LC-MS-Based Metabolomics Study of Marine Bacterial Secondary Metabolite and Antibiotic Production in Salinispora arenicola

An LC-MS-based metabolomics approach was used to characterise the variation in secondary metabolite production due to changes in the salt content of the growth media as well as across different growth periods (incubation times). We used metabolomics as a tool to investigate the production of rifamycins (antibiotics) and other secondary metabolites in the obligate marine actinobacterial species Salinispora arenicola, isolated from Great Barrier Reef (GBR) sponges, at two defined salt concentrations and over three different incubation periods. The results indicated that a 14 day incubation period is optimal for the maximum production of rifamycin B, whereas rifamycin S and W achieve their maximum concentration at 29 days. A “chemical profile” link between the days of incubation and the salt concentration of the growth medium was shown to exist and reliably represents a critical point for selection of growth medium and harvest time.     

Marine-Derived Quorum-Sensing Inhibitory Activities Enhance the Antibacterial Efficacy of Tobramycin against Pseudomonas aeruginosa

Bacterial epiphytes isolated from marine eukaryotes were screened for the production of quorum sensing inhibitory compounds (QSIs). Marine isolate KS8, identified as a Pseudoalteromonas sp., was found to display strong quorum sensing inhibitory (QSI) activity against acyl homoserine lactone (AHL)-based reporter strains Chromobacterium violaceum ATCC 12472 and CV026. KS8 supernatant significantly reduced biofilm biomass during biofilm formation (−63%) and in pre-established, mature P. aeruginosa PAO1 biofilms (−33%). KS8 supernatant also caused a 0.97-log reduction (−89%) and a 2-log reduction (−99%) in PAO1 biofilm viable counts in the biofilm formation assay and the biofilm eradication assay respectively. The crude organic extract of KS8 had a minimum inhibitory concentration (MIC) of 2 mg/mL against PAO1 but no minimum bactericidal concentration (MBC) was observed over the concentration range tested (MBC > 16 mg/mL). Sub-MIC concentrations (1 mg/mL) of KS8 crude organic extract significantly reduced the quorum sensing (QS)-dependent production of both pyoverdin and pyocyanin in P. aeruginosa PAO1 without affecting growth. A combinatorial approach using tobramycin and the crude organic extract at 1 mg/mL against planktonic P. aeruginosa PAO1 was found to increase the efficacy of tobramycin ten-fold, decreasing the MIC from 0.75 to 0.075 µg/mL. These data support the validity of approaches combining conventional antibiotic therapy with non-antibiotic compounds to improve the efficacy of current treatments.     

The endophytic fungus Piriformospora indica protects wheat from fusarium crown rot disease in simulated UK autumn conditions

The root endophytic fungus Piriformospora indica (Sebacinacea) forms mutualistic symbioses with a broad range of host plants, increasing their biomass production and resistance to fungal pathogens. This study evaluated the effect of P. indica on fusarium crown rot disease of wheat, under in vitro and glasshouse conditions. Interaction of P. indica and Fusarium isolates under axenic culture conditions indicated no direct antagonistic activity of P. indica against Fusarium isolates. Seedlings of wheat were inoculated with P. indica and pathogenic Fusarium culmorum or F. graminearum and grown in sterilized soil‐free medium or in a non‐sterilized mix of soil and sand. Fusarium alone reduced emergence and led to visible browning and reduced root growth. Roots of seedlings in pots inoculated with both Fusarium isolates and P. indica were free of visible symptoms; seed emergence and root biomass were equivalent to the uninoculated. DNA was quantified by real‐time polymerase chain reaction (qPCR). The ratio of FusariumDNA to wheat DNA rose rapidly in the plants inoculated with Fusarium alone; isolates and species were not significantly different. Piriformospora indica inoculation reduced the ratio of Fusarium to host DNA in the root systems. The reduction increased with time. The ratio of P. indica to wheat DNA initially rose but then declined in root systems without Fusarium. With Fusarium, the ratio rose throughout the experiment. The absolute amount of FusariumDNA in root systems increased in the absence of P. indica but was static in plants co‐inoculated with P. indica.     

Use of mycophenolate mofetil as a rescue agent in the treatment of severe generalized myasthenia gravis in three dogs

Objective – To describe the use of IV and oral mycophenolate mofetil (MMF) as adjunctive therapy in 3 dogs with severe generalized myasthenia gravis.
Case Series Summary – Three dogs suffering from severe generalized myasthenia gravis as confirmed by acetylcholine antibody titers were treated with MMF as part of their treatment regimens. All 3 dogs had radiographic evidence of megaesophagus and suffered from severe regurgitation. Each dog was initially treated with pyridostigmine and supportive agents. When clinical remission was not achieved, IV MMF was administered to all dogs. Signs of clinical remission were apparent within 48 hours and all dogs were later maintained on oral MMF following resolution of regurgitation.
New or Unique Information Provided – This is the first report of the use of IV MMF as adjunctive treatment in dogs with severe generalized myasthenia gravis. Outcome was favorable in all 3 dogs and no adverse effects were noted from the MMF.     

Canine leishmaniosis in the United Kingdom: A zoonotic disease waiting for a vector?

Leishmaniosis is an important sand fly transmitted protozoan disease of dogs and humans. In northern Europe, infection is mainly restricted to dogs that have travelled to and/or from endemic areas of the Mediterranean region during periods when there is high sand fly exposure, mostly between March and November. Infected dogs in these areas in northern latitudes are a potential reservoir should incursion of a competent vector occur. However, information on the scale of the potential reservoir in the UK is lacking. Confirmed cases of canine leishmaniosis entering the United Kingdom between 2005 and 2007 were identified using diagnostic samples submitted to the Department of Clinical Veterinary Science, University of Bristol and from collaborating laboratories (n = 257). All study dogs had clinico-pathological signs compatible with leishmaniosis, as typically reported in endemic countries and were leishmania positive in real time or conventional PCR tests, IFA serology and/or tissue microscopic examination for amastigote identification. Information obtained from each case included travel history, habitat, clinico-pathological findings and geographical location once located in the UK. The majority of dogs with complete travel history (n = 183) had spent at least 6 months in Spain (105/183), 28/183 were rescued from re-homing centres in the country of origin and 26/183 entered the UK with confirmed leishmaniosis. Once located in the UK, the majority of positive cases were resident in south and central England. The spectrum of clinico-pathological signs for this group of dogs is similar to that reported in endemic countries. These data confirm that a potentially significant reservoir of infected dogs is resident in areas where future climatic conditions may support introduction of competent vectors.     

Tepoxalin reduces pruritus and modified CADESI-01 scores in dogs with atopic dermatitis: a prospective, randomized, double-blinded, placebo-controlled, cross-over study

Thirty dogs with atopic dermatitis were given tepoxalin (Zubrin®, Intervet/Schering-Plough Animal Health, Boxmeer, the Netherlands) or placebo once daily for 4 weeks, followed by a wash-out period of 1 week before reversing the treatments. Pruritus was scored by the owners using the Edinburgh Pruritus Scale and one investigator employed a modification of the Canine Atopic Dermatitis Extent and Severity Index-01 (mCADESI-01) to score the physical lesions. After administration of tepoxalin there was a ≥ 50% reduction in pruritus and mCADESI-01 scores in 36% and 25% of the dogs, respectively, whereas following administration of the placebo there was a ≥ 50% reduction in pruritus and mCADESI-01 scores in only 25% and 16% of the dogs, respectively. Analysis of pooled data indicated that tepoxalin resulted in a significant reduction in pruritus ( P  = 0.012) and mCADESI-01 ( P  = 0.002) scores but there was no significant change after placebo. The median pruritus scores before and after tepoxalin were 2 (range 1–5) and 1 (range 0–5), respectively, and before and after placebo were 2 (range 0–4) and 2 (range 0–4), respectively. The median mCADESI scores before and after tepoxalin were 23 (range 0–68) and 16 (range 0–72), respectively, and before and after placebo were 18 (range 3–79) and 24 (range 0–65), respectively. At the dose used in this study (10.0–19.1 mg kg⁻¹), tepoxalin was well-tolerated and no adverse effects were noted.     

Retrospective analysis of the relationship between time of thoracostomy drain removal and discharge time

O bjectives : The objective of this study was to evaluate the relationship between the volume of fluid being produced at the time of thoracostomy drain removal and the time to hospital discharge in dogs and cats.
M ethods : Records of 101 dogs and 26 cats with thoracostomy drains were reviewed. Three subgroups were created according to the reason for thoracostomy drain placement: P (postsurgical), A (air) and F (fluid). A generalised linear model with Poisson Errors was performed to test the relationship between the volume of fluid produced at the time of thoracostomy drain removal and the time to discharge. The volume of fluid produced and the time to discharge were compared between species and subgroups.
R esults : No significant relationship was found between the volume of fluid produced at the time of thoracostomy drain removal and the time to discharge in either species or between the time to discharge and the reason for thoracostomy drain placement. Animals with a volume of fluid higher than 2 ml/kg/day at the time of thoracostomy drain removal did not have increased hospitalisation times.
C linical S ignificance : Thoracostomy drain can be removed, without clinical compromise, when the volume of fluid produced exceeds 2 ml/kg/day. However, other clinical parameters must be taken into consideration.     

Detecting defects in conifers with ground penetrating radar: applications and challenges

Our objective was to test ground penetrating radar (GPR) to non‐destructively estimate decay volumes in living coniferous trees. GPR is geophysical tool which uses an antenna to propagate short bursts of electromagnetic energy in solid materials and measure the two‐way travel time and amplitude of reflected signals. We compared estimates of bole decay from data collected with a SIR 3000 GPR system equipped with a 900 MHz antenna to measurements of decay from stem cross sections and increment cores for three conifer species (Pseudotsuga menziesii, Thuja plicata and Tsuga heterophylla). We found that near‐surface decay, air‐filled voids and desiccated boles had unique electromagnetic signatures, which could be separated from other defects. GPR successfully estimated the percent area of air‐filled cavities and was not significantly different than results from destructive sampling. However, separation of incipient to severe decay from benign reflectors (e.g. moisture gradient between sapwood and heartwood) in conifers was much less diagnostic than with angiosperms. A limited assessment of Acer saccharum showed that GPR has potential to detect defects in angiosperms; however, more research is needed to outline the full range of detectable defects. Based on the trees in this study, the potential for GPR to detect decay‐related defects in conifers seems limited. Despite problems detecting decay, reflections originating from the sapwood : heartwood boundary may prove useful to determine thickness of functional sapwood in conifers, but accurate quantification will require further technical development.     

Components of genetic variation for resistance of strawberry to Phytophthora cactorum estimated using segregating seedling populations and their parent genotypes

Strawberry ( Fragaria × ananassa ) seedlings from 50 bi-parental crosses among 20 elite genotypes were evaluated for resistance to Phytophthora cactorum after artificial inoculation. Plots of seedlings or runner plants were rated using a disease severity score and the percentage of stunted plants per plot. The distribution of cross means for percentages of plants with stunting was highly skewed; 79% of the inoculated seedlings showed some level of stunting compared to non-inoculated control seedlings, and all but one of the crosses had 50% or more stunted plants. Disease severity scores for the bi-parental crosses were normally distributed and expressed a range of variation not reflected by the percentage of visibly stunted plants. Factorial analysis based on seedling plot means demonstrated significant additive genetic variance for the disease severity score, and the additive genetic variance was 1·9 times greater than the estimated dominance variance. The cross-mean heritability was     for the severity score. Estimates of the additive genetic variance component using the covariance of severity scores obtained from the seedling analysis and with severity scores for their parents evaluated in a commercial environment were similar,     and 0·30, respectively. Most of the selection response obtained through genotypic selection would thus be transferred to segregating offspring.