Use of DNA in forensic entomology

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Forensic entomology has three sub-fields: urban, stored product and medico-criminal entomologies. This article focuses on medico-criminal entomology and how DNA is analyzed with various blood-feeding insects.

Contents

Forensic entomology can be an important aspect for law enforcement. With the magnitude of information that can be gathered, investigators can more accurately determine time of death, location, how long a body has been in a specific area, if it has been moved, and other important factors.

Blood meal extraction

To extract a blood meal from the abdomen of an insect to isolate and analyze DNA, the insect must first be killed by placing it in 96% ethanol. The killed insect can be stored at -20 °C until analysis. When it is time for analysis, the DNA must then be extracted by dissecting the posterior end of the abdomen and collecting 25 mg of tissue. The cut in the abdomen should be made with a razor blade as close to the posterior as possible to avoid the stomach. [1] Using a DNA extraction kit, the DNA is extracted from the tissue. If the DNA is mixed with samples from more than one individual, it is separated using a species specific primer. Once extracted and isolated, the DNA sample goes through a polymerase chain reaction (PCR), is amplified and identified.

PCR works by analyzing species specific mitochondrial DNA. PCR is currently the most commonly used method of species identification. This results from the fact that it is very sensitive in that it requires only a small amount of biological material, and can also utilize material that is not particularly fresh. The sample can be frozen and stored while still remaining usable for later PCR.

DNA requires one hour to reach the abdomen of an insect, so DNA can be amplified one to forty-four hours after an insect feeds. Some research suggests that the source of a blood meal can be determined up to two months post feeding.

To amplify DNA, it must first be denatured by exposing it to a 95 °C temperature for one minute, followed by thirty cycles of thirty-second 95 °C exposures. Then denatured DNA is mixed with a specific primer. A chromatograph is conducted on 2% agarose gel, stained, and viewed with UV fluorescence. The DNA is identified by looking for genome specific repetitive elements and by comparing it with known examples.

Haematophagous insects of forensic importance

Humans are constantly fed on by haematophagous (blood feeding) insects. The ingested blood can be recovered and used to identify the person from which it was taken. Bite marks and reactions to bites can be used to place a person in an area where those insects are found.

Order Diptera

The following among the flies (Diptera) have been utilized: [2]

Order Siphonaptera

Listed here are fleas commonly encountered by humans that could potentially be used for DNA identification.

Order Hemiptera

Cimex lectularius is an obligate parasite of humans. Testing a sample of a residence's bed bug population and screening for bites could reveal possible recent visitors to the structure, as they have been observed to feed approximately once a week in temperate conditions. [8] A recent re-emergence of bedbug populations in North America as well as growing interest in the field of forensics may prove bedbugs to be useful investigative tools. [9] Recent studies have revealed that human DNA can be recovered from bed bugs for up to 60 days after feeding, thus demonstrating the potential use of this insect in forensic entomology [10] [11]

Order Phthiraptera

Lice can be indicators of contact with another person. Many species closely associated with humans can be easily transferred between individuals. DNA identification of multiple individuals using blood meals from body and head lice has been demonstrated in laboratory settings. [12]

Suborder Anoplura

Other Arthropods

Order Ixodida

Due to the low probability of a tick detaching and falling to the ground at the scene of the crime, these may not be highly useful regardless of the large amount of blood and lymph they ingest. However, should an engorged tick be found in an area of interest, it would likely contain sufficient genetic material for identification.

Analysis of collected DNA

DNA identification of species can be a useful tool in forensic entomology. [13] Although it does not replace conventional identification of species through visual identification, it can be used to differentiate between two species of very similar or identical physical and behavioral characteristics. [14] A thorough identification of the species through conventional methods is needed before an attempt at DNA analysis. This DNA can be obtained from practically any part of the insect, including the body, leg, setae, antennae, etc. There are about one million species described in the world and many more that have still not been identified. A project termed "the barcode of life" was launched by Dr. Paul D. N. Hebert, where he identified a gene that is used in cellular respiration by all species, but is different in every species. This difference in sequence can help entomologists easily identify two similar species.

DNA sequencing is basically done in three steps: polymerase chain reaction (PCR), followed by a sequencing reaction, then gel electrophoresis. PCR is a step that cleaves the long chain of chromosomes into much shorter and workable pieces. These pieces are used as patterns to create a set of fragments. These fragments are different in length from each other by one base which is helpful in identification. Those sets of fragments are then separated by gel electrophoresis. [15] This process uses electricity to separate DNA fragments by size as they move through a gel matrix. With the presence of an electric current the negative DNA strand marches toward the positive pole of the current. The smaller DNA fragments move through the gel pores much more easily/faster than larger molecules. At the bottom of the gel the fragments go through a laser beam that emits a distinct color according to the base that passes through.

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<span class="mw-page-title-main">Mosquito</span> Family of flies

Mosquitoes are a family, the Culicidae, of some 3,600 species of small flies. The word "mosquito" is Spanish for "little fly". Mosquitoes have a slender segmented body, one pair of wings, three pairs of long hair-like legs, and specialized, highly elongated, piercing-sucking mouthparts. All mosquitoes drink nectar from flowers; females of some species have in addition adapted to drink blood. Evolutionary biologists view mosquitoes as micropredators, small animals that parasitise larger ones by drinking their blood without immediately killing them. Medical parasitologists view mosquitoes instead as vectors of disease, carrying protozoan parasites or bacterial or viral pathogens from one host to another.

<i>Anopheles</i> Genus of mosquito

Anopheles is a genus of mosquito first described by J. W. Meigen in 1818. Its members are sometimes called nail mosquitoes or marsh mosquitoes. Many are vectors of the parasite Plasmodium, which causes malaria in birds, reptiles, and mammals including humans. Anopheles gambiae is the best-known species, as it transmits one of the most dangerous human malarial parasites, Plasmodium falciparum. No other mosquito genus is a vector of human malaria.

<span class="mw-page-title-main">Pyrethroid</span> Class of insecticides

A pyrethroid is an organic compound similar to the natural pyrethrins, which are produced by the flowers of pyrethrums. Pyrethroids are used as commercial and household insecticides.

<span class="mw-page-title-main">Bed bug</span> Type of insect that feeds on human blood

Bed bugs are parasitic insects from the genus Cimex, who are micropredators that feed on blood, usually at night. Their bites can result in a number of health impacts including skin rashes, psychological effects, and allergic symptoms. Bed bug bites may lead to skin changes ranging from small areas of redness to prominent blisters. Symptoms may take between minutes to days to appear and itchiness is generally present. Some individuals may feel tired or have a fever. Typically, uncovered areas of the body are affected. Their bites are not known to transmit any infectious disease. Complications may rarely include areas of dead skin or vasculitis.

<span class="mw-page-title-main">Cimicidae</span> Family of parasitic blood-feeding insects

The Cimicidae are a family of small parasitic bugs that feed exclusively on the blood of warm-blooded animals. They are called cimicids or, loosely, bed bugs, though the latter term properly refers to the most well-known member of the family, Cimex lectularius, the common bed bug and its tropical relation Cimex hemipterus. The family contains over 100 species. Cimicids appeared in the fossil record in the Cretaceous period. When bats evolved in the Eocene, Cimicids switched hosts and now feed mainly on bats or birds. Members of the group have colonised humans on three occasions.

<i>Cimex lectularius</i> Species of true bug

Cimex lectularius is a species of Cimicidae. Its primary hosts are humans, and it is one of the world's major "nuisance pests".

<span class="mw-page-title-main">Black fly</span> Family of insects

A black fly or blackfly is any member of the family Simuliidae of the Culicomorpha infraorder. It is related to the Ceratopogonidae, Chironomidae, and Thaumaleidae. Over 2,200 species of black flies have been formally named, of which 15 are extinct. They are divided into two subfamilies: Parasimuliinae contains only one genus and four species; Simuliinae contains all the rest. Over 1,800 of the species belong to the genus Simulium.

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<span class="mw-page-title-main">Culicinae</span> Subfamily of flies

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<i>Cimex</i> Genus of true bugs

Cimex is a genus of insects in the family Cimicidae. Cimex species are ectoparasites that typically feed on the blood of birds and mammals. Two species, Cimex lectularius and Cimex hemipterus, are known as bed bugs and frequently feed on humans, although other species may parasitize humans opportunistically. Species that primarily parasitize bats are known as bat bugs.

<i>Aedes vexans</i> Species of fly

Aedes vexans, the inland floodwater mosquito or tomguito, is a cosmopolitan and common pest mosquito. This species has been collected in southern California.

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<i>Cimex hemipterus</i> Species of true bug

Cimex hemipterus, known as the tropical bed bug, is a species of bed bugs within the family Cimicidae that primarily resides in tropical climates. However, it has been reported that this species can live in more temperate climates along with the closely related bed bug species C. lectularius.C. hemipterus is a hematophagous, obligate parasite of humans. This means that it requires blood meals from their human hosts in order to survive. When bitten, humans experience itchiness, wheals, and lesions around the affected areas on the skin. This species typically resides in human domiciles within cracks, crevices, or mattresses, and are more prevalent in developing countries. Like other bed bugs, C. hemipterus is primarily active during the night time.

<i>Aedes taeniorhynchus</i> Species of fly

Aedes taeniorhynchus, or the black salt marsh mosquito, is a mosquito in the family Culicidae. It is a carrier for encephalitic viruses including Venezuelan equine encephalitis and can transmit Dirofilaria immitis. It resides in the Americas and is known to bite mammals, reptiles, and birds. Like other mosquitoes, Ae. taeniorhynchus adults survive on a combination diet of blood and sugar, with females generally requiring a blood meal before laying eggs.

References

  1. ↑ Pizarro, Juan et al. "A method for the identification of guinea pig blood meal in the Changas disease vector, Triatoma infestans" in Kinrtoplastid Biol Dis., V. 6, 2007.
  2. ↑ Gibson G, Torr S.J. (1999) Visual and Olfactory Responses of Haematophagous Diptera to Host Stimuli. Medical and Veterinary Entomology: Volume 13:pp 2-23
  3. ↑ S. Spitaleri, C. Romano, E. Di Luise, E. Ginestra, L. Saravo, Genotyping of human DNA recovered from mosquitoes found on a crime scene, Int. Congr. Ser. 1288 (2006) 574–576.
  4. ↑ Curic, Goran; Hercog, Rajna; Vrselja, Zvonimir; Wagner, Jasenka (2014). "Identification of person and quantification of human DNA recovered from mosquitoes (Culicidae)". Forensic Science International: Genetics. 8 (1): 109–112. doi:10.1016/j.fsigen.2013.07.011. PMID   24315597.
  5. ↑ Chow-Shaffer, E; Sina, B; Hawley, WA; De Benedictis, J; Scott, TW (2000). "Laboratory and Field Evaluation of Polymerase Chain Reaction-Based Forensic DNA Profiling for Use in Identification of Human Blood Meal Sources of Aedes aegypti (Diptera: Culicidae)". Journal of Medical Entomology. 37 (4): 492–502. doi: 10.1603/0022-2585-37.4.492 . PMID   10916289. S2CID   11153465.
  6. ↑ Curic, Goran; Hercog, Rajna; Vrselja, Zvonimir; Wagner, Jasenka (2014). "Identification of person and quantification of human DNA recovered from mosquitoes (Culicidae)". Forensic Science International: Genetics. 8 (1): 109–112. doi:10.1016/j.fsigen.2013.07.011. PMID   24315597.
  7. ↑ Spitaleri, S.; Romano, C.; Luise, E. Di; Ginestra, E.; Saravo, L. (2006). "Genotyping of human DNA recovered from mosquitoes found on a crime scene". Int. Congr. Ser. 1288: 574–576. doi:10.1016/j.ics.2005.11.055.
  8. ↑ Reinhardt K, Siva-Jothy M (2007) Biology of the Bed Bugs (Cimicidae). Annu. Rev. Entomol.: Volume 52, pp. 351–74
  9. ↑ "Bed Bug Information (Identification, Biology, and Control)" Harvard University, Environmental Health and Safety 2005. Accessed March 19, 2008
  10. ↑ Szalanski, A.L; Austin, J.W.; McKern, J.A.; McCoy, T.; Steelman, C.D.; Miller, D.M. (2006). "Time course analysis of bed bug, "Cimex lectularius" L., (Hemiptera: Cimicidae) blood meals using PCR". Journal of Agricultural and Urban Entomology. 23: 237–241.
  11. ↑ Szalanski, A.L.; Austin, J.W.; McKern, J.A.; Steelman, C.D.; Miller, D.M.; Gold, R.E. (2006). "Isolation and characterization of human DNA from bed bug, "Cimex lectularius" L., (Hemiptera: Cimicidae) blood meals". Journal of Agricultural and Urban Entomology. 23: 189–194.
  12. ↑ Mumcuoglu, KY; Gallili, N; Reshef, A; Brauner, P; Grant, H (2004). "Use of Human Lice in Forensic Entomology". Journal of Medical Entomology. 41 (4): 803–806. doi: 10.1603/0022-2585-41.4.803 . PMID   15311479.
  13. ↑ Wells, J.D.; Stevens, J.R. (2008). "Application of DNA-Based Methods in Forensic Entomology". Annual Review of Entomology. 53: 103–120. doi:10.1146/annurev.ento.52.110405.091423. PMID   17685848.
  14. ↑ Mansfield, Betty. "DNA forensics." Human Genome Project Information. 21 Feb 2008. 2 March 2008.
  15. ↑ Klug, William, and Michael Cummings. Essential of Genetics. 6th ed. Upper Saddle River, NJ: Prentice Hall, 2007.
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