Discriminating lineages of Batrachochytrium dendrobatidis using quantitative PCR

Abstract

The ability to detect and monitor infectious disease in a phylogenetically informative manner is critical for their management. Phylogenetically informative diagnostic tests enable patterns of pathogen introduction or changes in the distribution of genotypes to be measured, enabling research into the ecology of the pathogen. Batrachochytrium dendrobatidis (Bd), a causative agent of chytridiomycosis in amphibian populations, emerged worldwide in the 21st century and is composed of six lineages which are display varying levels of virulence in their hosts. Research into the distribution, ecology and pathogenicity of these lineages has been hampered by an inability to type lineage efficiently. Here, we describe a lineage-specific TaqMan qPCR assay that differentiates the two lineages of Bd most commonly associated with chytridiomycosis: BdGPL and BdCAPE. We demonstrate how this assay can be used for the surveillance of wild populations of amphibians in Southern Africa using skin swabs, tissue samples and cultured isolates.

Not just a pathogen: The importance of recognizing genetic variability to mitigate a wildlife pandemic

“…Recent advances in genetics have made novel tools for pathogen detection and characterization more accessible and reliable (Boyle et al, 2004;Byrne et al, 2019). In this issue of Molecular Ecology Resources, Ghosh et al (2021) report the development of a new genotyping qPCR assay targeting mitochondrial DNA (mtDNA) of Bd, and based on noninvasive swab samples (Figure 1), discriminate between the two most globally widespread and pathogenic genetic lineages of Bd. Having a better understanding of how the genetic diversity of a pathogen is distributed is crucial to understand their spread patterns and develop timely mitigation strategies.…”

“…Diagnostic assays targeting pathogen variants are highly desirable when seeking further insights into the molecular epidemiology, history of pathogen introduction and pathogen's ecology (Byrne et al, 2019). Ghosh et al (2021) AZAT that performed with high sensitivity, specificity and repeatability, to discriminate between the two most widespread and virulent Bd lineages: BdGPL and BdCAPE (Byrne et al, 2019;O'Hanlon et al, 2018; Figure 2). In addition, as with Boyle et al (2004), Ghosh et al (2021) mtDNA lineage specific qPCR is able to detect Bd at very low burdens (i.e., from one genomic equivalent in the sample).…”

“…Ghosh et al (2021) AZAT that performed with high sensitivity, specificity and repeatability, to discriminate between the two most widespread and virulent Bd lineages: BdGPL and BdCAPE (Byrne et al, 2019;O'Hanlon et al, 2018; Figure 2). In addition, as with Boyle et al (2004), Ghosh et al (2021) mtDNA lineage specific qPCR is able to detect Bd at very low burdens (i.e., from one genomic equivalent in the sample). With qPCR detection systems becoming more widely accessible and around hundreds of thousands DNA extractions from amphibian noninvasive swabs cryopreserved in laboratories around the globe, the application of Ghosh et al (2021) methodology, and its expansion to develop further primers and probes targeting mtDNA SNPs of other major Bd genotypes, will allow much more precise description of Bd lineages distributions globally and a greater understanding of the ecology of Bd.…”

“…In addition, as with Boyle et al (2004), Ghosh et al (2021) mtDNA lineage specific qPCR is able to detect Bd at very low burdens (i.e., from one genomic equivalent in the sample). With qPCR detection systems becoming more widely accessible and around hundreds of thousands DNA extractions from amphibian noninvasive swabs cryopreserved in laboratories around the globe, the application of Ghosh et al (2021) methodology, and its expansion to develop further primers and probes targeting mtDNA SNPs of other major Bd genotypes, will allow much more precise description of Bd lineages distributions globally and a greater understanding of the ecology of Bd. Also, with hybrids potentially being more virulent than their parental lineages (Greenspan et al, 2018;Schloegel et al, 2012), identifying the concurrent presence of two or more genotypes within a location with the potential of genetic recombination can help to implement conservation management actions on time, such as rescue and translocation of highly susceptible species.…”

“…However, a limitation of Ghosh et al (2021) qPCR assay is the detection of hybrids that can still be problematic as mtDNA is likely to be inherited from only one parent, cases in which the use of WGS or MLST might still be needed as a complementary methodology.…”