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"DNA forensik adalah teknik identifikasi individu berdasarkan profil DNA STR. Bercak darah merupakan barang bukti utama dalam kejadian kriminal. Paparan lingkungan diketahui merusak DNA, oleh karena itu penelitian ini menganalisis pengaruh lingkungan terhadap marka STR untuk kepentingan forensik. Bercak darah dari 12 individu diletakkan pada kain katun steril dikelompokkan menjadi 3 kelompok perlakuan lingkungan (dalam, luar dan tanah); 4 kelompok perlakuan lama paparan (1, 7, 14, dan 28 hari). DNA diekstraksi, dikuantifikasi menggunakan PCR real time kuantitatif, dilanjutkan amplifikasi 24 lokus STR. Stabilitas DNA diukur berdasarkan tingkat keberhasilan amplifikasi alel pada 24 lokus STR. Hasil penelitian menunjukkan hubungan bermakna (P<0.05) antara stabilitas DNA dengan pengaruh lingkungan, yang ditemukan pada lama paparan hari ke-7, 14 dan 28. Paparan lingkungan dalam tanah menunjukkan pengaruh kuat terhadap stabilitas DNA. Hasil korelasi Gamma menunjukkan paparan lingkungan memiliki tingkat korelasi yang tinggi terhadap stabilitas DNA, yaitu semakin ekstrim lingkungan dan semakin lama paparan lingkungan tersebut, stabilitas DNA semakin menurun ditunjukkan dengan terjadinya beberapa loss of heterozigosity dan allelic drop-out. Analisis pada tiap lokus menunjukkan semakin panjang fragmen alel pada lokus semakin mudah alel tersebut terdegradasi. Pada penelitian ini lokus CSF1PO (<350 pb), SE33 (360 pb) dan TPOX (250 pb) memiliki kecenderungan allelic drop-out paling tinggi.

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Forensic DNA is an identification technique based on STR DNA profile. Blood stain is the main evidence in criminal act. Recent researches showed that environment have role to DNA degradation, therefore this research was designed to show environmental effect on DNA STR for forensic application of blood stain. Blood from 12 people dropped on sterile cottons and divided on 3 groups environmental (room, outdoor, soil). Each group has 4 sub-treatments time of environmental exposure (1, 7, 14, 28 days). DNA extracted from treated blood stain, quantified with real time qPCR, then 24 STR locus amplified on multiplex PCR. DNA stability measured based on amplification success rate of 24 STR locus. The results show that there were significant association (P<0.05) between stability DNA and environment effect. DNA on soil has the lowest stability among three environment condition. Significant differences were shown more significant at 28 days. Based on Gamma correlation analysis, the more extreme an environment and the longer DNA being exposed, the more decrease stability of DNA. It was shown by LOH and allelic drop-out. The longer allele, the more increase the rate of DNA degradation. In this research CSF1PO (<350bp), SE33 (<360bp), and TPOX (<250bp) have the lowest stability."

"Infeksi malaria saat kehamilan telah dilaporkan berasosiasi dengan peningkatan risiko lahirnya bayi berat badan lahir rendah (BBLR, berat badan lahir < 2500 g) di negara-negara endemis malaria, termasuk di Timika, Papua, Indonesia. Infeksi Plasmodium falciparum (P. falciparum) mengakibatkan penumpukkan eritrosit terinfeksi di plasenta, sehingga berkontribusi pada gangguan fungsi plasenta dan terhambatnya petumbuhan janin. Penelitian bertujuan untuk mengetahui pola ekspresi transkrip mRNA yang terlibat dalam aksis insulin-like growth factor (IGF) dan persinyalan leptin yang berperan dalam mengatur fungsi plasenta selama kehamilan. Plasenta ibu hamil terinfeksi malaria falciparum yang melahirkan bayi tunggal sebanyak 55 sampel digunakan untuk isolasi RNA total. RNA hasil isolasi kemudian ditranskripsi balik menjadi complementary DNA (cDNA) dengan reverse transcription polymerase chain reaction (RT PCR), kemudian diukur ekspresinya dengan quantitative real-time PCR (qPCR). Ekspresi IGF-I (r = 0,232, p = 0,089) dan reseptor leptin isoform pendek (OBRa) (r = 0,215, p = 0,115) pada plasenta cenderung berkorelasi positif terhadap skor-z berat badan lahir. Ekspresi OBRa juga berkorelasi negatif secara terhadap umur kehamilan ( = -0,294, p = 0,029). Sedangkan, ekspresi IGFBP-1 cenderung berkorelasi negatif terhadap berat plasenta (r = -0,237, p = 0,081). Ekspresi leptin dan reseptor leptin isoform panjang (OBRb) plasenta menunjukkan korelasi yang lemah terhadap berat badan lahir, skor-z berat badan lahir, berat plasenta, maupun umur kehamilan. Faktor berat plasenta dan ekspresi OBRa menunjukkan kontribusi yang nyata terhadap skor-z berat badan lahir dibandingkan variabel lainnya. Keterkaitan antara ekspresi komponen aksis IGF-I dan persinyalan leptin pada plasenta dari kehamilan terinfeksi malaria menunjukkan respon plasenta terhadap kondisi intrauterin yang merugikan akibat infeksi malaria.

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Malaria infection during pregnancy has been reported to be associated with an increased risk for delivering low birth weight (LBW, birth weight < 2500 g) infants in malaria-endemic area, including in Timika, Papua, Indonesia. Plasmodium falciparum (P. falciparum) infection leads to placental sequestration of infected erythrocytes, causing impaired placental function and altered fetal growth. This study was aimed to investigate the expression pattern of mRNA transcripts involved in insulin-like growth factor (IGF) axis and leptin signaling which play a role in modulating placental function during pregnancy. A total of 55 placenta samples collected from falciparum malaria-infected mothers who delivered singleton infant were employed for total RNA isolation. The isolated RNA was reverse transcribed into complementary DNA (cDNA) using reverse transcription polymerase chain reaction (RT PCR), followed by measurement of gene expression using quantitative real-time PCR (qPCR). Placental expressions of IGF-I (r = 0,232, p = 0,089) and long isoform of leptin receptor (OBRa) (r = 0,215, p = 0,115) were tend to be positively correlated with birth weight z-score. The expression of OBRa was also negatively correlated with gestational age ( = - 0,294, p = 0,029). Meanwhile, the expression of IGFBP-1 showed a tendency to be negatively correlated with placental weight (r = -0,237, p = 0,081). Placental leptin and long isoform of leptin receptor (OBRb) expressions showed weak correlations with birth weight, placental weight, and gestational age. Placental weight and OBRa expression represent a significant contribution to determination of birth weight z-score as compared to the others variables. Correlation between placental expression of IGF axis and leptin signaling in malaria-infected pregnancies might reflect placental response to adverse intrauterine condition due to malaria infection."

"ABSTRAKLatar belakang: Zoonosis malaria telah menjadi perhatian komunitas kesehatan dunia setelah adanya laporan kasus di Sarawak pada tahun 2004. Penyakit ini disebabkan oleh parasit malaria satwa primata Plasmodium knowlesi dengan inang alami Macaca fascicularis dan M. nemestrina. Baku emas diagnosis parasit malaria masih berdasarkan pada identifikasi mikroskopik. Selain membutuhkan keahlian yang tinggi, teknik ini terkadang sulit menentukan spesies parasit bila terjadi infeksi campuran dan parasitemia yang sangat rendah. Belakangan diusulkan DNA barcoding, suatu metode identifikasi menggunakan penanda gen sitokrom c oksidase subunit I COI DNA mitokondria untuk spesiasi. Penelitian yang dilakukan bertujuan untuk mengembangkan metode identifikasi spesies parasit menggunakan gen COI sebagai penanda molekul dan mengungkap dasar molekul transmisi zoonosis parasit malaria dengan mempelajari peran gen penyandi protein DARC Duffy Antigen Receptor for Chemokines dan DBP Duffy Binding Protein yang berhubungan dengan invasi sel darah merah.Metode: Verifikasi potensi barcode COI sebagai penanda identifikasi spesies parasit malaria dilakukan dengan studi in-silico, sedangkan validasi penggunaan barcode COI dilakukan dengan analisis sensitivitas dan spesifisitas. Teknologi molekuler PCR-Sequencing dilakukan untuk mengaplikasikan barcode COI pada penapisan parasit malaria di populasi manusia dan satwa primata, serta identifikasi variasi genetik gen penyandi protein DARC dan DBP terutama pada daerah pengikatan ligan parasit dan reseptor inang.Hasil: Studi in-silico menunjukkan bahwa DNA barcoding berpotensi sebagai penanda identifikasi parasit malaria. Primer yang dirancang mengamplifikasi daerah COI sepanjang 670 pb berhasil mengidentifikasi parasit malaria dengan sensitivitas 1 ndash; 3 parasit/ l. Pada penapisan parasit malaria di populasi manusia di Kalimantan Tengah ditemukan 3,34 78/2309 kasus malaria, di mana dua diantaranya adalah kasus malaria knowlesi, yang secara statistik berbeda bermakna bila dibandingkan dengan mikroskopik 2,82 dan 18S rRNA 1,82 . Pada daerah yang sama, penapisan parasit malaria di populasi satwa primata, ditemukan 52,01 168/323 sampel orangutan dan 23,25 10/43 sampel monyet Macaca positif malaria. Spesies parasit yang ditemukan pada orangutan adalah P. species tipe parasit ovale, P. species tipe vivax-cynomolgi, P. species tipe vivax-hylobati dan P. species tipe malariae-inui, sedangkan pada monyet Macaca meliputi P. knowlesi, P. coatneyi, P. inui, juga P. spesies tipe malariae-inui, spesies parasit yang sama ditemukan di orangutan. Studi ini juga menemukan keanekaragaman genetik pada gen penyandi protein Duffy Antigen Receptor for Chemokines manusia maupun satwa primata dan Duffy Binding Protein parasit malaria yang memainkan peran penting dalam invasi parasit malaria.Kesimpulan: Barcode COI dapat secara spesifik dan sensitif mengidentifikasi spesies parasit malaria dan dapat diaplikasikan sebagai alat identifikasi zoonosis malaria. Terdapat variasi genetik gen penyandi protein Duffy Antigen Receptor for Chemokines dan Duffy Binding Protein yang berhubungan dengan invasi sel darah merah.

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ABSTRACTBackground Zoonotic case of malaria had just come to the attention of public health communities after the Sarawak study in 2004. Zoonotic malaria is caused by Plasmodium knowlesi, primarily a simian malaria parasite in wild long tail macaque Macaca fascicularis and pig tail macaque M. nemestrina as the reservoir hosts. The diagnosis of malaria parasites has mainly relied on the microscopic examination. However, this method is labor intensive, requires an experienced microscopist and difficult in identifying mixed infections in very low parasitemia cases. Recently, DNA barcoding system, which is based on the PCR amplification of a short and highly conserved region of mitochondrial cytochrome c oxidase sub unit I COI has shown to be an invaluable tool for diagnosing and differentiating the species of wide range of organisms. This study was aimed to develop identification tools of malaria parasite by using mtDNA COI gene as a molecular marker and reveal the molecular basis of zoonotic malaria by identifying the genetic variation of protein coding gene of DARC Duffy Antigen Receptor for Chemokines and DBP Duffy Binding Protein that are related to receptor ligand interaction in red blood cell invasion.Methods In silico study was carried out for verifying the potential of DNA barcoding based on the mtDNA COI gene sequence as a marker identification. Sensitivity and specificity analyses were carried out to validate the use of DNA barcoding for medical diagnosis of parasitic infection. Molecular technology of PCR Sequencing was carried out for screening malaria parasit in human and non human primate population and identifying the genetic variation within protein coding gene of DARC and DBP. Results We have initiated a study to explore the use of DNA barcoding for malaria parasite diagnosis through in silico study. We have thus designed primers spanning a 670 bp fragment of the 5 rsquo region of COI gene that could detect parasite isolates as low as 1 3 parasite per l. DNA barcode was used to detect malaria parasite in human population in Central Kalimantan. Of the 2309 subjects, 78 3.34 subjects were malaria positive of which two samples were determined as P. knowlesi infection. The detection rate of COI barcode was significantly higher as compared to microscopic 2.82 and 18S rRNA 1.82 analyses. Of the 366 non human primate samples that include 323 orangutan and 43 macaque 168 orangutan were found to be positive for either P. species ovale type, P. species vivax cynomolgi type, P. species vivax hylobati type and P. species malariae inui type. In macaque, 10 samples were positive for P. knowlesi, P. coatneyi, P. inui and P. species malariae inui type similar to that found in orangutan. The study has also found genetic variation in both human and non human primates Duffy Antigen Receptor for Chemokines and malaria parasite Duffy Binding Protein.Conclusions The study showed that mtDNA COI can be used to diagnose malaria parasites at very low parasitemia level and applied as a diagnosis tool for identification of zoonotic malaria. There is genetic variation in both human and non human primates Duffy Antigen Receptor for Chemokines and malaria parasite Duffy Binding Protein as major determinants for the invasion of malaria parasite."