Strain JP2^T (= DSM 5205 = ATCC 35100) is the type strain of Thiothrix nivea [¹,²] which is the type species of the genus Thiothrix [¹,²]. Cultures of the species were first observed and classified as “Beggiatoa nivea” in 1865 by Rabenhorst [³] and later (1888) placed into the novel genus Thiothrix by Winogradsky [²]. The species was included on the Approved List of Bacterial Names Amended edition in 1980 [⁴]. Axenic cultures isolated from sulfide-containing well water became available in 1980 through the work of J. M. Larkin [⁵], with the formal description of strain JP2^T as the neotype strain of the species T. nivea in 1983 [¹], as well as strain JP1 as a reference strain within the species [¹]. The generic name derives from the Neo-Greek words theion, sulfur, and thrix, hair [⁶]. The species epithet is derived from the Latin word nivea snow-white [⁶]. The species became well known for its sulfur granules, the gliding motility and the typical rosettes [¹], which were first observed by Winogradsky [²]. Here we present a summary classification and a set of features for T. nivea JP2^T, together with the description of the complete genomic sequencing and annotation.

A representative genomic 16S rRNA sequence of T. nivea JP2^T was compared using NCBI BLAST [⁷] under default settings (e.g., considering only the high-scoring segment pairs (HSPs) from the best 250 hits) with the most recent release of the Greengenes database [⁸] and the relative frequencies of taxa and keywords (reduced to their stem [⁹]) were determined, weighted by BLAST scores. The most frequently occurring genus was Thiothrix (100.0%, 12 hits in total). Regarding the single hit to sequences from members of the species, the average identity within HSPs was 99.5%, whereas the average coverage by HSPs was 99.4%. Regarding the four hits to sequences from other members of the genus, the average identity within HSPs was 94.2%, whereas the average coverage by HSPs was 96.0%. Among all other species, the one yielding the highest score was, Thiothrix fructosivorans (GU269554) which corresponded to an identity of 94.5% and an HSP coverage of 100.0%. (Note that the Greengenes database uses the INSDC (= EMBL/NCBI/DDBJ) annotation, which is not an authoritative source for nomenclature or classification.) The highest-scoring environmental sequence was AM490765 ('Linking and functional nutrient spiraling mats (USA) microbial mat sulfidic cave spring Lower Kane Cave Big Horn LKC22 clone SS LKC22 UB32'), which showed an identity of 96.7% and an HSP coverage of 100.0%. The most frequently occurring keywords within the labels of environmental samples which yielded hits were 'sulfid' (4.2%), 'microbi' (4.0%), 'biofilm' (3.4%), 'cave' (2.8%) and 'karst' (2.7%) (238 hits in total). Environmental samples which yielded hits of a higher score than the highest scoring species were not found. These keywords reflect the ecological properties reported for the species and strain JP2^T in the original description [¹,²].

Figure 1 shows the phylogenetic neighborhood of T. nivea in a 16S rRNA based tree. The sequences of the two identical 16S rRNA gene copies in the genome do not differ from the previously published 16S rRNA sequence (L40993), which contains six ambiguous base calls.

Cells of strain JP2^T are rod shaped with various lengths (Figure 2). Cultures of T. nivea contain gliding gonidia, filaments and rosettes (= aggregations of gonidial cells, not visible in Figure 2) [¹]. The presence of a sheath was first reported in the 19^th century [²] and later confirmed for the neotype strain [¹]. The sheath contains several separate layers [¹] of so far unknown structure. Motility was observed, but no flagella [¹]. Numerous genes allocated to the functional role category motility were identified in the genome (see below). Many of these genes might be involved in the formation of the polar located fimbriae [³²]. The typical rosettes generated by T. nivea are known from sulfide-containing waters [¹,²]. Sulfur granules are invaginated by the cells, as reported in detail by Larkin and Shinabarger [¹]. Strain JP2^T stains Gram-negative, and grows only aerobically, best within a temperature range of 20 – 30°C [¹]. Both the neotype strain and reference strain JP1 produce oxidase, but not catalase. The strains also produce poly-β-hydroxybutyrate [¹]. Strain JP2^T uses only four carbon sources; acetate, malate, pyruvate and oxalacetate [¹]. Ammonia and nitrate (but not nitrite) are used as sole nitrogen sources [¹]. The sole sulfur sources are sulfide and thiosulfate. What remains unresolved, based on the literature is whether or not T. nivea is autotrophic, obtaining carbon from CO₂ and energy via oxidation of sulfide as reported by Winogradsky [¹³] or not, as reported by Larkin and Shinabarger [¹]. In the case in which strain JP2^T could use CO₂ as a carbon source as well as acetate, malate, pyruvate and oxalacetate, while oxidizing the reduced sulfur compounds, it could be considered to be a mixotroph [¹].

The genome consists in the current assembly of 15 contigs in four scaffolds with a length of 5,599 bp, 7,015 bp, 40,927 bp, and 4,638,170 bp, respectively, and a G+C content of 54.9% (Table 3). Of the 4,594 genes predicted, 4,542 were protein-coding genes, and 52 RNAs; 213 pseudogenes were also identified. The majority of the protein-coding genes (98.8%) were annotated as hypothetical proteins. The distribution of genes into COGs functional categories is presented in Table 4.

The genomic basis for gliding motility is not yet completely resolved, but the requirement of the genes gldA, gldF and gldG was described [⁴⁴]. Genes gldA, gldF and gldG exhibit a high degree of sequence similarity to components of ABC transporters [⁴⁴]. A closer examination of the JP2^T genome revealed a region of three genes (Thini_0004.00016790, Thini_0004.00016780, Thini_0004.00016770, currently annotated as ABC-type uncharacterized transport system), for which the derived protein sequences show high similarity to GldA, GldF and GldG of Bdellovibrio bacteriovorus HD100 (Bd1023, Bd1024 and Bd1025) [⁴⁵]. The requirement of gldA for gliding motility was experimentally shown for Flavobacterium johnsoniae. A non-motile mutant lacking the intact gldA gene was complemented by a vector carrying an intact gldA gene. The motility of the mutant was restored [⁴⁶].

While we were able to locate a phosphoenolpyruvate carboxylase gene in the genome, (Thini_0004.00035050), we could not identify a gene for malate dehydrogenase. Unless T. nivea encodes a malate dehydrogenase that is not homologous to other malate dehydrogenases, we can not confirm for the neotype strain the genomic basis for the speculation that the original T. nivea culture can fix CO₂ as reported by Winogradsky [¹³] and is a mixotroph [¹].

We would like to gratefully acknowledge the help of Anja Frühling (DSMZ) for growing T. nivea cultures. This work was performed under the auspices of the US Department of Energy Office of Science, Biological and Environmental Research Program, and by the University of California, Lawrence Berkeley National Laboratory under contract No. DE-AC02-05CH11231, Lawrence Livermore National Laboratory under Contract No. DE-AC52-07NA27344, and Los Alamos National Laboratory under contract No. DE-AC02-06NA25396, UT-Battelle and Oak Ridge National Laboratory under contract DE-AC05-00OR22725, as well as German Research Foundation (DFG) INST 599/1-1.