The Systematics Of Family Thermo-Actinomycetaceae And The Multi-omics Research For Its Hyphae Differentiation

Members of Thermoactinomycetaceae are a kind of gram-positive,heterotrophic,aerobic microorganisms that can produce mycelium and live under high temperature conditions.The taxonomic status of this group is Bacillales,however,its special hyphae differentiation is similar to the actinomycetes.Members of this group has diverse physiological,biochemical and morphological characteristics.Its habitat is widely distributed and its metabolic products have unique properties because of endospores resistance.Few studies have been conducted on the phylogenomics of this group,however the mechanism in the genome and transcriptome level have not been studied.Therefore,the present study was designed to investigate the phylogenetic genomes and hyphae differentiation mechanism and others underlying science problems,based on big data of whole family genome in addition to the strain transcriptome analysis by genomic data to explore phylogenetic problems.At the same time,transcriptome analysis was performed to reveal the difference in key genes for mycelia differentiation and special mechanism between Thermoactinomycetaceae and actinomycetes.First,30 Thermoactinomycetaceae strains were obtained from the world’s preservation centers(CCTCC、CGMCC、DSMZ、JCM、KCTC、NBRC),which were cultured and screened for transcriptome analysis.We collected genome data of Thermoactinomycetaceae strains from NCBI and JGI database.The 16 strains that were successfully cultured which had no previously published genome data were sequenced for genome.The strains Laceyella sediminis CCTCC AA 208058 and Laceyella sacchari JCM 3137 were sequenced using PacBio.Preliminary comparative analysis of sequencing 18 strains including Laceyella sacchari JCM 3137 were selected as the target group for the transcriptome analysis.Total of 57 genomes were cultured or obtained,which are comprised of 18 sequenced strains of Thermoactinomycetaceae,the genomes of Thermoactinomycetaceae strains available in NCBI and JGI databases and the genomes of strains closely related to Thermoactinomycetaceae,based on 16S rRNA gene sequences.The collective data comprised the unified prediction and annotation.Homologous gene analysis was performed by using orthoMCL and GeneContent and SuperMatrix trees were built and compared against phylogenetic tree,based on 16S rRNA gene sequence.Based on the combined the characteristics of the strains and the ICNP rules,we proposed to rename Laceyella sacchari,Laceyella sediminis,Laceyella tengchongensis,Laceyella thermophila,Laceyella putida to Thermoactinomyce sacchari,Thermoactinomyce sediminis,Thermoactinomyce tengchongensis,Thermoactinomyce putida,Thermoactinomyce thermophile respectively.Thermoactinomyces guangxiensis should be removed from Thermoactinomyces to Lihuaxuella genus and be renamed Lihuaxuella guangxiensis.Novibacillus thermophilus should be removed from Thermoactinomycetaceae and to be considered as a new family named Novibacillaceae,Novibacillus as the type genus,Novibacillus thermophilus as the type species.Strain Laceyella sacchari JCM 3137 was used as the transcriptome analysis material for the design of experiements.By using the PacBio genome data of strain JCM 3137 as reference genome to analyze,we found the gene clusters:for example,orf767-orf771 and the genes orf 1546、orf1046、orfO 170(gerPC)、orf 1139(spoⅢAB)、orfl 145(spoⅢAH)are very important for hyphae differentiation within the strains of the family Thermoactinomycetaceae,The results also showed that the hyphae differentiation of Thermoactinomycetaceae strains was different from Streptomyces,with independent hyphae differentiation mechanism.It was confirmed that the hyphae differentiation of Thermoactinomycetaceae was closely related to the spo series gene,which indicates that this family is more closely related to bacillus in the classification status.At the same time,the transcriptome analysis revealed that Tryptone and Mannitol can promote mycelium growth and directly lead to the up-regulation of most metabolic pathways,particularly in the late stage growth of bacteria.In contrast,K2HPO4 can directly inhibit the growth of mycelium,the genes for signaling,organelle and structural molecule activity.When K2HPO4 was added at 18 hours,After 30 hours some genes about membrane-enclosed lumen and protein binding transcription factor activity was down-regulated.In this paper,phylogenetic genomics was used to study the Thermoactinomycetaceae and the classification status of some strains was suggested.At the same time,hyphae differentiation mechanism was studied using a combination of genome and transcriptome analysis of Thermoactinomycetaceae.The important genes and gene clusters associated with Thermoctinomycetaceae hyphae differentiation were identified.It is concluded that the Thermoactinomycetaceae hyphae differentiation mechanism is different from Streptomyces hyphae differentiation,having independent hyphae differentiation mechanism,which has a direct relationship with the gene of the Bacillus spores’ formation.