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Background  In recent years, interest in  Bacillus velezensis  has increased significantly due to its role in many industrial water bioremediation processes. In this study, we isolated and assessed the transcriptome of  Bacillus velezensis  LG37 (from an aquaculture pond) under different nitrogen sources. Since  Bacillus  species exhibit heterogeneity, it is worth investigating the molecular mechanism of LG37 through ammonia nitrogen assimilation, where nitrogen in the form of molecular ammonia is considered toxic to aquatic organisms.    Results  Here, a total of 812 differentially expressed genes (DEGs) from the transcriptomic sequencing of LG37 grown in minimal medium supplemented with ammonia (treatment) or glutamine (control) were obtained, from which 56 had Fold Change ≥2. BLAST-NCBI and UniProt databases revealed 27 out of the 56 DEGs were potentially involved in NH 4  +  assimilation. Among them, 8 DEGs together with the two-component regulatory system GlnK/GlnL were randomly selected for validation by quantitative real-time RT-PCR, and the results showed that expression of all the 8 DEGs are consistent with the RNA-seq data. Moreover, the transcriptome and relative expression analysis were consistent with the transporter gene  amtB  and it is not involved in ammonia transport, even in the highest ammonia concentrations. Besides, CRISPR-Cas9 knockout and overexpression  glnK  mutants further evidenced the exclusion of  amtB  regulation, suggesting the involvement of alternative transporter. Additionally, in the transcriptomic data, a novel ammonium transporter  mnrA  was expressed significantly in increased ammonia concentrations. Subsequently, OE mnrA  and Δ mnrA  LG37 strains showed unique expression pattern of specific genes compared to that of wild-LG37 strain.    Conclusion  Based on the transcriptome data, regulation of nitrogen related genes was determined in the newly isolated LG37 strain to analyse the key regulating factors during ammonia assimilation. Using genomics tools, the novel MnrA transporter of LG37 became apparent in ammonia transport instead of AmtB, which transports ammonium nitrogen in other  Bacillus  strains. Collectively, this study defines heterogeneity of  B. velezensis  LG37 through comprehensive transcriptome analysis and subsequently, by genome editing techniques, sheds light on the enigmatic mechanisms controlling the functional genes under different nitrogen sources also reveals the need for further research.

Bacillus velezensis LG37: transcriptome profiling and functional verification of GlnK and MnrA in ammonia assimilation