Editing Forensic entomology (section)

==Modern techniques==
Many new techniques have been developed<ref>Villet MH, Amendt J, 2011. Advances in entomological methods for estimating time of death. In: Turk EE, ed. Forensic Pathology Reviews. Heidelberg: Humana Press, pp. 213-238</ref> and are used in order to more accurately [[entomological evidence collection|gather evidence]], or reevaluate old information. The use of these newly developed techniques and evaluations have become relevant in litigation and appeals. Forensic entomology not only uses arthropod biology, but it pulls from other sciences, introducing fields like chemistry and genetics, exploiting their inherent synergy through the [[use of DNA in forensic entomology]]. In order to improve the precision and dependability of insect-based evidence analysis, forensic entomologists nowadays use a variety of cutting-edge technologies, such as stable isotope analysis and DNA analysis. These methods have broadened the field of forensic entomology by making it possible to identify insect species more precisely, pinpoint their geographic origins, and draw important conclusions about the circumstances surrounding a death. Some of the most important contemporary forensic entomology techniques are examined in this section along with how they are applied to criminal investigations.

===Scanning electron microscopy===
Fly larvae and fly eggs are used to aid in the determination of a PMI. In order for the data to be useful the larvae and eggs must be identified down to a species level to get an accurate estimate for the PMI. There are many techniques currently being developed to differentiate between the various species of forensically important insects. A study in 2007 demonstrates a technique that can use [[Scanning electron microscope|scanning electron microscopy]] (SEM) to identify key morphological features of eggs and maggots.<ref name="pmid18353656">{{cite journal | vauthors = Mendonça PM, dos Santos-Mallet JR, de Mello RP, Gomes L, de Carvalho Queiroz MM | title = Identification of fly eggs using scanning electron microscopy for forensic investigations | journal = Micron | volume = 39 | issue = 7 | pages = 802–807 | date = October 2008 | pmid = 18353656 | doi = 10.1016/j.micron.2008.01.014 }}</ref> Some of the morphological differences that can help identify the different species are the presence/absence of anastomosis, the presence/absence of anterior and posterior spiracles,<ref>{{cite journal | vauthors = Ahmad Firdaus MS, Marwi MA, Syamsa RA, Zuha RM, Ikhwan Z, Omar B | title = Morphological descriptions of second and third instar larvae of Hypopygiopsis violacea Macquart (Diptera:Calliphoridae), a forensically important fly in Malaysia | journal = Tropical Biomedicine | volume = 27 | issue = 1 | pages = 134–137 | date = April 2010 | pmid = 20562824 | url = https://pubmed.ncbi.nlm.nih.gov/20562824/ }}</ref> the cephalopharyngeal skeleton as well as the shape and length of the median area.

The SEM method provides an array of morphological features for use in identifying fly eggs; however, this method does have some disadvantages. The main disadvantage is that it requires expensive equipment and can take time to identify the species from which the egg originated, so it may not be useful in a field study or to quickly identify a particular egg.<ref name=Sukontason>{{cite journal | vauthors = Sukontason K, Sukontason KL, Piangjai S, Boonchu N, Kurahashi H, Hope M, Olson JK | title = Identification of forensically important fly eggs using a potassium permanganate staining technique | journal = Micron | volume = 35 | issue = 5 | pages = 391–395 | year = 2004 | pmid = 15006363 | doi = 10.1016/j.micron.2003.12.004 }}</ref>
The SEM method is effective provided there is ample time and the proper equipment and the particular fly eggs are plentiful. The ability to use these morphological differences gives forensic entomologists a powerful tool that can help with estimating a post mortem interval, along with other relevant information, such as whether the body has been disturbed post mortem.

===Potassium permanganate staining===
When scanning electron microscopy is not available, a faster, lower cost technique is [[potassium permanganate]] staining. The collected eggs are rinsed with a [[Saline (medicine)|normal saline]] solution and placed in a glass petri dish. The eggs are soaked in a 1% potassium permanganate solution for one minute and then dehydrated and mounted onto a slide for observation.<ref name=Sukontason/> These slides can be used with any [[Optical microscope|light microscope]] with a calibrated eyepiece to compare various morphological features. The most important and useful features for identifying eggs are the size, length, and width of the plastron, as well as the morphology of the plastron in the area around the micropyle.<ref name=Sukontason/> The various measurements and observations when compared to standards for forensically important species are used to determine the species of the egg.

===Mitochondrial DNA===
In 2001, a method was devised by Jeffrey Wells and Felix Sperling to use [[mitochondrial DNA]] to differentiate between different species of the subfamily Chrysomyinae. This is particularly useful when working to determine the identity of specimens that do not have distinctive morphological characteristics at certain life stages.<ref name = "Wells_2001" />

===Mock crime scenes===
A valuable tool that is becoming very common in the training of forensic entomologists is the use of mock crime scenes using pig carcasses. The pig carcass represents a human body and can be used to illustrate various environmental effects on both arthropod succession and the estimate of the post mortem interval.<ref>{{cite journal | vauthors = Schoenly KG, Haskell NH, Mills DK, Bieme-Ndi C, Larsen K, Lee Y | title = Recreating death's acre in the school yard: using pig carcasses as model corpses to teach concepts of forensic entomology & ecological succession. | journal = The American Biology Teacher | date = September 2006 | volume = 68 | issue = 7 | pages = 402–410 | url = http://www.nabt.org/sites/S1/index.php?p=2 | doi = 10.1662/0002-7685(2006)68[402:RDAITS]2.0.CO;2 | access-date = 1 April 2008 | archive-date = 2 July 2022 | archive-url = https://web.archive.org/web/20220702194633/https://nabt.org/sites/S1/index.php?p=2 | url-status = dead }}</ref> Pigs are the most frequently utilised model in an attempt to gather data regarding forensic experimental analysis. The latter is highly proportionate to human nature due to our overlapping characteristics with the mentioned species. These interrelated components include: subcutaneous fat stores, skin thickness, range of adult body mass, hair covering, and omnivorous diets.<ref>{{cite book | veditors = Pokines JT, Symes SA |title=Manual of Forensic Taphonomy |publisher=CRC Press |year=2013 | isbn = 978-1-4398-7843-9 }}</ref>

===Gene expression studies===
Although physical characteristics and sizes at various [[instars]] have been used to estimate fly age, a more recent study has been conducted to determine the age of an egg based on the expression of particular genes. This is particularly useful in determining developmental stages that are not evidenced by change in size; such as the egg or pupa and where only a general time interval can be estimated based on the duration of the particular developmental stage. This is done by breaking the stages down into smaller units separated by predictable changed in [[gene expression]].<ref name=Tarone>{{cite journal | vauthors = Tarone AM, Jennings KC, Foran DR | title = Aging blow fly eggs using gene expression: a feasibility study | journal = Journal of Forensic Sciences | volume = 52 | issue = 6 | pages = 1350–1354 | date = November 2007 | pmid = 18093065 | doi = 10.1111/j.1556-4029.2007.00587.x | s2cid = 32533922 | citeseerx = 10.1.1.497.3287 }}</ref> Three genes were measured in an experiment with ''Drosophila melanogaster'': bicoid (bcd), slalom (sll), and chitin synthase (cs). These three genes were used because they are likely to be in varied levels during different times of the egg development process. These genes all share a linear relationship in regards to age of the egg; that is, the older the egg is the more of the particular gene is expressed.<ref name=Tarone/> However, all of the genes are expressed in varying amounts. Different genes on different loci would need to be selected for another fly species. The genes expressions are mapped in a control sample to formulate a developmental chart of the gene expression at certain time intervals. This chart can then be compared to the measured values of gene expression to accurately predict the age of an egg to within two hours with a high [[Confidence interval|confidence level]].<ref name=Tarone/> Even though this technique can be used to estimate the age of an egg, the feasibility and legal acceptance of this must be considered for it to be a widely utilized forensic technique.<ref name=Tarone/> One benefit of this would be that it is like other DNA-based techniques so most labs would be equipped to conduct similar experiments without requiring new capital investment. This style of age determination is in the process of being used to more accurately find the age of the instars and pupa; however, it is much more complicated, as there are more genes being expressed during these stages.<ref name=Tarone/> The hope is that with this and other similar techniques a more accurate PMI can be obtained.

=== DNA analysis ===
Modern forensic entomology now relies heavily on DNA analysis as a fundamental tool for accurately identifying insect species and gaining important insights into their interactions with human remains. Using this method, DNA is extracted from insect specimens discovered at crime scenes and compared to databases containing known DNA sequences. Forensic entomologists can verify species identification, identify the existence of particular species linked to decomposition, and even establish a connection between insects and particular geographical areas through the examination of insect DNA.<ref>{{cite journal | vauthors = Wells JD, Stevens JR | title = Application of DNA-based methods in forensic entomology | journal = Annual Review of Entomology | volume = 53 | issue = 1 | pages = 103–120 | date = 2008 | pmid = 17685848 | doi = 10.1146/annurev.ento.52.110405.091423 }}</ref>

The identification of insect stomach contents is one of the main uses of DNA analysis in forensic entomology. Investigators can ascertain the insect's most recent meal by sequencing the DNA contained in the gut of maggots or other insect larvae discovered on a body. When determining the postmortem interval (PMI) or locating possible sources of contamination or infection, this information might be extremely important.<ref name="Amendt_2007" />

The study of insect dispersal patterns and colonization behaviour has also been transformed by DNA research. Researchers can deduce patterns of travel and colonization by examining the genetic variety of insect populations. This can provide important insights into the origin of insect specimens found at crime scenes or the transportation of a body.<ref name = "Wells_2001">{{cite journal | vauthors = Wells JD, Sperling FA | title = DNA-based identification of forensically important Chrysomyinae (Diptera: Calliphoridae) | journal = Forensic Science International | volume = 120 | issue = 1–2 | pages = 110–115 | date = August 2001 | pmid = 11457617 | doi = 10.1016/S0379-0738(01)00414-5 }}</ref>

In general, DNA analysis has improved forensic entomology's accuracy and dependability significantly, allowing investigators to obtain previously unobtainable specific information from insect evidence.

=== Stable isotope analysis ===
A contemporary method that is being used more and more in forensic entomology to shed light on the ecology and life cycle of insects connected to human remains is stable isotope analysis. Using this technique, the stable isotopic composition of elements found in insect tissues, including carbon, nitrogen, hydrogen, and oxygen, is measured. The environment in which the insect developed, including its diet and place of origin, is reflected in these isotopes.<ref>{{cite journal | vauthors = Farmer NL, Meier-Augenstein W, Kalin RM | title = Stable isotope analysis of safety matches using isotope ratio mass spectrometry--a forensic case study | journal = Rapid Communications in Mass Spectrometry | volume = 19 | issue = 22 | pages = 3182–3186 | date = 2005 | pmid = 16220465 | doi = 10.1002/rcm.2088 | bibcode = 2005RCMS...19.3182F }}</ref>

Stable isotope analysis provides information on the travel of the body or the insect itself, and can be used in forensic investigations to assist identify the geographic origin of insects found on a body. Furthermore, features of the insect's diet, such as whether it consumed organic matter or human remains, can be inferred by stable isotope analysis. This information can be vital to comprehending the insect's function in the decomposition process.<ref>{{cite journal | vauthors = Wang LM, Wu H, Lin GH |date=2015 |title=Technology to Determining Geographical Origins of Traditional Chinese Medicines  |journal=Tong Wei Su |volume=28 |issue=4 |pages=225–232|doi=10.7538/tws.2015.28.04.0225 }}</ref>

When using traditional methods to identify insect species or estimate age, this methodology has proven quite helpful. Stable isotope analysis contributes to more accurate and thorough forensic studies by improving the precision and depth of forensic entomological investigations by offering a distinct chemical signature that represents the insect's surroundings.