UNIVERSITY OF CALIFORNIA IRVINE MALARIA INITIATIVE

(UCIMI)

Year 3 and 4 updates 

Hard copy Safe Drop Box https://webfiles.uci.edu/sentelle/UCIMI/Project%20Plan%20and%20Gantt

/sentelle/UCIMI/Project Plan and Gantt

UNIVERSITY OF CALIFORNIA, IRVINE, MALARIA INITIATIVE (UCIMI)

PROJECT PLAN 20190227

Executive summary:

The mission of the University of California, Irvine, Malaria Initiative (UCIMI) is to promote the discovery and development of novel science for the goal of malaria eradication. This mission is accomplished by providing the necessary intellectual, resource and infrastructure support to test novel genetics-based, sustainable technologies to prevent malaria transmission. The featured activities are to develop and field-test genetically-engineered strains of malaria vector mosquitoes in collaboration with scientists and public health personnel from disease-endemic countries.

Vector population modification strategies (also known as population replacement or alteration) employ genes designed to interfere with malaria parasite transmission coupled with highly efficient gene-drive systems. These will play a crucial role in the malaria eradication agenda by providing resistance to parasite and competent vector reintroduction and allow resources to be focused on new sites while at the same time providing confidence that treated areas remain malaria-free. Our goal is to conduct a field trial of a population modification strain with the objective of local malaria elimination in Africa. We also will help build capacity that allows local and regional scientists and public health personnel to adapt and deploy population modification strategies. Strategic development is needed in (1) the development of population modification mosquito strains, (2) field-site selection, (3) trial design and implementation, and 4) community engagement.  The UCIMI provides the organizational structure to successfully complete these activities. Working hand-in-hand with thought-leaders, scientists, public health personnel government, we expect to lay the foundation for eradicating a disease that affects hundreds of millions of people globally every year.

TABLE OF CONTENTS

Executive summary…

Introduction and background…    

Goals and activities…

Organizational Structure of UCIMI…

Project Plan Annotation…

OBJECTIVE 1: Project Administration…

Specific Aim 1.1 Recruit Project Personnel…

Specific Aim 1.2. Develop comprehensive Project Plan…

Specific Aim 1.3. Develop decision-making protocol….

Specific Aim 1.4. Develop communication strategy… .

Specific Aim 1.5. Develop central web portal… .

Specific Aim 1.6. Develop regulatory plan… .

Specific Aim 1.7. Annual meeting of project participants… .

OBJECTIVE 2. Develop and test Anopheles gambiae strains carrying autonomous gene-drive systems linked anti-pathogen effector genes.

Specific Aim 2.1 Recruit Project Personnel.

Specific Aim 2.2 Develop CRISPR/Cas9 gene-drive systems for Anopheles gambiae.

Specific Aim 2.3 Engineer additional anti-parasite effector genes.

Specific Aim 2.4 Generate transgenic mosquitoes carrying gene-drive elements and parasite- resistance genes.

Specific Aim 2.5 develop and maintain detailed laboratory standard operating procedures, experimental records, budgets, and provide reports to appropriate components of the administrative core.

Specific Aim 2.6 Test the gene-drive performance characteristics of the AgNosCd-1 strain with no included effector molecules.

Specific Aim 2.7 Test the gene-drive performance characteristics of the AgNosCd-1  strain with included effector molecules.

Specific Aim 2.8 Test the effector performance characteristics of the strains.

Specific Aim 2.9 Mathematical modeling to support target product profile recommendation

Specific Aim 2.10 Make recommendation for release strain.

Specific Aim 2.11 Develop Target Product Profile for recommended strain(s)

Specific Aim 2.12 Perform risk assessment for recommended strain(s).

OBJECTIVE 3. Evaluate field site(s) for population modification trials...       

Specific Aim 3.1 Recruit Project Personnel…

Specific Aim 3.2 Evaluation of potential field sites for selection of four candidate sites…

Specific Aim 3.3 Evaluation of candidate field sites for selecting final field trial site…

Specific Aim 3.4. Assess “colonizability” of  An. gambiae from candidate field sites…

Specific Aim 3.5 Selection of final population modification field trial site…

Specific Aim 3.6 Model Evaluation of Candidate field site

Specific Aim 3.7 Research and training Activities at UCI MI locations

Specific Aim 3.8 Develop timeline for continued research with field site collaborators

Specific Aim 3.9 Develop Field station at selected site

Specific Aim 3.10 Recruit hire and train postdoctoral biologist(s) to direct field work at each site

Specific Aim 3.11 Conduct 2^nd season of mosquito sampling at field site.

Specific Aim 3.12 First release of TPP

Specific Aim 3.13 Post release evaluations

OBJECTIVE 4. Community Engagement

Specific Aim 4.1 Build Relationships and Establish Collaborations

Specific Aim 4.2 Site and Community Assessment

Specific Aim 4.3 Education and Capacity Building

Specific Aim 4.4 Evaluation and Communication Development

Specific Aim 4.5 Communication Management

Introduction and background

      Malaria is a disease caused by protozoan parasites in the genus Plasmodium and is transmitted to humans by the bites of infected Anopheles mosquitoes. The World Health Organization (WHO) estimates that there are ~219 million cases and ~435,000 deaths due to malaria in 2017 (World Malaria report 2018, WHO) and more than 90% of these were in Africa. No effective vaccine exists yet for this disease, and parasite resistance to prophylactic and therapeutic drugs requires the discovery of at least one new chemical every five years (REF MalEra GET).

Mosquito vector control measures have the greatest success in reducing the number of malaria infections and deaths (Walker et al., 2016). Insecticide-treated nets can reduce severe disease by 60% and other vector-targeted approaches also contribute significantly.  However, these measures alone are not enough to achieve elimination in highly-endemic areas and new technologies are needed urgently. Among the more promising are genetic strategies that will control vector mosquitoes and block parasite transmission as these strategies are not subject to the human compliance issues that compromise other strategies.

Genetic strategies seek to eliminate vector mosquitoes or reduce their densities below thresholds needed for stable parasite transmission (population suppression), or make them incapable of transmitting parasites (population modification) (Macias and James 2016). Transgenesis technologies can produce mosquito strains that carry genes that contribute to both strategies. However, long-term, cost-effective and sustainable malaria elimination requires the development of genetic strategies that are resilient to the immigration of parasite-infected mosquitoes and people, and the lack of such tools represents a significant unmet need in the malaria eradication agenda.

Mosquito population modification strains carrying genes conferring parasite resistance have the appropriate design features for this purpose. Wild, parasite-susceptible mosquitoes invading a region populated by an altered strain acquire the parasite-resistance genes by mating with the local insects, and persons with parasites moving into the same region cannot infect the resident vectors, and therefore are not a source of parasites for infection of other people. Population modification also shares with other genetic control strategies the exploitation of the ability of male mosquitoes to find females, and this is expected to offer access to vector populations that would be unreachable using conventional tools. Release of a population modification strain alone or in conjunction with other tools should make elimination possible in carefully-selected endemic areas. Population modification strategies can be used as early as the control phase of an elimination campaign alongside other measures that reduce disease incidence. As the efforts progress, this strategy takes on a larger role and ultimately is the mainstay of the prevention of reintroduction phase. As this elimination is achieved, the released altered mosquitoes facilitate consolidation of this success by allowing resources to be moved to another region with the confidence that the area just cleared will remain so. Thus, population modification offers a real chance to achieve sustainable elimination and therefore contributes significantly to malaria eradication.

A promising newly-emerging strategy for combating malaria is based on harnessing gene drive systems to spread anti-malarial genes throughout mosquito populations rendering them unable to transmit the parasites.  The James laboratory (UC Irvine) developed highly-effective anti-malarial gene cassettes that can result in a 100% block of P. falciparum, the deadliest human malaria parasite, in mosquitoes (Isaacs et al., 2011, 2012).  Working in collaboration with Ethan Bier and Valentino Gantz (UC San Diego), these effector gene cassettes coupled to a gene-drive system based on CRISPR/Cas9 biology were shown to spread with 99.5% efficiency in An. stephensi, the major malarial vector mosquito in urban India (Gantz et al., 2015).  We will adapt this new technology to a major African vector, An. gambiae, and perform a confined field trial in a well-chosen site to demonstrate proof-of-principle for how this system will eliminate malaria locally. If successful, this strategy then could be incorporated as part of a broader initiative to achieve malaria elimination throughout Africa within the following decade.

Goals and activities

            The primary goal of UCIMI is to achieve local elimination of malaria caused by P. falciparum transmitted by An. gambiae in a well-chosen field site.  The proposed trial is based on a phased approach outlined in WHO guidelines (WHO 2014) and contains specific ‘go/no-go’ evaluation points and milestones. The overall timetable for this effort is to produce a field-ready modification gene-drive strain (year one); test it in the laboratory population cages (year 2); review all potential African field trial sites and select two candidate sites from among them (year 1); conduct site visits to candidate field sites to engage officials and obtain permission to initiate collection of baseline data (year 2); . In years 3 and 4 we will collect baseline entomological data from our candidate field sites in São Tomé & Princípe and the Comoros Islands. Finally, pending community and regulatory approval, to release the modification-drive mosquitoes into the environment and follow the course of transgene spread and parasite reduction over the course of two consecutive seasons (year 5 and beyond). Once this objective is met, mosquitoes carrying the validated modification-drive could be scaled to a capacity appropriate for use in conjunction with current anti-malarial strategies to eliminate malaria regionally and ultimately throughout Africa.

Project Plan activities for the first two years were to answer to the following questions:

1)     Can we adapt coupled anti-parasite effector genes and Cas9-based, gene-drive systems for population modification to African malaria vector mosquitoes?

2)     Can we identify a field site in Africa at which it is possible to conduct a safe and ethically-sound release  trial of a population modification strain?

Project Plan activities for the next three years are:

3)     Have a field ready product that has been validated in a Target Product Profile and undergone product-specific risk assessment,

4)     Complete collection of baseline data in both Sao Tome & Principe and The Union of the Comoros. Data collection will span two malaria transmission seasons. (years 3 and 4),

5)      Following analysis of baseline data, development of a logistical plan based on these data, and pending regulatory approval, we will conduct initial release of our field ready product and initiate post release surveillance. (years 5 and beyond).

6)     To engage with the site community and stakeholders and develop a variety of educational tools to build support and understanding for the project goals and approach and encourage open lines of communication among all groups for the duration of the project.

Organizational Structure of UCIMI:

Four interacting organizational components are established for the proposed project: 1) an administrative core, 2) a laboratory component, 3) a field component and 4) a community engagement component. A single principal investigator (PI, Anthony A. James, UC Irvine) acts as the overall director of the activities. Research supervision may be delegated to project coordinators (PC) based on their expertise.

Administrative component:  This component is based at UC Irvine and has a highly-qualified, top-level administrator who functions as the project manager, Sentelle Eubanks (PM), reporting to the PI. The PM carries out necessary core administrative responsibilities essential for the successful completion of the project. This includes regulatory management to ensure that appropriate approvals are completed, requirements are met, and community consents are obtained and maintained. This component will continue after the field release of the test product and manage long-term follow-up of post project events or concerns. 

Laboratory component:  The laboratory component is carried out at UC Irvine and UC San Diego, and its goals are to develop new CRISPR/Cas9-based gene-drive systems, engineer additional anti-parasite effector genes, generate transgenic mosquitoes carrying gene-drive elements and the parasite-resistance genes, and model and test the performance characteristics of the strains (drive- and parasite-challenge assays) in controlled laboratory settings.

Field component:  The field component is led by an expert in malarial field studies (Drs. Gregory C. Lanzaro and collaborating with Anthony Cornel, and Yoosook Lee UC Davis) who works closely with the PI and in-country scientists and public health personnel to conduct site-selection activities. Field component will also be assisted by Dr. John Marshall (UC Berkeley) for modeling. 

Community Engagement (CE):  Community Engagement is intended to build relationships of trust with the collaborating country community and stakeholder groups, maintain open lines of communication, and provide education and information that result in support and understanding of the project and goals.  The community engagement components will identify, and map existing international and local regulatory requirements and framework.   The CE Specialist will work with the administrative team stakeholder community members and local government to obtain appropriate approvals and consents.  Active contact and communication with project stakeholders will be maintained and this component will continue after the field release of the test product. 

These four components will facilitate the primary project goal of local elimination of malaria in selected test sites, where conditions were chosen to be optimal for success.  During these activities, we will contribute to the education and infrastructure that allows our collaborating scientists to apply the UCIMI approach at our field sites. This would also enhance tackling the larger and far more complex problem of malaria elimination at regional scale in continental Africa.  This landscape is challenging in that new gene-drive systems would need to be developed to target additional mosquito species and possibly an additional parasite species.  It may be necessary to integrate application of gene-drive population modification systems with other well-validated anti-malarial strategies to achieve sustainable elimination of malaria at many key sites as a prelude to launching the final effort at eradication. Achieving these goals requires training of scientists both regionally and in the USA to carry out the elimination agenda.

Project Plan Annotation

Below is a synopsis of the Project Plan for years 3-5 including the Objectives, Specific Aims, Activities, Decision points and Milestones.  Project Coordinators are listed below each Objective. There are three different dates associated with each decision point: a completion date, a reporting date and a review date.  The completion dates are based on a starting date of April 1, 2019.  The reporting date refers to the date that Project Coordinators must have their reports submitted for review.  The review date refers to the dates that the Principal Investigator, Project Manager and Project Coordinators evaluate each Activity based on the “Decision Making Criteria.”

OBJECTIVE 1: PROJECT ADMINISTRATION.

Project coordinators: Anthony A. James (UC Irvine) Principal Investigator and Sentelle Eubanks (UC Irvine) Project Manager.

This Objective provides the central administration of the project. Core operational activities include: 1) preparing budgets for the entire project (principal budget) and subcontracts for each component as well as reports and audits of these efforts; 2) communication: both proactive (e.g., establishing and maintaining an updated website describing project objectives and current status) and reactive (e.g., responding to unanticipated events impacting the project or to community concerns); 3) establishing regulatory guidelines and monitoring their implementation (which involves cycles of interaction with relevant government and local representatives, institutional review boards [IRBs], institutional biosafety committees [IBCs] and their national equivalents); 5) managing legal issues (e.g., intellectual property [both project-generated and outside IP] required to meet project objectives, leases, land purchases, crisis management); 6) interaction with in-country government or research administrators, and 7) organizing participation of ancillary scientists to evaluate project outcomes in companion efforts (e.g., effects of elimination of malaria in local on education, income, political involvement).

Specific Aim 1.1 Recruit Project Personnel

This Specific Aim is to recruit the appropriate personnel for Objective 1.

James - Starts 4/1/17

              Activity 1.1.1.          Write job descriptions and job announcements to be advertised.

Finalize personnel needs and prepare draft of job descriptions. The job description for each position will be drafted to be consistent with UC approved job titles and descriptions.

Completed.

James - Starts 4/1/17

              Activity 1.1.2.          Advertise positions.

Job advertisements will be posted in multiple job list sites and appropriate scientific journals to maximize candidate pools.

Completed.

James - Starts 4/1/17

              Activity 1.1.3.          Interview Candidates.

Candidates will be interviewed by James via an online meeting platform or in person at UCI. Interviewees will be ranked and individuals will be selected for each available position and notified.

Completed.

James - Starts 4/1/17

              Activity 1.1.4.          Fill positions.

Administrative procedures for hiring will be completed. Each new hiree will take the required UCI training, such as lab safety, cyber security, etc. The requisite permits, visas, permissions and facility keys will be obtained.

Completed.

Specific Aim 1.2. Develop comprehensive Project Plan.

This Specific Aim produces and maintains the document that describes the goals, objectives and operational aspects of the UCIMI, as well as their timelines. It also includes as an Appendix I the definitions, terms and conditions of the UCIMI. Appendix II will be developed to provide in-depth information on the Specific Aims and Activities in Objectives 2 AND 3.

James - Starts 4/1/17

Activity 1.2.1. Develop draft Project Plan.

Continuing.

Specific Aim 1.3. Develop decision-making protocol.

The decision-making plan is an explicit part of the comprehensive Project Plan and is included in Appendix I.

James - Starts 4/1/17

Activity 1.3.1. Develop decision-making protocol.

Completed.

Specific Aim 1.4. Develop communication strategy.  

This Specific Aim establishes the reactive and proactive communication plans for the project.

James - Starts 4/1/17

Activity 1.4.1. Reactive plan

Develop both internal and external communication framework to give or exchange information when an unexpected event occurs.

Decision point 1.4.1a: Completion date: 6/30/19; Reporting date: Quarterly   Have decided on means to communicate when an unexpected event occurs.

James - Starts 4/1/17

Activity 1.4.1. Proactive plan

Devise strategy to communicate with both within group members and outside groups.

Decision point 1.4.1a: Completion date: 6/30/19; Reporting date: Quarterly   Set a long-term communication goals and decide on means to achieve those goals.

Specific Aim 1.5. Develop central web portal.

This Specific Aim provides for the UCIMI interface with the public and a private posting site for project members and documents.  

James - Starts 4/1/17

Activity 1.5.1. Develop central website.

Host a website that will serve communication channel with public about UCIMI.

Completed.

James - Starts 10/1/17

Activity 1.5.2. Update website.

Update project progress, project personnel, contact information as needed.

Completed.

Decision points 1.5.2a-c:  Completion date: quarterly or as needed for duration   These decision points involve evaluating the effectiveness of the website and updating it as required by progress on the project.

Specific Aim 1.6. Develop regulatory plan. 

 This Specific Aim provides the structure for maintaining regulatory compliance for the project.

James - Starts 4/1/17

Activity 1.6.1. Implement procedures for tracking research-related regulatory requirements.

Have system in place to keep track of research-related regulatory requirements for various agencies such as IBC, IACUC, CDC, USDA, and others.

Decision points 1.6.1a-h:  Completion date: quarterly or as needed for duration Complete any necessary permits. Document the contact information, website, and process to obtain such permits. Have legal consultant(s) review documents.

James, Lanzaro and Cornel - Starts 4/1/17

Activity 1.6.2. Review and develop knowledge base for regulatory requirements for potential field sites.

Research on regulatory requirements at four potential field sites recommended by UCD.

Decision points 1.6.2a-h:  Completion date: quarterly or as needed for duration Documents laws, contact information, website, and process to obtain permit to conduct genetically engineered mosquitoes in four potential field sites. Have legal consultant(s) review documents.

James, Lanzaro and Cornel - Starts 4/1/17

Activity 1.6.3. Work with field team to obtain necessary permits.

Obtain permits/permission letters needed to carry out mosquito collections in the potential field sites as well as import permits to bring mosquitoes (both live and dead) into UCI and UCD laboratories.

Decision points 1.6.3a-i:  Completion date: quarterly or as needed for duration. Obtain permit, renew or resubmission as needed.

Activity 1.6.4. Hire Consultants.

Hire Community Engagement Consultant and Regulatory Consultant to strategize and implement engagement and regulatory requirements.

Decision points 1.6.3a-b:  Completion date: as needed for duration. Hire and train as needed.

Specific Aim 1.7. Annual meeting of project participants

This Specific Aim provides for an Annual Meeting of project participants.

James - Starts 10/1/17

Activity 1.7.1. Host annual meeting.

Organize annual meeting (location to be determined) where all parties involved in the project can report their progress and discuss plans for the following year. Any revision on activity or timeline based on the current progress will be discussed.

Decision points 1.5.1a-b: Establish meeting date These decision points are the actual holding of the meeting in the last quarter of the annual cycle. These meeting will cover both scientific and administrative issues as required by the progress in the program.

Decision point  1.5.1a:   Completion date: 08/08/2017  Have the first annual meeting.

Completed.

Decision point  1.5.1b:   Completion date: 01/24/2018  Have the second annual meeting.

Decision point  1.5.1c:   Completion date: 01/31/2019  Have the third annual meeting.

Decision point  1.5.1d:   Completion date: 3/30/20  Have the fourth annual meeting.

Decision point  1.5.1e:   Completion date: 3/30/21  Have the fifth annual meeting.

OBJECTIVE 2. DEVELOP AND TEST ANOPHELES GAMBIAE STRAINS CARRYING AUTONOMOUS GENE-DRIVE SYSTEMS LINKED TO ANTI-PATHOGEN EFFECTOR GENES.

Project coordinators: Anthony A. James (UC Irvine) Principal Investigator, Sentelle Eubanks (UC Irvine) Project Manager, Ethan Bier (UC San Diego) and John Marshall (UCB).

This component provides the Phase 1-validated insect strains for population modification. Laboratory activities include: 1) link gene-drive systems to anti-P. falciparum effector genes and insert them in An. gambiae into fitness-neutral mosquito chromosomal loci, chromosomal loci conferring decreased mosquito fitness, and into chromosomal loci that affect parasite propagation in the vector; 2) conduct parasite challenge assays; 3) develop and test new gene-drive strategies; 4) identify new reagents to inactivate P. falciparum; 5) calibrate mathematical models of gene drive systems to data from laboratory studies and use these models to contribute to the discussion on construct design; 6) development of target product profiles; 7) conduct risk assessment of specific strains in reference with field sites and 8) develop and maintain detailed laboratory standard operating procedures, experimental records, budgets, and provide reports to appropriate components of the administrative core.

Specific Aim 2.1 Recruit Project Personnel

This Specific Aim is to recruit the appropriate personnel for Objective 1.

James and Bier - Starts 4/1/17

              Activity 2.1.1.          Write job descriptions and job announcements to be advertised.

Finalize personnel needs and prepare draft of job descriptions. The job description for each position will be drafted to be consistent with UC approved job titles and descriptions.

Completed.

James and Bier - Starts 4/1/17

              Activity 2.1.2.          Advertise positions.

Job advertisements will be posted in multiple job list sites and appropriate scientific journals to maximize candidate pools.

Completed.

James and Bier - Starts 4/16/17

              Activity 2.1.3.          Interview Candidates.

Candidates will be interviewed by James and/or Bier via an online meeting platform or in person at UCI or UCSD. Interviewees will be ranked and individuals will be selected for each available position and notified.

Completed.

James and Bier - Starts 4/1/17

              Activity 2.1.4.          Fill positions.

Administrative procedures for hiring will be completed. Each new hiree will take the required UCI or UCSD training, such as lab safety, cyber security, etc. The requisite permits, visas, permissions and facility keys will be obtained.

Completed.

James, Bier, Lanzaro, Marshall , Cornel, Lee, Eubanks Starts 04/01/2019 and 04/01/2021

Activity 2.1.5 Perform serial Risk Assessment(s) of project and final construct. Assess the risks associated with the project, concept and site(s).   As TPP is complete perform risk assessment on final construct at selected site.

Specific Aim 2.2 Develop CRISPR/Cas9 gene-drive systems for Anopheles gambiae

James and Bier - Starts 4/1/17

Activity 2.2.1 Identify and clone appropriate promoter sequences (for example, β2-tubulin, U6, vitellogenin, carboxypeptidase, G12) for expressing CRISPR/Cas9 gene-drive system components.

Multiple promoter regions will be investigated to improve the design of gRNA and/or the efficacy of anti-malarial gene expressions.

Completed.

James and Bier – Starts -

Avtivity 2.2.1a Build alternative gene-drive constructs for insertion into new candidate loci paralleling those already built and awaiting injection into An. stephensi (e.g., Rab5, SPO11, zpg, mre11, rad51, msh4, syn7).

Decision point 2.2.1 a Completion Date 06/01/2019

James and Bier - Starts 4/1/17

Activity 2.2.2 Identify An. gambiae fitness-neutral mosquito chromosomal loci, chromosomal loci conferring decreased mosquito fitness, and chromosomal loci that affect parasite propagation in the vector

Multiple chromosomal loci will be investigated to evaluate effects on mosquito fitness and parasite propagation in the vector.

Completed.

James Bier

Activity 2.2.2a Inject the recoded-kh construct already built by Gerard into An. gambiae to establish a transgenic gene-drive line comparable to that being tested in An. stephensi.

Decision Point 2.2.2a Completion date 06/01/2019

James Bier

Activity 2.2.2b Build tagging construct (and inject to establish a transgenic line) to modify nos-Cas9 gene drive inserted into the cardinal locus such that it carries a recoded functional version of the cardinal gene.

James and Bier

Activity 2.2.2c CRISPR targeted transgenesis to insert an attP recombination target site into an orthologous locus of a known fitness neutral target site in An. stephensi. (e.g., 80.9 or 44C).

James and Bier - Starts 7/1/17

Activity 2.2.3 Design and synthesize gene-drive constructs and verify sequence.

Multiple gene-drive constructs will be tested for efficacy in gene conversion rate, mosquito fitness and parasite propagation.

In progress.

Specific Aim 2.3 Engineer additional anti-parasite effector genes

James and Bier - Starts 4/1/17

Activity 2.3.1 Develop additional single-chain (scFv) immunological reagents to inactivate P. falciparum (Pfs47 with Barrillas-Mury, NIH; Pfs230 with Narum, NIH; Ashley Birkett, PATH).

Additional anti-parasite effector genes will be developed to optimize efficacy of ‘no parasite’ phenotype.

Completed/Modified (moved to Specific Aim 2.7).

James and Bier - Starts 4/1/17

Activity 2.3.2 Develop alternative effector molecules based on engineered synthetic receptors (SM1).

Multiple alternative effector molecules will be engineered and tested for parasite propagation.

        Terminated.

James and Bier - Starts 7/1/17

Activity 2.3.3 Make An. gambiae transgenic lines to test additional effector genes using transposon-mediated transgenesis.

Generate An. gambiae transgenic lines via transposon-mediated transgenesis to test for engineered alternative effector molecules.

       Modified (moved to Specific Aim 2.7).

James and Bier - Starts 10/1/18

Activity 2.3.4 Perform challenge assays with P. falciparum to determine efficacy of novel effector constructs.

Challenge An. gambiae transgenic lines with P. falciparum parasite to determine efficacy of alternative effector molecules.

        Modified (moved to Specific Aim 2.7).

Specific Aim 2.4 Generate transgenic mosquitoes carrying gene-drive elements and the parasite- resistance genes.

            Modified.

This specific aim will result in transgenics mosquito lines carrying multiple effector molecules designed to give complete blocking of parasite transmission.

James and Bier - Starts 12/1/18

Activity 2.4.1 Engineer effector cassettes with multiple molecules targeting different stages of the parasites.

Design and build effector cassettes with multiple effector mechanisms.

Decision point 2.4.1a-c:  Completion date: 03/31/2019 or every 6 months for duration. At least two An. gambiae transgenic lines that are validated to have integrated gene-drive and multiple effector genes.

Activity 2.4.2 Perform ‘swap’ experiments to integrate effector molecules into the AgNosCd-1 gene-drive line.

Perform microinjection experiments, screen resulting insects, and validate integrated DNA to generate gene-drive lines with linked anti-parasite effector genes

Decision point 2.4.2a-c:  Completion date: 07/30/2019 or every 6 months for duration. At least two An. gambiae transgenic lines that are validated to have integrated gene-drive and multiple effector genes.

Specific Aim 2.5 Develop and maintain detailed laboratory standard operating procedures, experimental records, budgets, and provide reports to appropriate components of the administrative core. This Specific Aim establishes and maintains the laboratory documents to validate strain development and other standard operating procedures.

James and Bier - Starts 4/1/17

Activity 2.5.1 Develop and maintain detailed laboratory standard operating procedures (SOPs)

Decision point 2.5.1a-c:  Completion date: every 6 months for duration. Have in place a fully functional project file repository. Add new SOP or update existing SOPs as required by progress on the project and upload to project file repository. SOPs are available at: the James Lab Website

Ongoing.

Specific Aim 2.6 Test the gene-drive performance characteristics of the AgNOSCd-1 strain with no included effector molecules. This Specific Aim tests the drive components in the absence of the effector molecules along with several other activities that will affect the drive parameters.  Started 12/01/2018

Modified, In progress

James, and Marshall - Starts 12/01/18

Activity 2.6.1 Conduct small cage trials with first gene-drive line (nanos, cardinal).

Drive features of the AgNOSCd-1 strain tested in small cage trials.

Decision point  2.6.1a-c:  Completion date: 10/31/2019. Drive characteristic of line established.

Activity 2.6.2 Develop method for accurately measuring NHEJ rate.

Decision point  2.6.2 XX:  Completion date: 04/30/2019. Have straightforward approach to measuring NHEJ rate.

Activity 2.6.3 Assay in vitro the effects of gRNA single nucleotide polymorphisms on Cas9-gRNA nuclease activity.

Decision point  2.6.3 XX:  Completion date: 12/31/2018. Know impact of SNPs on cutting.

Completed

Activity 2.6.4 Assay in vivo the effects of gRNA single nucleotide polymorphisms on Cas9-gRNA nuclease activity.

Decision point  2.6.4 XX:  Completion date: 05/31/2020. Know impact of SNPs on introgression in small cage experiments. (Field sample dependent)

Activity 2.6.5 Determine life-table parameters of the AgNosCd-1 strain using fitness components. Started 11/01/2018

Male: competitiveness with wild-type, adult survival, (no. possible progeny); female: no eggs laid, no. eggs hatching, no. females laying eggs over three bloodmeals; larval to adult survival both sexes.

Decision point  2.6.5 XX:  Completion date: 06/30/2019. Know values of fitness components.

Activity 2.6.5a Inject the Ag8341Cas9 expression construct that Zhiquin Li has built to drive male specific expression of Cas9 in An. gambiae.

Decision point 2.6.5 a Completion date 04/01/2019

Activity 2.6.5b Build and inject a CHACR element into An. gambiae that is inserted into the FREP1 coding locus and carries the Q13545L allele as part of a recoded frep1 gene construct.  This construct would be comparable to the one Gerard has already built for An. stephensi in which the equivalent amino acid change has been made (Q671L). 

Activity 2.6.6 Refine life-table parameter estimates of AgNosCd-1 strain using cage trial data. Using a Bayesian model framework, we will incorporate prior knowledge of life-table parameters based on the results of Activity 2.6.5, and update these by fitting models of gene drive dynamics to laboratory cage gene drive experiments. From this analysis, we will produce posterior estimates of life-table parameters with associated credible intervals.

Decision point  2.6.6 a-e:  Completion date: Quarterly. Model-inferred values of homing efficiency, resistant allele generation and fitness parameters.

James, Lanzaro, Bier and Marshall - Starts 7/1/18

Activity 2.6.7 Use fitted model of gene drive system and its effects on An. gambiae life-table parameters to predict behavior of system in the environment.

Decision point 2.6.6a-m:  Completion date: every 6 months for duration.

Utilize the MGDrivE modeling framework (https://marshalllab.github.io/MGDrivE/), parameterized for the gene drive system fitted in Activity 2.6.6, An. gambiae, and the landscapes of interest to the UCI Malaria Initiative, to explore the expected behavior in the environment of the gene drive systems studied in the laboratory. Use results from these model predictions to advise future molecular development of the gene drive systems. 

Modify 

Specific Aim 2.7 Test the gene-drive performance characteristics of the AgNOSCd-1 strain with included effector molecules. This Specific Aim tests the drive components with effector molecules for Gene Drive Stability, Ideal release ratios, frequency of resistant alleles, phenotypes and genotypes, new male specific promoter and natural variation from field strains.

Modified

James, Bier and Marshall - Starts 08/01/2019

Activity 2.7.1 Conduct small cage trials with first gene-drive line (nanos, cardinal).

Drive features of the AgNosCde-1 strain tested in small cage trials.

Decision point  2.6.1a-c:  Completion date: 06/30/2020. Drive characteristic of line established.

Activity 2.7.2 Refine life-table parameter estimates of AgNosCde-1 strain using cage trial data. Using a Bayesian model framework, we will incorporate prior knowledge of life-table parameters based on the results of Activity 2.6.5, and update these by fitting models of gene drive dynamics to laboratory cage gene drive experiments based on the results of Activity 2.7.1. From this analysis, we will produce posterior estimates of life-table parameters with associated credible intervals.

Decision point  2.7.2a-c:  Completion date: every three months  06/21/2020 Model-inferred values of homing efficiency, resistant allele generation and fitness parameters, after the inclusion of an effector molecule.

Activity 2.7.3 Use fitted model of gene drive system and its effects on An. gambiae life-table parameters to predict behavior of system in the environment.

Decision point 2.7.3a-c:  Completion date: every 6 months for duration. 

Utilize the MGDrivE modeling framework (https://marshalllab.github.io/MGDrivE/), parameterized for the gene drive system fitted in Activity 2.7.2, An. gambiae, and the landscapes of interest to the UCI Malaria Initiative, to explore the expected behavior in the environment of the gene drive systems studied in the laboratory, after the inclusion of an effector molecule. Use results from these model predictions to advise future molecular development of the gene drive systems and effector molecules.

Specific Aim 2.8 Test the effector performance characteristics of the strains. This Specific Aim tests the efficacy of the effector molecule combinations to prevent sporozoites in the salivary glands of transgenic mosquitoes.

James and Bier - Starts 08/01/2019

Modified.

Activity 2.8.1 Conduct parasite challenge assays on transgenic An. gambiae lines

Developed strains from Specific Aim 2.4 will be challenged with P. falciparum to measure the efficacy of parasite propagation.

Decision point  2.8.1a-e:  Completion date: every 6 months for duration. At least two An. gambiae transgenic lines will be challenged with P. falciparum and measure parasite propagation.

Specific Aim 2.9.1 Mathematical modeling to support target product profile recommendation (epidemiology). Develop an epidemiological module for the MGDrivE modeling framework. Using the combined framework, in a randomly mixing population, determine the combinations of gene drive system parameters required to reduce the R₀ for malaria to below 1 in a range of transmission settings.

James, Marshall and Bier - Starts 1/1/19

Specific Aim 2.9.2. Mathematical modeling to support target product profile recommendation (entomology). Apply the MGDrivE framework to a landscape of interest to the UCI Malaria Initiative (e.g. Sao Tome & Principe, Grand Comore). Determine the combinations of gene drive system parameters required for the system to spread to a level required for local malaria elimination throughout the landscape.

James, Marshall and Bier - Starts 1/1/19

Specific Aim 2.10 Make recommendation for release strain(s). This Specific Aim has the milestone of recommending a specific product to start the Target Product Profile and Risk Assessment components. Based on gene-drive success rate, fitness, and modeling results, make a short list of release strain(s) for field trial.

Milestone 2.10.1 Completion Date: XXX, Reporting date: XXX; Review date: Annual meeting. A short list of release strain(s) for field trial will be decided.

Specific Aim 2.11 Develop target product profile for recommended strains. This Specific Aim provides the target product profile to be used.

Starts 1/1/19

Activity 2.11.1 Work with FNIH and other non-UCI MI projects to develop consensus on TPP parameters and develop values for ‘ideal’ and ‘minimally acceptable’.

Drive features of the AgNOSCd-1 strain tested in small cage trials.

Decision point  2.11.1a-c:  Completion date: XXX. Attend and contribute to FNIH meetings.

Activity 2.11.2 Adopt consensus TPP parameters for ‘ideal’ and ‘minimally acceptable’ and assure AgNosCd-1 fits profile.

Decision point  2.11.2 XX:  Completion date: XXX. Have product that meets TPP prfile for Phase 1 parameters.

Activity 2.11.3 Report TPP and test results to WHO Vector Control Advisory Group (VCAG).

Decision point  2.10.2 XX:  Completion date: XXX. Provide Phase 1 results to VCAG as part of their review process.

Specific Aim 2.12 Perform risk assessment for recommended strain.

Activity 2.12.1 Work with field site to identify regulatory structure of the proposed field site.

Decision point  2.10.1a-c:  Completion date: XXX. Know what is required for risk assessment at the proposed field site

Activity 2.12.2 Work with Site Partners to Identify Risks. Identify credible independent entity to conduct site-relevant risk assessment. 

Decision point  2.10.2 XX:  Completion date: XXX. Have identified an independent partner with which to work.

Activity 2.12.3 Conduct site-relevant risk assessment of product.

Decision point  2.10.2 XX:  Completion date: XXX. Risk assessment completed.

OBJECTIVE 3. EVALUATE FIELD SITE(S) FOR POPULATION MODIFICATION TRIALS.           

Project coordinators: Anthony A. James (UC Irvine) Principal Investigator, Sentelle Eubanks (UC Irvine) Project Manager, Gregory C. Lanzaro (UC Davis), Anton Cornel (UCD), Yoosook Lee (UCD), John Marshall (UCB).

YEARS 1 and 2: This Objective conducts the site-selection process and identifies a field trial site for evaluation of population modification of Anopheles gambiae. The overall goal of this component is to answer the question of whether or not one or more field trial sites exist that maximize the prospects for success, minimize the risk and serve as a fair, valid and convincing test of the efficacy of the population modification strain being evaluated.

We refer to "potential field sites" as all possible field sites; "candidate field sites" as four sites selected for closer evaluation from among the potential sites; "village" as a sub-site within a candidate site. Villages will represent the sampling units from which mosquito specimens will be collected.

Field activities in the first two years include: [MODIFIED²]:1) evaluate one or more sites for the field trials;; 2) identify required facilities for laboratory aspects at the field trials and establish an on-site presence. Successful completion of these efforts is expected to be followed in subsequent years by  4) generate transgenic mosquitoes carrying gene drive; 5) production of mosquitoes to be tested in the field; 6) conduct on-site open confined trials; 7) develop protocols for staged open-field releases including mitigation plans in case trials have to be halted; 8) monitor mosquito and parasite parameters during field trials; 9) report results to administrative core; 10) interact with consultants, regulatory personnel on-site, and administrative staff dedicated to regulatory responsibilities in the administrative core; 11) contribute to community engagement and interact with staff tasked with community relations in administrative core; 12) interact with the laboratory component by sending personnel to labs as necessary and communicating regularly with laboratory researchers regarding results obtained by both components.

² Field activities in years 1 and 2 were substantially modified. Our original plan was to conduct site visits as part of our site evaluation process. When this was attempted in the Union of the Comoros, we were denied access. Officials there required more information about the UCIMI program. At this point we established a Community Engagement program under the leadership of Ana Kormos. In collaboration with the CE program we organized one day workshops in both São Tomé & Príncipe and The Comoros. These workshops were highly successful and in hindsight should have been part of our original plan. However, the time required to organize and conduct these workshops and then obtain confirmation from our collaborators to proceed with the first year of field work necessitate a change in the original plan. At this point we are confident that STP and the Comoros are in fact the best choices for field sites, therefore we plan to initiate the collection of baseline data starting early in Phase II (early 2019). This modification to the original plan puts us on schedule with the original plan.

Specific Aim 3.1 Recruit Project Personnel

Lanzaro and Cornel - Starts 4/1/17

Activity 3.1.1 Write job descriptions and job announcements to be advertised.

Organize meeting of Objective 3 key personnel to finalize personnel needs and prepare draft of job descriptions. The job description for each position will be drafted to be consistent with UC approved job titles and descriptions.

Decision point 3.1.1a Completion Date: 4/15/17, Reporting date: 6/30/17; Review date: Annual meeting. Final version of job descriptions will be completed.

Completed.

Lanzaro and Cornel - Starts 4/1/17

Activity 3.1.2 Advertise positions.

Job advertisements will be posted in multiple job list sites and appropriate scientific journals to maximize candidate pools.

Decision point  3.1.2a Completion Date: 5/15/17, Reporting date: 6/30/17; Review date: Annual meeting. Applications will be collected, reviewed by Lanzaro, Cornel and Lee and a short list of candidates will be completed.

Completed.

Lanzaro, Cornel and Lee - Starts 5/17/17

Activity 3.1.3 Interview Candidates.

Candidates will be interviewed by Lanzaro, Lee and Cornel via an online meeting platform or in person at UC Davis. Interviewees will be ranked and individuals will be selected for each available position and notified.

Decision point 3.1.3a Completion Date: 6/17/17, Reporting date: 6/30/17; Review date: Annual meeting. Final list of candidates will be notified and provided with timeline to respond with their decision. All positions will remain open until filled.

Completed.

Lanzaro, Lee  and Cornel - Starts 7/1/17

Activity 3.1.4 Fill positions.

Administrative procedures for hiring will be completed. Each new hiree will take the required UC Davis training, such as lab safety, cyber security, online sexual harassment training etc. The requisite permits, visas, permissions and facility keys will be obtained. Assistance in housing etc. will be provided.

Decision point  3.1.4a Completion Date: 8/30/17, Reporting date: 9/30/17; Review date: Annual meeting. Hiring process will be completed.

Completed.

Specific Aim 3.2 Evaluation of Potential Field Sites for Selection of Four Candidate Sites

Lanzaro, Cornel and Lee - Starts 4/1/17

              Activity 3.2.1.          Literature Survey for the Evaluation of 13 Potential Field Trial Sites

We will conduct literature searches to identify geographically isolated island sites and obtain detailed information for each including: geophysical and biological features; malaria vector species and parasite species present; identify on-site university or government institutions as potential collaborators to assist with all field studies; health infrastructure and logistics of travel and shipping. Literature search will include, but will not be limited to, WHO country profile, PMI country profile, CIA handbook, US Department of State notes, Pubmed, local public health reports, and personal communications.

Decision point 3.2.1a. Completion Date: 6/1/17, Reporting date: 6/30/17; Review date: Annual meeting. Written report of information for 13 potential field sites under consideration.

Completed.

Lanzaro, Haddad, and Lee - Starts 4/1/17

              Activity 3.2.2.          Analysis of remotely sensing data for 13 potential field sites.

For each of the 13 potential field sites under consideration, the time series (or static values, for topography) of several environmental variables will be extracted from remote sensing observations and from model reanalysis (specifically, NASA's MERRA-2). The analysis will quantify the spatial variability of the environment in potential field site, as well as the intra-annual and inter-annual variability at the spatial resolution of each variable. Using a set of well-defined criteria we will select four sites as candidate field sites for this program. The results will provide objective spatio-temporal selection criteria and will inform the timing of site visits to facilitate mosquito collection.

Decision point 3.2.2a. Completion Date: 6/1/17, Reporting date: 6/30/17; Review date: Annual meeting. Written report with maps and graphics on the environmental data for each of 13  potential field sites.

Completed.

Lanzaro, Cornel, Lee, Marshall and Haddad - Starts 6/2/17

              Activity 3.2.3.          Select, based on set criteria, four candidate sites.

Based on the compiled reports of entomology, epidemiology and environmental data, four candidate sites will be selected. Temporal remote sensing data will be used to determine the timing of site visits to maximize Anopheles collections.

Decision point 3.2.3a. Completion Date: 6/30/17, Reporting date: 6/30/17; Review date: Annual meeting. Four sites and a tentative schedule for site visits will be determined.

Completed.

Specific Aim 3.3 Evaluation of Candidate Field Sites for Selecting Final Field Trial Site

Lanzaro, Cornel, Lee and Marshall - Starts 7/1/17

              Activity 3.3.1.          Prepare site visits to four candidate field sites and nearest mainland. [MODIFIED³]

We will contact potential collaborators at each of the four candidate field sites to introduce ourselves, outline the project and schedule site visits. Travel arrangements including visa applications will be concluded prior to site visit. Potential collaborators at nearest mainland sites will also be contacted at this time for the purpose of scheduling travel and planning work.

³ Site visits were conducted in the context of the Community Engagement workshops. These did include extensive field work that included mosquito surveillance, however at this point we did not have permission to collect specimens or conduct research. These permissions have now been obtained for STP and are pending with the Comoros. We did obtain specimens from mainland sites including Equatorial Guinea, Gabon, Cameroon and Angola for STP and Tanzania for the Comoros. We are working on obtaining permits to collect specimens from northwest Madagascar.

Decision point  3.3.1a-c: Completion Date: 8/30/17, Reporting date: every 6 months for duration starting 9/30/17. Establish contacts, collaborative arrangements and site visit schedule for candidate field sites.

Modified/Completed.

Lanzaro, Cornel and Lee - Starts 8/15/17

              Activity 3.3.2.          Visit villages on each candidate site and nearest mainland to collect Anopheles specimens. [⁴MODIFIED]

An. gambiae s.l. will be collected from at least 3 villages for 9 days (3 days/village x 3 villages) at each of the four candidate field site. The ovaries from all half gravid An. gambiae s.l. will be preserved for cytogenetic analysis. The remaining carcasses and all other mosquitoes will be preserved individually in 80% ethanol with a unique accession number to track the specimen’s location (site and village) and date of collection. All mosquito specimens will be cataloged and archived at UC Davis for downstream longitudinal studies. A collection of gravid female An. gambiae will be made from each village. These will be held to collect eggs. Eggs and early stage larvae will be returned to UC Davis and reared to the adult stage to be used for insecticide susceptibility testing and to evaluate the potential for establishing colonies , as described below.

Decision point 3.3.2a. Completion Date: 9/30/17, Reporting date: 12/15/17; Review date: Annual meeting. Collections of mosquitoes from site 1 and nearest mainland. Written categorical evaluation of infrastructure, collaborators, and epidemiological data and data keeping system for each candidate site
Decision point 3.3.2b. Completion Date: 1/30/18, Reporting date: 3/30/18; Review date: Annual meeting. Collections of mosquitoes from site 2 and nearest mainland. Written categorical evaluation of infrastructure, collaborators, and epidemiological data and data keeping system for each candidate site
Decision point 3.3.2c. Completion Date: 6/30/18, Reporting date: 9/30/18; Review date: Annual meeting. Collections of mosquitoes from site 3 and nearest mainland. Written categorical evaluation of infrastructure, collaborators, and epidemiological data and data keeping system for each candidate site
Decision point 3.3.2d. Completion Date: 8/30/18, Reporting date: 12/15/18; Review date: Annual meeting. Collections of mosquitoes from site 4 and nearest mainland. Mosquito collections from 4 candidate sites and nearest mainland for a total of 2,700 adults and 2,700 larvae. Written categorical evaluation of infrastructure, collaborators, and epidemiological data and data keeping system for each candidate site.

⁴ Because of delays in obtaining permission to conduct fieldwork in STP and The Comoros this work will not be initiated as part of our evaluation of sites. We have determined, without conducting these activities, that STP and the Comoros are in fact the best field sites. Both are archipelagos, two islands in STP and three in the Comoros. We have determined, based on earlier work and our own re-analysis that these islands are not only genetically isolated from the mainland but are also isolated from one another. We have two manuscripts in preparation that describe a preliminary description of the genetics of Anopheles gambiae (Comoros) and Anopheles coluzzii (STP) populations on these islands. The work in Activity 3.3.2 will now be conducted as part of our effort to collect baseline data for these sites in Phase II, starting early in 2019.

Lanzaro, and Lee - Starts 10/1/17

⁵MODIFIED

              Activity 3.3.3.          Analysis of genotype data to assess entomological parameters

The divergence island SNP (DIS) assay (Lee et al. 2014, 2015) will be used for each specimen to determine species, insecticide resistance genotypes, Plasmodium species infection status (adult) and sex (larvae).  DNA will be extracted from other Anopheles species to determine their malaria infection status. Maps of species composition, insecticide resistance genotypes, Plasmodium species infection status will be generated. Statistical analyses will be conducted to examine differences between indoor vs outdoor collections.

Decision point 3.3.3a-c: Completion Date: 11/1/18, Reporting date: every 6 months for duration starting 3/30/18. Genotypes of 5,400 An. gambiae s.l. specimens entered into online database. Plasmodium infection status of An. gambiae s.l. and other Anopheles species will be entered into the online database. Written report of species composition, insecticide resistance genotypes, Plasmodium infection prevalence with maps and statistical analysis.

Lanzaro, Cornel and Lee - Starts 10/1/17

              Activity 3.3.4.          Analysis of insecticide resistance

Wild An. gambiae reared from the eggs of field collected females from each of the four candidate field sites will be tested for their phenotypic susceptibility to commonly used insecticides. These tests will be conducted at UC Davis where standard susceptible An. gambiae lines are available. Mosquitoes will also be screened for target site knock-down mutations L1014S(kdr-E), L1014F(kdr-w)

Decision point 3.3.4a-c: Completion Date: 11/1/18, Reporting date: every 6 months for duration starting 3/30/18.  Bottle bioassays for F1 progeny from each of the four candidate field sites. Genotype for knock-down mutations of F1 progeny from each candidate field site. Written report of insecticide resistance phenotype/genotype with maps and statistical analysis.

Lanzaro, and Lee - Starts 10/1/17

              Activity 3.3.5.          Analysis of genome data to assess degree of genetic isolation of sites

A total of 192 An. gambiae (8 mosquitoes/village x 3 villages x 8 sites) will be subjected to 150bp paired-end read HiSeq 4000 sequencing with a goal of 10X coverage per individual. Freebayes software (Garrison and Marth 2012) will be used for variant calling from 192 An. gambiae sequences. Principal Component Analysis (PCA) and SpaceMix (Bradburd et al. 2013) will be used to examine population structures and genetic isolation qualitatively. Polymorphism analysis will be conducted for known transgene insertion sites.

Decision point 3.3.5a-c: Completion Date: 11/1/18, Reporting date: every 6 months for duration starting 3/30/18. Whole genome sequences of 192 individuals at 10X coverage/sample. One sample/village (N=24) will be sequenced at 30X coverage/sample for increasing confidence in calling variants. Written report of population structure and qualitative genetic isolation test with PCA and geo-genetic map figures.

Lanzaro, and Marshall - Starts 7/1/17

              Activity 3.3.6.          Model evaluation of candidate field sites

Mathematical model frameworks will be developed to assess the suitability of candidate field sites for trials of population modification strains with Cas9-based gene drive systems. Models will be designed to assess: (a) confinability of a transgenic release; (b) ability to measure potential trial end-points (various entomological measures, and epidemiological measures if possible); (c) feasibility of gene drive in the presence of drive-resistant alleles and local population structure; and (d) ability to remediate transgenes from the environment using insecticide-based campaigns.

Decision point 3.3.6a-c: Completion Date: 11/1/18, Reporting date: every 6 months for duration. Review date: Annual meeting. Development of the model framework for the first 6 months. Integration of data from the other activities described here, and modeling of field trials in the four candidate field sites after model development.

⁵ Activities 3.3.3 through 3.3.7 could not be completed in Phase I because of the delay in obtaining access to field sites. Analysis of existing data and material we were able to obtain from colleagues around the world provided adequate information to convince us, with a high degree of confidence, that STP and the Comoros are the best available sites to conduct field trials of our GEM. These activities will be conducted as part of the baseline data collection effort in Phase II.

Specific Aim 3.4. Assess “Colonizability” of  An. gambiae from Candidate Field Sites

Lanzaro, and Cornel - Starts 5/1/17

              Activity 3.4.1.          Establish mosquito rearing protocols & insectary at UC Davis

Applications for the appropriate live mosquito import permits (e.g. CDC and USDA) will be initiated. Appropriate insectary set up will be prepared prior to USDA inspection. Appropriate mosquito rearing supplies will be ordered.

Completed. (we have all these at UCI?)

Decision point 3.4.1a-c. Completion Date: 7/1/17, Reporting date: every 6 months or as needed for duration starting 9/30/17. Review date: Annual meeting. Receive copy of CDC and/or USDA permit for live mosquito import at UC Davis insectary. Written protocol with visual aids for mosquito rearing and data record sheet to log life table for each mosquito strain.

Lanzaro, Cornel and Lee - Starts 10/1/17

              Activity 3.4.2.          Assess ease of colonization for An. gambiae for each site. MODIFIED.

Gravid females will be collected and placed individually in microtubes with water and filter paper for oviposition. Egg clutches separated by single females will be transported back to UC Davis as checked baggage or via shipping. Egg numbers, hatching rate, emergence rate, sex ratio, mating status in cage (spermatheca dissections), feeding preference (and whether they laid eggs in cage conditions) and adult longevity will be recorded for each strain.

Decision point 3.4.2a-c: Completion Date: 11/1/18, Reporting date: every 6 months for duration starting 3/30/18. Review date: Annual meeting. 100 gravid  females/site will be collected to ensure 2,800 mosquitoes for colony establishment. Receive eggs from 4 candidate sites. Complete species identification of females for “colonizability” tests. Written document of comparing strains from multiple locations with respect to ease of rearing efforts.

Specific Aim 3.5. Select Field Trial Site

Lanzaro, Cornel, Marshall and Lee - Starts 11/2/18

              Activity 3.5.1.          Prepare a written evaluation of each of the four candidate sites.

Candidate sites will be evaluated on the basis of the following criteria:

1. Genetic isolation from mainland
2. Species composition and presence of population structure within species
3. Degree of polymorphism present in target transgene insertion sites
4. “Colonizability” (ability to establish and maintain laboratory colonies)of local An.
               gambiae strain
5. Level of insecticide resistance of local An. gambiae populations
6. Malaria parasite prevalence
7. Malaria incidence
8. Environmental complexity of An. gambiae habitat,
9. Willingness for collaboration from local institutions and collaborators
10. Acceptance level for genetically engineered mosquito release trials
11. Insectary setup or potential to set up local insectary for the project
12. Laboratory setup or potential for setting up laboratory dedicated for the project
13. Logistics of transportation between US and the field site and within the field site.
14. Logistics of financial transactions
15. Local personnel resources
16. Availability for housing for an on-site project manager, a junior specialist and
                UCD/UCI project personnel visiting field site
17. Availability of health clinic or hospital proximal to base station for on-site      personnel
18. Security for foreign personnel for both lab and housing
19. Ease of communication with local personnel
20. Cost of running local facility and hiring project personnel

Decision point  3.5.1a Completion Date: 1/3/18, Reporting date: 3/30/19; Review date: Annual meeting. Final report on evaluation of 4 candidate sites.

COMPLETE-with exception of items 4, 6, 16, 17 18 and 20. Written evaluation in the form of three manuscripts currently being prepared for publication.

Lanzaro and James - Starts 2/1/19

              Activity 3.5.2.          Make final decision on the project field site

A team meeting will be organized at UC Davis to present the progress toward site evaluation and make recommendations for a field site to PI (James).

Milestone 3.5.2a Completion Date: 2/28/19, Reporting date: 3/30/19; Review date: Annual meeting. Decision on the field site.

COMPLETE 01/31/2019.

PHASE II- YEARS 3, 4 and 5 – 2019-2021

Specific Aim 3.6 Mosquito Collection and Research Activities at Field Site Union of Comoros

Activity 3.6.1 Mosquito collections (larvae and adults) for population genetics evaluation and genotyping.

Larval mosquitoes will be sampled from a variety of water sources in multiple villages on all islands, and adults will be collected using multiple trapping methods from specific sites on all islands, over a 12 month period during 3 separate visits.  The samples will be used for genotyping and population genomic analysis.  The genotyping includes assays for species identification, insecticide resistance genes, plasmodium infection status, blood meal identification (in blood fed females), and sex determination of larvae (sex cannot be determined by visual examination in this stage.

Milestone 3.6.1a Completion Date: 12/30/19, Reporting Date: 1/30/20; Review date: 01/31/2019 Annual meeting.

Sampling completed in Year 3 at all sites.

Activity 3.6.2 Determine feasibility of establishing An. coluzzii colony and An. gambiae colony in Grande Comore.

In partnership with the National Malaria Control Program (NMCP) at both sites, rear collected larvae to adults in insectary, and attempt to collect eggs and hatched larvae from reared adults to determine species. 

Decision Point 3.6.2a Completion Date: 12/30/19, Reporting Date: 1/30/20; Review Date: Annual meeting.

Feasibility of establishing appropriate colonies in Grande Comore known.

Activity 3.6.3 Conduct preliminary WHO insecticide resistance assays. 

In partnership with the site NMCP, rear collected larvae to adults in insectary to conduct WHO insecticide resistance assays.

Decision Point 3.6.3a Completion Date: 12/30/19, Reporting Date: 1/30/20; Review Date: Annual meeting.

Insecticide susceptibility profiles for collected mosquitoes completed.

Specific Aim 3.7 Research and Training Activities at UCI MI locations.

            Activity 3.7.1 DNA extraction, sequencing and analysis of site collected mosquitoes.

DNA extraction, genotyping and genome sequencing will be performed for site collected mosquitoes. Genotyping and sequencing data will be analyzed.  Data will be reported using a shared online data base and presented to stakeholders and collaborators at field site(s) as well as written as manuscripts for publication.

Milestone 3.7.1a-b Completion Date: 12/30/19, Reporting Date: 1/30/20; Review date: Annual meeting.

Data Analysis complete and results shared with key stakeholders and collaborators.

Activity 3.7.2 Provide training at UCI MI locations for field site collaborators

Select individuals from field site will travel to UCI MI campuses for capacity building training onsite in the laboratory.

Milestone 3.7.2a Completion Date: 12/30/19, Reporting Date: 1/30/20; Review date: Annual meeting.

Training complete.

Activity 3.7.3 Develop plan for field station at field site(s).

Identify selected site, location of lab, and names of field personnel.  Identify areas for housing and begin rental or purchase agreements for locations.   

Assessment of needs and resources is complete, and development of field station plans at field sites (Sao Tome and Grande Comore) is created.

Milestone 3.7.3a-b Completion Date: 12/30/19, Reporting Date: 1/30/20; Review date: Annual meeting.

List of resources and supplies needed for field stations is complete, and plan has been developed for the implementation of a field station at field site(s).

Activity 3.7.4 Provide training at UCI MI in construct and microinjection.

Assessment of needs and resources is complete

Milestone 3.7.4 Completion Date: 12/30/19, Reporting Date: 1/30/20; Review date: Annual meeting. List of resources and supplies needed for field stations is complete, and plan has been developed for the implementation of a field station at field site(s).

Specific Aim 3.8 Develop timeline/plan for continued research with field site collaborators

Activity 3.8.1 Conduct follow up workshop for site collaborators and stakeholders

We will conduct a workshop to distribute data and results of our field collections to site stakeholders and have discussion about next steps and going collaboration.

Milestone 3.8.1a-b Completion Date:2/30/20, Reporting Date: 3/30/20; Review date: Annual meeting.

Workshop is complete.

Activity 3.8.2 Create and implement strategy for next phase of research at field site(s) with collaborators

Obtain permission for next phase of research and develop a work plan with collaborators.

Milestone 3.8.2a-b Completion Date: 3/30/20, Reporting Date: 4/30/20; Review date: Annual meeting.

Have permission for next phase and a completed work plan.

Specific Aim 3.9 Develop Field Station at Field Site

Activity 3.9.1 Develop Field Station  Obtain permissions, resources, and supplies needed for onsite field station and complete set up. 

Milestone 3.9.1a-b Completion Date: 3/30/20, Reporting Date: 4/30/20; Review date: Annual meeting.

            Supplies and permissions completed, and field station is set up.

Activity 3.9.2 Upgrade insectary facilities at Grande Comore.

Construction or purchase of shelves to hold larval rearing trays and adult cages in existing rooms.  Install and test temperature/climate control equipment. 

Milestone 3.9.2a Completion Date: 6/30/20, Reporting Date: 7/30/20; Review date: Annual meeting.

            Functional insectary for rearing mosquitoes.

Activity 3.9.3 Upgrade laboratory facilities at Grande Comore

Upgrade existing electrical or solar in existing labs.  Purchase basic equipment such as PCR thermocyclers for routine laboratory work.

Milestone 3.9.3a Completion Date: 6/30/20, Reporting Date: 7/30/20; Review date: Annual meeting.

            Functional laboratory for the conduct of routine work. .

Activity 3.9.4 Rent or purchase suitable housing for onsite UCI MI staff

Arrange for of a minimum 4 bedroom house, and arrange hiring of security and household personnel. House should include room for an office and guest facilities for visiting UC personnel.  

Milestone 3.9.4a-b Completion Date: 6/30/20, Reporting Date: 7/30/20; Review date: Annual meeting.

Contract signed for housing through end of 2021, and contract signed for household and             security personnel.

Activity 3.9.5 Acquire necessary furnishings for field station and hire security.

Purchase all furniture and supplies needed for housing. 

Milestone 3.9.5a Completion Date: 6/30/20, Reporting Date: 7/30/20; Review date: Annual meeting.

            Furnishings and supplies purchased.

Specific Aim 3.10 Hire Field Site Personnel

Activity 3.10.1  Recruit, Hire and Train post-doctoral biologist(s) to direct field work at each field site.

Recruit for and hire onsite personnel who will conduct ongoing field research at field site(s) and provide them with orientation required to successfully complete work.

Milestone 3.10.1a-b Completion Date: 1/30/20, Reporting Date: 2/30/20; Review date: Annual meeting.

Onsite research personnel are hired and trained

Activity 3.10.2  Recruit, Hire and Train administrative personnel to assist with administrative duties at each field site.

Recruit for and hire junior personnel to assist with administrative duties at field site, and provide them with orientation required to successfully complete work.

Milestone 3.10.2 a-b Completion Date: 1/30/20, Reporting Date: 2/30/20; Review date: Annual meeting.

Onsite administrative assistant hired and trained

Activity 3.10.3  Recruit, Hire and Train two National/local to site biologists who will work as field station staff.

Recruit for and hire two local biologists who will assist with the field research at field site(s) and we provide them with orientation required to successfully complete work.

Milestone 3.10.3a-b Completion Date: 1/30/20, Reporting Date: 2/30/20; Review date: Annual meeting.

Local field biologists are hired and trained.

Specific Aim 3.11 Conduct Second Season of Mosquito Sampling at Field Site(s).

Activity 3.11.1  Conduct mark release recapture studies

Standard mark, release recapture studies will be conducted at two time points, and at two sites, at both Sao Tome and Grande Comore. .

Milestone 3.11.1a-b Completion Date: 12/30/20, Reporting Date: 1/30/20; Review date: Annual meeting.

            Estimates of dispersal and population size will be derived from data from the studies.

Activity 3.11.2  Sample aquatic organisms co-inhabiting larval mosquito breeding sites

We will conduct sampling of aquatic organisms in a sub-sample of initial collection sites in both Sao Tome and Comoros.

Milestone 3.11.1a-b Completion Date: 12/30/20, Reporting Date: 1/30/20; Review date: Annual meeting.

            Sampling completed.

Specific Aim 3.12.1 First release of TPP

Activity 3.11.2a – create sample database, collect samples and document.

Specific Aim 3.13.1 Recapture

Activity 3.13.1a – Recapture and sequence altered mosquitoes.

Specific Aim 3.13.2 Assess parasites

Activity 3.13.2 a – dissect and count existing parasites in altered mosquitoes

Specific Aim 3.13.3 Community Assessment of TPP

Activity 3.13.3a – Assess community and stakeholders regarding the project, document and report summary of project.

OBJECTIVE 4. COMMUNITY ENGAGEMENT

Project coordinators: Anthony A. James (UC Irvine) Principal Investigator, Sentelle Eubanks (UC Irvine) Project Manager, Ana Kormos (UC Irvine) Community Engagement Specialist, Gregory Lanzaro (UC Davis), Ethan Bier (UC San Diego) and John Marshall (UCB).

This objective facilitates community engagement activities required to support the success of objectives 2 and 3.  It includes 4 specific phases over three years, the last phase in year 3 designed specifically to manage ongoing communication (internal and external) related to all activities associated with this plan.  Hire and train local CE staff and engage with health ministries.

Specific Aim 4.1 Build Relationships and Establish Collaborations

Kormos and Lanzaro – starts June 2018

Activity 4.1.1 Conduct workshops at potential field sites

In partnership with key stakeholders, conduct informative and collaborative workshop at each potential field site to determine decision making steps needed in order to conduct initial research.

Complete 11/01/2018.

Activity 4.1.2 Conduct stakeholder assessment of all sites

Determine who field site stakeholder members/groups are and begin relationship development with them.  Understand stakeholder involvement, and interest in, public health education, malaria control, and genetically modified mosquitoes. 

Milestone 4.1.2a Completion Date: 3/30/19, Reporting date: 4/30/19; Review date: Annual meeting. Stakeholder groups will be identified and engaged.

Activity 4.1.3 Draft Regulatory Framework for any/all field sites

Draft regulatory framework and communication pathways for potential field sites using existing regulatory documentation, and information from key stakeholders.

Milestone 4.1.3a-b Complete for STP 07/30/2019 reporting date 10/30/2019.  Completion Date for Comoros: 12/1/19

Activity 4.1.4 Documentation and data collection of all communication and engagement

Create activity timelines for each community engagement activity and document/collect all progress and communication associated with each activity.

Milestone 4.1.4a Completion Date: 12/30/2019, Complete activities for 2018. Completed activity timeline documenting completion dates of activities in 2019

Specific Aim 4.2 Site and Community Assessment

Kormos and Lanzaro – starts January 2019

            Activity 4.2.1 Develop and Implement community assessment(s)

Develop and implement community assessment of needs: understanding of malaria and malaria control, collaborative capacity, education level, cultural beliefs, etc. in order to develop communication strategies best suited to the needs of the communities where research is conducted.

4.2.1a Completion Date: 10/1/19, Reporting date: every 6 months; Review date: Annual meeting.

Completed assessment at field site(s).  Summary of findings will help guide ongoing education, needed resources, and grievance process.

Activity 4.2.2 Develop Educational Tools

Develop educational tools and resources that can be used at the field site to educate community members and stakeholder groups about UCIMI team, their work, the collection process, and general information about malaria and malaria transmission. Completion date 03/31/2019

Milestone 4.2.2a Completion Date: 7/1/19, Reporting date: every 30 days until complete and review every 3 months after; Review date: Annual meeting.

Completed UCIMI brochure, and educational materials about Year 1 work and malaria transmission and control methods. In progress

Activity 4.2.3 Develop Process for Responding to Community Questions and Concerns

Create a formal and standardized process for receiving community questions, concerns, and comments related to the project and activities at field site.  Training on process to be provided to anyone at the site that is working with CE team who is communicating directly with community members.

Milestone 4.2.3a-b Completion Date: 11/30/19, Reporting date: 6/1/19; Review date: Annual meeting.

Activity 4.2.4 Regulatory Planning with Field Site Collaborators– starts June 2019

Initiate conversations to clarify existing regulatory framework and guidelines at field site country in partnership with collaborators .  Determine who regulatory contacts are at field site(s).

Milestone 4.2.4a Completion Date: 12/30/19, Reporting date: every 6 months; Review date: Annual meeting.

Documented regulatory strategy for each field site(s), and list of contacts, and organizational charts.

Activity 4.2.5 Refine Communication Plan

In coordination and collaboration with UCIMI team develop a long-term CE communication strategy for both internal and external stakeholder groups.

Completion Date: 03/31/19

Specific Aim 4.3 Education and Capacity Building

Kormos and Lanzaro – starts June 2019

            Activity 4.3.1 Public Education

Begin delivery of public education to community members and community leaders about the project work and goals (focus groups, community meetings, workshops, Q&A sessions, individual meetings).

Milestone 4.3.1a Completion Date: 12/30/19, Reporting date: every 3 months or as needed; Review date: Annual meeting.

Documented completion of community meetings, outreach and education at all collection sites at field site(s).

Activity 4.3.2 Education/Training program for local educators/CE staff

Develop training strategy and action plan for local, site specific CE personnel who will conduct training and education activities in the communities.  Lecture/workshops presented to University faculty and students about research and gene drive technology.

Milestone 4.3.2a-b Completion Date: 12/1/19, Reporting date: every 3 months; Review date: Annual meeting.

Training for CE staff and focal points onsite is complete.  Workshops at University completed.

Activity 4.3.3 Recruit and Train Onsite CE Personnel

Identify and hire a point of lead CE point of contact for field site(s) who will be responsible for conducting ongoing education, outreach, and community assessments.  Identify focal point contacts at each field collection site who can act as a local liaison for onsite CE contact.

Decision Point 4.3.3a Completion Date: 6/1/19, Reporting date: every 3 months; Review date: Annual meeting.

Hire CE Personnel and have agreements with community focal points.

Activity 4.3.4 Site Regulatory Plan and Preparation- starts December 2019

Facilitate communication for the development of a regulatory process and plan in partnership with UCIMI team and field site government and regulatory officials. 

Milestone 4.3.4a Completion Date: 12/30/19, Reporting date: every 6 months; Review date: Annual meeting.

Identified regulatory contacts, organizational charts and completed regulatory framework for next phase of project.

Specific Aim 4.4 Evaluation and Communication Development

Eubanks – starts January 2020

            Activity 4.4.1 Assessment/Evaluation of Public Education and Tools

Assess the utility and success of tools used to educate communities about project work.  Determine what changes need to be made to tools and adjust accordingly.  Evaluate public understanding and support of our research, goals, and project plan. 

Milestone 4.4.1a-c Completion Date: 7/1/20, Reporting date: 07/01/20; Review date: Annual meeting.

Report of community and stakeholder support for project plan and understanding of goals.  Strategic plan to address concerns and questions. 

Decision Point 4.4.2  Completion Date: 1/30/20, Reporting date: every 6 months; Review date: Annual meeting.

 Activity 4.4.3 Education Tools for Population Modification – starts July 2020

Develop education tools that will be used to explain modified mosquitoes, the release and evaluation process, and next steps. 

Milestone 4.4.3a Completion Date: 7/1/20, Reporting date: 06/01/2020 Review date: Annual meeting.

Education tools for gene drive and modified mosquitoes complete.

Activity 4.4.4 Evaluation of Stakeholder Groups

Assess understanding and support of UCIMI work among project stakeholders.  Determine gaps in knowledge and understanding and strategize how to address them. 

Milestone 4.4.4a-c Completion Date: 7/1/20, Reporting date: every 6 months; Review date: Annual meeting.

Assessment complete and knowledge gaps identified.  Plan completed to address gaps and incorporate into education tools.

Specific Aim 4.5 Communication Management

Eubanks/Kormos – starts 2020

            Activity 4.5.1 Refinement of Communication Plan

Review and adjust CE communication plan and strategy to continue to meet goals of UCIMI project.  Adapt communication strategies to feedback and assessments of stakeholders and community members. 

Milestone 4.5.1a Completion Date: 06/01/2019, Reporting date: as needed; Review date: Annual meeting.

Adjusted communication strategy for ongoing communication needs of community and stakeholder groups.

APPENDIX I: UCIMI DEFINITIONS, TERMS AND CONDITIONS.

Decision-Making Process

Decisions will be made by a consensus of the Principal Investigator, Project Manager and Project Coordinators.  If necessary, the PI will arbitrate disputes to facilitate final decisions.  Evaluation of Activity progress and project plans will be made semi-annually and be based on progress presented at the meeting, draft progress reports (in anticipation of annual reports) and discussions.  The action options available are to: 

1)  Continue the Activity (progress is good).

2)  Modify the Activity (progress is good but unanticipated outcomes or unknowns require changes in the experimental design).

3)  Start a six month re-evaluation period (unsatisfactory progress which, if not remedied, will result in termination of Activity and dependent Activities).

4)  Terminate the Activity even though progress was good because the specific approach no longer contributes to the ultimate goals.

5)  Terminate the Activity because the progress is unsatisfactory and corrections would exceed the project’s timeline.

6)  Conclude the Activity due to its successful completion.

Project Management

Administrative management is the responsibility of the principal investigator (PI, Dr. Anthony A. James - aajames@uci.edu), the project manager (Sentelle Eubanks sentelle@uci.edu ),  office personnel (Rhodell Valdez – rhodellv@uci.eud ) and the professional grants administration at UC Irvine.  Bookkeeping for the subcontractors is handled by the subcontractors’ institutions, with overall review and monitoring by UC Irvine (Sentelle Eubanks).  Scientific Management of Objectives 2 and 3 is carried out by the Principal Investigator, Project Manager and Project Coordinators.  They are responsible for review of scientific goals and progress evaluation.  Review and evaluation criteria are based on the research descriptions and timelines provided in the Project Plan.  An annual meeting in which all scientists participate will facilitate the review process and provide the basis for decision-making about resource allocation.  Programmatic and financial decisions are made by consensus agreement of the Principal Investigator, Project Manager and Project Coordinators with disputes mediated by Principal Investigator.  Resources are allocated initially based on the budgets submitted by the Principal Investigator, Project Manager and Project Coordinators.  At each annual review, progress will be indexed against budget allocations.  Disparities will be corrected by reallocation of funds.  The PI will communicate semiannually with designated administrative officers at UC Irvine to update progress and the project plan.  Information flow among project researchers will be facilitated via the distribution of the semi-annual progress reports as well as through data posting on a secure web site (data postings are announced via email alerts).  Redacted versions of the progress reports will be distributed as hard copies and posted on the secure website. 

Project Management Committee

The Project Management Committee (PMC) has the responsibility of overseeing the protection of Intellectual Property (IP) matters as they develop during the course of the project.  A major goal of the UCIMI is to make new technologies available to the developing world for public benefit.  However, it is beyond the scope of this project to take a product to development.  It is more likely that a successful outcome will be attractive and worthy of investment by the nonprofit and for-profit companies that will be better able to take the project from the trials to the next stages.  It is therefore important to protect the coordinated patentability of the components developed so that a “package” of IP rights (as opposed to numerous individual patents) will be available.  To this end the PMC will review all publications/ presentations of data developed during the course of the project for potential IP material.

Drafts of all publications and presentations of data will be submitted at least 20 days prior to submission for publication or presentation to the Committee for review.  In the event the PMC determines a patent is necessary to protect any discoveries present, the publication/presentation that discloses such inventions will be withheld to allow for the filing of applicable patent applications.  The Committee is not expected to have to meet in person on a regular schedule but should be able to conduct its business electronically on an “as needed” basis. 

PMC permanent members:

            Anthony James ex officio

Sentelle Eubanks Project Manager ex officio

Kevin Kennan - Assistant Director of Intellectual Property Administration at the Office of Technical Alliances, UC Irvine

Members ad hoc:

PCs from the IP originating lab(s)

IP personnel from the originating institution(s)

In the case of the unexpected unavailability of any member at a scheduled meeting they will be replaced by an appropriate (someone having the knowledge, responsibility and authority to act on the IP matter at hand) substitute who is endorsed by the rest of the committee.

Regulatory Compliance

We will maintain regulatory compliance in all areas that apply to project-related activities.  We will monitor and adhere to intramural regulations and guidelines for research protocols and working conditions, as well as extramural regulations for shipping and receiving organisms and permits required to conduct research internationally.  Institutional Review Boards, Institutional Animal Care and Use Committees, and Institutional Biosafety Committees oversee research protocol compliance regarding human subjects, use of vertebrate animals and the use of pathogens and recombinant DNA, respectively.  Working conditions and worker safety are monitored by Environmental Health and Safety programs and certified through Biological Use Authorization.  Records of current protocol approvals are maintained by the project manager.  Copies of all assurances for Select Agents and Conflict of Interest Statements also are to be provided to the Project Manager. The project manager should be notified each time these protocols are reviewed and/or modified.

        The movement of genetically engineered vectors into and within the USA is regulated by the United States Department of Agriculture/Animal and Plant Health Inspection Service (USDA/APHIS).  A shipping permit from the Biotechnology Regulatory Services (BRS) (http://www.aphis.usda.gov/) is required for both inter-state transport as well as importation of genetically-modified (GM) vectors into the USA (APHIS form 2000 - http://www.aphis.usda.gov/brs/pdf/2000.pdf ).  To help keep a record of regulatory compliance, each shipping application (APHIS form 2000) that is associated with the project should be completed with the assistance of and submitted by the Project Manager.  This policy will assure that changes in the USDA application requirements result in an update of the template application.  A two-way permit will cover the possibility that the receiver may want to send vectors back one day.  It should be noted that the application procedure requires an inspection of the recipient’s insectary facilities by the USDA.  The inspection will be the slowest step to the approval process.  If the receiving party has been inspected previously and the sending party has not, it would be quicker to apply for a one-way permit. 

Importation of exotic organisms into the USA is regulated by the Etiologic Agent Import Permit Program of the CDC (http://www.cdc.gov/od/eaipp).   A permit to import a vector of human disease is required according to USPHS 42 CFR - Part 71 Foreign Quarantine.

Part 71.54 Etiologic agents, hosts, and vectors.

(a) A person may not import into the United States, nor distribute after importation, any etiologic agent or any arthropod or other animal host or vector of human disease, or any exotic living arthropod or other animal capable of being a host or vector of human disease unless accompanied by a permit issued by the Director.

(b) Any import coming within the provisions of this section will not be released from custody prior to receipt by the District Director of U.S. Customs Service of a permit issued by the Director (Centers for Disease Control and Prevention).

Applications filed with the CDC for permits to import mosquitoes have, at times, elicited responses indicating that importation permits for this species are unnecessary.  Minimally, we ask that project members who anticipate importing mosquitoes from outside of the US keep these letters on file (with a copy provided to the Project Manager) and have a copy accompany any mosquito shipments into the country.  Preferentially, we ask that you request that your application be considered regardless of the CDC’s interpretation of its necessity. 

The California Department of Public Health (CDPH) also requires anyone wishing to import exotic vectors should first write to the Vector-Borne Disease Section (VBDS), 1616 Capitol Avenue MS 7307, P.O. Box 997413, Sacramento, CA 95899-7413 and provide information about the vector and any pathogens that the vector may carry. In addition, the letter shall include the sources of the vector, the reason for the importation, the time period you will have the vector, security and contingency provisions, plans for disposal of the vector upon completion of the time period, and any other pertinent information. VBDS will write a letter on behalf of the Director of the CDPH authorizing the importation. VBDS may request that a facility be inspected prior to authorization.

California Health and Safety code 116120.

(a) It shall be unlawful for any person to import into the state any exotic vector without written approval from the state department.

(b) The state department shall issue an applicant written authority to import into the state any exotic vector upon a determination by the state department that the public health and safety will not be endangered thereby.

(c) "Exotic vector" means a vector species that is not native to California and is not commonly found in the state.

(d) Any violation of this section is a misdemeanor.

Publicity

Public announcements about the UCIMI must be cleared by UC Irvine communications personnel.

1.  Materials will be reviewed and approved by Tom Vasich, (949) 824-6455, tmvasich@uci.edu.

2.  Correspondence or publicity efforts must indicate that they are being funded by the grantee institution (UC Irvine).  Suggested language to describe affiliation with the project may include: “… supported by the University of California Irvine Malaria Initiative” or “…the University of California Irvine Malaria Initiative.”

3. Additional conditions apply to UCIMI work funded by the Bill and Melinda Gates Foundation (BMGF). Specifically:

PUBLICITY BY UCIMI: The BMGF’s prior written approval must be obtained before: (a) issuing a press release or other public announcement regarding the grant; and (b) any other public use of the Foundation’s name or logo. Requests for permission may be emailed to: grantee.comms@gatesfoundation.org two weeks in advance to provide the BMGF an opportunity to review and comment. Detailed guidelines are available at: www.gatesfoundation.org/grantseeker/documents/guidelines_communications_for_grantees.doc.

PUBLICITY BY OTHERS: The UCIMI, its subgrantees, subcontractors, contingent workers, agents, or affiliates may not state or otherwise imply to third parties that the BMGF directly funds or otherwise endorses their activities.

Publications

Manuscripts, research reports, or other publication resulting from work performed on this project must be reviewed by the Project Management Committee for the evaluation of potential IP property.  Manuscripts intended to be published in peer reviewed journals should be submitted to the Project Manager upon submission of the manuscript to the journal.  Other articles should be submitted to the Project Manager as soon as possible following the creation of a final draft.  Submission of articles to the Project Management Committee should be accompanied by your opinion regarding the potential for IP within the submitted work. All manuscripts will be published as ‘open access’.

The following language for acknowledging support in manuscripts is suggested: “Funded by University of California Irvine Malaria Initiative.” This should be included along with acknowledgements to other sources as needed.

Progress Reports

All researchers are responsible for communicating progress to the Project Manager.  Semi-annual progress reports will be posted on the secure website for review.   Annual progress reports will be posted before the annual meeting for review by all to prepare for the decision making process.  In addition to written progress reports, data from each activity will be presented during the Annual Meetings.

References Cited

Walker PW et al. (2016) Estimating the most efficient allocation of interventions to achieve reductions in Plasmodium falciparum malaria burden and transmission in Africa: a modeling study. Lancet Glob Health 4: e474-e484.

WHO. Guidance framework for testing of genetically modified mosquitoes. World Health Organization on behalf of the Special Programme for Research and Training in Tropical Diseases; 2014. ISBN 9789241507486. ,http://www.who.int/tdr/publications/year/2014/ Guidance_framework_mosquitoes.pdf.

WHO Working Group. Proceedings: Working Group on Strategic Plan to Bridge Laboratory and Field Research in Disease Vector Control. 14-16 July 2004, ICIPE, Nairobi, Kenya. UNICEF/UNDP/World Bank/WHO Special Programme for Research and Training In Tropical Diseases. Frontis Press, Wageningen, The Netherlands, 2004.

World Malaria Report 2018. Geneva: World Health Organization; 2018. Licence: CC BY-NC-SA 3.0 IGO.

Transgenic Anopheles stephensi coexpressing single-chain antibodies resist Plasmodium falciparum development

Alison T. Isaacs, Nijole Jasinskiene, Mikhail Tretiakov, Isabelle Thiery, Agnès Zettor, Catherine Bourgouin, Anthony A. James

Proc Natl Acad Sci U S A. 2012 Jul 10; 109(28): E1922-E1930. Published online 2012 Jun 11. doi: 10.1073/pnas.1207738109

PMCID: PMC3396534

Gantz VM, Jasinskiene N, Tatarenkova O, et al. Highly efficient Cas9-mediated gene drive for population modification of the malaria vector mosquito Anopheles stephensi. Proc Natl Acad Sci U S A. 2015;112(49):E6736-43.

Estimating the most efficient allocation of interventions to achieve reductions in Plasmodium falciparum malaria burden and transmission in Africa: a modelling study Dr. Patrick G T Walker, PhD  Jamie T Griffin, PhD Prof Neil M Ferguson, DPhil

Prof Azra C Ghani, PhD Open Access Published: June 03, 2016DOI:https://doi.org/10.1016/S2214-109X(16)30073-0

CDC https://www.cdc.gov/cpr/ipp/index.htm

CDPH https://www.cdph.ca.gov/Programs/CEH/Pages/CLPR.aspx

USDA https://www.aphis.usda.gov/aphis/resources/permits