Pyomelanin-Producing sp. nov., Isolated From Grape ( L.).

A novel endophytic bacterial strain, designated GR-TSA-9, was isolated from surface-sterilized grape ( L.). 16S rRNA gene sequence analyses showed that the isolate was grouped within the genus , displaying the highest similarity with DS-18 (97.9%) and KSL-102 (97.8%) and less than 97.5% similarity with other members of . The strain GR-TSA-9 was a gram negative, rod shaped, facultatively anaerobic, catalase and oxidase positive, and motile bacterium. Its growth occurred at 10-37°C (optimally 25-30°C), at pH 7.0-8.0, and in NaCl 0-1% (optimally 0%). It contained ubiquinone-10 as a respiratory quinone, and the major cellular fatty acids (>10% of the total) were C (14.2%) and summed feature 8 (Cω7 and/or Cω6c, 65.6%). The polar lipids present in the strain were phosphoglycolipids, phosphatidylglycerol, 1,2-di-acyl-3-O-[d-glucopyranosyl-(1→4)-α-d-glucopyranuronosyl]glycerol, and unidentified lipids (L1, L2, and L4). The strain had one 2,976,716bp circular chromosome with a G+C content of 66.4%. The digital DNA-DNA hybridization value between strain GR-TSA-9 and DS-18 was 20.9%, while the average nucleotide identity value was 76.7%. In addition, the dDDH and ANI values to other members in this genus, whose genome sequences are available, are less than 21.1 and 77.6%. Genome annotation predicted the presence of some gene clusters related to tyrosine degradation and pyomelanin formation. Strain GR-TSA-9 produced a brown melanin-like pigment in the presence of L-tyrosine-containing media. The highest pigment production (0.19g/L) was observed in tryptic soy broth with 1.0mg/ml L-tyrosine at 25°C for 6days of culture. Biophysical characterization by ultraviolet (UV)-visible spectroscopy, Fourier-transform infrared spectroscopy, and electrospray ionization mass spectrometry confirmed that the pigment was pyomelanin. Additionally, melanized GR-TSA-9 cells could protect the cells against UVC exposure. The phylogenetic, genomic, phenotypic, and chemotaxonomic features indicated that strain GR-TSA-9 represents a novel melanin-producing species of . The type strain was GR-TSA-9 (KCTC 82386=CGMCC 1.18820).