A04_-__Statement_of_Work_-_Bat_Genomic_Assessment.pdf

STATEMENT OF WORK

Commercial Service (NAICS 541715)

Genomic Assessment of the Federally Endangered Indiana Bat

PR Number: 0044051176

1. General

1.1 Project Description.

Who: Indiana Ecological Services Field Office (INFO) 620 South Walker Street Bloomington, IN 47403

What: Genomic Assessment of the Federally Endangered Indiana Bat Type of Services: Genomic analysis of Indiana bat, interpretation of analysis, and results report

Where: Using samples collected throughout the Indiana bat’s range by various partners, the successful recipient will complete analyses at a suitable genetics and isotope laboratory(s).

1.2 Purpose/End Application:

The U.S. Fish and Wildlife Service (FWS) requires a genomic analysis to better assess genetic diversity, population structure, connectivity, candidate adaptive differentiation, and potential impacts of stressors on the Indiana bat. These data will inform conservation planning and support the potential identification of biologically meaningful representation (i.e., adaptive capacity) units. Currently, we are reviewing the status of the Indiana bat, a species listed as ‘endangered’ under the Endangered Species Act of 1973 (ESA). Up-to-date information on the species’ genetic diversity, population structure, and potential adaptive differentiation is important for understanding the current and future condition of the species. As a part of this project, an updated range-wide genomic analysis will inform our evaluation of the species’ resiliency (ability to withstand to stochastic events), redundancy (ability to withstand catastrophic events), and representation (ability to adapt to environmental change), which are foundational to a species status assessment. In addition, the genomic assessment will 1) generate range-wide, baseline population genomic estimates of genetic diversity, population differentiation, and (if possible) contemporary effective population size, 2) assess the biological accuracy of established recovery units, and 3) perform foundational exploratory research of the genomic health of this species aimed at improving our understanding of impacts from white-nose syndrome.

1.3 Contractor Qualifications:

The contractor shall demonstrate:

• Documented experience preparing, extracting and sequencing DNA from small biological samples and conducting genomic analyses utilizing the RADseq technique or similar approaches.

• Ability to report, archive, and document analytical procedures, code, and workflows consistent with scientific best practices.

1.4 Other:

The statutory authority for this work is provided under the Endangered Species Act of 1973 (ESA), which establishes protections for fish, wildlife, and plants that are classified as threatened and endangered species. The ESA requires preparation and implementation of plans for the recovery of each listed species. This research specifically supports Recovery Action 3.5.3 Conduct additional studies of Indiana bat population genetics based on sampling at hibernacula from the 2007 Draft Indiana Bat Recovery Plan.

2. Place of Performance and Period of Performance Place of Performance: Suitable genetics lab with qualified wildlife geneticists Period of Performance: October 2026 – December 2027

2.1 Place of Performance

Work is to be performed at the contractor site.

2.2 Period of Performance

Performance Start Date: October 1, 2026 Performance End Date: December 31, 2027

Over the past two years, INFO, in coordination with partners, has actively collected genetic samples (i.e., wing punches) throughout the range of the Indiana bat. The successful recipient would begin processing and analyzing available samples in early October 2026. A limited number of additional samples may continue to be collected by FWS biologists and partners at targeted hibernacula through March 2027.

2.3 Normal duty hours are Monday through Friday 8:00 a.m. - 4:30 p.m. In the event of emergencies, Contractor shall provide contact information.

2.4 Non-duty hours include week nights, weekends, and Government holidays (New Year’s Day, Martin Luther King Jr.s’s Birthday, Washington’s Birthday, Memorial Day, Juneteenth, Independence Day, Labor Day, Columbus Day, Veterans Day, Thanksgiving Day, Christmas Day).

3. Scope

3.1 The contractor shall extract total genomic DNA from all samples provided by the FWS and cooperating partners, generate sequence reads via restriction site-associated sequencing (RADseq) library prep, sequencing via the Illumina NovaSeq X+ platform, conduct bioinformatic processing, analyze the resultant data, and submit a report utilizing their expertise, services and training.

Task 1: DNA Extraction and Quality Control

• Extract high-quality total genomic DNA from approximately 200-400 tissue or DNA samples (e.g., wing punches from 20 individual bats collected from approximately 10 different sites across the range).

• Perform DNA quantification and quality assessments to ensure suitability for sequencing.

Task 2: RADseq Library Preparation and Sequencing

• Prepare RADseq libraries using a protocol appropriate for the target taxa.

• Sequence libraries on the Illumina NovaSeq X Plus platform or equivalent, generating sufficient depth for individual-level inference (i.e., >15-20x coverage per individual).

Task 3: Bioinformatic Processing

• Process raw sequencing reads, including demultiplexing, adapter removal, and quality filtering.

• Perform de novo or reference-guided alignment and SNP discovery.

• Filter SNPs based on quality metrics and prepare datasets suitable for downstream analyses.

Task 4: Genomic Analysis

• Conduct PCA and STRUCTURE/Admixture (or similar population assignment analysis) to identify potential distinct genetic clusters.

• Estimate gene flow and connectivity among populations using appropriate statistical models

• Estimate genetic diversity metrics and contemporary effective population sizes of populations.

• Evaluate genetic signatures of stress or reduced diversity potentially attributable to threats such as habitat loss, disease, or environmental change.

• Estimate potential adaptive differentiation based on environmental factors such as climate using genotype-environment association tests.

4. Deliverables and/or Delivery Schedule

4.1 Required deliverables include a comprehensive genomic assessment of the Indiana bat genetic diversity, including, but not limited to, nucleotide diversity, expected and observed heterzygosity, inbreeding coefficients, and contemporary effective population size estimates for the purposes of learning fundamental insights on the genomic structuring and veracity of the species. Additionally, assessment of population differentiation, gene flow and connectivity, and potential adaptive differentiation, as discussed above. These assessments shall specifically address baseline population genomic estimates, relevance to white-nose syndrome impacts, as well as biological relevance of current recovery units (see 2007 draft recovery plan for the Indiana bat).

Deliverable Schedule:

Deliverable Description Due Date

DNA extraction and QC report Summary of extraction and quality metrics Dec. 31, Raw sequence data FASTQ files submitted to INFO or uploaded to a secure repository

Apr. 31, Final technical report Includes full analysis and interpretation Dec. 31, Data files and scripts All processed data, scripts, and metadata Dec. 31, In addition, the contractor shall provide brief monthly updates on the progress of deliverables to the primary POC at the Indiana Ecological Services Field Office via email and calls, with a final written technical report on file by the close of award.

4.2 Identify the type of documentation (printed and electronic-such as manuals) to be provided, as well as those quality indicators desired by the Government.

The contractor shall provide an electronic scientific technical report (PDF) detailing the methods, results, and relevant discussion of the Genomic Assessment of the Federally Endangered Indiana Bat. This report should be written in a similar style and format as a manuscript being prepared for publication in a peer-reviewed scientific journal. The findings within the report may be published at some point, but this is not a requirement under this award.

5. Data Management Plan

Raw sequence data will be permanently archived in the NCBI Sequence Read Archive before project completion. Filtered VCFs, metadata, and all code used for data filtering, analysis, and report production will be uploaded to Dryad or equivalent publically-available data archive system before project completion.

6. Government-Furnished Equipment (GFE) / Government-Furnished Information (GFI)

6.1 N/A

7. Evaluation and Acceptance

7.1 The contractor will be evaluated as detailed herein. Government acceptance is contingent upon:

Satisfactory fulfillment of deliverables identified in section 4.1.

8. Quality Control/Quality Assurance:

The contractor shall adhere to the following QA/QC measures or similar alternatives…

1. Sample Handling & Chain-of-Custody

• Maintain clear chain-of-custody documentation for all samples.

• Record sample condition on receipt and ensure proper storage (−20°C or −80°C).

• Handle degraded or low-yield samples using standardized procedures.

2. DNA Extraction Quality Control

• Quantify DNA using fluorometric methods and document purity (A260/280, A260/230).

• Verify DNA integrity (e.g., TapeStation/Bioanalyzer).

• Re-extract or flag samples below quality thresholds.

3. Library Preparation QC

• Randomize individuals included in each library across sampling locations to avoid correlations between location and library.

• Include technical replicates (≈5%) to assess genotyping consistency.

• Verify library size and integrity using fragment analysis tools.

• Quantify libraries using qPCR-based methods.

• Ensure adapter-dimer levels and insert sizes meet minimum standards.

4. Sequencing Quality Control

• Include internal sequencing controls (e.g., PhiX).

• Report run metrics: Q30 scores, cluster density, error rates.

• Meet or exceed defined minimum sequencing depth per sample.

• Re-sequence samples or batches that fail quality metrics.

5. Bioinformatics QC

• Document versions and parameters for all software and pipelines.

• Conduct read-quality checks (FastQC/MultiQC).

• Remove low-quality reads, adapters, and contaminants.

• Report mapping rates (if using reference), missing data levels, and duplicate read summaries.

6. Variant Calling & Filtering QC

• Use recognized variant callers with transparent filtering thresholds.

• Apply minimum depth, genotype quality, and missing-data criteria.

• Flag loci failing basic population genetic assumptions (e.g., extreme heterozygosity).

7. Reproducibility & Replication

• Conduct replicate extractions or library preparations for a subset of samples.

• Demonstrate consistency between replicates within defined tolerance limits.

• Provide workflow scripts or pipeline documentation.

8. Data Management & Deliverables QC

• Deliver all raw FASTQ files, BAM/VCF files, metadata, and QC summaries.

• Provide checksums (e.g., MD5) for file integrity verification.

• Confirm all data are complete, readable, and correctly labeled.

9. Reporting Requirements

• Summarize all QA/QC metrics in the final report.

• Identify any sample failures and reasons.

• Document all deviations from standard protocols and corrective actions taken.

9. Technical Coordinators:

9.1 Primary technical point of contact

Indiana Bat Recovery Lead:

Andrew King Deputy Field Supervisor Indiana Ecological Service Field Office 620 South Walker Street Bloomington, IN 47404 Email: andrew_king@fws.gov Phone: 812-902-1809

9.2 Other cooperating and technical contacts

USFWS BSSS Geneticist (primary genomic technical advisor/POC) mailto:andrew_king@fws.gov

Dr. Brenna Forester Branch of Species Status Assessment Science Support

Division of Endangered Species U.S. Fish and Wildlife Service Email: brenna_forester@fws.gov

Region 3 White-nose Syndrome Coordinator:

Richard Geboy Fish and Wildlife Biologist Indiana Ecological Service Field Office Email: richard_geboy@fws.gov Phone: 812-902-1796

Program Manager:

Susan Cooper Field Supervisor Indiana Ecological Service Field Office Email: susan_e_cooper@fws.gov Phone: 812-902-1792 mailto:brenna_forester@fws.gov mailto:richard_geboy@fws.gov mailto:susan_e_cooper@fws.gov