Commercial Testing Laboratories Research Articles

The Contribution of Legionella anisa to Legionella Contamination of Water in the Built Environment.

Legionella bacteria can proliferate in poorly maintained water systems, posing risks to users. All Legionella species are potentially pathogenic, but Legionella pneumophila (L. pneumophila) is usually the primary focus of testing. However, Legionella anisa (L. anisa) also colonizes water distribution systems, is frequently found with L. pneumophila, and could be a good indicator for increased risk of nosocomial infection. Anonymized data from three commercial Legionella testing laboratories afforded an analysis of 565,750 water samples. The data covered July 2019 to August 2021, including the COVID-19 pandemic. The results confirmed that L. anisa commonly colonizes water distribution systems, being the most frequently identified non-L. pneumophila species. The proportions of L. anisa and L. pneumophila generally remained similar, but increases in L. pneumophila during COVID-19 lockdown suggest static water supplies might favor its growth. Disinfection of hospital water systems was effective, but re-colonization did occur, appearing to favor L. pneumophila; however, L. anisa colony numbers also increased as a proportion of the total. While L. pneumophila remains the main species of concern as a risk to human health, L. anisa's role should not be underestimated, either as a potential infection risk or as an indicator of the need to intervene to control Legionella's colonization of water supplies.

Comparison of Water and DTPA Extractants for Testing of Greenhouse Soilless Substrates

ABSTRACT The main purpose of this study was to improve the accuracy and reliability of nutrient analysis on soilless substrates by comparing two different saturated medium extract (SME) methods, using either water (SMEW) or DTPA (diethylenetriamine-pentaacetic acid, SMED) as extractants. A series of analyses were performed to compare ion levels quantified from either SMEW or SMED. Substrate pH, electrical conductivity (EC) and nutrient data were analyzed from a survey of substrate components (coconut coir, peat, bark, perlite, vermiculite), and blended greenhouse propagation substrates using the SMEW and SMED methods. In addition, a series of SMEW and SMED extracted samples were sent to three commercial soil testing laboratories for nutrient analysis. The objectives of these analyses were to (1) validate that published relationships between SMEW and SMED methods were consistent with correlation curves for pH, EC and nutrient levels across a wide range of soilless substrates, (2) compare statistical variability of results from SMEW and SMED methods and (3) compare variability in results between different laboratories. The results validated that previously published standards for interpreting SMEW extractions can be used for SMED method for macronutrients and EC due to an approximately 1:1 relationship for test results between extraction methods. Up to 2 orders of magnitude higher levels of micronutrients were extracted for iron (Fe), manganese (Mn), zinc (Zn), and copper (Cu) using SMED compared with SMEW, with low correlation between extractants. Concentrations of ions from SMED extractions were less variable than SMEW for micronutrient analysis but were comparable for macronutrient analysis. Results from this study were combined with published research to provide suggested ranges using SMED for soilless substrates.

Familial Polycythemia Associated with JAK2 Mutation Previously Known As Variant of Unknown Significance

Introduction Mutations in the Janus Kinase 2 gene (JAK2) are commonly encountered in patients with myeloproliferative neoplasms (MPN) including polycythemia vera (PV) and essential thrombocythemia. JAK2 mutations particularly V617F is associated with PV in 98% cases. Gain of function mutations in the JAK2 exon 12 are reported to be associated with isolated erythropoiesis. Atypical mutations in JAK2 including (c.3323A>G, p.N1108S) were reported in patients with MPN who tested negative for the established driver mutations including exon 14 JAK 2, exon 9 CALR and exon 10 MPL genes. Primary familial congenital polycythemia (PFCP) is a rare disease inherited in an autosomal dominant fashion caused by hypersensitivity of EPO receptor. In such cases, an EPO receptor mutation is only found in 12-15% of affected patients. Due to associated increased risk of thrombosis, bleeding, leukemic involvement and myelofibrosis in PV, bone marrow evaluation is essential in this patient population. In patients with polycythemia, the goals of treatment are to maintain appropriate level of hemoglobin/hematocrit and to reduce severity of symptoms. We present a family affected by polycythemia and leukocytosis associated with an atypical heterozygous variant of JAK2 [c.3323A>G (p.Asn1108Ser)] currently reported as benign or variant of unknown significance (VUS) in literature. Methods Case series, retrospective electronic medical record review with institutional board review exemption Results We report a case series of three Caucasian family members affected with JAK2 VUS mutation (mother and two sons). The proband is a 17-year-old male who presented to an outside facility at the age of 14 with complaints of fatigue, headache, and intense pruritis after hot showers. Initial workup was significant for marked erythrocytosis and reticulocytosis without evidence of abnormal cells on peripheral film. Bone marrow evaluation showed trilineage hematopoiesis with no morphological features of a myeloproliferative neoplasm. Genetic testing was negative for JAK2 V617F and exon 12 mutations. However, whole exome sequencing (WES) revealed a heterozygous mutation of JAK2 gene, c.3323A>G (p.Asn1108Ser). Additional investigation on proband's younger brother, who complained of similar symptoms, also revealed the same JAK2 variant. The proband's mother reported non-specific symptoms including headache, flushing, and occasional blurry vision that she previously attributed to hypertension. Her labs were significant for leukocytosis, monocytosis and thrombocytosis in the presence of normal red blood cell count and WES showed the same mutation. Bone marrow evaluation of the brother and the mother showed normal trilineage hematopoiesis and was negative for malignancy (Table 1). Proband's father is reportedly negative for the aforesaid mutation and is clinically asymptomatic. The proband and his brother are currently managed with phlebotomy every 3 to 4 weeks to maintain hematocrit below 45%, leading to improvement in symptoms. Cytoreductive therapies have been discussed but not initiated. Proband's mother is managed by an adult hematologist. Scheduled phlebotomies with close surveillance is recommended for clinical practice along with interval research sequencing. Conclusion Our case series reports a family with clinical features and laboratories suggestive of a familial form of polycythemia, WES detected the presence of an atypical JAK2 mutation: c.3323A>G, (p.Asn1108Ser). The phenotypical and clinical significance of this mutation has been considered non canonical per reports and classified as likely benign by reputable commercial genetic testing laboratories. However, other reports identify this JAK2 variant as having a significant increased prevalence in patients with MPNs and AML compared to the general population. It may result in a gain of function of the JAK2 protein and subsequent development of MPNs. Enrollment in research sequencing protocols is encouraged in such patients to closely follow the pathologic potential and spectrum of genetic variants rarely reported in literature. Additional research to determine the pathogenicity of this genetic variant is highly warranted.

Calculating soil titratable acidity from routine soil analyses of cultivated South African soils using various model structures

Soil titratable acidity (TA) is determined through extraction with an acetate-buffered salt solution (potassium sulphate, K2SO4) and is routinely used to predict the soil lime requirement using the Eksteen method. Soil TA is not always reported by commercial soil testing laboratories, but rather exchangeable acidity (1M KCl) is determined. Therefore, a need exists to use other routinely determined soil properties in a model to derive TA. In this study, routine soil-analysis results from a commercial laboratory (n = 5 915 measurements) were used to generate various model structures to calculate a theoretical value of TA and to evaluate the accuracy thereof. Measured TA was found to be significantly correlated to the calculated TA using the Eksteen R-value, soil pH, organic carbon percentage, exchangeable Ca2+ and Mg2+, and a function of the interaction between pH and organic carbon. Soil TA could be calculated most accurately using these factors in multivariate adaptive regression splines model (r 2 = 0.69, mean absolute error = 0.16) at TA values of < 2 cmolc kg−1. However, the exponential model calculations remained stable over a larger range of actual TA values up to 4 cmolc kg−1. Given the ease of use and interpretability, it is recommended that an exponential function model is used to calculate TA.

Abstract P5-03-03: Variant Classification Discordance: A real-world experience of genetic test results in a community-based setting

Abstract BACKGROUND Accurate interpretation of hereditary cancer germline genetic variants is critical to ensuring appropriate care. Myriad Genetics has developed tools that are instrumental in the accurate classification of variants, including a previously described history-weighting algorithm (Pheno), mutation co-occurrence statistical analysis (MCO), and in trans haplotype analysis. Differences in classification are known to occur among commercial testing laboratories, however the rate at which a single provider may observe variants with a different classification has not been reported. Here, we compared genetic test results from multiple laboratories ordered by a single surgical community-based practice with the classifications from Myriad’s testing laboratory. METHODS Variants initially reported as a “variant of uncertain significance” (VUS) on hereditary cancer test results from multiple commercial laboratories ordered by a single surgeon at a community-based, comprehensive breast center from June 2013 to May 2021 were evaluated and compared to the classifications from Myriad. In total, 212 variants were submitted for comparison. After review, 42 variants were excluded because they had not been observed previously in Myriad-tested patients. Therefore, 170 variants were eligible for comparison. Variants were classified as pathogenic/likely pathogenic, VUS, and benign/likely benign for comparison. Descriptive statistics were used for analysis. RESULTS Discordant classification was observed between Myriad and other testing laboratories for 28.2% (48/170) of the variants compared. Initially, all 170 variants were classified and reported by other testing laboratories as VUS, however, 15 variants (8.8%) were subsequently reclassified by other testing laboratories (N=13 were reclassified as benign/likely benign; N=2 were reclassified as pathogenic/likely pathogenic). Among all variants compared, 23.5% (40/170) were definitively classified by Myriad as benign/likely benign. For 90.0% (36/40) of these variants, evidence driving the classification relied upon Pheno (32 variants), MCO (4 variants), and in trans haplotype analysis (8 variants), with some variants having multiple lines of evidence. Other in-house specific rules were used for the remaining classifications (4/40;10%). Fifty-five percent (22/40) of the discordant classifications were seen in high-risk genes including APC, BRCA1/2, MLH1, MSH2, MSH6, PMS2, PALB2 and STK11, and 45% (18/40) were in moderate-risk genes including ATM, BARD1, BRIP1, CHEK2, and RAD51C. Of the 13 variants reclassified by other testing laboratories as benign/likely benign, six were classified as VUS by Myriad. Notably, two variants classified by Myriad as VUS were reclassified by other laboratories as pathogenic/likely pathogenic. CONCLUSIONS These data indicate that even in a single practice, significant discordance in variant classification exists based on the chosen laboratory. Myriad definitively classified nearly one-quarter of variants classified as VUS by other laboratories, likely due to the use of Myriad’s laboratory-developed classification tools. The degree of discordance observed here reflects the need for continuous laboratory investment in variant classification tools and evaluation of genetic variants, enabling physicians and patients to receive accurate results to facilitate appropriate medical management decisions. Citation Format: Shelly Cummings, Erin Mundt, Ann Marie Miller, TinaMarie Bauman, Thomas Slavin, Robert Maganini. Variant Classification Discordance: A real-world experience of genetic test results in a community-based setting [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr P5-03-03.

Genetic Testing to Inform Epilepsy Treatment Management From an International Study of Clinical Practice

It is currently unknown how often and in which ways a genetic diagnosis given to a patient with epilepsy is associated with clinical management and outcomes. To evaluate how genetic diagnoses in patients with epilepsy are associated with clinical management and outcomes. This was a retrospective cross-sectional study of patients referred for multigene panel testing between March 18, 2016, and August 3, 2020, with outcomes reported between May and November 2020. The study setting included a commercial genetic testing laboratory and multicenter clinical practices. Patients with epilepsy, regardless of sociodemographic features, who received a pathogenic/likely pathogenic (P/LP) variant were included in the study. Case report forms were completed by all health care professionals. Genetic test results. Clinical management changes after a genetic diagnosis (ie, 1 P/LP variant in autosomal dominant and X-linked diseases; 2 P/LP variants in autosomal recessive diseases) and subsequent patient outcomes as reported by health care professionals on case report forms. Among 418 patients, median (IQR) age at the time of testing was 4 (1-10) years, with an age range of 0 to 52 years, and 53.8% (n = 225) were female individuals. The mean (SD) time from a genetic test order to case report form completion was 595 (368) days (range, 27-1673 days). A genetic diagnosis was associated with changes in clinical management for 208 patients (49.8%) and usually (81.7% of the time) within 3 months of receiving the result. The most common clinical management changes were the addition of a new medication (78 [21.7%]), the initiation of medication (51 [14.2%]), the referral of a patient to a specialist (48 [13.4%]), vigilance for subclinical or extraneurological disease features (46 [12.8%]), and the cessation of a medication (42 [11.7%]). Among 167 patients with follow-up clinical information available (mean [SD] time, 584 [365] days), 125 (74.9%) reported positive outcomes, 108 (64.7%) reported reduction or elimination of seizures, 37 (22.2%) had decreases in the severity of other clinical signs, and 11 (6.6%) had reduced medication adverse effects. A few patients reported worsening of outcomes, including a decline in their condition (20 [12.0%]), increased seizure frequency (6 [3.6%]), and adverse medication effects (3 [1.8%]). No clinical management changes were reported for 178 patients (42.6%). Results of this cross-sectional study suggest that genetic testing of individuals with epilepsy may be materially associated with clinical decision-making and improved patient outcomes.

Performance of Hair Testing for Cocaine Use-Comparison of Five Laboratories Using Blind Reference Specimens.

The purpose of this study was to compare results from five commercial hair testing laboratories conducting workplace drug testing with regard to bias, precision, selectivity and decontamination efficiency. Nine blind hair specimens, including cocaine-positive drug user specimens (some contaminated with methamphetamine) and negative specimens contaminated with cocaine, were submitted in up to five replicates to five different laboratories. All laboratories correctly identified cocaine in all specimens from drug users. For an undamaged hair specimen from a cocaine user, within-laboratory Coefficients of Variation (CVs) of 5-22% (median 8%) were reported, showing that it is possible to produce a homogenous proficiency testing sample from drug user hair. Larger CVs were reported for specimens composed of blended hair (up to 29%) and curly/damaged hair (19-67%). Quantitative results appeared to be method-dependent, and the reported cocaine concentrations varied up to 5-fold between the laboratories, making interlaboratory comparisons difficult. All laboratories reported at least one positive result in specimens contaminated with cocaine powder, followed by sweat and shampoo treatments. Benzoylecgonine, norcocaine, cocaethylene and hydroxylated cocaine metabolites were all detected in cocaine powder-contaminated specimens. This indicates that current industry standards for analyzing and reporting positive cocaine results are not completely effective at identifying external contamination. Metabolite ratios between meta- or para-hydroxy-cocaine and cocaine were 6- and 10-fold lower in contaminated specimens compared to those observed in cocaine user specimens, supporting their potential use in distinguishing samples positive due to contamination and drug use.

COL4A4 variant recently identified: lessons learned in variant interpretation—a case report

BackgroundAlport syndrome is a hereditary kidney disease characterized by hematuria and proteinuria. Although there have been reports of autosomal dominant COL4A4 variants, this is likely an underdiagnosed condition. Improved access to affordable genetic testing has increased the diagnosis of Alport syndrome. As genetic testing becomes ubiquitous, it is imperative that clinical nephrologists understand the benefits and challenges associated with clinical genetic testing.Case PresentationWe present a family of Mexican descent with a heterozygous COL4A4 variant (c.5007delC, ClinVar accession numbers: SCV001580980.2, SCV001993731.1) not previously discussed in detail in the literature. The proband received a biopsy diagnosis suggestive of Fabry disease 18 years after she first developed hematuria and progressed to chronic kidney disease stage III. One year later, the proband was provisionally diagnosed with Alport syndrome after a variant of uncertain significance in the COL4A4 gene was identified following targeted family variant testing of her daughter. Upon review of the medical histories of the proband’s children and niece, all but one had the same variant. Of the four with the variant, three display clinical symptoms of hematuria, and/or proteinuria. The youngest of the four, only months old, has yet to exhibit clinical symptoms. Despite these findings there was a considerable delay in synthesizing this data, as patients were tested in different commercial genetic testing laboratories. Subsequently, understanding this family’s inheritance pattern, family history, and clinical symptoms, as well as the location of the COL4A4 variant resulted in the upgrade of the variant’s classification. Although the classification of this variant varied among different clinical genetic testing laboratories, the consensus was that this variant is likely pathogenic.ConclusionsThis COL4A4 variant (c.5007delC) not yet discussed in detail in the literature is associated with Alport syndrome. The inheritance pattern is suggestive of autosomal dominant inheritance. This report highlights the intricacies of variant interpretation and classification, the siloed nature of commercial genetic testing laboratories, and the importance of a thorough family history for proper variant interpretation. Additionally, the cases demonstrate the varied clinical presentations of Alport syndrome and suggest the utility of early screening, diagnosis, monitoring, and treatment.

Implementing proactive quality assurance quality control (QAQC) and quality management (QM) practices at coal testing laboratories

Many commercial coal testing laboratories are accredited to ISO 17025 – General requirements for the competence of testing and calibration laboratories. There is an expected reliance in the Coal mining industry that the laboratory adheres to all elements of this standard in between successive accreditation audits conducted every 18 months by the independent accreditation body, National Association of Testing Authorities (NATA). In the absence of proactive QAQC &amp; QM practices monitoring the quality of the information reported by laboratories, potential issues impacting production decisions and reconciliation results are only determined in a reactive manner. In addition, for mining companies working with several internal and external laboratories across the supply chain, the management of the logistics, practices and information becomes very challenging and time-consuming, impacting the company’s ability to track laboratory results as key inputs in a production and reconciliation perspective. The absence of proactive QAQC &amp; QM practices results in a sub-optimal/reactive approach in the Coal industry, increasing the risk of short-term unaware production gaps related to quality, increased time required for quality breach investigations, the absence of a holistic approach/monitoring in the value chain, and the financial impact for the business performing under sub-optimal conditions. This paper aims to show the journey towards the implementation of a new proactive QAQC and QM program, where now the quality of many different laboratories across the supply chain can be monitored and linked with global reconciliation results, as an improvement opportunity to complement the current industry standard ISO 17025 accreditation and Proficiency Round Robin approach.

Multiresidue analysis of pesticides, polyaromatic hydrocarbons and polychlorinated biphenyls in poultry meat and chicken eggs by GC-MS/MS: method development and validation

The study uses gas chromatography with tandem mass spectrometry (GC-MS/MS) to develop a reliable analytical approach for detecting multiclass pesticides, polycyclic aromatic hydrocarbons (PAHs) and polychlorinated biphenyls (PCBs) in poultry meat and chicken eggs. The meat (2 g) and egg (4 g) samples were extracted with acidified acetonitrile (10 mL) as part of the optimized sample preparation technique. The cleanup consisted of freezing an aliquot of the extract (5 mL) at −20 °C, followed by dispersive solid phase extraction using 50 mg PSA + 100 mg C18+150 mg MgSO4. The matrix co-extractives were effectively removed and the method performance met the European Commission’s analytical quality control criteria (SANTE/12682/2019). The method was validated at two spiking levels (10 and 20 ng/g of 225 pesticides, 9 PAHs and 8 PCBs), and good recoveries (70–120%) and precision-RSDs (≤20%) were achieved for 90% of the targeted pesticide residues. For 80% of the compounds, the LOQs were ≤10 ng/g. The results of the intra-laboratory (involving six analysts) and inter-laboratory validation studies (involving eight ISO 17025 accredited laboratories) established satisfactory ruggedness and reproducibility. It created potential applications in commercial residue testing laboratories for regulatory compliance check purposes.

Quantifying the Effects of Modifications to the Sikora Buffer pH Method for Oregon Soils

ABSTRACT The Sikora buffer pH method is a non-hazardous alternative to the Shoemaker, McLean, and Pratt (SMP) method for lime requirement estimation (LRE) for Oregon soils. Commercial soil testing laboratories may implement method modifications in order to streamline the method for large-scale testing, however the potential effects of these modifications on buffer pH results are currently unknown. Modifications that artificially increase buffer pH are of particular concern, as they could potentially result in under-application of lime and crop damage. The goal of this project was to evaluate and quantify five Sikora buffer pH method modifications for five Oregon agricultural soils. Soil type did not show a significant interaction effect with method modifications. Modifications related to mixing method, measurement in supernatant instead of slurry, and soil:water:buffer ratio were found to significantly increase Sikora buffer pH results by 0.063, 0.065, and 0.058 units, respectively, in comparison to the control. Mean Sikora buffer pH decreased by 0.02–0.05 units when equilibration time was increased from 0 to 180 min. Our findings show how even minor modifications can alter Sikora buffer pH results, illustrating the importance of avoiding deviations from the original method. Additionally, comparing results among labs may help minimize over- or under-recommending of lime to agricultural fields.

EP517: Differences in genetic testing uptake in a large metropolitan pediatric outpatient genetics clinic over time

Genetic testing is an important diagnostic tool in pediatric genetics that can end a diagnostic odyssey and help a patient receive prompt and appropriate medical care. Historically, prior authorizations and approvals for genetic tests often occurred and were initiated by the clinic directly and testing could only be completed on peripheral blood samples. It often took several weeks to months to obtain prior authorization. This introduced barriers to completing the recommended testing. More recently, commercial genetic testing laboratories work directly with payers to obtain prior authorizations for genetic tests rather than the clinic. In addition, buccal swabs and saliva samples are widely utilized and accepted as an alternative, non-invasive sample type for a majority of genetic tests. This has allowed our genetics clinic at Texas Children's Hospital (TCH) to send genetic tests for patients during the clinical encounter or shortly after. In the context of updates to commercial laboratory billing and specimen type updates, we sought to describe genetic tests completed and compare the difference in uptake of genetic testing in our outpatient genetics clinic between 2018 and 2021. We completed a retrospective review of all genetic tests recommended for pediatric patients seen in the outpatient Genetics Clinic at TCH from March 1, 2021 through August 31, 2021. All patients had private or public insurance accepted in Texas or the United States. Data was compared to similar, previously collected genetic testing data in the same period in 2018 at TCH including type of payer, genetic tests recommended, and outcomes of completed genetic tests. In 2018, 1,173 unique tests were recommended; 702 (59.8%) tests were sent on patients with a public payer and 471 (40.2%) tests had private payers. All tests had prior authorizations completed by the institution and required a peripheral blood sample. The 2021 data included 903 unique recommended genetic tests of which 403 (44.6%) of the tests were sent on patients who had a private payer and 500 (55.4%) had a public payer. Buccal samples were collected during the clinical encounter for 293/903 (32.4%) tests. Of those, four tests were not completed and eight tests were sponsored and not billed to a payer. The remaining 281 tests were billed directly to the performing genetic testing laboratory. Tests billed to public payers were more likely (p<0.0001) to have been completed in 2021 compared to 2018. Tests billed to private payers were not more likely to be completed in the same period (p=0.83). In 2021, 791 (87.6%) of the recommended tests were billed directly to the performing commercial laboratory. Of those, 645 (81.5%) were completed. In 2018, when prior authorizations were obtained by the institution, 665 (57.6%) were completed (p<0.0001). Genetic testing uptake has improved in our pediatric outpatient genetics clinic between 2018 and 2021. Increased uptake in genetic testing can shorten the diagnostic odyssey for pediatric patients and help them more quickly and efficiently access necessary medical care. The improved uptake in genetic testing in our clinic may be attributed to recent updates to billing practices and the use of alternative sample types accepted at commercial genetic testing laboratories. Some genetic tests can now be completed the same day as the clinical encounter or at home with buccal swabs. This reduces the potential barriers patients may face related to limited transportation, availability during business hours, financial constraints, and invasive testing. Additional investigation into alternative billing practices and how it impacts genetic testing and patient satisfaction should be completed.

Rapid and Quantitative Analysis of Aflatoxin M1 From Milk Using Atmospheric Pressure-Matrix Assisted Laser Desorption/Ionization (AP-MALDI)-Triple Quadrupole Selected Reaction Monitoring.

Aflatoxin M1 (AFM1) is a carcinogenic hydroxylated metabolite commonly found in milk. It is relatively stable toward decontamination procedures posing a major health risk, and it requires an international regulatory mandate of detection at trace levels. To develop a high-throughput, reliable, and compliant method for the identification of AFM1 in milk samples using atmospheric pressure-matrix assisted laser desorption/ionization (AP-MALDI) selected reaction monitoring (SRM) quantitation. The milk sample was diluted in water and cleaned with immunoaffinity chromatography (IAC), followed by analysis using AP-MALDI hyphenated with a triple quadrupole mass spectrometer for SRM. A fast and reliable AP-MALDI SRM quantitative method was developed for the determination of AFM1 with analysis time of 1 min per sample. The diagnostic product ions of AFM1 at 273.1 u and 229.2 u were monitored during the SRM. The calibration curves yielded excellent linearity (R2 = 0.99) with good recoveries for quality control samples (97-106%). The ion ratios of the qualifier to quantifier displayed excellent RSD (1-7.8%) for n = 3. The developed method provided rapid quantification for AFM1. The fast AP-MALDI SRM method can allow analysis of AFM1 in a large number of milk samples. Given the time required for analysis, cost-effectiveness, and superior analytical performance, this method can be adopted in commercial food testing laboratories. Aflatoxins (AF) are a major health risk. Speedy analysis of large sample sizes from food is a risk mitigation strategy but remains an unmet need. Quantitative, chromatography-free, and internal standard-free AP-MALDI SRM based analysis of AF is a high-throughput and cost-efficient alternative. Satisfactory performance was achieved for quantitative AP-MALDI SRM analysis of AFM1 in milk subsequent to a simple sample clean-up step.

Hereditary Cancer Risk Using a Genetic Chatbot Before Routine Care Visits.

To examine user uptake and experience with a clinical chatbot that automates hereditary cancer risk triage by collecting personal and family cancer history in routine women's health care settings. We conducted a multicenter, retrospective observational study of patients who used a web-based chatbot before routine care appointments to assess their risk for hereditary breast and ovarian cancer, Lynch syndrome, and adenomatous polyposis syndromes. Outcome measures included uptake and completion of the risk-assessment and educational section of the chatbot interaction and identification of hereditary cancer risk as evaluated against National Comprehensive Cancer Network criteria. Of the 95,166 patients invited, 61,070 (64.2%) engaged with the clinical chatbot. The vast majority completed the cancer risk assessment (89.4%), and most completed the genetic testing education section (71.4%), indicating high acceptability among those who opted to engage. The mean duration of use was 15.4 minutes (SD 2 hours, 56.2 minutes) when gaps of inactivity longer than 5 minutes were excluded. A personal history of cancer was reported by 19.1% (10,849/56,656) and a family history of cancer was reported by 66.7% (36,469/54,652) of patients who provided the relevant information. One in four patients (14,850/54,547) screened with the chatbot before routine care appointments met National Comprehensive Cancer Network criteria for genetic testing. Among those who were tested, 5.6% (73/1,313) had a disease-causing pathogenic variant. A chatbot digital health tool can help identify patients at high risk for hereditary cancer syndromes before routine care appointments. This scalable intervention can effectively provide cancer risk assessment, engage patients with educational information, and facilitate a path toward preventive genetic testing. Implementation of the chatbot in clinics was funded by industry support from commercial genetic testing laboratories Ambry, Invitae, and Progenity.

Nearly half of TP53 variants are misattributed to Li-Fraumeni syndrome: A clinical evaluation of individuals with TP53 variants detected by hereditary cancer panel assays on blood or saliva.

10501 Background: Multigene panel testing (MGPT) has identified TP53 pathogenic or likely pathogenic (P/LP) variants in patients with diverse phenotypes from no cancer to classic Li-Fraumeni syndrome (LFS). There is increasing recognition of variants at low allelic fraction (VAF) for TP53 in particular, which can be suggestive of post-zygotic mosaicism or aberrant clonal expansion (ACE), comprising clonal hematopoiesis of indeterminate potential (CHIP) or occult hematologic neoplasia. Distinguishing among these categories is essential because of widely different cancer risk and management implications for patients and their relatives. We report an evaluation of TP53 positive probands to determine germline versus somatic status from a cancer genetics clinic. Methods: We reviewed probands with TP53 P/LP variants by MGPT on blood (N = 83) or saliva (N = 1) samples from 2012-2019. Available VAFs were collected from commercial testing laboratories. Probands positive for a known familial variant, who met LFS testing criteria without indication of low VAF, or who carried the Brazil founder p.R337H variant were considered germline. For those with uncertain germline status, data was obtained from ancillary testing of family members, cultured skin fibroblasts, and other somatic benign or tumor tissues. TP53 variants were further categorized based on all available data. Results: Of the 84 probands, 28 (33%) had germline TP53 P/LP variants determined by above initial criteria; 18 (21%) were confirmed germline through ancillary testing. Seven (8%) individuals were classified as having constitutional mosaicism. In eleven (13%) individuals, the TP53 variants were consistent with ACE, in 9 (11%) with CHIP and in 2 (2%) with a hematologic malignancy (1 CLL, 1 NHL). Five (6%) cases could not be categorized despite ancillary testing. Fifteen (18%) probands declined any further workup. Conclusions: A TP53 P/LP variant found on peripheral blood or saliva MGPT does not always originate in the germline. In a clinical cancer genetics cohort, only 54% of patients had TP53 P/LP germline variants; these patients plus those with constitutional mosaicism (8%) require intensified surveillance. Assessment of VAF, family member testing, and analysis of TP53 in cultured fibroblasts or other tissue samples may distinguish germline and constitutional mosaic variants from the ACE spectrum. Expanding use of MGPT will increase this clinical challenge, which may motivate the modification of lab reports to include VAF and possible non-germline explanations. The findings of this study support a framework of multiple strategies to discern true constitutional status of a TP53 P/LP variant.

Real-world integration of genomic data into the electronic health record: the PennChart Genomics Initiative

Technologies in genomic medicine have rapidly evolved and transformed the ability to deliver precision medicine in nearly every field of health care. As genomic medicine has advanced, the electronic health record (EHR) has simultaneously been adopted into routine practice. A recent Points to Consider Statement by the American College of Medical Genetics and Genomics (ACMG) provides a framework for the optimal integration of genomic data into the EHR.1 The PennChart Genomics Initiative (PGI) at the University of Pennsylvania is a multidisciplinary collaborative effort including Penn Medicine clinicians, researchers, pathologists, legal staff, and information services with input and efforts from Epic Systems Corporation (Wisconsin) and Ambry Genetics Corporation (California), a commercial genetic testing laboratory.

Atypical Genotypes for Canine Agouti Signaling Protein Suggest Novel Chromosomal Rearrangement.

Canine coat color is a readily observed phenotype of great interest to dog enthusiasts; it is also an excellent avenue to explore the mechanisms of genetics and inheritance. As such, multiple commercial testing laboratories include basic color alleles in their popular screening panels, allowing for the creation of genotyped datasets at a scale not before appreciated in canine genetic research. These vast datasets have revealed rare genotype anomalies that encourage further exploration of color and pattern inheritance. We previously reported the simultaneous presence of greater than two allele variants at the Agouti Signaling Protein (ASIP) locus in a commercial genotype cohort of 11,790 canids. Here we present additional data to characterize the occurrence of anomalous ASIP genotypes. We document the detection of combinations of three or four ASIP allele variants in 17 dog breeds and Dingoes, at within-breed frequencies of 1.32–63.34%. We analyze the potential impact on phenotype that these allele combinations present, and propose mechanisms that could account for the findings, including: gene recombination, duplication, and incorrect causal variant identification. These findings speak to the accuracy of industry-wide protocols for commercial ASIP genotyping and imply that ASIP should be analyzed via haplotype, rather than using only the existing allele hierarchy, in the future.

Association of a Polygenic Risk Score With Breast Cancer Among Women Carriers of High- and Moderate-Risk Breast Cancer Genes

To date, few studies have examined the extent to which polygenic single-nucleotide variation (SNV) (formerly single-nucleotide polymorphism) scores modify risk for carriers of pathogenic variants (PVs) in breast cancer susceptibility genes. In previous reports, polygenic risk modification was reduced for BRCA1 and BRCA2 PV carriers compared with noncarriers, but limited information is available for carriers of CHEK2, ATM, or PALB2 PVs. To examine an 86-SNV polygenic risk score (PRS) for BRCA1, BRCA2, CHEK2, ATM, and PALB2 PV carriers. A retrospective case-control study using data on 150 962 women tested with a multigene hereditary cancer panel between July 19, 2016, and January 11, 2019, was conducted in a commercial testing laboratory. Participants included women of European ancestry between the ages of 18 and 84 years. Multivariable logistic regression was used to examine the association of the 86-SNV score with invasive breast cancer after adjusting for age, ancestry, and personal and/or family cancer history. Effect sizes, expressed as standardized odds ratios (ORs) with 95% CIs, were assessed for carriers of PVs in each gene as well as for noncarriers. The median age at hereditary cancer testing of the population was 48 years (range, 18-84 years); there were 141 160 noncarriers in addition to carriers of BRCA1 (n = 2249), BRCA2 (n = 2638), CHEK2 (n = 2564), ATM (n = 1445), and PALB2 (n = 906) PVs included in the analysis. The 86-SNV score was associated with breast cancer risk in each of the carrier populations (P < 1 × 10-4). Stratification was more pronounced for noncarriers (OR, 1.47; 95% CI, 1.45-1.49) and CHEK2 PV carriers (OR, 1.49; 95% CI, 1.36-1.64) than for carriers of BRCA1 (OR, 1.20; 95% CI, 1.10-1.32) or BRCA2 (OR, 1.23; 95% CI, 1.12-1.34) PVs. Odds ratios for ATM (OR, 1.37; 95% CI, 1.21-1.55) and PALB2 (OR, 1.34; 95% CI, 1.16-1.55) PV carrier populations were intermediate between those for BRCA1/2 and CHEK2 noncarriers. In this study, the 86-SNV score was associated with modified risk for carriers of BRCA1, BRCA2, CHEK2, ATM, and PALB2 PVs. This finding supports previous reports of reduced PRS stratification for BRCA1 and BRCA2 PV carriers compared with noncarriers. Modification of risk in CHEK2 carriers associated with the 86-SNV score appeared to be similar to that observed in women without a PV. Larger studies are needed to provide more refined estimates of polygenic modification of risk for women with PVs in other moderate-penetrance genes.

Development of feed composition tables using a statistical screening procedure

Millions of feed composition records generated annually by testing laboratories are valuable assets that can be used to benefit the animal nutrition community. However, it is challenging to manage, handle, and process feed composition data that originate from multiple sources, lack standardized feed names, and contain outliers. Efficient methods that consolidate and screen such data are needed to develop feed composition databases with accurate means and standard deviations (SD). Considering the interest of the animal science community in data management and the importance of feed composition tables for the animal industry, the objective was to develop a set of procedures to construct accurate feed composition tables from large data sets. A published statistical procedure, designed to screen feed composition data, was employed, modified, and programmed to operate using Python and SAS. The 2.76 million data received from 4 commercial feed testing laboratories were used to develop procedures and to construct tables summarizing feed composition. Briefly, feed names and nutrients across laboratories were standardized, and erroneous and duplicated records were removed. Histogram, univariate, and principal component analyses were used to identify and remove outliers having key nutrients outside of the mean ± 3.5 SD. Clustering procedures identified subgroups of feeds within a large data set. Aside from the clustering step that was programmed in Python to automatically execute in SAS, all steps were programmed and automatically conducted using Python followed by a manual evaluation of the resulting mean Pearson correlation matrices of clusters. The input data set contained 42, 94, 162, and 270 feeds from 4 laboratories and comprised 25 to 30 nutrients. The final database included 174 feeds and 1.48 million records. The developed procedures effectively classified by-products (e.g., distillers grains and solubles as low or high fat), forages (e.g., legume or grass-legume mixture by maturity), and oilseeds versus meal (e.g., soybeans as whole raw seeds vs. soybean meal expellers or solvent extracted) into distinct sub-populations. Results from these analyses suggest that the procedure can provide a robust tool to construct and update large feed data sets. This approach can also be used by commercial laboratories, feed manufacturers, animal producers, and other professionals to process feed composition data sets and update feed libraries.

Application of Automated Mini-Solid-Phase Extraction Cleanup for the Analysis of Pesticides in Complex Spice Matrixes by GC-MS/MS.

Pesticide residues are routinely tested in spices for trade compliance. This results in a huge sample load for food testing laboratories and demands automation in sample preparation. Although there exists a method for the analysis of pesticides in fruits using an automated sample cleanup by mini-solid-phase extraction (mini-SPE) technique, no study is available to date on spices. This study aims to develop an automated sample cleanup method using mini-SPE technique in a range of spices, including chili powder, turmeric, black pepper, cumin, coriander, and cardamom. This automated sample preparation workflow involved an X-Y-Z instrument autosampler, and a set of mini-SPE cartridges comprising cleanup sorbents. Spice samples were extracted by acetonitrile, and the extract was put into an autosampler vial for automated mini-SPE cleanup before analysis by GC tandem MS. For an efficient cleanup, three different sorbent compositions were compared along with various automated workflows. For the relatively simple matrixes (e.g., coriander, cumin, and cardamom), the LOQ for the target pesticides was 10 ng/g with acceptable recovery, and precision. The method provided an LOQ of 10 ng/g for around 77% of the compounds in the relatively complex matrixes (e.g., turmeric, chili powder, and black pepper). The remainder of the compounds had satisfactory recoveries at 20 ng/g and higher levels. Given its time effectiveness and efficient analytical performance, this method can be adopted in commercial food testing laboratories for time-bound analysis of a large volume of samples. The study describes effectiveness of the automated mini-SPE cleanup in multiresidue analysis of pesticides in a range of spice matrixes. The method facilitates high-throughput residue analysis in compliance with the regulatory requirements of sensitivity and method performance.