Judul: Development of DNA Vaccine Encoding Glycoprotein of Koi Herpes Virus Using Local Isolate.
Abstrak: The aim of this research was to develop DNA vaccine of Koi Herpesvirus in carp. Glycoprotein gene originated from ORF 25 KHV virus that is 1.8 kbp in size, is a potential gene in DNA vaccine construction. The sequencing and similarity analysis of the gene using BLAST software showed that this gene is a GP25 gene and was found to be very similar (99%) to GP25 of KHV virus from the USA and Israel. When this GP25 gene was ligated with pGEMT Easy (pT, 3 kbp in size) vector, plasmid size was increasing up to 5 kb. Following this, a digestion of pT-GP25 plasmid using Sal I enzyme was applied resulted in a GP25 fragment (1.8 kbp). Subsequently, pAct-D6 with a size of 8.5 kbp was also cut using the same enzyme to eleminate a D6 gene with a size of 1.6 kbp. GP25 gene was then ligated with pAct resulted a pAct-GP25 plasmid with a size of 8.8 kbp. As there was only one restriction site applied, ligation can be occurred in two different directions. Thus, in order to determine the plasmid with the desired direction PCR analysis was carried out, and plasmid with the right direction was the one that sized approximately 2.1 kbp (no. 6, 17 and 20). High level of GFP gene expression was detected in all tissues analyzed, i.e. kidney, gil, spleen, and muscle 24 hours after injection which indicated that β-actin promoter of Japanese medaka could be active in common carp. Although not as high as on 24 hours, expression was still detected until day 7 and disappeared after day 28 post injection indicating that pAct-GFP can be remained and β-actin promoter of Japanese medaka can be active in muscle, gills and kidney at least a week after injection. The result of RT-PCR analysis on pAct-GP25 injected common carp showed that GP25 gene expression can be detected 14 days after injection in all tissues observed. The expression level differences between GFP and GP25 genes possibly related to the primer sensitivity on annealing process to cDNA template and the size of PCR targeted DNA fragment, as smaller PCR targeted product will be easier to be amplified than a bigger one. The length of PCR targeted DNA with GP25 primer was 1.8 kbp whereas GFP gene was 0.6 kbp.The challenge test on common carp results showed that all unvaccinated and negative control (not infected with KHV) was alive until the end of the experiment. Whereas, the survival of fish vaccinated with a dose of 12.5 was ranged from 60.9 and 96.7%. In relation to the activity of β-actin promoter that was used in this DNA vaccine, gene expression was occured 24 hours after vaccination and remained until 2 weeks. The control fish that was not injected by GP25 showed no GP25 expression either in the observed tissues or organs and resulted in a lower survival, i.e. 23.3%. The challenge test results has indicated that the fish vaccinated with GP25 at doses of 7.5 and 12.5 μg/100μL were able to produce KHV glycoprotein in their body which would be
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Judul: Efikasi Vaksin DNA Penyandi Glikoprotein Koi Herpesvirus pada Ikan Mas Stadia Benih melalui Perendaman
Abstrak: Koi herpesvirus (KHV) is a herpesvirus which especially infects and causes mass mortality to koi and common carp. Therefore the protection of carp from KHV infection is urgently needed. In this study, we developed an application of DNA vaccine containing gene encoding KHV glycoprotein by immersion method to increase survival of common carp against KHV infection. As many as 400 of 30-day-old common carp juveniles were immersed in the water containing 1.3 x 108 CFU/ml of the killed Escherichia coli cells carrying DNA vaccine. Treatments performed were variation of duration and frequency of DNA vaccine immersion; as follows: 1x 30 minutes, 1x 60 minutes, 1x 90 minutes, 2x 90 minutes, and 3x 90 minutes of immersion treatments. Reverse transcription polymerase chain reaction (RT-PCR) analysis showed that DNA vaccine was successfully expressed among the vaccinated fish. Twenty eight days post vaccination, the fish were challenged by injecting 10-4 ml/fish of KHV. The result demonstrated that vaccination by 30 minutes immersion; allowed the 61% of fish survived, this was significantly different (P<0.05) compared to control (without vaccination), but insignificantly different (P>0.05) compared to those of other treatments. The relative percent survival of vaccinated fish were also similar among different treatments (P>0.05). Vaccination by immersion have increased the survival rate about 2 fold compared to unvaccinated fish control (26.67%). Thus, DNA vaccine was effective delivered by immersion for 1x 30 minutes, and this technique can be useful to level up the resistance to KHV infection in common carp juveniles.
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Judul: Pengaruh Intensitas Curah Hujan, Kemiringan Lereng dan Sifat Fisik Tanah terhadap Erosi pada Berbagai Jenis Tanah
Abstrak: Fungsi tanah bagi tanaman adalah sebagai penyedia unsur hara, matrik tempat akar berjangkar dan tempat penyimpanan air tanah. Erosi tanah merupakan salah satu penyebab mundurnya fungsi tanah. Erosi terjadi karena pukulan butir-butir , I - . hu j an pada permukaan tdn'aah 'dm' "a.iran 'per- mukaan gang mengangkut butir-butir tanah sebagai hasil dispersi oleh butir hujan tersebut. Pada daerah-daerah gurun pasir, faktor penyebab erosi tanah adalah oleh'angin
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