The Genome of Caenorhabditis bovis.

Lewis Stevens,Mark Blaxter,Maurice K Murungi,Kelvin Momanyi,Joseph Ogola,Christian O Odinga,Eunice M Machuka,Eric M Fèvre,Stefan Rooke,Allan Ogendo,Laura C Falzon,Samuel M Njoroge

doi:10.1016/j.cub.2020.01.074

Abstract

The free-living nematode Caenorhabditis elegans is akey laboratory model for metazoan biology. C.elegans has also become a model for parasitic nematodes despite being only distantly related to most parasitic species. All of the ∼65 Caenorhabditis species currently in culture are free-living, with most having been isolated from decaying plant or fungal matter. Caenorhabditis bovis is a particularly unusual species that has been isolated several times from the inflamed ears of Zebu cattle in Eastern Africa, where it is associated with thedisease bovine parasitic otitis. C.bovis is therefore of particular interest to researchers interested in the evolution of nematode parasitism. However, as C.bovis is not in laboratory culture, it remains little studied. Here, by sampling livestock markets and slaughterhouses in Western Kenya, we successfully reisolated C.bovis from the ear of adult female Zebu. We sequenced the genome of C.bovis using the Oxford Nanopore MinION platform in a nearby field laboratory and used the data to generate a chromosome-scaledraft genome sequence. We exploited this draft genome sequence to reconstruct the phylogenetic relationships of C.bovis to other Caenorhabditis species and reveal the changes in genome size and content that have occurred during its evolution. We also identified expansions in several gene families that have been implicated in parasitism in other nematode species. The high-quality draft genome and our analyses thereof represent a significant advancement in our understanding of this unusual Caenorhabditis species.

Highlights

In collaboration with local veterinarians and scientists, we sampled cattle at livestock markets and slaughterhouses i
We sequenced the genome of C. bovis in a nearby field laboratory using the Oxford Nanopore MinION platform and used the data to generate a high-quality, chromosome-scale draft genome sequence
Reisolation of C. bovis We sampled a total of 44 cattle of various ages and breeds at livestock markets and slaughterhouses in three counties in Western Kenya (Figure 1A; Table S1)

Summary

Methods

DNA extraction We harvested nematodes by washing each plate with phosphate-buffered saline (PBS) supplemented with 0.01% Tween. The nematodes were washed three times with clean PBS and subsequently centrifuged to form a pellet. We added 600 mL of Cell Lysis Solution (QIAGEN) and 20 mL of proteinase K (20 mg/mL) to each frozen pellet and incubated for four h at 56C. We added 200 ml of Protein Precipitation Solution (QIAGEN) and centrifuged at 15,000 rpm for 3 min. The supernatant was collected in a new tube and 600 mL of isopropanol added to precipitate the DNA. We centrifuged each tube at 15,000 rpm for 3 min and discarded the supernatant. The resulting DNA pellets were washed twice with 70% ethanol and briefly allowed to dry before being resuspended in 100 mL of elution buffer (10 mM Tris-Cl).

Results

Discussion

Conclusion