Indonesian Journal of Biotechnology
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Decolorization and detoxification of batik dye effluent containing Indigosol Blue-04B using fungi isolated from contaminated dye effluent
No full textFungi are capable of treating various synthetic dye effluents. Previously, we isolated seven strains of fungi from contaminated batik dye effluent at Banyumas, Central Java. The aims of this study were to screen the ability of these fungi to decolorize batik dye effluents containing Indigosol Blue-04B and to investigate the phytotoxicity effects of biodegraded effluent on the germination of corn seeds Zea mays L. and green bean seeds Vigna radiata (L.) Wilczek. In addition, the decolorized effluents were tested for toxic effect on the agriculturally important gram-positive and gram-negative soil bacteria Bacillus cereus and Azotobacter sp., Staphylococcus aureus and Escherichia coli, respectively. Study of decolorization showed that fungi were able to decolorize Indigosol Blue-04B batik dye effluents by 21.04% to 99.89% at room temperature after three days of incubation. The assay of phytotoxicity showed that both plumule and radicle length of Z. mays and V. radiata grown on the decolorized effluent was longer than on untreated effluent. The percentage of Z. mays and V. radiata seed germination in decolorized effluent was higher than in untreated effluent. There was no inhibition zone found around the decolorized effluent samples after incubating the bacteria for 48 hours. Aspergillus sp. 3 was the most effective for degradation and could be used for batik effluent mycoremediation processes
Amylolytic ability of bacteria isolated from termite (Coptotermes sp.) gut
No full textBSR 2, BSR 3, BSR 8, and BSR 9, different bacteria isolated from the termite gut, have been shown to possess cellulolytic activities, but their amylolytic ability has heretofore been unknown. This study attempted to fill in this knowledge gap. The formation of a clear zone using the iodine test showed that the bacteria were able to produce and secrete amylase. Based on the results, the best cultivation times for strains BSR 2, BSR 3, BSR 8, and BSR 9 were 6, 3, 2, and 2 d, respectively, yielding amylase activities of 2.59 ± 0.13 U/mg, 2.00 ± 0.08 U/mg, 1.67 ± 0.10 U/mg, and 1.55 ± 0.12 U/mg, respectively. BSR 2 had the highest amylase activity compared with the other bacterial isolates. The optimum ph for bacterial amylase activity of BSR 2 was 7.0, and the optimum temperature was 40°C. The molecular characterization of isolates BSR 2, BSR 3, BSR 8, and BSR 9 was based on 16S rRNA gene sequences. Isolates BSR 8 and BSR 9 were thus identified as Brevibacillus parabrevis and Brevibacillus sp. With similarities amounting to 92.48% and 95.91%, while the BSR 3 isolate was identified as Pseudomonas alcaligenes with a similarity of 94.29%, and the BSR 2 isolate could not be identified yet
Whole genome sequencing of Indonesian dengue virus isolates using next-generation sequencing
No full textIndonesia is a tropical country and hyperendemic for dengue. The disease prevalently affected Indonesian and it caused high morbidity and substantial economic burden. This vector-borne viral disease is caused by infection of dengue viruses (DENVs), which are the member of Flaviviridae family. While most of dengue studies in Indonesia focused on the epidemiology, the clinical aspects, the vectors, and to certain extent the virology, there were still gaps in the DENVs genomic aspects. Considering their high mutation rate, the DENVs were known for their high genetic diversity and it might affect the characteristics of the viruses. Comprehensive DENV genomic data were thus important for many aspects of disease management, including virus surveillance, pathogenesis, diagnostics, antiviral drug design, and vaccine development. We established in this study a method for DENV whole genome sequencing using the advanced Next-Generation Sequencing (NGS) and Nextera XT DNA library preparation kit, coupled with simplified bioinformatic analysis methods. The Indonesian DENVs from four serotypes were isolated from patients’ sera, while library was prepared from enriched templates and sequenced using Illumina NGS. Our study highlighted the potential of a robust NGS method in producing whole genome sequence of DENVs, which would be important for future dengue studies
Generation of recombinant scFv antibody against Ochratoxin A (OTA)
No full textOchratoxin A (OTA) is a mycotoxin commonly found in agricultural products and can accumulate in the blood and tissues after that consuming contaminated food. Recombinant single-chain antibody fragments (scFv) against OTA were selected from phage display libraries. After of one round of biopanning against BSA-conjugated OTA (OTA-BSA), 52 and 6 phage clones displaying scFv antibodies were isolated from human (Yamo I.3) and rabbit (Bozmix I.2) libraries. Two phage clones (one from each libraries, i.e., yOTA1e3 and bOTA2a9) showed binding to free toxin by competitive ELISA. The soluble scFv antibodies were produced by superinfecting phage clones into E. coli suppressor strain HB2151. The scFv genes from these two phage clones were sub-cloned into pKP300ΔIII vectors to generate scFv-AP fusions. The binding affinity (IC50) of antibody derived from human library was higher than those from rabbit library. The binding property of recombinant antibody in the form of scFv-AP was better than those of soluble scFv form. Cross-reactivity analysis indicated that the two recombinant antibodies did not cross-react with other soluble toxins, namely AFB1, DON, ZEN and FB. The ability to use the recombinant scFv-AP to detect contaminated toxins in agricultural product (corn) was demonstrated
Isolation of actinomycetes from maize rhizosphere from Kupang, East Nusa Tenggara Province, and evaluation of their antibacterial, antifungal, and extracellular enzyme activity
No full textActinomycetes are the one of the components of the rhizospheric microbial population and useful for producing secondary metabolites such as lytic enzymes, antibiotics, and antifungal. The aim of the study was to isolate the actinomycetes from maize rhizosphere collected from Kupang, East Nusa Tenggara. The screening was focused on the actinomycetes that showed the ability to produce antibacterial, antifungal, and extracellular enzymes such as amylase, cellulase, and protease. The actinomycetes were isolated using Humic-Acid Vitamin B (HV) agar media. The antagonistic assay was tested against Escherichia coli, Staphylococcus aureus, Sclerotium rolfsii and Fusarium oxysporum. Isolate JKP-8 was an isolate that showed the highest activity in inhibiting the growth of E. coli and S. aureus bacteria. Isolate JKP-5 showed the highest activity in inhibiting the growth of F.oxysporum. There were no actinomycetes isolates that showed an ability to inhibit the growth of S. rolfsii fungus based on dual culture assay. JKP-3 and JKP-4 isolates exhibited the highest ability to hydrolyze amylum, while JKP-5 and JKP-8 isolates exhibited the highest ability to hydrolyze CMC. The results of the amplification of 16S rRNA gene in selected potential isolates JKP 5 and JKP 8 indicated that both isolates belong to the genus Streptomyces
Determination of secondary and tertiary structures of cervical cancer lncRNA diagnostic and siRNA therapeutic biomarkers
No full textCervical cancer is one of the primary causes of mortality in women due to human papilloma virus (HPV) infection. The fingerprint of an HPV infection could be detected using a long non-coding RNA (lncRNA) biomarker, enabling it to be utilized in molecular diagnostics. The primary structure or sequences of RNA should be annotated within conventional bioinformatics tools. Therefore, this study aimed to determine the fine-grained 2D and 3D structures of lncRNA PVT1 and its respective siRNA inhibitors. lncRNA PVT1 sequences from Homo sapiens, Mus musculus, and Rattus norvegicus were retrieved from Genbank (NCBI). Prediction of the 2D structure and analysis of the interactions of the lncRNA and siRNA were performed using the Vienna RNA package. The 3D structure of the RNA was computed using the SimRNA and ModeRNA software programs. The results showed that lncRNA PVT1 from H. sapiens and M. musculus had a conserved region. However, the lncRNA from both H. sapiens and M. musculus showed a low conserved region, and the 2D structure could not be determined; thus, the annotation and 2D model focused only on H. sapiens. Both of their lncRNA PVT1 also had a short half-life in the cell. Based on the 3D modeling pipeline, the 3D model of lncRNA PVT1 showed the stability and possible function as molecules, while the PVT1 siRNA-lncRNA interaction analysis revealed that the molecules could bind well. Based on these findings, the structures of both lncRNA PVT1 and its siRNA have the potential to be utilized as biomarkers
Low cost and comprehensive pork detection in processed food products with a different food matrix
No full textThe adulteration of processed beef-based meat products with pork is a sensitive issue in Indonesia. In this study, we developed a detection method for the low cost identification of pork in processed meat products. We used the cost-efficient Taq DNA polymerase, DreamTaq Green PCR master mix (2x), and duplex PCR method to recognize pork simultaneously with 18S rRNA detection. A positive control containing a pork gene inserted into pGEM®-T easy was prepared, along with a negative control. The results of the duplex PCR were used to assess its specificity, detection limit, and its ability to recognize pork in processed meat products with a different food matrix. 18S rRNA detection was for confirming DNA integrity of DNA extracted from the processed food, while the positive control confirmed that the reagents were working well and the negative control confirmed a non-contamination problem. Following this, the duplex PCR was optimized and the optimum concentration primer for duplex PCR detection was found to be 3 µm for pork and 0.2 µm for 18S rRNA. As little as 3.125 ng of the DNA template could be used to detect whether a sample contained pork. Duplex PCR is a simple, fast, sensitive, specific, and low cost method of detecting pork in processed meat products
Evaluation of N-benzoylthiourea derivatives as possible analgesic agents by predicting their physicochemical and pharmacokinetic properties, toxicity, and analgesic activity
No full textThis study aimed to predict the physicochemical properties, pharmacokinetic properties (ADME), toxicity, and analgesic activity of 30 compounds of N-benzoylthiourea derivatives that are potential analgesic drugs. One of the mechanisms of action of N-benzoylthiourea derivatives is the inhibition of the cyclooxygenase-2 (COX-2) isoenzyme. An in silico test was performed by docking a compound that would predict its activity with the target COX-2 isoenzyme, PDB ID: 1PXX, using the MVD (Molegro Virtual Docker) program. The result of the docking was a form of energy bond indicated by the value of the rerank score (RS), where compounds that had lower RS values were predicted to have a higher activity. The pkCSM and Protox online tools were used to predict various physicochemical properties. Based on the RS values, the N-benzoylthiourea derivatives can be predicted to have lower analgesic activity than diclofenac, the reference ligand. Three of the N-benzoylthiourea derivatives—N-(2,4-bis-trifluoromethyl)-benzoylthiourea, N-(3,5-bis-trifluoromethyl)benzoylthiourea, and N-(3-trifluoromethoxy)-benzoylthiourea—had RS values of -90.82, -94.73, and -92.76, respectively, suggesting that these compounds were predicted to have analgesic activity relatively similar to diclofenac (RS value = -95.16). Furthermore, the majority of the N-benzoylthiourea derivatives were predicted to have good pharmacokinetic properties (ADME), and cause relatively low toxicity
NMR metabolite comparison of local pigmented rice in Yogyakarta
No full textPigmented rice may have a black or red color due to higher anthocyanin content in its grain. A natural antioxidant, many studies on anthocyanin have reported its positive effects on human health. This fact has spurred the development of pigmented rice as a functional food. This study aimed to compare the metabolite profiles of black and red rice. Three black rice cultivars, namely Melik, Pari Ireng, and Cempo Ireng Sleman, and two red rice cultivars, Inpari 24 and RC 204, were used. After husk removal, grain samples were ground in liquid nitrogen and dried with a freeze dryer. The dried samples were extracted using 50% MeOD4 (in a D2O phosphate buffer pH 6 containing 0.01% TSP as an internal standard). Metabolomic analysis was performed using 500 MHz NMR followed by multivariate data analysis. An orthogonal partial least squares-discriminant analysis (OPLS-DA) model ađer PCA was constructed to discriminate between the five different cultivars. The resulting OPLS-DA score plot revealed a clear separation between black rice and red rice. The metabolites that could influence the separation of red rice and black rice were valine, threonine, alanine, glutamate, galactinol, β-glucose, α-glucose, raffinose, and fumaric acid
The aqueous extract of Gerrardanthus macrorhizus caudex enhanced doxorubicin activity in MCF-7 human breast cancer cells
No full textGerrardanthus macrorhizus (GM) caudex, is traditionally used in cancer therapy by the Tetun people in Belu District, East Nusa Tenggara Province, Indonesia, where it is known as “akar batu”. This study aimed to explore the cytotoxic effects of G. macrorhizus caudex aqueous extract, as well as its combination with doxorubicin, on MCF-7 cells. Also investigated were the possible mechanisms of interaction through cell cycle progression and apoptosis induction. Single treatments of 5–320 µg/mL of the extract showed morphological alterations in MCF-7 cells, but did not show any cytotoxic effect. Combining the extract with doxorubicin resulted in a synergistic cytotoxic effect. Doxorubicin concentrations equivalent to 1/12, 1/8, and 1/5 fold of the IC50 combined with 20 µg/mL decreased viability to 48%. We then explored the combination effect of doxorubicin 0.4 µM with GM 5 and 20 µg/mL using a flow cytometer. A low concentration of the extract (5 µg/mL) combined with 0.4 µM of doxorubicin resulted in slight cell cycle modulation by G1, G2M arrested and apoptosis induction. The combination of doxorubicin and a higher concentration of the extract (20 µg/mL) did not show cell cycle modulation, and led to necrosis. Therefore, G. macrorhizus caudex at low concentrations has the potential to be developed further as a co-chemotherapeutic agent