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The burden of colorectal cancer (CRC) is high in the Asia‐Pacific region, and several countries in this region have among the highest and/or fastest growing rates of CRC in the world. A significant proportion of patients will present with or develop metastatic CRC (mCRC), and BRAFV600E‐mutant mCRC represents a particularly aggressive phenotype that is less responsive to standard chemotherapies. In light of recent therapeutic advances, an Asia‐Pacific expert consensus panel was convened to develop evidence‐based recommendations for the diagnosis, treatment, and management of patients with BRAFV600E‐mutant mCRC. The expert panel comprised nine medical oncologists from Australia, Hong Kong, Singapore, and Taiwan (the authors), who met to review current literature and develop eight consensus statements that describe the optimal management of BRAFV600E‐mutant mCRC in the Asia‐Pacific region. As agreed by the expert panel, the consensus statements recommend molecular testing at diagnosis to guide individualized treatment decisions, propose optimal treatment pathways according to microsatellite stability status, advocate for more frequent monitoring of BRAFV600E‐mutant mCRC, and discuss local treatment strategies for oligometastatic disease. Together, these expert consensus statements are intended to optimize treatment and improve outcomes for patients with BRAFV600E‐mutant mCRC in the Asia‐Pacific region. [ABSTRACT FROM AUTHOR]

Accurate detection of Helicobacter pylori and its antimicrobial resistance is essential for eradication of the infections. The aim of this study was to compare five different CE‐IVD marked assays in detection of H. pylori from 268 clinical stool samples. Samples were considered positive for H. pylori when at least three of the five tests were positive. Amplified IDEIA Hp StAR (Oxoid) and Premier Platinum HpSA PLUS (Meridian Bioscience Inc.) assays showed sensitivity of 100% [95% CI (confidence interval): 87–100] and LIAISON® Meridian H. pylori SA (DiaSorin) of 83.3% (95% CI: 66–93). Specificities of the assays were 94.5% (95% CI: 91–97), 95.4%; (95% CI: 92–97), and 97.1% (95% CI: 94–99) respectively. Amplidiag® H. pylori + ClariR (Mobidiag) assay showed 93.3% (95% CI: 78–99) and Allplex™ H. pylori & ClariR Assay (Seegene Inc.) 36.7% (95% CI: 22–55) sensitivity, while specificity of both was 97.9% (95% CI: 95–99). The Amplidiag® and Allplex™ assays concordantly detected clarithromycin resistance in positive for H. pylori samples. The Amplidiag® assay showed the highest accuracy, namely 97.4% (95% CI: 95–99). These data provide helpful information for planning laboratory diagnostics of H. pylori and detection of clarithromycin resistance from stool samples. [ABSTRACT FROM AUTHOR]

Background: Screening for anaplastic lymphoma kinase (ALK) rearranged non‐small cell lung cancer (NSCLC) is crucial for identifying patients eligible for targeted therapy. The FDA‐approved ALK (D5F3) immunohistochemistry (IHC) assay, used with the OptiView Amplification Kit, demonstrates excellent sensitivity and specificity in detecting these patients. However, the clinical significance of resulting focal positivity remains unclear, and ALK (D5F3) expression unrelated to ALK fusion is observed in some cases of neuroendocrine differentiation. This study aims to validate these findings with molecular testing and contribute to the accurate interpretation of ALK (D5F3) IHC results. Methods: A total of 1619 patients diagnosed with NSCLC and neuroendocrine carcinoma were evaluated using ALK (D5F3) IHC. For cases with strong but focal expression and those with diffuse strong positivity in neuroendocrine differentiation, ALK fluorescence in situ hybridization (FISH) and/or next‐generation sequencing (NGS) tests were performed. Results: Seven out of 1109 adenocarcinomas (0.6%) and six out of 289 squamous cell carcinomas (2.1%) exhibited strong focal ALK (D5F3) expression. Nine out of 209 neuroendocrine carcinomas (4.3%) showed homogeneously strong ALK (D5F3) expression. All these cases, including adenocarcinoma with neuroendocrine differentiation and combined small cell carcinoma, were negative for ALK fusions by FISH and/or NGS. Conclusion: This study demonstrates that strong but focal ALK (D5F3) immunostaining and strong expression in neuroendocrine differentiation may not indicate ALK fusion. By considering these findings, we can improve the accuracy of patient selection for targeted therapy by minimizing false‐positive interpretations of ALK (D5F3) staining. [ABSTRACT FROM AUTHOR]

Background: Facioscapulohumeral dystrophy (FSHD) is a myopathy characterized by the loss of repressive epigenetic features affecting the D4Z4 locus (4q35). The assessment of DNA methylation at two regions (DUX4-PAS and DR1) of D4Z4 locus proved to be an effective method to detect epigenetic signatures compatible with FSHD. The present study aims at validating the employment of this method into clinical practice and improving the protocol by refining the classification thresholds of 4qA/4qA patients. To this purpose, 218 subjects with clinical suspicion of FSHD collected in 2022–2023 were analyzed. Each participant underwent in parallel the traditional FSHD molecular testing (D4Z4 sizing) and the proposed methylation assay. The results provided by both analyses were compared to evaluate the concordance and calculate the performance metrics of the methylation test. Results: Among the 218 subjects, the 4q variant type distribution was 54% 4qA/4qA, 43% 4qA/4qB and 3% 4qB/4qB. The methylation analysis was performed only on carriers of at least one 4qA allele. After refining the classification threshold, the test reached the following performance metrics: sensitivity = 0.90, specificity = 1.00 and accuracy = 0.93. These results confirmed the effectiveness of the methylation assay in identifying patients with genetic signature compatible with FSHD1 and FSHD2 based on their DUX4-PAS and DR1 profile, respectively. The methylation data were also evaluated with respect to the clinical information. Conclusions: The study confirmed the ability of the method to accurately identify methylation profiles compatible with FSHD genetic signatures considering the 4q genotype. Moreover, the test allows the detection of hypomethylated profiles in asymptomatic patients, suggesting its potential application in identifying preclinical conditions in patients with positive family history and FSHD genetic signatures. Furthermore, the present work emphasizes the importance of interpreting methylation profiles considering the patients' clinical data. [ABSTRACT FROM AUTHOR]

Background: The objective of the following retrospective review was to perform a cost-effectiveness analysis of the use of molecular testing of indeterminate thyroid nodules compared to current management practices in Nova Scotia, Canada. Methods: All cases of cytologically indeterminate thyroid nodules from January 1st, 2014 to December 31st, 2018 were reviewed. All interventions related to an indeterminate thyroid nodule were recorded. Patients were excluded if less than 18 years old if no further information regarding medical management was electronically available beyond the diagnosis of an indeterminate thyroid nodule, history of radiation, or previous thyroid surgery prior to diagnosis of an indeterminate thyroid nodule in the remaining lobe. Microcosting was performed to determine the cost of all relevant interventions including repeat fine needle aspiration biopsy, ultrasound, thyroid surgery(s), and molecular testing. Institution-specific transition state probabilities were calculated and used to build a cost-effectiveness analysis model. Model output was an incremental cost-effectiveness ratio, defined as the ratio of cost difference to effectiveness difference between routine molecular testing and the current management strategy, yielding cost per surgery avoided. Results: The mean effectiveness of the current management of indeterminate thyroid nodules in Nova Scotia based on the American Thyroid Association guidelines is 64% at a mean cost of $6431, while the simulated mean effectiveness of routine molecular testing is 89% at a mean cost of $8414. Differences in management strategies generated an incremental cost-effectiveness ratio of $7876 per surgery avoided. Conclusion: Routine molecular testing is the more effective strategy for the appropriate management of indeterminate thyroid nodules; however, it comes at a higher mean cost compared to the current management strategy. As the cost of molecular testing continues to decrease, and the cost of OR resources continues to rise, molecular testing is likely to become the optimal strategy in Nova Scotia. [ABSTRACT FROM AUTHOR]

Introduction: Congenital hypopituitarism (CH) is characterized by the deficiency of pituitary hormones. Among CH patients, 85% lack a molecular diagnosis. Whole Exome Sequencing (WES) identified a homozygous variant (c.865G>A, p.Val289Ile) in the CDH2 gene, responsible for N-Cadherin production, crucial for cell-cell adhesion. Predicted to be likely pathogenic, the variant was found in a patient deficient in GH, TSH, ACTH, and LH/FSH. Its impact on cell adhesion was confirmed in L1 fibroblast cell lines. Objective: Create a cdh2 knockdown in zebrafish for investigating its role in pituitary development through growth hormone and transcription factors expression. Methods: Utilized pET28B-RfxCas13d-His plasmid for Cas13 mRNA production via in vitro transcription, guiding Cas13 to cdh2 with three RNAs. Injected the complex into single-cell embryos for analysis up to 96 hpf. Assessed gene expression of cdh2, prop1, pit1, and gh1 using RT-qPCR. Evaluated cdh2 protein expression through the western blot technique. Results: Knockdown animals displayed developmental delay. The cdh2 expression decreased by 75% within 24 hours, rebounded by 48 hours, and reached wild-type levels by 96 hpf. gh1 expression decreased at 48h but increased by 96 hpf, aligning with WT. No significant differences in prop1 and pit1 expression were observed. Conclusion: Our findings underscore cdh2's role in pituitary development and hormonal regulation, offering insights for developmental biology research. [ABSTRACT FROM AUTHOR]

Gastrointestinal cancer is a common malignant tumor with a high incidence worldwide. Despite continuous improvements in diagnosis and treatment strategies, the overall prognosis of gastrointestinal tumors remains poor. Carcinoembryonic antigen (CEA) is highly expressed in various types of cancers, especially in gastrointestinal cancers, making it a potential target for therapeutic intervention. Therefore, the expression of CEA can be used as an indication of the existence of tumors, chosen as a target for molecular imaging diagnosis, and effectively utilized in the targeted therapy of gastrointestinal cancers. In this study, we report the selection and characterization of affibody molecules (ZCEA539, ZCEA546, and ZCEA919) specific to the CEA protein. Their ability to bind to recombinant and native CEA protein has been confirmed by surface plasmon resonance (SPR), immunofluorescence, and immunohistochemistry assays. Furthermore, Dylight755-labeled ZCEA affibody showed accumulation within the tumor site 1 h post injection and was continuously enhanced for 4 h. The Dylight755-labeled ZCEA affibody exhibited high tumortargeting specificity in CEA+ xenograft-bearing mice and possesses promising characteristics for tumor-targeting imaging. Overall, our results suggest the potential use of ZCEA affibodies as fluorescent molecular imaging probes for detecting CEA expression in gastrointestinal cancer. [ABSTRACT FROM AUTHOR]

A biomarker is defined as a characteristic that is measured as an indicator of a normal biological or pathological process, a response to an exposure or intervention. Biomarkers with a diagnostic approach must identify not only the presence but also the absence of the disease with high precision, so having the biological source of the said marker is of vital importance to ensure precision and accuracy; the aim was to carry out a review of its diagnostic potential. The search strategy was carried out in three databases: PubMed, ScienceDirect, and Scopus. The keywords that were used were as follows: "gingival crevicular fluid", "Biomarker", and "Diagnosis", using the Boolean operator "AND". The filter was used at 10 years. Within the type of molecules most studied, the cytokine family was the most abundant with 25.42% of the studies, followed by metalloproteinases and proteins with 16.9% each one. Studies that included RNA‐type genetic material were less frequently found. As has been demonstrated, the use of GCF as a source of biomolecules for diagnostic use has been increasing, both for oral diseases, which reflects the local conditions of the disease; it also has the ability to reflect the development of distant diseases; and this is because GCF is a blood ultrafiltrate. [ABSTRACT FROM AUTHOR]

Background: Cutaneous leishmaniasis (CL) in Ethiopia and some parts of Kenya is predominantly caused by Leishmania aethiopica. While skin-slit (SS) microscopy is routinely used for CL diagnosis, more sensitive molecular tests are available. The Loopamp™ Leishmania detection kit (Loopamp) is a robust loop-mediated isothermal amplification (LAMP) assay with the potential for implementation in primary healthcare facilities. In this study, we comparatively assessed the diagnostic accuracy of four methods currently used to diagnose CL: Loopamp, kinetoplast DNA (kDNA) PCR, spliced leader RNA (SL-RNA) PCR and SS microscopy. Methods: A study on 122 stored tape disc samples of suspected CL patients was conducted in Gondar, northwestern Ethiopia. Routine SS microscopy results were obtained from all patients. Total nucleic acids were extracted from the tapes and subjected to PCR testing targeting kDNA and SL-RNA, and Loopamp. Diagnostic accuracy was calculated with SS microscopy as a reference test. The limit of detection (LoD) of Loopamp and kDNA PCR were determined for cultured L. aethiopica and Leishmania donovani. Results: Of the 122 patients, 64 (52.5%) were identified as CL cases based on SS microscopy. Although the PCR tests showed a sensitivity of 95.3% (95% confidence interval [CI] 91.6–99.1), Loopamp only had 48.4% (95% CI 39.6–57.3) sensitivity and 87.9% (95% CI 82.1–93.7) specificity. The LoD of Loopamp for L. donovani was 100-fold lower (20 fg/µl) than that for L. aethiopica (2 pg/µl). Conclusions: The Loopamp™ Leishmania detection kit is not suitable for the diagnosis of CL in Ethiopia, presumably due to a primer mismatch with the L. aethiopica 18S rRNA target. Further research is needed to develop a simple and sensitive point-of-care test that allows the decentralization of CL diagnosis in Ethiopia. [ABSTRACT FROM AUTHOR]

Background: There are no studies assessing the evolution and patterns of genetic studies performed at diagnosis in acute myeloid leukemia (AML) patients. Such studies could help to identify potential gaps in our present diagnostic practices, especially in the context of increasingly complex procedures and classifications. Methods: The REALMOL study (NCT05541224) evaluated the evolution, patterns, and clinical impact of performing main genetic and molecular studies performed at diagnosis in 7285 adult AML patients included in the PETHEMA AML registry (NCT02607059) between 2000 and 2021. Results: Screening rates increased for all tests across different time periods (2000–2007, 2008–2016, and 2017–2021) and was the most influential factor for NPM1, FLT3‐ITD, and next‐generation sequencing (NGS) determinations: NPM1 testing increased from 28.9% to 72.8% and 95.2% (p <.001), whereas FLT3‐ITD testing increased from 38.1% to 74.1% and 95.9% (p <.0001). NGS testing was not performed between 2000–2007 and only reached 3.5% in 2008–2016, but significantly increased to 72% in 2017–2021 (p <.001). Treatment decision was the most influential factor to perform karyotype (odds ratio [OR], 6.057; 95% confidence interval [CI], 4.702–7.802), and fluorescence in situ hybridation (OR, 2.273; 95% CI, 1.901–2.719) studies. Patients ≥70 years old or with an Eastern Cooperative Oncology Group ≥2 were less likely to undergo these diagnostic procedures. Performing genetic studies were associated with a favorable impact on overall survival, especially in patients who received intensive chemotherapy. Conclusions: This unique study provides relevant information about the evolving landscape of genetic and molecular diagnosis for adult AML patients in real‐world setting, highlighting the increased complexity of genetic diagnosis over the past 2 decades. A retrospective analysis of 7285 cases over 2 decades. This study explores the evolution of diagnostic practices in acute myeloid leukemia patients, highlighting factors influencing test implementation and their association with patient outcomes. [ABSTRACT FROM AUTHOR]

Cas12 and Cas13 are extensively utilized in molecular diagnostics for their trans-cleavage activities, yet their activation characteristics remain partially understood. Here, we conduct an in-depth investigation of Cas12a, Cas12f1, and Cas13a, uncovering the characteristics of their trans-DNase and trans-RNase activities with noncanonical activators. Our findings reveal that DNA can serve as a direct target for CRISPR-Cas13a, markedly increasing the detection sensitivity for single-base mismatches. Moreover, the trans-cleavage activities of Cas12a and Cas13a can be activated by diverse RNA:DNA and RNA:RNA duplexes, respectively, indicating that the presence of stem–loop structures in crRNAs is not essential for their activation. Notably, Cas12f1, unlike Cas12a, exhibits intrinsic RNase activity independently of activation. Leveraging these insights, we have improved the accuracy of a dual-gene target detection approach that employs the CRISPR-Cas12f1 and Cas13a systems. Our research advances the understanding of the noncanonical activation characteristics of Cas12 and Cas13a, contributing to the field of CRISPR-based diagnostics. Uncovering noncanonical trans-cleavage characteristics of Cas12 and Cas13a improves the accuracy of CRISPR-based dual-gene diagnostics and enhances detection sensitivity for single base mismatches through newly discovered activation mechanisms [ABSTRACT FROM AUTHOR]

Background: Strongyloidiasis a neglected tropical disease is known to cause severe disease among immunosuppressed and has not been studied extensively in Sri Lanka. Parasitological diagnostic approaches based on faecal microscopy and culture often fail to detect low-intensity infections. This study investigates the presence of strongyloidiasis among selected immunocompromised individuals using parasitological, molecular and serological techniques. Methods: Adult patients with immunocompromising conditions admitted to three tertiary care hospitals in Sri Lanka were recruited. A faecal sample and 2 ml of venous blood were collected. The faecal samples were subjected to direct faecal smear and cultures (agar plate, charcoal and Harada-Mori) and polymerase chain reaction (PCR) using species specific primers designed for Strongyloides stercoralis. The presence of Strongyloides IgG antibodies was tested in the collected serum samples using DRG Strongyloides IgG enzyme-linked immunosorbent assay (ELISA) kits. The PCR products of the positive samples were sequenced using Sanger sequencing method. Results: A total of 260 patients were recruited to this study, out of which 160 provided faecal samples and 122 provided blood samples. Out of the 160 faecal samples, none were positive for strongyloidiasis by direct smear, charcoal and Harada-Mori cultures. Only one sample (0.6%) was positive by agar plate culture. Out of the 123 samples subjected to PCR, 14 (11.4%), including the culture positive patient, were positive for S. stercoralis. Sequencing results of the PCR products indicated 100% similarity to S. stercoralis. Out of the 122 serum samples subjected to ELISA, 20 (16.4%), including the culture positive patient, were positive for Strongyloides IgG antibodies. However, sociodemographic, exposure factors, clinical features were not significantly associated with the presence of strongyloidiasis infection. Conclusions: Strongyloidiasis is present among the immunocompromised population in Sri Lanka, even in the absence of a significant relationship with associated factors. It is advisable to screen such patients with highly sensitive tests such as PCR for early diagnosis and treatment. [ABSTRACT FROM AUTHOR]

Multiple endocrine neoplasia (MEN) is a group of rare genetic diseases characterized by the occurrence of multiple tumors of the endocrine system in the same patient. The first MEN described was MEN1, followed by MEN2A, and MEN2B. The identification of the genes responsible for these syndromes led to the introduction of family genetic screening programs. More than twenty years later, not all cases of MENs have been resolved from a genetic point of view, and new clinicogenetic entities have been described. In this review, we will discuss the strategies and difficulties of genetic screening for classic and newly described MENs in a clinical setting, from limitations in sequencing, to problems in classifying variants, to the identification of new candidate genes. In the era of genomic medicine, characterization of new candidate genes and their specific tumor risk is essential for inclusion of patients in personalized medicine programs as well as to permit accurate genetic counseling to be proposed for families. [ABSTRACT FROM AUTHOR]

Immuno-positron emission tomography (immunoPET) enables imaging of specific targets that play a role in targeted therapy and immunotherapy, such as antigens on cell membranes, targets in the disease microenvironment, or immune cells. The most common immunoPET applications use a monoclonal antibody labeled with a relatively long-lived positron emitter such as 89Zr (T1/2 = 78.4 h), but smaller antibody-based constructs labeled with various other positron emitting radionuclides are also being investigated. This molecular imaging technique can thus guide the development of new drugs and may have a pivotal role in selecting patients for a particular therapy. In early phase immunoPET trials, multiple imaging time points are used to examine the time-dependent biodistribution and to determine the optimal imaging time point, which may be several days after tracer injection due to the slow kinetics of larger molecules. Once this has been established, usually only one static scan is performed and semi-quantitative values are reported. However, total PET uptake of a tracer is the sum of specific and nonspecific uptake. In addition, uptake may be affected by other factors such as perfusion, pre-/co-administration of the unlabeled molecule, and the treatment schedule. This article reviews imaging methodologies used in immunoPET studies and is divided into two parts. The first part summarizes the vast majority of clinical immunoPET studies applying semi-quantitative methodologies. The second part focuses on a handful of studies applying pharmacokinetic models and includes preclinical and simulation studies. Finally, the potential and challenges of immunoPET quantification methodologies are discussed within the context of the recent technological advancements provided by long axial field of view PET/CT scanners. [ABSTRACT FROM AUTHOR]

Background: Short stature is a complex disorder with phenotypic and genetic heterogeneity. This study aimed to investigate clinical phenotypes and molecular basis of a cohort of patients with short stature. Methods: Trio whole-exome sequencing (Trio-WES) was performed to explore the genetic aetiology and obtain a molecular diagnosis in twenty Chinese probands with syndromic and isolated short stature. Results: Of the twenty probands, six (6/20, 30%) patients with syndromic short stature obtained a molecular diagnosis. One novel COMP pathogenic variant c.1359delC, p.N453fs*62 and one LZTR1 likely pathogenic variant c.509G>A, p.R170Q were identified in a patient with short stature and skeletal dysplasia. One novel de novo NAA15 pathogenic variant c.63T>G, p.Y21X and one novel de novo KMT2A pathogenic variant c.3516T>A, p.N1172K was identified in two probands with short stature, intellectual disability and abnormal behaviours, respectively. One patient with short stature, cataract, and muscle weakness had a de novo POLG pathogenic variant c.2863 T>C, p.Y955H. One PHEX pathogenic variant c.1104G>A, p.W368X was identified in a patient with short stature and rickets. Maternal uniparental disomy 7 (mUPD7) was pathogenic in a patient with pre and postnatal growth retardation, wide forehead, triangular face, micrognathia and clinodactyly. Thirteen patients with isolated short stature had negative results. Conclusion: Trio-WES is an important strategy for identifying genetic variants and UPD in patients with syndromic short stature, in which dual genetic variants are existent in some individuals. It is important to differentiate between syndromic and isolated short stature. Genetic testing has a high yield for syndromic patients but low for isolated patients. [ABSTRACT FROM AUTHOR]

Background: Prenatal ultrasound findings of fetal bilateral echogenic kidneys accompanied by oligohydramnios can be highly stressful for both pregnant women and physicians. The diversity of underlying causes makes it challenging to confirm a prenatal diagnosis, predict postnatal outcomes, and counsel regarding recurrence risks in future pregnancies. Case Presentation: We report two cases of abnormal fetal echogenic kidneys with oligohydramnios detected in the early third trimester. Autosomal recessive polycystic kidney disease (ARPKD), a rare genetic syndrome, was initially suspected in both cases. However, postnatal diagnoses differed: the first case was confirmed as glomerulocystic kidney disease (GCKD) through renal pathology, while the second case was diagnosed as ARPKD with a compound heterozygous likely pathogenic PKHD1 mutation. Conclusion: Prenatal diagnosis of fetal echogenic kidneys with oligohydramnios should prioritize accurate diagnosis. Given the differences in the clinical spectrum, GCKD should be considered a differential diagnosis for this condition, particularly ARPKD. This study highlights the importance and benefits of molecular diagnosis and postnatal renal pathology for precise diagnosis and effective counseling. [ABSTRACT FROM AUTHOR]

ABSTRACT Melanocytic tumors are a diverse group of lesions and are traditionally classified based on a combination of clinical presentation as well as histological examination. More recently, molecular diagnostics has become an increasingly important part of differentiating different melanocytic lesions in the current WHO standards. This molecular testing, however, can result in unexpected findings. In this report, we describe that molecular testing of a clinical atypical melanocytic lesion showed a mutation in the MAP2K1 gene as well as an unexpected germline mutation in PTPN11, indicative of Noonan syndrome. Based on these findings we concluded that the patient had a MAP2K1 associated melanocytic lesion with Noonan syndrome as an incidental finding. Melanomas are classically not associated with Noonan syndrome. However, we hypothesized that the germline mutations of PTPN11 and the somatic second hit mutation in the MAP2K1 genes might be involved in the formation of the aforementioned lesion. As they are both part of the RAS‐MAPK pathway. Furthermore, with the expansion of molecular diagnostics in melanomas, we expect to find an increase in unexpected (germline) mutations. [ABSTRACT FROM AUTHOR]

Introduction: Genomic variant testing of tumors is a critical gateway for patients to access the full potential of personalized oncology therapeutics. Current methods such as next-generation sequencing are costly and challenging to interpret, while PCR assays are limited in the number of variants they can cover. We developed ASPYRE® (Allele-Specific PYrophosphorolysis RE action) technology to address the urgent need for rapid, accessible and affordable diagnostics informing actionable genomic target variants of a given cancer. The targeted ASPYRE-Lung panel for non-small cell carcinoma covers 114 variants in 11 genes (ALK, BRAF, EGFR, ERBB2, KRAS, RET, ROS1, MET & NTRK1/2/3) to robustly inform clinical management. The assay detects single nucleotide variants, insertions, deletions, and gene fusions from tissue-derived DNA and RNA simultaneously. Methods: We tested the limit of detection, specificity, analytical accuracy and analytical precision of ASPYRE-Lung using FFPE lung tissue samples from patients with non-small cell lung carcinoma, variant-negative FFPE tissue from healthy donors, and FFPE-based contrived samples with controllable variant allele fractions. Results: The sensitivity of ASPYRE-Lung was determined to be ≤ 3% variant allele fraction for single nucleotide variants and insertions or deletions, 100 copies for fusions, and 200 copies for MET exon 14 skipping. The specificity was 100% with no false positive results. The analytical accuracy test yielded no discordant calls between ASPYRE-Lung and expected results for clinical samples (via orthogonal testing) or contrived samples, and results were replicable across operators, reagent lots, runs, and real-time PCR instruments with a high degree of precision. Conclusions: The technology is simple and fast, requiring only four reagent transfer steps using standard laboratory equipment (PCR and qPCR instruments) with analysis via a cloud-based algorithm. The ASPYRE-Lung assay has the potential to be transformative in facilitating access to rapid, actionable molecular profiling of tissue for patients with non-small cell carcinoma. [ABSTRACT FROM AUTHOR]

Sensing and imaging of biomolecules are crucial to disease diagnosis, prognosis, and therapy where optical techniques have essential utility. Untethered and remotely controlled micro/nanorobots have shown promising sensing and imaging capabilities, especially in complex biological environments. In this review, how micro/nanorobots are used for optical biosensing and imaging while highlighting the significant developments in the field is discussed. Starting is done by exploring colorimetric biosensing methods enabled by micro/nanorobots. Significant advancements in surface‐enhanced Raman spectroscopy‐integrated micro/nanorobots are reviewed. Further, state‐of‐the‐art optical bio‐imaging applications by micro/nanorobots at in vitro intracellular level are highlighted. Novel in vivo bio‐imaging assisted by optical micro/nanorobot sensors is examined. Furthermore, innovations in micro/nanorobots are assessed where motion augmentation is used as a detection mechanism, with applications in point‐of‐care molecular diagnostics. Finally, the challenges associated with micro/nanorobots‐assisted advanced optical biosensing and imaging while discussing insights about potential research directions for this rapidly progressing field are summarized. [ABSTRACT FROM AUTHOR]

Given the high degree of variability and complexity of cancer, precise monitoring and logical analysis of different nucleic acid markers are crucial for improving diagnostic precision and patient survival rates. However, existing molecular diagnostic methods normally suffer from high cost, cumbersome procedures, dependence on specialized equipment and the requirement of in‐depth expertise in data analysis, failing to analyze multiple cancer‐associated nucleic acid markers and provide immediate results in a point‐of‐care manner. Herein, we demonstrate a transistor‐based DNA molecular computing (TDMC) platform that enables simultaneous detection and logical analysis of multiple microRNA (miRNA) markers on a single transistor. TDMC can perform not only basic logical operations such as "AND" and "OR", but also complex cascading computing, opening up new dimensions for multi‐index logical analysis. Owing to the high efficiency, sensing and computations of multi‐analytes can be operated on a transistor at a concentration as low as 2×10−16 M, reaching the lowest concentration for DNA molecular computing. Thus, TDMC achieves an accuracy of 98.4 % in the diagnosis of hepatocellular carcinoma from 62 serum samples. As a convenient and accurate platform, TDMC holds promise for applications in "one‐stop" personalized medicine. [ABSTRACT FROM AUTHOR]

Mesophilic Argonautes (Agos) from microbial resources have received significant attention due to their potential applications in genome editing and molecular diagnostics. This study characterizes a novel Ago from Pseudobutyrivibrio ruminis (PrAgo), which can cleave single-stranded DNA using guide DNA (gDNA). PrAgo, functioning as a multi-turnover enzyme, effectively cleaves DNA using 5′-phosphate gDNA, 14–30 nucleotides in length, in the presence of both Mn2+ and Mg2+ ions. PrAgo demonstrates DNA cleavage activity over a broad pH range (pH 4–12), with optimal activity at pH 11. As a mesophilic enzyme, PrAgo cleaves efficiently DNA at temperatures ranging from 25 to 65 °C, particularly at 65 °C. PrAgo does not show strong preferences for the 5′-nucleotide in gDNA. It shows high tolerance for single-base mismatches, except at positions 13 and 15 of gDNA. Continuous double-nucleotide mismatches at positions 10–16 of gDNA significantly reduce cleavage activity. Furthermore, PrAgo mediates DNA-guided DNA cleavage of AT-rich double stranded DNA at 65 °C. Additionally, molecular dynamic simulations suggest that interactions between the PAZ domain and different nucleic acids strongly influence cleavage efficiency. These findings expand our understanding of Protokaryotic Agos and their potential applications in biotechnology. [ABSTRACT FROM AUTHOR]

Summary: Diagnosis of essential thrombocythaemia (ET) is challenging in patients lacking JAK2/CALR/MPL mutations. In a retrospective evaluation of 320 patients with 'triple‐negative thrombocytosis', we assessed utility of bone marrow histology (90.9% of patients) and myeloid gene panel (MGP, 55.6%). Supportive histology ('myeloproliferative neoplasm‐definite/probable', 36.8%) was associated with higher platelet counts and varied between centres. 14.6% MGP revealed significant variants: 3.4% JAK2/CALR/MPL and 11.2% other myeloid genes. Final clinical diagnosis was strongly predicted by histology, not MGP. 23.7% received cytoreduction (17.6% under 60 years). Real‐world 'triple‐negative' ET diagnosis currently depends heavily on histology; we advocate caution in MGP‐negative cases and that specific guidelines are needed. [ABSTRACT FROM AUTHOR]

This study aimed to carry out a molecular screening for the presence of Giardia, Cryptosporidium, and/or Entamoeba in the feces of pet and stray/feral cats in Jordan. G. duodenalis was found in 27.9% (95% CI, 23.2–32.9) of the 348 sampled cats overall; E. histolytica was found in only 0.6% (95% CI, 0.1–2.1) of the cats, while none of the sampled cats had Cryptosporidium infections. The infection rate of G. duodenalis among indoor cats (32.3%) did not differ significantly from that among outdoor cats (24.1%). There were significantly more infections (p = 0.0004) geographically in the cold semiarid areas (67%) than in the cold desert areas (24%). Multilocus sequence typing analysis of amplicons based on the bg, tpi, and gdh genes revealed that the majority of G. duodenalis infections were zoonotic assemblage B (65.9%; 64 of 97 positive samples); followed by feline-specific assemblage F (18.5%, 18/97); cattle-specific assemblage E (5.2%, 5/97); and then assemblage C that was shared with canids (1.0%; 1/97). Within Giardia isolates, a substitution mutation (A/G) was found at position 297 of the complete protein coding sequence (cds) of tpi-assemblage B, which may represent a new spreading mutation within this gene among the cat population in Jordan. The results of the present study suggest that close human-cat interactions could play a role in zoonotic transmission of Giardia, but further research is needed to determine the possible contribution of cats to the transmission of other protozoa to humans. [ABSTRACT FROM AUTHOR]

Pancreatic cancer is one of the most aggressive and lethal forms of cancer, with a five-year survival rate of less than 10%. Despite advances in treatment modalities, the prognosis for pancreatic cancer patients remains poor, highlighting the urgent need for innovative approaches for early diagnosis and targeted therapies. In recent years, there has been significant progress in understanding the molecular mechanisms underlying pancreatic cancer development and progression. This paper reviews the current knowledge of molecular targets for the diagnosis and treatment of pancreatic cancer. [ABSTRACT FROM AUTHOR]

Simple Summary: With advances in radiological imaging and its increased use, small nodules are being detected more frequently. Surgical resection is often the final option for diagnosis and treatment; however, small nodules may be too small to be detected during surgery. The lungs are soft, deformable organs that can change shape during respiratory phases and surgical procedures. Thus, surgeons rely on preoperative or intraoperative markings for nodule identification and precise resection. Furthermore, two randomized clinical trials found that sublobar resection could be an alternative treatment for early-stage non-small-cell lung cancer. Therefore, the demand for preoperative or intraoperative marking techniques is increasing, particularly for wedge resection or segmentectomy. In this study, we provide a narrative review of the current status and future perspectives of preoperative and intraoperative markings in thoracic surgery. The widespread implementation of lung cancer screening and thin-slice computed tomography (CT) has led to the more frequent detection of small nodules, which are commonly referred to thoracic surgeons. Surgical resection is the final diagnostic and treatment option for such nodules; however, surgeons must perform preoperative or intraoperative markings for the identification of such nodules and their precise resection. Historically, hook-wire marking has been performed more frequently worldwide; however, lethal complications, such as air embolism, have been reported. Therefore, several surgeons have recently attempted to develop novel preoperative and intraoperative markers. For example, transbronchial markings, such as virtual-assisted lung mapping and intraoperative markings using cone-beam computed tomography, have been developed. This review explores various marking methods that have been practically applied for a better understanding of preoperative and intraoperative markings in thoracic surgery. Recently, several attempts have been made to perform intraoperative molecular imaging and dynamic virtual three-dimensional computed tomography for the localization, diagnosis, and margin assessment of small nodules. In this narrative review, the current status and future perspectives of preoperative and intraoperative markings in thoracic surgery are examined for a better understanding of these techniques. [ABSTRACT FROM AUTHOR]

Simple Summary: Our research integrates two key areas of focus. We first focus on a comparative biostatistical analysis of the agreement between three laboratory methods for genotyping gene copy number variations, and then we explore new characteristics of the distribution of this type of genetic aberration among cancer-associated genes. The results could be used to help molecular oncologists pick the smallest set of methods that will allow for accurate CNV genotyping in personalized cancer prognosis and drug-response prediction. We see the following strengths of our research: (1) We employ various methods for the detection of CNVs, such as CoreExome microarrays, nCounter v2 Cancer CN Assay panel, and digital droplet PCR; (2) This study sets out an interpretable comprehensive biostatistical analysis of the agreement using the PABAK score and Passing–Bablok regression; (3) This study presents the detection of CNVs in 12 paired tumor ovarian cancer/normal samples with clinical data; and (4) The research has translational potential. Objective: The goal of this study was to compare the results of CNV detection by three different methods using 13 paired carcinoma samples, as well as to perform a statistical analysis of the agreement. Methods: CNV was studied using NanoString nCounter v2 Cancer CN Assay (Nanostring), Illumina Infinium CoreExome microarrays (CoreExome microarrays) and digital droplet PCR (ddPCR). Results: There was a good level of agreement (PABAK score > 0.6) between the CoreExome microarrays and the ddPCR results for finding CNVs. There was a moderate level of agreement (PABAK values ≈ 0.3–0.6) between the NanoString Assay results and microarrays or ddPCR. For 83 out of 87 target genes studied (95%), the agreement between the CoreExome microarrays and NanoString nCounter was characterized by PABAK values < 0.75, except for MAGI3, PDGFRA, NKX2-1 and KDR genes (>0.75). The MET, HMGA2, KDR, C8orf4, PAX9, CDK6, and CCND2 genes had the highest agreement among all three approaches. Conclusions: Therefore, to get a better idea of how to genotype an unknown CNV spectrum in tumor or normal tissue samples that are very different molecularly, it makes sense to use at least two CNV detection methods. One of them, like ddPCR, should be able to quantitatively confirm the results of the other. [ABSTRACT FROM AUTHOR]

The article focuses on the molecular analysis of two-and-a-half-year-old twins diagnosed with odontohypophosphatasia due to a missense mutation in the alkaline phosphatase biomineralization associated (ALPL) gene. Topics discussed include the clinical presentation and diagnosis, identification of a novel heterozygous missense mutation (c.224G>A) in ALPL, and its predicted impact on tissue-nonspecific alkaline phosphatase (TNSALP) function.

Objective: This study aimed to compare the effectiveness of reflex molecular testing at identifying thyroid malignancy in indeterminate thyroid nodules (ITNs) since its implementation at our institution. Method: Identified all ITNs at our institution from January 2010 to October 2020. Calculated the sensitivity, specificity, negative predictive value (NPV), and positive predictive value (PPV) of ThyroSeq since the initiation of universal reflex testing of all first-time ITNs beginning in March 2016. Analyze effect on frequency of diagnostic surgeries. Results: Study group: March 2016 to October 2020, 378 ITNs underwent ThyroSeq (318 Bethesda III and 60 Bethesda IV). Mean age 52 years, 35.9% male, 61.1% female. 145 surgically excised with overall resection rate of 38.4% (III: 32.7%; IV 68%). Final histology: 49 malignant with overall rate of malignancy (ROM) of ITNs at 33.8% (III: 31.7%; IV: 39%). ThyroSeq sensitivity for ITNs at 84% (III 78.8%, IV 93.8%). ThyroSeq NPV for ITNs at 86% (III 84%, IV 93.3%). ThyroSeq specificity for ITNs at 52% (III 50.7%, IV 56%). ThyroSeq PPV for ITNs at 47% (III 42.6%, IV 57.7%). Control group: From January 2010 to February 2016 there were 242 ITNs (152 Bethesda III, 90 Bethesda IV). Mean age 52.6 years, 25.8% male, 74.2% female. 157 cases were surgically excised, with an overall resection rate of 64.9% (III: 57.2%; IV: 77.8%). Final histology: 32 malignant, with overall ROM of ITNs at 20.4% (III: 27.6%; IV: 11.4%). Conclusion: The initiation of universal Thyroseq (sensitivity 84% and NPV 86%) of ITNs at our institution has significantly decreased our percentage of diagnostic lobectomies, with a decreased resection rate of 26.5%. [ABSTRACT FROM AUTHOR]

Background: Platelet‐derived growth factor A (PDGFA) has been shown to be upregulated in several tumors, contributing to their malignant phenotypes. However, its expression and function in head and neck squamous cell carcinoma (HNSC) are not clearly understood. Thus, we aimed to evaluate this issue using bioinformatic analyses and primary experimental validation. Methods: The expression of PDGFA was analyzed using popular bio‐databases and further validated by RT‐PCR and immunohistochemical staining. Survival analyses were then performed. The association between PDGFA expression levels and immune cell infiltration in the immune microenvironment was assessed. Results: PDGFA has been found to be significantly upregulated in a variety of cancers, including HNSC, and increased PDGFA expression may be an independent prognostic factor associated with immune cell infiltration in HNSC. Conclusion: Overexpression of PDGFA in HNSC is significantly associated with poor prognosis and immune cell infiltration in the tumor microenvironment (TME). PDGFA has potential as a molecular indicator for diagnosis, prognosis, and immune processes in HNSC. [ABSTRACT FROM AUTHOR]

Introduction: Neonatal diabetes mellitus (NDM) is a rare non‐immunological monogenic disorder characterized by hyperglycemic conditions primarily occurring within the first 6 months of life. The majority of cases are attributed to pathogenic variants in genes affecting beta‐cell survival, insulin regulation, and secretion. This study aims to investigate the genetic landscape of NDM in Iran. Methods: We recruited a total of 135 patients who were initially diagnosed with diabetes at <12 months of age in Iran and referred to pediatric endocrinology clinics across the country. These patients underwent genetic diagnostic tests conducted by the Exeter Molecular Genetics Laboratory in the UK. The pathogenic variants identified were sorted and described based on type, pathogenicity (according to ACMG/AMP criteria), novelty, and the affected protein domain. Results: Genetic defects were identified in 93 probands, presenting various pathogenic abnormalities associated with NDM and its associated syndromes. 76% of the patients were born as a result of consanguineous marriage, and a familial history of diabetes was found in 43% of the cases. A total of 58 distinct variants in 14 different genes were discovered, including 20 variants reported for the first time. Causative variants were most frequently identified in EIF2AK3, KCNJ11, and ABCC8, respectively. Notably, EIF2AK3 and ABCC8 exhibited the highest number of novel variants. Discussion: These findings provide valuable insights into the genetic landscape of NDM in the Iranian population and contribute to the knowledge of novel pathogenic variants within known causative genes. [ABSTRACT FROM AUTHOR]

Since the World Health Organization (WHO) 2016 revision, the number of molecular markers required for diffuse gliomas has increased, placing a burden on clinical practice. We have established an in‐house, molecular diagnostic platform using Senshin‐Iryo, a feature of Japan's unique healthcare system, and partially modified the analysis method in accordance with the WHO 2021 revision. Herein, we review over a total 5 years of achievements using this platform. Analyses of IDH, BRAF, and H3 point mutations, loss of heterozygosity (LOH) on 1p/19q and chromosomes 10 and 17, and MGMT methylation were combined into a set that was submitted to Senshin‐Iryo as "Drug resistance gene testing for anticancer chemotherapy" and was approved in August 2018. Subsequently, in October 2021, Sanger sequencing for the TERT promoter mutation was added to the set, and LOH analysis was replaced with multiplex ligation‐dependent probe amplification (MLPA) to analyze 1p/19q codeletion and newly required genetic markers, such as EGFR, PTEN, and CDKN2A from WHO 2021. Among the over 200 cases included, 54 were analyzed after the WHO 2021 revision. The laboratory has maintained a diagnostic platform where molecular diagnoses are confirmed within 2 weeks. Initial expenditures exceeded the income from patient copayments; however, it has gradually been reduced to running costs alone and is approaching profitability. After the WHO 2021 revision, diagnoses were confirmed using molecular markers obtained from Senshin‐Iryo in 38 of 54 cases (70.1%). Among the remaining 16 patients, only four (7.4%) were diagnosed with diffuse glioma, not elsewhere classified, which was excluded in 12 cases where glioblastoma was confirmed by histopathological diagnosis. Our Senshin‐Iryo trial functioned as a salvage system to overcome the transition period between continued revisions of WHO classification that has caused a clinical dilemma in the Japanese healthcare system. [ABSTRACT FROM AUTHOR]

To study and compare the preimplantation genetic testing for monogenic disorders (PGT-M) results, and to evaluate the treatment cycle outcomes of embryos derived from a single pronucleus (1PN) vs. two pronuclei (2PN). A retrospective cohort study from January 2018 to December 2022 involving in vitro fertilization (IVF)-PGT-M treatment cycles. Single, academically affiliated fertility center. A total of 244 patients underwent 351 IVF-PGT-M treatment cycles. Embryo biopsy with molecular testing for a monogenic disorder. The molecular diagnosis results and clinical outcomes after the transfer of embryos derived from 1PN and 2PN in IVF-PGT-M treatment cycles. Embryos derived from 1PN have a significantly low developmental potential with a lower rate of embryos that underwent biopsy compared with 2PN-derived embryos; 1PN-derived embryos demonstrated a significantly lower number of blastocysts (24% vs. 37.9%) and top-quality blastocysts (22.3% vs. 48.1%) compared with 2PN-derived embryos. Lower successfully completed and unaffected PGT-M results were achieved in 1PN compared with 2PN-derived embryos (47.1% vs. 65.5% and 18.7% vs. 31.6%, respectively), with significantly higher abnormal molecular results (39.6% vs. 22.7%). The embryo transfer of 24 1PN-derived embryos with no affected genetic disorder resulted in 5 (20.8%) clinical pregnancies and 4 (16.7%) live births (LBs). Within the limits of fewer embryos derived from 1PN that yielded unaffected embryos suitable for transfer, the clinical pregnancy and LB rate of 1PN embryos undergoing PGT-M are reassuring. We, therefore, suggest applying PGT-M to embryos derived from 1PN embryos to improve the cumulative clinical pregnancy and LB rates. [ABSTRACT FROM AUTHOR]

We advocate for the widespread adoption of molecular diagnostics, alongside traditional methods, to support sustainable nematode management. Tangible examples utilising molecular diagnostics as proof-of-concept studies are urgently needed, particularly those involving longitudinal multiple grazing season studies. While absolute quantification of parasite DNA in samples is a priority, it is equally important to define thresholds for treatment and to develop and optimise tools that consider the host and its specific immune response. Facilitating open dialogue between farmers and veterinarians regarding sustainable management is essential, encouraging joint planning and collaboration instead of treating them as distinct entities with separate agendas. The management of parasitic nematodes calls for a shift from conventional, indiscriminate, anthelmintic use to a more precise approach, directed by diagnostics. We should accept those parasite infection intensities that have minimal impact on production and welfare rather than attempt to eliminate them. The diagnostic toolbox for gastrointestinal nematodes (GINs) faces challenges due to anthelmintic resistance (AR), which is species-specific, drug-class-specific, and varies by region. We discuss which traditional tools may become obsolete and which tools need development to gain widespread use. Social science research highlights the need for dialogue between farmers and veterinarians that emphasises effective parasite management and upskilling the veterinary workforce for more sustainable practices centred on diagnostics to be adopted in practice by 2030. [ABSTRACT FROM AUTHOR]

Purpose: Primary central nervous system lymphoma (PCNSL) is a rare but deadly malignancy that principally affects adults in the fifth and sixth decades of life. Despite diagnostic advances in analyses of cerebral spinal fluid and neuroimaging, definitive diagnosis of PCNSL requires primary brain tissue biopsy. While small neurosurgical biopsy volumes are pursued to minimize removal of normal brain tissue, the spatial margins to precisely biopsy pathologic tissue are narrow and can result in missed diagnoses. Furthermore, prior steroid treatment can significantly reduce tumor burden increasing the likelihood of a non-diagnostic biopsy. Methods: A retrospective case report from a tertiary referral center using a combination of neuroradiological studies, sterotactic tissue biopsy, and molecular testing for genome mutations. Results: A 72-year-old woman with strong suspicion for PCNSL clinically and radiologically, but cerebral spinal fluid and primary brain tissue biopsy were negative for tumor. However, vitreous liquid biopsy molecular testing for a MYD88 mutation as well as B-cell clonality (IGH/IGK rearrangement) were positive, indicating the presence of secondary vitreoretinal lymphoma from PCNSL. Only after autopsy of her brain was histopathological and immunohistochemical evidence of PCNSL confirmed. Conclusion: This case illustrates the unique contribution of liquid biopsy neuropathology-oriented molecular testing in a challenging case with high clinical suspicion of PCNSL in which gold-standard diagnostic testing failed to yield a diagnosis. [ABSTRACT FROM AUTHOR]

Genetic epilepsy diagnosis is increasing due to technological advancements. Although the use of molecular diagnosis is increasing, chromosomal microarray analysis (CMA) remains an important diagnostic tool for many patients. We aim to explore the role and indications of CMA in epilepsy, given the current genomic advances. We obtained data from 378 epileptic described patients, who underwent CMA between 2015 and 2021. Different types of syndromic or nonsyndromic epilepsy were represented. After excluding patients who were undertreated or had missing data, we included 250 patients with treated epilepsy and relevant clinical information. These patients mostly had focal epilepsy or developmental and epileptic encephalopathy, with a median start age of 2 years. Ninety percent of the patients had intellectual disability, more than two thirds had normal head size, and 60% had an abnormal magnetic resonance imaging. We also included 10 patients with epilepsy without comorbidities. In our cohort, we identified 35 pathogenic copy number variations (CNVs) explaining epilepsy with nine recurrent CNVs enriched in patients with epilepsy, 12 CNVs related to neurodevelopmental disorder phenotype with possible epilepsy, five CNVs including a gene already known in epilepsy, and nine CNVs based on size combined with de novo occurrence. The diagnosis rate in our study reached 14% (35 of 250) with first-line CMA, as previously reported. Although targeted gene panel sequencing could potentially diagnose some of the reported epilepsy CNVs (34% [12 of 35]). CMA remains a viable option as the first-line genetic test in cases where other genetic tests are not available and as a second-line diagnostic technique if gene panel or exome sequencing yields negative results. [ABSTRACT FROM AUTHOR]

This review examines the advancements in magnetic resonance imaging (MRI) techniques and their pivotal role in diagnosing and managing gliomas, the most prevalent primary brain tumors. The paper underscores the importance of integrating modern MRI modalities, such as diffusion-weighted imaging and perfusion MRI, which are essential for assessing glioma malignancy and predicting tumor behavior. Special attention is given to the 2021 WHO Classification of Tumors of the Central Nervous System, emphasizing the integration of molecular diagnostics in glioma classification, significantly impacting treatment decisions. The review also explores radiogenomics, which correlates imaging features with molecular markers to tailor personalized treatment strategies. Despite technological progress, MRI protocol standardization and result interpretation challenges persist, affecting diagnostic consistency across different settings. Furthermore, the review addresses MRI's capacity to distinguish between tumor recurrence and pseudoprogression, which is vital for patient management. The necessity for greater standardization and collaborative research to harness MRI's full potential in glioma diagnosis and personalized therapy is highlighted, advocating for an enhanced understanding of glioma biology and more effective treatment approaches. • Modern MRI techniques are key in diagnosing and guiding glioma treatment. • Modern MRI techniques are key in diagnosing and guiding glioma treatment. • Standardizing MRI protocols remains challenging, needing more research and collaboration. • Differentiating tumor recurrence from pseudoprogression is crucial; advanced MRI often needs biopsy validation. [ABSTRACT FROM AUTHOR]

The most common antibiotic-resistant bacteria in Korea are methicillin-resistant Staphylococcus aureus (MRSA) and vancomycin-resistant enterococci (VRE). Pathogen identification in clinical laboratories can be divided into traditional phenotype- and genotype-based methods, both of which are complementary to each other. The genotype-based method using multiplex real-time polymerase chain reaction (PCR) is a rapid and accurate technique that analyzes material at the genetic level by targeting genes simultaneously. Accordingly, we aimed to develop a rapid method for studying the genetic characteristics of antibiotic-resistant bacteria and to provide an experimental guide for the efficient antibiotic resistance gene analysis of mecA detection for MRSA and vanA or vanB detection for VRE using a one-step multiplex qPCR assay at an early stage of infection. As a result, the sensitivity and specificity of the mecA gene for clinical S. aureus isolates, including MRSA and methicillin-susceptible S. aureus, were 97.44% (95% CI, 86.82–99.87%) and 96.15% (95% CI, 87.02–99.32%), respectively. The receiver operating characteristic area under the curve for the diagnosis of MRSA was 0.9798 (*** p < 0.0001). Therefore, the molecular diagnostic method using this newly developed one-step multiplex qPCR assay can provide accurate and rapid results for the treatment of patients with MRSA and VRE infections. [ABSTRACT FROM AUTHOR]

Leptospirosis is a widespread zoonosis caused by spirochaetes belonging to the pathogenic species of Leptospira, which are classified into more than 25 serogroups and 250 serovars. Vaccination can prevent the disease in dogs but offers incomplete efficacy because of a lack of cross-protection between serogroups. The aim of this study was to validate a robust recruitment and sampling process, with the objectives of isolating and typing circulating Leptospira pathogenic strains and then selecting those of proven virulence and pathogenicity for vaccine development. Blood and urine samples from dogs with clinical syndromes compatible with acute leptospirosis were sterilely collected and transported to a reference laboratory for a micro-agglutination test (MAT), PCR, and bacterial isolation. Isolated strains underwent molecular typing using RNA16S, variable-number tandem repeat (VNTR), and pulsed-field gel electrophoresis (PFGE). Subtyping was performed using core genome multilocus sequence typing (CgMLST). Among 64 included dogs, 41 had MAT and/or PCR results compatible with Leptospira infection, and 14 Leptospira strains were isolated. Based on molecular typing, 11 isolates were classified as L. interrogans serogroup Australis, serovar Bratislava, and 3 as serogroup Icterohaemorrhagiae, serovar Icterohaemorrhagiae. CgMLST subtyping revealed a diversity of clonal groups (CGs) distributed in several regional clusters. Besides validating a robust recruitment and sampling process, this study outlines the value of combining PCR and serological testing when suspecting leptospirosis and the usefulness of implementing molecular typing methods to identify circulating field strains. It also confirms the epidemiological importance of the Australis serogroup and allows for the collection of different highly pathogenic strains for vaccine development. [ABSTRACT FROM AUTHOR]

Background: Renal hypokalemia is associated with mutation. This study aimed to investigate the clinical features and pathogenic mutations in patients with renal hypokalemia. Methods: The patients with hypokalemia were enrolled, and the renal function, thyroid function, renin-aldosterone system, urinary potassium excretion, and exome sequencing were performed. The correlation between the clinical phenotypes and causative genes was assessed. Results: Five patients with hypokalemia were enrolled and diagnosed as tubular hypokalemia. The patients with common clinical manifestations were difficult to differentiate based on atypical laboratory findings. The results of the genetic analysis were as follows: both patient 1 and patient 2 were heterozygous for the c.C625T mutation of the KCNJ1 gene, which is responsible for Bartter syndrome. Patient 3 was heterozygous for the c.G298A mutation of the ATP6V1B1 gene, which is responsible for renal tubular acidosis. Patient 4 had a compound heterozygous mutation of c.G893A of the BSND gene, responsible for Bartter syndrome, and c.1029+5G>A, the ATP6V0A4 gene responsible for distal renal tubular acidosis. Patient 5 had Gitelman syndrome and carried the compound heterozygous mutations c.C1963T and c.G2029A of the SLC12A3 gene. All the above loci were known heterozygous mutations. Conclusions: The unusual heterozygous mutations were identified in five renal hypokalemia patients. Molecular diagnosis of tubular hypokalemia was conducive to accurate diagnosis and treatment. [ABSTRACT FROM AUTHOR]

Objective: Thalassemia is a Mendelian‐inherited blood disorder with severe consequences, including disability and mortality, making it a significant public health concern. Therefore, there is an urgent need for precise diagnostic technologies. We introduce two innovative diagnostic techniques for thalassemia, SNPscan and CNVplex, designed to enhance molecular diagnostics of thalassemia. Methods: The SNPscan and CNVplex assays utilize variations in PCR product length and fluorescence to identify multiple mutations. In the SNPscan method, we designed three probes per locus: two 5′ and one 3′, and incorporated allele identification link sequences into one of the 5′ probes to distinguish the alleles. The detection system was designed for 67 previously reported loci in the Chinese population for a specific genetic condition. CNVplex identifies deletion types by analyzing the specific positions of probes within the globin gene. This innovative approach enables the detection of six distinct deletional mutations, enhancing the precision of thalassemia diagnostics. We evaluated and refined the methodologies in a training cohort of 100 individuals with confirmed HBA and HBB genotypes. The validation cohort, consisting of 1647 thalassemia patients and 100 healthy controls, underwent a double‐blind study. Traditional diagnostic techniques served as the control methods. Results: In the training set of 100 samples, 10 mutations (Hb QS, Hb CS, Hb Westmead, CD17, CD26, CD41‐42, IVS‐II‐654, ‐‐SEA, −α3.7 and −α4.2) were identified, consistent with those identified by traditional methods. The validation study showed that SNPscan/CNVplex offered superior molecular diagnostic capabilities for thalassemia, with 100% accuracy compared to 99.43% for traditional methods. Notably, the assay identified three previously undetected mutations in 10 cases, including two deletion mutations (Chinese Gγ(Aγδβ)0 del and SEA‐HPFH), and one non‐deletion mutation (Hb Q‐Thailand). Conclusions: The SNPscan/CNVplex assay is a cost‐effective and user‐friendly tool for diagnosing thalassemia, demonstrating high accuracy and reliability, and showing great potential as a primary diagnostic method in clinical practice. [ABSTRACT FROM AUTHOR]

Fragile X syndrome is a genetic condition causing a range of developmental problems, with males more severely affected compared to female patients. The main features include a long and narrow face, large ears, and a prominent jaw and forehead. Males develop enlarged testicles after puberty, and carrier females are expected to show fragile X-associated primary ovarian insufficiency (FXPOI). Fragile X Syndrome (FXS) was suspected in a consanguineous family referred to a Medical Genetics center because of a family history of intellectual disability and primary ovarian insufficiency in their small village population. The cytosine guanine guanine (CGG) repeat expansion of the FMRI gene in the 65-year-old proband was amplified and then analyzed by Gene Marker software. The female proband showed two expanded alleles, including one full mutation allele and one premutation allele with an accurate size of 74 (CGG) repeats. Despite having two mutant FMRI alleles and manifesting some symptoms of FXS, she was fertile. Consanguineous marriages and, in more unfavorable conditions, marrying Fragile X-affected or premutation-carrying males with female carriers is not uncommon in such genetically isolated populations. Therefore, the need for Fragile X syndrome examination in suspected patients with similar features and screening their relatives is highly emphasized [ABSTRACT FROM AUTHOR]

Molecular diagnostic technologies empower new clinical opportunities in precision medicine. However, existing approaches face limitations with respect to performance, operation and cost. Biological molecules including proteins and nucleic acids are being increasingly adopted as tools in the development of new molecular diagnostic technologies. In particular, leveraging their complementary properties—the functional diversity of proteins and the precision programmability of nucleic acids—a wide range of protein-nucleic acid hybrid nanostructures have been developed. These hybrid structures take diverse forms, ranging from one-dimensional to three-dimensional hybrids, as static assemblies to dynamic machines, and possess myriad functions to recognize target biomarkers, encode vast information and execute catalytic activities. Motivated by recent advances in this area of molecular nanotechnology, we review the state-of-art design and application of various types of protein-nucleic acid hybrid nanostructures for molecular diagnostics, and present an outlook on the challenges and opportunities for emerging pre-clinical and clinical applications, highlighting the promise for earlier detection, more refined diagnosis and highly tailored treatment decision that ultimately lead to improved patient outcomes. [ABSTRACT FROM AUTHOR]

Genetic diagnosis plays a crucial role in rare diseases, particularly with the increasing availability of emerging and accessible treatments. The International Rare Diseases Research Consortium (IRDiRC) has set its primary goal as: "Ensuring that all patients who present with a suspected rare disease receive a diagnosis within one year if their disorder is documented in the medical literature". Despite significant advances in genomic sequencing technologies, more than half of the patients with suspected Mendelian disorders remain undiagnosed. In response, IRDiRC proposes the establishment of "a globally coordinated diagnostic and research pipeline". To help facilitate this, IRDiRC formed the Task Force on Integrating New Technologies for Rare Disease Diagnosis. This multi-stakeholder Task Force aims to provide an overview of the current state of innovative diagnostic technologies for clinicians and researchers, focusing on the patient's diagnostic journey. Herein, we provide an overview of a broad spectrum of emerging diagnostic technologies involving genomics, epigenomics and multi-omics, functional testing and model systems, data sharing, bioinformatics, and Artificial Intelligence (AI), highlighting their advantages, limitations, and the current state of clinical adaption. We provide expert recommendations outlining the stepwise application of these innovative technologies in the diagnostic pathways while considering global differences in accessibility. The importance of FAIR (Findability, Accessibility, Interoperability, and Reusability) and CARE (Collective benefit, Authority to control, Responsibility, and Ethics) data management is emphasized, along with the need for enhanced and continuing education in medical genomics. We provide a perspective on future technological developments in genome diagnostics and their integration into clinical practice. Lastly, we summarize the challenges related to genomic diversity and accessibility, highlighting the significance of innovative diagnostic technologies, global collaboration, and equitable access to diagnosis and treatment for people living with rare disease. [ABSTRACT FROM AUTHOR]

Preeclampsia is a potentially life-threatening condition for both mother and baby, characterized by hypertension and potential organ damage. Early diagnosis is crucial to mitigate its adverse health effects. Traditional diagnostic methods, which focus on late-manifesting symptoms like hypertension and proteinuria, underscore the need for molecular diagnostic approaches for timely detection. This study successfully designs and evaluates novel aptamers with high specificity and affinity for Vascular Endothelial Growth Factor (VEGF) and Placental Growth Factor (PlGF), biomarkers closely associated with preeclampsia. Using molecular docking, molecular dynamics simulations, and BioLayer Interferometry (BLI), we identified aptamers that demonstrated strong binding affinities, comparable or superior to traditional antibodies. Our findings suggest that these aptamers have the potential to be integrated into cost-effective, point-of-care diagnostic tools, significantly improving early detection and intervention strategies for preeclampsia. The robust performance of these aptamers marks a pivotal step toward the development of more reliable and accessible diagnostic solutions, with implications for better maternal and fetal health outcomes. [ABSTRACT FROM AUTHOR]

The present study focused on three autochthonous varieties of apple trees belonging to the old Tuscany germplasm (central Italy), Chitignano, Mora and Ruggine, and aimed to (a) evaluate the presence of the four main latent viruses of apple trees (apple chlorotic leaf spot virus [ACLSV], apple mosaic virus [ApMV], apple stem grooving virus [ASGV] and apple stem pitting virus [ASPV]) and (b) elucidate the biochemical and physiological responses of apple leaves and fruits challenged by ACLSV (the only virus detected). Indeed, despite no visible symptoms being reported, more than half of the assayed trees tested positive for ACLSV, although it occurred only in Mora and Ruggine. ACLSV impaired the photosynthetic performance of apple trees, especially in Ruggine leaves, but variety‐specific responses in terms of reorganization of photosynthetic pigment and primary metabolites were reported. Mora showed a higher ability to trigger the de‐epoxidation state of xanthophylls and to accumulate α‐tocopherol, while d‐glucose and sucrose increased only in infected Ruggine leaves, probably as a signalling response and/or an osmotic adjustment of this variety under ACLSV infection. Unexpectedly, ACLSV infection slightly increased pome size, but it dramatically reduced their quality (i.e., soluble solids content), again especially in Ruggine, the only variety where reductions of d‐fructose, d‐glucose and malic acid contents were reported. Overall, the evidence here reported could offer valuable insights to effectively cope with the detrimental viral diseases affecting this species and to protect the production of one of the most consumed and appreciated fruits in the world. [ABSTRACT FROM AUTHOR]