Evaluation of the rapid analyte measurement platform (RAMP) for the detection of Bacillus anthracis at a crime scene

The aim of this study was to evaluate the accuracy and reliability of the rapid analyte measurement platform (RAMP) for presumptive identification of Bacillus anthracis spores. Test samples consisted of serial dilutions of spore preparations of several Bacillus species, including B. anthracis, which were tested, using the RAMP Anthrax test cartridge, according to the manufacturer's instructions. The fluorescence labelled antibody-antigen complexes were detected in the portable reader after 15 min following sample addition. Dilutions of common environmental and household powders were also tested to identify possible false positive results. B. anthracis spores were identified reliably in test samples containing more than 6000 spores. The test kits were highly specific, showing no cross reactivity with other Bacillus species or any environmental powders tested. The RAMP system for detection of B. anthracis spores, from environmental samples, showed consistent results under a variety of analytical conditions, enabling the trained user to provide a rapid, accurate preliminary risk assessment of a suspected bioterrorism incident.     

Effect of Sterilants on Amplification and Detection of Target DNA from Bacillus cereus Spores

To conceal criminal activity of a bioterrorist or agroterrorist, the site of pathogen generation is often treated with sterilants to kill the organisms and remove evidence. As dead organisms cannot be analyzed by culture, this study examined whether DNA from sterilant‐treated Bacillus cereus spores was viable for amplification. The spores were exposed to five common sterilants: bleach, Sterilox®, oxidizer foam (L‐Gel), a peroxyacid (Actril®), and formaldehyde vapor. The spores were inoculated on typical surfaces found in offices and laboratories to test for environmental effects. It was found that the surface influenced the efficiency of recovery of the organisms. The DNA isolated from the recovered spores was successfully detected using RT‐qPCR for all treatments except for formaldehyde, by amplifying the phosphatidylinositol phospholipase C and sphingomyelinase genes. The results demonstrated that evidence from sites treated with sterilants can still provide information on the uncultured organism, using DNA amplification.     

Gamma Irradiation as a Biological Decontaminant and Its Effect on Common Fingermark Detection Techniques and DNA Profiling

Abstract:  The use of disease-causing organisms and their toxins against the civilian population has defined bioterrorism and opened forensic science up to the challenges of processing contaminated evidence. This study sought to determine the use of gamma irradiation as an effective biological decontaminant and its effect on the recovery of latent fingermarks from both porous and nonporous items. Test items were contaminated with viable spores marked with latent prints and then decontaminated using a cobalt 60 gamma irradiator. Fingermark detection was the focus with standard methods including 1,2-indanedione, ninhydrin, diazafluoren-9-one, and physical developer used during this study. DNA recovery using 20% Chelex extraction and quantitative real-time polymerase chain reaction was also explored. Gamma irradiation proved effective as a bacterial decontaminant with D -values ranging from 458 to 500 Gy for nonporous items and 797–808 Gy for porous ones. The results demonstrated the successful recovery of latent marks and DNA establishing gamma irradiation as a viable decontamination option.     

Forensic Identification of Blood in the Presence of Contaminations Using Raman Microspectroscopy Coupled with Advanced Statistics: Effect of Sand,Dust, and Soil

Body fluid traces recovered at crime scenes are among the most common and important types of forensic evidence. However, the ability to characterize a biological stain at a crime scene nondestructively has not yet been demonstrated. Here, we expand the Raman spectroscopic approach for the identification of dry traces of pure body fluids to address the problem of heterogeneous contamination, which can impair the performance of conventional methods. The concept of multidimensional Raman signatures was utilized for the identification of blood in dry traces contaminated with sand, dust, and soil. Multiple Raman spectra were acquired from the samples via automatic scanning, and the contribution of blood was evaluated through the fitting quality using spectroscopic signature components. The spatial mapping technique allowed for detection of “hot spots” dominated by blood contribution. The proposed method has great potential for blood identification in highly contaminated samples.     

犯罪现场生物检材的发现、提取策略

在改进现场生物检材的发现和提取技术的同时,也要进行理论创新和体系构建,才能够更好地指导实践。全局的观点、具体案件具体分析和犯罪现场重建对于现场勘查中有效地发现、提取和利用生物检材是十分重要的。     

Forensic palynology: why do it and how it works

Forensic palynology has been a law enforcement tool for over 50 years. Forensic palynology is the application of pollen and spores in solving legal issues, either civil or criminal. Pollen and spores can be obtained from an extremely wide range of items, including bodies. Pollen and spores provide clues as to the source of the items and the characteristics of the environments from which the material on them is sourced. Their usefulness lies in a combination of their abundance, dispersal mechanisms, resistance to mechanical and chemical destruction, microscopic size, and morphology. Their often complex morphology allows identification to an individual parent plant taxon that can be related to a specific ecological habitat or a specific scene. Pollen and spore assemblages characterise different environments and scenes and can easily be picked up and transported away from scenes of interest without providing any visual clue to a suspect as to what has occurred. With so many publications and high-profile cases involving forensic palynology and environmental analysis now receiving publicity, the future of this branch of forensic science is assured. Furthermore, with the development of multi-disciplinary approaches to environmental analyses of crime scenes, far more detailed information is now available to law enforcement agencies, enabling them to determine with greater accuracy what may have happened during the commission of criminal activities.     

Bioterrorism: processing contaminated evidence, the effects of formaldehyde gas on the recovery of latent fingermarks

In the present age of heightened emphasis on counter terrorism, law enforcement and forensic science are constantly evolving and adapting to the motivations and capabilities of terrorist groups and individuals. The use of biological agents on a population, such as anthrax spores, presents unique challenges to the forensic investigator, and the processing of contaminated evidence. In this research, a number of porous and nonporous items were contaminated with viable anthrax spores and marked with latent fingermarks. The test samples were then subjected to a standard formulation of formaldehyde gas. Latent fingermarks were then recovered post decontamination using a range of methods. Standard fumigation, while effective at destroying viable spores, contributed to the degradation of amino acids leading to loss of ridge detail. A new protocol for formaldehyde gas decontamination was developed which allows for the destruction of viable spores and the successful recovery of latent marks, all within a rapid response time of less than 1 h.     

Public health law in the new century

Public health law is a broad and sometimes nebulous field which has undergone extensive reform and rethinking over the past decade. This article provides a survey of current issues in public health law, highlighting these reforms and the potential for public health legislation to deal with upcoming threats, notably bioterrorism. While recognising the anxieties bioterrorism brings, public health responses must be grounded in a coherent philosophy of risk management. Its administrators must also be aware that large-scale threats to public health are not unprecedented and that past experience can provide an important guide for future strategies.     

Trace detection of meglumine and diatrizoate from Bacillus spore samples using liquid chromatography/mass spectrometry

Following the September 11, 2001 terrorist attacks, letters containing Bacillus anthracis were distributed through the United States postal system killing five people. A complex forensic investigation commenced to identify the perpetrator of these mailings. A novel liquid chromatography/mass spectrometry protocol for the qualitative detection of trace levels of meglumine and diatrizoate in dried spore preparations of B. anthracis was developed. Meglumine and diatrizoate are components of radiographic imaging products that have been used to purify bacterial spores. Two separate chromatographic assays using multiple mass spectrometric analyses were developed for the detection of meglumine and diatrizoate. The assays achieved limits of detection for meglumine and diatrizoate of 1.00 and 10.0 ng/mL, respectively. Bacillus cereus T strain spores were effectively used as a surrogate for B. anthracis spores during method development and validation. This protocol was successfully applied to limited evidentiary B. anthracis spore material, providing probative information to the investigators.     

Remote spectroscopic identification of bloodstains

Blood detection and identification at crime scenes are crucial for harvesting forensic evidence. Unfortunately, most tests for the identification of blood are destructive and time consuming. We present a fast and nondestructive identification test for blood, using noncontact reflectance spectroscopy. We fitted reflectance spectra of 40 bloodstains and 35 nonbloodstains deposited on white cotton with spectroscopic features of the main compounds of blood. Each bloodstain was measured 30 times to account for aging effects. The outcome of the blood measurements was compared with the reflectance of blood-mimicking stains and various body fluids. We found that discrimination between blood and nonblood deposited on white cotton is possible with a specificity of 100% and a sensitivity of 98%. In conclusion, a goodness of fit between the sample's reflectance and the blood component fit may allow identification of blood at crime scenes by remote spectroscopy.     

The use of simulated crime scenes in teaching undergraduate forensic sciences: Implementing an active learning approach to forensics

Incorporating a simulated crime scene into one’s pedagogy in forensic science undergraduate courses allows students the opportunity to experience a realistic scenario while demonstrating their knowledge and enhancing their critical thinking skills. The purpose of this paper is to examine an active learning approach to using simulated crime scenes to provide an impactful learning experience for students. While potentially challenging for the instructor, constructing a crime scene scenario can provide students with hands-on practical experience while helping to dispel forensic science misconceptions. Through many years of creating crime scenes, best practices for using such high impact activities in forensic science courses are described in detail, including preparatory exercises that culminate in the final crime scene scenario, considerations in preparing and constructing a crime scene activity, and supervising and assessing students once the activity begins. Three major challenges to implementing active learning exercises such as simulated crime scenes in undergraduate forensic science programs include high workload for instructors, limited supplies, and lack of suitable facilities. Workload solutions include instructors and departments considering the cost and benefit of course releases to improve curriculum and student enrollment. Supplies and facilities solutions involve innovation in reuse and repurposing of supplies, and instructor flexibility in using classroom and outdoor spaces.     

Measuring the impact of forensic science in detecting burglary and autocrime offences

As UK investment in forensic science has increased, the government has taken a fresh interest in how far this has led to dividends in terms of the detection of crime and its reduction. The Home Office funded 'Pathfinder Project' sought to monitor and document the complex relationships between the collection and use of forensic material (looking at a range of forensic science techniques) and its impact on crime detection. The project specifically targeted the 'volume' crimes of burglary and vehicle crime. Detailed data was gathered on all stages of the process between the collection and use of forensic material and crime detection. The model falls into two conceptual phases--scene attendance to suspect identification and identification to detection. From the analysis it was found that approximately one third of burglary and autocrime scenes are visited by SOCOs. While scientific identifications are only made in a minority of burglary and autocrime offences overall, it belies their importance. About one in ten of burglary and autocrime cases are cleared up by the police and it is estimated that fingerprints and SGMPlus were a contributory factor in achieving one third of these clear ups.     

Use of footwear impressions in crime scene investigations assisted by computerised footwear collection system

Crime scene footwear classification systems can be utilized to look for possible suspects, to get models and brand names for crime scene impressions and to link crime scenes. In this work a computerised footwear classification system is presented. The classification coding of the system has been designed especially for partial footwear impressions which are the most typical in crime scenes. The system is flexible because it is possible to agree within a crime scene investigation unit the way in which the system is to be applied. If outsole patterns are classified accurately in processing for the database, time will be saved in the search process and vice versa. The accurate classification coding is propounded by the fact that only a few experienced users are needed to perform the classification and data storage and all police officers returning from crime scenes are able to perform searches very easily. The users only have to follow the defined rules for geometric shapes. The crime scene investigation unit of the Turku police tested the system for a period of 1.5 years. The system has proved to be of valuable assistance in investigation when several of the hints lead to early solving of a crime. Practical experiences are described.     

Palynology and mycology as biological evidence in a homicide case

Criminal cases are studied from several disciplines to link a suspect with a criminal act. In this case, a man was reported missing in a coastal area in Buenos Aires Province, Argentina. The victim's relatives pointed to a possible suspect, and the local police carried out the investigation. We contributed to this research by applying palynological and mycological techniques. Palynomorphs and fungal spores offer valuable trace evidence, as they can be easily transferred between objects and crime scenes due to their minute size and persist on them for a long time. The victim was found 25 days later, lying on sandy soil, which partially covered the body, 35 km from where the suspect was arrested. Comparative samples were collected from the crime scene and the suspect's home and belongings (clothes, footwear, and seized vehicle). The palynological associations obtained from the crime scene and the defendant's belongings were dominated by diatoms and acritarchs (Acantomorphitae), all elements of marine origin, and a high CFU number of Bipolaris cynodontis, which allowed the defendant's clothing to be related to the place of corpse discovery. Soil from the defendant's home had an entirely continental composition, and the fungal biota was characteristic of prairie areas which were different from those of the crime scene.     

Development of a nanoplate-based digital PCR assay for species identification with mixture deconvolution

In crime scenes, not all biological stains are human in origin. Some exhibits can be from pets living on the premises or from animal products used in food consumption. In addition, it could be necessary to test animal carcasses for other forensic purposes. Often such stains can include mixtures involving humans or other species. Thus, identifying and deconvoluting mixtures of species commonly found in and around a household can be crucial in forensic casework. Different molecular techniques have been employed for species identification such as immunoprecipitation, qPCR, and DNA sequencing.In this project, a nanoplate-based digital PCR assay for species identification was developed, targeting Homo sapiens, canine, feline, bovine swine, pisces, and gallus in two multiplexes. An internal positive control was included in the design. The assay is simple, rapid, and can determine a wide variety of different vertebrates from biological exhibits, as well as in mixtures. Because the assay utilizes digital PCR, the procedure shows sensitivity down to a few copies, even in the presence of larger amounts of a major contributor, making the assay particularly useful in mixture deconvolution. Overall, this assay presents the forensic community with a novel application in which digital PCR can provide a sensitive and specific determination of species.     

Elimination of Bioweapons Agents from Forensic Samples During Extraction of Human DNA

Collection of DNA for genetic profiling is a powerful means for the identification of individuals responsible for crimes and terrorist acts. Biologic hazards, such as bacteria, endospores, toxins, and viruses, could contaminate sites of terrorist activities and thus could be present in samples collected for profiling. The fate of these hazards during DNA isolation has not been thoroughly examined. Our goals were to determine whether the DNA extraction process used by the Royal Canadian Mounted Police eliminates or neutralizes these agents and if not, to establish methods that render samples safe without compromising the human DNA. Our results show that bacteria, viruses, and toxins were reduced to undetectable levels during DNA extraction, but endospores remained viable. Filtration of samples after DNA isolation eliminated viable spores from the samples but left DNA intact. We also demonstrated that contamination of samples with some bacteria, endospores, and toxins for longer than 1 h compromised the ability to complete genetic profiling.     

Shoeprint image retrieval and crime scene shoeprint image linking by using convolutional neural network and normalized cross correlation

A shoeprint image retrieval process aims to identify and match images of shoeprints found at crime scenes with shoeprint images from a known reference database. It is a challenging problem in the forensic discipline of footwear analysis because a shoeprint found at the crime scene is often imperfect. Recovered shoeprints may be partial, distorted, left on surfaces that do not mark easily, or perhaps come from shoes that do not transfer marks easily. In this study, we present a shoeprint retrieval method by using a convolutional neural network (CNN) and normalized cross-correlation (NCC). A pre-trained CNN was used to extract features from the pre-processed shoeprint images. We then employed NCC to compute a similarity score based on the extracted image features. We achieved a retrieval accuracy of 82% in our experiments, where a “successful” retrieval means that the ground truth image was returned in the top 1% of returned images. We also extend our shoeprint retrieval method to the problem of linking shoeprints recovered from crime scenes. This new method can provide a linkage between two crime scenes if the two recovered shoeprints originated from the same shoe. This new method achieved a retrieval accuracy of 88.99% in the top 20% of returned images.     

The evaluation of methods for soil analysis under simulated scenes of crime conditions

Five simulated scenes of crime have been studied in which the soils adhering to shoes, car tyres and plant roots have been compared with multiple control samples using dry colour, ashed colour, pH, saccharide levels and particle size analysis. Colour and pH were found to be as reproducible under simulated scenes of crime conditions as for true duplicates from homogeneous soil, whereas the variation in saccharide levels was slightly greater than for true duplicates. The interpretational scheme developed for the silt and sand fractions of homogeneous soils was applied to the samples from each simulated scene of crime and every comparison was found to be similar at the 99% confidence limit.     

Influence of the luminol chemiluminescence reaction on the confirmatory tests for the detection and characterization of bloodstains in forensic analysis

Preliminary tests for the detection of stains at crime scenes aim to focus the police work making them more efficient in the combat of criminality. The application of the luminol chemiluminescence reaction (3-aminoftalhidrazida) in presumptive tests for the detection of bloodstains is known for more than 40 years in forensic science. This reaction is based on the emission of light through the chemical reaction of luminol mixed with hydrogen peroxide and a hydroxide in the presence of a catalytic molecule (iron from the hemoglobin) (Laux [1]).This work evaluates the luminol interference and its effect on subsequent serological and DNA testing. Samples prepared with blood and different concentrations of luminol solution containing luminol, peroxide of hydrogen and sodium carbonate, were analyzed. Additionally, samples of serial dilutions of standard DNA mixed with luminol solution were also analyzed. Although presumptive tests with luminol do not establish the characterization and identification of stains at crime scenes, preliminary results indicated that it is suitable for the detection of invisible bloodstains for forensic analysis, with few detrimental effects on the serological tests and subsequent DNA recovery and typing.     

The Extraction and Recovery Efficiency of Pure DNA for Different Types of Swabs

The extraction and recovery efficiency of swabs used to collect evidence at crime scenes is relatively low (typically <50%) for bacterial spores and body fluids. Cell‐free deoxyribonucleic acid (DNA) is an interesting alternative compared to whole cells as a source for forensic analysis, but extraction and recovery from swabs has not been tested before using pure DNA. In this study cotton, foam, nylon flocked, polyester and rayon swabs are investigated in order to collect pure DNA isolated from saliva samples. The morphology and absorption capacity of swabs is studied. Extraction and recovery efficiencies are determined and compared to the maximum theoretical efficiency. The results indicate that a substantial part of DNA is not extracted from the swab and some types of swab seem to bind effectively with DNA. The efficiency of the different types of swab never exceeds 50%. The nylon flocked 4N6FLOQSwab used for buccal sampling performs the best.