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The National Centre for Genetic Engineering and Biotechnology (BIOTEC) was established under the Ministry for Science, Technology and Energy in September 1983. In 1991, Thailand established the National Science and Technology Development Agency (NSTDA) and BIOTEC became one of the NSTDA centres, operating autonomously outside the normal framework of civil service and state enterprises. This enabled it to operate more effectively to support and transfer technology for the development of industry, agriculture, natural resources, the environment and the socio-economy (Sriwatanapongse et al., 2000).

BIOTEC policy provides the resources for the country to develop the critical mass of researchers necessary to achieve Thailand's national R&D requirements in biotechnology. This is achieved through R & D projects, the facilitation of transfer of advanced technologies from overseas, human resource development at all levels, institution building, information services and the development of public understanding of the benefits of biotechnology.

BIOTEC is both a granting and implementing agency. It allocates approximately 70% of its R & D budget to several universities and research institutes in Thailand and 30% for in-house research projects. The facilities of national and specialized laboratories are made available for in-house research programmes as well as for visiting researchers. The construction of a Science and Technology Park will be completed in 2001 and will house BIOTEC's main laboratories.

Several research programmes have been undertaken by a BIOTEC-appointed committee of recognized experts in the field. The major biotechnology programmes and activities are described below.

Shrimp

Basic knowledge about the major cultivated shrimp species has lagged behind technical innovations that have led to successful intensification of culture and to ever-increasing world production. Sustaining high production levels will require innovation to minimize adverse environmental impacts. Biotechnology will play a central role in helping us to know more about shrimp and thereby improve rearing practices. BIOTEC's support will focus on issues dealing with shrimp diseases and with improvement of the seed supply. The disease work has so far emphasized the characterization, diagnosis and control of serious shrimp pathogens, particularly yellow-head disease (YHD) and white-spot syndrome (WSS) disease. Luminescent bacterial infections have contributed to the declining production to a lesser degree. These diseases become progressively more serious threats to the industry as it has grown and intensified. Indeed, the work on YHD virus and WSS virus (WSSV) supported by BIOTEC has been instrumental in substantially reducing losses in Thailand. The losses to YHD (probably exceeding US$40 million in 1995) and those to WSS (probably exceeding US$500 million in 1996) could have been much more serious without the basic knowledge and the DNA diagnostic probes made available to the industry by Thai researchers. Checking for subclinical WSSV infections by polymerase chain reaction (PCR) has been a common practice in Thailand, to help farmers in screening out WSSV +ve postlarvae (PL) before stocking (Flegel, 1997).

The Shrimp Biotechnology Service Laboratory was established in July 1999 at BIOTEC to summarize the reference PCR methods for viral disease detection in Thai shrimp farming. Laboratory objectives are to serve as the reference laboratory for major shrimp pathogen diagnosis based on molecular techniques, to conduct research and to provide assistance for the molecular detection of various shrimp viruses.

It has been reported that WSSV can be vertically transmitted and widespread among wild brood-stock. In addition to the disease problem, a decline in the growth rate of shrimp produced from currently available wild brood-stock has also been observed. Production of specific pathogen-free animals and the development of specific pathogen-resistant (SPR) strains are now being carried out in the USA, Venezuela and French Polynesia with Penaeus stylirostris and Penaeus vannamei. This could be considered a breakthrough, since production of P. vannamei more than doubled during 1992-1994. Currently the most important programme involves the domestication and genetic improvement of Penaeus monodon stocks (Withyachumnarnkul et al., 1998). The project will lead to the development of SPR stocks and improved growth performance through selective breeding. The first domesticated stocks from this programme were to be ready for pond production tests in 1999. BIOTEC is also supporting advanced studies on DNA characterization and DNA tagging of the shrimp stocks. These studies are providing the tools that will be important for rapid genetic improvement strategies.

BIOTEC is dedicated to the principle that the players in the shrimp industry should take an active role in the R & D effort for their industry, in both planning and finance. BIOTEC took an active part in promoting the formation in 1996 of an industry consortium (the Shrimp Culture Research and Development Company) dedicated to solving problems common to the shrimp aquaculture industry as a whole. This consortium serves the industry directly and also serves as a bridge to other public and private institutions involved in relevant research, not only in Thailand, but throughout the world.

Cassava and starch

About 70% of the 16 million t of cassava root produced in 1998 was used in the production of pellets and chips, and the remaining 30% was mainly used to produce flour and starch. A production shortage in 1997/98 prompted the Thai Tapioca Development Institute (TTDI) and Kasetsart University to develop a new strain with a higher yield. Kasetsart 50, a new strain, has an average yield of 26.4 t of roots ha-1 and a starch content of 26.7%, compared with 13.75 t ha-1 and 18% starch content of the best strain available.

The tapioca starch industry is one of the largest in Thailand. In 1998, tapioca starch was worth about US$120 million. About 40% of starch was used domestically for the production of modified starch, sweetener and monosodium glutamate. Most of the remaining 60% was exported. Efficient production, low production costs and the development of value-added products are vital to the starch industry and the farming sector (total of 1.3 million ha planted in cassava). The programme on starch and cassava products was established to provide support and funding for R & D. The programme is funded jointly by BIOTEC and TTDI to carry out R & D in three core activities. The short-term project aims to improve the processing efficiency of starch production, in particular to minimize water and energy consumption. This will reduce water use and costs and also reduce waste-water treatment. Waste-water discharge varies from 13 to 50 m3 t-1 of starch produced, with an average of 20m3. A benchmark on water use is a priority for the Thai starch industry.

Biotechnology can play an important role in waste utilization. Solid waste (after starch extraction) still contains 50% starch (dry weight) and has been utilized as animal feed. Tapioca, however, is not suitable for the production of feed requiring high protein content. Attempts have been made for protein enrichment using various microorganisms, such as Aspergillus and Rhizopus. Nevertheless, the economic feasibility is still in doubt and further technological development is needed. In contrast, turning waste water into energy through high-rate anaerobic digestion is promising. Though the technology is proven, an adaptation to such high-strength waste water and low buffering capacity is required to ensure stability of the system. In comparison with the upflow anaerobic sludge blanket (UASB) technology, the fixed bed is easier to control and operate. R & D, however, is focused on increasing loading efficiency. Based on calculations, methane generated from anaerobic treatment of starch waste water from 60 factories would be approximately 630 million m3 annually. This could be substituted for fuel oil used in drying, saving energy costs of about US$4 million annually. There is also the environmental cost of large land areas required for conventional pond systems. In addition to native starch, production of modified starch is increasing, leaving an excessive amount of sulphate in waste water. This may interfere with the anaerobic digestion intended for energy production. A number of papers have been published recently on the interactions between the sulphate-reducing bacteria (SRB) and the methanogenic bacteria (MGB). Molecular diagnosis has been developed and applied for the mixed cultured system. A better understanding of these anaerobic microbes could lead to the biological removal of sulphate, which is the main problem of various industries.

The European Union (EU) has set a quota for exported tapioca pellets. Product diversification is part of the second core research activity. As a result, production of biodegradable plastic from cassava starch is being investigated. Increasing use of cassava as a raw material for fermentation industries, such as amino acids and organic acids, must proceed, expanding the development of value-added products. To reduce costs of production, however, research is orientated towards the production of good-quality cassava chips as a starting material to replace the starch.

Finally, basic research on cassava starch structure and properties will add to our knowledge and help increase the use of cassava starch. The Cassava and Starch Technology Unit, a specialized BIOTEC laboratory established in 1995 at Kasetsart University, has been engaged in studying the physicochemical properties of cassava. The unit is well equipped and provides regular service and training on instrument analysis of starch properties for the private sector and government agencies.

Rice

Rice yields in Thailand are low. One of the major constraints in cultivation is blast disease, especially in high-quality rice cultivars, such as the aromatic Khao Dawk Mali. In northern Thailand, about 200,000 ha of rice were affected by blast in 1993, causing serious economic loss and resulting in government intervention of about US$10 million to assist disease-struck farms.

Another US$1.2 million was spent on fungicides (Disthaporn, 1994). Attempts have been made to breed higher resistance levels to blast in Thai rice. Limiting factors, however, are lack of insight and information on resistance genes and the complex structure of the pathogen populations. Genetic analysis provides an efficient tool to identify useful resistance genes in the host while analysing the race composition of the pathogen population. Recent research activities applying molecular genetic methods (DNA fingerprinting of a blast isolate collection at Ubon Ratchathani Rice Research Station, mapping of host resistance genes by the DNA Fingerprinting Unit at Kamphaengsaen campus of Kasetsart University) are providing baseline data on the interaction between rice and blast. The project is working on three closely related areas as follows:

• Establishment of a suitable differential cultivar series; identification of resistance genes conferring complete and partial resistance to blast disease in rice.

• Pathotype and molecular genetic characterization of the blast pathogen population in Thailand. So far, more than 500 monospore isolates have been deposited with the BIOTEC specialized culture collection.

• The special case of fertile isolates: the potential of using Thai isolates of Magnaporthe grisea for the development of a molecular diagnostic tool for pathogen race analysis. The degree of fertility can be assessed from the timing and number of perithecia that develop. BIOTEC has the capacity to test the mating type of about 80 isolates per month.

This project is a nationwide, network-type collaboration combining molecular genetics and classical approaches to help scientists breed rice cultivars with improved blast resistance.

BIOTEC provided US$1.5 million in 1999 to fund the Rice Genome Project Thailand. On behalf of Thailand, BIOTEC has joined an International Collaboration for Sequencing the Rice Genome (ICSRG) by sequencing 1 Mb annually of chromosome 9 for the next 5 years. BIOTEC is expected to provide about US$3.7 million to cover this work. Chromosome 9 was selected based on the previous extensive work on the fine genetic and physical maps surrounding the submergence-tolerance quantitative trait locus (QTL), the prospect of gene richness and the small chromosome size. Joining ICSRG will allow Thai scientists to directly access the rest of the genome sequence made available by the other collaborating members. Gene discovery from wild rice germ-plasm will be undertaken in parallel to efficiently use the genome sequence data. The project will bring Thailand into the international scientific arena, incorporate state-of-the-art technology and improve Thailand's competitive edge in the international rice market.

Dairy cows

In 1997, Thai milk consumption was 12 l person-1 year-1. Milk production is still insufficient to meet local demand and Thailand has to import more than

50% (worth US$305 million) of the dairy products consumed in the country. To meet the national demand, we need an additional 130,000 cows.

Reproductive efficiency is a primary determinant of dairy herd production profitability. Milk yield (10 kg day-1) is still far below the average (30 kg day-1) of most developing countries. It is therefore important to promote an increase in dairy production through science and technology. The major programmes are breeding and feeding. The lack of proper management is another major contributing factor to an underproductive dairy industry.

Traditional breeding practices in Thailand have been too slow to meet national requirements and importing pregnant heifers and/or young quality-bred calves from abroad is too costly. Cutting-edge technologies, such as embryo transfer, in vitro fertilization, embryo sexing and semen sexing, have been studied by Thai scientists for more than 10 years. Nevertheless, the technologies have not yet been adopted. Technology transfer and training of Thai researchers at the leading laboratories/companies are now under discussion. The goal is to increase production of high-quality heifer calves at the most economical cost.

Gene engineering

By the mid-1970s, with biotechnology centred in genetic engineering and molecular biology, Thailand was ready to adopt the new tools and apply them to various practical problems, first in the biomedical field and later in agriculture and other areas. A few specific examples will be given here to highlight the application of molecular biology and genetic engineering to agricultural development. Efforts in agricultural biotechnology and genetic engineering have been focused on three main areas: plant transformation, DNA fingerprinting and molecular diagnosis of plant and animal diseases. The first area should lead to the production of transgenic plants with superior properties, including resistance to diseases, insect pests and abiotic stresses.

The Plant Genetic Engineering Unit (PGEU), the specialized laboratory of BIOTEC at Kasetsart University, Kamphaengsaen campus, was established in 1985 to carry out work on plant biotechnology and genetic engineering. A transgenic tomato plant carrying the coat-protein gene of tomato yellow-leaf-curl virus was first developed to control this serious virus disease of tomato (Attathom et al., 1990). The same approach was taken to develop transgenic papaya and pepper for resistance to papaya ringspot virus and chilli vein-banding mottle virus, respectively (Chaopongpang et al., 1996; Phaosang et al., 1996). Sri Somrong 60, a Thai cotton variety, was successfully transformed with cryIA[b] gene expressing a toxin from Bacillus thuringiensis (Bt). Development of transgenic rice varieties has been supported by the Rice Biotechnology Programme launched by BIOTEC and the Rockefeller Foundation. An example is the transformation of Khao Dawk Mali 105, an aromatic Thai rice with A1 -pyrroline-5-carboxylate synthetase (P5CS) for salt and drought tolerance. Most transgenic plants are now being tested under greenhouse conditions in accordance with the Biosafety Guidelines (Attathom and Sriwatanapongse, 1994; Attathom et al., 1996). Field testing of trans-genic plants developed in Thailand will begin in 2001.

DNA fingerprinting

Each living creature has a unique DNA sequence. Using DNA fingerprinting and PCR, scientists can identify organisms and genes. Important genes can be located (genetic maps). Moreover, the availability of DNA probes and specific sequences has made it possible to develop appropriate molecular methods for the diagnosis of plant and animal diseases. Molecular mapping of genes in rice involving flooding tolerance, rice blast, aroma, cooking quality and fertility restoration were accomplished using three mapping populations. A back-cross breeding programme for the improvement of Jasmine rice was initiated. In the first stage, resistance to bacterial leaf blight, flooding tolerance, resistance to brown planthopper/gall midge and photoperiod insensitivity were the main areas of focus. Restriction fragment length polymorphism (RFLP)-based markers were an important limiting factor for high throughput and cost-effectiveness. The PCR-based marker for Xa21 is the most reliable for marker-assisted back-crossing in rice.

Tomato production in the tropics and subtropics faces serious constraints due to bacterial wilt (BW), a disease caused by the bacterial pathogen recently reclassified as Ralstonia solanacearum (previously Pseudomonas solanacearum). In Thailand, an endemic outbreak of BW in tomato, potato, pepper, ginger and groundnut occurs each year, causing a yield loss of approximately 50-90% depending on growing conditions. BW-resistant varieties cannot easily be developed, due to the nature of the (quantitatively inherited) resistance, which involves several genes. Marker-assisted selection (MAS), a breeding method of selecting individuals based on markers linked to target genes, in addition to phenotypic measurement, is essential and useful only for enhanced resistance to diseases. At this time, three putative OTLs corresponding to BW resistance have been found, using amplified fragment length polymorphism (AFLP) markers. Once markers closely linked to BW-related OTLs are well established, they can be used for marker-assisted breeding for enhanced resistance to bacterial wilt in tomato. A tomato consortium has been set up to extend public-private collaboration.

BIOTEC has set up the DNA Fingerprinting Service Unit at Kasetsart University. The unit has provided services for public and private concerns for more than 2 years. The main services are DNA fingerprinting and DNA diagnosis.

Biocontrol

In 1996, Thailand imported 38,000 t of chemicals, mainly insecticides and herbicides. The global trend of going organic is an opportunity for Thai farmers to supply fresh organic produce, especially fruit and vegetables, to the world. Over the past decade, the developmental work on biocontrol in Thailand has continued to receive active support from BIOTEC and the Thailand

Research Fund. Two companies are now producing commercially grown Trichoderma to control Sclerotium rolfsii Sacc. and Chaetomium to control soil fungi, such as Phytophthora (Yuthavong, 1999). BIOTEC and the Department of Agriculture have set up a pilot-scale production facility to produce nuclear polyhedrosis virus (NPV), Bt and Bacillus sphericus. NPV is widely used to control Spodoptera moth in grapes. Bt produced locally has gained popularity over the last few years. The capacities of pilot plants at Mahidol University and King Mongkut's University of Technology (Thonburi) are taken up with Bt production. Commercial production may begin soon. A project at Mahidol University to transfer the chitinase gene into B. thuringiensis subsp. israelensis has received support from BIOTEC.

Biosafety

Biosafety issues are being debated in Thailand. The National Biosafety Committee (NBC) was established in January 1993 under BIOTEC. The NBC has introduced two biosafety guidelines: one for laboratory work and the other for fieldwork and the release of genetically improved organisms (GIOs) into the environment. The establishment of institutional biosafety committees (IBCs) at various public institutes and private companies was also strongly recommended and, in many cases, these recommendations have been implemented.

The importation of prohibited materials under Plant Quarantine Law BE 2507 implemented by the Department of Agriculture controls to a certain degree the use of GIOs. Ministry regulation II (1994 [A.D.]) identifies certain prohibited transgenic plants. Permission from the Ministry of Agriculture is required to perform field testing of transgenic plants brought into Thailand. The following have received permission to perform the test: the Flavr Savr tomato produced by Calgene for the production of seeds (1994); a field trial of Monsanto Bt cotton was carried out under restricted containment in a netted house in 1996; in 199 7, a Bt maize field trial was approved to be carried out by Novartis at their experiment station in a netted screen house.

People seem to pay more attention to the introduction of GIOs into the country by the multinational companies than to considerations of technological information. An issue never discussed or debated, in particular at the political level, is whether or not Thailand should be more aggressive on the development of transgenic organisms. Thailand is rich in biodiversity and several genes resistant to biotic and abiotic stresses embedded in wild plants and other bioresources need to be discovered and utilized. This illustrates the potential benefits of biotechnology and genetic engineering. In the 1980s, when genetic engineering and biotechnology first made their impact felt, genetic engineering capability was present in only two or three institutions in Thailand (Yuthavong, 1987). Ten institutions now have genetic engineering capability. Nevertheless, the most important challenge for the future of GIOs is not technical in nature, but the attitude of the public towards the technology. These issues need to be studied and debated among the scientists, the public and the policy-makers, and an optimal policy developed. BIOTEC realizes that genetic engineering depends critically on public support, so the Centre has emphasized public education, with information programmes on GlOs being introduced to the public and to industry.

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