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"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."

"Antimicrobial Resistance (AMR) menyebabkan penurunan efektivitas pengobatan dan dapat dideteksi dengan keberadaan Antibiotik Resistance Genes (ARG) seperti bla_CTX-M yang paling banyak ditemukan dan memberikan resistansi terhadap antibiotik sefotaksim. Hingga saat ini, belum ada penelitian yang bertujuan untuk mendeteksi keberadaan gen bla_CTX-M pada Sungai Bekasi. Penelitian ini bertujuan untuk menganalisis konsentrasi bakteri Escherichia coli non-selektif dan resistan pada hilir Sungai Bekasi, menganalisis keberadaan ARG bla_CTX-M pada bakteri E. coli tersebut, serta memberikan rekomendasi lokasi sampling bakteri E. coli untuk mendeteksi AMR. Metode Polymerase Chain Reaction pada penelitian ini digunakan untuk mengamplifikasi DNA dan dilanjutkan dengan elektroforesis untuk pembacaan ukuran DNA. Berdasarkan hasil penelitian, hilir Sungai Bekasi memiliki rata-rata konsentrasi bakteri E. coli non-selektif sebesar 261 CFU/mL dengan titik sampling 1/3 dari pinggir sungai dan sebesar 207 CFU/mL dengan titik sampling di pinggir sungai. Sedangkan, rata-rata konsentrasi bakteri E. coli resistansi antibiotik sefotaksim sebesar 26 CFU/mL dengan titik sampling 1/3 dari pinggir sungai dan sebesar 20 CFU/mL dengan titik sampling di pinggir sungai. Rasio perbandingan bakteri E. coli resisten antibiotik dengan bakteri E. coli non-selektif adalah 9,78% dengan titik sampling 1/3 dari pinggir sungai dan 9,27% dengan titik sampling di pinggir sungai. Hasil penelitian mendapatkan bahwa adanya keberadaan gen resistansi antibiotik bla_CTX-M pada hilir Sungai Bekasi sebanyak 80% dari sampel yang diambil. Dimana gen yang mendominasi adalah CTX-M grup 1 yang beranggotakan gen CTX-M-1, CTX-M-3, dan CTX-M-15. Hasil penelitian juga menunjukan bahwa sampel dari pinggir sungai memiliki hasil yang lebih homogen dan lebih mudah untuk dilakukan.

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Antimicrobial Resistance (AMR) causes a decrease in the effectiveness of treatment and can be detected by the presence of Antibiotic Resistance Genes (ARG) such as bla_CTX-M which is the most commonly found and provides resistance to the antibiotic cefotaxime. Until now, there has been no research aimed at detecting the presence of the bla_CTX-M gene in the Bekasi River. This study aims to analyze the concentration of non-selective and resistant Escherichia coli bacteria at downstream of the Bekasi River, analyze the presence of ARG bla_CTX-M in the E. coli bacteria, and provide recommendations for sampling locations for E. coli bacteria to detect AMR. The Polymerase Chain Reaction method in this study was used to amplify DNA and was followed by electrophoresis to read the size of the DNA. Based on the research results, the downstream Bekasi River has an average concentration of non-selective E. coli bacteria of 261 CFU/mL with a sampling point 1/3 from the riverbank and 207 CFU/mL with a sampling point on the riverbank. Meanwhile, the average concentration of cefotaxime-resistant E. coli bacteria was 26 CFU/mL with a sampling point of 1/3 from the riverbank and 20 CFU/mL with a sampling point on the riverbank. The ratio of antibiotic resistant E. coli bacteria to non-selective E. coli bacteria was 9.78% with a sampling point of 1/3 from the riverbank and 9.27% with a sampling point on the riverbank. The results of the study found that the presence of the bla_CTX-M antibiotic resistance gene in the downstream of the Bekasi River was as much as 80% of the samples taken. Where the dominant gene was CTX-M group 1 consisting of the CTX-M-1, CTX-M-3, and CTX-M-15. The results of the study also shown that samples from the riverbank have more homogeneous results and are easier to carry out."

"ABSTRACTVibrio cholera, Vibrio parahaemolyticus and Vibrio vulnificus are opportunistic pathogens causing disease in weak shrimp and possible food poisoning in shrimp consumers. In this study, three pairs of primers were designed to amplify the target DNA fragments of three Vibrio spp. and together with one pair for internal amplification control. The PCR condition was optimized to detect three Vibrio spp. in one reaction tube. The specificity and sensitivity of the reaction were evaluated. The results showed that this technique can detect V. cholera, V. parahaemolyticus and V. vulnificus in the same reaction tube with high specificity. Sensitivity is moderate, 0.5 ng-10 pg. In the future, this technique can be used to detect these bacteria in shrimp. It is potentially useful for shrimp farmers and shrimp consumers."

"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."