Identification of HPV genotypes causing cervical precancer using tissue-based genotyping.

Identification of high-risk human papillomavirus genotypes causing cervical precancer is crucial for informing HPV vaccine development and efficacy studies, and for determining which types to include in next-generation genotyping assays. Co-occurrence of hrHPV infections is common and complicates carcinogenicity assessment; accurate attribution requires tissue-based genotyping of precancers. We included all women with cervical intraepithelial neoplasia Grade 2 or worse (CIN2+) from the Biopsy Study, an observational study of 690 women enrolled between 2009 and 2012 at the University of Oklahoma. Tissue-based genotyping, including whole tissue sections (WTS) and laser-capture microdissection (LCM), was performed on all precancers with multiple hrHPV infections detected in cytology, totaling over 1,800 HPV genotyping assays. Genotype attribution was compared to hierarchical and proportional hrHPV-type attribution models. Of 276 women with CIN2+, 122 (44.2%) had multiple hrHPV genotypes in cytology. Of 114 women with genotyping data, 94 had one or more hrHPV detected in tissue. Seventy-one women (75.5%) had a single causal hrHPV genotype, while 23 women had multiple hrHPV genotypes causing CIN2+. Ten women had multiple causal infections in a single biopsy, contrary to the previous notion that each lesion is caused by a single type only. While HPV16 was the predominant causal hrHPV genotype using all approaches, the hierarchical model overattributed HPV16, whereas other causal hrHPV genotypes, particularly HPV18 and HPV35, were underattributed. Understanding true causal genotypes is important for the evaluation of vaccine efficacy, to estimate the extent of unmasking, and for type-specific risk assessment in screening and management.

Primer site polymorphisms: potential implications for bone marrow engraftment monitoring.

Assessment of chimerism after allogeneic bone marrow transplant frequently relies on PCR amplification of donor- and recipient-specific polymorphic short tandem repeats (STR) with subsequent fluorescent detection of amplicons. In vitro amplification of STR loci is often achieved through commercially available, multiplex PCR kits. While originally developed for forensic purposes, these kits are increasingly being used in clinical laboratories for early detection of graft failure and relapse of disease. Despite the obvious benefits of sensitivity, accuracy, reproducibility, and ease-of-use of these commercial kits, laboratories must understand the technological shortcomings of such assays to avoid misinterpretation of patient results. The current case illustrates how primer site polymorphisms associated with specific STR alleles can potentially lead to erroneous interpretation of engraftment analysis using one of these multiplex STR kits.