A novel metabarcoded deep amplicon sequencing tool for disease surveillance and determining the species composition of Trypanosoma in cattle and other farm animals

Yasein, Ghulam; Zahid, Osama; Minter, Emily; Ashraf, Kamran; Rashid, Imran; Shabbir, Muhammad Zubair; Betson, Martha; Sargison, Neil; Chaudhry, Umer

doi:10.1016/j.actatropica.2022.106416

Cited by 4 publications

(4 citation statements)

References 61 publications

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“…to demonstrate species composition in coinfection of blood protozoa such as piroplasms (Theileria parva, T. annulate, Babesia bigemina, and Bartonella bovis) and Trypanosoma spp. (Trypanosoma brucei, T. theileri, T. congolense, and Tabanus vivax) (Chaudhry et al, 2019;Yasein et al, 2022). Further, because of deep amplicon sequencing ability to detect minor population parasites, this method was used to detect low-density Plasmodium falciparum infection (Early et al, 2019).…”

Section: Discussionmentioning

confidence: 99%

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New insight into genetic diversity of zoonotic‐potential Ancylostoma ceylanicum in stray cats living in Bangkok, Thailand, based on deep amplicon sequencing

Jitsamai,

Kamkong,

Traub

et al. 2023

Zoonoses and Public Health

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AimsThis study aimed to characterize feline hookworms from stray cats living in Bangkok.Methods and ResultsA total of 56 hookworm‐positive faecal samples were identified for hookworm species by using PCR targeting the ITS1, 5.8S, and ITS2 fragment and qPCR targeting ITS2. Of 56 samples, 96.4% (54/56) were identified as Ancylostoma ceylanicum and 1.8% (1/56) as Ancylostoma caninum. With qPCR, 89.3% (50/56) were identified as single A. ceylanicum infection and 5.4% (3/56) as coinfection of A. ceylanicum and A. caninum. For genetic characterization of A. ceylanicum, 10 samples were pooled, and the partial COI gene was amplified, followed by deep amplicon sequencing. Five pooled samples were analysed, and 99.73% were identified with A. ceylanicum sequences, which were allocated into 19 haplotypes (AC01‐AC19). Genetic diversity findings for A. ceylanicum in Asia revealed that three of eight haplotypes considered of zoonotic significance occurred in humans, dogs, and cats, including haplotypes H01, H20, and H21. The predominant haplotype in this study, AC01, was clustered with H01‐a zoonotic haplotype.ConclusionsThe diversity obtained by deep amplicon sequencing supported that the A. ceylanicum community had high genetic variation. Deep amplicon sequencing was a useful method to determine source, zoonotic potential, and host–parasite relationship.

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Section: Discussionmentioning

confidence: 99%

“…theileri , T . congolense , and Tabanus vivax ) (Chaudhry et al., 2019; Yasein et al., 2022). Further, because of deep amplicon sequencing ability to detect minor population parasites, this method was used to detect low‐density Plasmodium falciparum infection (Early et al., 2019).…”

Section: Discussionmentioning

confidence: 99%

New insight into genetic diversity of zoonotic‐potential Ancylostoma ceylanicum in stray cats living in Bangkok, Thailand, based on deep amplicon sequencing

Jitsamai,

Kamkong,

Traub

et al. 2023

Zoonoses and Public Health

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“…High throughput deep amplicon sequencing using the Illumina Mi-Seq platform is relatively low-cost and potentially less error-prone [35]. The method has transformed the study of veterinary and human haemoprotozoa [17][18][19], and clade V parasitic nematodes [36][37][38], and has the potential to improve surveillance of Dicrocoelium species as previously demonstrated by the concept of a 'tremobiome' for the quantification of the Fasciola and Calicophoron species [25,26,39].…”

Section: Plos Onementioning

confidence: 99%

“…The sensitivity and specificity of molecular approaches have improved in recent years. Still, the reliability of infection reports varies considerably between affected areas because of the different molecular-based methods used [ 19 ]. Traditional PCR and Sanger sequencing methods amplify fragments of nuclear ribosomal genes, and their internal transcribed spacers have been developed for dicrocoeliid parasites [ 6 ].…”

Section: Introductionmentioning

confidence: 99%

Validation of deep amplicon sequencing of Dicrocoelium in small ruminants from Northern regions of Pakistan

Khan,

Afshan,

Firasat

et al. 2024

PLoS ONE

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Dicrocoelium lancet flukes cause significant production loss in ruminant livestock. Although co-infection with multiple Dicrocoelium species within a host is common, techniques for studying the composition of these complex parasite communities are lacking. The pathogenicity, epidemiology, and therapeutic susceptibility of different helminth species vary, and little is known about the interactions that take place between co-infecting species and their hosts. Here, we describe the first applicationof metabarcoding deep amplicon sequencing method to studythe Dicrocoelium species in sheep and goats. First, rDNA ITS-2 sequences of four Dicrocoelium species (Dicrocoelium dendriticum, Dicrocoelium hospes, Dicrocoelium orientalis, and Dicrocoelium chinensis) were extracted from the NCBI public database. Phylogenetic analysis revealed separate clades of Dicrocoelium species; hence, molecular differentiation between each species is possible in co-infections. Second, 202 flukes belonging to seventeen host populations (morphologically verified as belonging to the Dicrocoelium genus) were evaluated to determine the deep amplicon sequencing read threshold of an individual fluke for each of the four species. The accuracy of the method in proportional quantification of samples collected from single hosts was further assessed. Overall, 198 (98.01%) flukes were confirmed as D. dendriticum and 1.98% produced no reads. The comparison of genetic distances between rDNA ITS-2 revealed 86% to 98% identity between the Dicrocoelium species. Phylogenetic analysis demonstrated a distinct clustering of species, apart from D. orientalis and D. chinensis, which sit very close to each other in a single large clade whereas D. hospes and D. dendriticum are separated into their own clade. In conclusion each sample was identified as D. dendriticum based on the proportion of MiSeq reads and validated the presence of this group of parasites in the Gilgit Baltistan and Khyber Pakhtunkhwa provinces of Pakistan. The metabarcoding deep amplicon sequencing technology and bioinformatics pathway have several potential applications, including species interactions during co-infections, identifying the host and geographical distribution of Dicrocoelium in livestock, drug therapy response evaluation and understanding of the emergence and spread of drug resistance.

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A Method for Isolating and Sequencing Trypanosome Cells to Investigate Species Associations in Multiple Morphotype Infections

Shannon,

Bolek

2023

Journal of Parasitology

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A novel metabarcoded deep amplicon sequencing tool for disease surveillance and determining the species composition of Trypanosoma in cattle and other farm animals

Cited by 4 publications

References 61 publications

New insight into genetic diversity of zoonotic‐potential Ancylostoma ceylanicum in stray cats living in Bangkok, Thailand, based on deep amplicon sequencing

New insight into genetic diversity of zoonotic‐potential Ancylostoma ceylanicum in stray cats living in Bangkok, Thailand, based on deep amplicon sequencing

Validation of deep amplicon sequencing of Dicrocoelium in small ruminants from Northern regions of Pakistan

A Method for Isolating and Sequencing Trypanosome Cells to Investigate Species Associations in Multiple Morphotype Infections

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