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TRANSPOSITION AND EXPRESSION OF GEP GENE IN THE GENOME OF Vibrio harveyi TO MONITOR ITS ADHERENCE IN SHRIMP LARVAE Suwanto, Antonius
BIOTROPIA Vol. 13 No. 1 (2006): BIOTROPIA Vol. 13 No. 1 June 2006
Publisher : SEAMEO BIOTROP

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (534.675 KB) | DOI: 10.11598/btb.2006.13.1.74

Abstract

Expression of green fluorescent protein encoded by GFP gene in Vibrio harveyi was investigated to understand the ability of the gene as a molecular marker for adherence of this pathogenic Vibrio in shrimp larvae. The GFP gene was inserted into pUC18Not and pUTmini-Tn5 to generate a recombinant plasmid pWGO2 and pWGO3, respectively, which was transferred into the three isolates of V. harveyi employing diparental mating. Recombinant E. coli carrying pWGO2 and pWGO3 resulted in green-fluorescent colonies and cells due to the production of GFP. However, al1 of mini-Tn5, including mini-Tn5-gfp were not successful1y transferred to V. harveyi. Therefore, we used mini-Tn10 (pLOFKm-gfp) for inserting of gfp gene into V. harveyi genome. Although we could obtain relatively high (l0 pangkat -8) transconjugans employing Tn10, only one of Tnl0 derived isolate of V. harveyi G3 (G3-Tn1Ogfp) showed gfp expression and was further employed for adherence assay. Fluorescent 03-TnlOgfp cells could be observed inside the digestive tract of shrimp larvae and could be distinguished from vibrio that naturally exist in shrimp larvae.
PRESENCE OF hemA-LIKE AND hemT-LlKE GENES IN A JSUMBER OF ANOXYGENIC PHOTOSYNTHETIC BACTERIAL ISOLATES FROM INDONESIA AND SOIL SAMPLES FROM BOGOR AREA AINI, NURUL; SUWANTO, ANTONIUS
BIOTROPIA No. 15 (2000)
Publisher : SEAMEO BIOTROP

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (162.845 KB) | DOI: 10.11598/btb.2000.0.15.158

Abstract

The Rhodobacter sphaeroides hemA and hemT are known to encode a distinct 5-aminolevulinic acid (ALA)-synthase isozyme. This enzyme catalyzes the first and rate limiting step in ALA biosynthesis through the C4 pathway. This study was carried out to detect hemA-\\ke and hemT-\\ke genes in twenty Anoxygenic Photosynthetic Bacterial (APB) isolates from several wetland areas in Indonesia, and four DNA samples that were isolated from four soil samples obtained from Bogor area. Hybridization techniques of Southern and dot blot were used, using hemA and hemT fragment as probes. Southern hybridization analyses indicated the presence of hemA-\\ke gene in five of APB isolates, i.e., MB15, MB16, MB21.2, MB55 and MB6, whereas hemT-\\ke gene was detected only in MB15. Dot blot hybridization analyses suggested that the soil samples from waterlogged paddy-field, dry paddy-field as well as a mud pond were predominantly occupied by prokaryotic organisms which harboured hemA-]\ke gene. However, /iem7"-like sequences were also found in soil sample from dry paddy-field. Key words:    hemA-\\ks gene / hemT-\\ke gene / Southern hybridization analysis / dot blot hybridization analysis.
Enhanced Lipase Production in Pichia pastoris via Multiple Copies of Bacterial Lipase Genes and Co-expression of the HAC1 Gene Puspitasari, Esti; Rustam, Yepy Hardi; Satya, Andreas Adhi; Suwanto, Antonius; Wahyudi, Aris Tri; Astuti, Rika Indri
HAYATI Journal of Biosciences Vol. 33 No. 2 (2026): March 2026
Publisher : Bogor Agricultural University, Indonesia

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.4308/hjb.33.2.288-296

Abstract

The hac1 gene, a key regulator of the untranslated protein response (UPR), was co-expressed in Pichia pastoris GS115 to enhance the production of a lipase from Geobacillus stearothermophilus. Multicopy lipase constructs (1X and 4X) were transformed with pPICZAwbe_hac1, generating GS115/T1.2RQ(1X)_hac1 and GS115/T1.2RQ(4X)_hac1 strains. The GS115/T1.2RQ(1X)_hac1 strain showed an 186% lipase activity after 120 hours versus the control (100%), while the GS115/T1.2RQ(4X)_hac1 strain showed a faster initial increase (38% at 48 hours) and 28% at 120 hours, which was beneficial for efficient enzyme production. Overexpression of the hac1 gene enhances lipase production because it activates UPR genes when the endoplasmic reticulum is stressed due to a large number of recombinant proteins and forms proteins that are not appropriately folded. SDS-PAGE and tributyrin plate assays confirmed extracellular lipase expression (~43 kDa). These results demonstrate that hac1 co-expression significantly (p = 0.01)  enhances lipase production in Pichia pastoris, especially in lower-copy constructs. This is the first report of co-expressing hac1 with Geobacillus stearothermophilus lipase genes in yeast. The findings are expected to contribute to developing more efficient microbial cell factories for producing industrial enzymes.
Expression of An Immunogenic Intimin Fragment of EHEC O157:H7 in Escherichia coli Periplasm under The Control of A Rhamnose-Based Regulated Promoter Hariyatun, Hariyatun; Suwanto, Antonius; Kusharyoto, Wien
Annales Bogorienses Vol. 18 No. 1 (2014): Annales Bogorienses
Publisher : BRIN

Show Abstract | Download Original | Original Source | Check in Google Scholar

Abstract

Intimin is the main adhesin of Enterohemorrhagic E. coli (EHEC) O157:H7 bacteria which are the most common leading infectious cause of bloody diarrhea and acute kidney failure in children who develop hemolytic uremic syndrome (HUS). Intimin is required for persistent bacterial colonization to eukaryotic host cell and its receptor-binding activity is localized at the C-terminus 282 amino acids (Intimin282). Thus, Intimin282 is an attractive antigen candidate that could be useful in vaccine and diagnostic systems against EHEC infections. Previous studies had reported expression of Intimin in E. coli cytoplasm using commonly used prokaryotic expression systems. However, it usually encountered several problems, i.e. low expression level, leaky expression, inclusion body formation, and truncated protein. The pRHA vector, which is tightly regulated by Lrhamnose and D-glucose, represents a viable alternative E. coli expression system to overcome such problems. Moreover, E. coli periplasm has an advantage of maintaining protein functionality by providing an oxidative environment that is more efficient than cytoplasm. However, to date there is no study about Intimin expressionusing pRHA expression system and/or in E. coli periplasm. Accordingly, we constructed a recombinant pRHA vector harbouring the respective gene to investigate the expression of an immunogenic Intimin fragment of EHEC O157:H7 in E. coli periplasm. The gene encoding His6-tagged Intimin282 (Int282) together with pelB signal sequence was cloned into the pRHA vector, subsequently expressed in E. coli JM109 and purified. Expression and purification of Int282 were verified by SDS-PAGE and Western blot. The result showed that Int282 was successfully expressed in E. coli periplasm with a protein size of approximately 32 kDa, which corresponded with the predicted size of the protein based on its amino acid sequence.
DYNAMICS OF MICROBIAL COMMUNITY DURING TEMPEH FERMENTATION Suwanto, Antonius
BIOTROPIA Vol. 28 No. 1 (2021): BIOTROPIA Vol. 28 No. 1 April 2021
Publisher : SEAMEO BIOTROP

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (524.271 KB) | DOI: 10.11598/btb.0.0.0.820

Abstract

Tempeh is sliceable soybean-cake fermented by Rhizopus oligosporus. Various bacteria were detected in tempeh employing cultivation technique with limited information about their origin or sources. The present study aimed to examine the source/s of bacterial community in tempeh by combining metagenomics analysis and cultivation technique. Samples were obtained from a number of steps in tempeh production employing two-times boiling of soybean (WJB tempeh production). All samples were plated on Enterobacteriaceae and Lactic Acid Bacteria medium. Total DNA were extracted directly from tempeh for metagenomics analysis, employing High-Throughput Sequencing (HTS) and cloned 16S rRNA genes. Firmicutes and Proteobacteria were the predominant and second dominant bacteria existed in fresh tempeh (FT) obtained by metagenomics analysis. In contrast, cultivation technique showed that Proteobacteria was the predominant phylum, suggested that most of the Firmicutes were not culturable. FT was dominated by Lactobacillus and Acetobacter. Both FT and soaking water (SW) were dominated by same species of Lactobacillus, i.e. L. delbreuckii and L. mucosae, indicated that SW was probably the source of bacterial community established in the final product of fermentation. Predominant bacteria in starter culture (SC), Acinetobacter, was not detected in FT, indicating that bacteria in SC might not play significant role in bacterial community development in FT.
GENETIC DIVERSITY OF AMPICILLIN-RESISTANT Vibrio ISOLATED FROM VARIOUS STAGES OF TIGER SHRIMP LARVAE DEVELOPMENT WIDANARNI, WIDANARNI; SUWANTO, ANTONIUS
BIOTROPIA No. 15 (2000)
Publisher : SEAMEO BIOTROP

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (219.218 KB) | DOI: 10.11598/btb.2000.0.15.160

Abstract

This research was carried  out to  study  genetic  diversity of ampicillin-resistant Vibrio  from various  stages of tiger  shrimp larvae  (Penaeus Monodon) development from,Tambak Inti Rakyat hatchery, near Labuan, West Java, Indonesia. A total of 25 ampicillin-resistant Vibrio isolates were isolated using thiosulphate  citrate bile-salt  sucrose  agar (TCBS-Agar) and seawater  complete agar (SWC-Agar). Physiological and biochemical characterization showed that the isolates could be grouped into only two species, i.e. V. harveyi from the egg stage; and V. metschnikovii from larvae and post-larval stage (i.e nauplius, zoea, mysis, PLi, PL5, PL,0, and PL,5). These isolates were also present in their respective rearing water  of  each stage  and  some natural feed.  Schizotyping analysis employing restriction endonuclease Noll (5'-GC4GGCCGC) indicated that the isolates could be grouped into  at least  13 different  genotypes.  Therefore,  schizotyping  was  more discriminative than physiological characterization. This study showed that particular groups of Vibrio colonized all stages of shrimp larvae and demonstrated closed phylogenetic relationship. These groups of Vibrio might be  the dominant microbiota which could suppress the development of other Vibrio including the pathogenic Vibrio. Key words : Shrimp/ampicillin-resistant K/fcno/schizotyping
A NOVEL INTEGRON IN THE GENOME OF ESCHERICHIA COLI ISOLATED FROM INDONESIAN MONITOR LIZARD (VARANUS SPP). PUSPITASARI, ESTI; SUWANTO, ANTONIUS; MALIK, AMARILA; ERDELEN, WALTER
BIOTROPIA No. 16 (2001)
Publisher : SEAMEO BIOTROP

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (97.942 KB) | DOI: 10.11598/btb.2001.0.16.163

Abstract

The genotype of antibiotic resistance  in natural isolates of Escherichia coli was determined  through integron detection and  characterization of the associated antibiotic  resistance. E. coli SG2 isolated from Varanus salvator  of Java demonstrated  resistance  to spectinomycin (50ng/ml)  and streptomycin (SOng/ml). Integron detection indicated  that eight isolates out of nine E. coli  isolates possessed a conserved segment of the integron. Amplification of  the inserted cassette of the integron in this SG2 isolate yielded a 1-kb DNA fragment. Sequence analyses indicated that this fragment was homologous with aad gene, which confirmed  the resistance  to spectinomycin/streptomycin. This is the first report on the presence of integron in the E. coli isolated from the environment. Key words: Integron / antibiotic resistance / Escherichia coli
CHARACTERIZATION OF THREE BENZOATE DEGRADING ANOXYGENIC PHOTOSYNTHETIC BACTERIA ISOLATED FROM THE ENVIRONMENT SURYANTO, DWI; SUWANTO, ANTONIUS; MERYANDINI, ANJA
BIOTROPIA No. 17 (2001)
Publisher : SEAMEO BIOTROP

Show Abstract | Download Original | Original Source | Check in Google Scholar | Full PDF (152.215 KB) | DOI: 10.11598/btb.2001.0.17.166

Abstract

Three  anoxygenic photosynthetic bacteria, DS-1, DS-4 and Cas-13, have been examinated  for  themorphological and physiological  properties. All strains were rod-shape cells with  a swollen terminal  endGram  negative, motile, non-halophilic, non-alkalophilic  and non-acidophilic,  and capable of utilizinbenzoate aerobically and photo-anaerobically. Sequence analysis of part of 16S rRNA genes showed that DS1 and Cas-13 were closely related to Rhodopseudomonas palustris Strain 7 with a similarity of 97%, whereaDS-4 may not be closely related to the former two strains with a similarity of 78% based on the constructephylogenic  tree. Spectral analysis indicated that the three  bacteria  had  bacteriochlorophyl  a  and normaspirilloxanthin series. Growth in medium enriched with vitamin and supplemented with benzoate as their sole C-sources wabetter than in medium without vitamin. Benzoate degradation in medium with vitamin was accelerated. Thability  to grow on benzoate without added vitamins indicated  that  the bacteria were able to synthesize  theown vitamins. Key words: anoxygenic photosynthetic bacteria/ benzoate degradation/ 16S rRNA gene.
CHARACTERIZATION OF XYLANASE FROM A XYLANOLYTIC- THERMOPHILIC BACTERIUM ISOLATED FROM GUNUNG PANCAR HOT SPRING, WEST JAVA SUWANTO, ANTONIUS; SURYANTO, DWI; MERYANDINI, ANJA
BIOTROPIA No. 17 (2001)
Publisher : SEAMEO BIOTROP

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.11598/btb.2001.0.17.167

Abstract

A xylanolytic-thermophilic bacterium (IT-08) was isolated from Gunung Pancar Hot Spring after two days of enrichment in Modified Thermus Medium (MTM) supplemented with 0.5% oat spelt xylan. 16S-rRNA sequence analysis indicated that IT-08 resembles Bacillus thermoleovorans, a species of thermophilic bacteria. When grown on xylan containing media, IT-08 produces a thermoactive xylanase. Xylanase from IT-08 was active at temperatures between 40 and 100°C, at pH values between 4.0 and 9.0 with optimum values obtained at 80°C and pH 6.0, respectively. SDS-PAGE and zymogram analysis demonstrated that a crude xylanase complex of IT-08 comprised two active bands with molecular masses of 78 and 60 kDa. Keywords: xylanase / xylanolytic-thermophilic bacteria
ADHERENCE AND PATHOGENICITY ASSAY OF VIBRIO HARVEYI IN TIGER SHRIMP (PENAEUS MONODON) LARVAE FOR SCREENING BIOCONTROL AGENT SUWANTO, ANTONIUS; HALA, YUSMINAH; AFFANDI, RIDWAN; Jr., MUHAMMAD ZAIRIN
BIOTROPIA No. 18 (2002)
Publisher : SEAMEO BIOTROP

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.11598/btb.2002.0.18.171

Abstract

Rifampicin-resistant marker was employed as a reporter to detect the adherence and colonization of V. harveyi  in shrimp larvae. Vibrio harveyi P1B  and YA32.2 were isolated from dead shrimp larvae in Besuki, Northern Coast of East Java, while V. harveyi HB3, was isolated from pristine sea water in Pacitan, Southern Coast of East Java. Vibrio metschnikovii used as biocontrol agent was isolated from healthy shrimp larvae in Serang, West Java. Spontaneous mutation was conducted to generate V. harveyi P1B, YA32.2 and HB3 resistant to rifampicin. These mutants exhibited similar survival ability to their parental (wild type) strains. Significant larval mortality was observed in shrimp larvae inoculated with YA32.2 than that of larvae inoculated with P1B. Larvae  inoculated with HB3 showed the lowest mortality. Bacterial cell count of Vibrio Rf*  in dead  larvae were 103-104 cells/larvae. Isolates of Vibrio metschnikovii Z and M as biocontrol candidates effectively reduced the growth and adherence ability of YA32.2 to shrimp larvae. Larval mortality in rearing water inoculated simultaneously with YA32.2 and V. metschnikovii was lower than the one inoculated with YA32.2 alone. Therefore, Vibrio metschnikovii Z or M could be developed as an effective probiotic or biocontrol agent for V. harveyi in shrimp hatcheries. Key words :   Biological control/Vibrio metschnikovii/shrimp \arvae/Penaeus mwu«fon/pathogenicity assay/Vibrio harveyi
Co-Authors . EFRIWATI . Melani . NURHAIMI-HARIS A A Hermosaningtyas A A Hermosaningtyas, A A ADI YULANDI Agus PURWANTARA Agus Purwito Agustin Wydia Gunawan Agustina, Delia Aini Nurul Alfred Michael ALINA AKHDIYA RUSMANA Amarila Malik Andi Khaeruni Andi Khaeruni Andreas Adhi Satya Andreas Adhi Satya Anja Meryandini ARI SUSILOWATI Arifudin Rafif , Muhammad Ghildan Arild Ranlym Arifin Arina Amalia Putri Aris Tjahjoleksono Aris Tri Wahyudi ARTINI PANGASTUTI Asmini Budiani Basil J NIKOLAU BIBIANA W LAY Budi Tjahjono Budinarta, Widyah C Hanny Wijaya Cahya Prihatna CAHYA PRIHATNA Cahya Prihatna CECILIA ANNA SEUMAHU CECILIA ANNA SEUMAHU DEBORA HADISUSANTO Delia Agustina Desniar - - DIAH ISKANDRIATI DIANA ELIZABETH WATURANGI Djoko Santoso Dondin Sajuthi DWI SURYANTO Dyah Kusuma Anggraini EDI HUSEN Edi Husen Edi Husen ERDELEN, WALTER ERNIN HIDAYATI Esti Puspitasari ESTI PUSPITASARI Esti Utarti Eunice Limantara EVELINE AYU Felicia Felicia Felicia Kartawidjajaputra Ferymon Mahulette Ferymon Mahulette Ferymon Mahulette, Ferymon G. A. Wattimena Gayuh Rahayu Griselda Griselda Griselda Herman Natadiputri Griselda, Griselda Hajrial ASWIDINNOO HAJRIAL ASWIDINNOOR Hariyatun Hariyatun Hariyatun, Hariyatun Hariyatun, Hariyatun HARLING, ROB HENAWIDJAJA, MAGGY T Hermosaningtyas, Anastasia Aliesa Iman Rusmana Inez H.S. Loeddin Suharsono It Jamilah IVAN HANJAYA JOANITA SADELI Jr., Muhammad Zairin KATHARINA JESSICA Kusharyoto, Wien Kusharyoto, Wien Lilis Nuraida Linda Wati LISTYA UTAMI KARMAWAN Ludovika Jessica Virginia Maggy T Suhartono Maggy T Suhartono MAGGY T. SUHARTONO Maggy T. Suhartono MAGGY THENAWIJAYA SUHARTONO Maria Dita Febriani Lumban Gaol Maria Indah Purnamasari Maria Indah Purnamasari Maria Indah Purnamasari Maria Sugiharti Meity S. Sinaga MEITY SURADJI SINAGA Muhamad Azwar Syah Muhammad Agus Muljanto Ni Nyoman Tri Puspaningsih NISA RACHMANIA MUBARIK NURITA TORUAN-MATHIUS PRIHASTO SETYANTO Prihasto Setyanto Prihatna , Cahya Puspitasari, Esti QURROTA A’YUN R. Sapto Hendri Boedi Soesatyo Raden Ajie Syahbarie RAHAYU WIDYASTUTI RASTI SARASWATI Rasti Saraswati RASTI SARASWATI Rasti Saraswati RATIH DEWI HASTUTI RATNA SETYANINGSIH RATNA SETYANINGSIH RIDWAN AFFANDI Rika Indri Astuti ROHANI CINTA BADIA GINTING Rustam, Yepy Hardi Satya, Andreas Adhi Sheila Sutanto Sheila Sutanto, Sheila Sumardi Suryo Wiyono SUSAN SOKA SUSILOWATI1 SUSILOWATI1 Tan Watumesa Agustina TARUNI SRI PRAWAST MIEN KAOMINI ANY ARYANI DEDY DURYADI SOLIHIN Tati Barus URWADARIA, T RESNAWATI P VICKY MEICY Widanarni Widanarni WIDANARNI WIDANARNI Wien Kusharyoto Yasinta Ratna Esti Wulandari Yepy Hardi Rustam Yogiara Yogiara YULIN LESTARI Yusminah Hala