Effect of chelating and antioxidant agents on morphology and DNA methylation in freeze-drying rabbit (Oryctolagus cuniculus) spermatozoa

Freeze-drying (FD) has been exhaustively tried in several mammalian species as an alternative technique to sperm cryopreservation, but few studies have been done in rabbits (Oryctolagus cuniculus). The main objective of this study was to compare the protective effect of various antioxidants added to EDTA medium on structural and functional components of FD rabbit spermatozoa and on their status of global DNA methylation. FD media used were composed of basic FD medium (10 mM Tris-HCl buffer and 50 mM NaCl) supplemented with either 50 mM EDTA alone (EDTA) or added with 105 µM of rosmarinic acid (RA, EDTA-RA) or 10 µM of melatonin (MLT, EDTA-MLT). The effect of each medium on the preservation of FD spermatozoon structure was evaluated with light and scanning electron microscopy (SEM). Global DNA methylation was quantified in all FD sperm samples as well as in fresh spermatozoa. Morphologically, fracture points were evidenced in the neck, mid and principal piece of the spermatozoon tail. No differences in spermatozoon fracture points were evidenced among FD treatments: intact spermatozoa were the largest (p < .01) category, whereas the most frequent (p < .01) injury was the neck fracture, resulting in tailless heads. At SEM, the head of spermatozoa showed a well-conserved shape and intact membrane in all treatments. DNA methylation status was the same in all FD treatments. In conclusion, supplementation of EDTA, EDTA-RA and EDTA-MLT during FD preserved rabbit sperm morphological integrity and methylation status as well. Therefore, the difficulty of getting viable offspring using FD semen is likely unrelated to the impact of the lyophilization process on DNA methylation and morphology of lyophilized spermatozoa.     

Modelling habitat use by the Guiana dolphin,Sotalia guianensis,in south‐eastern Brazil: Effects of environmental and anthropogenic variables,and the adequacy of current management measures

1. Species distribution modelling has been used to identify critical habitats for the delimitation of Marine Protected Areas. Although Marine Protected Areas may often overlap with the distribution of key marine species, illegal human activities often continue within these areas, causing negative impacts on the local biodiversity.
2. A generalized linear model with spatial eigenvector mapping was used to investigate, for the first time, the influence of environmental variables and anthropic activities on the number of Guiana dolphin, Sotalia guianensis sightings/grid, and to determine whether a Marine Reserve, in south‐eastern Brazil, is adequate for the protection of the local population. Data were collected between May 2007 and October 2013 onboard a 7.5‐m vessel with an inboard engine.
3. The models for the different periods (annual, dry, and rainy seasons) all indicated that depth, the distance to seafood farms and fishing grounds, and mean sea surface temperatures influenced the distribution of the dolphins within the study area. The annual and seasonal models predicted that the dolphins prefer a continuous area between Cedro and Pico Islands, a large area that lies outside the limits of the Marine Reserve.
4. Although the habitat of Ilha Grande bay is still much less degraded than that inhabited by other nearby Guiana dolphin populations, the results of the present study indicate that anthropic activities in this area influence habitat use by the dolphins. Less than 30% of the area used by the dolphins is protected by the Tamoios Ecological Station (ESEC Tamoios).
5. Sotalia guianensis is classified as ‘Vulnerable’ in Brazil, although the marine reserve (ESEC Tamoios) does not protect the core area used by the local dolphin population. The results of the present study provide specific locations for the creation of a new multiple‐use MPA, as suggested by the Brazilian National Action Plan for the Conservation of Small Cetaceans, or the inclusion of a special management programme for the area between Cedro and Pico islands to better protect the dolphins in the ESEC Tamoios buffer zone. The continuation of surveys to better understand the current and future impacts of human activities, and the development of a closer interaction with both the local community and local stakeholders will help to safeguard Ilha Grande Bay and the local Guiana dolphin population.

     

Ship noise and cortisol secretion in European freshwater fishes

Underwater noise pollution is a growing problem in aquatic environments and as such may be a major source of stress for fish. In the present study, we addressed the effects of ship noise and continuous Gaussian noise on adrenal activity in three European freshwater species. Underwater ship noise recorded in the Danube River and two Austrian lakes was played back to fish at levels encountered in the field (153 dB re 1 μPa, 30 min). Post exposure cortisol secretion was compared with control situations. Cortisol was measured with enzyme immunoassay techniques (EIA, ng cortisol/l water/g fish) in extracted aquarium water with corrections for fish mass. In the first series, two hearing specialists, the common carp (Cyprinus carpio) and the gudgeon (Gobio gobio) and one hearing generalist, the European perch (Perca fluviatilis) were exposed to ship noise. The noise level was well above hearing thresholds in these species. In a second series, fish were exposed to continuous Gaussian noise at a similar level (156 dB) which is known to induce temporary hearing loss in hearing specialists. All three species responded with increased cortisol secretion when exposed to ship noise. Interestingly, no elevation was observed when fish were exposed to continuous Gaussian noise. Our results indicate that ship noise characterized by amplitude and frequency fluctuations, constitutes a potential stressor in contrast to continuous noise. Surprisingly, the data also demonstrate no apparent differences between species possessing excellent hearing abilities (hearing specialists) and species with poor hearing abilities like perch.     

Antioxidant mechanisms of caseinophosphopeptides and casein hydrolysates and their application in ground beef

Caseinophosphopeptides (CPP) and casein hydrolysates have been shown to bind prooxidant metals such as iron, but their effectiveness as metal chelators to inhibit lipid oxidation in foods has still not been fully investigated. Thus, the antioxidant activity of CPP and casein hydrolysates was studied in phosphatidylcholine liposome model systems. CPP (< 1.0 mg/mL) and casein hydrolysates (0.3-1.7 mg/mL) were effective inhibitors of TBARS development when oxidation was promoted by ferric/ascorbate. High amounts of CPP (> 1.0 mg/mL) were prooxidant, whereas casein hydrolysates were observed to be only antioxidative. In the presence of peroxyl radicals, casein hydrolysates were more effective scavengers than enriched CPP (3-15 mM). In cooked ground beef, TBARS formation was inhibited 75, 39, and 17% by 0.5% enriched CPP, casein hydrolysates, and low molecular weight casein hydrolysates, respectively, after 4 days of storage. The results show that CPP and casein hydrolysates are promising sources of natural antioxidants for foods.     

Monoclonal antibodies against the leptospiral immunoglobulin-like proteins A and B conserved regions

Leptospirosis is an infectious disease caused by pathogenic spirochetes of the genus Leptospira that affects humans and a wide variety of animals. Recently the genomes of Leptospira interrogans, Leptospira borgpetersenii and Leptospira biflexa species were sequenced allowing the identification of new virulence factors involved in survival and pathogenesis of bacteria. LigA and LigB are surface-exposed bacterial adhesins whose expression is correlated with the virulence of Leptospira strains. In this study, we produced and characterized five monoclonal antibodies (MAbs) against a recombinant fragment of LigB (rLigBrep) with approximately 54 kDa that comprise the portions of LigA and LigB (domains 2–7). The 5 MAbs obtained were of the IgG1 (2) and IgG2b (3) isotypes and their affinity constants for rLigBrep ranged from 7 × 10⁷ M⁻¹ to 4 × 10⁸ M⁻¹. The MAbs were able to react with the native antigen on the L. interrogans, L. borgpetersenii and Leptospira noguchii surfaces by indirect immunofluorescence, immunoblotting and immunoelectron microscopy. These results demonstrate that the MAbs anti-rLigBrep can be useful to complement genetic studies and to aid studies aiming understanding the role of Lig proteins in Leptospira pathogenesis and the development of Lig-based vaccines and improved diagnostic tests for leptospirosis.     

Surgical stress and postoperative complications related to regional and radical mastectomy in dogs

Background

Surgery is the treatment of choice for regional control of mammary neoplasms in female dogs. Various surgical techniques may be used, as long as mammary gland anatomy, lymphatic drainage, and known prognostic factors are respected. The purpose of this study was to compare surgical stress—including duration of surgery, nociception and hematological changes—and postoperative complications in dogs undergoing regional and unilateral radical mastectomy. Eighteen dogs were selected for each technique. Postoperative pain (nociception), hematological changes, and postoperative complications were compared between the two groups.

Results

The group treated with radical mastectomy had a longer surgical duration, showed more intense physiological changes, achieved higher scores on nociception scales, and experienced more postoperative complications.

Conclusion

Compared to regional mastectomy, radical mastectomy was associated with longer surgical duration, greater nociceptive stimulus, greater surgical stress, and higher incidence of postoperative complications in dogs. Although evaluation of long-term results was not a goal of this study, it is suggested that postoperative recovery and patient quality of life should be considered when choosing a surgical approach for treating mammary tumors in dogs.     

Soil particle size range correction for improved calibration relationship between the laser-diffraction method and sieve-pipette method

Particle size fraction(clay, silt, and sand) is an important characteristic that influences several soil functions. The laser-diffraction method(LDM) provides a fast and cost-effective measurement of particle size distribution, but the results usually differ from those obtained by the traditional sieve-pipette method(SPM). This difference can persist even when calibration is applied between the two methods. This partly relates to the different size ranges of particles measured by the two methods as a result of different operational principles, i.e., particle sedimentation according to Stokes’ Law vs. Mie theory for laser beam scattering. The objective of this study was to identify particle size ranges of LDM equivalent to those measured by SPM and evaluate whether new calibration models based on size range correction can be used to improve LDM-estimated particle size fractions, using 51 soil samples with various texture collected from five soil orders in New Zealand. Particle size distribution was determined using both LDM and SPM. Compared with SPM, original data from LDM underestimated the clay fraction(< 2 μm), overestimated the silt fraction(2–53 μm), but provided a good estimation of the sand fraction(53–2 000 μm).Results from three statistical indices, including Pearson’s correlation coefficient, slope, and Lin’s concordance correlation coefficient, showed that the size ranges of < 2 and 2–53 μm defined by SPM corresponded with the < 5 and 5–53 μm size ranges by LDM, respectively. Compared with the traditional calibration(based on the same particle size ranges), new calibration models(based on the corrected size ranges of these two methods) improved the estimation of clay and silt contents by LDM. Compared with soil-specific models(i.e., different models were developed for different soils), a universal model may be more parsimonious for estimating particle size fractions if the samples to be assessed represent multiple soil orders.     

Impact of the Textile Dye Acid Blue 40 on the Periphyton of a Simulated Microecosystem

Textile industry is responsible for a large amount of wastewater inappropriate for both human consumption and aquatic species. Aquatic ecosystems are way more sensitive to the release of textile wastewater, and the usage of Winogradsky columns is interesting, once they are a simulated aquatic ecosystem in which the growth of algae and other microorganisms can be observed. In this research, simulated textile effluents with the dye Acid Blue 40 were treated with an electrolytic reactor, for a later ecotoxicological evaluation using Winogradsky columns. The algal and microbial population and primary production were measured. The results have shown that the electrolytic treatment was satisfactory when it comes to color removal, but the presence of the treated effluent in the Winogradsky columns changed the microecosystem. The number of algae identified decreased when exposed to certain effluents, and some algae groups even disappeared, while others such as Cyanophyceae were benefited.     

Wild almonds gone wild: revisiting Darwin’s statement on the origin of peaches

There is a general consensus that cultivated almond was domesticated in western Asia while cultivated peach was domesticated in China; however, the lack of a comprehensive phylogeny of cultivated peaches and almonds and their wild relatives has until recently rendered the controversy over the origin of these plants, their interrelationships and their domestication difficult to decipher. This shortcoming has made Darwin’s hypothesis that peaches are almonds modified in a marvelous manner difficult to test. In this work, Darwin’s hypothesis is revisited based on the first comprehensive molecular and morphological phylogeny of almonds, peaches and their wild relatives. The analysis reveals that cultivated peaches and cultivated almonds are closely related but not sister species. It also reveals that Darwin’s hypothesis was not entirely wrong, particularly considering the species concept he was employing.