Detection of Ganciclovir-Resistant Cytomegalovirus in a Prospective Cohort of Kidney Transplant Recipients Receiving Subtherapeutic Valganciclovir Prophylaxis.

Cytomegalovirus infection during antiviral prophylaxis occurs in transplant recipients despite individualized regimens based on renal function. Fifty kidney transplant recipients were assessed between 2016 and 2019 for valganciclovir dosing, ganciclovir exposure, cytomegalovirus infection, and genotypic resistance markers during the first year posttransplant. Ganciclovir plasma concentrations were measured using mass spectrometry. Population pharmacokinetics was used to determine individual ganciclovir exposure and to evaluate the ability of manufacturer dosing guidelines to meet therapeutic target daily area under the curve (AUC24) of 40 to 50 µg·h/mL. Full-length UL54 and UL97 were assessed using high-throughput sequencing in cytomegalovirus DNA-positive patient specimens. Valganciclovir doses administered to recipients with creatinine clearance of <40 mL/min were higher than specified by guidelines, and they were lower for recipients with creatinine clearance of ≥40 mL/min. The mean ganciclovir AUC24 was 33 ± 13 µg·h/mL, and 82% of subjects did not attain the therapeutic target. Pharmacokinetic simulations showed that the guidelines similarly could not attain the therapeutic target in 79% of individuals. Cytomegalovirus breakthrough occurred in 6% (3/50) of recipients, while 12% (6/50) developed late-onset infection. The mean AUC24s of recipients with (n = 3) and without (n = 47) infection were not significantly different (P = 0.528). However, one recipient with an AUC24 of 20 µg·h/mL acquired two UL97 ganciclovir resistance mutations. Current prophylaxis guidelines resulted in subtherapeutic ganciclovir exposure in several study recipients, including the emergence of resistance genotypes. IMPORTANCE This study examined the pharmacokinetics and viral genomic data from a prospective cohort of kidney transplant recipients undergoing valganciclovir prophylaxis for cytomegalovirus (CMV) prevention. We showed for the first time using high-throughput sequencing the detection of ganciclovir resistance mutations in breakthrough CMV infection during subtherapeutic plasma ganciclovir as indicated by the pharmacokinetic parameter daily area under the curve (AUC24). In addition, we found that current valganciclovir dosing guidelines for CMV prophylaxis are predicted to attain therapeutic targets in only 21% of recipients, which is consistent with previous pharmacokinetic studies. The novel findings of resistance mutations during subtherapeutic ganciclovir exposure presented here can inform future studies investigating the dynamics of drug selection pressure and the emergence of resistance mutations in vivo.

Assessment of Inter-Laboratory Differences in SARS-CoV-2 Consensus Genome Assemblies between Public Health Laboratories in Australia.

Whole-genome sequencing of viral isolates is critical for informing transmission patterns and for the ongoing evolution of pathogens, especially during a pandemic. However, when genomes have low variability in the early stages of a pandemic, the impact of technical and/or sequencing errors increases. We quantitatively assessed inter-laboratory differences in consensus genome assemblies of 72 matched SARS-CoV-2-positive specimens sequenced at different laboratories in Sydney, Australia. Raw sequence data were assembled using two different bioinformatics pipelines in parallel, and resulting consensus genomes were compared to detect laboratory-specific differences. Matched genome sequences were predominantly concordant, with a median pairwise identity of 99.997%. Identified differences were predominantly driven by ambiguous site content. Ignoring these produced differences in only 2.3% (5/216) of pairwise comparisons, each differing by a single nucleotide. Matched samples were assigned the same Pango lineage in 98.2% (212/216) of pairwise comparisons, and were mostly assigned to the same phylogenetic clade. However, epidemiological inference based only on single nucleotide variant distances may lead to significant differences in the number of defined clusters if variant allele frequency thresholds for consensus genome generation differ between laboratories. These results underscore the need for a unified, best-practices approach to bioinformatics between laboratories working on a common outbreak problem.