Comparative genomics reveals a novel genetic organization of the sad cluster in the sulfonamide-degrader 'Candidatus Leucobacter sulfamidivorax' strain GP

BACKGROUND: Microbial communities recurrently establish metabolic associations resulting in increased fitness and ability to perform complex tasks, such as xenobiotic degradation. In a previous study, we have described a sulfonamide-degrading consortium consisting of a novel low-abundant actinobacterium, named strain GP, and Achromobacter denitrificans PR1. However, we found that strain GP was unable to grow independently and could not be further purified. RESULTS: Previous studies suggested that strain GP might represent a new putative species within the Leucobacter genus (16S rRNA gene similarity <¿97%). In this study, we found that average nucleotide identity (ANI) with other Leucobacter spp. ranged between 76.8 and 82.1%, further corroborating the affiliation of strain GP to a new provisional species. The average amino acid identity (AAI) and percentage of conserved genes (POCP) values were near the lower edge of the genus delimitation thresholds (65 and 55%, respectively). Phylogenetic analysis of core genes between strain GP and Leucobacter spp. corroborated these findings. Comparative genomic analysis indicates that strain GP may have lost genes related to tetrapyrrole biosynthesis and thiol transporters, both crucial for the correct assembly of cytochromes and aerobic growth. However, supplying exogenous heme and catalase was insufficient to abolish the dependent phenotype. The actinobacterium harbors at least two copies of a novel genetic element containing a sulfonamide monooxygenase (sadA) flanked by a single IS1380 family transposase. Additionally, two homologs of sadB (4-aminophenol monooxygenase) were identified in the metagenome-assembled draft genome of strain GP, but these were not located in the vicinity of sadA nor of mobile or integrative elements. CONCLUSIONS: Comparative genomics of the genus Leucobacter suggested the absence of some genes encoding for important metabolic traits in strain GP. Nevertheless, although media and culture conditions were tailored to supply its potential metabolic needs, these conditions were insufficient to isolate the PR1-dependent actinobacterium further. This study gives important insights regarding strain GP metabolism; however, gene expression and functional studies are necessary to characterize and further isolate strain GP. Based on our data, we propose to classify strain GP in a provisional new species within the genus Leucobacter, 'Candidatus Leucobacter sulfamidivorax'.This work was financially supported by (i) Swiss National Science Foundation (Grant No. 160332); (ii) project UID/EQU/00511/2019 by FCT/MCTES (PIDDAC); and (iii) project “LEPABE-2-ECO-INNOVATION” – NORTE-01-0145-FEDER-000005 by NORTE 2020 through the European Regional Development Fund (ERDF)