Volume 25, Number 8—August 2019
Prolonged Zika Virus RNA Detection in Semen of Immunosuppressed Patient
|Sample no.||Days after symptom onset||RT-PCR Ct value†||Sequence coverage, %‡||Average read depth||Sequencing platform§||Mutations detected||Culture result|
|1||13||19||99.9 (min depth 2), 88.4 (min depth 40)||386.9||MinION||Reference||Frozen sample: unsuccessful|
|2||46||26||ND||NA||NA||Fresh sample: unsuccessful|
|4||194||No RNA detected¶||ND||NA||NA||ND|
|6||257||34||44 (min depth 2), 0 (min depth 40)||3.3||MinION||None||Frozen sample: unsuccessful|
|7||278||No RNA detected¶||ND||NA||NA||ND|
|8||326||26||76 (min depth 2), 6 (min depth 40)||13.2||MinION||None||Fresh sample: unsuccessful|
|10||515||24||98.1 (min depth 5)||33.3||MiSeq||K3272E, Syn2921||Fresh sample: unsuccessful|
*Ct, cycle threshold; min, minimum; NA, not applicable; ND, not done; RT-PCR, reverse transcription PCR.
†Before PCR, nucleic acid was extracted from samples 1–7 using the EZ1 Virus Mini Kit (QIAGEN, https://www.qiagen.com). Samples 8–12 were extracted using the MagNA Pure 96 DNA and Viral NA Small Volume Kit (LifeScience-Roche Diagnostics Corporation, https://lifescience.roche.com). Ct values <40 with acceptable amplification curves are interpreted as positive, but results for samples with Ct values >35 are confirmed by re-extraction and repeat PCR in triplicate, where possible.
‡Conservative read-depth thresholds were selected for comparative analyses of the day 13 (sample 1) and day 515 (sample 10) consensus genomes.
§MinION (Oxford Nanopore Technologies, https://nanoporetech.com); MiSeq (Illumina, https://www.illumina.com).
¶Confirmed on reextraction and repeat PCR testing.