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The Effect of Bone Marrow Transplants on DNA profiles; a case example

  • Principal Forensic Services, UK

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A case was received in which a woman suspected that she had been sexually assaulted or raped while unconscious, with a request to establish whether a profile suitable for the National DNA Database could be obtained. Mixed DNA profiles were obtained from the external vaginal swab, body fluids on the knickers and a reference buccal swab taken from the complainant. Samples of blood and hair from her gave DNA profiles that were different to each other. Further enquiries revealed that she had received a bone marrow transplant (BMT) as a treatment for leukaemia. This article describes the results obtained in this case, together with the possible effect of a BMT on casework and the impact on the NDNAD.
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... A patient with successful bone-marrow transplantation will possess a variation in their DNA profile, i.e., blood will possesses the complete profile of the donor while the nail and buccal fluids are will contain a mixed DNA profile and hair will show an unmixed profile of the receiver (Pope and Chapman 2006). Aforesaid points are of paramount importance for DNA profiling and must be put into consideration from a legal viewpoint. ...
Background: The scientific and technological advances along with the changing socio-economical standards of society have posed new challenges to the criminal justice system. With the changes in society and technology, there is also an increase in crime rate. Forensics provides the scientific proofs beyond the shadow of reasonable doubt and thus significantly contributes in criminal as well as civil investigations and legal matters. Main body of the abstract: Discovery of DNA has opened new avenues and the advancement of DNA technology and its introduction into the court of law has provided extensive aid in the resolution of civil and criminal disputes. In India, the DNA technology was first introduced in a paternity dispute in 1989. However, the need for legislation and guidelines to support grounds for the use of DNA profiling for forensic purposes in India has been recognized for some time now. In July 2019, the DNA Technology (Use and Application) Bill 2019 was introduced in the Indian parliament. Short conclusion: Herein, this article is focused on the current aspects of DNA based evidence in the Indian Criminal Justice system along with the associated issues, highlighting the need for specific DNA based legislation.
... This study holds more importance in situations where the subjects providing a DNA reference sample have undergone a bone marrow transplant (BMT) or frequent blood transfusion (37), as well as in some touch DNA studies which may need a reference sample from the skin surface of subjects (38). ...
An acceptable area for collecting DNA reference sample is a part of the forensic DNA analysis development. The aim of this study was to evaluate skin surface cells (SSC) as an alternate source of reference DNA sample. From each volunteer (n = 10), six samples from skin surface areas (forearm and fingertips) and two traditional samples (blood and buccal cells) were collected. Genomic DNA was extracted and quantified then genotyped using standard techniques. The highest DNA concentration of SSC samples was collected using the tape/forearm method of collection (2.1 ng/μL). Cotton swabs moistened with ethanol yielded higher quantities of DNA than swabs moistened with salicylic acid, and it gave the highest percentage of full STR profiles (97%). This study supports the use of SSC as a noninvasive sampling technique and as a extremely useful source of DNA reference samples among certain cultures where the use of buccal swabs can be considered socially unacceptable.
... This change of genetic profile, in which donor and recipient hematopoietic cells can coexist, is called chimerism. 2 One of the methods used to verify chimerism is short tandem repeat (STR) testing. STRs, or microsatellites, are repetitive sequences of three to seven base pairs that identify different alleles by the number of copies of repetitive sequences contained in the DNA region analyzed. ...
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Hematopoietic stem cell transplantation is the curative option for patients with myelodysplastic syndrome; however, it requires a long post-transplantation follow-up. A 53-year-old woman with a diagnosis of myelodysplastic syndrome underwent related donor allogeneic hematopoietic stem cell transplantation in July 2006. Three months after transplantation, a comparative short tandem repeat analysis between donor and recipient revealed full chimerism, indicating complete, healthy bone marrow reconstitution. Three years and ten months after hematopoietic stem cell transplantation, the patient developed leukopenia and thrombocytopenia. Another short tandem repeat analysis was carried out which showed mixed chimerism (52.62%), indicating relapsed disease. A donor lymphocyte infusion was administered. The purpose of donor lymphocyte infusion is to induce a graft-versus-leukemia effect; in fact, this donor's lymphocyte infusion induced full chimerism. Successive short tandem repeat analyses were performed as part of post-transplantation follow-up, and in July 2010, one such analysis again showed mixed chimerism (64.25%). Based on this finding, a second donor lymphocyte infusion was administered, but failed to eradicate the disease. In September 2011, the patient presented with relapsed disease, and a second related donor allogeneic hematopoietic stem cell transplantation was performed. Subsequent short tandem repeat analyses revealed full chimerism, indicating complete bone marrow reconstitution. We conclude that quantitative detection of mixed chimerism is an important diagnostic tool that can guide early therapeutic intervention.
... His physician was contacted and informed us that his patient suffered from a lymphoma but had not received blood transfusions or a bone marrow transplant. Although blood transfusion is not thought to affect DNA profiling [2], this medical information allowed to exclude chimerism associated with medical intervention (i.e., artificial chimerism) see, e.g., [3][4][5][6] as being the explanation of the observed pattern. Natural chimerisms resulting from the fusion of zygotes [7], from exchanges of blood between fetuses in utero, e.g., [8][9][10], or from double parental contribution, e.g., [11][12][13][14] as well as mosaicism are compatible with our data. ...
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Nuclear DNA markers, such as short tandem repeats (STR), are widely used for crime investigation and paternity testing. STR were used to determine whether a piece of tissue regurgitated by a dog was part of the penis of a dead, emasculated, man. Unexpectedly, when analyzing the recovered material and a blood sample from the deceased, five out of the 18 loci differed. According to the results, one could have concluded that these samples originated from two different persons. However, taking into account contextual information and data from complementary genetic analyses, the most likely hypothesis was that the deceased was a genetic mosaic or a chimera. Within a forensic genetic context, such genetic peculiarities may prevent associating the perpetrator of an offense with a stain left at a crime scene or lead to false paternity exclusions. Fast recognition of mosaics or chimeras, adapted sampling scheme, as well as careful interpretation of the data should allow avoiding such pitfalls.
DNA testing is a powerful tool in criminal investigation. The advent of DNA profiling has revolutionized the criminal justice system. Since the discovery of DNA till now, it has provided the cogent evidential base to corroborate guilt or proving innocence in the court of law. The anticipation of technological advancement in the criminal justice system puts two significant concerns that are to protect the fundamental rights of the citizen, and other is eliminate the probability of misuse of technique. Many countries have enacted DNA legislation to provide the legal background to DNA testing along with the establishment of DNA databases, while India is in the process of adopting the DNA Technology (Use and Application) bill 2019. Legal perspectives of DNA testing in different nations are discussed in this chapter as to build an understanding of the status of DNA in the court of law, its admissibility, and ethical concerns.
Biological vestiges are used in forensic science to resolve a large number of cases by typing the genetic profile and identifying the individual to whom it belongs. However, chimeric persons that possess cells with two or more different DNA make these types of analyses difficult. This situation can occur naturally, by errors in the fertilization or early embryogenesis, or in an artificial way, for example after hematopoietic stem cell transplantation (HSCT), when host and donor cells coexist in the patient. In this paper, we will specially focus on the latter. The vestiges from transplant patients represent a challenge from a forensic perspective since the interpretation of the genetic fingerprint can be misleading because of the presence of chimerism. Due to the high number of transplant patients (and their increase over the years) and the existence of natural chimeras (probably many of them hidden), it is necessary to consider whether we are facing a possible chimeric person or someone who has been a donor of hematopoietic stem cells in a forensic context. In this review, the presence of donor bone marrow derived cells in some tissues of forensic interest will be discussed. Finally, to emphasize the importance of chimerism after HSCT in forensic genetics, some real-life cases will be examined.
As criminalistics is taking advantage today of the progress of sciences, mainly dedicated to sensibility and sensitivity, crime scene management remains purely practical, led by the less educated people of the forensic chain, although the whole interpretation process is highly depending on it. Instead of favouring the optimization of the judicial system through the primacy of biometric traces tending to their exclusive collection, won't it be time to recognize the global concept of forensic science to found these crucial choices, which are surpassing the simple good justice deal?.
Organ transplantation is one of the most important services of modern medicine to the humanity. In judicial death cases the interaction between judicial needs and transplantation needs is inevitable and both should be provided in a short time before the decomposition of the body. Thus, the description of this interaction and the algorithm which should be carried out to manage these cases are important. Aim of this study is to determine the problems confronted in forensic autopsies and to determine what to do for both judicial processes’ and cadaveric organ donations’ not becoming limited due to each other. With these aims, autopsy case archive of the Council of Forensic Medicine Istanbul Morgue Department was reviewed, between the years 2009 and 2011, to reveal the number of organ donors among autopsy cases and also to find out the judicial problems confronted during autopsies. Among 12016 judicial death cases referred to Istanbul Morgue Department in 3 years, 35 cases were found to have undergone cadaveric solid organ harvesting procedure and 307 cases cornea-only harvesting procedure. Manner of deaths for organ donor cases were blunt trauma due to traffic accident in 20 cases, firearm injury in 3 cases, stabbing in 2 cases, suspicious criminal battery in 4 cases and fatal falls in 5 cases. Only 1 case was suspected to have died due to high dose insulin administration. Through the whole data presented in this study, it can be concluded that consulting with the Forensic Medicine Expert not only for the autopsies but also during the clinical process of a judicial case, who is a candidate to be an organ donor, is absolutely important. The early contribution of the Forensic Medicine Expert would provide help to plan both the judicial process and the transplantation process which needs urgent decisions. A Forensic Medicine Expert may be an organ harvest team member performing initial investigations on the cause of death and collecting some of the toxicological screening samples when needed.
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Chronic myeloid leukaemia (CML) can be treated successfully with allogeneic bone marrow transplantation (BMT) leading to long-term disease-free survival. Leukemia relapse, however, remains a significant clinical problem. Relapse following BMT presumably results from the expansion of small numbers of recipient leukaemic cells which have survived the conditioning therapy. In order to define patients who are at a high risk of leukaemia relapse, a variety of techniques have been employed to detect persistence of host haemopoiesis (mixed chimaerism, MC) or residual leukaemia (minimal residual disease, MRD). However, the precise relationship between the detection of MC and MRD post-BMT is unknown. We have investigated chimaerism and MRD status in 22 patients who were in clinical and haematological remission post-allogeneic BMT for chronic phase CML. Chimaerism was assessed using short tandem repeat PCR (STR-PCR) while BCR-ABL mRNA detection using reverse transcriptase polymerase chain reaction (RT-PCR) was performed to detect the presence of MRD. Seventeen patients received unmanipulated marrow (non-TCD) while in five patients a T cell-depleted transplant (TCD) was performed as additional GVHD prophylaxis. Chimaerism was evaluated in 18 patients (14 non-TCD, four TCD). Mixed chimaerism was an uncommon finding in recipients of unmanipulated BMT (21%) when compared to TCD BMT (100%). No evidence of MRD, as identified using the BCR-ABL mRNA RT-PCR assay, was detected in those patients who were donor chimaeras. Early and transient MC and MRD was detected in four patients (two non-TCD, two TCD) who have subsequently converted to a donor profile. One patient has stable low-level MC but remains MRD negative 4 years post-BMT. Late MC and MRD was observed in two patients who relapsed >6 years after TCD BMT for CML. We conclude that mixed chimaerism is a rare event in recipients of unmanipulated BMT and that donor chimaerism as detected by STR-PCR assay is consistent with disease-free survival and identifies patients with a low risk of leukaemic relapse post-BMT for CML.
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Monitoring the engraftment of donor cells after allogeneic blood stem cell transplantation (BSCT) may be important for the early diagnosis of graft failure or relapse of disease. Several techniques have been reported for this purpose. PCR-based assays analyzing polymorphic short tandem repeat (STR) markers are attractive because they are sensitive and can be performed rapidly. The intent of the present study was to test a novel approach for the quantification of mixed chimerism using a commercial multiplex STR assay with fluorescence-based detection for forensic purposes. The feasibility of this assay and the accuracy of quantitative results was tested using serial cell mixtures of unrelated individuals. Sample preparation was optimized to obtain information from minute amounts of starting material, eg from patients with aplasia or from sorted cell populations. Using the STR-PCR, discrimination between donor and recipient was possible in all patients analyzed (n = 25). Cell dilution experiments showed a linear correlation between the cell numbers added and the proportions found, with the limit of detection for a minor cell population being 5%. Comparison of values obtained with standard FISH analysis in patients transplanted from sex-mismatched donors showed an excellent correlation with the STR-PCR results. Taken together, this procedure allows the rapid, versatile and accurate quantification of mixed chimerism, even with minuscule numbers of cells.
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Detection of chimerism by PCR analysis of short tandem repeats (STR) in blood samples of patients who received allogeneic bone marrow transplantation (BMT) has proved to be an important method for early detection of relapse. The prerequisite for this type of analysis is knowledge of donor and recipient pretransplantation genotypes. In some cases, recipient cells from time points prior to BMT are not available and the pretransplant fingerprint cannot be determined. As BM recipients only alter their genotype in blood cells, we attempted to identify patient's pretransplantation genotypes after transplantation in mouthwash samples that contain easily accessible epithelial cells. Of 17 patients who had undergone BMT between one week and 45 months prior to analysis, DNA was isolated from mouthwash cell pellets or from epithelial cells obtained from mouthwashes. PCR analysis of STR loci in the von Willebrand and the tyrosine hydroxylase genes were performed. Even though the mouthwash cell pellets contained about 75% epithelial cells (presumably of recipient origin) and only about 25% leukocytes (presumably of donor origin), three of five patients showed donor genotype and only two patients exhibited chimeric DNA patterns, when cellular DNA was obtained by boiling of mouthwash cell pellets. Following phenol/chloroform extraction, eight of 10 DNA samples exhibited a chimeric pattern, while two of 10 DNAs showed only donor genotype. Of three patients, epithelial cells were attached to magnetic beads prior to DNA isolation. Even this DNA contained donor and recipient material. From our results it appears that blood cells serve as preferential DNA source in mouthwash samples and cannot be removed by epithelial cell separation.
Bone Marrow Transplantation is a high quality, peer-reviewed journal covering all aspects of clinical and basic haemopoietic stem cell transplantation.
The AMPFlSTR((R)) SGM Plus system is a commercially available STR multiplex produced by Applied Biosystems, a division of Perkin Elmer, Foster City, California, USA that supersedes SGM. The multiplex contains the six SGM loci, amelogenin and four additional loci. These additional loci are D3S1358, D19S433, D16S539 and D2S1338. Consequently, the match probability is significantly improved (conservatively quoted as 1 in 10(9) for reporting a full profile match). The system was subjected to validation. For example, ageing and degradation studies demonstrated semen stains to be the most stable evidence type, whereas buccal scrapes were the least stable. An apparent rise in the sensitivity increases the chance of obtaining successful results from the more difficult samples submitted for analysis. Two of the new loci (D3S1358 and D19S433) are low molecular weight (between 100 and 150 base pairs); this improved the success rates of the degraded samples where high molecular weight loci may drop out. Of 26 non-primates tested, four gave results that appeared as single peaks and were unlikely to cause interpretation problems. None of the 19 micro-organisms tested gave discernible results. Extensive casework and simulated casework studies demonstrated that SGM and SGM plus results were comparable. There was one example of a null allele (primer binding site mutation) recorded at the HUMFIBRA locus (in both systems). However, a concordance study of 1000 samples using both SGM and SGM plus did not demonstrate any differences in typing.
Ehninger G and Neubaur A Rapid quantification of mixed chimerism using multiplex amplification of short tandem repeat markers and fluorescence detection
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Thide C, Uorek M, Bomb-M, Ritter M, Mohr B, Brmdei C. Ehninger G and Neubaur A Rapid quantification of mixed chimerism using multiplex amplification of short tandem repeat markers and fluorescence detection. Bone Marrow Transplantat~on 1999: 23: 1055-1 060.
Buccal swabs but not mouthwash samples can be used to obtain pretransplant DNA fingerpr~nts from recipients of allogeneic bone marrow transplants
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T h i i C. PrmgeXmx G, Frelkg-Richter J and hninger G Buccal swabs but not mouthwash samples can be used to obtain pretransplant DNA fingerpr~nts from recipients of allogeneic bone marrow transplants. Letter to Bone Marrow Transplantation 2000: 25: 575-577.
Short tandem repeat analysis to monitor chimerism after bone marrow transplantation
  • Hasskamp