Background:To advance the use of embryo vitrification in veterinary practice,we developed a system in which embryo vitrification,warming and dilution can be performed within a straw.Ovine in vitro produced embryos(IVE...Background:To advance the use of embryo vitrification in veterinary practice,we developed a system in which embryo vitrification,warming and dilution can be performed within a straw.Ovine in vitro produced embryos(IVEP)were vitrified at either early(EBs:n=74)or fully expanded blastocyst stage(FEBs:n=195),using a new device named"E.Vit",composed by a 0.25-m L straw with a 50-μm pore polycarbonate grid at one end.Embryos at each stage(EBs and FEBs)were vitrified by either Two-step(TS)or Multi-step(MS;6 different concentrations of vitrification solutions)protocol.Non-vitrified embryos(n=102)were maintained in in vitro culture as a control.Warming consisted of placing the straws directly into 1.5 m L tubes containing a TCM-199 solution with three decreasing concentrations of sucrose.Blastocyst re-expansion,embryo survival and hatching rate were evaluated at2,24 and 48 h post warming.The number of apoptotic cells was determined by TUNEL assay.Results:Blastocyst re-expansion(2 h)after warming was higher(P<0.05)in FEBs group,vitrified with the MS and TS methods(77.90%and 71.25%,respectively)compared with the EBs group(MS:59.38%and TS:48.50%,respectively).Survival rates of vitrified FEBs after 24 h IVC were higher(P<0.001)in both methods(MS and TS)than vitrified EBs(MS:56.25%;TS:42.42%)and was higher(P<0.05)in the MS method(94.19%)compared with those in TS(83.75%).After 48 h of culture the hatching rate for FEBs vitrified in MS system(91.86%)was similar to control(91.89%),but higher than FEB TS(77.5%)and EBs vitrified in MS(37.5%)and TS(33.33%).Number of apoptotic cells were higher in EBs,irrespective of the system used,compared to FEBs.The number of apoptotic cells in FEBs vitrified with MS was comparable to the control.Conclusions:A high survival rate of IVP embryos can be achieved by the new"E.Vit"device with hatching rates in vitro comparable with control fresh embryos.This method has the potential for use in direct embryo transfer in field conditions.展开更多
Background： Current research to enrich cattle feed has primarily focused on treatment using white rot fungi, while there are scarce reports using the enzyme tannase, which is discussed only in reviews or in the form ...Background： Current research to enrich cattle feed has primarily focused on treatment using white rot fungi, while there are scarce reports using the enzyme tannase, which is discussed only in reviews or in the form of a hypothesis. In this context, the aim of the present study was to evaluate the effect of tannase on wheat straw （WS） and also the effect of lyophilized tannase at concentrations of 0.1%, 0.2%, and 0.3% （w/w） on WS followed by fermentation with Ganodermo sp. for 10 d and compared in relation to biochemical parameters, crude protein （CP） content, and nutritional value by calculating the C/N ratio in order to improve the nutritional value of cattle feed. Results： Penicillium charlesii, a tannase-producing microorganism, produced 61.4 IU/mL of tannase in 54 h when 2% （w/v） tannic acid （TA） was initially used as a substrate in medium containing （% w/v） sucrose （1.0）, NaNO3 （1.0）, and MgSO4 （0.08 pH, 5.0） in a 300-L fermentor （working volume 220 L）, and concomitantly fed with 1.0% （w/v） TA after 24 h. The yield of partially purified and lyophilized tannase was 5.8 IU/mg. The tannin-free myco-straw at 0.1% （w/w） tannase showed 37.8% （w/w） lignin degradation with only a 20.4% （w/w） decrease in cellulose content and the in vitro feed digestibility was 32.2%. An increase in CP content （up to 1.28-fold） along with a lower C/N ratio of 25.0%, as compared to myco-straw, was obtained. Conclusions： The use of tannin-free myco-straw has potential to improve the nutritional content of cattle feed. This biological treatment process was safe, eco-friendly, easy to perform, and was less expensive as compared to other treatment methods.展开更多
基金supported by Regione Autonoma della Sardegna.-L.R.7-MIGLIOVINGENSAR ProjectBando competitivo Fondazione di Sardegna–2016,CUP J86C18000800005“Progetto FAR2019LEDDAS Una Tantum 2019,University of Sassari”.
文摘Background:To advance the use of embryo vitrification in veterinary practice,we developed a system in which embryo vitrification,warming and dilution can be performed within a straw.Ovine in vitro produced embryos(IVEP)were vitrified at either early(EBs:n=74)or fully expanded blastocyst stage(FEBs:n=195),using a new device named"E.Vit",composed by a 0.25-m L straw with a 50-μm pore polycarbonate grid at one end.Embryos at each stage(EBs and FEBs)were vitrified by either Two-step(TS)or Multi-step(MS;6 different concentrations of vitrification solutions)protocol.Non-vitrified embryos(n=102)were maintained in in vitro culture as a control.Warming consisted of placing the straws directly into 1.5 m L tubes containing a TCM-199 solution with three decreasing concentrations of sucrose.Blastocyst re-expansion,embryo survival and hatching rate were evaluated at2,24 and 48 h post warming.The number of apoptotic cells was determined by TUNEL assay.Results:Blastocyst re-expansion(2 h)after warming was higher(P<0.05)in FEBs group,vitrified with the MS and TS methods(77.90%and 71.25%,respectively)compared with the EBs group(MS:59.38%and TS:48.50%,respectively).Survival rates of vitrified FEBs after 24 h IVC were higher(P<0.001)in both methods(MS and TS)than vitrified EBs(MS:56.25%;TS:42.42%)and was higher(P<0.05)in the MS method(94.19%)compared with those in TS(83.75%).After 48 h of culture the hatching rate for FEBs vitrified in MS system(91.86%)was similar to control(91.89%),but higher than FEB TS(77.5%)and EBs vitrified in MS(37.5%)and TS(33.33%).Number of apoptotic cells were higher in EBs,irrespective of the system used,compared to FEBs.The number of apoptotic cells in FEBs vitrified with MS was comparable to the control.Conclusions:A high survival rate of IVP embryos can be achieved by the new"E.Vit"device with hatching rates in vitro comparable with control fresh embryos.This method has the potential for use in direct embryo transfer in field conditions.
基金supported by a grant from the Council of Scientific and Industrial Research of India to S.R.(grant no.:9/45(1190)/2012-EMR-1)
文摘Background： Current research to enrich cattle feed has primarily focused on treatment using white rot fungi, while there are scarce reports using the enzyme tannase, which is discussed only in reviews or in the form of a hypothesis. In this context, the aim of the present study was to evaluate the effect of tannase on wheat straw （WS） and also the effect of lyophilized tannase at concentrations of 0.1%, 0.2%, and 0.3% （w/w） on WS followed by fermentation with Ganodermo sp. for 10 d and compared in relation to biochemical parameters, crude protein （CP） content, and nutritional value by calculating the C/N ratio in order to improve the nutritional value of cattle feed. Results： Penicillium charlesii, a tannase-producing microorganism, produced 61.4 IU/mL of tannase in 54 h when 2% （w/v） tannic acid （TA） was initially used as a substrate in medium containing （% w/v） sucrose （1.0）, NaNO3 （1.0）, and MgSO4 （0.08 pH, 5.0） in a 300-L fermentor （working volume 220 L）, and concomitantly fed with 1.0% （w/v） TA after 24 h. The yield of partially purified and lyophilized tannase was 5.8 IU/mg. The tannin-free myco-straw at 0.1% （w/w） tannase showed 37.8% （w/w） lignin degradation with only a 20.4% （w/w） decrease in cellulose content and the in vitro feed digestibility was 32.2%. An increase in CP content （up to 1.28-fold） along with a lower C/N ratio of 25.0%, as compared to myco-straw, was obtained. Conclusions： The use of tannin-free myco-straw has potential to improve the nutritional content of cattle feed. This biological treatment process was safe, eco-friendly, easy to perform, and was less expensive as compared to other treatment methods.
