The Actinobacteriota (previously Actinobacteria (Whitman et al., 2018) is a bacterial phylum comprising species characterized by gram-positive cell walls and high (≥55%) genomic G+C content (Nouioui et al., 2018). Actinobacteriota are globally distributed across both terrestrial and aquatic environments (Kurtbӧke, 2017) and have been found as symbionts of diverse eukaryotic organisms, from insects to humans (Kaltenpoth, 2009, Grice et al., 2009). Actinobacteriota have also commonly been cultivated from marine sponges (phylum Porifera), where they have been found to be a rich source of antibiotic and antitumor compounds (Sheikh et al., 2018).

The Actinobacteriota consists of eight characterized classes: Acidimicrobiia, Actinomycetia (previously Actinobacteria, (Salam et al., 2020), Aquicultoria, Coriobacteriia, Geothermincolia, Humimicrobiia, Rubrobacteria, and Thermoleophilia (Parks et al., 2022). Amongst sponge-associated Actinobacteriota, research attention has previously predominantly focused on the Actinomycetia due to evidence from cultivation studies that suggests they dominate the sponge-associated actinobacteriotal diversity (Selvin et al., 2009). However, 16S rRNA gene-based studies have rather found that Acidimicrobiia comprise much of the sponge-associated actinobacteriotal abundance and diversity, sometimes to the apparent exclusion of all other Actinobacteriota (Pires et al., 2018, Steinert et al., 2016, Cleary et al., 2015, Jeong et al., 2015, Ruiz et al., 2020, Souza et al., 2017).

Previous studies of Acidimicrobiia from non-sponge sources have found them to be particularly diverse in their ecological functions, metabolic pathways, and habitats (Norris and Class, 2012). In the Genome Taxonomy database (GTDB) (release 07-RS207), the order Acidimicrobiales contains three families, namely the Acidimicrobiaceae, Illumatobacteraceae, and Candidatus Microtrichaceae, with multiple cultured, characterized species (Parks et al., 2022). Species of the family Acidimicrobiaceae are often characterized as thermotolerant, acidophilic microorganisms capable of both oxidizing or reducing iron-containing compounds (Davis-Belmar and Norris, 2009, Jones and Johnson, 2015, Itoh et al., 2011, Johnson et al., 2009). Some are also capable of oxidizing sulfur-containing compounds, such as pyrite (Davis-Belmar and Norris, 2009, Johnson et al., 2009). In contrast, Ilumatobacteraceae species have generally been cultivated from marine environments and have been found to grow optimally at temperatures ranging from 10°C to 30°C and a pH of 6 to 10 (Matsumoto et al., 2013, Matsumoto et al., 2009). Ca. Microtrichaceae species are often cultivated from activated sludge in wastewater management plants (Muller et al., 2012). So far, all characterized Ca. Microtrichaceae species have been found to be organotrophic rather than lithotrophic (Muller et al., 2012).

The order Acidimicrobiales also contains in the GTDB three singleton cultured, functionally characterized species: Aquihabitans daechungensis, Rhabdothermincola sediminis, and Actinomarinicola tropica (Parks et al., 2022). Isolated from reservoir water, A. daechungensis was found to be a non-motile, halophobic chemoorganotroph capable of reducing nitrate to nitrite (Jin et al., 2013). R. sediminis was isolated from hot springs and found to have an optimal growth temperature of 45°C (Liu et al., 2021). Lastly, A. tropica was isolated from a marine sediment and is characterized as non-motile, halophilic, chemoorganotrophic, and like A. daechungensis, also capable of nitrate reduction (He et al., 2020). According to phylogenetic studies, A. tropica is closely related to both R. sediminis and Iamia majanohamensis, the latter is however not present in the GTDB (Liu et al., 2021, He et al., 2020). I. majanohamensis has been the only characterized acidimicrobiial species that has been isolated from a eukaryotic host (sea cucumber Holothuria edulis) and has been described as an aerobic chemoorganotroph, capable of assimilating diverse organic compounds and respiring nitrate to dinitrogen gas (Kurahashi et al., 2009).

Sponge-associated members of the class Acidimicrobiia however remain uncultured and uncharacterized, and thus little is known about their functional characteristics and taxonomic affiliation. Only recently did one study (Engelberts et al., 2020) identify the capacity for taurine catabolism and vitamin biosynthesis for Acidimicrobiia associated with the sponge Ircinia ramosa. Towards an in-depth exploration of the diversity and functions of uncultured sponge-associated Acidimicrobiia, we characterized and classified 22 acidimicrobiial metagenome-assembled genomes (MAGs) from three marine sponge species - Carteriospongia foliascens, Cymbastela coraliophila, and Rhopaloeides odorabile.