🧬 Metagenomic Profiling Reveals Lung Multi-Kingdom Microbes as Forensic Markers in Aquatic Death Investigation
Introduction
Forensic investigation of bodies recovered from water remains one of the most complex challenges in medico-legal science. Determining whether a victim truly drowned, identifying the drowning site, and accurately estimating the postmortem submersion interval (PMSI) are critical but often difficult tasks. Traditional autopsy findings may be inconclusive, especially in advanced decomposition.
Recent advances in forensic microbiology suggest that the lung microbiome can provide valuable biological evidence in aquatic deaths. While previous studies mainly focused on bacteria, the broader multi-kingdom microbiome—including archaea, eukaryotes, and viruses—has not been fully explored.
Why Metagenomics?
Conventional amplicon sequencing limits microbial detection due to insufficient sequencing depth. In contrast, shotgun metagenomic sequencing enables comprehensive profiling of all microbial kingdoms present in tissue samples. This approach allows:
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High-resolution species-level identification
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Detection of non-bacterial microbes
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Functional insights into microbial communities
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Improved biomarker discovery
Study Overview
In this study, researchers analyzed lung tissues from murine carcasses submerged in water for up to 10 days using metagenomic sequencing. Potential biomarkers were then validated using quantitative PCR (qPCR) in both murine and human forensic samples.
Machine learning models based on the Random Forest (RF) algorithm were constructed to evaluate diagnostic performance.
Key Findings
1️⃣ Multi-Kingdom Lung Microbiome Composition
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Bacteria were the dominant microbial group.
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Eukaryotes were the second most abundant kingdom.
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Archaea and viruses were present but less prominent.
2️⃣ Biomarkers for Drowning Diagnosis
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17 bacterial species identified as potential biomarkers
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9 eukaryotic species identified as potential biomarkers
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Detection of Aeromonas hydrophila showed strong diagnostic significance
The presence of Aeromonas hydrophila in lung tissue provides compelling molecular evidence supporting drowning diagnosis.
3️⃣ Drowning Site Inference
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14 bacterial species were identified as markers capable of differentiating drowning environments
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The bacterial RF model achieved 100% accuracy in site inference
4️⃣ Estimation of Postmortem Submersion Interval (PMSI)
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17 bacterial species contributed to PMSI prediction
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Mean Absolute Error (MAE): 0.66 ± 0.097 days
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This demonstrates high temporal precision
5️⃣ Model Performance
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Bacterial model accuracy for drowning diagnosis: 89.29%
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Eukaryotic model accuracy: 87.5%
Forensic Significance
This study expands forensic microbiology beyond bacterial profiling by incorporating multi-kingdom microbial analysis. The findings demonstrate that lung microbial signatures can:
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Confirm drowning as the cause of death
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Differentiate drowning sites
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Provide accurate PMSI estimation
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Strengthen molecular forensic evidence
Such advancements contribute toward more objective, data-driven forensic investigations in aquatic environments.
Future Perspectives
Multi-kingdom metagenomic profiling holds great promise for:
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Standardization in forensic casework
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Integration with virtopsy and molecular autopsy techniques
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Development of rapid diagnostic kits for drowning cases
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AI-driven predictive forensic modeling
Further large-scale human validation studies will enhance its admissibility in court and practical forensic applications.
Conclusion
Metagenomic analysis of lung multi-kingdom microbes represents a transformative approach in aquatic forensic investigations. The identification of specific bacterial and eukaryotic biomarkers—particularly Aeromonas hydrophila—provides powerful molecular evidence for drowning diagnosis, drowning site inference, and PMSI estimation.
This research marks an important step toward precision forensic microbiology and improved investigative reliability in aquatic death cases.
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