Amplification of the <Emphasis Type="Italic">Phytophthora infestans</Emphasis> RG57 loci Facilitates <Emphasis Type="Italic">in planta</Emphasis> T-RFLP Identification of Late Blight Genotypes

Late blight, caused by the oomycete Phytophthora infestans (Mont.) de Bary, is a devastating disease in potato and tomato and causes yield and quality losses worldwide. The disease first emerged in central America and has since spread in North America including the United States and Canada. Several new genotypes of P. infestans have recently emerged, including US-22, US-23 and US-24. Due to significant economic and environmental impacts, there has been an increasing interest in the rapid identification of P. infestans genotypes. In addition to providing details regarding the various phenotypic characteristics such as fungicide resistance, host preference, and pathogenicity associated with various P. infestans genotypes, information related to pathogen movement and potential recombination may also be determined from the genetic analyses. Restriction fragment length polymorphism (RFLP) analysis with the RG57 loci is one of the most reliable procedures used to genotype P. infestans. However, the RFLP procedure requires propagation and isolation of the pathogen and relatively large amounts of DNA. Isolation of the late blight pathogen is sometimes impossible due to the poor condition of the infected tissues or the presence of fungicide residues. In this study, we describe a procedure to identify P. infestans at the molecular level in planta using terminal restriction fragment length polymorphism (T-RFLP) of the RG57 loci. This T-RFLP assay is sufficiently sensitive to detect and differentiate P. infestans genotypes directly in planta without propagation and isolation of the pathogen, to facilitate the timely implementation of best management practices.     

Photodynamic therapy of feline cutaneous squamous cell carcinoma using a newly developed liposomal photosensitizer: preliminary results concerning drug safety and efficacy

BACKGROUND: Squamous cell carcinomas are common skin tumors in cats. We investigated photodynamic therapy (PDT) using a new liposomal photosensitizer as a minimally invasive, effective treatment that can be easily performed while achieving good cosmetic results. AIM: The goal of this study was to assess and describe possible toxicities using a liposomal formulation of the photosensitizer meta-(Tetrahydroxyphenyl)Chlorin (m-THPC) and investigate if favorable pharmacokinetics translate into favorable tumor response and control. ANIMALS: Eighteen client-owned cats with 20 spontaneous cutaneous squamous cell carcinomas were included in the study. METHODS: PDT was performed using a new, liposomal formulation of the photosensitizer. Toxicity, tumor response, and tumor control were evaluated retrospectively. RESULTS: No general adverse effects were observed in cats treated with the new liposomal formulation. Mild local toxicity such as erythema and edema were seen in 15% of the patients. All cats responded to therapy, with a complete response rate of 100%. The overall 1-year control rate was 75%. The tumor recurrence rate was 20% with a median time to recurrence of 172.25 +/-87.1) days. CONCLUSIONS AND CLINICAL IMPORTANCE: A new liposomal photosensitizer was successfully used for squamous cell carcinoma in cats and was well tolerated. There were no systemic adverse effects observed with the liposomal formulation. The favorable pharmacokinetics of the liposomal drug resulted in a favorable tumor response.     

Quantifying uncertainty in the wild‐caught fisheries goal of the Ocean Health Index

Sustainability indices are proliferating, both to help synthesize scientific understanding and inform policy. However, it remains poorly understood how such indices are affected by underlying assumptions of the data and modelling approaches used to compute indicator values. Here, we focus on one such indicator, the fisheries goal within the Ocean Health Index (OHI), which evaluates the sustainable provision of food from wild fisheries. We quantify uncertainty in the fisheries goal status arising from the (a) approach for estimating missing data (i.e., fish stocks with no status) and (b) reliance on a data‐limited method (catch‐MSY) to estimate stock status (i.e., B/B_MSY). We also compare several other models to estimate B/B_MSY, including an ensemble approach, to determine whether alternative models might reduce uncertainty and bias. We find that the current OHI fisheries goal model results in overly optimistic fisheries goal statuses. Uncertainty and bias can be reduced by (a) using a mean (vs. median) gap‐filling approach to estimate missing stock scores and (b) estimating fisheries status using the central tendency from a simulated distribution of status scores generated by a bootstrap approach that incorporates error in B/B_MSY. This multitiered approach to measure and describe uncertainty improves the transparency and interpretation of the indicator and allows us to better understand uncertainty around our OHI fisheries model and outputs for country‐level interpretation and use.     

Host resistance to lung infection mediated by major vault protein in epithelial cells

The airway epithelium plays an essential role in innate immunity to lung pathogens. Ribonucleoprotein particles primarily composed of major vault protein (MVP) are highly expressed in cells that encounter xenobiotics. However, a clear biologic function for MVP is not established. We report here that MVP is rapidly recruited to lipid rafts when human lung epithelial cells are infected with Pseudomonas aeruginosa, and maximal recruitment is dependent on bacterial binding to the cystic fibrosis transmembrane conductance regulator. MVP was also essential for optimal epithelial cell internalization and clearance of P. aeruginosa. These results suggest that MVP makes a substantial contribution to epithelial cell-mediated resistance to infection.     

Ectomycorrhizal communities in a Mediterranean forest ecosystem dominated by <Emphasis Type="Italic">Quercus ilex</Emphasis>: seasonal dynamics and response to drought in the surface organic horizon

• Introduction   

Millions of hectares of Quercus ilex forests dominate disturbed landscapes in the western part of the Mediterranean basin. Although these forests are very widespread, little is known about the composition and structure of their associated ectomycorrhizal fungal communities.     

Binding Sites for Ca2+-Channel Effectors and Ryanodine in Periplaneta americana—Possible Targets for New Insecticides

The calcium channel and the ‘calcium release channel’ of muscle membrane of the cockroach Periplaneta americana have been characterized. Biological assays with calcium channel blockers and ryanodine on different insects and acari revealed pronounced insecticidal effects with ryanodine, but not with calcium channel blockers, at concentrations between 0·1 and 300 μg ml⁻¹. Skeletal muscle membranes derived either from the tubular network or from the sarcoplasmatic reticulum of P. americana were characterized with respect to the binding of the dihydropyridine (DHP) [³H]isradipine (PN 200-110), the phenyl-alkylamine [³H]verapamil and the alkaloid [³H]ryanodine. Preliminary binding studies with the benzothiazepine [³H]diltiazem suggest a low-affinity binding site with a IC₅₀ value of 3·3 μM . All binding sites tested were sensitive to treatment with proteinase K. Optimal conditions for binding of the radioligand ryanodine revealed the highest specific binding at pH 8 and at calcium chloride concentrations between 100 and 500 μM . EGTA at 10 μM abolished 95% of the ryanodine binding. Binding studies with calcium channel binding sites revealed a pronounced effect of low Ca²⁺ concentrations on specific isradipine binding, whereas verapamil and diltiazem binding were only reduced by the presence of 200 μM EGTA. With respect to high Ca²⁺ concentrations, specific binding of diltiazem, isradipine and verapamil was reduced by 73, 40 and 20%, respectively, at 5 mM Ca²⁺. Radioligand binding experiments showed high-affinity binding sites for ryanodine and isradipine. K_D values of 0·95 nM (B_max=550 fmol mg⁻¹ protein) and 0·75 nM (B_max=213 fmol mg⁻¹ protein) were determined respectively. A lower-affinity binding site was identified in binding studies with verapamil (K_D=7·4 nM and B_max=27 fmol mg⁻¹ protein). [³H]isradipine displacement studies with several dihydropyridines revealed the following ranking of affinity: nitrendipine>isradipine>Bay K8664≪nicardipine. Displacement of [³H]verapamil binding by effectors of the phenylalkylamine binding site showed that bepridil and S(-)verapamil had the highest affinities of the compounds tested followed by (±)verapamil, nor-methylverapamil and R(+)verapamil.