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Cross-contamination is a risk associated with short-term insect specimen storage

Published online by Cambridge University Press:  19 November 2020

Jennifer A. Zettler Open the ORCID record for Jennifer A. Zettler [Opens in a new window] ,
Brian Kohler ,
Traci Ness ,
Heather Buske  and
Ariel Joy Lipat
Jennifer A. Zettler*
Affiliation:
Department of Biology, Georgia Southern University, 11935 Abercorn Street, Savannah, Georgia, 31419, United States of America
Brian Kohler
Affiliation:
Department of Biology, Georgia Southern University, 11935 Abercorn Street, Savannah, Georgia, 31419, United States of America
Traci Ness
Affiliation:
Department of Biology, Georgia Southern University, 11935 Abercorn Street, Savannah, Georgia, 31419, United States of America
Heather Buske
Affiliation:
Department of Biology, Georgia Southern University, 11935 Abercorn Street, Savannah, Georgia, 31419, United States of America
Ariel Joy Lipat
Affiliation:
Department of Biology, Georgia Southern University, 11935 Abercorn Street, Savannah, Georgia, 31419, United States of America
*
*Corresponding author. Email: [email protected]
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Abstract

Molecular analyses of DNA samples necessitate careful attention to experimental protocols to prevent contamination of samples during the specimen collection and DNA extraction, amplification, and visualisation stages. When these studies involve insects that are collected in the field, standard procedures require samples to be individually sorted and stored in single tubes to prevent cross-contamination of DNA between specimens. This additional step can be time consuming and impractical when the survey population is large. The focus of our study was to experimentally determine if this assumed contamination risk is valid and if it varies with storage conditions. To test this, we co-stored a single intact fruit fly (Drosophilia melanogaster) (Diptera: Drosophilidae) known to be infected with the bacterial endosymbiont Wolbachia (Anaplasmataceae) with uninfected flies in tubes for three-hour, 24-hour, and one-week durations. For each treatment time, replicates of five tubes contained one infected fly and either one or five uninfected flies (1:1 and 1:5 ratios). Overall, eight of 30 tubes had contamination regardless of storage duration, and storing samples in crowded (1:5 ratio) conditions significantly increased the risk of Wolbachia DNA transference. Our results suggest that researchers should make efforts to sample and store specimens individually if cross-contamination is a concern in molecular analyses.

Type
Scientific Notes
Information
The Canadian Entomologist , Volume 153 , Issue 3 , June 2021 , pp. 368 - 373
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Copyright
© The Author(s), 2020. Published by Cambridge University Press on behalf of the Entomological Society of Canada

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Footnotes

Subject Editor: Dezene Huber

References

Bordenstein, S.R., Brothers, C., Wolfe, G., Bahr, M., Minckley, R.L., Clark, M.E., et al. 2010. Using the Wolbachia bacterial symbiont to teach inquiry-based science: a high school laboratory series. The American Biology Teacher, 72: 478–483.CrossRefGoogle Scholar
Dobson, S.L., Bourtzis, K., Braig, H.R., Jones, B.F., Zhou, W., Rousset, F., and O’Neill, S.L. 1999. Wolbachia infections are distributed throughout insect somatic and germ line tissues. Insect Biochemistry and Molecular Biology, 29: 153–160.CrossRefGoogle ScholarPubMed
Duploy, A., Vermenot, C., Davies, N., Roderick, G., Hurst, G.D., and Charlat, S. 2009. Assessing risks of Wolbachia DNA cross-specimen contamination following mass collection and ethanol storage. Molecular Ecology Resources, 9: 46–50.CrossRefGoogle Scholar
Dutra, H.L., Rocha, M.N., Dias, F.B., Mansur, S.B., Caragata, E.P., and Moreira, L.A. 2016. Wolbachia blocks currently circulating Zika virus isolates in Brazilian Aedes aegypti mosquitoes. Cell Host & Microbe, 19: 771–774.CrossRefGoogle ScholarPubMed
Frentiu, F.D., Zakir, T., Walker, T., Popovici, J., Pyke, A.T., van den Hurk, A., et al. 2014. Limited dengue virus replication in field-collected Aedes aegypti mosquitoes infected with Wolbachia. PLOS Neglected Tropical Diseases, 8: e2688. https://doi.org/10.1371/journal.pntd.0002688.CrossRefGoogle ScholarPubMed
Giordano, R., O’Neill, S.L., and Robertson, H.M. 1995. Wolbachia infections and the expression of cytoplasmic incompatibility in Drosophila sechellia and D. mauritiana. Genetics, 140: 1307–1317.CrossRefGoogle ScholarPubMed
Hertig, M. and Wolbach, S.B. 1924. Studies on Rickettsia-like micro-organisms in insects. The Journal of Medical Research, 44: 329–374.Google ScholarPubMed
Hertig, M. 1936. The Rickettsia, Wolbachia pipientis (gen. et sp.n.) and associated inclusions of the mosquito, Culex pipiens. Parasitology, 28: 453.CrossRefGoogle Scholar
Hilgenboecker, K., Hammerstein, P., Schlattmann, P., Telschow, A., and Werren, J.H. 2008. How many species are infected with Wolbachia? A statistical analysis of current data. FEMS Microbiology Letters, 281: 215–220.CrossRefGoogle ScholarPubMed
Mains, J.W., Kelly, P.H., Dobson, K.L., Petrie, W.D., and Dobson, S.L. 2019. Localized control of Aedes aegypti (Diptera: Culicidae) in Miami, FL, via inundative releases of Wolbachia-infected male mosquitoes. Journal of Medial Entomology, 56: 1296–1303.CrossRefGoogle ScholarPubMed
Martin, O.Y., Puniamoorthy, N., Gubler, A., Wimmer, C., Germann, C., and Bernasconi, M.V. 2013. Infections with the microbe Cardinium in the Dolichopodidae and other Empidoidea. Journal of Insect Science, 13: 47. https://doi.org/10.1673/031.013.4701.CrossRefGoogle ScholarPubMed
Mee, P.T., Weeks, A.R., Walker, P.J., Hoffmann, A.A., and Duchemin, J. 2015. Detection of low-level Cardinium and Wolbachia infections in Culicoides . Applied and Environmental Microbiology, 81: 6177–6188.CrossRefGoogle ScholarPubMed
Miller, W.J. and Riegler, M. 2006. Evolutionary dynamics of wAu-like Wolbachia variants in neotropical Drosophila spp. Applied and Environmental Microbiology, 72: 826–835.CrossRefGoogle ScholarPubMed
Nakamura, Y., Kawai, S., Yukuhiro, F., Ito, S., Gotoh, T., Kisimoto, R., et al. 2009. Prevalence of Cardinium bacteria in planthoppers and spider mites and taxonomic revision of “Candidatus Cardinium hertigii” based on detection of a new Cardinium group from biting midges. Applied and Environmental Microbiology, 75: 6757–6763.CrossRefGoogle ScholarPubMed
Phillips, K., McCallum, N., and Welch, L. 2012. A comparison of methods for forensic DNA extraction: Chelex-100 and the QIAGEN DNA Investigator Kit (manual and automated). Forensic Science International: Genetics, 6: 282–285.CrossRefGoogle Scholar
Sazama, E.J., Bosch, M.J., Shouldis, C.S., Ouellette, S.P., and Wesner, J.S. 2017. Incidence of Wolbachia in aquatic insects. Ecology and Evolution, 7: 1165–1169.CrossRefGoogle ScholarPubMed
Skaljac, M., Zanic, K., Ban, S.G., Kontsedalov, S., and Ghanim, M. 2010. Co-infection and localization of secondary symbionts in two whitefly species. BMC Microbiology, 10: 142 https://doi.org/10.1186/1471-2180-10-142.CrossRefGoogle ScholarPubMed
Weinert, L.A., Araujo-Jnr, E.V., Ahmed, M.Z., and Welch, J.J. 2015. The incidence of bacterial endosymbionts in terrestrial arthropods. Proceedings of the Royal Society B: Biological Sciences, 282: 1471–2954. https://doi.org/10.1098/rspb.2015.0249.Google ScholarPubMed
Werren, J.H. 1997. Biology of Wolbachia . Annual Review of Entomology, 42: 587–609.CrossRefGoogle ScholarPubMed
White, J.A., Kelly, S.E., Perlman, S.J., and Hunter, M.S. 2011. Endosymbiont costs and benefits in a parasitoid infected with both Wolbachia and Cardinium. Heredity, 106: 585–591.CrossRefGoogle Scholar
Willard, J.M., Lee, D.A., and Holland, M.M. 1998. Recovery of DNA for PCR amplification from blood and forensic samples using a chelating resin. Methods in Molecular Biology, 98: 9–18.Google ScholarPubMed
Wu, M., Sun, L.V., Vamathevan, J., Riegler, M., Deboy, R., Brownlie, J.C., et al. 2004. Phylogenomics of the reproductive parasite Wolbachia pipientis wMel: a streamlined genome overrun by mobile genetic elements. PLOS Biology, 2: e69. https://doi.org/10.1371/journal.pbio.0020069.CrossRefGoogle ScholarPubMed
Yen, J.H. and Barr, A.R. 1971. New hypothesis of the cause of cytoplasmic incompatibility in Culex pipiens L. Nature, 232: 657–658.CrossRefGoogle ScholarPubMed
Zug, R. and Hammerstein, P. 2012. Still a host of hosts for Wolbachia: analysis of recent data suggests that 40% of terrestrial arthropod species are infected. PLOS One, 7: e38544. https://doi.org/10.1371/journal.pone.0038544.CrossRefGoogle Scholar