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Halolitoreus marinus gen. nov., sp. nov. and Halolitoreus rarus sp. nov., halophilic archaea isolated from diverse coastal tidal flats, and proposal of the novel family Halolitoreaceae fam. nov. in the order Halobacteriales within the class Halobacteria.

Coastal tidal flats represent dynamic saline environments that harbor largely unexplored haloarchaeal communities. In this study, amplicon sequencing, metagenomic analyses, and cultivation-based approaches revealed substantial haloarchaeal diversity in tidal flats from four provinces of eastern China despite their relatively low salinity. Five haloarchaeal strains, designated YSMS36T, DYSN1, QDMS2, CMSO5T, and ZSTT2, were isolated from diverse tidal flats. Theses strains shared 16S rRNA gene sequence similarities of 92.1-92.2% with their closest validly named relative, Salinilacihabitans rarus AD-4T. Phylogenetic analyses based on 16S rRNA and rpoB' gene sequences showed that the five strains formed a distinct and well-supported monophyletic lineage, separated from currently recognized members of the class Halobacteria. Average amino acid identity (AAI), average nucleotide identity (ANI), and digital DNA-DNA hybridization (dDDH) values between these five strains and the related Halobacteria representatives were 49.3-62.7%, 66.9-74.9%, and 16.2-29.5%, respectively, and well below the accepted thresholds for species and genus delineation. Phylogenomic analyses further supported their placement within a novel family of the order Halobacteriales. Based on phylogenetic, genomic, chemotaxonomic, and phenotypic analyses, these five strains represent two novel species of a novel genus within a novel family. The names, Halolitoreaceae fam. nov., Halolitoreus marinus gen. nov., sp. nov., and Halolitoreus rarus sp. nov. are herein proposed.

Coastal tidal flat

Halorubrum marinum sp. nov., Halorubrum rarum sp. nov., Halorubrum wangae sp. nov., Halorubrum shenae sp. nov., and Halorubrum zhoui sp. nov., halophilic archaea from coastal tidal flats, a saline lake, and a marine solar saltern.

Five novel halophilic archaeal strains, designated DTA46T, DTA98T, HHNYT27T, N11T, and SY-15T, were isolated from diverse saline environments across various regions of China. Amplicon and metagenome analyses revealed that three amplicon reads were affiliated with strains DTA46T, HHNYT27T, and N11T while two MAGs related to strains N11T and SY-15T. The sequence similarities among these five strains and current species of the genus Halorubrum were 93.1%-99.1% and 86.0%-95.9% judged by 16S rRNA and rpoB' genes, respectively. Phylogenomic and comparative genomic analyses revealed their close affiliation with Halorubrum. The average nucleotide identity (ANI), digital DNA-DNA hybridization (dDDH), and average amino acid identity (AAI) values between these strains and existing Halorubrum species ranged from 74.9%-93.6%, 22.3%-58.3%, and 68.3%-93.7%, respectively. All are below the recommended thresholds for species delineation, which supports their classification as novel taxa. The growth characteristics of strains DTA46T, DTA98T, HHNYT27T, N11T, and SY-15T were determined as follows: temperature range 20-60 °C (optima: 35, 37-42, 37, 35, and 42 °C), NaCl concentration 1.4-5.5 M (optima: 2.6, 3.1, 3.1, 3.1, and 5.1 M), and pH range 5.5-9.5 (optima: 8.0, 8.0, 7.0, 7.5, and 7.0). Based on the polyphasic characterization integrating phenotypic, chemotaxonomic, phylogenetic, and phylogenomic evidence, strains DTA46T, DTA98T, HHNYT27T, N11T, and SY-15T are proposed to represent five novel species of the genus Halorubrum, for which the names Halorubrum marinum sp. nov., Halorubrum rarum sp. nov., Halorubrum wangae sp. nov., Halorubrum shenae sp. nov., and Halorubrum zhoui sp. nov. are designated, respectively.

Phylogeny

Land use conversion to uplands significantly increased the risk of antibiotic resistance genes in estuary area.

Land use conversion in estuary wetlands may affect the transmission of antibiotic resistance genes (ARGs), while the risk rank of the ARGs and the change of clinically relevant ARGs under various land-use types are not well understood. This study used metagenomics to reveal the diversity and abundance of ARGs across five distinct land uses: reed wetland, tidal flat, grassland, agricultural land and fallow land, as well as their distribution and potential health risks. Results showed that high numbers of ARG subtypes and classes were detected irrespective of land-use types, notably higher in agricultural land (144 ARG subtypes). The most shared ARG subtypes were multidrug resistance genes across all the land uses (29 subtypes, 4.7 × 10-2-1.5 × 10-1 copies per 16S rRNA gene copy). Proteobacteria and Actinobacteria were primary ARG hosts, with 18 and 15 ARGs were found in both of them, respectively. The ARG subtype mdtB was the most dominant clinical ARG detected with 90 % amino acid identity. The change of ARGs exhibited a consistent trend across land uses in terms of health risk ranks, with the highest observed in fallow land and the lowest in reed wetland. This study reveals the distribution pattern of ARGs across various land-use types, and enhances our understanding of the potential health risks associated with ARGs in the context of coastal wetland conversion in estuary areas.

Estuaries