bluuebunny/crossref_metadata_2025_split huggingface dataset:
B-274 Determining Novel Biomarker Candidates Associated With Heroin and Cocaine Exposure Using Urine Metabolomics
Authors: Tamama K , Patel S , Vanderschelden R A 
Published: October 2024

Abstract

Background
Substance abuse poses an escalating health risk in the United States. While acute drug exposure can be tracked using drugs and their metabolites as biomarkers in clinical settings, there is a notable absence of biomarkers for chronic drug exposure or addiction. The development and availability of such biomarkers could significantly enhance clinicians’ ability to identify patients at high risk of addiction, and more accurately predict disease progression and treatment responses.

Methods
Our study aimed to discover novel biomarkers associated with substance abuse by analyzing raw mass spectrometry (MS) data from urine comprehensive drug screening (UCDS) conducted at the University of Pittsburgh Medical Center (UPMC). The MS data, acquired through untargeted data collection using LC-qToF-MS at the UPMC Clinical Toxicology Laboratory, was initially preprocessed (including peak identification, alignment, and putative annotation) using MS-DIAL to generate a data matrix. This matrix, consisting of objects (cases) and features (m/z and retention time), was further normalized and analyzed using MetaboAnalyst. A combination of statistical methods, including point biserial correlation analysis, volcano plot, significance analysis of microarrays and metabolites (SAM), empirical Bayesian analysis of microarrays and metabolites (EBAM), partial least squares discriminant analysis (PLS-DA), and random forest (RF), were employed to select features significantly associated with the EIA-results. These features were subsequently evaluated for analyte identification using MS-FINDER.

Results
Our analysis identified nine significant features associated with 6MAM-EIA for heroin abuse, including 6-monoacetylmorphine itself and norfentanyl. For features associated with cocaine metabolite-EIA, 37 significant features were selected, including multiple cocaine metabolites, nicotine metabolites, and norfentanyl. Some of these selected features are likely products of in-source fragmentation or endogenous metabolites.

Conclusions
Our study has identified a set of potential biomarker candidates for illicit drug exposure. Notably, norfentanyl was found to be significantly associated with both 6MAM-EIA and cocaine metabolite-EIA, reflecting current trends in illicit drug use. Further chemical identification of these biomarkers is planned for future work.
B-275 Trends of urine drug testing findings in a pediatric hospital from 2019 to 2023
Authors: Narasimhan M , Jones P M , Poling A , Jones V , Fisher S , Forssell D , Duong T , Muthukumar A , Hashim I , Cao J 
Published: October 2024

Abstract

Background
Semi-quantitative immunoassays (IA) are commonly used as the initial urine drug screen (UDS), with positive results confirmed by additional testing. However, substantial false negative (FN) rates have been reported in the classes of opioids and amphetamines. In our pediatric hospital, a semi-quantitative comprehensive urine drug testing by liquid chromatography quadrupole time-of-flight mass spectrometry (LC-QTOF) MS (referred as QTOF) is offered as either a direct order or a reflexive order following positive IA UDS. The lab also reflexes to run an IA UDS with every direct QTOF order. This study presents paired urine drug IA and QTOF results in the classes of opiate, synthetic opioid, and amphetamine/methamphetamine between 2019 and 2023 to understand trends and to evaluate assay accuracy.

Methods
Paired results from IA (Siemens Atellica) and QTOF (Sciex 4600) were analyzed with reporting cutoff concentrations from representative drugs in each class listed in Figure 1. The opiate class reported on QTOF includes 6-monoacetylmorphine, morphine, hydrocodone, codeine and metabolites. The synthetic opioid class includes buprenorphine, fentanyl, levorphanol, methadone, naloxone, oxycodone, tramadol and metabolites. Simultaneous findings of opiate and synthetic opioid were counted as the opiate class. FN rate of IA was calculated using QTOF as the reference.

Results
As shown in Figure 1, QTOF testing nearly doubled from 2019 to 2023, with an expected high FN rate in the synthetic opioid class and alarmingly rising FN rates in opiate and amphetamine classes. Within the synthetic opioid class, percentage of positive fentanyl results increased from 37.6% to 59.0% in 5 years, and ∼10% are only positive for norfentanyl, which is detected in very few commercial fentanyl IAs.

Conclusions
Our study clearly demonstrates the gap in pediatric toxicology investigation, reporting high FN rates in common drug classes by urine IA, calling for more comprehensive and sensitive UDS to be broadly adopted.
B-278 Evaluation of serum copper, zinc and selenium reference intervals using The National Health and Nutrition Examination Survey (NHANES) data
Authors: Boyd J , Yu L , Burkhead J 
Published: October 2024

Abstract

Background
Serum copper, zinc and selenium, are commonly ordered for assessment of nutrient status or investigation of deficiency or overload. Review of reference intervals for these analytes is challenging as most laboratories do not have the resources to do a de novo reference interval study and many literature references are not recent. The National Health and Nutrition Examination Survey (NHANES) survey presents a unique opportunity to access a comprehensive database that includes biochemical parameters and health information in order to review reference intervals regularly, especially for more esoteric tests such as trace elements. In this study, copper, zinc and selenium reference intervals were calculated using NHANES data.

Methods
NHANES data for individuals 18 years of age and older was obtained from three releases between 2012-2016. Copper, selenium and zinc 95% confidence intervals were evaluated by age, sex and body mass index (BMI) partitions using quantile regression. All calculations were performed in R using the quantreg package.

Results
The calculated reference intervals for serum copper, zinc and selenium are shown in Table 1. Serum copper had the largest changes observed between sexes, with females having a much wider reference interval on the high end. Oral contraceptive use is known to increase serum copper levels which likely explains this trend. BMI over 30 also increased the serum copper reference interval, more clearly in women than in men. Zinc reference intervals were more aligned between women and men, with slightly higher intervals observed in men. BMI did not have much effect in either sex. For selenium, men had slightly higher reference intervals than women.

Conclusions
NHANES data provides a comprehensive data set for reference interval evaluation, especially for more esoteric analytes such as trace elements.
B-292 Detection of natural toxins derived from potatoes by biotin-streptavidin direct ELISA
Authors: Okada K 
Published: October 2024

Abstract

Background
α-solanine (SO) and α-chaconine (CHA) are natural toxins of potato and often cause food poisoning worldwide. We have previously developed a polyclonal antibody (anti-Sold antibody) that bind to SO and CHA and used it in an enzyme-linked immunosorbent assay (ELISA). Our previously constructed ELISA (conventional ELISA) has a weak point of low sensitivity in detecting SO and CHA, we have tried to improve it in this study.

Methods
Solanidine, a chemical compound found in both SO and CHA, was conjugated with bovine albumin, and this complex was repeatedly injected into two rabbits for two months. Anti-Sold antibody was then purified from rabbit serum using affinity column chromatography. In our ELISA method, named “B-S direct ELISA”, biotin-labeled anti-Sold antibody is bound to SO and CHA in the sample, followed by the binding of peroxidase-labeled streptavidin. To simulate biological samples from patients with potato food poisoning, SO and CHA powders were mixed with commercially available human serum and urine, and SO and CHA levels were then measured using ELISA. Lastly, SO and CHA levels were detected in extracts from rotten potato (Irish Cobbler) sprouts, peels, and tubers.

Results
The absorbance of the samples containing SO and CHA prepared in 10 mM phosphate buffer saline (PBS) was about 5-fold higher than that of the conventional ELISA. Furthermore, the absorbance of the samples containing SO and CHA prepared in human serum and urine was enhanced about 2.5- and 1.6-fold, respectively, compared to that of the conventional ELISA. The limit of detection in B-S direct ELISA was 0.32 ng/mL in 10 mM PBS, 2.08 ng/mL in serum, and 4.88 ng/mL in urine, respectively. Using this ELISA, it was also possible to measure the content of SO and CHA in extracts of potato tubers, peels, and sprouts. Both SO and CHA in the extracts from rotten potato tubers and peels were detected as approximately 1.4- and 2.2-fold higher, respectively, in each part than those in the extracts from fresh potatoes. Furthermore, the potato sprout extract contained approximately 50-fold more SO and CHA than the tuber and 9-fold more than the peel. On the other hand, B-S direct ELISA cross-reacted with solanidine and solasodine, which have similar chemical structures to SO and CHA, and also with cholesterol and vitamin D3 in serum.

Conclusions
Based on our results, B-S direct ELISA presented greater performance in detecting SO and CHA than our conventional ELISA. The non-specific reactions in the serum samples and influence of urea in the urine samples may underlie the reduced detection performance of SO and CHA in ELISA, respectively. Although this ELISA has a detection performance applicable to human biological samples, cross-reactivity should be fully considered when judging the results.
B-294 Case report: Postmortem ethanol production following traumatic injury
Authors: Whitt A , Alquran B , Jortani S A 
Published: October 2024

Abstract

Background
Ethanol is the most commonly encountered psychoactive substance in forensic toxicology. Accurate interpretation of ethanol concentrations from postmortem samples is fraught with potential hurdles, including decomposition, redistribution, and postmortem production of ethanol. Through the fermentation of substrates like glucose, pyruvate, lactate, and amino acids, microorganisms present in the gut and/or environment can generate ethanol in contaminated human tissues, especially following traumatic injury. The degree of fermentation varies widely depending on the time between death and autopsy, temperature, humidity, tissue glucose concentrations, and the specimen type. Vitreous humor (VH) is the preferred specimen for postmortem toxicological analysis of ethanol because it accurately reflects the antemortem ethanol concentration in serum while being resistant to microbial contamination. This case highlights the utility of VH in an extreme instance of postmortem ethanol production.

Methods
Postmortem toxicological analysis of liver tissue and VH (NMS Labs, Horsham, PA) was performed by headspace-gas chromatography (HS-GC)

Results
Case presentation: A man in his sixties with a history of type 1 diabetes mellitus and hypertension was involved in a fatal mining accident while operating heavy machinery in which the equipment fell down a cliff, causing the man to be ejected onto the ground nearby. The decedent was discovered hours after the fall with extensive traumatic injuries to the left side of his body and head. Seven days later, an autopsy was performed by the medical examiner’s office. Postmortem toxicological analysis of liver tissue by HS-GC revealed ethanol at a concentration of 690 mg/100g. The liver tissue also tested positive for amlodipine, nicotine, cotinine, and caffeine. A vitreous humor (VH) sample tested negative (&lt;10 mg/dL) for ethanol.

Conclusions
The toxicological findings reported herein are inconsistent with antemortem alcohol consumption and are instead an example of severe postmortem ethanol production by microbial contamination. Several factors likely led to the stark discrepancy between the concentrations of ethanol in VH and liver tissue. First, the decedent suffered traumatic injuries, creating conditions favorable for extensive microbial contamination. Second, the time between death and autopsy was seven days, allowing ample time for fermentation to occur, even under refrigerated conditions. Third, postmortem glycogenolysis in addition to the patient’s underlying T1DM likely provided a high concentration of glucose to fuel postmortem fermentation. Ultimately, this case underscores the importance of forensic specimen selection in accurately interpreting toxicology findings following traumatic injury.
B-303 Stability of Antipsychotic Drugs in Patient Serum and Plasma Clinical Samples
Authors: Konrath M S , Drisgill A , Kozo D , Hilaire M , Salamone S J 
Published: October 2024

Abstract

Background
Therapeutic drug monitoring (TDM) of antipsychotic blood levels is strongly recommended (clozapine and olanzapine) and recommended (risperidone, aripiprazole, and quetiapine) by ASCP/AGNP Consensus Statement. For effective TDM, it is paramount to understand both the stability of the antipsychotic drugs in serum and plasma, as well as any differences in levels due to sample matrix to ensure robust concentration determinations. While clozapine levels are considered equivalent in serum and plasma, we included in our study four additional antipsychotics.

Methods
Whole blood samples from ten patients taking clozapine, risperidone, aripiprazole, olanzapine, or quetiapine were collected in one red top tube and in one purple top tube according to an IRB-approved protocol. Samples were then processed within 2 hours of collection to obtain matched serum and K2EDTA plasma samples from each of the patients. CE marked immunoassays for the measurement of clozapine (also FDA cleared), total risperidone/paliperidone, total aripiprazole, olanzapine, and quetiapine were used to quantitate drug concentrations. Stability of the samples was assessed at 4°C, ambient room temperature (ART), and frozen at -80°C, with respect to their Day 0 concentration by measurement of six replicates at each timepoint using the immunoassays.

Results
Comparison of Day 0 values between serum and plasma for the same patient showed ±9% bias between sample matrices for all drugs, except olanzapine which showed differences up to ±15%. Average bias between serum and plasma was ≤-5% (olanzapine: -9%). All drugs were stable (±15% deviation from Day 0; olanzapine up to ±23%) in serum and in plasma for 7 days at both 4°C and ART and for up to 8 months frozen.

Conclusions
This study demonstrated that (1) the antipsychotic drugs clozapine, risperidone, aripiprazole, olanzapine, and quetiapine have sufficient stability in both serum and plasma for effective TDM and (2) for TDM, serum or plasma may be used.
B-349 Diagnostic Performance of the Atellica IM High-Sensitivity PSA Assay in Men Undergoing Prostate Cancer Screening
Authors: Olson Nissen K , Snyder J , Pagano M , Owusu A , Grosett M , Birmingham N , Christenson R , deFilippi C 
Published: October 2024

Abstract

Background
This study evaluated the diagnostic performance of the Atellica® IM High-Sensitivity PSA (hsPSA)* assay in men &gt;50 years undergoing prostate cancer (PC) screening in conjunction with digital rectal exam (DRE).

Methods
Serum samples were collected prospectively from 2671 subjects &gt;50 years referred to a urologist for evaluation of PC from 26 sites across the United States. To establish PC versus benign disease, all subjects underwent a transrectal prostate biopsy ≤60 days after blood draw. At the cutoff of 4.0 ng/mL for the hsPSA assay, the percentage of PCs detected (sensitivity) and corresponding percentage of biopsies avoided in men without PC (specificity) were determined.

Results
Cancer prevalence (malignant biopsy) was 58.5% (1562/2671) among the diagnostic population. Preliminary results demonstrated that for DRE alone, sensitivity and specificity were 30.9% (482/1562) and 83.0% (920/1109) compared to biopsy. For hsPSA alone at the cutoff of 4.0 ng/mL, sensitivity was 88.7% (1386/1562). The table provides incidence of cancer and positive biopsies by subgroups defined by DRE and hsPSA. hsPSA detected 61.1% (955/1562) of PCs that DRE did not. Of those DRE+, 71.8% (482/671) had positive biopsy. Of those DRE+ and hsPSA ≥4.0 ng/mL, 78.6% (431/548) had positive biopsy, which significantly increased PC detection by 7.0% (95% CI:1.4%-12.5%). Similarly, in the DRE- population, while 54.0% of subjects had positive biopsy, subjects both DRE- and hsPSA &lt;4.0 ng/mL had only 38.5% positive biopsies, a significant difference of 16.5% (95% CI: 6.3%-27.8%).

Conclusions
Using DRE alone detected only 3 out of 10 patients with PC. It was observed that when used in conjunction with DRE, the hsPSA assay improved both sensitivity and specificity of PC detection over DRE alone.
CAUTION: Investigational Device. Not available for sale. Future availability cannot be guaranteed. The results are preliminary and were achieved in unique setting with no expected timeline for completion.
B-352 Effects of COVID-19 on Breast Cancer Positivity: A Retrospective and Predictive Study
Authors: de Andrade S S , Moreno Rodrigues de Souza G , Pagan Cilotti H , Ribeiro Ramadan-Boscolo D , Tufik S 
Published: October 2024

Abstract

Background
The COVID-19 pandemic has had a significant impact on the healthcare system, resulting in overall reduction in access to healthcare services, including the suspension of screening programs and postponement of surgeries and medical treatments. The aim of this study was to evaluate the effects of the pandemic on breast cancer positivity from cases received at a laboratory in São Paulo, Brazil.

Methods
A retrospective analysis of a database from a pathology laboratory in São Paulo, Brazil, was performed during the 2017-2023 period and classified as following: pre-pandemic (January 2017-February 2020), pandemic (March 2020 - April-2022), and post-pandemic (May 2022-December 2023). Cases were classified as negative or positive for malignancies. Positive anatomopathological examinations (breast biopsies and surgical specimens) considered primary, metastatic mesenchymal or endothelial malignancies. The Mann-Kendall test was used to evaluate the trend, and the PACF (Partial Autocorrelation Function) correlogram was used to analyze historical memory for definition of the autoregressive model. The augmented Dickey-Fuller hypothesis test was applied to analyze whether the series is stationary or not, and the first differences test was applied. The ARIMA model was used for the estimates (1,1,1). Statistics were performed using the gretl software and pymannkendall library in the Python language.

Results
A total of 74,973 samples were evaluated during the 2017-2023 period. In the three-year pre-pandemic period, 32,644 tests were recorded, with an average of 29.34% positive cases. From the analysis of the pre-pandemic time series, there was a positive trend towards an increase in cases (p &lt; 0.0001, z=6.1360, Tau=0.6955). In a non-pandemic scenario, from March 2020 to February 2021, the number of new tests expected would be 11,535; however, in the pandemic period, only 8,904 were observed, a 22.81% reduction on expected cases. According to the historical average positivity rate in the pre-pandemic period, the expected and observed total were, respectively, 3,384 and 2,566 positive cases for malignancy.

Conclusions
This study reveals a significant decrease of approximately 22.81% in new breast anatomopathological diagnoses during the COVID-19 pandemic in a laboratory in São Paulo, Brazil. For comparison purposes, we identified a 24.17% of missed opportunities on diagnoses of new malignancy-positive cases, compared to the pre-pandemic period. These findings highlight the need for strategies to mitigate the negative effects of the pandemic on the early detection and treatment of breast cancer.
Two leucine-rich repeat receptor-like kinases initiate herbivory defense responses in tea plants
Authors: Jiang Qi , Ding Changqing , Feng Lingjia , Wu Zhenwei , Liu Yujie , He Lintong , Liu Chuande , Wang Lu , Zeng Jianming , Huang Jianyan , Ye Meng 
Published: October 2024

Abstract
Leucine-rich repeat receptor-like kinases (LRR-RLKs) have emerged as key regulators of herbivory perception and subsequent defense initiation. While their functions in grass plants have been gradually elucidated, the roles of herbivory-related LRR-RLKs in woody plants remain largely unknown. In this study, we mined the genomic and transcriptomic data of tea plants (Camellia sinensis) and identified a total of 307 CsLRR-RLK members. Phylogenetic analysis grouped these CsLRR-RLKs into 14 subgroups along with their Arabidopsis homologs. Gene structure and conserved domain analyses revealed notable similarities among subgroup members. Among the identified CsLRR-RLKs, we focused on two plasma membrane-localized LRR-RLKs, CsLRR-RLK44, and CsLRR-RLK239, which do not form homodimers or heterodimers with each other. Both respond strongly to herbivory, and their expression patterns significantly correlate with herbivore resistance phenotypes across different tea accessions. CsLRR-RLK44 and CsLRR-RLK239 act upstream of mitogen-activated protein kinase (MPK) cascades and modulate the expression of defense-related MPKs and WRKY transcription factors. Additionally, silencing CsLRR-RLK44 or CsLRR-RLK239 reduced the levels of herbivory-induced jasmonates, thereby weakening the plant resistance to tea geometrid larvae (Ectropis obliqua). Our work is the first to demonstrate that in woody plants, LRR-RLKs are essential for enhancing herbivore resistance through the activation of the canonical signaling, including MPKs, WRKYs, and jasmonates. Furthermore, our study extends mechanistic insights into how LRR-RLKs initiate plant defenses from grasses to economically important tree species.
SlSTOP1-regulated SlHAK5 expression confers Al tolerance in tomato by facilitating citrate secretion from roots
Authors: Zhu Huihui , Chen Weiwei , Yang Zheng’an , Zeng Congfang , Fan Wei , Yang Jianli 
Published: October 2024

Abstract
SENSITIVE TO PROTON RHIZOTOXICITY 1 (STOP1) is a core transcription factor that regulates the expression of aluminum (Al) resistance genes to manage Al toxicity in plants. However, the genome-wide roles of SlSTOP1 in the Al stress response of tomato (Solanum lycopersicum) remain largely unknown. Here, we report that SlSTOP1 is crucial for Al tolerance in tomato, as loss-of-function mutants of SlSTOP1 displayed hypersensitivity to Al stress. Aluminum stress had no effect on SlSTOP1 mRNA expression, but promoted accumulation of SlSTOP1 protein in the nucleus. Through integrated DNA affinity purification sequencing and RNA sequencing analysis, we identified 39 SlSTOP1-targeted Al-responsive genes, some of which are homologous to known Al resistance genes in other plant species, suggesting that these SlSTOP1-targeted genes play essential roles in Al resistance in tomato. Furthermore, using peak enrichment analysis of SlSTOP1-targeted sequences, we identified a cis-acting element bound by SlSTOP1 and validated this finding via dual-luciferase reporter and electrophoretic mobility shift assay (EMSA). Additionally, we demonstrated SlHAK5 is one of direct targets of SlSTOP1 and functionally characterized it in terms of Al stress tolerance. Compared with wild-type plants, Slhak5 mutants developed by CRISPR/Cas9 technology presented increased sensitivity to Al stress, which was associated with reduced citrate secretion from the roots. Together, our findings demonstrate that SlSTOP1 directly interacts with cis-acting elements located in the promoters of target genes involved in diverse pathways contributing to Al resistance in tomato.