Rapid Turnaround Time Research Articles

Improving Turnaround Times for Routine Antimicrobial Sensitivity Testing Following European Committee on Antimicrobial Susceptibility Testing Methodology in Patients with Bacteraemia.

Background/Objectives: Bacteraemia can be fatal without antibiotic intervention. Antibiotic Susceptibility Testing (AST) provides the necessary information for targeted antibiotic therapy; however, the traditional method using disc diffusion can take over two days from a positive blood culture. Inappropriate empiric therapy is associated with increased mortality and increased antibiotic resistance, highlighting the need for more rapid turnaround times for AST. By making changes to an established method, turnaround times can be reduced. Methods: Eighty-two patient positive blood culture samples were collected from January to April 2022, representing the range of common bacteria causing sepsis. This followed the normal methodology in the laboratory of inoculating agar from positive blood cultures in preparation for European Committee on Antimicrobial Susceptibility Testing (EUCAST) disc diffusion AST method. EUCAST methodology outlines that disc diffusion should be performed on isolates from an overnight culture of 16-24 h. This study looked at comparing disc diffusion results from cultures with 6 h of incubation to those with incubation times of 24 h, after organism identification by MALDI-ToF. Results from 6-h and 24-h cultures were compared by disc zone sizes and by interpreted susceptibility reading following EUCAST guidelines of sensitive, resistant, susceptible with increased exposure, or an area of technical uncertainty. Results: A total of 99.65% interpreted susceptibility readings matched across all organisms to all relevant antibiotics, with an average zone size difference of 1.08 mm between results from 6 h versus 24 h cultures. Conclusions: This method offers a non-automated way of using the traditional disc diffusion method, reducing turnaround times while still producing reliable and accurate results. This would mean validated ASTs can be set up in the same day as a blood culture flags positive rather than waiting for a longer culture. As this method is widely used within the laboratory already, it would mean that additional training is not required, as the process is the same, and only incubation time varies. This would positively impact patient outlook due to the shorter use of empiric therapy, and benefit antimicrobial stewardship (AMS).

Target-triggered catalytic hairpin assembly of miR-21/155/1 coupled with dsDNA-reporter amplified detection for prediction of clinically significant coronary artery disease

Multiple microRNAs (miRNAs) have demonstrated potential as early biomarkers of coronary artery disease (CAD), but diagnosis outcome is challenging because of the limitations of clinical application. In this study, we reported a general and machine learning-enhanced diagnostic strategy for miRNA analysis: catalytic hairpin assembly (CHA) and dsDNA-reporter-assisted cascade reaction for isothermal amplification. The sensor achieved high sensitivity and rapid turnaround time (<30min) using micro samples of patient serum in a one-pot assay without extraction and enrichment. Additionally, the system demonstrated robust multiplex capability in simultaneously detecting three miRNA targets. By training a machine learning model on a large dataset of miR-21/155/1, the assay was superior in differentiating CAD patients (n=115) from non-CAD individuals (n=30) achieving an enhanced accuracy of 87.5%, surpassing the individual performance of miR-1 (18.9%), miR-21 (75.6%), and miR-155 (51.1%). This reliable and user-friendly approach provides a powerful tool for multiple miRNA analysis, holding significant promise for point-of-care CAD diagnosis.

Seq-Scope: repurposing Illumina sequencing flow cells for high-resolution spatial transcriptomics.

Spatial transcriptomics technologies aim to advance gene expression studies by profiling the entire transcriptome with intact spatial information from a single histological slide. However, the application of spatial transcriptomics is limited by low resolution, limited transcript coverage, complex procedures, poor scalability and high costs of initial setup and/or individual experiments. Seq-Scope repurposes the Illumina sequencing platform for high-resolution, high-content spatial transcriptome analysis, overcoming these limitations. It offers submicrometer resolution, high capture efficiency, rapid turnaround time and precise annotation of histopathology at a much lower cost than commercial alternatives. This protocol details the implementation of Seq-Scope with an Illumina NovaSeq 6000 sequencing flow cell, allowing the profiling of multiple tissue sections in an area of 7 mm × 7 mm or larger. We describe the preparation of a fresh-frozen tissue section for both histological imaging and sequencing library preparation and provide a streamlined computational pipeline with comprehensive instructions to integrate histological and transcriptomic data for high-resolution spatial analysis. This includes the use of conventional software tools for single-cell and spatial analysis, as well as our recently developed segmentation-free method for analyzing spatial data at submicrometer resolution. Aside from array production and sequencing, which can be done in batches, tissue processing, library preparation and running the computational pipeline can be completed within 3 days by researchers with experience in molecular biology, histology and basic Unix skills. Given its adaptability across various biological tissues, Seq-Scope establishes itself as an invaluable tool for researchers in molecular biology and histology.

NanoString nCounter-Based Assay for Detection of Fusion-Associated Salivary Gland Tumors.

Salivary gland tumors include numerous subtypes that vary from benign to highly aggressive, with many showing overlapping histopathological features that can make diagnosis challenging. Most subtypes express driver fusion genes that are tumor specific, and detection of such fusions is useful for differentiating amongst specific diagnoses, determining appropriate tumor grading, and guiding effective treatment. Currently, fusions can be detected by FISH, RT-PCR or through next-generation sequencing approaches, all of which are highly effective methodologies but can be costly or time consuming. We developed a rapid NanoString nCounter platform-based assay to detect salivary gland tumor fusions using a combination of fusion junction-specific probes and an approach through differential exon expression analysis. The assay includes 68 junction-specific probes and analysis of exon expression across 9 fusion-associated genes in a single multiplex assay. Out of 55 retrospective and 171 prospective cases assayed, we accurately detected the majority of cases of pleomorphic adenoma, adenoid cystic carcinoma, cribriform adenocarcinoma, clear cell carcinoma, secretory carcinoma and NUT-rearranged carcinoma, including cases of these tumor types arising in non-salivary gland sites, with the major drawback being an inability to detect MAML2-containing mucoepidermoid samples. With mucoepidermoid carcinoma excluded, the assay shows an overall sensitivity of 96.1% and specificity of 100%. We show that the majority of salivary gland tumor fusions can be effectively detected with a single rapid NanoString based assay, which can serve as a useful adjunctive tool for routine diagnostic head and neck pathology. The assay is low cost with a rapid turnaround time (30h total assay time per sample batch, with minimal technician input required) compared to alternate detection methods.

Evaluation of the CARBA PAcE test, a colorimetric imipenem hydrolysis test for rapid detection of carbapenemase activity.

There is an urgent need for accurate and fast diagnostic tests to identify carbapenemase-producing bacteria. Here, we evaluated a colorimetric imipenem hydrolysis test, called the CARBA PAcE test, to detect carbapenemase-producing Gram-negative bacteria (GNB). We tested a collection of 270 GNB isolates with a characterized carbapenemase content. Our testing set included 205 carbapenemase-producing, carbapenemase-resistant Enterobacterales (CP CRE) with 40 Klebsiella pneumoniae carbapenemase (KPC), 49 New Delhi metallo beta lactamase (NDM), 49 OXA-48-like, 15 Verona integron-mediated metallo-β-lactamase (VIM), three IMP, 43 IMI, six isolates producing more than one carbapenemase, and 65 non-carbapenemase-producing Enterobacterales (20 ESBL producers, 35 non-carbapenemase-producing, carbapenem-resistant [non-CP CRE], and 10 carbapenem-susceptible Enterobacterales [non-CP, non-CSE, third-generation cephalosporin and carbapenem susceptible]). We compared the performances of the CARBA PAcE test, the qualitative colorimetric β-CARBA test, and the modified CarbaNP test to a gold standard of carbapenemase gene detection by PCR. Specificities of all tests were high: 95.4% (62/65) for CARBA PAcE test, 98.5% (64/65) for β-CARBA test, and 100% (65/65) for the modified CarbaNP test. Sensitivity varied by carbapenemase: all three tests had a sensitivity of 100% for NDM, VIM, and IMP and 97.5% for KPC. Sensitivity to detect IMI was 0% for the CARBA PAcE and β-CARBA tests and 11.6% for the modified CarbaNP test. Sensitivity to detect OXA-48-like was 89.7% for the CARBA PAcE test, 87.7% for the β-CARBA test, and 14.2% for the modified CarbaNP test. Reading the results of the CARBA PAcE assay was difficult. The CARBA PAcE assay is highly sensitive for detecting NDM, VIM, IMP, and KPC, but slightly less sensitive for OXA-48-like. It does not detect IMI. It is highly specific, and its overall diagnostic accuracy is similar to that of β-CARBA. Its operational advantages are rapid turnaround time, ease of use, and long shelf life, but reading of results is subjective.IMPORTANCEWe evaluated the ability of the CARBA PAcE test to detect carbapenemases in 274 Gram-negative isolates with a known carbapenemase content. Specificity was high for all carbapenemases tested (96.9%). Sensitivity was high for KPC, NDM, VIM, and IMP (97.5-100%); but lower for OXA-48-like (89.7%). Activity of IMI could not be detected. Taken together, our results indicate that CARBA PAcE is a useful alternative in regions where NDM and KPC are predominant. The limitations of the test are difficulty in reading results and incompatibility with mSuperCARBA.

Rapid detection of human adenovirus by multiple cross displacement amplification combined with nanoparticle-based biosensor platform

Human adenoviruses (HAdV), particularly serotypes 3 and 7 (HAdV-3 and HAdV-7), are significant respiratory pathogens that contribute to high morbidity rates and severe pneumonia in infants and children. The lack of distinct clinical presentations and effective treatments highlights the urgent need for rapid and reliable diagnostic methods for HAdV-3 and HAdV-7. This study devises a novel detection assay, termed HAdV-MCDA-LFB, which combines isothermal multiple cross displacement amplification (MCDA) with a nanoparticle-based lateral flow biosensor (LFB). Targeting the conserved hexon gene, HAdV-MCDA-LFB demonstrated high sensitivity, detecting low to 10 fg of hexon-containing plasmid per reaction without cross-reaction under the optimized conditions. Moreover, HAdV-MCDA-LFB exhibited comparable diagnostic accuracy to real-time PCR in clinical sample analysis, indicating its practical applicability. The whole procedure, including rapid template preparation (15 min), MCDA reaction (40 min at 67 °C) and result interpretation (<5 min), can be completed within one hour. Together, this rapid turnaround time, coupled with its simplicity and accuracy, makes HAdV-MCDA-LFB a promising point-of-care diagnostic tool for HAdV-3 and HAdV-7, particularly in resource-limited settings.

High-Throughput Hybridization Assay as First-Line Diagnostic Test for Sarcomas: Clinical Assessment in a Tertiary Referral Center.

Sarcomas are rare and highly heterogeneous mesenchymal tumors with deceptive morphologic features that pose a challenge for precise diagnostics. Chromosomal rearrangements generating pathognomonic gene fusions are useful diagnostic markers, traditionally tested using single-plex standard of care assays with limited diagnostic yield. NanoString nCounter technology has emerged as a robust solution with multiplexing capabilities. To optimize NanoString effective coverage of specific entities and conduct a validation study to support its clinical implementation. We reconfigured a NanoString's codeset by including a set of probes for detecting gene fusion variants of solitary fibrous tumors, low-grade fibromyxoid sarcomas/sclerosing epithelioid fibrosarcomas, and undifferentiated small round cell sarcomas, totaling 188 probes. A technical validation study was conducted with 96 retrospective samples. Additionally, 76 prospective samples were evaluated to assess the assay's clinical performance. Both technical and clinical validation studies showed that NanoString's codeset reached >88% sensitivity and 100% specificity, compared with standard of care methods, and superior diagnostic yield as a first-line test. Our design enabled the detection of almost all fusion variants of NGFI-A binding protein 2 (NAB2) with signal transducer and activator of transcription 6 (STAT6) in solitary fibrous tumors, as well as cAMP responsive element binding protein 3 like 1/2 (CREB3L1/2) rearrangements in all low-grade fibromyxoid sarcoma/sclerosing epithelioid fibrosarcoma cases. Identification of specific gene fusions of undifferentiated small round cell sarcoma was also improved, but additional strategies are necessary to attain full coverage. The NanoString platform demonstrated good sensitivity, specificity, and superior diagnostic yield. It is a cost-effective assay with rapid turnaround time, low sample consumption, streamlined analysis, and easy customization. Therefore, it is a promising alternative first-line diagnostic tool for routine sarcoma testing.

Clinical performance evaluation of the BioFire Joint Infection Panel.

The BioFire Joint Infection (JI) Panel offers a significant advancement in the rapid diagnosis of joint infections by facilitating the simultaneous detection of multiple bacterial and fungal pathogens, as well as resistance markers, directly from synovial fluid samples. An article published in the Journal of Clinical Microbiology by Moran et al. (J Clin Microbiol 62:e00182-24, 2024, https://doi.org/10.1128/jcm.00182-24) presents both prospective and retrospective analyses of the panel's real-world clinical application. The study highlights the panel's benefits, such as its rapid turnaround time and ability to identify challenging pathogens, while also discussing its limitations, particularly in detecting certain off-panel organisms.

Ultrasensitive Rapid Antigen Test by Geometric Lateral Flow Assays and Highly Doped Upconversion Nanoparticles.

The paper-based lateral flow assay (LFA) testing strips are currently the most widely used for point-of-care testing (POCT), valued for their rapid result turnaround times in a few minutes. However, their sensitivity has been limited. Upconversion nanoparticles (UCNPs), especially highly doped ones, have emerged as promising luminescent reporters to enhance the LFA sensitivity. These UCNPs exhibit a nonlinear enhancement in luminescence with excitation power density, necessitating higher power densities for higher brightness. In this study, we utilized a geometric paper strip design to minimize the immune reaction area and maximize the excitation power density, enabling ultrasensitive detection of the SARS-CoV-2 nucleoprotein antigen. This design also slowed the antigen flow on the paper strip, extending the reaction time between antigen and antibody, thereby enhancing the efficiency of the immune reaction. Through this design, our approach achieved over a 100-fold enhancement in the limit of detection (LOD) compared with the widely used LFAs, based on gold colloidal nanoparticles and europium nanoparticles. This innovation expands the scope of LFA applications that require a low LOD.

A-153 Study of the evaluation of fragment flag and percentage schistocyte count in MINDRAY® BC-6800 PLUS in a public pediatric hospital in Argentina

Abstract Background Schistocytes are fragmented red blood cells (FRCs) that arise from the mechanical destruction of red blood cells (RBCs). They indicate the possible presence of a thrombotic microangiopathy (TMA). TMA includes hemolytic uremic syndrome (HUS) and thrombotic thrombocytopenic purpura(TTP). ICSH defined schistocytes as 4 well stablish shapes always smaller than intact RBCs. The gold standard for the identification of schistocytes is microscopic evaluation. A schistocyte count should be considered clinically meaningful if it represent the main morphological RBCs abnormality in the periferic blood smear (PBS). The results should be expressed as a percentage, after counting at least 1000 RBCs in optimal areas of the PBS. The cut off for TMA was previously stablished in above 1% (adult population) and a higher percentaje in pre term newborns (above 5%). Some blood cell count analyzers have a flag for FRC, and some of them also provide methods for an automated FRC count, through the measurement of forward scatter and the intensity of fluorescence or 2-dimensional optical analysis. The Mindray® BC 6800 PLUS provides a flag for fragments and the research FRC% parameter that quantifies fragmented RBCs. When the flag and the FRC% are combined, can bring value to diagnostic TMA. Objetives Evaluate the sensitivity and specificity of the fragment flag to predict schistocyte on the PBS of pediatric samples. Evaluate the utility of FRC% as TMA diagnostic tool and compare with the gold standard. Methods 725 pediatric samples were analyzed (0-18 years).16 of them was patient samples diagnosed with TMA. All blood cell counts were determined using Mindray®BC-6800 PLUS. Cut off 1% was considered for FRC% in TMA. Inclusion criteria for TMA samples include thrombocytopenia, increased lactate dehydrogenase and decreased hemoglobin concentration. Schistocyte counting was carried out from May-Grunwald-Giemsa-stained PBS according to the recommendations proposed by ICSH. Only ICSH established shapes were kept as valid. Results Regarding the sensitivity of the fragment flag, it was 84.62% with a specificity of 91.13% to detect the presence of schistocytes in PBS samples. In the Bland-Altman analysis, the FRC% shows a difference with a tendency to underestimation with respect to the observation on the PBS, which was less at lower percentages of schistocytes. The sensitivity of the FRC% to TMA was 75% while the specificity was 94.36% and a negative predictive value (NPV) of 99.41%. Conclusions Fragments flag represents a potentially powerful tool for screening of squistocytes in pediatric samples. Although sensibility and NPV for FRC% authomatical counts were acceptable, a few false-negative samples were found. Presence of FRC% below a threshold of 1% could exclude TMA.However PBS microscopic observation is still the gold standard to determine squistocytes presence. For the purpose of complete our study and to get further larger scale prospective data, we started collecting all data for all samples entering the laboratory in order to improve an authomatical method that should have a more rapid turn-around time at lower costs.

Should Routine Diagnostics Implement Gut Microbiota Analysis?

The gut microbiota plays a key role in human health. Dominated by the phyla Firmicutes and Bacteroidetes, its composition is highly individualized and influenced by diet, age, genetics, and the environment. The gut-liver axis highlights the bidirectional re-lationship between the gut and the liver, impacting metabolic homeostasis and immune reg-ulation. Gut dysbiosis, an imbalance in the gut microflora, contributes to liver diseases by disrupting gut barrier function and bile acid metabolism, leading to inflammation and fibro-genesis. Advancements in omics approaches, such as metagenomics, metatranscriptomics, metaproteomics, and metabolomics, have enhanced our understanding of the gut microbi-ota. These approaches offer insights into microbial composition and function, although they vary in cost, efficiency, and complexity. Metagenomics is widely used for its cost-effec-tiveness and rapid turnaround time despite limitations in taxonomic resolution, while meta-transcriptomics, metaproteomics, and metabolomics offer functional and metabolic insights but require sophisticated techniques and expertise. The Firmicutes/Bacteroidetes (F/B) ratio is a potential biomarker of gut dysbiosis linked to obesity, type 2 diabetes mellitus, and liver diseases. However, its diagnostic reliability is debated due to variations in individual factors and a lack of data on its associations with several diseases. Future research should focus on integrating multi-omics approaches so as to provide a holistic view of the gut microbiota and its role in health and disease, aiming for applications in precision medicine. While promising, the F/B ratio should be used cautiously alongside other diagnostic measures. In addition, renewed efforts are needed to develop cost-effective and rapid analysis methods for clinical use.

Nanoparticle-Supported, Rapid, and Electronic Detection of SARS-CoV-2 Antibodies and Antigens at Attomolar Level.

A gold nanoparticle (AuNP)-supported rapid, electronic detection (NasRED) platform is demonstrated with high specificity to differentiate SARS-CoV-2 from human coronaviruses while achieving high sensitivity. Uniquely, active fluidic force is exerted on AuNPs in a microcentrifuge tube, through engineered centrifugation and vortex agitation, to accelerate signal transduction from biochemical protein binding to physical AuNP aggregation and precipitation and finally to electronic signals in a customized, stabilized circuitry. The limits of detection are found <80 aM (12 fg/mL) and ∼700 aM (105 fg/mL) to detect antibodies in human pooled serum and 20% diluted whole blood, and <900 aM (45 fg/mL) and 4.1 fM (200 fg/mL) to detect nucleocapsid proteins in human saliva and nasal fluid, respectively. With a small test volume (6 µL), a rapid turnaround time (as fast as <15 min), low reagent cost (estimated ∼$2), and a small system footprint, this novel digital platform can be utilized for accurate diagnosis and screening of a wide range of infectious and chronic diseases.

Evaluation of Lyme serologic quantitative test indexes: High first-tier test index values predict positive second-tier result in standard and modified 2-tier Lyme testing algorithms.

In this study, we evaluated the potential utility of reporting a quantitative Lyme serologic test index to improve the utility of results from first-tier Lyme assays. Serum from consecutive samples sent to our laboratory for Lyme testing were tested on 2 commercial first-tier Lyme assays and evaluated to determine the probability of second-tier confirmation based on the serologic index value. For both assays, we identified an index value above which 100% of samples confirmed on second-tier testing using both standard and modified 2-tier testing algorithms. Lower rates of confirmation were observed for positive or equivocal samples with lower index values. The use of a Lyme test index value may eliminate the need for confirmatory testing on many positive first-tier samples, providing more rapid turnaround time to a definitive result. This practice would also increase efficiency in the clinical laboratory.

Molecular identification of Malassezia from the skin scrapings of clinically suspected cases of pityriasis versicolor.

Pityriasis versicolor (PV) is caused by a lipophilic fungus belonging to the genus Malassezia. Potassium hydroxide (KOH) mount is most frequently used for screening of cases and culture is the gold standard. KOH lacks sensitivity, and culture is time-consuming and technically demanding. A cross-sectional study was conducted at a tertiary care teaching institution. We aimed to use multiplex-PCR for faster and accurate identification of Malassezia spp directly from skin scrapings of suspected cases of PV. The study was conducted on suspected cases of PV over a period of 12 months. The clinical and demographic details were recorded. The skin scrapings were subjected to KOH mount and cultured on Sabouraud's dextrose agar with an olive oil overlay. Multiplex-PCR targeting 11 Malassezia spp was performed on DNA extracted from skin scrapings. A total of 69 suspected cases of PV were studied. Most patients belonged to metro cities and worked in hot and humid climates. The mean duration of lesions was 18 months, and most had macular and patchy lesions. The sensitivity of KOH and culture was found to be 82.6% and 91.3%, respectively. M. globosa (n = 60, 87%) and M. restricta (n = 3, 4.3%) were isolated in culture. Multiplex PCR detected 85.5% of M. globosa, 5.8% of M. restricta, and 8.7% of mixed infection with M. globosa and M. restricta. M-PCR detected Malassezia in all the samples. M-PCR could identify Malassezia species directly from skin specimens, eliminating the need for culture. M-PCR was accurate, dependable, and exhibited a rapid turnaround time.

Integrating loop-mediated isothermal amplification with lateral flow assay to achieve a highly sensitive method for detecting Streptococcus suis Genome in raw pork

Streptococcus suis (S.suis), a zoonotic foodborne pathogen prevalent in Southeast Asia, poses a substantial threat to human and animal health because of its ability to cause severe and life-threatening illnesses. To address this challenge, a rapid and highly sensitive detection platform for S. suis in raw pork was developed by integrating loop-mediated isothermal amplification (LAMP) and a lateral flow assay (LFA), S. suis LAMP-LFA. LAMP reactions targeting the S. suis glutamate dehydrogenase (gdh) gene were optimized for specific detection of S. suis within 45 min at an isothermal temperature of 65 °C. The assay exhibited marked sensitivity, with a detection limit of 100 fg for genomic DNA extracted from S. suis cultures. Notably, this method showed no cross-reactivity with other bacterial contaminants commonly found in raw pork. The resulting LAMP amplicons were effectively detected using LFA, with a test limit of 101 CFU per 25 g of raw pork. S. suis LAMP-LFA proved to be highly specific and reliable, with no false-positives detected in spiked pork samples or pork samples containing other bacterial contaminants. Due to its high sensitivity, specificity, and rapid turnaround time, the proposed technique has immense potential as a field-deployable screening test for S. suis detection in raw pork, contributing to enhanced food safety and public health protection.

Correlation between the prothrombin time values obtained by point-of care prothrombin time meter and clinical laboratory in paediatric intensive care unit of a developing country

Background: Monitoring of prothrombin time (PT) is essential for adequate management of children in pediatric intensive care unit (PICU) who require serial and repeated PT measurements. In contrast to laboratory testing, point-of-care (POC) PT meters have been configured especially for bedside that are cost effective with rapid turnaround time. Our study aimed to assess its feasibility for routine use in PICU by assessing the correlation between PT values obtained through the POC PT meter and those acquired from the laboratory. Methods: A total of 200 children of either gender aged between 1 month to 12 years, admitted to PICU of Indus hospital Karachi Pakistan from February to August 2023 were analysed in this cross-sectional study. Venous blood samples were collected in PT bottles and transported to the pathology department within a time frame of 20-30 minutes for analysis. Concurrently, a drop of blood was obtained via finger prick and promptly assessed using the POC device-Coaguchek INRange system, (Roche diagnostics Ltd, USA). Pearson’s correlation was applied. Results: 200 children (96 males, 104 females) having a mean age of 6.42±3.55 years. The mean PT values with POC meter and Laboratory were 2.588±1.299 and 2.574±1.304 respectively (p=0.100). Very strong correlation of PT between POC PT meter and laboratory evaluation was observed (r=0.996, p&lt;0.001). Conclusions: The strong correlation observed between POC PT meter and laboratory supports the use of these devices for timely coagulation monitoring, potentially improving patient outcomes in PICUs.

Performance of three multiplex real-time PCR assays for simultaneous detection of 12 infectious pathogens in mice affected with respiratory and digestive diseases.

Research quality can be improved with reliable and reproducible experimental results when animal experiments are conducted using laboratory animals with guaranteed microbiological and genetic quality through health monitoring. Therefore, health monitoring requires the rapid and accurate diagnosis of infectious diseases in laboratory animals. This study presents a performance evaluation of a commercially available multiplex real-time PCR (mRT-PCR) assay for the rapid detection of 12 infectious pathogens (Set 1: Sendai virus [SeV, formally murine respirovirus], Mycoplasma spp., Rodentibacter pneumotropicus, and Rodentibacter heylii; Set 2: Helicobacter spp., Murine norovirus [MNV], Murine hepatitis virus [MHV], and Salmonella spp.; Set 3: Staphylococcus aureus, Streptobacillus moniliformis, Corynebacterium kutscheri, and Pseudomonas aeruginosa). To evaluate the efficacy of the mRT-PCR assay, 102 clinical samples encompassing fecal and cecal specimens were analyzed. The resulting data were then compared with the findings from sequence analysis for validation. The assay's detection limit ranged from 1 to 100 copies per reaction. Specificity testing involving various viruses and bacteria indicated no cross-reactivity between strains. Additionally, the assay exhibited good reproducibility, with mean coefficients of variation for inter- and intra assay variation below 3%. The overall positive rate was 52.9% (n = 54), with the mRT-PCR assay findings matching sequence analysis results (κ = 1). MHV (n = 29, 28.4%) was the most prevalent pathogen, followed by Helicobacter spp. (n = 28, 27.5%), R. heylii (n = 18, 17.6%), Mycoplasma spp. (n = 14, 13.7%), MNV (n = 12, 11.8%), S. aureus (n = 9, 8.8%), P. aeruginosa (n = 4, 3.9%), and R. pneumotropicus (n = 1, 0.9%). This assay offers a rapid turnaround time of 100 min, including 30 min for DNA preparation and 70 min for target DNA/RNA amplification. It ensures accuracy, minimizing false positives or negatives, making it a convenient tool for the simultaneous detection of infectious diseases in many samples. Overall, the propose‑d assay holds promise for the effective detection of the most important pathogens in laboratory animal health monitoring.

FPGA-assisted Design Space Exploration of Parameterized AI Accelerators: A Quickloop Approach

FPGAs facilitate prototyping and debug, and recently accelerate full-stack simulations due to their rapid turnaround time (TAT). However, this TAT is restrictive in exhaustive design space explorations of parameterized RTL generators, especially DNN accelerators that unleash an explosive full-stack search space. This paper presents Quickloop, an efficient and scalable framework to enable FPGA-accelerated exploration. Quickloop first abstracts away the cumbersome flow of RTL generation, software stack, FPGA toolflow, workload execution and metrics extraction by wrapping these stages into isolated Quicksteps, featuring cascadability, scalability, and replay. Then, we analytically minimize the FPGA toolflow TAT via a novel, data-driven strategy that intelligently utilizes build fragments from previous iterations, enhancing the loop efficiency and simultaneously lowering the toolflow’s compute utilization.Quickloop is built around the OpenAI Gym environment framework and thus supports drop-in regression and reinforcement learning explorations. With a Quickloop around a reference Berkeley’s Gemmini DNN accelerator, we exhaustively explore its parameter space and discover complex performance patterns, based on full-stack simulation of Imagenet benchmarks as a workload. Compared to conventional FPGA toolflow, we further show that Quickloop effectively reduces episodal time by above 30%, as the episode approaches realistic lengths.

Early identification of the nosocomial spread of vancomycin-resistant Enterococcus faecium by Fourier-transform infrared spectroscopy and performance comparison with PFGE and WGS

ABSTRACT Early detection of disseminating vancomycin-resistant Enterococcus faecium (VREfm) in ICU wards is crucial for outbreak identification and the implementation of prompt infection control measures. Genotypic methods like pulsed-field gel electrophoresis (PFGE) and whole-genome sequencing (WGS) are costly and time-consuming, hindering rapid response due to batch dependency. Fourier-transform infrared spectroscopy (FT-IR) offers the potential for real-time outbreak detection and reliable strain typing. We utilized FT-IR to identify clonal VREfm dissemination and compared its performance to PFGE and WGS. Between February through October 2023, an unusually high number of VREfm were recovered at a tertiary hospital in Barcelona. Isolates were examined for antimicrobial susceptibility, carriage of vanA/vanB genes and clonality was also studied using FT-IR, PFGE, and WGS. Routine FT-IR inspections revealed recurring VREfm clustering during the outbreak's initial weeks. In total, 104 isolates were recovered from 75 patients and from multiple wards. However, only one isolate was recovered from an environmental sample, suggesting the absence of environmental reservoirs. An ST80 vancomycin-resistant (vanA) E. faecium strain was the main strain responsible for the outbreak, although a few additional VREfm strains were also identified, all belonging to CC17. PFGE and cgMLST (WGS) yielded identical clustering results to FT-IR, and WGS confirmed vanA/vanB gene carriage in all VREfm isolates. Infection control measures led to a rapid decline in VREfm isolates, with no isolates detected in November. FT-IR spectroscopy offers rapid turnaround times, sensitivity, and reproducibility, comparable to standard typing methods. It proved as an effective tool for monitoring VREfm dissemination and early outbreak detection.

Cross-sectional evaluation of online direct-to-public calprotectin testing

Objective Why about a quarter of patients with inflammatory bowel disease (IBD) suffer symptoms for more than a year before their diagnosis made is unclear. Low public awareness, embarrassment and...