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Microorganisms 08 01132

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34 views1 page

Microorganisms 08 01132

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joshua lara
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Microorganisms 2020, 8, 1132 4 of 14

and the NCBI prokaryotic genome annotation pipeline (PGAP) (2020-03-30.build4489—https://github.


com/ncbi/pgap) with default parameters and databases for all of the five software/pipelines [25–29].
The functional annotation of predicted coding DNA sequences (CDSs) was further blasted with NCBI
(v2.10.0+), and UniProtKB database (release 2020_04). Hydrogenase classification was checked using
the HydDB webtool (https://services.birc.au.dk/hyddb/) [30].

2.3. Clustered Regularly Interspaced Short Palindromic Repeats (CRISPRs) and Genomic Islands
Identification and classification of the CRISPR–Cas systems were performed by using the
CRISPRCas Finder webserver, with default parameters (https://crisprcas.i2bc.paris-saclay.fr/) [31].
The prediction of laterally transferred gene clusters (genomic islands) was performed with the
IslandViewer4 webserver (http://www.pathogenomics.sfu.ca/islandviewer/) based on an EMBL file
generated by Dfast [32].

2.4. Geographical Distribution


The geographical distribution of D. thermomarina was studied at species and genus level within
the 16S rRNA gene sequences available in the databases and in the public metagenomes deposited
at the GBIF (Global Biodiversity Information Facility) facility (https://www.gbif.org/) and in the
NCBI database.

2.5. Taxonomical Analyses and Comparative Genomics


To study the taxonomic position of the strain, we used GTDB-Tk (v1.1.1—https://github.com/
Ecogenomics/GTDBTk) to place the genome on a tree made of concatenated reference proteins, we
compared by blast the 16S rRNA CDS obtained from genomic assembly to the sequences in NCBI
(v2.10.0+) and performed a tetra correlation comparison search with the JSpecies webserver against its
own database (http://jspecies.ribohost.com/jspeciesws/).
The genome of D. thermomarina was compared by subtractive comparative genomics to the
genomes of the hydrothermal bacteria Thermosulfurimonas marina (ASM1231758v1), Thermosulfuriphilus
ammonigenes (ASM1120745v1), Dissulfuribacter thermophilus (ASM168733v1), and Thermosulfurimonas
dismutans (ASM165258v1) to identify potential genetic markers of DNRA, and of thiosulfate
disproportionation, two physiological properties absent in D. thermomarina, explored by excluding
D. thermomarina’s genome. These genomes were compared by using the MaGE platform Pan-genome
Analysis tool (https://mage.genoscope.cns.fr/microscope/home/index.php), based on the clustering
algorithm SiLiX (http://lbbe.univ-lyon1.fr/-SiLiX-.html) which clustered genomic CDSs by 50% amino
acid identity and 80% amino acid alignment coverage, with permissive parameters. Resulting
CDSs were blasted on the UniprotKB database and hypothetical protein CDSs were analyzed with
InterProScan webserver (https://www.ebi.ac.uk/interpro/) for functional predictions.
Finally, to evaluate the hypothesis of Ward et al. (2020) [21] suggesting that the tail truncation of the
AprB protein could be a molecular marker of the disproportionation capacity of sulfur, we extracted
the CDS encoding the AprB protein (based on Prokka annotation) from the genomes of characterized
sulfur disproportioners or sulfate reducers: Thermosulfurimonas marina, Thermosulfuriphilus ammonigenes,
Thermosulfurimonas dismutans, Dissulfuribacter thermophilus, Thermodesulfatator atlanticus, Thermodesulfatator
autotrophicus and Thermodesulfatator indicus, in addition to that of D. thermomarina [4–8,33–35]. The AprB
putative protein sequences were then aligned and their length were calculated.

3. Results and Discussions

3.1. General Genome Properties and Genomic Islands


The complete genome sequence of Dissulfurirhabdus thermomarina strain SH388T was deposited in
GenBank databases under the accession number JAATWC000000000. The strain is available in the
DSMZ culture collection under the accession number DSM 100025T and in All-Russian Collection

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