Keystone Flap Type IV in Breast Reconstruction: A Case Report.

OBJECTIVE: The objective of this paper is to present and document a specific case of breast reconstruction using an adapted Type IV Keystone Flap technique, with a droplet-shaped design with a reduced flap ratio, and to identify the qualities of this method. CASE REPORT: A 41-year-old woman, with a history of myocardial infarction and low ejection fraction, underwent a lumpectomy, resulting in a lower medial quadrant deficit in her left breast. After she developed skin and tissue necrosis and infection, implementing the Type IV Keystone Flap effectively addressed the deficit, ensuring sufficient coverage. The flap extended dropwise beneath the deficit, progressing anteriorly towards the upper rectus abdominis, with a ratio of 2.5:1. The flap's novel droplet shape allowed for the utilization of fewer perforators, while ensuring adequate blood supply and tissue coverage, leading to improved perfusion and aesthetic outcome. CONCLUSION: The application of the adapted Type IV Keystone Flap highlights its capacity as a versatile and effective method for breast reconstruction post-lumpectomy. With the advantages of a short learning curve, easy execution, and acceptable risk profile, it offers a valuable alternative for patients who may not be suitable for more complex surgeries. Further research is recommended to confirm its broader applicability and to conduct a comparative analysis with other techniques.

Performance of Five Metagenomic Classifiers for Virus Pathogen Detection Using Respiratory Samples from a Clinical Cohort.

Viral metagenomics is increasingly applied in clinical diagnostic settings for detection of pathogenic viruses. While several benchmarking studies have been published on the use of metagenomic classifiers for abundance and diversity profiling of bacterial populations, studies on the comparative performance of the classifiers for virus pathogen detection are scarce. In this study, metagenomic data sets (n = 88) from a clinical cohort of patients with respiratory complaints were used for comparison of the performance of five taxonomic classifiers: Centrifuge, Clark, Kaiju, Kraken2, and Genome Detective. A total of 1144 positive and negative PCR results for a total of 13 respiratory viruses were used as gold standard. Sensitivity and specificity of these classifiers ranged from 83 to 100% and 90 to 99%, respectively, and was dependent on the classification level and data pre-processing. Exclusion of human reads generally resulted in increased specificity. Normalization of read counts for genome length resulted in a minor effect on overall performance, however it negatively affected the detection of targets with read counts around detection level. Correlation of sequence read counts with PCR Ct-values varied per classifier, data pre-processing (R2 range 15.1-63.4%), and per virus, with outliers up to 3 log10 reads magnitude beyond the predicted read count for viruses with high sequence diversity. In this benchmarking study, sensitivity and specificity were within the ranges of use for diagnostic practice when the cut-off for defining a positive result was considered per classifier.

Taxonomic classification and abundance estimation using 16S and WGS-A comparison using controlled reference samples.

The assessment of microbiome biodiversity is the most common application of metagenomics. While 16S sequencing remains standard procedure for taxonomic profiling of metagenomic data, a growing number of studies have clearly demonstrated biases associated with this method. By using Whole Genome Shotgun sequencing (WGS) metagenomics, most of the known restrictions associated with 16S data are alleviated. However, due to the computationally intensive data analyses and higher sequencing costs, WGS based metagenomics remains a less popular option. Selecting the experiment type that provides a comprehensive, yet manageable amount of information is a challenge encountered in many metagenomics studies. In this work, we created a series of artificial bacterial mixes, each with a different distribution of skin-associated microbial species. These mixes were used to estimate the resolution of two different metagenomic experiments - 16S and WGS - and to evaluate several different bioinformatics approaches for taxonomic read classification. In all test cases, WGS approaches provide much more accurate results, in terms of taxa prediction and abundance estimation, in comparison to those of 16S. Furthermore, we demonstrate that a 16S dataset, analysed using different state of the art techniques and reference databases, can produce widely different results. In light of the fact that most forensic metagenomic analysis are still performed using 16S data, our results are especially important.

Retrospective Validation of a Metagenomic Sequencing Protocol for Combined Detection of RNA and DNA Viruses Using Respiratory Samples from Pediatric Patients.

Viruses are the main cause of respiratory tract infections. Metagenomic next-generation sequencing (mNGS) enables unbiased detection of all potential pathogens. To apply mNGS in viral diagnostics, sensitive and simultaneous detection of RNA and DNA viruses is needed. Herein, were studied the performance of an in-house mNGS protocol for routine diagnostics of viral respiratory infections with potential for automated pan-pathogen detection. The sequencing protocol and bioinformatics analysis were designed and optimized, including exogenous internal controls. Subsequently, the protocol was retrospectively validated using 25 clinical respiratory samples. The developed protocol using Illumina NextSeq 500 sequencing showed high repeatability. Use of the National Center for Biotechnology Information's RefSeq database as opposed to the National Center for Biotechnology Information's nucleotide database led to enhanced specificity of classification of viral pathogens. A correlation was established between read counts and PCR cycle threshold value. Sensitivity of mNGS, compared with PCR, varied up to 83%, with specificity of 94%, dependent on the cutoff for defining positive mNGS results. Viral pathogens only detected by mNGS, not present in the routine diagnostic workflow, were influenza C, KI polyomavirus, cytomegalovirus, and enterovirus. Sensitivity and analytical specificity of this mNGS protocol were comparable to PCR and higher when considering off-PCR target viral pathogens. One single test detected all potential viral pathogens and simultaneously obtained detailed information on detected viruses.

The respiratory virome and exacerbations in patients with chronic obstructive pulmonary disease.

INTRODUCTION: Exacerbations are major contributors to morbidity and mortality in patients with chronic obstructive pulmonary disease (COPD), and respiratory bacterial and viral infections are an important trigger. However, using conventional diagnostic techniques, a causative agent is not always found. Metagenomic next-generation sequencing (mNGS) allows analysis of the complete virome, but has not yet been applied in COPD exacerbations. OBJECTIVES: To study the respiratory virome in nasopharyngeal samples during COPD exacerbations using mNGS. STUDY DESIGN: 88 nasopharyngeal swabs from 63 patients from the Bergen COPD Exacerbation Study (2006-2010) were analysed by mNGS and in-house qPCR for respiratory viruses. Both DNA and RNA were sequenced simultaneously using an Illumina library preparation protocol with in-house adaptations. RESULTS: By mNGS, 24/88 samples tested positive. Sensitivity and specificity, as compared with PCR, were 96% and 98% for diagnostic targets (23/24 and 1093/1120, respectively). Additional viral pathogens detected by mNGS were herpes simplex virus type 1 and coronavirus OC43. A positive correlation was found between Cq value and mNGS viral normalized species reads (log value) (p = 0.002). Patients with viral pathogens had lower percentages of bacteriophages (p<0.001). No correlation was found between viral reads and clinical markers. CONCLUSIONS: The mNGS protocol used was highly sensitive and specific for semi-quantitative detection of respiratory viruses. Excellent negative predictive value implicates the power of mNGS to exclude any pathogenic respiratory viral infectious cause in one test, with consequences for clinical decision making. Reduced abundance of bacteriophages in COPD patients with viral pathogens implicates skewing of the virome during infection, with potential consequences for the bacterial populations, during infection.

Cyanobacteria of Greece: an annotated checklist.

BACKGROUND: The checklist of Greek Cyanobacteria was created in the framework of the Greek Taxon Information System (GTIS), an initiative of the LifeWatchGreece Research Infrastructure (ESFRI) that has resumed efforts to compile a complete checklist of species reported from Greece. This list was created from exhaustive search of the scientific literature of the last 60 years. All records of taxa known to occur in Greece were taxonomically updated. NEW INFORMATION: The checklist of Greek Cyanobacteria comprises 543 species, classified in 130 genera, 41 families, and 8 orders. The orders Synechococcales and Oscillatoriales have the highest number of species (158 and 153 species, respectively), whereas these two orders along with Nostocales and Chroococcales cover 93% of the known Greek cyanobacteria species. It is worth mentioning that 18 species have been initially described from Greek habitats. The marine epilithic Ammatoidea aegea described from Saronikos Gulf is considered endemic to this area. Our bibliographic review shows that Greece hosts a high diversity of cyanobacteria, suggesting that the Mediterranean area is also a hot spot for microbes.