Diversity of Coat Protein Gene of Rice tungro bacilliform tungrovirus (RTBV) and Rice grassy stunt tenuivirus (RGSV) Isolates from Several Districts in Java Island
Keragaman Gen Protein Selubung Isolat Rice tungro bacilliform tungrovirus (RTBV) dan Rice grassy stunt tenuivirus (RGSV) dari beberapa Kabupaten di Pulau Jawa;
Keragaman Gen Protein Selubung Isolat Rice tungro bacilliform tungrovirus (RTBV) dan Rice grassy stunt tenuivirus (RGSV) dari beberapa Kabupaten di Pulau Jawa
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Date
2013Author
Astuti, Dwi
Nurhayati, Endang
Hendrastuti, Sri
Nurhayati, Endang
Hendrastuti, Sri
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Indonesia is the largest consumer of rice in the world. International Rice Research Institute (IRRI) estimated that Indonesia may require an increase in rice production for the next 25 years. Among many constrains in rice production increement is plant diseases including tungro and grassy stunt diseases. Tungro disease is caused by infection of two viruses, Rice tungro bacilliform tungrovirus (RTBV) and Rice tungro spherical waikavirus (RTSV). These viruses are transmitted by the rice leaf hopper vectors which mainly is green leafhoppers (Nephotettix virescens). RTBV is a determinant of the disease symptoms eventhough it could not infect rice without the presence of RTSV. Infected plants showed typical tungro symptoms involving stunting, yellow or orange-yellow discoloration of the leaves, reduced number of tillers and rust-colored spots on the leaves. In 2006 a new symptoms was found in paddy crop in Indonesia, i.e. yellow-orange discoloration of the leaves, reduced number of tillers without any reduction in plant height. It was reported that the disease is caused by infection of Rice grassy stunt tenuivirus (RGSV) the cause of grassy stunt disease. Symptoms similarity of tungro and RGSV infection may cause difficulty in disease diagnosis and cause confusion in determining diseases control strategies. Therefore research on symptoms and genetic diversity of RTBV and RGSV is necessary to determine the appropriate control strategy. The genetic diversity of RTBV isolates from South Asia and Southeast Asia as well as from some tungro endemic area in Indonesia based on the symptoms variation and diversity of coat protein gene have been reported. The genetic diversity of the isolates RGSV from two areas in Taiwan and Philippines based on symptoms variation have also been reported. Genetic and symptoms diversity of RTBV and RGSV could be analysed through the diversity of coat protein gene. Coat protein gene is known as a determinant factor of infection, virulence, and symptoms induction. The research is conducted to study the diversity of symptoms and coat protein gene of RTBV and RGSV from several districts in Java. The areas in Bogor, Subang, Pandeglang, Klaten, Batang, Sleman, Jember and Banyuwangi were selected as disease survey location because these regions were reported as endemic areas for tungro disease and brown leafhopper. The study includes three main activities, i.e. observation of symptoms in the field and sampling of infected plants, detection of RTBV and RGSV, and analysis of diversity of coat protein gene of RTBV and RGSV. Most infected plants in the fields showed tungro-like symptoms. Two variations of symptoms can be classified by the number of tillers and plant height. These symptoms variations can be caused by RTBV or RGSV infection or mix infection of RTBV and RGSV as indicated by detection results using PCR. Identification of total 64 samples showed infection of RTBV in 13 samples, RGSV in 36 samples and mix of RTBV and RGSV in 2 samples. The results showed that tungro-like symptoms were positive from infection of RGSV. Therefore identification of diseases based only on symptoms are not sufficient. The whole coat protein gene of RTBV was amplified using DA-F and DA-R primers with the amplicon of 1224 bp. Part of coat protein gene of RGSV amplified using F3 and B3 primers with the amplicon of 243 bp. Amplified DNAs were then used for sequencing and further analysis of genetic diversity among isolates using phylogenetic analysis by Neighbor-Joining method. Analysis of nucleotide sequence of coat protein gene from 10 isolates of RTBV showed 40% to 100% similarity, whereas analysis of amino acid sequence of the isolates showed 88% to 100% similarity. Analysis of nucleotide sequence of coat protein gene from 20 isolates of RGSV showed 30% to 98% similarity, whereas analysis of amino acid sequence of the isolates showed 20% to 100% similarity. The sequence analysis indicated that genetic diversity of RTBV occurred only in nucleotide level, whereas genetic diversity of RGSV is high on nucleotide as well as amino acid level. Phylogenetic trees based on nucleotide sequence of coat protein gene showed that 10 isolates of RTBV can be differentiated into 3 groups whereas 20 isolates of RGSV can be differentiated into 4 groups. The diversity of RTBV and RGSV isolates from Java is not location specific. Genetic value between RTBV isolates from Java with isolates outside Indonesia explained that RTBV isolates from Java is similar strain with RTBV isolates that have been reported. Genetic distance‟s value between RGSV isolates from Java with isolates outside Indonesia explained that RGSV isolates from Java is different strain with RGSV isolates that have been reported. Further research is required to determine the genetic and symptoms diversity of RTBV and RGSV isolates from other tungro and grassy stunt endemic areas in Indonesia.
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