Tasman-PCR: A genetic diagnostic assay for Tasmanian devil facial tumour diseases

Abstract

Tasmanian devils have spawned two transmissible cancer clones, known as devil facial tumour 1 (DFT1) and devil facial tumour 2 (DFT2). DFT1 and DFT2 are transmitted between animals by the transfer of allogeneic contagious cancer cells by biting, and both cause facial tumours. DFT1 and DFT2 tumours are grossly indistinguishable, but can be differentiated using histopathology, cytogenetics or genotyping of polymorphic markers. However, standard diagnostic methods require specialist skills and equipment and entail long processing times. Here, we describe Tasman-PCR: a simple PCR-based diagnostic assay that distinguishes DFT1 and DFT2 by amplification of DNA spanning tumour-specific interchromosomal translocations. We demonstrate the high sensitivity and specificity of this assay by testing DNA from 557 tumours and 818 normal devils. A temporal-spatial screen confirmed the reported geographic ranges of DFT1 and DFT2 and did not provide evidence of additional DFT clones. DFT2 affects disproportionately more males than females, and devils can be co-infected with DFT1 and DFT2.Overall, we present a PCR-based assay that delivers rapid, accurate and high-throughput diagnosis . CC-BY 4.0 International license It is made available under a (which was not peer-reviewed) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity.

The Origins and Vulnerabilities of Two Transmissible Cancers in Tasmanian Devils

“…DFT1 tumors (95T, 88T, 812T2, and 813T1) and DFT2 tumors (202T2, 203T3, 812T1, and 637T1) are labeled in blue and red, respectively, and DFT1 hosts (95H, 88H, 812H, and 813H) and DFT2 hosts (202H1, 203H, 812H, and 637H) are displayed in black. The upper panel shows SRY product and the lower panel shows positive control ( RPL13A ) and diagnostic amplification product for confirmation of DFT1 or DFT2 ( Kwon et al., 2018 ). Red arrows highlight presence (812T1) or absence (637T1) of an SRY band in DFT2 tumors infecting female Tasmanian devils.…”

“…DFT2 faces a further potential immunological challenge due to its possession of the Y chromosome. This lineage arose in a male devil and has, to date, usually been observed in males (of the 11 reported cases of DFT2, 9 involve a male host [ Kwon et al., 2018 ]). This apparent bias toward male hosts raises the possibility that females may be less susceptible to DFT2 due to immunogenicity of antigens derived from the Y chromosome.…”

“…As expected, Y chromosome DNA was not detected in DFT1, which is derived from a female founder devil, regardless of the gender of the host ( Figure 4 E). In DFT2, Y chromosome DNA was present in DFT2 tumors in male hosts, as well as in one DFT2 tumor in a female host, Devil 812 (Devil 812 also carried two DFT1 tumors [ Kwon et al., 2018 ]). However, the Y chromosome locus could not be detected in the DFT2 tumor derived from the second female host, Devil 637 ( Figure 4 E).…”

“…A multiplex DFT diagnostic PCR has been developed to confirm DFT1 or DFT2 diagnosis ( Kwon et al., 2018 ). Briefly, the PCR incorporates three primer sets, respectively targeting a DFT1-specific structural variant, a DFT2-specific structural variant, and the RPL13A locus, which acts as an internal positive control.…”

“…Indeed, karyotype evidence suggests that DFT2 arose from the somatic cells of a male animal, in contrast to the female origin of DFT1 ( Pye et al., 2016b ). To date, DFT2 has been confirmed in only 11 devils, all located on the Channel Peninsula in Tasmania's south-east ( Kwon et al., 2018 ).…”