A real-time polymerase chain reaction assay for the detection of Mycoplasma agalactiae

AIM: To develop a real-time PCR for the detection of Mycoplasma agalactiae, using PCR primers targeting the ma-mp81 gene.

METHODS: A group of 15 M. agalactiae isolates, 21 other Mycoplasma spp. isolates and 21 other bacterial isolates was used in evaluation of the assay.

RESULTS: All M. agalactiae isolates were detected by the assay and none of the non-target isolates was amplified. The analytical detection limit of the assay was 10 fg of purified genomic DNA and 10⁴ cfu/ml milk inoculated with M. agalactiae. When applied to goat-milk samples collected from three herds free of M. agalactiae infection, the assay had a specificity of 100%.

CONCLUSIONS: The assay would be useful in a diagnostic laboratory, providing specific, sensitive and rapid detection of M. agalactiae.     

In planta detection used to define the distribution of the European lineages of Phytophthora ramorum on larch (Larix) in the UK

Two genetically distinct evolutionary lineages of the oomycete pathogen Phytophthora ramorum are responsible for the major epidemic on larch (Larix spp.) in the UK: EU1 (historically widespread) and the recently identified EU2 (reported only from Northern Ireland and a small area in southwest Scotland). Methods for lineage discrimination have required pure cultures of P. ramorum but, as the pathogen is challenging to isolate from infected larch tissue, only limited data have been available on the distribution of EU2. In this study a protocol was developed to determine the lineage of P. ramorum in infected larch tissue without the need for isolation. The protocol was applied to 134 UK samples collected during 2013–14. In addition, lineage testing was applied to over 300 P. ramorum isolates cultured from a wide range of hosts between 2002 and 2012. Combined data confirmed that EU2 is restricted to Northern Ireland and a small area of southwest Scotland where it is the dominant lineage. There was no evidence of EU2 spread into England and Wales where only EU1 was found. However, EU2 was more widely distributed in southern and eastern parts of Scotland than previously reported. Furthermore, EU1 and EU2 were detected <10 km apart in larch plantations. This study provides the first reports of natural EU2 infection on European larch (Larix decidua), hybrid larch (Larix × eurolepis), beech (Fagus sylvatica), noble fir (Abies procera) and western hemlock (Tsuga heterophylla).     

Canine calvarial subperiosteal hematomas are fluid to soft tissue attenuating mass‐like lesions with smoothly marginated peripheral mineralization on CT

Subperiosteal hematomas are accumulations of blood between a bone and the periosteum leading to elevation of the periosteum. When ossified they have a mineralized outer rim. For this retrospective, multi‐institutional case series, medical records were searched to identify dogs that underwent CT for focal calvarial swellings noted acutely after trauma. A total of four dogs were included. Computed tomography images were reviewed for each case. The focal swellings had progressed in size during the weeks after the head trauma until the time of imaging. Findings in all cases included a fluid to soft tissue attenuating mass‐like lesion with smoothly marginated peripheral mineralization. Diagnosis was confirmed in two cases by cytology and/or histopathology. Therefore, authors recommend that subperiosteal hematoma be included in the differential diagnosis list for dogs with these clinical and CT characteristics. Based on our review of the literature, this is the first report to describe the CT features of calvarial subperiosteal hematomas in dogs.     

Treatment of patent ductus arteriosus by placement of two intravascular embolisation coils in a puppy

A 14-week-old border collie with a history of exercise intolerance was confirmed to have a patent ductus arteriosus (PDA) from colour flow Doppler echocardiography. Under general anaesthesia, angiography was performed to gauge accurately the width of the ductus, and then two intravascular embolisation coils were introduced into the ductus via percutaneous catheterisation of the femoral vein. The coils were both 8 mm in diameter, each with four loops. This led to an immediate disappearance of the murmur. Some slight residual flow was detected at the time of the procedure by angiography, but by 10 days postoperatively there was no PDA flow detectable. The time taken to complete the procedure was 55 minutes, with a total fluoroscopy time of 15 minutes. Placement of intravascular embolisation coils represents a viable alternative to traditional surgical methods of ductus closure.     

Limited collaborative study of an optional method for the isolation of light filth from fig and fruit paste.

The present official first action method for the isolation of light filth from fig and fruit paste, 44.083(a), occasionally yields excessive plant debris on filter papers, which causes difficulty in effectively counting insect filth. The proposed method provides the option of an additional flotation of trapped off material in a Corning percolator. This revision has been adopted as official first action as an alternative to 44.083(a).     

Branch growth and biomass allocation in Abies amabilis saplings in contrasting light environments

Aboveground biomass allocation, and height and branch growth were studied in saplings of the shade-tolerant conifer, Abies amabilis Dougl. ex Forbes growing in large openings and in the understory of an old-growth forest in western Oregon. The presence of annual overwintering budscale scars was used to infer extension growth histories; annual growth rings in branches and stems were used in combination with extension histories to compute partitioning of new biomass among leaves, branches and stems. Saplings growing in large gaps had conical crowns, whereas understory saplings had umbrella shaped crowns as a result of much greater rates of branch extension than stem extension. Understory saplings grew slowly in height because of low rates of biomass production and low allocation of biomass to stem extension. About 40% of new biomass was allocated to foliage in both groups, but understory saplings allocated more of the remaining growth increment to branches and less to stem than did saplings growing in large gaps. These results differ from the patterns observed in shade-tolerant saplings of tropical forests, where allocation to foliage increases with shading and branch allocation is much lower than observed here. This difference in allocation may reflect mechanical constraints imposed by snow loads on the evergreen A. amabilis crowns, particularly on flat-crowned understory saplings.     

Effects of elevated concentrations of atmospheric CO2 and tropospheric O3 on decomposition of fine roots

Rising atmospheric carbon dioxide (CO2) concentration ([CO2]) could alter terrestrial carbon (C) cycling by affecting plant growth, litter chemistry and decomposition. How the concurrent increase in tropospheric ozone (O3) concentration ([O3]) will interact with rising atmospheric [CO2] to affect C cycling is unknown. A major component of carbon cycling in forests is fine root production, mortality and decomposition. To better understand the effects of elevated [CO2] and [O3] on the dynamics of fine root C, we conducted a combined field and laboratory incubation experiment to monitor decomposition dynamics and changes in fine root litter chemistry. Free-air CO2 enrichment (FACE) technology at the FACTS-II Aspen FACE project in Rhinelander, Wisconsin, elevated [CO2] (535 microl 1-1) and [O3] (53 nl 1-1) in intact stands of pure trembling aspen (Populus tremuloides Michx.) and in mixed stands of trembling aspen plus paper birch (Betula papyrifera Marsh.) and trembling aspen plus sugar maple (Acer saccharum Marsh.). We hypothesized that the trees would react to increased C availability (elevated [CO2]) by increasing allocation to C-based secondary compounds (CBSCs), thereby decreasing rates of decomposition. Because of its lower growth potential, we reasoned this effect would be greatest in the aspen-maple community relative to the aspen and aspen-birch communities. As a result of decreased C availability, we expected elevated [O3] to counteract shifts in C allocation induced by elevated [CO2]. Concentrations of CBSCs were rarely significantly affected by the CO2 and O3 treatments in decomposing fine roots. Rates of microbial respiration and mass loss from fine roots were unaffected by the treatments, although the production of dissolved organic C differed among communities. We conclude that elevated [CO2] and [O3] induce only small changes in fine root chemistry that are insufficient to significantly influence fine root decomposition. If changes in soil C cycling occur in the future, they will most likely be brought about by changes in litter production.     

The ‘European Apple Genome Mapping Project’-developing a strategy for mapping genes coding for agronomic characters in tree species

Summary A recently initiated collaborative project involving apple breeders in seven European countries is described. The objective is to improve the European apple crop by molecular-aided breeding to increase efficency and reduce the time-scale in breeding for resistance, tree habit and fruit quality. The strategy adopted provides a model for similar studies in fruit, forest and other woody species. The project is based on progenies from a small number of crosses involving many important agronomic genes. Replication of these reference progenies by vegetative propagation will enable studies to be carried out simultaneously in each country. By developing a range of molecular markers, including isozymes, RFLPs and sequence-tagged DNA probes, an integrated molecular map is being constructed for use in a wide range of breeding and genetic studies. Construction of a database recording many mapped molecular markers will enable efficient exploitation of data in future genetic, breeding and physiological studies of apple. Aspects of the adopted strategy, techniques and management are discussed in the context of mapping genes in perennial crop genomes.     

Relationships between water-use traits and photosynthesis in Brassica oleracea resolved by quantitative genetic analysis

The genetic control of water‐use and photosynthetic traits in Brassica oleracea is resolved by genetic analysis of quantitative trait loci (QTL). Variations in leaf conductance, photosynthetic assimilation rate, leaf thickness and leaf nitrogen content were assessed in a segregating population of F₁‐derived doubled haploid (DH) B. oleracea lines. In addition, stable carbon isotope ratios in leaf organic material were used as a surrogate measure of plant water‐use efficiency. Analysis of an existing linkage map for the population revealed significant QTL on seven linkage groups. Single significant QTL explained between 3.4% and 36.6% of the phenotypic variance in each of the traits measured. The locations of QTL for several traits were found to coincide in a physiologically meaningful way; stable carbon isotope discrimination had QTL co‐locating with leaf level water‐use efficiency, photosynthetic capacity with leaf thickness and nitrogen content and stomatal density with leaf thickness. Taken together, these results suggest that single genes or clusters of genes at these loci may have an influence on the expression of physiologically related traits controlling water‐use and photosynthesis.     

Robenacoxib in the dog: target species safety in relation to extent and duration of inhibition of COX-1 and COX-2

The safety of robenacoxib, a nonsteroidal anti-inflammatory drug with high selectivity for inhibition of the cyclooxygenase (COX)-2 isoform of COX, was investigated in the dog in two randomized, placebo-controlled, parallel group studies. Robenacoxib was administered orally once daily to healthy young beagle dogs at 0 (placebo), 10, 20 and 40 mg/kg for 1 month (Study 1) and at 0 (placebo), 2, 4, 6 and 10 mg/kg for 6 months (Study 2). Relative to placebo treatment, no significant adverse effects of robenacoxib were recorded in either study for clinical observations, haematological and clinical chemistry variables or macroscopic or microscopic lesions at necropsy. In Study 2, additional examinations identified no adverse effects of robenacoxib on buccal bleeding time, electrocardiographic and ophthalmoscopic examinations, urinalysis or stifle joint tissues. Pharmacokinetic-pharmacodynamic simulations indicated that all dosages of robenacoxib were associated with marked inhibition of COX-2 (median Emax 74-99% inhibition). For the highest dosage of robenacoxib (40 mg/kg in Study 1), the upper limit of the 90% tolerance interval was associated with 71% inhibition of COX-1 at Emax, but 50% inhibition persisted for only 3.5 h. This level of inhibition of COX-1 with robenacoxib was not associated with any detectable toxicity, suggesting that the high safety index of robenacoxib in dogs is a function of both its high COX-2 selectivity and short residence time in the central compartment.