Informativeness of NGS Analysis for Vaginal Fluid Identification

Abstract

The identification of vaginal fluids in forensic examinations plays an important role in crime scene reconstruction. Molecular detection of vaginal bacterial communities can lead to the correct discrimination of body fluids. These kinds of studies can be performed through multiplex real-time PCR using primers for a specific selection of bacteria. The availability of next-generation sequencing (NGS) protocols provided for the extension of the analysis to evaluate the prokaryotes present in specimens. In this study, DNA was extracted from 18 samples (vaginal, oral, fecal, yoghurt) and analyzed by real-time PCR and NGS. The comparison between the two approaches has demonstrated that the information developed through NGS can augment the more conventional real-time PCR detection of a few key bacterial species to provide a more probative result and the correct identification of vaginal fluid from samples that are more forensically challenged.

Changes of vaginal microbiota during cervical carcinogenesis in women with human papillomavirus infection

“…NGS method directly amplifies and sequences a particular region of 16S rRNA gene using DNA extracted from the clinical specimen without the need for cell culture [ 7 ]. NGS based methods have allowed massive parallelization of bacterial DNA sequencing and facilitated the rapid identification of thousands of vaginal microbiota taxa within a few days [ 8 ].…”

Potential testing of reprocessing procedures by real-time polymerase chain reaction: A multicenter study of colonoscopy devices

“…The assays allowed identification of positivity thresholds (CT < 33) and the acquisition of a microbial signature typical of that biological fluid, providing rapid and specific information on the contamination source and its traceability. 28 Indeed, this strategy was effective in covering the naturally occurring microbial variability, as confirmed by the NGS data showing a clear presence of these and other species belonging to the intestinal microbiota.…”

“…The PCR amplicon was obtained using primers (containing overhang adapters) as previously described. 27,28 Libraries were quantified through PicoGreen dsDNA Quantitation Assay (ThermoFisher Scientific, Waltham, MA) and validated on Bioanalyzer DNA 1000 chip (Agilent, Santa Clara, CA) for the presence of the expected amplicon. Sequencing was performed on a MiSeq desktop sequencer (Illumina).…”

Microbial forensics: new breakthroughs and future prospects

“…It has been proposed that each body fluid type presents a specific composition of microorganisms that can be used as bioindicators, being possible to infer the type of body fluid present based on its microbial composition (Hanssen et al 2017). For instance, saliva could be identified by detection of specific bacteria, such as Veillonella atypica, Streptococcus mutans and Streptococcus salivarius; while vaginal fluid could be identified by detection of specific bacteria, such as Lactobacillus crispatus and Lactobacillus gasseri (Choi et al 2014;Giampaoli et al 2017).…”