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The availability of high-quality germline DNA is an important prerequisite for a variety of genetic analyses. We have shown previously that fingernail clippings provide an optimal source of autologous, constitutional DNA for PCR-based applications. However, most existing protocols for nucleic acid purification from nails do not provide sufficiently high yields of pure and intact DNA for more demanding downstream analyses such as next generation sequencing (NGS). We have extensively tested and systematically modified a number of different protocols for DNA purification from nail material to optimize the yield and quality. The integrity of DNA was determined by PCR amplification of short (<300 bp), mid-range (>400 bp), and long-range (>2 kb) sequences using different target genes. Among the methods tested, the Prepfiler Forensic DNA Extraction kit was identified as the most appropriate approach to isolation of high-quality DNA from nail clippings. A standardized input of 20 mg nail material (1 to 10 pieces of fingernail clippings) yielded a mean of 1 μg DNA (range, 0.5 to 2.3 μg). Subsequent PCR-analysis revealed efficient amplifiability of short and mid-range targets in 93% and 90%, and long-range fragments in 60% of the samples tested. The adequacy for next generation sequencing applications was demonstrated by successful high-resolution HLA-typing in ten transplant recipients. Hence, the protocol presented facilitates the exploitation of fingernail material even for demanding genomic analyses both in research and diagnostics., (Copyright © 2014 American Society for Investigative Pathology and the Association for Molecular Pathology. Published by Elsevier Inc. All rights reserved.)

Adenovirus (AdV) infection in the course of allogeneic stem cell transplantation (SCT) is associated with high transplant-related morbidity and mortality. Disseminated AdV disease is lethal in most instances. Early detection of AdV infection and identification of patients carrying a high risk of disseminated disease therefore remain a major challenge. In view of the large number of existing AdV types, we have established real-time polymerase chain reaction (PCR) assays permitting sensitive detection and quantification of all 51 currently known human AdV serotypes. In a series of 132 consecutive pediatric patients undergoing SCT, more than 5000 samples derived from peripheral blood (PB), stool, urine, and throat were screened for adenovirus infection by PCR during the posttransplantation period. Thirty-six patients (27%) tested positive by PCR, revealing AdV types of the subgenera A, B, C, D, and F. Except for enteritis in some patients with AdV positivity in stool, detection of the virus at sites other than PB was not associated with clinical signs of virus disease, and transplant-related mortality was not significantly different from AdV-negative patients. By contrast, 82% of patients who had detectable AdV in PB died from infectious complications (P <.001). Monitoring of PB specimens by real-time PCR permitted early diagnosis of invasive AdV infection in all instances. In patients who developed disseminated AdV disease, detection of the virus in PB preceded onset of clinical symptoms by a median of more than 3 weeks. The observation of AdV in peripheral blood may therefore serve as a basis for early initiation of preemptive antiviral treatment.