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Adaptation to life at different oxygen tensions plays a role in protozoan ecology and controls the distribution of different species in anoxic habitats. The ciliate genus  Spirostomum  inhabiting fresh or low salinity water globally where these species are considered as bioindicators. Under anaerobic or low oxygen conditions, the rhodoquinol-dependent pathway has been reported in the species from the class Heterotrichea. With the help of RNA sequencing (RNAseq) data,  Spirostomum  spp., are suitable for deep molecular investigations on  rquA  for rhodoquinone (RQ) biosynthesis. In this study,  Spirostomum ambiguum ,  Spirostomum subtilis , and  Spirostomum teres  collected from densely vegetated freshwater habitat in Fuzhou, China, explored the evidence of  rquA . Based on transcriptome analysis, two to three RquA proteins were identified in  S. ambiguum ,  S. teres , and  S. subtilis , respectively. The presence of a key Motif-I of RquA and mitochondrial targeting signals (MTS), also confirmed the identity of these as RquA. Furthermore,  Spirostomum  RquA proteins could be sorted into two groups based on their conserved amino acid (CAA) residues. Phylogenetic analysis also exhibited RquA division into two subclades contained RquA1 and RquA2/RquA3 and supports two to three paralogs of  rquA  genes in the genomes  Spirostomum  spp. Additional transcriptomes and genomes analysis of  Blepharisma  spp., and  Stentor  spp., respectively, also revealed at least two paralogs of  rquA  in members of the class Heterotrichea. The present study provides evidence for the presence of RquA and rhodoquinol dependent fumarate reduction pathway in  Spirostomum  species potentially use to respire in the oxygen-depleted habitats and two to three diverse  rquA  genes.

Transcriptome Profiling Revealed Multiple rquA Genes in the Species of Spirostomum (Protozoa: Ciliophora: Heterotrichea)