Vector soup: high‐throughput identification of Neotropical phlebotomine sand flies using metabarcoding

Kocher, Arthur; Gantier, Jean-Charles; Gaborit, Pascal; Zinger, Lucie; Holota, Hélène; Valière, Sophie; Dusfour, Isabelle; Girod, Romain; Bañuls, Anne‐Laure; Murienne, Jérôme

doi:10.1111/1755-0998.12556

Cited by 35 publications

(42 citation statements)

References 62 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…[30]). Because of the short length of the sequences, genetic distances are computed based on pairwise alignments rather than on the multiple sequence alignment of all sequences.…”

Section: Methodsmentioning

confidence: 99%

“…This approach, referred to as metabarcoding, allows the simultaneous identification of multiple specimens from a single bulk-DNA extraction [29; 30]. While the COI marker has been used as a standard in barcoding applications, it is not the best choice when it comes to metabarcoding [31] and a shorter fragment in the 16S ribosomal gene has been specifically designed for metabarcoding applications for insects [32].…”

Section: Introductionmentioning

confidence: 99%

“…While the COI marker has been used as a standard in barcoding applications, it is not the best choice when it comes to metabarcoding [31] and a shorter fragment in the 16S ribosomal gene has been specifically designed for metabarcoding applications for insects [32]. It was recently successfully used to analyze samples of Phlebotomine sandflies [30]. …”

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

DNA reference libraries of French Guianese mosquitoes for barcoding and metabarcoding

et al. 2017

Self Cite

View full text Add to dashboard Cite

The mosquito family (Diptera: Culicidae) constitutes the most medically important group of arthropods because certain species are vectors of human pathogens. In some parts of the world, the diversity is so high that the accurate delimitation and/or identification of species is challenging. A DNA-based identification system for all animals has been proposed, the so-called DNA barcoding approach. In this study, our objectives were (i) to establish DNA barcode libraries for the mosquitoes of French Guiana based on the COI and the 16S markers, (ii) to compare distance-based and tree-based methods of species delimitation to traditional taxonomy, and (iii) to evaluate the accuracy of each marker in identifying specimens. A total of 266 specimens belonging to 75 morphologically identified species or morphospecies were analyzed allowing us to delimit 86 DNA clusters with only 21 of them already present in the BOLD database. We thus provide a substantial contribution to the global mosquito barcoding initiative. Our results confirm that DNA barcodes can be successfully used to delimit and identify mosquito species with only a few cases where the marker could not distinguish closely related species. Our results also validate the presence of new species identified based on morphology, plus potential cases of cryptic species. We found that both COI and 16S markers performed very well, with successful identifications at the species level of up to 98% for COI and 97% for 16S when compared to traditional taxonomy. This shows great potential for the use of metabarcoding for vector monitoring and eco-epidemiological studies.

show abstract

“…[30]). Because of the short length of the sequences, genetic distances are computed based on pairwise alignments rather than on the multiple sequence alignment of all sequences.…”

Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

DNA reference libraries of French Guianese mosquitoes for barcoding and metabarcoding

et al. 2017

Self Cite

View full text Add to dashboard Cite

show abstract

“…Studies have shown that metabarcoding can be used to identify arthropod taxa in bulk arthropod samples (e.g. Kocher et al, ; Liu et al, ; Morinière et al, ), and for example be applied to assess how arthropod diversity changes to different land‐uses (Beng et al, ). Metabarcoding has also been used to assess how invertebrates might leak DNA to their preservative ethanol which then potentially can serve as a mean to obtain sample diversity information (Erdozain et al, ; Hajibabaei, Spall, Shokralla, & Konynenburg, ).…”

Section: Introductionmentioning

confidence: 99%

A simplified DNA extraction protocol for unsorted bulk arthropod samples that maintains exoskeletal integrity

et al. 2019

View full text Add to dashboard Cite

High‐throughput DNA sequencing offers an efficient tool for assessing the taxonomic content of bulk arthropod samples. Many current DNA extraction protocols however require extensive handling of samples, like specimen‐based DNA extractions, or sorting of samples and are thus unsuitable for large scale studies. Furthermore, protocols often include homogenization and thus imply partial or complete destruction of the sample constituents. The aim of this study was therefore to investigate steps related both to sample pre‐processing and DNA extraction of unsorted bulk arthropod samples and explore possibilities for simplifying sample processing and thus increase sample handling efficiency without losing taxonomic information. Using mock bulk arthropod samples, we compare laboratory steps related to DNA extraction and semi‐automatic handling. Specifically, we (a) assess whether aliquots of digest buffer from bulk arthropod samples adequately describe the community composition; (b) compare a non‐destructive and a destructive DNA extraction method; (c) compare a phenol/chloroform inhibitor removal method with the exclusion of the same; and (d) compare manual purification to automated DNA purification on a QIAcube laboratory robot. Using DNA metabarcoding and mock bulk arthropod samples, we show that it is possible to efficiently process unsorted arthropod bulk samples with a non‐destructive DNA extraction approach. We found that homogenizing samples yielded more DNA but also generally produced more inconsistent results when compared to non‐destructive extraction. When assessing the recovered taxonomic content of samples (operational taxonomic units, OTUs), intact samples performed at least comparable to, if not better, than homogenized samples. Additionally, we show that sample processing can be further simplified from using a defined volume of digest, no phenol/chloroform purification and automated DNA purification. This approach can be a way to process hundreds of unsorted bulk samples effectively, consistently and with a minimum loss of valuable morphologic information.

show abstract

“…; Kocher et al . ). In addition, it has the great advantage to be applicable on degraded DNA present in the environment such as soil (Andersen et al .…”

Section: Introductionmentioning

confidence: 97%

Evaluation of short mitochondrial metabarcodes for the identification of Amazonian mammals

et al. 2017

Self Cite

View full text Add to dashboard Cite

Summary DNA barcoding and metabarcoding are increasingly used as alternatives to traditional morphological identifications. For animals, the standard barcode is a c. 658‐bp portion of the COI gene, for which reference libraries now cover a large proportion of described mammal species. Unfortunately, because its sequence is too long and does not contain highly conserved primer binding sites, this marker is not adapted for metabarcoding. Although alternative metabarcodes have been developed, their performances are generally seldom assessed. We evaluate the reliability of a short metabarcode located in the mitochondrial 12S ribosomal RNA for the identifications of Amazonian mammals. We (i) constitute a nearly exhaustive reference library for species found in French Guiana, (ii) assess the taxonomic resolution of the marker and validate its use with dipteran blood meal analyses, (iii) assess the conservation of the primer binding sites, and (iv) compare its theoretical performances with that of a newly designed metabarcode located within the standard COI barcode. About 576 specimens representing 164 species were gathered and sequenced. We show that the 12S marker allows remarkably accurate taxonomic assignations despite its very short size, and that primer binding sites are highly conserved, which is important to avoid PCR amplification bias potentially leading to detection failure. Additionally, our results stress that the identifications should only be considered at the generic level when they are based on incomplete reference libraries, even when a stringent similarity cut‐off is used. A new short COI metabarcode was designed based on 569 reference sequences of mammals retrieved on BOLD. Our results clearly show that, while both markers provide similar taxonomic resolution, much higher rates of primer mismatches are found with COI. Besides demonstrating the accuracy of the short 12S marker for the identification of Amazonian mammals and providing a reliable molecular reference database, this study emphasizes that the accuracy of taxonomic assignations highly depends on the comprehensiveness of the reference library and that great caution should be taken for interpreting metabarcoding results based on scarce reference libraries. The comparison with a short COI metabarcode also provides novel evidence in support for the use of ribosomal markers in metabarcoding studies.

show abstract

Vector soup: high‐throughput identification of Neotropical phlebotomine sand flies using metabarcoding

Cited by 35 publications

References 62 publications

DNA reference libraries of French Guianese mosquitoes for barcoding and metabarcoding

DNA reference libraries of French Guianese mosquitoes for barcoding and metabarcoding

A simplified DNA extraction protocol for unsorted bulk arthropod samples that maintains exoskeletal integrity

Evaluation of short mitochondrial metabarcodes for the identification of Amazonian mammals

Contact Info

Product

Resources

About