Amorim, Gabriela Branco, Diana Noronha Nunes, Emmanuel Dias-Neto, Helano C. Freitas, NIRVANA team, Ricardo Armisén. Tumor genomic heterogeneity in non-small cell lung cancer (NSCLC) patients from Latin America [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6158.

Nanozyme-assisted amplification-free CRISPR/Cas system realizes visual detection.

The CRISPR (clustered regularly interspaced short palindromic repeats)/Cas (CRISPR associated) system has proven to be a powerful tool for nucleic acid detection due to its inherent advantages of effective nucleic acid identification and editing capabilities, and is therefore known as the next-generation of molecular diagnostic technology. However, the detection technologies based on CRISPR/Cas systems require preamplification of target analytes; that is, target gene amplification steps through isothermal amplification or PCR before detection to increase target analyte concentrations. This creates a number of testing limitations, such as extended testing time and the need for more sophisticated testing instruments. To overcome the above limitations, various amplification-free assay strategies based on CRISPR/Cas systems have been explored as alternatives, which omit the preamplification step to increase the concentrations of the target analytes. Nanozymes play a pivotal role in enhancing the sensitivity of CRISPR-based detection, enabling visual and rapid CRISPR assays. The utilization of nanozyme exceptional enzyme-like catalytic activity holds great promise for signal amplification in both electrochemical and optical domains, encompassing strategies for electrochemical signal sensors and colorimetric signal sensors. Rather than relying on converting a single detection target analyte into multiple analytes, these methods focus on signal amplification, the main mechanism of which involves the ability to form a large number of reporter molecules or to improve the performance of the sensor. This exploitation of nanozymes for signal amplification results in the heightened sensitivity and accuracy of detection outcomes. In addition to the strategies that improve sensor performance through the application of nanozymes, additional methods are needed to achieve visual signal amplification strategies without preamplification processes. Herein, we review the strategies for improving CRISPR/Cas systems that do not require preamplification, providing a simple, intuitive and preamplification-free CRISPR/Cas system detection platform by improving in-system one-step amplification programs, or enhancing nanozyme-mediated signal amplification strategies.

REVIEW AND ANALYSIS OF ANTHRAX DIAGNOSTIC TOOLS ON THE TERRITORY OF EURASIAN ECONOMIC UNION

This review examines anthrax diagnostic tools registered in the national Veterinary Drugs Registers and Medicines and Medical Devices Registers of the Eurasian Economic Union (EAEU). Actually, throughout the entire territory of the EAEU, there are natural soil foci of anthrax, which causes annual cases of infection of animals and people. Continuous monitoring of the epidemiological situation in the EAEU countries should be ensured by modern diagnostic tools. All diagnostic tools used for epidemiological surveillance must be registered in the national registers of the EAEU countries. For all test systems and diagnostic kits included in the registers of the EAEU countries, the authors provide comparative characteristics that reflect the sensitivity and specificity of diagnostic tools. An analysis of various methods for identifying the anthrax pathogen, focused on modern molecular diagnostic technologies, is presented. The authors discuss the most important problems associated with improving the efficiency of modern methods for detecting the bacterium Bacillus anthracis. Some considerations about the shortcomings in certain areas of early diagnosis of the anthrax microbe are also provided. The authors present a number of proposals that should improve the efficiency of the currently available anthrax monitoring system.

Droplet Digital PCR (ddPCR) Does Not Enhance the Sensitivity of Detection of Cytomegalovirus (CMV) DNA in Newborn Dried Blood Spots Evaluated in the Context of Newborn Congenital CMV (cCMV) Screening.

Congenital cytomegalovirus (cCMV) infection is a leading cause of sensorineural hearing loss (SNHL) and neurodevelopmental disabilities in children worldwide. Some regions in the United States and Canada have implemented universal newborn screening for cCMV, which requires molecular diagnostic technologies for identifying cCMV, such as PCR testing of newborn dried blood spots (DBS). This study aimed to evaluate the sensitivity of droplet digital PCR (ddPCR) compared to quantitative real-time PCR to detect CMV DNA in newborn DBS. The limit of detection of various ddPCR primer/probe combinations (singleplex UL55-HEX, singleplex UL83-FAM, and multiplex UL55-HEX/UL83-FAM) was evaluated using the National Institute of Standards and Technology's (NIST) CMV quantitative standard. Singleplex UL55-HEX ddPCR exhibited the lowest limit of detection among the primer/probe combinations tested for ddPCR. UL55 ddPCR was then compared to real-time PCR in 49 infants with confirmed cCMV identified through newborn screening for CMV in saliva swabs and confirmed by a urine test. The results showed that ddPCR was only positive for 59% (29 out of 49) of the cCMV infants, while real-time PCR was positive for 80% (39 out of 49). Due to its lower sensitivity and throughput, ddPCR may not be suitable for cCMV newborn screening.

Follow that Spore! Airborne Inoculum Detection as a Decision Aid

Plant disease management requires early intervention, which has traditionally required extensive scouting and surveillance to guide decision-making. Visual inspection, unfortunately, requires conditions for disease symptom expression and expertise in identifying causal organisms and may delay management strategies that prevent losses associated with a given plant pathogen. Advances in spore trap technology and molecular diagnostic technologies provide opportunities for greater confidence in pathogen detection and implementation by scientists, farmers, and regulatory agencies. [Formula: see text] Copyright © 2024 The Author(s). This is an open access article distributed under the CC BY 4.0 International license .

Research advances on allergen component-resolved diagnosis in respiratory allergic diseases

Allergen component-resolved diagnosis (CRD) is an emerging molecular diagnostic technology, which can further clarify the protein profile of allergen components in allergic patients, achieve accurate detection of allergens, and have great significance and value for the precise prevention and treatment of allergic diseases. In this article, the CRD technology and its research progress in respiratory allergic diseases are introduced, and the importance of CRD in the evaluation, prevention and treatment of respiratory allergic diseases are discussed.

#5158 AUTOSOMAL DOMINANT TUBULAR KIDNEY DISEASE: FROM PHENOTYPE TO GENOTYPE

Abstract Background and Aims Autosomal dominant tubulointerstitial kidney disease (ADTKD) represent a recently described group of kidney diseases, characterized by chronic interstitial nephritis (CIN), in most cases hereditary, with an autosomal dominant inheritance pattern, subclassified according to its genetic cause, when identified. Mutations in UMOD, MUC1, REN and HNF1B genes have been implicated in their pathogenesis. Recent molecular diagnostic technologies, such as Next-Generation Sequencing (NGS), and particularly the use of restriction and Snapshot method for the detection of a cytosine insertion on the coding variable-number tandem repeat (VNTR) sequence in the MUC1 gene encoding mucin 1, have allowed the nephrologists to diagnose renal diseases of previously unknown cause, with a real prevalence probably underestimated. We present the cohort of patients identified with ADTKD in the Nephrogenetics’ clinic from the Nephrology department of our hospital from 2015 to 2022. Method Patients with chronic kidney disease (CKD) of unknown etiology and renal phenotype suggesting CIN (bland urine analysis, hyperuricemia and/or anemia unrelated to the degree of CKD, hypo/hypercalcemia, acidemia, urinary concentration defects, absent hypertension, renal cysts), especially with a family history of similar CKD, were referred for the Nephrogenetics’ clinic and ADTKD was investigated. In these cases, we performed a genetic study of ADTKD by the following steps: 1) UMOD, REN, HNF1B mutations studied through NGS; 2) if negative, search for the insertion of a single cytosine of the repeat unit comprising the extremely long (∼1.5-5 kb), GC-rich (>80%) coding variable-number tandem repeat (VNTR) sequence in the MUC1 gene encoding mucin 1 (see Kirby et al Nat Genet. 2013). It is used a restriction and Snapshot method. 3) if negative, search for HNF1B Large deletion, detected by MLPA (kit P241-D2, MRC-Holland). Pre-genetic counselling was performed by the nephrologist, and post-genetic counselling was performed by the nephrologist and geneticist, namely to proceed with family screening of potentially affected family members when a pathogenic mutation was identified or when segregation studies were necessary. Results 31 families were studied and a confirmed genetic diagnosis was obtained in 11 (35%), allowing the identification of 30 patients with ADTKD; the mutations identified were in MUC1 in 4 families (13 patients), UMOD in 4 families (11 patients), HNF1B (2 families, 5 patients) and REN: 1 patient with a phenotype compatible with ADTKD but with a genotype compatible with renal tubular dysgenesis (two mutations in REN gene from each of the parents). One patient with HNF1B deletion also presented with a heterozygous variant of unknown significance in PKD1 gene, and a pathogenic variant of SLC12A3, heterozygous. Three families are still under study: 2 families are being studied with segregation studies to evaluate the pathogenicity of mutations in UMOD (1 family) and HNF1B (1 family), and 1 family with a phenotype of ADTKD (3 young patients with positive family history) with negative results for mutations in ADTKD genes is in study, with a negative result in WES. The study will be pursued in a specialized laboratory to better study MUC1, but at this moment we classify this family as ADTKD-NOS. If positive results are obtained for these 3 families, the percentage of families identified using this protocol will increase to 43,3% of the families studied through this model. Conclusion In our cohort, a genetic diagnosis was established in 35% of the families studied. MUC1 and UMOD are the most prevalent genes implicated in ADTKD at our center, similarly to what has been reported in other cohorts of ADTKD. Given the rarity of these diseases and the fact that they require specific molecular techniques for accurate diagnosis, we believe that Nephrogenetics’ clinics with dedicated nephrologists and geneticists in close cooperation are critical for an accurate diagnosis.

From bottle to bedside: Implementation considerations and antimicrobial stewardship considerations for bloodstream infection rapid diagnostic testing.

Antimicrobial stewardship (AMS) programs have been quick to adopt novel molecular rapid diagnostic technologies (mRDTs) for bloodstream infections (BSIs) to improve antimicrobial management. As such, most of the literature demonstrating the clinical and economic benefits of mRDTs for BSI is in the presence of active AMS intervention. Leveraging mRDTs to improve antimicrobial therapy for BSI is increasingly integral to AMS program activities. This narrative review discusses available and future mRDTs, the relationship between the clinical microbiology laboratory and AMS programs, and practical considerations for optimizing the use of these tools within a health system. Antimicrobial stewardship programs must work closely with their clinical microbiology laboratories to ensure that mRDTs are used to their fullest benefit while remaining cognizant of their limitations. As more mRDT instruments and panels become available and AMS programs continue to expand, future efforts must consider the expansion beyond traditional settings of large academic medical centers and how combinations of tools can further improve patient care.

Invasive Mold Infections in Children: Navigating Troubled Waters with a Broken Compass.

Incidence of invasive mold infections in children, while rare, is increasing as the population of high-risk patients expands, including premature infants, pediatric patients undergoing treatment for hematological malignancies, or recipients of allogeneic hematologic stem cell transplants. The infectious agents, including Aspergillus spp., Mucorales, and other molds, are especially difficult to treat and have serious morbidity and high mortality. Clinicians must maintain a high index of suspicion for invasive mold infections in at-risk patients. Diagnosis of invasive mold infections is complicated by difficulties isolating pathogens on culture, but progress is being made in immunological and molecular diagnostic technologies. Treatment in children is challenging; no randomized controlled trials exist. There is a growing body of data on treatment, specifically on safer antifungal agents, including indications for treatment, spectrum of coverage, pharmacokinetics for different ages, and pharmacodynamic targets associated with therapeutic success. However, pediatricians must often extrapolate from adult data. In this review, we aim to harmonize the existing body of literature on invasive mold infections in children, covering epidemiology, clinical presentations, diagnostic methods, and principles of management.

Advances in the application of molecular diagnostic techniques for the detection of infectious disease pathogens (Review).

Infectious diseases are a major global cause of morbidity and mortality, seriously affecting public health and socioeconomic stability. Since infectious diseases can be caused by a wide variety of pathogens with similar clinical manifestations and symptoms that are difficult to accurately distinguish, selecting the appropriate diagnostic techniques for the rapid identification of pathogens is crucial for clinical disease diagnosis and public health management. However, traditional diagnostic techniques have low detection rates, long detection times and limited automation, which means that they do not meet the requirements for rapid diagnosis. Recent years have seen continuous developments in molecular detection technology, which has a higher sensitivity and specificity, shorter detection time and increased automation, and performs an important role in the early and rapid detection of infectious disease pathogens. The present study summarizes recent progress in molecular diagnostic technologies such as PCR, isothermal amplification, gene chips and high‑throughput sequencing for the detection of infectious disease pathogens, and compares the technical principles, advantages and disadvantages, applicability and costs of these diagnostic techniques.