ABSTRACT
Liquid biopsy based on cell-free DNA (cfDNA) enables non-invasive dynamic assessment of disease status in patients with cancer, both in the early and advanced settings. The analysis of DNA-methylation (DNAm) from cfDNA samples holds great promise due to the intrinsic characteristics of DNAm being more prevalent, pervasive, and cell- and tumor-type specific than genomics, for which established cfDNA assays already exist. Herein, we report on recent advances on experimental strategies for the analysis of DNAm in cfDNA samples. We describe the main steps of DNAm-based analysis workflows, including pre-analytics of cfDNA samples, DNA treatment, assays for DNAm evaluation, and methods for data analysis. We report on protocols, biomolecular techniques, and computational strategies enabling DNAm evaluation in the context of cfDNA analysis, along with practical considerations on input sample requirements and costs. We provide an overview on existing studies exploiting cell-free DNAm biomarkers for the detection and monitoring of cancer in early and advanced settings, for the evaluation of drug resistance, and for the identification of the cell-of-origin of tumors. Finally, we report on DNAm-based tests approved for clinical use and summarize their performance in the context of liquid biopsy.
Subject(s)
Cell-Free Nucleic Acids/genetics , DNA Methylation/genetics , Medical Oncology/methods , Animals , Computational Biology , Humans , Liquid BiopsyABSTRACT
Morpho-biological notes and histopathology, based on LM and SEM observations, of the fig cyst nematode Heteroderafici isolated from Ficuscarica roots, collected in home and public gardens of Apulia region, southern Italy, are described and illustrated. Seventy-five localities throughout the Apulia region were sampled and one-quarter of the sampled localities had fig roots infested with H.fici, with population densities ranging from 44 to 180 cysts/100 ml of soil. All attempts to detect H.fici on ornamental Ficus spp. as well as on imported bonsai in Italy were unsuccessful. Morphometric characters of the Italian population conform to those of the type and re-description populations reported for H.fici. Molecular analysis using ITS, D2-D3 expansion domains of the 28S rRNA, and the partial 18S rRNA sequences of H.fici newly obtained in this study matched well with the corresponding sequences of H.fici present in the GenBank database. Phylogenetic trees confirmed and supported the grouping of H.fici in the Humuli group. Heteroderafici completes its embryogenic development in 14-16 days at 25 °C. Post-invasion development and maturity in the roots of F.carica seedlings is completed in 64-68 days at 25-28 °C with juveniles and adults showing different parasitic habits, being endoparasitic and semi-endoparasitic respectively. The establishment of permanent feeding sites that consist of the formation of large syncytia causes anatomical modification of vascular elements and general disorder in the root stelar structures. Syncytia structures associated with mature females showed different degrees of vacuolisation, numbers of syncytial cells, and contained nuclei and nucleoli which were constantly hypertrophied.
ABSTRACT
Longidorus piceicola, a new geographical and host record from Romania, was described and illustrated on the basis of two populations originating from a coniferous and a deciduous forest. The main morphological characters of specimens from Romania correspond very well with the type material collected from the soil around Picea abies L. (Slovakia) except for the shorter body and tail. The D2-D3 fragment of 28S rDNA from both populations was amplified and sequenced, and the sequences were identical to L. piceicola sequence from Slovakia. The partial 18S-ITS1-5.8S-ITS2 rDNA regions from one of the populations were sequenced for the first time. The evolutionary relationships between L. piceicola and the closest species L. intermedius based on D2-D3 sequence divergence and single-nucleotide polymorphisms are discussed. Although having very low sequence dissimilarity (0.3-0.9 %) both species have distinct morphology and biology. Longidorus piceicola differs from L. intermedius in having a much longer odontostyle, body, distance anterior end - guide ring, a wider lip region, more ventromedian supplements (11 vs 5-7) in the male, and develops through four rather than three juvenile stages. Furthermore, L. piceicola occurs more frequently in association with conifers, while L. intermedius is found mainly in oak forests.
ABSTRACT
A description is provided of Longidorus cholevae sp. n., a bisexual species associated with wild cherry (Prunus avium L.) from the Rila Mountains, Bulgaria. The position of L. cholevae sp. n. among other species of the genus was elucidated by using morphological and molecular data. Phylogenetic analyses were performed of D2-D3 expansion domains of the 28S rRNA and the partial ITS1 containing regions by Neighbor-Joining, Maximum Likelihood and Bayesian Inference methods. The species is characterised by a female body length of 6.1-8.1 mm; long odontostyle (106-129 µm); lip region wide (21.5-24 µm) rounded and continuous with the body profile; amphidial pouches short and wide, funnel-shaped; a posteriorly situated guide ring (30-37 µm); normal arrangement of pharyngeal glands, and short bluntly rounded to hemispherical tail. Four juvenile stages indentified, first stage with elongate conoid tail. Males with 2-4 adanal pairs and a row of 11-13 single ventromedian supplements, spicules 96-120 µm long. Based both on morphological and molecular data the new species appearred to be the most similar witha group of species distributed in Europe sharing common charcters such as amphidial fovea, lip region and tail shapes, and having similar odontostyle and body length: L. poessneckensis, L. caespiticola, L. macrososma, L. helveticus, L. carniolensis and L. pius. An updated list of Longidorus species and a partial polytomous keys to the Longidorus species with long odontostyle (code A45) and short tail (code H1) are provided.
