Apoptosis in the Antral Follicles of Swamp Buffalo and Cattle Ovary: TUNEL and Caspase-3 Histochemistry

The present study was carried out to investigate the pattern of apoptosis in the healthy antral and atretic follicles of Philippine swamp buffaloes (BU) in comparison with Holstein-Friesian (HF) cows. Paraffin sections of healthy follicles and various stages of atretic follicles were stained using the terminal deoxynucleotidyl transferase (TdT)-mediated biotinylated deoxyuridine triphosphates (dUTP) nick end-labelling (TUNEL) method to detect DNA fragmentation and cleaved caspase-3 antibody to detect cells committed to undergo apoptosis. Five equidistant areas of a follicle were counted for the presence of TUNEL- and caspase-3-positive cells. Healthy follicles of BU and HF contained no TUNEL-positive cells in the granulosa and theca layer but showed some caspase-3 positivity. The granulosa layer of advanced atretic follicles showed a significantly higher frequency of caspase-3 positivity than the healthy and early atretic follicles in both breeds. The frequency of caspase-3-positive cells of BU was significantly higher than HF in the granulosa layer of healthy, early atretic and advanced atretic follicles. In the theca interna layer, BU and HF showed a significantly lower and higher frequency of TUNEL-positive cells in the late atretic follicles compared with advanced atretic follicle, respectively. However, the frequency of caspase-3-positive cells of both BU and HF in the late atretic follicles was significantly higher than the advanced atretic follicles in the theca interna layer. These results indicate that caspase-3 and DNA fragmentation is involved in the buffalo ovarian apoptotic process.     

Cell Proliferation in the Atretic Follicles of Buffalo and Cattle Ovary

The present study was carried out to describe the proliferative activity of granulosa and theca cells in healthy antral and atretic follicles of Philippine buffaloes (BU) and Holstein-Friesian (HF) cows. Paraffin sections of ovary were immunostained with mouse monoclonal antibody to proliferating cell nuclear antigen (PCNA). Then the follicles were classified into healthy and various stages of atretic follicles. The granulosa layer of healthy follicles had a significantly higher frequency of PCNA-positive cells than the early and advanced atretic follicles in both breeds. In the theca interna, significantly reduced populations of the PCNA-positive cells were found in both breeds as atresia progressed. Moreover, HF had significantly higher PCNA-positive cells in the theca interna of healthy, early atretic and advanced atretic follicles than BU. A reduction of PCNA-positive cells during atresia was also noted in the theca externa in both animals although differences were not significant. The results of the present work suggest that the proliferative activity of granulosa and theca cells decreases in association with follicular atresia in the BU similar to HF. Furthermore, a significantly deficient cell proliferative activity of theca interna was found in BU compared with HF.     

Growth- and leaf-temperature effects on photosynthesis of sweet orange seedlings infected with Xylella fastidiosa

The effects of growth and leaf temperature on photosynthesis were evaluated in sweet orange seedlings ( Citrus sinensis cv. Pera) infected with Xylella fastidiosa (the bacterium that causes citrus variegated chlorosis, CVC). Measurements of leaf gas exchange and chlorophyll  a fluorescence were taken at leaf temperatures of 25, 30, 35 and 40°C in healthy and infected (without visible symptoms) seedlings submitted to two temperature regimes (25/20 or 35/20°C, day/night), not simultaneously. The CO₂ assimilation rates ( A ) and stomatal conductance ( g _s) were higher in healthy plants in both temperature regimes. Values for A and g _s of infected and healthy plants were higher in the 35/20°C regime, decreasing with leaf temperature increase. In addition, differences between healthy and infected plants were higher at 35/20°C, while no differences in chlorophyll  a fluorescence parameters were observed except for potential quantum efficiency of photosystem II, which was higher in infected plants. Low A values in infected plants were caused by low g _s and probably by biochemical damage to photosynthesis. The high alternative electron sink of infected plants was another effect of reduced A . Both high growth and high leaf temperatures increased differences in A between healthy and infected plants. Therefore this feature may be partially responsible for lower growth and/or productivity of CVC-affected plants in regions with high air temperature.     

Ergovaline does not alter the severity of ryegrass staggers induced by lolitrem B

AIM: To investigate a possible interaction between lolitrem B and ergovaline by comparing the incidence and severity of ryegrass staggers in sheep grazing ryegrass (Lolium perenne) containing lolitrem B or ryegrass containing both lolitrem B and ergovaline.

METHODS: Ninety lambs, aged approximately 6 months, were grazed on plots of perennial ryegrass infected with either AR98 endophyte (containing lolitrem B), standard endophyte (containing lolitrem B and ergovaline) or no endophyte, for up to 42 days from 2 February 2010. Ten lambs were grazed on three replicate plots per cultivar. Herbage samples were collected for alkaloid analysis and lambs were scored for ryegrass staggers (scores from 0–5) weekly during the study. Any animal which was scored ≥4 was removed from the study.

RESULTS: Concentrations of lolitrem B did not differ between AR98 and standard endophyte-infected pastures during the study period (p=0.26), and ergovaline was present only in standard endophyte pastures. Ryegrass staggers was observed in sheep grazing both the AR98 and standard endophyte plots, with median scores increasing in the third week of the study. Prior to the end of the 42-day grazing period, 22 and 17 animals were removed from the standard endophyte and AR98 plots, respectively, because their staggers scores were ≥4. The cumulative probability of lambs having scores ≥4 did not differ between animals grazing the two pasture types (p=0.41).

CONCLUSIONS AND CLINICAL RELEVANCE: There was no evidence for ergovaline increasing the severity of ryegrass staggers induced by lolitrem B. In situations where the severity of ryegrass staggers appears to be greater than that predicted on the basis of concentrations of lolitrem B, the presence of other tremorgenic alkaloids should be investigated.     

Meta‐analysis of the energy and protein requirements of hair sheep raised in the tropical region of Brazil

The objective of this study was to estimate, through mathematical models, energy and protein requirements for maintenance and gain of hair sheep raised in the tropical region of Brazil. To determine the equation parameters, a meta‐analysis of seven independent experiments of nutrient requirements was performed, comprising a total of 243 experimental units (animals), which were conducted under tropical conditions, using hair sheep in growing and finishing phases and endowed of the following quantitative data for each animal: body weight (BW ), empty body weight (EBW ), average daily gain (ADG ), empty body gain (EBG ), heat production (HP ), metabolizable energy intake (MEI ), retained energy (RE ), metabolizable protein intake (MPI ) and body protein content. The regression equations generated were as follows: for Net Energy for maintenance, (NE _m): ; for Net Energy for gain, (NE _g): ; for Metabolizable Protein for maintenance,(MP _m): MPI _(g/day) = 24.8470 (±7.3646) + 560.28 (±99.6582) × EBG _(kg/day); for Net Protein for gain, (NP _g): . The NE _m requirement was 0.246 MJ EBW^?0.75 day^?1. The metabolizable energy for maintenance requirement was 0.391 MJ EBW^?0.75 day^?1. Considering an ADG of 100 g, the NE _g requirement ranged from 0.496 to 1.701 MJ/day for animals with BW ranging from 10 to 40 kg respectively. The efficiencies of use of the metabolizable energy for maintenance and gain were 0.63 and 0.36 respectively. The MP _m requirement was 3.097 g EBW^?0.75 day^?1. Considering an ADG of 100 g, the NP _g requirement ranged from 12.4 to 10.5 g/day for animals with BW ranging from 10 to 40 kg respectively. The total metabolizable energy and protein requirements were lower than those reported by the NRC and AFRC systems. Thus, our results support the hypothesis that nutrient requirements of hair sheep raised in tropical regions differ from wool sheep raised in temperate regions. Therefore, the use of the equations designed in this study is recommended.     

Transgenerational epigenetic variance for body weight in meat quails

We aimed to estimate transgenerational epigenetic variance for body weight using genealogical and phenotypic information in meat quails. Animals were individually weighted from 1 week after hatching, with weight records at 7, 14, 21, 28, 35 and 42 days of age (BW7, BW14, BW21, BW28, BW35 and BW42, respectively). Single‐trait genetic analyses were performed using mixed models with random epigenetic effects. Variance components were estimated by the restricted maximum likelihood method. A grid search for values of autorecursive parameter (λ) ranging from 0 to 0.5 was used in the variance component estimation. This parameter is directly related to the reset coefficient (ν) and the epigenetic coefficient of transmissibility (1‐ν). The epigenetic effect was only significant for BW7. Direct heritability estimates for body weight ranged in magnitude (from 0.15 to 0.26), with the highest estimate for BW7. Epigenetic heritability was 0.10 for BW7, and close to zero for the other body weights. The inclusion of the epigenetic effect in the model helped to explain the residual and non‐Mendelian variability of initial body weight in meat quails.     

Melatonin reduces apoptotic cells,SOD2 and HSPB1 and improves the in vitro production and quality of bovine blastocysts

Effects of adding different concentrations of melatonin (10^?7, 10^?9 and 10^?11 M) to maturation (Experiment 1; Control, IVM  + 10^?7, IVM  + 10^?9, IVM  + 10^?11) and culture media (Experiment 2; Control, IVC  + 10^?7, IVC  + 10^?9, IVC  + 10^?11) were evaluated on in vitro bovine embryonic development. The optimal concentration of melatonin (10^?9 M) from Experiments 1–2 was tested in both maturation and/or culture media of Experiment 3 (Control, IVM  + 10^?9, IVC  + 10^?9, IVM /IVC  + 10^?9). In Experiment 1, maturated oocytes from Control and IVM  + 10^?9 treatments showed increased glutathione content, mitochondrial membrane potential and percentage of Grade I blastocysts (40.6% and 43%, respectively). In Experiment 2, an increase in the percentage of Grade I blastocysts was detected in IVC  + 10^?7 (43.5%; 56.7%) and IVC  + 10^?9 (47.4%; 57.4%). Moreover, a lower number and percentage of apoptotic cells in blastocysts were observed in the IVC  + 10^?9 group compared to Control (3.8 ± 0.6; 3.6% versus 6.1 ± 0.6; 5.3%). In Experiment 3, the IVC  + 10^?9 treatment increased percentage of Grade I blastocysts with a lower number of apoptotic cells compared to IVM /IVC  + 10^?9 group (52.6%; 3.0 ± 0.5 versus 46.0%; 5.4 ± 1.0). The IVC  + 10^?9 treatment also had a higher mRNA expression of antioxidant gene (SOD 2) compared to the Control, as well as the heat shock protein (HSPB 1) compared to the IVM  + 10^?9. Reactive oxygen species production was greater in the IVM /IVC  + 10^?9 treatment group. In conclusion, the 10^?9 M concentration of melatonin and the in vitro production phase in which it is used directly affected embryonic development and quality.     

Aboveground biomass,transpiration and water use efficiency in eucalypt plantation fertilized with KCl,NaCl and phonolite rock powder

Potassium has important physiological functions in eucalypt plantations, increasing their productivity when applied to soil via mineral fertilizers. There is interest in identifying alternative sources to KCl owing to its high cost and limited reserves. The aim of the study was to test the effect of replacing KCl with NaCl and phonolite rock powder. Two comparisons were made: (1) application of 283 kg ha^?1 of KCl compared with that of 2125 kg ha^?1 of phonolite rock powder (equivalent to 170 kg ha^?1 of K₂O in both treatments); (2) application of 139 kg ha^?1 of NaCl compared with that of 183 kg ha^?1 of KCl (equivalent to 2.33 kmol Na and K, respectively). Radial growth, soil water content, leaf water potential (Ψ), accumulated transpiration, stem volume and biomass increment, as well as water use efficiency (WUE) were evaluated. In the first comparison, both fertilizations presented equal values for all characteristics evaluated. In the second, the accumulated transpiration in trees fertilized with KCl was 17% higher than that in plants fertilized with NaCl. In contrast, the WUE was 20% higher in the trees fertilized with NaCl than in those fertilized with KCl, reflecting the lower water consumption for the same increment in stem volume and biomass. We conclude that phonolite rock powder and NaCl are possible substitutes for conventional K fertilization performed with KCl.

     

Feed preference and growth response of juvenile Litopenaeus vannamei to supplementation of marine chemoattractants in a fishmeal‐challenged diet

This study compared the feed preference and growth response of Litopenaeus vannamei to chemoattractants. A diet with 3% fishmeal was supplemented with either 3% salmon meal (POS), 3% soy protein concentrate (NEG), 3% krill meal (KRM), 3% squid meal (SQM), 3% shrimp head meal (SHM), 3% shrimp meal (SM), 3% squid liver meal (SLM), or 5% liquid sardine hydrolysate (SAH). Shrimp with a body weight (BW) of 0.99 ± 0.08 g were stocked at 100 animals/m² in 56 tanks of 1 m³ and fed 10 times daily for 74 days. Feed preference was evaluated by feeding shrimp of 10.87 ± 1.82 g in excess twice a day for 10 days in two separate feeding trays allocated in 50 tanks of 0.5 m³. Survival reached 93.3 ± 5.80% and was unaffected by supplementation. Final BW was the highest for shrimp fed the KRM‐supplemented diet (11.97 ± 0.93 g), followed by POS (11.11 ± 0.77 g) and SQM (11.01 ± 1.17 g). Diets SHM, SM, SLM, and NEG showed a lower shrimp BW than POS, but were not statistically different among them. Shrimp fed the SAH diet achieved the lowest BW (10.06 ± 1.02 g). The highest gained yield was obtained with diets KRM and POS. No statistical difference was observed in shrimp yield among other diets. The lowest feed conversion ratio (FCR) was achieved with shrimp fed KRM (1.31 ± 0.05) when compared to diets SHM (1.47 ± 0.05), SAH (1.47 ± 0.07), and SLM (1.45 ± 0.17). Two‐by‐two comparisons indicated that shrimp preferred SHM and KRM, except when these were compared to SQM and SLM. No difference in feed preference was found between diets with SQM and SLM. SAH was the least preferred raw material in all comparisons. Results indicated that KRM acts as a powerful feeding effector and growth enhancer in fishmeal‐challenged diets for whiteleg shrimp. A dietary supplementation with 3% KRM is more effective than the same dose of any other chemoattractant evaluated.