Characterizing the Diversity of Oomycetes Associated with Diseased Cotton Seedlings in Alabama.

Many oomycete species are associated with the seedlings of crops, including upland cotton (Gossypium hirsutum L.), which leads to annual threats. The diversity of oomycete species in Alabama needs to be better understood since the last survey of oomycetes associated with cotton in Alabama was 20 years ago-before significant updates to taxonomy and improvements in identification of oomycetes using molecular tools. Our current study aimed to identify oomycetes associated with Alabama cotton seedlings, correlate diversity with soil edaphic factors, and assess virulence toward cotton seed. Thirty symptomatic cotton seedlings were collected independently from 25 fields in 2021 and 2022 2 to 4 weeks after planting. Oomycetes were isolated by plating root sections onto a semiselective medium. The internal transcribed spacer (ITS) region was sequenced to identify the resulting isolates. A seed virulence assay was conducted in vitro to verify pathogenicity, and 347 oomycete isolates were obtained representing 36 species. Northern Alabama soils had the richest oomycete communities and a greater silt and clay concentration than sandier soils in the central and southern coastal plains. Globisporangium irregulare and Phytophthora nicotianae were consistently recovered from cotton roots in both years. Globisporangium irregulare was pathogenic and recovered from all Alabama regions, whereas P. nicotianae was pathogenic but recovered primarily in areas with lower sand content in northern Alabama. Many oomycete species have not been previously reported in Alabama or the southeastern United States. Altogether, this knowledge will help facilitate effective management strategies for cotton seedling diseases caused by oomycetes in Alabama and the United States.

Update on the state of research to manage Fusarium head blight.

Fusarium head blight (FHB) is one of the most devastating diseases of cereal crops, causing severe reduction in yield and quality of grain worldwide. In the United States, the major causal agent of FHB is the mycotoxigenic fungus, Fusarium graminearum. The contamination of grain with mycotoxins, including deoxynivalenol and zearalenone, is a particularly serious concern due to its impact on the health of humans and livestock. For the past few decades, multidisciplinary studies have been conducted on management strategies designed to reduce the losses caused by FHB. However, effective management is still challenging due to the emergence of fungicide-tolerant strains of F. graminearum and the lack of highly resistant wheat and barley cultivars. This review presents multidisciplinary approaches that incorporate advances in genomics, genetic-engineering, new fungicide chemistries, applied biocontrol, and consideration of the disease cycle for management of FHB.

A ß-lactamase gene of Fusarium oxysporum alters the rhizosphere microbiota of soybean.

The rhizosphere is a multitrophic environment, and for soilborne pathogens such as Fusarium oxysporum, microbial competition in the rhizosphere is inevitable before reaching and infecting roots. This study established a tritrophic interaction among the plant growth-promoting rhizobacterium Burkholderia ambifaria, F. oxysporum and Glycine max (soybean) to study the effects of F. oxysporum genes on shaping the soybean microbiota. Although B. ambifaria inhibited mycelial growth and increased bacterial propagation in the presence of F. oxysporum, F. oxysporum still managed to infect soybean in the presence of B. ambifaria. RNA-Seq identified a putative F. oxysporum secretory ß-lactamase-coding gene, FOXG_18438 (abbreviated as Fo18438), that is upregulated during soybean infection in the presence of B. ambifaria. The ∆Fo18438 mutants displayed reduced mycelial growth towards B. ambifaria, and the complementation of full Fo18438 and the Fo18438 ß-lactamase domain restored mycelial growth. Using the F. oxysporum wild type, ∆Fo18438 mutants and complemented strains with full Fo18438, Fo18438 ß-lactamase domain or Fo18438 RTA1-like domain for soil inoculation, 16S rRNA amplicon sequencing revealed that the abundance of a Burkholderia operational taxonomic unit (OTU) was increased in the rhizosphere microbiota infested by the strains with Fo18438 ß-lactamase domain. Non-metric multidimensional scaling and PICRUSt2 functional analysis revealed differential abundance for the bacterial ß-lactam-related functions when contrasting the genotypes of F. oxysporum. These results indicated that the Fo18438 ß-lactamase domain provides F. oxysporum with the advantage of growing into the soybean rhizosphere, where ß-lactam antibiosis is involved in microbial competition. Accordingly, this study highlights the capability of an F. oxysporum gene for altering the soybean rhizosphere and taproot microbiota.

Endophytic Fungi as a Promising Biocontrol Agent to Protect Wheat from Fusarium graminearum Head Blight.

The search for beneficial endophytes that can be part of a constructed microbial community has increased in recent years. We characterized three endophytic fungi previously isolated from wheat for their in vitro and in planta antagonism toward the Fusarium head blight pathogen, Fusarium graminearum. The endophytes were phylogenetically characterized and shown to be Alternaria destruens, Fusarium commune, and Fusarium oxysporum. Individual fungal endophytes significantly increased seed weight and lowered the accumulation of the mycotoxin deoxynivalenol compared with F. graminearum-infected wheat heads without endophyte pretreatment. Investigation into the mechanism of competition in vitro showed that endophytes competitively excluded F. graminearum by preemptive colonization and possible inhibition over a distance. Investigations on the use of these endophytes in the field are in progress. Identification of these three endophytes highlights a common quandary in searching for beneficial microbes to use in agriculture: species definitions often do not separate individual isolates' lifestyles. A greater understanding of the risks in using intraspecies variants for biocontrol is needed and should be examined in the context of the ecology of the individuals being investigated.

Phytophthora sojae Pathotype Distribution and Fungicide Sensitivity in Michigan.

Identifying the pathotype structure of a Phytophthora sojae population is crucial for the effective management of Phytophthora stem and root rot of soybean (PRR). P. sojae has been successfully managed with major resistance genes, partial resistance, and fungicide seed treatments. However, prolonged use of resistance genes or fungicides can cause pathogen populations to adapt over time, rendering resistance genes or fungicides ineffective. A statewide survey was conducted to characterize this pathotype structure and fungicide sensitivity of P. sojae within Michigan. Soil samples were collected from 69 fields with a history of PRR and fields having consistent plant stand establishment issues. Eighty-three isolates of P. sojae were obtained, and hypocotyl inoculations were performed on 14 differential soybean cultivars, all of which carry a single Rps gene or no resistance gene. The survey identified a loss of effectiveness of Rps genes 1b, 1k, 3b, and 6, compared with a previous survey conducted in Michigan from 1993 to 1997. Three effective resistance genes were identified for P. sojae management in Michigan; Rps 3a, 3c, and 4. Additionally, the effective concentration of common seed treatment fungicides to inhibit mycelial growth by 50% (EC50) was determined. No P. sojae isolates were insensitive to the tested chemistries with mean EC50 values of 2.60 × 10-2 µg/ml for ethaboxam, 3.03 × 10-2 µg/ml for mefenoxam, 2.88 × 10-4 µg/ml for oxathiapiprolin, and 5.08 × 10-2 µg/ml for pyraclostrobin. Results suggest that while there has been a significant shift in Rps gene effectiveness, seed treatments are still effective for early season management of this disease.

Statin gap in patients living with HIV: assessing dose appropriateness.

OBJECTIVES: Patients living with HIV (PLWH) are predisposed to atherosclerotic cardiovascular disease (ASCVD), resulting in concomitant antiretroviral and statin use. A statin prescribing gap for PLWH has been reported, but appropriateness of statin selection and dosing (ASD) has not been described. METHODS: This is a comparative, retrospective study reviewing ASD in PLWH vs. uninfected patients at two outpatient clinics within an academic medical centre. Adults > 21 years old indicated for statin therapy were included. The primary outcome was percentage of PLWH prescribed an appropriately dosed statin, accounting for clinical- and patient-related variables, compared with uninfected patients. The secondary outcome was to identify patient characteristics associated with inappropriately dosed statins. RESULTS: After propensity score matching, 879 PLWH and 879 uninfected patients were included for analysis. Fewer PLWH (27.8%, n = 244) were prescribed an ASD compared with uninfected patients (40.5%, n = 356, P < 0.001). Similar rates of statin omission were seen in both populations (P = 0.11). More PLWH received too low a dose compared with the uninfected population (P < 0.0064). There were lower ASD rates in PLWH for subgroups of patients with clinical ASCVD (P = 0.00013) and 10-year ASCVD risk ≥7.5% (P = 0.00055), but not in patients with low-density lipoprotein cholesterol ≥190 mg/dL or diabetes. CONCLUSIONS: Although a statin gap exists in both PLWH and uninfected patients, the clinical significance may be greater for PLWH given the increased risk of ASCVD. This study confirms a larger statin gap in PLWH, particularly when underdosing of statin medications is considered. Additional analysis is warranted to investigate reasons for the ASD gap and beneficial clinical interventions.

Walk the Line-The Importance of Well-Informed Interpretation of QT Prolongation.

Acute care pharmacists play an integral role in identifying drug-drug interactions that may predispose patients to QT prolongation. Although most pharmacists are equipped with a baseline understanding of drug interactions and the risks of QTc prolongation, few understand the limitations of QTc calculation and interpretation. In this commentary, we put forth the notion that at times health care providers, including pharmacists, place an overemphasis on the QTc interval. In the context of using the QTc to guide pharmacotherapy decisions, unintended consequences may include a cascade of effects leading to delays in treatment, suboptimal medication selection, alert fatigue, and overutilization of resources.

Influence of Soybean Tissue and Oomicide Seed Treatments on Oomycete Isolation.

Soybean seedlings are vulnerable to different oomycete pathogens. Seed treatments containing the two antioomycete (oomicide) chemicals, metalaxyl-M (mefenoxam) and ethaboxam, are used for protection against oomycete pathogens. This study aimed to evaluate the influence of these two oomicides on isolation probability of oomycetes from soybean taproot or lateral root sections. Soybean plants were collected between the first and third trifoliate growth stages from five Midwest field locations in 2016 and four of the same fields in 2017. Oomycetes were isolated from taproot and lateral root. In 2016, 369 isolation attempts were completed, resulting in 121 isolates from the taproot and 154 isolates from the lateral root. In 2017, 468 isolation attempts were completed, with 44 isolates from the taproot and 120 isolates from the lateral roots. In three of nine site-years, the probability of isolating an oomycete from a taproot or lateral root section was significantly different. Seed treatments containing a mixture of ethaboxam and metalaxyl significantly reduced the probability of oomycete isolation from lateral roots in Illinois in 2016 and 2017, but not in other locations, which may have been related to the heavy soil type (clay loam). Among the 439 isolates collected from the two years sampled, 24 oomycete species were identified, and community compositions differed depending on location and year. The five most abundant species were Pythium sylvaticum (28.9%), P. heterothallicum (14.3%), P. ultimum var. ultimum (11.8%), P. attrantheridium (7.9%), and P. irregulare (6.6%), which accounted for 61.7% of the isolates collected. Oomicide sensitivity to ethaboxam and mefenoxam was assessed for >300 isolates. There were large differences in ethaboxam sensitivity among oomycete species, with effective concentrations to reduce optical density at 600 nm by 50% compared with the nonamended control (EC50 values) ranging from <0.01 to >100 µg/ml and a median of 0.65 µg/ml. Isolates with insensitivity to ethaboxam (>12 µg/ml) belonged to the species P. torulosum and P. rostratifingens but were sensitive to mefenoxam. Oomicide sensitivity to mefenoxam ranged from <0.01 to 0.62 µg/ml with a median of 0.03 µg/ml. The mean EC50 value of the five most abundant species to ethaboxam ranged from 0.35 to 0.97 µg/ml of ethaboxam and from 0.02 to 0.04 µg/ml of mefenoxam. No shift in sensitivity to mefenoxam or ethaboxam was observed as a result of soybean seed treatment or year relative to the nontreated seed controls. In summary, this study contributed to the understanding of the composition of oomycete populations from different soybean root tissues, locations, years, and seed treatments. Finally, seed treatments containing mefenoxam or metalaxyl plus ethaboxam can be effective in reducing the probability of oomycete isolation from soybean roots.

hagis, an R Package Resource for Pathotype Analysis of Phytophthora sojae Populations Causing Stem and Root Rot of Soybean.

Phytophthora sojae is a significant pathogen of soybean worldwide. Pathotype surveys for Phytophthora sojae are conducted to monitor resistance gene efficacy and determine if new resistance genes are needed. Valuable measurements for pathotype analysis include the distribution of susceptible reactions, pathotype complexity, pathotype frequency, and diversity indices for pathotype distributions. Previously the Habgood-Gilmour Spreadsheet (HaGiS), written in Microsoft Excel, was used for data analysis. However, the growing popularity of the R programming language in plant pathology and desire for reproducible research made HaGiS a prime candidate for conversion into an R package. Here we report on the development and use of an R package, hagis, that can be used to produce all outputs from the HaGiS Excel sheet for P. sojae or other gene-for-gene pathosystem studies.

Convergent Evolution of C239S Mutation in Pythium spp. ß-Tubulin Coincides with Inherent Insensitivity to Ethaboxam and Implications for Other Peronosporalean Oomycetes.

Ethaboxam is a benzamide antioomycete chemical (oomicide) used in corn and soybean seed treatments. Benzamides are hypothesized to bind to ß-tubulin, thus disrupting microtubule assembly. Recently, there have been reports of corn- and soybean-associated oomycetes that are insensitive to ethaboxam despite never having been exposed. Here, we investigate the evolutionary history and molecular mechanism of ethaboxam insensitivity. We tested the sensitivity of 194 isolates representing 83 species across four oomycete genera in the Peronosporalean lineage that were never exposed to ethaboxam. In all, 84% of isolates were sensitive to ethaboxam (effective concentration to reduce optical density at 600 nm by 50% when compared with the nonamended control [EC50] < 5 µg ml-1), whereas 16% were insensitive (EC50 > 11 µg ml-1). Of the insensitive isolates, two different transversion mutations were present in the 239th codon in ß-tubulin within three monophyletic groups of Pythium spp. The transversion mutations lead to the same amino acid change from an ancestral cysteine to serine (C239S), which coincides with ethaboxam insensitivity. In a treated soybean seed virulence assay, disease severity was not reduced on ethaboxam-treated seed for an isolate of Pythium aphanidermatum containing a S239 but was reduced for an isolate of P. irregulare containing a C239. We queried publicly available ß-tubulin sequences from other oomycetes in the Peronosporalean lineage to search for C239S mutations from other species not represented in our collection. This search resulted in other taxa that were either homozygous or heterozygous for C239S, including all available species within the genus Peronospora. Evidence presented herein supports the hypothesis that the convergent evolution of C239S within Peronosporalean oomycetes occurred without selection from ethaboxam yet confers insensitivity. We propose several evolutionary hypotheses for the repeated evolution of the C239S mutation.

A High-Throughput Microtiter-Based Fungicide Sensitivity Assay for Oomycetes Using Z'-Factor Statistic.

Current methods to quantitatively assess fungicide sensitivity for a diverse range of oomycetes are slow and labor intensive. Microtiter-based assays can be used to increase throughput. However, many factors can affect their quality and reproducibility. Therefore, efficient and reliable methods for detection of assay quality are desirable. The objective of this study was to develop and validate a robust high-throughput fungicide phenotyping assay based on spectrophotometric quantification of mycelial growth in liquid culture and implementation of quality control with Z' factor and growth curves. Z' factor was used to ensure that each isolate grew enough in the absence of fungicides compared with the negative control, and growth curves were used to ensure active growth at the time of concentration of a fungicide that reduces growth by 50% (EC50) estimation. EC50 and relative growth values were correlated in a side-by-side comparison with values obtained using the amended medium (gold standard) assay. Concordance correlation indicated that the high-throughput assay is accurate but may not be as precise as the amended medium assay. To demonstrate the utility of the high-throughput assay, the sensitivity of 216 oomycete isolates representing four genera and 81 species to mefenoxam and ethaboxam was tested. The assay developed herein will enable high-throughput fungicide phenotyping at a population or community level.

Predicting Soybean Yield and Sudden Death Syndrome Development Using At-Planting Risk Factors.

In the United States, sudden death syndrome (SDS) of soybean is caused by the fungal pathogen Fusarium virguliforme and is responsible for important yield losses each year. Understanding the risk of SDS development and subsequent yield loss could provide growers with valuable information for management of this challenging disease. Current management strategies for F. virguliforme use partially resistant cultivars, fungicide seed treatments, and extended crop rotations with diverse crops. The aim of this study was to develop models to predict SDS severity and soybean yield loss using at-planting risk factors to integrate with current SDS management strategies. In 2014 and 2015, field studies were conducted in adjacent fields in Decatur, MI, which were intensively monitored for F. virguliforme and nematode quantities at-planting, plant health throughout the growing season, end-of-season SDS severity, and yield using an unbiased grid sampling scheme. In both years, F. virguliforme and soybean cyst nematode (SCN) quantities were unevenly distributed throughout the field. The distribution of F. virguliforme at-planting had a significant correlation with end-of-season SDS severity in 2015, and a significant correlation to yield in 2014 (P < 0.05). SCN distributions at-planting were significantly correlated with end-of-season SDS severity and yield in 2015 (P < 0.05). Prediction models developed through multiple linear regression showed that F. virguliforme abundance (P < 0.001), SCN egg quantity (P < 0.001), and year (P < 0.01) explained the most variation in end-of-season SDS (R2 = 0.32), whereas end-of-season SDS (P < 0.001) and end-of-season root dry weight (P < 0.001) explained the most variation in soybean yield (R2 = 0.53). Further, multivariate analyses support a synergistic relationship between F. virguliforme and SCN, enhancing the severity of foliar SDS. These models indicate that it is possible to predict patches of SDS severity using at-planting risk factors. Verifying these models and incorporating additional data types may help improve SDS management and forecast soybean markets in response to SDS threats.

Annotation resource of tandem repeat-containing secretory proteins in sixty fungi.

Fungal secretory proteins that interact with host plants are regarded as effectors. Because fungal effectors rarely contain conserved sequence features, identification and annotation of fungal effectors from predicted secretory proteins are difficult using outward comparison methods such as BLAST or hidden Markov model. In desire of more sequence features to prioritize research interests of fungal secretory proteins, this study developed a pipeline to identify tandem repeat (TR) domain within putative secretory proteins and tested a hypothesis that at least one type of TR domain in non-orthologous secretory proteins has emerged from convergent evolution for plant pathogenicity. There were 2804 types of TR domains and a total of 2925 TR-containing secretory proteins found from 60 fungi. There was no conserved type of TR domain shared only by plant pathogens, indicating functional divergence for different types of TR domain and TR-containing secretory proteins. The annotation resource of putative fungal TR-containing secretory proteins provides new sequence features that will be useful for the community interested in fungal effector biology.

Focused Update on Pharmacologic Management of Hypertensive Emergencies.

PURPOSE OF REVIEW: Hypertensive emergency is defined as a systolic blood pressure > 180 mmHg or a diastolic blood pressure > 120 mmHg with evidence of new or progressive end-organ damage. The purpose of this paper is to review advances in the treatment of hypertensive emergencies within the last 5 years. RECENT FINDINGS: New literature and recommendations for managing hypertensive emergencies in the setting of pregnancy, stroke, and heart failure have been published. Oral nifedipine is now considered an alternative first-line therapy, along with intravenous hydralazine and labetalol for women presenting with pre-eclampsia. Clevidipine is now endorsed by guidelines as a first-line treatment option for blood pressure reduction in acute ischemic stroke and may be considered for use in intracranial hemorrhage. Treatment of hypertensive heart failure remains challenging; clevidipine and enalaprilat can be considered for use in this population although data supporting their use remains limited.

Significant Influence of EC50 Estimation by Model Choice and EC50 Type.

The effective control to 50% growth inhibition (EC50) is a standard statistic for evaluating dose-response relationships. Many statistical software packages are available to estimate dose-response relationships but, recently, an open source package ("drc") in R has been utilized. This package is highly adaptable, having many models to describe dose-response relationships and flexibility to describe both hormetic relationships and absolute and relative EC50. These models and definitions are generally left out of phytopathology literature. Here, we demonstrate that model choice and type of EC50 (relative versus absolute) can matter for EC50 estimation using data from Pythium oopapillum and Fusarium virguliforme. For some P. oopapillum isolates, the difference between absolute and relative EC50 was significant. Hormetic effects changed F. virguliforme EC50 distributions, leading to higher estimates than when using four- or three-parameter log-logistic models. Future studies should pay careful attention to model selection and interpretation in EC50 estimation and clearly indicate which model and EC50 measure (relative versus absolute) was used. We provide guidelines for model choice and interpretation for those wishing to set up experiments for accurate EC50 estimation.

Comparison of High-Dose Versus Standard Dose Oseltamivir in Critically Ill Patients With Influenza.

BACKGROUND: Limited data support high-dose oseltamivir in critically ill patients with influenza. In several recent influenza seasons, there were oseltamivir drug shortages. METHODS: This was a retrospective cohort analysis of 57 patients admitted to the intensive care unit (ICU) with confirmed influenza. Patients receiving high-dose oseltamivir were compared to those receiving standard dosing. RESULTS: When adjusted for clinically relevant predictors of disease severity, including age, duration of therapy, Acute Physiology and Chronic Health Evaluation II score, and receipt of extracorporeal membrane oxygenation, there was no difference in the duration of mechanical ventilation, oxygenation, ICU length of stay, or hospital length of stay between the high-dose and standard dose groups. CONCLUSIONS: As compared to the standard doses of oseltamivir, higher-dose (ie, double dose) oseltamivir was not associated with improvement in any clinical outcomes. Using higher doses empirically on all patients during influenza season may exacerbate local drug shortages.

Old Habits Die Hard: Regression in Learning and Study Strategies Inventory Scores in PharmD Students.

OBJECTIVE: Prior studies have demonstrated that "learning to learn" (L2L) courses can lead to significant improvements in students' Learning and Study Strategies Inventory (LASSI) scores immediately following the course. This study aimed to analyze whether improvements in LASSI scores are sustained 1 year following an L2L elective course. METHODS: First-year pharmacy students in the classes of 2024 and 2025 completed the LASSI at the start of the fall semester and again immediately following an L2L course. One year later, during the second professional year, students completed the LASSI a third time. Repeated-measures multivariate analysis of variance was used to analyze within-participant differences in LASSI scores across each of the 10 LASSI scales. Univariate analysis of variance with Bonferroni correction was used for pairwise comparison. RESULTS: A total of 119 students completed all 3 LASSI assessments. LASSI scores improved in all 10 scales following completion of the L2L course. However, 1 year after the completion of the course, there was a statistically significant regression in all 10 scale scores (Wilks' Λ [20,98] = 8.7). Among the 10 scales, scores for the Attitude and Concentration scales were statistically significantly lower during the second professional year relative to baseline at the start of the first professional year. Selecting Main Ideas was the only scale with a higher score during the second professional year relative to baseline. CONCLUSION: Despite marked improvements in LASSI scores following the implementation of a "learning to learn" course for first-year pharmacy students, the improvements were not sustained after 1 year.

Ready or Not: A Crossover Study of (Un)graded Individual Readiness Assurance Tests in Team-Based Learning.

OBJECTIVE: This study aimed to measure the effects of graded vs ungraded individual readiness assurance tests (iRATs) on the students' test scores and achievement goals in a team-based learning classroom. METHODS: A 2 × 2 crossover study was conducted in a required second-year pharmacotherapy course. Teams 1 to 8 were assigned to a UG iRAT during the first half of the course, followed by a G iRAT the second half of the course (G/UG group). Teams 9 to 16 were assigned to the opposite grading sequence (ie, UG/G). A multivariate analysis of variance was used to analyze the differences in test scores, as measured using iRAT and examination scores. A separate multivariate analysis of variance was used to examine the differences in achievement goals. RESULTS: There was a significant difference in test scores based on the iRAT grading condition. Individual readiness assurance tests were higher in the G condition (72.51% vs 67.99%); however, the examination scores were similar in the G and UG conditions (81.07% vs 80.32%). There was no statistically significant difference in the achievement goals based on the iRAT grading condition. CONCLUSION: In a required second-year pharmacotherapy course that uses team-based learning, student performance on the iRAT was modestly lower in the UG iRAT condition; however, the students' examination scores were unchanged. Achievement goals were unchanged based on the iRAT grading condition.