PONE-D-25-41106-->-->Fed the Same Way? Exploring the Influence of Breastfeeding, Weaning, and Childhood Diet on Adult Sex Ratios Through Stable Isotope Analysis of Incremental Dentine in Medieval Tuscany, Italy (11th – 15th c. CE)-->-->PLOS ONE?>

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Additional Editor Comments:

Dear Mr Amaro,

Please review the paper according to the reviewers' comment, and take this chance to double-check, for a last time, if all references have been accurately formatted according to the journal style. I look forward to read the reviewed manuscript.

Reviewers' comments:

Comments to the Author

1. Is the manuscript technically sound, and do the data support the conclusions?

Reviewer #1: Yes

Reviewer #2: Yes

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2. Has the statistical analysis been performed appropriately and rigorously? -->?>

Reviewer #1: Yes

Reviewer #2: Yes

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3. Have the authors made all data underlying the findings in their manuscript fully available??>

The PLOS Data policy

Reviewer #1: Yes

Reviewer #2: Yes

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4. Is the manuscript presented in an intelligible fashion and written in standard English??>

Reviewer #1: Yes

Reviewer #2: Yes

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Reviewer #1:  This is a really well written piece of research with solid questions focusing on infant feeding practices in medieval Tuscany and their possible effects on the sex ratio imbalance. The paper is well structured and arranged, with great historical background, meticulous referencing, as well as proper English.

However, there are some minimal aspects that I would advise to reconsider to improve the clarity of the paper. These are also reported in the pdf as comments in specific highlighted sections.

1. I feel that sometimes, during the train of thoughts, the authors tend to make logical leaps in the discourse, assuming that the reader is aware of the reasoning behind some particular statements. For instance, it is not clearly stated at the beginning for which actual reasons you decided to concentrate on the infant feeding strategies as the main cause of an imbalanced sex ratio in this specific population. Although it is indeed a possible explanation, it wasn't stated why you chose to give priority to this hypothesis rather than others such as nutritional imbalance in adolescence or adulthood, as well as labour or cultural choices. A similar dissonance occurs in the infanticide section, in which it is again unclear how the cited authors arrived to such extreme statements from solely census documents. I would advise to recheck these points, perhaps adding a couple more statements giving more clarity.

2. It is not clear from the practical point of view how you define the weaning process: how and where you see it starting and especially how you see it ending in the isotopic data. You give very specific ages for the end of the process (and not an age range), which give the impression of an abrupt moment while the actual process is quite gradual and often moves back and forth. It would be good to show these clearly in the graphs, perhaps using one as a model, and explain them very clearly in the text.

3. There can be several reasons for the raising of N values, also when considering incremental dentine sections. Although here it is probable that the rise was influenced by breastfeeding, I think that also diet, stress and pathology should be mentioned as possible concomitant factors. Are some of these individuals showing sings of childhood stress such as enamel hypoplasia on other teeth, or cribra orbitalia? It could be interesting to check this and see if the timing is influenced by this as well.

4. In the introduction, the authors correctly report the limitations of using a cross-sectional approach for studying infant feeding patterns. However, I think the paper misses an essential section reporting the limitations of incremental dentine sampling in general (which unfortunately are several), as well as the actual limitations of this specific study, namely the very few individuals of each sex that are funneled down when you pick the age categories per site. It is very briefly mentioned in the results, but I think it should also be clearly stated in a dedicated paragraph.

5. I'm not sure if this is a problem of the pdf for reviewers, but the images look really blurry. Please re-check them.

With this said, I think that with these few modifications, the paper is ready for publication.

Reviewer #2:  Peer review report on “Fed the Same Way? Exploring the Influence of Breastfeeding, Weaning, and Childhood Diet on Adult Sex Ratios Through Stable Isotope Analysis of Incremental Dentine in Medieval Tuscany, Italy (11th – 15th c. CE)”

1. Recommendation

Minor revision

2. Comments to Author:

Title: “Fed the Same Way? Exploring the Influence of Breastfeeding, Weaning, and

Childhood Diet on Adult Sex Ratios Through Stable Isotope Analysis of Incremental

Dentine in Medieval Tuscany, Italy (11th – 15th c. CE)”

Authors: Alessio Amaro, Antonio Fornaciari, Valentina Giuffra, Sang-Tae Kim, Martyn Knyf Paul Szpak, Bonnie Kahlon, Tracy L. Prowse

Overview and general recommendation:

Amaro et al. present a valuable study addressing two key issues: (1) the phenomenon of missing female individuals in the funerary context, and (2) differences in infant feeding and children dietary practices between male and female individuals from medieval populations in Aulla, Badia Pozzeveri, and Montescudaio, Italy. The dataset generated represents an important contribution to the field and is suitable for publication. The results are presented clearly and in organized manner; however, the manuscript would benefit from the inclusion of additional plots, such as plots of individual nitrogen and carbon isotope values by age, to improve data accessibility. The interpretation that females were breastfed for a longer duration than males across all three sites requires reconsideration, as the small sample sizes – particularly when subdivided by phase, with the exception of Montescudaio phase 2 – limit the strength of this conclusion. A more nuanced discussion of possible reasons for longer breastfeeding and weaning practices among females would strengthen the argument. As these revisions are relatively minor and can be implemented without restructuring, I recommend the manuscript for publication pending minor revisions.

Comments and suggestions:

Introduction

Page 2, lines 50-51: the transition into the paragraph of nitrogen stable isotope feels a little abrupt. The flow could be improved by adding a short introductory sentence to indicate that the following section will focus on the application of stable isotope analysis to infant feeding practices in archaeological populations.

Page 3, line 59: ‘has turned to the analysis of the sequential dentine to investigate breastfeeding and weaning patterns in those individuals who survived childhood (e.g., [23, 24]).’ I suggest being more specific here by emphasizing the use of sequential or incremental dentine analysis. While dentine as a window into childhood diet was already explored in the early 2000s through bulk dentine analysis, the latest studies apply a different approach, and it would be useful to highlight this methodological distinction.

Page 4, line 95: The phrasing ‘investigations into gender imbalances in contemporary populations’ should be reconsidered. Since sex and gender are distinct concepts, and the text later refers to ‘female survival’, it would be clearer and more accurate to frame this specifically in terms of sex rather than gender.

In Section 2.1. It would be helpful to clarify whether there is any evidence or discussion regarding female exogamy. Including this information could provide additional context for interpreting the sex-specific patterns observed in the populations.

Materials and Methods

Page 11, lines 276-286: It would be helpful to include the approximate number of individuals uncovered in the site description. Since other sites report these numbers, adding this information here would ensure consistency and enhance clarity regarding the material analyzed.

Page 11, line 293: There is an inconsistency in site names. Elsewhere in the text, as well as in plots and tables, the site is referred to as Aulla rather than San Caprasio.

Page 11, line 300: If you choose to use ‘innominate’, it would be clearer to use the more precise and formal term ‘innominate bone’. Alternatively, following current anatomical terminology, the preferred academic term is ‘os coxae’ or ‘pelvic bone’.

Results

Page 14, line 354: Could you please provide information on how many subsamples were obtained per tooth on average using the biopsy punch, as well as the maximum and the minimum number?

Page 15, lines 379¬-382: The statement here is confusing. Based on the bolded means provided in Table 2, there does not appear to be a statistically significant difference in carbon values between males and females in phase 2 within the 0-2.9 year age group or ‘males exhibiting more positive δ13C values’ – the reported means are exactly the same (-16.8). Could you please rephrase or clarify this section to make the intended point clearer?

Page 16, Table 2. Could you please comment on the nitrogen values in Badia Pozzeveri phase 2 between males and females within the 0-2.9 year age group? The difference is slightly more than 2 ‰. What are your thoughts on the potential significance or interpretation of this difference?

Page 17, lines 408-412: Would you consider other possible explanations for the higher average nitrogen values observed in females in the first increment, apart from weaning? Plots for individual carbon and nitrogen curves would be very helpful to include. They could allow readers to better assess whether any evidence of catabolic activity or potential “stress bubbles” is present in the data.

Discussion:

Page 20, lines 473-484: This paragraph would be more suitable for the Introduction section, where it could be framed as a general problem in reconstructing infant and childhood diet.

Page 21, lines 514-517: Is there any evidence of non-specific stress indicators, such as enamel hypoplasia, that could point to health restrictions during the first years of life? Such indicators might provide useful context for interpreting infant feeding practices, particularly if they were adapted to individual-specific needs.

Page 21, from 518 line: I would recommend including some additional references on dietary differences between male and female children. Although these works are not specifically focused on Medieval contexts, they could still be useful for discussing your results within a broader comparative framework.

• Henderson et al., 2014 https://doi.org/10.1002/ajpa.22554

• Henderson, 2015 (PhD thesis)

• Miller et al., 2020 https://doi.org/10.1002/ajpa.24033

Page 22, lines 543-545: Is there any adult or animal isotopic data available that could support or contextualize?

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Reviewer #1: No

Reviewer #2: No

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Fed the Same Way? Exploring the Influence of Breastfeeding, Weaning, and Childhood Diet on Adult Sex Ratios Through Stable Isotope Analysis of Incremental Dentine in Medieval Tuscany, Italy (11th – 15th c. CE)

PONE-D-25-41106R1

Dear Alessio,

We’re pleased to inform you that your manuscript has been judged scientifically suitable for publication and will be formally accepted for publication once it meets all outstanding technical requirements.

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Kind regards,

Dario Piombino-Mascali, Ph.D.

Academic Editor

PLOS ONE

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