Effects of permanent magnetic fields on the proliferation of Phalaenopsis protocorm-like bodies using liquid medium

Magnetic fields (MFs) have been applied for the first time in orchid micropropagation. Protocorm-like bodies (PLBs) – approximately 3 mm in diameter – first derived from leaf segment culture of Phalaenopsis Gallant Beau ‘George Vazquez’, were subcultured every 2 months, and served as initial explants. The proliferation of Phalaenopsis PLBs in liquid medium in the Miracle Pack^® culture system was affected by the action of different intensities and polarities of MFs: 0.1, 0.15 and 0.2 Tesla (T) at North (N) and South (S) poles. The MF of 0.1 T – S resulted in the greatest fresh weight of regenerated PLBs. The average number of neo-PLBs formed per clusters in the PLB treated by MF: 0.1–0.2 T was decreased compared to the control exposed to natural MF (5 × 10⁻⁶ T). The proliferation of PLBs under 0.15 T – MF at both N and S poles for 2 and 7 weeks demonstrated that a longer duration of exposure to an MF of 0.15 T, regardless of the polarity, resulted in greater biomass of newly formed PLBs and smaller average number of newly formed PLBs. The S pole of MF had stronger effects on Phalaenopsis PLBs proliferation than the N pole did in all treatments.     

Agrobacterium-mediated transformation of the CrDAT gene and selection of transgenic periwinkle lines with a high vincristine accumulation

Catharanthus roseus contains vincristine and vinblastine, which are outstanding drugs for cancer. In the biosynthetic pathways of terpenoid indole alkaloids (TIAs) in C. roseus, deacetylvindoline 4-O-acetyltransferase (DAT) is a key enzyme that catalyses the last reaction of vindoline biosynthesis to form vinblastine and vincristine. In this study, the CrDAT transgene was transferred into the periwinkle by Agrobacterium-mediated transformation and generated transgenic periwinkle lines with an increase in vincristine accumulation. The C. roseus DAT gene was introduced into C. roseus plants and it was confirmed that CrDAT was successfully transferred into the genome of periwinkle plants and efficiently translated to synthesise recombinant DAT protein. Four transgenic periwinkle lines in T1 generation, T1-1, T1-3, T1-6, and T1-7, expressed recombinant DAT protein with the total protein content in the range of 2.86 μg.mg^?1 to 5.12 μg.mg^?1. Moreover, the vincristine contents of four transgenic lines increased by 1.63?2.48-fold compared to non-transgenic plants, ranging from 6.91 µg.g^?1 (fresh weight) to 10.53 µg.g^?1 (fresh weight). The T1-1 line had the highest vincristine content. Hence, the overexpression of the recombinant DAT protein can improve the vincristine accumulation of transgenic C. roseus plants.

Abbreviation: CrDAT - Catharanthus roseus Deacetylvindoline-4-O-Acetyl Transferase; D4H - Deacetoxyvindoline 4-hydroxylase; ELISA - Enzyme-Linked Immunosorbent Assay Monoterpene indole alkaloid; T0, T1 - Generations of transgenic plants; TIAs - Terpenoid indole alkaloids; WT- The wild-type tobacco plants (non transgenic plant); 35S - Cauliflower mosaic virus 35S promoter     

Optimization of in vitro embryo production and zygote vitrification for the indigenous Vietnamese Ban pig: The effects of different in vitro oocyte maturation systems

The Vietnamese Ban pig is a precious genetic resource that needs to be preserved. In vitro embryo production from in vitro matured (IVM) oocytes is an important tool for the utilization of cryopreserved porcine sperm. The aim of this study was to compare two media for the IVM of Ban pig oocytes. Immature oocytes were subjected to IVM either in a non‐defined (TCM‐199 + pig follicular fluid) or in a defined base medium (POM + epidermal growth factor). At the end of IVM, the oocytes were in vitro fertilized (IVF) with frozen Ban sperm. Ten hours after IVF, the oocytes were either subjected to orcein staining to check fertilization and maturation status or cultured in vitro for 7 days. There was no difference between the two IVM media in terms of percentages of oocyte maturation and blastocyst production. However, the percentage of male pronuclear formation after IVF and the total cell numbers in blastocysts were higher with the defined system. Zygotes obtained by the two IVM systems survived vitrification at similar rates. In conclusion, the two IVM systems were both effective for the production of Ban pig embryos; however, better embryo quality was achieved with the defined one.     

交替饲喂整粒小麦或含蛋白质及矿物质浓缩的饲料提高蛋鸡饲料的利用效率

对16～46周龄的蛋鸡生产性能的研究表明,交替饲喂或喂混合饲料的效果是不同的。一种方法是利用同等比例的整粒小麦或营养均衡的含蛋白质及矿物质浓缩的饲料交替饲喂ISA布朗母鸡,另一种方法是饲喂混合饲料。对照组喂传统的饲料。每组16笼,每笼中5只鸡。每天光照时间16h。控制饲料的饲喂量,121g·只-·1d-1,每天饲喂2次,分别为开灯后4h和11h。在交替饲喂时,仅第1次喂食小麦,第2次喂食营养的均衡饲料。喂混合饲料时,两种定量配给的饲料混合喂2次。在下一次喂食前拿走剩余的饲料。与喂混合饲料和对照组相比,交替饲喂减少总采食量,小麦减少9g·只-·1d-1,但喂均衡饲料增多1.7g·只-·1d·-1。在3种方式中,鸡蛋的生产,蛋的大小,蛋的重量均相似。交替饲喂与喂混合饲料相比,饲料利用率的提高分别为10%和5%。与喂混合饲料和对照组相比,交替饲喂ME低,CP摄取量减少。交替饲喂时蛋鸡体重轻,其肌胃、胰腺和肝脏较重,肝脏脂质类似。喂混合饲料肝糖最高。试验结论为,交替饲喂整粒小麦和营养均衡的饲料或喂混合饲料对蛋鸡生产无负面影响。因此,两种饲喂方法是可以替代传统的喂养方式。与喂混合饲料相比,交替饲喂饲料的利用效率高,因而是可行的。     

Genotyping canine distemper virus (CDV) by a hemi-nested multiplex PCR provides a rapid approach for investigation of CDV outbreaks

CDV is a highly contagious viral pathogen causing a lethal systemic disease in dogs and other carnivores. Several lineages or genotypes of CDV exist that are variously distributed throughout several continents. Legal or uncontrolled trading of animals may modify the epidemiology of CDV, introducing novel strains in CDV-na?ve areas or accounting for the resurgence of CDV in areas where vaccine prophylaxis was effective and successful to control the disease. A hemi-nested PCR system was developed to genotype strains of the major CDV lineages, America-1, Europe, Asia-1, Asia-2 and Arctic. The assay was tested using a collection of 27 laboratory and vaccine strains and of 36 field CDV strains. Distinct lineages could be differentiated by specific primers targeted to the H gene. The method could be useful for molecular epidemiological studies of CDV, providing a tool for large-scale studies, and for the diagnosis of vaccine-related disease.     

Sucrose assists selection of high‐quality oocytes in pigs

We investigated whether high‐quality in vitro matured (IVM) oocytes can be distinguished from poor ones based on the morphological changes after treatment with hyperosmotic medium containing 0.2 mol/L sucrose in pigs. We hypothesize that IVM oocytes maintaining round shape have higher quality than mis‐shapened oocytes following dehydration. Oocyte quality was verified by determining embryonic developmental competence using in vitro fertilization, nuclear transfer and parthenogenetic activation. In all cases, the round oocytes had greater (p < .05) developmental competence than that of mis‐shapened oocytes in terms of blastocyst rate and total cell number in blastocysts obtained after 6 days of in vitro culture. We also confirm that round aged oocytes are higher in quality than mis‐shapened aged oocytes. In an attempt to find out why high‐quality oocytes maintain a round shape whereas poorer oocytes become mis‐shapened following sucrose treatment, we examined the arrangement of actin microfilaments and microtubules. Abnormal organization of these cytoskeletal components was higher (p < .05) in mis‐shapened oocytes compared to round oocytes after 52 hr of IVM. In conclusion, sucrose treatment helps selection of high‐quality oocytes, including aged oocytes, in pigs. Abnormal cytoskeleton arrangements partly explain for low developmental competence of mis‐shapened oocytes.     

Studies of the digestibility of pigs fed dietary sucrose, fructose or glucose.

Digestion of sucrose, fructose and glucose was studied in pigs with reentrant cannulas in the distal ileum or intact pigs fed diets formulated with sucrose, fructose and glucose as the major energy source and torula yeast. Sucrose and glucose were completely digested in the small intestine (apparent digestibility, 98.3 and 98.3%, respectively but a fraction of the dietary fructose escaped to the large intestine (adjusted value, 11.7%). There was no carbohydrate in rectal contents of the pigs. There was no carbohydrate influence on the pre-caecal or total digestibility of nitrogen.     

Distribution of Salmonella Serovars and Antimicrobial Susceptibility from Poultry and Swine Farms in Central Vietnam

This study was conducted to estimate the prevalence of Salmonella spp. and their antimicrobial susceptibilities on poultry and swine farms, sampled in 2 regions in Central Vietnam. A total of 67 poultry farms and 46 swine farms were sampled in a period of 5 months (from September 2012 to January 2013). Salmonella spp. was prevalent in 46.3% and 71.7% of poultry and swine farms, respectively. Altogether, 99 non‐typhoidal Salmonella were isolated and the most common serovars were Salmonella Weltevreden (19%), followed by Salmonella Typhimurium (12%) and Salmonella 4,[5],12:i:‐ (11%). Overall, 71 of 99 (72%) Salmonella isolates were resistant to at least one of the 14 antimicrobial agents tested. Both in poultry and swine farms, high levels of resistance were observed for ampicillin, chloramphenicol, ciprofloxacin, sulphamethoxazole and tetracycline. The presence of Salmonella isolates from poultry and swine farms which were resistant to different classes of antimicrobials suggests that alternative control measures to antimicrobials should be implemented. Moreover, an effective policy should be promoted to encourage a prudent use of these agents in animal farming in Vietnam.     

The effect of ovary storage and in vitro maturation on mRNA levels in bovine oocytes; a possible impact of maternal ATP1A1 on blastocyst development in slaughterhouse-derived oocytes

Since BSE testing of slaughtered cattle is obligatory in Japan, storage of ovaries at 15-20 C overnight in phosphate buffered saline has become a routine protocol in in vitro production (IVP) of cattle embryos. Ovary storage is known to reduce developmental competence of oocytes; however, its effects on oocyte gene expression have not been clarified yet. This study compared oocytes collected from stored slaughterhouse-derived ovaries with those collected by Ovum Pick-Up (OPU) in terms of the expression of 20 selected genes to determine if ovary storage affects cellular processes at the molecular level. Expression of mRNA in oocytes was assayed before and after in vitro maturation (IVM) by real-time quantitative PCR. Maternal mRNA levels of genes were investigated in 2-cell stage embryos obtained from slaughterhouse oocytes to assess their roles for blastocyst formation. In immature OPU oocytes, genes related to metabolism (GAPDH), transporters (GLUT8, ATP1A1) and stress resistance protein (HSP70) showed significantly higher expression compared with oocytes derived from stored ovaries. During IVM, the expression of GDF9, GLUT8, CTNNB1 and PMSB1 was significantly decreased irrespective of oocyte source. Two-cell stage embryos cleaving at 22-25 h after in vitro fertilization (IVF) showed a significantly higher blastocyst formation rate and ATP1A1 gene expression level compared with those cleaving at 27-30 h after IVF. Our results reveal that storage of ovaries alters mRNA levels in oocytes. Correlation of Na/K ATPase ATP1A1 expression in IVP embryos at the 2-cell and 8-cell stages with their developmental ability to the blastocyst stage may suggest the importance of maternal mRNA of this gene during blastulation in embryos derived from slaughterhouse oocytes.