A new chromosome-level genome assembly and annotation of Cryptosporidium meleagridis

Abstract

Cryptosporidium spp. are medically and scientifically relevant protozoan parasites that cause severe diarrheal illness in infants, immunosuppressed populations and many animals. Although most human Cryptosporidium infections are caused by C. parvum and C. hominis, there are several other human-infecting species including C. meleagridis, which are commonly observed in developing countries. Here, we annotated a hybrid long-read Oxford Nanopore Technologies and short-read Illumina genome assembly for C. meleagridis (CmTU1867) with DNA generated using multiple displacement amplification. The assembly was then compared to the previous C. meleagridis (CmUKMEL1) assembly and annotation and a recent telomere-to-telomere C. parvum genome assembly. The chromosome-level assembly is 9.2 Mb with a contig N50 of 1.1 Mb. Annotation revealed 3,919 protein-encoding genes. A BUSCO analysis indicates a completeness of 96.6%. The new annotation contains 166 additional protein-encoding genes and reveals high synteny to C. parvum IOWA II (CpBGF). The new C. meleagridis genome assembly is nearly gap-free and provides a valuable new resource for the Cryptosporidium community and future studies on evolution and host-specificity.

Fig. 1

DNA synteny plot mapping the contigs of CmUKMEL1 to the eight chromosome-level contigs of CmTU1867. Jupiterplot between the previous CmUKMEL1 genome sequence and the new CmTU1867 genome sequence. Ribbons are colored with respect to the reference CmTU1867 chromosome.

Fig. 2

Protein synteny analysis of the eight chromosome-level contigs of CmTU1867 and Cryptosporidium parvum, CpBGF. Circos plot rings, moving from the center to the exterior illustrate shared ortholog clusters between CmTU1867 and CpBGF, number of base pairs in 50,000 bp increments, GC content histogram, and gene density. Locations of rRNA genes are as indicated.

Fig. 3

Portion of the 18SrRNA gene sequence and the putative ORF contained within it. Multiple sequence alignment of 18S rRNAs representing the guanine SNV and the putative ORFs contained in these sequences.

Fig. 4

Venn diagram of orthogroup search results following manual validation. Orthogroup comparison among the new CmTU1867, the previous CmUKMEL1, and the newly released reference genome, CpBGF. See Fig. 6 for the pre-validation results. Arrows link the list of gene IDs found in the smaller orthogroups that are unique to a species or shared by two species.

Fig. 5

Experimental workflow for genome sequencing, assembly, annotation, and validation. Bioinformatics workflow for assembly and annotation of the DNA derived from CmTU1867 WGA. Green boxes represent initial steps as well as new data used for parts of the pipeline and blue boxes represent subsequent downstream analyses of the data generated. The illustration was generated in BioRender.

Fig. 6

Ortholog search results shown in a Venn diagram. Orthogroup comparison among the new CmTU1867, the previous CmUKMEL1, and CpBGF prior to validation and correction. Arrows link the list of gene IDs found in the smaller orthogroups that are unique to a species or shared by two species.