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All Journal Jurnal Kimia (Journal of Chemistry) Jurnal Veteriner Jurnal Kedokteran Syiah Kuala Folia Medica Indonesiana Qanun Medika - Medical Journal Faculty of Medicine Muhammadiyah Surabaya ANNALES BOGORIENSES Biomolecular and Health Science Journal JUXTA: Jurnal Ilmiah Mahasiswa Kedokteran Universitas Airlangga Indian Journal of Forensic Medicine & Toxicology Berkala Penelitian Hayati Jurnal Natural Journal of Clinical Microbiology and Infectious Diseases Jurnal Widya Medika
Eddy Bagus Wasito
Departemen Mikrobiologi Kedokteran, Fakultas Kedokteran, Universitas Airlangga
Agriculture, Biological Sciences & Forestry Astronomy Biochemistry, Genetics & Molecular Biology Chemical Engineering, Chemistry & Bioengineering Chemistry Earth & Planetary Sciences Energy Health Professions Immunology & microbiology Materials Science & Nanotechnology Medicine & Pharmacology Neuroscience Physics Public Health Veterinary Other

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Construction of pY-Af Vector for Expression of Thermostable α-L-Arabinofuranosidase in Saccharomyces cerevisiae Wirajana, I Nengah; Puspaningsih, Ni Nyoman Tri; Wasito, Eddy Bagus; Kusuma, Soekry Erfan; Kimura, Tetsuya; Sakka, Kazuo
ANNALES BOGORIENSES Vol 14, No 2 (2010): Annales Bogorienses
Publisher : Research Center for Biotechnology - Indonesian Institute of Sciences (LIPI)

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.1234/51

Abstract

In this research,  construction  of expression  vector  for thermostable α -L-arabinofuranosidase  in Saccharomyces cerevisiae was conducted. BJ1824 was conducted The  Escherichia coli/S. cerevisiae  shuttle vector, pYES2 was  used  as  parental  vector  in  construction.  The  abfA  gene  encoding  α-L- arabinofuranosidase  from Geobacillus  thermoleovorans  IT-08  was  amplified  by  PCR,  in  which  the  plasmid  pTP510 was  used  as  a template. The amplification product was treated with  SacI and XhoI and then subcloned to the pYES2 vector, which was previously digested with  SacI and  XhoI. The recombinant plasmid was designated as pY-Af and propagated  first  in  E.  coli  Top 10,  and  then  transformed  into  S.  cerevisiae  BJ1824.  For  α- Larabinofuranosidase (AbfA) production, the yeast transformants were grown in YNBG selective medium and YPG rich medium, using galactose as an inducer. The AbfA activity was assayed by measuring the amount of p-nitrophenol (pNP) released  from  p-nitrophenyl-α-L-arabinofuranoside  (pNPA) substrate at pH 6.0 and 70 C  for  30  min.  The  recombinant  AbfA  activity  was  detected  in  either  of  culture  medium  (0.98%),  cellassociated (14.17%) and intracellular (84.85%) when recombinant yeast was grown in YPG rich medium.Key words: α-L-arabinofuranosidase; Saccharomyces cerevisiae; expression vector
Mechanism of Corneal Epithelial Cells Death by Infection of pseudomonas Aeruginosathrough Analysis Expression of Caspase-1, TNF, RIPK1, RIPK3, Caspase-3 in Rats Model Ismi Zuhria; Nurwasis; Eddy Bagus Wasito; Winarto; I Ketut Sudiana; Reny I’tishom; Nyilo Purnami
Indian Journal of Forensic Medicine & Toxicology Vol. 15 No. 3 (2021): Indian Journal of Forensic Medicine & Toxicology
Publisher : Institute of Medico-legal Publications Pvt Ltd

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.37506/ijfmt.v15i3.15802

Abstract

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Perbandingan kecepatan pertumbuhan Escherichia coli non ESBL dengan Escherichia coli ESBL Branandito Putra; Linda Dewanti; Eddy Bagus Wasito
Jurnal Kedokteran Syiah Kuala Vol 20, No 2 (2020): Volume 20 Nomor 2 Agustus 2020
Publisher : Universitas Syiah Kuala

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.24815/jks.v20i2.18498

Abstract

Abstrak. Penelitian ini bertujuan untuk mengidentifikasi perbedaan kecepatan pertumbuhan antara kelompok bakteri Escherichia coli NON ESBL dengan kelompok Escherichia coli ESBL. Jenis dan rancangan penelitian ini adalah penelitian eksperimental. Besar sampel sebanyak 13 isolat bakteri Escherihcia coli. Teknik sampling yang digunakan non random sampling. Variabel dalam penelitian ini meliputi Isolat Escherichia coli, media cair, fotometer, suhu inkubasi. Kecepatan pertumbuhan bakteri Escherichia coli NON ESBL, yaitu 2,206 ± 0,789. Sedangkan kecepatan pertumbuhan bakteri Escherichia coli ESBL, yaitu 1,963 ± 0,674. Nilai signifikasi Uji T sampel independen perbandingan kecepatan pertumbuhan bakteri Escherichia coli NON ESBL dengan kecepatan pertumbuhan bakteri Escherichia coli ESBL (p = 0,257) yang artinya tidak ada perbedaan yang signifikan.Hal tersebut terjadi karena jumlah sampel penelitian yang kurang dari jumlah minimal sampel yang diperlukan.Kata kunci: Escherichia coli, ESBL, Pertumbuhan.Abstract. This study aims to identify a difference of growth’s velocity between NON ESBL Escherichia coli with ESBL Escherichia coli. Type and design of this study was experimental. The sample size was 13 isolate of Escherichia coli. Sampling technique used was non random sampling. Variables in this study were Isolate of Escherichia coli, liquid media, photometer, temperature of incubation. The velocity of growth in NON ESBL Escherichia coli, was 2,206 ± 0,789. Meanwhile, The velocity of growth in ESBL Escherichia coli, was 1,963 ± 0,674. Significant value of T independent sample test  comparison of growth velocity between NON ESBL Escherichia coli  with ESBL Escherichia coli (p =  0,257) which means there’s no different significant. .It happens because an amount of total sample less than minimum standards amount of sample. Keywords: Escherichia coli, ESBL, Growth.
SUBKLONING GEN -L-ARABINOFURANOSIDASE (abfA) DALAM VEKTOR EKSPRESI pYES2 I Nengah Wirajana; Eddy Bagus Wasito; H.M. Soekry Erfan Kusuma; Ni Nyoman Tri Puspaningsih
Jurnal Kimia (Journal of Chemistry) Vol. 4, No. 2 Juli 2010
Publisher : Program Studi Kimia, FMIPA, Universitas Udayana (Program of Study in Chemistry, Faculty of Mathematics and Natural Sciences, Udayana University), Bali, Indonesia

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (512.632 KB)

Abstract

A Gene encoding -L-arabinofuranosidase (abfa) that had been cloned into recombinant E. coliDH5a/pTP510 was subcloned into Saccharomyces cerevisiae yeast system. The aim of subcloning of abfA gene intoS. cerevisiae is to express the thermostable -L-arabinofuranosidase (AbfA) enzyme in the host that is often termedas Generally Recognized As Safe (GRAS), so that this enzyme earn the broader application like in food and beverageindustries. The gene of abfA was subcloned by the amplification of Polymerase Chain Reaction (PCR) from plasmidpTP510 templat. A pair of primers, pFSacI-Af (forward) and pXhoI-Af (reverse) from designed this research wasused for the amplifcation. The PCR condition was performed as follows : beginning denaturation at 94oC for 5 min;PCR cycle 30 times that consisted of denaturation (94oC for 1 min), annealing (55oC for 30 s), andpolymerization/extension at 72oC for 2 min; and than final extension at 72oC for 7 min. The abfA gene, resulted wasinserted between GAL1 promoter and CYT1 terminator in the pYES2 expression vector. This ligation product wastransformed into E. coli TOP10 host. Restriction analysis of recombinant plasmid from this construction, thedesignated as plasmid pY-Af, showed the expected size, about 7,4 kb, which was the summation of sizes of parentalplasmid ( 5,9 kb) and fragment of DNA insert (1,5 kb).
Prevalensi dan Pola Gen Extended Spectrum B-lactamase Bakteri Usus Sapi Perah dan Penduduk Sekitar Peternakan di Surabaya Triffit Imasari; Wiwiek Tyasningsih; Eddy Bagus Wasito; Kuntaman Kuntaman
Jurnal Veteriner Vol 19 No 3 (2018)
Publisher : Faculty of Veterinary Medicine, Udayana University and Published in collaboration with the Indonesia Veterinarian Association

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (126.766 KB)

Abstract

Since identified in 1980s, the multiple drug resistant organisms such as Extended Spectrum Beta Lactamases (ESBL) producing bacteria is increasing. These bacteria Enterobacteriaceae strain are mostly resistant against third and also fouth generation cephalosporin. ESBL-producing bacteria are identified in both of human, environment and also in animal. There are three main ESBL genes that are commonly found namely SHV, TEM and CTX-M. The aims of this study were to explore the prevalence and pattern of ESBL gene among dairy cows and people around the farm. The faecal samples were collected from dairy cows and people around the farm, cultured on MacConkey agar supplemented with cefotaxim 1 mg/L, incubated at 37oC for 24 hours. Then the growing colony were tested for ESBL producer by Double Disk Synergy Test (DDST), then followed by Polymerase Chain Reaction (PCR) for ESBL gene. Total sampling technique with inclusion and exclusion criteria, Total 49 samples were collected, consisting of 25 dairy cows faeces and 24 people faeces. Among these, were identified 18 samples (72%) positive in dairy cows and 19 samples (79.1%) positive results in the people around the dairy farm. The ESBL gene, SHV, TEM, CTX-M were identified dairy cows were zero for SHV, TEM (12%), CTX-M (72%) while in people around the farm SHV (25%), TEM (16.7%), CTX-M (66.7%). There were significant different (p < 0.05) between dairy cows and people around the farm, of SHV ESBL gene and not different (p>0.05) of TEM and CTX-M ESBL gene respectively. The ESBL genes have spread among dairy cows and people around the farm.
Comparation of Phenotypic and Genotypic Profile of Carbapenemase Producing Escherichia coli Silvia Sutandhio; Budiono Budiono; Hardiono Hardiono; Kuntaman Kuntaman; Eddy Bagus Wasito; Maria Inge Lusida
Folia Medica Indonesiana Vol. 54 No. 1 (2018): March
Publisher : Faculty of Medicine, Universitas Airlangga

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (221.592 KB) | DOI: 10.20473/fmi.v54i1.8045

Abstract

Carbapenemase-producing Escherichia coli (E. coli) has caused trouble in therapeutic antibiotic selection. Carbapenemase screening procedure in laboratories is usually based on inacurate semi-automatic system. Confirmation and classification of carbapenemases according to Ambler can be done with combination of phenotypic methods, i.e., Modified Hodge Test (MHT), Sodium Mercaptoacetic Acid (SMA), and 3-Aminophenylboronic Acid (PBA). This study aimed to compare profiles of carbapenemase-producing E. coli which were confirmed and classified phenotypically with the genotypic profiles. E. coli isolates from urine specimens which were potential as carbapenemase-producers according to semi-automatic system BD Phoenix were phenotypically tested with MHT, SMA, and PBA. Isolates were grouped as carbapenemase-producers and non carbapenemase-producers. Phenotypic carbapenemase-producer isolates were classified based on Ambler criteria. All isolates were then tested with Polymerase Chain Reaction (PCR) for the presence of OXA-48, IMP1, IMP2, GES, VIM, NDM, KPC genes. Out of 30 isolates, 6 isolates (20.0%) were MHT positive, and 25 isolates (83.3%) were SMA positive, which indicated that most isolates produced were carbapenemase Ambler B. PCR confirmed 12 isolates (40.0%) had VIM gene which were classified as carbapenemase Ambler B. Phenotypic confirmatory test had 100% sensitivity and 22.2% specificity. Classification with phenotypic confirmatory test had 91.7% match with PCR. Phenotypic confirmatory test detected more carbapenemase than PCR. This low specificity may be caused by inappropriate use of diagnostic gold standard. PCR should not be used for routine carbapenemase confirmation because of vast diversity of carbapenemases. Phenotypic confirmatory test can classify carbapenemase according to Ambler classification.
The Carrier Rate of Extended Spectrum Beta Lactamase (ESBL) Producing Bacteria in Cockroaches (Periplaneta americana) in Hospital and Community Ardhiya Puspita; Radita Yuniar Arizandy; Eddy Bagus Wasito; Kuntaman
Folia Medica Indonesiana Vol. 57 No. 4 (2021): December
Publisher : Universitas Airlangga

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (1304.96 KB) | DOI: 10.20473/fmi.v57i4.17590

Abstract

Highlight:Bacteriologically for colonization of  ESBL producing Enterobacteriaceae in cockroaches (Periplaneta americana) were analyzed.The prevalence of ESBL producing bacteria among cockroaches in hospitals is bigger than in households.Abstract: Cockroach (Periplaneta americana) is one of the vectors in the environment that can transmit disease. Cockroaches can act as potential mechanical vectors of antibiotic resistant bacteria. Enterobacteriaceae is a gram-negative bacteria that has natural habitats in the digestive tract of humans and animals. Enterobacteriaceae that produce Extended Spectrum β-lactamases (ESBLs) have emerged as major pathogens in hospitals. The study analyzed the prevalence of ESBL producing bacteria in cockroaches that lived in hospitals and residential homes. In this study, a total of 200 cockroaches consisting of 100 cockroaches from the hospital environment and 100 cockroaches from the residential environment were analyzed bacteriologically for colonization of  ESBL producing Enterobacteriaceae. The specimen of the alimentary tract was taken and sub-cultured in MacConkey agar supplemented with cefotaxime 2 ug/ml. Growth colonies were suggested as an ESBL-producing bacteria, then were confirmed as ESBL producers by the Double Disk Synergy Test (DDST). The ESBL gene was detected by Polymerase Chain Reaction (PCR). Among 100 household cockroach samples, 14 (14%) were identified as ESBL producers, while 100 hospital cockroaches were 26 (26%) positive ESBL. The ESBL gene, in hospital cockroach were identified of CTXM 19 (19%), SHV 7 (7%), and not any TEM gene, while among household cockroaches were identified CTXM 2 (2%), SHV 11 (11%), and also not detected TEM ESBL gene. Among ESBL genes, only the CTXM gene was significantly different between household and hospital cockroaches.
Gram Negative Bacteria (Escherichia coli) Win Against Gram Positive Bacteria (Staphylococcus aureus) in The Same Media Neisya Intan Cahyaningtyas Agung Putri; Ramadhani Ramadhani; Eddy Bagus Wasito
Biomolecular and Health Science Journal Vol. 4 No. 2 (2021): Biomolecular and Health Science Journal
Publisher : Faculty of Medicine, Universitas Airlangga, Surabaya, Indonesia

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.20473/bhsj.v4i2.30177

Abstract

Introduction: Biodiversity of the microorganism in Indonesia lead to the large amount of patient with infection. Human can get infected in two different place, with different kind of bacteria that cause the infection. This may lead to bacteremia without knowing which bacteria type whose causing it, either the Gram positive or Gram negative bacteria, whereas the treatment of this two types of bacteria are different. The aim of this study is to determine the doubling time of the Gram positive and Gram negative bacteria when they are grown in the same lesion and the kinds of bacteria that we need to eliminate first.Methods: Staphylococcus aureus and Escherichia coli bacteria were used as samples in this study. Bacterial culture in nutrient broth with 0.5 OD turbidity were mixed then incubated in incubator with 35˚C. Every one hour within 24 hour, 0.01 ml of bacterial culture was taken in serial dilutionover time, varying between 106 – 1012, . It was then planted in nutrient agar plate with droplets technique. After it had been incubated for 24 hours, we counted the Colony Forming Unit per ml (CFU/ml) to time, then the doubling time of the bacteria. The result were then compared between the Staphylococcus aureus and Escherichia coli group.Results: Two tailed t-test result of the doubling time between Staphylococcus aureus dan Escherichia coli was < 0,05 (p=0,000) wich means that there is significant difference of the doubling time between Staphylococcus aureus (24,35 ± 2,23 munites), and Escherichia coli (18,37 ± 0,50 minutes). When grown in the same media, Gram positive bacteria (Staphylococcus aureus) had slower doubling time than Gram negative bacteria (Escherichia coli) as much as 1.32 times.Conclusion: In bacteremia with two possible kinds of bacterial suspect, we need to eliminate the Gram negative bacteria first.
PENGUJIAN IN VITRO XILOOLIGOSAKARIDA SEBAGAI KANDIDAT PREBIOTIK Asnia Zainuddin; Eddy Bagus Wasito; Ni Nyoman Tri Puspaningsih
JURNAL PENELITIAN BIOLOGI BERKALA PENELITIAN HAYATI Vol 14 No 1 (2008): December 2008
Publisher : The East Java Biological Society

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.23869/314

Abstract

The objectives of this study were to fi nd the in-vitro effect of xyloolygosaccharide on the cell count of Lactobacillus casei Shirota strain, to identify the effect of xyloolygosaccharide on the production of lactic, acetic, propionic and butyric acid (short chain fatty acid = SCFA), and to prove the effect of xyloolygosaccharide on the change of pH value.This was a laboratory experimental study using complete randomized design with 4 treatments, i.e, MRS broth media added xyloolygosaccharide with concentrations of 0% (control), 1%, 3%, and 5%, which were then inoculated with Lactobacillus casei Shirota strain and incubated for 6,12, 18, and 24 hours. The Lactobacillus casei Shirota strain cell count was counted using Drop plate. Data obtained were analyzed statistically using two way ANOVA with signifi cance level of 5%. The type and level of organic acids, i.e., lactic, acetic, propionic, and butyric acids (SCFA), formed in incubation time of 12 hours, were measured using gas chromatography and the change of pH value during incubation time was measured using pH paper. Results showed that xyloolygosaccharide addition MRS Broth media provided highly signifi cant interaction effect on the cell count of Lactobacillus casei Shirota strain (p<0.05). The result of gas chromatography showed that the addition of xyloolygosaccharide in MRS Broth media could increase Lactobacillus casei Shirota strain metabolism activity that produced lactic, acetic, propionic, and butyric acid. The reduction of pH value showed that the lowest pH value of 3.8 after the addition of 5% xyloolygosaccharide with incubation time of 24 hours. In conclusion, the addition of xyloolygosaccharide with concentrations of 1%, 3%, and 5% in MRS Broth media with incubation periods of 6, 12, 18, and 24 hours can increase Lactobacillus casei Shirota strain cell count, increase the metabolism activity of Lactobacillus casei Shirota strain, capable in producing SCFA in incubation time of 12 hours, and results in the reduction of pH value.
Construction of pY-Af Vector for Expression of Thermostable α-L-Arabinofuranosidase in Saccharomyces cerevisiae I Nengah Wirajana; Ni Nyoman Tri Puspaningsih; Eddy Bagus Wasito; Soekry Erfan Kusuma; Tetsuya Kimura; Kazuo Sakka
ANNALES BOGORIENSES Vol 14, No 2 (2010): Annales Bogorienses
Publisher : Research Center for Biotechnology - Indonesian Institute of Sciences (LIPI)

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (822.516 KB) | DOI: 10.14203/ann.bogor.2010.v14.n2.15-20

Abstract

In this research,  construction  of expression  vector  for thermostable α -L-arabinofuranosidase  in Saccharomyces cerevisiae was conducted. BJ1824 was conducted The  Escherichia coli/S. cerevisiae  shuttle vector, pYES2 was  used  as  parental  vector  in  construction.  The  abfA  gene  encoding  α-L- arabinofuranosidase  from Geobacillus  thermoleovorans  IT-08  was  amplified  by  PCR,  in  which  the  plasmid  pTP510 was  used  as  a template. The amplification product was treated with  SacI and XhoI and then subcloned to the pYES2 vector, which was previously digested with  SacI and  XhoI. The recombinant plasmid was designated as pY-Af and propagated  first  in  E.  coli  Top 10,  and  then  transformed  into  S.  cerevisiae  BJ1824.  For  α- Larabinofuranosidase (AbfA) production, the yeast transformants were grown in YNBG selective medium and YPG rich medium, using galactose as an inducer. The AbfA activity was assayed by measuring the amount of p-nitrophenol (pNP) released  from  p-nitrophenyl-α-L-arabinofuranoside  (pNPA) substrate at pH 6.0 and 70 C  for  30  min.  The  recombinant  AbfA  activity  was  detected  in  either  of  culture  medium  (0.98%),  cellassociated (14.17%) and intracellular (84.85%) when recombinant yeast was grown in YPG rich medium.Key words: α-L-arabinofuranosidase; Saccharomyces cerevisiae; expression vector
Co-Authors Achmad Basori, Achmad Agung Dwi Wahyu Widodo Alimsardjono, Lindawati Ardhiya Puspita Arthur Pohan Kawilarang Asnia Zainuddin Bambang Hermanto Branandito Putra Budi Utomo Budiono Budiono Eko Budi Koendhori, Eko Budi Erwin Astha Triyono H.M. Soekry Erfan Kusuma Hanindita, Meta Herdiana Hardiani, Kartika Hardiono Hardiono I Nengah Wirajana I'tishom, Reny Isnaeni Isnaeni Ivanna Kartuti Debora Kartuti Debora, Kartuti Kazuo Sakka Kimura, Tetsuya Kuntaman Kuntaman Kusuma, Soekry Erfan Linda Dewanti Lindarto, Wira W Margaretha Nathania Maria Inge Lusida Muhammad Amin Nasronudin Nasronudin Neisya Intan Cahyaningtyas Agung Putri Ni Nyoman Tri Puspaningsih Nisa Amnifolia Niazta Nur Aisiyah Widjaja, Nur Aisiyah Nurina Hasanatuludhhiyah Nurwasis Nurwasis Nyilo Purnami Radita Yuniar Arizandy Ramadhani Ramadhani Ratna Kusumawati Roedi Irawan S.Pd. M Kes I Ketut Sudiana . Sakka, Kazuo Silvia Sutandhio Soekry Erfan Kusuma SOETJIPTO . Suharyadi Sasmanto Tetsuya Kimura Triffit Imasari WINARTO Wiwiek Tyasningsih Yulia Sari Ismail Zuhria, Ismi