Strategic considerations for invasive species managers in the utilization of environmental DNA (eDNA): steps for incorporating this powerful surveillance tool

Morisette, Jeffrey T.; Burgiel, Stanley W.; Brantley, Kelsey A.; Daniel, Wesley M.; Darling, John A.; Davis, Jeanette; Franklin, Thomas W.; Gaddis, Keith D.; Hunter, Margaret E.; Lance, Richard F.; Leskey, Tracy; Passamaneck, Yale J.; Piaggio, Antoinette J.; Rector, B. G.; Sepulveda, Adam J.; Smith, M. R.; Stepien, Carol A.; Wilcox, Taylor M.

doi:10.3391/mbi.2021.12.3.15

Cited by 34 publications

(34 citation statements)

References 43 publications

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“…As such, both TaqMan and LAMP assays were tested against a single environmental matrix for the purpose of assessing assay reproducibility. Real Time PCR has been routinely used to detect eDNA from a broad range of surveillance applications (15) and was shown by this study to successfully amplify T. granarium eDNA in a biosecurity context. Similarly, LAMP assays have also been developed to amplify DNA from microscopic pathogens and parasites in water samples and other fluid matrices (29)(30)(31)(32), with one example of LAMP eDNA detection for a marine mollusc in ex-situ conditions (33).…”

Section: Discussionmentioning

confidence: 89%

“…For these methods to be adopted, the sampling, processing, and analyzing samples must be technically feasible, precise, and repeatable (24). Surveillance applications using eDNA-based detection have been demonstrated to reliably inform end users on the presence and activity of exotic species to better manage biosecurity risks and improve detection probability (15,(24)(25)(26). Border control and onshore biosecurity responses stand out as unique applications due to their importance in trade and legislation.…”

Section: Discussionmentioning

confidence: 99%

“…As such, there is a need to test and develop molecular methods that complement the needs of biosecurity officers during biosecurity responses. Environmental DNA (eDNA)-based methods have been shown to be cost-effective reliable methods to inform users on the presence of target species in surveillance applications (15), with studies showing how eDNA-based detection using soil and airborne dust samples can provide valuable data on species presence and diversity (16)(17)(18). When used with portable, pointof-care technologies, eDNA-based detection can offer sensitive detection tools to support management of biosecurity risks (19), ADDIN EN.CITE (16)(17)(18)but no studies have so far tested suitable extraction methods of dust samples and the utility of portable technologies during biosecurity applications.…”

Section: Introductionmentioning

confidence: 99%

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Detection of Khapra Beetle Environmental DNA Using Portable Technologies in Australian Biosecurity

et al. 2022

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Khapra beetle, Trogoderma granarium Everts, 1898, is a serious pest of stored grain products globally. Environmental DNA (eDNA)-based methods offer sensitive detection tools used to inform biosecurity officers on the presence of high-risk pests. This study tested laboratory and portable molecular technologies to detect khapra beetle environmental DNA extracted from dust samples collected during biosecurity responses (Tuggeranong and Fyshwick) to khapra beetle incursions in Australia. Airborne and floor dust samples were collected opportunistically using handheld vacuum cleaners and eDNA was extracted using either field or laboratory-based extraction methods and analyzed using laboratory benchtop real time PCR machines and portable machines with two TaqMan and one LAMP-based assay. We successfully collected, extracted, and amplified khapra beetle eDNA from dust samples by qPCR, but failed to amplify T. granarium eDNA using LAMP. The Laboratory qPCR machine showed significantly higher mean Ct values (p < 0.001) and significantly higher positive detections for both assays (p < 0.001) compared to the portable thermocycler. DNA yield was significantly higher in samples extracted using laboratory-based kits compared to field kits (p < 0.001) for both vacuumed and airborne samples (Mean DNA ± S.D. = 5.52 ± 4.45 and 4.77 ± 1.68 ng/μL, respectively), compared to field kits, (1.75 ± 1.17 and 1.36± 1.29 ng/μL for vacuumed and airborne samples, respectively). There were no significant differences in DNA yield between collection methods or differences in amplification associated to extraction or collection methods in either platform tested in this study. Portable technologies tested in this study (Franklin™ Real Time Thermocycler and Genie III) accurately amplified all tissue derived DNA during assay optimisation and field testing, highlighting the capacity of these technologies to complement biosecurity in confirming specimen ID. There was a high incidence of positive detections in field negative controls (Tuggeranong = 12.3 % and Fyshwick = 50 %), mostly attributed to the use of contaminated vacuum cleaners. We discuss suitable methods to minimize sample cross-contamination, the potential of portable molecular technologies as tools for biosecurity applications, and the suitability of eDNA-based molecular detection methods to complement global trade biosecurity for one of the most invasive and important grain pests worldwide.

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Section: Discussionmentioning

confidence: 89%

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Detection of Khapra Beetle Environmental DNA Using Portable Technologies in Australian Biosecurity

et al. 2022

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“…Monitoring and related control programs have recently focussed on the role of eDNA in invasive species detection (Hunter et al 2015;Morisette et al 2021), with several studies focusing on LAMP as a potential eDNA monitoring tool (Vythalingam et al 2021;Williams et al 2017). The emphasis on monitoring primarily concerns invertebrate pests, as demonstrated by the development of LAMP assays as a means of point of entry detection or field-based detection.…”

Section: Invasive Species Monitoringmentioning

confidence: 99%

Shining a LAMP on the applications of isothermal amplification to monitoring illegal wildlife trade

Deliveyne¹,

Young²,

Austin³

et al. 2022

Preprint

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Environmental biosecurity risks associated with the Illegal Wildlife Trade (IWT) include the loss of biodiversity, threats to public health, and the proliferation of invasive alien species. To assist enforcement agencies in identifying trafficked species, rapid forensic techniques enable the detection of trace Environmental DNA (eDNA) where physical identification is not possible. Loop Mediated Isothermal Amplification (LAMP) is an emerging technique with recent applications in biosecurity and forensic sciences, and with potential to function as a field-based detection tool. Here we provide an overview of current research that applies LAMP to human and wildlife forensic science, including identification of ornamental wildlife parts, consumer products, and invasive species monitoring and biosecurity detection. We discuss the current scope of LAMP as applied to various wildlife crime scenarios and biosecurity checkpoint monitoring, highlight the specificity, sensitivity, and robustness for these applications, and review the potential utility of LAMP for rapid field-based detection within the IWT. Based on our assessment of the literature we recommend broader interest, research, and investment in LAMP as an appropriate field-based species detection method for a wide range of environmental biosecurity scenarios.

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“…However, use of eDNA results for decision‐making has been impeded by the uncertainty of eDNA detections since multiple sources of error can give rise to observation and site‐level false positives (Darling et al, 2021). Resource managers require greater confidence that eDNA detections are not erroneous because there can be high economic, social and political costs associated with these decisions (Darling & Mahon, 2011; Kelly et al, 2014; Morisette et al, 2021; Sepulveda et al, 2020).…”

Section: Introductionmentioning

confidence: 99%

Time to get real with qPCR controls: The frequency of sample contamination and the informative power of negative controls in environmental DNA studies

Hutchins

Simantel

Sepulveda

2021

Molecular Ecology Resources

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Environmental (e)DNA methods have enabled rapid, sensitive and specific inferences of taxa presence throughout diverse fields of ecological study. However, use of eDNA results for decision‐making has been impeded by uncertainties associated with false positive tests putatively caused by sporadic or systemic contamination. Sporadic contamination is a process that is inconsistent across samples and systemic contamination occurs consistently over a group of samples. Here, we used empirical data and laboratory experiments to (i) estimate the sporadic contamination rate for each stage of a common, targeted eDNA workflow employing best practice quality control measures under simulated conditions of rare and common target DNA presence, (ii) determine the rate at which negative controls (i.e., “blanks”) detect varying concentrations of systemic contamination, and (iii) estimate the effort that would be required to consistently detect sporadic and systemic contamination. Sporadic contamination rates were very low across all eDNA workflow steps, and, therefore, an intractably high number of negative controls (>100) would be required to determine occurrence of sporadic contamination with any certainty. Contrarily, detection of intentionally introduced systemic contamination was more consistent; therefore, very few negative controls (<5) would be needed to consistently alert to systemic contamination. These results have considerable implications to eDNA study design when resources for sample analyses are constrained.

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Strategic considerations for invasive species managers in the utilization of environmental DNA (eDNA): steps for incorporating this powerful surveillance tool

Cited by 34 publications

References 43 publications

Detection of Khapra Beetle Environmental DNA Using Portable Technologies in Australian Biosecurity

Detection of Khapra Beetle Environmental DNA Using Portable Technologies in Australian Biosecurity

Shining a LAMP on the applications of isothermal amplification to monitoring illegal wildlife trade

Time to get real with qPCR controls: The frequency of sample contamination and the informative power of negative controls in environmental DNA studies

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