Evaluation of Next-Generation Amplicon Sequencing to Identify Eimeria spp. of Chickens

Genetic and biological characterisation of three cryptic Eimeria operational taxonomic units that infect chickens (Gallus gallus domesticus)

“…Subsequent locus-specific PCR and amplicon sequencing reiterated their cryptic status ( Cantacessi et al, 2008 , Morgan and Godwin, 2017 ), while molecular surveillance suggested an unexpectedly polarised global distribution ( Clark et al, 2016 ). Recent reports describing detection of all three OTU genotypes in the United States suggest a far wider prevalence ( Hauck et al, 2019 ), prompting renewed interest in their status.…”

“…First identified by capillary electrophoretic resolution of internal transcribed spacer (ITS)-2 PCR amplicons ( Morris et al, 2007 ), subsequent deep-amplicon sequencing of more isolates revealed 10 sequence types that resolved into three distinct phylogenetic clusters, termed OTUx, y, and z ( Cantacessi et al, 2008 ). Occurrence of these three OTU genotypes was initially thought to be restricted to the southern hemisphere ( Clark et al, 2016 , Jatau et al, 2016 ), but recent next-generation ITS-2 amplicon sequencing has detected all three in North America ( Hauck et al, 2019 ). While short amplicon sequence-based surveillance is informative, it is difficult to assess the true provenance of candidate new Eimeria spp.…”

Exploring Eimeria Genomes to Understand Population Biology: Recent Progress and Future Opportunities

“…These latter results indicate a requirement for additional validation and/or more specific primers. The second poultry study also targeted the Eimeria 18S rDNA, supplemented by COI, but followed a nested PCR approach using inner and outer primers, followed by MiniSeq NGS, in an attempt to increase specificity and sensitivity [ 27 ]. Application of the nested 18S and COI NGS assays to live anticoccidial vaccines and field samples collected from backyard and commercial chickens revealed best results for the 18S rDNA, confirmed by quantitative TaqMan PCR.…”

“…Application of the nested 18S and COI NGS assays to live anticoccidial vaccines and field samples collected from backyard and commercial chickens revealed best results for the 18S rDNA, confirmed by quantitative TaqMan PCR. Analysis of the 18S rDNA and COI datasets identified the presence of the cryptic Eimeria OTUs x, y and z in North America for the first time [ 27 ]. However, as for the publication by Hinsu et al [ 81 ], a series of additional sequences associated with non-chicken hosts were also detected including ferrets, rodents and rock partridge.…”

“…Preliminary phylogenetic inference suggests that each cryptic Eimeria OTU is related to, but distinct from, recognized Eimeria species with close affinities to E. maxima , E. brunetti and E. mitis , respectively [ 26 ]. Until recently the cryptic OTU genotypes appeared to be geographically restricted, primarily within the southern hemisphere [ 25 ], although recent studies have suggested a wider spatial occurrence [ 27 ]. Attempts to resolve questions around the heritage of these and other OTU Eimeria genotypes will benefit from recent and future advances in genomics and genetics.…”