Genome assemblers are a critical component of genome science, but the choice of assembly software and protocols can be daunting. Here, we investigate genome assembly variation and its implications for gene discovery across three nematode species—Caenorhabditis bovis, Haemonchus contortus, and Heligmosomoides bakeri—highlighting the critical interplay between assembly choice and downstream genomic analysis. Selecting commonly used genome assemblers, we generated multiple assemblies for each species, analyzing their structure, completeness, and effect on gene family analysis. Our findings demonstrate that assembly variations can significantly affect gene family composition, with notable differences in gene families important in anthelmintic discovery and immunomodulation. Despite broadly similar performance using various assembly metrics, comparisons of assemblies with a single species revealed underlying structural rearrangements and inconsistencies in gene content, which would affect downstream analyses. This emphasizes the need for continuous refinement of genome assemblies and their annotations.

The assemblies were generated using different software.

Caenorhabditis bovis and Haemonchus contortus

1. Redbean2.5
2. Flye
3. Canu
4. SMARTdenovo
5. Falcon
6. Falcon-unzip

Heligmosomoides bakeri

1. Flye
2. HiCanu
3. Hifiasm

The gene models were generated using the software BRAKER3