June 1997, Volume 18 No. 2

Research Updates

Biogeography and Biotechnology: Fighting Dutch Elm Disease

Dutch elm disease is on the rampage again, but new methods being developed for its control rely on understanding the origins of the fungal pathogen, and manipulating its pathogenicity and the resistance of the tree. Clive Brasier of the UK Forestry Commision says that the elm and the pathogen are seriously out of balance: the pathogen is too aggressive for its host, and the traditional `front line' controls - quarantine, sanitation and chemical control - have been largely overwhelmed by the sheer momentum of the epidemic.

Unheard of before the beginning of the 20th century, two major pandemics of Dutch elm disease, a fungal wilt spread by Scolytus bark beetles, have since destroyed most of the elms (Ulmus spp.) in the epidemic areas which carved huge swathes across the northern hemisphere. The first pandemic was caused by a fungus identified as Ophiostoma ulmi and started in north-west Europe in about 1910. By the time it died down in the 1940s it had killed 10-40% of the elms in the region, notably the English elm, U. procera, and the wych elm, U. glabra. The pathogen appeared in North America in the 1920s with similar devastating effects, but here it did not die down, partly because of the greater susceptibility of the American elm, U. americana. The second pandemic began in the 1960s and was caused by a second and even more aggressive fungus, O. novo-ulmi. This caused a new and more virulent form of the disease which decimated the elm populations of Europe (killing some 25 million of Britain's 30 million elms, for example) and subsequently spread across Europe to central Asia, and into North America where it replaced O. ulmi, although without quite such disastrous results as in Europe because O. ulmi had never ceased being active there. The spread of the disease, as often the case with forest health problems, was helped in part by the international timber trade, and this allowed it eventually to reach even New Zealand (see BNI 14(3)). With so many trees dead, the pandemic died down, but then in the early 1990s, evidence of new epidemics appeared, as a new generation of elms grew big enough for the beetles that spread the disease to breed in.

The need to understand the origins of the pathogens was urgent, both to try and explain why such epidemics had occurred, and because identifying the area of origin would give an opportun-ity to look for possible biological control agents. But the origins of the two species and the relationship between them remained obscure despite extensive studies and worldwide surveys in the suspected area of origin in Asia and particularly China. Then in 1993 a fungus was found in the western Indian Himalayas, associated with beetle-feeding galleries on indigenous U. wallichiana. This proved to be a new species, O. himal-ulmi, and to be as pathogenic as O. novo-ulmi; but the relationship between all three species has remained elusive. However, the fact that O. himal-ulmi is quiescent in its natural habitat and produces no wilt symptoms in the indigenous elm raises hopes that there may be competitors or predators of the fungus in the Himalayas that are missing from the Dutch elm disease system elsewhere. Clive Brasier suggests that there is also much to be gained from studying the natural balance between elm, fungus and beetle in an endemic situation.

One novel approach to controlling the disease has been made possible by advances in biotechnology which have allowed researchers to focus on reducing the pathogenicity of the fungi. D-factors, first identified in 1983, are naturally occurring virus-like agents composed of double-stranded RNA segments. They occur in the fungal cytoplasm and are spread by hyphal fusion. They can reduce the aggressiveness of O. novo-ulmi; in the laboratory they slow growth and reduce sporulation and germination, and in the field in the UK they can increase the number of spores needed to infect an English elm (U. procera) to beyond the number carried by most vector beetles. They also change the behaviour and population structure of O novo-ulmi at epidemic fronts. Population studies conducted in Europe and North America have suggested that they may have played a role in the decline of the first Dutch elm disease epidemic in Europe in the 1940s. In any event, they are a fourth com-ponent in the system: tree-beetle-fungus-fungal virus. They exert an effect as natural biocontrol agents, and are also candidates for use, either unmodified or genetically man-ipulated, as artificial agents.

Studies currently underway at the Forestry Commission, supported by the Pilkington Trust, are aimed at characterizing the virulence of different d-factors, assessing the most appropriate ones for release, and studying interactions between them. Molecular studies are also being conducted (with Imperial College, London and funded by the Gatsby Trust), and one objective of these is to insert the d-factor into the fungal nucleus. The problem with cyto-plasmic transmission is that it only works between genetically similar individuals. At disease fronts, the fungus functions as a clone reproduc-ing by asexual reproduction, and cytoplasmic d-factors are easily transmitted. But as the disease becomes more established, the population begins to reproduce sexually, and as it thus becomes genetically more diverse, d-factor transmission wanes. There also appears to be a mechanism for eliminating cytoplasmic d-factors during sexual reproduction. So while cytoplasmic d-factors might impede the spread of the disease, they would be ineffective where the disease had really taken hold. The aim is that by inserting the d-factor into the nucleus, it would be spread more effectively in nature and these problems would be overcome.

The question of how introduced d-factors would affect the disease in the wild remains to be answered. Sites have been identified in New Zealand and the USA where there are single clones of O. novo-ulmi free of d-factors, which are thus suitable for studying the effects of released d-factors. It is hoped that experimental releases may be made soon. Clive Brasier argues that d-factors could be a significant factor in any eventual reduction in the aggressiveness of Dutch elm disease, whether this occurs naturally or artificially.

Another line of research is concerned with improving the resistance of the elm. Traditional breeding techniques using resistant trees from Asia may produce trees unsuited to other climates. However, progress in techniques of genetic manipulation allows an alternative to be pursued: indigenous elms with anti-fungal or anti-beetle genes incorporated. The Forestry Commission, in collaboration with HRI (Horticultural Research International) and Abertay University, Scotland have investigated the potential for manipulation of the English elm. Using Agrobacterium, marker genes have been successfully inserted, and the tree is proving to be an excellent model system for genetic manipulation in broad-leaved trees. A pilot anti-fungal gene (chosen from a range supplied by Zeneca, UK assayed for activity against O. novo-ulmi) may be inserted into the elm soon, and anti-beetle genes are also under consideration.

Although new hopes for controlling Dutch elm disease may seem brighter than they have ever been, Clive Brasier sounds a warning note: if such a highly aggressive tree pathogen as O. himal-ulmi can exist unsuspected in a major forest region such as the Himalayas, how many more exotic pathogen threats do we have yet to discover? Conquering Dutch elm disease may be just the beginning.

Source: Brasier, C. (1996) New horizons in Dutch elm disease control. In: Report on Forest Research 1996. Edinburgh, UK; Forestry Commision, pp. 20-28.

Whitefly Research and Control

Research on Bemisia argentifolii (also known as B. tabaci biotype B), the silverleaf whitefly, control in the USA has made great strides since the pest was first identified in Florida on Poinsettia in 1986 and a five-year plan of research and action was adopted, according to a report in Agricultural Research (February 1997). The whitefly, which spread quickly through Florida and into Texas, California and Arizona, attacks some 500 species of plant including cotton and vegetable crops. It is estimated to cause losses worth billions of dollars from direct damage and by spreading viral disorders.

The whitefly is not just an American problem. It also severely damages tomato crops in southern Europe and Israel. According to a report in New Scientist (21/28 December 96), it has been kept out of the UK so far by the rigorous application of plant import regulations, but Ministry of Agriculture, Fisheries and Food scientists have predicted that an outbreak there could destroy 20% of tomato crops in England and Wales and halve growers' profits.

Research by the US Department of Agriculture's Agricultural Research Service (USDA-ARS) together with universities, state departments of agriculture and industry has led to the identification of numerous new disorders in a multitude of crops from viruses spread by the whitefly. With few insecticides to choose from, delaying the development of resistance is seen as vital, and in California, biweekly monitoring of whiteflies and screening for chemical resistance allows mortality data to be reported to vegetable farmers via newsletters, news media and the Internet so they can decide the most appropriate insecticide. Rapid sampling pro-cedures and treatment thresholds have been developed for cotton which are used not only in the USA. In cotton, the whitefly honeydew causes crop contamination which lowers its value, and methods to deal with this are being developed, using enzymes to break down the honeydew oligosaccharides which cause stickiness. Research on crop resistance is a long-term process, but a cotton variety with whitefly resistance has been available for the last two years.

There has been a strong focus on biological control methods. Initially, indigenous US predatory Delphastus sp., Collops vitatus and Serangium parcesetosum beetles were studied, but latterly the ARS European Control Laboratory at Montpellier (France) has been a primary source of exotic natural enemies. More than 30 species of parasitic and predatory insects and hundreds of isolates of pathogenic fungi, collected during surveys of 25 countries in Africa, Asia, Europe and South America, have been dispatched for study. From among these, an Eretmocerus sp. from Spain proved to have a promising attack rate and was relatively more tolerant of some pesticides. Both this species and Eretmocerus spp. from India and Pakistan have persisted for one to two years in the field in California. Meanwhile, a native Eretmocerus sp. from Arizona was found to be effec-tive in greenhouses, and this is now being mass-reared in Europe for use in the greenhouse industry there.

Research into the use of pathogenic fungi formulated as bioinsecticides has also been productive. Mycotrol, a Beauveria bassiana-based product developed under a cooperative research and development agreement (CRADA) between the ARS and Mycotech (Montana) was given US Environmental Protection Agency approval in 1995 and registration is being sought in Europe. Meanwhile, Intrachem (Switzerland) have been granted the first provisional regist-ration in Europe for another B. bassiana-based product, Naturalis-L (manufactured by Troy Biosciences of Arizona).

A further CRADA between ARS, Weslaco (Texas) and Peoria (Illinois) and Mycotech is developing methods for mass production of Paecilomyces. Research at the University of Florida led to the discovery that a native species, P. fumosoroseus, was highly effective against not only whiteflies, but also spider mites, aphids and diamondback moths (Plutella xylostella). Under licence from the University, Thermo Trilogy (Maryland) has developed a P. fumosoroseus-based product for the European greenhouse market and has applied for US registration. In Europe, the first biopesticide is likely soon to receive EU approval. The registration dossier by the Belgian company Biobest
for the provisional approval of Paecilomyces fumosoroseus was dec-lared complete by the European Union Standing Committee on Plant Health in February, although it still needs to be approved by the European Commission.

American research has demonstrated that the entomopathogenic fungi are generally compatible with the ben-eficial insect species, and results indicate that Beauveria and a Texan Eretmocerus sp. are a promising combination. Further trials are assessing the impact of crop fungicides on Beauveria and of plant growth promoters and plant extracts (including neem biopesticides) on boosting crop defences.