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Oophila is monophyletic within a three-taxon eukaryotic microbiome in egg masses of the salamander Ambystoma maculatum

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Abstract

Tests of hypotheses of vertical transmission and competitive exclusion for particular resource-exchange mutualisms require assessments of the number and identity of partners. Unicellular green algae form a symbiosis with embryos of the northeastern yellow spotted salamander Ambystoma maculatum. The extent to which the composition of egg mass-associated taxa depends on temporal or spatial factors is unknown and the discovery of an endosymbiotic component to this symbiosis has increased the importance of knowing the specificity of the association. We assessed the diversity and relative abundance of algal and other unicellular eukaryotes in egg masses by subjecting DNA harvested from cells in egg capsules or water from breeding habitat to 18S amplicon sequencing. A nested sampling regime for two breeding seasons allowed comparisons among capsules within egg masses, among egg masses in ponds, between breeding locales, between pond water and capsule fluid, and between years. Based upon 4.33 X106 reads, we found no evidence inside fifty-eight egg capsules of algal lineages outside the Oophila clade and report that, whereas non-algal eukaryotic sequence diversity was low, cercozoan protists and chytridomycete fungi were sometimes abundant. There were weak or inconsistent egg mass, year or locale effects, but there was a very strong localization effect, indicating that capsules are rarefied spaces for eukaryotic aquatic taxa. This work provides a foundation for comparing microbial community structure within and among egg masses across the geographic range of the host.

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Acknowledgements

This work began during a sabbatical to CDB, which was gratefully held in John Archibald’s laboratory (Dalhousie University) and was supported by a Discovery Grant to CDB from the Natural Sciences and Engineering Research Council of Canada. EJ conducted DNA extractions and PCR, prepared some graphs and was supported by a Undergraduate Student Research Assistantship from the Natural Sciences and Engineering Research Council of Canada; LG first developed methods to extract IF from egg capsules and then did so for all samples; CDB collected egg masses, conducted DNA extractions and PCR, performed data analyses and wrote the manuscript. All authors reviewed and revised the manuscript. The authors declare no conflicts.

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  1. E. Jurga

    Present address: Dept. Molecular Genetics, University of Toronto, Toronto, Canada

Authors and Affiliations

  1. Department of Biology, St. Francis-Xavier University, 2320 Notre Dame Avenue, Antigonish, NS, B2G 2W5, Canada

    E. Jurga, L. Graham & C.D. Bishop

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Electronic supplementary material

Online Resource 1

18S Eukarya standard operating procedure, v1 (PDF 88 kb)

Online Resource 2

Sequencing statistics as a function of treatment of samples prior to sequencing (raw vs. PCR) for 2016 (a) and 2017 (b) (PDF 25 kb)

Online Resource 3

A table of F-statistics describing differences among groups (a) and a binning scheme to describe the relative magnitude of effect (b) (PDF 43 kb)

Online Resource 4

Efficacy of detection of non-Oophila algal cells. Known quantities of cells of C. vacuolatum were injected into, then harvested from, A. maculatum egg capsules. Harvested cells were then subjected to amplicon sequencing and recovery was expressed as the percentage of sequences detected, relative to the quantity of cells injected. (PDF 82 kb)

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Jurga, E., Graham, L. & Bishop, C. Oophila is monophyletic within a three-taxon eukaryotic microbiome in egg masses of the salamander Ambystoma maculatum. Symbiosis 81, 187–199 (2020). https://doi.org/10.1007/s13199-020-00693-w

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