Unlocking the story in the swab: A new genotyping assay for the amphibian chytrid fungus Batrachochytrium dendrobatidis

Allison Q. Byrne, Andrew P. Rothstein, Thomas J. Poorten, Jesse Erens, Matthew L. Settles, Erica Bree Rosenblum*

*Corresponding author for this work

Research output: Contribution to journalArticleAcademicpeer-review

6 Citations (Scopus)

Abstract

One of the most devastating emerging pathogens of wildlife is the chytrid fungus, Batrachochytrium dendrobatidis (Bd), which affects hundreds of amphibian species around the world. Genomic data from pure Bd cultures have advanced our understanding of Bd phylogenetics, genomic architecture and mechanisms of virulence. However, pure cultures are laborious to obtain and whole-genome sequencing is comparatively expensive, so relatively few isolates have been genetically characterized. Thus, we still know little about the genetic diversity of Bd in natural systems. The most common noninvasive method of sampling Bd from natural populations is to swab amphibian skin. Hundreds of thousands of swabs have been collected from amphibians around the world, but Bd DNA collected via swabs is often low in quality and/or quantity. In this study, we developed a custom Bd genotyping assay using the Fluidigm Access Array platform to amplify 192 carefully selected regions of the Bd genome. We obtained robust sequence data for pure Bd cultures and field-collected skin swabs. This new assay has the power to accurately discriminate among the major Bd clades, recovering the basic tree topology previously revealed using whole-genome data. Additionally, we established a critical value for initial Bd load for swab samples (150 Bd genomic equivalents) above which our assay performs well. By leveraging advances in microfluidic multiplex PCR technology and the globally distributed resource of amphibian swab samples, noninvasive skin swabs can now be used to address critical spatial and temporal questions about Bd and its effects on declining amphibian populations.

Original languageEnglish
Pages (from-to)1283-1292
Number of pages10
JournalMolecular Ecology Resources
Volume17
Issue number6
DOIs
Publication statusPublished - 1 Nov 2017

Fingerprint

Chytridiomycota
Amphibians
amphibian
genotyping
amphibians
Fungi
fungus
assay
fungi
skin (animal)
assays
skin
genomics
genome
virulence
sampling
topology
wildlife
Genome
pathogen

Keywords

  • Batrachochytrium dendrobatidis
  • genotype
  • microfluidic multiplex PCR
  • swab

Cite this

Byrne, A. Q., Rothstein, A. P., Poorten, T. J., Erens, J., Settles, M. L., & Rosenblum, E. B. (2017). Unlocking the story in the swab: A new genotyping assay for the amphibian chytrid fungus Batrachochytrium dendrobatidis. Molecular Ecology Resources, 17(6), 1283-1292. https://doi.org/10.1111/1755-0998.12675
Byrne, Allison Q. ; Rothstein, Andrew P. ; Poorten, Thomas J. ; Erens, Jesse ; Settles, Matthew L. ; Rosenblum, Erica Bree. / Unlocking the story in the swab: A new genotyping assay for the amphibian chytrid fungus Batrachochytrium dendrobatidis. In: Molecular Ecology Resources. 2017 ; Vol. 17, No. 6. pp. 1283-1292.
@article{24c6553e95774e288f21c82769553d53,
title = "Unlocking the story in the swab: A new genotyping assay for the amphibian chytrid fungus Batrachochytrium dendrobatidis",
abstract = "One of the most devastating emerging pathogens of wildlife is the chytrid fungus, Batrachochytrium dendrobatidis (Bd), which affects hundreds of amphibian species around the world. Genomic data from pure Bd cultures have advanced our understanding of Bd phylogenetics, genomic architecture and mechanisms of virulence. However, pure cultures are laborious to obtain and whole-genome sequencing is comparatively expensive, so relatively few isolates have been genetically characterized. Thus, we still know little about the genetic diversity of Bd in natural systems. The most common noninvasive method of sampling Bd from natural populations is to swab amphibian skin. Hundreds of thousands of swabs have been collected from amphibians around the world, but Bd DNA collected via swabs is often low in quality and/or quantity. In this study, we developed a custom Bd genotyping assay using the Fluidigm Access Array platform to amplify 192 carefully selected regions of the Bd genome. We obtained robust sequence data for pure Bd cultures and field-collected skin swabs. This new assay has the power to accurately discriminate among the major Bd clades, recovering the basic tree topology previously revealed using whole-genome data. Additionally, we established a critical value for initial Bd load for swab samples (150 Bd genomic equivalents) above which our assay performs well. By leveraging advances in microfluidic multiplex PCR technology and the globally distributed resource of amphibian swab samples, noninvasive skin swabs can now be used to address critical spatial and temporal questions about Bd and its effects on declining amphibian populations.",
keywords = "Batrachochytrium dendrobatidis, genotype, microfluidic multiplex PCR, swab",
author = "Byrne, {Allison Q.} and Rothstein, {Andrew P.} and Poorten, {Thomas J.} and Jesse Erens and Settles, {Matthew L.} and Rosenblum, {Erica Bree}",
year = "2017",
month = "11",
day = "1",
doi = "10.1111/1755-0998.12675",
language = "English",
volume = "17",
pages = "1283--1292",
journal = "Molecular Ecology Resources",
issn = "1755-098X",
publisher = "Wiley",
number = "6",

}

Unlocking the story in the swab: A new genotyping assay for the amphibian chytrid fungus Batrachochytrium dendrobatidis. / Byrne, Allison Q.; Rothstein, Andrew P.; Poorten, Thomas J.; Erens, Jesse; Settles, Matthew L.; Rosenblum, Erica Bree.

In: Molecular Ecology Resources, Vol. 17, No. 6, 01.11.2017, p. 1283-1292.

Research output: Contribution to journalArticleAcademicpeer-review

TY - JOUR

T1 - Unlocking the story in the swab: A new genotyping assay for the amphibian chytrid fungus Batrachochytrium dendrobatidis

AU - Byrne, Allison Q.

AU - Rothstein, Andrew P.

AU - Poorten, Thomas J.

AU - Erens, Jesse

AU - Settles, Matthew L.

AU - Rosenblum, Erica Bree

PY - 2017/11/1

Y1 - 2017/11/1

N2 - One of the most devastating emerging pathogens of wildlife is the chytrid fungus, Batrachochytrium dendrobatidis (Bd), which affects hundreds of amphibian species around the world. Genomic data from pure Bd cultures have advanced our understanding of Bd phylogenetics, genomic architecture and mechanisms of virulence. However, pure cultures are laborious to obtain and whole-genome sequencing is comparatively expensive, so relatively few isolates have been genetically characterized. Thus, we still know little about the genetic diversity of Bd in natural systems. The most common noninvasive method of sampling Bd from natural populations is to swab amphibian skin. Hundreds of thousands of swabs have been collected from amphibians around the world, but Bd DNA collected via swabs is often low in quality and/or quantity. In this study, we developed a custom Bd genotyping assay using the Fluidigm Access Array platform to amplify 192 carefully selected regions of the Bd genome. We obtained robust sequence data for pure Bd cultures and field-collected skin swabs. This new assay has the power to accurately discriminate among the major Bd clades, recovering the basic tree topology previously revealed using whole-genome data. Additionally, we established a critical value for initial Bd load for swab samples (150 Bd genomic equivalents) above which our assay performs well. By leveraging advances in microfluidic multiplex PCR technology and the globally distributed resource of amphibian swab samples, noninvasive skin swabs can now be used to address critical spatial and temporal questions about Bd and its effects on declining amphibian populations.

AB - One of the most devastating emerging pathogens of wildlife is the chytrid fungus, Batrachochytrium dendrobatidis (Bd), which affects hundreds of amphibian species around the world. Genomic data from pure Bd cultures have advanced our understanding of Bd phylogenetics, genomic architecture and mechanisms of virulence. However, pure cultures are laborious to obtain and whole-genome sequencing is comparatively expensive, so relatively few isolates have been genetically characterized. Thus, we still know little about the genetic diversity of Bd in natural systems. The most common noninvasive method of sampling Bd from natural populations is to swab amphibian skin. Hundreds of thousands of swabs have been collected from amphibians around the world, but Bd DNA collected via swabs is often low in quality and/or quantity. In this study, we developed a custom Bd genotyping assay using the Fluidigm Access Array platform to amplify 192 carefully selected regions of the Bd genome. We obtained robust sequence data for pure Bd cultures and field-collected skin swabs. This new assay has the power to accurately discriminate among the major Bd clades, recovering the basic tree topology previously revealed using whole-genome data. Additionally, we established a critical value for initial Bd load for swab samples (150 Bd genomic equivalents) above which our assay performs well. By leveraging advances in microfluidic multiplex PCR technology and the globally distributed resource of amphibian swab samples, noninvasive skin swabs can now be used to address critical spatial and temporal questions about Bd and its effects on declining amphibian populations.

KW - Batrachochytrium dendrobatidis

KW - genotype

KW - microfluidic multiplex PCR

KW - swab

U2 - 10.1111/1755-0998.12675

DO - 10.1111/1755-0998.12675

M3 - Article

VL - 17

SP - 1283

EP - 1292

JO - Molecular Ecology Resources

JF - Molecular Ecology Resources

SN - 1755-098X

IS - 6

ER -