The goal of this study was to identify novel highly specific RNA biomarkers for the identification of vaginal secretions (VS).
The inability to definitively determine the tissue source of origin of forensically relevant biological fluids could result in a failure to provide crucial information necessary to the investigation and prosecution of the case; for example, in instances of sexual assault with a foreign object or digital penetration, the identification of vaginal secretions (VS) transferred to such objects or the perpetrators might be critical in establishing the circumstances of the assault. Conventional serological and immunological methods for body fluid identification can confirm the presence of human blood and semen; however, none of the routinely used biochemical tests can currently definitively identify the presence of human saliva or VS. It has been demonstrated that mRNA (or miRNA) profiling of body fluid stains can provide a degree of identification specificity of tissue and body fluids heretofore unattainable by conventional means. Early promising VS candidate RNA biomarkers, however, failed to exhibit the required degree of specificity or sensitivity and thus it is not currently possible for the forensic scientist to definitively identify VS using molecular genetics techniques. In the current study, whole transcriptome profiling (RNA-Seq) of vaginal swab samples from different donors resulted in the identification of some putative VS-specific mRNA candidates. After detailed evaluation of > 200 candidates from the tens of thousands of mRNA species found in VS, six promising candidates were identified. From these, two gene transcripts, namely CYP2B7P1 and MYOZ1, consistently demonstrated high specificity and sensitivity for VS when used in a qualitative capillary electrophoresis-based assay. Importantly, these two biomarkers differentiated between VS and other body fluids containing significant numbers of epithelia, particularly saliva and skin. Significantly, CYP2B7P1 is exceedingly specific with no detectable cross reactivity with other forensically relevant body fluids/tissues noted to date. The other four putatively specific biomarkers are expressed at higher levels in VS compared with saliva and will be more suitable for use with a quantitative (i.e. qRT-PCR) assay format. (publisher abstract modified)