Genomic analysis of salinity-tolerant bacteria

Genome sequencing of the strains was performed to obtain the completed map of the strain genome. Combined with the genomic information of the remaining five strains, the genomes of the six salinity-tolerant bacteria were annotated and analyzed using the RAST database to find the similarities and differences in the genomes of the six strains in which the salinity tolerance was reflected.

1.Salinity-tolerant bacterial genome annotation

Brevibacterium casei G20, Bacillus haynesii P19, Micrococcus luteus R17, Halomonas sp. TD01, Halomonas campaniensis LS21, and Enterobacter cloacae RS35 genomes were annotated online using RAST (Rapid Annotation using Subsystem Technology). TD01, Halomonas campaniensis LS21, and Enterobacter cloacae RS35 genomes, and compared with the Subsystems library for functional gene annotation.The results of the annotation of Brevibacterium casei G20 are shown in Figure 2.5.The total number of annotated genes for Brevibacterium casei G20 is shown in Figure 2.5.The total number of annotated genes for Brevibacterium casei G20 is shown in Figure 2.5. casei G20 annotated a total of 3871 genes, associated with 25 Subsystems. Among them, Amino Acids and Derivatives (AAD) had the highest proportion of 436 genes, followed by Carbohydrates (Carbohydrates) with 342 genes, Cofactors, Vitamins, Prosthetic Groups, Pigments (Cofactors, Prosthetic Groups, Pigments, Pigments) with 436 genes, and Cofactor, Prosthetic Groups, Pigments with 342 genes. Cofactors, Vitamins, Prosthetic Groups, Pigments (Cofactors, Vitamins, Prosthetic Groups, Pigments) systematically annotated 309. The remaining systems with more genes annotated were Protein Metabolism with 245 genes, Fatty Acids, Lipids, and Isoprenoids with 186 genes, and Membrane Transport with 115 genes. Transport, Membrane Transport, Stress Response, and Stress Response, and 103 related genes.

Bacillus haynesii P19 genome is 4,235,749 bp in length, with 46.04% GC content, and a total of 4,348 genes were annotated, which were related to 26 Subsystems (Figure 2.6). Among them, Carbohydrates (Carbohydrates) system annotated the highest proportion of related genes of 599, followed by Amino Acids and Derivatives (Amino Acids and Derivatives) system annotated 474 related genes, Cofactors, Vitamins, Prosthetic Groups, Pigments (Cofactors, Prosthetic Groups, Pigments), and other related genes. Cofactors, Vitamins, Prosthetic Groups, Pigments) were systematically annotated to 244. The remaining systems with more genes annotated were Protein Metabolism with 244 genes, RNA Metabolism with 159 genes, Membrane Transport with 90 genes, and Stress Response with 104 genes. Stress Response system annotated 104 related genes.

The Micrococcus luteus R17 genome was 2,597,893 bp in length with 73.1% GC content, and a total of 2,313 genes were annotated, associated with 26 Subsystems (Figure 2.7). Among them, amino acids and derivatives (Amino Acids and Derivatives) system annotated 313 related genes, followed by carbohydrates (Carbohydrates) system annotated 197 related genes, cofactors, vitamins, repair groups, pigments (Cofactors, Vitamins, Prosthetic Groups, Pigments). Cofactors, Vitamins, Prosthetic Groups, Pigments (Cofactors, Vitamins, Vitamins, Prosthetic Groups, Pigments) systematically annotated 193 related genes. The remaining systems with more genes annotated were Protein Metabolism with 211 genes, Fatty Acids, Lipids, and Isoprenoids with 131 genes, Membrane Transport with 97 genes, and Vitamins, Vitamins, Prosthetic Groups, Pigments with 193 genes. Transport, Membrane Transport, and Stress Response, with 131 genes, 97 genes, and 135 genes.

Halomonas sp. TD01 genome is 5,809,624 bp in length, with 52.4% GC content, and a total of 5,602 genes were annotated, which were associated with 25 Subsystems (Figure 2.8). Among them, Amino Acids and Derivatives (AAD) system annotated 529 genes, followed by Cofactors, Vitamins, Prosthetic Groups, Pigments (Cofactors, Vitamins, Prosthetic Groups, Pigments) system annotated 352 genes The number of genes annotated in the Carbohydrates system was 293. The other systems with more genes annotated were Protein Metabolism with 230 genes, Nucleosides and Nucleotides with 148 genes, Membrane Transport with 97 genes, and Stress Response with 97 genes. The Stress Response system annotated 135 related genes.

Halomonas campaniensis LS21 genome is 4,074,048 bp in length, with 52.6% GC content, and a total of 3,767 genes were annotated, which were related to 26 Subsystems (Figure 2.9). Among them, amino acids and derivatives (Amino Acids and Derivatives) system annotated to 419 related genes, followed by carbohydrates (Carbohydrates) system annotated to 372 related genes. Cofactors, Vitamins, Prosthetic Groups, Pigments (Cofactors, Vitamins, Prosthetic Groups, Pigments) systematically annotated 272 related genes. The remaining systems with more genes annotated were Protein Metabolism with 172 genes, RNA Metabolism with 171 genes, Membrane Transport with 159 genes, and Stress Response with 159 genes. RNA Metabolism (RNA Metabolism), Membrane Transport (Membrane Transport), 112 genes were annotated.

The Enterobacter cloacae RS35 genome is 5,112,849 bp in length, with 54.25% GC content, and a total of 4657 genes were annotated, which were associated with 26 subsystems (Figure 2.10). Among them, 654 genes were annotated in the Carbohydrates system, followed by 508 genes in the Amino Acids and Derivatives system. The Protein Metabolism system annotated 298 genes. The other systems with more genes annotated were Cofactors, Vitamins, Prosthetic Groups, Pigments (Cofactors, Vitamins, Prosthetic Groups, Pigments) with 262 genes annotated , RNA Metabolism (RNA Metabolism) with 236 genes annotated , Membrane Transport (Membrane Transport) with 236 genes annotated , and Membrane Transport (Membrane Transport) with 262 genes annotated. Membrane Transport (209 genes) and Stress Response (183 genes).

RAST annotation revealed that the six strains had high annotations in four systems: amino acids and derivatives, carbohydrates, cofactors, vitamins, prosthetic groups, pigments, and protein metabolism, which are essential for the strains to maintain their normal growth and development. Pigments), and protein metabolism (Protein Metabolism) are highly annotated in all four systems, which are genes necessary for the strain to maintain its normal growth and development. In addition, the genes were highly annotated in Membrane Transport and Stress Response systems, which are essential for the strain to maintain its normal growth and development. In addition, Micrococcus luteus R17 has the shortest genome length and the highest GC content among the six strains of salinity-tolerant bacteria, probably because it was screened from the radiation environment. The short genome length and high GC content may enable Micrococcus luteus R17 to maintain its DNA strand stability in the radiation environment.Brevibacterium casei G20 also had a high GC content, but Bacillus haynesii P19, which is also an endophytic bacterium, had a low GC content. The GC content of Gram-negative bacteria screened from the sodium salt environment was similar at around 52%.

2.Analysis of membrane transport and stress response genes

In order to understand the genomic similarities and differences in the strain's tolerance to saline and alkaline environments, we further explored the relevant genes annotated to the strain's Membrane Transport system and Osmotic stress system.

Comparison of Gram-positive and Gram-negative membrane transport and stress response systematic annotation results are shown in Tables 2.5-2.6. It can be found that the number of genes annotated for choline and betaine uptake and betaine biosynthesis is higher in osmotic stress in Brevibacterium casei G20 and Halomonas campaniensis LS21. The number of genes annotated for Choline and Betaine Uptake and Betaine Biosynthesis was higher in Brevibacterium casei G20 and Halomonas campaniensis LS21. Also tetrahydropyrimidine biosynthesis and regulation was annotated in both strains. Both betaine and tetrahydropyrimidine are compatible solutes possessing strong osmotic pressure homeostasis. Therefore, it is hypothesized that one of the strategies of Brevibacterium casei G20 and Halomonas campaniensis LS21 to cope with saline environments is to accumulate betaine and tetrahydropyrimidine.The number of genes annotated in osmoregulation was greater in the three Gram-negative than in the Gram-positive strains. All strains annotated ABC transporter protein-related genes, with Enterobacter cloacae RS35 annotating the highest number of genes. Gram-positive bacteria had a greater number of genes annotated in Cation transporters than Gram-negative bacteria. In addition six strains had more genes annotated in Uni- Sym- and Antiporters, Membrane Transport - no subcategory.ABC transporter is a protein that binds ATP and transports a variety of molecules across membranes in an active transporter mode. A variety of molecules are transported across membranes in an active transporter mode. Cation transporter proteins are involved in the transport of a wide range of cationic ions.There are three subsystems under Uni-Sym- and retrotransporter proteins: proton-dependent peptide transporter, multisubunit cation retrotransporter, NhaA, NhaD, and sodium-dependent phosphate transporter. Notably the TRAP transport (TRAP transporte) system was only annotated to a high number of genes in the genus Salmonella.

It can be hypothesized that six strains of saline-tolerant bacteria maintain intracellular osmotic pressure and pH balance through a series of ion transport mechanisms in response to saline environments, and Brevibacterium casei G20 and Halomonas campaniensis LS21 may accumulate compatible solutes such as betaine and tetrahydropyrimidine in saline environments.

3.Discussion

Comparison of the RAST database revealed that six bacterial strains had a high percentage of genes annotated in the Amino Acids and Derivatives (Amino Acids and Derivatives), Carbohydrates (Carbohydrates), Cofactors, Vitamins, Prosthetic Groups, Pigments, and Protein Metabolism subsystems (Protein Metabolism). Pigments), and protein metabolism (Protein Metabolism) subsystems annotated to a high percentage of genes. This is related to the genes necessary for the strains to maintain their own growth and development. The next six strains had a high proportion of genes annotated in the Membrane Transport and Osmotic stress subsystems, including ABC transporters, Cation transporters, Uni-Sym- and Reverse-Sym- transporters. transporters, Uni-Sym- and Antiporters, and Membrane Transport-no subcategory annotated to more genes. It is hypothesized that the six strains of saline-tolerant bacteria maintain intracellular osmotic pressure and pH balance in saline environments through a series of ion transport mechanisms, such as Na+ /H+ reverse transporter proteins. The genomes of Brevibacterium casei G20, Halomonas campaniensis LS21, and Halomonas sp. TD01 were shown to be in the system of choline and betaine uptake and betaine biosynthesis (Choline and Betaine Uptake and Betaine Biosynthesis (Choline and Betaine Uptake and Betaine Biosynthesis) system, a series of enzymes ranging from choline to betaine could be annotated, which possessed the ability to synthesize betaine. The subsequent mining of the metabolites of these three strains also revealed the presence of betaine in the metabolites of the strains.