Polyphasic Taxonomy Of Two Potential Novel Actinomycetes Species

Two potential novel species of actinomycetes, one of the strains K55was isolated fromVicia cracca soil sample which was polluted by nickel in lueyang county Shaanxi Province,the other one D34was isolated from Stipa grandis vegetation rhizosphere soil in yuwushan. Apolyphasic study was carried out to confirm the taxonomic position of the strains.Morphological, physiological and biochemical, chemotaxonomic characteristics,16SrRNA phylogeny, combined with DNA-DNA hybridization were used to identify theirtaxonomic status.The Morphological characteristics indicated that the aerial mycelium and substratemycelium of K55both grow well, have widely branches, orangered mycelium, wet bacterialcolony, produce spores and black pigment. The mycelium has no break, no transverse septa,the spore has single, round and smooth surface which is born at the surface ofmycelium.These morphological characteristics coincided with the typical characteristics ofMicromonospora. So K55was tentatively assigned to Micromonospora. The other one D34,aerial mycelium and substrate mycelium both grow well, have widely branches,well-developed aerial mycelium, white aerial mycelium, comparatively wet bacteria colony,produce spores and faint yellow pigment. The mycelium has no break, no transverse septa,and developed spiral spore chains. The spore is spheroidicity, with non-motile noduler towarty spore surfaces. These morphological characteristics coincided with the typiclecharacteristics of Saccharothrix. Therefor D34was tentatively assigned to Saccharothrix.The chemotaxonomic characteristics showed that cell wall of K55contains alanine,glutamic acid and meso-DAP, the whole cell sugar contains xylose and arabinose, which istypical of the patten D. Phosphatidyl ethanolamine is the diagnostic phospholipid, which istypical of the type PII, the major menaquinone of strain is MK-12, the main cellular fatty acidis15:0iso(20.53%),17:0iso(12.74%),16:0iso(12.15%). The chemotaxonomiccharacteristics are also accord with chemotaxonomic characteristics of Micromonospora. Thecell wall of D34contains Alanine, glutamate, meso-DAP, the whole cell sugar containsMannose, rhamnose and galactose. Phosphatidyl ethanolamine is the diagnostic phospholipid,which is typical of the type PII, the major menaquinone is MK-9(H4), the main cellular fatty acid is15:0iso(24.06%),16:0iso(15.89%). The chemotaxonomic characteristics are alsoaccord with chemotaxonomic characteristics of Saccharothrix.Phylogenetic analysis based on the16S rRNA sequences indicated that strain k55formeda coherent and monophyletic clade with the closest relatives Micromonospora coxensis2-30-b/28T(98.92%), Micromonospora haikouensis232617T(98.85%), Micromonosporapurpureochromogenes DSM43821T(98.78%), Micromonospora matsumotoense IMSNU22003T(98.78%), Micromonospora siamensis TT2-4T(98.62%). The strain D34is most closely relatedto Saccharothrix espanaensis DSM44229T(98.99%), Saccharothrix variisporea NRRLB-16296T(98.56%), initially assessed that the two potential actinomycetes are novel species.The result of G+C content showed that G+C content of K55is71%, the differences ofG+C content between K55and the reference strains is within10%. The G+C content of D34is64%, the differences of G+C content between D34and the reference strains is within10%.In conclusion, all of these could illustrate that K55is a novel species in the genusMicromonospora, and D34is a novel species in the genus Saccharothrix.