title: Development of a modified molecular diagnostic procedure for the identification and quantification of naturally occurring strongylid larvae on pastures.
abstract: A molecular procedure was developed to detect and quantify larvae of different strongylid parasite species recovered from pasture samples. Two lamb flocks (L and S) grazed separate paddocks with different natural larvae challenges (one low [Paddock L] and one high [Paddock S] challenge) on a commercial farm in Western Australia. Pasture samples were collected and analysed for larvae on 9 separate occasions from each paddock. Pregnant Merino ewes were sampled on 3 separate occasions (2 pre-partum and 1 post-partum). Following lambing, 203 female crossbred lambs were identified, from which faecal samples were collected across five separate samplings. Lamb production and faecal attributes were recorded. Genomic DNA was extracted directly from lamb faeces, in addition to the genomic DNA extracts from strongylid larval species recovered from pastures. Faecal worm egg counts (FWECs) were undertaken. Species-specific qPCRs and conventional PCRs (ITS-2 nuclear ribosomal DNA) were used to screen samples for strongylid species (Teladorsagia circumcincta, Trichostrongylus spp., Haemonchus contortus, Chabertia ovina and Oesophagostomum venulosum). Negative correlations (r(2)>0.91) were found between qPCR C(q) values and log-transformed pasture larval counts for Trichostrongylus spp. and T. circumcincta. Moderate levels of agreement between pasture larval counts and qPCR results were observed (67%). A clear difference in pasture larval challenge levels was observed between the two flocks using both qPCR and conventional pasture larval counts. It is difficult to draw conclusions on the production performances of lambs from the two experimental flocks, as no further replicates were able to be conducted following this experiment. Flock L had higher dressing percentages than Flock S (P=0.038), along with significantly higher faecal consistency and breech fleece faecal soiling scores at successive samplings. The molecular procedures utilised in this study have the potential to be beneficial for livestock grazing management strategies and parasite surveillance, however further investigation is necessary before they can become part of routine diagnostics.
authors.affiliation: School of Veterinary and Biomedical Sciences, Murdoch University, 90 South Street, Murdoch, Western Australia 6150, Australia. joshua.sweeny@agric.wa.gov.au
authors.forename: Joshua P A
authors.initials: JP
authors.lastname: Sweeny
authors.forename: Una M
authors.initials: UM
authors.lastname: Ryan
authors.forename: Ian D
authors.initials: ID
authors.lastname: Robertson
authors.forename: Dominic
authors.initials: D
authors.lastname: Niemeyer
authors.forename: Peter W
authors.initials: PW
authors.lastname: Hunt
mesh_terms: D000818:Animals; D018509:DNA, Helminth / Q000235:genetics / Q000302:isolation & purification; D005260:Female; D005838:Genotype; D007814:Larva / Q000145:classification; D016133:Polymerase Chain Reaction / Q000379:methods / Q000662:veterinary; D011247:Pregnancy; D012756:Sheep; D012757:Sheep Diseases / Q000469:parasitology*; D013045:Species Specificity; D017183:Strongylida / Q000145:classification* / Q000502:physiology*
chemical_list: D018509:DNA, Helminth