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"ABSTRAK Lesi fokal otak merupakan komplikasi neurologi pada pasien HIV yang ditandai oleh lesi desak ruang (Space Occupying Lesion) yang membutuhkan penanganan cepat dan tepat. Di beberapa negara, lesi ini dapat disebabkan oleh toksoplasma ensefalitis dan limfoma otak primer. Lesi yang disebabkan oleh toksoplasmosis dan limfoma otak primer yang disebabkan oleh Epstein Barr virus sulit untuk dibedakan menggunakan CT scan ataupun MRI. Pemeriksaan gold standar untuk membedakan keduanya yaitu dengan biopsi otak, namun hal ini merupakan tindakan invasif dan dapat menimbulkan komplikasi. Penelitian ini bertujuan untuk memperoleh uji deteksi untuk diagnosis cepat infeksi Toxoplasma gondii dan Epstein Barr virus. Desain yang dipakai pada penelitian adalah studi eksperimental laboratorium. Uji deteksi yang dikembangkan adalah dupleks real-time PCR yang dapat mendeteksi T.gondii dan EBV atau kombinasi keduanya dalam satu reaksi pada sampel pasien HIV dengan gejala klinis tersangka infeksi otak. Tahap pertama dilakukan optimasi dupleks real-time PCR meliputi suhu annealing, konsentrasi primer dan probe, uji volume elusi dan volume cetakan. Penentuan ambang batas deteksi dilakukan untuk mengukur minimal T.gondii dan EBV yang dapat dideteksi. Reaksi silang untuk mengetahui spesifisitas teknik dilakukan menggunakan bakteri dan virus sebagai berikut Staphylococcus aureus, Klebsiella pneumonia, Pseudomonas aeruginosa, Mycobacterium tuberculosis H37Rv, Candida spp, Cytomegalo virus, Herpes zoster virus, dan Varicella zoster virus. Dupleks real-time PCR yang telah optimal diaplikasi pada sampel pasien. Sampel yang digunakan adalah darah dan cairan serebrospinal dari pasien HIV dengan gejala klinis infeksi otak yang dirawat di bagian neurologi RSCM. Hasil optimasi dupleks real-time PCR diperoleh suhu annealing untuk T.gondii dan EBV 58°C, konsentrasi primer forward dan reverse untuk T.gondii dan EBV adalah 0,2 µM, konsentrasi probe T.gondii 0,4µM, konsentrasi probe EBV 0,2 µM. Deteksi ambang batas minimal DNA untuk T.gondii 5,68 copy /ml, sedangkan EBV 1,31 copy/ml. Uji yang dikembangkan pada penelitian ini termasuk uji yang sensitif dibandingkan hasil penelitian lain. Uji reaksi silang primer dan probe dupleks real-time PCR terhadap beberapa bakteri dan virus lain, menunjukkan tidak bereaksi silang dengan primer dan probe yang digunakan untuk mendeteksi T.gondii dan EBV. Hasil pemeriksaan dupleks real-time PCR pada sampel darah diperoleh 16% positif T.gondii, 40% positif Epstein Barr virus, sebanyak 16% positif Epstein Barr virus dan T.gondii dan pada sampel cairan serebrospinal diperoleh hasil 20% positif T.gondii, sebanyak 28% positif Epstein Barr virus dan 4% positif terhadap Epstein Barr Virus dan T.gondii. ABSTRACT Focal brain lesion is neurology complication in HIV that marked with Space Occupying Lesion (SOL), that need rapid and effective handling. In most country, this lesion could be cause by encephalitis toxoplasma and Primary Central Nervous System Lymphoma that related to Epstein Barr virus infection that was difficult to distinguished using CT scan or MRI. Gold standard to distinguished was brain biopsy, but this examination was invasive procedure that cause complication. Therefore, we need a reliable and rapid examination to distinguished it. This study aimed to get detection for rapid diagnosis of T.gondii and EBV infection. This study was an experimental laboratory. First step was optimation of dupleks real-time PCR include annealing temperature, primer andprobe consentration, elution volume and template volume. Minimal detection of DNA to measured minimal T.gondii and EBV that could be detected. Cross reaction to know technique spesivisity using bacterial and virus Staphylococcus aureus, Klebsiella pneumonia, Pseudomonas aeruginosa, Mycobacterium tuberculosis H37Rv, Candida spp, Cytomegalo virus, Herpes zoster virus, and Varicella zoster virus. Dupleks real-time PCR has been optimally applied to patient. The sample from blood and cerebrospinal fluid of HIV patients who admitted in the neurology department of RSCM then examined to duplex real-time PCR to detect T.gondii and EBV. The optimation of duplex real-time PCR, the annealing temperature for T.gondii and EBV were 58°C, consentration of primer forward and reverse for T.gondii and EBV were 0,2 µM, consentration of probe for T.gondii was 0,4µM and EBV was 0,2µM.. Minimal DNA detection for T.gondii was 5,68 copy/ml and EBV was 1,31 copy /ml. This study was sensitive like the others. Spesivisity technique of real-time PCR, there was not cross reaction between another bacteria and virus that used as primer and probe for T.gondii and EBV. From the results of the duplex real-time PCR on blood samples, 16 % was positive T.gondii, 40% Epstein Barr virus, and 16% were positive Epstein Barr virus and T.gondii and from cerebrospinal fluid samples 20% was positive T.gondii, 28% was positive Epstein Barr virus and 4% were positive for Epstein Barr Virus and T.gondii.;Focal brain lesion is neurology complication in HIV that marked with Space Occupying Lesion (SOL), that need rapid and effective handling. In most country, this lesion could be cause by encephalitis toxoplasma and Primary Central Nervous System Lymphoma that related to Epstein Barr virus infection that was difficult to distinguished using CT scan or MRI. Gold standard to distinguished was brain biopsy, but this examination was invasive procedure that cause complication. Therefore, we need a reliable and rapid examination to distinguished it. This study aimed to get detection for rapid diagnosis of T.gondii and EBV infection. This study was an experimental laboratory. First step was optimation of dupleks real-time PCR include annealing temperature, primer andprobe consentration, elution volume and template volume. Minimal detection of DNA to measured minimal T.gondii and EBV that could be detected. Cross reaction to know technique spesivisity using bacterial and virus Staphylococcus aureus, Klebsiella pneumonia, Pseudomonas aeruginosa, Mycobacterium tuberculosis H37Rv, Candida spp, Cytomegalo virus, Herpes zoster virus, and Varicella zoster virus. Dupleks real-time PCR has been optimally applied to patient. The sample from blood and cerebrospinal fluid of HIV patients who admitted in the neurology department of RSCM then examined to duplex real-time PCR to detect T.gondii and EBV. The optimation of duplex real-time PCR, the annealing temperature for T.gondii and EBV were 58°C, consentration of primer forward and reverse for T.gondii and EBV were 0,2 µM, consentration of probe for T.gondii was 0,4µM and EBV was 0,2µM.. Minimal DNA detection for T.gondii was 5,68 copy/ml and EBV was 1,31 copy /ml. This study was sensitive like the others. Spesivisity technique of real-time PCR, there was not cross reaction between another bacteria and virus that used as primer and probe for T.gondii and EBV. From the results of the duplex real-time PCR on blood samples, 16 % was positive T.gondii, 40% Epstein Barr virus, and 16% were positive Epstein Barr virus and T.gondii and from cerebrospinal fluid samples 20% was positive T.gondii, 28% was positive Epstein Barr virus and 4% were positive for Epstein Barr Virus and T.gondii.;Focal brain lesion is neurology complication in HIV that marked with Space Occupying Lesion (SOL), that need rapid and effective handling. In most country, this lesion could be cause by encephalitis toxoplasma and Primary Central Nervous System Lymphoma that related to Epstein Barr virus infection that was difficult to distinguished using CT scan or MRI. Gold standard to distinguished was brain biopsy, but this examination was invasive procedure that cause complication. Therefore, we need a reliable and rapid examination to distinguished it. This study aimed to get detection for rapid diagnosis of T.gondii and EBV infection. This study was an experimental laboratory. First step was optimation of dupleks real-time PCR include annealing temperature, primer andprobe consentration, elution volume and template volume. Minimal detection of DNA to measured minimal T.gondii and EBV that could be detected. Cross reaction to know technique spesivisity using bacterial and virus Staphylococcus aureus, Klebsiella pneumonia, Pseudomonas aeruginosa, Mycobacterium tuberculosis H37Rv, Candida spp, Cytomegalo virus, Herpes zoster virus, and Varicella zoster virus. Dupleks real-time PCR has been optimally applied to patient. The sample from blood and cerebrospinal fluid of HIV patients who admitted in the neurology department of RSCM then examined to duplex real-time PCR to detect T.gondii and EBV. The optimation of duplex real-time PCR, the annealing temperature for T.gondii and EBV were 58°C, consentration of primer forward and reverse for T.gondii and EBV were 0,2 µM, consentration of probe for T.gondii was 0,4µM and EBV was 0,2µM.. Minimal DNA detection for T.gondii was 5,68 copy/ml and EBV was 1,31 copy /ml. This study was sensitive like the others. Spesivisity technique of real-time PCR, there was not cross reaction between another bacteria and virus that used as primer and probe for T.gondii and EBV. From the results of the duplex real-time PCR on blood samples, 16 % was positive T.gondii, 40% Epstein Barr virus, and 16% were positive Epstein Barr virus and T.gondii and from cerebrospinal fluid samples 20% was positive T.gondii, 28% was positive Epstein Barr virus and 4% were positive for Epstein Barr Virus and T.gondii."

"Lesi fokal otak merupakan komplikasi neurologi pada pasien HIV yang ditandai oleh lesi desak ruang (Space Occupying Lesion) yang membutuhkan penanganan cepat dan tepat. Di beberapa negara, lesi ini dapat disebabkan oleh toksoplasma ensefalitis dan limfoma otak primer. Lesi yang disebabkan oleh toksoplasmosis dan limfoma otak primer yang disebabkan oleh Epstein Barr virus sulit untuk dibedakan menggunakan CT scan ataupun MRI. Pemeriksaan gold standar untuk membedakan keduanya yaitu dengan biopsi otak, namun hal ini merupakan tindakan invasif dan dapat menimbulkan komplikasi. Penelitian ini bertujuan untuk memperoleh uji deteksi untuk diagnosis cepat infeksi Toxoplasma gondii dan Epstein Barr virus. Desain yang dipakai pada penelitian adalah studi eksperimental laboratorium. Uji deteksi yang dikembangkan adalah dupleks real-time PCR yang dapat mendeteksi T.gondii dan EBV atau kombinasi keduanya dalam satu reaksi pada sampel pasien HIV dengan gejala klinis tersangka infeksi otak. Tahap pertama dilakukan optimasi dupleks real-time PCR meliputi suhu annealing, konsentrasi primer dan probe, uji volume elusi dan volume cetakan. Penentuan ambang batas deteksi dilakukan untuk mengukur minimal T.gondii dan EBV yang dapat dideteksi. Reaksi silang untuk mengetahui spesifisitas teknik dilakukan menggunakan bakteri dan virus sebagai berikut Staphylococcus aureus, Klebsiella pneumonia, Pseudomonas aeruginosa, Mycobacterium tuberculosis H37Rv, Candida spp, Cytomegalo virus, Herpes zoster virus, dan Varicella zoster virus. Dupleks real-time PCR yang telah optimal diaplikasi pada sampel pasien. Sampel yang digunakan adalah darah dan cairan serebrospinal dari pasien HIV dengan gejala klinis infeksi otak yang dirawat di bagian neurologi RSCM. Hasil optimasi dupleks real-time PCR diperoleh suhu annealing untuk T.gondii dan EBV 58°C, konsentrasi primer forward dan reverse untuk T.gondii dan EBV adalah 0,2 µM, konsentrasi probe T.gondii 0,4µM, konsentrasi probe EBV 0,2 µM. Deteksi ambang batas minimal DNA untuk T.gondii 5,68 copy /ml, sedangkan EBV 1,31 copy/ml. Uji yang dikembangkan pada penelitian ini termasuk uji yang sensitif dibandingkan hasil penelitian lain. Uji reaksi silang primer dan probe dupleks real-time PCR terhadap beberapa bakteri dan virus lain, menunjukkan tidak bereaksi silang dengan primer dan probe yang digunakan untuk mendeteksi T.gondii dan EBV. Hasil pemeriksaan dupleks real-time PCR pada sampel darah diperoleh 16% positif T.gondii, 40% positif Epstein Barr virus, sebanyak 16% positif Epstein Barr virus dan T.gondii dan pada sampel cairan serebrospinal diperoleh hasil 20% positif T.gondii, sebanyak 28% positif Epstein Barr virus dan 4% positif terhadap Epstein Barr Virus dan T.gondii.

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Focal brain lesion is neurology complication in HIV that marked with Space Occupying Lesion (SOL), that need rapid and effective handling. In most country, this lesion could be cause by encephalitis toxoplasma and Primary Central Nervous System Lymphoma that related to Epstein Barr virus infection that was difficult to distinguished using CT scan or MRI. Gold standard to distinguished was brain biopsy, but this examination was invasive procedure that cause complication. Therefore, we need a reliable and rapid examination to distinguished it. This study aimed to get detection for rapid diagnosis of T.gondii and EBV infection. This study was an experimental laboratory. First step was optimation of dupleks real-time PCR include annealing temperature, primer andprobe consentration, elution volume and template volume. Minimal detection of DNA to measured minimal T.gondii and EBV that could be detected. Cross reaction to know technique spesivisity using bacterial and virus Staphylococcus aureus, Klebsiella pneumonia, Pseudomonas aeruginosa, Mycobacterium tuberculosis H37Rv, Candida spp, Cytomegalo virus, Herpes zoster virus, and Varicella zoster virus. Dupleks real-time PCR has been optimally applied to patient. The sample from blood and cerebrospinal fluid of HIV patients who admitted in the neurology department of RSCM then examined to duplex real-time PCR to detect T.gondii and EBV. The optimation of duplex real-time PCR, the annealing temperature for T.gondii and EBV were 58°C, consentration of primer forward and reverse for T.gondii and EBV were 0,2 µM, consentration of probe for T.gondii was 0,4µM and EBV was 0,2µM.. Minimal DNA detection for T.gondii was 5,68 copy/ml and EBV was 1,31 copy /ml. This study was sensitive like the others. Spesivisity technique of real-time PCR, there was not cross reaction between another bacteria and virus that used as primer and probe for T.gondii and EBV. From the results of the duplex real-time PCR on blood samples, 16 % was positive T.gondii, 40% Epstein Barr virus, and 16% were positive Epstein Barr virus and T.gondii and from cerebrospinal fluid samples 20% was positive T.gondii, 28% was positive Epstein Barr virus and 4% were positive for Epstein Barr Virus and T.gondii."

"Latar belakang: Infeksi intrakranial merupakan masalah yang menjadi tantangan berat bagi setiap dokter yang merawat. Tingkat kematian saat rawat inap pasien infeksi intrakranial sangat tinggi. Walaupun pasien infeksi intrakranial dapat keluar dari rumah sakit dalam keadaan hidup, namun berbagai komplikasi dan masalah paska rawat inap yang kompleks dapat menyebabkan kematian pasien saat rawat jalan. Metode penelitian: Penelitian ini merupakan penelitian kohort retrospektif untuk mengetahui kesintasan 180 hari pada pasien infeksi otak yang dirawat di RS Cipto Mangunkusumo. Populasi penelitian ini adalah subjek dari penelitian Optimization of Diagnosis and Treatment of Tuberculous Meningitis ODT-TBM selama periode Januari-Desember 2015. Keluaran 180 hari subjek diketahui dengan penelusuran data kunjungan rawat jalan melalui rekam medis, telepon, pesan singkat atau kunjungan rumah. Analisis kesintasan Total survival rate dilakukan dengan menggunakan analisis cox regression baik univariat maupun multivariat. Penyajian data kesintasan dilakukan dengan menggunakan kurva kaplan meier. Hasil: Didapatkan 218 pasien dengan diagnosis akhir infeksi intrakranial. Berdasarkan status HIV, didapatkan 47,7 subjek HIV positif dan 52,3 HIV negatif. Tingkat kesintasan 180 hari pasien infeksi intrakranial di RSCM secara umum adalah 43,5. Kesintasan pada kelompok HIV positif 32,7 secara bermakna p 0,005; Rasio Hazard 1,695 1,177-2,442 lebih buruk daripada HIV negatif 53,5. Faktor lain yang mempengaruhi kesintasan adalah usia, papiledema, suhu aksila awal, SKG awal, anemia, hiponatremia, gambaran herniasi serebri pada pencitraan otak, rasio glukosa CSS/serum, dan kadar protein CSS. Kesimpulan : Tingkat kesintasan 180 hari pasien infeksi intrakranial pada penelitian ini rendah. Infeksi HIV secara bermakna mempengaruhi kesintasan pasien infeksi intrakranial.

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Background: Managing brain infection patients is a challenge for every physician. Beside a very high in hospital mortality, many complexes problems and complications can cause patient die after discharge.

Methods: This is a retrospective cohort research to find 180 days outcomes of brain infection patients that admitted in Cipto Mangunkusumo Hospital. The study population is Optimization of Diagnosis and Treatment of Tuberculous Meningitis ODT TBM research subject that admitted in 2015. Health records, phone calls, short message or home visit is done to find patient rsquo s outcome. Total survival rate analysis is done with univariate and multivariate cox regression analysis. The comparison of survival rates between 2 groups is presented by Kaplan Meier curve.

Results: A total of 218 subjects were included in this study. There were 47,7 subjects with HIV positive and 52,3 HIV negative. Overall 180 days survival rates is 43,5. HIV status is strongly influenced the survival rate of brain infection patients in this study p value 0,005 Hazard Ratio 1,695 1,177 2,442. The survival rate of HIV negative subjects was 53,5 that significantly higher than HIV positive subjects 32,7. Other factors that influenced the survival rate in this research are age, papil edema, early axial temperature, Glasgow coma scale, anemia, hyponatremia, imaging of brain herniation, blood CSF glucose ratio and CSF protein.

Conclusion: The survival rate of brain infection patients in this research is low. HIV infection significantly influenced patients rsquo survival rates."

"Latar Belakang: Meskipun kontroversial, hospital readmission (HR) dapat mencerminkan keadaan pasien saat dipulangkan dan sebagai indikator untuk mengevaluasi mutu perawatan rumah sakit (RS). Penelitian ini bertujuan untuk mengetahui insidensi dan faktor risiko HR pada pasien infeksi intrakranial.Metode Penelitian: Studi kohort retrospektif pasien infeksi intrakranial periode April 2019-November 2021, menggunakan data Indonesian Brain Infection Study dan telusur rekam medis. Analisis bivariat menggunakan uji Chi Square dan Mann Whitney, dilanjutkan dengan analisis multivariat regresi logistik.Hasil: Insidensi HR pasien infeksi intrakranial sebesar 28,45%. Mayoritas subjek mengalami HR sebelum 30 hari (64,7%). Penyebab HR terbanyak adalah penyakit lain yang berbeda dengan diagnosis awal (55,9%). Komorbid penyakit ginjal meningkatkan risiko HR (aOR=7,2, IK 95%=2,2-23,8,p=0,000). Gejala klinis saat perawatan awal berupa kelemahan motorik dan kejang juga meningkatkan risiko HR (aOR=2,27,IK 95%=1,28-4,01, p=0,001) dan (aOR=1,93,IK 95%=1,02-3,62, p=0,037). Sedangkan ketersediaan pelaku rawat dapat menurunkan risiko HR (aOR=0,07,IK 95%=0,03-0,45, p=0,002).Kesimpulan: Insidensi HR pada pasien infeksi intrakranial dalam waktu 6 bulan sebesar 28,45%. Penyakit ginjal, gejala klinis kelemahan motorik dan kejang pada perawatan awal merupakan faktor yang dapat meningkatkan risiko HR, sedangkan ketersediaan pelaku rawat merupakan faktor yang dapat menurunkan risiko HR. 

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Background: Although controversial, hospital readmission (HR) can reflect the patient's condition at discharge and as an indicator to evaluate the quality of hospital care. This study aims to determine the incidence and risk factors for HR in intracranial infections.

Method: A retrospective cohort study of intracranial infection patients, in period April 2019-November 2021, using secondary data from the Indonesian Brain Infection Study and tracing medical records. Bivariate analysis using Chi Square and Mann Whitney test, followed by multivariate logistic regression analysis.

Results: The incidence of HR in patients with intracranial infections was 28.45%. The majority of subjects experienced HR before 30 days (64.7%). The most common cause of HR was other diseases that were different from the initial diagnosis (55.9%). Kidney disease comorbidity increased HR risk (aOR=7.2;95%CI=2.2-23.8;p=0.000). Clinical symptoms during initial treatment such as motor weakness and seizures also increased the risk of HR (aOR=2.27;95%CI=1.28-4.01;p=0.001) and (aOR=1.93;95%CI=1.02-3.62;p=0.037). Meanwhile, the availability of caregivers can reduce HR risk (aOR=0.07;CI 95=0.03-0.45;p=0.002).  

Conclusion: The incidence of HR in patients with intracranial infection within 6 months was 28.45%. Kidney disease, motor weakness and seizures are factors that can increase the risk of HR, while the availability of caregivers is a factor that can reduce the risk of HR."

"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."

"ABSTRAKToksoplasmosis merupakan penyakit yang disebabkan oleh Toxoplasma gondii. Penyakit infeksi ini dapat menyebabkan kondisi fatal bila terjadi pada pasien imunokompromis, misalnya adalah toksoplasma ensefalitis (TE) yang menyerang sistem saraf pusat. Untuk menegakkan diagnosis Toxoplasma sebagai penyebab kelainan SSP (sistem saraf pusat) pada pasien HIV sangat sulit, sehingga diperlukan metode pemeriksaan lain sebagai alternatif, salah satunya adalah pemeriksaan PCR mendeteksi gen B1 dari Toxoplasma gondii. Penelitian ini bertujuan untuk mengembangkan metode PCR pada sampel CSS (cairan serebrospinal) yang sesuai untuk menegakkan diagnosis TE dan mengetahui keunggulan dan kelemahan pemeriksaan PCR dibandingkan pemeriksaan IgG anti-Toxoplasma pada cairan serebrospinal. Penelitian dilakukan pada cairan serebrospinal pasien HIV/AIDS dengan gangguan serebral. Hasil pemeriksaan PCR dari 88 sampel CSS pasien HIV yang datang ke Laboratorium Parasitologi FK UI, adalah 23 (26,1%) positif dan 65 (73,9%) negative T. gondii. Ada hubungan postif bermakna antara pemeriksaan PCR dengan pemeriksaan IgG anti-Toxoplasma dari CSS.

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ABSTRACT

Toksoplasmosis is a disease caused by infection of Toxoplasma gondii. This infection can caused a life threatening condition in immunocompromised patients, for example toxoplasma ensefalitis (TE) which attack central nervous system. It is very difficult to diagnose Toxoplasma as a cause of CNS infection in HIV patient, so we need another methods as alternative, one of which is one of which is a PCR detection of Toxoplasma gondii B1 gene. This research aims to develop a PCR method on samples Cerebrospinal Fluid (CSF) that suitable for TE diagnosis and determine the advantages and disadvantages PCR methods compared to detection of anti-Toxoplasma IgG from CSF. The study was conducted in the cerebrospinal fluid of patients with HIV / AIDS with cerebral disorders. PCR examination results of 88 samples CSS HIV patients who came to the Laboratory of Parasitology FK UI, was 23 (26.1%) positive and 65 (73.9%) negative T. gondii. There is a significant positive relationship between PCR and detection anti-Toxoplasma IgG from CSF."

"Latar belakang: Difteri merupakan penyakit infeksi bakteri yang diperantarai olehtoksin. Corynebacterium diphtheriae adalah penyebab tersering difteri. Diagnostiklaboratorium harus dilakukan dengan cepat untuk menunjang diagnosis klinis difteri.Pemeriksaan mikroskopik tidak direkomendasikan karena tidak spesifik, kultur dan ujitoksin yang merupakan uji baku emas cukup memakan waktu, membutuhkanketerampilan dan pengalaman serta hanya dilakukan di laboratorium rujukan. PCRmerupakan metode pemeriksaan yang cepat, sensitif dan spesifik. Duplex real-time PCRdapat mendeteksi bakteri penyebab tersering dan gen pengkode toksin secara simultan.Tujuan penelitian: Melakukan optimasi uji duplex real time PCR untuk deteksi C.diphtheriae potensial toksigenik dan menerapkannya pada spesimen usap tenggorokpasien tersangka difteri.Metode: Duplex real-time PCR menggunakan dua pasang primer dan probe dengantarget gen rpoB C.diphtheriae dan toksin difteri subunit A Tox. Parameter yangdioptimasi adalah suhu penempelan, konsentrasi masing-masing primer dan probe,inhibitor, reaksi silang dengan patogen lain dan ambang batas deteksi uji. Kemudian ujidiaplikasikan pada spesimen usap tenggorok pasien tersangka difteri yang dirawat diRSPI Sulianti Saroso pada periode 2018-2019. Sebagai perbandingan dilakukan uji Elekuntuk konfirmasi toksigenitas dan analisa data klinis pasien.Hasil: Kondisi optimal uji didapat pada suhu penempelan 55oC, konsentrasi primer Cd0,4 μM, primer Tox 0,6 μM, probe Cd 0,5 μM dan probe Tox 0,625 μM, volume elusiekstraksi DNA 50 μL, volume cetakan DNA 5 μL dan ambang batas deteksi 2 CFU/ml.Uji tidak bereaksi silang dengan mikroorganisme lain yang dicobakan. Dari 89 sampel,proporsi positif C.diphtheriae potensial toksigenik dengan uji duplex real-time PCRadalah 21,3%, sedangkan proporsi positif C.diphtheriae toksigenik menggunakan ujibaku emas adalah 11,2%.Kesimpulan: Duplex real time PCR untuk deteksi C.diphtheriae potensial toksigeniktelah dioptimasi dan diaplikasikan pada pasien tersangka difteri. Diharapkan uji inidapat meningkatkan diagnosis laboratorium kasus difteri.

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Background: Diphtheria is toxin-mediated bacterial infection. The most common

etiology is Corynebacterium diphtheriae. Laboratory diagnostic should be done

immediately to support clinical diagnosis. Microscopic examination is not

recommended, culture followed by toxin test is consider gold standard but timeconsuming,

require experience and only done in referral laboratory. PCR is fast,

sensitive and specific. Duplex real-time PCR can detect bacteria and toxin-encoding

gene simultaneously.

Objective: Optimizing duplex real-time PCR assay for detection of potentially

toxigenic C.diphtheriae and applicate the assay on throat swab of suspected diphtheria

patient.

Method: Two pair of primers and specific probe targeting rpoB gene of C.diphtheriae

and A-subunit of diphtheria toxin gene were used in this study. Parameters including

annealling temperature, concentration of primers and probes, inhibitors, cross reaction

and detection limit were being optimized to receive optimal condition. The optimized

assay was applicated on throat swab of suspected diphtheria patient in Sulianti Saroso

Infectious Disease Hospital at 2018-2019. Elek toxigenity test was used for comparison

and clinical data of the patient were analyzed.

Result: The optimum condition for duplex real-time PCR was received upon the

annealing temperature 60oC, concentration of Cd primer 0,4 μM, Tox primer 0,6 μM,

Cd probe 0,5 μM, Tox probe 0,625 μM, DNA elution volume 50 μL, DNA template

volume 5 μL and detection limit 2 CFU/ml. There was no cross reaction found with

other tested microorganisms. Of 89 samples, proportion of potentially toxigenic

C.diphtheriae was 21,3% and proportion of toxigenic C.diphtheriae confirmed by gold

standard was 11,2%.

Conclusion: Duplex real time PCR has been optimized for detection of potentially

toxigenic C.diphtheriae. This method can be used to detect C.diphtheriae and Tox

simultaneosly and increase supporting diagnosis."

"Latar belakang: Infeksi virus Epstein-Barr (EBV) dapat menjadi infeksi oportunistik pada anak dengan HIV. Gejala infeksi EBV sulit dibedakan dengan infeksi HIV dan bersifat laten. Infeksi EBV laten dapat reaktivasi mulai dari gangguan limfoproliferatif hingga terjadinya keganasan. Di Indonesia belum ada data mengenai infeksi EBV pada anak dengan HIV. Tujuan: Mengetahui proporsi, karakteristik dan faktor-faktor yang berhubungan dengan terjadinya infeksi EBV pada anak dengan HIV di RSUPN Dr. Cipto Mangunkusumo Jakarta. Metode: Penelitian potong lintang untuk melihat karakteristik infeksi EBV pada anak dengan HIV dan faktor-faktor yang berhubungan di RSUPN Dr. Cipto Mangunkusumo Jakarta, periode bulan September 2020 hingga Februari 2021. Sampel darah diambil untuk dilakukan pemeriksaan PCR EBV kualitatif (whole blood), darah perifer lengkap, kadar CD4 dan viral load HIV. Hasil: Total subyek 83 anak dengan HIV. Proporsi subyek terinfeksi EBV sebesar 28,9%, dengan rerata usia 9,58 tahun. Limfadenopati merupakan gejala terbanyak, meskipun tidak dapat dibedakan dengan infeksi lain. Dua anak mengalami keganasan akibat EBV yaitu Limfoma Non Hodgkin dan leiomiosarkoma. Sebanyak 75% subyek terinfeksi EBV yang berusia di bawah 12 tahun mengalami anemia (rerata Hb 10,68 ± 2,86 g/dL), dapat disebabkan infeksi EBV atau penyebab lain. Hasil analisis bivariat menunjukkan kadar viral load HIV > 1000 kopi/mL berhubungan dengan terjadinya infeksi EBV pada subyek (OR 2,69 (1,015-7,141); P = 0,043). Simpulan: Proporsi anak dengan HIV yang terinfeksi EBV di RSUPN Dr. Cipto Mangunkusumo Jakarta adalah 28,9%, dengan kadar viral load HIV > 1000 kopi/mL berhubungan dengan terjadinya infeksi EBV pada anak dengan HIV.

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Background: Epstein-Barr virus (EBV) infection can be an opportunistic infection in HIV-infected children. EBV infection is difficult to be differentiated from HIV infection, and it can be latent. Latent EBV infection can reactivate into lymphoproliferative disorders and malignancy. There is no data on EBV infection in HIV-infected children in Indonesia.

Objective: To identify the proportion, manifestations and factors associated with EBV infection in HIV-infected children in Dr. Cipto Mangunkusumo National Central Hospital Jakarta.

Methods: Cross-sectional study to examine the manifestations of EBV infection in HIV-infected children and it’s associated factors in Dr. Cipto Mangunkusumo National Central Hospital Jakarta, during September 2020 to February 2021. Blood samples were taken to examine qualitative EBV PCR (whole blood), complete blood count, CD4 levels and HIV viral load.

Results: Total subjects were 83 HIV-infected children. The proportion of children infected with EBV was 28.9%, with mean age 9.58 years. Lymphadenopathy was the most common symptoms, although it was difficult to differentiate from other infections. Two children have malignancy due to EBV, namely Non-Hodgkin's lymphoma and leiomyosarcoma. Total 75% of EBV-infected subjects under 12 years of age were anemic (mean Hb 10.68 ± 2.86 g/dL), could be due to EBV infection or other causes. Bivariate analysis showed HIV viral load levels > 1000 copies/mL were associated with EBV infection in subjects (OR 2.69 (1.015-7.141); P = 0.043).

Conclusion: The proportion of EBV infection in HIV-infected children in Dr. Cipto Mangunkusumo National Central Hospital Jakarta is 28.9%, with HIV viral load levels > 1000 copies/mL were associated with the EBV infection in HIV-infected children."

"Demam akut yang disertai gejala mirip demam Dengue nyeri kepala, nyeri sendi, ruam, perdarahan perifer seperti mimisan, ptekie adalah gejala yang paling sering dikeluhkan oleh pasien. Gejala-gejala tersebut seringkali diakibatkan infeksi arbovirus yang sangat endemik di Indonesia sebagai negara tropis. Deteksi agen penyebab infeksi tersebut sangat diperlukan untuk penatalaksanaan yang tepat. Studi ini melakukan uji optimasi untuk deteksi molekuler virus penyebab demam mirip demam Dengue, meliputi DENV, ZIKV, WNV, JEV, YFV, CHIKV, dan Hantavirus menggunakan RT PCR. Primer pada studi ini dirancang menggunakan perangkat lunak online Primer-BLAST dari NCBI dan primer-primer tersebut memenuhi kriteria untuk reaksi RT PCR. Kontrol positif pada real time RT-PCR menggunakan DNA sintetik yang dirancang sesuai dengan amplicon target virus. DNA sintetik sepanjang 1.047 pasang basa dirancang untuk digunakan pada virus ZIKV, JEV, YFV, WNV, CHIKV, dan Hantavirus. Suhu penempelan optimum pada primer-primer adalah 600C kecuali primer flavivirus universal yaitu 560C. Limit deteksi primer JEV mencapai 4.355 salinan DNA setiap reaksi real time RT PCR. Tidak terdapat reaksi silang maupun positif palsu pada sampel RNA DENV serotipe 2 maupun pada sampel orang sehat yang digunakan pada studi ini. Sebagai kesimpulan, studi ini menghasilkan primer dan protokol real time RT-PCR yang berpotensi untuk dikembangkan lebih lanjut untuk digunakan dalam uji diagnostik pada sampel pasien demam akut menyerupai gejala demam Dengue.

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Acute fever with Dengue like fever symptoms headache, rash, joint pain, perifer bleeding like ptechie, rinhorrhea is general symptoms that often being complained by patient. Usually the etiology agent of the symptoms are arbovirals which are endemic in Indonesia as a tropical country. In this case, molecular detection is very important to confirm the etiology of disease for prompt and adequate management. This study optimized viral molecular detection as an etiology agent for Dengue like fever symptoms using real time RT PCR. The viruses that were investigated were DENV, ZIKV, WNV, JEV, YFV, CHIKV, and Hantavirus. Primer were designed used Primer BLAST software from NCBI. Those primers fulfilled the good primer requirements and could be used in real time RT PCR reaction. Synthetic DNA with 1.047 base pairs was designed based on amplicon target to be used as control positive for ZIKV, JEV, YFV, WNV, CHIKV, and Hantavirus. The optimal annealing temperature for all primers were at 600C except for flavivirus universal primer was at 560C. The limit of detection of JEV primer was 4355 copies DNA per reaction. Cross reactivity between all primers with DENV serotype 2 RNA and healthy person sample were not found. This study still need RNA viruses as negative or positive control and clinical sample to determine the sensitivity and specificity. As a conclusion, this study provided primers and real time RT PCR protocol that potentially be further developed as diagnostic tools for patient with Dengue like fever symptoms. "

"Tujuan: Mengetahui konsentrasi dari virus Epstein-Barr pada saliva dengan teknik Real-Time PCR pada RS Kramat 128 Jakarta dan korelasinya dengan terapi antiretroviral, Limfosit T CD4 dan viral load HIV. Metode: Penelitian ini merupakan penelitian analitik deskriptif dengan metode potong lintang. Data didapatkan dari pasien HIV yang berkunjung ke RS Kramat 128 pada periode bulan September-Oktober 2019 dengan kelompok kontrol pegawai RS Kramat pada periode tersebut. Seluruh subjek penelitian (77 subjek, 53 HIV dan 24 non-HIV sebagai kelompok kontrol) yang bersedia berpartisipasi diminta untuk mengisi kuesioner, diperiksa rongga mulutnya, serta dikumpulkan salivanya dalam kondisi terstimulasi dan tidak terstimulasi. Saliva yang terkumpul kemudian diekstraksi DNA nya dan dilakukan pemeriksaan real-time PCR dengan menggunakan diagnostik kit untuk EBV pada Pusat Riset Virologi dan Kanker Patobiologi FKUI RSCM. Hasil: Konsentrasi virus Epstein-Barr pada saliva pasien HIV di RS Kramat 128 Jakarta secara statistik lebih tinggi daripada kelompok kontrol dengan median (min-maks) pada pasien HIV 13.950 (0-38.550.000) dan 680 (0-733.000) pada kelompok kontrol. Tipe antiretroviral memiliki korelasi rendah dengan konsentrasi EBV, namun penggunaan ART jangka panjang memiliki korelasi sedang dalam menurunkan konsentrasi EBV (korelasi negatif dengan r=0,295). Kenaikan jumlah EBV saliva pada pasien HIV secara signifikan memiliki korelasi sedang (korelasi positif dengan r=0,295), namun memiliki korelasi rendah dengan jumlah Limfosit T CD4. Kesimpulan: Terdapat perbedaan signifikan antara konsentrasi EBV pada pasien HIV dan kelompok kontrol. Penggunaan ART jangka panjang dan viral load HIV secara signifikan memiliki korelasi sedang dengan konsentrasi EBV pada saliva.

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Objective: To reveal concentration of salivary Epstein-Barr Virus with real-time PCR Technique in Kramat 128 General Hospital HIV patient and its correlation with antiretroviral therapy, CD4 and HIV viral load.

Method: This is an analytic descriptive cross-sectional study on HIV outpatient of Kramat 128 General Hospital in September-Oktober 2019 and employees of Kramat 128 as control group. All subjects (77 subject, with 53 HIV positive respondent and 24 non-HIVcontrol) willing to participate were asked to fill out a questionnare, followed by oral examination and saliva colection in stimulated and unstimulated method. The collected saliva then extracted and EBV concentration were count by real-time PCR using an EBV diagnostic kit at Center for Research on Institute of Human Virology and Cancer Biology Universitas Indonesia.

Result: The concentrations of salivary EBV were significantly higher in HIV patients than non-HIV controls, with median (min-max) values in HIV patient 13.950 (0-38.550.000) and 680 (0-733.000) in non-HIV controls. The type of ART has low correlation with EBV concentrations, but long-term ART has medium correlation in reducing EBV concentrations (negative correlation with r=0,279). Increase amount of EBV in HIV patient were significantly has medium correlation with HIV viral load (positive correlation with r=0,295) but has low correlation with CD4 cell count.

Conclusion: There are significant differences of salivary EBV concentrations in HIV patients and control group. Long term ART and HIV viral load significantly has medium correlation with EBV concentration."