Bacterial Assassins

Wednesday April 13, 2005

Executives

Bacterial assassins The battle against malaria is now being fought at the cellular level with mass-produced bacteria being drafted into the war. Maore Ithula reports on ICIPE’s pilot trials with the Bacillus thuringiensis bacterium in Kenya. Every passing second, malaria kills a child somewhere in the world, according to World Health Organisation (WHO) estimates. Researchers at the International Centre for Insect Physiology and Ecology (ICIPE) estimate that malaria infection rate in Kenya can be reduced by at least 30 per cent by using larvacides. The procedure involves killing mosquito larvae by poisoning them using a bacterium called Bacillus thuringiensis var. israelensis (Bti). Bti, which is cultured and massively produced in factories throughout the world, is now being produced at ICIPE’s Duduville facility.

Eric Zhang, a technician with Chinese company Kernel, which is partnering with ICIPE on the project, at the Duduville demonstration factory.

Dr Jean Nguya Maniania, an insect pathologist at the centre, points out that pilot trials conducted in Mwea, Suba, Kisii and the coast region, have shown 100 per cent destruction of mosquito larvae at their breeding grounds before fresh re-invasion by the insects. "Larvaciding has been in existence for nearly 30 years in the western world with a 90 per cent efficiency (rate)," Maniania says. "Bti control exploits the vulnerability, concentration and accessibility of larvae at the breeding point. This is the Dr Jean Ngaya Maniania stage when the insect is naturally most vulnerable." The anopheles mosquito (which carries the malariacausing pathogen) breeds continuously in still clear water. A single bite by an infected mosquito is sufficient to make one suffer a bout of malaria within 10 days. The insect is mature enough to mate and lay eggs at least eight days after developing into an adult. It has a lifespan of between 14 to 30 days. It breeds faster in temperatures above 26 degrees Centigrade, taking a minimum of seven days to release an adult. The incubation period is twice as long in cooler environments. Since mosquitoes feed on water microorganisms at the larval stage, Bti is introduced into this medium in liquid or pellet forms. "Bti is a stomach toxin to the juvenile mosquito," Maniania says. "Each Bti organism can produce five different microscopic protein toxins or pro-toxins that are packaged into a protein container or a crystal. The crystal form of

the toxin (delta-endotoxin) is swiftly ingested by the larvae. The five proteins are released into the insect’s gut where they work alone or together to break down the gut wall. This leads to paralysis and eventually death for the mosquito larvae. In less than a day the insect dies of starvation and/or inflammation in the gut." Bti is produced in large quantities in America and Europe where mosquitoes are only killed because they are a nuisance. The bacteria are also modified to control other pests in these countries. Currently the technology has brought malaria under control in China’s Yangtze River Valley. However, Maniania says, importation of the finished product to control malaria in Kenya is uneconomical. It is cheaper to produce Bti locally using cheap raw materials. "Besides, it is another avenue of job creation." Local production will reduce the cost of the bacteria by a third of its price were it to be imported. "What we have here (at ICIPE) is actually a demonstration factory as way of showing the government, members of the public and the local business community how easy, beneficial and cheap it is to produce the product commercially," he explains. Locally, the bacteria can be produced through a fermentation process using a mixture of powdered fresh water sardines (omena) and soya beans. Three tons of raw materials can produce one ton of Bti with a potency of 1,000 International Toxic Units per millilitre (ITU/ml), which can treat 400,000 square metres of water area for seven days. The mixture is soaked in water and boiled at 120 degrees Centigrade to sterilise it. It is then cooled to room temperature and Bti is introduced. The mix is then left to ferment for 24 hours before it is passed through a centrifuge to separate the bacteria from the medium. The bacteria can also be formulated into pellets. "We appreciate that it is not possible to eradicate the existence of mosquitoes in totality," says Dr John Githure, head of the Human Health Division at ICIPE. That said, the country needs to embrace an integrated vector management approach where biological control methods such as larvacide are used to supplement the existing methods. Currently, the popular approaches that are in place include the use of bed nets, indoor residue sprays and environmental management. The latter involves the drainage of any unnecessary small man made water receptacles and holes or filling them altogether. Studies conducted by ICIPE have shown that more than 90 per cent of mosquito breeding sites are made by humans.

Indoor residual spraying is the most disastrous method of pest control, because the chemicals used persist in the environment. The pesticides also kill other non-target organisms, thus destroying the ecosystem. For this reason, the use of DDT has been proscribed. "For a long time governments in the tropics where malaria is prevalent have emphasised curative remedies of malaria, rather than controlling the source," Githure notes. As malaria has resisted all available treatments, it has become necessary to have more input in preventative approaches. "In this line we need to recognise that the mosquito has developed various survival mechanisms over the years. The most fundamental is that the insect has adapted to biting humans (who are the primary source of their meal) when they are asleep. The creature will in the same way realise that bed nets are a threat to its existence and therefore resort to feeding before bedtime. We need to open more fronts in this war just in case," Githure says. Bti is a naturally-occurring bacterium that was discovered and isolated by Professor Joel Margalit of Ben-Gurion University-Israel in 1977. The researcher had bumped into the bacterium while investigating other microorganisms found in a pool of rainwater near the institution. The US Centre for Development Research (CDR) has funded Israeli-Less Developed Countries (LDC) teams to adapt and extend Bti technology throughout the world. They are currently in Thailand, Sri Lanka, the Philippines, Kenya and Guinea. Available information on the advancement of this technology indicate that CDR-sponsored biotechnologists have successfully cloned the Bti toxin gene, expressed it in more persistent bacterial strains, adapted it to local conditions and introduced a variety of other improvements in the USA and EU. "In this regard the next stop is to increase the longevity of Bti in the regional climate," Githure says. Although the scientists are confident that controlling mosquitoes at the larval stage is one of the most effective ways, the plan is dogged by the short lifespan of Bti in the tropics. Like other microorganisms, Bti is swiftly killed by excess UVradiation in the tropics. Consequently Bti can only survive for between three and five days as opposed to a maximum of 10 days in temperate climates. Under the prevailing conditions, Bti needs to be applied once a fortnight while in temperate regions only a single application is needed each month.

"ICIPE is now working closely with the Valent Bioscience Company of the USA (the world’s largest Bti producer) in developing a variety that is able to resist UV-destruction," says Githure. The researcher says that the programme has passed through environmental impact assessment together with efficacy tests and expresses optimism that the government will soon clear the way for the official launch of Bti use in the country. The demonstration factory at Duduville was constructed two years ago at a cost of more than Sh40 million. It was funded through a partnership between the Kenya and Chinese governments, ICIPE and the Chinese company Kernel. The factory, which has a capacity to produce 700 tons of Bti annually, is the first of its kind on the continent. The Bti produced is sufficient to serve most of the Common Market for Eastern and Southern Africa (Comesa) member countries with the product. The facility can also produce other Bti strains — from a gene bank of over 200 strains — capable of controlling pests in the horticultural sub-sector. The facility can also produce other microorganisms for use in plant pest and disease vector controls, including bacteria such as Bacillus sphaericus, and fungi such as Metarhizium anisopliae. In the mid 1990s, ICIPE and Oy GAC of Finland imported limited amounts of a Bti strain that was employed to kill filth flies in refuse dumps and latrines. The pilot trials were used by the UNHCR to treat sanitary facilities in refugee camps in Northern Kenya, Ethiopia and the Kibera slums. "With little genetic modification, several Bti strains can be produced that are capable of controlling other crop pests," Maniania says. "These improvements are in intense horticultural production application in Europe and the Americas." Maniania appeals to investors with brewing experience to embrace the plan as another method of business expansion, job creation and enhancing their social responsibility. "Bti is efficient in dipteran control (control of insects belonging to the order Diptera) in the continent as has long been confirmed by its effectiveness when it was first used alongside pesticides in the war against the black fly in West Africa a decade ago," Maniania explains. "Bti is specific on its attack of organisms. It kills only two-winged insects at the larval stage with abandon. In this case, it only tackles the black fly (which causes river blindness) and the mosquito because they are the only dipterans that breed in water masses — a place that is accessible and convenient to apply the bacterium." Bti is ideal because it is a highly biodegradable protein

that does not persist in the environment for long and it does not attack any other organisms. This character makes it environmental and user friendly, Maniania points out. Since it is uneconomical for individuals to apply Bti in homesteads, the researchers say ICIPE will need to partner with community-based organisations, the government and NGOs in making the plan a reality. "Apart from being a real nuisance to humans and being a vector of malaria, the mosquito plays only an insignificant role on the food chain in the ecosystem," Githure says. "Sparrows and lizards eat it. Period. Otherwise it is one organism that any conservationist will agree that we all can do without." The use of Bti in pest control, Maniania says, is the future for agricultural production as the EU market (on which Kenya mainly relies as an outlet for her goods) puts more stringent measures in ensuring that finished products — especially for horticultural goods — meet minimum residual levels (MRLs) for pesticides. Not everyone is as enthusiastic about the possibility of Bti use in Kenya. Some consumer rights advocates are opposed to the adoption of this technology without taking into account safety issues. These issues, says Dorcas Mwangi, the Project Officer in charge of Food, Health and Trade with the Consumer Information Network of Kenya, arise from the introduction of genetically modified cotton and maize containing B thuringiensis genes by the Kenya Agricultural Research Institute. Last year, Kenya imported Bt cottonseed from the United States for field trials at KARI farms in central Kenya. Bt maize varieties were first planted in Kenya a few months ago in an open quarantine site under KARI’s Insect Resistant Maize for Africa (IRMA) project. Both Bt-cotton and Bt-maize have B thuringiensis genes that make them resistant to stalk-borers. "We cannot rule out the possibility of the Bt in maize making other pests resistant to other strains of Bt in the environment," Mwangi says. As it may be another few years before either Bti is commercially produced or Btcrops are harvested in farmers’ fields, there is still time to ensure that all these issues are addressed. Home

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