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. 2021 Apr 26;11(1):8918.
doi: 10.1038/s41598-021-87937-x.

Systematic STR analysis of old post-vasectomy seminal fluid stains to examine evidence stored for 16 years

Julianna Kesselring Romero  1 , Eloisa Auler Bittencourt  1   2 , José Arnaldo Soares-Vieira  3 , Ana Claudia Pacheco  4 , Alexandre Learth Soares  4 , Edna Sadayo Miazato Iwamura  5
Affiliations

Systematic STR analysis of old post-vasectomy seminal fluid stains to examine evidence stored for 16 years

Julianna Kesselring Romero et al. Sci Rep. .

Abstract

To understand stored evidence and the insertion in genetic databases is important in forensic investigations. Blood, pre- and post-vasectomy semen from 90 fertile male individuals, aged 24 to 45, were donated for research after informed consent. The semen samples were stored in the form of 30 µL stains on cotton fabric, for 16 years at room temperature in the laboratory. As well as the seminal fluid post vasectomy stains, which were performed after microscopy analyzes and certainty of the absence of spermatozoon. The pre vasectomy stains contained mainly haploid spermatozoon and the post vasectomy stains diploid epithelial cells and leukocytes. DNA extraction was performed with magnetic resin, followed by quantification and analysis of degradation of DNA. In this study we analyze these genetic profiles of DNA from stains on cotton fabric, using two Short Tandem Repeat multiplex systems, the PowerPlex Fusion 6C and Y23. Electrophoresis was performed on a 3500xL and analyzed using the Gene Mapper ID-X software. The genetic profiles of the 90 individuals were fully amplified in pre-vasectomy and partially in post-vasectomy stain samples, using the both multiplex systems. The results provide information about 0.25 cm2 semen stains on cotton fabric from 90 individuals, correlating concentration, degradation, and allele analysis. It also provides an understanding of the cells present in semen stains and the implications of individual factors. In the stains of post-vasectomy samples the small quantity of DNA was one of the limiting factors, in addition to degradation. Considering that all evaluations were carried out in a laboratory that has a quality control certificate and audited for being part of the national genetic profile database, the results were very consistent. Many aspects of the semen samples stored in the form of stains on cotton fabric have been clarified. The performance and sensitivity of the amplification systems used in the genotyping of azoospermic individuals were assessed. Conclusions: Genetic profiles were satisfactorily amplified in pre-vasectomy stain samples, and partially amplified in post-vasectomy stain samples, stored for almost two decades at room temperature in a tropical country. The small amount of DNA was one of the limitations in post-vasectomy stain samples, in addition to degradation and fragmentation. There are no publications in the literature on PowerPlex Fusion 6C and Y23 analyses using blood, sperm, and seminal fluids of the same individual, much less in the form of stains. This study can serve as a benchmark for the tracking analyses of stored samples. In addition, it anticipates a few social issues related to the analysis of post-vasectomy samples in forensic cases, most notably sex crimes.

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Conflict of interest statement

The authors declare no competing interests.

Figures

Graph 1
Graph 1
Absolute frequency per DNA degradation index ranges, from 0–1, 1–2, 2–4 and > 4, of blood, pré- and post-vasectomy seminal fluid stains samples (n = 90 individuals).
Graph 2
Graph 2
Boxplots of the degradation index range and genetic profile of PowerPlex Y23 System of the pre- vasectomy samples (n = 90 individuals).
Graph 3
Graph 3
Boxplots of the degradation index range and genetic profile of the PowerPlex Y23 System of the post- vasectomy samples (n = 90 individuals).
Graph 4
Graph 4
Boxplots of the degradation index range and genetic profile of PowerPlex Fusion 6C System of pre- vasectomy samples (n = 90 individuals).
Graph 5
Graph 5
Boxplots of the degradation index range and genetic profile of PowerPlex Fusion 6C System of post- vasectomy samples (n = 90 individuals).
Graph 6
Graph 6
Relationship between semen DNA concentrations (μg/ml) pre-and post-vasectomy (n = 90 individuals).
Graph 7
Graph 7
Relationship between the logarithms of DNA concentrations (μg/ml) pre- and post-vasectomy.
See this image and copyright information in PMC

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