Adenovirus type 5 (Ad5) is one of the vaccine vectors, including the COVID-19 vaccine. Pre-existing immunity to Ad5 may suppress the immunogenicity and efficacy of adenovirus vectored vaccine. The neutralizing antibodies are directed specifically toward seven hypervariable regions (HVR) of hexon proteins located on the outer surface of the capsid. This study aims to design an Ad5 vector that may circumvent anti-Ad5 immunity by designing a chimera Ad5 vector with the sequence of Ad26 HVR (Ad5HVR26) using in silico approach. Substitution of the Ad5 HVR DNA sequence may affect the alternative splicing process of adenovirus mRNA, which then influence the protein product. The splice site prediction of Ad5HVR26 chimera vector was found at HVR5, 6, and 7. The codon change in the splice site was performed to decrease the possibility of incorrect splicing, while retaining the original amino acid sequence. The HVR substitution in chimera vector Ad5HVR26 may also affect the interaction of hexon in the capsid. The HVR2 and HVR4 hexon proteins individually interact with other hexon proteins and IX protein. Thus, two designs of the Ad5HVR26 chimera vector were created in this research. The first design was the Ad5 chimera vector with complete substitution of HVR hexon by Ad26 sequence, with codon modification on the splice site. The second design was Ad5HVR26 chimera vector without the HVR2 and HVR4 substitution to maintain the hexon protein interaction with the capsid proteins. Production of the designed vectors are needed to prove the reduction of vector neutralization by pre-existing immunity.
Cases of food poisoning often occur due to food contamination caused by pathogenic bacteria. One of the pathogenic bacteria is Vibrio parahaemolyticus which is found in seafood. Thus, a fast, accurate and specific detection method is needed. The purpose of this study was to quickly detect Vibrio parahaemolyticus bacteria in seafood samples targeting the ToxR gene using Real Time PCR. In a previous study, gradient PCR was used to optimize ideal annealing temperature ranges from 53-62°C and revealed that 58°C produced the best outcomes for the ToxR primer with a size of 171 base pairs. Real-Time PCR was utilized to amplify, specify, and test for sensitivity under the ideal conditions from the PCR Gradient. The confirmation results show that the primer pairs could amplify ToxR of Vibrio parahaemolyticus with the amount of concentration as much as 50 ng/µL with Ct 10,69 and 10,32 and melting curve at temperature 82,18°C and 82,23°C. This primer pair can also distinguish non-target bacteria with different Ct and melting curve temperature. The sensitivity assay for this primer can amplify DNA templates at concentration 0,0032 ng/µL. Shrimp samples that are contaminated artificially can still be detected at Ct 13,02 and Ct 13,09. Based on these results, it can be concluded that Real Time PCR with ToxR primer can be applied to develop a detection kit for Vibrio parahaemolyticus in seafood.
This study investigates the effect of fermentation using lactic acid bacteria and yeast as starters on the formation of flavonoid compounds and the antioxidant activity of cacao beans. The fermentation process were divided into 4 groups: F1: spontaneous fermentation, F2: fermentation using Lactic Acid Bacteria (LAB), F3: fermentation using yeast and F4: fermentation using LAB and yeast. The extraction process was done using ethanol. Flavonoid content was analysis using spectrophotometer assay. The antioxidant activity was analyzed by 1,1-difenil-2-pikrilhidrazil (DPPH) method. All ethanol extract samples of fermented cacao beans contained alkaloids, polyphenols, flavonoids, and tannins. The flavonoid compounds from ethanol extract of cacao beans in F1 is 4.35 ± 0.20 mg/L, F2 (5.64 ± 0.05), F3 (5.37 ± 0.17), and F4 (5.99 ± 0.23 mg/L). The antioxidant activity of cacao bean fermentation extracts using starter were increase compared to the spontaneous fermentation extract (F1). The antioxidant activity in F2 increased to 46.45 ± 2.00%, F3 (49.05 ± 0.58%), and F4 (50.33 ± 0.43%), while the antioxidant activity of F1 was 42.31 ± 0.66%. IC50 value as the ability of the extract to reduce 50% DPPH radical on the ethanol extract of cacao beans from spontaneous fermentation (F1) was 141.67 mg/L. The IC50 value of the fermented cacao bean extract with the addition of starter was obtained at F2 at 109.30 mg/L, F3 (97.51), and F4 is 88.15 mg/L.
Antimicrobial resistance in bacteria has become a leading global public health issue. Staphylococcus sp. has an efficient mechanism to deal with antimicrobial agents that make them hard to treat in hospital-acquired and community-acquired infections. This study was conducted due to limited data about multidrug resistance and extensively drug resistance in Staphylococcus sp. in Indonesia. This study was a descriptive retrospective study using a cross-sectional design to get the prevalence and antimicrobial susceptibility of S. haemolyticus, S. aureus, and S. epidermidis. The data was secondary data extracted from WHONET 2022 software. This study’s data were from bacteria from samples sent to UKK LMK FKUI, Jakarta from 2017 to 2021 for routine diagnostic. In this study, we found that the prevalence of methicillin-resistant S.aureus was 24,9%, methicillin-resistant S.epidermidis was 65,5%, and methicillin-resistant S.haemolyticus was 86,8%. The prevalence of MDR S.aureus is less than S.epidermidis and S.haemolyticus, respectively. MDR S.haemolyticus was consistently above 85% each year, while S.epidermidis was above 50% and S.aureus was below 50%. XDR Staphylococcus was only found in S.aureus and S.haemolyticus, i.e. three and seven XDR isolates of S.aureus and S.haemolyticus respectively during 2017-2021. Although we could not find any pan-resistant isolates from all samples, we found methicillin-resistant S.aureus and S.haemolyticus isolates that were also resistant to vancomycin and linezolid. S.haemolyticus dan S. epidermidis were an important coagulase-negative Staphylococcus species that can’t be neglected due to the high percentage of MDR and the discoveries of XDR in S.haemolyticus so that they have the potential to disseminate resistance plasmids to the more virulent bacteria. Therefore we need to control the use of antimicrobial agent to prevent this resistance.
Antimicrobial resistance is one of the top 10 global health threats. The hospital wastewater (HWW) potentially becomes the reservoir and dissemination of antibiotic resistance gene (ARG) and bacterial pathogens. In Indonesia, the protocol to monitor the ARGs form HWW has not been established. This study aimed to detect the presence and find the relative abundance of P. aeruginosa and aadA2 genes from Dr. RSUPN. Cipto Mangungkusumo (RSCM) inlet and outlet wastewater through qPCR assay. The primers used were supported by Resistomap. The study revealed that the qPCR assay was able to detect the Ct value of P. aeruginosa and aadA2. The aadA2 gene was found in all waste water samples, meanwhile P. aeruginosa was only found in some of inlet samples. aadA2 had the highest relative abundance and this gene’s mobility uses plasmids and integrons that potentially enhance the acquired antimicrobial resistance (AMR) mechanism. This study implicated that qPCR assay was capable to detect pathogenic bacteria and ARG, and ARG could be released to the environment even though the wastewater samples have been proceeded in wastewater treatment plants (WWTP). The qPCR assay can be used as the method to monitor the AMR status in a hospital and the spreading potency to the environment using the HWW.
Abstract. Chitinolytic bacteria can produce chitinase, reported as a biocontrol agent against plants. This research aims to see chitinolytic activity in inhibiting the growth of Rhizoctonia solani and Fusarium oxysporum. Anti fungal testing in dual culture test by growing each of the chitinolytic bacteria, Lysinibacillus fusiformis and Brevibacillus reuszeri, with the pathogenic fungi, F. oxysporum and R. solani, in Petri dishes containing Chitin Agar Media facing a distance of 3 cm. The results showed that chitinolytic bacterial isolates were capable inhibit the fungus by having the activity of each index inhibition of L. fusiformis isolates (30%), B. reuszeri (77%) against F. oxysporum, and R. solani fungi isolates (100%) for each chitinolytic bacterial isolate. Keywords : Anti fungal, Chitinolytic bacteria, Pathogenic fungi.
Background: Dengue fever is still a serious health problem in the world. DENV consists of 11 kb of single positive-stranded RNA encoding three structural proteins and seven non-structural proteins. PrM and E proteins are the main targets of the antibody response that rich of epitopes and able to induce protective immunity. There are four DENV serotypes that have similar antigenic structures in the amino acid sequence of protein E. In our previous study, we successfully constructed a recombinant tetravalent DNA vaccine candidate consisting pUMVC4a-based expression plasmid for prM-E protein of all DENV serotypes (pUMD1, pUMD2, pUMD3 and pUMD4). It has been proved that the vaccine candidate was able to induced anti-dengue IgG as well as neutralization antibody to all DENV serotypes. This study aims to determine IgG subclasses of immunized mice with recombinant tetravalent DNA vaccine candidates based on prM-E genes of all serotypes. Methods: Mice (Balb/c) were immunized with a dose of 100 μg 100 uL/mouse in triplicate, at three weeks interval. Blood was drawn two weeks post immunization as well as termination blood. IgG subclasses titre were measured using in-house indirect ELISA. Results: The titer of IgG2a subclass was the highest levels with optical density of 1.004±0.154 followed by IgG1,IgG2b, and IgG3 to DENV-2, respectively. Conclusion: The data demonstrate the humoral immune response IgG subclasses of this recombinant tetravalent DNA vaccine candidates based on prM-E genes of all serotypes, supporting further translational studies to advance the development of this candidate in response to DENV infection. Keywords: dengue vaccine, DNA vaccine, recombinant, IgG subclass, Tetravalent
Mycoprotein is food with high protein content, fiber, and low in cholesterol made from fungal mycelium. In this research, mycoprotein was produced by spp. isolated fr Rhizopus om tempeh with soymilk as growth media. This research aims to determine the best strain of spp. and op Rhizopus timum carbon to nitrogen ratio for mycoprotein production Two parameters were applied which w . ere inoculum selection and carbon to nitrogen ratio treatment in media. The best inoculum was selected from four strains of Rhizopus spp., ATH 1,ATH 24,ATH 40, and ATH 53. On the other hand, carbon to nitrogen ratio treatment used were as follows 20:1, 20:2, and 40:2. Mycelium dry weight and protein content were measured, as well as reduction sugar, dissolved protein and total volatile base nitrogen concentration in media. The best strain for producing biomass was ATH 24 with 0.6 g of 0 mycelium dry weight per 50 mL of media and the protein content was 0.236 g. The best carbon to nitrogen ratio treatment was 20:1 with 0.57 gram of mycelium dry weight per 50 mLof media and the protein content was 0.20 g. Thus, our data indicate that strain ATH 24 with 20:1 of carbon to nitrogen ratio in media were highly potential for producing mycoprotein. Keywords: fermentation, mycelium, mycoprotein, Rhizopus, soymilk
Tapai singkong is one of the popular fermented foods in Indonesia, which is processed from cassava tubers (Manihot utilissima ). The bacteria present during the fermentation process de Manihot utilissima determines the quality of Tapaisingkong. However, information about the bacteria of Tapai singkong is still limited. Therefore, this study aimed to analyze the bacterial community of based on culturing techniques Tapai singkong and based on metagenomic sequencing with Next-Generation Sequencing (NGS) techniques. Five types of samples were Tapai singkong obtained from producers in Jakarta, Bogor, Tangerang, Band Tapai singkong ung, and Kediri-Indonesia. The bacterial community in this study was studied in from Kediri Tapai singkong because the taste was most favored by the panelists based on the hedonic test. Based on the culture technique using De Man Rogosa and Sharp Agar media, the two most abundant bacterial isolates were found. Based on the 16S rRNA gene sequence, both isolates were the same lactic acid bacteria (LAB), namely Pediococcus acidilactici DSM 20284, with 99.6% similarity. Based on metagenomic sequencing, it was found that the bacteria in the consisted of Firmicutes Tapai singkong (82%), Bacteriodetes (10%), unidentified bacteria (5%), and Verrucomicrobia (1%). The genus of Firmicutes was dominated by the LAB group, namely Pediococcus (61.23%), Weissella (4.8%), Lactobacillus (3.9%), Sporolactobacillus (2.2%), and Staphyloccocus (2.1%). The results of this study showed that the LAB group was most abundant in Tapai singkong . Therefore, the role of each LAB needs to be studied further to determine its role in the quality of Tapai singkong.
Lipase has an important role in industry. The KC4J mutated isolates from oil palm waste had 100% similarity to Aspergillus fumigatus strain RA204, a fungus known produce lipase. The study aimed to increase lipase activity of KC4J mutant through media optimization using Response Surface Methodology (RSM) and partial characterization. The three variables of media composition (olive oil, soy flour, and pH were optimized using Central Composite Design (CCD). The lipase characterization measured the influence of pH, temperature and metal ions. The pH tested on range 6 to 12, while the temperature variation tested on 30 to 70 °C. The metal ions tested were Mg2+, Ca2+, Zn2+, Mn2+, Fe2+ and K+ with concentrations of 1 mM and 10 mM. The production medium containing 1.25% of olive oil, 3.5% of soy flour and 7.5 pH resulting 11.25 U/mL of Lipase activity, which was higher than the previous media composition (10.00 U/mL). The results of CCD and quadratic analysis showed that the source of carbon, nitrogen and pH had an effect on lipase activity which showed R2 0.93. The optimum lipase activity produced at pH 6 and on 60 °C, and the lipase stable at pH 6-8 and on 30-70 °C. All metal ions tested were able to increase lipase activity with Ca 2+ ion gave the highest result. Keywords: Lipase, KC4J mutant, Central Composite Design, Oil Palm Waste.
Streptomyces genera plays important role in lignocellulose degradation. Many research founds Streptomyces has cellulolytic and ligninolytic enzymes that sufficient to degrade lignocellulosic materials. However, minimum lignocellulosic material condition that can efficiently degraded by Streptomyces sp. has not been fully understood. In this research, three pretreament conditions (physical, alkaline-hydrotermal, and hydrogen-peroxide chemical treatments) of sugarcane bagasse used as lignocellulosic material, to further degraded by Streptomyces sp. S2. Lignocellulose component measurement conclude that raw (physical treated only) bagasse wasn’t efficiently degraded by Streptomyces sp S2. Hydrogen-peroxide was effective on reducing both syringil and guaiacyl lignin, meanwhile alkaline-hydrotermal pretreatment was very effective on reducing syringil lignin. This study suggest that hydrogen-peroxide pretreatment can be used in many type of lignocellulosic material, which can be further degraded by Streptomyces sp. S2. Alkaline-hydrotermal preteatment on the other hand is best suited to degrade lignocellulosic material that have high percentage of syringil lignin.
Ginger is a rhizomatous perennial herb that grows abundantly in tropical areas. It has been used around the world as a spice, flavoring agent, and ingredient in traditional medicine. Ginger essential oils (GEOs) are derivatives of ginger that can be found in various products used in daily life, such as food, pharmaceutical, and cosmetics. The present study analyzed the chemical compositions, antioxidant, and antibacterial activities of three commercially available GEOs. The compositions of GEOs were identified using the gas chromatography method. The antioxidant activity was evaluated using 2,2-diphenyl-1-picryl-hydrazyl (DPPH) and 2,2’-azinobis- (3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) assay methods. The antibacterial activity was determined using a disc diffusion assay based on the diameter of the inhibition zone (DIZ). The main compounds identified from the samples were zingiberene, α-curcumene, β-sesquiphellandrene, camphene, α-farnesene, β-bisabolene, α-pinene, and 3-carene. The IC50 values were found to be 5.3023 and 1.4504 mg/mL for GEO1; 0.9249 and 0.5276 mg/mL for GEO2; and 10.4463 and 3.3535 mg/mL for GEO3 when evaluated using DPPH and ABTS assay methods, respectively. All samples showed antibacterial activity against Staphylococcus aureus ATCC 13420 and Bacillus subtilis (collection of Indonesian Institute of Sciences), while only GEO2 and 3 displayed inhibitory effect against Escherichia coli ATCC 9637.
Manganese superoxide dismutase (MnSOD) from bacteria shares high amino acid sequence homology and nearly identical structure. Despite of that, their characteristics are diverse, which likely due to their bacterial origin and adaptation to the environment. Most importantly, their structural similarity extends to eukaryotic MnSOD, i.e. human. Therefore, structural study of bacterial MnSOD is relevant to its human SOD and henceforth for its use in human as a therapeutic agent or a cosmetic ingredient. Further, eukaryotic MnSOD occurs as a tetramer while almost all of the prokaryotic are dimeric. In this review, relationship between the amino acid sequences and structures of MnSOD as well as their origin and evolution is discussed. The structures of FeSOD and cambialistic SOD, which are MnSOD closest homologs, are visited as the comparison. This study provides an insight to potential safe application of bacterial MnSOD, including necessary modifications to obtain desired characteristics for applications in human.
Salmonella typhimurium bacteria could cause gastroenteritis and its growth could be controlled by the active compounds from natural products, which is Portulaca oleracea L. herb. Portulaca oleracea contained tannin, saponins and flavonoids compounds which had different characteristics towards temperature extraction. This study aims to determine the antibacterial activity of ethanol extract of the Portulaca oleracea herb from various extraction methods against S. typhimurium bacteria. Extraction of Portulaca oleracea herb was carried out with four variations methods which were the cold method (maceration and percolation) and heat method (soxhlet and refluxs) using 96% ethanol solvent. The four types of extract were tested for their antibacterial activity by disk diffusion at concentrations of 30%, 35%, 40%, 45% and 50% (b/v). The positive control was chloramphenicol 30 µg/disk, while the negative control was DMSO solvent. The results of antibacterial activity test in the form of zone of inhibition were statistically analyzed by Two Way Anova. The results showed that the ethanol extract of the Portulaca oleracea herb from various extraction methods had antibacterial activity against S. typhimurium. There was a significantly difference in the antibacterial activity of ethanol extract of the Portulaca oleracea herb obtained from the reflux method with other methods (maceration, percolation and soxhlet) against S. typhimurium. Keywords: ethanol extract of Portulaca oleracea L. herb, antibacterial, Salmonella typhimurium, various extraction methods
The aim of this research was to obtain and determine the identity of Hg-resistant bacteria in soil contaminated with gold processing waste and test its ability to reduce mercury contamination. Soil samples as a source of Hg resistant bacterial isolates were obtained from the gold processing location in Ilangata Village, Anggrek District, North Gorontalo Regency. The research was conducted at the Microbiology Laboratory, Department of Biology, Faculty of Mathematics and Natural Sciences. Mercury analysis was carried out at the Laboratory of Fisheries Product Quality Development and Testing (LPPMHP), Gorontalo Province, and bacterial identification was carried out at the Hasanuddin University Medical Research Center Research Unit. The parameters observed were the types of Hg resistant bacteria and the ability of the bacteria to reduce mercury contamination. The results showed that there were four bacterial isolates on the soil contaminated with 4.5 ppm mercury, which were named ILb01, ILB02, ILb03, and ILb04. Molecular identification showed that ILb01 was closely related to Stenotrophomonas sp. SB67 and ILb02 close to Enterobacter cloacae strain CM 1, these strains were not resistant to mercury contamination; while ILb03 which is similar to strain BS0591 and ILb04 which is similar to Bacterium Bacillus albus strain SQ30 16S could be resistant and was able to reduce mercury contamination by 99% at 10 ppm levels. Key words: heavy metals, Hg-resistant bacteria, molecular identification, mercury
In the antibiotic era, Tuberculosis (TB) drugs resistance especially Rifampicin (RIF) is highly reported around the world. Resistance of RIF is caused by the mutation of genes that associated with RIF receptor. The aims of this study are detecting the Single Nucleotide Polymorphism of Rifampicin resistant genes using Whole Genome Sequencing (WGS) and analysing the profile of protein changing caused by SNP. Twenty Mycobacterium tuberculosis culture samples were passed on WGS procedure and 19 samples were adequate to further bioinformatics analysis. Single Nucleotide Polymorphisms Analysis was done using TBprofiler. Based on TBProfiler, seventeen samples were resistant to rifampicin. The mutations that cause the resistance are S450L, D435Y, H445Y, 430P, Q432K. Other Single Nucleotide Polymorphisms H835R, V534M and R224C were also found. The H835R mutants are present together with the S450L, V534M with S450L mutants, and R224C with Q432K mutants. Native protein for RNA Polymerase Subunit β used was the result of separation from the crystal structure of Mycobacterium tuberculosis H37Rv RNA polymerase (PDB: 5UHB). Binding affinity RIF to RNA Polymerase Subunit β calculated using AutoDock vina. Construction of mutant 3D structures using FoldX5. From the analysis, it was found that seventeen samples were resistant to rifampicin and two samples did not contain SNP which could cause resistance to rifampicin.
Dermatophytosis cases have increased significantly in various countries, more than 20-25% of the population were infected by superficial fungal infections. Malabero urban village is a coastal area in Bengkulu city with high temperature and humidity. The daily activities of residents in the area are mostly fishermen, so they are in a watery or wet environment for approximately 12 hours every day. So, this research aimed to identify dermatophyte fungi that caused tinea pedis and tinea unguium from these coastal communities. The diagnosis of tinea was based on symptoms and physical examination of the lesion area. Furthermore, specimen collection was carried out by scraping the lesion area, then examined with 20% KOH and fungal culture on sabouraud dextrose agar media. Subjects included 79 people who had symptoms of tinea pedis and 33 people with symptoms of tinea unguium. The results of dermatophyte fungi identification were obtained, Trichophyton mentagrophytes, Trichophyton rubrum, Trichophyton tonsurans, and Aspergillus sp. Our conclusion is the most identified dermatophyte species is Trichophyton mentagrophytes. Keywords: Coastal, dermatophyte, tinea pedis, tinea unguium
Ramie fiber is a potential raw material to substitute imported raw materials such as cotton. Due to its higher hemicellulose content, ramie fiber required hydrolysis in a process called degumming. Enzymatic degumming is environmentally friendly compared to traditional process which using chemicals. Alkalithermophilic xylanase have high ability in hemicellulose hydrolysis. The production of xylanase was conducted by submerged fermentation of Bacillus halodurans CM1 in 20L bioreactor using Mamo and corncob medium with optimum conditions at 50°C, pH 9, 150 RPM and 1 vvm. The optimum specific activity of xylanase measured by Bailey method at 70°C and pH 9 is 475.41 U/mg. Xylanase was stable at 50°C, pH 9 and relatively stable to K+, Na2+, Co2+ and Ca2+ metal ions and Triton-X, Saba dan Tween-80 surfactants. Degumming process was carried out by immersing ramie fibers in formulated degumming solution with vlot 1:20 at 50°C, 150 RPM and 180 minutes. The enzymatic degumming process may substitute or reduce the use of chemicals due to its significant effect on ramie fiber quality. Enzymatic and chemical degumming process reduce the weight of Ramie Fiber to 7.23 %, and 7.72 %, slightly higher than enzymatic degumming 7.15%. Enzymatic degumming maintains tensile strength at 27.51 %. Whiteness index enhanced to 2.99% enzymatically and 3.49% chemically. Keywords: Bacillus halodurans CM1, enzymatic degumming, ramie fiber, textile industry, thermoalkaliphilic xylanase
Background: Dengue Hemorrhagic Fever (DHF) is an infectious disease caused by the dengue virus (DENV) which spread widely in tropical and subtropical regions of the world. DENV is a single-positive strand RNA virus with a genome size of ± 11kb which encodes three structural proteins, seven non-structural proteins, and two untranslated regions (UTR). The non-structural protein-1 (NS1) of DENV is known to have important role in dengue pathogenesis also promising to be developed as dengue vaccine. Lately, novel vaccine approach by DNA immunization have given new perspective for a safe, stable, and immunogenic vaccine platform. Previously, we have successfully construct DNA vaccine encoding NS1 protein of DENV2 (pUNS1) which express recombinant NS1 protein in-vitro. Thus, in this current study the ability of pUNS1 to induce humoral immune response will be further analyzed by in mice immunization. Methods: Sixteen BALB/c mice aged of 4 weeks were immunized 3 times with 100 µg of pUNS1 or pUMVC4a on 2 week time interval. Blood sampling was carried out just before immunization and termination was done 2 week after last immunization. Titer from individual mice sera against DENV-2 were measure with in-house ELISA. Results: IgG against NS1 protein of DENV2 titer from mice group immunized with recombinant pUNS1 shown high ELISA absorbancies, 5 times higher than pUMVC4a group. This result suggest the ability of pUNS1 to induce humoral immune response against NS1 DENV-2 in-vivo. Conclusion: Recombinant pUNS1 can induce humoral immune response in mice.
