Forensic evidence recovered from crime scenes often contains a mixture of human and bacterial DNA. Although short tandem repeat (STR) profiling of genomic DNA (gDNA) is widely used for human identification, its effectiveness can be limited in cases involving highly degraded DNA. In such cases, human mitochondrial DNA (mtDNA) and microbiome analysis may serve as alternative methods. In this study, we developed a multiplex quantification assay targeting the bacterial 16S rRNA V7 region and the human mitochondrial NADH-dehydrogenase subunit 5 (ND5) gene. Quantification was performed using TaqMan-based real-time PCR (Human-Bacteria qPCR; HBQ) and droplet digital PCR (Human-Bacteria ddPCR; HBD). Optimal primer and probe concentrations were at 7 μM for the HBQ assay, and 5 μM bacterial primer set, 7 μM human mtDNA primer set, and 700 nM probes for the HBD assay. Sensitivity testing showed that the HBQ assay detected all DNA samples-except G147A-down to 20 fg, while the HBD assay detected both bacterial and human DNA at 20 fg, demonstrating higher analytical sensitivity than the real-time PCR method. Moreover, mock forensic samples were analyzed to confirm the assay applicability, and PCR inhibitor tolerance tests using humic acid and tannic acid were conducted to further validate their performance. Furthermore, the HBQ and HBD assays may be used in quality control processes for samples potentially affected by bacterial DNA or human mtDNA contamination and could also be applied to other fields such as food safety, environmental science, and biological research involving microbial DNA and human mtDNA.

Sex crime investigations often rely on evidence involving minimal amounts of seminal material, making it necessary to use sensitive biomarkers to detect semen. Thanks to its high concentration, prostate-specific antigen (PSA) has been extensively utilized as a forensic marker, but there remains a lack of consensus regarding its diagnostic cut-off value. The technique proposed in this study applies a tiered diagnostic algorithm that combines a highly sensitive screening assay with a highly specific assay. The objective was to validate PSA quantification as a screening tool and establish an optimal cut-off value based on the receiver operating characteristic curve. A total of 460 forensic samples from sex crime investigations were analyzed for PSA quantification using electrochemiluminescence immunoassay (ECLIA). Optical microscopy was used as the reference standard to detect spermatozoa. Receiver operating characteristic curve analysis established a cut-off value of 0.085 ng/mL, with an area under the curve (AUC) of 0.848, a sensitivity of 82.8%, and a negative predictive value of 92.8%, showing diagnostic performance in line with international standards. The established cut-off value was lower than those previously documented and made it possible to increase the detection of potential semen in samples, doubling the number of positive identifications. In child victims, PSA detection is particularly relevant, given that endogenous secretion begins around the age of 9. Presence in children, even at minimal levels, may be indicative of adult male semen. These findings confirm the role of PSA as a sensitive and reliable screening test in forensic diagnostics.

Sex estimation methods from the pelvis have been well-studied in research settings to estimate accuracy, error, and bias. However, patterns in casework are minimally described. We uniquely examine forensic anthropology casework in the United States retrospectively for the Phenice and Klales etal.'s sex estimation methods. Our hypothesis is that casework patterns will reflect the greater literature derived from research settings that show Phenice's method is more accurate and has lower error and sex bias. We use the publicly available Forensic Anthropology Database for Assessing Methods Accuracy. A sample of 229 cases from the United States reported the outcomes of applying these methods. McNemar's tests evaluate whether estimated sex is consistent with documented sex, and a Fisher's exact test compared the performance of the two methods. We further calculated accuracy, error, and sex biases of the methods. The McNemar's and Fisher's exact tests were not statistically significant, which indicates that both methods estimated sex at a rate close to the documented sex and to each other. Phenice's method displayed an accuracy of 99.4%, an error of 0.6%, and a sex bias of -2.4%. Alternatively, the Klales etal.'s method performed slightly lower with a 97.5% accuracy, 2.5% error, and 3.5% sex bias. Forensic anthropology casework in the United States reflects broader patterns in accuracy, error, and bias in the research setting literature, where Phenice outperforms the Klales etal.'s method, despite the values from casework probably reflecting practitioners using information beyond the method reported to make a final sex estimate.

The demand for analyzing images from sources such as closed-circuit television cameras has increased significantly. Conventional analyses, including gait and soft biometrics, typically require the comparison of two video footage clips, as these methods are predicated on video-to-video comparisons. Moreover, numerous prerequisites often limit their applicability, particularly in the field of gait biometrics. To address these limitations, this paper introduces a simple yet effective image-to-person comparison method, leveraging image reproduction from a structure from motion (SfM)/photogrammetry-based three-dimensional (3D) computer graphics reference virtual avatar. This avatar is generated from a reference real person. It is demonstrated that the proposed method, by applying 3D joint manipulations to the reference virtual avatar, qualitatively reproduces a person captured in a target image with high fidelity. Furthermore, quantitative silhouette comparisons successfully confirm distributions for forensic image-to-person comparison. The proposed method holds promise as a body shape-based forensic image-to-person comparison tool in scenarios where a real person can be used as a reference.

Volatile organic compounds (VOCs) released during human decomposition are chemically diverse and can provide forensic evidence indicating the prior presence of a corpse. In July 2023, the Michigan City Police Department received a report from an individual claiming to have murdered his roommate and stored the body in a basement cellar for 57 days before dismemberment and disposal. Concrete core samples from the basement were analyzed using headspace solid-phase microextraction coupled with gas chromatography-mass spectrometry (HS-SPME GC-MS). Three concrete samples contained six VOCs that are known to originate from the decomposition process. This leads to the conclusion that the decomposing body of someone or something was present in the room for enough time for the decomposition VOCs to collect on, in, and under the floor. This case represents the first successful legal introduction of VOC analysis evidence from concrete substrates within the State of Indiana.

The rapid advancement of deepfake technology poses a significant threat to digital content authenticity and public trust. Deepfakes leverage artificial intelligence to generate realistic yet manipulated images and videos, often for deceptive purposes. This study introduced an enhanced version of the MesoNet convolutional neural network tailored for deepfake detection. The model incorporates two additional convolutional layers, resulting in substantial performance gains across various metrics. It achieved a precision of 96.60%, recall of 95.33%, F1-score of 95.96%, accuracy of 95.59%, and a Matthews Correlation Coefficient (MCC) of 91.11%, outperforming baseline models such as ResNet-50, VGG variants, and AlexNet. Additionally, a real-time detection system was developed using a React frontend and Flask backend, demonstrating the model's potential for practical deployment. This research contributed a robust and scalable approach to deepfake detection and lays the groundwork for real-world applications in digital forensics and content authenticity verification.

Facial identification examiners assess whether two facial images-such as an image of an unknown person from surveillance footage and a controlled image of a known individual-depict the same person or different people. To communicate their observations, they rely on predefined verbal articulation scales that sometimes have associated numeric equivalents. However, these terms have not been calibrated against the actual strength of the evidence except indirectly through proficiency tests and black box studies. The present research reanalyzes the findings of face comparisons from the most comprehensive facial identification black box study to date, as well as multiple facial examination proficiency tests, to generate a quantitative measure of the strength of the evidence for each comparison. We used an ordered probit model to summarize the distribution of responses of both individual examiners and examiner teams to produce a set of likelihood ratios for each group and test. The likelihood ratios can be lower than values implied by the evaluative statements, which do not seem to justify the strengths of evidence implied by current articulation scales used in facial comparisons. Our analyses suggest that examiners are using language that overstates the strength of the evidence by several orders of magnitude.

Juvenile firesetting remains underexamined in non-Western forensic populations. This study investigated the psychiatric, motivational, and familial characteristics of 55 adolescents (mean age = 15.05 years; 92.7% male) referred for court-ordered forensic psychiatric assessment in Turkey between 2019 and 2025. Structured coding captured motivational subtypes, family adversity, psychiatric diagnoses, co-occurring offending, and incident characteristics. Motivations included antisocial/criminal, anger- or revenge-driven, impulsive, curiosity-related, and distress-linked acts. Family adversity-particularly fragmented caregiving and neglect-was common, alongside high rates of conduct disorder, attention-deficit/hyperactivity disorder, and depression. Fires most frequently occurred at home or outdoors and were typically ignited with lighters. Recurrence occurred in 14.5% of cases and was more strongly associated with delinquent behavior patterns than with psychiatric diagnoses; co-occurring offending independently predicted recurrence (odds ratio = 7.78, p = 0.046). Findings highlight heterogeneous externalizing pathways shaped by cumulative family adversity and extend the international literature by providing forensic evidence from a non-Western context. Results may inform structured assessment and tailored intervention strategies within juvenile justice systems.

This case study reports the forensic identification of N-methylaniline (NMA) in an automotive gasoline sample seized from a gas station in Brazil, with a semi-quantitative estimate of ca. 1.9% (v/v), indicating a case of fuel adulteration. Although NMA has been investigated in experimental fuel formulations as a potential nonmetallic anti-knock additive capable of increasing octane rating, to the best available knowledge, no previous studies have reported its detection in real-world gasoline samples. In Brazil, the National Petroleum Agency (ANP) authorizes only previously approved additives, meaning that the presence of NMA constitutes a noncompliant fuel component. The sample was analyzed using gas chromatography-mass spectrometry (GC-MS), tandem mass spectrometry (MS/MS), and attenuated total reflectance Fourier-transform infrared spectroscopy (ATR-FTIR). The compound was unambiguously identified and characterized, demonstrating the effectiveness of these complementary analytical techniques for detecting unapproved or unconventional additives in complex hydrocarbon matrices. This case underscores the relevance of forensic fuel analysis for regulatory compliance and highlights the need for continuous monitoring to mitigate health, environmental, and operational risks associated with aromatic amine contamination.