Spermatic and oxidative profile of domestic cat (Felis catus) epididymal sperm subjected to different cooling times (24, 48 and 72 hours)

Cooling stored epididymal samples for several days allows facilities to transport and process genetic material post‐mortem. Improvements to this practice allow the preservation of sperm from domestic cats, which are the ideal study model for wild felids. However, the modifications in spermatic features and the oxidative profile are not fully understood in cats. This information is necessary for the development of biotechniques, such as new extenders for cryopreservation. Therefore, the purpose of this study was to evaluate the spermatic and oxidative profile in samples from the epididymal cauda of domestic cats cooled at 5°C for 24, 48 and 72 hr. Spermatozoa were collected from the epididymis cauda. Evaluations consisted of computer‐assisted sperm analysis (CASA), plasma membrane integrity (eosin/nigrosin), acrosome integrity (fast green/rose bengal), sperm morphology, sperm DNA integrity (toluidine blue), mitochondrial activity (3′3 diaminobenzidine), activity of the antioxidant enzymes glutathione peroxidase (GPx) and superoxide dismutase (SOD), measurement of lipid peroxidation (TBARS) and protein oxidation. A decrease in sperm motility parameters was observed after 72 hr of cooling (i.e. total and progressive) with a higher percentage of minor (37.7 ± 6.3%) and total defects (53.4 ± 6.3%). Additionally, a decrease in high mitochondrial activity (Class I: 16.6 ± 2.2%) occurred after 72 hr. The decrease in motility rates after a long cooling time probably was caused by the increase in sperm abnormalities. A long cooling time causes cold shock and mitochondrial exhaustion, but there was no observed change with the oxidative stress condition. Therefore, cat epididymal sperm stored at 5°C appear to maintain a high quality for up to 48 hr of cooling time.     

A 3-year assessment of nitrous oxide emission factors for urine and dung of grazing sheep in a subtropical ecosystem

Purpose

Grazing livestock has strong impact on global nitrous oxide (N₂O) emissions by providing N sources through excreta. The scarcity of information on factors influencing N₂O emissions from sheep excreta in subtropical ecosystems such as those of Southern Brazil led us to conduct field trials in three different winter pasture seasons on an integrated crop–livestock system (ICL) in order to assess N₂O emission factors (EF-N₂O) in response to variable rates of urine and dung.

Materials and methods

The equivalent urine-N loading rates for the three winter seasons (2009, 2010, and 2013) ranged from 96 to 478 kg ha^?1, and the dung-N rates applied in 2009 and 2010 were 81 and 76 kg ha^?1, respectively. Air was sampled from closed static chambers (0.20 m in diameter) for approximately 40 days after excreta application and analyzed for N₂O by gas chromatography.

Results and discussion

Soil N₂O-N fluxes spanned the ranges 4 to 353 μg m^?2 h^?1 in 2009, ??47 to 976 μg m^?2 h^?1 in 2010, and 46 to 339 μg m^?2 h^?1 in 2013. Urine addition resulted in N₂O-N peaks within for up to 20–30 days after application in the 3 years, and the strength of the peaks was linearly related to the N rate used. Emission factors of N₂O (EF-N₂O, % of N applied that is emitted as N₂O) of urine ranged from 0.06 to 0.34% and were essentially independent of N rate applied. By considering a ratio of N excreted by urine and dung of 60:40, a single combined excretal EF-N₂O of 0.14% was estimated.

Conclusions

Our findings showed higher mean EF-N₂O for sheep urine than that for dung (0.21% vs 0.03%), irrespective of the occurrence or not of urine patches overlap. This value is much lower than default value of 1% of IPCC’s Tier 1 and reinforces the needs of its revision.

     

Method of determination of nitrosamines in sausages by CO2 supercritical fluid extraction (SFE) and micellar electrokinetic chromatography (MEKC)

In this paper, the use of supercritical fluid extraction (SFE) and micellar electrokinetic capillary chromatography (MEKC) is proposed for the complete analysis of volatile nitrosamines in sausages. The extraction fluid used was CO2 and variables such as density, temperature of thimbles, extraction time, modifier, fluid flow, and kind of traps were investigated. Several experiments were carried out to obtain the most favorable conditions for analysis of volatile nitrosamines in sausages. The recoveries ranged from 21 to 82% for the five nitrosamines studied. The optimal condition of extraction was 0.2 g of sample fortified with 10 mg/kg, using dynamic extraction during 20 min and with adsorbent Florisil in the trap. The solvent selected for the elution of the analytes was methanol.     

Drivers and risk factors for circulating African swine fever virus in Uganda, 2012–2013

We explored observed risk factors and drivers of infection possibly associated with African swine fever (ASF) epidemiology in Uganda. Representative sub-populations of pig farms and statistics were used in a case-control model. Indiscriminate disposal of pig viscera and waste materials after slaughter, including on open refuse dumps, farm-gate buyers collecting pigs and pig products from within a farm, and retention of survivor pigs were plausible risk factors. Wire mesh-protected windows in pig houses were found to be protective against ASF infection. Sighting engorged ticks on pigs, the presence of a lock for each pig pen and/or a gate at the farm entrance were significantly associated with infection/non-infection; possible explanations were offered. Strict adherence to planned within-farm and community-based biosecurity, and avoidance of identified risk factors is recommended to reduce infection. Training for small-scale and emerging farmers should involve multidimensional and multidisciplinary approaches to reduce human-related risky behaviours driving infection.     

Stabilization of soybean oil with heated quercetin and 5-caffeoylquinic acid in the presence of ferric ion

The effect of heated quercetin (400 mg/kg of oil) or 5-caffeoylquinic acid (5-CQA) and the presence of ferric ion (2.2 mg/kg of oil) on the stability of soya oil oxidized in an oxidative stability index (OSI) instrument was investigated. After heating the phenolic at 100 degrees C or 150 degrees C, the OSI values of treated oils were not significantly (p < 0.001) different, whereas, at 200 degrees C, the values decreased significantly with addition of quercetin, but not with 5-CQA. However, the antioxidative activity of quercetin remained significantly greater than that of 5-CQA. The antioxidative potency of quercetin was reduced significantly by addition of ferric palmitate (FP), but that of 5-CQA was not. Reaction between the ortho-dihydroxy groups of the quercetin and ferric ion may reduce the number of hydroxyls available to react with free radicals. Chelating action of 5-CQA might be provided by ortho-dihydroxy grouping of the quinic acid moiety.     

A fast,low‐cost and efficient method for the diagnosis of sperm DNA fragmentation in several species

Sperm DNA fragmentation is a condition that interferes directly in the reproductive efficiency. Currently, there are several methods for assessing the sperm DNA integrity, such as Alkaline Comet, TUNEL and Sperm Chromatin Structure Assay. However, many of these techniques are laborious and require high‐precision equipment. Thus, the development of new techniques can optimize the evaluation of sperm DNA damage. Therefore, the aim of this study was to standardize the toluidine blue (TB) stain technique for the analysis of DNA fragmentation of dog, cat, bull, stallion and ram spermatozoa. For this purpose, we used six animals of each specie (n = 30), in reproductive age. Sperm was collected by different methods according to the particularities of each species, and such samples were divided into two aliquots: a sperm sample was kept at 5°C (considered as intact sperm DNA), and the remaining samples were submitted to the induction of DNA fragmentation by exposure to ultraviolet light for 4 hr. Samples were then mixed with the intact sample to obtain known and progressive proportions of sperm with fragmented DNA (0%, 25%, 50%, 75% and 100%). Semen smears were performed and subjected to staining with TB. Blue‐stained spermatozoa were considered to have DNA fragmentation. We observed high linear regression coefficients between the expected proportion of damaged DNA and the results of TB for dog, cat, ram, bull and stallion samples. In conclusion, TB stain was considered a fast and effective technique for the study of spermatozoa DNA in several species.     

Development of cotton pests exhibiting different feeding strategy on water-stressed and kaolin-treated cotton plants

Consistent data demonstrate the positive response of sap-sucking insects to water-stressed plants, but there is a lack of information about the performance of chewing species, including whether their responses vary according to their feeding specializations. We tested the hypothesis that herbivores with distinct feeding strategies and host specialization will respond differently to two plant conditions, i.e. water deficit and kaolin treatment, where the latter is used to reduce abiotic and biotic plant stressors. We determined the development and reproduction of four major cotton pest species with different feeding strategies (chewing and sap sucking) and with specializations ranging from monophagous to polyphagous using well-watered or water-stressed cotton plants, which were treated or not treated with kaolin. Three chewing (lepidopteran) species, Alabama argillacea (Hüb.), Heliothis virescens (Fabr.) and Chrysodeixis includens (Walk), as well as one sap-sucking species, Bemisia tabaci (Gen.), were used as models. The neonate larvae of all three chewing species exhibited lower survival when fed the leaves of water-stressed plants. In addition, the neonate larvae fed leaves from water-stressed and kaolin-treated plants did not complete their development or they exhibited low survival. Older larvae also exhibited significantly lower survival on kaolin-treated plants. The effects on life history traits were more obvious in monophagous chewing species compared with polyphagous species when fed leaves from water-stressed plants. The performance of B. tabaci was superior on water-stressed and untreated kaolin plants. Thus, water-stressed and kaolin-treated plants negatively affected all chewing species, whereas water-stressed plants only positively affected the sap-sucking species.     

Roles of bootstrap resampling,measure of central tendency,and method for calculating thermal units when developing phenology models for pest management

The successful development of phenology models from field studies depends on many factors, some of which are entirely under the control of pest managers. For example, one such factor is the choice of method for calculating thermal units. In this study, we have demonstrated that four methods for calculating thermal units provided for acceptable predictions of one phenological event of one insect species, while another method for calculating thermal units did not. The measure of central tendency (mean or median) that is used to estimate lower developmental temperatures and required thermal summations is another factor that pest managers can control when developing phenology models from field studies. Here, we show that predictions that were made when using phenology models based on median lower developmental temperatures and median required thermal summations were superior to predictions that were made when using phenology models based on mean lower developmental temperatures and mean required thermal summations. The use of bootstrap vs. non-bootstrap estimates of lower developmental temperatures and required thermal summations is yet another factor that pest managers can control when developing phenology models from field studies. In this study, we found that calculating and using bootstrap estimates of lower developmental temperatures and required thermal summations in phenology models did not improve the predictions of one phenological event for one insect species. The implications of these and other findings are discussed.     

Influence of murine Toxocara canis infection on plasma and bronchoalveolar lavage fluid eosinophil numbers and its correlation with cytokine levels

Toxocara canis is a nematode of the Ascaridae family that normally parasites the small intestine of canid species. Humans are accidentally infected upon ingestion of embryonated eggs, and can manifest several clinical alterations such as fever, hepatomegaly, splenomegaly, respiratory symptoms, muscle pain and anorexia. In the present work, we investigated the kinetics of tissue distribution of L2 larva in lungs, liver, kidney, brain, skeletal muscle and myocardium. Also, we analyzed the blood and bronchoalveolar lavage fluid (BAL) for levels of IL-6, IFN-gamma, eotaxin and Regulated on Activation Normal T Cell Expressed and Secreted (RANTES) in experimental murine T. canis infection. We observed liver, lung and kidney lesions correlated to larva migration as early as the first day of infection. After the seventh post-infection day, larva could also be detected in brain, skeletal muscle and heart, as an indicator of biphasic migration pattern. Increased inflammatory activity was detected in BAL and plasma of infected animals, as was an intense eosinophil migration associated with an increase in the levels of all the cytokines studied. In conclusion, our results establish a tight correlation between tissue lesions caused by larva migration and increased plasma levels of pro-inflammatory and eosinophil chemotactic cytokines. Thus, murine T. canis infection may prove to be useful in understanding the role of cytokines in infection.