DENGUE FEVER

Dengue is a viral disease transmitted bymosquitoes, sometimes referred to as“breakbone fever”. “Classic” dengue is an

excruciatingly painful illness with high fever, agonizingbone, muscle and eye pain and an intensely-itchingrash that can extend from the head to the soles of thefeet. The acute phase lasts less than two weeks butvictims may suffer months of post-infection symptomsthat include mental depression, physical exhaustion,distressing eye problems and even temporarybaldness. About 5% of clinical cases are classed as“severe” and are potentially fatal (dengue shocksyndrome/dengue haemorrhagic fever).

The global burden of dengue is formidable: nearlyhalf the world’s population lives in dengue-proneregions, there are an estimated 390 million infectionsper year, and incidence is continually increasing. Atpresent there is no dengue vaccine and there are noeffective antiviral drugs; mosquito control is the onlyavailable counter-measure.

The principal vector of dengue is the mosquito Aedesaegypti, a species that originated in the African forestbut has adopted the human domestic environment. Inits original habitat it breeds in natural water-holdingcontainers – holes in trees and rocks, leaf axils, fruithusks, etc. – and feeds exclusively on roaming bandsof monkeys. In the peri-domestic environment, wehumans are the perfect hosts. Instead of tree holes weprovide an abundance of artificial containers: water-barrels, old buckets, flower-pot saucers, discardedtyres and so on. Instead of an environment rich inpredators we provide safe, clean quarters for them tohide; clothes cupboards are a favorite. And of course,an abundant supply of fresh blood on tap.

Control of Ae. aegypti dates back to the beginning ofthe 20th century, when it was demonstrated thatyellow fever – a terrifying disease caused by a virusclosely related to dengue – is transmitted by themosquito. The approach, conducted as a military-styleoperation, was simple and effective: infested

containers were either discarded or covered. The firsttest of the method, in Havana, Cuba, was a resoundingsuccess: the disease was wiped out in four months.Within a few years, similar results had been achievedin most major seaports of the Americas.

In the 1950s, “source reduction” was replaced by anew approach: infested sites were treated with amagic powder, DDT. The method was incredibly easyto apply and within 10 years the species was declared“eradicated” in 22 countries of the Americas.Unfortunately, for reasons that are now well-known,DDT fell out of favour and from the mid-1960sonwards there was swift return of the mosquito,quickly followed by dengue, in much of thehemisphere. At the same time, new technology wasdeveloped to dispense insecticidal aerosols by hand-held or truck-mounted “fogging machines”. Thesemachines are a familiar sight in most regions wheremosquitoes are a nuisance or public health problem.

Fogging machines are highly visible, make lots ofnoise and produce an unpleasant smell. They can beeffective against mosquitoes in open areas but havevery little impact on Ae. aegypti, mainly because themosquito rests in sheltered places, inaccessible by theaerosol. Nevertheless, despite their proven lack ofefficacy and their high cost, the machines are widelyused, particularly during outbreaks of dengue. Thebest that can be said is that the passing of a machinereassures the public that their government isresponding to a problem.

From the 1960s onwards, the burgeoning increase inthe size and population density of many tropical cities,coupled with an increasingly pervasive “throw-awaysociety”, furnished an expanding paradise for Ae.aegypti and an enormous increase in the prevalenceand incidence of dengue. By the 1980s it was clearthat conventional interventions were far too costly andimpractical for most governments to consider. Inresponse, the idea arose that educational campaignscould be used to transfer responsibility for source-

The need for innovation…

…to counter the global dengue pandemic

reduction to the population as a whole; oncehouseholders would be aware of their responsibilities,government efforts could be limited to informationand enlightenment. The popular phrase was “BottomUp” rather than “Top Down”.

Unfortunately, despite numerous sociological studiesand valiant, often highly creative efforts by localcommunities, there is no evidence that the strategyhas ever had a sustainable impact on the mosquitopopulation. Thus, today, we are helpless in the face ofan ever-burgeoning mosquito problem, exacerbatedby the rapidly increasing mobility of the virusengendered by global air-travel.

The reality is that apart from the early source reductionprogrammes and the eradication campaign,epidemiologically meaningful control has rarely beenachieved and never sustained. Quasi-militarystrategies are no longer acceptable, the heady days ofDDT-optimism have passed, reliance on aerosols hasnever been the answer and community-basedapproaches have proven fruitless. To deny all this isostrich science.

Once we confront this reality we can accept the needto innovate, to explore new and imaginativeapproaches, from the simplest, low-tech ideas to novelmethods of using insecticides, transgenics, cellularpathogens and other pioneering technology. None ofthese approaches is likely to succeed on its own: weneed to devise ways to synergise them with oneanother. Moreover, to develop these synergies wemust eliminate important gaps in our knowledge ofmosquito biology and our understanding of dengueepidemiology

Finally, dengue vectors will never be controlled on ashoe-string. Even if the community assumes a majorportion of responsibility, support is still needed atvarious levels of government. For this there must becontinuity in management, immune from frequent andabrupt changes in leadership. The need for high-

quality vector control personnel implies a soundcareer structure and the recognition that continuedvigilance can never be terminated; as a colleagueonce remarked, we would never think of stoppingrubbish collections. These requirements may seemimpracticable or impossible when ranked againstmany other disease priorities but when set against thecost of repeated dengue and dengue haemorrhagicfever epidemics in terms of morbidity, mortality,medical resources, work absenteeism, impact ontourism and other criteria, the benefits of effectiveprevention are clear.

Paul Reiter, MPhil, DPhil, FRES

October, 2013

Paul Reiter is a professor of medical entomology at the Pasteur Institute in the

city of Paris, France. He is a member of the World Health Organization Expert

Advisory Committee on Vector Biology and Control. Previously he was an

employee of the Center for Disease Control (Dengue Branch). He is a Fellow

of the Royal Entomological Society. He is a specialist in the natural history,

epidemiology and control of arboviral diseases such as dengue fever, West

Nile Virus, and chikungunya.

The principal vector of dengue fever, Aedesaegypti, has only recently adapted its lifestylefrom the forests of Africa to now living in close

proximity to humans in urban and semi urban areasacross the world. During its aquatic larval stage, itthrives in small, frequently transient containers filledwith rain water – common around our homes and indisposed rubbish – and in stored water when tap wateris not available. Aedes aegypti is an anthropophilicmosquito: the female almost exclusively bites humanswhen taking a blood meal, dramatically increasingrates of virus transmission between people. Onlyfemale mosquitoes bite – they need the blood proteins

for egg maturation – whereas males only requirecarbohydrates, which they obtain from natural sugarsources such as nectar from plants and flowers.

The female mosquito that takes a blood meal from adengue virus-infected person becomes itself infectedwith the virus. About 8-10 days later, the virus spreadsto the mosquito's salivary glands and is subsequentlyreleased into its saliva while biting another host. Thefemale mosquito remains infected for life and cantransmit the virus to a new person each time she bites.

In the absence of an effective vaccine or specific

A new solution

Dr Renaud Lacroix describes Oxitec’s new approach to combat Aedes mosquitoes

The Oxitec approach. Females that mate with wild males canproduce up to 500 offspring during their life. However, if they matewith an Oxitec male the offspring die before adulthood.

therapeutic or prophylactic drugs against dengue, theonly way to tackle dengue transmission is by targetingits vector, the mosquito. Conventional mosquitocontrol relies largely on insecticides: sprayed orfogged insecticides to kill adults, larvicides spread inbreeding sites against larvae. As described byProfessor Paul Reiter in the accompanying article,these strategies do not reach all of the breeding andresting sites that Aedes aegypti favours, and have off-target ecological effects. Targeting the larval breedingsite of Aedes aegypti is a major challenge, as they areso dispersed. Widespread resistance to insecticidesfurther limits their effectiveness. These strategies havenot been able to stem the rise in dengue cases overthe years. Novel control methods are desperatelyneeded to fight this burden.The “gold standard” mosquito control method wouldhave several attributes: • Ubiquitous: ability to reach every corner wheremosquitoes breed and rest, i.e. hidden discardedcontainers, humid resting places in houses, etc…

• Safe to people: it will mostly be used in residentialareas.

• Species-specific: no off-target effects.

• Environmentally friendly: no long-lasting side effectson the environment.

• Effective: suppress Aedes aegypti population belowthe disease transmission threshold.

• Sustainable:maintain the level of control at areasonable cost.

Recent breakthroughs in genetic engineering havemade possible new control methods. Oxitec’s Aedesaegypti product is an advancement of the SterileInsect Technique (SIT), in which released radiation-sterilised male insects mate with wild females andthereby reduce the population in the nextgeneration. SIT has been widely used since the1950s against a number of major agriculture pestsand has enabled control at a continental level of theNew World screwworm and Mediterranean fruit fly.Unfortunately, mosquitoes have proven toovulnerable to the irradiation process – required forsterilisation – to compete effectively against the wildmales for mates.

Oxitec’s solution overcomes this limitation usinggenetically engineered sterile male mosquitoes

Oxitec release male mosquitoes as males do not bite

(males can’t bite or transmit diseases) whose offspringwill die. The Oxitec males, which to wild females areindistinguishable from wild males, are released inlarge numbers to out-number the wild males.Consequently, a majority of females will encounterand mate with Oxitec males and produce no viableoffspring. Sustained releases over several months willcause the local Aedes aegypti population to collapse.The releases need to be carefully planned so that theOxitec males are evenly distributed in the area; highlyinfested areas will initially require higher numbers tobe released – or other control methods can beemployed prior to the releases to provide an initialknock-down. Oxitec males provide a means ofreducing the population much further thanconventional methods, as they are able to spread inthe treated area and find the females wherever theyhide: mosquito biology turned against the mosquitoesthemselves. The releases are adaptable and can beconducted by hand, truck or even boat, depending onthe local situation.

The Oxitec lethal gene can be suppressed by achemical antidote, tetracycline, which is not significantlypresent in the environment. The males can thereforebe mass-produced at an affordable cost in a dedicatedproduction facility, which can be established locallyeither in dedicated refurbished buildings or in mobilerearing units developed by Oxitec. The technology hasalready been transferred successfully in Brazil, Cayman

Islands and Malaysia and only a few months arerequired to deliver the facility and train the new staff.

A fluorescent marker gene is also integrated in theOxitec insects to provide easy and efficientmonitoring. Indeed, a simple fluorescence microscopecan be used to monitor the presence of the gene inmosquito larvae collected in the environment, thusproviding information on the proportion of matedfemales, dispersal of the Oxitec males, or disappearanceof the gene when releases are stopped. Oxitectechnology is self-limiting as, like wild mosquitoes, themales can’t survive for long in the environment. Themarker also enables in-programme monitoring toensure releases are tailored to the local populationlevel of Aedes aegypti.

After gaining regulatory approval, Oxitec malemosquitoes were first released in the Cayman Islands in2009. The experience was a success and demonstratedthat they mated with wild females and competed wellagainst wild males. Subsequently, sustained releases ofmales in the Cayman Islands and in two locations innorth-eastern Brazil have achieved reductions in localAedes aegypti population of over 80%. In the last ofthese studies, in the village of Mandacaru in Bahia, themosquito population was reduced by 96% (see graph).These levels of effectiveness contrast with those ofconventional strategies, offering a tool that is uniquelyeffective against Aedes aegypti.

The Moscamed production unit in Juazeiro, Brazil. Courtesy of Moscamed

A vital component of Oxitec’s mosquito controlprogrammes is interaction with local communities,which needs to be adapted to local cultures and socialstructures. The projects mentioned above included

various vehicles for the dissemination of information.In Brazil, where Oxitec’s partner Moscamed led theprojects, after being advertised by flyers, posters,radio broadcasts and newspapers, open meetingswith the local residents were organised with the localauthorities to answer all questions regarding thetechnology. In the same location, house-to-house visitswere also conducted to inform and discuss with thelocals without the pressure of talking in front of acrowd: 88% of the houses were visited in one area andnone of the householders opposed the technology. Ingeneral, the local response has been very positive andcommunities show great support for the technology.

After these initial successes, the Brazilian state of Bahiacommissioned Moscamed to conduct a pilot-scalecontrol programme in the city of Jacobina, home to50,000 people. For this purpose, Moscamed isexpanding its production facility for Oxitec mosquitoes,aiming to produce several million males per week.The releases started in the north-west of Jacobina inJune 2013 and the programme will roll out across theentire city over two or three years. This programmeoffers new hope for populations throughout the tropicsthat live with the daily threat of dengue infection andprovides authorities with a new tool in their fightagainst disease vectors like Aedes aegypti.

The Oxitec lethal gene can besuppressed by a chemicalantidote, tetracycline, which isnot significantly present in theenvironment. The males cantherefore be mass-produced at anaffordable cost in a dedicatedproduction facility…

“

Oxitec releases in Mandacaru resulted in a reduction in the dengue mosquito population by 96%. Reduced release maintainedthis control even into the rainy season when neighbouring areas experience a huge increase

In 1992, at the Earth Summit in Rio de Janeiro, theworld’s nations were for the first time united in theirrecognition that mankind’s activities were having a

potentially irreversible impact on the environment andbiodiversity. More than two decades later, the UnitedNations announced that the years 2011-2020 wouldbe the UN Decade on Biodiversity, reflecting a broadinternational consensus that the decline and loss ofspecies remains one of the most pernicious threats toour natural environment.

Of course, since people first developed the capacity tomanipulate the world around them, our attempts toshape our environment according to our own needsand comforts have engendered tensions between ourlifestyle and industry, and the welfare of species whoshare that environment. And nowhere are these battlelines more clearly drawn than in our attempts tocontain, control, or altogether eradicate species thatmany of us feel we might be better off living without.

Mosquitoes are the deadliest animal on the planet.Each year, they claim the lives of some 2 millionpeople: victims of mosquito-borne diseases such asmalaria, yellow fever and dengue fever. From earliesttimes, the war that humanity has waged against thesetraffickers in pestilence has had a profound impact onour natural environment; from the drainage of themalaria-infested marshes of ancient Rome to thewidespread use of DDT around much of the tropicalworld this century.

Yet it is an often under-appreciated fact that this mostancient of mankind’s enemies (and mosquitoes wereplaguing the dinosaurs long before people wereadded to their list of victims) is nevertheless a relativenewcomer in many of the areas where it is todayproving most troublesome.

Take the dengue mosquito, Aedes aegypti, as a primeexample. Thought to have originated in central Africa,its global spread over the last few decades has beenremarkably rapid. An urban-dweller, which thrives in

densely-populated human habitation, its fortunes haverisen as inexorably as the high-rise apartment blocks inrapidly-expanding towns and cities; so much so that itis now endemic throughout the world’s tropicalregions (see below).

Aedes aegypti is not alone in embarking on a globalmarch. The Asian tiger mosquito, Aedes albopictus –aegypti’s partner-in-crime where dengue transmissionis concerned – has likewise been spreading at analarming rate, capitalising in particular on the globaltrade in used tyres to colonise territories as diverse asRussia, La Réunion and Italy. Environmentalists inEurope face a deluge of rogue immigrants, which arethreatening native species and the livelihoods offarmers. A changing climate, and an era of globaltrade and travel, has resulted in the unprecedentedmobility, not only of our own species, but that of manyothers who have hitch-hiked upon our transportnetworks. These ‘rogue’ species add a new dimensionto the challenge of protecting human health andwelfare, alongside that of preserving naturalecosystems.

Faced with a threat as severe as that of the denguemosquito, inaction is unthinkable. Recent estimates

Squaring the circle in pest control

Protecting people while preserving our environment

Aedes aegypti has spread from Africa around the world,aided mainly by the movement of humans and freight. Thedengue virus has followed though not all countries reportdengue cases. The WHO estimate that 2.5bn people are atrisk from dengue.

place the number of annual dengue cases at 350million1; more than three times the figure previouslyestimated by the World Health Organization. Withincreased consciousness of the importance ofbiodiversity, however, comes a need for interventionsthat are precise, targeted, limited, and reversible;calculated to minimise wider species impacts, andcause the smallest possible ecological disturbance.

Evaluated against this scorecard, Oxitec’s novelapproach to pest control is uniquely well-suited. Itinvolves the release of male mosquitoes modified topass on a lethal gene to their offspring. Any wildfemale that mates an Oxitec male will fail to produceprogeny capable of surviving and breeding, thusreducing the population. Only male mosquitoes arereleased, which don’t bite. Crucially, from theperspective of environmental impact, the males willonly mate with females of their own species. In doingso, they deliver a precisely targeted form of control.

After mating with a female, the released males will die,as will their offspring: there is no persistence in theenvironment. If it remains true that any attempt tocontrol or contain invasive species risks leaving holes inthe delicate fabric of a habitat’s ecosystem, then Oxitec’ssolution nonetheless provides a means to elegantly un-

pick the rogue thread, with minimum risk of unravellingthe complex weave of wider ecological networks.

Solutions like this are urgently needed if we are tocombat the dual threat of invasive pest species andthe decline in biodiversity. More traditionalapproaches, such as the use of insecticides, willcontinue to play an important role. But chemicalcontrols almost invariably have ‘off-target’ effects onother species, and are faced with concerns overhuman health impacts: for this reason manyjurisdictions, such as the EU, are re-evaluating whichagents and what manner of use they consideracceptable. Biological controls, too, such asdeployment of natural predators, can have unintendedconsequences for the habitats in which they aredeployed. Two decades of study on biodiversityteaches that the use of all pest control approachesshould be governed with caution; moreover, thattraditional methods may be both safer and moreeffective when used in combination with alternativeapproaches.

This is the niche in which Oxitec’s method can play avital role. Other alternative approaches – trapping,breeding site removal, community interventions –need also be brought into play. Above all, effectiveenvironmental monitoring to assess the impact of anyintervention will be a central part of a comprehensiveand more environmentally conscious ethos of pestcontrol.

Today, the global disease-burden resulting frommosquito-borne illnesses may already exceed 600million people each year; by the close of the UN’sDecade of Biodiversity, that number could beconsiderably higher. There are no straightforwardsolutions to meeting the threat of invasive, disease-carrying pests while managing the parallel challengesto biodiversity that might result. Yet neither are thesechallenges insoluble: by embracing new technology,making use of the full range of tools at our disposal,and by judging potential solutions without prejudice,there is no reason why the needs of humanity andthose of our environment cannot find common ground– in this decade and beyond.

1 Bhatt et al. 2013 The global distribution and burden of dengue. Nature.

Michael Conway, PhD student

Larvae that have an Oxitec male as a parent display afluorescent colour when viewed under a fluorescentmicroscope. Using the marker allows Oxitec scientists notonly to monitor the efficacy of the approach but todemonstrate there is no persistence of the released males orgene in the environment

“Today, dengue ranks as the mostimportant mosquito-borne viral diseasein the world. Everywhere the humanand economic costs are staggering”

Dr Margaret Chan, 2012 Director General, World Health Organization

Dengue is currently the fastest-spreading humandisease, driven by increasing levels ofurbanisation and international trade that

facilitate the proliferation of the disease vectormosquitoes. This fuels demand for effective andenvironmentally acceptable mosquito control solutions.

However, conventional insecticides are beingwithdrawn due to environmental or human healthconcerns and resistance is developing in the targetinsect populations. In fact, for the dengue vectorAedes aegypti, current control options are simply noteffective enough. We find ourselves in a position ofincreasing need with decreasing solutions. There isnow the real prospect of being left ‘empty-handed’with no viable alternative for controlling some of theworld’s most devastating pest species.

The pressure is on to find novel technologies that candeliver long-term protection from vector-bornediseases, while minimising the ecological footprintassociated with pest management practices. Oxitechave developed a control strategy that offers levels ofAedes aegypti control that have not been possiblethrough conventional means.

To appreciate the context and significance of this newtool, it helps to understand a little of the evolution ofpest control approaches.

Historically, agricultural pest management practiceshave focused almost exclusively on efficacy andeconomics, with compromises between ‘how much itcosts’ and ‘how well it performs’. Over-reliance onfavoured insecticides has inevitably shortened theireffective life-span through the development ofresistance. Contemporary approaches attempt tocombine complementary management techniques(chemical, biological, physical, cultural and genetic)into a more effective and sustainable systemsapproach, known as integrated pest management.

For control of vectors of human disease, such as

Aedes aegypti, the situation is altogether different.New, effective active ingredients have been few andfar between, dramatically limiting the control optionsavailable. Many control programmes still rely onconventional chemistries, such as synthetic pyrethroids,some of which have been in use since the 1970s.Chemical applications targeting adult life-stagesfrequently come from these older, broad-spectrumchemical classes with poor environmental profiles. Atthe same time, the efficacy of products acting onwater-borne larvae is limited by our inability to findand treat a high enough proportion of breeding sites.To date, the desperately narrow range of tools thathave been available to vector control teams has madeeffective long-term control of Aedes aegyptipractically impossible.

For the success of future Aedes aegypti control it willbe important to maintain a focus on communityengagement that seeks to educate and empower thepublic, reducing the number of potential breedingsites within local environments. Although sustainingimpetus and public activity levels has previously beena significant hurdle to providing long-term benefits inthis manner, community involvement offers a firmfoundation for an integrated approach to diseasevector control.

The current toolset of insecticides and publiceducation has been insufficient to halt the globalproliferation of Aedes aegypti and the associatedincrease in dengue fever to over 350 million casesper year. A holistic approach can deliver sustained,environmentally friendly control and Oxitec’stechnology is the final piece of the mosquito controlpuzzle.

Oxitec has conducted a series of suppression trialsagainst Aedes aegypti, to reduce wild populations inareas of dengue incidence by 80% to 96%, andrecently sustained this level of population suppressionthrough an entire high season, with minimal releasesof Oxitec male mosquitoes.

A holistic approach to vector management

Dr Kevin Gorman describes the implementation ofcontemporary vector control

Furthermore, the Oxitec mosquito is safe,environmentally friendly and practical to use:• The males cannot bite and so do not transmitdisease;

• Targets only Aedes aegypti;• Does not persist in the environment;• Built-in marker for simplicity of monitoring;• Ability to tailor to pest infestation level.

These strengths come together with existing tools ina complementary system that offers a sustainablesolution to public health problems that havepreviously been unmanageable, revolutionisingvector control.

Truly sustainablemanagement of a pest,whether in a public health,agricultural or veterinarycontext, has repeatedlyproven problematic withouta cohesive approach.Adaptation of pests tochallenging environments isan on-going process and, inresponse, managementstrategies need to beflexible and dynamic.Oxitec’s approach is tailoredto pest infestation levels soareas of greatest infestationreceive correspondinglyhigher rates of control,minimising redundancy andmaximising efficiency.

Oxitec has developed a programme of control forAedes aegypti that is safe, widely applicable,environmentally friendly, cost-efficient, and highlyeffective. The approach is underpinned by area-widemosquito monitoring based upon retrieving samplesof eggs from the wild population; the data from whichare used to assess and adjust the range of controlmethods being employed. A fundamental componentis community-based breeding site removal, which notonly helps to minimise mosquito habitats but alsoraises levels of public awareness. For large mosquitopopulations, initial short-term use of insecticides canbe utilized to provide a rapid ‘knock down’. Long-term suppression of the population is then providedby repeated releases of Oxitec’s genetically sterilisedmale insects that are capable of sustained andsubstantial suppression of the pest population, withpotential for local elimination and protection fromre-infestation.

Invasive species such as Aedes aegypti are generallyhighly adaptable and prolific when conditions arefavourable, which makes them such successful colonisersand renders them difficult to control. The prospect ofmaking headway against this globally important diseasevector is now a genuine one, with potential for enormoussocietal benefits. Best use of current and new vectorcontrol tools will be to employ them in strategiccombinations to exploit the strengths of each.

Dengue mosquitoes can breed in any water containingvessel, community participation is essential to reduce thenumber of breeding sites that are available.

Holistic Approach different but complimentary features areprovided by the four central elements

Oxitec Ltd. 71 Innovation Drive, Milton Park, Abingdon, UK OX14 4RQtel: +44 (0) 1235 832 393 fax: +44 (0) 1235 861 138 email: info@oxitec.com