Susceptibility of Selected Ericaceous Ornamental Host Species to Phytophthora ramorum

We assessed disease reactions of 51 species or varieties of ericaceous ornamental hosts to two isolates of Phytophthora ramorum, the causal agent of sudden oak death. Inoculation was performed with an A2 mating type U.S. isolate from rhododendron and the P. ramorum type culture of A1 mating type from Germany. For only one host were statistically significant differences in disease observed between the two isolates. Several different inoculation methods were compared. The 51 hosts tested varied widely in susceptibility, ranging from 0% to over 90% leaf area infected. Two cultivars of Vaccinium macrocarpon (cranberry) showed no disease, while three cultivars of Kalmia latifolia (mountain laurel) were all highly susceptible. The results indicate that many ornamental hosts grown in the United States are susceptible to P. ramorum under artificial inoculation conditions. Inoculum density studies with two susceptible host species showed that P. ramorum is capable of producing disease symptoms over sporangium concentrations ranging from 100 to 5,000 sporangia per ml. Mean numbers of chlamydospores forming in host tissue of 21 hosts ranged from 2 to over 900 chlamydospores per 6-mm-diameter leaf disk. Whether hosts showing susceptiblity under the experimental conditions used in this study would become infected with P. ramorum in the presence of inoculum under natural conditions is unknown

Survival of Phytophthora ramorum hyphae after exposure to temperature extremes and various humidities

We examined the effect of short-term exposure to high and low temperatures and a range of relative humidity (RH) on survival of Phytophthora ramorum hyphae. Spore-free hyphal colonies were grown on dialysis squares atop V8 medium. Colonies were transferred to water agar plates positioned at 27.5-50 C on a thermal gradient plate and incubated 2.5-480 min. For low temperature trials colonies were transferred to vials of distilled water and incubated in a water bath at 25 to 225 C for 1-24 h. In the relative humidity trials hyphal colonies were transferred to sealed humidity chambers containing various concentrations of glycerin for 1-8 h. Relative humidity was 41-93% at 20 C and 43-86% at 28 C. Survival in all trials was characterized by growth from dialysis squares into V8 medium. Temperatures of 37.5- 40 C were lethal to P. ramorum hyphae within several hours, and temperatures of 42.5-50 C were lethal within minutes. Exposure to 32.5 and 35 C resulted in reduced survival over 8 h, while 30 C had no effect on three of four isolates. Hyphal colonies demonstrated considerable tolerance to cold, with all isolates surviving a 24 h exposure to 25 C. Survival diminished over time at lower temperatures, however a few colonies survived 24 h exposure to 225 C. Temperature also affected the ability of hyphal colonies to withstand reduced humidity. A RH of 41-43% was lethal in 2 h at 28 C compared to 8 h at 20 C. Three of four isolates were unaffected by an 8 h exposure to 81 and 95% RH at 20 C, and 73 and 86% RH at 28 C. Isolate differences were apparent in tolerance to freezing temperatures and reduced humidity. From these results it is apparent that the cold temperatures found in the northeastern USA are not likely to prevent the establishment of P. ramorum. There is also the potential for hyphae, and presumably spores,to survive periods of high humidity on the leaf surface in the absence of free water. © 2008 by The Mycological Society of America

Troponin Regulatory Function and Dynamics Revealed by H/D Exchange-Mass Spectrometry*

Muscle contraction is tightly regulated by Ca2+ binding to the thin filament protein troponin. The mechanism of this regulation was investigated by detailed mapping of the dynamic properties of cardiac troponin using amide hydrogen exchange-mass spectrometry. Results were obtained in the presence of either saturation or non-saturation of the regulatory Ca2+ binding site in the NH2 domain of subunit TnC. Troponin was found to be highly dynamic, with 60% of amides exchanging H for D within seconds of exposure to D2O. In contrast, portions of the TnT-TnI coiled-coil exhibited high protection from exchange, despite 6 h in D2O. The data indicate that the most stable portion of the trimeric troponin complex is the coiled-coil. Regulatory site Ca2+ binding altered dynamic properties (i.e. H/D exchange protection) locally, near the binding site and in the TnI switch helix that attaches to the Ca2+-saturated TnC NH2 domain. More notably, Ca2+ also altered the dynamic properties of other parts of troponin: the TnI inhibitory peptide region that binds to actin, the TnT-TnI coiled-coil, and the TnC COOH domain that contains the regulatory Ca2+ sites in many invertebrate as opposed to vertebrate troponins. Mapping of these affected regions onto the troponin highly extended structure suggests that cardiac troponin switches between alternative sets of intramolecular interactions, similar to previous intermediate resolution x-ray data of skeletal muscle troponin

Nesprin 1α2 is essential for mouse postnatal viability and nuclear positioning in skeletal muscle

The position of the nucleus in a cell is controlled by interactions between the linker of nucleoskeleton and cytoskeleton (LINC) complex and the cytoskeleton. Defects in nuclear positioning and abnormal aggregation of nuclei occur in many muscle diseases and correlate with muscle dysfunction. Nesprin 1, which includes multiple isoforms, is an integral component of the LINC complex, critical for nuclear positioning and anchorage in skeletal muscle, and is thought to provide an essential link between nuclei and actin. However, previous studies have yet to identify which isoform is responsible. To elucidate this, we generated a series of nesprin 1 mutant mice. We showed that the actin-binding domains of nesprin 1 were dispensable, whereas nesprin 1α2, which lacks actin-binding domains, was crucial for postnatal viability, nuclear positioning, and skeletal muscle function. Furthermore, we revealed that kinesin 1 was displaced in fibers of nesprin 1α2-knockout mice, suggesting that this interaction may play an important role in positioning of myonuclei and functional skeletal muscle