Seasonal prevalence of fungi in the conjunctival fornix of healthy cows during a 2‐year study

Objective The aims of the present paper were to: (i) identify and quantify conjunctival fungi isolated from healthy cows; (ii) verify the influence of different methods of farm management on the prevalence (percentage of positive cultures for each fungal species per farm) of conjunctival fungi. Material and methods Forty Friesian and twenty Limousin female cows aged 1–10 years stabled in three farms with different managements (farm 1: cows housed strictly indoors; farm 2: cows housed outside during the day and inside the stall during the night; farm 3: cows housed strictly outdoors) were investigated for conjunctival fungal flora. Air and food were also tested. Specimens were collected every season during a 2‐year study. Identification of colonies of filamentous fungi was achieved to the genus level on the basis of macro‐ and microscopic features. Results The total number of eyes positive for fungi ranged from 85 to 100% at farm 1, from 65 to 95% at farm 2, and from 55 to 95% at farm 3. Fungi most frequently isolated from conjunctival fornix were Cladosporium spp. and Penicillium spp. Statistical analysis did not show any differences in fungal prevalences among the three farms during the same season. Some fungal species were consistently isolated while others were intermittently isolated. Conclusions Fungi found in the conjunctival fornix of cows might represent transient seeding from the environment, as suspected in other species. The prevalence of conjunctival fungal organisms is not different in cattle housed indoors vs. outdoors.     

Ocotea quixos Lam. essential oil: In vitro and in vivo investigation on its anti-inflammatory properties

Here we investigated the anti-inflammatory properties of Ocotea quixos essential oil and of its main components, trans-cinnamaldehyde and methyl cinnamate, in in vitro and in vivo models. Ocotea essential oil and trans-cinnamaldehyde but not methyl cinnamate significantly reduced LPS-induced NO release from J774 macrophages at non-toxic concentrations, inhibited LPS-induced COX-2 expression and increased forskolin-induced cAMP production. The essential oil (30–100 mg/kg os) and trans-cinnamaldehyde (10 mg/kg os) in carrageenan-induced rat paw edema showed anti-inflammatory effect without damaging gastric mucosa. In conclusion we provide the first evidence of a significant anti-inflammatory gastro-sparing activity of O.quixos essential oil.     

Induction of hypericins in Hypericum perforatum in response to chromium

Seedlings of Hypericum perforatum were grown with 0.01 and 0.1 mM of chromium added to the nutrient media. A treatment with 0.01 mM Cr(VI) for seven days resulted in an increased production of protopseudohypericin (+135%), hypericin (+38%) and pseudohypericin (+5%). Treatment with 0.1 mM Cr(VI) for two days also caused an increase of protopseudohypericin (+167%), hypericin (25%) and pseudohypericin (+5%). The greatest effect of chromium treatment was observed at a concentration of 0.1 mM for seven days: protopseudohypericin increased +404% and pseudohypericin to +379%. Hypericin was not affected by this treatment.     

Traceability of plant contribution in olive oil by amplified fragment length polymorphisms

Application of DNA molecular markers to traceability of foods is thought to bring new benefit to consumer's protection. Even in a complex matrix such as olive oil, DNA could be traced with PCR markers such as the amplified fragment length polymorphisms (AFLPs). In this work, fluorescent AFLPs were optimized for the characterization of olive oil DNA, to obtain highly reproducible, high-quality fingerprints, testing different parameters: the concentrations of dNTPs and labeled primer, the kind of Taq DNA polymerase and thermal cycler, and the quantity of DNA employed. It was found that correspondence of fingerprinting by comparing results in oils and in plants was close to 70% and that the DNA extraction from olive oil was the limiting step for the reliability of AFLP profiles, due to the complex matrix analyzed.     

Wild artichoke prevents the age-associated loss of vasomotor function

Endothelial dysfunction, which is more often observed in conduit arteries such as the aorta, carotid, femoral, and brachial arteries, is largely due to alterations in cellular signal transduction initiated by an escalating cycle of damage triggered by oxidative stress. This phenomenon is exacerbated in the elderly, where a progressive loss of vascular endothelial function and concurrent loss of vasomotor control is frequent. In a previous study, we demonstrated that the wild artichoke (Cynara cardunculus) is able to increase the production of the vasorelaxant factor nitric oxide by cultured aortic endothelial cells. We now extended that study to verify (1) the vasorelaxant potential of C. cardunculus on isolated rat aortic rings and (2) whether the vasomodulating properties of C. cardunculus are maintained in vivo, after administration to aged rats. The results demonstrate that the wild artichoke and its main components, namely, luteolin and apigenin, improve aortic relaxation when added to the incubation bath. Moreover, the feeding of wild artichoke [10 mg (kg of polyphenols)(-1) day(-1)] to aged rats significantly restores proper vasomotion, to a degree similar to that observed in young animals. This study provides further justification to the advice to consume wild greens as part of a balanced diet and suggests that close attention should be paid to the diet of the elderly, because it can effectively modulate important parameters of cardiovascular risk.     

Limited fluid volume resuscitation (LFVR) in severe shock unresponsive to initial fluid challenge: A pilot study in 10 cats

Objective

To evaluate the effect of limited fluid volume resuscitation (LFVR) administration in cats with severe shock that was unresponsive to initial conventional resuscitation (CR) with isotonic crystalloids.

A Comparative Study on the Flocculation of Silica and China Clay with Chitosan and Synthetic Polyelectrolytes

Flocculation is still one of the most important and efficient processes for water treatment. However, most industrial processes, such as in water treatment plants, still use huge amounts of synthetic polyelectrolytes for the flocculation process. Here we compare the flocculation of two different suspended particles, i.e., silica particles and china clay, with the biopolymer chitosan and two common strong synthetic polyelectrolytes. As a flocculant, chitosan featured a minimum uptake rate of 0.05 mg/g for silica and 1.8 mg/g for china clay. Polydiallyldimethylammonium chloride (PDADMAC) for comparison possessed a minimum uptake rate of 0.05 mg/g for silica and 2.2 mg/g for china clay. Chitosan as an environmentally friendly biopolymer competes with the synthetic polyelectrolytes and thus represents a beneficial economic alternative to synthetic flocculants.     

Differential in Gel Electrophoresis (DIGE) Comparative Proteomic Analysis of Macrophages Cell Cultures in Response to Perthamide C Treatment

Secondary metabolites contained in marine organisms disclose diverse pharmacological activities, due to their intrinsic ability to recognize bio-macromolecules, which alter their expression and modulate their function. Thus, the identification of the cellular pathways affected by marine natural products is crucial to provide important functional information concerning their mechanism of action at the molecular level. Perthamide C, a marine sponge metabolite isolated from the polar extracts of Theonella swinhoei and endowed with a broad and interesting anti-inflammatory profile, was found in a previous study to specifically interact with heat shock protein-90 and glucose regulated protein-94, also disclosing the ability to reduce cisplatin-mediated apoptosis. In this paper, we evaluated the effect of this compound on the whole proteome of murine macrophages cells by two-dimensional DIGE proteomics. Thirty-three spots were found to be altered in expression by at least 1.6-fold and 29 proteins were identified by LC ESI-Q/TOF-MS. These proteins are involved in different processes, such as metabolism, structural stability, protein folding assistance and gene expression. Among them, perthamide C modulates the expression of several chaperones implicated in the folding of proteins correlated to apoptosis, such as Hsp90 and T-complexes, and in this context our data shed more light on the cellular effects and pathways altered by this marine cyclo-peptide.     

Phenotypic and genotypic identification of streptococci and related bacteria isolated from bovine intramammary infections

Background

Streptococcus spp. and other Gram-positive, catalase-negative cocci (PNC) form a large group of microorganisms which can be found in the milk of cows with intramammary infection. The most frequently observed PNC mastitis pathogens (major pathogens) are Streptococcus uberis, Strep. dysgalactiae, and Strep. agalactiae. The remaining PNC include a few minor pathogens and a large nonpathogenic group. Improved methods are needed for the accurate identification and differentiation of PNC.A total of 151 PNC were collected from cows with intramammary infection and conclusively identified by 16S rRNA sequencing as reference method. Nine phenotypic microbiological tests (alpha-hemolysis, CAMP reaction, esculin hydrolysis, growth on kanamycin esculin azide agar and on sodium chloride agar, inulin fermentation, hippurate hydrolysis, leucine aminopeptidase and pyrrolidonyl peptidase activity), multiplex PCR for the three major pathogens (target genes for Strep. uberis, Strep. dysgalactiae and Strep. agalactiae: pauA, 16S rRNA, and sklA3, respectively), and mass spectroscopy using the matrix-assisted laser desorption ionization-time of flight (MALDI-TOF MS) were evaluated for the diagnosis and discrimination of the three clinically most relevant PNC.

Results

The probability that a strain of Strep. uberis, Strep. dysgalactiae and Strep. agalactiae was correctly identified by combining the results of the 9 phenotypic tests was 92%, 90%, and 100%, respectively. Applying the multiplex PCR, all strains of the three major pathogens were correctly identified and no false positive results occurred. Correct identification was observed for all strains of Strep. uberis and Strep. agalactiae using MALDI-TOF MS. In the case of Strep. dysgalactiae, some variability was observed at the subspecies level, but all strains were allocated to one single cluster.

Conclusions

The results of the present study show that reliable identification of the clinically most relevant PNC (Strep. uberis, Strep. agalactiae and Strep. dysgalactiae) can be obtained by use of a combination of colony morphology, hemolysis type and catalase reaction, and a multiplex PCR with specific primers restricted to these 3 pathogens. The MALDI-TOF MS is a fast method that shows promising results, although identification of Strep. dysgalactiae at the subspecies level is not yet satisfactory.     

Effect of different forms of alkali treatment on specific fermentation inhibitors and on the fermentability of lignocellulose hydrolysates for production of fuel ethanol

Treatment with alkali, particularly overliming, has been widely used as a method for the detoxification of lignocellulose hydrolysates prior to ethanolic fermentation. However, the mechanisms behind the detoxification effect and the influence of the choice of cation have not been well understood. In this study, a dilute acid hydrolysate of spruce and an inhibitor cocktail consisting of six known inhibitors were used to investigate different alkali detoxification methods. The various treatments included the addition of calcium hydroxide, sodium hydroxide, potassium hydroxide, and ammonia to pH 10.0 and subsequent adjustment of the pH to 5.5 with either sulfuric or hydrochloric acid as well as treatment with the corresponding amounts of calcium, sodium, and potassium as sulfate or chloride salts at pH 5.5. An RP-HPLC method was developed for the separation of 18 different inhibitors in the hydrolysate, including furaldehydes and phenolics. Detection and quantification were carried out by means of UV, DAD, and ESI-MS in negative mode. Treatment of the spruce hydrolysate with alkali resulted in up to approximately 40% decrease in the concentration of furaldehydes. The effects on the aromatic compounds were complex. Furthermore, SFE was performed on the precipitate formed during alkali treatment to evaluate the inhibitor content of the precipitate, and the following RP-HPLC analysis implied that potential inhibitors were removed mainly through conversion rather than through filtration of precipitate. Parallel experiments in which sulfuric acid or hydrochloric acid was used for acidification to pH 5.5 after alkali treatment indicated that the choice of anion did not affect the removal of inhibitors. Detoxification with calcium hydroxide and ammonia resulted in better fermentability using Saccharomyces cerevisiae than detoxification with sodium hydroxide. The results from the experiments with the inhibitor cocktail indicated that the positive effects of alkali treatment are difficult to explain by removal of the inhibitors only and that possible stimulatory effects on the fermenting organism warrant further attention.