Comparative Genomics Provides Insight into the Function of Broad-Host Range Sponge Symbionts.
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.
- ✓Any deviation was negligible
- 🟡Could not use the authors’ exact input data
- 🟡Reported values were only indirectly comparable
- 🟡A deviation arose in the data or preprocessing
- 🟡A deviation was attributed to the published material
- 🟡Reported values were not (fully) derivable from the shared data
- 🟡The central claim did not (fully) hold under reproduction
- 🟡Overall, the reproduction showed a material discrepancy
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
REPRODUCED 1:1. Named code = third-party barrnap 0.9 (https://github.com/tseemann/barrnap), applied per P16 to the paper's own deposited Table-1 genome AqS2 = GCA_001750625.1 (Candidatus Amphirhobacter heronislandensis, the A. queenslandica symbiont). All four checkable pipeline-derived claims reproduced EXACTLY on «our HPC» SLURM: barrnap recovers 1 full-length 16S (1536 bp) + 23S + 5S (C1); genome size 1,608,671 bp = 1.61 Mbp matches Table 1 (C2); CheckM lineage_wf completeness 71.13% (C3) and contamination 1.52% (C4) match Table 1 TO THE DIGIT, despite a CheckM v1.0.12->v1.2.2 version bump (same 2015 marker DB). Dataset PRJNA508092 profiled: 94 ENA runs = 85 Illumina 16S-amplicon + 9 Ion-Torrent WGS metagenome, all metagenomic; only 61 runs belong to this 2021 paper (33 are an unrelated 2025 extension grafted onto the same BioProject), and the paper's analysed MAGs live in JGI/IMG/NCBI-Assembly not in this raw-reads deposit (delivers_promised=partial). NOT attempted (honest, no completeness claim): the multi-genome comparative analyses (JGI-gated MAGs) and stochastic de-novo binning.
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 75assessed: 2026-06-18 ⛓ e085ae4dd63d
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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
- 2026-06-25
- Rubric version
- v1.0
- Assessed by
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🤖 AI curator · claude (ai-curator room) · v1.0 · run #1 2026-06-19no 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: sonnetThe paper tests whether the broad-host-range sponge symbiont order Tethybacterales has functionally diversified across its constituent families and lineages since an ancient association with sponge hosts, and how its functional repertoire and host preference compare to the well-characterized Entoporibacteria/Poribacteria.
- ★ Eleven new genomes were added to the Tethybacterales order and a novel family (Polydorabacteraceae) was identified finding
- ★ Functional potential differs between the three Tethybacterales families finding
- ★ Tethybacterales preferentially associate with low-microbial-abundance (LMA) sponges while Entoporibacteria preferentially associate with high-microbial-abundance (HMA) sponges finding
- ★ Tethybacterales and Poribacteria have distinct functional repertoires and these families can coexist within a single sponge host finding
- ★ Functional potential tracks taxonomic ranking rather than host-specific adaptation, suggesting multiple independent association events rather than a single ancestral association followed by coevolution finding
- ★ Tethybacterales may represent a more ancient lineage of ubiquitous sponge-associated symbionts finding
- The Sp02-1 (bin 003B_4) genome shows hallmarks of genome reduction, with an abundance of pseudogenes and low coding density finding
- Shared genes across all 17 Tethybacterales genomes, including chorismate synthase, suggest a common role as tryptophan producers for sponge hosts unable to synthesize tryptophan mechanism
| Assay | System | Perturbation | Readout | Platform |
|---|---|---|---|---|
| Genome binning/metagenomic assembly | Tsitsikamma favus sponge metagenomes and 36 additional sponge SRA data sets (14 sponge species) | none | Metagenome-assembled genomes (MAGs)/bins | — |
| 16S rRNA gene phylogenetic analysis (UPGMA) | Tethybacterales Sp02-1 and related 16S rRNA sequences | none | Phylogenetic relatedness/tree topology | MEGA X |
| Multi-locus phylogenomics using single-copy marker genes (autoMLST) | 17 Tethybacterales genomes/MAGs | none | Phylogenetic tree and family delineation | autoMLST with IQ-TREE and ModelFinder |
| Average amino acid identity (AAI) and pairwise 16S identity comparison | Tethybacterales genomes across three families | none | Pairwise sequence identity scores | — |
| Genome quality assessment (completeness/contamination, MIMAG standards) | 27 putative Tethybacterales bins/genomes | none | Completeness %, contamination %, quality tier (medium/low) | — |
| Comparative genome annotation/metabolic pathway reconstruction | Tethybacterales Sp02-1 (bin 003B_4) genome | none | Presence/absence of genes for glycolysis, PRPP biosynthesis, amino acid biosynthesis and transport | — |
| Orthologous gene clustering and hierarchical clustering of gene presence/absence | 17 Tethybacterales genomes | none | Orthogroup counts and clustering pattern by family | — |
| Unique gene identification (comparative genomics) | Bin 003B_4 versus four T. favus metagenomes | none | Genes unique to the Sp02-1 symbiont | — |
- – Bin 003B_4 genome is ~2.95 Mbp, medium quality, with ~25% pseudogenes and 65.27% coding density
- – New family Polydorabacteraceae identified, sharing on average 80% AAI within the family 80% AAI
- – The three Tethybacterales families share less than 89% 16S rRNA sequence similarity, with intraclade differences of less than 92% <89%/<92%
- – 4,306 orthologous gene groups identified among 17 Tethybacterales genomes, but only 18 genes were common to all 18/4,306 orthogroups
- – Chorismate synthase genes were found across all 17 genomes, suggesting shared tryptophan production capacity
- – Gene presence/absence pattern of bin 003B_4 most closely resembles other Persebacteraceae genomes (from Crambe crambe, Crella incrustans, Scopalina sp.)
- – Thirteen genes were unique to Sp02-1 relative to other T. favus metagenomes, including phage-associated genes and the antirestriction protein ArdA 13 genes
- – 16S rRNA gene of bin 003B_4 shares 99.86% identity with the Sp02-1 clone (HQ241787.1) 99.86%
- other 99.86% (16S rRNA identity between bin 003B_4 and Sp02-1 clone HQ241787.1)
- other 2.95 Mbp (Genome size of bin 003B_4)
- count ~25% pseudogenes (Pseudogene content of bin 003B_4)
- other 65.27% (Coding density of bin 003B_4)
- other 80% AAI (Average amino acid identity within newly proposed Polydorabacteraceae family)
- other <89% (interfamily), <92% (intraclade) (16S rRNA sequence similarity among/within Tethybacterales families)
- count 4,306 orthogroups; 18 shared genes (Orthologous gene clustering across 17 Tethybacterales genomes)
- count 13 unique genes (Genes unique to Sp02-1 relative to other genes in four T. favus metagenomes)
Statistical methods review
Model: sonnetA 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 of Tethybacterales sponge symbionts. The primary analytical approaches were phylogenetic inference (UPGMA with 10,000 bootstrap replicates for 16S rRNA trees in MEGA X; maximum-likelihood via IQ-TREE with ModelFinder in autoMLST for multi-locus marker gene trees), pairwise average amino acid identity (AAI) and 16S rRNA sequence identity comparisons, and hierarchical clustering of orthologous gene presence/absence data. Genome bins were filtered by quality tier using MIMAG standards; results were reported as percentage identity values, bootstrap support values, and genome metadata (completeness, contamination, size).
| Test | Applied to | n | Assumptions |
|---|---|---|---|
| UPGMA phylogenetic inference with bootstrap support (10,000 replicates) | 16S rRNA gene phylogeny of Tethybacterales Sp02-1 bin relative to closest relatives (Fig. S2) | 17 16S rRNA gene sequences, 1,291 positions | not stated |
| Maximum composite likelihood (evolutionary distance calculation) | Branch lengths in 16S rRNA UPGMA tree (Fig. S2) | 17 sequences | not stated |
| Maximum-likelihood phylogenetic inference (IQ-TREE with ModelFinder, via autoMLST de novo pipeline) | Whole-genome marker gene phylogeny of 17 Tethybacterales genomes (Fig. 1) | 17 medium-quality genomes/MAGs | not stated |
| Pairwise average amino acid identity (AAI) | Family-level classification of Tethybacterales genomes (Table S6) | 17 genomes | na |
| Pairwise 16S rRNA gene sequence identity | Family- and class-level boundary assessment (Table S6; <89% between families, <92% intraclade) | — | na |
| Hierarchical clustering of orthologous gene presence/absence | Gene content similarity among 17 Tethybacterales genomes (Fig. 2A) | 4,306 ortholog groups across 17 genomes | not stated |
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The 16S rRNA gene tree was inferred with the UPGMA method↳ Could also: Maximum-likelihood (e.g., IQ-TREE with ModelFinder) or Bayesian inference (e.g., MrBayes, BEAST) could also be applied to the 16S rRNA alignment — UPGMA assumes a molecular clock (equal evolutionary rates across lineages); ML and Bayesian methods relax this assumption and are generally preferred for reconstructing microbial phylogenies where rate variation among lineages is likely
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Branch support for the 16S rRNA tree was assessed with 10,000 bootstrap replicates↳ Could also: Bayesian posterior probabilities (via MrBayes or BEAST) or ultrafast bootstrap approximation (UFBoot2 in IQ-TREE) could also quantify branch support — Bayesian posteriors and UFBoot2 often converge faster and can provide complementary or more calibrated confidence measures; reporting both bootstrap and posterior support is common practice in phylogenomics
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Family-level boundaries were assessed using a fixed pairwise AAI threshold (~80% within-family) and 16S rRNA identity thresholds (<89% between families)↳ Could also: Percentage of conserved proteins (POCP) or whole-genome average nucleotide identity (ANI) could also be used as complementary boundary criteria — POCP is increasingly used alongside AAI for higher-rank prokaryotic taxonomy and can distinguish genus- from family-level relationships; ANI is widely adopted for species-level delineation and together these metrics provide a multi-criterion view of genomic relatedness
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Gene content similarity across genomes was visualized via hierarchical clustering of ortholog presence/absence↳ Could also: Ordination methods such as principal coordinates analysis (PCoA) on a Jaccard or Bray-Curtis dissimilarity matrix of gene presence/absence could also represent the same data — PCoA/NMDS displays the full multivariate structure in a low-dimensional space that can be directly inspected for clustering patterns, and dissimilarity metrics can be chosen to weight shared-absence differently from shared-presence, which matters for incomplete MAGs
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Genome bins were retained or excluded based on MIMAG completeness/contamination tiers without a stated sensitivity analysis on the quality cutoff↳ Could also: Supplementary analyses retaining low-quality bins or applying GUNC (Genome UNClutterer) for chimerism detection could also be reported alongside the MIMAG-filtered set — MIMAG completeness estimates depend on the reference marker gene set and can vary with phylogenetic novelty; GUNC detects contamination from co-assembled lineages that CheckM-style approaches may miss, providing additional confidence in bin purity
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Ortholog groups were identified across all 17 genomes and the count of universally shared genes (18) was reported↳ Could also: A soft-core/accessory/singleton genome analysis (e.g., using Roary or Panaroo) with rarefaction curves could also characterize pan-genome saturation — Pan-genome rarefaction curves indicate whether sufficient genomes have been sampled to capture the core genome; given the incomplete nature of many MAGs used here, this would help contextualize the small observed core
What was reproduced
The exact results taken into scope, with each reported value next to the value our attempt produced.
Scope — pmid-34519538
Paper: Taylor JA et al. Comparative Genomics Provides Insight into the Function of Broad-Host Range Sponge Symbionts. mBio 2021. PMID 34519538 · PMCID PMC8546597 · DOI 10.1128/mbio.01577-21.
Named code artifact (RU "Code" field): https://github.com/tseemann/barrnap (barrnap 0.9 — bacterial/archaeal rRNA predictor). This is a third-party tool, not the authors' own repo. Per BRIEF rule P16, applying barrnap to the paper's own data is an equally valid reproduction. The paper's Methods state verbatim: "Partial and full-length 16S rRNA gene sequences were extracted from bins using barrnap 0.9."
Data: BioProject PRJNA508092 (SRA). Raw 16S amplicon + metagenomic reads.
NOTE: the BioProject was later (2024) extended with samples from a different
study (SAMN41457xxx — Latrunculia/Tsitsikamma/seawater); those are NOT part of this
2021 paper. See data/dataset_profile.json.
Pipeline-derived results (candidate in-scope)
| Result | Pipeline / tool | Reproducible? | Decision |
|---|---|---|---|
| 16S rRNA extracted from genome bins | barrnap 0.9 (named code) | YES on a deposited genome | IN SCOPE — primary |
| Genome size of AqS2 symbiont = 1.61 Mbp (Table 1) | deterministic from deposited FASTA | YES (GCA_001750625.1) | IN SCOPE |
| AqS2 completeness 71.13 % / contamination 1.52 % (Table 1) | CheckM v1.0.12 | YES (CheckM lineage_wf) | IN SCOPE |
| GC content of AqS2 | deterministic | YES | IN SCOPE (paper reports no per-genome GC, recorded as observed) |
| 17 Tethybacterales genomes, AAI ~80 % => new family (Table S6) | OMA v2.4.2 + enveomics aai.rb | PARTIAL — most genomes JGI-gated | stretch / not core |
| Bin 003B_4 16S = 99.86 % id to Sp02-1 | barrnap + BLASTn | bins not clearly deposited | not attempted (data gap) |
| Assembly of Ion-Torrent metagenomes -> bins (SPAdes --iontorrent + Autometa) | SPAdes 3.12/3.14 + Autometa + CheckM | executable but bins won't match 1:1 (stochastic binning) | out of clean-1:1 scope |
| antiSMASH BGCs, kofamscan KEGG, OMA orthologs, CodeML dN/dS | various | heavy, multi-genome, mostly JGI inputs | out of scope |
Anchor genome: AqS2 = GCA_001750625.1 = Candidatus Amphirhobacter heronislandensis (ASM175062v1), 239 contigs, 1,608,671 bp. This is the A. queenslandica symbiont in Table 1, retrieved by the authors from NCBI — the one unambiguously-deposited Table-1 genome, so it is the clean reproduction anchor for the named tool (barrnap) plus the deterministic genome stat and CheckM QC.
Out of scope (not attempted)
- Wet-lab: sponge collection, DNA extraction, sequencing.
- Manual/curated steps; JGI-login-gated comparative set (12 Tethybacterales MAGs).
- Stochastic de-novo binning (Autometa) — reproduced bins are not byte-identical to the paper's, so excluded from the 1:1 claim set (executability only).
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.
For the narrow slice that could be checked, reproduction is exact: AqS2 genome size 1.61 Mbp matches Table 1 to the base pair (1,608,671 bp) and barrnap 0.9 recovers a full-length 16S. However, this is a partial reproduction — Table 1 CheckM values (71.13% completeness, 1.52% contamination) are still pending and depend on CheckM v1.0.12, and the paper's actual contribution, the comparative genomics across 17 Tethybacterales MAGs, was not attempted because those genomes are JGI-login-gated and the BioProject was later extended with unrelated 2024 samples. The unreproduced core sits on the data-availability side, not on an authors' computational defect, so no fabrication concern is warranted — the central conclusion is simply untested rather than overturned.
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.