Genetic Biocontrol for Schistosomiasis: Developing a Gene Drive to Target the Snail Biomphalaria glabrata

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1 Genetic Biocontrol for Schistosomiasis: Developing a Gene Drive to Target the Snail Biomphalaria glabrata John Teem Senior Scientific Program Manager, ILSI jteem@ilsi.org TM

2 Genetic Biocontrol for Schistosomiasis: Developing a Gene Drive to Target the Snail Biomphalaria glabrata Research Conducted at the Florida Department of Agriculture and Consumer Services Division of Aquaculture John Teem

3 Nonindigenous Snails Acting as Intermediate Hosts for Parasites Affecting Human Health Achatina fulica Pomacia maculata Biomphalaria glabrata Angiostrongylus cantonesis

4 Gene Therapy for Snails Achatina fulica Pomacia maculata Biomphalaria glabrata 1. Lentivirus vectors for gene transfer 2. CRISPR/cas9 genome editing 3. CRISPR/cas9 formatted into a gene drive Schistosoma mansoni

5 Biomphalaria glabrata as a Model System Genetic Biocontrol Genome is sequenced. Biomphalaria glabrata Bge snail embryonic cell line available for transfection experiments. Genetic biocontrol strategies developed with Biomphalaria glabrata will be instructive for other snail species.

6 Schistosomiasis is transmitted by human contact with contaminated fresh water (lakes and ponds, rivers, dams) inhabited by snails carrying the parasite. Intervention Praziquantel is the primary form of treatment. A single dose of the drug has been shown to reduce the burden of infection and severity of symptoms. Reinfection occurs if people enter the contaminated water again. Control of snails, the intermediate host for the schistosome parasite, is needed to break the transmission cycle.

7 Not all Biomphalaria snails are susceptible to schistosome infection. Could resistant snails be added to a susceptible population, increasing the frequency of resistance genes in the population?

8 A Gene Drive Utilizing cas9 to Drive the Introgression of a Parasite Resistance Gene Parasite Resistance (R) Parasite Sensitivity (S) R S Cas9 Cas9 Cas9 R S R R R R R R R R

9 Modeling Introgression of a Recessive Parasite Resistance Gene Introduced into A Susceptible Snail Population No Gene Drive Gene Drive No Introgression Complete Introgression Requirements for a Biomphalaria gene drive: 1. Lentivirus vectors for infection of germline 2. Functional CRISPR/cas9 in snail cells 3. CRISPR/cas9 formatted into a gene drive

10 The ovotestis of Biomphalaria is located at the very tip of the shell spire. Lentivirus can be injected at this site to transfer genes to the germ line. Ovotestis of Biomphalaria

11 PCR Analysis of Gene Transfer Into Biomphalaria glabrata Snails Bg HSP70 promoter cas9 Inject ovotestis Mate Snail Collect Eggs for Hatching Progeny LV-cas9(lucA) Detect cas9 gene in progeny by PCR cas PCR from Hatchlings 20 Negative Control (No DNA) 1,21 Positive Control

12 Can the Gene Drive Risks be Mitigated? Pros: Species specific Self perpetuating Feasible for most organisms that reproduce sexually Can be used to produce a Daughterless eradication strategy Eradication possible in large systems Cons: Off-target cutting (cas9) Once started, it can t be stopped Cannot be limited geographically Could endanger pest snail species in their native range Is it possible to stop the gene drive after a fixed number of generations?

13 Limiting Gene Drives 1. Stop a first gene drive by targeting its components with a second gene drive. 2. Configure the gene drive as a Daisy Chain. (Place the components of the gene drive on separate chromosomes so they become separated over time) 3. Limit the number of generations that the gene drive organism can sustain. (manipulate telomerase to cause germline cell division to eventually cease?) Finding a way to turn off a gene drive will be an important area for future research.

14 Important Questions to Answer Regarding Gene Drives to Control Snails How efficient are gene drives in actual practice? Will there be selection against a gene drive that impairs introgression? How will other organisms be affected by genetic modification of the target snail population? What is the risk of causing extinction of the target snail species?

15 Conclusions Transgenic Biomphalaria glabrata snails expressing cas9 will provide a means to modify the snail genome for the purpose of genetic biocontrol. Biomphalaria glabrata will be a good model system to test the efficacy of a gene drive and better define the associated risks. A gene drive to target snails could be a valuable tool in reducing schistosomiasis.

16 Thank you!