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Articles by issue : Vol 10, No 2 (2016): June 2016
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Articles
Cloning, Expression, and Functional Characterization of Autoactivated Human Prethrombin-2 Synthetic Gene by Using Pichia pastoris SMD1168 As a Host

SUBROTO, TOTO, PERTIWI, WULAN, FADHILLAH, MUHAMMAD, HASAN, KHOMAINI, BUDIANTORO, OGI, ENUS, SUTARYA, SOEMITRO, SOETIJOSO

Microbiology Indonesia Vol 10, No 2 (2016): June 2016
Publisher : Indonesian Society for microbiology

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Abstract

Prethrombin-2 is a thrombin precursor that has important role in blood coagulation. It is the smallest precursor which is activated into thrombin by FXa prior to coagulation process. However, as a commercial theurapetic protein in fibrin sealant component, prethrombin-2 must be activated by ecarin before used. Thus, the production process of this protein needs further purification. In order to eliminate ecarin activation step and to increase production efficiency, we designed, cloned and expressed the recombinant autoactivated human prethrombin-2 in Pichia pastoris SMD1168. The variant was designed with 4 mutations, E40A, D47A, G48P, and E52A, following the result of a previous study. The synthetic variant gene was first optimized to conform with P. pastoris codon preference. The optimized synthetic gene was cloned in pD912 plasmid using XhoI and SacII restriction enzymes. The transformed P. pastoris was selected on agar plate supplemented with 1,000 µg.mL-1 Zeocin as a selection marker. This study showed that autoactivated prethrombin-2 was succesfully expressed extracellularly by P. pastoris SMD1168. The activity of recombinant autoactivated prethrombin-2 using a chromogenic substrate S-2238 was 0.540 unit/mg. Taken together, these results demonstrated that autoactivated human prethrombin-2 was successfully produced extracellularly in P. pastoris.

ITA registration form and back cover

Helianti, Is ( 1. BPPT 2. Indonesian Society for Microbiology )

Microbiology Indonesia Vol 10, No 2 (2016): June 2016
Publisher : Indonesian Society for microbiology

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Abstract

Primary Tupaia javanica Hepatocytes Cultures As In Vitro Replication System for Ape Hepatitis B Viruses

SURYA, MARYATI, ISKANDRIATI, DIAH, MARIYA, SILMI, SAEPULOH, UUS, PERMANAWATI, PERMANAWATI, SAJUTHI, DONDIN, PAMUNGKAS, JOKO

Microbiology Indonesia Vol 10, No 2 (2016): June 2016
Publisher : Indonesian Society for microbiology

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Abstract

Hepatitis B virus (HBV) is a DNA virus with liver as primary target organ.This virus caused chronic infection that can progress to cirrhosis, liver cancer and even death. In vitro model system of hepatocyte cultures is important and widely used to study a variety aspects of hepatitis B. Development of small animal Tupaia sp. for the in vitro model system is an alternative to the existinghepatocyte cultures. The specific purpose of the studyis to develop Tupaia javanica hepatocytes culture for HBV replication, and in a broader spectrum to answer the need for in vitro model of hepatocytes. Primary T. javanica hepatocytes (PTH) culture was successfully maintained for 14 days to reach 80% confluence, and infection of Javan gibbon HBV (GiHBV) and orangutan HBV (OuHBV) onto the culture on day 15 showed viral replication for up to eight days as measured by polymerase chain reaction (PCR). PCR quantification indicated that the highest copy number of DNA virus was detected onday two anddecreased until day 8 after infection. Cell receptor for HBV attachment, known as sodium taurocholate cotransporting polypeptide was expressed on the surface of PTH and shown as green luminenscent when observed by immunofluorescence assay. Sequence of partialS gene from the apes HBVs after the viruses have been infected to the PTH showed amino acid identity to their wildtype as high as 99.29% for GiHBV and 95.71% for OuHBV. This study suggested that the PTH can support the replication of GiHBV and  OuHBV. 

Diversity of Culturable Bacterial in Various Parts of Luwak’s (Paradoxurus hermaprodithus javanica) Gastrointestinal Tract

SUHANDONO, SONY, SETIADI, HERI, KRISTIANTI, TATI, KUSUMA, ALI BUDHI, WEDARINGTYAS, ANDINI WARIH, DJAJADI, DEMI TRISTAN, ARYANTHA, I NYOMAN PUGEG

Microbiology Indonesia Vol 10, No 2 (2016): June 2016
Publisher : Indonesian Society for microbiology

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Abstract

Luwak coffee is a highly-priced coffee produced exclusively by the palm civet or luwak (Paradoxurus hermaphrodites ssp.). The purpose of this study was to determine the diversity of culturable bacteria in the gastro intestinal tract of luwak. The bacterial isolates were phenotypically characterized by their morphology and molecularly by analysis of their1,500bp 16s rDNA sequence. The results showed that Enterobacter cloacae and Lactobacillus brevis were found all over luwak’s digestive tract. Enterobacter cloacae was the most common species. The most diverse bacterial population was found in small intestine. Seven bacterial generawere successfully identified from the small intestine and colon, compared to only five genera found in the stomach.

Nitrogen Fixing Potential of Endophytic Bacteria Isolated from Aloe barbadensis Miller and Aloe sp.

WANGSA PUTRIE, RAHAYU FITRIANI, WIDOWATI, TIWIT, LEKATOMPESSY, SYLVIA J R, SUKIMAN, HARMASTINI

Microbiology Indonesia Vol 10, No 2 (2016): June 2016
Publisher : Indonesian Society for microbiology

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Abstract

Aloe is a crassulacean acid metabolism (CAM) species that are known to live in extreme enviroment such as drought condition. Nitrogen fixation procces influenced by the ability of plants to adapt in drought condition. Endophytic bacteria from Aloe and their ability for nitrogen fixation were little reported, but potential and its relationship between the ability for nitrogen fixing with resistance to drought conditions have not been reported. This research aimed study the endophytic bacteria from two varieties of aloe, namely Aloe barbadensis Miller and Aloe sp. in their ability on conducting the nitrogen fixing process and its relationship with resistance to drought. Characterization of endophytic bacteria were carried out by morphological observation of colony, Gram staining and molecular identification. Screening of nitrogen fixation was done using nitrogen-free semisolid NFb malate medium. Endophytic bacteria from Aloe sp. more than A. barbadensis in their potency of nitrogen fixation which related with habitat where their planted. A total of 40% of the endophytic bacteria isolates from the leaves of the aloe var. A. barbadensis and 62.5% of isolates from var. Aloe sp. are known to have a better ability to fixing nitrogen than the others. Isolates A barbadensis AB 12 and Aloe sp. AS 8 were the best isolates from each varieties on ability for nitrogen fixation. Based on 16S rRNA gene analysis those two selected isolates belonged to Bacillus methalotropicus strain DA 16-5 and Bacillus aryabhattai strain B8W22. 

Antimicrobial Activity of Lactic Acid Bacteria from Bamboo Shoot Pickles Fermented at 15 oC

HARTAYANIE, LAKSMI ( Fakultas Teknologi Pertanian UNIKA Soegijapranata Semarang ) , LINDAYANI, LINDAYANI, MURNIATI, MONIKA PALUPI

Microbiology Indonesia Vol 10, No 2 (2016): June 2016
Publisher : Indonesian Society for microbiology

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Abstract

Lactic Acid Bacteria (LAB) produces natural antimicrobial compounds that can inhibit and prevent the growth of spoilage bacteria. LAB can be isolated from fermented food such as pickles which fermented at  cool temperature. The objective of this research to isolate and to obtain  LAB from yellow bamboo (Dendrocalamus asper) shoots pickles that has antimicrobial activity against Escherichia coli  and Staphylococcus aureus.  It was made by submerged yellow bamboo shoots in 2.5%   of brine solution and kept into sealed container then fermented at chiller (15oC) temperatures for 10 days. LAB was isolated using MRS agar and identified base on their morphological, physiological and biochemical characteristics. The result showed that LAB isolates identified as Lactobacilli and had antimicrobial activity against Escherichia coli  and Staphylococcus aureus.  All Lactobacilli (21 isolates) that was isolated from fermentation at 15oC were homofementative.