Faecal metagenomes of great tits and blue tits provide insights into host, diet, pathogens and microbial biodiversity
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Background The vertebrate gut microbiome plays crucial roles in host health and disease. However, there is limited information on the microbiomes of wild birds, most of which is restricted to barcode sequences. We therefore explored the use of shotgun metagenomics on the faecal microbiomes of two wild bird species widely used as model organisms in ecological studies: the great tit (Parus major) and the Eurasian blue tit (Cyanistes caeruleus). Results Short-read sequencing of five faecal samples generated a metagenomic dataset, revealing substantial variation in composition between samples. Reference-based profiling with Kraken 2 identified key differences in the ratios of reads assigned to host, diet, and microbes. Some samples showed high abundance of potential pathogens, including siadenoviruses, coccidian parasites and the antimicrobial-resistant bacterial species Serratia fonticola. From metagenome assemblies, we obtained complete mitochondrial genomes from the host species and from Isospora spp., while metagenome-assembled genomes (MAGs) documented new prokaryotic species. Conclusions Here, we have shown the utility of shotgun metagenomics in uncovering microbial diversity beyond what is possible with 16S rRNA gene sequencing. These findings provide a foundation for future hypothesis testing and microbiome manipulation to improve fitness in wild bird populations. The study also highlights the potential role of wild birds in the dissemination of antimicrobial resistance. Impact statement The community of microorganisms that lives in an animal’s gut is crucial for the host’s health. However, in wild birds, such as the great tit and Eurasian blue tit, the role of this community is poorly understood. Here, we have applied advanced DNA sequencing and computational techniques to faecal samples from these birds, revealing a rich diversity of microbes, including previously unknown species and potential pathogens. We discovered antibiotic resistance genes, raising concerns about the potential spread of resistance from wild birds to humans. This research highlights the importance of understanding gut microbes in wild birds and lays the groundwork for future efforts to manipulate their gut microbial communities to enhance their health and resilience in changing environments. Data summary The authors confirm all supporting data, code and protocols have been provided within the article or through supplementary data files.
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Thank you for submitting your revised manuscript. I'm pleased to inform you it has now been accepted for publication. Thank you for choosing ACMI and please consider us for future publications. Best wishes, John.
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Dear Mark, Thank you for your submission. Following our previous correspondence, I have now secured reviews. I have selected minor revisions based the comments but once addressed we will be pleased to accept. Best wishes, John.
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Comments to Author
A very well-written manuscript. The project is very interesting, although it is unfortunate that the accuracy of the data is somewhat limited by current technology. Regardless, the data remains a valuable contribution to the field and a good basis for future investigations. There are a few grammatical errors that need rectified. Authors have commonly missed oxford commas. The initial section in methods utilises 'we', whereas latter sections utilises the passive voice. This should be made consistent. Line 307 'using DRAM, allowing to provide' does not make grammatical sense. Figure 2 contained text to small to read. Overall, very interesting and only needing minor tweaks.
Please rate the manuscript for methodological rigour
Good
Please rate the quality of the …
Comments to Author
A very well-written manuscript. The project is very interesting, although it is unfortunate that the accuracy of the data is somewhat limited by current technology. Regardless, the data remains a valuable contribution to the field and a good basis for future investigations. There are a few grammatical errors that need rectified. Authors have commonly missed oxford commas. The initial section in methods utilises 'we', whereas latter sections utilises the passive voice. This should be made consistent. Line 307 'using DRAM, allowing to provide' does not make grammatical sense. Figure 2 contained text to small to read. Overall, very interesting and only needing minor tweaks.
Please rate the manuscript for methodological rigour
Good
Please rate the quality of the presentation and structure of the manuscript
Very good
To what extent are the conclusions supported by the data?
Partially support
Do you have any concerns of possible image manipulation, plagiarism or any other unethical practices?
No
Is there a potential financial or other conflict of interest between yourself and the author(s)?
No
If this manuscript involves human and/or animal work, have the subjects been treated in an ethical manner and the authors complied with the appropriate guidelines?
Yes
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Comments to Author
The manuscript of Pallen et al describes a metagenomic study into the faecal metagenomes of Blue and Great Tits, which are both used widely in ecological studies of wild birds. The manuscript is well written and the authors are to be commended on the detail and organisation they provide in their supplementary data files (Zenodo). The use of metagenomics vs 16S amplicon sequencing is a large advance on current data and this allowed the authors to have greater insight from the data obtained. The work is sound, well written and presented and will provide a basis for future studies. Paragraph starting line 216 - Oak seems like an unlikely food plant for both species - given that Oak Tortrix Moth caterpillars are a particular favourite for breeding tits - may the abundance of oak reads reflect prey …
Comments to Author
The manuscript of Pallen et al describes a metagenomic study into the faecal metagenomes of Blue and Great Tits, which are both used widely in ecological studies of wild birds. The manuscript is well written and the authors are to be commended on the detail and organisation they provide in their supplementary data files (Zenodo). The use of metagenomics vs 16S amplicon sequencing is a large advance on current data and this allowed the authors to have greater insight from the data obtained. The work is sound, well written and presented and will provide a basis for future studies. Paragraph starting line 216 - Oak seems like an unlikely food plant for both species - given that Oak Tortrix Moth caterpillars are a particular favourite for breeding tits - may the abundance of oak reads reflect prey species gut contents? One minor points for the authors to address line 141 & 143 - Parus major and Cyanistes caeruleus should be in italics.
Please rate the manuscript for methodological rigour
Very good
Please rate the quality of the presentation and structure of the manuscript
Very good
To what extent are the conclusions supported by the data?
Strongly support
Do you have any concerns of possible image manipulation, plagiarism or any other unethical practices?
No
Is there a potential financial or other conflict of interest between yourself and the author(s)?
No
If this manuscript involves human and/or animal work, have the subjects been treated in an ethical manner and the authors complied with the appropriate guidelines?
Yes
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