Investigation of biofilm production and icaA and icaD genes in Staphylococcus aureus isolated from heifers and cows with mastitis

Biofilm formation and antimicrobial resistance of Staphylococcus aureus are important virulence factors in cases of mastitis in dairy cows. However, few studies have investigated mastitis strains isolated from heifers. Within this context, the objective of the present study was to investigate biofilm formation on Congo red agar, the presence of the icaA and icaD genes by polymerase chain reaction (PCR), and the percentage of in vitro antimicrobial resistance of 110 S. aureus isolates from mammary gland secretions of heifers and cows with mastitis. PCR detected the icaA and icaD genes in 98% and 100% of isolates, respectively. However, only 55.5% of all isolates produced a biofilm on Congo red agar. Antimicrobial susceptibility testing revealed that 47.0% of isolates from heifers and 70.4% of isolates from cows were resistant to at least one of the antimicrobial agents tested. Resistance to penicillin and/or ampicillin was the most frequent (44.5%). These results indicate the need to implement prophylactic and control measures of mastitis for heifers. Heifers and cows can carry resistant strains with the capacity of biofilm production, a fact representing a threat to public health and animal well‐being and generating losses to dairy farmers.     

Impact of short nutrient stimuli with different energy source on follicle dynamics and quality of oocyte from hormonally stimulated goats

The aim of this study was to verify the effect of the energy source for a short‐term diet supplementation on follicular dynamics, ovarian response and oocyte recovery in goats. Thirty Anglo Nubian crossbred does received a diet for 4 weeks to satisfy the nutritional requirements of breeding for adult non‐dairy goats. Seven days prior to oocyte recovery (OR), a group of does (n = 10) was supplemented with ground full‐fat linseed in the diet (Diet A), whereas a second group of does (n = 10) received crude glycerine in the diet (Diet B). The total mixed ration (TMR) diet was maintained as the Control Diet (n = 10). All animals were oestrous‐synchronized by the use of a progesterone insert for 12 days prior to OR. Follicles were stimulated by using pFSH (five 40‐mg/ml doses) during the supplementation time. At OR, follicles were counted and recovered oocytes were classified as viable or degenerated. Follicular dynamics was monitored by ultrasonography, and plasma glucose, cholesterol and triglyceride levels were measured during supplementation. Glucose was higher in Diet B and cholesterol in Diet A. Diet B had a lower proportion of small (<3 mm) and large follicles (≥3 mm; p = 0.01). The follicular growth rate was higher in Diet A (p < 0.01), with follicles emerging in the 5th day of supplementation. No differences were observed for follicles counted and oocytes recovered. Thus, the type of energy source supplemented for a short term was capable to alter the follicular dynamics, without affecting the proportion of morphologically viable oocytes upon recovery.     

Impact of Degree of Ionization and PEGylation on the Stability of Nanoparticles of Chitosan Derivatives at Physiological Conditions

Nowadays, the therapeutic efficiency of small interfering RNAs (siRNA) is still limited by the efficiency of gene therapy vectors capable of carrying them inside the target cells. In this study, siRNA nanocarriers based on low molecular weight chitosan grafted with increasing proportions (5 to 55%) of diisopropylethylamine (DIPEA) groups were developed, which allowed precise control of the degree of ionization of the polycations at pH 7.4. This approach made obtaining siRNA nanocarriers with small sizes (100–200 nm), positive surface charge and enhanced colloidal stability (up to 24 h) at physiological conditions of pH (7.4) and ionic strength (150 mmol L⁻¹) possible. Moreover, the PEGylation improved the stability of the nanoparticles, which maintained their colloidal stability and nanometric sizes even in an albumin-containing medium. The chitosan-derivatives displayed non-cytotoxic effects in both fibroblasts (NIH/3T3) and macrophages (RAW 264.7) at high N/P ratios and polymer concentrations (up to 0.5 g L⁻¹). Confocal microscopy showed a successful uptake of nanocarriers by RAW 264.7 macrophages and a promising ability to silence green fluorescent protein (GFP) in HeLa cells. These results were confirmed by a high level of tumor necrosis factor-α (TNFα) knockdown (higher than 60%) in LPS-stimulated macrophages treated with the siRNA-loaded nanoparticles even in the FBS-containing medium, findings that reveal a good correlation between the degree of ionization of the polycations and the physicochemical properties of nanocarriers. Overall, this study provides an approach to enhance siRNA condensation by chitosan-based carriers and highlights the potential of these nanocarriers for in vivo studies.     

Comparison of Photocatalytic and Photosensitized Oxidation of Paraben Aqueous Solutions Under Sunlight

It is well-established that aquatic wildlife is exposed to natural and synthetic endocrine disrupting compounds which are able to interfere with the hormonal system. Although advanced oxidation processes (AOPs) have shown to be effective, their application is limited by a relatively high operational cost. In order to reduce the cost of energy consumed in the AOPs, widely available solar energy instead of UV light may be applied either as photocatalytic oxidation or as photosensitized oxidation. The main goal of the present study was to investigate the sunlight photodegradation of paraben mixture. Two processes, namely the photocatalytic oxidation with modified TiO₂ nanoparticles and photosensitized oxidation with photosensitive chitosan beads, were applied. The oxidants were identified as singlet oxygen and hydroxyl radicals for photosensitized and photocatalytic oxidation, respectively. The toxicity, as well as ability to water disinfection of both processes under natural sunlight, has been investigated. Application of sunlight for the processes led to degradation of parabens. The efficiency of both processes was comparable. Despite the fact that singlet oxygen is weaker oxidant than hydroxyl radicals, the photosensitized oxidation seems to be more promising for wastewater purification, due to the possibility of chitosan bead reuse and more effective water disinfection.

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Effect of spores of saprophytic fungi on phytoalexin accumulation in seeds of frog-eye leaf spot and stem canker-resistant and -susceptible soybean (Glycine max L.) cultivars

Two saprophytic fungi (Mucor ramosissimus and Rhizopus sp.) were tested for their ability to induce phytoalexin production by seeds of frog-eye leaf spot and stem canker-resistant and -susceptible soybean (Glycine max L.) cultivars. Only M. ramosissimus was shown to elicit a response and qualitative differences in phytoalexin accumulation were found between the susceptible and resistant cultivars. Glyceollins I, II, and III and glycinol were isolated from the susceptible cultivar, whereas Glyceollins I, II, and III, glycinol, glyceocarpin, genistein, isoformononetin, and N-acetyltyramine accumulated in the resistant cultivar in response to the same fungal elicitor. Genistein was found to be an inducibly formed isoflavonoid instead of a constitutive metabolite in the resistant cultivar, whereas N-acetyltyramine is described for the first time as a soybean phytoalexin. All the compounds, except genistein, showed fungitoxic activity against Cladosporium sphaerospermum. Spectral data of the pterocarpan phytoalexins, genistein, and N-acetyltyramine are also given in this work.     

Effect of top-dressed potassium fertilization on the yield and quality of cucumber

The current study aimed to evaluate the effect of top-dressed potassium (K) application on the production of hybrid cucumber “Sapphire.” The experimental design was a randomized complete block, with five blocks of 0.80 × 0.40 m² each and eight replicate plants per block. The five fertilization rates of K used were 0, 45, 90, 135, and 180 kg K₂O ha^?1. Data collection consisted of the estimation of fruit diameter, fruits length, fruit fresh and dry weights, the number of fruits per plant, and the weight of fruits per plant. The number of fruits per hectare and the fruit fresh weight per hectare were calculated. Fruit tissue was analyzed for determination of macronutrient concentrations, pH, titratable acidity, soluble solids, sugar contents, and protein content. The data were statistically analyzed using regression analysis and analysis of variance (ANOVA). There was a significant effect of the fertilization rate of K on fruit diameter, fruit fresh and dry weights, macronutrients concentrations in fruits, titratable acidity, soluble solids, and reducing sugars. A quadratic equation was adjusted for the number of fruits per plant and per hectare; fruit yield per plant and per hectare, with maximum top-dressing doses, was estimated to be between 60 and 95 kg K₂O ha^?1. A linear increase was obtained in the pH. An increase in the K fertilization rate caused a linear decrease in the fruit length.     

Surface Water Quality Assessment of Lis River Using Multivariate Statistical Methods

This study presents the application of multivariate statistical tools for the evaluation of spatial variations and the interpretation of water quality data obtained in a monitoring program of Lis river basin surface water, Portugal. Twenty-seven physicochemical and microbiological parameters were determined in six water sampling campaigns at 16 monitoring sites during the period from September 2003 to November 2006. Correlation analysis, principal component analysis, and cluster analysis were performed to evaluate the main water pollution sources and to characterize the spatial distribution of water pollution profiles in river basin. The results achieved with the statistical methodologies led to distinguish natural and anthropogenic pollution sources. Additionally, monitoring sites with similar water pollution profile were identified, indicating that some monitoring locations can be changed to improve the spatial characterization of water quality in the river basin. CBO, CQO, P, and N were identified as significant variables affecting spatial variations, namely in the Lis river middle reach. Besides the identification of main pollution sources, the applied statistical tools were able to identify spatial patterns of water pollution in Lis river basin, which further helps in the reassessment of the number and location of monitoring sites.     

Germplasm characterization of Sesbania accessions based on isozyme analyses

Twenty-two accessions belonging to seven species of Sesbania Scop. (S. emerus, S. exasperata, S. grandiflora, S. rostrata, S. sesban, S. tetrapteraand S. virgata), were evaluated in order to characterize the isozyme electrophoretic patterns and to estimate their genetic variability and outcrossing rates. Eight isozyme systems were used: acid phosphatase (ACP), isocitrate dehydrogenase (IDH), phosphoglucomutase (PGM), malate dehydrogenase (MDH), phosphoglucoisomerase (PGI), glutamate oxaloacetate transaminase (GOT), peroxidase (PRX) and catalase (CAT), with two gel/electrode buffers and distilled water as the extraction buffer. Cotyledons were chosen for enzyme extraction. The accessions were compared considering the presence/absence of bands for each system. A dendrogram was obtained using the Jaccard similarity index and the UPGMA clustering method, classifying the accessions into eight groups, evidencing both inter- and intraspecific variability. Annual species were classified in one major group separate from the perennials (S. grandiflora, S. sesban, S. virgata), which were allocated into two other main groups. S. virgata accessions (subgenus Daubentonia) were clearly separated from the other species. Isozyme banding patterns for the accessions are presented for each system. A high incidence of intraspecific monomorphism was observed. Interpretation in terms of loci and alleles was made for S. sesbanand S. virgataaccessions, which presented polymorphism for the systems PGI, GOT (S. sesban) and MDH (S. virgata). The average expected heterozygosity estimates varied from 0.0 to 0.231 for S. sesban and 0.0 to 0.20 for S. virgata. A mean number of 2.0 alleles per locus was observed for both species. Population multilocus outcrossing rate (t _m) of 0.62 was estimated for S. sesban, indicating a partial allogamy mode of reproduction. Individual families presented multilocus rates varying from 0.00 to 1.00, indicating both self-pollination and cross-pollination.