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Double trouble: untangling mixed sequence signals in bird samples with avian haemosporidian co-infections

Published online by Cambridge University Press:  04 March 2022

Huiqing Yeo Open the ORCID record for Huiqing Yeo [Opens in a new window] ,
Denise Nastaya Harjoko  and
Frank E. Rheindt
Huiqing Yeo
Affiliation:
Department of Biological Sciences, National University of Singapore, 16 Science Drive 4, Block S3 Level 4, 117558 Singapore, Singapore
Denise Nastaya Harjoko
Affiliation:
Department of Biological Sciences, National University of Singapore, 16 Science Drive 4, Block S3 Level 4, 117558 Singapore, Singapore
Frank E. Rheindt*
Affiliation:
Department of Biological Sciences, National University of Singapore, 16 Science Drive 4, Block S3 Level 4, 117558 Singapore, Singapore
*
Author for correspondence: Frank E. Rheindt, E-mail: [email protected]
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Abstract

Blood parasites comprise some of the most prevalent pathogens in nature, and their detection and identification are major objectives in varied fields such as ecology and biomedicine. Two approaches were compared, one based on Sanger sequencing and the other next-generation sequencing (NGS) based, in terms of their performance in detecting avian blood parasites across tropical Southeast Asian birds. Across a panel of 528 bird individuals, 43 birds were ascertained to be infected with avian haemosporidians using a polymerase chain reaction-based detection method. Among these samples, NGS-based barcoding confirmed co-infections by multiple blood parasites in all eight cases where Sanger sequencing produced double peaks. Importantly however, the NGS-based method produced another five diagnoses of co-infections (62.5%) in which Sanger-based barcoding remained equivocal. In contrast to Sanger sequencing, the NGS-based method was able to identify co-infecting haemosporidian lineages via their barcodes. The accuracy of avian haemosporidian lineage identification was not compromised by the shorter length of NGS sequences, with ~94% of NGS barcodes producing matches identical to those of the Sanger barcodes. The application of NGS-based barcoding methods promises to enhance parasite identification and reduce erroneous inferences based on artefacts.

Keywords

Barcodingblood parasitesmixed infectionsnext-generation sequencing
Type
Research Article
Information
Parasitology , Volume 149 , Issue 6 , May 2022 , pp. 799 - 810
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Copyright
Copyright © The Author(s), 2022. Published by Cambridge University Press

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References

Altschul, SF, Gish, W, Miller, W, Myers, EW and Lipman, DJ (1990) Basic local alignment search tool. Journal of Molecular Biology 215, 403–410.CrossRefGoogle ScholarPubMed
Andrews, S (2010) FastQC: a quality control tool for high throughput sequence data [Online]. Available at Babraham Bioinformatics website http://www.bioinformatics.babraham.ac.uk/projects/fastqc/ (Accessed 16 December 2021).Google Scholar
Asghar, M, Hasselquist, D and Bensch, S (2011) Are chronic avian haemosporidian infections costly in wild birds? Journal of Avian Biology 42, 530–537.CrossRefGoogle Scholar
Bensch, S, Stjernman, M, Hasselquist, D, Örjan, Ö, Hannson, B, Westerdahl, H and Pinheiro, RT (2000) Host specificity in avian blood parasites: a study of Plasmodium and Haemoproteus mitochondrial DNA amplified from birds. Proceedings of the Royal Society of London B, Biological Sciences 267, 1583–1589.CrossRefGoogle ScholarPubMed
Bensch, S, Waldenström, J, Jonzén, N, Westerdahl, H, Hansson, B, Sejberg, D and Hasselquist, D (2007) Temporal dynamics and diversity of avian malaria parasites in a single host species. Journal of Animal Ecology 76, 112–122.CrossRefGoogle Scholar
Bensch, S, Hellgren, O and Pérez-Tris, J (2009) MalAvi: a public database of malaria parasites and related haemosporidians in avian hosts based on mitochondrial cytochrome b lineages. Molecular Ecology Resources 9, 1353–1358.CrossRefGoogle ScholarPubMed
Bernotienė, R, Palinauskas, V, Iezhova, T, Murauskaitė, D and Valkiūnas, G (2016) Avian haemosporidian parasites (Haemosporida): a comparative analysis of different polymerase chain reaction assays in detection of mixed infections. Experimental Parasitology 163, 31–37.CrossRefGoogle ScholarPubMed
Boyer, F, Mercier, C, Bonin, A, Le Bras, Y, Taberlet, P and Coissac, E (2016) obitools: a unix-inspired software package for DNA metabarcoding. Molecular Ecology Resources 16, 176–182.CrossRefGoogle ScholarPubMed
Callahan, BJ, McMurdie, PJ, Rosen, MJ, Han, HW, Johnson, AJ and Holmes, SP (2016) DADA2: high-resolution sample inference from Illumina amplicon data. Nature Methods 13, 581–583.CrossRefGoogle ScholarPubMed
Ciloglu, A, Ellis, VA, Bernotienė, R, Valkiūnas, G and Bensch, S (2019) A new one-step multiplex PCR assay for simultaneous detection and identification of avian haemosporidian parasites. Parasitology Research 118, 191–201.CrossRefGoogle ScholarPubMed
Dmitriev, DA and Rakitov, RA (2008) Decoding of superimposed traces produced by direct sequencing of heterozygous indels. PLoS Computational Biology 4, e1000113.CrossRefGoogle ScholarPubMed
Edgar, RC (2010) Search and clustering orders of magnitude faster than BLAST. Bioinformatics (Oxford, England) 26, 2460–2461.CrossRefGoogle ScholarPubMed
Fecchio, A, Collins, MD, Bell, JA, García-Trejo, EA, Sánchez-González, LA, Dispoto, JH, Rice, NH and Weckstein, JD (2019) Bird tissues from museum collections are reliable for assessing avian haemosporidian diversity. The Journal of Parasitology 105, 446–453.CrossRefGoogle ScholarPubMed
Françoso, E and Arias, MC (2013) Cytochrome c oxidase I primers for corbiculate bees: DNA barcode and mini-barcode. Molecular Ecology Resources 13, 844–850.CrossRefGoogle ScholarPubMed
Galen, SC, Nunes, R, Sweet, PR and Perkins, SL (2018) Integrating coalescent species delimitation with analysis of host specificity reveals extensive cryptic diversity despite minimal mitochondrial divergence in the malaria parasite genus Leucocytozoon. BMC Evolutionary Biology 18, 128.CrossRefGoogle ScholarPubMed
Hellgren, O, Waldenström, J and Bensch, S (2004) A new PCR assay for simultaneous studies of Leucocytozoon, Plasmodium, and Haemoproteus from avian blood. Journal of Parasitology 90, 797–802.CrossRefGoogle ScholarPubMed
Hellgren, O, Atkinson, CT, Bensch, S, Albayrak, T, Dimitrov, D, Ewen, JG, Kim, KS, Lima, MR, Martin, L, Palinauskas, V, Ricklefs, R, Sehgal, RNM, Valkiūnas, G, Tsuda, Y and Marzal, A (2014) Global phylogeography of the avian malaria pathogen Plasmodium relictum based on MSP1 allelic diversity. Ecography 38, 842–850.CrossRefGoogle Scholar
Katoh, K and Standley, DM (2013) MAFFT multiple sequence alignment software version 7: improvements in performance and usability. Molecular Biology and Evolution 30, 772–780.CrossRefGoogle ScholarPubMed
Larsson, A (2014) AliView: a fast and lightweight alignment viewer and editor for large datasets. Bioinformatics (Oxford, England) 30, 3276–3278.CrossRefGoogle ScholarPubMed
Little, DP (2013) A DNA mini-barcode for land plants. Molecular Ecology Resources 14, 437–446.CrossRefGoogle ScholarPubMed
Martin, M (2011) Cutadapt removes adapter sequences from high-throughput sequencing reads. EMBnet Journal 17, 10–12.CrossRefGoogle Scholar
Martinsen, ES, Paperna, I and Schall, JJ (2006) Morphological versus molecular identification of avian Haemosporidia: an exploration of three species concepts. Parasitology 133, 279–288.CrossRefGoogle ScholarPubMed
Marzal, A, Bensch, S, Reviriego, M, Balbontin, J and De Lope, F (2008) Effects of malaria double infection in birds: one plus one is not two. Journal of Evolutionary Biology 21, 979–987.CrossRefGoogle Scholar
Meier, R, Wong, W, Srivathsan, A and Foo, M (2016) $1 DNA barcodes for reconstructing complex phenomes and finding rare species in specimen-rich samples. Cladistics 32, 100–110.CrossRefGoogle ScholarPubMed
Merino, S, Moreno, J, José Sanz, J and Arriero, E (2000) Are avian blood parasites pathogenic in the wild? A medication experiment in blue tits. Proceedings of the Royal Society B 267, 2507–2510.CrossRefGoogle Scholar
Meusnier, I, Singer, GAC, Landry, J, Hickey, DA, Hebert, PDN and Mehrdad, H (2008) A universal DNA mini-barcode for biodiversity analysis. BMC Genomics 9, 214.CrossRefGoogle ScholarPubMed
Pacheco, MA, Cepeda, AS, Bernotienė, R, Lotta, IA, Matta, NE, Valkiūnas, G and Escalante, AA (2018 a) Primers targeting mitochondrial genes of avian haemosporidians: PCR detection and differential DNA amplification of parasites belonging to different genera. International Journal for Parasitology 48, 657–670.CrossRefGoogle ScholarPubMed
Pacheco, MA, Matta, NE, Valkiūnas, G, Parker, PG, Mello, B, Stanley, CE Jr., Lentino, M, Garcia-Amado, MA, Cranfield, M, Pond, SLK and Escalante, AA (2018 b) Mode and rate of evolution of haemosporidian mitochondrial genomes: timing the radiation of avian parasites. Molecular Biology and Evolution 35, 383–403.CrossRefGoogle ScholarPubMed
Palinauskas, V, Valkiūnas, G, Bolshakov, CV and Bensch, S (2008) Plasmodium relictum (lineage P-SGS1): effects on experimentally infected passerine birds. Experimental Parasitology 120, 372–380.CrossRefGoogle ScholarPubMed
Palinauskas, V, Bernotienė, R, Žiegytė, R, Bensch, S and Valkiūnas, G (2017) Experimental evidence for hybridization of closely related lineages in Plasmodium relictum. Molecular and Biochemical Parasitology 217, 1–6.CrossRefGoogle ScholarPubMed
Palinauskas, V, Žiegytė, R, Šengaut, J and Bernotienė, R (2018) Different paths – the same virulence: experimental study on avian single and co-infections with Plasmodium relictum and Plasmodium elongatum. International Journal for Parasitology 48, 1089–1096.CrossRefGoogle Scholar
Pérez-Tris, J and Bensch, S (2005) Diagnosing genetically diverse avian malarial infections using mixed-sequence analysis and TA-cloning. Parasitology 131, 15–23.CrossRefGoogle ScholarPubMed
Rheindt, FE, Gwee, CY, Baveja, P, Ferasyi, TR, Nurza, A, Rosa, TS and Haminuddin, (2020) A taxonomic and conservation re-appraisal of all the birds on the island of Nias. Raffles Bulletin of Zoology 68, 496–528.Google Scholar
Ricklefs, RE, Outlaw, DC, Svensson-Coelho, M, Medeiros, MCI, Ellis, VA and Latta, S (2014) Species formation by host shifting in avian malaria parasites. Proceedings of the National Academy of Sciences 111, 14816–14821.CrossRefGoogle ScholarPubMed
Sadanandan, KR, Tan, DJX, Schjølberg, K, Round, PD and Rheindt, FE (2015) DNA reveals long-distance partial migratory behavior in a cryptic owl lineage. Avian Research 6, 25.CrossRefGoogle Scholar
Santiago-Alarcon, D, Palinauskas, V and Schaefer, HM (2012) Diptera vectors of avian haemosporidian parasites: untangling parasite life cycles and their taxonomy. Biological Reviews 87, 928–964.CrossRefGoogle ScholarPubMed
Sehgal, RNM (2015) Manifold habitat effects on the prevalence and diversity of avian blood parasites. International Journal for Parasitology: Parasites and Wildlife 4, 421–430.Google ScholarPubMed
Silva-Iturriza, A, Ketmaier, V and Tiedemann, R (2012) Prevalence of avian haemosporidian parasites and their host fidelity in the central Philippine islands. Parasitology International 61, 650–657.CrossRefGoogle ScholarPubMed
Tamura, K, Stecher, G and Kumar, S (2021) MEGA11: molecular evolutionary genetics analysis version 11. Molecular Biology and Evolution 38, 3022–3027.CrossRefGoogle ScholarPubMed
Tenney, AE, Wu, JQ, Langton, L, Klueh, P, Quatrano, R and Brent, MR (2007) A tale of two templates: automatically resolving double traces has many applications, including efficient PCR-based elucidation of alternative splices. Genome Research 17, 212–218.CrossRefGoogle ScholarPubMed
Valkiūnas, G (2005) Avian Malaria Parasites and Other Haemosporidia. Boca Rotan: CRC Press. Available at https://doi.org/10.1201/9780203643792.Google Scholar
Valkiūnas, G, Kazlauskienė, R, Bernotienė, R, Bukauskaitė, D, Palinauskas, V and Iezhova, TA (2014) Haemoproteus infections (Haemosporida, Haemoproteidae) kill bird-biting mosquitoes. Parasitology Research 113, 1011–1018.CrossRefGoogle ScholarPubMed
Xuan, MNT, Kaewlamun, W, Saiwichai, T, Thanee, S, Poofery, J, Tiawsirisup, S, Channumsin, M and Kaewthamasorn, M (2021) Development and application of a novel multiplex PCR assay for the differentiation of four haemosporidian parasites in the chicken Gallus gallus domesticus. Veterinary Parasitology 293, 109431.CrossRefGoogle ScholarPubMed
Yeo, D, Srivathsan, A and Meier, R (2020) Longer is not always better: optimising barcode length for large-scale species discovery and identification. Systematic Biology 69, 999–1015.CrossRefGoogle ScholarPubMed