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All Journal HAYATI Journal of Biosciences VALENSI JSMARTech : Journal of Smart Bioprospecting and Technology Jurnal Bioteknologi & Biosains Indonesia (JBBI)
Azzahra, Maharanianska
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Agriculture, Biological Sciences & Forestry Biochemistry, Genetics & Molecular Biology Chemistry Earth & Planetary Sciences Environmental Science Health Professions Materials Science & Nanotechnology Medicine & Pharmacology Neuroscience

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Development of Bacillus subtilis Detection Method Targeting Genes codY, narH, and ureC Using Polymerase Chain Reaction Nurjayadi, Muktiningsih; Berkahingrum, Ayu; Putri, Adinda Myra Amalia; Rahmawati, Atikah Nur; Anggraeni, Rosita Gio; Fahriza, Tiara; Declan, Jefferson Lynford; Putri, Gladys Indira; Krisdawati, Ismaya; Juliansyah, Dandy Akbar; Azzahra, Maharanianska; Maulana, Irvan; Kartika, Irma Ratna; Kurniadewi, Fera; Sukmawati, Dalia; Rahayu, Sri; Saamia, Vira; Wiranatha, I Made; Abomoelak, Bassam; El Enshasy, Hesham
JSMARTech: Journal of Smart Bioprospecting and Technology Vol. 5 No. 1 (2024): JSMARTech Volume 5, No. 1, 2024
Publisher : JSMARTech

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.21776/ub.jsmartech.2024.005.01.26

Abstract

Bacillus subtilis, a causative agent of foodborne illness in bread, cakes, cereals, and cheese leading to symptoms like diarrhea and nausea, was investigated for its role in nosocomial infections, including endocarditis, bacteremia, septicemia, and meningitis. The research focused on three genes: codY, narH, and ureC, which respectively contribute to bacterial survival through biofilm and spore formation, acid resistance, and adaptation to anaerobic conditions. The objective of this study was to evaluate the codY, narH, and ureC genes using the Gradient PCR method for Bacillus subtilis detection in bread and cheese through Real-Time PCR. During the research, bacterial growth was observed with an OD600 of 1.438 in Tryptic Soy Broth (TSB), and colonies of medium size, smooth, cream-coloured, and round shape were successfully isolated on Tryptic Soy Agar (TSA). The DNA template used had a 71.5 ng/µL concentration with an A260/280 ratio of approximately 1.830. The annealing temperature for Gradient PCR used was 53-62°C. The primers successfully amplified codY (175 bp), narH (222 bp), and ureC (153 bp) gene amplicons. The optimal annealing temperature for the primers used was 60°C, as indicated by the presence of a single bright band in electrophoresis. Using these three different genes, testing can also be conducted with the Multiplex PCR method. The next step involves developing a detection kit using optimized primers and annealing temperatures for the identification of Bacillus subtilis in bread and cheese samples using Real-Time PCR.
Real-time PCR-based Detection of Foodborne Pathogen Cronobacter sakazakii DNA in Infant Formula Milk with Specific Targeting on the hfq Gene Nurjayadi, Muktiningsih; Juliansyah, Dandy Akbar; Declan, Jefferson Lynford; Putri, Gladys Indira; Krisdawati, Ismaya; Rahmawati, Atikah Nur; Azzahra, Maharanianska; Maulana, Irvan; Putri, Gusti Angieta; Kurniadewi, Fera; Kartika, Irma Ratna; Saamia, Vira; Wiranatha, I Made; Abomoelak, Bassam; El-Enshasy, Hesham Ali
HAYATI Journal of Biosciences Vol. 32 No. 6 (2025): November 2025
Publisher : Bogor Agricultural University, Indonesia

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.4308/hjb.32.6.1597-1607

Abstract

Cronobacter sakazakii has been linked to cause meningitis, necrotizing enterocolitis, and sepsis in infants and newborns, with case fatality rates ranging from 40 to 80%. The most common source of infection has been identified as Cronobacter sakazakii-contaminated infant formula. With a relatively specific target hfq gene, this study aims to develop a real-time PCR method to identify Cronobacter sakazakii in infant formula milk. Real-time PCR is used as a detection method because rt- PCR has higher specificity and sensitivity compared to conventional PCR methods. The real-time PCR method also has a higher level of effectiveness and time efficiency compared to conventional PCR. Cronobacter sakazakii ATCC 29544 genomic DNA was isolated and used in a real-time PCR assay. Cronobacter sakazakii DNA was amplified using a primer targeting the hfq gene, yielding a 145 bp amplicon. The results of the real-time PCR test showed that Cronobacter sakazakii DNA with a concentration of 53 ng/µL could be amplified by the primer pairs of hfq gene with Ct values of 11 respectively then had Tm values of 81.7°C±0.5. The specificity test showed that the hfq primer pairs could differentiate between the target and some non-target bacteria. The sensitivity test showed the ability of the primer to detect the smallest concentration of 3.392 pg/µL with a Ct of 26.16. Based on the results obtained, it can be concluded that the hfq primer has the potential to be used as a fast detection method for Cronobacter sakazakii bacteria in infant formula using real-time PCR.
Efficacy Test of Prototype Kit for Detection Bacillus cereus and Listeria monocytogenes in Processed Meat using Real-time PCR Method Nurjayadi, Muktiningsih; Fahriza, Tiara; Putri, Adinda Myra Amalia; Rahmawati, Atikah Nur; Berkahingrum, Ayu; Anggraeni, Rosita Gio; Putri, Gladys Indira; Declan, Jefferson Lynford; Akbar, Dandy; Maulana, Irvan; Azzahra, Maharanianska; Shangkara, Muhammad Arkent; Kartika, Irma Ratna; Kurniadewi, Fera; Sukmawati, Dalia; Rahayu, Sri; Saamia, Vira; Wiranatha, I Made; Abumoelak, Bassam; El Enshasy, Hesham
Jurnal Kimia Valensi Vol. 11 No. 1 (2025): Jurnal Kimia VALENSI
Publisher : Department of Chemistry, Faculty of Science and Technology Syarif Hidayatullah Jakarta State Islamic University

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.15408/jkv.v11i1.44280

Abstract

According to the World Health Organization (WHO), harmful agents such as Bacillus cereus and Listeria monocytogenes are responsible for 600 million cases of disease and 420,000 deaths annually. This research aims to test the effectiveness of the real-time PCR method for developing a prototype kit to detect pathogenic bacteria in processed meat. As a comparison, the and conventional PCR methods were used to obtain the accuracy, specificity, sensitivity, and effectiveness of the real-time PCR method. All the samples were cultured in solid media agar, performed amplification using specific primers cyt-K 2 and hly using PCR and real-time PCR. Meatballs, nuggets, and sausages, five samples each, were found to be positive positively contaminated with all targeted bacteria. However, they did not provide specific results using solid media culture and the PCR method. In addition, the real-time PCR method using prototype kit formulas accomplished that all contaminated samples had a Ct value smaller than the negative control, NTC (No Template Control), and had a similar melting curve to the positive control. This establishes that the real-time PCR method clarifies that all samples were contaminated with target bacteria. A formula was developed in the prototype kit with real-time PCR methods that can be used and applied on a commercial scale efficiently and precisely.
DETERMINATION OF OPTIMAL ANNEALING TEMPERATURE Vibrio alginolyticus PRIMERS USING POLYMERASE CHAIN REACTION METHOD Putri, Gladys Indira; Nurjayadi, Muktiningsih; Declan, Jefferson Lynford; Krisdawati, Ismaya; Juliansyah, Dandy Akbar; Fahriza, Tiara; Azzahra, Maharanianska; Maulana, Irvan; Kartika, Irma Ratna; Kurniadewi, Fera; Sukmawati, Dalia; Saamia4, Vira; Saputro, Dwi Anna Oktaviani; Wiranatha, I Made; Abomoelak, Bassam; El-Enshasy, Hesham Ali
Jurnal Bioteknologi & Biosains Indonesia (JBBI) Vol. 10 No. 2 (2023)
Publisher : BRIN - Badan Riset dan Inovasi Nasional

Show Abstract | Download Original | Original Source | Check in Google Scholar | DOI: 10.55981/jbbi.2023.2976

Abstract

Food poisoning is a global issue of grave concern. If food is not properly cooked, it can be a medium for the spread of pathogenic bacteria. Vibrio alginolyticus is one of the pathogenic bacteria that can cause food poisoning. real-time Polymerase Chain Reaction (rt-PCR) can detect pathogenic bacteria in food, so it is necessary to determine the optimal annealing temperature. This research aims to obtain the optimal annealing temperature of the Va_Chr1_FR primer using Gradient PCR. The DNA concentration used was 174.5 with an A260/A280 purity of 1.94. The temperature range tested, 53°C-62°C, corresponds to the melting temperature of the Va_Chr1_FR primers. The primers designed were F5'-TTCTTCTGTTGTAGGTTCCG-F3' and R5'-CCAGCCCTCACATCTAATAC-R3'. Based on these results, a temperature of 60°C is deemed as the most optimal annealing temperature because it produces one of the brightest bands on electrophoresis with an amplicon length of 146 bp. The findings of this study will be beneficial to the development of Va_Chr1_FR Vibrio alginolyticus primers testing on food samples using the real-time PCR method. 
Co-Authors Abomoelak, Bassam Abumoelak, Bassam Akbar, Dandy Anggraeni, Rosita Gio Berkahingrum, Ayu Dalia Sukmawati El Enshasy, Hesham El-Enshasy, Hesham Ali Fahriza, Tiara Jefferson Lynford Declan Juliansyah, Dandy Akbar Kartika, Irma Ratna Krisdawati, Ismaya Kurniadewi, Fera Maulana, Irvan Muktiningsih Nurjayadi Putri, Adinda Myra Amalia Putri, Gladys Indira Putri, Gusti Angieta Rahmawati, Atikah Nur Saamia, Vira Saamia4, Vira Saputro, Dwi Anna Oktaviani Shangkara, Muhammad Arkent SRI RAHAYU Wiranatha, I Made