LUBILOSA was the name of a research programme that aimed at developing a biological alternative to the chemical control of locusts. This name is an acronym of the French title of the programme: Lutte Biologique contre les Locustes et les Sauteriaux (biological control of locusts and grasshoppers). During its 13-year life (November 1989 to December 2002), the programme identified an isolate of an entomopathogenic fungus belonging to the genus Metarhizium and virulent to locusts, and went through all the necessary steps to develop the commercial biopesticide product Green Muscle based on its spores.
Collaborators The programme was conceived by Chris Prior and David Greathead of the International Institute of Biological Control (IIBC), a former research institute of CAB International that was based at Silwood Park in the UK at that time. IIBC requested and obtained the collaboration of the former Biological Control Centre for Africa in Cotonou, Benin, that was owned and managed by the International Institute of Tropical Agriculture (IITA). When IIBC asked for financial support from the Dutch Directoraat Generaal voor Internationale Samenwerking (DGIS: Directorate General of International Cooperation), DGIS contributed by paying for a Dutch locust expert based at the Département de Formation en Protection des Végétaux (DFPV: Department of Crop Protection Training) of the AGRHYMET Regional Centre (ARC) in Niamey, Niger, which is part of the Comité permanent Inter-états de Lutte contre la Sécheresse dans le Sahel (CILSS: Permanent Interstate Committee for Drought Control in the Sahel). Since DFPV was given its own budget within LUBILOSA, it became a full partner within the programme. For a number of years, LUBILOSA collaborated with GTZ's locust programme. Permanent collaboration was established with the crop protection agencies of Niger, Benin, Burkina Faso, Chad, Mali, Senegal and The Gambia.
Donors The LUBILOSA programme was supported financially by the following donors: CIDA: Canadian International Development Agency DFID: Department for International Development, UK DGIS: Dutch Directorate General of International Cooperation SDC: Swiss Agency for Development and Cooperation USAID: United States Agency for International Development
Over the life of the programme, these donors contributed UK£10,200,000 (approximately US$17,000,000 at the time).
Four phases The LUBILOSA programme has been funded in phases of three or four years each: First phase 1990–1992 Second phase 1993–1995 Third phase 1996–1998 Fourth phase 1999–2002
First phase The project was started late in 1989, with Chris Prior and David Greathead obtaining funding and forming a team to develop a biological means of controlling locusts and grasshoppers. While examining the various options for biological control, it soon became apparent that oil formulations of the spores of certain fungi belonging to the form phylum Deuteromycota (Anamorphic fungi) offered the most promising option. Such fungi will grow on artificial substrates and so can be mass-produced relatively quickly in large quantities. Their spores are lipophilic and therefore suspend much more readily in oils than in water. As most locust control is carried out using oil formulations at ultra-low volume (ULV) rates of application, developing an oil formulation of fungal spores enabled operators to use the same equipment for applying a chemical pesticide and maximise efficacy.
Usually, fungi are most active under moist conditions and in early attempts to use them as control agents, they were therefore applied in water formulations. However, the results in practice were often disappointing and the approach was largely abandoned during most of the 20th century. When interest in microbial control was revived, water formulations of fungi started to be used successfully in greenhouses, but the results in open fields continued to disappoint. In the 1980s, Prior discovered that some anamorphic fungi are more effective when applied in oil.
During the first phase of LUBILOSA, the technical feasibility of using such oil formulations against locusts was demonstrated in the laboratory, then field cage and "arena" trials. An extensive survey was launched in West Africa and the Arabian Peninsula to look for isolates virulent to locusts and grasshoppers, because few such isolates were available in public collections. The survey made use of a network of collaborators and came up with some 180 isolates, many of which belonged to Metarhizium acridum, some to other Metarhizium species and to Beauveria bassiana with a few records of Syngliocladium acridiorum (syn. Sorosporella). Laboratory screening of these isolates showed that M. acridum was easily the most virulent species under warm conditions (30 °C), and confirmed the selection of isolate IMI 330189 for further development. Subsequent trials in cages and arenas confirmed that oil formulations were infective even under very dry conditions. IMI 330189 and similar isolates were assigned to Metarhizium flavoviride in early papers, then Metarhizium anisopliae var. acridum, but are now described as Metarhizium acridum (Driver & Milner) J.F. Bisch., Rehner & Humber.
Second phase During phase 2, the oil formulation was tested in the field and shown to be effective. Field trials proved to be difficult to carry out on such highly mobile insects as grasshoppers and locusts. With locusts, fixed plots, unless several square kilometres in size, cannot be used, so it is necessary to follow hopper bands. Although there were many locust infestations during this phase, of both desert locusts (Schistocerca gregaria) and brown locusts (Locustana pardalina), the chemical treatment teams were also very active and little headway was therefore made in developing the techniques necessary to measure the effect of Metarhizium on unrestrained locusts. However, it was possible to scale up treatments against the variegated grasshopper (Zonocerus variegatus) in the forest zone and against Sahelian grasshoppers, especially the Senegalese grasshopper (Oedaleus senegalensis).
Contacts were made with several internationally renowned biopesticide producers to ascertain whether they would be able and willing to reliably meet LUBILOSA's spore requirements during phase 3, but none were able to make that commitment. Accordingly, part of the IITA station in Cotonou was converted into a spore production unit and the human and technical capacity enhanced to meet the expected spore requirements. The facility proved an excellent research unit and made it possible to refine realistic technical specifications and to test production, contamination control, spore separation, drying and packaging.
Third phase
Field trials The LUBILOSA team continued to look for opportunities to test Metarhizium against locusts and grasshoppers. Field trials were conducted against Senegalese grasshoppers, African rice grasshoppers (Hieroglyphus daganensis), variegated grasshoppers, Sahelian tree locusts (Anacridium melanorhodon), brown locusts and desert locusts. These trials demonstrated that Metarhizium at a dosage of 50 g/ha could reduce grasshopper populations and those of tree locusts by 80-90% within two to three weeks. Population level control was more difficult to achieve with other locusts, but a significant effect was demonstrated on hopper bands notwithstanding the great difficulties in tracking individual bands. Some trials against Senegalese grasshoppers included the chemical insecticide fenitrothion for comparison. Because of its rather short persistence, this chemical turned out to be much less effective than commonly believed. It was highly effective in producing an immediate sharp reduction in population densities, but continued hatching and remigration into the treated plots caused these densities to start increasing again after less than a week, until they reached or exceeded the original levels after about two to three weeks. Metarhizium, on the other hand, reduced population levels at a slower rate, but maintained low levels for at least one and a half months. The half-life of its spores on the vegetation was estimated at more than 7 days under rainy season conditions in the Sahel and, of course, some conidia must survive between seasons.
Grasshopper behaviour In the course of these field trials, it became clear that locusts and grasshoppers are able to detect that they have become infected and alter their behaviour accordingly. In particular, they spend more time basking in the sun, even in the middle of the day. Uninfected grasshoppers also bask in the sun when their body temperature is below the preferred 38-40 °C. However, detailed investigations showed that infected grasshoppers increase their body temperature to up to 4° higher. This effect has been dubbed “behavioural fever” and is thought to slow down the infection just like a fever in warm-blooded animals does. This basking behaviour poses a potential problem for the fungus. The latter's preferred temperature is 28-30 °C and it stops growing above 35°. Measurements of body temperatures in the field have shown that the basking behaviour reduces the time available for growth by many hours per day. Under certain conditions, this enables the insects to lay at least one egg pod before dying. Healthy locusts and grasshoppers, especially females, take some time after fledging to reach maturity and lay eggs, during which time they build up sufficient fat to produce up to three egg pods. However, infected ones usually dispense with this fattening period and go on to produce eggs immediately after fledging. This generally wears them out so much that they die after laying one pod. So even if Metarhizium does not kill before or shortly after fledging, it usually allows the formation of only a single egg pod and thereby reduces the fecundity of infected females.
(Eco)toxicology During phase 3, the programme started investigating the environmental impact of the chosen Metarhizium isolate (IMI 330189). Standard toxicological tests carried out by certified laboratories showed that the strain had practically no effect on mammals, birds and fish (e.g. for rats >2000 mg active ingredient per kg of body weight). Other toxicological and ecotoxicological tests were performed by the LUBILOSA team. An important goal was to show that its isolate only infected locusts and grasshoppers. Infection tests were therefore performed on a large number of invertebrates. The great majority could not be infected even by high doses. However, some species, like termites, honey bees, certain parasitoid wasps and silk worms, did get infected under laboratory conditions. Further experiments were carried out under semi-field and full field conditions, which showed that only silk worms suffered significant infection under those conditions. The conclusion from all these tests was that the LUBILOSA strain of Metarhizium was incapable of infecting vertebrates and infected only a small number of insect species under artificial conditions. The available studies indicate that under natural conditions, the isolate only infects species belonging to the Acrididea and the domesticated silk worm Bombyx mori.
Mass production