Interaction of four monoterpenes contained in essential oils with model membranes: implications for their antibacterial activity

The present article reports the antimicrobial efficacy of four monoterpenes (thymol, carvacrol, p-cymene, and gamma-terpinene) against the Gram-positive bacterium Staphylococcus aureus and the Gram-negative bacterium Escherichia coli. For a better understanding of their mechanism of action, the damage caused by these four monoterpenes on biomembranes was evaluated by monitoring the release, following exposure to the compounds under study, of the water-soluble fluorescent marker carboxyfluorescein (CF) from large unilamellar vesicles (LUVs) with different lipidic composition (phosphatidylcholine, PC, phosphatidylcholine/phosphatidylserine, PC/PS, 9:1; phosphatidylcholine/stearylamine, PC/SA, 9:1). Furthermore, the interaction of these terpenes with dimyristoylphosphatidylcholine multilamellar vesicles as model membranes was monitored by means of differential scanning calorimetry (DSC) technique. Finally, the results were related also with the relative lipophilicity and water solubility of the compounds examined. We observed that thymol is considerably more toxic against S. aureus than the other three terpenes, while carvacrol and p-cymene are the most inhibitory against E. coli. Thymol and carvacrol, but not gamma-terpinene and p-cymene, caused a concentration-dependent CF leakage from all kinds of LUVs employed; in particular, thymol was more effective on PC and PC/SA LUVS than on PC/PS vesicles, while carvacrol challenge evoked a CF leakage from PC/PS LUVs similar to that induced from PC/SA LUVs, and lower than that measured with PC vesicles. Concerning DSC experiments, these four terpenes caused a decrease in Tm and (especially carvacrol and p-cymene) DeltaH values, very likely acting as substitutional impurities. Taken together, our findings lead us to speculate that the antimicrobial effect of thymol, carvacrol, p-cymene, and gamma-terpinene may result, partially at least, from a gross perturbation of the lipidic fraction of the plasmic membrane of the microorganism. In addition to being related to the physicochemical characteristics of the compounds (such as lipophilicity and water solubility), this effect seems to be dependent on the lipidic composition and net surface charge of the microbic membranes. Furthermore, the compounds might cross the cell membranes, thus penetrating into the interior of the cell and interacting with intracellular sites critical for antibacterial activity.     

Variation of DNA methylation and phenotypic traits following unilateral sexual polyploidization in <Emphasis Type="Italic">Medicago</Emphasis>

Sexual hybridization is an important generator of biodiversity and a powerful breeding tool. Hybridization can also overcome ploidy barriers when it involves 2n gametes, as in the case of unilateral sexual polyploidization (USP) that has been utilized in several crops, among which alfalfa. This research was aimed at gaining insights into the effects of USP on genome methylation and on phenotypic traits in alfalfa, an important forage species. The Methylation-Sensitive Amplified Polymorphism technique was used to estimate the cytosine methylation changes occurring in a tetraploid (2n = 4x = 32) USP progeny from crosses between a diploid Medicago sativa subsp. falcata genotype that produces 2n eggs and a cultivated tetraploid Medicago sativa subsp. sativa variety. De novo methylation or demethylation in the USP progeny were observed for 13% of the detected genomic sites, indicating that methylation changes can be relevant. USP plants showed larger surface area of the leaf epidermis cells than both parents, but this did not result in larger leaf size or higher plant biomass. They displayed significant higher ovule sterility than the tetraploid parent, but normal fertility was observed in crosses with unrelated male testers. We conclude that hybridization and sexual polyploidization resulted in novel variation in terms of remodeling of the methylation landscape as well as changes in phenotypic traits in alfalfa.     

Lipid Utilization Pathways Induced by Early Training in Standardbred Trotters and Thoroughbreds

Controversial results on lipid utilization as an energy source during training in horses are found in the literature. The objective of this study was to assess blood lipid profile during different training programs in horses. Seventeen Standardbred horses (400 ± 50 kg) and 17 Thoroughbred horses (380 ± 15 kg) followed different training programs. Blood lipid profile, including triglycerides (TGs), total cholesterol, high-density lipoprotein (HDL), and low-density lipoprotein (LDL), was analyzed, and very-low-density lipoprotein (VLDL) concentration was calculated. Data were analyzed using Student t test, and linear regressions were done. Cholesterol and LDL decreased during training programs in Standardbred trotters (P = .0001 and P = .0053, respectively), and VLDL was found to be close to the significance level (P = .053). Blood lipid profile, including TGs (P = .0014), cholesterol (P = .0057), HDL (P = .0128), LDL (P = .0091), and VLDL (P = .0014), varied throughout the training program in Thoroughbred horses. Negative slope of blood lipids and positive slope of TG linear regression in Standardbred trotters were significant for all parameters (P ≤ .05), whereas cholesterol and LDL regression showed poor P and r² values and HDL P value was slightly above the significance level (P = .069) in Thoroughbred horses. TGs and VLDL showed a positive linear trend in Thoroughbred horses (P = .002). Exercise and different training programs lead to significant variations of lipid profile and lipid utilization in horses. Lipid utilization as an energy source improved with training in Standardbred trotters, whereas this was not the case in Thoroughbred horses. Further studies on the effect of training programs using different conditions and horse breeds would be necessary to understand lipid utilization as an energy source in athletic horses.     

Effect of fertilization of a biochar and compost amended technosol: Consequence on Ailanthus altissima growth and As- and Pb-specific root sorption

The remediation of metal(loid) polluted soil using plants (i.e. phytoremediation) often requires the application of amendments, as well as chemical fertilizer. However, such fertilizers can have negative effects when applied alone and can thus be applied together with other organic amendments to diminish this negative effect. Finally, plants to be used in phytoremediation should be selected based on their adaptive capacity and tolerance to poor and highly contaminated soils, characteristics that possesses Ailanthus altissima. The objective of this study was to evaluate the effects of osmocote fertilization on the amended mining technosol properties and plant growth parameters, as well as to study the accumulation pattern of As and Pb in plant roots. Results showed that osmocote ameliorated soil conditions, but increased Pb mobility. It also greatly improved plant growth. Finally, different behaviours of metal(loid) accumulation were observed in the roots: As was absorbed mainly in the roots because of its similarity with P, with very low amounts in the leaves, whereas Pb was adsorbed mainly on the root surface, with lesser proportion absorbed inside the root system.     

Interactions between temperature and size on the growth,size heterogeneity,mortality and cannibalism in cultured larvae and juveniles of the Asian catfish,Pangasianodon hypophthalmus (Sauvage)

Temperature effects vary throughout the ontogeny, but are proportionally more variable during the early life stages than in older fish. The larvae of a few species have been studied but contrasting trajectories were observed, thereby making it difficult to predict how temperature impacts on the growth, survival and size heterogeneity in a particular species. This study examined these interactions in young [0.9 mg–1 g wet mass (WM)] Pangasianodon hypophthalmus, one of the most extensively cultured tropical fish. Fish were raised at five temperatures from 23 to 33 °C in a recirculated water system, fed in excess with a high‐energy feed (Artemia then a formulated feed, >50% protein), and examined at 1‐day (first feeding days) or 4‐day intervals (older fish). The temperature that produced the fastest growth (T°_opt) was 31 °C at the start of exogenous feeding (0.9 mg); it increased to 32.7 °C at 8 mg then decreased by 0.7 °C for each 10‐fold increase of WM. Size heterogeneity was lower and survival was higher (70–85% from 0.9 mg to 1 g) when the temperature was close to T°_opt. Comparisons with other species suggest that the slope of the relationship between T°_opt and fish size has a strong latitudinal component, and may also reflect the fish sensitivity to oxygen.     

Epidemiology of bovine tick-borne diseases in southern Italy

This investigation was carried out in an area covering part of three southern Italian regions: Campania, Basilicata and Apulia. Eighty-one farms were involved using the formula suggested by Thrusfield; they were equally distributed over the area which was subdivided into 81 geo-referenced sub-areas. In May and June 1999 from a total of 506 cattle, older than 18 months, blood-samples were taken and ticks were collected and identified. Serum samples were tested for antibodies of Bahesia bigemina, Babesia bovis and Anaplasma marginale with an ELISA technique. Eight farms (9.8%) out of the 81 examined were positive for B. bigemina only, 3 (3.7%) for A. marginale only, and 70 (86.4%) for both. None of the animals of any farm was found to be positive for B. bovis. Out of the 506 sera tested, 117 (23.1 %) were positive for B. bigemina only, 58 (11.5%) forA. marginale only and 250 (49.4%) for both species; 81 (16.0%) were negative for all of them. Ticks were collected on animals on 62 (76.5%) out of the 81 farms. Adult ticks (1 410) were collected and identified; the highest number belonged to the Rhipicephalus bursa species (65.5%), followed by Rhipicephalus turanicus (8.6) and Haemaphysalis punctata (8.4). The results showed that B. bigemina, A. marginale and their potential vectors are common in the area examined and indicated that there is a risk for animals imported from tick-borne disease-free areas.     

Copromicroscopic and molecular assays for the detection of cancer-causing parasitic nematode Spirocerca lupi

Spirocerca lupi (Nematoda, Spirurida) is a life-threatening parasitic nematode of dogs that is presently emerging in several countries. Nonetheless, canine spirocercosis is neglected and underestimated, mainly due to diagnostic limitations inherent to clinico-pathologic, diagnostic imaging and laboratory methodologies. Given the significant benefit of improved diagnosis, the present work evaluated the reliability of a recently described copromicroscopic approach, the FLOTAC technique, as well as a PCR-based assay with that of traditional coproscopic techniques to diagnose S. lupi infection. Ninety-four faecal field samples were collected from two endemic areas (i.e. 29 and 65 from Kenya and Israel, respectively) and processed using different coproscopic examination techniques. In particular, set I (Kenyan samples) comprised the modified flotation with Sheather's sugar solution and merthiolate-iodine-formalin technique, while set II (Israeli samples) comprised a flotation technique with zinc sulphate solution, a modified sugar flotation procedure and the FLOTAC method. All samples were also subjected to a semi-nested PCR protocol specific for a region internal to the mitochondrial cytochrome c oxidase subunit 1 gene of S. lupi. The coproscopic examinations showed low sensitivity and high variability, demonstrating the unreliability of the conventional methods for detecting S. lupi eggs. Nonetheless, the FLOTAC technique scored the highest number of positives and significantly higher number of S. lupi eggs per microscopic field compared to the other coproscopic methods. Additionally, of the coproscopically negative samples, 9 (45%) Kenyan and 21 (38.2%) Israeli samples scored molecularly positive using the PCR-based approach. The potential implications and perspectives for canine spirocercosis of these coproscopic and molecular diagnostic methodologies evaluated herein are discussed.