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DNA barcoding is broadly used to analyse the diet of both invertebrate and vertebrate organisms[1][2] to detect and describe their trophic interactions[3][4]. This approach is based on the identification of consumed species by characterization of DNA present in dietary samples[5], e.g. individual food remains, regurgitates, gut and fecal samples, homogenized body of the host organism (for example with insects[6]).

The DNA sequencing approach to be adopted depends on the diet breadth of the target consumer. For organisms feeding on one or only few species, traditional Sanger sequencing techniques can be used. For polyphagous species with diet items more difficult to identify, it is conceivable to determine all consumed species using NGS methodology[7]. The barcode markers utilized for amplification will differ depending on the diet of the organism. For herbivore diets, the standard DNA barcode loci will differ significantly depending on the plant taxonomic level[8]. Therefore, for identifying plant tissue at the taxonomic family or genus level, the markers rbcL and trn-L-intron are used, which differ from the loci ITS2, matK, trnH-psbA (noncoding intergenic spacer) used to identify diet items to genus and species level[9]. For animal prey, the most broadly used DNA barcode markers to identify diets are the mitochondrial Cytocrome C oxydase (COI) and Cytochrome b (cytb)[10]. When the diet is broad and diverse, DNA metabarcoding is used to identify most of the consumed items[11].


Advantages

A major benefit of using DNA barcoding in diet assessment is the ability to provide high taxonomic resolution of consumed species[12]. Indeed, when compared to traditional morphological analysis, DNA barcoding enables a more reliable separation of closely related taxa[13]. Moreover, DNA barcoding enables to detect highly digested items, not recognisable through morphological identification[14]. For example, Arachnids feed on pre-digested bodies of insects or other small animals and their stomach contenct is too decomposed and morphologically unrecognizable using traditional methods such as microscopy[15]. Another advantage of DNA barcoding is to reduce the observer bias compared to morphological analysis, mostly when distinguishing between closely related taxa that can be mixed up [16][17][18]. When investigating herbivores diet, DNA metabarcoding enables detection of highly digested plant items with a higher number of taxa identified compared to macroscopic analysis[19][20]. For instance, Nichols et al. (2016) highlighted the taxonomic precision of metabarcoding on rumen contents, with on average 90% of DNA-sequences being identified to genus or species level in comparison to 75% of plant fragments recognised with macroscopy.

Challanges

With DNA barcoding it is impossible to retrieve information about sex or age of prey species, which can be crucial. This limitation can anyway be overcome with an additional step in the analysis by using microsatellite polimorphism and Y-chromosome amplification[21]. Moreover, DNA provides detailed information of the most recent events (e.g. 24–48 hr) but it is not able to provide a longer dietary prospect unless a continuous sampling is conducted[22]. When using generic primers that amplify ‘barcode’ regions from a broad range of food species, the amplifiable host DNA may largely outnumber the presence of prey DNA, complicating prey detection. However, a strategy to prevent the host DNA amplification has been developed by the addition of a predator-specific blocking primer[23][24] [25]. Indeed, blocking primers for suppressing amplification of predator DNA allows the amplification of the other vertebrate groups and produces amplicon mixes that are predominately food DNA[26][27].

Further, DNA barcoding secondary predation cannot be detected using DNA barcoding i.e. items that were consumed by a prey of the predator and true prey (Harms-Tuohy et al. 2016), but that is a problem also in traditional stomach content analysis.  

Eglė Jakubavičiūtė et al 2017

The quantitative interpretation of DNA barcoding results is not straightforward [28]. There have been attempts to use the number of sequences recovered to estimate the abundance of prey species in digested contents (e.g. gut, faeces). For example, if the wolf ate more moose than wild boar, there should be more moose DNA in their gut, and thus, more moose sequences are recovered. Despite the evidence for general correlations between the sequence number and the biomass, actual evaluations of this method have been unsuccessful[29] (Deagle et al. 2013; Pinol et al. 2014). This can be explained by the fact that tissues originally contain different densities of DNA and can be digested differently[30].

Examples

Mammals

Within mammals there are three different feeding regimes: herbivores, carnivores, generalists.

herbivores: Through its fine resolution in plant determination, this molecular approach represents a crucial tool in wildlife management to identify the feeding habits of endangered species and animals that can cause feeding damages to the environment[31].

voles small herbivores Soininen 2009

bear (De Barba et al., 2014)

leopard cat (Shehzad et al., 2012)

seal (Casper et al., 2007; Meheust et al., 2015)

red bat (Clare et al., 2009)

Fish

Birds

Arthropods

See also

  • DNA barcoding of fish
  • DNA barcoding of invertebrates

Reference

  1. ^ King, R. A.; Read, D. S.; Traugott, M.; Symondson, W. O. C. (2008-01-14). "INVITED REVIEW: Molecular analysis of predation: a review of best practice for DNA-based approaches: OPTIMIZING MOLECULAR ANALYSIS OF PREDATION". Molecular Ecology. 17 (4): 947–963. doi:10.1111/j.1365-294X.2007.03613.x.
  2. ^ Pompanon, Francois; Deagle, Bruce E.; Symondson, William O. C.; Brown, David S.; Jarman, Simon N.; Taberlet, Pierre (2012). "Who is eating what: diet assessment using next generation sequencing". Molecular Ecology. 21 (8): 1931–1950. doi:10.1111/j.1365-294X.2011.05403.x. ISSN 1365-294X.
  3. ^ Sheppard, S. K.; Harwood, J. D. (2005-10). "Advances in molecular ecology: tracking trophic links through predator-prey food-webs". Functional Ecology. 19 (5): 751–762. doi:10.1111/j.1365-2435.2005.01041.x. ISSN 0269-8463. {{cite journal}}: Check date values in: |date= (help)
  4. ^ Erickson, David L.; Uriarte, Maria; García-Robledo, Carlos; Kress, W. John (2015-01-01). "DNA barcodes for ecology, evolution, and conservation". Trends in Ecology & Evolution. 30 (1): 25–35. doi:10.1016/j.tree.2014.10.008. ISSN 0169-5347. PMID 25468359.
  5. ^ POMPANON, FRANCOIS; DEAGLE, BRUCE E.; SYMONDSON, WILLIAM O. C.; BROWN, DAVID S.; JARMAN, SIMON N.; TABERLET, PIERRE (2011-12-15). "Who is eating what: diet assessment using next generation sequencing". Molecular Ecology. 21 (8): 1931–1950. doi:10.1111/j.1365-294x.2011.05403.x. ISSN 0962-1083.
  6. ^ HARWOOD, JAMES D.; DESNEUX, NICOLAS; YOO, HO JUNG S.; ROWLEY, DANIEL L.; GREENSTONE, MATTHEW H.; OBRYCKI, JOHN J.; O′NEIL, ROBERT J. (2007-10). "Tracking the role of alternative prey in soybean aphid predation byOrius insidiosus: a molecular approach". Molecular Ecology. 16 (20): 4390–4400. doi:10.1111/j.1365-294x.2007.03482.x. ISSN 0962-1083. {{cite journal}}: Check date values in: |date= (help)
  7. ^ POMPANON, FRANCOIS; DEAGLE, BRUCE E.; SYMONDSON, WILLIAM O. C.; BROWN, DAVID S.; JARMAN, SIMON N.; TABERLET, PIERRE (2011-12-15). "Who is eating what: diet assessment using next generation sequencing". Molecular Ecology. 21 (8): 1931–1950. doi:10.1111/j.1365-294x.2011.05403.x. ISSN 0962-1083.
  8. ^ Kress, W. John; García-Robledo, Carlos; Uriarte, Maria; Erickson, David L. (2015-01). "DNA barcodes for ecology, evolution, and conservation". Trends in Ecology & Evolution. 30 (1): 25–35. doi:10.1016/j.tree.2014.10.008. ISSN 0169-5347. {{cite journal}}: Check date values in: |date= (help)
  9. ^ Kress, W. John; García-Robledo, Carlos; Uriarte, Maria; Erickson, David L. (2015-01). "DNA barcodes for ecology, evolution, and conservation". Trends in Ecology & Evolution. 30 (1): 25–35. doi:10.1016/j.tree.2014.10.008. ISSN 0169-5347. {{cite journal}}: Check date values in: |date= (help)
  10. ^ Tobe, Shanan S.; Kitchener, Andrew; Linacre, Adrian (2009-12). "Cytochrome b or cytochrome c oxidase subunit I for mammalian species identification—An answer to the debate". Forensic Science International: Genetics Supplement Series. 2 (1): 306–307. doi:10.1016/j.fsigss.2009.08.053. ISSN 1875-1768. {{cite journal}}: Check date values in: |date= (help)
  11. ^ Jakubavičiūtė, Eglė; Bergström, Ulf; Eklöf, Johan S.; Haenel, Quiterie; Bourlat, Sarah J. (2017-10-23). "DNA metabarcoding reveals diverse diet of the three-spined stickleback in a coastal ecosystem". PLOS ONE. 12 (10): e0186929. doi:10.1371/journal.pone.0186929. ISSN 1932-6203.{{cite journal}}: CS1 maint: unflagged free DOI (link)
  12. ^ Garnick, Sarah; Barboza, Perry S.; Walker, John W. (2018-07). "Assessment of Animal-Based Methods Used for Estimating and Monitoring Rangeland Herbivore Diet Composition". Rangeland Ecology & Management. 71 (4): 449–457. doi:10.1016/j.rama.2018.03.003. ISSN 1550-7424. {{cite journal}}: Check date values in: |date= (help)
  13. ^ Santos, Teresa; Fonseca, Carlos; Barros, Tânia; Godinho, Raquel; Bastos-Silveira, Cristiane; Bandeira, Victor; Rocha, Rita Gomes (2015-05-20). "Using stomach contents for diet analysis of carnivores through DNA barcoding". Wildlife Biology in Practice. 11 (1). doi:10.2461/wbp.2015.11.4. ISSN 1646-2742.
  14. ^ Piñol, J.; San Andrés, V.; Clare, E. L.; Mir, G.; Symondson, W. O. C. (2013-08-20). "A pragmatic approach to the analysis of diets of generalist predators: the use of next-generation sequencing with no blocking probes". Molecular Ecology Resources. 14 (1): 18–26. doi:10.1111/1755-0998.12156. ISSN 1755-098X.
  15. ^ Agustí, N.; Shayler, S. P.; Harwood, J. D.; Vaughan, I. P.; Sunderland, K. D.; Symondson, W. O. C. (2003). "Collembola as alternative prey sustaining spiders in arable ecosystems: prey detection within predators using molecular markers". Molecular Ecology. 12 (12): 3467–3475. doi:10.1046/j.1365-294X.2003.02014.x. ISSN 1365-294X.
  16. ^ Robeson, Michael S.; Khanipov, Kamil; Golovko, George; Wisely, Samantha M.; White, Michael D.; Bodenchuck, Michael; Smyser, Timothy J.; Fofanov, Yuriy; Fierer, Noah (2017-11-26). "Assessing the utility of metabarcoding for diet analyses of the omnivorous wild pig (Sus scrofa )". Ecology and Evolution. 8 (1): 185–196. doi:10.1002/ece3.3638. ISSN 2045-7758. {{cite journal}}: line feed character in |title= at position 96 (help)
  17. ^ Valentini, Alice; Pompanon, François; Taberlet, Pierre (2009-02). "DNA barcoding for ecologists". Trends in Ecology & Evolution. 24 (2): 110–117. doi:10.1016/j.tree.2008.09.011. ISSN 0169-5347. {{cite journal}}: Check date values in: |date= (help)
  18. ^ Clare, Elizabeth L. (2014-10-29). "Molecular detection of trophic interactions: emerging trends, distinct advantages, significant considerations and conservation applications". Evolutionary Applications. 7 (9): 1144–1157. doi:10.1111/eva.12225. ISSN 1752-4571.
  19. ^ Nichols, Ruth V.; Åkesson, Mikael; Kjellander, Petter (2016-06-20). "Diet Assessment Based on Rumen Contents: A Comparison between DNA Metabarcoding and Macroscopy". PLOS ONE. 11 (6): e0157977. doi:10.1371/journal.pone.0157977. ISSN 1932-6203.{{cite journal}}: CS1 maint: unflagged free DOI (link)
  20. ^ Soininen, Eeva M; Valentini, Alice; Coissac, Eric; Miquel, Christian; Gielly, Ludovic; Brochmann, Christian; Brysting, Anne K; Sønstebø, Jørn H; Ims, Rolf A (2009). "Analysing diet of small herbivores: the efficiency of DNA barcoding coupled with high-throughput pyrosequencing for deciphering the composition of complex plant mixtures". Frontiers in Zoology. 6 (1): 16. doi:10.1186/1742-9994-6-16. ISSN 1742-9994.{{cite journal}}: CS1 maint: unflagged free DOI (link)
  21. ^ GRIFFITHS, R.; TIWARI, B. (1993-12). "Primers for the differential amplification of the sex-determining region Y gene in a range of mammal species". Molecular Ecology. 2 (6): 405–406. doi:10.1111/j.1365-294x.1993.tb00034.x. ISSN 0962-1083. {{cite journal}}: Check date values in: |date= (help)
  22. ^ Thomsen, Philip Francis; Kielgast, Jos; Iversen, Lars Lønsmann; Møller, Peter Rask; Rasmussen, Morten; Willerslev, Eske (2012-08-29). "Detection of a Diverse Marine Fish Fauna Using Environmental DNA from Seawater Samples". PLoS ONE. 7 (8): e41732. doi:10.1371/journal.pone.0041732. ISSN 1932-6203.{{cite journal}}: CS1 maint: unflagged free DOI (link)
  23. ^ Vestheim, Hege; Jarman, Simon N (2008). "Blocking primers to enhance PCR amplification of rare sequences in mixed samples – a case study on prey DNA in Antarctic krill stomachs". Frontiers in Zoology. 5 (1): 12. doi:10.1186/1742-9994-5-12. ISSN 1742-9994.{{cite journal}}: CS1 maint: unflagged free DOI (link)
  24. ^ SHEHZAD, WASIM; RIAZ, TIAYYBA; NAWAZ, MUHAMMAD A.; MIQUEL, CHRISTIAN; POILLOT, CAROLE; SHAH, SAFDAR A.; POMPANON, FRANÇOIS; COISSAC, ERIC; TABERLET, PIERRE (2012-01-17). "Carnivore diet analysis based on next-generation sequencing: application to the leopard cat (Prionailurus bengalensis) in Pakistan". Molecular Ecology. 21 (8): 1951–1965. doi:10.1111/j.1365-294x.2011.05424.x. ISSN 0962-1083.
  25. ^ Jarman, Simon N.; McInnes, Julie C.; Faux, Cassandra; Polanowski, Andrea M.; Marthick, James; Deagle, Bruce E.; Southwell, Colin; Emmerson, Louise (2013-12-16). "Adélie Penguin Population Diet Monitoring by Analysis of Food DNA in Scats". PLoS ONE. 8 (12): e82227. doi:10.1371/journal.pone.0082227. ISSN 1932-6203.{{cite journal}}: CS1 maint: unflagged free DOI (link)
  26. ^ Vestheim, Hege; Jarman, Simon N (2008). "Blocking primers to enhance PCR amplification of rare sequences in mixed samples – a case study on prey DNA in Antarctic krill stomachs". Frontiers in Zoology. 5 (1): 12. doi:10.1186/1742-9994-5-12. ISSN 1742-9994. PMC 2517594. PMID 18638418.{{cite journal}}: CS1 maint: PMC format (link) CS1 maint: unflagged free DOI (link)
  27. ^ Vestheim, Hege; Deagle, Bruce E.; Jarman, Simon N. (2010-09-29), "Application of Blocking Oligonucleotides to Improve Signal-to-Noise Ratio in a PCR", Methods in Molecular Biology, Humana Press, pp. 265–274, ISBN 9781607619437, retrieved 2019-03-29
  28. ^ Valentini, Alice; Pompanon, François; Taberlet, Pierre (2009-02). "DNA barcoding for ecologists". Trends in Ecology & Evolution. 24 (2): 110–117. doi:10.1016/j.tree.2008.09.011. ISSN 0169-5347. {{cite journal}}: Check date values in: |date= (help)
  29. ^ POMPANON, FRANCOIS; DEAGLE, BRUCE E.; SYMONDSON, WILLIAM O. C.; BROWN, DAVID S.; JARMAN, SIMON N.; TABERLET, PIERRE (2011-12-15). "Who is eating what: diet assessment using next generation sequencing". Molecular Ecology. 21 (8): 1931–1950. doi:10.1111/j.1365-294x.2011.05403.x. ISSN 0962-1083.
  30. ^ Deagle, Bruce E.; Chiaradia, André; McInnes, Julie; Jarman, Simon N. (2010-06-17). "Pyrosequencing faecal DNA to determine diet of little penguins: is what goes in what comes out?". Conservation Genetics. 11 (5): 2039–2048. doi:10.1007/s10592-010-0096-6. ISSN 1566-0621.
  31. ^ Ando, Haruko; Fujii, Chieko; Kawanabe, Masataka; Ao, Yoshimi; Inoue, Tomomi; Takenaka, Akio (2018-10-22). "Evaluation of plant contamination in metabarcoding diet analysis of a herbivore". Scientific Reports. 8 (1). doi:10.1038/s41598-018-32845-w. ISSN 2045-2322.
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