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"Penelitian ini mengembangkan metode deteksi spesies babi (Sus scrofa) pada sampel daging campuran menggunakan automasi ekstraksi DNA magLEAD gC. DNA dianalisis menggunakan PCR dan TaqMan probe RT-PCR dengan primer spesifik untuk gen Cytochrome c oxidase I (COI), Cytochrome b (Cytb), dan NADH5 dehydrogenase 5 (ND5). Hasil menunjukkan bahwa ekstraksi DNA otomatis menghasilkan konsentrasi DNA 129,4–388,5 ng/μL pada daging mentah dan 66,4–89,5 ng/μL pada bakso dengan rasio kemurnian A260/A280 dan 260/A230 > 1,8. Primer COI, Cytb dan ND5 dapat mendeteksi DNA babi. PCR dan RT-PCR in vitro menunjukkan ketiga primer hanya mendeteksi DNA babi. Efisiensi amplifikasi RT-PCR primer COI, Cytb, dan ND5 adalah 144,14% (R2=0,982), 88,05% (R2=0,998), dan 81,25% (R2=0,997) dengan batas deteksi 0,0001 ng/μL, 0,001 ng/μL, dan 0,001 ng/μL. Primer/probe Cytb dan ND5 mendeteksi bakso dengan campuran daging babi hingga 0,1% (w/w).

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This study developed a method to detect pig species (Sus scrofa) in mixed meat samples using automated DNA extraction with the magLEAD gC. DNA was analyzed using PCR and TaqMan probe RT-PCR with specific primers for the genes Cytochrome c oxidase I (COI), Cytochrome b (Cytb), and NADH5 dehydrogenase 5 (ND5). Results showed that automated DNA extraction produced DNA concentrations of 129.4–388.5 ng/μL in raw meat and 66.4–89.5 ng/μL in processed meatballs with purity ratios A260/A280 dan 260/A230 > 1.8. The COI, Cytb and ND5 primers could be used to detect pig DNA. In vitro PCR and RT-PCR showed that all three primers only detected pig DNA. The RT-PCR amplification efficiency for COI, Cytb, and ND5 primers were 144,14% (R2=0,982), 88,05% (R2=0,998), dan 81,25% (R2=0,997) with detection limits of 0.0001 ng/μL, 0.001 ng/μL, and 0.001 ng/μL. The Cytb and ND5 primers/probes detected meatballs with pig meat content as low as 0.1% (w/w)."

"Latar Belakang. Penyakit COVID-19 yang disebabkan oleh SARS-CoV-2 dengan cepat menyebar dan menjadi Pandemi serta menimbukan kerugian yang sangat besar pada masyarakat di seluruh dunia. Deteksi virus yang cepat dan akurat memegang peranan penting untuk mengendalikan penyebaran di masyarakat dan membantu pasien untuk menghindari perkembangan penyakit lebih lanjut. Saat ini real-time Reverse Transcriptase Polymerase Chain Reaction (real-time RT-PCR) merupakan reference standard diagnostic test dalam mendeteksi SARS-CoV-2 di seluruh dunia. Real-time Reverse Transcriptase Loop Mediated Isothermal Amplification (RT-LAMP) merupakan metode amplifikasi asam nukleat isotermal yang memiliki sensitivitas dan spesifisitas tinggi dan waktu pengerjaan yang jauh lebih cepat dibandingkan real-time RT-PCR. Tujuan. Penelitian bertujuan untukiDetectTM SARS-CoV-2 Detection KitSARS-CoV-2.Metode. Penelitian ini merupakan uji kesesuaian dengan studi potong lintang dan menggunakan metode pengumpulan sampel secara consecutive sampling. Subjek penelitian yaitu spesimen swab nasofaring dan orofaring dalam VTM (N=80) yang dianalisis di Laboratorium Mikrobiologi Klinik Fakultas Kedokteran Universitas Indonesia. iDetectTM SARS-CoV-2 Detection Kit menggunakan uji kesesuaian Kappa aplikasi SPSS versi 25.Hasil. Dari 72 sampel valid yang diperiksa dengan real-time RT-LAMP iDetectTM SARS- CoV-2 Detection Kit dan real-time RT-PCR, 24 sampel terdeteksi positif oleh real-time RT-PCR dan hanya tiga sampel yang terdeteksi positif oleh real-time RT-LAMP. Tiga sampel yang terdeteksi positif oleh real-time RT-LAMP termasuk ke dalam sampel - sampel yang terdeteksi positif oleh real-time RT-PCR. Secara statistik, uji reliabilitas / uji kesesuaian dari penelitian kedua alat diagnostik ini menunjukkan nilai Kappa yang sangat rendah, yaitu 0,16. Uji kesesuaian Kappa kedua alat ini menunjukkan bahwa hasil pemeriksaan alat real-time RT-LAMP iDetectTM SARS-CoV-2 Detection Kit tidak sesuai dengan alat real-time RT-PCR dalam mendeteksi SARS-CoV-2. Kesimpulan. Real-time RT-LAMP iDetectTM SARS-CoV-2 Detection Kit tidak sesuai dengan alat real-time RT-PCR dan tidak dapat digunakan sebagai alat diagnostik dalam mendeteksi SARS-CoV-2.

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Introduction. COVID-19 caused by SARS-CoV-2 quickly spread and became Global Pandemic and caused enormous losses to people around the world. Rapid and accurate virus detection plays an important role in controlling spread in the community and helping patients to avoid further disease progression. Currently, real-time Reverse Transcriptase Polymerase Chain Reaction (real-time RT-PCR) is determined as the reference standard diagnostic test for detecting SARS-CoV-2 worldwide. Real-time Reverse Transcriptase Loop Mediated Isothermal Amplification (RT-LAMP) is an isothermal nucleic acid amplification method that has high sensitivity and specificity and provide faster result than real-time RT-PCR. Aim. The research aims to compare real-time RT-LAMP iDetectTM SARS-CoV-2 Detection Kit and real-time RT-PCR in detecting SARS-CoV-2. Method. This research is a comparison test with a cross-sectional study and uses a consecutive sampling method to collect samples. The research subjects were nasopharyngeal and oropharynx swab specimens in VTM (N=80) which were analyzed at the Clinical Microbiology Laboratory, Faculty of Medicine, Universitas Indonesia. The data obtained from the real-time RT-LAMP iDetectTM SARS-CoV-2 Detection Kit and real-time RT-PCR test results were analyzed using Kappa test SPSS version 25.

Results. Of the 72 valid samples examined by the real-time RT-LAMP iDetectTM SARS- CoV-2 Detection Kit and real-time RT-PCR, 24 samples were detected positive by real- time RT-PCR and only three samples were detected positive by real-time RT-LAMP. Three samples that were detected positive by the real-time RT-LAMP were included in the samples that were detected positive by the real-time RT-PCR. Statistically, the comparison test of the research of these two diagnostic tools showed a very low Kappa value, which was 0.16. The Kappa suitability test of these two tools showed that the real- time RT-LAMP iDetectTM SARS-CoV-2 Detection Kit were not compatible with the real- time RT-PCR in detecting SARS-CoV-2. Summary. Real-time RT-LAMP iDetectTM SARS-CoV-2 Detection Kit is not compatible with real-time RT-PCR and cannot be used as a diagnostic tool in detecting SARS-CoV-2."

"ABSTRAKBenih bawal bintang memperlihatkan perubahan tingkah laku seperti kehilangan kemampuan berenang dan berkumpul di dasar kolam, diduga terinfeksi RSIVD. Real time PCR dengan SBYR green telah diaplikasikan secara luas untuk diagnosis penyakit. Kesederhanaan, sensitifitas, rentang deteksi yang dinamis, reproduktifitas, dan jaminan skrining dengan kecepatan waktu yang tinggi membuat real time PCR sesuai untuk mendeteksi virus (Niesters, 2002). Oleh karena itu dilakukan aplikasi metode real time PCR dengan SYBR green untuk mendeteksi RSIV pada benih bawal bintang. Penelitian ini menggunakan primer 1F dan 1R untuk skrining Megalocityvirus, grunt fin cell line untuk kultur RSIV, pengklonaan menggunakan vektor pGEM-T easy dan primer MCPspecR697-F4 dan MCP-specR888-R6 untuk deteksi RSIV dengan metode real time PCR menggunakan SYBR green. Karakterisasi dari CPE menunjukkan sel GF menjadi berbentuk bundar dan sel-sel GF terlihat berpendar pada inokulasi RSIV pada hari ke lima sampai ke tujuh. Kurva standar menghasilkan R2 0,99999, slope -2,41675 dan y-intercept 38,68938. Limit deteksi 10 salinan DNA. Spesimen klinis menunjukkan hasil positif pada jaringan hati, limpa dan ginjal. Jumlah salinan DNA paling banyak dari ekstraksi limpa yaitu: 6054 dan 4182 salinan DNA sedangkan pada organ ginjal sebanyak 72 dan 101 salinan DNA dan hati 1 dan 2 salinan DNA. Metode real time PCR menggunakan SYBR green berhasil diaplikasikan untuk mendeteksi RSIV pada benih ikan bawal bintang.

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ABSTRACTSnubnose pompano juvenile showed behavioral changes such as losed the ability to swim and congregated at the bottom of the pool, suspected of being infected RSIVD. Real time PCR with green SBYR has been widely applied to the diagnosis of the disease. Simplicity, sensitivity, dynamic range of detection, reproducibility, and the assurance screening with a high speed makes real time PCR according to detect viruses (Niesters, 2002). Therefore, it is done in real time application method with SYBR green PCR to detect RSIV on Snubnose pompano juvenile. This study using the primers 1F and 1R for screening Megalocityvirus, grunt fin cell line for RSIV culture, cloning using pGEM-T easy vector and primer MCPspecR697-F4 and MCP-specR888-R6 for RSIV detection by real-time PCR method using SYBR green.GF cells infected by RSIV showed round and flourescence as a result of CPE at day 5 to 7. Subnose pompano juvenile positive infected by RSIV at 191 bp. Standard curve equation R2: 0.99999, slope: -2.41675 and y-intercept: 38.68938. qPCR using primers MCP-specR674-F4 and MCP-specR888 R6 primer assay showed detection limit of 10 copies of the. Liver, spleen and kidneys of Subnose Dart juvenile were infected by RSIV, positively. The highest of copy number of DNA were shown in the spleen (6054 and 4182 copies DNA, respectively), while in kidney were 101 and 72 copies DNA respectively. The lowest copy number DNA were shown in the liver (1 to 2 copies DNA, respectively). SYBR green quantitative PCR method can be applied to detect RSIV on Subnose pompano juvenile."

"Deteksi Infeksi Submikroskopis Necator americanus, Ancylostoma duodenale, dan Ascaris lumbricoides dari Sampel Feses di Nangapanda, Ende, Menggunakan Real-Time Polymerase Chain Reaction. Infeksi dari Soil-Transmitted Helminthes (STH) (N. americanus, A. duodenale (Hookworm), dan A. lumbricoides) dapat menyebabkan anemia, kekurangan zat besi, bahkan malnutrisi. Pemeriksaan infeksi STH dapat dilakukan menggunakan mikroskop, tetapi metode tersebut masih kurang sensitif. Penelitian bertujuan mendeteksi dan mengetahui persentase infeksi submikroskopis STH dari sampel feses anak (usia 5-18 tahun) di Nangapanda, Ende menggunakan metode real-time polymerase chain reaction (PCR). Sampel feses dikoleksi sebanyak dua kali, yaitu sebelum dan sesudah pemberian albendazole 400 mg. Total sampel yang diperoleh adalah 242 tetapi hanya 45 sampel yang negatif secara mikroskopis yang diuji dengan real-time PCR. DNA sampel diisolasi dan diamplifikasi menggunakan primer dari daerah internal transcribed spacer (ITS-1 dan ITS-2) rDNA. Deteksi dengan real-time PCR menghasilkan kurva amplifikasi pada fluorophore VIC, FAM, dan Texas Red. Sebanyak tiga sampel (6,7%) pada pre treatment termasuk low load of DNA (N. americanus and A. lumbricoides) (Ct > 35), empat sampel (9,1%) termasuk low load of DNA untuk N. americanus saja (Ct > 35), dan lima sampel (11,4%) termasuk moderate load of DNA untuk A. lumbricoides saja (30 < Ct < 35) pada post treatment. Hasil penelitian menunjukkan bahwa real-time PCR dapat mendeteksi infeksi submikroskopis dari Hookworm dan A. lumbricoides.

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Soil-transmitted helminth (STH) infections (Necator americanus (hookworm), Ancylostoma duodenale (hookworm), and Ascaris lumbricoides) can lead to anemia, malnutrition, and iron deficiency. Traditionally, STH infections have been diagnosed using microscopy to detect eggs in human fecal samples. However, there are several limitations of this method. The aim of this research was to detect the percentage of submicroscopic STH infections from human fecal samples (children, 5?18 years old) in Nangapanda, Ende, using the real-time polymerase chain reaction (PCR) method. The fecal samples were collected in two time periods, which were before and after treatment, using 400 mg of Albendazole. There were 242 samples in total, but only 45 negative samples from microscopic detection were tested with real-time PCR. The DNA samples were isolated and amplified wih primers of internal transcribed spacer (ITS-1 and ITS-2) region of rDNA. The detection of samples with real-time PCR generated an amplification curve in VIC, FAM, and Texas Red fluorophore. Three samples (6.7%) in pre-treatment were low load of DNA (N. americanus and A. lumbricoides) (Ct > 35). Four samples (9.1%) were low load of DNA (N. americanus) (Ct > 35) in post-treatment. Five samples (11.4%) were moderate load of DNA (A. lumbricoides) (30 < Ct < 35) in post-treatment. real-time PCR could detect submicroscopic infections from specific species of hookworm and A. lumbricoides."

"Deteksi Infeksi Submikroskopis Necator americanus, Ancylostoma duodenale, dan Ascaris lumbricoides dari Sampel Feses di Nangapanda, Ende, Menggunakan Real-Time Polymerase Chain Reaction. Infeksi dari Soil-Transmitted Helminthes (STH) (N. americanus, A. duodenale (Hookworm), dan A. lumbricoides) dapat menyebabkan anemia, kekurangan zat besi, bahkan malnutrisi. Pemeriksaan infeksi STH dapat dilakukan menggunakan mikroskop, tetapi metode tersebut masih kurang sensitif. Penelitian bertujuan mendeteksi dan mengetahui persentase infeksi submikroskopis STH dari sampel feses anak (usia 5-18 tahun) di Nangapanda, Ende menggunakan metode real-time polymerase chain reaction (PCR). Sampel feses dikoleksi sebanyak dua kali, yaitu sebelum dan sesudah pemberian albendazole 400 mg. Total sampel yang diperoleh adalah 242 tetapi hanya 45 sampel yang negatif secara mikroskopis yang diuji dengan real-time PCR. DNA sampel diisolasi dan diamplifikasi menggunakan primer dari daerah internal transcribed spacer (ITS-1 dan ITS-2) rDNA. Deteksi dengan real-time PCR menghasilkan kurva amplifikasi pada fluorophore VIC, FAM, dan Texas Red. Sebanyak tiga sampel (6,7%) pada pre treatment termasuk low load of DNA (N. americanus and A. lumbricoides) (Ct > 35), empat sampel (9,1%) termasuk low load of DNA untuk N. americanus saja (Ct > 35), dan lima sampel (11,4%) termasuk moderate load of DNA untuk A. lumbricoides saja (30 < Ct < 35) pada post treatment. Hasil penelitian menunjukkan bahwa real-time PCR dapat mendeteksi infeksi submikroskopis dari Hookworm dan A. lumbricoides.

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Soil-transmitted helminth (STH) infections (Necator americanus (hookworm), Ancylostoma duodenale (hookworm), and Ascaris lumbricoides) can lead to anemia, malnutrition, and iron deficiency. Traditionally, STH infections have been diagnosed using microscopy to detect eggs in human fecal samples. However, there are several limitations of this method. The aim of this research was to detect the percentage of submicroscopic STH infections from human fecal samples (children, 5?18 years old) in Nangapanda, Ende, using the real-time polymerase chain reaction (PCR) method. The fecal samples were collected in two time periods, which were before and after treatment, using 400 mg of Albendazole. There were 242 samples in total, but only 45 negative samples from microscopic detection were tested with real-time PCR. The DNA samples were isolated and amplified wih primers of internal transcribed spacer (ITS-1 and ITS-2) region of rDNA. The detection of samples with real-time PCR generated an amplification curve in VIC, FAM, and Texas Red fluorophore. Three samples (6.7%) in pre-treatment were low load of DNA (N. americanus and A. lumbricoides) (Ct > 35). Four samples (9.1%) were low load of DNA (N. americanus) (Ct > 35) in post-treatment. Five samples (11.4%) were moderate load of DNA (A. lumbricoides) (30 < Ct < 35) in post-treatment. real-time PCR could detect submicroscopic infections from specific species of hookworm and A. lumbricoides."

"Latarbelakang:Sebagian besar wanita hamil mengalami kolonisasi Streptococcus haemolyticus grup B (SGB) di saluran urogenital yang mempengaruhi kesehatan ibu hamil dan bayi. Deteksi SGBintrapartum perlu pemeriksaan yang cepat dan sensitif. Pemeriksaan mikrobiologi untuk mendeteksi SGB menggunakan metode kultur dan real time polymerase chain reaction(RT-PCR) telah digunakan untuk mendukung diagnosis, namun penggunaannya untuk skrining pada ibu hamil belum pernah diuji keakuratannya di Indonesia. Penelitian ini bertujuan untuk mencari metode terbaik untuk mendeteksi kolonisasi SGB pada ibu hamil sekaligus menilai akurasi uji RT-PCR.Metode: Penelitian dengan metode potong lintang padawanita hamil kurang dari 37 minggu dengan ketuban pecah diRumah Sakit Umum Pusat Nasional Cipto Mangunkusumo(RSCM), Rumah Sakit Umum Daerah Pasar Rebo, dan Rumah Sakit Budi Kemuliaan. serta bayi baru lahir dengan tersangka sepsis neonatorum awitan dini yang lahir dari ibu tersebut di RSCM. Swab rektovaginal pada ibu dan darah pada bayi untuk pemeriksaan tes RT-PCRdan kultur pada 3 media: agar darah (AD), agar darah Columbia (ADC), dan CHROMagar (CA).Hasil: Ada 50 ibu dan 25 bayi direkrut dalam penelitian ini.Prevalensi SGB pada ibu hamil 24%, 2 bayi meninggal. Dibandingkan dengan kultur dengan media ADC, tes RT-PCR mempunyai sensitivitas 83,33%, spesifisitas 86,84 %, NPP 66,67 %, NPN 94,29 %, dan akurasi 86,00 %.Media CAmenunjukkan hasil yang lebih tinggi dalam hal sensitivitas 100%, spesifisitas 100%, NPP 100%, NPN 100%, dan akurasi 100 %, dengan hasil lebih singkat, lebih praktis, dan lebih murah.Simpulan: Pemeriksaan RT-PCR menjadi pilihan dalam skrining SGB intrapartum, dengan alternatif media CA.

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Background: Most pregnant women were colonized of group B Streptococcus haemolyticus (GBS) in urogenital tract which affects the health of pregnant women and babies. Detection of intrapartum GBS requires rapid and sensitive examination. Microbiological examination to detect GBS using culture and real time polymerase chain reaction (RT-PCR) has been used to support the diagnosis, but its use for screening in pregnant women has never been tested for accuracy in Indonesia. This study aims to find the best method for detecting GBS colonization in pregnant women as well as assessing the accuracy of the RT-PCR test.

Methods: This was a cross-sectional study in pregnant women less than 37 weeks with ruptured membranes at the Cipto Mangunkusumo National Central General Hospital (RSCM), Pasar Rebo Regional General Hospital, and Budi Kemuliaan Hospital, And newborns with suspected early-onset neonatal sepsis born to these mothers at RSCM. Rectovaginal swab in mother and blood in infant for RT-PCR assay and culture on 3 media: blood agar (BA), Columbia blood agar (CBA), and CHROMagar (CA).

Results: There were 50 mothers and 25 infants recruited in this study. The prevalence of GBS in pregnant women was 24%, 2 neonates died. Compared with culture with CBA media, the RT-PCR test had a sensitivity of 83,33%, specificity 86,84%, PPN 66,67%, NPN 94,29%, and accuracy 86,00%. CA media showed higher results in terms of 100% sensitivity, 100% specificity, 100% NPP, 100% NPN, and 100% accuracy, with shorter results, more practical, and cheaper.

Conclusions: RT-PCR examination is an option in intrapartum GBS screening, with CA media as an alternative. "

"ABSTRAK Latar belakang: Diare masih merupakan masalah kesehatan masyarakat dinegara berkembang karena morbiditas dan mortalitasnya yang masih tinggi. Diaredapat disebabkan oleh virus, bakteri dan parasit yang penting diketahui untukmemberikan tatalaksana yang tepat, namun saat ini belum ada data mengenaibakteri penyebab diare di Indonesia. Tujuan: Mengetahui gambaran klinis anak dengan diare akut dan mengetahui jenis bakteri enteropatogen penyebab diare akut dengan menggunakan real-timePCR di Rumah Sakit Cipto Mangunkusumo. Metode: Penelitian potong lintang pada anak dengan diare akut berusia 1-15tahun di Rumah Sakit Cipto Mangunkusumo. Hasil: Subyek penelitian ini terdiri dari 60 subyek dengan diare akut. Sebagian besar berusia 1-3 tahun, status gizi baik, berasal dari ibu dengan pendidikansedang dengan status ekonomi keluarga menengah rendah, sebagian besar belummendapat antibiotik sebelum ke rumah sakit tetapi sudah mendapat cairanrehidrasi oral. Gambaran klinis diare akut akibat infeksi bakteri yaitu frekuensidiare ≤5X/hari (p=0,018), tanpa leukositosis feses (p=0,015) dan malabsorpsilemak (p=0,031). Sebaran infeksi bakteri patogen penyebab diare akutberdasarkan real-time PCR sebagai berikut: Campylobacter jejuni 7 subyek,Escherichia coli patogen 17 subyek yang terdiri dari EPEC 9 subyek, EIEC 5subyek dan ETEC 3 subyek. Infeksi bakteri campuran pada subyek sebagaiberikut: EPEC+EIEC 2 subyek , C.jejuni+EPEC 1 subyek, C.jejuni+EPEC+EIEC1 subyek dan C.jejuni+EPEC+ETEC 1 subyek. Simpulan: Sebagian besar diare terjadi pada usia 1-3 tahun dengan status pasiengizi baik dengan status keluarga menengah rendah. Sekitar 48% anak dengandiare akut didapatkan bakteri dari hasil real-time PCR feses dengan proporsiterbanyak yaitu EPEC, diikuti Campylobacter jejuni, EIEC dan ETEC.ABSTRACT Background: Diarrhea is still a public health problem in developing countriesdue to it?s morbidity and mortality. Diarrhea can be caused by viruses, bacteriaand parasites. It is important to know the etiology to provide proper management,but there is currently no data on the bacteria that causes diarrhea in Indonesia. Objective: To characterize the clinical manifestations of children with acutediarrhea and determine the type of enteropathogens bacteria causing acutediarrhea using real-time PCR in Cipto Mangunkusumo Hospital. Methods: This was a cross-sectional study, done in June-November 2015. Stoolspecimens were collected from patients aged 1-15 years with acute diarrhea andtested for bacterial enteropathogens using real-time PCR.Results: Of the 60 children enrolled, mostly aged 1-3 years, good nutritionalstatus, from low income families and secondary education mothers, most have notreceived antibiotics prior to hospital admission but had received oral rehydrationfluids. The clinical features of acute diarrhea caused by bacterial infection isdiarrhea frequency ≤5X / day without fecal leukocytosis and fat malabsorption.From 60 subjects, 29 (48,3%) children excreted bacteria in their feces prooved byreal-time PCR. Distribution of pathogenic bacterial infection causes acute diarrheaby real-time PCR as follows: Campylobacter jejuni 7 subjects, pathogenicEscherichia coli 17 subjects which consists of EPEC 9 subjects, EIEC 5 subjectsand ETEC 3 subjects. Multiple infections in subjects as follows: EPEC+EIEC 2subjects, EPEC+C.jejuni 1 subject, C.jejuni+EPEC+EIEC 1 subject andC.jejuni+EPEC+ETEC 1 subject. Conclusions: Most diarrhea occurs at the age of 1-3 years with good nutritional status of patients with low-medium family status. Approximately 48% of children with acute diarrhea excreted bacteria in their feces prooved by real-time PCRstool with the highest proportion is EPEC, followed by Campylobacter jejuni,EIEC and ETEC.;Background: Diarrhea is still a public health problem in developing countriesdue to it?s morbidity and mortality. Diarrhea can be caused by viruses, bacteriaand parasites. It is important to know the etiology to provide proper management,but there is currently no data on the bacteria that causes diarrhea in Indonesia. Objective: To characterize the clinical manifestations of children with acutediarrhea and determine the type of enteropathogens bacteria causing acutediarrhea using real-time PCR in Cipto Mangunkusumo Hospital. Methods: This was a cross-sectional study, done in June-November 2015. Stoolspecimens were collected from patients aged 1-15 years with acute diarrhea andtested for bacterial enteropathogens using real-time PCR.Results: Of the 60 children enrolled, mostly aged 1-3 years, good nutritionalstatus, from low income families and secondary education mothers, most have notreceived antibiotics prior to hospital admission but had received oral rehydrationfluids. The clinical features of acute diarrhea caused by bacterial infection isdiarrhea frequency ≤5X / day without fecal leukocytosis and fat malabsorption.From 60 subjects, 29 (48,3%) children excreted bacteria in their feces prooved byreal-time PCR. Distribution of pathogenic bacterial infection causes acute diarrheaby real-time PCR as follows: Campylobacter jejuni 7 subjects, pathogenicEscherichia coli 17 subjects which consists of EPEC 9 subjects, EIEC 5 subjectsand ETEC 3 subjects. Multiple infections in subjects as follows: EPEC+EIEC 2subjects, EPEC+C.jejuni 1 subject, C.jejuni+EPEC+EIEC 1 subject andC.jejuni+EPEC+ETEC 1 subject. Conclusions: Most diarrhea occurs at the age of 1-3 years with good nutritional status of patients with low-medium family status. Approximately 48% of children with acute diarrhea excreted bacteria in their feces prooved by real-time PCRstool with the highest proportion is EPEC, followed by Campylobacter jejuni,EIEC and ETEC.;Background: Diarrhea is still a public health problem in developing countriesdue to it?s morbidity and mortality. Diarrhea can be caused by viruses, bacteriaand parasites. It is important to know the etiology to provide proper management,but there is currently no data on the bacteria that causes diarrhea in Indonesia. Objective: To characterize the clinical manifestations of children with acutediarrhea and determine the type of enteropathogens bacteria causing acutediarrhea using real-time PCR in Cipto Mangunkusumo Hospital. Methods: This was a cross-sectional study, done in June-November 2015. Stoolspecimens were collected from patients aged 1-15 years with acute diarrhea andtested for bacterial enteropathogens using real-time PCR.Results: Of the 60 children enrolled, mostly aged 1-3 years, good nutritionalstatus, from low income families and secondary education mothers, most have notreceived antibiotics prior to hospital admission but had received oral rehydrationfluids. The clinical features of acute diarrhea caused by bacterial infection isdiarrhea frequency ≤5X / day without fecal leukocytosis and fat malabsorption.From 60 subjects, 29 (48,3%) children excreted bacteria in their feces prooved byreal-time PCR. Distribution of pathogenic bacterial infection causes acute diarrheaby real-time PCR as follows: Campylobacter jejuni 7 subjects, pathogenicEscherichia coli 17 subjects which consists of EPEC 9 subjects, EIEC 5 subjectsand ETEC 3 subjects. Multiple infections in subjects as follows: EPEC+EIEC 2subjects, EPEC+C.jejuni 1 subject, C.jejuni+EPEC+EIEC 1 subject andC.jejuni+EPEC+ETEC 1 subject. Conclusions: Most diarrhea occurs at the age of 1-3 years with good nutritional status of patients with low-medium family status. Approximately 48% of children with acute diarrhea excreted bacteria in their feces prooved by real-time PCRstool with the highest proportion is EPEC, followed by Campylobacter jejuni,EIEC and ETEC."

"Pada akhir tahun 2019 dilaporkan beberapa kasus pneumonia di Wuhan, Cina yang disebabkan oleh Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2). Penyakit yang ditimbulkan oleh virus ini disebut sebagai coronavirus disease 2019 (COVID-19). Jumlah kasus COVID-19 terus mengalami peningkatan dan penyebarannya terjadi pada seluruh kelompok usia termasuk anak-anak. Pemeriksaan real-time reverse transcription-polymerase chain reaction (rRT-PCR) telah diotorisasi oleh Food and Drug Administration (FDA) dan pada pemeriksaan ini dikenal istilah cycle threshold (Ct). Nilai Ct sering dijadikan acuan dalam menentukan tingkat keparahan penyakit pada anak, akan tetapi masih terdapat kontroversi apakah nilai Ct berhubungan dengan tingkat keparahan penyakit. Penelitian ini bermaksud mencari hubungan bermakna antara nilai Ct khususnya gen ORF1ab dan gen N dengan tingkat keparahan COVID-19 pada anak yang dibagi menjadi tingkat keparahan ringan dan sedang sampai kritis. Didapat 52 responden anak dalam penelitian ini, dengan 24 responden terdeteksi gen ORF1ab dan 49 responden terdeteksi gen N. Rerata nilai Ct gen ORF1ab kelompok ringan (33,5 ± 4,4) lebih tinggi dibandingkan dengan kelompok sedang sampai kritis (31,0 ± 6,0). Median nilai Ct gen N kelompok ringan (34,8 [21,3 – 39,4]) lebih tinggi dibandingkan dengan kelompok sedang sampai kritis (31,7 [19,4 – 38,9]). Tidak didapatkan hubungan bermakna baik antara nilai Ct gen ORF1ab (nilai p = 0,25) maupun gen N (nilai p = 0,159) dengan tingkat keparahan COVID-19 pada anak. Berdasarkan hasil penelitian ini, diperlukan berbagai pertimbangan dalam menginterpretasi nilai Ct.

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At the end of 2019, several cases of pneumonia were reported in Wuhan, China caused by Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2). The disease caused by this virus is known as Coronavirus Disease 2019 (COVID-19). The number of cases of COVID-19 continues to increase and its spread occurs in all age groups including children. Real-time reverse transcription-polymerase chain reaction (rRT-PCR) has been authorized by the Food and Drug Administration (FDA) and in this method a cycle threshold (Ct) value were obtained. The Ct value is often used as a reference in determining the clinical severity in children, but there is still controversy whether the Ct value is related to the clinical severity. This study intends to find a significant relationship between the Ct values, especially the ORF1ab gene and the N gene, with the COVID-19 clinical severity in children which is divided into mild and moderate to critical severity. There were 52 children in this study, with 24 children have ORF1ab gene detected and 49 children have N gene detected. The mean of ORF1ab gene Ct value in mild group (33.5 ± 4.4) was higher than moderate to critical group (31.0 ± 6.0). The median of N gene Ct value ​​in mild group (34.8 [21.3 – 39.4]) was higher than moderate to critical group (31.7 [19.4 – 38.9]). There was no significant relationship between the Ct value of the ORF1ab gene (p value = 0.25) and the N gene (p value = 0.159) with COVID-19 clinical severity in children. Based on the results of this study, various considerations are needed in interpreting the Ct value."

"ABSTRAK Latar belakang : Cytomegalovirus (CMV) merupakan salah satu infeksi oportunistikpada pasien dengan sindrom immunodefisiensi (AIDS). Gejala klinis dan CT scantidak dapat menegakkan diagnosa definitif ensefalitis CMV. Oleh karena itudiperlukan uji alternatif untuk menegakkan diagnosis infeksi CMV pada pasien HIVdengan infeksi otak. Salah satu uji yang sensitif dan spesifik adalah Real TimePolymerase Chain Reaction (rPCR).Tujuan : Mendapatkan uji deteksi molekular CMV pada pasien HIV dengantersangka infeksi otak.Metode : Penelitian dilakukan dalam 3 tahap. Tahap 1 adalah optimasi konsentrasiprimer, probe, suhu annealing, volume elusi ekstraksi DNA, dan volume cetakan.Tahap 2 adalah uji spesifisitas (reaksi silang) dan uji sensitivitas (ambang batasdeteksi DNA) rPCR dan tahap 3 adalah penerapan uji rPCR yang sudah dioptimasiterhadap sampel plasma, urin, dan LCS.Hasil : Kondisi optimal uji rPCR telah diperoleh dengan konsentrasi primer danprobe 0,1 μM, dengan kondisi suhu reaksi rPCR: aktivasi enzim pada 950C selama 3menit; 45 siklus pada 950C selama 15 detik (denaturasi) dan 560C selama 1 menit(annealing dan ekstensi). Volume elusi ekstraksi DNA yang optimal untuk ketigajenis sampel (LCS, plasma dan urin) adalah 40 μL, dan volume cetakan rPCR untukLCS, plasma, dan urin, masing-masing adalah 5, 4, dan 3 μL. Uji rPCR mampumendeteksi DNA pada 50.000 jumlah kopi/mL dan tidak bereaksi silang denganStaphylococcus aureus, Staphylococcus epidermidis, Staphylococcus saprophyticus,Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, Mycobacteriumtuberculosis, Candida spp, Toxoplasma gondii, EBV,HSV,dan VZV. Penerapan ujirPCR pada sampel klinis memberikan hasil negatif pada semua sampel LCS, 72,22%positif pada sampel plasma, dan 72,22% positif pada sampel urin.Kesimpulan: Telah dilakukan optimasi uji rPCR dengan minimal deteksi DNACMV 50.000 jumlah kopi/mL dan tidak bereaksi silang dengan mikroorganisme yangberpotensi menyebabkan positif palsu (false positive).ABSTRACT Background: Cytomegalovirus (CMV) is one of opportunistic infections in patientswith Aquired Immunodeficiency Syndrome (AIDS). Clinical manifestations are nottypical, and CT scans can not define encephalitis CMV specifically. Therefore, it isimportant to apply an alternative assay for sensitive and specific detection of CMVinfection in HIV patients with suspected central nervous system (CNS) infections.One of the assays is real time polymerase chain reaction (rPCR).Objective: To obtain a molecular assay for detection of CMV in HIV patients withsuspect CNS infections.Methods: This study was conducted in three phases. The first is optimization ofconcentrations of primers, probe, annealing temperature, final elution of DNAextraction, and volume of PCR template. The second is determinations of sensitivity(minimal detection of DNA) and specificity (cross-reaction) of the optimized rPCR,and the third is application of the rPCR for clinical samples of plasma, urine, andliquor cerebrospinal (LCS).Results: The rPCR reaction showed optimal concentrations of primers and probe at0.1 μM, with thermal cycler: 950C for 3 min (enzyme activation), followed by 45cycles of 950C for 15 sec (denaturation) and 560C for 1 min (annealing andextension). Final elution of DNA extraction was 40 μL and volume of PCR templatesfor urine, plasma, and LCS was 3, 4, and 5 μL, respectively. The rPCR had minimaldetection of DNA at 50,000 copies/mL and was not cross-reacted withStaphylococcus aureus, Staphylococcus epidermidis, Staphylococcus saprophyticus,Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, Mycobacteriumtuberculosis, Candida spp, Toxoplasma gondii, Epstein-Bar Virus (EBV), HerpesSimplex Virus (HSV) and Varicella Zoster Virus (VZV). Application of rPCR forclinical samples showed that the rPCR yielded 72.22% positive for plasma or urine,and negative for all LCS samples.Conclusion: The rPCR has been optimized in this study with minimal DNA detectionat 50,000 copies/mL and was not cross-reacted with other microorganisms that arepotential to cause false positive results.;Background: Cytomegalovirus (CMV) is one of opportunistic infections in patientswith Aquired Immunodeficiency Syndrome (AIDS). Clinical manifestations are nottypical, and CT scans can not define encephalitis CMV specifically. Therefore, it isimportant to apply an alternative assay for sensitive and specific detection of CMVinfection in HIV patients with suspected central nervous system (CNS) infections.One of the assays is real time polymerase chain reaction (rPCR).Objective: To obtain a molecular assay for detection of CMV in HIV patients withsuspect CNS infections.Methods: This study was conducted in three phases. The first is optimization ofconcentrations of primers, probe, annealing temperature, final elution of DNAextraction, and volume of PCR template. The second is determinations of sensitivity(minimal detection of DNA) and specificity (cross-reaction) of the optimized rPCR,and the third is application of the rPCR for clinical samples of plasma, urine, andliquor cerebrospinal (LCS).Results: The rPCR reaction showed optimal concentrations of primers and probe at0.1 μM, with thermal cycler: 950C for 3 min (enzyme activation), followed by 45cycles of 950C for 15 sec (denaturation) and 560C for 1 min (annealing andextension). Final elution of DNA extraction was 40 μL and volume of PCR templatesfor urine, plasma, and LCS was 3, 4, and 5 μL, respectively. The rPCR had minimaldetection of DNA at 50,000 copies/mL and was not cross-reacted withStaphylococcus aureus, Staphylococcus epidermidis, Staphylococcus saprophyticus,Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, Mycobacteriumtuberculosis, Candida spp, Toxoplasma gondii, Epstein-Bar Virus (EBV), HerpesSimplex Virus (HSV) and Varicella Zoster Virus (VZV). Application of rPCR forclinical samples showed that the rPCR yielded 72.22% positive for plasma or urine,and negative for all LCS samples.Conclusion: The rPCR has been optimized in this study with minimal DNA detectionat 50,000 copies/mL and was not cross-reacted with other microorganisms that arepotential to cause false positive results.;Background: Cytomegalovirus (CMV) is one of opportunistic infections in patientswith Aquired Immunodeficiency Syndrome (AIDS). Clinical manifestations are nottypical, and CT scans can not define encephalitis CMV specifically. Therefore, it isimportant to apply an alternative assay for sensitive and specific detection of CMVinfection in HIV patients with suspected central nervous system (CNS) infections.One of the assays is real time polymerase chain reaction (rPCR).Objective: To obtain a molecular assay for detection of CMV in HIV patients withsuspect CNS infections.Methods: This study was conducted in three phases. The first is optimization ofconcentrations of primers, probe, annealing temperature, final elution of DNAextraction, and volume of PCR template. The second is determinations of sensitivity(minimal detection of DNA) and specificity (cross-reaction) of the optimized rPCR,and the third is application of the rPCR for clinical samples of plasma, urine, andliquor cerebrospinal (LCS).Results: The rPCR reaction showed optimal concentrations of primers and probe at0.1 μM, with thermal cycler: 950C for 3 min (enzyme activation), followed by 45cycles of 950C for 15 sec (denaturation) and 560C for 1 min (annealing andextension). Final elution of DNA extraction was 40 μL and volume of PCR templatesfor urine, plasma, and LCS was 3, 4, and 5 μL, respectively. The rPCR had minimaldetection of DNA at 50,000 copies/mL and was not cross-reacted withStaphylococcus aureus, Staphylococcus epidermidis, Staphylococcus saprophyticus,Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, Mycobacteriumtuberculosis, Candida spp, Toxoplasma gondii, Epstein-Bar Virus (EBV), HerpesSimplex Virus (HSV) and Varicella Zoster Virus (VZV). Application of rPCR forclinical samples showed that the rPCR yielded 72.22% positive for plasma or urine,and negative for all LCS samples.Conclusion: The rPCR has been optimized in this study with minimal DNA detectionat 50,000 copies/mL and was not cross-reacted with other microorganisms that arepotential to cause false positive results.;Background: Cytomegalovirus (CMV) is one of opportunistic infections in patientswith Aquired Immunodeficiency Syndrome (AIDS). Clinical manifestations are nottypical, and CT scans can not define encephalitis CMV specifically. Therefore, it isimportant to apply an alternative assay for sensitive and specific detection of CMVinfection in HIV patients with suspected central nervous system (CNS) infections.One of the assays is real time polymerase chain reaction (rPCR).Objective: To obtain a molecular assay for detection of CMV in HIV patients withsuspect CNS infections.Methods: This study was conducted in three phases. The first is optimization ofconcentrations of primers, probe, annealing temperature, final elution of DNAextraction, and volume of PCR template. The second is determinations of sensitivity(minimal detection of DNA) and specificity (cross-reaction) of the optimized rPCR,and the third is application of the rPCR for clinical samples of plasma, urine, andliquor cerebrospinal (LCS).Results: The rPCR reaction showed optimal concentrations of primers and probe at0.1 μM, with thermal cycler: 950C for 3 min (enzyme activation), followed by 45cycles of 950C for 15 sec (denaturation) and 560C for 1 min (annealing andextension). Final elution of DNA extraction was 40 μL and volume of PCR templatesfor urine, plasma, and LCS was 3, 4, and 5 μL, respectively. The rPCR had minimaldetection of DNA at 50,000 copies/mL and was not cross-reacted withStaphylococcus aureus, Staphylococcus epidermidis, Staphylococcus saprophyticus,Escherichia coli, Klebsiella pneumoniae, Pseudomonas aeruginosa, Mycobacteriumtuberculosis, Candida spp, Toxoplasma gondii, Epstein-Bar Virus (EBV), HerpesSimplex Virus (HSV) and Varicella Zoster Virus (VZV). Application of rPCR forclinical samples showed that the rPCR yielded 72.22% positive for plasma or urine,and negative for all LCS samples.Conclusion: The rPCR has been optimized in this study with minimal DNA detectionat 50,000 copies/mL and was not cross-reacted with other microorganisms that arepotential to cause false positive results."