scholarly journals ANALISIS MOLEKULER FILOGENETIK DAN STRUKTUR ANTIGENIC VIRUS AVIAN INFLUENZA SUBTIPE H5N1 ISOLAT LAMPUNG TAHUN 2008-2013

2015 ◽  
Vol 9 (1) ◽  
Author(s):  
Eko Agus Srihanto ◽  
Widya Asmara ◽  
Michael Haryadi Wibowo
Keyword(s):  
Amino Acids ◽  
Avian Influenza ◽  
Animal Health ◽  
Antigenic Site ◽  
Multiple Alignment ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Phylogenic Tree ◽  
Tree Analysis ◽  
Polymerase Chain

Penelitian ini bertujuan melakukan karakterisasi molekuler antigenic site terhadap isolat virus avian influenza (AI) Balai Penyidikan dan Pengujian Veteriner (BPPV) Regional III Lampung dari tahun 2008-2013. Amplifikasi RNA dilakukan dengan teknik reverse transcription polymerase chain reaction (RT-PCR) menggunakan 4 pasang primer referens dari Australian Animal Health Laboratory (AAHL) Geelong Australia (HA10, HA20, dan HA30) dan dilanjutkan dengan proses pengurutan. Analisis hasil pengurutan dengan menggunakan perangkat lunak MEGA versi 5.05 yang meliputi multiple alignment, deductive amino acids prediction, dan phylogenic tree analysis diperoleh hasil perbedaan genetik antar isolat Lampung dari tahun 2003-2013 ditemukan berkisar 1,1-9,1% dengan tingkat homologi mencapai 90,9-98,9%. Variasi genetik ditemukan adanya substitusi pada posisi 53 (R53K), 126 (D126E), 136 (P136), 138 (H138Q, dan H138L), 140 (R140K, R140S, dan R140N), 141 (S141P), dan 189 (K189R). Berdasarkan analisis filogenic tree isolat Lampung tahun 2008-2011 termasuk ke dalam clade 2.1.3. Analisis filogenik isolat AI tahun 2012-2013 yang menginfeksi unggas air mempunyai homologi sekitar 98,5-99,1% dibandingkan dengan isolat AI yang menginfeksi unggas air asal Jawa dan termasuk ke dalam clade 2.3.2.1.

scholarly journals Evaluation of a 384–Well Format for High-Throughput Real-Time Reverse Transcription Polymerase Chain Reaction for Avian Influenza Testing

2009 ◽  
Vol 21 (5) ◽  
pp. 679-683 ◽  
Author(s):  
Pamela J. Ferro ◽  
Jason Osterstock ◽  
Bo Norby ◽  
Geoffrey T. Fosgate ◽  
Blanca Lupiani
Keyword(s):  
Polymerase Chain Reaction ◽  
Avian Influenza ◽  
Real Time ◽  
High Throughput ◽  
Reverse Transcription ◽  
Virus Isolation ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Response Planning ◽  
Polymerase Chain

As concerns over the global spread of highly pathogenic avian influenza H5N1 have heightened, more countries are faced with increased surveillance efforts and incident response planning for handling a potential outbreak. The incorporation of molecular techniques in most diagnostic laboratories has enabled fast and efficient testing of many agents of concern, including avian influenza. However, the need for high-throughput testing remains. In this study, the use of a 384–well format for high-throughput real-time reverse transcription polymerase chain reaction (real-time RT-PCR) testing for avian influenza is described. The analytical sensitivity of a real-time RT-PCR assay for avian influenza virus matrix gene with the use of both 96– and 384–well assay formats and serial dilutions of transcribed control RNA were comparable, resulting in similar limits of detection. Of 28 hunter-collected cloacal swabs that were positive by virus isolation, 26 (92.9%) and 27 (96.4%) were positive in the 96– and 384–well assays, respectively; of the 340 hunter-collected swabs that were negative by virus isolation, 45 (13.2%) and 23 (6.8%) were positive in the 96– and 384–well assays, respectively. The data presented herein supports the utility of the 384–well format in the event of an avian influenza outbreak for high-throughput real-time RT-PCR testing.

scholarly journals DETEKSI VIRUS AVIAN INFLUENZA SUBTIPE H5N1 DI BEBERAPA PASAR UNGGAS HIDUP DALAM WILAYAH PROVINSI JAWA BARAT SEKITARNYA

2015 ◽  
Vol 9 (1) ◽  
Author(s):  
Atik Ratnawati ◽  
NLP Indi Dharmayanti
Keyword(s):  
Polymerase Chain Reaction ◽  
Avian Influenza ◽  
Reverse Transcriptase ◽  
Ribonucleic Acid ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Polymerase Chain

Pada penelitian ini dilakukan identifikasi virus avian influenza (AI) subtipe H5N1 pada unggas dan lingkungan pasar untuk mengetahui peran pasar sebagai sumber penularan virus. Metode yang digunakan dalam penelitian ini adalah pengambilan sampel swab kloaka unggas dan lingkungan di beberapa pasar di wilayah Jawa Barat dan Tangerang. Sampel selanjutnya dilakukan isolasi ribonucleic acid (RNA) dan dilakukan reverse transcriptase polymerase chain reaction (RT-PCR) dengan menggunakan primer AI subtipe H5N1. Hasil penelitian menunjukkan bahwa virus AI/H5N1 terdeteksi pada unggas dan lingkungan pasar. Disimpulkan bahwa pasar dapat menjadi sumber penularan virus AI subtipe H5N1 terhadap unggas lainnya.

scholarly journals Development of multiplex reverse transcription polymerase chain reaction (RT-PCR) for simultaneous detection of matrix, haemagglutinin and neuraminidase genes of H5N1 avian influenza virus

2012 ◽  
Vol 28 (2) ◽  
pp. 55-59 ◽  
Author(s):  
R. Mojumder ◽  
E. H. Chowdhury ◽  
R Parvin ◽  
J. A. Begum ◽  
M. Giasuddin ◽  
...  
Keyword(s):  
Polymerase Chain Reaction ◽  
Influenza Virus ◽  
Avian Influenza ◽  
Matrix Protein ◽  
Simultaneous Detection ◽  
High Fidelity ◽  
Rt Pcr ◽  
Chain Reaction ◽  
One Step ◽  
Polymerase Chain

Influenza A virus, subtype H5N1 causes a fatal disease in domestic poultry and could spread directly from poultry to humans. The aim of this study was to develop a multiplex reverse transcription polymerase chain reaction (mRT-PCR) for simultaneous detection of Type A influenza virus-specific matrix protein (M) gene as well as H5 and N1 genes of highly pathogenic avian influenza (HPAI) viruses. Finnzymes Phusion-Flash High- Fidelity PCR Master Mix (Finnzymes Oy, Finland) and Qiagen one-step RT-PCR enzyme mix (Qiagen, Germany) were used in a one-step RT-PCR. RNA was extracted from two known positive samples using Qiagen RNA extraction kit. RT-PCR was carried out with a mixture of primers specific for the Type A influenza virus matrix protein (M), and H5 and N1 genes of H5N1 HPAI viruses in a single reaction system. The mRT-PCR cDNA products were visualized by gel electrophoresis. The mRT-PCR yielded fragments of 245 bp for M, 545 bp for H5 and 343 bp for N1 genes of HPAI virus, which were clearly distinguishable. The mRT-PCR using the Finnzymes Phusion-Flash High-Fidelity PCR Master Mix (Finnzymes Oy, Finland) with Qiagen one-step RT-PCR Enzyme Mix (Qiagen, Germany) required only one hour and 20 minutes. (Bangl. vet. 2011. Vol. 28, No. 2, 55 – 59)DOI: http://dx.doi.org/10.3329/bvet.v28i2.10653

scholarly journals Deteksi Antigen Virus Avian Influenza pada Ayam Kampung di Pasar Hewan Beringkit dan Pasar Umum Galiran, Bali

2020 ◽  
Vol 9 (5) ◽  
pp. 757-772
Author(s):  
Annisa Musdalifa ◽  
Gusti Ayu Yuniati Kencana ◽  
I Nyoman Suartha
Keyword(s):  
Polymerase Chain Reaction ◽  
Avian Influenza ◽  
Real Time ◽  
Highly Pathogenic Avian Influenza ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Chi Square ◽  
Highly Pathogenic ◽  
Pathogenic Avian Influenza ◽  
Polymerase Chain

Avian Infuenza termasuk ke dalam kelompok penyakit menular strategis dan bersifat zoonosis. Penyebabnya adalah virus Highly Pathogenic Avian Influenza (HPAI) subtipe H5N1. Introduksi virus Avian Infuenza subtipe H5N1 dapat melalui jalur pasar karena berpotensi dalam penyebarannya penyakit Avian Influenza pada ayam kampung secara alamiah melalui kontak langsung dengan unggas lain. Penelitian ini dilakukan di Pasar Hewan Beringkit, Kabupaten Badung dan Pasar Umum Galiran, Kabupaten Klungkung. Tujuan penelitian ini untuk mengetahui keberadaan virus Avian Influenza subtipe H5N1 pada ayam kampung di Pasar Hewan Beringkit dan Pasar Umum Galiran. Sampel yang digunakan sebanyak 120 sampel swab kloaka dan swab trakea ayam kampung, dengan jumlah masing-masing pasar sebanyak 60 sampel. Periode pengambilan sampel dilakukan selama dua bulan dengan periode pengambilan setiap dua minggu sekali sebanyak empat kali. Pengambilan sampel dilakukan pada ayam kampung berumur di atas tiga bulan yang tidak divaksinasi. Isolasi virus pada telur ayam berembrio (TAB) berumur sembilan hari dan identifikasinya dengan uji hemaglutinasi (HA), uji hambatan hemaglutinasi (HI) serta Real Time Polymerase Chain Reaction (RT-PCR). Analisis data hasil uji antigen Avian Influenza dengan uji statistika non-parametrik Chi-square (?2). Hasil uji sampel swab kloaka dan trakea ayam kampung asal Pasar Beringkit positif virus Avian Influenza sebesar 5%, sedangkan sampel positif virus Avian Influenza asal Pasar Galiran sebesar 6,7%. Berdasarkan hasil penelitian maka dapat disimpulkan bahwa virus Avian Influenza terdeteksi pada ayam kampung di Pasar Hewan Beringkit dan Pasar Umum Galiran, virus AI masih bersikulasi pada daerah asal peternakan dan penyakit AI masih mewabah di Bali.

scholarly journals Analysis of pFQ12, a 22.4-kbFrankiaplasmid

2001 ◽  
Vol 47 (7) ◽  
pp. 608-617 ◽  
Author(s):  
Theodore R John ◽  
Jeffrey M Rice ◽  
Jerry D Johnson
Keyword(s):  
Amino Acids ◽  
Woody Plants ◽  
Codon Bias ◽  
Open Reading Frames ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Rich Region ◽  
Filter Hybridization ◽  
Polymerase Chain ◽  
Reading Frames

Frankia are gram-positive, filamentous bacteria capable of fixing atmospheric dinitrogen in symbiosis with a wide variety of woody plants and shrubs. Some isolates of Frankia harbor plasmids of 8.5 (pFQ11) and 22.4 kb (pFQ12) that have no known function but are transmitted through many generations in culture. We have sequenced the 22 437-bp pFQ12 plasmid that is present in isolates CpI1 and ArI3. This sequence, with 76% G+C, is almost totally unrelated to that of pFQ11 found in the same cells. However, four regions of identity, 40-90 bp each, are dispersed around the plasmids. The 22.4-kb plasmid has >50 open reading frames (ORFs) that encode putative proteins of more than 100 amino acids, with the largest being 2226 amino acids. Twenty of these ORFs are likely to encode proteins based on their codon bias as determined by two different algorithms. Transcripts from nine of these regions have been identified by reverse transcriptase-polymerase chain reaction (RT-PCR) or filter hybridization. The two Frankia plasmids each encode a protein similar to the korSA protein that regulates transmission of pSAM2 in Streptomyces. The origin of replication (ORI) region of pFQ12 was localized by intrastrand AT and GC equivalence switch. It includes a 40-bp, intergenic, A+T-rich region that has a strong identity in pFQ11.Key words: ORI analysis, RT-PCR, Glimmer, DNA sequence.

LOW SPECIFICITY OF A NESTED REVERSE TRANSCRIPTION POLYMERASE CHAIN REACTION TO DETECT AVIAN INFLUENZA VIRUS NUCLEOPROTEIN GENE

2016 ◽  
Vol 43 (02) ◽  
pp. 75-79
Author(s):  
Lih-Chiann Wang ◽  
Wei-En Hsu ◽  
Wei Thong ◽  
Ting-Yen Chao ◽  
Ching-Ho Wang
Keyword(s):  
Polymerase Chain Reaction ◽  
Influenza Virus ◽  
Avian Influenza ◽  
Avian Influenza Virus ◽  
Reverse Transcription ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Nucleoprotein Gene ◽  
Polymerase Chain ◽  
Np Gene

Reverse transcription polymerase chain reaction (RT-PCR) was used routinely to detect the avian influenza virus (AIV) nucleoprotein (NP) gene. The purpose of the present study was to compare the correctness of a nested RT-PCR (nRT-PCR), one conventional RT-PCR with its outer primer (oRT-PCR) and the other conventional RT-PCR with its inner primer (iRT-PCR) to detect AIV NP gene. A total of 365 AI-free fecal swabs (73 pools), 7 tracheal swabs and anllantoic fluid from 25 chicken embryos were used to determine the analytic specificities of those tests. Compared with the iRT-PCR, the nRT-PCR was more sensitive for AIV detection. However, the specificities of nRT-PCR, oRT-PCR and iRT-PCR were 48.6% (35/72), 100% (67/67) and 91.3% (84/92), respectively. The amplifying band was sequenced and confirmed to be the AIV NP gene as the positive control. The specificity of this nRT-PCR is too low to be used for the AIV screening test.

scholarly journals PENENTUAN SUBTIPE VIRUS AVIAN INFLUENZA DENGAN METODE SINGLE STEP MULTIPLEX REVERSE TRANSCRIPTASE- POLYMERASE CHAIN REACTION (RT-PCR) ISOLAT ASAL PROVINSI ACEH

2014 ◽  
Vol 8 (1) ◽  
Author(s):  
Teuku Zahrial Helmi ◽  
Rini Widayanti ◽  
Aris Haryanto
Keyword(s):  
Polymerase Chain Reaction ◽  
Avian Influenza ◽  
Reverse Transcriptase ◽  
Influenza A ◽  
Deoxyribonucleic Acid ◽  
Single Step ◽  
Rt Pcr ◽  
Chain Reaction ◽  
Polymerase Chain ◽  
Virus Influenza

Tujuan dari penelitian ini adalah mengidentifikasi keberadaan gen M, H5, dan N1 virus avian influenza (AI) melalui metode single step multiplex reverse transcriptase-polymerase chain reaction (RT-PCR), sebagai acuan untuk peneguhan diagnosis secara molekuler virus AI di Provinsi Aceh. Penelitian ini menggunakan 11 isolat virus AI asal Provinsi Aceh yang diperoleh dari Laboratorium Balai Penyidikan dan Pengujian Veteriner (BPPV) Regional I Medan di Sumatera Utara dari tahun 2006-2008. Penelitian dilakukan di Laboratorium Virologi BPPV Regional I di Medan, Sumatera Utara. Amplifikasi terhadap gen matriks (M) virus AI menggunakan metode simplex RT-PCR. Hasil simplex RT-PCR terhadap gen M diperoleh 10 isolat yang menunjukkan pita deoxyribonucleic acid (DNA) pada 276 bp dan satu isolat yang tidak muncul, kemudian dilanjutkan dengan metode single step multiplex RT-PCR menggunakan pasangan primer gen penyandi protein N1, H5, dan M. Produk PCR 131 bp (N1), 189 bp (H5), dan 276 bp (M) muncul sebagai hasil elektroforesis dari semua isolat virus AI. Semua virus AI yang mewabah dari tahun 2006-2008 di Provinsi Aceh termasuk ke dalam virus influenza A subtipe H5N1.

scholarly journals TRACING THE KINSHIP OF BRUCELLA SP. BY USINGA POLYMERASE CHAIN REACTION TECHNIQUE WITH A SHORT PRIMER

2020 ◽  
Vol 21 (4) ◽  
pp. 493-502
Author(s):  
Difa Widyasari ◽  
Eko Sugeng Pribadi ◽  
Fachriyan Hasmi Pasribu ◽  
Widya Septiningtyas ◽  
Jati Adiputra
Keyword(s):  
Animal Health ◽  
Small Ruminants ◽  
Chain Reaction ◽  
Cloning And Sequencing ◽  
Phylogenic Tree ◽  
Cloning Technique ◽  
Polymerase Chain ◽  
Relationship Of ◽  
Pcr Techniques ◽  
Sequencing Result

Brucellosis is a zoonotic disease that has spread throughout the world and has an important impacton both human and animal health. The four species of Brucella that cause disease in humans are Brucellaabortus, B. suis, B. melitensis and B. canis, and B. melitensis as the most pathogenic species. This Researchused 46 samples were collected from Government Small Ruminants Abattoir in Bogor Regency. Thirty twospleen samples were examined by previous research and showed a positive result when were tested withCFT and PCR techniques, but sequencing has not yet been done. Fourteen serum and spleen samples wereexamined by the similar techniques. The Research aimed to determined the genetic relationship of Brucellasp. using a PCR technique with a specific short primer to B. mellitensis. Cloning technique was appliedpreviously to five PCR positive samples before sequencing. Cloning and sequencing result of the Sample91 showed higher homology to B. melitensis and B. abortus for 127 nucleotide lengths, 97.6% -100% and99.2% -100% respectively. In the phylogenic tree, the Sample 91 was part of B melitensis sequences 1, 2,and 3 with accession numbers LT962930.1 and LT962936.1, B abortus sequences 1 and 2 with accessionnumbers CP033079.1 and B. abortus sequence 1 with accession number CP034695.1. Sample of 95, 97, 7,and 13 have lower homologies than Sample 91.

Sign in / Sign up

Export Citation Format

Share Document