Deep sequencing of DNA from urine of kidney allograft recipients to estimate the donor-specific DNA fraction

Abstract

Renal transplantation is the method of choice for patients with end stage kidney failure. But transplanted allograft could be affected by viral and bacterial infections and immune rejections. The standard test for the diagnosis of acute pathologies in kidney transplants is the renal biopsy. However, noninvasive tests would be desirable. Various methods using different techniques have been developed by the transplantation community. But these methods expect improvements. We present here a cost-effective method based on estimating donor-specific DNA fraction in recipient urine based on sequencing of recipient urine DNA only. We hypothesized that in the no-pathology stage, the largest tissue types present in recipient urine are donor kidney cells and in case of rejection, a larger number of recipient immune cells would be observed. Extensive in-silico simulation was used to tune the sequencing parameters: number of variants and depth of coverage. Sequencing of DNA mixture from 2 healthy individuals showed the method high prediction accuracy (maximum error < 0.04). We then demonstrated the insignificant impact of familial relationship and ethnicity using an in-house and public database. Lastly, we performed recipient deep urine DNA sequencing in 32 samples representing two pathology groups: acute rejection (AR, 12 samples) and acute tubular injury (ATI, 11 samples) and 9 samples with no pathology. We found a significant association between the donor-specific DNA fraction in the two pathology groups compared to no pathology (P = 0.0064 for AR and P = 0.026 for ATI). We conclude that deep DNA sequencing of recipient urine offers a noninvasive means of diagnosing and prognosticating acute pathologies in the human kidney allograft.