Taxogenomic and Metabolic Insights into Marinobacterium ramblicola sp. nov., a New Slightly Halophilic Bacterium Isolated from Rambla Salada, Murcia

A Gram-negative, motile, rod-shaped bacteria, designated D7T, was isolated by using the dilution-to-extinction method, from a soil sample taken from Rambla Salada (Murcia, Spain). Growth of strain D7T was observed at 15–40 °C (optimum, 37 °C), pH 5–9 (optimum, 7) and 0–7.5% (w/v) NaCl (optimum, 3%). It is facultatively anaerobic. Phylogenetic analysis based on 16S rRNA gene sequence showed it belongs to the genus Marinobacterium. The in silico DDH and ANI against closest Marinobacterium relatives support its placement as a new species within this genus. The major fatty acids of strain D7T were C16:0, summed feature 3 (C16:1 ω7c/C16:1 ω6c) and summed feature 8 (C18:1 ω7c/C18:1 ω6c). The polar lipid profile consists of phosphatidylethanolamine, phosphatidylglycerol and two uncharacterized lipids. Ubiquinone 8 was the unique isoprenoid quinone detected. The DNA G + C content was 59.2 mol%. On the basis of the phylogenetic, phenotypic, chemotaxonomic and genomic characterization, strain D7T (= CECT 9818T = LMG 31312T) represents a novel species of the genus Marinobacterium for which the name Marinobacterium ramblicola sp. nov. is proposed. Genome-based metabolic reconstructions of strain D7T suggested a heterotrophic and chemolitotrophic lifestyle, as well as the capacity to biosynthetize and catabolize compatible solutes, and to degrade hydrocarbon aromatic compounds.


Introduction
The genus Marinobacterium belongs to the family Alteromonadaceae, order Alteromonadales, class Gammaproteobacteria. At the time of writing, the genus Marinobacterium comprises 18 different species, from which the type species is Marinobacterium georgiense, a later heterotypic synonym of Marinobacterium iners [1,2].
Cells of members of this genus are Gram-stain-negative rods, motile by a single or two polar flagella [3]. They are slightly halophilic microorganisms, requiring NaCl for growth [3]. Some members are able to grow on aromatic compounds or to fixate nitrogen under limiting oxygen conditions [3]. Major fatty acids include C 16:0 , C 16:1 ω6c and/or C 16:1 ω7c, and C 18:1 ω6c and/or C 18:1 ω7c. Ubiquinone (Q-8) is the major respiratory quinone [3].
Using culture-independent techniques, members of the genus Marinobacterium were also detected in a wide range of habitats including soil samples from tidal freshwater wetlands in China [20], from mangrove river sediment in Taiwan [21], surface sediments

Bacterial Strains
Strain D7 T was isolated in this study from a soil sample taken from Rambla Salada, a hypersaline steep-sided river (rambla), located in the Province of Murcia, southeast Spain, 38 • 07 27.1 N 1 • 07 01.4 W. The physicochemical parameters of pH, oxygen (mg L −1 ) and salinity (g L −1 ) of the sampling location in Rambla Salada were 6.3, 10.2 and 44.4, respectively.
The sample was collected using a sterile polycarbonate tube, taken immediately to the laboratory and stored at 4 • C until study. The pH of the sample was close to neutral and salinity was around 40 g L −1 . For the isolation, we used S3 medium, a lownutrient medium [40] supplemented with 3% (w/v) sea-salt solution [41] and the dilution-toextinction approach as a cultivation method, described previously by Castro et al. [38]. This approach is a technique that improves the isolation of slow-growing species or apparently uncultivable species [42][43][44]. The extinction cultures were incubated at 25 • C for 30 days and after, the contents of wells were then re-isolated on Reasoner's 2A (R2A) medium plates [45]. The isolated strain was maintained and routinely grown in R2A with 3% (w/v) sea-salt solution at 30 • C as well as on marine agar (MA; 2216, Difco, Sparks, MD, USA).
For taxonomic comparison purposes, Marinobacterium zhoushanense KCTC 42782 T was used in this study and routinely grown in the same media as strain D7 T .

DNA Extraction, Purification and Sequencing
Genomic DNA was extracted using an X-DNA purification kit (Xtrem Biotech, Granada, Spain) from an overnight culture of strain D7 T in R2A medium. The 16S rRNA gene was amplified by PCR using the universal primers for bacteria 16F27 and 16R1488 [46]. The obtained PCR product was then purified, cloned into the pGEM-T vector (Promega, Madison, WI, USA), and sequenced by direct sequencing using the ABI prism dye-terminator, cyclesequencing ready-reaction kit and the ABI prism 377 sequencer according to PerkinElmer's instructions. The GenBank/EMBL/DDBJ accession number for the sequence of the 16S Microorganisms 2021, 9, 1654 3 of 16 rRNA gene is MG773714. The genome of strain D7 T was sequenced by using the Illumina MiSeq methodology (PE 150 × 2).

Phylogenetic Analysis Based on 16S rRNA Gene Sequence Comparison
Phylogenetic analyses based on the 16S rRNA gene were conducted as described previously [38,47]. The phylogenetic neighbors' identification and the pairwise 16S rRNA gene sequence similarities calculations were carried out by using the EzBiocloud server "www.ezbiocloud.net" [48] (accessed on April 2021). Phylogenetic and molecular evolutionary analyses were conducted using Mega v. 7 [49]. Clustering was determined using the neighbor-joining and maximum-likelihood algorithms and the evolutionary distances were computed using the Jukes-Cantor method [50]. The analysis involved 34 nucleotide sequences and the stability of the clusters was determined by a bootstrap analysis (1000 replications).
The genome of strain D7 T was annotated using BlastKOALA [55] and metabolic pathways were analyzed using KEGG [56].
InteractiVenn software [60] was used to display the Venn diagram.

Phylogenomic Reconstruction
Predicted protein sequences were compared using an all-versus-all BLAST search [61]. A total of 134 proteins were shared between all studied genomes and aligned using MUS-CLE v3.8.31 [62]. The concatenated and aligned orthologous genes were used to build the phylogenomic tree in Mega v. 7 [49].
Gram staining was performed according to the method described by Komagata [64]. Growth under anaerobic conditions was determined in an anaerobic jar using AnaeroGen (Oxoid) and an anaerobic indicator (Oxoid, Hampshire, UK) using marine agar (MA; Difco). Motility was observed using log-phase culture according to the hanging-drop method [65]. Oxidase activity was determined with 1% (v/v) tetramethyl-p-phenylenediamine [66] and catalase activity was examined by bubble production with 3% (v/v) H 2 O 2 solution [65]. The reduction of nitrate and nitrite and gas production were detected by adding the Griess-Ilosvay's reagent (Merck) in cultures grown in peptone broth supplemented with 1% KNO 3 . Other biochemical character, carbon utilization, sugar fermentation and enzymatic tests were carried out by using the GEN III MicroPlate TM system (Biolog), API 20NE, API 50CH Scanning electron microscope images of strain D7 T were produced on an FIB-FESEM (CrossBeam NVision 40, Carl Zeiss SMT) to determine bacterium size and type of flagella.

Chemotaxonomic Characterization
The fatty acids of strain D7 T were analyzed at the Spanish Type Culture Collection (CECT). Cells were grown on MA for 48 h, incubated at 30 • C. The whole-cell composition of the fatty acids was determined by GC using the midi microbial identification system [67]. The fatty acid profile was obtained with an Agilent 6850 gas chromatograph using the database TSBA6 145 [68].
Analysis of polar lipids and respiratory quinones of strain D7 T was carried out by the Identification Service of DSMZ, Braunschweig, Germany. Polar lipids were extracted following the protocol described by Bligh and Dyer [69]. Polar lipids were separated by two-dimensional silica-gel thin-layer chromatography (Macherey-Nagel Art. No. 818135) following the protocol described by Tindall et al. [70]. The two-stage method described by Tindall [71,72] was used to first extract respiratory lipoquinones followed by polar lipids.

Phylogenetic Analysis
During the course of the study of a soil sample from Rambla Salada (38 • 07 27.1 N 1 • 07 01.4 W), a hypersaline river located in Murcia (southeast of Spain), a novel strain designated D7 T was isolated and selected for further studies.
Based on the 16S rRNA gene sequence analysis, strain D7 T (1490 bp) was most closely related to the genus Marinobacterium, exhibiting the highest 16S rRNA gene sequence similarity to Marinobacterium zhoushanense WM3 T (98.0%), followed by Marinobacterium lutimaris DSM 22012 T (95.7%), Marinobacterium mangrovicola Gal22 T (95.3%) and Marinobacterium litorale IMCC 1877 T (95.1%). Moreover, the 16S rRNA gene sequence similarities to other genera, such as Neptunomonas and Nitrincola, were always equal or lower than 93.5%. The phylogenetic analysis based on the multiple sequence alignment of the 16S rRNA gene using the neighbor-joining algorithm (Figure 1), indicated that strain D7 T belongs to the genus Marinobacterium clustering with Marinobacterium zhoushanense WM3 T but was located in an independent branch with a high bootstrap value (Figure 1), indicating that the new strain could represent a new member of the genus Marinobacterium. A phylogenetic tree devised using the maximum-likelihood algorithm exhibited similar topologies.

Phylogenomic Analysis
According to the minimal standards for the use of genome data for the taxonomy of prokaryotes [73] and to confirm the phylogenomic relationships previously obtained by 16S rRNA gene sequence comparison, a phylogenomic tree based on core orthologous translated genes from strain D7 T and closely related members of the genus Marinobacterium was also obtained. A total of 134 single-copy orthologous genes were shared between all studied genomes and the phylogenomic tree reconstruction (Figure 3), clearly reflects that strain D7 T constitutes a monophyletic clade distinct from any other previously described species of the genus Marinobacterium, and therefore, supporting the placement of strain D7 T as a new species within this genus.

Genomic Characteristics
The draft genome sequence of strain D7 T was obtained and compared with that of the closest phylogenetic species, Marinobacterium zhoushanense CGMCC 1.15341 T , and with those of other members of the genus Marinobacterium with available genomes ( Table 1). The draft genome of strain D7 T was de novo assembled into a total of 69 contigs, with a N50 value of 150,886 bp, a sequencing depth of 844X and a completeness of 99.9%. This genome sequence is in accordance with the minimal standards for the use of genome data for the taxonomy of prokaryotes [73]. The G+C content and genome size of strain D7 T were 59.2 mol% and 4,897,523 bp, respectively; those values were within the range of the genomes of the genus Marinobacterium, which ranged from 54.9 to 62.1 mol%, and from 3,653,655 to 5,637,742 bp, respectively (Table 1). Additional genomic characteristics are detailed in Table 1. Besides, the 16S rRNA gene sequence of strain D7 T obtained from the draft genome sequence was identical to that from the PCR, verifying the authenticity of this genome.

Genomic Characteristics
The draft genome sequence of strain D7 T was obtained and compared with that of the closest phylogenetic species, Marinobacterium zhoushanense CGMCC 1.15341 T , and with those of other members of the genus Marinobacterium with available genomes ( Table 1). The draft genome of strain D7 T was de novo assembled into a total of 69 contigs, with a N50 value of 150,886 bp, a sequencing depth of 844X and a completeness of 99.9%. This genome sequence is in accordance with the minimal standards for the use of genome data for the taxonomy of prokaryotes [73]. The G+C content and genome size of strain D7 T were 59.2 mol% and 4,897,523 bp, respectively; those values were within the range of the genomes of the genus Marinobacterium, which ranged from 54.9 to 62.1 mol%, and from 3,653,655 to 5,637,742 bp, respectively (Table 1). Additional genomic characteristics are detailed in Table 1. Besides, the 16S rRNA gene sequence of strain D7 T obtained from the draft genome sequence was identical to that from the PCR, verifying the authenticity of  In addition, a Venn diagram displaying the number of genes shared between strain D7 T , Marinobacterium zhoushanense CGMCC 1.15341 T and Marinobacterium lutimaris DSM 22012 T was obtained ( Figure 2). A total of 645 genes were shared between M. zhoushanense CGMCC 1.15341 T , M. lutimaris DSM 22012 T and strain D7 T , while 462 genes were shared between M. lutimaris DSM 22012 T and strain D7 T , and 561 genes between M. zhoushanense CGMCC 1.15341 T and strain D7 T (Figure 2). A total of 2780 genes were identified as unique to strain D7 T (Figure 2). These results indicate that strain D7 T was unique from its closely related species. In addition, a Venn diagram displaying the number of genes shared between strain D7 T , Marinobacterium zhoushanense CGMCC 1.15341 T and Marinobacterium lutimaris DSM 22012 T was obtained (Figure 2). A total of 645 genes were shared between M. zhoushanens CGMCC 1.15341 T , M. lutimaris DSM 22012 T and strain D7 T , while 462 genes were shared between M. lutimaris DSM 22012 T and strain D7 T , and 561 genes between M. zhoushanens CGMCC 1.15341 T and strain D7 T (Figure 2). A total of 2780 genes were identified as unique to strain D7 T (Figure 2). These results indicate that strain D7 T was unique from its closely related species.

Phylogenomic Analysis
According to the minimal standards for the use of genome data for the taxonomy o prokaryotes [73] and to confirm the phylogenomic relationships previously obtained by 16S rRNA gene sequence comparison, a phylogenomic tree based on core orthologous translated genes from strain D7 T and closely related members of the genus Marinobacte rium was also obtained. A total of 134 single-copy orthologous genes were shared between all studied genomes and the phylogenomic tree reconstruction (Figure 3), clearly reflects that strain D7 T constitutes a monophyletic clade distinct from any other previously de scribed species of the genus Marinobacterium, and therefore, supporting the placement o strain D7 T as a new species within this genus. CGMCC 1.15341 T , M. lutimaris DSM 22012 T and strain D7 T , while 462 genes were shared between M. lutimaris DSM 22012 T and strain D7 T , and 561 genes between M. zhoushanense CGMCC 1.15341 T and strain D7 T (Figure 2). A total of 2780 genes were identified as unique to strain D7 T (Figure 2). These results indicate that strain D7 T was unique from its closely related species.

Phylogenomic Analysis
According to the minimal standards for the use of genome data for the taxonomy of prokaryotes [73] and to confirm the phylogenomic relationships previously obtained by 16S rRNA gene sequence comparison, a phylogenomic tree based on core orthologous translated genes from strain D7 T and closely related members of the genus Marinobacterium was also obtained. A total of 134 single-copy orthologous genes were shared between all studied genomes and the phylogenomic tree reconstruction (Figure 3), clearly reflects that strain D7 T constitutes a monophyletic clade distinct from any other previously described species of the genus Marinobacterium, and therefore, supporting the placement of strain D7 T as a new species within this genus.

Average Nucleotide Identity (ANI) and In Silico DNA-DNA Hybridization (DDH)
To elucidate if strain D7 T may constitute a new species within the genus Marinobacterium, the genome-based sequence similarity analysis (ANI and in silico DDH) between Microorganisms 2021, 9, 1654 8 of 16 strain D7 T and members of this genus was performed. For species delineation, the proposed and accepted boundaries for ANI and DDH are 95-96% and 70%, respectively [58,74,75].
The ANI and DDH values between strain D7 T and Marinobacterium zhoushanense CGMCC 1.15341 T , the closest phylogenetic neighbor, were 86.7% and 31.3%, respectively ( Figure 4). In addition, the ANI and DDH estimations of strain D7 T in comparison to those of the other members of the genus Marinobacterium with available genomes were in all cases lower than the established cutoff values (Figure 4). These results support the conclusion that strain D7 T represents a novel species of the genus Marinobacterium. Figure 3. Neighbor-joining core gene phylogenetic tree including available genomes of the genus Marinobacterium and strain D7 T . The tree was recovered from the alignment of 134 single-copy orthologous translated genes shared between these genomes. Bar, 0.1 substitutions per amino acid position.

Average Nucleotide Identity (ANI) and In Silico DNA-DNA Hybridization (DDH)
To elucidate if strain D7 T may constitute a new species within the genus Marinobacterium, the genome-based sequence similarity analysis (ANI and in silico DDH) between strain D7 T and members of this genus was performed. For species delineation, the proposed and accepted boundaries for ANI and DDH are 95-96% and 70%, respectively [58,74,75].
The ANI and DDH values between strain D7 T and Marinobacterium zhoushanense CGMCC 1.15341 T , the closest phylogenetic neighbor, were 86.7% and 31.3%, respectively ( Figure 4). In addition, the ANI and DDH estimations of strain D7 T in comparison to those of the other members of the genus Marinobacterium with available genomes were in all cases lower than the established cutoff values (Figure 4). These results support the conclusion that strain D7 T represents a novel species of the genus Marinobacterium.  Table 1.

Chemotaxonomic Characterization
To taxonomically describe strain D7 T as a new species, the complete chemotaxonomic characterization of this strain was performed. The polar lipid profile of strain D7 T includes phosphatidylethanolamine (PE), phosphatidylglycerol (PG) and two uncharacterized lipids (L) ( Figure S1).  Table 1.

Chemotaxonomic Characterization
To taxonomically describe strain D7 T as a new species, the complete chemotaxonomic characterization of this strain was performed. The polar lipid profile of strain D7 T includes phosphatidylethanolamine (PE), phosphatidylglycerol (PG) and two uncharacterized lipids (L) ( Figure S1).
The fatty acids composition of strain D7 T and the type strains of related species of the genus Marinobacterium are shown in Table 2. In accordance with members of the genus Marinobacterium [3], the major cellular fatty acids (>10%) of strain D7 T were C 16:0 (28.7%), summed feature 3 (C 16:1 ω7c/C 16:1 ω6c) (26.6%) and summed feature 8 (C 18:1 ω7c/C 18:1 ω6c) (25.2%). The fatty acid profile of strain D7 T was similar to those of the reference strains, but differing in their proportion (Table 2); hence, reaffirming its condition as a different species. The respiratory quinone of strain D7 T was ubiquinone-8 (Q-8), which is consistent with the rest of the members of the genus Marinobacterium [3].

Phenotypic Characterization
The phenotypic characteristics of strain D7 T were described and compared with those of Marinobacterium zhoushanense KCTC 42782 T . Cells were Gram-staining-negative, short motile rods (0.5-0.6 × 1.0-1.8 µm) with a single polar flagellum ( Figure S2). They were catalase and oxidase positive. Nitrate and nitrite were reduced. Glucose was not fermented. Weak growth was observed under anaerobic conditions. When tested on MA, growth of strain D7 T was observed at 15-40 • C (optimum, 37 • C), pH 5-9 (optimum, 7) and 0-7.5% (w/v) NaCl (optimum, 3%). Other characteristics of strain D7 T are given in the species description and those that differ from the strain type of the closest related species of the genus Marinobacterium are shown in Table 3.

Metabolism of Strain D7 T
Metabolic insights after the in-depth genomic analysis of strain D7 T suggest a heterotrophic and chemolithotrophic lifestyle for this strain. In relation to its heterotrophic capabilities, central carbohydrate pathways such as glycolysis, gluconeogenesis, pentose phosphate, Entner-Doudoroff, tricarboxylic acid and glyoxylate cycle were detected ( Figure 5). For pyruvate oxidation to acetyl-CoA, genes encoding pyruvate dehydrogenase (aerobic route) and pyruvate ferredoxin oxidoreductase (anaerobic route) were present. On the other side, a large number of ABC transporters for carbohydrate uptake (i.e., multiple sugar, glucose/mannose or fructose) were also identified in the studied genome ( Figure 5). Table 3. Differential phenotypic features of strain D7 T in comparison to those of the closest related species of the genus Marinobacterium.
As part of the nitrogen metabolism of strain D7 T , complete pathways for nitrogen fixation, dissimilatory nitrate reduction and denitrification were detected on its genome ( Figure 5). While genomic evidence for nitrogen fixation and nitrate reduction to ammonia via dissimilatory nitrate reduction pathway has been previously suggested for other Marinobacterium species [3], the whole set of genes encoding all steps of the denitrification was not identified before in any other members of this group [3]. Only Marinobacterium jannaschii exhibited the almost complete route, but lacked the key enzyme (NirK) [3]. Moreover, in the specific case of strain D7 T , the nitrate reduction capacity of this strain was also detected during the nitrate reduction phenotypic test. Several transporters for exogenous nitrogen-rich organic compounds uptake, such as Amt for ammonia, ABC for amino acids, putrescine or urea, with others, were found ( Figure 5). In addition, various complete aminoacidic biosynthetic routes (i.e., serine, threonine, cysteine, valine/isoleucine, leucine, lysine, ornithine, arginine, proline, histidine and tryptophan) were encountered in this genome.  Like many halotolerant microorganisms, to cope with osmotically varying conditions, D7 T encode genes for the synthesis and uptake of different compatible solutes. Glycine betaine is one of the most important osmoprotectants in prokaryotes that could also serve as an energy and carbon source in hypersaline environments [76]. The key enzymes choline dehydrogenase (BetA) and glycine betaine aldehyde dehydrogenase (BetB), involved in the biosynthesis of glycine betaine from choline, were recognized during the genomic analysis of strain D7 T ( Figure 5). The gene clusters gbcAB, dgcAB and soxBDAG for its further catabolism to glycine were also identified. This strain also possesses transporters for the uptake of this compound, such as the glycine betaine BCCT family and ABC transporters ( Figure 5). No evidence for the alternative route for glycine betaine biosynthesis from glycine (via glycine and sarcosine methyltransferase and dimethylglycine methyltransferase) was identified in the studied genome.
Ectoine is another osmotic solute widely synthetized by halophilic bacteria [76]. The presence of the complete biosynthesis pathway in this strain suggests its capacity to additionally synthetize ectoine as a compatible solute ( Figure 5). Besides, the ectoine hydroxylase (EctD) enzyme and the doeBDAC gene cluster, coding for Nα-acetyl-L-2,4-diaminobutyrate deacetylase (DoeB), diaminobutyrate transaminase (DoeD), ectoine hydrolase (DoeA) and aspartate semialdehyde dehydrogenase (DoeC), were also identified in this genome, indicating the ability to likewise synthetize its hydroxylated derivate, 5-hydroxyectoine, and to degrade ectoine, respectively ( Figure 5). Considering ectoine biosynthesis is energetically more expensive than the betaine one, it would be reasonable to believe that this strain only synthetizes ectoine under starving betaine or choline concentrations.
In addition, other several transporters related to osmotic stress for potassium uptake and sodium extrusion were also found in the genome.
Entire pathways for the catabolism of aromatic hydrocarbons (such as benzoate, benzene and anthranilate) to catechol were encountered in the genome of strain D7 T , fueling the catechol meta-cleavage pathway for its further breaking down ( Figure 5). In the same way, other Marinobacterium representatives (i.e., M. aestuarii, M. stanieri, M. profundum and/or M. jannaschii) were predicted to degrade benzoate or benzene, although in some cases through the ortho-cleavage pathway [3,16].
Noteworthily, strain D7 T encodes a sulfide:quinone oxidoreductase (SQR) which is predicted to oxidize sulfide to elemental sulfur, and thus exhibits a potential chemolithotrophic energetic metabolism for this strain. The SQR enzyme was also previously identified in the genomes of M. jannaschii and M. litorale [3]. Several others sulfur-based lithotrophy genes were also encoded by diverse Marinobacterium species [3]. On the other hand, the assimilatory sulfate reduction pathway via 3 -phosphoadenosine-5 -phosphosulfate (PAPS) was identified in the genome of strain D7 T ( Figure 5), reflecting its capacity to reduce sulfate to sulfide with the aim of satisfying sulfur nutritional requirements.
Acetate is a major product of the metabolism formed during fermentation by many bacteria and other organisms [77]. In strain D7 T , acetate is produced from Acetyl-CoA catalyzed by the phosphate acetyltransferase (Pta) and acetate kinase (AckA) enzymes ( Figure 5). In addition to acetate, this reaction generates ATP, hence contributing to the energy metabolism of the cell.
Finally, consistently with its microscopical visualization, genes encoding flagellum were found during the genomic analysis.

Conclusions
As a conclusion of the polyphasic taxogenomic analyses performed in this study, strain D7 T represents a new species within the genus Marinobacterium, for which the name Marinobacterium ramblicola sp. nov. is proposed. We enclose below the taxonomic description of this new species. The detailed genomic analysis of Marinobacterium ramblicola D7 T inferred a versatile energetic metabolism for this new taxon, characterized by a typical aerobic electron transport chain, oxidation of sulfur compounds, and nitrogen assimilation and fixation pathways. Likewise, based on the genomic inspection, it was suggested that the new species has the ability to biosynthetize or catabolize compatible solutes (i.e., ectoine or glycine betaine) and to degrade several aromatic hydrocarbons (i.e., benzene, benzoate and anthranilate).
The type strain is D7 T (= CECT 9818 T = LMG 31312 T ), isolated from a hypersaline river located in Murcia, Spain.
The GenBank/EMBL/DDBJ accession numbers for the 16S rRNA gene sequence and for the draft genome are MG773714 and JAHREP000000000, respectively.