Exploration of plant genomes in the FLAGdb++ environment

Background

Efficient high throughput screening systems of useful mutants are prerequisite for study of plant functional genomics and lots of application fields. Advance in such screening tools, thanks to the development of analytic instruments. Direct analysis in real-time (DART)-mass spectrometry (MS) by ionization of complex materials at atmospheric pressure is a rapid, simple, high-resolution analytical technique. Here we describe a rapid, simple method for the genetic discrimination of intact Arabidopsis thaliana mutant seeds using metabolic profiling by DART-MS.

Results

To determine whether this DART-MS combined by multivariate analysis can perform genetic discrimination based on global metabolic profiling, intact Arabidopsis thaliana mutant seeds were subjected to DART-MS without any sample preparation. Partial least squares-discriminant analysis (PLS-DA) of DART-MS spectral data from intact seeds classified 14 different lines of seeds into two distinct groups: Columbia (Col-0) and Landsberg erecta (Ler) ecotype backgrounds. A hierarchical dendrogram based on partial least squares-discriminant analysis (PLS-DA) subdivided the Col-0 ecotype into two groups: mutant lines harboring defects in the phenylpropanoid biosynthetic pathway and mutants without these defects. These results indicated that metabolic profiling with DART-MS could discriminate intact Arabidopsis seeds at least ecotype level and metabolic pathway level within same ecotype.

Conclusion

The described DART-MS combined by multivariate analysis allows for rapid screening and metabolic characterization of lots of Arabidopsis mutant seeds without complex metabolic preparation steps. Moreover, potential novel metabolic markers can be detected and used to clarify the genetic relationship between Arabidopsis cultivars. Furthermore this technique can be applied to predict the novel gene function of metabolic mutants regardless of morphological phenotypes.     

Mutations in the juxtamembrane domain of c-KIT are associated with higher grade mast cell tumors in dogs

Mast cell tumors are among the most commonly seen tumors of the skin in dogs and are more highly aggressive than mast cell tumors of other species. Some breeds display a markedly higher incidence of mast cell tumor development than others and appear to have some genetic predisposition. Recently, mutations have been found in canine mast cell tumor tissues and cell lines within the juxtamembrane domain of the protooncogene c-KIT In previous studies utilizing a small number of cases, no association between the presence of a mutation and the breed of dog or grade of the tumor could be identified. An expanded study with a larger sample set was performed to explore this possibility. The juxtamembrane domain of c-KIT was amplified using the polymerase chain reaction from genomic DNA preparations of 88 paraffin-embedded mast cell tumors from selected breeds. Mutations, consisting of duplications and deletions, were found in 12 of the tumors. A significant association was found between the presence of a mutation and a higher grade of tumor but not between breed and grade or between breed and the presence of a mutation.     

A three‐dimensional cell culture system as an in vitro canine mammary carcinoma model for the expression of connective tissue modulators

In this study, derived complex carcinoma (CC) and simple carcinoma (SC) cell lines were established and cultured under two‐dimensional (2D) and three‐dimensional (3D) conditions. The 3D was performed in six‐well AlgiMatrix? (LifeTechnologies®, Carlsbad, CA, USA) scaffolds, resulting in spheroids sized 50–125 µm for CC and 175–200 µm for SC. Cell viability was demonstrated up to 14 days for both models. Epidermal growth factor receptor (EGFR) was expressed in CC and SC in both systems. However, higher mRNA and protein levels were observed in SC 2D and 3D systems when compared with CC (P < 0.005). The connective tissue modulators, metalloproteinases‐1, ‐2, ‐9 and ‐13 (MMPs), relaxin receptors 1 and 2 (RXR1 and RXR2) and E‐cadherin (CDH1) were quantitated. All were upregulated similarly when canine mammary tumour (CMT)‐derived cell lines were cultured under 3D AlgiMatrix, except CDH1 that was downregulated (P < 0.005). These results are promising towards the used of 3D system to increase a high throughput in vitro canine tumour model.     

Assessment of the efficacy of composite surgery for the treatment of dorsal displacement of the soft palate in a group of 53 racing Thoroughbreds (1990-1996)

REASONS FOR PERFORMING STUDY: There has been no objectively assessed case-control study of the efficacy of surgery to correct dorsal displacement of the soft palate (DDSP) previously reported. HYPOTHESIS: Composite surgery has a beneficial result on racing performance in horses affected with DDSP as compared to a matched control population. METHODS: Race records were obtained for 53 racing Thoroughbreds which underwent composite staphylectomy, sternothyrohyoideus myectomy and ventriculectomy for correction of idiopathic DDSP at the University of Bristol between 1990 and 1996. Each surgical case was matched for age, sex and training yard with 2 control horses. The racing performance, based on prize money won, of surgical cases and control horses were compared for 3 races run before and after the date of surgery. RESULTS: Ninety-two percent of the surgical cases returned to racing after surgery. There was a significant increase in earnings of the surgical group before and after surgery (P = 0.011), but there was no significant difference in earnings of the control group before and after the date of surgery (P = 0335). Sixty percent of the surgical group had higher earnings after surgery than before, compared to 40% of controls. When horses which underwent surgery were ranked relative to their 2 matched controls, surgical cases did not significantly change in rank (P = 033), whereas control horses significantly decreased in rank (P = 0.012). Additionally, horses within the surgical group were more likely (P < 0.01) to start in 3 post operative races than those in the control group. CONCLUSIONS AND POTENTIAL RELEVANCE: Composite surgery had a beneficial effect on racing performance of horses afflicted with idiopathic DDSP, and further studies to evaluate objectively the usefulness of other surgical techniques are warranted.     

Vertical Distribution of Arthropod Interactions Within Turfgrass

Arthropod predators are abundant in turfgrass systems, and they play an important role in managing pests. Understanding the vertical distribution of predation is critical to developing cultural strategies that enhance and conserve predatory services. However, little is known on how the predation is vertically distributed within the turfgrass canopy. Thus, the objective of this study was to determine the vertical distribution of predation within the turfgrass canopy. Clay models were used to emulate the general appearance of Noctuidae caterpillars, to estimate the predatory activity. The choice and no-choice experiments were conducted by placing clay models at 2.54, 5.08, and 7.62 cm from the thatch surface and denoted as lower, intermediate, and upper levels, respectively, within turfgrass canopy. The predator-mediated impressions, paired mark, scratch, deep cut mark, deep distortion, prick, dent, stacked surface impression, scooped mark, granulation, and U-shaped mark, were identified on clay models. The incidence and severity of impressions were significantly greater on clay models placed at the lower canopy level than on those placed at the intermediate and upper canopy levels in the choice and no-choice experiments (P < 0.05). Thus, predators are more likely to find their prey at the soil level. This information can be used to refine management strategies, such as mowing height and insecticide use for effectively managing soil-borne and foliar-feeding arthropod pests and beneficial arthropods.     

Validity of a health-related quality-of-life scale for dogs with signs of pain secondary to cancer

OBJECTIVE: To develop and validate a health-related quality-of-life scale for dogs with pain secondary to cancer. DESIGN: Questionnaire development. ANIMALS: 40 healthy dogs with no history or signs of pain, 20 dogs with dermatologic disease but no signs of pain other than mild pruritus, and 20 dogs with cancer. PROCEDURE: Owners of all dogs completed a questionnaire containing 12 questions with 4 options for each question, and a quality-of-life score ranging from 0 to 36 was calculated. Scores for dogs with cancer were compared with scores for healthy dogs and dogs with dermatologic disease. RESULTS: All owners indicated that the questionnaire was easy to complete. Scores for healthy dogs were significantly different from scores for dogs with cancer and scores for dogs with dermatologic disease. Scores for dogs with dermatologic disease were significantly different from scores for dogs with cancer. CONCLUSIONS AND CLINICAL RELEVANCE: Results suggested that a simple questionnaire may be useful in assessing health-related quality of life in dogs with pain secondary to cancer, in that dogs with cancer had significantly lower scores than did healthy dogs and dogs with dermatologic disease.     

Fertility of Holstein cows and heifers submitted to timed artificial insemination and receiving one or two doses (12 h apart) of semen

The objective of this retrospective study was to assess the effect of receiving a single (n = 50,285) or double (n = 4392) artificial insemination (AI), 12 h apart, within a timed artificial insemination protocol on pregnancy per AI (P/AI) in nulliparous heifers (inseminated with either sex-sorted or conventional semen) and pluriparous Holstein cows in a commercial dairy herd. Also, this study aimed to investigate the relationship between temperature-humidity index (THI) and time of the first AI and fertility. Fertility of cows receiving two AI with normothermia (THI <68) was higher (p < .05) than cows receiving a single AI (42.9% vs. 36.4%). P/AI of cows receiving two AI with severe heat stress (THI >85) was higher (p < .05) than cows receiving a single AI (21.0% vs. 12.6%). Regardless of heat stress conditions, applying the first AI in the morning increased (p < .05) P/AI in cows with double AI than in cows whose first AI occurred in the afternoon (38.4 vs. 33.3%). With moderate heat stress, and sexed-sorted semen, P/AI to timed AI was higher (65.0 vs. 51.9%; p < .05) in heifers receiving double AI than those serviced once. It was concluded that double AI, 12 h apart, enhanced fertility at timed AI than herd mates with a single AI, particularly with heat stress at breeding.     

Ovicidal activity of seven <Emphasis Type="Italic">Pochonia chlamydosporia</Emphasis> fungal isolates on <Emphasis Type="Italic">Ascaris suum</Emphasis> eggs

The ovicidal effect of the nematophagous fungus Pochonia chlamydosporia on eggs of Ascaris suum was tested under laboratory conditions. A. suum eggs were plated on 2% water–agar with seven fungal isolates (Isol. 5, Isol. 31, Isol. 1, VC1, Isol. 12, Isol. 22 and VC4) and control without fungus. After 5, 7, 10, 14, 15 and 21 days of incubation, approximately 100 eggs were removed from the plates and classified according to the following parameters: type 1, biochemical and physiological effect without morphological damage to the eggshell, type 2, lytic effect with morphological alteration of the eggshell and embryo and type 3, lytic effect with morphological alteration of eggshell and embryo showing hyphal penetration and internal egg colonization. The isolates effectively destroyed A. suum eggs and all types of effects were observed during the experiment. There was no variation in ovicidal capacity (type 3 effect) among the isolates (p > 0.05) throughout the experiment. After 21 days, isolate 5 showed the highest percentages of type 3 effect (58.33%). The results indicated that P. chlamydosporia (Isol. 5, Isol. 31, Isol. 1, VC1, Isol. 12, Isol. 22 and VC4) can destroy A. suum eggs and is, therefore, a potential biological control agent of nematodes.     

Juvenile mortality in captive lesser kudu (Tragelaphus imberbis) at Basle Zoo and its relation to nutrition and husbandry.

Since 1956, when the Basle Zoo (Switzerland) initiated the breeding of lesser kudu (Tragelaphus imberbis), 43% of the lesser kudu juveniles died before reaching an age of 6 mo. In this study, the objective was to obtain the pathological findings, nutritional history, and family tree information in order to evaluate the influence of husbandry on juvenile mortality in these animals. The main cause of death was white muscle disease (WMD), diagnosed in 14 cases (26%) of the deceased juveniles. Although enclosure size had remained constant and animal accessibility to the public was constantly high, both herd size and juvenile mortality had increased from 1956-2004. The diet consumed by the whole group in 2004 had deficient levels of vitamin E and selenium. The increasing linear trend of the mortality rate since the 1960s was significant, and there was a significant correlation between herd size and overall juvenile mortality. In contrast, there was no correlation between herd size and the occurrence of juvenile mortality associated specifically with WMD. Other investigated factors (sex, inbreeding, and season) had no significant effect on overall mortality up to 6 mo of age or on mortality associated with WMD. These results characterize both a dietary and a husbandry problem, and are supported by a lack of similar juvenile mortality in another facility where the diet was supplemented with vitamin E, animal numbers were kept low, and the enclosure structure offered more retreat options for the animals.