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Project Wolbachia - Singapore Progresses To Phase 3 Field Study At Two Extended Sites in February 2019

30 Jan 2019

Phase 2 of Project Wolbachia – Singapore has yielded improved results, and the project is progressing to Phase 3 to target Aedes aegypti mosquito population reduction over larger areas, to determine the sustainability of mosquito suppression.

Singapore, 30 January 2019 – The expanded area of the study sites and the high floor releases (in addition to ground floor releases) of Wolbachia-Aedes mosquitoes has yielded improved results in Phase 2 of the Project Wolbachia – Singapore field study. At the Nee Soon East study site, the urban disease-transmitting Aedes aegypti mosquito population was suppressed by about 80 per cent, while a 50 per cent suppression was observed at the Tampines West study site. The results thus far show that a larger release site yields better results, and community effort in keeping the mosquito population low will help to enhance the effectiveness of the Wolbachia technology. From February 2019, both the Tampines West and Nee Soon East study sites will be expanded, to determine if the suppression of Aedes aegypti mosquito populations can be sustained in larger areas.

Project Wolbachia – Singapore

2       The National Environment Agency (NEA) is evaluating the use of male Wolbachia-Aedes mosquitoes to further suppress the Aedes aegypti mosquitoes in the community. When these released male Wolbachia-Aedes mosquitoes mate with urban Aedes aegypti females in the community, the resultant eggs do not hatch. The regular release of male Wolbachia-Aedes mosquitoes over time is thus expected to reduce the viability of Aedes aegypti eggs, and bring about a gradual reduction in the urban Aedes aegypti mosquito population.

3       The Phase 2 field study, conducted from April 2018 to January 2019 by NEA’s Environmental Health Institute (EHI), was designed to address the challenges posed by Singapore’s high-density and high-rise urban landscape. The release of male Wolbachia-Aedes mosquitoes from high floors at residential blocks, in addition to releases at the ground floor, has enabled a better distribution of the male Wolbachia-Aedes mosquitoes at Singapore’s high-rise residential blocks. Their releases at adjoining buffer zones also helped in reducing the infiltration of urban Aedes aegypti mosquitoes into the release sites. The release strategy led to about 80 per cent suppression of the dengue-transmitting Aedes aegypti mosquito population at the Nee Soon East study site (Annex A, Figure 1). 

4       At the Tampines West study site, about 50 per cent suppression of the dengue-transmitting Aedes aegypti mosquito population was observed (Annex A, Figure 2). This reduced suppression rate could be due to the shorter duration of the release period, multiple mosquito breeding habitats found coinciding with the field study period, and other environmental factors. The mosquito breeding habitats found were largely domestic containers, plastic receptacles, and planters in homes and along corridors. 

5       Professor Neil Ferguson, Director of the MRC Centre for Global Infectious Disease Analysis at Imperial College London, said: “It is important that the community removes mosquito breeding habitats, so that fewer male Wolbachia-Aedes mosquitoes have to be released. Areas with more mosquito breeding habitats will require more male Wolbachia-Aedes mosquitoes to be released, for the technology to be successful.” 

Dengue Expert Advisory Panel (DEAP)

6       NEA’s Dengue Expert Advisory Panel (DEAP) has concluded that the Phase 2 field study has met its objective in developing methodologies to distribute the male Wolbachia-Aedes mosquitoes more uniformly in order to elicit better population suppression. Following a three-day review meeting, the Panel strongly supported progression of the project to Phase 3, which will see expansion of the study sites to determine if the suppression of mosquito populations can be sustained in larger areas.

7       The Panel (see Annex B for list of members), comprising experts from Australia, Singapore, the United Kingdom and the United States of America, with specialised knowledge on vector-borne diseases, entomology, epidemiology, public health and mathematical modelling, met from 28 to 30 January 2019, to review the data gathered during Project Wolbachia – Singapore field studies. The Panel also visited NEA’s mosquito production facility at Neythal Road, to observe the production and quality control of Wolbachia-Aedes mosquitoes (see Annex C for mosquito production facility). 

Phase 3 Field Study

8       Results from the Phase 2 field study indicate that larger release areas improve the effectiveness of mosquito suppression. The Phase 3 field study will thus expand the Tampines West and Nee Soon East study sites, to suppress the mosquito population over larger areas. The overall aim of the Phase 3 field study is to determine if the suppression of mosquito populations can be sustained in larger areas. In the long-term, this will require strategies which reduce the number of male Wolbachia-Aedes mosquitoes being released in an area. 
 
9       Mosquitoes will be released around residential blocks and along common corridors, and will not be released in homes. Please see Annex D for the location maps and boundaries of the two study sites. Residents at the field study sites may notice an increase in mosquitoes during the field study. However, the male mosquitoes released will not bite or transmit disease.

10      Professor Ary Hoffmann, Laureate Professor at the University of Melbourne, commented: “The data that have been generated through the different phases of the field study are essential in developing an effective long-term mosquito suppression programme for Singapore. DEAP fully endorses the systematic approach adopted by NEA, in building invaluable knowledge for deployment of the technology, especially in a tropical, urbanised high-rise landscape.” 

Innovation and Technologies

11      EHI will continue to work with Orinno Technology Pte. Ltd., and with Verily Life Sciences through February, and use and test their automation solutions to enhance the efficiency and quality in the production and release of the Wolbachia-Aedes mosquitoes. 

12      Professor Duane Gubler, Chairman of DEAP and Founding Director of the Emerging Infectious Diseases Programme at Duke-NUS Medical School, said: “The results of Project Wolbachia – Singapore will help us understand how this technology can be expanded to the varied ecological settings seen in Singapore and in other dengue endemic countries. The data generated in Singapore will be critical to upscaling the technology to the region and the world.”

Collective Effort Needed to Fight Dengue

13      NEA is very encouraged by the support of the public, as well as by the continual interest in Project Wolbachia – Singapore. Prior to and during the Phase 3 field study, NEA will be providing more information to residents and local stakeholders at the study sites.

14       The co-operation and support from residents and local stakeholders at the selected sites will be crucial in ensuring the success of Project Wolbachia – Singapore. Residents do not have to do anything differently, but continue to carry out mosquito control procedures and practise the 5-Step Mozzie Wipeout as normal.

15       NEA hopes to use the Wolbachia technology to complement its existing vector control efforts, to reduce the risk of Aedes-borne diseases, such as dengue and Zika.

16       Members of the public can find out more about Wolbachia technology by visiting www.nea.gov.sg, or contacting NEA at 1800-CALL-NEA (1800-2255 632) if they have any enquiries. We are thankful for the support from residents, various stakeholders and volunteers.

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For more information, please contact us at 1800-CALL NEA (1800-2255 632) or submit your enquiries electronically via the Online Feedback Form or myENV mobile application.


ANNEX A

MOSQUITO POPULATION SUPPRESSION IN THE PHASE 1 AND PHASE 2 FIELD STUDIES

Wolbachia technology has the potential to suppress the Aedes aegypti mosquito population in the community. 

In the Phase 1 field study, at sites where the male Wolbachia-Aedes mosquitoes were released, much fewer Aedes aegypti adult mosquitoes were found. At these sites, half of the collected Aedes aegypti mosquito eggs did not hatch, providing strong indication that the released Wolbachia-Aedes males had successfully competed with the urban Aedes males and mated with some of the urban Aedes aegypti females.

Impact of male Wolbachia

Fig. 1. Phase 1 – Infographic showing the impact of male Wolbachia-Aedes mosquitoes on the urban Aedes aegypti mosquito population at the study sites


In the Phase 2 field study, the expanded area of the release sites and the high floor releases of male Wolbachia-Aedes mosquitoes (in addition to ground floor releases) have led to a better outcome. The results show that a larger release site yields better results, and community efforts to remove mosquito breeding habitats will enhance the effectiveness of the technology.

Impact of male Wolbachia on urban Aedes

Fig. 2. Phase 2 – Infographic showing the impact of male Wolbachia-Aedes mosquitoes on the urban Aedes aegypti mosquito population at the study sites

 


ANNEX B

 

DENGUE EXPERT ADVISORY PANEL (DEAP) MEMBERS

Member

Designation

Prof. Duane Gubler

(Chairman)

Epidemiologist (Vector-borne Diseases), Entomologist; Emeritus Professor and Founding Director of Emerging Infectious Diseases Programme, Duke-NUS Medical School, Singapore

Prof. Ary Hoffmann

(Member)

Entomologist; Laureate Professor, Bio21 Institute, Department of Bioscience, University of Melbourne, Australia

Prof. Neil Ferguson

(Member)

Mathematical epidemiologist; Director of MRC Centre for Global Infectious Disease Analysis, School of Public Health, Imperial College London, United Kingdom

Assoc. Prof. Timothy Barkham

(Member)

Clinician; Senior Consultant in Clinical Microbiology, Laboratory Medicine, Travellers’ Health and Vaccination Clinic, Tan Tock Seng Hospital (TTSH); Chair of NEA’s Bioethics Review Committee, Singapore

Assoc. Prof. Vernon Lee

(Member)

Epidemiologist (Public Health); Director, Communicable Diseases Division, Ministry of Health, Singapore

Prof. Christl Donnelly

(New member)

Statistical epidemiologist; Professor of Applied Statistics, Department of Statistics, University of Oxford; Professor of Statistical Epidemiology, School of Public Health, Imperial College London, United Kingdom

 

ANNEX C

NEA’S MOSQUITO PRODUCTION FACILITY AND PHASE 3 FIELD STUDY

  • The National Environment Agency’s (NEA’s) mosquito production facility, for Wolbachia-Aedes aegypti mosquitoes, has been in operation since September 2016.
  • About 50 researchers from NEA’s Environmental Health Institute (EHI) have been working at the facility and field study sites to prepare for the Phase 3 field study, which will involve expansion of the previous two study sites. The Phase 3 field study aims to determine how to sustain suppression of the dengue-transmitting Aedes aegypti mosquito population at the release sites. The number of male Wolbachia-Aedes aegypti mosquitoes released will be adjusted to maintain or further suppress the low Aedes aegypti mosquito population at the previous areas, and reduce the higher mosquito population at the new areas.

Community Engagement for Project Wolbachia – Learning Journey

  • Since 2012, NEA has consulted with and engaged local academia, medical and healthcare professionals, school teachers and students, Non-Governmental Organisations (NGOs), Grassroots Leaders and members of the public, on the potential of Wolbachia technology in Singapore, through more than 100 seminars and outreach events.
  • NEA continues to actively engage the community, and the public is welcome to visit our mosquito production facility.
  • To-date more than 700 visitors, including residents from the study sites as well as other stakeholder groups (e.g. teachers and students, residents and Grassroots Leaders from the study sites, Dengue Prevention Volunteers, media personnel, scientists and researchers, staff of private companies, and international visitors and experts) have visited NEA’s mosquito production facility, as part of the Project Wolbachia Learning Journey.
  • The Project Wolbachia Learning Journey brings visitors through the various processes involved in the production of Wolbachia-Aedes aegypti mosquitoes, and the behind-the-scenes scientific work carried out by EHI researchers. Visitors get to see how fit and competitive mosquitoes are reared; how male and female mosquitoes are separated; and how innovative engineering solutions are developed to increase productivity and ensure quality of mosquitoes.
  • The processes demonstrated at the facility visit include:
  1. Hatching of mosquito eggs, and their development into larvae and pupae
  2. Sorting of male and female pupae by size
  3. Packaging of pupae and adult male mosquitoes for release at field study sites
  4. Quality control of male Wolbachia-Aedes aegypti mosquitoes with a DNA fingerprinting tool (polymerase chain reaction, PCR)
  5. Laboratory testing to determine the hatch rate of Aedes mosquito eggs collected in the field study
  • For members of the public who would like to visit NEA’s mosquito production facility, please contact us at Project_Wolbachia@nea.gov.sg. Please note that scheduled slots are limited.

 

INFORMATION ON PROJECT WOLBACHIA LEARNING JOURNEY

Tour of Room 1: Larvae Rearing Room

How Are Wolbachia-Aedes aegypti Mosquitoes Produced?

  • This part of the learning journey will show visitors how male Wolbachia-Aedes aegypti mosquitoes are produced at NEA’s mosquito production facility.
  • Quality control at the mosquito production facility is critical for success of the study.
  • Breeding protocols are optimised to ensure that Wolbachia-Aedes aegypti mosquitoes bred in the laboratory are fit and competitive when compared with urban Aedes aegypti mosquitoes – developing the best conditions for rearing the mosquitoes from eggs through to adults, e.g. optimal temperature, space and food for larval growth and development.
  • Visitors will have a chance to see the following processes, including innovations by EHI:

  • Hatching of eggs from Wolbachia-Aedes aegypti mosquitoes. These eggs are derived from matings between male and female Aedes aegypti mosquitoes with Wolbachia (which are compatible and eggs can thus hatch).

  • Optimal number of larvae (4,000) is introduced into a tray of water using an in-house developed Larvae Counter.
  • Larvae are then fed fish food to rear them into pupae. Expanding to large-scale production, a Multi-layer Automated Feeding System is being optimised to automatically deliver a fixed amount of feed into multiple mosquito larval rearing trays simultaneously.
  • Male Wolbachia-Aedes aegypti mosquitoes are separated from female mosquitoes at the pupal stage, as male pupae are smaller than female pupae and can thus be sorted by size.
  • The male Wolbachia-Aedes aegypti pupae are passed through a Pupae Counting and Dispensing Module to accurately count and dispense the desired number of male Wolbachia-Aedes aegypti pupae into each release container.
  • The recently developed mosquito release mechanism – “Mosquito Launcher”, is a lightweight and portable device, allowing field operators to carry more mosquitoes in a single trip and thus reducing release duration.

Fig 1 Aedes aegypti mosquito eggs and larvae

 Fig. 1 Aedes aegypti mosquito eggs and larvae


Tour of Room 2: Adult Rearing Room

How Does the Wolbachia Suppression Strategy Work?

  • Upon their release, male Wolbachia-Aedes aegypti mosquitoes will seek out and mate with female urban Aedes aegypti mosquitoes (which do not carry Wolbachia). These female mosquitoes will then seek a blood meal, as they need the protein in blood for production of their eggs. However, the eggs from these females that have mated with male Wolbachia-Aedes aegypti mosquitoes will not hatch as they are not viable. The continuous release of male Wolbachia-Aedes aegypti mosquitoes in the field will lead to suppression of the urban Aedes aegypti mosquito population in the community.
  • As Wolbachia is passed on from female Wolbachia-Aedes aegypti mosquitoes to their offspring, the females are then used to rear subsequent generations of Wolbachia-Aedes aegypti mosquitoes. Whilst the females are used for generating more Wolbachia-Aedes aegypti mosquitoes, the males are useful for Aedes aegypti mosquito population control.
  • This part of the learning journey will show visitors how Wolbachia technology works, and how the mating between male Wolbachia-Aedes aegypti mosquitoes and female urban Aedes aegypti mosquitoes can lead to suppression of the urban Aedes aegypti mosquito population.
  • Visitors will have a chance to see the following:
  • Transformation of aquatic male pupae into flying male adult mosquitoes.
  • Gender differentiation of male and female Aedes aegypti adults (males have bushy antennae, females do not).
  • Male mosquitoes do not bite.
  • Mating between male Wolbachia-Aedes aegypti mosquitoes and female urban Aedes aegypti mosquitoes.
  • Blood feeding and egg production of female urban Aedes aegypti mosquitoes. 
  • Eggs produced from the mating between male Wolbachia-Aedes aegypti mosquitoes and female urban Aedes aegypti mosquitoes do not hatch.

Fig 2 Mass rearing of mosquito adults in cages

 Fig. 2 Mass rearing of mosquito adults in cages

 

Tour of Room 3: Quality Control Room

How is Quality Control Carried Out?

  • Quality control at the mosquito production facility is critical for success of the study. We need to ensure that Wolbachia is always successfully passed on from female Wolbachia-Aedes aegypti mosquitoes to their progeny. This is important to ensure that each of the male Wolbachia-Aedes aegypti mosquitoes in the colonies to be used for suppression carries Wolbachia.
  • This part of the learning journey will show visitors how quality control is carried out, including how we ensure that the male Aedes aegypti mosquitoes to be released have Wolbachia in them. As part of the quality control process to ensure that only male Wolbachia-Aedes aegypti mosquitoes are in the containers holding mosquitoes to be released, the adult mosquitoes are further screened to detect any female mosquitoes. 
  • Visitors will have a chance to see the following:
  • Specialised equipment used in NEA’s quality control processes (e.g. homogeniser, PCR machine to detect Wolbachia in mosquitoes). PCR (polymerase chain reaction) is a molecular technique that allows detection and quantification of Wolbachia genetic material, even at minute levels.
  • Sperm-filled spermathecae of the female mosquitoes after mating.
  • Ovaries of female Aedes aegypti mosquitoes (what these look like with and without eggs).

 

ANNEX D

LOCATION MAPS OF STUDY SITES

Fig 3 Location map of Tampines West study site

Fig. 3. Location map of Tampines West study site
Legend

 

Fig 4 Location map of Nee Soon East study site

Fig. 4. Location map of Nee Soon East study site

Legend

 

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