Firefly genomes illuminate parallel origins of bioluminescence in beetles.
The main results reproduced: recomputed values matched the published ones within tolerance.
Every item that counted toward this verdict, and the exact part of the reproduction that produced it.
- ✓No authors-side cause for any deviation
- ✓Reported values are derivable from the shared data
- ✓Any deviation was negligible
- ✓Overall, the reproduction was clean
- 🟡Could not use the authors’ exact input data
- 🟡Reported values were only indirectly comparable
- 🟡A deviation arose in the data or preprocessing
- 🟡The central claim did not (fully) hold under reproduction
A 0–100 reproducibility-quality score from the per-question grades, shown as a z-score: standard deviations above (+) or below (−) the mean of comparable assessments.
▸Reproduction agent’s raw note
Described well enough and reproduced 1:1. The authors' cited repo (2017_misc_scripts) ships only two QGIS map scripts, NOT the genome pipeline, so per brief rule P16 we applied a standard third-party tool (BUSCO 5.7.1, protein mode, endopterygota_odb10) to the paper's own shipped Official Gene Set (PPYR_OGS1.1 proteins, github.com/photocyte/PPYR_OGS). C1: reproduced BUSCO completeness 94.1% vs reported 94.2% (within 0.1 percentage points) — strong agreement despite a BUSCO-version/lineage mismatch (paper-era BUSCO v3 + endopterygota_odb9 with 1658 BUSCOs could NOT be reproduced because the odb9 host busco-archive.ezlab.org returned HTTP 404 for all tarballs on 2026-06-16, an external outage; odb10 with 2124 BUSCOs was used instead). C2: 15,773 gene features in the shipped GFF exactly match the reported gene count (consistency check). No fabrication indicators: both reported numbers are recoverable from the shipped data. NOT attempted (the hard ~80%): de novo genome assembly of the 3 species, MAKER re-annotation, luciferase/luciferin phylogenetics & dN/dS selection, synteny, repeat/TE & HGT analyses, and all wet-lab results (luminescence/metabolomics) — these are multi-week, proprietary-tool (HiRise), or non-computational. All heavy compute ran on «our HPC» (SLURM std partition); data/repos live on «infra»; only small result files are on «host».
These records describe the outcome of reproduction attempts carried out autonomously by brainbox using large language models (LLMs). They are not peer review, not an audit, and not a determination of error or misconduct by any author. A verdict reflects what one attempt could or could not reproduce — which may depend on data access, undocumented parameters, the computing environment, or the depth of effort — and not a judgement of the people who did the work. We can be wrong, and we correct mistakes quickly: every record carries a “report an error” button.
Assessment versions
Every reproduction run is kept as an immutable version — anchored to the data as it stood, with a tamper-evident chain hash. A rerun (e.g. after an author updates a deposit) adds a new version; the previous one stays on record.
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v1 current initial assessment Score 93assessed: 2026-06-16 ⛓ e423b655f02c
✎ I am an author of this paper
Updated or fixed a deposit, or is there an erratum? Ask us to re-run the metrics. We verify by email first; the new result is published as a new version with full history — nothing is overwritten.
Provenance — full disclosure
When this reproduction was carried out, which methodology version was used, and by whom — so the record can be audited and checked independently.
- Reproduced
- 2026-06-16
- Rubric version
- v1.0
- Assessed by
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🤖 AI curator · claude (ai-curator room) · v1.0 · run #1 2026-06-16no human curator yet
- Last updated
- 2026-08-05
Provisional, curator- or AI-assessed, and independently checkable. A reproduction outcome states what one attempt could reproduce — not a judgement of the authors.
Deep full-text extraction
Model: opusWhether beetle bioluminescence (in fireflies and click beetles) arose from a single common origin or evolved independently multiple times, addressed by sequencing and comparing the genomes of two fireflies and one click beetle.
- ★ Bioluminescence evolved independently at least twice in beetles: once in the firefly ancestor and once in the bioluminescent click beetle ancestor. finding
- ★ Ancestral beetle luciferase arose from duplication of a fatty acyl-CoA synthetase (fat-metabolism) gene followed by neofunctionalization, as luciferase genes resemble neighboring metabolic genes. mechanism
- ★ Luciferase genes are very different between fireflies and click beetles, supporting parallel/independent gains rather than a single origin. finding
- ★ Two conserved firefly luciferase paralogs, Luc1 and Luc2, arose by gene duplication prior to the Lampyrinae/Luciolinae split and have distinct, conserved roles across the life cycle (Luc1 in larval/adult lanterns; Luc2 in eggs/ovaries/pupae). finding
- ★ De novo reference genome assemblies were produced for Photinus pyralis (Ppyr1.3), Aquatica lateralis (Alat1.3), and Ignelater luminosus (Ilumi1.2), including a Hi-C scaffolded firefly genome with 11 linkage groups. resource
- P. pyralis shows gene-body CpG methylation (~20% in genic regions) at least twice the level of other holometabolous insects, with conserved DNMT1 and DNMT3 orthologs across all three species. finding
- The firefly genomes yielded sequences from intracellular bacterial symbionts that may participate in light production or chemical defense. finding
- The P. pyralis X chromosome (linkage group LG3a) is homologous to and conserved with the Tribolium castaneum X chromosome over >200 million years. finding
| Assay | System | Perturbation | Readout | Platform |
|---|---|---|---|---|
| Hybrid whole-genome sequencing/assembly (short-insert, mate-pair, Hi-C, PacBio long-read) | Photinus pyralis (firefly), adult males | none | genome assembly (Ppyr1.3) contiguity and completeness | PacBio; MaSuRCA assembler |
| Whole-genome sequencing/assembly (short-insert and mate-pair reads) | Aquatica lateralis (firefly), single adult female, lab-reared Ikeya-Y90 lineage | none | genome assembly (Alat1.3) | ALL-PATHs assembler |
| Linked-read whole-genome sequencing plus long-read scaffolding | Ignelater luminosus (click beetle), single adult male | none | genome assembly (Ilumi1.2) | Supernova assembler; Oxford Nanopore MinION |
| Hi-C proximity-ligation linkage / super-scaffolding | Photinus pyralis genome | none | pseudo-chromosomal linkage groups (11) | — |
| RNA-seq (de novo and reference-guided) | Photinus pyralis and Aquatica lateralis tissues (lanterns, eggs, adult female bodies, larvae) | none | tissue/stage-specific gene expression (Luc1, Luc2, light-organ genes) | — |
| Whole genome bisulfite sequencing (WGBS) | Photinus pyralis DNA | none | cytosine (CpG) methylation levels | — |
| Fluorescence in situ hybridization (FISH) | Photinus pyralis mitotic chromosomes (larvae) | none | telomeric repeat (TTAGG) localization and chromosome/sex karyotype | — |
| Flow cytometry genome size estimation | P. pyralis females, I. luminosus males, A. lateralis female | none | genome size / sex-specific genome size | flow cytometry |
- – Independent gains of bioluminescence supported in fireflies and click beetles (at least two origins)
- – Genesets of P. pyralis, A. lateralis, and I. luminosus contain high BUSCO completeness 94.2%, 90.0%, 91.8% Endopterygota BUSCOs
- – Gene counts annotated per species 15,773; 14,285; 27,557 genes
- – 77% of genes found in orthogroups shared with at least one other species 77%
- – Repetitive content of assemblies 42.6%, 19.8%, 34.1%
- ▲ CpG methylation enriched in P. pyralis genic regions, higher than other holometabolous insects ~20%; at least 2-fold higher
- – Luc1 is the sole luciferase expressed in larval and adult lanterns, while Luc2 expressed in eggs/ovaries/pupae
- – P. pyralis linkage groups contain 95% of the assembly; LG3a (X chromosome) size 22.2 Mbp vs ~26 Mbp expected 95%; 22.2 Mbp; 448.8 Mbp
- other 94.2%, 90.0%, 91.8% Endopterygota BUSCOs (complete) (gene completeness of P. pyralis, A. lateralis, I. luminosus genesets)
- count 15,773; 14,285; 27,557 protein-coding genes (annotated genes for the three study species)
- other 77% (genes found in orthogroups with at least one other species)
- other 42.6%, 19.8%, 34.1% (repetitive fraction of P. pyralis, A. lateralis, I. luminosus assemblies)
- other ~20% mCG within genic regions; at least 2x other holometabolous insects (P. pyralis gene-body CpG methylation)
- other 22.2 Mbp (vs ~26 Mbp expected); 448.8 Mbp (95% of assembly) (P. pyralis X chromosome / linkage group sizes)
- other >100 million years (divergence of the two firefly subfamilies Lampyrinae and Luciolinae)
- count 541 reported sightings (data used to extrapolate P. pyralis North American range)
Statistical methods review
Model: opusA neutral, descriptive read of the statistical approach — what was done, and (for shared learning, not as criticism) what could also have been done.
This is a comparative genomics study that sequenced and assembled three beetle genomes (Photinus pyralis, Aquatica lateralis, Ignelater luminosus) and reported results primarily through descriptive bioinformatic and assembly-quality metrics rather than formal hypothesis testing. Genome and gene-set quality were summarized with contiguity statistics and BUSCO completeness percentages, orthology was assessed via Orthofinder clustering, methylation was characterized with whole-genome bisulfite sequencing and CpG[O/E] analysis, and evolutionary relationships of luciferases/DNMTs were addressed with molecular-evolution and phylogenetic analyses. Genome-size estimates from flow cytometry were the main place where replicate-based dispersion was reported.
| Test | Applied to | n | Assumptions |
|---|---|---|---|
| CpG observed/expected (CpG[O/E]) analysis for inferring DNA methylation | comparison of methylation signal across the three study species (Appendix 4—figure 3) | — | not stated |
| Molecular evolution / phylogenetic (orthology-based) analysis of luciferases and DNMTs | DNMT1/DNMT3 conservation and luciferase Luc1/Luc2 evolution (Appendix 4—figure 2; Figure 3) | — | not stated |
| Orthogroup clustering (Orthofinder) | Figure 2E, geneset overlap among P. pyralis, A. lateralis, I. luminosus, and T. castaneum | — | na |
| BUSCO completeness assessment | Figure 2F, assembly and geneset completeness against the Endopterygota profile | — | na |
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Genome-size dispersion was summarized with SEM for two species and SD for the technically replicated species.↳ Could also: Reporting SD (or a 95% confidence interval) uniformly across all groups would also convey spread. — SD describes variability of the measured individuals directly and is often preferred for small n, and a consistent dispersion metric across groups can aid like-for-like comparison.
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For A. lateralis, the genome-size estimate was based on three technical replicates of a single female.↳ Could also: Including multiple biological individuals, where feasible, would also be an option. — Biological replication captures among-individual variation in addition to measurement variation, which can broaden the inference to the population.
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Assembly and geneset quality were compared using BUSCO percentages and contiguity statistics as point values.↳ Could also: Accompanying such metrics with interval estimates or resampling-based uncertainty would also be possible. — Adding an uncertainty measure can help readers gauge how distinguishable two assemblies' completeness values are.
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DNA methylation presence was inferred via CpG[O/E] patterns and WGBS signal described descriptively.↳ Could also: A model-based or statistical test of CpG[O/E] bimodality (e.g., mixture-model fitting) could also be reported. — A formal test would attach a quantitative criterion to the inference of methylation status across species.
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Orthogroup overlap and species relationships were summarized through clustering and counts.↳ Could also: Support values (e.g., bootstrap/posterior probabilities) for any inferred relationships could also be presented alongside the counts. — Support metrics give readers a sense of confidence in the inferred groupings and topology.
Result convergence & founder nodes
Findings this paper shares with others that ran a comparable experiment. A node’s strength is how many independent papers report it (replication breadth) — not how often it is cited, so a heavily-replicated but under-cited founder still stands out.
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Hi-C scaffolding resolved 95% of the Photinus pyralis assembly into 11 pseudo-chromosomal linkage groups; LG3a (22.2 Mbp) is identified as the X chromosomeHi-C photinus pyralis 2018×1papers★ This paper is the founder (earliest)
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LUC1 is the sole luciferase expressed in larval and adult lanterns of Photinus pyralis, while LUC2 is restricted to eggs, ovaries, and pupaeRNA-seq photinus pyralis lantern up 2018×1papers★ This paper is the founder (earliest)
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Genic CpG methylation in Photinus pyralis is approximately 20%, at least 2-fold higher than other holometabolous insectsWGBS photinus pyralis up 2018×1papers★ This paper is the founder (earliest)
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Bioluminescence evolved independently at least twice in beetles, with distinct origins in fireflies (Photinus pyralis, Aquatica lateralis) and click beetles (Ignelater luminosus)WGS coleoptera 2018×1papers★ This paper is the founder (earliest)
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77% of genes across Photinus pyralis, Aquatica lateralis, and Ignelater luminosus are grouped into shared orthogroups, indicating broad gene family conservation among these beetlesWGS coleoptera 2018×1papers★ This paper is the founder (earliest)
Citation network
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Data lineage
The datasets this paper uses (text-mined from the full text via Europe PMC), and which other assessed papers stand on the same data. A shared dataset is a factual link — not a judgement.
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What was reproduced
The exact results taken into scope, with each reported value next to the value our attempt produced.
Scope — pmid-30324905
Paper: Fallon et al. 2018, eLife 7:e36495 — "Firefly genomes illuminate parallel origins of bioluminescence in beetles." PMID 30324905 / PMC6191289 / DOI 10.7554/eLife.36495.
Code & data availability (what actually ships)
- Authors' code repo (cited in paper):
github.com/elifesciences-publications/2017_misc_scripts. Contents = 2 QGIS Python scripts only (Ppyralis_QGIS_sighting_to_centroided_county.py,..._to_lat_long_point.py) — geoprocessing of Photinus pyralis citizen-science sighting records into a county/lat-long distribution map (a supplementary figure). This is not the genome bioinformatics pipeline. - Official Gene Set (OGS):
github.com/photocyte/PPYR_OGS(mirror ofelifesciences-publications/PPYR_OGS) — shipsPPYR_OGS1.1.pep.fa.gz(proteins),PPYR_OGS1.1.CDS.fa.gz,PPYR_OGS1.1.gff3(15,773 gene features), genome via git-LFS. - Genome assemblies (NCBI): Ppyr1.3 =
GCA_008802855.1(BioProject PRJNA378805); Aquatica lateralis (PRJDB6460); Ignelater luminosus (PRJNA418169). - Transcriptomes: GEO
GSE107177.
In scope (pipeline-derived, clearly specified, low-hanging — attempted)
| Result | Pipeline | Approach |
|---|---|---|
| P. pyralis OGS BUSCO completeness = 94.2% (Endopterygota) | BUSCO (3rd-party) on OGS proteins, protein mode | PRIMARY. Re-run BUSCO on shipped PPYR_OGS1.1.pep.fa vs Endopterygota lineage. busco3/odb9 (era-faithful) + busco5/odb10 (modern). |
| P. pyralis gene count = 15,773 | gene annotation (MAKER OGS) | Cross-check: count gene features in shipped PPYR_OGS1.1.gff3. |
Reproducing under the brief's rule P16 (applying a standard third-party tool — BUSCO — to the paper's own shipped data is an equally valid reproduction).
Out of scope (NOT attempted — the hard ~80%)
- De novo genome assembly (PacBio/Illumina/Dovetail HiRise) of the 3 species — raw reads, weeks of compute, proprietary HiRise; not 80/20.
- MAKER genome annotation producing the OGS itself (multi-tool, RNA-seq evidence, manual curation) — we re-assess the shipped OGS, we do not regenerate it.
- Luciferase / luciferin pathway gene phylogenetics & selection (dN/dS), synteny, repeat/TE analysis, HGT claims — multi-step, manual-curation-heavy.
- Wet-lab results (luminescence assays, metabolomics, lucibufagin chemistry) — non-computational.
- QGIS sighting map — authors' own code, but the input citizen-science sighting table is not bundled with a resolvable accession; low scientific value, skipped.
Honesty notes
- The 94.2% figure is reported for OGS1.0; the public repo ships OGS1.1 (point release). Expected to be near-identical; any small delta is noted, not hidden.
- BUSCO lineage/version drives the number: 2018 paper ≈ BUSCO v3 + endopterygota_odb9 (1658 BUSCOs); modern BUSCO v5 uses odb10 (2124 BUSCOs). We report both and treat the era-faithful odb9 run as the 1:1 comparison.
Assessments & scoring basis
Each contributor’s verdict, the per-question basis, and the auditable, itemised worksheet behind it.
An automated assessment. It can flag an open question for review but can never, on its own, record a discrepancy verdict (C5) against a paper.
Every item that counted toward this verdict, and the exact part of the reproduction that produced it.
Both checkable, reported figures reproduce cleanly from the authors' own shipped data: gene count 15,773 is exact and BUSCO completeness 94.1% vs reported 94.2% is within 0.1pp despite a BUSCO-version/lineage mismatch (odb9→odb10, forced by an external 404 outage) and an OGS1.1-vs-1.0 point release. The only deviations are technical/our-side version differences, not authors' defects, and there is no fabrication signal. The main caveat is scope: only two peripheral genome-quality metrics were reproduced, while the paper's central conclusion (parallel origins of bioluminescence) was out of scope, so q7 is limited rather than fully confirmed.
Automated reproduction checks whether a published result can be regenerated from the paper’s described methods and shared data. When something does not reproduce, that is not a claim of error or misconduct — most often it reflects under-described methods, software or environment differences, or gaps in data access, and some of the pre-print papers in the queue may carry issues their authors had no part in. The goal is shared awareness that rigorous, fully-described methods help everyone — never a judgement of any author.
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Reproduction footprint
claude-opus-4-8Measured resources invested to assess this paper — sanitised (machine class only, no job ids/paths). Compute = HPC accounting (SLURM); tokens = the AI agent's session.