Isolation and characterization of immortalized porcine aortic endothelial cell lines

Primary porcine endothelial cells have a limited life span in culture. After four to five passages, they tend to de-differentiate and eventually reach senescence. The aim of this work was to establish immortalized porcine aortic endothelial cell lines (AOCs) to facilitate in vitro studies of different pathological process involving the endothelium. Primary porcine aortic endothelial cells (PAECs) were transfected with a plasmid containing the SV40 genome and selected on the basis of morphological and phenotypical features. Flow cytometry analysis demonstrated uptake of acetylated low density lipoproteins (Ac-LDL) and constitutive expression of SLA class I, CD29, CD31, CD41/61, CD80/86, CD46, SWC3, and LAMP-1 antigens by all analyzed lines and showed little differences to primary cells. The functional similarity between primary and immortalized endothelial cells was demonstrated in a cytotoxicity assay using a human natural killer cell line (NKL) as effector. The AOCs cell lines should be valuable tools for in vitro study of the human immune response against pig endothelial cells. In addition, they would be very useful to gain insight in the pathogenesis of some viral haemorrhagic diseases of pig such as African swine fever (ASF) or classical swine fever (CSF).     

Osteomalacia as a result of phosphorus deficiency in beef cattle grazing subtropical native pastures in Uruguay

We investigated 2 outbreaks of osteomalacia as a result of phosphorus (P) deficiency in herds of lactating beef cows grazing subtropical native pastures in Uruguay. Cows exhibited pica, difficulty to stand and walk, rib fractures, and body weight loss even with adequate forage availability. Osteopenia and severe osteomalacia were observed on gross and histologic examination. The concentrations of bicarbonate-extractable P in soil (4.0, 4.1 mg P/kg), total P in pasture (0.9, 1.1 g P/kg), inorganic P in serum (1.0, 0.71 mmol P/L), and P in bone (73 mg P/mL) were all low. Although injectable and mineral salt supplements provided additional P in both outbreaks, these supplementary amounts were insufficient to prevent P deficiency. The P ingested by the cows from the pasture and supplements would have provided 20–55% of their daily P requirements of ~21 g P/d. Osteomalacia occurred in cattle at the 2 ranches as a result of severe P deficiency in the soil and forage, and inadequate P supplementation. Following diagnosis, control of P deficiency in beef cattle requires estimation of the amount of pasture P ingested and provision of sufficient additional supplementary P to meet the animals’ requirements.     

Phylogenetic and structural studies of a novel equine papillomavirus identified from aural plaques

Papillomaviruses (PVs) infect a wide range of animal species and show great genetic diversity. To date, excluding equine sarcoids, only three species of PVs were identified associated with lesions in horses: Equus caballus papillomavirus 1 (EcPV1-cutaneous), EcPV2 (genital) and EcPV3 (aural plaques). In this study, we identified a novel equine PV from aural plaques, which we designated EcPV4. Cutaneous samples from horses with lesions that were microscopically diagnosed as aural plaques were subjected to DNA extraction, amplification and sequencing. Rolling circle amplification and inverse PCR with specific primers confirmed the presence of an approximately 8 kb circular genome. The full-length EcPV4 L1 major capsid protein sequence has 1488 nucleotides (495 amino acids). EcPV4 had a sequence identity of only 53.3%, 60.2% and 51.7% when compared with the published sequences for EcPV1, EcPV2 and EcPV3, respectively. A Bayesian phylogenetic analysis indicated that EcPV4 clusters with EcPV2, but not with EcPV1 and EcPV3. Using the current PV classification system that is based on the nucleotide sequence of L1, we could not define the genus of the newly identified virus. Therefore, a structural analysis of the L1 protein was carried out to aid in this classification because EcPV4 cause lesion similar to the lesion caused by EcPV3. A comparison of the superficial loops demonstrated a distinct amino acid conservation pattern between EcPV4/EcPV2 and EcPV4/EcPV3. These results demonstrate the presence of a new equine PV species and that structural studies could be useful in the classification of PVs.     

Induction of an antioxidant enzyme system and other oxidative stress markers associated with compatible and incompatible interactions between chickpea (Cicer arietinum L.) and Fusarium oxysporum f. sp.ciceris

To ascertain if active oxygen species play a role in fusarium wilt of chickpea caused by Fusarium oxysporum f. sp. ciceris, the degree of lipid peroxidation (malondialdehyde formation) and the activity levels of diamine oxidase (DAO), an apoplastic H₂O₂-forming oxidase, and several antioxidant enzymes, namely ascorbate peroxidase (APX), catalase (CAT), glutathione reductase (GR), guaiacol-dependent peroxidase (GPX) and superoxide dismutase (SOD), were determined spectrophotometrically in roots and stems of ‘WR315’ (resistant) and ‘JG62’ (susceptible) chickpea cultivars inoculated with the highly virulent race 5 of the pathogen. Moreover, APX, CAT, GPX and SOD were also analysed in roots and stems by gel electrophoresis and activity staining; and the protein levels of APX and SOD in roots were determined by Western blotting. In roots, infection by the pathogen increased lipid peroxidation and CAT and SOD activities, although such responses occurred earlier in the incompatible compared with the compatible interactions. APX, GPX and GR activities were also increased in infected roots, but only in the compatible interaction. In stems, infection by the pathogen increased lipid peroxidation and APX, CAT, SOD and GPX activities only in the compatible interaction, and DAO activity only in the incompatible one. In general, electrophoregrams agreed with the activity levels determined spectrophotometrically and did not reveal any differences in isoenzyme patterns between cultivars or between infected and non-infected plants. Further, Western blots revealed an increase in the root protein levels of APX in the compatible interaction and in those of SOD in both compatible and incompatible interactions. In conclusion, whereas enhanced DAO activity in stems, and earlier increases in lipid peroxidation and CAT and SOD activities in roots, can be associated with resistance to fusarium wilt in chickpea, the induction of the latter three parameters in roots and stems along with that of APX, GR (only in roots) and GPX (only in stems) activities are rather more associated with the establishment of the compatible interaction.     

A common near infrared—based partial least squares regression model for the prediction of wood density of Pinus pinaster and Larix?×?eurolepis

Wood density is defined as the ratio of mass to volume and therefore in principle it should be possible to calculate a unique partial least squares regression (PLS-R) model for several species. PLS-R models for wood density based on X-ray microdensity data were calculated for each species Pinus pinaster and Larix × eurolepis and for both species together. After cross-validation and test set validation the data sets were combined and final models were calculated. The common model gave a residual prediction deviation (RPD) of 3.1, a range error ratio (RER) of 11.7, and a SEP/SEC of 1.06. The single models for Pinus pinaster and Larix?×?eurolepis gave RPD’s of 3.5 and 3.2, RER’s of 13 and 11, and a SEP/SEC of 1.2. To the best knowledge of the authors all obtained PLS-R models are the first ones that fulfil the requirements according to AACC Method 39-00 (AACC in AACC Method, 39-00:15, 1999) to be used at least for screening (RPD?≥?2.5). Although this method and the defined limits were developed for the analysis of grains they can be used as a rough rule of thumb until limits for wood are available. The improvement of the PLS-R models, compared to published results, might be due to three facts (1) the higher number of scans collected for a single spectrum, (2) that the samples were better represented by the NIR spectra and X-ray microdensity values, and (3) that the sites for the measurement of NIR spectra and X-ray microdensity were coincided as strictly as possibly.     

Ecological and eco‐social models for the introduction of the abalone Haliotis discus hannai into benthic systems of north‐central Chile: sustainability assessment

• 1. Loop models of ecological and socio‐economic systems were developed to analyse and predict the impact of a possible accidental introduction (escapes) of the abalone Haliotis discus hannai into a benthic community of north‐central Chile.
• 2. Although the ‘new’ ecological system resulting from a successful invasion of abalone would be locally stable, the establishment of a self‐enhanced dynamic of recruits would transform this into an unstable system.
• 3. The harvest of the kelp, Lessonia trabeculata and other macroalgae is not recommended because this destabilizes the system. The harvest of abalone adults is only sustainable if they do not exert a negative effect upon other native invertebrates.
• 4. The eco‐social model showed three important results: (1) if the variables ‘Price’ and ‘Farming’ are in expansion and stationary dynamics, then the models were found unstable; (2) a self‐enhanced dynamic of abalone recruits tends toward instability; and (3) the harvest of the kelp L. trabeculata and other macroalgae would be non‐sustainable.
• 5. Based on our results, the sustainable development of extensive farming of H. discus hannai in the sea would be not reached. If it is done, an intensive monitoring of the community after introduction into the system is strongly recommended. Likewise, the farming of macroalgal species (source of food for abalone) should be promoted in order to avoid harvesting of macroalgae from natural systems.

Copyright © 2006 John Wiley & Sons, Ltd.