Photobacterium kishitanii sp. nov., a luminous marine bacterium symbiotic with deep-sea fishes

Abstract

Six representatives of a luminous bacterium commonly found in association with deep, colddwelling marine fishes were isolated from the light organs and skin of different fish species. These bacteria were Gram-negative, catalase-positive, and weakly oxidase-positive or oxidase-negative. Morphologically, cells of these strains were coccoid or coccoid-rods, occurring singly or in pairs, and motile by means of polar flagellation. After growth on seawater-based agar medium at 22 6C for 18 h, colonies were small, round and white, with an intense cerulean blue luminescence. Analysis of 16S rRNA gene sequence similarity placed these bacteria in the genus Photobacterium. Phylogenetic analysis based on seven housekeeping gene sequences (16S rRNA gene, gapA, gyrB, pyrH, recA, rpoA and rpoD), seven gene sequences of the lux operon (luxC, luxD, luxA, luxB, luxF, luxE and luxG) and four gene sequences of the rib operon (ribE, ribB, ribH and ribA), resolved the six strains as members of the genus Photobacterium and as a clade distinct from other species of Photobacterium. These strains were most closely related to Photobacterium phosphoreum and Photobacterium iliopiscarium. DNA-DNA hybridization values between the designated type strain, Photobacterium kishitanii pjapo.1.1 T , and P. phosphoreum LMG 4233 T , P. iliopiscarium LMG 19543 T and Photobacterium indicum LMG 22857 T were 51, 43 and 19 %, respectively. In AFLP analysis, the six strains clustered together, forming a group distinct from other analysed species. The fatty acid C 17 : 0 cyclo was present in these bacteria, but not in P. phosphoreum, P. iliopiscarium or P. indicum. A combination of biochemical tests (arginine dihydrolase and lysine decarboxylase) differentiates these strains from P. phosphoreum and P. indicum. The DNA G+C content of P. kishitanii pjapo.1.1 T is 40.2 %, and the genome size is approximately 4.2 Mbp, in the form of two circular chromosomes. These strains represent a novel species, for which the name Photobacterium kishitanii sp. nov. is proposed. The type strain, pjapo

Crystal Structures of the Lumazine Protein fromPhotobacterium kishitaniiin Complexes with the Authentic Chromophore, 6,7-Dimethyl- 8-(1′-d-Ribityl) Lumazine, and Its Analogues, Riboflavin and Flavin Mononucleotide, at High Resolution

“…The sequence was identical to that of P. phosphoreum. This is not surprising, because all strains of luminous bacteria share very high nucleotide sequence homology (2,20). LumP from Photobacterium kishitanii (LumP PK ) has been expressed as inclusion bodies in E. coli, solubilized in a 1.5 M urea buffer containing RBF, refolded by removing the urea, and finally, purified by column chromatography.…”

Natural Merodiploidy of thelux-ribOperon ofPhotobacterium leiognathifrom Coastal Waters of Honshu, Japan

“…To test for recombination between the genes of lux-rib 1 and lux-rib 2 in lnuch.13.1, lelon.2.1, and lnuch.21.1, we developed a partition congruence analysis, as described here (script to produce PAUP* commands are available on request). Included in the analysis were lux-rib 1 of P. leiognathi ATCC 25521 T , the lux-rib operon of P. mandapamensis ATCC 27561 T , and, as an outgroup, the lux-rib operon of P. kishitanii ATCC BAA-1194 T (also known as pjapo.1.1 T ) (4). Omitted from the analysis were noncoding intergenic spacer regions, the luxF gene in P. mandapamensis and P. kishitanii, and the first 341 bp of the luxC sequence, which is not available for the lux-rib 2 operons of lelon.2.1 and lnuch.21.1 (Fig.…”

“…Linked to the luminescence genes in some Photobacterium species, and apparently cotranscribed with them, are genes involved in the synthesis of riboflavin, forming an operon of 10 or 11 genes, luxCDAB(F)EG-ribE-BHA, which we refer to here as the lux-rib operon (23, 26, 27, 34, 37; this study). Upstream of the lux-rib operon in P. mandapamensis, a species closely related to P. leiognathi, are lumQ and lumP (encoding proteins of the lumazine operon), and these genes are located adjacent to the putA gene (encoding proline dehydrogenase) (29,30,31).Phylogenetic analysis of lux and rib genes, together with housekeeping genes such as the16S rRNA gene, gyrB, pyrH, recA, rpoA, and rpoD, has proven helpful in defining evolutionary relationships among luminous bacteria and in the identification of new species (2,3,4,12,23). Phylogenetic analysis based on lux and other genes has also proven effective in providing the bacterial species-and clade-level resolution necessary for testing hypotheses of symbiont-host specificity and evolutionary codivergence in bioluminescent symbioses (16,23).…”

“…The genes for bacterial light production are present as an operon, luxCDABEG: luxA and luxB encode the ␣ and ␤ subunits of luciferase; luxC, luxD, and luxE specify the enzymatic components of a fatty acid reductase complex necessary for synthesis and recycling of the aldehyde substrate; and luxG encodes a flavin reductase (14). Most luminous Photobacterium species, i.e., Photobacterium phosphoreum, Photobacterium kishitanii, and Photobacterium mandapamensis, also carry luxF, which encodes a nonfluorescent flavoprotein, with a lux operon gene order of luxCD-ABFEG (2,3,4,23,28,37). Linked to the luminescence genes in some Photobacterium species, and apparently cotranscribed with them, are genes involved in the synthesis of riboflavin, forming an operon of 10 or 11 genes, luxCDAB(F)EG-ribE-BHA, which we refer to here as the lux-rib operon (23, 26, 27, 34, 37; this study).…”

“…Phylogenetic analysis of lux and rib genes, together with housekeeping genes such as the16S rRNA gene, gyrB, pyrH, recA, rpoA, and rpoD, has proven helpful in defining evolutionary relationships among luminous bacteria and in the identification of new species (2,3,4,12,23). Phylogenetic analysis based on lux and other genes has also proven effective in providing the bacterial species-and clade-level resolution necessary for testing hypotheses of symbiont-host specificity and evolutionary codivergence in bioluminescent symbioses (16,23).…”