Intermediate Sensitivity Research Articles

Identification of Escherichia coli and Related Enterobacteriaceae and Examination of Their Phenotypic Antimicrobial Resistance Patterns: A Pilot Study at A Wildlife-Livestock Interface in Lusaka, Zambia.

A cross-sectional study was used to identify and assess prevalence and phenotypic antimicrobial resistance (AMR) profiles of Escherichia coli and other enterobacteria isolated from healthy wildlife and livestock cohabiting at a 10,000 acres game ranch near Lusaka, Zambia. Purposive sampling was used to select wildlife and livestock based on similarities in behavior, grazing habits and close interactions with humans. Isolates (n = 66) from fecal samples collected between April and August 2018 (n = 84) were examined following modified protocols for bacteria isolation, biochemical identification, molecular detection, phylogenetic analysis, and antimicrobial susceptibility testing by disc diffusion method. Data were analyzed using R software, Genetyx ver.12 and Mega 6. Using Applied Profile Index 20E kit for biochemical identification, polymerase chain reaction assay and sequencing, sixty-six isolates were identified to species level, of which Escherichia coli (72.7%, 48/66), E. fergusonii (1.5%, 1/66), Shigella sonnei (22.7%, 14/66), Sh. flexinerri (1.5%, 1/66) and Enterobacteriaceae bacterium (1.5%, 1/66), and their relationships were illustrated in a phylogenetic tree. Phenotypic antimicrobial resistance or intermediate sensitivity expression to at least one antimicrobial agent was detected in 89.6% of the E. coli, and 73.3% of the Shigella isolates. The E. coli isolates exhibited the highest resistance rates to ampicillin (27%), ceftazidime (14.3%), cefotaxime (9.5%), and kanamycin (9.5%). Multidrug resistance (MDR) was detected in 18.8% of E. coli isolates while only 13.3% Shigella isolates showed MDR. The MDR was detected among isolates from impala and ostrich (wild animals in which no antimicrobial treatment was used), and in isolates from cattle, pigs, and goats (domesticated animals). This study indicates the possible transmission of drug-resistant microorganisms between animals cohabiting at the wildlife–livestock interface. It emphasizes the need for further investigation of the role of wildlife in the development and transmission of AMR, which is an issue of global concern.

Abstract PS5-18: Association of factors of the obesity phenotype with breast cancer recurrence risk measured by oncotype recurrence score and short-term response to neoadjuvant endocrine therapy

Abstract Background:Increased body mass index (BMI) and metabolic syndrome are associated with an increased risk of breast cancer recurrence. The mechanism(s) responsible for this increased risk are not known, though multiple mechanisms including resistance to endocrine therapy have been proposed. We sought to determine whether obesity and/or metabolic syndrome were associated with intrinsic tumor risk as determined by the Oncotype recurrence score (RS), or with resistance to neoadjuvant endocrine therapy, as determined by Ki67 response after short-term presurgical treatment. Methods:To investigate whether BMI was associated with increased intrinsic recurrence risk, we analyzed a cohort of 779 patients treated at Vanderbilt for early stage breast cancer and who had Oncotype test results. Medical records were reviewed to record BMI at the time of breast cancer diagnosis. We compared the Oncotype RS between patients with BMI greater or less than 27, and in normal, overweight, and obese ranges. To investigate whether the increase in breast cancer recurrence risk associated with obesity could be identified by a surrogate marker of short-term response to endocrine therapy, we analyzed data from a presurgical trial of endocrine therapy in early stage breast cancer patients previously completed at Vanderbilt. In this study patients were treated with 1-2 weeks of letrozole prior to surgery, then the surgical specimen was analyzed for Ki67. The original goal of the study was to compare endocrine sensitive tumors with endocrine resistant tumors as a platform for discovery of resistance mechanisms to endocrine therapy. We retrospectively analyzed medical records for 143 patients in that study who had post-treatment Ki67 values available, and for whom BMI at the time of surgery could be obtained. We also reviewed medical records for components of the metabolic syndrome (hypertension, hyperglycemia or anti-hyperglycemic medications, low HDL, high triglycerides or statin use, and BMI > 30 as a surrogate for waist circumference). Metabolic syndrome was defined as 3 or more of the 5 components. For endocrine therapy response, a cutoff of 1 for the natural log Ki67 expression was defined as sensitive (approximately 2.5%) and tumors with natural log of Ki67 expression between 1 and 2 (approximately 7.5%) were categorized as intermediate sensitivity. Results:We did not find any significant association between BMI and recurrence score and the distribution of risk scores was similar across all BMI groups. We did not observe any correlation between RS and BMI. However we did observe a difference in the proportion of endocrine sensitive and resistant tumors in overweight and obese groups (only a minority of patients in this cohort, 18%, had normal weight). In patients with overweight, 70% of the tumors had a post-treatment Ki67 level classified as sensitive, whereas only 40% of the tumors in patients with obesity were endocrine sensitive (chi square = 0.0518). Similarly, 72% of patients without metabolic syndrome had sensitive tumors, but only 51% of patients with metabolic syndrome had sensitive tumors (relative risk 1.408, chi square = 0.0154). Conclusions:Our findings suggest that the association between obesity and/or metabolic syndrome and increased recurrence risk is not reflected in tumor-intrinsic properties present at the time of diagnosis. Rather, the effect of obesity and metabolic syndrome appears to be in response to treatment, at least as measured by a short-term surrogate marker of long-term endocrine sensitivity. Importantly, this suggests that interventions to reverse obesity and/or metabolic syndrome may be effective in improving outcomes for breast cancer recurrence, at least in part by improving sensitivity to endocrine therapy. Citation Format: Riley Bergman, Yvonne Berko, Brent Rexer. Association of factors of the obesity phenotype with breast cancer recurrence risk measured by oncotype recurrence score and short-term response to neoadjuvant endocrine therapy [abstract]. In: Proceedings of the 2020 San Antonio Breast Cancer Virtual Symposium; 2020 Dec 8-11; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2021;81(4 Suppl):Abstract nr PS5-18.

Assessing Douro Vineyards Exposure to Tropospheric Ozone

Tropospheric ozone (O3) can strongly damage vegetation. Grapevines (Vitis vinifera L.), in particular, have intermediate sensitivity to ozone. Wine production is an important economic activity, as well as a pillar to the cultural identity of several countries in the world. This study aims to evaluate the risk of Douro vineyards exposure to ozone, by estimating its concentration and deposition in the Demarcated Region of Douro in Portugal. Based on an assessment of the climatology of the area, the years 2003 to 2005 were selected among the hottest years of the recent past, and the chemical transport model CHIMERE was used to estimate the three-dimensional field of ozone and its dry deposition over the Douro region with 1 km2 of horizontal resolution. Model results were validated by comparison with measured data from the European air quality database (AirBase). The exposure indicator AOT40 (accumulated concentration of ozone above 40 ppb) was calculated and an exposure–response function was applied to determine the grapevine risk to ozone exposure. The target value for the protection of vegetation established by the Air Quality Framework Directive was exceeded on most of the Douro region, especially over the Baixo Corgo and Cima Corgo sub-regions. The results of the exposure–response functions suggest that the productivity loss can reach 27% and that the sugar content of the grapes could be reduced by 32%, but these values are affected by the inherent uncertainty of the used methodology.

Sensitivity of HER2-amplified colorectal organotypic cancer spheroids at ex vivo resistance to panitumumab and trastuzumab.

68 Background: HER2 amplification is an emerging biomarker in colorectal cancer (CRC) with increased copy number associated with improved clinical outcomes to HER2 targeting. RAS/RAF wildtype CRC also benefit from use of epidermal growth factor receptor inhibition (EGFRi). The sequencing of EGFRi versus HER2 inhibition in low copy number HER2 amplified CRC remains uncertain. Patient-derived cancer organoids (PDCOs) allow an ex vivo method to assess treatment sensitivity. We examined treatment sensitivity of a HER2 amplified PDCO at baseline and following resistance to panitumumab and trastuzumab. Methods: Following IRB-approval, fresh CRC tissue was cultured to maturation. After expansion, subcultures were treated with stepwise (20%) increase to physiologic Cmax of panitumumab (230ug/mL) and trastuzumab (180ug/mL). Threshold for escalation was median relative growth of +20% at 96h. Sensitivity was assessed on primary culture (RC1), panitumumab resistance (RC1-P) and trastuzumab resistance (RC1-T) using 96h of physiologic Cmax panitumumab, trastuzumab, and combination trastuzumab/pertuzumab. Individual sphere response was assessed for change in mean NADH autofluorescence intensity and ratio of NADH/FAD signal. Response was assessed at 96h in comparison to control using effect size of Glass’s Delta (GΔ). Results: Molecular profiling revealed HER2 copy number of 14 with no concurrent alterations in RAS, RAF, or PIK3CA. Time to resistance was similar between panitumumab (55 days) and trastuzumab (51 days). RC1 had baseline growth (+116%) which was reduced with single agent panitumumab (+17%, GΔ=1.40) with intermediate sensitivity to trastuzumab (+48%, GΔ=0.95) and trastuzumab/pertuzumab (46%, GΔ=0.99). Normalized NADH/FAD ratio revealed significant metabolic response to panitumumab (-20%, GΔ=0.66) and trastuzumab/pertuzumab (-35%, GΔ=1.16) with insignificant effect of single agent trastuzumab (-14%, GΔ=0.46). Following resistance to panitumumab, RC1-P had persistent growth with trastuzumab (+68%) which improved in combination trastuzumab/pertuzumab (+34%, GΔ=1.16). Following resistance to trastuzumab, RC1-T was insensitive to EGFRi with panitumumab including persistent growth (+58%, GΔ=0.70) and unchanged metabolism (+2%, GΔ=-0.10). Conclusions: Therapeutic dose escalation in a single PDCO of HER2 amplified CRC suggests improved sensitivity to EGFRi and dual HER2 targeting with trastuzumab/pertuzumab. Resistance to EGFRi resulted in persistent sensitivity to dual HER2 inhibition using trastuzumab/pertuzumab, however resistance to single agent trastuzumab. Resistance to trastuzumab resulted in future insensitivity to EGFRi. Molecular profiling at resistance revealed no pathologic alterations in EGFR or ERBB2 signaling, with ongoing analysis of transcriptional changes by RNAseq.

60. Evaluation of Outcomes Associated with Intermittent Versus Extended Infusion of Piperacillin/tazobactam in Acutely Ill Veterans

BackgroundAntibiotic dosing optimization is a key principle of antimicrobial stewardship. This study evaluated the impact of an extended infusion piperacillin/tazobactam dosing protocol on clinical outcomes in acutely ill veterans treated for infections at VA San Diego.MethodsThis retrospective cohort study looked at veterans admitted to the medical-surgical unit who were treated with piperacillin/tazobactam for at least 48 hours. The control group included patients who received treatment between 12/14/2017 to 7/22/2018, and the “protocol” or after protocol implementation group included patients who received treatment between 7/23/2018 to 2/28/2019. Excluded from the study were veterans with microbiological cultures showing intermediate sensitivity or resistance to piperacillin/tazobactam, those who experienced interruption in therapy, or those who required dialysis. Primary clinical outcomes included in-hospital mortality rate, 30-day mortality rate, hospital length of stay (LOS), and 30-day readmission rates. Rates of adverse effects such as elevated liver enzymes, thrombocytopenia, acute kidney impairment (AKI), and Clostridium difficile infection were also collected. χ 2, Fisher’s exact, and Mann-Whitney U tests were used for statistical analysis.Results260 veterans were included in the final analysis: 96% male, mean age 65 years, mean BMI 29, 84 met SIRS criteria for sepsis, and 55% received at least 48 hours of concomitant vancomycin. Groups had similar outcomes for median LOS, in-hospital mortality, and 30-day mortality. The incidence of AKI was significantly lower in the protocol group (39.2% vs. 56.9%, p=0.004), in veterans on concomitant vancomycin (42.3% vs. 63.2%, p=0.011), and in veterans with obesity (36.4% vs. 70.8%, p=0.001). Rates of liver enzyme elevation, thrombocytopenia, and C. difficile infection were lower in the protocol group though these were not significant.ConclusionThere was a significantly lower rate of AKI with EI dosing which supports enhanced patient safety. This may be the preferred method of administration for obese patients and/or those receiving vancomycin concurrently. This is the first study to demonstrate that EI piperacillin/tazobactam dosing significantly reduces rates of AKI in patients on concomitant vancomycin.Disclosures All Authors: No reported disclosures

Diphtheria Toxin Fused to Granulocyte-Macrophage Colony-Stimulating Factor Is Toxic to Blasts From Patients With Juvenile Myelomonocytic Leukemia and Chronic Myelomonocytic Leukemia

AbstractWe have previously demonstrated that human granulocyte-macrophage colony-stimulating factor fused to a truncated diphtheria toxin (DT388-GM-CSF) is toxic to patient acute myeloid leukemia progenitors bearing the GM-CSF receptor, but not normal marrow progenitors. We now report that exposure of mononuclear cells from five of seven (71%) juvenile myelomonocytic leukemia (JMML) patients and from 12 of 20 (60%) adult chronic myelomonocytic leukemia (CMML) patients to 10-9 mol/L DT388-GM-CSF for 48 hours in culture reduces the number of cells capable of forming colonies in semisolid medium (colony-forming units–leukemia) 10-fold to 300-fold (1 to 2.5 log decrease). In contrast, normal myeloid progenitors (colony-forming unit–granulocyte-macrophage) from six different donors treated and assayed under identical conditions were consistently insensitive to the same fusion toxin even when treated as highly purified CD34+ cells. The leukemic progenitors from the two other JMML patients showed intermediate sensitivity to DT388-GM-CSF and the leukemic progenitors from eight of the 20 (40%) CMML patients were not different from normal progenitors. Parallel measurements of the number and affinity of GM-CSF receptors on cells from the same samples showed no consistent differences between JMML, CMML, and normal light density or CD34+ bone marrow cells. The increased sensitivity of leukemic progenitors from all JMML progenitors and some CMML patients to the fusion toxin is therefore not likely to be explained by an increased density of GM-CSF receptors on these cells. We also examined the DT388-GM-CSF sensitivity of two murine cell lines transfected with cDNAs encoding varying portions of the human GM-CSF receptor α and/or β chains. These studies showed that high-affinity ligand binding was sufficient for DT388-GM-CSF–induced toxicity, as this could occur even in the absence of functional signal transduction and that the background of the host cell had a major influence on the degree to which this decreased the toxicity of DT388-GM-CSF. The selective sensitivity to DT388-GM-CSF of leukemic progenitors from a majority of JMML and CMML patients suggests that this agent could have therapeutic potential for some patients with these diseases.

Variabilidad fenotípica de aislados de Phytophthora capsici en Guanajuato

Phytophthora capsici, es el agente causal de la ‘marchitez del chile’ una de las enfermedades más importantes de este cultivo. Para establecer estrategias de control eficientes es necesario el conocimiento de la diversidad del patógeno, para esto, se realizó esta investigación cuyo objetivo fue determinar el grado de variabilidad de 29 aislados monozoospóricos obtenidos de chile (Capsicum annuum L.) de seis municipios del estado de Guanajuato y dos cepas de referencia de A1 y A2. Todos fueron caracterizados por la forma de crecimiento, tipo de compatibilidad, crecimiento a 35 °C, sensibilidad a mefenoxam y grado de virulencia. Los resultados obtenidos indican que los aislados, presentaron cinco formas de crecimiento: estrellada (62.5%), turuloso (15.62%), estolonífera (12.5%), filamentoso (6.25%) y petaloide (3.12%); su crecimiento a 35 °C fue menor a 30.7% respecto al control crecido a 27 °C, su tipo de compatibilidad fue 65.5% del tipo A1, se encontró sensibilidad intermedia al mefenoxam en 65.5% de los aislados, siendo el resto sensibles. El aislado más virulento fue el D3 y los avirulentos fueron D8, D11 y D12 todos originarios de Dolores Hidalgo. No se observa una relación directa entre la forma de crecimiento, el crecimiento a 35 °C, el tipo de compatibilidad, la resistencia a mefenoxam y el grado de virulencia.

Identification and characterization of resistance and pathogenicity of Enterococcus spp. in samples of donor breast milk.

BACKGROUNDBreast milk is the primary source of nutrition for newborns. Hospitalized babies frequently need nutritional support from Human Milk Banks. As bacterial species of the genus Enterococcus are part of the microbiota of healthy donors, they may contaminate samples of pumped breast milk. AIMTo identify and characterize the bacterial virulence and resistance in samples isolated from the nipple-areolar region, hands, and breast milk aliquots from donors at the Human Milk Bank of Municipal Hospital Esaú Matos in the city of Vitória da Conquista, Bahia State, Brazil.METHODSThe personal hygiene and sanitation of donors were analyzed with the aim of identifying possible reasons for contamination of pumped milk. Cutaneous samples as well as aliquots of unpasteurized and pasteurized milk from 30 participants were obtained. Each Enterococcus spp. isolate underwent a disk diffusion susceptibility test and molecular biology techniques to determine resistance and virulence genes.RESULTS Enterococcus spp. were identified in 30% of donors (n = 9), and 11 specimens were isolated. Resistance to tetracycline was highly prevalent, being detectable in 63% of the isolates (n = 7) and followed by intermediate sensitivity to ciprofloxacin, observed in 27% of the specimens (n = 3). The efaA gene was found in 63% (n = 7) of the isolates, while the ace gene was detected in 27% (n = 3). CONCLUSIONThis study illustrates the importance of microbiological monitoring by Human Milk Banks and the need for alternatives to prevent the presence of Enterococcus spp. in hospital settings.

NIRS-IVUS for Differentiating Coronary Plaque Rupture, Erosion, and Calcified Nodule in Acute Myocardial Infarction

ObjectivesThis study sought to investigate the ability of combined near-infrared spectroscopy and intravascular ultrasound (NIRS-IVUS) to differentiate plaque rupture (PR), plaque erosion (PE), or calcified nodule (CN) in acute myocardial infarction (AMI). BackgroundMost acute coronary syndromes occur from coronary thrombosis based on PR, PE, or CN. In vivo differentiation among PR, PE, and CN is a major challenge for intravascular imaging. MethodsThe study enrolled 244 patients with AMI who had a de novo culprit lesion in a native coronary artery. The culprit lesions were assessed by both NIRS-IVUS and optical coherence tomography (OCT). Maximum lipid core burden index in 4 mm (maxLCBI4mm) was measured by NIRS. Plaque cavity and convex calcium was detected by IVUS. The OCT diagnosis of PR (n = 175), PE (n = 44), and CN (n = 25) was used as a reference standard. ResultsIn the development cohort, IVUS-detected plaque cavity showed a high specificity (100%) and intermediate sensitivity (62%) for identifying OCT-PR. IVUS-detected convex calcium showed a high sensitivity (93%) and specificity (100%) for identifying OCT-CN. NIRS-measured maxLCBI4mm was largest in OCT-PR (705 [interquartile range (IQR): 545 to 854]), followed by OCT-CN (355 [IQR: 303 to 478]) and OCT-PE (300 [IQR: 126 to 357]) (p < 0.001). The optimal cutoff value of maxLCBI4mm was 426 for differentiating between OCT-PR and -PE; 328 for differentiating between OCT-PE and -CN; and 579 for differentiating between OCT-PR and -CN. In the validation cohort, the NIRS-IVUS classification algorithm using plaque cavity, convex calcium, and maxLCBI4mm showed a sensitivity and specificity of 97% and 96% for identifying OCT-PR, 93% and 99% for OCT-PE, and 100% and 99% for OCT-CN, respectively. ConclusionsBy evaluating plaque cavity, convex calcium, and maxLCBI4mm, NIRS-IVUS can accurately differentiate PR, PE, and CN.

Prevalence and Characterization of Carbapenem-Hydrolyzing Class D β-Lactamase-Producing Acinetobacter Isolates From Ghana

Multidrug resistance, especially carbapenem resistance in Acinetobacter bacteria is a global healthcare concern. However, available data on the phenotypic and genotypic characteristics of Acinetobacter isolates from West Africa, including Ghana is scanty. Our aim was to investigate the antibiotic resistance profile and genotypic characteristics of Acinetobacter isolates from Ghana and to characterize carbapenemase producers using whole-genome sequencing (WGS). A total of 36 Acinetobacter isolates collected at three hospitals in Ghana between 2016 and 2017 were analyzed. MICs were determined by commercial antibiotic plates. Acinetobacter baumannii MLST was determined using the Pasteur scheme. WGS of OXA-carbapenemase producers was performed using short- and long-read sequencing strategies. The resistance rate was highest for trimethoprim/sulfamethoxazole (n = 22; 61%). Six (16.7%) and eight (22.2%) isolates were resistant to ceftazidime and colistin, respectively. Two (5.6%) isolates were resistant and one (2.8%) isolate had intermediate sensitivity to three carbapenems. Fifteen STs were identified in 24 A. baumannii isolates including six new STs (ST1467 ∼ ST1472). ST78 was the predominant (n = 6) followed by ST1469 (n = 3). Four carbapenemase-producing A. baumannii isolates also were identified. Isogenic ST103 isolates Ab-B004d-c and Ab-D10a-a harbored blaOXA–23 within Tn2007 on identical plasmids, pAb-B004d-c_3, and pAb-D10a-a_3. ST1472 isolate Ab-C102 and ST107 isolate Ab-C63 carried blaOXA–58 and blaOXA–420, a rare blaOXA–58 variant, respectively, within novel genetic contexts. Our results show that A. baumannii isolates of diverse and unique genotypes, including OXA-carbapenemase producers, are circulating in Ghana highlighting the need for a wider surveillance of antimicrobial resistance.

Perspectives on Low Temperature Tolerance and Vernalization Sensitivity in Barley: Prospects for Facultative Growth Habit.

One option to achieving greater resiliency for barley production in the face of climate change is to explore the potential of winter and facultative growth habits: for both types, low temperature tolerance (LTT) and vernalization sensitivity are key traits. Sensitivity to short-day photoperiod is a desirable attribute for facultative types. In order to broaden our understanding of the genetics of these phenotypes, we mapped quantitative trait loci (QTLs) and identified candidate genes using a genome-wide association studies (GWAS) panel composed of 882 barley accessions that was genotyped with the Illumina 9K single-nucleotide polymorphism (SNP) chip. Fifteen loci including 5 known and 10 novel QTL/genes were identified for LTT—assessed as winter survival in 10 field tests and mapped using a GWAS meta-analysis. FR-H1, FR-H2, and FR-H3 were major drivers of LTT, and candidate genes were identified for FR-H3. The principal determinants of vernalization sensitivity were VRN-H1, VRN-H2, and PPD-H1. VRN-H2 deletions conferred insensitive or intermediate sensitivity to vernalization. A subset of accessions with maximum LTT were identified as a resource for allele mining and further characterization. Facultative types comprised a small portion of the GWAS panel but may be useful for developing germplasm with this growth habit.

Isolation and Antimicrobial Sensitivity of Mycoplasma synoviae and Mycoplasma gallisepticum from Vaccinated Hens in Mexico.

Mycoplasma synoviae (MS) and Mycoplasma gallisepticum (MG) strains were isolated at 39.5 °C to rule out temperature-sensitive strains (ts+) and identified using random amplification of polymorphic DNA. Then, their minimum inhibitory concentrations (MIC100) were calculated in isolated strains from broiler breeders and laying hens vaccinated with ts+ MS-H and ts+ MG TS-11 vaccines in Mexico. We sampled 631 lots of hens. A total of 28 of the 123 MS isolates and 12 of the 23 MG isolates were analyzed using random amplification of polymorphic DNA, of which 24 and 3 matched the DNA banding patterns of the MS-H and MG-F strains, respectively. The isolated MS and MG strains were sensitive to tiamulin and tylosin and showed intermediate sensitivity or resistance to lincomycin, florfenicol, erythromycin, enrofloxacin, and curcumin. Although both the MS and MG strains were sensitive to the same antibiotics (MIC100 lower than 1 mg mL−1), the MG strains were 5 to 10 times more sensitive than the MS strains. MS is the most frequently isolated mycoplasma in Mexican poultry production. The MS vaccine used (ts+ MS-H) could reverse its thermosensitivity and therefore could regain its virulence. MS was less sensitive to tiamulin and tylosin compared to MG.

Utility of Magnetic Resonance Imaging for Differentiating Necrotizing Fasciitis from Severe Cellulitis: A Magnetic Resonance Indicator for Necrotizing Fasciitis (MRINEC) Algorithm.

We developed a new magnetic resonance indicator for necrotizing fasciitis (MRINEC) algorithm for differentiating necrotizing fasciitis (NF) from severe cellulitis (SC). All adults with suspected NF between 2010 and 2018 in a tertiary hospital in South Korea were enrolled. Sixty-one patients were diagnosed with NF and 28 with SC. Among them, 34 with NF and 15 with SC underwent magnetic resonance imaging (MRI). The MRINEC algorithm, a two-step decision tree including T2 hyperintensity of intermuscular deep fascia and diffuse T2 hyperintensity of deep peripheral fascia, diagnosed NF with 94% sensitivity (95% confidence interval (CI), 80–99%) and 60% specificity (95% CI, 32–84%). The algorithm accurately diagnosed all 15 NF patients with a high (≥8) laboratory risk indicator for necrotizing fasciitis (LRINEC) score. Among the five patients with an intermediate (6–7) LRINEC score, sensitivity and specificity were 100% (95% CI, 78–100%) and 0% (95% CI, 0–84%), respectively. Finally, among the 29 patients with a low (≤5) LRINEC score, the algorithm had a sensitivity and specificity of 88% (95% CI, 62–98%) and 69% (95% CI, 39–91%), respectively. The MRINEC algorithm may be a useful adjuvant method for diagnosing NF, especially when NF is suspected in patients with a low LRINEC score.

Fluoride toxicity variably affects overall physiology and grain development in three contrasting rice genotypes, representing a potential biohazard.

Ingestion of fluoride through consumption of contaminated food grains has been regarded to be hazardous for consumer health. The current study indicated the possible occurrence of such biohazard due to fluoride bioaccumulation in rice grains and straw (cattle feed). The effects of fluoride toxicity at three stages of grain development in three rice genotypes, viz., IR-64, Gobindobhog (aromatic), and Khitish, were also studied. Irrigation with fluoride-infested water inhibited grain formation in IR-64 and reduced grain yield in Gobindobhog. Fluoride toxicity promoted seed sterility in IR-64 by triggering reactive oxygen species (ROS) production and cellular necrosis, suppressing genes like GIF1, DEP1, and SPL14 (positively controlling seed formation) and inducing GW2 (negatively mediating grain development). Gobindobhog showed intermediate fluoride sensitivity and accumulated high levels of proline, anthocyanins, flavonoids, and phenolics due to the induction of genes like P5CS, ANS, and PAL in developing grains. The agronomic attributes in Khitish were unaffected by fluoride stress due to regulated fluoride uptake and high expression of GIF1, DEP1, and SPL14 along with an increased synthesis of anthocyanins, flavonoids, and phenolics. Khitish also accumulated low ROS as a result of which lowest lipoxygenase expression (among selected cultivars) was observed in developing grains. Fluoride entry was accelerated in the straw of Khitish, possibly due to the absence of regulated uptake mechanism in dead seedlings. Furthermore, the ecological concerns regarding fluoride bioaccumulation and reduced grain yield at the varietal level were also established, based on statistical modelling.

Abstract 2701: Patient-derived colorectal cancer organoid therapeutic escalation: Tracking resistance to EGFR inhibition for predicting therapeutic sensitivity

Abstract Background: Precision oncology requires translational tools predictive of clinical benefit. Epidermal growth factor receptor inhibition (EGFRi) yields a survival advantage in RAS/RAF wildtype (wt) colorectal cancer (CRC), however clinical tools do not predict individual patient response or resistance mechanisms. Patient-derived cancer organoids (PDCOs) allow for an ex vivo method to track subclonal response. Here, we examine the development of PDCO EGFRi resistance as a novel biomarker to differentiate treatment sensitivities. Methods: Fresh CRC tissue was obtained following consent to an IRB-approved protocol. Molecular profiling was performed on expanded PDCOs and in parallel to clinical hotspot sequencing. Cultures were treated with stepwise (20%) increases in panitumumab (46ug/mL per escalation) up to the physiologic Cmax (230ug/mL). A HER2 amplified culture was also assessed by escalation of trastuzumab with 20% stepwise increase to physiologic Cmax (180ug/mL). Threshold for escalation to the next dose level was a median relative growth of 20% at 96h. Time to resistance (TTR) was defined from start to persistent PDCO growth at 100% Cmax. Treatment studies assessed the longest diameter of individual spheres, assessed at 48h and compared to control using effect size (Glass's (G) Δ). TTR was compared to molecular profiles of primary resistance and clinical outcomes. Results: CRC PDCOs were derived from 10 patients (5 localized and 5 metastatic samples). PDCOs had diverse molecular profiles: 3 KRAS mutant (mt), 1 PTEN mt, 7 TP53 mt, and 7 RAS/RAF wt. TTR ranged from 26-226 days, with 90% of PDCOs lines achieving resistance. RAS mt PDCOs had a homogeneous TTR of 27.0±1.4 days. RAS wt PDCOs displayed marked variation in therapeutic response with TTR of 51.5±61.7 days (p=0.06, compared to RAS mt). A HER2-amplified PDCO (copy number 14) showed intermediate sensitivity to EGFRi (TTR 55 days), similar to dose escalation of HER2-targeting trastuzumab (TTR of 51 days). This HER2-amplified PDCO showed intermediate sensitivity to single agent trastuzumab (GΔ = 0.56), yet no sensitivity to trastuzumab following EGFRi resistance (GΔ = 0.09). Prospective clinical correlation was possible for 1 PDCO line, a RAS wt PDCO derived prior to EGFRi which showed marked sensitivity with growth and metabolic arrest achieved at 40% CMax consistent with clinical response. Conclusions: We present therapeutic dose escalation using PDCOs as a translational tool to track the clonal dynamics of resistance to EGFRi in CRC. This technique provides an objective measure of TTR to differentiate culture specific sensitivity in a clinically relevant timeframe. Molecular profiling of resistant cultures is ongoing which may provide an ex vivo tool for predicting individual resistance as cancers evade therapeutic pressure of targeted selection. Citation Format: Jeremy D. Kratz, Aishwarya Sunil, Meghan Conroy, Cheri A. Pasch, Nicole Lassen, Hannah Houtler, Kayla K. Lemmon, Linda Clipson, Daniel E. Abbott, Evie H. Carchman, Elise H. Lawson, Noelle K. Loconte, Sam J. Lubner, Kristina A. Matkowskyj, Daniel L. Mulkerin, Nataliya Uboha, Dustin A. Deming. Patient-derived colorectal cancer organoid therapeutic escalation: Tracking resistance to EGFR inhibition for predicting therapeutic sensitivity [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 2701.

Characterization of Staphylococcus aureus Strains Isolated from Veterinary Hospital.

This study aims to detect Staphylococcus aureus (S. aureus) resistance in the veterinary hospital environment. S. aureus are one of the components of the microbiota, and they may be present in patients in a veterinary hospital environment. Methicillin resistance is determined by a chromosomal gene (mecA), which codes for modifications in the beta-lactam antibiotic receptor, where the penicillin-binding protein will have a low affinity for the antibiotic. Samples were collected through swabs of materials and equipment at the hospital. S. aureus was identified in 7.6% (21/276) of the samples collected, and of the 21 strains isolated, 4 (19.0%) carried the mecA gene. MRSA are all strains of S. aureus that express the mecA gene. Four strains harbor the mecA gene; however, only two expressed the phenotypic resistance to cefoxitin and were characterized as MRSA. An isolate (strain 18) present on a patient care table was identified as methicillin-resistant S. aureus with intermediate sensitivity to vancomycin (VISA). Our observations suggest the need for containment measures (good antisepsis practices) to avoid the possible transmission of resistant bacterial agents for the veterinary hospital environment.

Dealing with aflatoxin B1 dihydrodiol acute effects: Impact of aflatoxin B1-aldehyde reductase enzyme activity in poultry species tolerant to AFB1 toxic effects.

Aflatoxin B1 aldehyde reductase (AFAR) enzyme activity has been associated to a higher resistance to the aflatoxin B1 (AFB1) toxicity in ethoxyquin-fed rats. However, no studies about AFAR activity and its relationship with tolerance to AFB1 have been conducted in poultry. To determine the role of AFAR in poultry tolerance, the hepatic in vitro enzymatic activity of AFAR was investigated in liver cytosol from four commercial poultry species (chicken, quail, turkey and duck). Specifically, the kinetic parameters Vmax, Km and intrinsic clearance (CLint) were determined for AFB1 dialdehyde reductase (AFB1-monoalcohol production) and AFB1 monoalcohol reductase (AFB1-dialcohol production). In all cases, AFB1 monoalcohol reductase activity saturated at the highest aflatoxin B1 dialdehyde concentration tested (66.4 μM), whereas AFB1 dialdehyde reductase did not. Both activities were highly and significantly correlated and therefore are most likely catalyzed by the same AFAR enzyme. However, it appears that production of the AFB1 monoalcohol is favored over the AFB1 dialcohol. The production of alcohols from aflatoxin dialdehyde showed the highest enzymatic efficiency (highest CLint value) in chickens, a species resistant to AFB1; however, it was also high in the turkey, a species with intermediate sensitivity; further, CLint values were lowest in another tolerant species (quail) and in the most sensitive poultry species (the duck). These results suggest that AFAR activity is related to resistance to the acute toxic effects of AFB1 only in chickens and ducks. Genetic selection of ducks for high AFAR activity could be a means to control aflatoxin sensitivity in this poultry species.

Mapping Fluorescence Enhancement of Plasmonic Nanorod Coupled Dye Molecules.

Plasmonically enhanced fluorescence is a widely studied and applied phenomenon, however, only a comparative theoretical and experimental analysis of coupled fluorophores and plasmonic nanoresonators makes it possible to uncover how this phenomenon can be controlled. A numerical optimization method was applied to design configurations that are capable of resulting in an enhancement of excitation and emission, moreover, of both phenomena simultaneously in coupled Cy5 dye molecule and gold nanorod systems. Parametric sensitivity studies revealed how the fluorescence enhancement depends on the molecule’s location, distance and orientation. Coupled systems designed for simultaneous improvement exhibited the highest (intermediate directional) total fluorescence enhancement, which is accompanied by intermediate sensitivity to the molecule’s parameters, except the location and orientation sensitivity at the excitation wavelength. Gold nanorods with a geometry corresponding to the predicted optimal configurations were synthesized, and DNA strands were used to control the Cy5 dye molecule distance from the nanorod surface via hybridization of the Cy5-labelled oligonucleotide. State-of-the-art dSTORM microscopy was used to accomplish a proof-of-concept experimental demonstration of the theoretically predicted (directional) total fluorescence enhancement. The measured fluorescence enhancement was in good agreement with theoretical predictions, thus providing a complete kit to design and prepare coupled nanosystems exhibiting plasmonically enhanced fluorescence.

Blended Vision Achieved by Combining High and Low Addition Power Diffractive Intraocular Lenses with Micromonovision: A Clinical Outcome.

Purpose To evaluate the clinical outcome of blended vision combined with micromonovision (MMBV) using a +4 diopter (D) addition power (add) diffractive intraocular lens (MIOL) and a +2.75D add MIOL with a myopic target of −0.5 D. Methods One hundred twenty eyes of 60 cases were enrolled. The +4 D add MIOLs were placed in the nondominant eye, the +2.75 D add MIOLs were placed in the dominant eye with a myopic target of −0.5 D in 30 cases (the MMBV group), and the +4 D add MIOLs were placed in both eyes in another 30 cases (controls). Postoperative clinical outcomes were compared between the two groups. Results Compared with the controls, binocular uncorrected intermediate vision at 70 cm was significantly better in the MMBV group (p=0.02). Contrast sensitivity at 12 cycles per degree and the 6% and 12.5% low-contrast visual acuities were also significantly better in the MMBV group compared with the controls (p values = 0.05, 0.05, and 0.04, respectively). Uncorrected and corrected distance and near VA did not differ significantly between the two groups. Conclusion MMBV provided a better intermediate VA, contrast sensitivity, and low-contrast VA than bilateral implantation of the +4 D add MIOL, while preserving comparable near and distance vision.

Experimental Investigation of Uranium Volatility during Vapor Condensation.

The predictive models that describe the fate and transport of radioactive materials in the atmosphere following a nuclear incident (explosion or reactor accident) assume that uranium-bearing particulates would attain chemical equilibrium during vapor condensation. In this study, we show that kinetically driven processes in a system of rapidly decreasing temperature can result in substantial deviations from chemical equilibrium. This can cause uranium to condense out in oxidation states (e.g., UO3 vs UO2) that have different vapor pressures, significantly affecting uranium transport. To demonstrate this, we synthesized uranium oxide nanoparticles using a flow reactor under controlled conditions of temperature, pressure, and oxygen concentration. The atomized chemical reactants passing through an inductively coupled plasma cool from ∼5000 to 1000 K within milliseconds and form nanoparticles inside a flow reactor. The ex situ analysis of particulates by transmission electron microscopy revealed 2-10 nm crystallites of fcc-UO2 or α-UO3 depending on the amount of oxygen in the system. α-UO3 is the least thermodynamically preferred polymorph of UO3. The absence of stable uranium oxides with intermediate stoichiometries (e.g., U3O8) and sensitivity of the uranium oxidation states to local redox conditions highlight the importance of in situ measurements at high temperatures. Therefore, we developed a laser-based diagnostic to detect uranium oxide particles as they are formed inside the flow reactor. Our in situ measurements allowed us to quantify the changes in the number densities of the uranium oxide nanoparticles (e.g., UO3) as a function of oxygen gas concentration. Our results indicate that uranium can prefer to be in metastable crystal forms (i.e., α-UO3) that have higher vapor pressures than the refractory form (i.e., UO2) depending on the oxygen abundance in the surrounding environment. This demonstrates that the equilibrium processes may not dominate during rapid condensation processes, and thus kinetic models are required to fully describe uranium transport subsequent to nuclear incidents.