Periodic Reporting for period 1 - WolAntiS (Using antisense inhibition to understand Wolbachia symbiosis and antiviral protection)
Reporting period: 2019-03-01 to 2021-02-28
This proposal aimed at addressing key technological and knowledge deficits. The first objective was to develop an antisense strategy to target the function of Wolbachia genes. Second, to determine which Wolbachia genes are involved in animal-microbe symbiosis and antiviral protection.
This work is important due to the prevalence of Wolbachia in nature and its interaction with insects (eg. pests, pollinators) and pathogenic filarial nematodes causing human disease. Understanding Wolbachia-host interaction will allow us to understand insects and nematodes better, improve current efforts to control vector-borne and filarial-caused diseases.
Subsequently, 12 bp synthetic antisense molecules, spanning the ATG and ribosome-binging sites, were designed for two Wolbachia genes: wsp and ftsZ. These genes were selected, as antibodies against them were readily available, and we confirmed their robustness in the silencing protocol. We conjugated the antisense molecules with the cell-penetrating peptides targeting Wolbachia, and applied the onto Wolbachia-infected cells. The knock-down of selected Wolbachia genes was tested with the Western blot using Drosophila tubulin as a loading control. This work, together with the similar protocol using antisense molecules conjugated to cholesterol analogues, has led to the development of the first antisense inhibition protocol for selective and specific targeting of Wolbachia genes.
Although the results of the project were not deployed yet, they were disseminated to both, academic and non-academic audiences in a series of activities. During the project, I participated in the following conferences and scientific events:
1) The British Microbiology Society Annual Conference 2019, Belfast, UK, 08 – 11/04/2019
2) Royal Entomological Society – Special Interest Group: Insects: Infection & Immunity, Online Conference, 24-25 September 2020
3) Virtual Symbiosis Seminar, organized by prof. Greg Hurst and Prof. Nicole Gerardo (https://docs.google.com/spreadsheets/d/1DHkn2CjRXz9QbJPcSDuibYmn6e1khhgd2xPIxXjXVMY/edit#gid=943392522) 7th July 2020
The results of the project have been written up in the following publications/preprints:
- Chrostek E§, Martins NE§, Marialva MS, Teixeira L (2020) Wolbachia-conferred antiviral protection is determined by developmental temperature. bioRxiv, doi: 10.1101/2020.06.24.169169. (citations: 3)
§ Contributed equally
- Corbin C, Jones JE, Chrostek E, Fenton A, Hurst GDD (2020) Thermal sensitivity of the Spiroplasma-Drosophila hydei protective symbiosis: The best of climes, the worst of climes. Molecular Ecology doi: 10.1111/mec.15799.
The approaches and results of the action were disseminated further to non-scientific audience by the fellow invited to act as:
- An Invited author of the article: “Wolbachia – basic science and applications of an antiviral bacterium” for Microbiology Today (https://bit.ly/3eyZ03T) a magazine by the Microbiology Society with articles for all readers, including parliamentarians and policy-makers (06/08/2019).
- An Expert interviewed by “The Scientist Magazine”, a professional magazine providing coverage of recently published scientific papers (19/07/2019.
This project has many potential interdisciplinary impacts, which include: (i) Host-microbe interactions: Identification of mechanisms deployed by Wolbachia to interact with the hosts. This will enable design of new drugs and treatments for bacterial and viral diseases. (ii) Ecology: Dissecting consequences of the mechanisms of symbiosis and antiviral protection on insect population structure and infected individuals spread. This will allow to predict more accurately the outcomes of the current efforts using Wolbachia to limit the spread of dengue and Zika. (iii) Cell biology: Understanding of Wolbachia integration into host cells physiology and its engagement of host machinery. Wolbachia’s inconspicuousness within the host cell may uncover new biological pathways and phenomena. (iv) Vector control: Improvement of current efforts to control vector-borne diseases through understanding of the mechanism of virus-blocking. This will allow generation of new transgenic insects refractory to pathogens for the management of diseases in humans, livestock, and plants. (v) Biomedical sciences: Identification of novel Wolbachia drug targets for the treatment of filarial diseases and design of novel antifilarial treatments. (vi) Agriculture: Antiviral protection of beneficial insects and elimination of pest insects and worms.