Tildeniella torsiva Mai, J.R. Johansen and Pietrasiak

Kim, So-Won, Lee, Nam-Ju, Kim, Do-Hyun, Song, Ji-Ho & Lee, Hye-Ryeung Wang and Ok-Min, 2022, Five newly recorded species of cyanobacteria in Korea, Journal of Species Research 11 (4), pp. 296-309 : 303-307

Tildeniella torsiva Mai, J.R. Johansen and Pietrasiak ,

2018 ( Fig. 5 , Table 2)

Colony forming irregular fascicules, bright blue green in color, olive green with maturity. Filaments often entan- gled, form false branching in older cultures. Trichomes straight, curved, waved, or sometimes coiled, not attenuated towards ends, slightly constricted at cross-walls. Sheaths thin, firm, colorless. Cells mostly longer than wide, rarely isodiametric, ungranulated, 1.1-1.9 μm wide, 1.6-4.9 μm long. Apical cells rounded. Hormogonia and necridic cells absent.

Ecology. On a limestone wall ( Mai et al., 2018), planktonic in freshwater in Korea.

Distribution. Slovakia ( Mai et al., 2018).

Site collection. Wonuri, Hoehyeon-myeon, Gunsan-si , Jeollabuk-do (35°55′42.8″N / 126°46′22.1″E) .

Date of collection. April 24, 2019.

16S rRNA and phylogenetic affiliation

In this study, 16S rRNA gene sequences of cyanobacteria were obtained. Molecular phylogenetic analysis of ML and Bayesian was performed on 16S rRNA gene sequences of the most similar and closely related species registered in NCBI (National Center for Biotechnology Information). As a result, two phylogenetic trees were completed, one tree containing species belonging to order Nostocales , and the other containing species belonging to order Synechococcales ( Figs. 6 , 7 ). Additionally, nucleotide sequence similarity and genetic distance were analyzed. When the similarity of the 16S rRNA gene sequence is 98.5% or more, it can be determined as the same species ( Kim et al., 2014).

In the case of Sphaerospermopsis reniformis (FBCC- A194), molecular phylogenetic analysis was performed based on 16S rRNA gene sequences of S. reniformis and related taxa registered in NCBI. As a result, similar branch patterns were shown in all trees ( Fig. 6 ). S. reniformis (FB- CC-A194) collected in Korea was included in the same cluster as previously reported S. reniformis (06-01, 07- 01). Results of sequence similarity and genetic distance analysis revealed that it showed a sequence similarity of 99.9-100% and a genetic distance of 0.00% to S. reniformis (06-01, 07-01) ( Table 3). In this study, S. reniformis (FBCC-A194) also showed high sequence similarity and close genetic distance to S. kisseleviana and S. aphanizomenoides (similarity: 99.9% and 99.4%, respectively; genetic distance: 0.00% and 0.01%, respectively). However, for the morphology of trichomes, S. kisseleviana and S. aphanizomenoides are straight or slightly curved, whereas S. reniformis has a screw-like coil. Thus, they can be clearly distinguished morphologically ( Zapomělová et al., 2009). S. reniformis (FBCC-A194) can prove that it is S. reniformis because it has screw-like coiled trichomes. The high 16S rRNA gene sequence similarity of S. reniformis , S. kisseleviana , and S. aphanizomenoides supports the previously reported content that the presence or absence of coiling of trichomes is not divided in the phylogenetic tree ( Rajaniemi et al., 2005a; 2005b).

In the case of Pelatocladus maniniholoensis (FBCC- A1476), a phylogenetic analysis was performed through 16S rRNA gene sequences of P. maniniholoensis and related taxa registered in NCBI. All trees showed similar branching patterns ( Fig. 6 ). Since P. maniniholoensis (FBCC-A1476) collected from Ulleungdo was included in the same cluster as the previously reported P. maniniholoensis (HA4357-MV3), the species could be clearly distinguished from the phylogenetic tree based on 16S rRNA gene sequences. As a result of additional sequence similarity and genetic distance analysis, it showed a sequence similarity of 99.5% and a genetic distance of 0.00% to P. maniniholoensis (HA4357-MV3) ( Table 3). Also, Hapalosiphon hibernicus (BZ-3-1), which has the highest sequence similarity in the NCBI database, is 100% identical to 16S rRNA gene sequences of P. maniniholoensis (HA 4357-MV3). H. hibernicus (BZ-3-1) has been considered as a misidentification of P. maniniholoensis ( Miscoe et al., 2016; Casamatta et al., 2020).

In the case of Tolypothrix carrinoi (FBCC-A206, FBCC- A207, FBCC-A208), a molecular phylogenetic analysis was performed based on 16S rRNA gene sequences of T. carrinoi and related taxa registered in NCBI. As a result, all trees showed similar branching patterns ( Fig. 6 ). Tolypothrix carrinoi (FBCC-A206, FBCC-A207, FBCC- A208) collected in Korea was included in the same cluster as previously reported T. carrinoi (HA7290-LM1). As a result of additional sequence similarity and genetic distance analysis, it showed 98.9-99.2% similarity and 0.01% genetic distance with Tolypothrix carrinoi (HA7290-LM1) ( Table 3).

In the case of Myxacorys chilensis (FBCC-A216, FBCC-A220), molecular phylogenetic analysis based on 16S rRNA gene sequences of M. chilensis and related taxa registered in NCBI was performed. All trees showed similar branching patterns ( Fig. 7 ). M. chilensis (FBCC- A216, FBCC-A220) collected in this study was includ- ed in the same cluster as previously reported Myxacorys chilensis (ATA2-1-KO14). It is clearly separated from M. californica and M. almedinensis , other species included in the genus Myxacorys . Therefore, it could be clearly distinguished through the phylogenetic tree based on the 16S rRNA gene sequences. Additional nucleotide sequence similarity and genetic distance analysis results showed a similarity of 99.0-99.5% and a genetic distance of 0.00- 0.01% with M. chilensis ( Table 3).

In the case of Tildeniella torsiva (FBCC-A1474), molecular phylogenetic analysis was performed based on 16S rRNA gene sequences of T. torsiva and related taxa registered in NCBI. All trees showed similar branching patterns ( Fig. 7 ). The culture strain of this study, T. torsiva (FBCC-A1474), was included in the same cluster as previously reported T. torsiva (UHER1998/13D). It is clearly distinguished from T. alaskaensis included in the genus Tildeniella . Another species included in the genus Tildeniella , T. nuda , was excluded because it was included in a distant cluster from the genus Tildeniella . Additionally, as a result of analysis of nucleotide sequence similarity and genetic distance, it showed a similarity of 99.8% and a genetic distance of 0.00% with T. torsiva ( Table 3). Stackebrandt and Goebel (1994) have stated that a taxon is classified as a different genus when the 16S rRNA gene sequence similarity is less than 95%. In this study, T. nuda (Zehnder 1965/U140) showed 91.8% nucleotide sequence similarity and 0.07% genetic distance to T. torsiva (UHER1998/13D). Strunecky et al. (2020) have men- tioned that T. nuda shows 91.3% similarity to T. torsiva . It should be classified as a different genus because it has less than 95% sequence similarity to T. torsiva , a type species of genus Tildeniella . Mai et al. (2018) first reported T. nuda and stated that T. nuda 16S rRNA gene shared 99.1% sequence similarity with that of T. torsiva . This result is considered to be in error.

As described above, the five species collected in Korea were identified through morphological characteristics and phylogenetic analysis based on 16S rRNA gene sequenc- ing. They were added to unrecorded genus and species in Korea.

Figures

Fig. 5. Microphotographs of Tildeniella torsiva from the strain FBCC-A1474. (A) Arrangement of filament in the colony, (B) Curved terminal cells, (C) Apical cell, (D, E) Sheaths(sh), (F, G) False branching, (H-J) Morphologies of trichomes, (K) Diversity of cells morphology. Scale bars=(A) 20μm, (B-K) 10μm.

Fig. 6. Maximum-Likelihood (ML) phylogenetic tree based on 16S rRNA gene sequences of Sphaerospermopsis reniformis, Pelatocladus maniniholoensis, Tolypothrix carrinoi and other cyanobacterial strains. 16S rRNA gene sequences of genera (Gloeobacter violaceus PCC 7421, Synechococcus elongatus PCC 6301) were included as the outgroups.Additionally, the probability of Bayesian analysis was incorporated into the ML tree to support the strength of each branch. The first and second numbers at the nodes display the bootstrap proportions (>50%) in ML and posterior probabilities(>0.50) in Bayesian analysis, respectively.The branch lengths are proportional to the scale given.

Fig. 7. Maximum-Likelihood (ML) phylogenetic tree based on 16S rRNA gene sequences of Myxacorys chilensis, Tildeniella torsiva and other cyanobacterial strains. 16S rRNA gene sequences of genera (Gloeobacter violaceus PCC 7421, Pseudanabaena catenata PCC 7408) were included as the outgroups.Additionally, the probability of Bayesian analysis was incorporated into the ML tree to support the strength of each branch. The first and second numbers at the nodes display the bootstrap proportions (>50%) in ML and posterior probabilities (>0.50) in Bayesian analysis, respectively. The branch lengths are proportional to the scale given.