Reclassification of Halomicroarcula saliterrae Straková et al. 2024 and Halomicroarcula onubensis Straková et al. 2024 into the genus Haloarcula, as Haloarcula saliterrae comb. nov. and Haloarcula onubensis comb. nov., respectively

1 Reclassification of Halomicroarcula saliterrae Straková et al. 2024 and Halomicroarcula onubensis Straková et al. 2024 into the genus Haloarcula, as Haloarcula saliterrae comb. nov. and Haloarcula onubensis comb. nov.,respectively DášaStraková, CristinaSánchez- Porro, Rafael R.de la Haba and AntonioVentosa* RESEARCH ARTICLE Straková etal., Int. J. Syst. Evol. Microbiol. 2024;74:006510 DOI 10.1099/ijsem.0.006510 Author affiliations: 1Department of Microbiology and Parasitology, Faculty of Pharmacy, University of Sevilla, 41012 Sevilla, Spain. *Correspondence: Antonio Ventosa, ventosa@ us. es Keywords: Haloarcula; haloarchaea; Halomicroarcula; taxonomy; taxogenomic analysis. Abbreviations: AAI, average amino acid identity; dDDH, digital DNA–DNA hybridization; ICNP, International Code of Nomenclature of Prokaryotes; OGRI, overall genome relatedness index; OrthoANI, orthologous average nucleotide identity. A supplementary figure is available with the online version of this article. 006510 © 2024 The Authors This is an openaccess article distributed under the terms of the Creative Commons Attribution License. Abstract The haloarchaeal genera Halomicroarcula and Haloarcula, belonging to the family Haloarculaceae, order Halobacteriales, class Halobacteria, within the phylum Methanobacteriota, have previously exhibited significant phylogenetic and taxonomic overlaps. This issue was recently resolved by merging the two genera into a single genus, Haloarcula. However, Halomicroarcula saliterrae and Halomicroarcula onubensis were described almost simultaneously with the proposal to unify the genera Haloarcula and Halomicroarcula. Their names were validly published under the International Code of Nomenclature of Prokaryotes (ICNP) according to Validation List no. 217, alongside six Haloarcula species and the transfer of the existing Halomicroarcula species into the genus Haloarcula. Therefore a phylogenetic, phylogenomic, and comparative genomic analysis was carried out to clarify the taxonomic status of these two haloarchaeal species, Halomicroarcula saliterrae and Halomicroarcula onubensis, with lower priority than the six new species of the genus Haloarcula. Phylogenetic studies of 16S rRNA and rpoB′ gene sequences, along with phylogenomic reconstructions using singlecopy coreorthologous proteins, indicated that the two species clustered with the members of the genus Haloarcula. The overall genome relatedness indexes (OGRIs), comparative analyses of phenotypic features, and polar lipid profiles further supported their taxonomic reassignment as two separate species within the genus Haloarcula. Consequently, we propose the reclassification of Halomicroarcula saliterrae Straková et al. 2024 and Halomicroarcula onubensis Straková et al. 2024 into the genus Haloarcula, as Haloarcula saliterrae comb. nov. and Haloarcula onubensis comb. nov., respectively, in accordance with the ICNP. INTRODUCTION The genus Haloarcula belongs to the family Haloarculaceae, within the order Halobacteriales, class Halobacteria, within the phylum Methanobacteriota. It was proposed by Torreblanca et al. in 1986 [1] by reassigning a species, previously described as Halobacterium vallismortis, to the new genus Haloarcula, as Haloarcula vallismortis. The genus Halomicroarcula was proposed in 2013 by Echigo et al. [2] to accommodate the species Halomicroarcula pellucida. The study by Durán- Viseras et al. in 2021 [3] suggested merging the genera Halomicroarcula and Haloarcula into a single genus, Haloarcula, based on phylogenomic similarities among their species. They recommended a more exhaustive study, encompassing a larger set of strains and additional genomic analyses, to better reflect the evolutionary relationships and phenotypic features of species of these genera. Since then, four novel Halomicroarcula species and eight new Haloarcula species have been published under the International Code of Nomenclature of Prokaryotes (ICNP). In January 2024, Ma et al. [4] conducted phylogenetic, phylogenomic, and comparative genomic analyses to reassess the taxonomic status of species of these two genera. Their key findings supported the merging of Haloarcula and Halomicroarcula into a single genus, Haloarcula, resulting in a more coherent classification within the family Haloarculaceae. Currently, the genus Haloarcula includes 27 species according to the List of Prokaryotic names with Standing in Nomenclature [5]: Har. amylolytica [6], Har. amylovorans [4], Har. argentinensis [7], Har. halobia [4], Har. halophila [4], Har. hispanica [8], Har. japonica [9], Har. laminariae [4], Har. limicola [4], OPEN ACCESS 2 Straková etal., Int. J. Syst. Evol. Microbiol. 2024;74:006510 Har. litorea [4], Har. mannanilytica [10], Har. marina [4], Har. marismortui [11], Har. nitratireducens [4], Har. ordinaria [4], Har. pelagica [4], Har. pellucida [4], Har. rara [4], Har. rubra [4], Har. salina [12], Har. salinisoli [4], Har. sebkhae [13], Har. terrestris [14], Har. vallismortis [1], Har. quadrata [15], Har. salaria [16], and Har. tradensis [16]. The last three species have recently been proposed as later heterotypic synonyms of Har. argentinensis and Har. marismortui, respectively [12, 14]. On the other hand, two new Halomicroarcula species, Halomicroarcula saliterrae and Halomicroarcula onubensis, were described in 2024 by Straková et al. [17]. These species names were validly published under the ICNP according to Validation List no. 217 [18] concurrently with the six Haloarcula species described by Ma et al. [4] and the transfer of the existing Halomicroarcula species into the genus Haloarcula. Therefore, a comprehensive phylogenetic, phylogenomic, and comparative genomic analysis was conducted to elucidate the taxonomic status of these two haloarchaeal species. Together with comparative analysis of phenotypic features and polar lipid profiles, this analysis confirmed that these species should be reclassified into the genus Haloarcula, as Haloarcula saliterrae comb. nov. and Haloarcula onubensis comb. nov. METHODS Phylogenetic studies The 16S rRNA and rpoB′ gene sequences of the studied species, obtained from their respective genomes, were retrieved from the NCBI GenBank database and taxonomically associated with their phylogenetic neighbours using a blastn search [19]. Multiple sequence alignments of the 16S rRNA and rpoB′ genes from Halomicroarcula saliterrae S1CR25- 12 T , Halomicroarcula onubensis S3CR25- 11T, and their closest relatives were performed using the Fast Aligner tool in arb software [20]. The alignments were then visually inspected and corrected. Phylogenetic tree reconstructions based on the 16S rRNA and rpoB′ gene sequences were conducted using the maximumlikelihood [21], neighbourjoining [22], and maximumparsimony algorithms [23] implemented in arb software [20]. The ‘gitana’ script was used for formatting and visualizing the phylogenetic tree [24]. Phylogenomic analyses and genomic indexes For the taxogenomic analyses, publicly available genome sequences of Halomicroarcula saliterrae S1CR25- 12 T , Halomicroarcula onubensis S3CR25- 11 T , and closely related species were obtained from the NCBI GenBank database, adhering to the minimal standards for the use of genome data for taxonomic purposes [25, 26]. The features of these genomes have been published previously [3, 4, 17]. The Enveomics toolbox [27] was used to identify the clusters of orthologous proteins shared by all analysed species. The singlecopy coreorthologous proteins were individually aligned using muscle version 5.1 [28]. To determine the phylogenomic position of Halomicroarcula saliterrae and Halomicroarcula onubensis relative to the species of the genus Haloarcula and other related haloarchaea, an approximately maximumlikelihood phylogenomic tree was reconstructed using FastTreeMP version 2.1.8 [29]. Overall genome relatedness indexes (OGRIs) were calculated among Halomicroarcula saliterrae S1CR25- 12T, Halomicroarcula onubensis S3CR25- 11T, and the species of the genus Haloarcula and other related species of the family Haloarculaceae. Orthologous average nucleotide identity (OrthoANI) values were determined by OrthoANIu tool version 1.2 [30], digital DNA–DNA hybridization (dDDH) values were obtained using the Genome- to- Genome Distance Calculator (GGDC 3.0) from the Leibniz Institute DSMZ (Germany) [31, 32], and average amino acid identity (AAI) values were calculated using the ‘ aai. rb’ script from the Enveomics collection [27]. Comparative analysis of phenotypic features and chemotaxonomic profiles A comparative analysis of phenotypic features, including morphological, physiological, biochemical, and nutritional characteristics, as previously reported [3, 4, 12, 17], was conducted for Halomicroarcula saliterrae, Halomicroarcula onubensis and their closest related species. In addition, the characterization and differentiation of haloarchaea at the genus level have significantly benefited from the analysis of polar lipids, which serve as valuable taxonomic markers [1, 26, 33]. A comparative analysis of the lipid profiles of Halomicroarcula saliterrae and Halomicroarcula onubensis, as previously presented [17], with those of species of the genus Haloarcula was conducted to confirm the taxonomic affiliation of these two species within the genus Haloarcula. RESULTS AND DISCUSSION Phylogenetic studies incorporating six additional Haloarcula species We present an extended phylogenetic analysis including all current species of Haloarcula, that incorporate the six recently described Haloarcula species not included in our previous study conducted on Halomicroarcula saliterrae and Halomicroarcula onubensis [17]. The additional six Haloarcula species described by Ma et al. [4] exhibited 16S rRNA gene sequence identities ranging from 98.3 to 90.7%, and from 98.2 to 91.0% with Halomicroarcula saliterrae S1CR25- 12T and Halomicroarcula onubensis S3CR25- 11T, respectively. Among the six additional Haloarcula species, Haloarcula rara SHR3 T was identified as the most closely related to both Halomicroarcula species. Additionaly, an analysis of the rpoB′ gene sequences of Halomicroarcula saliterrae S1CR25- 12T and Halomicroarcula onubensis 3 Straková etal., Int. J. Syst. Evol. Microbiol. 2024;74:006510 S3CR25- 11T, compared with the rpoB′ gene sequences of the six additional Haloarcula species, demonstrated that the closest relatedness was to Haloarcula rara SHR3T, with sequence identities of 93.6 and 92.5%, respectively. Phylogenetic reconstruction based on the 16S rRNA (Fig.1) and rpoB′ (Fig. S1, available in the online version of this article) gene sequences indicated the affiliation of Halomicroarcula saliterrae S1CR25- 12T and Halomicroarcula onubensis S3CR25- 11T within the genus Haloarcula. Haloferax volcanii T NCIMB 2012 (AY425724) Haloglomus irregulare T F1660- (MH424600) Halosegnis rubeus T F1744- (NR_177380) Natronomonas pharaonis T JCM 8858 (D87971) Halococcoides cellulosivorans T HArcel1 (NR_177325) Halorhabdus utahensis T AX 2- (AF071880) Halorientalis regularis T JCM 16425 (AB663401) Halapricum salinum T CBA1105 (KF314042) Salinirussus salinus T YGH44 (JQ937358) T Halosimplex carlsbadense rrnA2−91 (NR_112206)- T Halosimplex carlsbadenserrnB2−91 (AB072815)- T Halomicrobium salinisolirrnBLT50 (MZ798423) T Halomicrobium urmianum DC8 (KM596805) T Halomicrobium salinisoli LT50 rrnA (KM496559) T Halomicrobium zhouii rrnATBN51(HM063952) T Halomicrobium zhouii rrnBTBN51(HQ215546) Haloarcula litorea GDY20 rrnA (MT023690) T Halomicrobium mukohataei rrnA JCM 9738EF645690 () Haloarcula halophila T DFY41 rrnA (MT023685) Haloarcula pelagica T YJ 61 S--rrnA (KC918823) T Haloarcula marina rrnADT1MW774245() T Haloarcula rubra rrnAF13(MH447277) Haloarcula halobia T XH51 rrnA (MK920147) Haloarcula ordinaria T ZS 22 S1-- rrnA (KC918828) T Halomicroarcula rrnAsaliterrae S1CR25-12025)(ON653 T Halomicroarcula rrnAonubensis S3CR25-11(ON653386) T Haloarcula laminariae rrnALYG108 MH062959- () T Haloarcula salinisoli rrnAF24A (MH447282) Haloarcula rara T SHR3 rrnA (KR012972) T Haloarcula nitratireducens rrnAF27 (MH447286) Haloarcula amylovorans T LR21 (MT649491)rrnA T Haloarcula limicola rrnAYGHS32 (KF582944) T Haloarcula pellucida rrnABNERC31 (AB766179) T Haloarcula japonica rrnAJCM 7785 (EF645684) T Haloarcula vallismortis CGMCC1.2048 rrnA (EF645687) T Haloarcula sebkhae rrnA SWO25(HQ844527) T Haloarcula argentinensis rrnAJCM 9737 (EF645680) Haloarcula mannanilytica T MD1301-(LC382261) Haloarcula terrestris T S1AR25 5A 0N653024-( ) rrnA T Haloarcula marismortui rrnACGMCC 1.1784 (EF645692) T Haloarcula amylolytica rrnA BD 3(DQ826512)- T Haloarcula salina rrnAYGHS18 (KF582943) T Haloarcula hispanica ATCC 33960 rrnB (AB090168) T Halomicrobium mukohataei rrnBJCM 9738 (EF645691 ) Haloarcula litorea T GDY20 rrnB (MT023703) T Haloarcula marina rrnBDT1OP435655() T Haloarcula rubra rrnB F13(MH447279) T Haloarcula nitratireducens rrnB F27(MH447284) T Haloarcula amylo rrnBvorans LR21 (MH620372) T Haloarcula limicola rrnB YGHS32 (JQ237128) Haloarcula halobia T XH51 rrnB (MK920151) Haloarcula ordinaria T ZS 22 S1-- rrnB (MF423468) Haloarcula halophila T DFY41 rrnB (MT023704) Haloarcula pelagica T YJ 61 S--rrnB (KM000049) T Haloarcula pellucida rrnB BNERC31 (AB766180) T Haloarcula laminariae rrnBLYG108 MH064348- () Halomicroarcula saliterrae T S1CR25 12- rrnB (ON653026) Haloarcula rara T SHR3 rrnB (KR012973) T Haloarcula salinisoli rrnBF24A (MH447281) Halomicroarcula onubensis T S3CR25 11-rrnB (ON653385) T Haloarcula salina rrnBYGHS18 (JQ237127) T Haloarcula amylolytica rrnBBD-3 Q826513)(D Haloarcula japonica JCM7785 rrnB (EF645685) T Haloarcula terrestr rnBis rS1AR25-5A ()ON653023 T Haloarcula hispanica rrnAATCC 33960 (AB090167) T Haloarcula vallismortis rrnB CGMCC 1.2048 (EF645688) T Haloarcula marismortui CGMCC 1.1784 rrnB (EF645693) T Haloarcula sebkhae rrnBSWO25(HQ641747) T Haloarcula argentinensis rrnB JCM 9737 (EF645681) 0.01 89 76 87 88 88 94 9183 99 80 86 83 94 95 76 93 81 81 97 98 90 98 84 T T Fig. 1. Neighbourjoining phylogenetic tree based on the 16S rRNA gene sequence comparison of the species Halomicroarcula saliterrae S1CR25- 12T and Halomicroarcula onubensis S3CR25- 11T with species of Haloarcula and other related species within the family Haloarculaceae. The species Haloferax volcanii NCIMB 2012T was used as an outgroup. Sequence accession numbers are shown in parentheses. Bootstrap values ≥70% (based on 1000 pseudoreplicates) are shown at branch points. Filled circles indicate branches that were also recovered in the trees generated with the maximumlikelihood and maximumparsimony algorithms. Bar, 0.01 expected substitutions per nucleotide position. 4 Straková etal., Int. J. Syst. Evol. Microbiol. 2024;74:006510 Phylogenomic analyses and genomic indexes validate reclassification to the genus Haloarcula Similar to the 16S rRNA gene sequence analysis, the phylogenomic and comparative analyses of genomic identities were expanded to include the six recently described Haloarcula species. The approximately maximumlikelihood phylogenomic tree reconstruction, based on the comparison of 153 singlecopy coreorthologous proteins, revealed that Halomicroarcula saliterrae S1CR25- 12T and Halomicroarcula onubensis S3CR25- 11T formed a cluster with Haloarcula salinisoli F24AT, Haloarcula laminariae LYG- 108T, and Haloarcula rara SHR3T (Fig.2). Additionally, to clarify the taxonomic positions of Halomicroarcula saliterrae S1CR25- 12T and Halomicroarcula onubensis S3CR25- 11 T , we calculated the OGRIs among the species of the family Haloarculaceae. The OrthoANI, dDDH, and AAI values among these two species and the current species of the genus Haloarcula were 86.0–77.4%, 30.9–21.6%, and 84.2–70.1 %, respectively, all significantly below the threshold values for prokaryotic species delineation (OrthoANI 95–96%, dDDH 70%, AAI 95- 96%) [34–37], and mostly above genus cutoff (AAI 65- 72%)[38] (Figs3 and 4). These results indicate unequivocally that Halomicroarcula saliterrae S1CR25- 12T and Halomicroarcula onubensis S3CR25- 11T should be reclassified within the genus Haloarcula. These results support that the two species, Halomicroarcula saliterrae and Halomicroarcula onubensis, show genotypic features compatible with those of the species of Haloarcula. Phenotypic features and chemotaxonomic analysis confirm the affiliation to the genus Haloarcula A comparative analysis of the phenotypic features of Halomicroarcula saliterrae S1CR25- 12 T , Halomicroarcula onubensis S3CR25- 11T, and their phylogenetically closest species, Haloarcula laminariae LYG- 108T, Haloarcula salinisoli F24AT, and Haloarcula rara SHR3T, as well as the type species of the genus, Haloarcula vallismortis ATCC 29715T, was conducted based on previous data T Halapricum salinum CBA1105(CP031310) Halorhabdus utahensis DSM 12940 (CP001687) T Halococcoides cellulosivorans HArcel1 (CP028858) T Salinirussus salinus YGH44 (WOWO00000000) Halorientalis regularis TNN28 (JALJCL000000000) Natronomonas pharaonis DSM 2160 (CR936257) T Haloglomus irregulare (QMDX00000000)F16-60 T Halosegnis rubeus F17-44 (QJOW00000000) T Halosimplex carlsbadense 2–9-1 (AOIU00000000) T Halomicrobium zhouii CGMCC 1.10457(FOZK00000000) T Halomicrobium mukohataei DSM 12286(CP001688) T Halomicrobium salinisoli LT50 (CP084463) T Halomicrobium urmianum IBRC-M 10911 (CP084090) T Haloarcula halophila DFY41 (CP119559–CP119562) T Haloarcula pelagica YJ-61-S (CP126161–CP126164) T Haloarcula litorea GDY20 (CP119779–CP119781) T Haloarcula amylovorans LR21 (SRIF00000000) T Haloarcula nitratireducens F27 (RKLT00000000) T Haloarcula limicola JCM 18640 (JAHQXF000000000) T Haloarcula marina DT1(CP100404) T Haloarcula pellucida CECT 7537 (RKLW00000000) Haloarcula rubra F13 (RKLR00000000) T Halomicroarcula saliterrae S1CR25-12(JAMQON000000000) T Halomicroarculaonubensis S3CR25-11(JAMQOS000000000) T Haloarcula laminariae LYG-108(JAMZFY000000000) T Haloarcula rara SHR3 (CP119785–CP119786) T Haloarculasalinisoli F24A (RKLQ00000000) T Haloarcula ordinaria ZS-22-S1 (CP119789–CP119790) T Haloarcula salina JCM 18369 (JAHQXE000000000) T Haloarculaterrestris S1AR25-5A (JAMQOM000000000) T Haloarcula mannanilytica MD130-1 (BIXZ00000000) T Haloarcula vallismortis ATCC 29715 (AOLQ00000000) T Haloarcula amylolytica JCM 13557 (AOLW00000000) T Haloarcula hispanica ATCC 33960(CP002921) T Haloarcula marismortui ATCC 43049 (CP039138) T Haloarcula japonica DSM 6131 (AOLY00000000) T Haloarcula sebkhae JCM 19018 (BMPD00000000) T Haloarcula argentinensis DSM 12282 (AOLX00000000) 100 100 100 100 100 100 100 100 100 100 100 100 100 100 100 100 100 100 83 100 100 99 100 100 98 100 100 100 100 100 100 100 100 100 98 0.05 Haloarcula halobia XH51 (CP119787–CP119788) T T T T T Fig. 2. Approximately maximumlikelihood phylogenomic tree based on the comparison of 153 singlecopy coreorthologous proteins showing the relationships of the species Halomicroarcula saliterrae S1CR25- 12T and Halomicroarcula onubensis S3CR25- 11T with species of Haloarcula and other related species within the family Haloarculaceae. Sequence accession numbers are shown in parentheses. Branch support values (%) are computed with the Shimodaira–Hasegawa test and are shown at branch points. Bar, 0.05 substitutions per amino acid position. 5 Straková etal., Int. J. Syst. Evol. Microbiol. 2024;74:006510 [3, 4, 12, 17, 39]. Their phenotypic characteristics are fully compatible with those of species of the genus Haloarcula. However, they presented some different characteristics, as shown in Table1. Furthermore, a comparative chemotaxonomic analysis of Halomicroarcula onubensis and Halomicroarcula saliterrae was conducted to confirm their affiliation within the genus Haloarcula. The major polar lipids of the two species included phosphatidylglycerol, phosphatidylglycerol phosphate methyl ester, phosphatidylglycerol sulfate, and a glycolipid chromatographically identical to sulfated diglycosyl diether [17]. These polar lipid profiles correspond to the currently established lipid patterns observed in species of the genus Haloarcula [4], validating the taxonomic reclassification of Halomicroarcula saliterrae and Halomicroarcula onubensis within this genus. TAXONOMIC CONCLUSIONS The species names Halomicroarcula saliterrae and Halomicroarcula onubensis were validly published under the ICNP according to Validation List no. 217 [18], concurrently with the six Haloarcula species described by Ma et al. [4] that also proposed the unification of the genera Haloarcula and Halomicroarcula. Section 7 of the ICNP [40] addresses changes in names of taxa as a result of transference, union, or change in rank, with actions depending on taxonomic judgement. With this study we Fig. 3. Heatmap displaying OrthoANI (upper right) and dDDH (lower left) percentages among the species Halomicroarcula saliterrae S1CR25- 12T and Halomicroarcula onubensis S3CR25- 11T, and members of the genus Haloarcula and other related species of the family Haloarculaceae. 6 Straková etal., Int. J. Syst. Evol. Microbiol. 2024;74:006510 confirm and support the proposal by Ma et al. [4] about the unification of the genera Haloarcula and Halomicroarcula. Moreover, following the phylogenetic, phylogenomic, and comparative genomic analysis to ascertain their taxonomic status, together with a comparative examination of phenotypic features and polar lipid profiles, we confirmed that Halomicroarcula saliterrae and Halomicroarcula onubensis should not be considered as separate species of another genus, since they belong to the expanded genus Haloarcula. Consequently, we propose the reclassification of Halomicroarcula saliterrae Straková et al. 2024 and Halomicroarcula onubensis Straková et al. 2024 into the genus Haloarcula, as Haloarcula saliterrae comb. nov. and Haloarcula onubensis comb. nov., respectively. DESCRIPTION OF HALOARCULA SALITERRAE COMB. NOV. Haloarcula saliterrae ( sa. li. ter’rae. L. masc. n. sal, salt; L. fem. n. terra, land; N.L. gen. n. saliterrae, of saline soil). Basonym: Halomicroarcula saliterrae Straková et al. 2024. Fig. 4. Heatmap showing AAI percentages among Haloarcula species, including the species Halomicroarcula saliterrae S1CR25- 12T and Halomicroarcula onubensis S3CR25- 11T, and other related species of the family Haloarculaeae. 7 Straková etal., Int. J. Syst. Evol. Microbiol. 2024;74:006510 The description is identical to that given for Halomicroarcula saliterrae by Straková et al. 2024. The type strain is S1CR25- 12T (=CECT 30620T=CCM 9252T). DESCRIPTION OF HALOARCULA ONUBENSIS COMB. NOV. Haloarcula onubensis ( o. nu. ben’sis. L. fem. adj. onubensis, of or belonging to Onuba, the ancient Latin name of Huelva, a city in Spain, where the type strain was isolated). Table 1. Differential characteristics between the species Halomicroarcula saliterrae S1CR25- 12T and Halomicroarcula onubensis S3CR25- 11T, and related species of the genus Haloarcula Characteristic Hma. saliterrae S1CR25- 12T* Hma. onubensis S3CR25- 11T* Har. rara SHR3T† Har. laminariae LYG- 108T‡ Har. salinisoli F24AT§ Har. vallismortis ATCC 29715T¶ Morphology Pleomorphic rods Pleomorphic rods Pleomorphic Rods Rods Pleomorphic Colony pigmentation Red Orange red Red Red Pink Pink NaCl requirement: Range (%, w/v) 15–30 12–30 8–28 8–28 15–30 >15 Optimum (%, w/v) 25 25 18 15 25 25 Temperature requirement: Range (°C) 20–50 25–55 25–55 20–50 25–50 20–45 Optimum (°C) 37 37 37 40 37 40 pH requirement: Range 6.0–9.0 6.0–9.0 5.5–8.5 5.0–9.5 6.0–8.5 5.5–8.5 Optimum 7.0–8.0 7.0–8.0 7.0 7.0 7.0–7.5 7.4–7.5 Hydrolysis of: Gelatin − − − − + − Tween 80 − − + − + − Reduction of nitrite − − − − + − Utilization as sole carbon and energy source: Citrate − − + − + nd d- Galactose + − + + − + d- Sorbitol − − − + + nd Fumarate − + + + + nd Glycerol − − + + − + Sucrose − + + + + + Lactose − − + − − − d- Xylose − + − − − − Maltose + − − − + + Glycine + − − + nd nd l- Alanine + − − − nd nd *Data from Straková et al. [17]. †Data from Ma et al. [4]. ‡Data from Ma et al. [12]. §Data from Durán- Viseras et al. [3]. ¶Data from Gonzalez et al. [39]. nd, Not determined. 8 Straková etal., Int. J. Syst. Evol. Microbiol. 2024;74:006510 Basonym: Halomicroarcula onubensis Straková et al. 2024. The description is identical to that given for Halomicroarcula onubensis by Straková et al. 2024. The type strain is S3CR25- 11T (=CECT 30621T=CCM 9254T). Funding information This study was supported by grant PID2020- 118136GB- I00, funded by MCIN/AEI/10.13039/501100011033 (to A.V. and C.S.-P.). Acknowledgements We thank David R. Arahal for his help with nomenclature issues and critical reading of the manuscript. Conflicts of interest The authors declare that there are no conflicts of interest. References 1. Torreblanca M, Rodriguez- Valera F, Juez G, Ventosa A, Kamekura M, et al. 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