title: Extracellular Onchocerca-derived small RNAs in host nodules and blood.
abstract: BACKGROUND
  microRNAs (miRNAs), a class of short, non-coding RNA can be found in a highly stable, cell-free form in mammalian body fluids. Specific miRNAs are secreted by parasitic nematodes in exosomes and have been detected in the serum of murine and dog hosts infected with the filarial nematodes Litomosoides sigmodontis and Dirofilaria immitis, respectively. Here we identify extracellular, parasite-derived small RNAs associated with Onchocerca species infecting cattle and humans.
  
  
  METHODS
  Small RNA libraries were prepared from total RNA extracted from the nodule fluid of cattle infected with Onchocerca ochengi as well as serum and plasma from humans infected with Onchocerca volvulus in Cameroon and Ghana. Parasite-derived miRNAs were identified based on the criteria that sequences unambiguously map to hairpin structures in Onchocerca genomes, do not align to the human genome and are not present in European control serum.
  
  
  RESULTS
  A total of 62 mature miRNAs from 52 distinct pre-miRNA candidates were identified in nodule fluid from cattle infected with O. ochengi of which 59 are identical in the genome of the human parasite O. volvulus. Six of the extracellular miRNAs were also identified in sequencing analyses of serum and plasma from humans infected with O. volvulus. Based on sequencing analysis the abundance levels of the parasite miRNAs in serum or plasma range from 5 to 127 reads/per million total host miRNA reads identified, comparable to our previous analyses of Schistosoma mansoni and L. sigmodontis miRNAs in serum. All six of the O. volvulus miRNAs identified have orthologs in other filarial nematodes and four were identified in the serum of mice infected with L. sigmodontis.
  
  
  CONCLUSIONS
  We have identified parasite-derived miRNAs associated with onchocerciasis in cattle and humans. Our results confirm the conserved nature of RNA secretion by diverse nematodes. Additional species-specific small RNAs from O. volvulus may be present in serum based on the novel miRNA sequences identified in the nodule fluid. In our analyses comparison to European control serum illuminates the scope for false-positives, warranting caution in criteria that should be applied to identification of biomarkers of infection.
authors.affiliation: Centre for Immunity, Infection and Evolution, Ashworth Laboratories, University of Edinburgh, West Mains Road, Edinburgh, UK. jquintan@staffmail.ed.ac.uk.
authors.forename: Juan F
authors.initials: JF
authors.lastname: Quintana
authors.affiliation: Institute of Infection and Global Health, University of Liverpool, Liverpool, Merseyside, UK. Blm1@liverpool.ac.uk.
authors.forename: Benjamin L
authors.initials: BL
authors.lastname: Makepeace
authors.affiliation: Institute of Biodiversity, Animal Health and Comparative Medicine, College of Medical, Veterinary and Life Sciences, University of Glasgow, Glasgow, UK. Simon.Babayan@glasgow.ac.uk.
authors.forename: Simon A
authors.initials: SA
authors.lastname: Babayan
authors.affiliation: Centre for Immunity, Infection and Evolution, Ashworth Laboratories, University of Edinburgh, West Mains Road, Edinburgh, UK. al.ivens@ed.ac.uk.
authors.forename: Alasdair
authors.initials: A
authors.lastname: Ivens
authors.affiliation: Institute of Medical Microbiology, Immunology and Parasitology, University Hospital of Bonn, Bonn, Germany. pfarr@microbiology-bonn.de.
authors.forename: Kenneth M
authors.initials: KM
authors.lastname: Pfarr
authors.affiliation: Centre for Immunity, Infection and Evolution, Ashworth Laboratories, University of Edinburgh, West Mains Road, Edinburgh, UK. mark.blaxter@ed.ac.uk.
authors.forename: Mark
authors.initials: M
authors.lastname: Blaxter
authors.affiliation: Kumasi Centre for Collaborative Research (KCCR) and Kwame Nkrumah University of Science and Technology, Kumasi, Ghana. yadebrah@yahoo.com.
authors.forename: Alexander
authors.initials: A
authors.lastname: Debrah
authors.affiliation: Research Foundation in Tropical Diseases and Environment and University of Buea, Buea, Cameroon. swanji@yahoo.fr.
authors.forename: Samuel
authors.initials: S
authors.lastname: Wanji
authors.affiliation: Institut de Recherche Agricole pour le Développement, Regional Centre of Wakwa, Ngaoundéré, Cameroon. ng_henrietta@yahoo.com.
authors.forename: Henrietta F
authors.initials: HF
authors.lastname: Ngangyung
authors.affiliation: Institut de Recherche Agricole pour le Développement, Regional Centre of Wakwa, Ngaoundéré, Cameroon. bahsohg2002@yahoo.com.
authors.forename: Germanus S
authors.initials: GS
authors.lastname: Bah
authors.affiliation: Cameroon Academy of Sciences, Yaoundé, Cameroon. vntanya@yahoo.com.
authors.forename: Vincent N
authors.initials: VN
authors.lastname: Tanya
authors.affiliation: Institute of Infection and Global Health, University of Liverpool, Liverpool, Merseyside, UK. David.W.Taylor@ed.ac.uk.|Division of Pathway Medicine, School for Biomedical Sciences, University of Edinburgh, Little France, Edinburgh, UK. David.W.Taylor@ed.ac.uk.
authors.forename: David W
authors.initials: DW
authors.lastname: Taylor
authors.affiliation: Institute of Medical Microbiology, Immunology and Parasitology, University Hospital of Bonn, Bonn, Germany. hoerauf@microbiology-bonn.de.
authors.forename: Achim
authors.initials: A
authors.lastname: Hoerauf
authors.affiliation: Centre for Immunity, Infection and Evolution, Ashworth Laboratories, University of Edinburgh, West Mains Road, Edinburgh, UK. a.buck@ed.ac.uk.
authors.forename: Amy H
authors.initials: AH
authors.lastname: Buck
mesh_terms: D000818:Animals; D001483:Base Sequence; D001826:Body Fluids / Q000469:parasitology*; D002417:Cattle; D002418:Cattle Diseases / Q000097:blood / Q000469:parasitology*; D006801:Humans; D035683:MicroRNAs / Q000097:blood* / Q000235:genetics; D008969:Molecular Sequence Data; D009854:Onchocerca / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D009855:Onchocerciasis / Q000097:blood / Q000469:parasitology* / Q000662:veterinary*; D018510:RNA, Helminth / Q000097:blood* / Q000235:genetics
chemical_list: D035683:MicroRNAs; D018510:RNA, Helminth