Search Results (380)

Fourth-generation immunoassays are widely used for HIV and syphilis screening; however, false-reactive results may increase confirmatory testing and operational burden in high-throughput laboratories. This study evaluated the comparative performance of automated chemiluminescent immunoassays (MAGLUMI^® HIV Ab/Ag Combi (Snibe Diagnostics Co. Ltd., Shenzhen, China), VITROS^® ECiQ HIV Combo (Ortho Clinical Diagnostics, Raritan, NJ, USA), MAGLUMI^® Syphilis (Snibe Diagnostics Co. Ltd., Shenzhen, China), and ARCHITECT^® Syphilis TP (Abbott Diagnostics, Abbott Park, IL, USA) within a routine diagnostic algorithm, incorporating antibody differentiation immunoassays (INNO-LIA^® HIV I/II Score (Fujirebio Europe N.V., Ghent, Belgium) and HIV-1 RNA PCR where applicable. A total of 240 archived serum samples for HIV testing and 180 for syphilis testing were analyzed. Agreement-based performance measures including sensitivity, specificity, overall percent agreement (OPA), and Cohen’s kappa (κ) were calculated as comparator-based estimates reflecting concordance within the routine diagnostic algorithm rather than absolute diagnostic accuracy against a universal reference standard. For comparisons with HIV-1 RNA PCR, positive and negative concordance rates are reported to reflect agreement between assays detecting different biological targets. Among samples with definitive (positive or negative) results, the MAGLUMI^® HIV Ab/Ag Combi assay showed complete agreement with INNO-LIA^® HIV I/II Score (κ = 1.00) and high agreement with PCR within the ARCHITECT^® HIV Ag/Ab Combo-reactive subset (κ = 0.90). The VITROS^® ECiQ HIV Combo assay demonstrated high agreement with INNO-LIA^® HIV I/II Score (κ = 0.916) and substantial agreement with PCR (κ = 0.715), with a lower negative concordance rate with PCR observed in the ARCHITECT-reactive subset. A parallel five-modality analysis of 11 discordant samples applying the CDC 2014 algorithm demonstrated that all three immunoassay platforms successfully detected confirmed HIV-seropositive individuals with controlled viremia despite negative PCR, while MAGLUMI^® HIV Ab/Ag Combi produced fewer false-reactive results than both ARCHITECT^® and VITROS^® in this discordant subset. Additionally, two cases showed INNO-LIA^® indeterminate results with positive PCR, consistent with acute HIV infection during the early seroconversion stage; all three immunoassay platforms produced signals above the non-reactive threshold in both cases. For syphilis testing, both MAGLUMI^® Syphilis and ARCHITECT^® Syphilis TP assays showed complete agreement with INNO-LIA^® Syphilis Score among samples with definitive results (κ = 1.00). In contrast, the RPR assay showed reduced positive predictive value (49.4%) and moderate agreement with INNO-LIA^® Syphilis Score (κ = 0.52). Automated chemiluminescent immunoassay (CLIA) platforms demonstrated high agreement within a structured diagnostic algorithm in a high-throughput screening setting. Differences in assay performance were observed across platforms, particularly with respect to discordant results in the ARCHITECT-reactive PCR-evaluated subset for HIV and non-treponemal concordance for syphilis. These platforms may support more efficient laboratory workflows; however, findings should be interpreted within the context of comparator-based classification rather than absolute diagnostic accuracy. Full article

Sensitive skin is characterized by hypersensitivity to normal stimuli, and objective diagnostic tools and treatments are still limited. Currently, cosmetics for sensitive skin are developed through the exclusion of known irritants rather than investigation into the underlying mechanisms of sensitivity. In this study, we developed an integrated pipeline combining transcriptome analysis via microneedle-based skin sampling (MISSM), bioinformatics, in vitro validation, and clinical assessment to identify sensitive skin-associated inflammatory biomarkers and cosmetic ingredients that regulate them. Candidate biomarkers and matched cosmetic ingredients were identified from transcriptomic data and validated in lactic acid-stimulated HaCaT and human dermal fibroblasts via qRT-PCR. A prototype emulsion was developed and evaluated in a 4-week open-label pilot clinical trial with longitudinal molecular monitoring via MISSM. After lactic acid stimulation, sensitive skin-associated biomarkers (MCOLN1, CYR61, PMAIP1, PTGS2, and HMGB2) were significantly upregulated in both cell types, and cosmetic ingredients that regulate these biomarkers were confirmed in vitro. The emulsion prototype demonstrated hypoallergenicity in a primary irritation test. In the pilot clinical trial, target biomarker expression was significantly reduced in MISSM-derived samples, with improvements in skin hydration, barrier function, redness, and sensory reactivity also observed. This integrated pipeline will enable the discovery of inflammatory biomarker-regulating cosmetic ingredients, with potential applicability to various inflammatory skin conditions. Full article

White pine blister rust disease (WPBR), caused by Cronartium ribicola, ranks among the most destructive pathogens of five-needle pines. We developed a hydroxynaphthol blue (HNB)-based Loop-mediated isothermal amplification (LAMP) assay enabling rapid, visual detection of C. ribicola directly following DNA extraction. LAMP primers targeting the internal transcribed spacer (ITS) region were designed and validated through in silico comparison with related Cronartium species and in vitro testing against sympatric forest fungi. The optimized 25 μL reaction contained 8.0 mM Mg²⁺, 1.0 mM dNTPs, and an inner-to-outer primer ratio of 8:1, with amplification conducted at 62 °C for 40 min. Positive amplification produced a distinctive color transition from purple to sky blue, enabling visual interpretation without instrumentation. Under the tested conditions, the assay achieved a detection limit of 460 ± 3.2 fg/μL genomic DNA—a 10-fold improvement over conventional PCR in concentration-based sensitivity. Assay applicability was evaluated using 211 field-collected Pinus armandii samples sourced from China. Detection efficiency varied significantly across tissue types. Symptomatic bark exhibited a substantially higher positive detection rate (68.97%, 95% CI: 49.2–84.7%) compared to needles from symptomatic trees (18.75%, 95% CI: 4.1–45.7%). Among asymptomatic samples, 3.75% of bark samples tested positive for C. ribicola DNA, whereas all needle samples were negative. Geographically, positive detections clustered at several discrete sampling sites in southwestern China, predominantly at elevated elevations. The established LAMP-HNB assay provides a rapid, visually interpretable diagnostic tool for early detection and quarantine monitoring of WPBR following DNA extraction. Beyond its practical utility, this assay establishes valuable baseline data for targeted disease surveillance in the context of evolving climate conditions. Full article

Background: The role of bacterial infection in canine extrahepatic biliary disease remains inconsistently characterized. This retrospective case series evaluated associations between ultrasonographic gallbladder findings and bile culture results in dogs. Materials and Methods: Sixty-seven dogs were screened between March 2024 and December 2025, of which 49 met inclusion criteria (clinical, biochemical, and ultrasonographic evidence of gallbladder disease) and underwent bile sampling by ultrasound-guided cholecystocentesis (n = 45) or intraoperative collection (n = 4). Samples were cultured aerobically and anaerobically; isolates identified by MALDI-TOF MS were further tested for in vitro antimicrobial susceptibility using Kirby–Bauer disk diffusion method. Results: Among the 49 included dogs, ultrasonography identified cholecystitis (75.5%), cholelithiasis (16.3%), and biliary mucocele (8.2%). Bile cultures were positive in 43/49 dogs (87.8%), yielding only aerobic bacteria. Escherichia coli (46.5%) and coagulase-positive Staphylococci (30.2%) were most frequently isolated organisms. However, these findings should be interpreted cautiously due to small subgroup sizes and the exploratory nature of the analysis. Conclusions: In this selected referral population, bacterial isolates were frequently recovered from bile samples, particularly in dogs with cholecystitis. Prospective studies involving larger populations are warranted to confirm these results, define the bacterial prevalence and clinical significance of bacterial colonization or infection, and further refine evidence-based diagnostic and treatment strategies for dogs with extrahepatic biliary disease. Full article

Mycobacterium chelonae is a rapidly growing nontuberculous mycobacterium (NTM) that can infect both immunocompetent and immunocompromised hosts. Cutaneous and soft tissue infections are the most common manifestations and occur more frequently in individuals with underlying immune dysfunction. Patients with chronic lymphocytic leukemia (CLL), particularly those receiving targeted therapies such as ibrutinib, may be at increased risk of opportunistic infections. The diagnostic workup, microbiological findings, antimicrobial susceptibility testing, and therapeutic approach adopted for a cutaneous M. chelonae infection arising in a CLL patient four months after the introduction of ibrutinib were described. Clinical course and surgical management are also reported. A 60-year-old beekeeper with B-cell CLL developed a progressive cutaneous lesion on the left lower limb within four months of starting ibrutinib. Culture of a skin biopsy identified M. chelonae. Antimicrobial therapy was initiated based on in vitro susceptibility testing, resulting in partial clinical improvement. Complete resolution required surgical excision of the infected tissue followed by skin grafting. The patient’s underlying hematologic disease, ongoing immunosuppression, and recent exposure to ibrutinib likely contributed to susceptibility and persistence of infection. This case highlights the increasing recognition of nontuberculous mycobacterial infections in immunocompromised individuals and underscores the importance of early diagnosis and susceptibility- guided therapy. Clinical response may be incomplete, and combined medical and surgical approaches may be required in selected cases. NTM infections should be considered in patients receiving Bruton’s tyrosine kinase inhibitors who present with persistent, atypical, or non-healing cutaneous lesions. However, the association between ibrutinib therapy and susceptibility to infection remains uncertain, as multiple predisposing factors may coexist. Increased awareness of this possible association, together with careful clinical evaluation, may facilitate earlier diagnosis and improved management. Full article

Background/Objectives: Reliable detection of residual root canal filling material after retreatment is essential for comparing retreatment protocols. However, available methods quantify different clinical–physical dimensions and may not yield comparable estimates. This in vitro study compared cone-beam computed tomography (CBCT) and digital microscopy (DGM) for detecting residual obturation material after retreatment, using microcomputed tomography (micro-CT) as the reference standard. Methods: Fifteen extracted human mandibular premolars with single, straight canals were instrumented, obturated with gutta-percha and a calcium silicate-based sealer (AH Plus Bioceramic), and retreated with ProTaper Universal Retreatment files. Residual material was assessed in the coronal, middle, and apical thirds using CBCT (voxel size 0.10 mm), micro-CT (voxel size 60 µm), and DGM after longitudinal root splitting. Surface-based (DGM) and volumetric (CBCT and micro-CT) outcomes were analyzed separately using Wilcoxon signed-rank tests, diagnostic accuracy metrics (sensitivity, specificity, predictive values), and Cohen’s kappa for agreement. Results: DGM showed low median residual surface percentages across thirds (0.34–1.52%), whereas CBCT yielded higher median residual volume percentages (10.20–14.20%) than micro-CT (3.27–5.04%). The difference in the middle third between CBCT and micro-CT remained significant after Bonferroni correction (p = 0.002). For binary detection, CBCT showed higher sensitivity but lower specificity (overclassification of positive thirds), whereas DGM showed high specificity but limited sensitivity in the coronal and middle thirds. Conclusions: Within the limitations of this laboratory study, micro-CT was the most reliable reference method. CBCT tended to overestimate residual material, suggesting that clinical decisions based solely on CBCT may lead to unnecessary retreatment. DGM underestimated remnants because it assesses only the exposed split surface. These method-specific limitations should guide both clinical interpretation and future research design. Full article

Synthetic biology is reshaping in vitro diagnostics (IVD) by enabling programmable and modular biosensing elements that can be integrated into point-of-care testing (POCT) platforms. Compared with conventional assays that depend on fixed chemistries and centralized instrumentation, synthetic biology-based systems offer adaptable molecular recognition, tunable signal processing, and flexible readout formats for decentralized diagnostics. In this review, we present synthetic biology-enabled IVD as programmable biosensing platforms organized into four functional layers: molecular recognition, signal transduction and amplification, output generation, and system integration. We discuss four major enabling modules, including cell-free protein synthesis (CFPS) systems, aptamer and riboswitch sensors, CRISPR-Cas diagnostic platforms, and microfluidic integration technologies. We summarize representative clinical applications from 2021 to 2025 in infectious disease detection, cancer biomarker analysis, and drug metabolism/toxicity screening. In addition, we examine practical considerations beyond analytical sensitivity, including matrix tolerance, workflow complexity, manufacturability, quantitative capability, and regulatory readiness. Finally, we highlight future directions for programmable diagnostics, including AI-assisted biosensor design, multimodal readouts, interoperable platform architectures, and real-world clinical validation. Full article

The global tilapia industry is threatened due to the mass die-off of farmed tilapia caused by an emerging virus, Tilapia Lake Virus (TiLV). The threat of TiLV disease demands for cost-effective diagnostics and preventive measures such as protein and antibody-based detection tools and even vaccines like protein subunit vaccines. In this study, we aimed to recombinantly express the entire S4 protein of TiLV in vitro and in vivo using an extended host-range baculovirus expression vector system (EHR-BEVS) and used the purified protein to generate polyclonal anti-S4 antibodies. The parental extended host-range ABM-eIF4E baculovirus previously developed in our laboratory was used to generate the recombinant ABM-eIF4E-S4-His baculovirus expression vector using homologous recombination and was purified through multiple rounds of plaque selection and end-point titration, eliminating the need for the use of antibiotic resistance gene selection marker. Different insect cell lines including Trichoplusia ni Hübner, 1802 larvae were infected with ABM-eIF4-S4-His to produce recombinant S4 protein. Western blotting was used to detect the recombinantly expressed S4 protein. We generated an antibiotic resistance gene-free EHR-BEV and observed the permissibility of all the insect cell lines tested including T. ni larvae to infection with the recombinant ABM-eIF4E-S4-His. Secondly, immunogenic S4 protein capable of eliciting rabbit polyclonal anti-S4 antibodies was recombinantly expressed in the infected insect cells and larvae. TiLV S4 protein was recombinantly expressed in vitro and in vivo using extended host-range ABM-eIF4E-S4-His baculovirus expression vector and was shown to induce rabbit antibody production that can recognize S4 protein supporting the potential of insect larvae as alternative biofactories in the production of recombinant TiLV protein subunit for the development of diagnostics and preventive vaccines against TiLV disease. Full article

Bst DNA polymerase is a biotechnologically modified thermostable enzyme from the thermophilic Gram-positive bacterium Geobacillus stearothermophilus. The unique structure of Bst DNA polymerase determines its thermal stability, ability to replace a DNA strand and specificity. The high specificity of Bst DNA polymerase ensures the efficiency, sensitivity, and high rate of loop-mediated isothermal amplification (LAMP), which is widely used in vitro biotechnology. The review reveals the structural and functional features of the enzyme, its application in LAMP and methods of improvement of thermal stability (including directed evolution, site-directed mutagenesis, fusion constructs, and chemical modifications). The terminal transferase activity and ab initio synthesis are discussed regarding problems of Bst DNA polymerase and the ways to eliminate them. The questions of introducing modified nucleotides and primers to expand the diagnostic capabilities of LAMP are also discussed. Modern advances in Bst DNA polymerase engineering pave the way for the creation of reliable, thermostable, and highly specific test systems suitable for widespread diagnostic applications. Full article

Over the past 200 years (1820–2020), global life expectancy has nearly tripled, increasing from 26 to 72.91 years, due to factors such as poverty reduction and public health initiatives. Today, society faces different challenges than it did centuries ago. In patient care and healthcare system priorities, the goal is to develop smart, feasible, long-lasting, cost-effective, readily available, adverse-reaction-free, adaptable, and personalized solutions that minimize patient discomfort, reduce caregiver effort, and decrease hospitalization duration and costs. In this context, biomaterials serve as versatile tools capable of performing a wide range of diagnostic, therapeutic, and theragnostic functions. Thanks to their biocompatibility, biodegradability, surface chemistry, and responsiveness, biomaterials are currently addressing issues such as patient compliance (through controlled drug-delivery systems and smart wound dressings), long transplant waiting lists, transplant rejection, non-adaptable prosthetics (artificial organs), oncology treatment efficacy (nano-formulations for theragnostics and multiple tumor targeting), and inconsistent in vitro drug-testing models (organs-on-a-chip). In this review, we focus on biomaterials’ smartness, then explore databases for efficient product design, and finally highlight their applications in the biomedical field, especially in drug delivery, tissue engineering, and regenerative medicine. Full article

Background: Biodegradable magnesium-based alloys are increasingly explored as emerging biomaterials for dental and maxillofacial applications due to their osteoconductive properties and potential to reduce long-term implant-related complications. However, early-stage evaluation requires predictive diagnostic screening methods capable of assessing cytocompatibility and cellular response under clinically relevant extract conditions. Objectives: In this study, Mg–0.5Ca alloys modified with increasing strontium concentrations (0.5–3 wt.%) were investigated through an in vitro diagnostic framework using MG-63 osteoblast-like cells. Methods: Cell viability was quantitatively assessed via MTT assays after 24 and 72 h of exposure, while fluorescence-based live-cell imaging provided complementary morphological insights. Results: demonstrated a composition-associated cytocompatibility profile, with Sr-enriched compositions showing improved cellular metabolic activity and adhesion patterns compared to lower-Sr compositions. Conclusions: These findings support the role of strontium as a functional alloying element and highlight the importance of standardized diagnostic screening workflows for emerging dental biomaterials. Overall, this study proposes a simplified predictive platform for early biocompatibility diagnostics, contributing to the integration of biomaterial evaluation into future digitalized dental regeneration workflows. Full article

Background/Objectives: Early embryonic arrest during the cleavage stage (days 2–4) accounts for a substantial proportion of developmental failure in in vitro fertilization. This phenomenon remains poorly understood at the molecular level, even in chromosomally normal embryos identified by preimplantation genetic testing. This review aims to redefine cleavage-stage arrest from a passive energy deficit to a checkpoint-regulated endpoint caused by inadequate coordination among metabolism, transcriptome integrity, and stress-response pathways. Methods: We integrate evidence from long-read transcriptomics, metabolomics, epigenetics, and immunobiology relevant to pre-blastocyst development. These data are assembled into a unifying mechanistic framework and a clinically oriented stratification model, together with candidate multimodal readouts for early classification. Results: We propose a three-axis model linking: (i) metabolic–epigenetic insufficiency, including defective histone lactylation and impaired alpha-ketoglutarate-dependent DNA demethylation; (ii) isoform-level abnormalities, including intron retention and retrotransposon activation within a hidden transcriptomic landscape better resolved by long-read sequencing; and (iii) stress-related immune signaling, in which NLRP7 links alternative splicing and DNA-damage-response dysfunction with mitochondrial stress and p53-associated arrest. Within this framework, we distinguish three molecular arrest states: an early transition failure marked by defective maternal-to-embryonic reprogramming and severe splicing disruption; a metabolically quiescent state that may retain a limited rescue window; and a later stress-associated state characterized by senescence-like features, oxidative stress, and broad transcriptomic and genomic instability. Conclusions: Early embryo arrest should no longer be viewed as a nonspecific developmental failure, but as a mechanistically stratifiable condition with distinct metabolic, transcriptomic, and stress-associated trajectories. A diagnostic platform combining fluorescence lifetime imaging microscopy, long-read sequencing, and digital polymerase chain reaction may improve early mechanistic classification, help identify embryos with possible reversibility, and reduce uncertainty in embryo selection during in vitro fertilization. Full article

Shigellosis remains a significant global cause of infectious colitis, increasingly complicated by multidrug-resistant strains and the microbiota-disrupting effects of broad-spectrum antibiotics. Although conventional antimicrobial therapy can reduce symptom duration and bacterial shedding, it also contributes to gut dysbiosis, loss of colonization resistance, and further selection for antimicrobial resistance. These challenges have renewed interest in precision antimicrobial strategies, particularly bacteriophage therapy, which provides strain-level specificity and preserves the gut microbiota. This narrative review evaluates the biological rationale, preclinical and early clinical evidence, safety considerations, and translational challenges associated with bacteriophage therapy targeting Shigella spp. The historical development and mechanistic basis of phage therapy are summarized, with emphasis on the advantages of obligately lytic phages, receptor-specific targeting, self-amplification at infection sites, and activity against both planktonic and biofilm-associated bacteria. Recent microbiota research indicates that shigellosis is closely associated with early and persistent disruption of gut ecology, including depletion of short-chain fatty acids-producing taxa and reduced microbial resilience. Phage-based approaches may reduce pathogen burden while preserving beneficial microbial communities. Evidence from in vitro systems, animal models, human intestinal organoids, and a Phase 1 clinical trial demonstrates targeted efficacy and favorable safety profiles for Shigella-specific phages and phage cocktails. Major barriers to clinical adoption include immune interactions, phage resistance dynamics, genomic safety screening, regulatory classification, and the need for standardized susceptibility testing. Future directions emphasize the development of personalized phage therapy platforms that integrate rapid diagnostics, phage libraries, metagenomics, and artificial intelligence-assisted matching to enable scalable, precision treatment. Full article

This study aimed primarily to evaluate the performance of two Conformité Européenne—In Vitro Diagnostic (CE-IVD) multiplex real-time PCR (rt-PCR) assays for the molecular identification of tick-borne pathogens (TBPs) of human interest on ticks removed from human skin and collected through a citizen science-based approach. As a secondary objective, the aggregated results were used to describe tick species distribution, developmental stages, and seasonal TBP circulation in 2024 in the considered area. The comparison was conducted on 116 tick samples collected in 2024 voluntarily delivered to a hospital in northeastern Italy. Detected TBPs were further confirmed with in-house-validated PCR methods and, where applicable, resolved to the species level. Clinically relevant pathogen species were identified as single infections or coinfections. Overall, 33.6% of tick samples tested positive for at least one TBP, and 6.9% showed coinfections. Kit B exhibited a higher detection rate for Borrelia spp. and Rickettsia spp. targets, partly reflecting its broader diagnostic specificity, while statistically significant differences in cycle threshold values were observed for Anaplasma phagocytophilum detection. The most frequently involved ticks were Ixodes ricinus nymphs, and the most represented area was Verona province. Late spring and early summer were identified as the periods with the highest tick conferment and pathogen diversity. Overall, the results support the use of multiplex real-time PCR commercial kits combined with citizen science-based tick collection as an effective approach for both diagnostic screening and regional surveillance of circulating ticks and TBPs. Full article

Rapid and actionable pathogen identification remains a major unmet need in the diagnosis of prosthetic joint infection (PJI). Current diagnostic approaches either provide rapid host response information without pathogen specificity or identify pathogens with delays of days to weeks. Here, we report a chemiresistive nanosensor array combined with machine learning analysis for same-day, pathogen-specific detection based on volatile organic compound (VOC) profiling. A 19-channel nanosensor array was first validated in vitro against a panel of ESKAPEE pathogens, achieving 96% mean classification accuracy using a radial-basis-function support vector machine (SVM) classifier. Data-driven optimization yielded a reduced six-sensor array with high signal-to-noise performance. The optimized platform was evaluated using pooled, uninfected human synovial fluid enriched 1:1 with nutrient media and spiked with Staphylococcus aureus, Staphylococcus epidermidis, or Pseudomonas aeruginosa across a range of 1–10⁶ CFU/mL. All infected samples were detected within 9 h, with distinct VOC signatures enabling accurate pathogen differentiation. Time-to-detection (TTD) demonstrated a strong inverse correlation with initial bacterial concentration, supporting semi-quantitative estimation of bacterial load. Negative controls remained at baseline throughout testing. This chemiresistive VOC-based biosensor platform demonstrates the potential to deliver rapid, integrated detection, identification, and burden estimation of metabolically active PJI pathogens, highlighting its promise for future point-of-care diagnostic applications. Full article