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"Latar Belakang: Mukopolisakaridosis tipe IV A (MPS IV A, Morquio A syndrome) merupakan kelainan autosomal resesif yang disebabkan adanya mutasi pada gen N-acetylgalactosamine-sulfate sulfatase (GALNS atau galactosamine (N-acetyl)-6-sulfate sulfatase; MIM #612222). Diagnosis MPS IV A dapat dicurigai melalui pemeriksaan penapisan GAGs (glukosaminogikans) urin dan ditegakkan dengan pemeriksaan dan aktifitas enzim GALNS pada leukosit atau kultur fibroblast. Pemeriksaan molekuler diperlukan karena bervariasinya gejala klinis yang berhubungan dengan variasi mutasi pada gen GALNS. Namun, lokasi mutasi hot spot berbeda-beda antar daerah dan etnis, Heterogenitas ini dapat menjadi tantangan bagi interpretasi pemeriksaan molekular pasien dengan MPS IV sehingga perlu strategi diagnostik yang efektif biaya untuk menemukan mutasi penyebab kelainan MPS tipe IVA di Indonesia. Tujuan: Mengetahui profil fenotipe-genotipe pasien MPS tipe IV A di Indonesia. Metode: Penelitian ini adalah studi potong lintang yang dilakukan di Departemen Ilmu Kesehatan Anak RSCM-FKUI dan Human Genetic Cluster (HGRC) IMERI FKUI sejak tanggal 1 Januari-13 Desember 2019. 3. Data terkait fenotipe, yaitu anamnesis, pemeriksaan fisis (PF), dan pemeriksaan penunjang, diambil dari data rekam medis. Selanjutnya dilakukan pengambilan darah dalam tabung EDTA sebanyak 5 mL. Tahapan pemeriksaan molekular meliputi isolasi DNA, desain primer, PCR, sequencing, dan analisis varian. Kategori varian baru (novel) yang ditemukan akan dibuat berdasarkan panduan dari American College of Medical Genetics and Genomics (ACMG). Hasil: Total subjek penelitian adalah 7 pasien MPS tipe IV A di Indonesia yang berasal dari 5 kota berbeda. Subjek terdiri dari 2 pasien lelaki dan 5 pasien perempuan. Rentang usia saat pemeriksaan antara 2-17 tahun. Terdapat riwayat keluhan serupa pada kakak subjek 1 dan 6. Pada kelima pasien lain, awitan gejala mukopolisakaridosis disadari pada usia antara 2-3 tahun. Tidak ada riwayat konsanguinitas pada orangtua subjek penelitian. Seluruh subjek pada penelitian ini diklasifikasikan sebagai tipe berat. Manifestasi klinis yang ditemukan pada seluruh subjek penelitian adalah leher pendek, genu valgum, pectus carinatum, sendi yang longgar, serta gangguan pada cara berjalan. Tiga dari 7 subjek saat ini masih dapat berjalan tanpa alat bantu. Data aktivitas enzim dan glikosaminoglikans pada subjek tidak seragam karena tempat pemeriksaan berbeda dan menggunakan metode yang berbeda pula. Sebanyak 24 varian ditemukan pada 7 subjek. Sebagian besar varian ditemukan pada ekson 7 (29,2%), diikuti ekson 5, 10, dan 12 (masing-masing 16,7%), ekson 13 (sebesar 12,5%). Sisanya varian ditemukan pada ekson 1, 11 dan ekson 14 (masing-masing 4,2%). Sebagian besar varian yang ditemukan merupakan varian missense (54,2%), diikuti varian silent (45,8%), dan hanya 1 (4,2%) varian yang ditemukan berupa varian nonsense. Varian tersering yang ditemukan adalah varian c.708C>T yang ditemukan pada 5 subjek, diikuti oleh varian c.510T>C dan c.1354 T>C yang ditemukan pada 3 subjek. Berdasarkan hasil temuan varian terdapat 12 varian benign, 4 VUS, dan 8 varian patogenik. Terdapat 3 varian novel pada subjek penelitian, satu di antaranya adalah varian likely pathogenic, yaitu varian c.1348 G>A. Simpulan: Dari tujuh subjek dalam penelitian ini, ditemukan 8 varian patogenik, di antaranya terdapat 1 varian likely pathogenic baru. Sebanyak 9 dari 14 alel (64,3%) dapat ditemukan varian patogenik, sedangkan 5 varian patogenik lainnya belum ditemukan. Fenotipe paling berat dialami oleh subjek 4 yang memiliki tinggi badan T.

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Background: Mucopolysaccharidosis type IVA (MPS IVA, Morquio A syndrome) is an autosomal recessive disease which is caused by defect in the N-acetylgalactosamine-6-sulfate sulfatase gene (GALNS, galactosamine (N-acetyl)-6-sulfate sulfatase; MIM #612222). After urine glycosaminoglycan is performed as a screening tool, diagnosis is confitmed through measuring GALNS enzyme activity in leucocyte or fibroblast. Molecular testing is needed because clinical symptoms are variable. Mutation in GALNS gene are many and can be different in each ethnicity and country. This heterogeneity poses a challenge to the diagnosis of MPS IVA especially in Indonesia. Therefore, data on clinical spectrum and genetic mutation of MPS IVA in Indonesian population is needed. Aim: To determine the phenotype-genotype correlation of MPS IVA in Indonesia Method: Subjects were recruited from Department of Pediatrics, Cipto Mangunkusumo Hospital, while molecular testing was performed in the Human Genetic Cluster (HGRC) IMERI Faculty of Medicine Universitas Indonesia between Januari 1st until December 13th, 2019. Data on phenotype was evaluated from medical records. A 5 ml EDTA whole blood was then collected from the subjects. Molecular testing consists of DNA isolation, primer design, PCR, sequencing, and variant analysis. Novel variants are then classified according to guidelines from the American College of Medical Genetics and Genomics (ACMG). Results: A total of 7 subjects from 5 different cities was included in this study, consisting of 2 boys and 5 girls. Age at recruitments was between 2 to 17 years-old. Two subjects had history of MPS IVA in older sibling. Age of onset were between 2-3 years-old. No history of consanguinity in the subjects parents. All subjects were classified as severe type. Clinical manifestations found in all patients were short neck, genu valgum, pectus carinatum, loose joint, and difficulty walking. Three out of 7 subjects were still able to walk. Data on enzyme activity and glycosaminoglycans could not be compared because they were performed with different methods. Twenty four variants were found in 7 subjects. Mostly located on exon 7 (29.2%), followed by exon 5, 10, dan 12 (each 16.7%), exon 13 (12.5%), and the rest were found in exon 1, 11, and 14. Missense variants are the most commonly found (54.2%), followed by silent variants (45.8%), and 1 nonsense variant (4,2%). The most common variants found was c.708C>T in 5 subjects, followed by c.510T>C and c.1354 T>C, each on 3 subjects. These variants are classified as benign variants (50%), VUS (1.7%), and pathogenic variants (33.3%). Three novel variants were found in this study, including one likely pathogenic variants, c.1348 G>A. Conclusion: Eight pathogenic variant were found including one novel likely pathogenic variant. Nine out of 14 alleles (64.3%) were found. The most sever phenotype was found in subject 4 who had nonsense homozygous pathogenic variant c.751 C>T."

"Introduksi. Mukopolisakaridosis tipe IVA (MPS IVA; Morquio A) merupakan kelainan genetik ditandai dengan adanya gangguan aktivitas enzim N-acetylgalactosamine-6-sulfatase (GALNS). Defisiensi enzim GALNS menyebabkan kegagalan degradasi KS dan C6S sehingga terakumulasi di urin.Metode. Data varian gen GALNS diperoleh dari Database Human Genetic Research Center IMERI FK UI. Perubahan struktur protein berdasarkan varian yang telah teridentifikasi divisualisasi menggunakan BIOVIA Discovery Studio. Pengukuran aktivitas enzim GALNS menggunakan sampel leukosit dengan metode fluorosensi 4-metilumbelliferone (4-MU). Pengukuran kadar KS dan C6S menggunakan sampel urin dengan metode ELISA kompetitif.Hasil. Varian missense yang teridentifikasi menyebabkan perubahan hilangnya ikatan hidrogen, jembatan disulfida, struktur hidrofobik dan salt bridge, sedangkan varian delesi mereduksi basa nukleotida yang mengakibatkan perubahan pemetaan asam amino Rerata nilai aktivitas spesifik GALNS pada pasien MPS IVA adalah 1,81 nmol/h/mg. Rerata kadar KS dan C6S pasien MPS IVA di Indonesia secara berturut-turut adalah 15,90 ng/mg kreatinin dan 2,14 ng/mg kreatinin.Kesimpulan. Pada keenam pasien MPS IVA varian yang telah teridentifikasi adalah varian missense dan delesi. Kedua tipe varian memengaruhi rendahnya nilai aktivitas spesifik GALNS (1,81 nmol/h/mg) dan meningkatnya kadar GAG urin pada pasien MPS IVA

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.Introduction. Mucopolysaccharidosis type IVA (MPS IVA; Morquio A) is a genetic disorder characterized by impaired activity of enzyme N-acetylgalactosamine-6-sulfatase (GALNS). Impaired activity of enzyme GALNS caused by failure degradation of glycosaminoglycans (GAG) including Keratan Sulfate (KS) and Chondroitin 6-Sulfate (C6S) and it leads to accumulating GAG in urine.

Methods. Data on GALNS gene variants was obtained from Human Genetic Research Center IMERI Universitas Indonesia. Changes in protein structure based on identified variants were visualized using BIOVIA Discovery Studio. Measurement of GALNS enzyme activity using leukocyte samples with the 4-methylumbelliferone (4-MU) fluorescence technique. KS and C6S levels were measured using urine samples using ELISA.

Results. The identified missense variant causes changes interaction of amino acid GALNS including loss of hydrogen bonds, disulfide bridges, hydrophobic structures, and salt bridges, while the deletion variant reduces nucleotide bases which results in changes in amino acid mapping. Mean specific activity of GALNS in MPS IVA patients is 1.81 nmol/h/mg. The mean levels of KS and C6S in MPS IVA patients in Indonesia were 15.90 ng/mgcreatinine and 2.14 ng/mgcreatinine, respectively.

Conclusions. Among six MPS IVA patients, the variants that were identified were missense and deletion variants. Both types of variants affected low value of GALNS specific activity (1.81 nmol/h/mg) and increased urinary GAG levels in MPS IVA patients."

"ABSTRAKMethicillin resistant Staphylococcus aureus (MRSA) merupakan bakteri yang resistenterhadap antibiotik methicillin dan antibiotik golongan β-laktam lainnya. MRSA adalahpatogen umum di rumah sakit dan masyarakat. Isolasi MRSA tidak mudah dilakukankarena seringkali bercampur atau terkontaminasi dengan flora normal seperti coagulasenegative Staphylococci (CoNS) yaitu Staphylococcus epidermidis dan Staphylococcushaemolyticus. Studi ini menggunakan metode fenotipik berupa pengamatan morfologi,pengecatan Gram, Uji biokimia, serta kepekaan antibiotik sedangkan uji genotipik(metode molekular) berupa PCR gen nuc dan mec, SCCmec typing, MLST dansekuensing. Subyek penelitian sebanyak 48 isolat tersimpan di LaboratoriumBakteriologi Molekular, Lembaga Eijkman Jakarta. Diperoleh sebanyak 33 sampel(68.75) memiliki tipe 5 ccr, 9 sampel (18.75) tipe 2 ccr dan 6 sampel (12.5 )nontypeable. Sequence type (ST) yang dominan pada penelitian ini adalah ST239 (2-3-1-1-4-4-3) dan merupakan strain yang multidrug resistant dominan. Pada penelitian inisemua isolat MRSA yang berjumlah 48 isolat telah dikonfirmasi memiliki ciri-cirifenotipik yang sesuai, yaitu Gram positif coccus menyerupai buah anggur, hemolisis,oksidase negatif, katalase positif dan koagulase positif. Sifat bakteri MRSA secaragenotipik mempunyai gen nuc dan gen mecA positif. Hubungan antara sifat genotipedan sifat fenotipe MRSA yang terlihat dalam penelitian ini adalah semua isolat MRSAyang multidrug resistant (uji secara fenotipik) juga merupakan sequence type yangdominan di rumah sakit (uji genotipik).

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ABSTRACTMethicillin resistant Staphylococcus aureus (MRSA) is a bacterium that is resistant tothe methicillin antibiotics and other β-lactam group antibiotics. MRSA is a commonpathogen in hospitals and communities. Isolation of MRSA is not easy to do because itis often mixed or contaminated with normal flora such as coagulase negativeStaphylococci (CoNS), namely Staphylococcus epidermidis and Staphylococcushaemolyticus. This study used phenotypic methods in the form of morphologicalobservations, Gram staining, biochemical tests, and antibiotic sensitivity whilegenotypic tests (molecular methods) in the form of nuc and mec PCR, SCCmec typing,MLST and sequencing. The research subjects were 48 isolates stored in the MolecularBacteriology Laboratory, Eijkman Institute Jakarta. Thirty three samples (68.75) hadtype 5 ccr, 9 samples (18.75) type 2 ccr and 6 samples (12.5) nontypeable. Thedominant sequence type (ST) in this study is ST239 (2-3-1-1-4-4-3) and is a multidrugresistant dominant strain. In this study, all isolates of MRSA, total of 48 isolates, wereconfirmed to have appropriate phenotypic features, which are Gram positive cocciresembling grapes,-hemolysis, negative oxidase, positive catalase and positivecoagulase. Genotypically all isolates have positive nuc gene and mecA gene. Therelationship between genotype features and MRSA phenotype seen in this study is thatMRSA isolates that are multidrug resistant (phenotypic test) are also the dominantsequence types in the hospital (genotypic test)."

"Rare-actinomycetes tersebar di berbagai habitat terutama di habitat ekstrem seperti kawasan geotermal. Penelitian mengenai rare-actinomycetes dilakukan terkait potensinya sebagai penghasil senyawa bioaktif baru yang bermanfaat dalam bidang kesehatan, industri dan farmasi. Tujuan dari penelitian ini adalah mengisolasi dan mengkarakterisasi secara fenotip dan genotip rare-actinomycetes dari sampel tanah di bawah batuan kuarsa (S3.5.3) di hutan kawasan geotermal Cisolok. Metode pengayaan sampel tanah dilakukan menggunakan medium 10% R2A (Reasoner’s 2A) cair dengan penambahan cycloheximide 100 ppm dan sodium azide 60 ppm, kemudian diinkubasi pada suhu 30°C selama 30 hari. Isolasi rare actinomycetes dilakukan dengan metode membran filter dan spread pada medium 10% R2A gellan gum yang diinkubasi pada suhu 45°C. Karakterisasi dilakukan secara genotipik (berdasarkan data sequence gen 16S rRNA, analisis homologi sequence, dan rekonstruksi pohon filogenetik dengan metode Neighbour-Joining, Maximum Likelihood, dan Minimum Evolution); dan karakterisasi fenotipik (morfologi, fisiologi, dan biokimia). Sebanyak 26 isolat diperoleh dari sampel S3.5.3. Lima isolat dengan karakter morfologi actinomycetes yang diisolasi dari suhu 45°C dengan membran filter, dipilih untuk diidentifikasi. Hasil analisis sequence gen 16S rRNA dari lima isolat menunjukkan persentase homologi sebesar 95,46-99,56% terhadap Micromonospora yasonensis DS3186T. Berdasarkan hasil analisis filogenetik dengan metode Neighbour-Joining, kelima isolat memiliki hubungan kekerabatan terdekat dengan Micromonospora yasonensis DS3186T. Kelima isolat merupakan anggota class Actinomycetes, ordo Micromonosporales, family Micromonosporaceae. Karakter fenotipik kelima isolat sesuai dengan Micromonospora yasonensis sebagai spesies terdekatnya. Kelima isolat merupakan bakteri termotoleran (tumbuh pada suhu 30-45°C dan suhu optimum 40°C), aerobik, Gram positif, menghasilkan miselium substrat tanpa adanya miselium aerial, positif katalase, dan menghasilkan soluble pigment. Penelitian ini mengungkapkan bahwa Micromonospora yasonensis dapat ditemukan di kawasan geotermal dan berasosiasi dengan batuan kuarsa.

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Rare-actinomycetes are distributed in various habitats, particularly in extreme environments such as geothermal areas. Research on rare-actinomycetes focuses on their potential as producers of new bioactive compounds beneficial in health, industrial, and pharmaceutical fields. The aims of this study were to isolate and characterize rare-actinomycetes from soil samples beneath quartz rocks (S3.5.3) in the geothermal forest of Cisolok. Soil sample enrichment was performed using 10% R2A (Reasoner’s 2A) liquid medium supplemented with 100 ppm cycloheximide and 60 ppm sodium azide, incubated at 30°C for 30 days. Rare actinomycetes isolation was carried out using the membrane filter method and spread on 10% R2A agar with gellan gum, incubated at 45°C. Characterization included genotypic analysis based on the sequence of 16S rRNA gene, supported by phenotypic characterization (morphology, physiology, and biochemistry). A total of 26 isolates were obtained from sample S3.5.3. Five isolates with actinomycetes morphology isolated at 45°C using the membrane filter method were selected for characterization. Analysis of the 16S rRNA gene sequences from these five isolates showed homology levels of 95.46-99.56% to Micromonospora yasonensis DS3186T. Based on phylogenetic analysis using the Neighbour-Joining method, the five isolates were most closely related to Micromonospora yasonensis DS3186T. These isolates belong to the class Actinomycetes, order Micromonosporales, and family Micromonosporaceae. Phenotypic characteristics of the five isolates were consistent with Micromonospora yasonensis as their closest species. These isolates are thermotolerant bacteria (growing at temperatures of 30-45°C; optimum temperature 40°C), aerobic, Gram-positive, produce substrate mycelium without aerial mycelium, positive for catalase, and produce soluble pigment. This study reveals that Micromonospora yasonensis can be found in geothermal areas associated with quartz rocks."

"ABSTRAKData genotype merupakan komponen yang penting pada bidang bioinformatik,namun data genotype sulit diperoleh. Selain sulitnya proses yang harus dilakukanuntuk memperoleh data genotype, proses tersebut juga hanya dapat dilakukan olehpara ahli yang memiliki kemampuan mengesktrak data genotype. Oleh karena haltersebut maka pada penelitian ini akan dibangun sebuah sistem yang dapatmembentuk rule phenotype-genotype yang digunakan untuk memprediksigenotype berdasarkan rule tersebut. Pembentukan rule akan menggunakan metodeassociation rules mining (ARM). Salah satu algoritma yang menggunakan dasarmetode ARM adalah algoritma classification predictive base on association rule(CPAR). Algoritma CPAR menjadi acuan untuk membentuk rule genotypephenotypepada riset ini. Proses prediksi dilakukan dengan menghitung nilaikemiripan antara phenotype pada rule dengan input phenotype yang akandiprediksi genotype-nya. Phenotype dengan nilai batas kemiripan < 0.05 akanmerujuk pada rule genotype dan menjadi hasil prediksi. Evaluasi dilakukandengan menghitung akurasi berdasarkan ground truth. Hasil prediksi mencapaiakurasi 48% dengan standar deviasi sebesar 30%.

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ABSTRACTGenotype data is an important component in bioinformatics research;unfortunately it is hard to get the data. An expert is also needed to extract the genesequence, so that the purpose of this research is to develop a system that can builda genotype-phenotype rules that can predict the gene. This research useassociation rules mining (ARM) to build the rules. Classification predictive baseon association rules (CPAR) is one of ARM algorithm. In this research, CPAR isan algorithm to build the genotype-phenotype rule that can be used to predict thegenotype data. The predictive process is based on the similarity betweenphenotype on the input and the rule. The input phenotype with similarity threshold< 0.05 will refer to the genotype rule. The result will be evaluated by the accuracybased on the ground truth. The average of the accuracy is 48% with standarddeviation is around 30%."

"Mukopolisakaridosis tipe II (MPS II) merupakan penyakit kelainan lisosomal langka yang disebabkan oleh mutasi pada gen iduronat 2-sulfatase (IDS) dapat menyebabkan disfungsi dari enzim I2S yang dihasilkan sehingga molekul heparan sulfat (HS) dan dermatan sulfat (DS) terakumulasi pada jaringan. Penelitian ini dilakukan untuk mengetahui dan menganalisis hubungan kadar HS dan DS urin dengan jenis mutasi gen IDS pada penderita MPS II di Indonesia. Data susunan nukleotida gen IDS dari tujuh pasien MPS II dianalisis untuk melihat jenis mutasi dan dibuat model 3D proteinnya. Analisis 3D protein akan dikorelasikan dengan kadar HS dan DS urin pasien tersebut yang diukur menggunakan metode Enzyme-Linked Immunosorbent Assay (ELISA). Hasil analisis mutasi ditemukan beberapa jenis mutasi, seperti mutasi nonsense (1/7), delesi (2/7), insersi (1/7), dan missense (3/7). Dari ketujuh pasien tersebut, tiga diantaranya (P2, P6, P7) telah menjalani terapi ERT. Kadar HS urin dari ketujuh pasien menunjukkan peningkatan yang beragam dibandingkan dengan kadar HS normal. Berbeda dengan HS, kadar DS urin sampel pasien ada yang mengalami sedikit peningkatan (P1, P2, P7) dan ada pula yang tetap berada pada rentang kadar DS normal (P3, P4, P5, P6). Keragaman kadar HS dan DS sampel pasien tersebut sangat dipengaruhi oleh letak mutasi, jenis mutasi, diagnosis dan prognosis yang ditegakkan sedini mungkin, terapi ERT yang telah dilakukan pasie, durasi ERT, dan respon masing-masing pasien terhadap pengobatan yang telah diberikan.

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Mucopolysaccharidosis type II (MPS II) is a rare lysosomal disorder caused by mutations in the iduronat 2-sulfatase (IDS) gene that can cause dysfunction of I2S enzyme so that the heparan sulfate (HS) and dermatan sulfate (DS) molecules accumulate in the tissue. This study was conducted to determine and analyze the relationship of urinary HS and DS levels with the type of IDS gene mutation in MPS II patients in Indonesia. The nucleotide of IDS genes sequences from seven MPS II patients were analyzed to see the type of mutation and the 3D protein model was made. 3D protein analysis will be correlated with urinary HS and DS levels of the patients measured by using the Enzyme-Linked Immunosorbent Assay (ELISA) method. Results of mutation analysis results found several types of mutations, such as nonsense mutations (1/7), deletions (2/7), insertions (1/7), and missense (3/7). From the seven patients, three of them (P2, P6, P7) had undergone ERT therapy. The urine HS level of the seven patients showed a varied increase compared to normal HS levels. In contrast to HS, the urine DS level of the sample of patients had a slight increase (P1, P2, P7) and some remained in the normal DS level range (P3, P4, P5, P6). The diversity of HS and DS levels of the patient's samples is strongly influenced by the location of the mutation, type of mutation, diagnosis and prognosis that is enforced as early as possible, ERT therapy has been carried out, ERT duration, and each patient's response to the treatment given."

"Latar belakang: Mukopolisakaridosis (MPS) tipe II adalah kelainan genetik yang ditandai dengan gangguan metabolik berupa defisiensi enzim iduronat-2-sulfatase (I2S) karena adanya mutasi pada gen iduronat-2-sulfatase (IDS), sehingga heparan sulfat dan dermatan sulfat tidak terdegradasi dan terakumulasi pada jaringan. Penelitian ini dilakukan untuk menganalisis aktivitas spesifik enzim I2S dan kaitannya dengan varian mutasi gen IDS pada pasien MPS II di Indonesia. Metode: Data sekuen nukleotida gen IDS dari enam pasien MPS II dianalisis untuk melihat jenis mutasi serta dibuat model konformasi proteinnya. Sel peripheral blood mononuclear cell (PBMC) diisolasi menggunakan metode sentrifugasi bertingkat dan dikultur menggunakan medium RPMI. Nilai aktivitas spesifik I2S diperoleh dengan mengukur aktivitas I2S per miligram konsentrasi total protein. Aktivitas enzim I2S diukur menggunakan metode fluorometri, sementara konsentrasi total protein diukur menggunakan bicinchoninic acid (BCA) protein assay. Hasil: Tiga varian mutasi yang ditemukan pada pasien MPS tipe II adalah missense (3/6), delesi (2/6), dan nonsense (1/6). Aktivitas enzim spesifik I2S pada pasien menunjukkan angka yang bervariasi. Mutasi dengan rata-rata aktivitas spesifik I2S paling rendah sampai paling tinggi secara berurutan adalah mutasi delesi (0,026 nmol/min/mg), missense (0,052 nmol/min/mg), dan nonsense (0,052 nmol/min/mg). Kesimpulan: Aktivitas spesifik enzim I2S pada pasien MPS tipe II di Indonesia 0,044 nmol/min/mg, sedangkan pada kontrol adalah 0,172 nmol/min/mg. Nilai rata-rata aktivitas spesifik I2S pada pasien menurun empat kali lipat dibandingkan pada kontrol.

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Background: Mucopolysaccharidosis type II (MPS II) is an inherited metabolic disorder that caused by iduronate-2-sulfatase (I2S) enzyme deficiency due to mutations in the iduronate-2-sulfatase (IDS) gene, so that heparan sulfate and dermatan sulfate do not be degrade and accumulate in tissues. This study was conducted to analyze the specific activity of I2S and its relationship to IDS variant mutation of MPS II patients in Indonesia.

Method: IDS gene nucleotide sequences from six MPS II patients were analyzed to see mutation type and protein conformation model was made. Peripheral blood mononuclear cell (PBMC) were isolated using a stratified centrifugation and cultured using the RPMI medium. I2S specific activity values are obtained by measuring I2S activity per milligram of total protein concentration. I2S enzyme activity was measured using fluorometry method, while total protein concentration was measured using bicinchoninic acid (BCA) protein assay.

Result: There were three variant mutation in MPS II patients, such as missense (3/6), deletion (2/6), and nonsense (1/6). I2S specific enzyme activity shows varying numbers. IDS mutation based on I2S specific activity from lowest to highest mean value are deletion (0.026 nmol/min/mg), missense (0.052 nmol/min/mg), and nonsense (0.052 nmol/min/mg).

Conclusion: I2S specific activity in MPS II patient is 0,044 nmol/min/mg, while the control is 0,172 nmol/min/mg based on the mean value. I2S specific activity in patients decreased four times compared to controls."

"Mukopolisakaridosis II MPS II, atau sindrom Hunter, adalah penyakit resesif terpaut-X langka yang disebabkan oleh gangguan penyimpanan lisosomal akibat defisiensi enzim lisosomal iduronate-2-sulfatase IDS. Gen IDS penting dalam proses degradasi lisosomal dermatan sulfat dan heparan sulfat, karena defisiensi gen IDS akan mengarah pada akumulasi Glikosaminoglikans GAG tersebut. Analisis spesifik ekson pada ekson 6 dan ekson 8 gen IDS dilakukan pada penderita MPS II di Rumah Sakit Umum Pusat Nasional Dr. Cipto Mangunkusumo, Jakarta, Indonesia. Dalam penelitian ini, sampel dari penderita MPS II dan sampel normal dianalisis menggunakan PCR dan metode sekuensing. Hasil yang diperoleh menunjukkan mutasi-mutasi novel pada dua penderita MPS II di Indonesia. Satu mutasi insersi sepanjang 14 pb c.792_793insCCCCTGTGGCCTAC ditemukan pada ekson 6 dari gen IDS pada satu penderita. Mutasi tersebut menyebabkan perubahan asam amino di p.Asn265ProfsTer20. Variasi delesi satu basa nukleotida c.1023delA yang menyebabkan perubahan susunan asam amino dengan notasi p.Glu341AspfsTer19 ditemukan pada seluruh sampel. Selain itu, ditemukan pula mutasi missense novel c.1033T>C yang mengubah asam amino triptofan menjadi arginin p.Trp345Arg, dan variasi delesi satu basa nukleotida c.1041delA yang menyebabkan perubahan susunan asam amino dengan notasi p. Lys347AsnfsTer13 pada ekson 8 dari gen IDS pada satu penderita lain.

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Mucopolysaccharidosis II MPS II, or Hunter syndrome, is a rare X linked recessive disease caused by lysosomal storage disorder due to the deficiency of the lysosomal enzyme iduronate 2 sulfatase IDS . The IDS gene is important for the lysosomal degradation process of dermatan sulfate and heparan sulfate, as the deficiency of the IDS gene will lead to the accumulation of these Glycosaminoglycans GAGs. Exon specific analyses on exon 6 and exon 8 of the IDS gene were done on patients with MPS II in Cipto Mangunkusumo National Referral Hospital, Jakarta, Indonesia. In this study, samples from MPS II patients and normal samples were analyzed using PCR and sequencing methods.

The results show novel mutations in Indonesian patients with MPS II. A 14 bp long insertion mutation c.792 793insCCCCTGTGGCCTAC were found on exon 6 of the IDS gene in one patient. This mutation leads to the alteration of amino acids at p.Asn265ProfsTer20. A single nucleotide deletion variant c.1023delA which leads to the alteration of amino acids in the p.Glu341AspfsTer19 was found in all patients. Other than that, a novel missense mutation c.1033T C which leads to the alteration of amino acid from tryptophan to arginine p.Trp345Arg and a single nucleotide deletion variant c.1041delA which leads to the alteration of amino acids at p.Lys347AsnfsTer13 on exon 8 of the IDS gene were found in another patient."

"Mukopolisakaridosis (MPS) tipe VI adalah kelainan genetik langka berupa defisiensi enzim arylsulfatase B (ARSB) akibat kemunculan varian pathogenic gen ARSB. Gejala MPS tipe VI meliputi kornea berkabut, fitur wajah kasar, abnormalitas tulang dan persendian, serta kelainan saluran pernapasan dan pembengkakan organ. Diagnosis MPS tipe VI dilakukan dengan pengukuran aktivitas enzim serta konfirmasi melalui analisis varian gen ARSB. Varian gen pathogenic yang paling umum dilaporkan pada pasien MPS tipe VI terletak di rentang ekson 5—8, yaitu c.962T>C (p.Leu321Pro), c.1197C>G (p.Phe399Leu), dan beberapa varian pada asam amino Arg315 di ekson 5. Analisis varian gen ARSB belum pernah dilakukan di Indonesia walapun sudah ada pelaporan kasus MPS tipe VI. Tujuan dari analisis varian gen ARSB di Indonesia adalah mengidentifikasi dan mengklasifikasi varian gen ARSB serta mendapatkan profil genetik gen ARSB pada pasien MPS tipe VI di Indonesia. Analisis varian gen ARSB dilakukan pada dua pasien MPS tipe VI dan sepuluh individu normal sebagai kelompok kontrol menggunakan sekuensing Sanger. Penelitian tidak menemukan adanya varian pathogenic pada gen ARSB ekson 5—8 pasien MPS tipe VI. Penelitian berhasil menemukan varian benign c.1072G>A (p.Val358Met) pada ekson 5, c.1142+233C>T dan c.1143-27A>C pada intron 5, dan c.1337-32C>G pada intron 7 serta satu varian likely benign c.1213+149C>G pada intron 6 yang sudah pernah dilaporkan sebelumnya. Ditemukan juga satu varian novel c.1142+213C>T pada intron 5 dengan klasifikasi variant of uncertain significance. Penambahan individu dalam kelompok kontrol disarankan agar frekuensi alel dalam populasi lebih tercerminkan dengan baik.

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Mucopolysaccharidosis (MPS) type VI is a rare genetic disorder due to arylsulfatase B (ARSB) enzyme deficiency caused by the presence of pathogenic variant in ARSB ene. Clinical symptoms of MPS type VI are corneal clouding, coarse facial features, joint and skeletal abnormalities, respiratory problems, and enlarged organs. Diagnosis of MPS type VI is done by evaluating ARSB enzyme activity and is confirmed by ARSB gene analysis. The most commonly reported pathogenic variants in MPS type VI patients are located in exon 5—8, such as c.962T>C (p.Leu321Pro), c.1197C>G (p.Phe399Leu), dan numerous variants involving Arg315 at exon 5. Analysis of ARSB gene has not been done in Indonesia although one MPS type VI case has been reported. Analysis of ARSB gene in Indonesia is done to identify and classify the ARSB gene variants and to also obtain genetic profile of MPS type VI patients in Indonesia. The analysis is done to two MPS type VI patients along with ten normal individuals as control group using Sanger sequencing. This study has found no pathogenic variants in exon 5—8 of ARSB gene. This study have identified benign variants c.1072G>A (p.Val358Met) at exon 5, c.1142+233C>T and c.1143-27A>C at intron 5, and c.1337-32C>G at intron 7 along with one likely benign variant c.1213+149C>G at intron 6 which have been previously reported before. One novel variant c.1142+213C>T at intron 5 was also found and classified as variant of uncertain significance. A larger control group is advised to better reflect the allele frequency in population."

"Mukopolisakaridosis tipe VI (MPS tipe VI) merupakan gangguan metabolisme yang diakibatkan defisiensi aktivitas enzim pengolah glycosaminoglycan (GAG) jenis dermatan sulfat, yaitu arylsulfatase B (ARSB). Prevalensi MPS tipe VI di dunia tercatat dalam rentang 0,03—7,85 per 100.000 kelahiran. Gejala MPS tipe VI meliputi coarse facies, dysostosis multiplex, gangguan pendengaran, pernapasan, dan penglihatan, serta penebalan katup jantung, tetapi tidak disertai kelainan sistem saraf pusat. Varian pathogenic gen Arylsulfatase B (ARSB) pada ekson 1—4 telah dilaporkan sebagai pemicu manifestasi MPS tipe VI pada pasien dari berbagai belahan dunia, salah satunya dari Thailand. Laporan varian gen ARSB pada ekson 1—4 pasien MPS tipe VI di Indonesia belum ditemukan sehingga penelitian ini bertujuan mengidentifikasi dan mengklasifikasikan tingkat patogenisitas varian gen ARSB pada ekson 1—4 pasien MPS tipe VI di Indonesia. Gen ARSB dua pasien MPS tipe VI dan 10 individu normal diamplifikasi menggunakan polymerase chain reaction (PCR) dan disekuensing menggunakan metode Sanger. Patogenisitas varian gen ARSB yang teridentifikasi diklasifikasikan menurut panduan yang diterbitkan American College of Medical Genetics (ACMG). Identifikasi varian gen ARSB pada ekson 1—4 pasien MPS tipe VI di Indonesia berhasil dilakukan dengan temuan sejumlah delapan varian. Satu varian novel berhasil diklasifikasikan sebagai varian likely pathogenic, yaitu c.235_236delinsCC (p.Gly79Pro) yang ditemukan pada ekson 1 kedua pasien MPS tipe VI. Enam varian reported yang ditemukan pada intron 1 individu-individu normal berhasil diklasifikasikan sebagai varian likely benign, yaitu c.312+167G>A, c.312+229C>A, c.312+304C>T, c.313-81G>A, c.313-77G>A, dan c.313-26T>C. Satu varian reported yang ditemukan pada ekson 1 dua individu normal diklasifikasikan sebagai variant of uncertain significance (VUS), yaitu c.181G>A (p.Gly61Ser). Penelitian lebih lanjut yang melibatkan lebih banyak individu normal diperlukan untuk memperoleh data frekuensi alel kedelapan gen ARSB tersebut dalam populasi normal di Indonesia sehingga spesifisitas klasifikasi varian dapat meningkat menjadi varian pathogenic atau benign.

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Mucopolysaccharide type VI (MPS tipe VI) is a metabolic disorder caused by deficient activity of arylsulfatase B (ARSB) enzyme, which processes a type of glycosaminoglycan (GAG) known as dermatan sulfate. Worldwide prevalence of MPS tipe VI ranges from 0.03—7.85 per 100,000 live births. Symptoms of MPS tipe VI include coarse facies, dysostosis multiplex, eyes, lungs, and ears disorders, as well as valvular stenosis, but without central nervous system abnormalities. Pathogenic variants of Arylsulfatase B (ARSB) gene in exons 1—4 has been reported to cause MPS tipe VI manifestation in patients from multiple countries, including Thailand. No report of ARSB gene variants in exons 1—4 of Indonesian MPS tipe VI patients have been found. This study aims to identify and classify the pathogenicity of ARSB gene variants in exons 1—4 of Indonesian MPS tipe VI patients. ARSB gene of two patients and 10 healthy individuals were amplified using polymerase chain reaction (PCR) and Sanger sequenced. Pathogenicity of identified ARSB gene variants were classified according to the American College of Medical Genetics (ACMG) guidelines. Identification of ARSB gene variants in exons 1—4 of MPS type VI patients in Indonesia was successfully carried out with a finding of eight gene variants. A novel variant found in exon 1 of both MPS type VI patients was classified as a likely pathogenic, notated as c.235_236delinsCC (p.Gly79Pro). Six reported variants found in intron 1 of healthy individuals were classified as likely benign, each notated as c.312+167G>A, c.312+229C>A, c.312+304C>T, c.313-81G> A, c.313-77G>A, and c.313-26T>C. One reported variant found in exon 1 of two healthy individuals was classified as a variant of uncertain significance (VUS), notated as c.181G>A (p.Gly61Ser). Further research involving more healthy individuals is required to obtain frequency allele data of the eight ARSB gene variants in the Indonesian normal population, which supports the increase of each variant’s classification specificity into pathogenic or benign."