Temporal dynamics of soil nematode communities in a grassland plant diversity experiment

We report here on an 8-year study examining links between plant and nematode communities in a grassland plant diversity experiment, located in the north of Sweden on previous agricultural soil. The examined plots contained 1, 4 and 12 common grassland plant species from three functional groups; grasses, legumes and forbs. The same plant species composition was maintained in the plots through weeding and resowing during the experimental period. The hypotheses were (i) that the nematode community would shift towards a more diverse and mature fauna over time and (ii) that the effects of both plant species identity and plant species richness would increase over time. As hypothesized, the Bongers’ Maturity Index (a measure of nematode responses to disturbance) increased over time, but not nematode diversity. Instead, the nematode community development in the present grassland experiment seemed to be more characterized by shifts in dominance patterns than by colonization of new taxa. Clear temporal trends were found for plant-feeders and Adenophorea bacterial-feeders which increased in abundance over time in almost all plant treatments. Rhabditidae bacterial-feeders decreased in abundance over time, in particular in plots with legumes. Fungal-feeders, omnivores/predators and the two nematode indices PPI (Bongers’ Plant Parasitic Index) and NCR (Nematode Channel Ratio) had significant interactions between plant composition and time giving some support to our second hypothesis. Our results highlight the need for long-term experiments to examine plant species effects on soil fauna, especially on taxa belonging to higher trophic levels. The results also stress the importance of plant composition for belowground soil faunal communities.     

Advanced ultrasound techniques in small animal reproduction imaging

Ultrasonography is the imaging technology of choice for the evaluation of the reproduction system and of pregnancy in both humans and animals. Over the past 10 years, there have been significant technological improvements of the equipment, while new technologies have been developed. Doppler, contrast-enhanced ultrasonography, elastography, and 3D/4D ultrasonography are advanced ultrasound techniques that have been designed as methods to increase the diagnostic sensitivity of two-dimensional (b-mode) ultrasound, and not as stand-alone tests. The basic physics as well as the advantages and limitations of these advanced ultrasound methods are briefly described. In the reproductive diagnostics of small animals, these techniques have gained an increased popularity as proved by the increased publication of several reports that are also briefly summarized in this review. Clinical applicability is to date limited because of a lack of research on the diagnostic value in concrete situations. Future research projects should focus also on standardization of the used techniques, on determination of thresholds to discriminate between healthy or diseases or fertile versus infertile and on the predictive value of advanced ultrasound findings. Continuing development and optimization of different ultrasound techniques as well as the increase of related scientific interest and worldwide research promises that the clinical interest on the use of advanced ultrasound techniques will increase in future.     

Granulosa cells in three-dimensional culture: A follicle-like structure for domestic cat vitrified oocytes

Follicle-like structures are three-dimensional matrices joint with living cells that allow the in vitro development of female gametes in more physiological conditions. They have been shown to be beneficial to fresh oocytes in different species, and in this study, domestic cat (Felis silvestris catus) granulosa cells were used to create a functional follicle-like structure aimed at supporting the in vitro maturation of conspecific vitrified oocytes, key players of fertility preservation programmes that usually struggle to acquire their full developmental competence after warming. Cat granulosa cells were cultured for up to 6 days in three-dimensional barium alginate microcapsules (i.e. follicle-like structures) or in two-dimensional monolayers, and their steroidogenic ability (estradiol and progesterone secretion) was assessed to confirm their functionality. The same systems were used (on day 2 or 6 of granulosa cells culture) for the in vitro maturation (IVM) of Cryotop^® vitrified immature cat oocytes and compared with microdrops of IVM medium without cells (control). Granulosa cells were able to maintain their functionality in vitro in both the conditions, even if with a different extent of hormonal secretion along culture (p = .02). Vitrified oocytes resumed meiosis at higher rates when cultured with 2 days old granulosa cells (p = .03), but full maturation rates slightly raised when granulosa cells were cultured longer, albeit without differences with the control group. This study paved the road for the creation of enriched culture systems in the domestic cat, but innovations are strongly needed for vitrified oocytes that deserve better chances to develop in vitro.     

In vitro and in vivo effects of toceranib phosphate on canine osteosarcoma cell lines and xenograft orthotopic models

Canine osteosarcoma (OSA) is the most common primary malignant bone tumour in dogs, and it has a high metastatic rate and poor prognosis. Toceranib phosphate (TOC; Palladia, Zoetis) is a veterinary tyrosine kinase inhibitor that selectively inhibits VEGFR‐2, PDGFRs and c‐Kit, but its efficacy is not yet fully understood in the treatment of canine OSA. Here, we evaluated the functional effects of TOC on six OSA cell lines by transwell, wound healing and colony formation assays. Subsequently, two cell lines (Wall and Penny) were selected and were inoculated in mice by intrafemoral injection to develop an orthotopic xenograft model of canine OSA. For each cell line, 30 mice were xenografted; half of them were used as controls, and the other half were treated with TOC at 40 mg/kg body weight for 20 days. TOC inhibited cell growth of all cell lines, but reduced invasion and migration was only observed in Penny and Wall cell lines. In mice engrafted with Penny cells and subjected to TOC treatment, decreased tumour growth was observed, and PDGFRs and c‐Kit mRNA were downregulated. Immunohistochemical analyses demonstrated a significant reduction of Ki67 staining in treated mice when compared to controls. The results obtained here demonstrate that TOC is able to slightly inhibit cell growth in vitro, while its effect is evident only in a Penny cell xenograft model, in which TOC significantly reduced tumour size and the Ki67 index without modifying apoptosis markers.     

Response of Cucurbita spp. to tomato leaf curl New Delhi virus inoculation and identification of a dominant source of resistance in Cucurbita moschata

Tomato leaf curl New Delhi virus is an emerging whitefly-borne species of begomovirus in Mediterranean regions that poses a severe threat to cucurbit crops of the genus Cucurbita. Until now, only two sources of resistance have been identified in Cucurbita spp., these being PI604506 (cv. Large Cheese) and PI381814 (Indian landrace), both of C. moschata. The resistance of cv. Large Cheese is conferred by a single recessive gene located on chromosome 8. The objective of the present investigation was to screen for tomato leaf curl New Delhi virus (ToLCNDV) resistance among 105 accessions drawn from five species of Cucurbita and, if high resistance was found in any of them, determine the mode of inheritance. Screening was conducted using whitefly-mediated inoculation on all 105 accessions. The accessions showing some resistance were further screened by mechanical inoculation as well as by quantitative PCR-based diagnostics. The results showed that, overall, the accessions of C. pepo and C. maxima were the most susceptible, those of C. argyrosperma and C. ecuadorensis intermediate, and those of C. moschata most resistant to ToLCNDV. Only one accession of C. moschata, BSUAL-252, originating from Japan, was highly resistant to ToLCNDV, showing no symptoms after either method of inoculation, and absence of virus accumulation. Upon crossing BSUAL-252 with a susceptible accession of C. moschata, BSUAL-265, the resistance was observed to be conferred by a single dominant gene. This gene is not linked to the genomic region on chromosome 8 where the locus of the previously identified recessive gene for ToLCNDV resistance resides.     

Isolation and genetic characterization of Acidovorax avenae from red stripe infected sugarcane in Northwestern Argentina

Red stripe is a bacterial disease of sugarcane causing important economic losses in Argentina that affects 30 % of the milling stems and consequently the juice quality. In this study, sugarcane leaves exhibiting red stripe symptoms were sampled in the 2008–09 growing season from 13 different sugarcane producing areas of Tucumán and Salta (northwest of Argentina). To achieve the identification and characterization of the causal agent of red stripe, bacterial isolation was performed. Species-specific PCR using Oaf1/Oar1 primers allowed the amplification of a fragment of 550 bp from approximately 50 % of the isolates; 16S rDNA sequences analysis displayed a similarity greater than 99 % with Acidovorax avenae subsp. avenae. By means of RAPD-PCR the presence of at least four different biotypes among the analyzed isolates was detected. Results of pathogenicity test allowed us to confirm A. avenae subsp. avenae as the pathogenic agent for red stripe. This study constitutes the first report on the identification and molecular characterization of this plant pathogen from the Argentina sugarcane production areas. The genetic diversity observed among A. avenae is an important factor to be considered to improve an accurate diagnosis and/or the selection of sugarcane tolerant clones.     

Nordic dairy farmers’ threshold for contacting a veterinarian and consequences for disease recording: Mild clinical mastitis as an example

Previous studies have addressed the differences in registered disease incidence between the Nordic dairy disease recording systems. The main objective of this study was to investigate whether Nordic dairy farmers have varying intention to contact a veterinarian the same day as detecting signs of mild clinical mastitis (MCM) in a lactating dairy cow. This is the first, and necessary, step in the process leading to a disease event being recorded. The second objective was to study underlying behavioural components influencing this threshold for action.A questionnaire-based survey was carried out in Denmark, Finland, Norway and Sweden. The questionnaire was based on the Theory of Planned Behaviour from the field of social psychology. After performing qualitative face-to-face elicitation interviews a set of statements about treatment of MCM was identified. These were grouped into behavioural, normative and control beliefs. The most frequently mentioned beliefs were rephrased as questions. Behavioural intention, a proxy for the behaviour of interest, was assessed using case scenarios. The target and eligible herds were in milk recording and had an average herd size of at least 15 cows. The questionnaire was distributed to 400 randomly sampled dairy producers per included country. The response rate was around 50% in all four countries.The hypothesis of differences in behavioural intention between the countries was tested using Wilcoxon's rank-sum tests. Multivariable linear regression was used to estimate the country-specific variability in behavioural intention as explained by attitude, subjective norm or perceived behavioural control alone, or in combination. The Spearman rank correlations between behavioural intention and each belief, weighted by its outcome evaluation or the motivation to comply, were estimated to find the most important drivers, constraints and social referents for the behaviour of interest.There were significant (p < 0.01) differences in behavioural intention between all countries except Denmark and Norway. Swedish farmers had the weakest behavioural intention and Finnish farmers the strongest. Attitude explained most of the variability in behavioural intention in all four countries. The most important driver in all countries was to achieve a quick recovery for the cow.The varying behavioural intention partly explain the differences in completeness of disease data in the Nordic countries: if farmers have different thresholds for contacting a veterinarian the registered incidence of clinical mastitis will be affected. Knowledge about the importance of attitudes and specific drivers may be useful in any communication about mastitis management in the Nordic countries.