Polymerase Chain Reaction (PCR) has a wide range of applications and usages in many disciplines of science. Some PCR failure can be attributed to the presence of inhibitors in the sample. Thirteen commonly encountered PCR inhibitors in forensic DNA analysis are investigated throughout this review. These inhibitors are humic substances, humin, humic acids, fulvic acids, hematin, hemoglobin, Immunoglobulin G, tannic acid, calcium, collagen, melanin, bile salts, and urea. PCR inhibitors either affect the amplification, known as amplification inhibitors, the fluorescent component, known as detection inhibitors, or a single inhibitor can produce effects through both mechanisms. In reviewing the current literature, three main methods to remove or mitigate PCR inhibition were identified; with the addition of an additive, using specialty coated magnetic beads, or using a spin column. This review discusses the advantages and disadvantages of each method and the success of each in regards to some of the inhibitors of interest.

Fingernail evidence is often collected in criminal cases of violent and/or sexual assault. In acts of aggression and self-defense, foreign deoxyribonucleic acid (DNA) can be transferred from the perpetrator to beneath the surface of the fingernail of the victim. It is possible to recover this foreign, exogenous DNA from the victim’s fingernails and potentially identify the perpetrator via DNA analysis. When attempting to recover this DNA from fingernail clippings, a couple of problems can occur. Often times, not enough exogenous DNA gets trapped underneath fingernails, so there is not usually much DNA to work with for recovery. The other major problem is the presence of endogenous DNA from the fingernail donor. Not only is there donor DNA in the fingernail itself, but the donor’s own DNA can build up underneath their nails simply by rubbing their face or combing their fingers through their hair. This means that there can be more donor DNA present that can mask the presence of the foreign DNA and cloud the results. In an attempt to improve the recovery of foreign DNA and produce a reportable, informative profile, a time course study was developed. Typically, when using forensicGEM as an extraction method, the samples would incubate for 15 minutes before going through protease inactivation. For this study, the extraction period was broken up into four 5-minute periods of incubating, for a total of 20 minutes, before inactivating the protease. This was done to pinpoint the time period at which more foreign DNA is being extracted from the surface of the nail before endogenous DNA is extracted in excess and clouds or even hides the presence of foreign DNA altogether. Female fingernail clippings were spiked with neat male saliva to observe the ratio of male to female DNA during quantitation and on the electropherograms. The quantitation results depicted a strong presence of male DNA through the entirety of the time course, and female DNA did not appear to be extracted in greater levels until the 15 minutes of incubation. The resulting profiles exhibited the male saliva profile as the major contributor for most of the samples, especially at the 5- and 10-minute markers. In 50% of the profiles, a minor female contributor could be identified as the nail donor. One sample produced a single, male profile for each time point with no indication of a female donor present in the extract; another sample produced a profile with a male major contributor with only 3 to 6 loci having additional detectable alleles of a minor contributor at each time point. These alleles could not be conclusively attributed to the female nail donor, but she could not be excluded. These preliminary results indicate that a shortening of the forensicGEM extraction period could be beneficial for improving the recovery ratio of exogenous to endogenous DNA from fingernail evidence.

Although the pubic symphysis remains the most commonly utilized osteological feature to ascertain age-at-death estimations by forensic anthropologists (Garvin and Passalacqua, 2012), these aging methods do not accurately age elderly individuals. Through the re-examination of a Balkan sample, Berg (2008) noted a morphological variant, which may be correlated to osteoporosis expression by the increasing presence of macroporosity, present on female individuals that had previously been unexplained. This morphological variant can assist in the application of the Suchey-Brooks method to age elderly female individuals with the inclusion of a seventh phase (Berg, 2008). Hartnett (2010) also re-examined the pubic symphysis to better estimate age for modern populations and noted morphological variants similar to those described by Berg (2008). Hartnett (2010) attributed this variant to a decrease in bone quality associated with age-related morphological change. The present study examined the 330 female skeletons housed at Lisbon Collection, with the specific aim to provide a comparison of pubic symphysis age estimation methods, including Suchey-Brooks (1990), Berg (2008), Hartnett (2010), and Boldsen et al. (2002), on a known modern skeletal collection geographically dissimilar from the collections originally examined by Berg (2008) and Hartnett (2010). This dissimilar population was important because Berg's original study noted regional differences in the appearance and applicability of the seventh phase. The morphological variants present in female elderly individuals in the Lisbon Skeletal Collection support the introduction of a seventh phase to the standard Suchey-Brooks pubic symphysis method. Using the seventh phase, the Berg (2008) and Hartnett (2010) method improved their accuracy rates for aging older individuals. However, when the entire female population sample is considered the established age-at-death estimation methods do not perform well. The relationship between bone quality, aging method estimation assessment, and known age are discussed with considerations made for the influencing factors on bone preservation. A major difficulty in this analysis was parsing out information regarding bone density loss that occurred as natural degeneration and had a relationship to age-related change. The most significant confounding factor for the analysis of bone density loss and its importance to age-related change is the influence of bone preservation. While it is clear that the seventh phase provides more valuable information for the age estimation of the elderly, the poor correlation of bone quality suggests that this feature is not particularly important for the assessment of elderly phases in this population. This research supports the induction of a seventh phase to help provide more accurate age estimations for elderly populations, as it has been found in various populations, including the Portuguese population.

The present research examined the effects of compound bows and crossbows on the remains of Sus scrofa and Odocoileus virginianus. Isolated pig heads and white-tailed deer necks were impacted by three forms of arrow heads: the broad-head tip, conical field-tip, and bullet field-tip from both the compound bow and the crossbow. The structural design of the arrowheads was examined to understand their level of impact, as well as, the velocities of the compound bow and crossbow were calculated and compared. The total number of impact marks for the experiment was 55. It was hypothesized that the compound bow would have a greater extent of trauma to bone than the crossbow due to the higher velocity created from a longer power stroke. It was also hypothesized that the broad-head arrow tip will create larger fracture patterns on bone due to the three-blade-prong design compared to the oval shape of both the conical field-tip and bullet field-tip. Through the use of one-way ANOVA and Pearson’s Chi-Square, the results show no direct correlation between the difference in the type of weapon used or the arrow tip used. The results show the vast majority of impacts are penetration with shapes that roughly resemble the cross-section of the type of tip used. The results, however, did not support both hypotheses due to the limited number of impact marks and sample size of the experiment. Further experiments are required to assess the extent to which it is possible to distinguish between arrow related osseous trauma.

Fundamentals of forensic deoxyribonucleic acid (DNA) typing for sexual assault samples require the successful application of a differential extraction. Gynecological swabs containing vaginal epithelial cells and sperm cells are commonly encountered in forensic casework. A priority in sexual assault casework is the identity of the male contributor and in order to identify the male contributor, separation of the vaginal epithelial cells and sperm cells must be achieved. Two considerations when separating the different cell-types from a substrate are cellular elution and purity of the DNA fractions. Maximizing DNA yield is directly proportional to the number of cells eluted off of the swab and the extraction method utilized. The trypsin-ZyGEM extraction method has shown results of increased DNA recovery on liquid mixture samples compared with the standard differential extraction, which uses proteinase K and dithiothreitol (DTT). The trypsin-ZyGEM differential extraction protocol calls for the use of proteases EA1, incorporated in the forensicGEM extraction kit, and trypsin, a serine proteinase that has been discovered to effectively digest the DNA bound protamines in sperm cells. The trypsin-ZyGEM protocol follows a similar preferential lysis procedure to the standard differential extraction; however, everything is incorporated into one tube, therefore, minimizing loss of DNA from transfer techniques in the trypsin-ZyGEM protocol. Initially, epithelial cells are lysed using the forensicGEM enzyme and removed from solution after centrifugation. Samples are subsequently treated with trypsin, digesting sperm cells. The resultant solution contains the sperm cell DNA and other cell components. The standard differential extraction commonly uses a Qiagen silica membrane column to purify DNA away from cellular proteins and other contaminants, ensuring successful downstream DNA testing with the polymerase chain reaction (PCR). However, treatment with the trypsin-ZyGEM method is followed by a second incubation with forensicGEM after sperm cells are lysed with trypsin. Extracted samples can be quantified and followed through to DNA typing. Expanding previous studies and optimizing conditions of the dual enzyme approach were explored in this study for semen samples on cotton swabs. When comparing DNA yields of extracted samples, tris-ethylenediaminetetraacetic acid (TE) buffer recovered more cells from the cotton swabs than phosphate buffered saline (PBS) buffer and Buffer ATL, used in the Qiagen QIAamp® DNA Investigator Kit. Relative to DNA recovery of the standard Qiagen differential protocol, the trypsin-ZyGEM method on the cotton swab samples appeared subpar and could be attributed to ineffective cellular elution. Additionally, carryover DNA into the non-sperm cell fraction was exhibited. Procedures with Accumax, a cell detachment solution, were implemented in an attempt to elute more cells. The resulting DNA yields were significantly lower in the presence of Accumax. Contrastingly, incorporating trypsin directly into the elution TE buffer exhibited significant increases in DNA recovery. The direct lysis of sperm cells on the swabs, rather than the two-phase method of eluting the cells from the swab and subsequently extracting the DNA, proved to be much more effective. The swab remains from the elution and subsequent trypsin-ZyGEM method were re-extracted using the direct lysis method. Whilst comparing DNA yields, it was discovered that approximately 90% of the cellular DNA was retained on the swab after the two-phase extraction method was performed. Experiments utilizing direct lysis with the trypsin-ZyGEM method, the standard Qiagen differential protocol, and the trypsin lysis followed by Qiagen extraction were performed on swab samples comprised of different semen concentrations resulting DNA yields and short tandem repeat (STR) DNA profiles were compared. Results obtained indicated that the trypsin-ZyGEM method provided substantially larger DNA recovery yields and provided full STR profiles to a target mass of 0.0625 ng and average peak height ratios above 60%. Successful implementation of this extraction procedure will require further studies with the addition of epithelial cells on the swab samples to mimic vaginal swab mixture samples. These samples will help to determine purity of the DNA fractions and effects of epithelial cells on the trypsin-ZyGEM protocol. / 2017-11-03T00:00:00Z

Processing sexual assault samples is a difficult time consuming task for the forensic analyst. Samples tend to be a mixture of the victim’s epithelial cells and the male suspect’s sperm cells that need to be separated prior to extraction of Deoxyribonucleic Acid (DNA). Without separation of the two cell types, the DNA extract would result in an uninterpretable mixture. In 1985, Peter Gill and colleagues outlined a procedure known as preferential lysis that would aid in the separation of female and male cells from sexual assault samples. The basis of this procedure, commonly referred to as a differential extraction, utilizes differences in the packaging of DNA between the two cell types to preferentially lyse the female epithelial cells and leave the sperm intact. By pelleting the sperm and removing the supernatant (termed the Non-Sperm Fraction) the Sperm Fraction can now be extracted without the contaminating epithelial cells. This procedure has been widely implemented in forensic laboratories and is still being used today over 30 years later. However, there are certain conditions under which this procedure does not perform sufficiently including excess of female epithelial cells and low amounts of sperm. Unfortunately, both of these conditions are common among sexual assault samples. The procedure also is quite long, and with the backlog of sexual assault samples continually growing in the United States, there is a need for a new procedure that is faster and performs optimally under the previously mentioned conditions. This research explores the use of two enzymes, EA1 (marketed by Zygem Corporation as ForensicGEM Saliva™) and Trypsin to separate the cells. Due to the inability of Zygem to cleave the disulfide bonds present in the sperm DNA packaging proteins, treatment of mixed samples with Zygem will lyse the epithelial cells and leave the sperm intact. The incubation time of Zygem is much faster than that of the Gill method and can be performed in one tube, minimizing the chances of DNA loss and contamination during transfers. Treatment of the pelleted Zygem extract with Trypsin effectively and rapidly lyses the sperm cells. Combining these methods into a differential extraction protocol has the potential to be a rapid, easily implemented procedure. Results from the Zygem-Trypsin differential extraction method showed incomplete separation of the two fractions due to the incomplete lysis of the epithelial cells by Zygem. The resultant profiles did show a major male contribution with a minor female component, however these results are not sufficient enough for real casework. While further research and development of the protocol are necessary, the Zygem-Trypsin differential extractions performed here show the potential for a rapid, easy differential extraction procedure that could be easily implemented in any laboratory. / 2017-11-03T00:00:00Z

Computer crimes have become very complex in terms of investigation and prosecution. This is mainly because forensic investigations are based on artifacts left oncomputers and other digital devices. In recent times, perpetrators of computer crimesare getting abreast of the digital forensics dynamics hence, capacitated to use someanti-forensics measures and techniques to obfuscate the investigation processes.Incases where such techniques are employed, it becomes extremely difficult, expensive and time consuming to carry out an effective investigation. This might causea digital forensics expert to abandon the investigation in a pessimistic manner.ThisProject work serves to practically demonstrate how numerous anti-forensics can bedeployed by the criminals to derail the smooth processes of digital forensic investigation with main focus on data hiding and encryption techniques, later a comparativestudy of the effectiveness of some selected digital forensics tools in analyzing andreporting shreds of evidence will be conducted.

The present study analyzes cranial nonmetric traits used in forensic ancestry estimation on contemporary skeletal remains of modern European Americans in order to determine if there are statistically significant differences between males and females in trait expression. Research on cranial nonmetric traits for ancestry estimation has largely ignored the effects of sexual dimorphism on trait expression; however, there is growing evidence that some traits may be impacted by sex, among other variables. The 17 macromorphoscopic traits described in Hefner and Linde (2018) and the six mandibular morphoscopic traits described in Berg (2008) were scored on 97 females and 113 males from the Texas State University Donated Skeletal Collection in San Marcos, Texas. Chi-square tests were used to analyze if there are statistically significant cranial nonmetric trait expressions between males and females. From these tests, the results indicate that 14 out of the 23 cranial and mandibular nonmetric traits are statistically significantly different between the sexes, with a p-value less than 0.05. Gonial angle flare is the most significant feature, while the zygomaticomaxillary suture is the least significant feature. Additionally, correspondence analyses (CA) show the relationship between each cranial nonmetric trait score, that demonstrated significance, and both sexes. Ultimately, this research demonstrates that several nonmetric traits used in ancestry estimation are affected by sex; thus, it may be beneficial to develop sex-specific ancestry models for nonmetric traits.

The Peganum harmala plant is a flowering shrub that produces small, dark brown seeds in pods. These seeds contain the hallucinogenic alkaloids harmine and harmaline. As such, they have been historically used for shamanic rites and folk medicine. Presently, P. harmala seeds are commercially available and subject to no legal restrictions in the United States. This has allowed for the recreational use and abuse of these hallucinogenic seeds or seed extracts made with household chemicals. Overdose cases from excessive consumption of seeds or seed extracts have been reported. Overdose patients present with hallucinations, tremors, agitation, tachycardia, and gastric distress. Severe overdose cases have resulted in hospitalization for respiratory depression and coma. The goal of this research was to develop a protocol for forensic analysis of suspected P. harmala seeds. Physical examination was selected as a quick, cost-effective preliminary method to screen seeds. P. harmala seeds are, on average, approximately 2.3 ± 0.3 mm long and 1.0 ± 0.2 mm thick, with an average Feret’s diameter of 2.8 ± 0.3 mm. The mean mass of one seed is 2.5 ± 0.2 mg. The seeds are dark brown, irregularly shaped, and have a pitted surface. Seeds matching these descriptors can be further analyzed to detect harmine and harmaline. Direct analysis in real time (DART) allows for very rapid mass spectral analysis of P. harmala seeds. Ions corresponding to harmine and harmaline can be detected when an intact seed is placed in front of the DART ion source, and higher levels of harmine and harmaline are observed when a seed cut in half to reveal interior surfaces is analyzed. Solvent extraction of crushed seeds using ethanol followed by gas chromatography – mass spectrometry allows for confirmation of the presence of harmine and harmaline in suspected seeds. When selected ion monitoring is used, this method is able to detect harmine and harmaline in a sample consisting of a single seed. Infrared spectra of harmine and harmaline standards, crushed P. harmala seeds, and solid material obtained from evaporating off the solvent from an extraction of crushed seeds were obtained. Infrared spectroscopy can be used to distinguish between pure harmine and harmaline, but is a poor choice for analysis of samples containing a mixture of harmine and harmaline, such as P. harmala seeds. In conclusion, physical characterization, direct analysis in real time, solvent extraction, and gas chromatography – mass spectrometry are recommended techniques for the forensic analysis of P. harmala seeds.

This article considers forensic anthropologists’ roles in negotiating the concept of criminality in biohistorical cases, those investigations of the famous and infamous dead that are driven by public interest rather than traditional medicolegal relevance. We review three biohistorical cases from the United States: the purported skull of a martyred Catholic priest from sixteenth century Georgia, the Mountain Meadows Massacre that occurred in Utah in the mid-nineteenth century, and the search for Billy the Kid’s grave in New Mexico. We find that anthropologists have active and passive roles in the manufacture, assignment, and sometimes denial of criminality in these cases. Additionally we explore how the analysis and discussion of violence in these biohistorical cases reflects two concepts that are distinctive to United States’ history, notably manifest destiny and the idea of closure in historical narratives. The perception that the present order is a natural culmination of history, and that the past is truly past underestimates the relevance and impact of labelling past personages as criminals to contemporary culture. As a result, forensic anthropologists’ negotiation of criminality in U.S. biohistorical cases is fraught with nebulous ethical challenges and tangible consequences.