Biological Control of Dengue Disease Using Symbiotic Wolbachia Bacteria: Environmentally Safe and Inexpensive Approach

• Project/Area Number: 19KK0107
• Research Category: Fund for the Promotion of Joint International Research (Fostering Joint International Research (B))
• Allocation Type: Multi-year Fund
• Review Section: Medium-sized Section 22:Civil engineering and related fields
• Research Institution: Ehime University
• Principal Investigator: 渡辺 幸三 愛媛大学, 沿岸環境科学研究センター, 教授 (80634435)
• Co-Investigator(Kenkyū-buntansha): 糠澤 桂 宮崎大学, 工学部, 助教 (20725642) ; 八重樫 咲子 山梨大学, 大学院総合研究部, 助教 (30756648) ; ガンボア・メンデス マリベット・アリアナ 愛媛大学, 理工学研究科(工学系), 助教 (30849477)
• Project Period (FY): 2019-10-07 – 2022-03-31
• Project Status: Granted (Fiscal Year 2020)
• Budget Amount: ¥18,460,000 (Direct Cost: ¥14,200,000、Indirect Cost: ¥4,260,000); Fiscal Year 2021: ¥5,850,000 (Direct Cost: ¥4,500,000、Indirect Cost: ¥1,350,000); Fiscal Year 2020: ¥6,760,000 (Direct Cost: ¥5,200,000、Indirect Cost: ¥1,560,000); Fiscal Year 2019: ¥5,850,000 (Direct Cost: ¥4,500,000、Indirect Cost: ¥1,350,000)
• Keywords: デング熱 / ボルバキア / 蚊媒介感染症 / 金ナノ粒子 / 集団遺伝解析

Outline of Research at the Start

近年，蚊からヒトへのデングウイルス感染を抑制する昆虫共生細菌ボルバキアが注目されている。本研究は，フィリピン・マニラを対象として，当地のネッタイシマカに自然感染したボルバキアを活用した安心・安価なデング熱制御技術を開発する。ボルバキアに自然感染した蚊幼虫を見つけ，施設で繁殖して都市に放虫するシステムである。この構築のため，自然感染した幼虫を金ナノ粒子で迅速・安価に選別する技術を米国・ミシガン州立大と，蚊の飛翔範囲を推定して最適放虫地点を決定する蚊ゲノム解析技術をドイツ・ベルリンゲノムセンターと開発する。蚊採取と繁殖実験はフィリピンで行い，4か国の国際研究拠点を形成する。

Outline of Annual Research Achievements

【研究項目１　金ナノ粒子によるボルバキア感染検査法の開発】：ボルバキアに特異的な配列を有する磁石プローブを使ったボルバキアDNAを濃縮する手法の開発を行った。
【研究項目２　蚊飛翔パターンの集団遺伝解析】：マニラの1ｋｍ×2kmの小さい範囲を対象にして200m幅の幹線道路で採取した376個体のネッタイシマカ成虫から抽出したDNAを使ってマイクロサテライトマーカーを使ったDNA多型実験を行った。その結果，道路跨ぐ蚊の移動が統計的有意に制限されていることが分かった。また，DNA多型データに基づく起源生息地を推定した結果，起源生息地から幹線道路を渡って採取された蚊は26個体（6.9％）に止まることも分かった。次世代シークエンサーを使った217個体のddRAD解析を行い，ゲノム全体から約1万の1塩基多型領域を検出することができた。
【研究項目３　ボルバキア感染蚊の繁殖と感染の検証】：メトロマニラで採取した378匹のシマカ属成虫と509匹の幼虫からボルバキアをPCRにより検出した。幼虫からは検出されなかったが，15匹の成虫からボルバキアを検出することができた。また，リアルタイムPCRにより蚊体内のボルバキアDNA濃度を定量する実験プロトコルを開発することもできた。
【交流実績】新型コロナウイルス感染症流行の影響により，海外共同研究者を訪問することができなかった。しかし，オンライン会議により，主にフィリピンやアメリカの共同研究者らと打ち合わせを継続的に行った。

Current Status of Research Progress

3: Progress in research has been slightly delayed.

Reason

新型コロナウイルス感染症流行の影響で海外渡航が制限されたため，海外共同研究者を訪問した形での共同研究を実施することができなかった。ただし，オンラインによる打ち合わせを継続して，それぞれの国において共同研究に必要な研究活動を実施した。

Strategy for Future Research Activity

新型コロナウイルス感染症への対応のため，海外への渡航が制限されているので，オンラインミーティングを行うことで共同研究を粛々と遂行すると共に，渡航制限が解除された後に速やかに交流を再開すべく準備を進めたい。

Research Products

• [Int'l Joint Research] デラサール大学/フィリピン大学マニラ校(フィリピン)
• [Int'l Joint Research] ライプニッツ淡水生態学・内水 漁業研究所(ドイツ)
• [Int'l Joint Research] ミシガン州立大学(米国)
• [Int'l Joint Research] デラサール大学/フィリピン大学マニラ校/フィリピン大学ロスバニョス校(フィリピン)
• [Int'l Joint Research] ライプニッツ淡水生態学・内水漁業研究所(ドイツ)
• [Int'l Joint Research] ミシガン州立大学(米国)
• [Journal Article] Intracellular Interactions between Arboviruses and Wolbachia in Aedes aegypti (2021)
  • Author(s): Reyes, J. I. L., Y. Suzuki, , T. M. Carvajal, M. N. M. Munoz, and K. Watanabe
  • Journal Title: prerprints Volume: n.a.
  • DOI: 10.20944/preprints202104.0151.v1
  • Open Access / Int'l Joint Research
• [Journal Article] Early Detection of Dengue Fever Outbreaks Using a Surveillance App (Mozzify): Cross-sectional Mixed Methods Usability Study (2021)
  • Author(s): Herbuela, V. R. D. M., T. Karita, T. M. Carvajal, H. T. Ho, J. D. Lorena, R. A. Regalado, G. D. Sobrepe?a, and K. Watanabe
  • Journal Title: JMIR Public Health and Surveillance Volume: 7 (3)
  • DOI: 10.2196/19034
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] The influence of roads on the fine-scale population genetic structure of the dengue vector Aedes aegypti (Linnaeus) (2021)
  • Author(s): Regilme, M. A. F., T. M. Carvajal, A. C. Honnen, D. M. Amalin, and K. Watanabe
  • Journal Title: PLOS Neglected Tropical Diseases Volume: 15(2)
  • DOI: 10.1371/journal.pntd.0009139
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Underdiagnosis does not account for the decrease in dengue cases in Bandung, Indonesia (2021)
  • Author(s): Faridah, L., S. Ekawardhani, H. L. Wiraswati, N. Fauziah, F. R. Rinawan, W. Mayasarid, and K. Watanabe
  • Journal Title: Systematic Reviews in Pharmacy Volume: 12(1) Pages: 1340-1342
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Spotted fever group Rickettsiae (SFGR) found in a follow-up investigation of in ticks after the occurrence of Japanese spotted fever human case in Niigata Prefecture, Japan (2021)
  • Author(s): Arai, R., M. Hayashi, M. Kato, J. Aoki, A. Nishida, K. Watanabe, C. Hirokawa, S. Ikeda, T. Tamura, K Watanabe, M. A. F. Regilme, M. O. Sato, and M. Sato
  • Journal Title: Scientific Reports Volume: 11 Pages: 2595-2595
  • DOI: 10.1038/s41598-021-81587-9
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Wing Geometry and Genetic Analyses Reveal Contrasting Spatial Structures between Male and Female Aedes aegypti Populations in Metropolitan Manila, Philippines (2021)
  • Author(s): Carvajal, T. M., D. M. Amalin, and K. Watanabe
  • Journal Title: Infection, Genetics and Evolution Volume: 87 Pages: 104676-104676
  • DOI: 10.1016/j.meegid.2020.104676
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Surveillance of Dengue Virus in Individual Aedes aegypti Mosquitoes Collected Concurrently with Suspected Human Cases in Tarlac City, Philippines (2020)
  • Author(s): Balingit,J. C., T. M. Carvajal, M. Saito-Obata, M. Gamboa, A. D. Nicolasora, A.K. Sy, H. Oshitani, and K. Watanabe
  • Journal Title: Parasites & Vectors Volume: 13 Pages: 594-594
  • DOI: 10.1186/s13071-020-04470-y
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Fine-Scale Population Genetic Structure of Dengue Mosquito Vector, Aedes aegypti in Metropolitan Manila, Philippines (2020)
  • Author(s): Carvajal, T. M., K. Ogishi, S. Yaegashi, L. F. T. Hernandez, K. M. Viacrusis, H. T. Ho, D. Amalin, and K. Watanabe
  • Journal Title: PLOS Neglected Tropical Diseases Volume: 4
  • DOI: 10.1371/journal.pntd.0008279
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Comparative population genetic structure of two Ixodidae ticks (Ixodes ovatus and Haemaphysalis flava) in Niigata Prefecture, Japan (2020)
  • Author(s): Regilme, M. A. F., M. Sato, T. Tamura, R. Arai, M. O. Sato, S. Ikeda, M. Doi, K. Tanaka, M. Gamboa, M. T. Monaghan, and K. Watanabe
  • Journal Title: bioRxiv Volume: n.a.
  • DOI: 10.1101/862904
  • Open Access
• [Journal Article] Understanding Hurdles in the Implementation of Health Information System (HIS) Through an Institutional Framework: A Case of Bandung, West Java Province, Indonesia (2020)
  • Author(s): Faridah, L, F. Rinawan, N. Fauziah, W. Mayasari, A. Dwiartama, K. Watanabe
  • Journal Title: bioRxiv Volume: 17(5) Pages: 1795-1795
  • DOI: 10.3390/ijerph17051795
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] An Integrated mHealth App for Dengue Reporting and Mapping, Health Communication, and Behavior Modification: Development and Assessment of Mozzify (2020)
  • Author(s): Herbuela, V. R. D. M., T. Karita, M. Francisco, and K. Watanabe
  • Journal Title: JMIR Formative Research Volume: 4(1)
  • Peer Reviewed / Int'l Joint Research
• [Journal Article] Depressive and Anxiety Symptoms among Pediatric In-patients with Dengue Fever: A Case-control Study (2019)
  • Author(s): Herbuela, V.R.D.M., F.S.d. Guzman, G. Sobrepe?a, A.B.F. Claudio, A.C.V. Tomas, C.M.A-d. Reyes, M.M. Teodoro, and K. Watanabe
  • Journal Title: International Journal of Environmental Research and Public Health Volume: 17 (1) Pages: 99-99
  • DOI: 10.3390/ijerph17010099
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Knowledge, Attitude and Practices on Dengue Fever among Paediatric and Adult In-patients in Metro Manila, Philippines (2019)
  • Author(s): Herbuela, V. R. D. M., F. S. d Guzman, G. D. Sobrepena, A. B. F. Claudio, A. C. V. Tomas, C. M. A-d Reyes, R. A. Regalado, M. M. Teodoro, and K. Watanabe
  • Journal Title: International Journal of Environmental Research and Public Health Volume: 16 (23) Pages: 4705-4705
  • DOI: 10.3390/ijerph16234705
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Detection of Wolbachia Infection in Field-collected Mosquito Vector, Aedes aegypti in Metropolitan Manila, Philippines (2019)
  • Author(s): Carvajal,T. M., K. Hashimoto, R. K. Harnandika, D. M. Amalin and K. Watanabe
  • Journal Title: Parasites & Vectors Volume: 12 (1) Pages: 361-361
  • DOI: 10.1186/s13071-019-3629-y
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Journal Article] Is Rapid Dengue Test Necessary at Primary Health Centre? A Study In Bandung City (2019)
  • Author(s): Faridah, L., N. Fauziah, S Ekawardhani, Riyadi, and K. Watanabe
  • Journal Title: Journal of Clinical and Diagnostic Research Volume: 13 (7)
  • DOI: 10.7860/jcdr/2019/41059.13004
  • Peer Reviewed / Open Access / Int'l Joint Research
• [Presentation] Inference of Spatial and Temporal Dataset Granularity Effect to the Performance of Machine Learning in Dengue Forecast (2021)
  • Author(s): Micanaldo Ernesto Francisco, Thaddeus M. Carvajal, and Kozo Watanabea
  • Organizer: 2nd JSPS Core-to-Core Symposium: Center of Excellence in Health Risk Assessment for Adaptation to Climate Change
  • Int'l Joint Research / Invited
• [Presentation] The influence of roads on the fine-scale population genetic structure of the dengue vector Aedes aegypti (Linnaeus) (2021)
  • Author(s): Maria Angenica Fulo Regilme, Thaddeus Carvajal , Ann?Christin Honnen, Divina Amalin, Kozo Watanabe
  • Organizer: 2nd JSPS Core-to-Core Symposium: Center of Excellence in Health Risk Assessment for Adaptation to Climate Change
  • Int'l Joint Research / Invited
• [Presentation] The Influence of Water Supply Conditions and Water Storage Containers toward Vector Prevalence and Abundance in Makassar City, Indonesia (2021)
  • Author(s): La Ane, R., I. Wahid, A. Susilawaty, H. Ishak, E. Ibrahim, M-C Chiu, V. R. D. M. Herbuela, and K. Watanabe
  • Organizer: 2nd JSPS Core-to-Core Symposium: Center of Excellence in Health Risk Assessment for Adaptation to Climate Change
  • Int'l Joint Research / Invited
• [Presentation] The Time Varying Effect of Weather Variables and its Interaction on Dengue Occurrence During 2009-2015, in Bandung City, Indonesia (2021)
  • Author(s): Agustian D, Jaya IGM, Permana YS, Faridah L, Agoes R, Watanabe K
  • Organizer: 5th International Conference on Climate Change 2021 (ICCC 2021)
  • Int'l Joint Research
• [Presentation] Comparative population genetic structure of Ixodes ovatus and Haempahysalis flava in Niigata Prefecture (2020)
  • Author(s): Maria Angenica Fulo Regilme , Megumi Sato , Tsutomu Tamura , Reiko Arai , Marcello Otake Sato , Sumire Ikeda , Masaya Doi, Kohki Tanaka , Maribet Gamboa , Michael T. Monaghan, Kozo Watanabe
  • Organizer: The 43rd Annual Meeting of the Molecular Biology Society of Japan (MBSJ online)
  • Int'l Joint Research
• [Presentation] Molecular Genetic Approaches to the Assessment of Dam Impact on Stream Macroinvertebrates and Dengue Virus Surveillance of Vector Mosquitoes (2020)
  • Author(s): Kozo Watanabe
  • Organizer: 47th Philippine Society of Biochemistry and Molecular Biology (PSBMB) Annual Convention
  • Int'l Joint Research / Invited
• [Presentation] Wing Geometry and Genetic Analyses Reveal Contrasting Spatial Structures between Male and Female Aedes aegypti (L.) (Diptera: Culicidae) Populations in Metropolitan Manila, Philippines (2020)
  • Author(s): Carvajal, Thaddeus; Amalin, Divina and Kozo Watanabe
  • Organizer: The Pan-African Mosquito Control Association (PAMCA) - Vectorbase Students and Young Investigators Vector Meeting
  • Int'l Joint Research
• [Presentation] The effect of roads on the population genetic structure of the dengue vector, Aedes aegypti (2020)
  • Author(s): Maria Angenica Fulo Regilme, Thaddeus Carvajal , Ann?Christin Honnen, Divina Amalin, Kozo Watanabe
  • Organizer: 2020 Annual Virtual Meeting of the American Society of Tropical Medicine and Hygiene
  • Int'l Joint Research
• [Presentation] Dengue Disease Dynamics are Modulated by Combinatory Influences of Precipitation and Landscapes: A Machine Learning ? based Approach (2020)
  • Author(s): Micanaldo Ernesto Francisco, Thaddeus M. Carvajal, Masahiro Ryo, Kei Nukazawa, Divina M. Amalin and Kozo Watanabea
  • Organizer: Water and Environment Technology Conference 2020 Online (WET2020-Online)
  • Int'l Joint Research
• [Presentation] The Status of Biological Control Approaches Against Aedes-borne Diseases: How Far Are We in Effectively Controlling this Mosquito Vector? (2020)
  • Author(s): Thaddeus Carvajal, Jayson Capistrano, Phoebe Diez, Paolo Pacheco, Jerica Isabel Reyes, Howell Ho, Divina Amalin and Kozo Watanabe
  • Organizer: Rundown International Webinar: Environmental Health Emerging Diseases
  • Int'l Joint Research / Invited
• [Presentation] "Development and assessment of Mozzify app: an integrated mhealth for real-time Dengue reporting and mapping, health communication and behavior modification reporting and mapping, health communication and behavior modification" (2019)
  • Author(s): Herbuela, Von Ralph Dane M., Karita Tomonori, Francisco, Micanaldo, Watanabe Kozo
  • Organizer: 1st JSPS Core-to-Core Symposium: Center of Excellence in Health Risk Assessment for Adaptation to Climate Change
  • Int'l Joint Research
• [Presentation] Designing and Developing a Nanobiosensor for the detection of important microbials in Ae. aegypti (2019)
  • Author(s): Carvajal Thadeus, Alocilja Evangeline, Divina Amalin and Watanabe Kozo
  • Organizer: 1st JSPS Core-to-Core Symposium: Center of Excellence in Health Risk Assessment for Adaptation to Climate Change
  • Int'l Joint Research / Invited
• [Presentation] Detection of Wolbachia in Field-collected Dengue Vector Mosquito, Aedes aegypti in the Philippines (2019)
  • Author(s): Kozo Watanabe
  • Organizer: 1st JSPS Core-to-Core Symposium: Center of Excellence in Health Risk Assessment for Adaptation to Climate Change
  • Int'l Joint Research / Invited
• [Presentation] A Bioclimate-Based Maximum Entropy Model for Comperiella calauanica Barrion, Almarinez & Amalin (Hymenoptera: Encyrtidae) and Its Utility in Predicting the Occurrence of Aspidiotus rigidus Reyne (Hemiptera: Diaspididae) in the Philippines (2019)
  • Author(s): Almarinez, BLM., Fadri, MJA., Lasina, R., Carvajal, TM., Watanabe, K., Legaspi, JC., and DM Amalin
  • Organizer: 1st JSPS Core-to-Core Symposium: Center of Excellence in Health Risk Assessment for Adaptation to Climate Change
  • Int'l Joint Research
• [Funded Workshop] International Symposium (Webinar) of jointly organized by Ehime University and De La Salle University - Philippines "Usable Science Resulting in Impact" (2020)