Genetic typing of classical swine fever virus

Three regions of the classical swine fever virus (CSFV) genome that have been widely sequenced were compared with respect to their ability to discriminate between isolates and to segregate viruses into genetic groups. Sequence data-sets were assembled for 55 CSFVs comprising 150 nucleotides of the 5' non-translated region, 190 nucleotides of the E2 envelope glycoprotein gene and 409 nucleotides of the NS5B polymerase gene. Phylogenetic analysis of each data-set revealed similar groups and subgroups. For closely related viruses, the more variable or larger data-sets gave better discrimination, and the most reliable classification was obtained with sequence data from the NS5B region. No evidence was found for intertypic recombination between CSFVs. A larger data-set was also analysed comprising 190 nucleotides of E2 sequence from 100 CSFVs from different parts of the world, in order to assess the extent and global distribution of CSFV diversity. Additional groups of CSFV are evident from Asia and the nomenclature of Lowings et al. (1996) [Lowings, P., Ibata, G., Needham, J., Paton, D., 1996. J. Gen. Virol. 77, 1311-1321] needs to be updated to accommodate these. A tentative assignment, adapting rather than overturning the previous nomenclature divides CSF viruses into three groups with three or four subgroups: 1.1, 1.2, 1.3; 2.1, 2.2, 2.3; 3.1, 3.2, 3.3, 3.4. The expanding data-base of CSFV sequences should improve the prospects of disease tracing in the future, and provide a basis for a standardised approach to ensure that results from different laboratories are comparable.     

Marine Cyclic Peptides: Antimicrobial Activity and Synthetic Strategies

Oceans are a rich source of structurally unique bioactive compounds from the perspective of potential therapeutic agents. Marine peptides are a particularly interesting group of secondary metabolites because of their chemistry and wide range of biological activities. Among them, cyclic peptides exhibit a broad spectrum of antimicrobial activities, including against bacteria, protozoa, fungi, and viruses. Moreover, there are several examples of marine cyclic peptides revealing interesting antimicrobial activities against numerous drug-resistant bacteria and fungi, making these compounds a very promising resource in the search for novel antimicrobial agents to revert multidrug-resistance. This review summarizes 174 marine cyclic peptides with antibacterial, antifungal, antiparasitic, or antiviral properties. These natural products were categorized according to their sources—sponges, mollusks, crustaceans, crabs, marine bacteria, and fungi—and chemical structure—cyclic peptides and depsipeptides. The antimicrobial activities, including against drug-resistant microorganisms, unusual structural characteristics, and hits more advanced in (pre)clinical studies, are highlighted. Nocathiacins I–III (91–93), unnarmicins A (114) and C (115), sclerotides A (160) and B (161), and plitidepsin (174) can be highlighted considering not only their high antimicrobial potency in vitro, but also for their promising in vivo results. Marine cyclic peptides are also interesting models for molecular modifications and/or total synthesis to obtain more potent compounds, with improved properties and in higher quantity. Solid-phase Fmoc- and Boc-protection chemistry is the major synthetic strategy to obtain marine cyclic peptides with antimicrobial properties, and key examples are presented guiding microbiologist and medicinal chemists to the discovery of new antimicrobial drug candidates from marine sources.     

Evidence for a relationship between bovine erythrocyte lipid membrane peculiarities and immune pressure from ruminal ciliates

Erythrocytes of bovines and other ruminants have a strikingly anomalous phospholipid composition, with low or absent phosphatidylcholine (PC) together with high sphingomyelin (SM) content. Here, we report the presence in normal bovine serum of high levels of anti-phospholipid antibodies of IgM isotype against, PC and the phosphono analogue of phosphatidylethanolamine, aminoethylphosphonolipid (AEPL), normally produced by rumen ciliates. In contrast, no antibodies were detected against SM or N-acyl-phosphatidylethanolamine (NAPE), the major components of bovine erythrocytes. In addition, we found that exposure of the ciliate Tetrahymena thermophila to bovine serum results in rapid lysis, an effect that was inhibited by adsorption of the serum with SM/AEPL liposomes. Furthermore, incubation with bovine serum had a similar effect on freshly obtained ruminal ciliates, and the lytic activity was eliminated by pre-adsorption of the serum with SM/PE liposomes. The ruminant mode of life with its concomitant ciliate fauna is hereby linked to the peculiar conformation of bovine erythrocyte membranes. We propose that the unique phospholipid composition of bovine erythrocytes appears as an evolutionary adaptation to tolerate the lytic effects of anti-phospholipid antibodies generated against AEPL, a membrane component of the huge mass of ruminal ciliates, necessary commensals of this group of mammals.     

Detection of bovine viral diarrhea virus (BVDV) in seropositive cattle

Detection of bovine virus diarrhoea virus (BVDV) in one vaccinated beef cattle and three non-vaccinated dairy herds was investigated on peripheral blood leukocytes (PBL) with or without previous treatment followed by a capture ELISA (cELISA). Using the combination of PHA and polycation treatment, PBL from 229 seropositive cattle were studied and could be classified in four different states of BVDV infection. Lysed PBL from four animals were directly positive in cELISA (Category I), PBL of 17 animals were positive after PHA stimulation (Category II), 15 animals were positive only after PHA stimulation plus polycation treatment (Category III), while virus could not be detected in 193 seropositive cattle. Wild-type BVDV strains were isolated by co-culture on polycation-treated MDBK cells from 11 of these seropositive animals. BVDV antibodies of these same animals were able to neutralize their own virus, indicating that virus persists in PBL in spite of strain-specific antibodies. No apparent change of leukocyte subpopulations could be detected in any category of virus-positive animals. Thus, BVDV may be present in the PBL of some cattle, even in the presence of a specific active immune response.     

Effect of dietary Schizochytrium microalga oil and fish oil on plasma cholesterol level in rats

The purpose of the study was to test the hypothesis that the dietary oils with different content of n‐3 polyunsaturated fatty acids (PUFA) eicosapentaenoic acid (EPA) + docosahexaenoic acid (DHA) affect plasma lipid level in rats in a different degree. The diets with 6% of fish oil (FO) and Schizochytrium microalga oil (SchO; EPA+DHA content in the diets 9.5 + 12.3 and 2.6 + 29.5% of the sum of total fatty acids, respectively) were used; the diet with 6% of safflower oil (high content of n‐6 PUFA linoleic acid, 65.5%; EPA+DHA content 0.7 + 0.9%) was used as a control. The difference between FO and SchO was established only in the case of plasma triacylglycerol (TAG) level: plasma TAG of the FO‐fed rats did not differ from the control rats (p > 0.05), while SchO decreased (p < 0.05) plasma TAG to 46% of the control. On the other hand, FO and SchO decreased (p < 0.05) total plasma cholesterol (TC) in rats in the same extent, to 73% of the control. Regarding the underlying mechanisms for the TC decrease, both SchO and FO up‐regulated hepatic Insig‐1 gene (181 and 133% of the control; p < 0.05), which tended (p = 0.15 and p = 0.19 respectively) to decrease the amount of hepatic nSREBP‐2 protein (61 and 66% of the control). However, neither SchO nor FO influenced hepatic 3‐hydroxy‐3‐methyl‐glutaryl‐CoA reductase gene expression (p > 0.05); SchO (but not FO) increased (p < 0.05) low‐density lipoprotein receptor mRNA in the liver. It was concluded that the decrease of total plasma cholesterol might be caused by an increased cholesterol uptake from plasma into the cells (in the case of SchO), but also by other (in the present study not tested) mechanisms.     

Studies on the various sugar-urea and sugar-starch-urea relationships with reference to the fermentation in the rumen and the digestibility of nutrients in fattening cattle]

Recommendations are given saying that sugar-to-crude fibre rations of not more than 1.7:1 and sugar-to-urea ratios of 12:1 should be used in the fattening of cattle fed urea-supplemented rations of roughages in which 40% to 50% of the crude protein content of the ration had been replaced by urea. The amount of sugar per kg live-weight should not exceed 4 g. Higher quantities reduce the digestibility of crude fibre and, additionally, decrease the NH3 level and the pH in the rumen. At the same time the amount of volatile fatty acids is increased while the content of ruminal acetic acid decreases. Variations in the sugar-to-starch ratio had no statistically significant influence on the nitrogen balance whereas it was found that increasing quantities of sugar significantly decreased the digestibility of crude fibre. The proportional content of sugar in the readily metabolisable carbohydrates contained in the ration should not exceed 20% to 50%.     

Loquat as a crop model for successful deficit irrigation

During three consecutive seasons, two different deficit irrigation strategies were compared with control fully irrigated trees regarding their capacity to induce early bloom and harvest in “Algerie” loquat. The first strategy, a continuous deficit irrigation strategy, consisted in a uniform reduction of 20% water needs through the entire season; the second strategy, a regulated deficit irrigation approach, while accounting for the same global reduction of 20% loquat water needs, concentrated water shortages after harvest from mid-May through the end of August. Regulated deficit irrigation resulted more successful. Postharvest regulated deficit irrigation advanced full bloom 10–20 days depending on the season. Such enhancement led to more precocious and valuable yield, with an average increase of fruit value of 0.21 € kg⁻¹. The effects of continuous deficit irrigation were less noticeable and average fruit value was increased 0.08 € kg⁻¹. Yield and fruit quality were not affected for the different deficit irrigation strategies. Water savings established around 1450 m³ ha⁻¹ year⁻¹. Deficit irrigation rose water use efficiency up to more than a 40%.     

Subolesin/akirin orthologs from Ornithodoros spp. soft ticks: cloning, RNAi gene silencing and protective effect of the recombinant proteins

Subolesin/akirin is a well characterized protective antigen highly conserved across vector species and thus potentially useful for the development of a broad-spectrum vaccine for the control of arthropod infestations including hard ticks, mosquitoes, sand flies and the poultry red mite Dermanyssus gallinae. Soft ticks could be also targeted by this vaccine if proved that the soft tick subolesin orthologs are conserved and induce protective immune responses too. However, to date no soft tick subolesin orthologs have been fully characterized nor tested as recombinant antigens in vaccination trials. The objectives of the present work were to clone and characterize the subolesin orthologs from two important vector species of soft ticks as Ornithodoros erraticus and O. moubata, to evaluate the effect of subolesin gene silencing by RNAi, and to test the protective value of the recombinant antigens in vaccination trials. The obtained results demonstrate that both soft tick subolesins are highly conserved showing more than 69% and 74% identity with those of hard ticks in their nucleotide and amino acid sequences, respectively. Additionally, we demonstrate that both soft ticks possess fully operative RNAi machinery, and that subolesin gene silencing by dsRNA injection inhibits oviposition indicating the involvement of subolesin in tick reproduction. Finally, vaccination with the recombinant soft tick subolesins induced a partial protective effect resulting in the reduction of the oviposition rate. These preliminary results encourage further studies on the use of recombinant subolesins as vaccines for the control of soft tick infestations, either alone or in combination with other specific molecules.     

Effect of Breeding Activity on the Microflora of the External Genitalia and in the Semen of Stallions,and the Relationship Between Micro‐organisms on the Skin and on the External Genitalia

A possible role of breeding activities in the composition of the microbial population in stallions' external genitalia (EG) and the relationship between micro‐organisms colonizing the skin of the abdomen and the ones colonizing the EG have not been studied. In experiment 1, EG microbiological samples were collected from 41 stallions used for both natural cover and semen collection (BST) and from 18 non‐breeding stallions (NBST). A higher (p < 0.05) frequency of isolation of potentially pathogenic species was found for BST. Age did not influence number of micro‐organism species isolated both in BST and NBST. In experiment 2, the microbial content of the EG and semen was compared in 23 BST. Most micro‐organisms isolated from the EG were present in semen, albeit with a numerically lower prevalence. In 7 stallions, six microbial species isolated from semen were absent from the EG cultures, suggesting contamination by the operator. In experiment 3, a numerically higher number of micro‐organism species was isolated from the EG of 31 stallions, than from their skin of the ventral abdomen in contact with the penis or from the skin of the thorax. With the sole exception of Escherichia coli, potentially pathogenic bacteria were only isolated from the EG but not from the skin. Results suggest that breeding activity increased the number of species colonizing the EG; most species isolated from the EG were also found in semen even if with a lower frequency, and additional semen contamination seemed to occur during its manipulation. Many micro‐organism species of the skin were also isolated from the penis, but independently of being or not in contact with the penis, skin did not seem to provide an adequate environment for the growth of potentially pathogenic bacteria that were isolated from EG, with the sole exception for E. coli.