Mohamad Agus Setiadi

Maturation and fertilisation of sheep oocytes cultured in serum-free medium containing silk protein sericin.

Sericin is a water-soluble component of silk and has been used as a biomaterial due to its antibacterial and ultraviolet radiation-resistant properties. This study was designed to evaluate the effect of sericin supplementation in a maturation medium on the meiotic competence and fertilisability of sheep oocytes. Cumulus-oocyte complexes (COCs) were cultured in TCM199 supplemented with sericin at various concentrations of 0 (control), 0.1, 0.25 and 0.5%, either with or without bovine serum albumin (BSA). When the COCs were matured without BSA, the supplementation of 0.1% sericin significantly increased the rates of maturation to metaphase II and the total fertilisation of oocytes compared with the other concentrations of sericin. When the COCs were matured with BSA, the beneficial effects of 0.1% sericin supplementation on the maturation and fertilisation of oocytes were not observed. Our findings indicate that supplementation with 0.1% sericin during maturation culture may improve the nuclear maturation and fertilisability of sheep oocytes. Moreover, it may be possible to replace BSA with sericin in chemically defined media without the risk of disease transmission.

Profil Progesteron Air Susu dan Tingkat Kebuntingan Sapi Perah Pascasinkronisasi Estrus Menggunakan Prostaglandin F2Alfa atau Progesteron-CIDR (MILK PROGESTERONE PROFILE AND PREGNANCY RATE ON DAIRY CATTLE AFTER ESTROUS SYNCHRONIZATION WITH PROSTAGLANDIN F

Research on estrus synchronization, progesterone profiles of post-synchronization and early pregnancyhas been conducted using 16 Holstein Frisian dairy cows. Treated cows were divided into 2 groups. Cowsin group I were synchronized with double injection of prostaglandin F2a (PGF2a, Lutalyse®) 25 mg / head at11 days apart, then were inseminated twice at 72 hours and 96 hours after the second injection of PGF2a(FTAI = fixed timed insemination). Group II were synchronized with progesterone implant-CIDR® for 11days. At the time of progesterone implant withdrawal the animals were injected with PGF2a at 25 mg /head then inseminated twice at 48 hours and 72 hours. Milk samples were collected on the day before, atthe treatment day and at day 1st, 3rd, 5th, 7th following the first insemination in order to determine theprofile of progesterone after synchronization, while for early pregnancy examinations, sampling of milkwere collected at day 21th, 24th and 27th after the first insemination. The milk samples were analyzed byRadioimmunoassay (RIA) method. Rectal palpation to confirm pregnancy was undertaken at day 60thafter the first insemination. The results showed a marked decrease in milk progesterone (nmol / l) at thefirst insemination (H0) in both group PGF2a and CIDR® (0.84; 0.49 vs. 0.92; 0.32), which indicated theoccurrence of estrus. Gradually increased of milk progesterone level (0.52; 0.68; 1.17; 1.69, respectively)started from day 1st, 3rd, 5th, 7th was seen in animals PGF2a group, whereas in the CIDR® group the milkprogesterone level was found fluctuate (0.21; 0.39; 0.33; 1.61). However, at day 7th the concentration ofprogesterone in both groups was significantly increased which indicated functional activity of the corpusluteum. Meanwhile the progesterone concentrations (nmol/l) of pregnant cows at day 21st, 24th and 27th ingroup PGF2a were 3.63; 3.51; 1.58 and in CIDR® group were 2.50; 2,79; 4.35, respectively. In non-pregnantcows, the progesterone concentrations (nmol/l) were lower (0.63; 0.42; 1.41 vs. 0.20; 0.27; 1.33), than thoseof pregnant cows. The results of rectal palpation after 60 days of the first Artificial Insemination (AI)confirmed that 5 cows with higher milk progesterone concentrations at day H21, H24, H27 from the firstinsemination were pregnant, with the possibilities at 62.5% in each group. It is concluded that estroussynchronization using either PGF2a or CIDR® in lactating dairy cows will give the same response and thiscould be detected using the milk progesterone profiles. Measurement of milk progesterone concentrationsby RIA began at day 21 of the first AI was effective for early pregnancy diagnosis.

The use of CR1aa for ovine in vitro embryo production

The aim of this study was to investigate the capacity of CR1aa as a simple medium for maturation, fertilization and culture of ovine embryo in vitro. Oocytes were collected by slicing method in Phosphate Buffer Saline (PBS) supplemented with 5% Fetal Bovine Serum (FBS) and 100 IU/ml penicillin streptomycin. Oocytes were matured in Tissue Culture Medium (TCM)-199 as control or CR1aa as treatment medium. Both maturation medium were supplemented with 10% Fetal Bovine Serum (FBS), 10 IU/ml Follicle Stimulating Hormone (FSH), 10 IU/ml Luteinizing Hormone (LH), 1 μg/ml Estradiol and 100 IU/ml penicillin-streptomycin. Oocytes were incubated in 5% CO 2 incubator, 38˚C for 24 h. Matured oocytes were fertilized in BO or CR1aa medium, supplemented with 2.5 mM caffeine benzoate and 20 mg /ml heparin. After 18 h in vitro fertilization, oocytes were cultured in TCM-199 or CR1aa medium, both supplemented with 5% FBS, 5 mg/ml insulin and 100 IU/ml penicillin streptomycin. Results showed that the highest maturation rate was found in TCM-199 medium (73.27%) and significantly different (P 0.05) between cleavage rate of ovine embryos in TCM-199 and CR1aa medium (39.45% vs 50.94%). In conclusion, optimum result on ovine in vitro embryo production can be achieved from a combination of TCM-199 as maturation medium and CR1aa as fertilization and culture medium. Key Words : CR1aa, TCM-199, Embryo, Ovine

Tingkat Pematangan Inti Oosit Domba dari Ovarium dengan Status Reproduksi dan Medium Maturasi yang Berbeda

The aim of the present study was to determine the number of follicles, oocyte quality and maturation rate of oocytes from pairs of ovary with different reproductive status in two maturation medium, TCM-199 as control and CR1aa as treatment. The pairs of ovary were classified into four groups: (i) ovaries with corpus luteum (CL) and dominant follicle (DF), (ii) ovaries with CL, without DF, (iii) ovaries with DF, without CL, (iv) ovaries without both CL and DF. Results of the experiment revealed that the greatest number of follicles was observed from ovary with CL without DF (15.88 ± 10.68), although not significantly different (P > 0.05) with other status of ovaries. The lowest number (P

Maturation Rate of Ovine Oocytes from Different Reproductive Status and Maturation Medium

A research has been conducted to optimize the rate of aeration and initial weight of cell aggregates in the production of ajmalicine in Catharanthus roseus cell culture in airlift bioreactor. Catharanthus roseus culture were grown in Zenk medium with the addition of 2.50 x 10 -6 M naphthalene acetic acid (NAA) and 10 -5 M benzyl amino purine (BAP). Cell aggregates were sub-cultured two times before transferring 20 and 30 g/fw of cell aggregates into bioreactor, respectively, and aerated with the rate of 0.25 l min -1 and 0.34 l min -1 , respectively. The pattern of ajmalicine production in bioreactor were observed in every three days within 24 days. Qualitative and quantitative analysis were conducted using HPLC connected to Cromatopac CL-7A Plus. The results showed that the cell aggregates and medium contain ajmalicine. The highest concentration was obtained in combination of 30 g/fw and 0.34 l min -1 aeration compare to 20 g/fw - 0.25 l min -1 , 20 g/fw - 0.34 l min -1 , as well as 30 g/fw A¢â‚¬â€œ 0.25 l min -1/sup>. The highest ajmalicine content in cell aggregates was obtained on the 12 days (79.23 A‚µg g - ) whilst in medium was obtained in the 18 thsup> days (981.15 A‚µg l - ). Key words: ajmalicine, cell aggregates culture, C. roseus, airlift bioreactorThe objectives of this experiment were to analyse physiological responses, such as oxidative burst reaction, peroxidase activity, and lignin content of healthy and S. rolfsii-infected peanut tissues. Differences in physiological responses among 24 peanut genotypes were determined, the disease severity was calculated and used to group resistance of tested genotypes. The regressions among observed peroxidase activity, lignin content and disease severity were used to determine the possible mechanisms of S. rolfsii resistance in peanut. Peanut seeds were grown in polybag and the growing plants were inoculated at the crown, stem, and leaf tissues. Results of the experiment indicated that infection of S. rolfsii in peanut did not induce oxidative burst. However, infection of the pathogen resulted in increased peroxidase activity and lignin content in the infected tissues. Regression analysis between peroxidase activity and disease severity showed negative slopes, indicating the more resistance the genotype, the more peroxidase activity in the tissue. Regression analysis between lignin content and disease severity was not significant. Key words: hypersensitive response (HR), resistance mechanisms, Sclerotium stem rot, disease response, Arachis hypogaeaHymenoptera is one of the four largest insect order (the other three are Coleoptera, Diptera, and Lepidoptera). There are curerently over 115 000 described Hymenoptera species. It is clear that Hymenoptera is one of the major components of insect biodiversity. However, Hymenoptera biodiversity is affected by ecology, environment, and ecosystem management. In an agricultural areas, the spatial structure, habitat diversity, and habitat composition may vary from cleared landscapes to structurally rich landscape. Thus, it is very likely that such large-scale spatial patterns (landscape effects) may influence local biodiversity and ecological functions. Therefore, the objective of this research were to study diversity and configuration elements of agricultural landscapes at Cianjur Watershed with geographical information sytems (GIS) and its influence on Hymenoptera biodiversity. The structural differences between agricultural landscapes of Nyalindung, Gasol, and Selajambe were characterized by patch analyst with ArcView 3.2 of digital land use data. Results indicated that class of land uses of Cianjur Watershed landscape were housing, mixed gardens, talun and rice, vegetable, and corn fields. Landscape structure influenced the biodiversity of Hymenoptera. Species richness and the species diversity were higher in Nyalindung landscape compare to Gasol and Selajambe landscape. Key words: diversity, Hymenoptera, landscape, watershed, GIS

Sperm Preservation using Freeze-Drying Method

Since the discovery of cryopreservation method for bull semen, cryopreservation become an alternative method for maintaining gamet resources of certain animal which is threatened or near extinction. This technology was then applied to the preservation of embryo, oocyte, ovary and testis. The application of intracytoplasmic sperm injection (ICSI) for which sperm motility is unnecessary had supported the effort to create simplified method such as freeze-drying for sperm preservation. Due to the benefit of ICSI over the conventional in vitro fertilization (IVF) the spermatozoon could be mechanically driven to pass through the zona pellucida and entering the cytoplasm of oocytes prior to fertilization. The freeze-drying method is an alternative method in sperm preservation which ignored the motility of sperm. The sperm resulted from this technique is in drying state, therefore, it might be stored in room temperature or in refrigerator. Many reports have claimed that freeze-dried sperm which is not motile but has an intact DNA was able to fertilize oocytes, even produced offspring in mouse.A screening for antiinflammatory effects was performed on several Indonesian Umbelliferae plants based on the contents of saponins and flavonoids. They were compared with Bupleurum falcatum L. as an introduced antiinflammatory plant. Roots and grains of each plant were collected, dried, and extracted with ethanol. The ethanolic extracts were then analyzed for their saponin and flavonoid contents by gravimetric and UV-vis spectrophotometric method. Antiinflammatory activity test was conducted on carragenin induced rat paw oedema. The results showed that the highest contents of saponin and flavonoid were found in the grains of Apium graveolens L. and showed antiinflammatory effect that was equivalent to that of the root of B. falcatum.

Pengawetan Spermatozoa Menggunakan Metode Pengeringbekuan

Since the discovery of cryopreservation method for bull semen, cryopreservation become an alternative method for maintaining gamet resources of certain animal which is threatened or near extinction. This technology was then applied to the preservation of embryo, oocyte, ovary and testis. The application of intracytoplasmic sperm injection (ICSI) for which sperm motility is unnecessary had supported the effort to create simplified method such as freeze-drying for sperm preservation. Due to the benefit of ICSI over the conventional in vitro fertilization (IVF) the spermatozoon could be mechanically driven to pass through the zona pellucida and entering the cytoplasm of oocytes prior to fertilization. The freeze-drying method is an alternative method in sperm preservation which ignored the motility of sperm. The sperm resulted from this technique is in drying state, therefore, it might be stored in room temperature or in refrigerator. Many reports have claimed that freeze-dried sperm which is not motile but has an intact DNA was able to fertilize oocytes, even produced offspring in mouse.