Nutrient availability and plant phenological stage influence the substrate microbiome in container-grown Impatiens walleriana 'Xtreme Red'.

BACKGROUND: The microbiome plays a fundamental role in plant health and performance. Soil serves as a reservoir of microbial diversity where plants attract microorganisms via root exudates. The soil has an important impact on the composition of the rhizosphere microbiome, but greenhouse ornamental plants are commonly grown in soilless substrates. While soil microbiomes have been extensively studied in traditional agriculture to improve plant performance, health, and sustainability, information about the microbiomes of soilless substrates is still limited. Thus, we conducted an experiment to explore the microbiome of a peat-based substrate used in container production of Impatiens walleriana, a popular greenhouse ornamental plant. We investigated the effects of plant phenological stage and fertilization level on the substrate microbiome. RESULTS: Impatiens plants grown under low fertilization rates were smaller and produced more flowers than plants grown under optimum and high fertilization. The top five bacterial phyla present in the substrate were Proteobacteria, Actinobacteria, Bacteriodota, Verrucomicrobiota, and Planctomycetota. We found a total of 2,535 amplicon sequence variants (ASV) grouped into 299 genera. The substrate core microbiome was represented by only 1.8% (48) of the identified ASV. The microbiome community composition was influenced by plant phenological stage and fertilizer levels. Phenological stage exhibited a stronger influence on microbiome composition than fertilizer levels. Differential abundance analysis using DESeq2 identified more ASVs significantly affected (enriched or depleted) in the high fertilizer levels at flowering. As observed for community composition, the effect of plant phenological stage on microbial community function was stronger than fertilizer level. Phenological stage and fertilizer treatments did not affect alpha-diversity in the substrate. CONCLUSIONS: In container-grown ornamental plants, the substrate serves as the main microbial reservoir for the plant, and the plant and agricultural inputs (fertilization) modulate the microbial community structure and function of the substrate. The differences observed in substrate microbiome composition across plant phenological stage were explained by pH, total organic carbon (TOC) and fluoride, and across fertilizer levels by pH and phosphate (PO4). Our project provides an initial diversity profile of the bacteria occurring in soilless substrates, an underexplored source of microbial diversity.

Examining barriers to implementing a surgical-site infection bundle.

BACKGROUND: Surgical-site infections (SSIs) can be catastrophic. Bundles of evidence-based practices can reduce SSIs but can be difficult to implement and sustain. OBJECTIVE: We sought to understand the implementation of SSI prevention bundles in 6 US hospitals. DESIGN: Qualitative study. METHODS: We conducted in-depth semistructured interviews with personnel involved in bundle implementation and conducted a thematic analysis of the transcripts. SETTING: The study was conducted in 6 US hospitals: 2 academic tertiary-care hospitals, 3 academic-affiliated community hospitals, 1 unaffiliated community hospital. PARTICIPANTS: In total, 30 hospital personnel participated. Participants included surgeons, laboratory directors, clinical personnel, and infection preventionists. RESULTS: Bundle complexity impeded implementation. Other barriers varied across services, even within the same hospital. Multiple strategies were needed, and successful strategies in one service did not always apply in other areas. However, early and sustained interprofessional collaboration facilitated implementation. CONCLUSIONS: The evidence-based SSI bundle is complicated and can be difficult to implement. One implementation process probably will not work for all settings. Multiple strategies were needed to overcome contextual and implementation barriers that varied by setting and implementation climate. Appropriate adaptations for specific settings and populations may improve bundle adoption, fidelity, acceptability, and sustainability.

Patients' experiences and compliance with preoperative screening and decolonization.

BACKGROUND: To improve adherence with pre-surgical screening for Staphylococcus aureus nasal carriage and decolonization, we need more information about patients' experiences with these protocols. METHODS: We surveyed patients undergoing orthopedic, neurosurgical, or cardiac operations at Johns Hopkins Hospitals (JHH), the University of Iowa Hospitals and Clinics (UIHC) at MercyOne Northeast Iowa Neurosurgery (MONIN) to assess patients' experiences with decolonization protocols. RESULTS: Five hundred thirty-four patients responded. Respondents at JHH were significantly more likely than those at the UIHC to report using mupirocin and were significantly more likely than those at the UIHC and MONIN to feel they received adequate information about surgical site infection (SSI) prevention and decolonization. Respondents at JHH were the least likely to not worry about SSI and they were more willing to do anything they could to prevent SSI. Few patients reported barriers to adherence and side effects of mupirocin or chlorhexidine. CONCLUSION: Respondents did not report either major side effects or barriers to adherence. Patients varied in their level of concern about SSI, their willingness to invest effort in preventing SSI, and their assessments of preoperative information. To improve patients' adherence, clinicians and hospitals should assess their patients' needs and desires and tailor their preoperative processes, education, and prophylaxis accordingly.

Virus-Induced Gene Silencing for Functional Analysis of Flower Traits in Petunia.

Virus-induced gene silencing (VIGS) uses recombinant viruses to knock down the expression of endogenous plant genes, allowing for rapid functional analysis without generating stable transgenic plants. The Tobacco rattle virus (TRV) is a popular vector for VIGS because it has a wide host range that includes Petunia × hybrida (petunia), and it induces minimal viral symptoms. Using reporter genes like chalcone synthase (CHS) in tandem with a gene of interest (GOI; pTRV2-PhCHS-GOI), it is possible to visually identify silenced flowers so that phenotyping is more accurate. Inoculation methods and environmental conditions need to be optimized for each host plant-virus interaction to maximize silencing efficiency. This chapter will provide detailed protocols for VIGS in petunia, with an emphasis on the investigation of flower phenotypes.

Exogenous abscisic acid enhances physiological, metabolic, and transcriptional cold acclimation responses in greenhouse-grown grapevines.

Previous studies have demonstrated that the freezing tolerance (FT) of grapevine was enhanced by foliar application of exogenous abscisic acid (exo-ABA), a treatment which might be incorporated into cultural practices to mitigate cold damage in vineyards. To investigate the underlying mechanisms of this response, a two-year (2017 and 2018) study was conducted to characterize the effects of exo-ABA on greenhouse-grown 'Cabernet franc' grapevine. In control grapevines, both physiological (deeper dormancy) and biochemical (sugar accumulation in buds) changes occurred, indicating that grapevines initiated cold acclimation in the greenhouse. Compared to control, exo-ABA decreased stomatal conductance 2 h after application. Two weeks post application, exo-ABA treated grapevines showed accelerated transition of grapevine physiology during cold acclimation (increased depth of dormancy, decreased bud water content and enhanced bud FT), relative to control. Exo-ABA induced the accumulation of several sugars in buds including the raffinose family oligosaccharides (RFOs), and the RFO precursor, galactinol. The expression of raffinose and galactinol synthase genes was higher in exo-ABA treated grapevine buds, compared to control. The new findings from this study have advanced our understanding of the role of ABA in grapevine FT, which will be useful to develop future strategies to protect grapevines from cold damage.

Identification of Pseudomonas Spp. That Increase Ornamental Crop Quality During Abiotic Stress.

The sustainability of ornamental crop production is of increasing concern to both producers and consumers. As resources become more limited, it is important for greenhouse growers to reduce production inputs such as water and chemical fertilizers, without sacrificing crop quality. Plant growth promoting rhizobacteria (PGPR) can stimulate plant growth under resource-limiting conditions by enhancing tolerance to abiotic stress and increasing nutrient availability, uptake, and assimilation. PGPR are beneficial bacteria that colonize the rhizosphere, the narrow zone of soil in the vicinity of the roots that is influenced by root exudates. In this study, in vitro experiments were utilized to screen a collection of 44 Pseudomonas strains for their ability to withstand osmotic stress. A high-throughput greenhouse experiment was then utilized to evaluate selected strains for their ability to stimulate plant growth under resource-limiting conditions when applied to ornamental crop production systems. The development of a high-throughput greenhouse trial identified two pseudomonads, P. poae 29G9 and P. fluorescens 90F12-2, that increased petunia flower number and plant biomass under drought and low-nutrient conditions. These two strains were validated in a production-scale experiment to evaluate the effects on growth promotion of three economically important crops: Petunia × hybrida, Impatiens walleriana, and Viola × wittrockiana. Plants treated with the two bacteria strains had greater shoot biomass than untreated control plants when grown under low-nutrient conditions and after recovery from drought stress. Bacteria treatment resulted in increased flower numbers in drought-stressed P. hybrida and I. walleriana. In addition, bacteria-treated plants grown under low-nutrient conditions had higher leaf nutrient content compared to the untreated plants. Collectively, these results show that the combination of in vitro and greenhouse experiments can efficiently identify beneficial Pseudomonas strains that increase the quality of ornamental crops grown under resource-limiting conditions.

Preoperative Depression, Smoking, and Employment Status are Significant Factors in Patient Satisfaction After Lumbar Spine Surgery.

STUDY DESIGN: Prospective cohort study. OBJECTIVE: To determine whether comorbidities and demographics, identified preoperatively, can impact patient outcomes and satisfaction after lumbar spine surgery. SUMMARY OF BACKGROUND DATA: The surgical treatment of lower back pain does not always eliminate a patient's pain and symptoms. Revision surgeries are costly and expose the patient to additional risk. We aim to identify patient characteristics that may suggest a greater or lesser likelihood of postsurgical success by examining patient-reported measures and outcomes after surgery. METHODS: Preoperative smoking status, self-reported depression, prevalence of diabetes, obesity, level of education, and employment status were assessed in the context of patient outcome and satisfaction after lumbar spine surgery. Patients were contacted before surgery, and at 3 and 12 months postoperatively, and responded to Oswestry Disability Index (ODI) and EuroQol-5 Dimensions (EQ-5D) self-assessment examinations, as well as a satisfaction measure. RESULTS: A total of 166 patients who underwent lumbar spine surgeries at Iowa Spine and Brain Institute, a department of Covenant Medical Center, and were included in the National Neurosurgery Quality and Outcomes Database were assessed preoperatively, and at 3 and 12 months postoperatively using self-assessment tools. Depression, smoking, and employment status were found to be significant factors in patient satisfaction. Depressed patients, smokers, and patients on disability at the time of surgery have worse ODI and EQ-5D scores at all of the timepoints (baseline, 3 months, and 12 months postsurgery). CONCLUSIONS: Depression, smoking, and employment status, specifically whether a patient is on disability at the time of surgery, are all significant factors in patient satisfaction after lumbar spine surgery. These factors are also shown in impact ODI and EQ-5D scores. Surgeons should consider these particular characteristics when developing a lower back pain treatment plan involving surgery.

RNA-sequencing reveals early, dynamic transcriptome changes in the corollas of pollinated petunias.

BACKGROUND: Pollination reduces flower longevity in many angiosperms by accelerating corolla senescence. This response requires hormone signaling between the floral organs and results in the degradation of macromolecules and organelles within the petals to allow for nutrient remobilization to developing seeds. To investigate early pollination-induced changes in petal gene expression, we utilized high-throughput sequencing to identify transcripts that were differentially expressed between corollas of pollinated Petunia × hybrida flowers and their unpollinated controls at 12, 18, and 24 hours after opening. RESULTS: In total, close to 0.5 billion Illumina 101 bp reads were generated, de novo assembled, and annotated, resulting in an EST library of approximately 33 K genes. Over 4,700 unique, differentially expressed genes were identified using comparisons between the pollinated and unpollinated libraries followed by pairwise comparisons of pollinated libraries to unpollinated libraries from the same time point (i.e. 12-P/U, 18-P/U, and 24-P/U) in the Bioconductor R package DESeq2. Over 500 gene ontology terms were enriched. The response to auxin stimulus and response to 1-aminocyclopropane-1-carboxylic acid terms were enriched by 12 hours after pollination (hap). Using weighted gene correlation network analysis (WGCNA), three pollination-specific modules were identified. Module I had increased expression across pollinated corollas at 12, 18, and 24 h, and modules II and III had a peak of expression in pollinated corollas at 18 h. A total of 15 enriched KEGG pathways were identified. Many of the genes from these pathways were involved in metabolic processes or signaling. More than 300 differentially expressed transcription factors were identified. CONCLUSIONS: Gene expression changes in corollas were detected within 12 hap, well before fertilization and corolla wilting or ethylene evolution. Significant changes in gene expression occurred at 18 hap, including the up-regulation of autophagy and down-regulation of ribosomal genes and genes involved in carbon fixation. This transcriptomic database will greatly expand the genetic resources available in petunia. Additionally, it will guide future research aimed at identifying the best targets for increasing flower longevity by delaying corolla senescence.

A mechanical-biochemical feedback loop regulates remodeling in the actin cytoskeleton.

Cytoskeletal actin assemblies transmit mechanical stresses that molecular sensors transduce into biochemical signals to trigger cytoskeletal remodeling and other downstream events. How mechanical and biochemical signaling cooperate to orchestrate complex remodeling tasks has not been elucidated. Here, we studied remodeling of contractile actomyosin stress fibers. When fibers spontaneously fractured, they recoiled and disassembled actin synchronously. The disassembly rate was accelerated more than twofold above the resting value, but only when contraction increased the actin density to a threshold value following a time delay. A mathematical model explained this as originating in the increased overlap of actin filaments produced by myosin II-driven contraction. Above a threshold overlap, this mechanical signal is transduced into accelerated disassembly by a mechanism that may sense overlap directly or through associated elastic stresses. This biochemical response lowers the actin density, overlap, and stresses. The model showed that this feedback mechanism, together with rapid stress transmission along the actin bundle, spatiotemporally synchronizes actin disassembly and fiber contraction. Similar actin remodeling kinetics occurred in expanding or contracting intact stress fibers but over much longer timescales. The model accurately described these kinetics, with an almost identical value of the threshold overlap that accelerates disassembly. Finally, we measured resting stress fibers, for which the model predicts constant actin overlap that balances disassembly and assembly. The overlap was indeed regulated, with a value close to that predicted. Our results suggest that coordinated mechanical and biochemical signaling enables extended actomyosin assemblies to adapt dynamically to the mechanical stresses they convey and direct their own remodeling.

Lateral communication between stress fiber sarcomeres facilitates a local remodeling response.

Actin stress fibers (SFs) are load-bearing and mechanosensitive structures. To our knowledge, the mechanisms that enable SFs to sense and respond to strain have not been fully defined. Acute local strain events can involve a twofold extension of a single SF sarcomere, but how these dramatic local events affect the overall SF architecture is not believed to be understood. Here we have investigated how SF architecture adjusts to episodes of local strain that occur in the cell center. Using fluorescently tagged zyxin to track the borders of sarcomeres, we characterize the dynamics of resting sarcomeres and strain-site sarcomeres. We find that sarcomeres flanking a strain site undergo rapid shortening that directly compensates for the strain-site extension, illustrating lateral communication of mechanical information along the length of a stress fiber. When a strain-site sarcomere extends asymmetrically, its adjacent sarcomeres exhibit a parallel asymmetric shortening response, illustrating that flanking sarcomeres respond to strain magnitude. After extension, strain-site sarcomeres become locations of new sarcomere addition, highlighting mechanical strain as a trigger of sarcomere addition and revealing a, to our knowledge, novel type of SF remodeling. Our findings provide evidence to suggest SF sarcomeres act as strain sensors and are interconnected to support communication of mechanical information.

Proteomic analysis of pollination-induced corolla senescence in petunia.

Senescence represents the last phase of petal development during which macromolecules and organelles are degraded and nutrients are recycled to developing tissues. To understand better the post-transcriptional changes regulating petal senescence, a proteomic approach was used to profile protein changes during the senescence of Petuniaxhybrida 'Mitchell Diploid' corollas. Total soluble proteins were extracted from unpollinated petunia corollas at 0, 24, 48, and 72 h after flower opening and at 24, 48, and 72 h after pollination. Two-dimensional gel electrophoresis (2-DE) was used to identify proteins that were differentially expressed in non-senescing (unpollinated) and senescing (pollinated) corollas, and image analysis was used to determine which proteins were up- or down-regulated by the experimentally determined cut-off of 2.1-fold for P <0.05. One hundred and thirty-three differentially expressed protein spots were selected for sequencing. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) was used to determine the identity of these proteins. Searching translated EST databases and the NCBI non-redundant protein database, it was possible to assign a putative identification to greater than 90% of these proteins. Many of the senescence up-regulated proteins were putatively involved in defence and stress responses or macromolecule catabolism. Some proteins, not previously characterized during flower senescence, were identified, including an orthologue of the tomato abscisic acid stress ripening protein 4 (ASR4). Gene expression patterns did not always correlate with protein expression, confirming that both proteomic and genomic approaches will be required to obtain a detailed understanding of the regulation of petal senescence.

Ethylene regulates phosphorus remobilization and expression of a phosphate transporter (PhPT1) during petunia corolla senescence.

The programmed degradation of macromolecules during petal senescence allows the plant to remobilize nutrients from dying to developing tissues. Ethylene is involved in regulating the timing of nucleic acid degradation in petunia, but it is not clear if ethylene has a role in the remobilization of phosphorus during petal senescence. To investigate ethylene's role in nutrient remobilization, the P content of petals (collectively called the corolla) during early development and senescence was compared in ethylene-sensitive wild type Petunia x hybrida 'Mitchell Diploid' (MD) and transgenic petunias with reduced sensitivity to ethylene (35S::etr1-1). When compared to the total P content of corollas on the day of flower opening (the early non-senescing stage), P in MD corollas had decreased 74% by the late stage of senescence (advanced wilting). By contrast, P levels were only reduced by an average of 32% during etr1-1 corolla (lines 44568 and Z00-35-10) senescence. A high-affinity phosphate transporter, PhPT1 (PhPht1;1), was cloned from senescing petunia corollas by RT-PCR. PhPT1 expression was up-regulated during MD corolla senescence and a much smaller increase was detected during the senescence of etr1-1 petunia corollas. PhPT1 mRNA levels showed a rapid increase in detached corollas (treated at 1 d after flower opening) following treatment with low levels of ethylene (0.1 microl l(-1)). Transcripts accumulated in the presence of the protein synthesis inhibitor, cycloheximide, indicating that PhPT1 is a primary ethylene response gene. PhPT1 is a putative phosphate transporter that may function in Pi translocation during senescence.

Farm Management Effects on Rhizosphere Colonization by Native Populations of 2,4-Diacetylphloroglucinol-Producing Pseudomonas spp. and Their Contributions to Crop Health.

ABSTRACT Analyses of multiple field experiments indicated that the incidence and relative abundance of root-colonizing phlD+ Pseudomonas spp. were influenced by crop rotation, tillage, organic amendments, and chemical seed treatments in subtle but reproducible ways. In no-till corn plots, 2-year rotations with soybean resulted in plants with approximately twofold fewer phlD+ pseudomonads per gram of root, but 3-year rotations with oat and hay led to population increases of the same magnitude. Interestingly, tillage inverted these observed effects of cropping sequence in two consecutive growing seasons, indicating a complex but reproducible interaction between rotation and tillage on the rhizosphere abundance of 2,4-diacetlyphloroglucinol (DAPG) producers. Amending conventionally managed sweet corn plots with dairy manure compost improved plant health and also increased the incidence of root colonization when compared with nonamended plots. Soil pH was negatively correlated to rhizosphere abundance of phlD+ pseudomonads in no-till and nonamended soils, with the exception of the continuous corn treatments. Chemical seed treatments intended to control fungal pathogens and insect pests on corn also led to more abundant populations of phlD in different tilled soils. However, increased root disease severity generally was associated with elevated levels of root colonization by phlD+ pseudomonads in no-till plots. Interestingly, within a cropping sequence treatment, correlations between the relative abundance of phlD and crop stand or yield were generally positive on corn, and the strength of those correlations was greater in plots experiencing more root disease pressure. In contrast, such correlations were generally negative in soybean, a difference that may be partially explained by difference in application of N fertilizers and soil pH. Our findings indicate that farming practices can alter the relative abundance and incidence of phlD+ pseudomonads in the rhizosphere and that practices that reduce root disease severity (i.e., rotation, tillage, and chemical seed treatment) are not universally linked to increased root colonization by DAPG-producers.

Distribution and Fungicide Sensitivity of Fungal Pathogens Causing Anthracnose-like Lesions on Tomatoes Grown in Ohio.

The primary causal agents of anthracnose-like fruit rots in Ohio and their potential resistance to fungicides commonly used to control these fungal pathogens were determined. Nineteen tomato production fields throughout the state were sampled in 2002 and 2003 for fruit with anthracnose-like lesions. Fungi were isolated from these samples, classified using restriction fragment length polymorphism analysis, and identified by internal transcribed spacer sequence analysis. Some of the fungi isolated may represent secondary invaders of preexisting wounds or lesions. Colletotrichum spp. were most abundant in our collection, representing 136 of the 187 isolates. In addition, there were 23 Alternaria, 12 Fusarium, 12 Phomopsis, and 4 Mucor isolates. Colletotrichum, Alternaria, and Fusarium spp. were found throughout the major tomato production areas in the state. In a laboratory investigation, a subset of the Colletotrichum, Alternaria, and Fusarium isolates caused symptoms similar to early development of anthracnose on wounded tomato fruit. In vitro inhibition assays indicated that most Colletotrichum isolates were sensitive to labeled rates of azoxystrobin, chlorothalonil, and mancozeb. However, some Alternaria isolates were less sensitive to azoxystrobin and chlorothalonil than the Colletotrichum isolates. In addition, most Fusarium isolates were also more insensitive to azoxystrobin and mancozeb, and most Phomopsis isolates were not inhibited by azoxystrobin at the levels tested. The patterns of insensitivity to azoxystrobin and chlorothalonil were also observed in situ with excised fruit. Because the fungicides tested are not currently labeled for control of tomato diseases caused by Fusarium or Phomopsis, these results indicate that some pathogen species that can cause anthracnose-like symptoms may not be entirely sensitive to fungicides commonly used in tomato production.