Gregarine single-cell transcriptomics reveals differential mitochondrial remodeling and adaptation in apicomplexans

Abstract

Background Apicomplexa is a diverse phylum comprising unicellular endobiotic animal parasites and contains some of the most well-studied microbial eukaryotes including the devastating human pathogens Plasmodium falciparum and Cryptosporidium hominis. In contrast, data on the invertebrate-infecting gregarines remains sparse and their evolutionary relationship to other apicomplexans remains obscure. Most apicomplexans retain a highly modified plastid, while their mitochondria remain metabolically conserved. Cryptosporidium spp. inhabit an anaerobic host-gut environment and represent the known exception, having completely lost their plastid while retaining an extremely reduced mitochondrion that has lost its genome. Recent advances in single-cell sequencing have enabled the first broad genome-scale explorations of gregarines, providing evidence of differential plastid retention throughout the group. However, little is known about the retention and metabolic capacity of gregarine mitochondria. Results Here, we sequenced transcriptomes from five species of gregarines isolated from cockroaches. We combined these data with those from other apicomplexans, performed detailed phylogenomic analyses, and characterized their mitochondrial metabolism. Our results support the placement of Cryptosporidium as the earliest diverging lineage of apicomplexans, which impacts our interpretation of evolutionary events within the phylum. By mapping in silico predictions of core mitochondrial pathways onto our phylogeny, we identified convergently reduced mitochondria. These data show that the electron transport chain has been independently lost three times across the phylum, twice within gregarines. Conclusions Apicomplexan lineages show variable functional restructuring of mitochondrial metabolism that appears to have been driven by adaptations to parasitism and anaerobiosis. Our findings indicate that apicomplexans are rife with convergent adaptations, with shared features including morphology, energy metabolism, and intracellularity.

PhyloFisher: A phylogenomic package for resolving eukaryotic relationships

“…This is useful for single-cell data or to generate a single, better-sampled "hybrid taxon" representing several closely related taxa. Many of these provided utility scripts have been designed to act as "stand-alone" programs and can be used on datasets generated outside of the main PhyloFisher workflow as was done in [8].…”

“…After sequences had been collected by fisher.py for all organisms, the resulting files were subjected to the workflow described in the "Automated Filtering, Alignment, Trimming, and Single-protein Tree Construction" section. The resulting homolog trees were examined manually with the included tool "ParaSorter" to select orthologous sequences, demarcate paralogs, and remove contaminating sequences using the logic outlined in the PLOS BIOLOGY section "Phylogenomic dataset construction and analyses" of [8]. Finally, the aligned orthologs of the 304 public taxa (S1 Table ) were used to produce the final profile HMMs shipped with PhyloFisher v. 1.0 as well as the sequences of the 240 orthologs and their paralogs that comprise the starting database.…”

The Elusive Mitochondrial Genomes of Apicomplexa: Where Are We Now?

“…Finally, the cyrptosporidians include all Cryptosporidium species; C. hominis, C. parvum, C. muris, among others. Recently, transcriptomic data allowed the assessment of the divergence pattern involving coccidiomorphs, gregarines and cryptosporidians and results support the placement of Cryptosporidium as the earliest diverging lineage of apicomplexans (Mathur et al, 2021;Salomaki et al, 2021). However, the history of early apicomplexan radiation remains highly contentious (Janouškovec et al, 2019;Mathur et al, 2019Mathur et al, , 2021Salomaki et al, 2021), which impacts our interpretation of evolutionary events within the phylum.…”

“…It is thought that the abundance of host-derived nutrients combined with anaerobic/hypoxic growth conditions support the massive reduction in the mitochondrial metabolic capability in cryptosporidians (Gagat et al, 2017). Two additional independent mitochondrial functional reductions and electron chain losses within Apicomplexa have been recently shown to have occurred amongst gregarines (Mathur et al, 2021;Salomaki et al, 2021). While these mitochondrial reductions might be related to the hypoxic conditions found within the digestive tract of their respective hosts, further studies are needed to proper characterize these organelles.…”

Nutrient Acquisition and Attachment Strategies in Basal Lineages: A Tough Nut to Crack in the Evolutionary Puzzle of Apicomplexa

“…It was recently proposed that Cryptosporidium spp. be included in Gregarinasina as Cryptogregarinorida [ 1 ], but shortly thereafter the proposal was questioned [ 7 ] and later abandoned [ 37 ].…”