Dichlorvos exposure impedes extraction and amplification of DNA from insects in museum collections

Espeland, Marianne; Irestedt, Martin; Johanson, Kjell Arne; Åkerlund, Monika; Bergh, Jan-Erik; Källersjö, Mari

doi:10.1186/1742-9994-7-2

Cited by 38 publications

(29 citation statements)

References 33 publications

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“…While past work has revealed that museum specimens are a challenging analytical target [21], [22], this study has provided new details on factors that impact DNA sequence recovery. Specimen age was confirmed as an important determinant of sequence recovery, but most specimens yielded a barcode compliant sequence if analyzed within 50 years of collection.…”

Section: Discussionmentioning

confidence: 96%

“…Specimens were selected for analysis based on their age and collection site, but sequencing success would have risen if collector had also been considered. The cause of these collector effects is uncertain, but may reflect differential killing agents, treatment during specimen preparation, and/or subsequent storage conditions [22]. The strength of these effects suggests the value of conducting a performance test on a subset of representative specimens from each major collector before launching large-scale efforts at barcode recovery.…”

Section: Discussionmentioning

confidence: 99%

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A DNA ‘Barcode Blitz’: Rapid Digitization and Sequencing of a Natural History Collection

et al. 2013

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DNA barcoding protocols require the linkage of each sequence record to a voucher specimen that has, whenever possible, been authoritatively identified. Natural history collections would seem an ideal resource for barcode library construction, but they have never seen large-scale analysis because of concerns linked to DNA degradation. The present study examines the strength of this barrier, carrying out a comprehensive analysis of moth and butterfly (Lepidoptera) species in the Australian National Insect Collection. Protocols were developed that enabled tissue samples, specimen data, and images to be assembled rapidly. Using these methods, a five-person team processed 41,650 specimens representing 12,699 species in 14 weeks. Subsequent molecular analysis took about six months, reflecting the need for multiple rounds of PCR as sequence recovery was impacted by age, body size, and collection protocols. Despite these variables and the fact that specimens averaged 30.4 years old, barcode records were obtained from 86% of the species. In fact, one or more barcode compliant sequences (>487 bp) were recovered from virtually all species represented by five or more individuals, even when the youngest was 50 years old. By assembling specimen images, distributional data, and DNA barcode sequences on a web-accessible informatics platform, this study has greatly advanced accessibility to information on thousands of species. Moreover, much of the specimen data became publically accessible within days of its acquisition, while most sequence results saw release within three months. As such, this study reveals the speed with which DNA barcode workflows can mobilize biodiversity data, often providing the first web-accessible information for a species. These results further suggest that existing collections can enable the rapid development of a comprehensive DNA barcode library for the most diverse compartment of terrestrial biodiversity – insects.

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Section: Discussionmentioning

confidence: 96%

Section: Discussionmentioning

confidence: 99%

A DNA ‘Barcode Blitz’: Rapid Digitization and Sequencing of a Natural History Collection

et al. 2013

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“…First, the specimens were pinned and dried to study external morphology; as such, the soft tissue inside exoskeleton was dried and decomposed by time without chemical preservative treatment (Dick, Bridge, Wheeler, & Desalle, ; Sutrisno, ), resulting in limited intact DNA source. Second, dichlorvos, a preservative chemical used to prevent collection pest, may negatively affect DNA amplification (Espeland et al, ; Werblow et al, ) and third, other physical factors might affect DNA integrity. For example, partial dehydration and exposure to air and light may potentially lead to DNA degradation, particularly deamination of cystidine residues (Mitchell, Willerslev, & Hansen, ).…”

Section: Discussionmentioning

confidence: 99%

A new cost‐effective and fast direct PCR protocol for insects based on PBS buffer

Thongjued

Chotigeat

Bumrungsri

et al. 2019

Molecular Ecology Resources

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Insect DNA barcoding is a species identification technique used in biodiversity assessment and ecological studies. However, DNA extraction can result in the loss of up to 70% of DNA. Recent research has reported that direct PCR can overcome this issue. However, the success rates could still be improved, and tissues used for direct PCR could not be reused for further genetic studies. Here, we developed a direct PCR workflow that incorporates a 2‐min sample preparation in PBS‐buffer step for fast and effective universal insect species identification. The developed protocol achieved 100% success rates for amplification in six orders: Mantodea, Phasmatodea, Neuroptera, Odonata, Blattodea and Orthoptera. High and moderate success rates were obtained for five other species: Lepidoptera (97.3%), Coleoptera (93.8%), Diptera (90.5%), Hemiptera (81.8%) and Hymenoptera (75.0%). High‐quality sequencing data were also obtained from these amplifiable products, allowing confidence in species identification. The method was sensitive down to 1/4th of a 1‐mm fragment of leg or body and its success rates with oven‐dried, ethanol‐preserved, food, bat guano and museum specimens were 100%, 98.6%, 90.0%, 84.0% and 30.0%, respectively. In addition, the pre‐PCR solution (PBS with insect tissues) could be used for further DNA extraction if needed. The workflow will be beneficial in the fields of insect taxonomy and ecological studies due to its low cost, simplicity and applicability to highly degraded specimens.

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“…The negative direct PCR results using legs as templates could be attributed to the conservation of the specimens. For the prevention of collection pests, dry preparations have often been treated with the insecticide dichlorvos, which affects the DNA of insects negatively (Espeland et al, 2010). Other parameters that may influence the amplification are the primer pairs and the polymerase used.…”

Section: Discussionmentioning

confidence: 99%

Direct PCR of indigenous and invasive mosquito species: a time‐ and cost‐effective technique of mosquito barcoding

Werblow

Flechl

Klimpel

et al. 2015

Medical Vet Entomology

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Millions of people die each year as a result of pathogens transmitted by mosquitoes. However, the morphological identification of mosquito species can be difficult even for experts. The identification of morphologically indistinguishable species, such as members of the Anopheles maculipennis complex (Diptera: Culicidae), and possible hybrids, such as Culex pipiens pipiens/Culex pipiens molestus (Diptera: Culicidae), presents a major problem. In addition, the detection and discrimination of newly introduced species can be challenging, particularly to researchers without previous experience. Because of their medical importance, the clear identification of all relevant mosquito species is essential. Using the direct polymerase chain reaction (PCR) method described here, DNA amplification without prior DNA extraction is possible and thus species identification after sequencing can be achieved. Different amounts of tissue (leg, head; larvae or adult) as well as different storage conditions (dry, ethanol, −20 and −80 °C) and storage times were successfully applied and showed positive results after amplification and gel electrophoresis. Overall, 28 different indigenous and non‐indigenous mosquito species were analysed using a gene fragment of the COX1 gene for species differentiation and identification by sequencing this 658‐bp fragment. Compared with standard PCR, this method is time‐ and cost‐effective and could thus improve existing surveillance and control programmes.

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Dichlorvos exposure impedes extraction and amplification of DNA from insects in museum collections

Cited by 38 publications

References 33 publications

A DNA ‘Barcode Blitz’: Rapid Digitization and Sequencing of a Natural History Collection

A DNA ‘Barcode Blitz’: Rapid Digitization and Sequencing of a Natural History Collection

A new cost‐effective and fast direct PCR protocol for insects based on PBS buffer

Direct PCR of indigenous and invasive mosquito species: a time‐ and cost‐effective technique of mosquito barcoding

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