DNA Barcoding in Forensic Entomology: A Molecular Tool That Increases Reliability in Estimating the Postmortem Interval

Mahmut Kabalak; Yavuz Turan

doi:10.35229/jaes.1840921

EN TR

DNA Barcoding in Forensic Entomology: A Molecular Tool That Increases Reliability in Estimating the Postmortem Interval

Abstract

Forensic entomology has become an indispensable tool for estimating the postmortem interval (PMI), particularly in advanced stages of decomposition where classical forensic methods are limited. Reliable PMI estimation requires accurate and precise identification of insect evidence. Traditional morphological approaches often fail when dealing with immature stages (eggs, larvae, pupae), closely related species, or partially damaged specimens. To overcome these limitations, DNA barcoding has been increasingly integrated into forensic entomology over the past two decades. DNA barcoding typically relies on sequencing a standardized fragment of the mitochondrial cytochrome c oxidase I (COI) gene and comparing the obtained sequence with reference databases such as BOLD and GenBank to assign species identity. In this review, we summarize the theoretical basis of DNA barcoding in forensic entomology, the genetic markers employed, and the main steps of sampling, DNA extraction, PCR amplification, and sequence analysis. We present application examples from Diptera and Coleoptera, and discuss how DNA barcoding contributes to PMI estimation and case reconstruction. Furthermore, we address recent developments such as mini-barcode strategies, Next Generation Sequencing (NGS)/metabarcoding approaches, multi-locus barcoding, and highlight key limitations including database gaps, the barcode gap concept, contamination, and NUMTs. We also evaluate the current situation in Türkiye and propose the establishment of a national barcode reference library for forensically relevant insect taxa. Overall, DNA barcoding emerges as a powerful molecular tool that complements rather than replaces morphology, thereby improving the reliability of species identification and PMI estimations in forensic casework.

Keywords

Forensic entomology , DNA barcoding , COI , postmortem interval , molecular species identification

Adli Entomolojide DNA Barkodlama: Ölüm Sonrası Zaman Aralığı Tahmininde Güvenirliği Artıran Bir Moleküler Araç

Öz

Adli entomoloji, özellikle yumuşak dokuların hızla bozulduğu ve klasik adli tıp yöntemlerinin yetersiz kaldığı durumlarda ölüm sonrası zaman aralığı (ÖSZA) tahmininde vazgeçilmez bir araç haline gelmiştir. Ancak böcek gelişim sürelerine dayalı güvenilir bir ÖSZA tahmini için, elde edilen entomolojik örneklerin türlerinin doğru ve kesin biçimde teşhis edilmesi gereklidir. Geleneksel morfolojik teşhis yöntemleri, özellikle yumurta, larva ve pupa gibi ergin olmayan dönemlerde, yakın akraba türlerde ve kısmen tahrip olmuş örneklerde çoğu zaman yetersiz kalmaktadır. Bu sınırlılığı aşmak üzere son 20 yılda DNA barkodlama yaklaşımı adli entomolojiye yaygın biçimde entegre olmuştur. DNA barkodlama, çoğunlukla mitokondriyal sitokrom c oksidaz I (COI) geninin standart bir bölgesinin dizilenmesine dayanmakta; elde edilen dizi, BOLD ve GenBank gibi referans veri tabanlarındaki kayıtlarla karşılaştırılarak tür tanısı yapılmaktadır. Bu derlemede, adli entomolojide DNA barkodlamanın kuramsal temeli, kullanılan genetik belirteçler, örnekleme–DNA izolasyonu–PCR–dizi analizi basamakları, Diptera ve Coleoptera başta olmak üzere çeşitli taksonomik gruplar üzerindeki uygulama örnekleri ve ÖSZA tahminine katkıları ele alınmıştır. Ayrıca mini-barkod stratejileri, Yeni Nesil Sekanslama (YNS)/metabarkodlama yaklaşımları, çoklu marker kullanımı, veri tabanı sorunları, barkod boşluğu (barcode gap), kontaminasyon ve NUMT gibi teknik sınırlılıklar tartışılmış; Türkiye’de adli entomoloji bağlamında DNA barkodlamanın mevcut durumu ve ulusal barkod veri tabanı oluşturulmasına yönelik öneriler sunulmuştur. Sonuç olarak DNA barkodlama, morfolojik teşhisi ikame etmekten çok onu tamamlayan, tür tayini ve PMI tahminini daha güvenilir hale getiren güçlü bir moleküler araç olarak öne çıkmaktadır.

Anahtar Kelimeler

Adli entomoloji , DNA barkodlama , COI , ölüm sonrası zaman aralığı , moleküler tür tanısı

Kaynakça

Abu El-Hassan, G.M.M., Abo El-Ela, R.H., Sawaby, R., El- Hamouly, H., El Sawaf, B.M. & Ghallab, E.H. (2025). Morphology, molecular phylogeny and record verification of Sarcophaga ruficornis Fabricius (Diptera: Sarcophagidae) from Egypt. Scientific Reports, 15, 40357. https://doi.org/10.1038/s41598- 025-25934-0
Adnan, A., Mona, S., Rakha, A., Nazir, S., Wang, H., & Ren, F. (2025). Molecular diversity of three forensically relevant dipterans from cadavers in Lahore, Pakistan. Insects, 16, 381. https://doi.org/10.3390/insects16040381
Alam, M., Abbas, K., Raza, M.T., & Husain, A. (2024). Forensic entomology: A comprehensive review on insect-based approaches in criminal forensics. Munis Entomology & Zoology, 19(1), 132-145.
Altunsoy, F., Turan, Y., Fırat, S., & Sert, O. (2017). Differences in succession of Coleoptera species attracted to pig carcasses in rural and urban habitats in Eskişehir Province, Turkey. Turkish Journal of Entomology, 41(2), 177-195. https://doi.org/10.16970/ted.65825
Amendt, J., Campobasso, C.P., Gaudry, E., Reiter, C., LeBlanc, H.N., & Hall, M.J.R. (2007). Best practice in forensic entomology-Standards and guidelines. International Journal of Legal Medicine, 121(2), 90– 104. https://doi.org/10.1007/s00414-006-0086-x
Bana, R., & Beyarslan, A. (2012). Determination of Coleoptera Species of Pig Carcasses and Internal Organs of Bovine in Edirne City of Turkey. BEU Journal of Science 1(2), 122-126.
Chimeno, C., Morinière, J., Podhorna J., Hardulak L., Hausmann, A., Reckel, F., Grunwald, J.E., Randolph P., & Haszprunar, G. (2019). DNA Barcoding in Forensic Entomology - Establishing a DNA Reference Library of Potentially Forensic Relevant Arthropod Species. Journal of Forensic Sciences, 64(2), 593-601. http://doi.org/10.1111/1556- 4029.13869
Ceylan, A.Z. (2020). Adli önemi olan Diptera türlerinin mtDNA COI gen analizi ile genetik teşhisi. Yüksek Lisans Tezi, Ankara Üniversitesi Sağlık Bilimleri Enstitüsü, Ankara, Türkiye, 81s.
Dupuis J.R., Roe A.D., & Sperling F.A. (2012). Multi-locus species delimitation in closely related animals and fungi: one marker is not enough. Mol. Ecol., 21(18), 4422-36. http://doi.org/10.1111/j.1365- 294X.2012.05642.x
Eenhakkattu Mana, A.S., Jiang, S., Abaffy, T., Matsunami,barcoding on digital microfluidic biochips. Journal of Hardware and Systems Security, 9, 136-148. https://doi.org/10.1007/s41635-025-00167-9.

Grzywacz, A., Wyborska, D., & Piwczyński, M. (2017). DNA barcoding allows identification of European Fanniidae (Diptera) of forensic interest. Forensic Science International, 275, 95-102 http://doi.org/10.1016/j.forsciint.2017.06.023
Hajibabaei, M., Smith, M.A., Janzen, D.H., Rodriguez, J.J., Whitfield, J.B., & Hebert, P.D.N. (2006). A minimalist barcode can identify a specimen whose DNA is degraded. Molecular Ecology Notes, 6(4), 959- 964. https://doi.org/10.1111/j.1471-8286.2006.01470.x
Haymer, D. (2024). State of the art and new directions using DNA and other molecular markers in forensic entomology. Academia Biology, 2, 1–8 https://doi.org/10.20935/AcadBiol7450.
Hebert, P.D.N., Cywinska, A., Ball, S.L., & deWaard, J.R. (2003). Biological identifications through DNA barcodes. Proceedings of the Royal Society B: Biological Sciences, 270(1512), 313-321 https://doi.org/10.1098/rspb.2002.2218
Karataş, D., &Darılmaz, M.C. (2018). A preliminary study on aquatic forensic entomology in Turkey. J. For. Med., 32(1), 18-29. https://doi.org/10.5505/adlitip.2018.48091
Khullar, N., & Singh, D. (2025). Advances in molecular techniques in forensic entomology. In Tanda A.S. (Ed), Advances in Forensic Entomology: Basic and Applied Aspects, 51–71p, TechSar Publishing.
Liu, J., Jiang, J., Song, S., Tornabene, L., Chabarria, R., Gavin, J.P.N. & Li, C. (2017). Multilocus DNA barcoding - Species Identification with Multilocus Data. Scientific Reports, 7, 16601. https://doi.org/10.1038/s41598-017-16920-2
López-Rubio, A., Amat, E., Pérez, A., Gómez, G., & Gómez, L.M. (2017). Assessing a DNA mini-barcode strategy for species identification in neotropical necrophagous blow flies (Diptera: Calliphoridae) of forensic importance. Genome 60(11), 965.
Lord, W.D., & Stevenson, JR. (1986). Directory of forensic entomologists. 2nd ed. Washington, DC: Defense Pest Management Information Analysis Center, Walter Reed Army Medical Center.
Meyer, C.P., & Paulay, G. (2005). DNA barcoding: Error rates based on comprehensive sampling. PLoS Biology, 3(12), e422 https://doi.org/10.1371/journal.pbio.0030422
Mutlu, E.Ç., Yılmaz, M.C., Balık, R., Onay, A.S., & Turan, Y. (2024). Determination of insect fauna and succession important for forensic entomology in İstanbul Province. Transactions of the American Entomological Society, 216-230. https://doi.org/10.3157/061.150.0202.
Özdemir, S., & Sert, O. (2009). Determination of Coleoptera fauna on carcasses in Ankara province, Turkey. Forensic Science International, 183, 24-32. https://doi.org/10.1016/j.forsciint.2008.09.018.
Park, S.H., Kim, H.J., Jang, H., Park, J., Kim, Y.H., & Ko, K.S. (2018). Molecular identification of forensically important flies (Diptera: Calliphoridae, Sarcophagidae) using COI and ITS2 gene regions. Journal of Asia- Pacific Entomology, 21, 1-10 https://doi.org/10.1016/j.forsciint.2017.12.017
Sagar, A., Singh, T.K., Harsh, N., Jha, N.K., Dode, P.S. & Podder, A. (2025). DNA barcoding: From taxonomy to technology - A review. Journal of Carcinogenesis, 24(7s), 835-842 https://doi.org/10.64149/J.Carcinog.24.7s.835-843
Schilthuizen, M., Scholte, C., van Wijk, R.E.J., Dommershuijzen, J., van der Horst, D., zu Schlochtern, M.M., Lievers, R., & Groenenberg, D.S.J. (2011). Using DNA-barcoding to make the necrobiont beetle family Cholevidae accessible for forensic entomology. Forensic Science International 210, 91-95 https://doi.org/10.1016/j.forsciint.2011.02.003
Scieuzo, C., Rinaldi, R., De Stefano, F., Di Fazio, A., & Falabella, P. (2025). The contribution of molecular biology to forensic entomology. Insects, 16, 694. https://doi.org/10.3390/insects16070694
Sert, O., Kabalak, M., & Şabanoğlu, B. (2012). Determination of forensically important Coleoptera and Calliphoridae (Diptera) species on decomposing dog (Canis lupus familiaris L.) carcass at Ankara province. Hacettepe Journal of Biology and Chemistry, 40(1), 99-103.
Sharawi, S.E. (2025). Forensic entomology in Jeddah: Molecular identification of emerging insect strains. Indian Journal of Animal Research, 59(1), 1-4. https://doi.org/10.18805/IJAR.BF-1881
Şabanoğlu, B., & Sert, O. (2010). Determination of Calliphoridae (Diptera) fauna and seasonal distribution on carrion in Ankara province. Journal of Forensic Sciences, 55(4), 1003-1007. https://doi.org/10.1111/j.1556-4029.2010.01366.x
Schilthuizen, M., Scholte, C., van Wijk, R.E.J., Dommershuijzen, J., van der Horst, D., Meijer zu Schlochtern, M., Lievers, R. & Groenenberg, D. S. J. (2011). Using DNA-barcoding to make the necrobiont beetle family Cholevidae accessible for forensic entomology. Forensic Science International, 210, 91-95. https://doi.org/10.1016/j.forsciint.2011.02.003
Sharma, S., & Dalal, J. (2025). in Tanda, A.S. (Ed.). Advances in forensic entomology: Basic and applied aspects. TechSar Publishing.
Taberlet, P., Coissac, E., Hajibabaei, M., & Rieseberg, L.H. (2012). Environmental DNA. Molecular Ecology, 21(8), 1789–1793. https://doi.org/10.1111/j.1365- 294X.2012.05542.x
Wang, B., Ha, S., Cai, J., Ma, Y., Li, D., Chen, J., & Deng, J. (2025). Exploratory study on DNA barcode combined with PCR-HRM technology for rapid and accurate identification of necrophilous fly species. Insects, 16, 590. https://doi.org/10.3390/insects16060590
Wells, J.D., & Stevens, J.R. (2008). Application of DNA-based methods in forensic entomology. Annual Review of Entomology, 53, 103-120.
Yavasoglu, S.İ., Yılmaz, C., Ulger, C., & Simsek, F.M. (2016). Molecular identification and genetic structure of Aedes phoeniciae (Diptera: Culicidae) in Northern Cyprus and Turkey. Biochemical Systematics and Ecology, 69, 6- 14. https://doi.org/10.1016/j.bse.2016.08.003
Yu, D.W., Ji, Y., Emerson, B.C., Wang, X., Ye, C., Yang, C., & Ding, Z. (2012). Biodiversity soup: Metabarcoding of arthropods for rapid biodiversity assessment and biomonitoring. Methods in Ecology and Evolution, 3(4), 613-623. https://doi.org/10.1111/j.2041- 210X.2012.00198.x
H., & Chakrabarty, K. (2025). Detecting partial sample forgery in DNA forensics using molecular barcoding on digital microfluidic biochips. Journal of Hardware and Systems Security, 9, 136-148. https://doi.org/10.1007/s41635-025-00167-9.
Grzywacz, A., Wyborska, D., & Piwczyński, M. (2017). DNA barcoding allows identification of European Fanniidae (Diptera) of forensic interest. Forensic Science International, 275, 95-102 http://doi.org/10.1016/j.forsciint.2017.06.023
Hajibabaei, M., Smith, M.A., Janzen, D.H., Rodriguez, J.J., Whitfield, J.B., & Hebert, P.D.N. (2006). A minimalist barcode can identify a specimen whose DNA is degraded. Molecular Ecology Notes, 6(4), 959- 964. https://doi.org/10.1111/j.1471-8286.2006.01470.x
Haymer, D. (2024). State of the art and new directions using DNA and other molecular markers in forensic entomology. Academia Biology, 2, 1–8 https://doi.org/10.20935/AcadBiol7450.
Hebert, P.D.N., Cywinska, A., Ball, S.L., & deWaard, J.R. (2003). Biological identifications through DNA barcodes. Proceedings of the Royal Society B: Biological Sciences, 270(1512), 313-321 https://doi.org/10.1098/rspb.2002.2218
Karataş, D., &Darılmaz, M.C. (2018). A preliminary study on aquatic forensic entomology in Turkey. J. For. Med., 32(1), 18-29. https://doi.org/10.5505/adlitip.2018.48091
Khullar, N., & Singh, D. (2025). Advances in molecular techniques in forensic entomology. In Tanda A.S. (Ed), Advances in Forensic Entomology: Basic and Applied Aspects, 51–71p, TechSar Publishing.
Liu, J., Jiang, J., Song, S., Tornabene, L., Chabarria, R., Gavin, J.P.N. & Li, C. (2017). Multilocus DNA barcoding - Species Identification with Multilocus Data. Scientific Reports, 7, 16601. https://doi.org/10.1038/s41598-017-16920-2
López-Rubio, A., Amat, E., Pérez, A., Gómez, G., & Gómez, L.M. (2017). Assessing a DNA mini-barcode strategy for species identification in neotropical necrophagous blow flies (Diptera: Calliphoridae) of forensic importance. Genome 60(11), 965.
Lord, W.D., & Stevenson, JR. (1986). Directory of forensic entomologists. 2nd ed. Washington, DC: Defense Pest Management Information Analysis Center, Walter Reed Army Medical Center.
Meyer, C.P., & Paulay, G. (2005). DNA barcoding: Error rates based on comprehensive sampling. PLoS Biology, 3(12), e422 https://doi.org/10.1371/journal.pbio.0030422
Mutlu, E.Ç., Yılmaz, M.C., Balık, R., Onay, A.S., & Turan, Y. (2024). Determination of insect fauna and succession important for forensic entomology in İstanbul Province. Transactions of the American Entomological Society, 216-230. https://doi.org/10.3157/061.150.0202.
Özdemir, S., & Sert, O. (2009). Determination of Coleoptera fauna on carcasses in Ankara province, Turkey. Forensic Science International, 183, 24-32. https://doi.org/10.1016/j.forsciint.2008.09.018.
Park, S.H., Kim, H.J., Jang, H., Park, J., Kim, Y.H., & Ko, K.S. (2018). Molecular identification of forensically important flies (Diptera: Calliphoridae, Sarcophagidae) using COI and ITS2 gene regions. Journal of Asia- Pacific Entomology, 21, 1-10 https://doi.org/10.1016/j.forsciint.2017.12.017
Sagar, A., Singh, T.K., Harsh, N., Jha, N.K., Dode, P.S. & Podder, A. (2025). DNA barcoding: From taxonomy to technology - A review. Journal of Carcinogenesis, 24(7s), 835-842 https://doi.org/10.64149/J.Carcinog.24.7s.835-843
Schilthuizen, M., Scholte, C., van Wijk, R.E.J., Dommershuijzen, J., van der Horst, D., zu Schlochtern, M.M., Lievers, R., & Groenenberg, D.S.J. (2011). Using DNA-barcoding to make the necrobiont beetle family Cholevidae accessible for forensic entomology. Forensic Science International 210, 91-95 https://doi.org/10.1016/j.forsciint.2011.02.003
Scieuzo, C., Rinaldi, R., De Stefano, F., Di Fazio, A., & Falabella, P. (2025). The contribution of molecular biology to forensic entomology. Insects, 16, 694. https://doi.org/10.3390/insects16070694
Sert, O., Kabalak, M., & Şabanoğlu, B. (2012). Determination of forensically important Coleoptera and Calliphoridae (Diptera) species on decomposing dog (Canis lupus familiaris L.) carcass at Ankara province. Hacettepe Journal of Biology and Chemistry, 40(1), 99-103.
Sharawi, S.E. (2025). Forensic entomology in Jeddah: Molecular identification of emerging insect strains. Indian Journal of Animal Research, 59(1), 1-4. https://doi.org/10.18805/IJAR.BF-1881
Şabanoğlu, B., & Sert, O. (2010). Determination of Calliphoridae (Diptera) fauna and seasonal distribution on carrion in Ankara province. Journal of Forensic Sciences, 55(4), 1003-1007. https://doi.org/10.1111/j.1556-4029.2010.01366.x
Schilthuizen, M., Scholte, C., van Wijk, R.E.J., Dommershuijzen, J., van der Horst, D., Meijer zu Schlochtern, M., Lievers, R. & Groenenberg, D. S. J. (2011). Using DNA-barcoding to make the necrobiont beetle family Cholevidae accessible for forensic entomology. Forensic Science International, 210, 91-95. https://doi.org/10.1016/j.forsciint.2011.02.003
Sharma, S., & Dalal, J. (2025). in Tanda, A.S. (Ed.). Advances in forensic entomology: Basic and applied aspects. TechSar Publishing.
Taberlet, P., Coissac, E., Hajibabaei, M., & Rieseberg, L.H. (2012). Environmental DNA. Molecular Ecology, 21(8), 1789–1793. https://doi.org/10.1111/j.1365- 294X.2012.05542.x
Wang, B., Ha, S., Cai, J., Ma, Y., Li, D., Chen, J., & Deng, J. (2025). Exploratory study on DNA barcode combined with PCR-HRM technology for rapid and accurate identification of necrophilous fly species. Insects, 16, 590. https://doi.org/10.3390/insects16060590
Wells, J.D., & Stevens, J.R. (2008). Application of DNA-based methods in forensic entomology. Annual Review of Entomology, 53, 103-120.
Yavasoglu, S.İ., Yılmaz, C., Ulger, C., & Simsek, F.M. (2016). Molecular identification and genetic structure of Aedes phoeniciae (Diptera: Culicidae) in Northern Cyprus and Turkey. Biochemical Systematics and Ecology, 69, 6- 14. https://doi.org/10.1016/j.bse.2016.08.003
Yu, D.W., Ji, Y., Emerson, B.C., Wang, X., Ye, C., Yang, C., & Ding, Z. (2012). Biodiversity soup: Metabarcoding of arthropods for rapid biodiversity assessment and biomonitoring. Methods in Ecology and Evolution, 3(4), 613-623. https://doi.org/10.1111/j.2041- 210X.2012.00198.x

Ayrıntılar

Birincil Dil

İngilizce

Konular

Adli Biyoloji , Entomoloji

Bölüm

Derleme

Yazarlar

Mahmut Kabalak
0000-0001-6073-2551
Türkiye

Yavuz Turan ^*
0000-0001-9727-040X
Türkiye

Yayımlanma Tarihi

28 Şubat 2026

Gönderilme Tarihi

12 Aralık 2025

Kabul Tarihi

27 Ocak 2026

Yayımlandığı Sayı

Yıl 2026 Sayı: 2026

DOI

https://doi.org/10.35229/jaes.1840921

IZ

https://izlik.org/JA37KF44ZY

APA

Kabalak, M., & Turan, Y. (2026). DNA Barcoding in Forensic Entomology: A Molecular Tool That Increases Reliability in Estimating the Postmortem Interval. Journal of Anatolian Environmental and Animal Sciences, 2026. https://doi.org/10.35229/jaes.1840921

JAES, 2016- 11.yıl

DNA Barcoding in Forensic Entomology: A Molecular Tool That Increases Reliability in Estimating the Postmortem Interval

Abstract

Keywords

Adli Entomolojide DNA Barkodlama: Ölüm Sonrası Zaman Aralığı Tahmininde Güvenirliği Artıran Bir Moleküler Araç

Öz

Anahtar Kelimeler

Kaynakça

Ayrıntılar

Birincil Dil

Konular

Bölüm

Yazarlar

Yayımlanma Tarihi

Gönderilme Tarihi

Kabul Tarihi

Yayımlandığı Sayı

DOI

IZ

Kaynak Göster