Forensic biology

Forensic biology is the use of biological principles and techniques in the context of law enforcement investigations. 

Forensic biology  mainly focuses on DNA sequencing of biological matter found at crime scenes. This assists investigators in identifying potential suspects or unidentified bodies.

Forensic biology has many sub-branches, such as forensic anthropology, forensic entomology, forensic odontology, forensic pathology, and forensic toxicology.

Disciplines

History 
The first known briefings of forensic procedures still used today are recorded as far back as the 7th century through the concept of utilizing fingerprints as a means of identification.

By the 7th century, forensic procedures were used to account criminals of guilt charges among other things.

Nowadays, the practice of autopsies and forensic investigations has seen a significant surge in both public interest and technological advancements. One of the early pioneers in employing these methods, which would later evolve into the field of forensics, was Alphonse Bertillon, who is also known as the "father of criminal identification". In 1879, he introduced a scientific approach to personal identification by developing the science of anthropometry. This method involved a series of body measurements for distinguishing one human individual from another.

Karl Landsteiner later made further significant discoveries in forensics. In 1901, he found out that blood could be categorized into different groups: A, B, AB, and O, and thus blood typing was introduced to the world of crime-solving. This development led to further studies and eventually, a whole new spectrum of criminology was added in the fields of medicine and forensics.

Dr Leone Lattes, a professor at the Institute of Forensic Medicine in Turin, Italy, has made significant additions into forensics as well. In 1915, he discovered a method to determine the blood group of dried bloodstains, which marked a significant advancement from prior techniques limited to analyzing liquid blood. This technique was adopted for criminal investigation.

In 1928, Albrecht. H.O, a German chemist, developed a chemical solution luminol, which can be used to detect trace amounts of blood stains at crime scenes.

Among others, Sir Alec Jeffreys further shaped forensics into what we know now. In 1984, he developed the DNA fingerprinting technique to examine the variations in the genetic code. This can be used to distinguish one individual from another. This method has become important in forensic science to assist police detective work, and it has also proved useful in resolving paternity and immigration disputes.

In 1983, Kary B. Mullis made a significant contribution to the fields of medicine and criminology  by developing the PCR (polymerase chain reaction) technique, which can amplify even trace amount of DNA segments in-vitro. Such DNA samples, often found in crime scenes in minute amounts, degraded states and sometimes mixed with various body fluids from multiple individuals, can be effectively amplified using PCR. Beyond forensics, PCR has been applied to a wide range of fields, including disease diagnosis and virus detection.

DNA analysis

DNA, or deoxyribonucleic acid, is one of the most popular pieces of evidence to recover at a crime scene. More often than not, evidence containing DNA is regarded as biological evidence. With all of the substantial advances that have been made regarding DNA, biological evidence is recognized to be the "golden standard" in forensic science.

At the scene, biological evidence must be initially visibly recognized. Sometimes this is not always possible and the aid of an alternative light source, or Advanced Light Source (ALS), is required. Once identified as a potential source, presumptive tests are conducted to establish the possibility of the specified biological presence (semen, saliva, blood, urine, etc.). If positive, samples are collected and submitted for analysis in the laboratory, where confirmatory tests and further tests are performed.

DNA analysis has numerous applications such as paternity testing, identifications of unknown human remains, cold case breakthroughs, as well as connecting suspects and/or victims to a piece(s) of evidence, a scene, or to another person (victim or suspect, respectively).Nuclear DNA evidence can be recovered from blood, semen, saliva, epithelial cells and hair (if the root is still intact). Furthermore, Mitochondrial DNA (mt DNA) can be recovered from the shaft of hair, bone and the roots of teeth.

For most forensic DNA samples, STR analysis of autosomal short tandem repeats (STR) is performed in an attempt to individualize the sample to one person with a high degree of statistical confidence. Here, STR markers for autosomal STR are used in forensic DNA typing to track down the missing, verify family connections, and potentially connect suspects to crime sites.

Laboratory analysis of DNA evidence involves the sample DNA being extracted, quantified, amplified, and visualized. There are several methods of DNA extraction possible including organic (phenol-chloroform) extraction, Chelex extraction, and differential extraction.

Quantitation is commonly conducted using a form of the polymerase chain reaction, known as real-time PCR, quantitative PCR (qPCR). qPCR is the preferred method of DNA quantitation for forensic cases because it is very precise, human-specific, qualitative, and quantitative. This technique analyses changes in fluorescence signals of amplified DNA fragments between each PCR cycle without needing to pause the reaction or open the temperature-sensitive PCR tubes. In addition to the components necessary for standard a PCR reaction (i.e. template DNA, carefully designed forward and reverse primers, DNA polymerase [usually Taq], dNTPs, and a buffer solution containing Mg2+), qPCR reactions involve fluorescent dye-labelled probes that complement and anneal to the DNA sequence of interest that lies between the two primers. A "reporter" (R) dye is attached at the 5’ end of the fluorescent probe, while a "quencher" (Q) dye is attached at the 3’ end. Before the DNA strands are extended by the polymerase, the reporter and quencher are close enough in space that no fluorescence is detected by the instrument (the quencher completely absorbs/masks the fluorescence of the reporter). As the polymerase begins to extend the strand, the 5' end of the probe is degraded by the polymerase due to its exonuclease activity. The reporter dye is released from the 5’ end. It is no longer quenched, thus enabling fluorescence detection. A graph is constructed for the sample DNA comparing the presence of fluorescence (y-axis) to cycle number (x-axis) of the qPCR process. This is then compared to a standard curve of the cycle fluorescence threshold (y-axis) versus the log of known DNA concentrations (x-axis). By comparing the sample data to the standard curve, one may extrapolate the DNA concentration in the sample, which is essential to move forward with PCR amplification and capillary electrophoresis to obtain a DNA profile. DNA profiles are produced as an electropherogram. The obtained profile can be compared to known samples in CODIS to identify a possible suspect. Based on known frequencies of the genotype found in the DNA profile, the DNA analyst may place a statistical measure of confidence on DNA match.

Mitochondrial DNA analysis 
Mitochondrial DNA (mtDNA) is used instead of nuclear DNA when forensic samples have been degraded, are damaged, or are in very small quantities. In many cases, there may be older human remains, sometimes ancient, and the only options for DNA collection are the body's bone, teeth, or hair.

mtDNA can be extracted from such degraded samples since its presence in cells is much higher than nuclear DNA. There can be more than 1,000 copies of mtDNA in a cell, while there are only two copies of nuclear DNA. Nuclear DNA is inherited from both the mother and the father but mtDNA is passed down from only the mother to all of her offspring. Due to this type of inheritance, mtDNA is useful for identification purposes in forensic work but can also be used for mass disasters, missing persons cases, complex kinship, and genetic genealogy.

The main advantage of using mtDNA is its high copy number. However, there are a few disadvantages of using mtDNA as opposed to nuclear DNA. Since mtDNA is inherited maternally and passed to each offspring, all members of the maternal familial line will share a haplotype. A haplotype "is a group of alleles in an organism that are inherited together from a single parent". Sharing this haplotype among family members can cause an issue in forensic samples because these samples are often mixtures that contain more than one DNA contributor. De convolution and interpretation of mtDNA mixtures is more difficult than that of nuclear DNA, and some laboratories choose not to attempt the process Since mtDNA does not recombine, the genetic markers are not as diverse as autosomal STRs are in the case of nuclear DNA. Another issue is that of heteroplasmy — when an individual has more than one type of mtDNA in their cells. This can cause an issue in interpreting data from questioned forensic samples and known samples that contain mtDNA. Having adequate knowledge and understanding of heteroplasmy can help ensure successful interpretation.

There are some ways to improve success of mtDNA analysis. Preventing contamination at all testing stages and using positive and negative controls is a priority. In addition, the use of mini-amplicons can be beneficial. When a sample of mtDNA is severely degraded or has been obtained from an ancient source, the use of small amplicons can be used to improve the success of amplification during PCR. In these cases primers amplifying smaller regions of HV1 and HV2 in the control region of mtDNA are used. This process has been referred to as the 'ancient DNA' approach.

The first use of mtDNA as evidence in court was in 1996 in State of Tennessee v. Paul Ware. There was only circumstantial evidence against Ware so the admittance of mtDNA from hairs found in the victim's throat and at the scene were key to the case.

In 2004, with the help of the National Center for Missing and Exploited Children and ChoicePoint, mtDNA was used to solve a 22-year-old cold case where the nuclear DNA evidence was not originally strong enough. After mtDNA analysis, Arbie Dean Williams was convicted of the murder of 15-year-old Linda Strait, which had occurred in 1982.

In 2012, mtDNA evidence allowed investigators to establish a link in a 36-year-old investigation into the murders of four Michigan children. Hair fibers found on the bodies of two of the children were tested and the mtDNA found to be the same for each sample. For the investigators this was a big break because it meant that the murders were likely connected.

Forensic anthropology 

Anthropology is applied to forensics most regularly through the collection and analysis of human skeletal remains. The primary goals of anthropological involvement include identification and aiding in scene reconstruction by determining details regarding the circumstances of the victim's death. In cases where conventional techniques are unable to determine the identity of the remains due to the lack of soft tissue, anthropologists are required to deduce certain characteristics based on the skeletal remains. Race, sex, age and possible ailments can often be determined through bone measurements and looking for clues throughout the skeletal structure.

This becomes necessary when conventional methods that use soft tissue  fail to establish the identity of remains. By examining bone measurements and other skeletal structure characteristics, anthropologists can often determine information such as race, sex, age, and potential health conditions.

Forensic botany 
Forensic botany deals with the study of plants, leaves, seeds, and pollen or plant properties (such as anatomy, growth, behavior, classification, population dynamics, and reproductive cycles) that would be considered as physical evidence.

Before a plant can aid the investigations, the plant must be identified first to see if it is from the location that the plant was found. Forensic botany can help investigators by providing a link between an individual and the crime scene by pinpointing the geographical location of missing bodies or estimate where the burial took place. Another way that is used is through the interrogation process where investigators compare the traces that the victim(s) are found and compare to the statements of the suspect(s). Forensic botany can also reveal whether a death is due to suicide, accident, or homicide. Another example of forensic botany aiding investigators is by moss, moss can establish the PMI of a human skeletal, through its growth rate.

For example, an elderly man fell a steep hillside and was found dead. They concluded that the man did not die from the fall but from hypertensive heart disease. Forensic botany concluded that the fall was approximately 10 m and there was a lane that joined the hill path around 9 m perpendicular form the body. It showed he had leaves on his left hand and sweater. There were also broken bushes in the scene. Forensic scientists specializing in forensic botany had grabbed five soil samples and some plant samples from the scene. The plant samples were analyzed through optical microscopy, and they also examined the victims' clothing since it did carry some plant elements. They did a macroscopic and microscopic analysis compared to the collected samples on the scene. It was concluded that there was no evidence of struggle caused by another person instead, the elderly man had lost his balance.

Subspecialties In Forensic Botany 
Subdisciplines within forensic botany examples include:

 forensic palynology (study of pollen and spores). Palynology can produce evidence of decomposition time, location of death or the time of year.
 dendrochronology (study of the growth of rings of trees stems and roots)
 Lichenology (study of lichens)
 Mycology (study of fungi)
 Bryology (study of bryophytes). Bryology is the easiest to find evidence since bryophyte (a species of plants) attaches to shoes and clothes easily. Bryophyte useful since even if it is ripped apart or broken down, DNA can still be analyzed. Ways in which DNA can be analyzed is through the microscope or other sophisticated DNA testing.

Forensic ornithology 
Forensic ornithology employs scientific methodologies to ascertain the bird species from traces of evidence such as a small feather, a fragment of bone, or a trace of blood. Bird remains can be identified, first and foremost from feathers as feathers exhibit distinctive characteristics specific to a particular bird species, which can be observed both at a macroscopic and microscopic level.

Forensic odontology 

Dentistry has helped law enforcement detect and solve cases in criminal and civil proceedings. Odontology did not get popular until the 1960s when an interest was brought up by the first instructional program in the United States at the Armed Forces Institute of Pathology. As a result of this program, the term "forensic odontology" is now widely known and understood by both dental and law enforcement professionals.

Odontologists or dental professionals can use dental science to establish a person's identity. The broad duties of a Forensic Odontologist include forensic identification of individuals, including in multi-casualty events, age estimation in living and deceased persons, interpreting injuries in the oral and perioral regions, bite mark analysis and comparison, and assisting forensic pathologists in determining the cause of death if there is a possible contributing dental condition.

Dental evidence is useful in establishing human identity by comparing the dental features of a deceased person with antemortem dental records.

Forensic pathology 
Forensic pathology is a subfield in forensics where experts examine persons (bodies) who die suddenly, unexpectedly or violently, and thus determine the cause of death. The function of a forensic autopsy is to provide information through a postmortem examination of the body and analysis of the fluids to determine the cause of death, manner of death, and mechanism of injury.

A forensic pathologist is a medical doctor who is an expert in both trauma and disease and is responsible for performing autopsies. He/she applies their extensive knowledge of the human body and possible internal and external inflections as he/she performs an autopsy, to ascertain the manner and cause of death hopefully.

Information derived from the autopsy often greatly assists investigative efforts and scene reconstruction.

Forensic toxicology 

Forensic toxicology is the use of toxicology and other disciplines such as analytical chemistry, pharmacology and clinical chemistry to aid medical or legal investigation of death, poisoning, and drug use. The primary concern for forensic toxicology is not the legal outcome of the toxicological investigation or the technology utilized but rather the obtainment and interpretation of results.

Forensic microbiology 
With recent advances in massive parallel sequencing (MPS), or next-generation sequencing (NGS), forensic microbiology has become an increasingly promising area of research. "Initial applications in circumstances of biocrime, bioterrorism and epidemiology use microorganisms for the following reasons: (i) as ancillary evidence in criminal cases; (ii) to clarify causes of death (e.g., drownings, toxicology, hospital-acquired infections, sudden infant death and shaken baby syndromes); (iii) to assist human identification (skin, hair and body fluid microbiomes); (iv) for geolocation (soil microbiome or microbiome of bodies of water); and (v) to estimate postmortem interval (PMI) via thanatomicrobiome and epinecrotic microbial community". In fact, scientists estimate the time elapsed since death or the PMI by analyzing the stage of decay from bacterial decomposition or the bacterial succession patterns.

Bioterrorism and epidemiology 
Bio terrorism occurs when biological components are used as warfare agents. Such biological weapons are microorganisms that  are deliberately dispersed to incapacitate or cause disease and death to humans, animals or plants. Biological warfare agents can be natural or genetically modified. Nevertheless, these biological agents - be it viruses, bacteria or fungi are highly contagious.

Forensic microbiology also contributes to the development of epidemiology. By examining microorganisms obtained from infected individuals, scientists can determine a suspected source of infection, the type of infection present, and the evolution or mutation pattern of the microorganism. The application of a forensic microbiologist would be to examine the microorganisms isolated from infected individuals and compare it to known sources of infectious pathogens.

"It is important to remember that biological agents that can be used as weapons are often found in the environment. For this reason, it is always difficult to determine whether infections associated with these bioagents are accidental or purposely started". While not the first, or only, incidence of bioterrorism, perhaps the most notable case in recent memory involved the sending of at least four anthrax-containing envelopes in the United States in September and October 2001. "At least 22 victims contracted anthrax due to the mailings: 11 individuals contracted inhalation anthrax, with 5 of these infections resulting in fatalities; another 11 individuals suffered cutaneous anthrax. In addition, 31 persons tested positive for exposure to B. anthracis spores". However, the advancements in PCR and whole-genome sequencing allowed scientists to collaborate with the FBI and identify the source of the letter spores.

Differentiating a biowarfare attack from a normal epidemiology outbreak 
The basic epidemiologic approach to evaluating a potential bioterrorist or biowarfare attack differs from any standard epidemiologic investigation. The first step uses laboratory and clinical findings to confirm that a disease outbreak has occurred. Once the case definition and attack rate have been determined, the outbreak can be characterized in the conventional context of time, place, and person. These data provide crucial information in determining the potential source of the outbreak. An epidemic curve can be calculated using data gathered on cases over time. The disease pattern is important to differentiate between a natural outbreak and an intentional attack. A bioterrorism attack is most likely caused by a point source, with everyone coming in contact with the agent at approximately the same time. Additional characteristics investigated in determining whether it is the result of a biological attack are: the presence of a large epidemic; more severe disease than expected for a given pathogen; a disease that is unusual for a given geographic area; and multiple simultaneous epidemics of different diseases.

Postmortem analysis 
"Post-mortem microbiology (PMM) aims to detect unexpected infections causing sudden deaths; confirm clinically suspected but unproven infection; evaluate the efficacy of antimicrobial therapy; identify emergent pathogens; and recognize medical errors. Additionally, the analysis of the thanatomicrobiome may help to estimate the post-mortem interval.". There is currently an extensive amount of research being performed, most notably using the famous "body farms" throughout the United States, to determine if there is a consistent microbial decomposition "clock" that could be used by itself, or in conjunction with other techniques (such as forensic entomology) to help estimate postmortem intervals. One such group has made extensive headway into describing such a microbial clock, and "believes she's within two to five years of testing her clock in a real crime scene scenario". However, if a reliable and consistent microbial clock is determined to exist, "it's too soon to know whether the microbial clock will pass scientific and legal muster," (Beans) and "a judge would also have to determine that the microbial clock meets the standard for admission of expert testimony".

Water sample analysis 
In cases involving a body of water at or near the scene of a crime, a sample of the water can be extracted and analyzed under a light microscope for microorganisms. One such microorganism that are analyzed within samples of fresh water are diatoms, microscopic algae of varying shapes. Different bodies of water have been found to contain unique sets of diatoms and therefore, a piece of evidence found in a specific body of water will contain unique diatoms on it found only in that specific body of water. Therefore, the diatoms on a questioned object or body can be compared to the diatoms from a body of water to determine whether it had been present in the water.

Current issues

Sexual assault kit backlog 
Prior to DNA testing, many sexual assault cases could only rely on "he said, she said" and possible witnesses. Even once DNA analysis was available, many sexual assault kits, or SAKs, were never tested and thrown into a backroom or storage facility, only to be forgotten until discovered. Since DNA analysis is frequently utilized in most cases, most SAKs are examined and analyzed. However, the issue remains about the preexisting SAKs that have never been tested. A prevalent issue then, that still extends to now, is the absence of funds to process and analyze these SAKs. Many districts would dedicate their funds to homicides or more high-profile cases and sexual assaults would be swept aside. The biggest concern about all of these SAKs, is how to go about processing all of them, especially as more and more are being found each year.

Cold cases 
With the considerable advances in DNA analysis, old, open cases with intact evidence can be examined for biological evidence. New profiles are uploaded to CODIS every day so the base population to search and compare to increases. Biological testing for cold cases, specifically homicides, encounters similar roadblocks as the SAKs - lack of funds or the DNA samples have not been properly stored; thus too much degradation has occurred for viable analyses.

Popular culture

In popular culture, forensic biology is frequently portrayed in shows like Law & Order, Hannibal, Bones, CSI, Dexter and Castle. However thanks to Hollywood's depiction of forensic science, the analysis of biological evidence has fallen prey to the CSI Effect, which results in the public's perception of its capabilities being severely distorted and its limits blurred.

See also
 DNA profiling
 Forensic chemistry 
 Forensic science

References

Branches of biology
Biology