Characterization of intracellular and extracellular saxitoxin levels in both field and cultured Alexandrium spp. samples from Sequim Bay, Washington

Traditionally, harmful algal bloom studies have primarily focused on quantifying toxin levels contained within the phytoplankton cells of interest. In the case of paralytic shellfish poisoning toxins (PSTs), intracellular toxin levels and the effects of dietary consumption of toxic cells by planktivores have been well documented. However, little information is available regarding the levels of extracellular PSTs that may leak or be released into seawater from toxic cells during blooms. In order to fully evaluate the risks of harmful algal bloom toxins in the marine food web, it is necessary to understand all potential routes of exposure. In the present study, extracellular and intracellular PST levels were measured in field seawater samples (collected weekly from June to October 2004–2007) and in Alexandrium spp. culture samples isolated from Sequim Bay, Washington. Measurable levels of intra- and extra-cellular toxins were detected in both field and culture samples via receptor binding assay (RBA) and an enzyme-linked immunosorbent assay (ELISA). Characterization of the PST toxin profile in the Sequim Bay isolates by pre-column oxidation and HPLC-fluorescence detection revealed that gonyautoxin 1 and 4 made up 65 ± 9.7 % of the total PSTs present. Collectively, these data confirm that extracellular PSTs are present during blooms of Alexandrium spp. in the Sequim Bay region.     

Biological Control of Sclerotinia Diseases of Rapeseed by Aerial Applications of the Mycoparasite Coniothyrium minitans

Indoor and field experiments were conducted to evaluate the efficacy of applying the mycoparasite Coniothyrium minitans to the aerial parts of rapeseed plants at the flowering stage to control sclerotinia diseases caused by Sclerotinia sclerotiorum. Under controlled conditions, a petal inoculation technique was used to determine the effect of conidial suspensions of C. minitans on suppression of sclerotinia leaf blight. Results showed that C. minitans was effective in inhibiting infection initiated by ascospores of S. sclerotiorum on flower petals by restricting mycelial growth of the pathogen. Suppression of lesion development was related to the conidial concentration of C. minitans, with larger lesions at low concentration (5×10³conidia ml⁻¹), but smaller lesions at high concentration (5×10⁴ conidia ml⁻¹ or higher). When C. minitans-treated rapeseed leaves were inoculated with mycelia of S. sclerotiorum, C. minitans failed to prevent infection of leaves, but caused a significant reduction in number of sclerotia produced on the diseased leaves. No significant difference in efficacy was detected between the two isolates of C. minitans, LRC 2137 and Chy-1, on the two rapeseed cultivars, Westar (spring type) and Zhongyou 821 (winter type). Results of field trials showed a significant reduction of stem rot of rapeseed in four (1997, 1999, 2003 and 2004) out of five years by aerial application of C. minitans, compared with controls. No significant difference in suppressive efficacy was observed between the treatments of C. minitans (10⁶ conidia ml⁻¹), C. minitans (10⁶ conidia ml⁻¹) + benomyl (50 μg ml⁻¹) and benomyl (100 μg ml⁻¹) in 2003, and between the treatments of C. minitans (10⁶ conidia ml⁻¹), C. minitans (10⁶ conidia ml⁻¹) + vinclozolin (100 μg ml⁻¹) and vinclozolin (500 μg ml⁻¹) in 2004. Sclerotia of S. sclerotiorum collected from diseased plants in plots treated with C. minitans in 1999, 2000 and 2003, or with C. minitans + benomyl in 2003 were infected by C. minitans at frequencies ranging from 21.3 to 54.5%. This study concludes that aerial spraying of C. minitans is an effective method for controlling sclerotinia diseases of rapeseed.     

Do Thoroughbred and Standardbred horses have similar increases in pulmonary vascular pressures during exertion?

To test the hypothesis that the pulmonary vascular pressures of Thoroughbred and Standardbred horses behave similarly during exertion. Measurements were made on 5 Thoroughbred and 5 Standardbred horses on a treadmill at rest and during 3-minute exercise intervals at speeds predicted to produce 75%, 90%, and 100% maximal heart rate. Left forelimb acceleration, heart rate, esophageal pressure, and pulmonary artery pressure were measured continuously. Pulmonary capillary and wedge pressures were measured during intermittent occlusion of the pulmonary artery. Breathing rate and gait frequency were the fundamental frequencies of the esophageal pressure and limb acceleration signals respectively. The ratio of speed:gait frequency gave stride length. The effects of exertion and breed were evaluated using two-way analysis of variance. Exertion produced significant increases in pulmonary artery (P = 0.001), capillary (P= 0.002), and wedge (P= 0.005) pressures. No significant effect of breed was detected on pulmonary artery pressure, but at exertion pulmonary capillary and wedge pressures were 15% (P= 0.03) and 23% (P= 0.04) greater in Thoroughbreds, respectively. Treadmill speed was ~12% greater (P= 0.04), stride length was ~25% greater (P= 0.0003), gait frequency was ~10% less (P= 0.006), breathing rate was ~10% less (P= 0.001), and heart rate was ~6% less (P= 0.06) for Thoroughbreds. There was no effect of breed on inspiratory or expiratory esophageal pressure although mean esophageal pressure was ~2 mmHg greater (P= 0.03) in exercising Standardbreds. In conclusion, pulmonary capillary and wedge pressures are greater in Thoroughbreds than in Standardbreds at similar fractions of maximal heart rate. This is compatible with the higher incidence of exercise-induced pulmonary hemorrhage observed in Thoroughbreds.     

Monoclonal antibodies for coagglutination of Streptococcus suis type 1

Monoclonal antibodies (MAbs) were sought that would identify Streptococcus suis. Spleen cells from Balb/c mice immunized with formalin-killed S. suis were fused with the murine cell-line SP-2/0. Hybridomas positive for S. suis were identified by a solid-phase radioimmunoassay with a mixture of all available serotypes of S. suis. On further screening with individual serotypes of S. suis (Types 1,2,1/2,3,4,5,7,8,9 and 12) and other related and unrelated bacteria, a hybridoma specific for Type 1 was identified. Another hybridoma was positive for all serotypes of S. suis. S. suis Type 1-specific MAbs in the form of ascites fluid was used in a coagglutination test. Agglutination with S. suis Type 1 was observed within 45 s, while there was no reaction with other serotypes of S. suis or other bacteria, during the 5-min observation period. A library of type-specific MAbs should help in rapid and accurate serotyping of S. suis isolates.     

The impact of direct‐fed microbials and enzymes on the health and performance of dairy cows with emphasis on colostrum quality and serum immunoglobulin concentrations in calves

Thirty‐six cows were blocked by calving date and randomly assigned to one of three treatments. Cows were on treatments 3 weeks prepartum through 8 weeks post‐partum. Treatments were as follows: (i) no direct‐fed microbial (DFM) or cellulase and amylase enzymes (C), (ii) 45.4 g/day of DFM (D) or (iii) 45.4 g/day of DFM and 18.2 g/day of enzyme (DE). Total mixed ration fed and refused were measured daily to determine dry matter intake (DMI). Blood samples were taken three times weekly and analysed for β‐hydroxybutyrate, glucose and non‐esterified fatty acids. Body weight (BW) was measured weekly. Colostrum was weighed and analysed for IgA and IgG concentration. Calves were fed 4 L of colostrum within 2 hr of birth. Calf blood samples were taken at 0 and 24 hr for analysis of IgA and IgG concentrations and apparent efficiency of absorption. Milk yield was measured daily and samples collected weekly. Initial BW was different among treatments with D being lesser than C or DE treatments. Body weight, weight gain, efficiency of gain, DMI and blood parameters were unaffected. Treatment did not affect colostrum yield. Ash percentage of colostrum tended to increase with D and DE, while IgA and total solids yield decreased with D. Colostrum fat yield was decreased in D and DE. Treatments did not impact BW, serum IgA and IgG concentrations or apparent efficiency of absorption of calves. Post‐partum BW, DMI, blood parameters, milk production and composition were unaffected by treatment. However, cows on D gained more BW and tended to have greater efficiency of gain compared to those on DE, but were similar to C. Somatic cell scores were greatest for D. Results indicate that DFM and enzyme supplementation did not improve health and performance of dairy cattle during the pre‐ and post‐partum periods under conditions of this study.     

Do Abrasion- or Temperature-Based Techniques More Effectively Relieve Physical Dormancy in Seeds of Cold Desert Perennials?

Seed dormancy can present a significant barrier to restoration outcomes in dryland systems. Physical and combinational (physical + physiological) dormancy are prevalent among seeds of many herbaceous perennials used in restoration of drylands throughout the western United States. Although many techniques designed to alleviate these dormancy traits exist, their efficacy is species specific, may result in embryo damage, and may have limited large-scale application. To identify the most effective means of dormancy alleviation with the potential to be used on an operational scale, we examined the effects of 16 temperature-based techniques (altering temperature and duration of wet heat, freezing and wet heat, and freeze-thaw cycles) and 6 abrasion-based techniques (altering pneumatic scarification length or using a single duration of manual scarification) on the enhancement of seed permeability among two physically dormant (western prairie clover [Dalea ornata {Douglas} Eaton & Wright] and Munro’s globemallow [Sphaeralcea munroana {Douglas} Spach]) and two combinationally dormant species (basalt milkvetch [Astragalus filipes Torr. ex A. Gray] and longspur lupine [Lupinus arbustus {Douglas} ex Lindl.]). We first assessed seed imbibition following exposure to all temperature- and abrasion-based techniques to identify those most successful at promoting seed permeability and then evaluated the effectiveness of those techniques through a series of germination experiments. For combinationally dormant species, we also tested whether exposure to GA₃ enhanced germination. Abrasion-based techniques were more effective than temperature-based techniques at improving water uptake across all species. Pneumatic scarification significantly improved germination, but optimal treatment durations were species specific. GA₃ did not enhance germination under the tested conditions. We conclude that pneumatic scarification is a fast, safe, and effective method for alleviating physical seed dormancy with a potential to be scaled up for operational use in restoration.     

Effect of fertilization on growth and ectomycorrhizal development of container-grown and bare-root nursery conifer seedlings

The effect of three levels of fertilizer on thegrowth of three species of containerized-grownconifer seedlings (Pinus contorta, Picea glauca, and Picea mariana) and twospecies of bare-root conifer seedlings (Pinus sylvestris and Larix sibirica),and on the colonization of these seedlings bysix species of ectomycorrhizal fungi (Hebeloma longicaudum, Laccaria bicolor,Paxillus involutus, Pisolithustinctorius, Rhizopogon vinicolor andSuillus tomentosus), was studied. Thegrowth of the seedlings in both container-grownand bare-root nurseries increased as the levelsof fertilizer increased. For better seedlinggrowth and environmental quality it may be possible to reduce the level of fertilizers in commercial nurseries upto 33% by using selected mycorrhizal fungi.Ectomycorrhizal colonization in all seedlingswas not affected by fertilizer levels. Hebeloma longicaudum, L. bicolor, P.involutus, and P. tinctorius formedwell-developed ectomycorrhizae, whereasectomycorrhizal development by R.vinicolor and S. tomentosus was poor.Native mycorrhizal fungi colonizednon-inoculated control seedlings; however,their colonization was always lower than withinoculated fungi.