Relevance of next generation sequencing (NGS) data re-analysis in the diagnosis of monogenic diseases leading to organ failure.

BACKGROUND: In 2018, our center started a program to offer genetic diagnosis to patients with kidney and liver monogenic rare conditions, potentially eligible for organ transplantation. We exploited a clinical exome sequencing approach, followed by analyses of in silico gene panels tailored to clinical suspicions, obtaining detection rates in line with what reported in literature. However, a percentage of patients remains without a definitive genetic diagnosis. This work aims to evaluate the utility of NGS data re-analysis for those patients with an inconclusive or negative genetic test at the time of first analysis considering that (i) the advance of alignment and variant calling processes progressively improve the detection rate, limiting false positives and false negatives; (ii) gene panels are periodically updated and (iii) variant annotation may change over time. METHODS: 114 patients, recruited between 2018 and 2020, with an inconclusive or negative NGS report at the time of first analysis, were included in the study. Re-alignment and variant calling of previously generated sequencing raw data were performed using the GenomSys Variant Analyzer software. RESULTS: 21 previously not reported potentially causative variants were identified in 20 patients. In most cases (n = 19), causal variants were retrieved out of the re-classification from likely benign to variants of unknown significance (VUS). In one case, the variant was included because of inclusion in the analysis of a newly disease-associated gene, not present in the original gene panel, and in another one due to the improved data alignment process. Whenever possible, variants were validated with Sanger sequencing and family segregation studies. As of now, 16 out of 20 patients have been analyzed and variants confirmed in 8 patients. Specifically, in two pediatric patients, causative variants were de novo mutations while in the others, the variant was present also in other affected relatives. In the remaining patients, variants were present also in non-affected parents, raising questions on their re-classification. CONCLUSIONS: Overall, these data indicate that periodic and systematic re-analysis of negative or inconclusive NGS data reports can lead to new variant identification or reclassification in a small but significant proportion of cases, with benefits for patients' management.

Toward a Dynamic Threshold for Quality Score Distortion in Reference-Based Alignment.

The intrinsic high-entropy sequence metadata, known as quality scores, is largely the cause of the substantial size of sequence data files. Yet, there is no consensus on a viable reduction of the resolution of the quality score scale, arguably because of collateral side effects. In this article, we leverage on the penalty functions of HISAT2 aligner to rebin the quality score scale in such a way as to avoid any impact on sequence alignment, identifying alongside a distortion threshold for "safe" quality score representation. We tested our findings on whole-genome and RNA-seq data, and contrasted the results with three methods for lossy compression of the quality scores.

Comparison of high-throughput sequencing data compression tools.

High-throughput sequencing (HTS) data are commonly stored as raw sequencing reads in FASTQ format or as reads mapped to a reference, in SAM format, both with large memory footprints. Worldwide growth of HTS data has prompted the development of compression methods that aim to significantly reduce HTS data size. Here we report on a benchmarking study of available compression methods on a comprehensive set of HTS data using an automated framework.

An Evaluation Framework for Lossy Compression of Genome Sequencing Quality Values.

This paper provides the specification and an initial validation of an evaluation framework for the comparison of lossy compressors of genome sequencing quality values. The goal is to define reference data, test sets, tools and metrics that shall be used to evaluate the impact of lossy compression of quality values on human genome variant calling. The functionality of the framework is validated referring to two state-of-the-art genomic compressors. This work has been spurred by the current activity within the ISO/IEC SC29/WG11 technical committee (a.k.a. MPEG), which is investigating the possibility of starting a standardization activity for genomic information representation.

Clinical and radiologic evaluation of 70 immediately loaded single implants in the maxillary esthetic zone: preliminary results after 1 year of functional loading.

PURPOSE: A key challenge in implant treatment in the esthetic zone is to provide patients with crowns and peri-implant mucosa that are in harmony with the adjacent teeth to restore both function and esthetics. The aim of the present study was to show the preliminary results of an immediate loading protocol in the maxillary esthetic zone that is designed to preserve the soft tissue. MATERIALS AND METHODS: Patients were treated with immediately loaded single-tooth implants (Seven, MIS). Implants were inserted in fresh postextraction sites or in single edentulous sites with clinically and radiologically healed bone. After 8 weeks of loading, the provisional crowns and abutments were modified as needed to enhance the esthetics of the gingival margins. Definitive restorations were delivered 4 to 6 months postoperatively. RESULTS: Seventy patients were treated; 25 implants were placed in fresh extraction sites, and the remaining 45 implants were inserted in healed single edentulous sites. All implants were stable, and radiographs showed complete osseointegration after 1 year of functional loading. CONCLUSIONS: The present study shows promising results for immediate loading of single implants. All implant sites fulfilled success criteria in terms of function and esthetics, and particularly promising results were seen following additional adjustments of the provisional restorations to preserve interdental papillae.

A comparative study of bone densitometry during osseointegration: piezoelectric surgery versus rotary protocols.

OBJECTIVES: To date, there have been no studies on the outcome of osseointegration of alveolar bone around dental implants inserted with piezoelectric osteotomy versus conventional osteotomy. The aim of this study was to compare the radiographic differences, through evaluation of peri-implant bone density, between implant insertion using traditional surgical technique and piezoelectric technique. METHOD AND MATERIALS: Forty patients were selected whose treatment consisted of a minimum of two implants placed in nonpathologic native bone. A single type of implant surface (SLA) was chosen. The implants were placed following the manufacturer protocol for traditional surgical technique and piezoelectric technique. Radiographs were taken following surgery and 30, 60, and 90 days after surgery. The bone density was studied with the densitometry application. RESULTS AND CONCLUSION: All patients completed the study period with success. Despite a limited number of treated patients, the results of this pilot study demonstrated that (1) piezoelectric implant site preparation promotes better bone density and osteogenesis, and (2) the piezoelectric technique is predictable, with a 100% success rate in this study.