TM

Genetic Biocontrol for

Schistosomiasis:

Developing a Gene Drive

to Target the Snail

Biomphalaria glabrata

John Teem

Senior Scientific

Program Manager, ILSI

jteem@ilsi.org

Genetic Biocontrol for Schistosomiasis:Developing a

Gene Drive to Target the SnailBiomphalaria glabrata

Research Conducted at the Florida Department of Agriculture and Consumer ServicesDivision of Aquaculture

John Teem

Achatina fulica

Nonindigenous Snails Acting as Intermediate Hosts for

Parasites Affecting Human Health

Pomacia maculata Biomphalaria glabrata

Angiostrongylus cantonesis

Achatina fulica

Gene Therapy for Snails

Pomacia maculata Biomphalaria glabrata

Schistosoma mansoni

1. Lentivirus vectors for gene transfer2. CRISPR/cas9 genome editing3. CRISPR/cas9 formatted into a gene drive

Biomphalaria glabrata as a Model SystemGenetic Biocontrol

Biomphalaria glabrataGenome is sequenced.

Bge snail embryonic cell line availablefor transfection experiments.

Genetic biocontrol strategiesdeveloped with Biomphalaria glabratawill be instructive for other snailspecies.

Schistosomiasis is transmitted by

human contact with contaminated fresh water (lakes and ponds, rivers, dams) inhabited by snails carrying the parasite .

InterventionPraziquantel 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 hostfor the schistosome parasite, is neededto break the transmission cycle.

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?

A Gene Drive Utilizing cas9 to Drive the Introgression of a Parasite Resistance Gene

Cas9Cas9Cas9

R

R R R R R R R R

S

Parasite Resistance (R)

Parasite Sensitivity (S)

R S

Modeling Introgression of a Recessive Parasite Resistance Gene Introduced into

A Susceptible Snail PopulationNo Gene Drive Gene Drive

No Introgression Complete Introgression

1. Lentivirus vectors for infection of germline2. Functional CRISPR/cas9 in snail cells3. CRISPR/cas9 formatted into a gene drive

Requirements for a Biomphalaria gene drive:

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

Ovotestis ofBiomphalaria

2-19 PCR from Hatchlings20 Negative Control (No DNA) 1,21 Positive Control

10 11 12 13 14 15 16 17 18 19 20 21

Mate

Snail

Collect Eggs

for Hatching

Progeny

Injectovotestis

Detect cas9 gene in progeny by PCR

cas9Bg HSP70promoter

1 2 3 4 5 6 7 8 9

LV-cas9(lucA)

PCR Analysis of Gene Transfer Into Biomphalaria glabrata Snails

cas9

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?

Can the Gene Drive Risks be Mitigated?

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 chromosomesso 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.

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 thetarget snail species?

Transgenic Biomphalaria glabrata snails expressing cas9 will provide a means to modify the snail genome for thepurpose 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.

Conclusions

Thank you!