Review Reports

Reviewer #1: We thank the reviewer 1 for providing an accurate and to the point summary of our research. We are pleased to see this good understanding and support to our work. We have carefully revised our manuscript as per reviewer 1 comments and suggestions.

Comment

Response

1

The experiment lacks a blank control, a sterile PBB control and an MPBB treatment without Cd contamination, making it impossible to isolate the synergistic effects of bacteria vs. the biochar carrier alone versus the combined stress.

Thanks a lot for good suggestions. In our study, the pot experiment was designed as follows: four treatments: (i) control (CK), consisting of 1 kg Cd-polluted soil with no amendment; (ii) 2% PBB; (iii) 2% MPBB; (iv) bacterial suspension (110 mL, 1.0×10⁸ CFU mL^-1). We acknowledge that it lacked no Cd-contaminated treatment. We have supplemented that it will be studied in the future at lines 355-356: Besides, the implication of MPBB in soil without Cd contamination would be studied in the future.

2

The pH decreases due to organic acids, this is not measured in the batch experiments, which is central to mechanistic claim.

Thanks a lot for good suggestions. We acknowledge that organic acid production was not directly measured. Accordingly, we have framed this interpretation as a literature‑supported hypothesis in the Discussion (Lines 223-226) for the link between pH decrease and phosphate solubilization.

3

The root cell wall extraction method (critical for the main mechanism) is only referenced, not described.

Thanks a lot for good suggestions. We have added the root cell wall extraction method at Lines 161-172.

4

PCA analysis is performed with only 4 data points (n=4), yielding p-values near 0.05, This lacks statistical power and the conclusions drawn from it are unreliable; this is a severe limitation.

Thanks a lot for good suggestions. We have removed the PCA analysis in pot experiment.

5

Enhanced root cell wall Cd retention is attributed to high IAA and Ca²⁺ release, this needs sufficient proof, which is missing.

Thanks a lot for good suggestions. We acknowledge that this attribution needs further validation and we have mentioned it at Line 349-351: "But more sufficient proofs that high IAA production by M and enhanced Ca²⁺ releasing from MPBB enhanced root cell wall Cd retention still need to be further explored in the future."

6

Errors like noting Pseudomonas as a phylum must have been avoided.

Thanks a lot for good suggestions. We have revised it.

7

English needs to be checked for syntax, grammar and completeness of some statements/ sentences. Lines 60; 70; 87; 259-260, are some to note.

Thanks a lot for good suggestions. We have revised it.

Reviewer #2: We are very thankful to the reviewer #2 for providing an accurate and to the point summary of our research. We are pleased to see this good understanding and support to our work

Comment

Response

1

The introduction is quite short and the literature framing is weak: refs [7,15,28] describe essentially identical approaches (PSB loaded on biochar for Cd/P remediation), yet the manuscript never clearly articulates what specifically differentiates pig-bonederived biochar from other animal-derived biochars already tested in the literature.

Thanks a lot for good suggestions. We have supplemented it at Lines: 66-69. 

2

Lacks quantitative context on the scale of the Cd contamination problem (e.g., extent of Cdcontaminated agricultural soils in China, relevant regulatory thresholds for Brassica napus).

Thanks a lot for good suggestions. We have added the following statement in the Introduction at Lines 40-41: "Nationally, Cd content in over 7% of agricultural soil exceed the soil quality standard for agricultural lands."

3

The two hypotheses (i, ii) are reasonably framed but should be tied more explicitly to measurable endpoints — notably, the absence of plant biomass data (see below) makes the "beneficial to crop growth" claim in the hypothesis untestable as written. Suggest adding a paragraph addressing the safety implications of using animal-bone-derived biochar (risk of co-contamination with heavy metals such as Pb or As present in bone), a topic never raised in the manuscript

Thanks a lot for good suggestions. We have added the risk of Pb/As co‑contamination from bone at Lines 62-66: “Although bone-derived biochar, especially that from heavy-contamined area, might pose a risk of co-contamination with heavy metals such as Pb or As present in bone, the safety of bone-derived biochar implication will be addressed to the greatest extent by strictly controlling the quality of bone feedback and high-temperature pyrolysis to reducing heavy metal bioavailability [12]”. We have add plant biomass data at Lines 322-327. 

4

 Pyrolysis conditions: "500 °C for 2h" is reported without heating ramp or inert gas flow rate — insufficient detail for reproducibility.

Thanks a lot for good suggestions. We have added the details of pyrolysis at Lines 105-107.

5

Strain identification: 16S sequencing is

mentioned, but GenBank accession numbers for F9 and F1-2-3 are not provided, nor is percent identity to the closest reference strain. Fig. 2 shows a phylogenetic tree, but the text stops at genus level — species-level identification with stated similarity percentage should be required

Thanks a lot for good suggestions. We have included the accession numbers, similarity percentages, and phylogenetic support at Lines 208-211.

6

Notable absence of plant growth data: the authors measure IAA, ACC deaminase, and siderophore production — all classic PGP traits — but never report Brassica napus biomass, shoot/root length, or dry weight. Since hypothesis (ii) explicitly claims benefit to crop growth, this is a critical missing dataset that should be requested.

Thanks a lot for good suggestions. We have added the biomass data at Line 322-327.

7

FTIR and SEM-EDS: it is unclear whether these analyses were replicated (technical/biological replicates) or performed on a single sample/single EDS spot. The EDS data (Cd Wt%: 0.15 vs 0.07 vs 0.04) are later described in Results using language implying significance without any statistical test — terminology should be corrected if no replication exists.

Thanks a lot for good suggestions. We have revised Line 216-219 to describe the observed trend more cautiously (without implying statistical significance), as the EDS data were determined for one sample for each treatment.

8

qPCR: amplification efficiency, standard curve details, and amplicon size are not reported.

Thanks a lot for good suggestions. We have added the qPCR parameters (including amplification efficiency, standard curve, and amplicon sizes) at Line 144-148.

9

Line 148: genus and species names must be italicized throughout the manuscript, including in the main text, figures, and tables. 

Thanks for your suggestion. We have carefully checked the entire manuscript and italicized all genus and species names throughout the text, figures, and tables.

10

The Results text is already heavily interpretive ("might be attributed to," "likely due to" — dozens of occurrences), language more appropriate for the Discussion. Results should remain descriptive.

Thanks a lot for good suggestions. We have separated the description of Results and Discussion.

11

 Unit inconsistency: IAA is reported in the abstract/text as "113.24–114.27 mg L⁻¹," while the Fig. 1C axis shows "µg mL⁻¹" — numerically equivalent, but the notation should be harmonized to avoid confusion.

Thanks a lot for your suggestion. We have revised it.

12

Minor numerical discrepancy between abstract/results (383.67–520.19) and conclusion (383.67–520.69). You should revise it.

Thanks a lot for your suggestion. We have revised it.

13

Alpha-diversity (Shannon, Chao1) and betadiversity (NMDS/PCoA) analyses of the rhizosphere community are entirely absent, and would substantially strengthen the communitylevel conclusions beyond simple phylum/genus relative abundance.

Thanks for your suggestion. We have added the Shannon diversity index (H′) analysis of the rhizosphere bacterial community in the Results and Discussion at Lines 261-264.

14

Translocation factor (TF, shoot/root Cd ratio) is never calculated, despite being a straightforward and informative complement to the existing data.

Thanks a lot for good suggestions. We have added the translocation factor (TF) data at Lines 331-333.

15

No discussion of the safety of the bone-derived biochar itself (potential residual heavy metals or pathogens from animal-derived material).No consideration of production cost/scalability of PBB relative to mineral phosphate fertilizers, despite the abstract framing MPBB as an "eco-friendly phosphate fertilizer."Limited comparison with field-scale (rather than pot) studies to contextualize the transferability of these findings.

Thanks a lot for good suggestions. We have added the description of the safety of the bone-derived biochar, economic analysis, and field-scale implication at Lines 352-356.

16

Please, improve the quality of the graphs showed in the figures 4-5-6. The labels are not clear (increase font size).

Thanks a lot for good suggestions. We have improved the quality of Figures 4, 5, and 6